Interacting signal pathways control defense gene expression in Arabidopsis in response to cell wall-degrading enzymes from Erwinia carotovora.

PubMed

Norman-Setterblad, C; Vidal, S; Palva, E T

2000-04-01

We have characterized the role of salicylic acid (SA)-independent defense signaling in Arabidopsis thaliana in response to the plant pathogen Erwinia carotovora subsp. carotovora. Use of pathway-specific target genes as well as signal mutants allowed us to elucidate the role and interactions of ethylene, jasmonic acid (JA), and SA signal pathways in this response. Gene expression studies suggest a central role for both ethylene and JA pathways in the regulation of defense gene expression triggered by the pathogen or by plant cell wall-degrading enzymes (CF) secreted by the pathogen. Our results suggest that ethylene and JA act in concert in this regulation. In addition, CF triggers another, strictly JA-mediated response inhibited by ethylene and SA. SA does not appear to have a major role in activating defense gene expression in response to CF. However, SA may have a dual role in controlling CF-induced gene expression, by enhancing the expression of genes synergistically induced by ethylene and JA and repressing genes induced by JA alone.

Gene Expression of Type VI Secretion System Associated with Environmental Survival in Acidovorax avenae subsp. avenae by Principle Component Analysis

PubMed Central

Cui, Zhouqi; Jin, Guoqiang; Li, Bin; Kakar, Kaleem Ullah; Ojaghian, Mohammad Reza; Wang, Yangli; Xie, Guanlin; Sun, Guochang

2015-01-01

Valine glycine repeat G (VgrG) proteins are regarded as one of two effectors of Type VI secretion system (T6SS) which is a complex multi-component secretion system. In this study, potential biological roles of T6SS structural and VgrG genes in a rice bacterial pathogen, Acidovorax avenae subsp. avenae (Aaa) RS-1, were evaluated under seven stress conditions using principle component analysis of gene expression. The results showed that growth of the pathogen was reduced by H2O2 and paraquat-induced oxidative stress, high salt, low temperature, and vgrG mutation, compared to the control. However, pathogen growth was unaffected by co-culture with a rice rhizobacterium Burkholderia seminalis R456. In addition, expression of 14 T6SS structural and eight vgrG genes was significantly changed under seven conditions. Among different stress conditions, high salt, and low temperature showed a higher effect on the expression of T6SS gene compared with host infection and other environmental conditions. As a first report, this study revealed an association of T6SS gene expression of the pathogen with the host infection, gene mutation, and some common environmental stresses. The results of this research can increase understanding of the biological function of T6SS in this economically-important pathogen of rice. PMID:26378528

Genome-wide gene expression pattern underlying differential host response to high or low pathogenic H5N1 avian influenza virus in ducks.

PubMed

Kumar, A; Vijayakumar, P; Gandhale, P N; Ranaware, P B; Kumar, H; Kulkarni, D D; Raut, A A; Mishra, A

The differences in the influenza viral pathogenesis observed between different pathogenic strains are associated with distinct properties of virus strains and the host immune responses. In order to determine the differences in the duck immune response against two different pathogenic strains, we studied genome-wide host immune gene response of ducks infected with A/duck/India/02CA10/2011 and A/duck/Tripura/103597/2008 H5N1 viruses using custom-designed microarray. A/duck/India/02CA10/2011 is highly pathogenic virus (HP) to ducks, whereas A/duck/Tripura/103597/2008 is a low pathogenic (LP) virus strain. Comparative lung tissue transcriptome analysis of differentially expressed genes revealed that 686 genes were commonly expressed, 880 and 1556 genes are expressed uniquely to infection with HP and LP virus, respectively. The up-regulation of chemokines (CCL4 and CXCR4) and IFN-stimulated genes (IFITM2, STAT3, TGFB1 and TGFB3) was observed in the lung tissues of ducks infected with HP virus. The up-regulation of other immune genes (IL17, OAS, SOCS3, MHC I and MHC II) was observed in both infection conditions. The expression of important antiviral immune genes MX, IFIT5, IFITM5, ISG12, β-defensins, RSAD2, EIF2AK2, TRIM23 and SLC16A3 was observed in LP virus infection, but not in HP virus infection. Several immune-related gene ontology terms and pathways activated by both the viruses were qualitatively similar but quantitatively different. Based on these findings, the differences in the host immune response might explain a part of the difference observed in the viral pathogenesis of high and low pathogenic influenza strains in ducks.

In Planta Stage-Specific Fungal Gene Profiling Elucidates the Molecular Strategies of Fusarium graminearum Growing inside Wheat Coleoptiles[W][OA

PubMed Central

Zhang, Xiao-Wei; Jia, Lei-Jie; Zhang, Yan; Jiang, Gang; Li, Xuan; Zhang, Dong; Tang, Wei-Hua

2012-01-01

The ascomycete Fusarium graminearum is a destructive fungal pathogen of wheat (Triticum aestivum). To better understand how this pathogen proliferates within the host plant, we tracked pathogen growth inside wheat coleoptiles and then examined pathogen gene expression inside wheat coleoptiles at 16, 40, and 64 h after inoculation (HAI) using laser capture microdissection and microarray analysis. We identified 344 genes that were preferentially expressed during invasive growth in planta. Gene expression profiles for 134 putative plant cell wall–degrading enzyme genes suggest that there was limited cell wall degradation at 16 HAI and extensive degradation at 64 HAI. Expression profiles for genes encoding reactive oxygen species (ROS)–related enzymes suggest that F. graminearum primarily scavenges extracellular ROS before a later burst of extracellular ROS is produced by F. graminearum enzymes. Expression patterns of genes involved in primary metabolic pathways suggest that F. graminearum relies on the glyoxylate cycle at an early stage of plant infection. A secondary metabolite biosynthesis gene cluster was specifically induced at 64 HAI and was required for virulence. Our results indicate that F. graminearum initiates infection of coleoptiles using covert penetration strategies and switches to overt cellular destruction of tissues at an advanced stage of infection. PMID:23266949

Expression Profiling of Castanea Genes during Resistant and Susceptible Interactions with the Oomycete Pathogen Phytophthora cinnamomi Reveal Possible Mechanisms of Immunity

PubMed Central

Santos, Carmen; Duarte, Sofia; Tedesco, Sara; Fevereiro, Pedro; Costa, Rita L.

2017-01-01

The most dangerous pathogen affecting the production of chestnuts is Phytophthora cinnamomi a hemibiotrophic that causes root rot, also known as ink disease. Little information has been acquired in chestnut on the molecular defense strategies against this pathogen. The expression of eight candidate genes potentially involved in the defense to P. cinnamomi was quantified by digital PCR in Castanea genotypes showing different susceptibility to the pathogen. Seven of the eight candidate genes displayed differentially expressed levels depending on genotype and time-point after inoculation. Cast_Gnk2-like revealed to be the most expressed gene across all experiments and the one that best discriminates between susceptible and resistant genotypes. Our data suggest that the pre-formed defenses are crucial for the resistance of C. crenata to P. cinnamomi. A lower and delayed expression of the eight studied genes was found in the susceptible Castanea sativa, which may be related with the establishment and spread of the disease in this species. A working model integrating the obtained results is presented. PMID:28443110

Pathogen Phytosensing: Plants to Report Plant Pathogens.

PubMed

Mazarei, Mitra; Teplova, Irina; Hajimorad, M Reza; Stewart, C Neal

2008-04-14

Real-time systems that provide evidence of pathogen contamination in crops can be an important new line of early defense in agricultural centers. Plants possess defense mechanisms to protect against pathogen attack. Inducible plant defense is controlled by signal transduction pathways, inducible promoters and cis-regulatory elements corresponding to key genes involved in defense, and pathogen-specific responses. Identified inducible promoters and cis-acting elements could be utilized in plant sentinels, or 'phytosensors', by fusing these to reporter genes to produce plants with altered phenotypes in response to the presence of pathogens. Here, we have employed cis-acting elements from promoter regions of pathogen inducible genes as well as those responsive to the plant defense signal molecules salicylic acid, jasmonic acid, and ethylene. Synthetic promoters were constructed by combining various regulatory elements supplemented with the enhancer elements from the Cauliflower mosaic virus (CaMV) 35S promoter to increase basal level of the GUS expression. The inducibility of each synthetic promoter was first assessed in transient expression assays using Arabidopsis thaliana protoplasts and then examined for efficacy in stably transgenic Arabidopsis and tobacco plants. Histochemical and fluorometric GUS expression analyses showed that both transgenic Arabidopsis and tobacco plants responded to elicitor and phytohormone treatments with increased GUS expression when compared to untreated plants. Pathogen-inducible phytosensor studies were initiated by analyzing the sensitivity of the synthetic promoters against virus infection. Transgenic tobacco plants infected with Alfalfa mosaic virus showed an increase in GUS expression when compared to mock-inoculated control plants, whereas Tobacco mosaic virus infection caused no changes in GUS expression. Further research, using these transgenic plants against a range of different pathogens with the regulation of detectable reporter gene could provide biological evidence to define the functional differences between pathogens, and provide new technology and applications for transgenic plants as phytosensors.

Expression of AtWRKY33 encoding a pathogen- or PAMP-responsive WRKY transcription factor is regulated by a composite DNA motif containing W box elements.

PubMed

Lippok, Bernadette; Birkenbihl, Rainer P; Rivory, Gaelle; Brümmer, Janna; Schmelzer, Elmon; Logemann, Elke; Somssich, Imre E

2007-04-01

WRKY transcription factors regulate distinct parts of the plant defense transcriptome. Expression of many WRKY genes themselves is induced by pathogens or pathogen-mimicking molecules. Here, we demonstrate that Arabidopsis WRKY33 responds to various stimuli associated with plant defense as well as to different kinds of phytopathogens. Although rapid pathogen-induced AtWRKY33 expression does not require salicylic acid (SA) signaling, it is dependent on PAD4, a key regulator upstream of SA. Activation of AtWRKY33 is independent of de novo protein synthesis, suggesting that it is at least partly under negative regulatory control. We show that a set of three WRKY-specific cis-acting DNA elements (W boxes) within the AtWRKY33 promoter is required for efficient pathogen- or PAMP-triggered gene activation. This strongly indicates that WRKY transcription factors are major components of the regulatory machinery modulating immediate to early expression of this gene in response to pathogen attack.

Transcriptome profiling during a natural host-parasite interaction.

PubMed

McTaggart, Seanna J; Cézard, Timothée; Garbutt, Jennie S; Wilson, Phil J; Little, Tom J

2015-08-28

Infection outcome in some coevolving host-pathogens is characterised by host-pathogen genetic interactions, where particular host genotypes are susceptible only to a subset of pathogen genotypes. To identify candidate genes responsible for the infection status of the host, we exposed a Daphnia magna host genotype to two bacterial strains of Pasteuria ramosa, one of which results in infection, while the other does not. At three time points (four, eight and 12 h) post pathogen exposure, we sequenced the complete transcriptome of the hosts using RNA-Seq (Illumina). We observed a rapid and transient response to pathogen treatment. Specifically, at the four-hour time point, eight genes were differentially expressed. At the eight-hour time point, a single gene was differentially expressed in the resistant combination only, and no genes were differentially expressed at the 12-h time point. We found that pathogen-associated transcriptional activity is greatest soon after exposure. Genome-wide resistant combinations were more likely to show upregulation of genes, while susceptible combinations were more likely to be downregulated, relative to controls. Our results also provide several novel candidate genes that may play a pivotal role in determining infection outcomes.

Tanscriptomic Study of the Soybean-Fusarium virguliforme Interaction Revealed a Novel Ankyrin-Repeat Containing Defense Gene, Expression of Whose during Infection Led to Enhanced Resistance to the Fungal Pathogen in Transgenic Soybean Plants

PubMed Central

Ngaki, Micheline N.; Wang, Bing; Sahu, Binod B.; Srivastava, Subodh K.; Farooqi, Mohammad S.; Kambakam, Sekhar; Swaminathan, Sivakumar

2016-01-01

Fusarium virguliforme causes the serious disease sudden death syndrome (SDS) in soybean. Host resistance to this pathogen is partial and is encoded by a large number of quantitative trait loci, each conditioning small effects. Breeding SDS resistance is therefore challenging and identification of single-gene encoded novel resistance mechanisms is becoming a priority to fight this devastating this fungal pathogen. In this transcriptomic study we identified a few putative soybean defense genes, expression of which is suppressed during F. virguliforme infection. The F. virguliforme infection-suppressed genes were broadly classified into four major classes. The steady state transcript levels of many of these genes were suppressed to undetectable levels immediately following F. virguliforme infection. One of these classes contains two novel genes encoding ankyrin repeat-containing proteins. Expression of one of these genes, GmARP1, during F. virguliforme infection enhances SDS resistance among the transgenic soybean plants. Our data suggest that GmARP1 is a novel defense gene and the pathogen presumably suppress its expression to establish compatible interaction. PMID:27760122

Identification of pathogenic genes related to rheumatoid arthritis through integrated analysis of DNA methylation and gene expression profiling.

PubMed

Zhang, Lei; Ma, Shiyun; Wang, Huailiang; Su, Hang; Su, Ke; Li, Longjie

2017-11-15

The purpose of our study was to identify new pathogenic genes used for exploring the pathogenesis of rheumatoid arthritis (RA). To screen pathogenic genes of RA, an integrated analysis was performed by using the microarray datasets in RA derived from the Gene Expression Omnibus (GEO) database. The functional annotation and potential pathways of differentially expressed genes (DEGs) were further discovered by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Afterwards, the integrated analysis of DNA methylation and gene expression profiling was used to screen crucial genes. In addition, we used RT-PCR and MSP to verify the expression levels and methylation status of these crucial genes in 20 synovial biopsy samples obtained from 10 RA model mice and 10 normal mice. BCL11B, CCDC88C, FCRLA and APOL6 were both up-regulated and hypomethylated in RA according to integrated analysis, RT-PCR and MSP verification. Four crucial genes (BCL11B, CCDC88C, FCRLA and APOL6) identified and analyzed in this study might be closely connected with the pathogenesis of RA. Copyright © 2017. Published by Elsevier B.V.

Concurrent host-pathogen gene expression in the lungs of pigs challenged with Actinobacillus pleuropneumoniae.

PubMed

Brogaard, Louise; Klitgaard, Kirstine; Heegaard, Peter M H; Hansen, Mette Sif; Jensen, Tim Kåre; Skovgaard, Kerstin

2015-05-28

Actinobacillus pleuropneumoniae causes pleuropneumonia in pigs, a disease which is associated with high morbidity and mortality, as well as impaired animal welfare. To obtain in-depth understanding of this infection, the interplay between virulence factors of the pathogen and defense mechanisms of the porcine host needs to be elucidated. However, research has traditionally focused on either bacteriology or immunology; an unbiased picture of the transcriptional responses can be obtained by investigating both organisms in the same biological sample. Host and pathogen responses in pigs experimentally infected with A. pleuropneumoniae were analyzed by high-throughput RT-qPCR. This approach allowed concurrent analysis of selected genes encoding proteins known or hypothesized to be important in the acute phase of this infection. The expression of 17 bacterial and 31 porcine genes was quantified in lung samples obtained within the first 48 hours of infection. This provided novel insight into the early time course of bacterial genes involved in synthesis of pathogen-associated molecular patterns (lipopolysaccharide, peptidoglycan, lipoprotein) and genes involved in pattern recognition (TLR4, CD14, MD2, LBP, MYD88) in response to A. pleuropneumoniae. Significant up-regulation of proinflammatory cytokines such as IL1B, IL6, and IL8 was observed, correlating with protein levels, infection status and histopathological findings. Host genes encoding proteins involved in iron metabolism, as well as bacterial genes encoding exotoxins, proteins involved in adhesion, and iron acquisition were found to be differentially expressed according to disease progression. By applying laser capture microdissection, porcine expression of selected genes could be confirmed in the immediate surroundings of the invading pathogen. Microbial pathogenesis is the product of interactions between host and pathogen. Our results demonstrate the applicability of high-throughput RT-qPCR for the elucidation of dual-organism gene expression analysis during infection. We showed differential expression of 12 bacterial and 24 porcine genes during infection and significant correlation of porcine and bacterial gene expression. This is the first study investigating the concurrent transcriptional response of both bacteria and host at the site of infection during porcine respiratory infection.

Ectopic Expression of the Wild Grape WRKY Transcription Factor VqWRKY52 in Arabidopsis thaliana Enhances Resistance to the Biotrophic Pathogen Powdery Mildew But Not to the Necrotrophic Pathogen Botrytis cinerea.

PubMed

Wang, Xianhang; Guo, Rongrong; Tu, Mingxing; Wang, Dejun; Guo, Chunlei; Wan, Ran; Li, Zhi; Wang, Xiping

2017-01-01

WRKY transcription factors are known to play important roles in plant responses to biotic stresses. We previously showed that the expression of the WRKY gene, VqWRKY52 , from Chinese wild Vitis quinquangularis was strongly induced 24 h post inoculation with powdery mildew. In this study, we analyzed the expression levels of VqWRKY52 following treatment with the defense related hormones salicylic acid (SA) and methyl jasmonate, revealing that VqWRKY52 was strongly induced by SA but not JA. We characterized the VqWRKY52 gene, which encodes a WRKY III gene family member, and found that ectopic expression in Arabidopsis thaliana enhanced resistance to powdery mildew and Pseudomonas syringae pv. tomato DC3000, but increased susceptibility to Botrytis cinerea , compared with wild type (WT) plants. The transgenic A. thaliana lines displayed strong cell death induced by the biotrophic powdery mildew pathogen, the hemibiotrophic P. syringe pathogen and the necrotrophic pathogen B. cinerea . In addition, the relative expression levels of various defense-related genes were compared between the transgenic A. thaliana lines and WT plants following the infection by different pathogens. Collectively, the results indicated that VqWRKY52 plays essential roles in the SA dependent signal transduction pathway and that it can enhance the hypersensitive response cell death triggered by microbial pathogens.

Antibiosis functions during interactions of Trichoderma afroharzianum and Trichoderma gamsii with plant pathogenic Rhizoctonia and Pythium.

PubMed

Zhang, Xinjian; Harvey, Paul R; Stummer, Belinda E; Warren, Rosemary A; Zhang, Guangzhi; Guo, Kai; Li, Jishun; Yang, Hetong

2015-09-01

Trichoderma afroharzianum is one of the best characterized Trichoderma species, and strains have been utilized as plant disease suppressive inoculants. In contrast, Trichoderma gamsii has only recently been described, and there is limited knowledge of its disease suppressive efficacies. Comparative studies of changes in gene expression during interactions of these species with their target plant pathogens will provide fundamental information on pathogen antibiosis functions. In the present study, we used complementary DNA amplified fragment length polymorphism (cDNA-AFLP) analysis to investigate changes in transcript profiling of T. afroharzianum strain LTR-2 and T. gamsii strain Tk7a during in vitro interactions with plant pathogenic Rhizoctonia solani and Pythium irregulare. Considerable differences were resolved in the overall expression profiles of strains LTR-2 and Tk7a when challenged with either plant pathogen. In strain LTR-2, previously reported mycoparasitism-related genes such as chitinase, polyketide synthase, and non-ribosomal peptide synthetase were found to be differentially expressed. This was not so for strain Tk7a, with the only previously reported antibiosis-associated genes being small secreted cysteine-rich proteins. Although only one differentially expressed gene was common to both strains LTR-2 and Tk7a, numerous genes reportedly associated with pathogen antibiosis processes were differentially expressed in both strains, including degradative enzymes and membrane transport proteins. A number of novel potential antibiosis-related transcripts were found from strains LTR-2 and Tk7a and remain to be identified. The expression kinetics of 20 Trichoderma (10 from strain LTR-2, 10 from strain Tk7a) transcript-derived fragments (TDFs) were quantified by quantitative reverse transcription PCR (RT-qPCR) at pre- and post-mycelia contact stages of Trichoderma-prey interactions, thereby confirming differential gene expression. Collectively, this research is providing information to elucidate the antibiosis mechanisms and disease suppressive activities of T. afroharzianum and T. gamsii against soilborne fungal and oomycete plant pathogens.

Transcriptome profiling of soybean (Glycine max) roots challenged with pathogenic and non-pathogenic isolates of Fusarium oxysporum.

PubMed

Lanubile, Alessandra; Muppirala, Usha K; Severin, Andrew J; Marocco, Adriano; Munkvold, Gary P

2015-12-21

Fusarium oxysporum is one of the most common fungal pathogens causing soybean root rot and seedling blight in U.S.A. In a recent study, significant variation in aggressiveness was observed among isolates of F. oxysporum collected from roots in Iowa, ranging from highly pathogenic to weakly or non-pathogenic isolates. We used RNA-seq analysis to investigate the molecular aspects of the interactions of a partially resistant soybean genotype with non-pathogenic/pathogenic isolates of F. oxysporum at 72 and 96 h post inoculation (hpi). Markedly different gene expression profiles were observed in response to the two isolates. A peak of highly differentially expressed genes (HDEGs) was triggered at 72 hpi in soybean roots and the number of HDEGs was about eight times higher in response to the pathogenic isolate compared to the non-pathogenic one (1,659 vs. 203 HDEGs, respectively). Furthermore, the magnitude of induction was much greater in response to the pathogenic isolate. This response included a stronger activation of defense-related genes, transcription factors, and genes involved in ethylene biosynthesis, secondary and sugar metabolism. The obtained data provide an important insight into the transcriptional responses of soybean-F. oxysporum interactions and illustrate the more drastic changes in the host transcriptome in response to the pathogenic isolate. These results may be useful in the developing new methods of broadening resistance of soybean to F. oxysporum, including the over-expression of key soybean genes.

Early indicators of exposure to biological threat agents using host gene profiles in peripheral blood mononuclear cells

PubMed Central

Das, Rina; Hammamieh, Rasha; Neill, Roger; Ludwig, George V; Eker, Steven; Lincoln, Patrick; Ramamoorthy, Preveen; Dhokalia, Apsara; Mani, Sachin; Mendis, Chanaka; Cummings, Christiano; Kearney, Brian; Royaee, Atabak; Huang, Xiao-Zhe; Paranavitana, Chrysanthi; Smith, Leonard; Peel, Sheila; Kanesa-Thasan, Niranjan; Hoover, David; Lindler, Luther E; Yang, David; Henchal, Erik; Jett, Marti

2008-01-01

Background Effective prophylaxis and treatment for infections caused by biological threat agents (BTA) rely upon early diagnosis and rapid initiation of therapy. Most methods for identifying pathogens in body fluids and tissues require that the pathogen proliferate to detectable and dangerous levels, thereby delaying diagnosis and treatment, especially during the prelatent stages when symptoms for most BTA are indistinguishable flu-like signs. Methods To detect exposures to the various pathogens more rapidly, especially during these early stages, we evaluated a suite of host responses to biological threat agents using global gene expression profiling on complementary DNA arrays. Results We found that certain gene expression patterns were unique to each pathogen and that other gene changes occurred in response to multiple agents, perhaps relating to the eventual course of illness. Nonhuman primates were exposed to some pathogens and the in vitro and in vivo findings were compared. We found major gene expression changes at the earliest times tested post exposure to aerosolized B. anthracis spores and 30 min post exposure to a bacterial toxin. Conclusion Host gene expression patterns have the potential to serve as diagnostic markers or predict the course of impending illness and may lead to new stage-appropriate therapeutic strategies to ameliorate the devastating effects of exposure to biothreat agents. PMID:18667072

RNA Sequencing Reveals Differences between the Global Transcriptomes of Salmonella enterica Serovar Enteritidis Strains with High and Low Pathogenicities

PubMed Central

2014-01-01

Salmonella enterica serovar Enteritidis is one of the important causes of bacterial food-borne gastroenteritis worldwide. Field strains of S. Enteritidis are relatively genetically homogeneous; however, they show extensive phenotypic diversity and differences in virulence potential. RNA sequencing (RNA-Seq) was used to characterize differences in the global transcriptome between several genetically similar but phenotypically diverse poultry-associated field strains of S. Enteritidis grown in laboratory medium at avian body temperature (42°C). These S. Enteritidis strains were previously characterized as high-pathogenicity (HP; n = 3) and low-pathogenicity (LP; n = 3) strains based on both in vitro and in vivo virulence assays. Using the negative binomial distribution-based statistical tools edgeR and DESeq, 252 genes were identified as differentially expressed in LP strains compared with their expression in the HP strains (P < 0.05). A majority of genes (235, or 93.2%) showed significantly reduced expression, whereas a few genes (17, or 6.8%) showed increased expression in all LP strains compared with HP strains. LP strains showed a unique transcriptional profile that is characterized by significantly reduced expression of several transcriptional regulators and reduced expression of genes involved in virulence (e.g., Salmonella pathogenicity island 1 [SPI-1], SPI-5, and fimbrial and motility genes) and protection against osmotic, oxidative, and other stresses, such as iron-limiting conditions commonly encountered within the host. Several functionally uncharacterized genes also showed reduced expression. This study provides a first concise view of the global transcriptional differences between field strains of S. Enteritidis with various levels of pathogenicity, providing the basis for future functional characterization of several genes with potential roles in virulence or stress regulation of S. Enteritidis. PMID:24271167

Comparative Temporal Transcriptome Profiling of Wheat near Isogenic Line Carrying Lr57 under Compatible and Incompatible Interactions

PubMed Central

Yadav, Inderjit S.; Sharma, Amandeep; Kaur, Satinder; Nahar, Natasha; Bhardwaj, Subhash C.; Sharma, Tilak R.; Chhuneja, Parveen

2016-01-01

Leaf rust caused by Puccinia triticina (Pt) is one of the most important diseases of bread wheat globally. Recent advances in sequencing technologies have provided opportunities to analyse the complete transcriptomes of the host as well as pathogen for studying differential gene expression during infection. Pathogen induced differential gene expression was characterized in a near isogenic line carrying leaf rust resistance gene Lr57 and susceptible recipient genotype WL711. RNA samples were collected at five different time points 0, 12, 24, 48, and 72 h post inoculation (HPI) with Pt 77-5. A total of 3020 transcripts were differentially expressed with 1458 and 2692 transcripts in WL711 and WL711+Lr57, respectively. The highest number of differentially expressed transcripts was detected at 12 HPI. Functional categorization using Blast2GO classified the genes into biological processes, molecular function and cellular components. WL711+Lr57 showed much higher number of differentially expressed nucleotide binding and leucine rich repeat genes and expressed more protein kinases and pathogenesis related proteins such as chitinases, glucanases and other PR proteins as compared to susceptible genotype. Pathway annotation with KEGG categorized genes into 13 major classes with carbohydrate metabolism being the most prominent followed by amino acid, secondary metabolites, and nucleotide metabolism. Gene co-expression network analysis identified four and eight clusters of highly correlated genes in WL711 and WL711+Lr57, respectively. Comparative analysis of the differentially expressed transcripts led to the identification of some transcripts which were specifically expressed only in WL711+Lr57. It was apparent from the whole transcriptome sequencing that the resistance gene Lr57 directed the expression of different genes involved in building the resistance response in the host to combat invading pathogen. The RNAseq data and differentially expressed transcripts identified in present study is a genomic resource which can be used for further studying the host pathogen interaction for Lr57 and wheat transcriptome in general. PMID:28066494

Effect of Periodontal Pathogens on the Metatranscriptome of a Healthy Multispecies Biofilm Model

PubMed Central

Duran-Pinedo, Ana

2012-01-01

Oral bacterial biofilms are highly complex microbial communities with up to 700 different bacterial taxa. We report here the use of metatranscriptomic analysis to study patterns of community gene expression in a multispecies biofilm model composed of species found in healthy oral biofilms (Actinomyces naeslundii, Lactobacillus casei, Streptococcus mitis, Veillonella parvula, and Fusobacterium nucleatum) and the same biofilm plus the periodontopathogens Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans. The presence of the periodontopathogens altered patterns in gene expression, and data indicate that transcription of protein-encoding genes and small noncoding RNAs is stimulated. In the healthy biofilm hypothetical proteins, transporters and transcriptional regulators were upregulated while chaperones and cell division proteins were downregulated. However, when the pathogens were present, chaperones were highly upregulated, probably due to increased levels of stress. We also observed a significant upregulation of ABC transport systems and putative transposases. Changes in Clusters of Orthologous Groups functional categories as well as gene set enrichment analysis based on Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways showed that in the absence of pathogens, only sets of proteins related to transport and secondary metabolism were upregulated, while in the presence of pathogens, proteins related to growth and division as well as a large portion of transcription factors were upregulated. Finally, we identified several small noncoding RNAs whose predicted targets were genes differentially expressed in the open reading frame libraries. These results show the importance of pathogens controlling gene expression of a healthy oral community and the usefulness of metatranscriptomic techniques to study gene expression profiles in complex microbial community models. PMID:22328675

cDNA-AFLP analysis reveals differential gene expression in compatible interaction of wheat challenged with Puccinia striiformis f. sp. tritici

PubMed Central

Wang, Xiaojie; Tang, Chunlei; Zhang, Gang; Li, Yingchun; Wang, Chenfang; Liu, Bo; Qu, Zhipeng; Zhao, Jie; Han, Qingmei; Huang, Lili; Chen, Xianming; Kang, Zhensheng

2009-01-01

Background Puccinia striiformis f. sp. tritici is a fungal pathogen causing stripe rust, one of the most important wheat diseases worldwide. The fungus is strictly biotrophic and thus, completely dependent on living host cells for its reproduction, which makes it difficult to study genes of the pathogen. In spite of its economic importance, little is known about the molecular basis of compatible interaction between the pathogen and wheat host. In this study, we identified wheat and P. striiformis genes associated with the infection process by conducting a large-scale transcriptomic analysis using cDNA-AFLP. Results Of the total 54,912 transcript derived fragments (TDFs) obtained using cDNA-AFLP with 64 primer pairs, 2,306 (4.2%) displayed altered expression patterns after inoculation, of which 966 showed up-regulated and 1,340 down-regulated. 186 TDFs produced reliable sequences after sequencing of 208 TDFs selected, of which 74 (40%) had known functions through BLAST searching the GenBank database. Majority of the latter group had predicted gene products involved in energy (13%), signal transduction (5.4%), disease/defence (5.9%) and metabolism (5% of the sequenced TDFs). BLAST searching of the wheat stem rust fungus genome database identified 18 TDFs possibly from the stripe rust pathogen, of which 9 were validated of the pathogen origin using PCR-based assays followed by sequencing confirmation. Of the 186 reliable TDFs, 29 homologous to genes known to play a role in disease/defense, signal transduction or uncharacterized genes were further selected for validation of cDNA-AFLP expression patterns using qRT-PCR analyses. Results confirmed the altered expression patterns of 28 (96.5%) genes revealed by the cDNA-AFLP technique. Conclusion The results show that cDNA-AFLP is a reliable technique for studying expression patterns of genes involved in the wheat-stripe rust interactions. Genes involved in compatible interactions between wheat and the stripe rust pathogen were identified and their expression patterns were determined. The present study should be helpful in elucidating the molecular basis of the infection process, and identifying genes that can be targeted for inhibiting the growth and reproduction of the pathogen. Moreover, this study can also be used to elucidate the defence responses of the genes that were of plant origin. PMID:19566949

A network approach of gene co-expression in the zea mays/Aspergillus flavus pathosystem to map host/pathogen interaction pathways

USDA-ARS?s Scientific Manuscript database

A gene co-expression network was generated using a dual RNA-seq study with the fungal pathogen A. flavus and its plant host Z. mays during the initial 3 days of infection. The analysis deciphered novel pathways and mapped genes of interest in both organisms during the infection. This network reveal...

Expressed sequence tags from the flower pathogen Claviceps purpurea.

PubMed

Oeser, Birgitt; Beaussart, François; Haarmann, Thomas; Lorenz, Nicole; Nathues, Eva; Rolke, Yvonne; Scheffer, Jan; Weiner, January; Tudzynski, Paul

2009-09-01

SUMMARY The ascomycete Claviceps purpurea (ergot) is a biotrophic flower pathogen of rye and other grasses. The deleterious toxic effects of infected rye seeds on humans and grazing animals have been known since the Middle Ages. To gain further insight into the molecular basis of this disease, we generated about 10 000 expressed sequence tags (ESTs)-about 25% originating from axenic fungal culture and about 75% from tissues collected 6-20 days after infection of rye spikes. The pattern of axenic vs. in planta gene expression was compared. About 200 putative plant genes were identified within the in planta library. A high percentage of these were predicted to function in plant defence against the ergot fungus and other pathogens, for example pathogenesis-related proteins. Potential fungal pathogenicity and virulence genes were found via comparison with the pathogen-host interaction database (PHI-base; http://www.phi-base.org) and with genes known to be highly expressed in the haustoria of the bean rust fungus. Comparative analysis of Claviceps and two other fungal flower pathogens (necrotrophic Fusarium graminearum and biotrophic Ustilago maydis) highlighted similarities and differences in their lifestyles, for example all three fungi have signalling components and cell wall-degrading enzymes in their arsenal. In summary, the analysis of axenic and in planta ESTs yielded a collection of candidate genes to be evaluated for functional roles in this plant-microbe interaction.

Host-Induced Silencing of Pathogenicity Genes Enhances Resistance to Fusarium oxysporum Wilt in Tomato.

PubMed

Bharti, Poonam; Jyoti, Poonam; Kapoor, Priya; Sharma, Vandana; Shanmugam, V; Yadav, Sudesh Kumar

2017-08-01

This study presents a novel approach of controlling vascular wilt in tomato by RNAi expression directed to pathogenicity genes of Fusarium oxysporum f. sp. lycopersici. Vascular wilt of tomato caused by Fusarium oxysporum f. sp. lycopersici leads to qualitative and quantitative loss of the crop. Limitation in the existing control measures necessitates the development of alternative strategies to increase resistance in the plants against pathogens. Recent findings paved way to RNAi, as a promising method for silencing of pathogenicity genes in fungus and provided effective resistance against fungal pathogens. Here, two important pathogenicity genes FOW2, a Zn(II)2Cys6 family putative transcription regulator, and chsV, a putative myosin motor and a chitin synthase domain, were used for host-induced gene silencing through hairpinRNA cassettes of these genes against Fusarium oxysporum f. sp. lycopersici. HairpinRNAs were assembled in appropriate binary vectors and transformed into tomato plant targeting FOW2 and chsV genes, for two highly pathogenic strains of Fusarium oxysporum viz. TOFOL-IHBT and TOFOL-IVRI. Transgenic tomatoes were analyzed for possible attainment of resistance in transgenic lines against fungal infection. Eight transgenic lines expressing hairpinRNA cassettes showed trivial disease symptoms after 6-8 weeks of infection. Hence, the host-induced posttranscriptional gene silencing of pathogenicity genes in transgenic tomato plants has enhanced their resistance to vascular wilt disease caused by Fusarium oxysporum.

Ectopic Expression of Xylella fastidiosa rpfF Conferring Production of Diffusible Signal Factor in Transgenic Tobacco and Citrus Alters Pathogen Behavior and Reduces Disease Severity.

PubMed

Caserta, R; Souza-Neto, R R; Takita, M A; Lindow, S E; De Souza, A A

2017-11-01

The pathogenicity of Xylella fastidiosa is associated with its ability to colonize the xylem of host plants. Expression of genes contributing to xylem colonization are suppressed, while those necessary for insect vector acquisition are increased with increasing concentrations of diffusible signal factor (DSF), whose production is dependent on RpfF. We previously demonstrated that transgenic citrus plants ectopically expressing rpfF from a citrus strain of X. fastidiosa subsp. pauca exhibited less susceptibility to Xanthomonas citri subsp. citri, another pathogen whose virulence is modulated by DSF accumulation. Here, we demonstrate that ectopic expression of rpfF in both transgenic tobacco and sweet orange also confers a reduction in disease severity incited by X. fastidiosa and reduces its colonization of those plants. Decreased disease severity in the transgenic plants was generally associated with increased expression of genes conferring adhesiveness to the pathogen and decreased expression of genes necessary for active motility, accounting for the reduced population sizes achieved in the plants, apparently by limiting pathogen dispersal through the plant. Plant-derived DSF signal molecules in a host plant can, therefore, be exploited to interfere with more than one pathogen whose virulence is controlled by DSF signaling.

Candidate innate immune system gene expression in the ecological model Daphnia

PubMed Central

Decaestecker, Ellen; Labbé, Pierrick; Ellegaard, Kirsten; Allen, Judith E.; Little, Tom J.

2011-01-01

The last ten years have witnessed increasing interest in host–pathogen interactions involving invertebrate hosts. The invertebrate innate immune system is now relatively well characterised, but in a limited range of genetic model organisms and under a limited number of conditions. Immune systems have been little studied under real-world scenarios of environmental variation and parasitism. Thus, we have investigated expression of candidate innate immune system genes in the water flea Daphnia, a model organism for ecological genetics, and whose capacity for clonal reproduction facilitates an exceptionally rigorous control of exposure dose or the study of responses at many time points. A unique characteristic of the particular Daphnia clones and pathogen strain combinations used presently is that they have been shown to be involved in specific host–pathogen coevolutionary interactions in the wild. We choose five genes, which are strong candidates to be involved in Daphnia–pathogen interactions, given that they have been shown to code for immune effectors in related organisms. Differential expression of these genes was quantified by qRT-PCR following exposure to the bacterial pathogen Pasteuria ramosa. Constitutive expression levels differed between host genotypes, and some genes appeared to show correlated expression. However, none of the genes appeared to show a major modification of expression level in response to Pasteuria exposure. By applying knowledge from related genetic model organisms (e.g. Drosophila) to models for the study of evolutionary ecology and coevolution (i.e. Daphnia), the candidate gene approach is temptingly efficient. However, our results show that detection of only weak patterns is likely if one chooses target genes for study based on previously identified genome sequences by comparison to homologues from other related organisms. Future work on the Daphnia–Pasteuria system will need to balance a candidate gene approach with more comprehensive approaches to de novo identify immune system genes specific to the Daphnia–Pasteuria interaction. PMID:21550363

Candidate innate immune system gene expression in the ecological model Daphnia.

PubMed

Decaestecker, Ellen; Labbé, Pierrick; Ellegaard, Kirsten; Allen, Judith E; Little, Tom J

2011-10-01

The last ten years have witnessed increasing interest in host-pathogen interactions involving invertebrate hosts. The invertebrate innate immune system is now relatively well characterised, but in a limited range of genetic model organisms and under a limited number of conditions. Immune systems have been little studied under real-world scenarios of environmental variation and parasitism. Thus, we have investigated expression of candidate innate immune system genes in the water flea Daphnia, a model organism for ecological genetics, and whose capacity for clonal reproduction facilitates an exceptionally rigorous control of exposure dose or the study of responses at many time points. A unique characteristic of the particular Daphnia clones and pathogen strain combinations used presently is that they have been shown to be involved in specific host-pathogen coevolutionary interactions in the wild. We choose five genes, which are strong candidates to be involved in Daphnia-pathogen interactions, given that they have been shown to code for immune effectors in related organisms. Differential expression of these genes was quantified by qRT-PCR following exposure to the bacterial pathogen Pasteuria ramosa. Constitutive expression levels differed between host genotypes, and some genes appeared to show correlated expression. However, none of the genes appeared to show a major modification of expression level in response to Pasteuria exposure. By applying knowledge from related genetic model organisms (e.g. Drosophila) to models for the study of evolutionary ecology and coevolution (i.e. Daphnia), the candidate gene approach is temptingly efficient. However, our results show that detection of only weak patterns is likely if one chooses target genes for study based on previously identified genome sequences by comparison to homologues from other related organisms. Future work on the Daphnia-Pasteuria system will need to balance a candidate gene approach with more comprehensive approaches to de novo identify immune system genes specific to the Daphnia-Pasteuria interaction. Copyright © 2011 Elsevier Ltd. All rights reserved.

Cell-Type–Specific Transcriptional Profiles of the Dimorphic Pathogen Penicillium marneffei Reflect Distinct Reproductive, Morphological, and Environmental Demands

PubMed Central

Pasricha, Shivani; Payne, Michael; Canovas, David; Pase, Luke; Ngaosuwankul, Nathamon; Beard, Sally; Oshlack, Alicia; Smyth, Gordon K.; Chaiyaroj, Sansanee C.; Boyce, Kylie J.; Andrianopoulos, Alex

2013-01-01

Penicillium marneffei is an opportunistic human pathogen endemic to Southeast Asia. At 25° P. marneffei grows in a filamentous hyphal form and can undergo asexual development (conidiation) to produce spores (conidia), the infectious agent. At 37° P. marneffei grows in the pathogenic yeast cell form that replicates by fission. Switching between these growth forms, known as dimorphic switching, is dependent on temperature. To understand the process of dimorphic switching and the physiological capacity of the different cell types, two microarray-based profiling experiments covering approximately 42% of the genome were performed. The first experiment compared cells from the hyphal, yeast, and conidiation phases to identify “phase or cell-state–specific” gene expression. The second experiment examined gene expression during the dimorphic switch from one morphological state to another. The data identified a variety of differentially expressed genes that have been organized into metabolic clusters based on predicted function and expression patterns. In particular, C-14 sterol reductase–encoding gene ergM of the ergosterol biosynthesis pathway showed high-level expression throughout yeast morphogenesis compared to hyphal. Deletion of ergM resulted in severe growth defects with increased sensitivity to azole-type antifungal agents but not amphotericin B. The data defined gene classes based on spatio-temporal expression such as those expressed early in the dimorphic switch but not in the terminal cell types and those expressed late. Such classifications have been helpful in linking a given gene of interest to its expression pattern throughout the P. marneffei dimorphic life cycle and its likely role in pathogenicity. PMID:24062530

Pathogen Phytosensing: Plants to Report Plant Pathogens

PubMed Central

Mazarei, Mitra; Teplova, Irina; Hajimorad, M. Reza; Stewart, C. Neal

2008-01-01

Real-time systems that provide evidence of pathogen contamination in crops can be an important new line of early defense in agricultural centers. Plants possess defense mechanisms to protect against pathogen attack. Inducible plant defense is controlled by signal transduction pathways, inducible promoters and cis-regulatory elements corresponding to key genes involved in defense, and pathogen-specific responses. Identified inducible promoters and cis-acting elements could be utilized in plant sentinels, or ‘phytosensors’, by fusing these to reporter genes to produce plants with altered phenotypes in response to the presence of pathogens. Here, we have employed cis-acting elements from promoter regions of pathogen inducible genes as well as those responsive to the plant defense signal molecules salicylic acid, jasmonic acid, and ethylene. Synthetic promoters were constructed by combining various regulatory elements supplemented with the enhancer elements from the Cauliflower mosaic virus (CaMV) 35S promoter to increase basal level of the GUS expression. The inducibility of each synthetic promoter was first assessed in transient expression assays using Arabidopsis thaliana protoplasts and then examined for efficacy in stably transgenic Arabidopsis and tobacco plants. Histochemical and fluorometric GUS expression analyses showed that both transgenic Arabidopsis and tobacco plants responded to elicitor and phytohormone treatments with increased GUS expression when compared to untreated plants. Pathogen-inducible phytosensor studies were initiated by analyzing the sensitivity of the synthetic promoters against virus infection. Transgenic tobacco plants infected with Alfalfa mosaic virus showed an increase in GUS expression when compared to mock-inoculated control plants, whereas Tobacco mosaic virus infection caused no changes in GUS expression. Further research, using these transgenic plants against a range of different pathogens with the regulation of detectable reporter gene could provide biological evidence to define the functional differences between pathogens, and provide new technology and applications for transgenic plants as phytosensors. PMID:27879840

Transgenic tobacco expressing a modified spider peptide inhibits the growth of plant pathogens and insect larvae

USDA-ARS?s Scientific Manuscript database

The gene encoding lycotoxin I, an amphipathic pore-forming peptide, was modified to increase oral toxicity to insects. One of the most active modified genes was then constitutively expressed in tobacco (Nicotiana tabacum) and transformants were evaluated for insect and disease resistance. Pathogenic...

Avian influenza rapidly induces antiviral genes in duck lung and intestine

PubMed Central

Vanderven, Hillary A.; Petkau, Kristina; Ryan-Jean, Kieran E. E.; Aldridge, Jerry R.; Webster, Robert G.; Magor, Katharine E.

2012-01-01

Ducks are the natural reservoir of influenza A and survive infection by most strains. To characterize the duck immune response to influenza, we sought to identify innate immune genes expressed early in an infection. We used suppressive subtractive hybridization (SSH) to construct 3 libraries enriched in differentially expressed genes from lung RNA of a duck infected with highly pathogenic avian influenza virus A/Vietnam/1203/04 (H5N1), or lung and intestine RNA of a duck infected with low pathogenic avian influenza A/mallard/BC/500/05 (H5N2) compared to a mock-infected duck. Sequencing of 1687 clones identified a transcription profile enriched in genes involved in antiviral defense and other cellular processes. Major histocompatibility complex class I (MHC I), interferon induced protein with tricopeptide repeats 5 (IFIT5), and 2′–5′oligoadenylate synthetase-like gene (OASL) were increased more than 1000-fold in relative transcript abundance in duck lung at 1 dpi with highly pathogenic VN1203. These genes were induced much less in lung or intestine following infection with low pathogenic BC500. The expression of these genes following infection suggests that ducks initiate an immediate and robust response to a potentially lethal influenza strain, and a minimal response a low pathogenic strain. PMID:22534314

DNA-Demethylase Regulated Genes Show Methylation-Independent Spatiotemporal Expression Patterns

PubMed Central

Schumann, Ulrike; Lee, Joanne; Kazan, Kemal; Ayliffe, Michael; Wang, Ming-Bo

2017-01-01

Recent research has indicated that a subset of defense-related genes is downregulated in the Arabidopsis DNA demethylase triple mutant rdd (ros1 dml2 dml3) resulting in increased susceptibility to the fungal pathogen Fusarium oxysporum. In rdd plants these downregulated genes contain hypermethylated transposable element sequences (TE) in their promoters, suggesting that this methylation represses gene expression in the mutant and that these sequences are actively demethylated in wild-type plants to maintain gene expression. In this study, the tissue-specific and pathogen-inducible expression patterns of rdd-downregulated genes were investigated and the individual role of ROS1, DML2, and DML3 demethylases in these spatiotemporal regulation patterns was determined. Large differences in defense gene expression were observed between pathogen-infected and uninfected tissues and between root and shoot tissues in both WT and rdd plants, however, only subtle changes in promoter TE methylation patterns occurred. Therefore, while TE hypermethylation caused decreased gene expression in rdd plants it did not dramatically effect spatiotemporal gene regulation, suggesting that this latter regulation is largely methylation independent. Analysis of ros1-3, dml2-1, and dml3-1 single gene mutant lines showed that promoter TE hypermethylation and defense-related gene repression was predominantly, but not exclusively, due to loss of ROS1 activity. These data demonstrate that DNA demethylation of TE sequences, largely by ROS1, promotes defense-related gene expression but does not control spatiotemporal expression in Arabidopsis. Summary: Ros1-mediated DNA demethylation of promoter transposable elements is essential for activation of defense-related gene expression in response to fungal infection in Arabidopsis thaliana. PMID:28894455

Engineering zucchini yellow mosaic potyvirus as a non-pathogenic vector for expression of heterologous proteins in cucurbits.

PubMed

Arazi, T; Slutsky, S G; Shiboleth, Y M; Wang, Y; Rubinstein, M; Barak, S; Yang, J; Gal-On, A

2001-04-27

Plant virus vectors provide an attractive biotechnological tool for the transient expression of foreign genes in whole plants. As yet there has been no use of recombinant viruses for the improvement of commercial crops. This is mainly because the viruses used to create vectors usually cause significant yield loss and can be transmitted in the field. A novel attenuated zucchini yellow mosaic potyvirus (AG) was used for the development of an environmentally safe non-pathogenic virus vector. The suitability of AG as an expression vector in plants was tested by analysis of two infectious viral constructs, each containing a distinct gene insertion site. Introduction of a foreign viral coat protein gene into AG genome between the P1 and HC-Pro genes, resulted in no expression in planta. In contrast, the same gene was stably expressed when inserted between NIb and CP genes, suggesting that this site is more suitable for a gene vector. Virus-mediated expression of reporter genes was observed in squash and cucumber leaves, stems, roots and edible fruit. Furthermore, AG stably expressed human interferon-alpha 2, an important human anti-viral drug, without affecting plant development and yield. Interferon biological activity was measured in cucumber and squash fruit. Together, these data corroborate a biotechnological utility of AG as a non-pathogenic vector for the expression of a foreign gene, as a benefit trait, in cucurbits and their edible fruit.

[Differentially expressed genes of cell signal transduction associated with benzene poisoning by cDNA microarray].

PubMed

Wang, Hong; Bi, Yongyi; Tao, Ning; Wang, Chunhong

2005-08-01

To detect the differential expression of cell signal transduction genes associated with benzene poisoning, and to explore the pathogenic mechanisms of blood system damage induced by benzene. Peripheral white blood cell gene expression profile of 7 benzene poisoning patients, including one aplastic anemia, was determined by cDNA microarray. Seven chips from normal workers were served as controls. Cluster analysis of gene expression profile was performed. Among the 4265 target genes, 176 genes associated with cell signal transduction were differentially expressed. 35 up-regulated genes including PTPRC, STAT4, IFITM1 etc were found in at least 6 pieces of microarray; 45 down-regulated genes including ARHB, PPP3CB, CDC37 etc were found in at least 5 pieces of microarray. cDNA microarray technology is an effective technique for screening the differentially expressed genes of cell signal transduction. Disorder in cell signal transduction may play certain role in the pathogenic mechanism of benzene poisoning.

Oral pathogens change proliferation properties of oral tumor cells by affecting gene expression of human defensins.

PubMed

Hoppe, T; Kraus, D; Novak, N; Probstmeier, R; Frentzen, M; Wenghoefer, M; Jepsen, S; Winter, J

2016-10-01

The impact of oral pathogens onto the generation and variability of oral tumors has only recently been investigated. To get further insights, oral cancer cells were treated with pathogens and additionally, as a result of this bacterial cellular infection, with human defensins, which are as anti-microbial peptide members of the innate immune system. After cell stimulation, proliferation behavior, expression analysis of oncogenic relevant defensin genes, and effects on EGFR signaling were investigated. The expression of oncogenic relevant anti-microbial peptides was analyzed with real-time PCR and immunohistochemistry. Cell culture experiments were performed to examine cellular impacts caused by stimulation, i.e., altered gene expression, proliferation rate, and EGF receptor-dependent signaling. Incubation of oral tumor cells with an oral pathogen (Porphyromonas gingivalis) and human α-defensins led to an increase in cell proliferation. In contrast, another oral bacterium used, Aggregatibacter actinomycetemcomitans, enhanced cell death. The bacteria and anti-microbial peptides exhibited diverse effects on the transcript levels of oncogenic relevant defensin genes and epidermal growth factor receptor signaling. These two oral pathogens exhibited opposite primary effects on the proliferation behavior of oral tumor cells. Nevertheless, both microbe species led to similar secondary impacts on the proliferation rate by modifying expression levels of oncogenic relevant α-defensin genes. In this respect, oral pathogens exerted multiplying effects on tumor cell proliferation. Additionally, human defensins were shown to differently influence epidermal growth factor receptor signaling, supporting the hypothesis that these anti-microbial peptides serve as ligands of EGFR, thus modifying the proliferation behavior of oral tumor cells.

Global differential gene expression in response to growth temperature alteration in group A Streptococcus.

PubMed

Smoot, L M; Smoot, J C; Graham, M R; Somerville, G A; Sturdevant, D E; Migliaccio, C A; Sylva, G L; Musser, J M

2001-08-28

Pathogens are exposed to different temperatures during an infection cycle and must regulate gene expression accordingly. However, the extent to which virulent bacteria alter gene expression in response to temperatures encountered in the host is unknown. Group A Streptococcus (GAS) is a human-specific pathogen that is responsible for illnesses ranging from superficial skin infections and pharyngitis to severe invasive infections such as necrotizing fasciitis and streptococcal toxic shock syndrome. GAS survives and multiplies at different temperatures during human infection. DNA microarray analysis was used to investigate the influence of temperature on global gene expression in a serotype M1 strain grown to exponential phase at 29 degrees C and 37 degrees C. Approximately 9% of genes were differentially expressed by at least 1.5-fold at 29 degrees C relative to 37 degrees C, including genes encoding transporter proteins, proteins involved in iron homeostasis, transcriptional regulators, phage-associated proteins, and proteins with no known homologue. Relatively few known virulence genes were differentially expressed at this threshold. However, transcription of 28 genes encoding proteins with predicted secretion signal sequences was altered, indicating that growth temperature substantially influences the extracellular proteome. TaqMan real-time reverse transcription-PCR assays confirmed the microarray data. We also discovered that transcription of genes encoding hemolysins, and proteins with inferred roles in iron regulation, transport, and homeostasis, was influenced by growth at 40 degrees C. Thus, GAS profoundly alters gene expression in response to temperature. The data delineate the spectrum of temperature-regulated gene expression in an important human pathogen and provide many unforeseen lines of pathogenesis investigation.

Resistance to Fusarium verticillioides and fumonisin accumulation in maize inbred lines involves an earlier and enhanced expression of lipoxygenase (LOX) genes.

PubMed

Maschietto, Valentina; Marocco, Adriano; Malachova, Alexandra; Lanubile, Alessandra

2015-09-01

Fusarium verticillioides causes ear rot in maize and contaminates the kernels with the fumonisin mycotoxins. It is known that plant lipoxygenase (LOX)-derived oxylipins regulate defence against pathogens and that the host-pathogen lipid cross-talk influences the pathogenesis. The expression profiles of fifteen genes of the LOX pathway were studied in kernels of resistant and susceptible maize lines, grown in field condition, at 3, 7 and 14 days post inoculation (dpi) with F. verticillioides. Plant defence responses were correlated with the pathogen growth, the expression profiles of fungal FUM genes for fumonisin biosynthesis and fumonisin content in the kernels. The resistant genotype limited fungal growth and fumonisin accumulation between 7 and 14 dpi. Pathogen growth became exponential in the susceptible line after 7 dpi, in correspondence with massive transcription of FUM genes and fumonisins augmented exponentially at 14 dpi. LOX pathway genes resulted strongly induced after pathogen inoculation in the resistant line at 3 and 7 dpi, whilst in the susceptible line the induction was reduced or delayed at 14 dpi. In addition, all genes resulted overexpressed before infection in kernels of the resistant genotype already at 3 dpi. The results suggest that resistance in maize may depend on an earlier activation of LOX genes and genes for jasmonic acid biosynthesis. Copyright © 2015 Elsevier GmbH. All rights reserved.

Characterization of two porcine macrophage cell lines for the expression of pathogen-recognition receptors, defensins, cytokines, chemokines, and surface sialic acid

USDA-ARS?s Scientific Manuscript database

Macrophages express various pathogen-recognition receptors (PRRs) which recognize pathogen-associated molecular patterns (PAMPs) and activate genes responsible for host defense. The aim of this study was to characterize two porcine macrophage cell lines (Cdelta+ and Cdelta–) for the expression of P...

Gene expression in the tanoak-Phytophthora ramorum interaction

Treesearch

Katherine J. Hayden; Matteo Garbelotto; Hardeep Fai; Brian Knaus; Richard Cronn; Jessica W. Wright

2012-01-01

Disease processes are dynamic, involving a suite of gene expression changes in both the host and the pathogen, all within a single tissue. As such, they lend themselves well to transcriptomic analysis. Here we focus on a generalist invasive pathogen (Phytophthora ramorum) and its most susceptible California Floristic Province native host, tanoak (...

Identification and Expression Profiles of Six Transcripts Encoding Carboxylesterase Protein in Vitis flexuosa Infected with Pathogens.

PubMed

Islam, Md Zaherul; Yun, Hae Keun

2016-08-01

Plants protect themselves from pathogen attacks via several mechanisms, including hypersensitive cell death. Recognition of pathogen attack by the plant resistance gene triggers expression of carboxylesterase genes associated with hypersensitive response. We identified six transcripts of carboxylesterase genes, Vitis flexuosa carboxylesterase 5585 (VfCXE5585), VfCXE12827, VfCXE13132, VfCXE17159, VfCXE18231, and VfCXE47674, which showed different expression patterns upon transcriptome analysis of V. flexuosa inoculated with Elsinoe ampelina. The lengths of genes ranged from 1,098 to 1,629 bp, and their encoded proteins consisted of 309 to 335 amino acids. The predicted amino acid sequences showed hydrolase like domains in all six transcripts and contained two conserved motifs, GXSXG of serine hydrolase characteristics and HGGGF related to the carboxylesterase family. The deduced amino acid sequence also contained a potential catalytic triad consisted of serine, aspartic acid and histidine. Of the six transcripts, VfCXE12827 showed upregulated expression against E. ampelina at all time points. Three genes (VfCXE5585, VfCXE12827, and VfCXE13132) showed upregulation, while others (VfCXE17159, VfCXE18231, and VfCXE47674) were down regulated in grapevines infected with Botrytis cinerea. All transcripts showed upregulated expression against Rhizobium vitis at early and later time points except VfCXE12827, and were downregulated for up to 48 hours post inoculation (hpi) after upregulation at 1 hpi in response to R. vitis infection. All tested genes showed high and differential expression in response to pathogens, indicating that they all may play a role in defense pathways during pathogen infection in grapevines.

Identification and Expression Profiles of Six Transcripts Encoding Carboxylesterase Protein in Vitis flexuosa Infected with Pathogens

PubMed Central

Islam, Md. Zaherul; Yun, Hae Keun

2016-01-01

Plants protect themselves from pathogen attacks via several mechanisms, including hypersensitive cell death. Recognition of pathogen attack by the plant resistance gene triggers expression of carboxylesterase genes associated with hypersensitive response. We identified six transcripts of carboxylesterase genes, Vitis flexuosa carboxylesterase 5585 (VfCXE5585), VfCXE12827, VfCXE13132, VfCXE17159, VfCXE18231, and VfCXE47674, which showed different expression patterns upon transcriptome analysis of V. flexuosa inoculated with Elsinoe ampelina. The lengths of genes ranged from 1,098 to 1,629 bp, and their encoded proteins consisted of 309 to 335 amino acids. The predicted amino acid sequences showed hydrolase like domains in all six transcripts and contained two conserved motifs, GXSXG of serine hydrolase characteristics and HGGGF related to the carboxylesterase family. The deduced amino acid sequence also contained a potential catalytic triad consisted of serine, aspartic acid and histidine. Of the six transcripts, VfCXE12827 showed upregulated expression against E. ampelina at all time points. Three genes (VfCXE5585, VfCXE12827, and VfCXE13132) showed upregulation, while others (VfCXE17159, VfCXE18231, and VfCXE47674) were down regulated in grapevines infected with Botrytis cinerea. All transcripts showed upregulated expression against Rhizobium vitis at early and later time points except VfCXE12827, and were downregulated for up to 48 hours post inoculation (hpi) after upregulation at 1 hpi in response to R. vitis infection. All tested genes showed high and differential expression in response to pathogens, indicating that they all may play a role in defense pathways during pathogen infection in grapevines. PMID:27493610

Differential expression of CPKs and cytosolic Ca2+ variation in resistant and susceptible apple cultivars (Malus x domestica) in response to the pathogen Erwinia amylovora and mechanical wounding.

PubMed

Kanchiswamy, Chidananda Nagamangala; Mohanta, Tapan Kumar; Capuzzo, Andrea; Occhipinti, Andrea; Verrillo, Francesca; Maffei, Massimo E; Malnoy, Mickael

2013-11-05

Plant calcium (Ca2+) signals are involved in a wide array of intracellular signalling pathways following pathogen invasion. Ca2+-binding sensory proteins such as Ca2+-dependent protein kinases (CPKs) have been predicted to mediate signalling following Ca2+ influx after pathogen infection. However, to date this prediction has remained elusive. We conducted a genome-wide identification of the Malus x domestica CPK (MdCPK) gene family and identified 30 CPK genes. Comparative phylogenetic analysis of Malus CPKs with CPKs of Arabidopsis thaliana (AtCPKs), Oryza sativa (OsCPKs), Populous trichocarpa (PtCPKs) and Zea mays (ZmCPKs) revealed four different groups. From the phylogenetic tree, we found that MdCPKs are closely related to AtCPKs and PtCPKs rather than OsCPKs and ZmCPKs, indicating their dicot-specific origin. Furthermore, comparative quantitative real time PCR and intracellular cytosolic calcium ([Ca2+]cyt) analysis were carried out on fire blight resistant and susceptible M. x domestica apple cultivars following infection with a pathogen (Erwinia amylovora) and/or mechanical damage. Calcium analysis showed an increased [Ca2+]cyt over time in resistant cultivars as compared to susceptible cultivars. Gene expression studies showed that 11 out of the 30 MdCPKs were differentially expressed following pathogen infection. We studied the genome-wide analysis of MdCPK gene family in Malus x domestica and analyzed their differential gene expression along with cytosolic calcium variation upon pathogen infection. There was a striking difference in MdCPKs gene expressions and [Ca2+]cyt variations between resistant and susceptible M. x domestica cultivars in response to E. amylovora and mechanical wounding. Our genomic and bioinformatic analysis provided an important insight about the role of MdCPKs in modulating defence responses in susceptible and resistant apple cultivars. It also provided further information on early signalling and downstream signalling cascades in response to pathogenic and mechanical stress.

Concurrent Drought Stress and Vascular Pathogen Infection Induce Common and Distinct Transcriptomic Responses in Chickpea

PubMed Central

Sinha, Ranjita; Gupta, Aarti; Senthil-Kumar, Muthappa

2017-01-01

Chickpea (Cicer arietinum); the second largest legume grown worldwide is prone to drought and various pathogen infections. These drought and pathogen stresses often occur concurrently in the field conditions. However, the molecular events in response to that are largely unknown. The present study examines the transcriptome dynamics in chickpea plants exposed to a combination of water-deficit stress and Ralstonia solanacearum infection. R. solanacearum is a potential wilt disease causing pathogen in chickpea. Drought stressed chickpea plants were infected with this pathogen and the plants were allowed to experience progressive drought with 2 and 4 days of R. solanacearum infection called short duration stress (SD stresses) and long duration stress (LD stresses), respectively. Our study showed that R. solanacearum multiplication decreased under SD-combined stress compared to SD-pathogen but there was no significant change in LD-combined stress compared to LD-pathogen. The microarray analysis during these conditions showed that 821 and 1039 differentially expressed genes (DEGs) were unique to SD- and LD-combined stresses, respectively, when compared with individual stress conditions. Three and fifteen genes were common among all the SD-stress treatments and LD-stress treatments, respectively. Genes involved in secondary cell wall biosynthesis, alkaloid biosynthesis, defense related proteins, and osmo-protectants were up-regulated during combined stress. The expression of genes involved in lignin and cellulose biosynthesis were specifically up-regulated in SD-combined, LD-combined, and LD-pathogen stress. A close transcriptomic association of LD-pathogen stress with SD-combined stress was observed in this study which indicates that R. solanacearum infection also exerts drought stress along with pathogen stress thus mimics combined stress effect. Furthermore the expression profiling of candidate genes using real-time quantitative PCR validated the microarray data. The study showed that down-regulation of defense-related genes during LD-combined stress resulted in an increased bacterial multiplication as compared to SD-combined stress. Overall, our study highlights a sub-set of DEGs uniquely expressed in response to combined stress, which serve as potential candidates for further functional characterization to delineate the molecular response of the plant to concurrent drought-pathogen stress. PMID:28382041

Gene expression profile of the plant pathogen Fusarium graminearum under the antagonistic effect of Pantoea agglomerans.

PubMed

Pandolfi, V; Jorge, E C; Melo, C M R; Albuquerque, A C S; Carrer, H

2010-07-06

The pathogenic fungus Fusarium graminearum is an ongoing threat to agriculture, causing losses in grain yield and quality in diverse crops. Substantial progress has been made in the identification of genes involved in the suppression of phytopathogens by antagonistic microorganisms; however, limited information regarding responses of plant pathogens to these biocontrol agents is available. Gene expression analysis was used to identify differentially expressed transcripts of the fungal plant pathogen F. graminearum under antagonistic effect of the bacterium Pantoea agglomerans. A macroarray was constructed, using 1014 transcripts from an F. graminearum cDNA library. Probes consisted of the cDNA of F. graminearum grown in the presence and in the absence of P. agglomerans. Twenty-nine genes were either up (19) or down (10) regulated during interaction with the antagonist bacterium. Genes encoding proteins associated with fungal defense and/or virulence or with nutritional and oxidative stress responses were induced. The repressed genes coded for a zinc finger protein associated with cell division, proteins containing cellular signaling domains, respiratory chain proteins, and chaperone-type proteins. These data give molecular and biochemical evidence of response of F. graminearum to an antagonist and could help develop effective biocontrol procedures for pathogenic plant fungi.

Usage of the Heterologous Expression of the Antimicrobial Gene afp From Aspergillus giganteus for Increasing Fungal Resistance in Olive.

PubMed

Narvaez, Isabel; Khayreddine, Titouh; Pliego, Clara; Cerezo, Sergio; Jiménez-Díaz, Rafael M; Trapero-Casas, José L; López-Herrera, Carlos; Arjona-Girona, Isabel; Martín, Carmen; Mercado, José A; Pliego-Alfaro, Fernando

2018-01-01

The antifungal protein (AFP) produced by Aspergillus giganteus , encoded by the afp gene, has been used to confer resistance against a broad range of fungal pathogens in several crops. In this research, transgenic olive plants expressing the afp gene under the control of the constitutive promoter CaMV35S were generated and their disease response against two root infecting fungal pathogens, Verticillium dahliae and Rosellinia necatrix , was evaluated. Embryogenic cultures derived from a mature zygotic embryo of cv. 'Picual' were used for A. tumefaciens transformation. Five independent transgenic lines were obtained, showing a variable level of afp expression in leaves and roots. None of these transgenic lines showed enhanced resistance to Verticillium wilt. However, some of the lines displayed a degree of incomplete resistance to white root rot caused by R. necatrix compared with disease reaction of non-transformed plants or transgenic plants expressing only the GUS gene. The level of resistance to this pathogen correlated with that of the afp expression in root and leaves. Our results indicate that the afp gene can be useful for enhanced partial resistance to R. necatrix in olive, but this gene does not protect against V. dahliae .

Heterologous expression of pathogen-specific genes ligA and ligB in the saprophyte Leptospira biflexa confers enhanced adhesion to cultured cells and fibronectin.

PubMed

Figueira, Cláudio Pereira; Croda, Julio; Choy, Henry A; Haake, David A; Reis, Mitermayer G; Ko, Albert I; Picardeau, Mathieu

2011-06-09

In comparison to other bacterial pathogens, our knowledge of the molecular basis of the pathogenesis of leptospirosis is extremely limited. An improved understanding of leptospiral pathogenetic mechanisms requires reliable tools for functional genetic analysis. Leptospiral immunoglobulin-like (Lig) proteins are surface proteins found in pathogenic Leptospira, but not in saprophytes. Here, we describe a system for heterologous expression of the Leptospira interrogans genes ligA and ligB in the saprophyte Leptospira biflexa serovar Patoc. The genes encoding LigA and LigB under the control of a constitutive spirochaetal promoter were inserted into the L. biflexa replicative plasmid. We were able to demonstrate expression and surface localization of LigA and LigB in L. biflexa. We found that the expression of the lig genes significantly enhanced the ability of transformed L. biflexa to adhere in vitro to extracellular matrix components and cultured cells, suggesting the involvement of Lig proteins in cell adhesion. This work reports a complete description of the system we have developed for heterologous expression of pathogen-specific proteins in the saprophytic L. biflexa. We show that expression of LigA and LigB proteins from the pathogen confers a virulence-associated phenotype on L. biflexa, namely adhesion to eukaryotic cells and fibronectin in vitro. This study indicates that L. biflexa can serve as a surrogate host to characterize the role of key virulence factors of the causative agent of leptospirosis.

Quantitative real time RT-PCR study of pathogen-induced gene expression in rock bream (Oplegnathus fasciatus): internal controls for data normalization.

PubMed

Zhang, Bao-cun; Sun, Li; Xiao, Zhi-zhong; Hu, Yong-hua

2014-06-01

Rock bream Oplegnathus fasciatus is an important economic fish species. In this study, we evaluated the appropriateness of six housekeeping genes as internal controls for quantitative real-time PCR (RT-qPCR) analysis of gene expression in rock bream before and after pathogen infection. The expression of the selected genes in eight tissues infected with Vibrio alginolyticus or megalocytivirus was determined by RT-qPCR, and the PCR data were analyzed with geNorm and NormFinder algorithms. The results showed that before pathogen infection, mediator of RNA polymerase II transcription subunit 8 and β-actin were ranked as the most stable genes across the examined tissues. After bacterial or viral infection, the stabilities of the housekeeping genes varied to significant extents in tissue-dependent manners, and no single pair of genes was identified as suitable references for all tissues for either of the pathogen stimuli. In addition, for the majority of tissues, the most stable genes during bacterial infection differed from those during viral infection. Nevertheless, optimum reference genes were identified for each tissue under different conditions. Taken together, these results indicate that tissue type and the nature of the infectious agent used in the study can all influence the choice of normalization factors, and that the optimum reference genes identified in this study will provide a useful guidance for the selection of internal controls in future RT-PCR study of gene expression in rock bream. Copyright © 2014 Elsevier B.V. All rights reserved.

Fire blight disease reactome: RNA-seq transcriptional profile of apple host plant defense responses to Erwinia amylovora pathogen infection.

PubMed

Kamber, Tim; Buchmann, Jan P; Pothier, Joël F; Smits, Theo H M; Wicker, Thomas; Duffy, Brion

2016-02-17

The molecular basis of resistance and susceptibility of host plants to fire blight, a major disease threat to pome fruit production globally, is largely unknown. RNA-sequencing data from challenged and mock-inoculated flowers were analyzed to assess the susceptible response of apple to the fire blight pathogen Erwinia amylovora. In presence of the pathogen 1,080 transcripts were differentially expressed at 48 h post inoculation. These included putative disease resistance, stress, pathogen related, general metabolic, and phytohormone related genes. Reads, mapped to regions on the apple genome where no genes were assigned, were used to identify potential novel genes and open reading frames. To identify transcripts specifically expressed in response to E. amylovora, RT-PCRs were conducted and compared to the expression patterns of the fire blight biocontrol agent Pantoea vagans strain C9-1, another apple pathogen Pseudomonas syringae pv. papulans, and mock inoculated apple flowers. This led to the identification of a peroxidase superfamily gene that was lower expressed in response to E. amylovora suggesting a potential role in the susceptibility response. Overall, this study provides the first transcriptional profile by RNA-seq of the host plant during fire blight disease and insights into the response of susceptible apple plants to E. amylovora.

Fire blight disease reactome: RNA-seq transcriptional profile of apple host plant defense responses to Erwinia amylovora pathogen infection

PubMed Central

Kamber, Tim; Buchmann, Jan P.; Pothier, Joël F.; Smits, Theo H. M.; Wicker, Thomas; Duffy, Brion

2016-01-01

The molecular basis of resistance and susceptibility of host plants to fire blight, a major disease threat to pome fruit production globally, is largely unknown. RNA-sequencing data from challenged and mock-inoculated flowers were analyzed to assess the susceptible response of apple to the fire blight pathogen Erwinia amylovora. In presence of the pathogen 1,080 transcripts were differentially expressed at 48 h post inoculation. These included putative disease resistance, stress, pathogen related, general metabolic, and phytohormone related genes. Reads, mapped to regions on the apple genome where no genes were assigned, were used to identify potential novel genes and open reading frames. To identify transcripts specifically expressed in response to E. amylovora, RT-PCRs were conducted and compared to the expression patterns of the fire blight biocontrol agent Pantoea vagans strain C9-1, another apple pathogen Pseudomonas syringae pv. papulans, and mock inoculated apple flowers. This led to the identification of a peroxidase superfamily gene that was lower expressed in response to E. amylovora suggesting a potential role in the susceptibility response. Overall, this study provides the first transcriptional profile by RNA-seq of the host plant during fire blight disease and insights into the response of susceptible apple plants to E. amylovora. PMID:26883568

Theobroma cacao L. pathogenesis-related gene tandem array members show diverse expression dynamics in response to pathogen colonization.

PubMed

Fister, Andrew S; Mejia, Luis C; Zhang, Yufan; Herre, Edward Allen; Maximova, Siela N; Guiltinan, Mark J

2016-05-17

The pathogenesis-related (PR) group of proteins are operationally defined as polypeptides that increase in concentration in plant tissues upon contact with a pathogen. To date, 17 classes of highly divergent proteins have been described that act through multiple mechanisms of pathogen resistance. Characterizing these families in cacao, an economically important tree crop, and comparing the families to those in other species, is an important step in understanding cacao's immune response. Using publically available resources, all members of the 17 recognized pathogenesis-related gene families in the genome of Theobroma cacao were identified and annotated resulting in a set of ~350 members in both published cacao genomes. Approximately 50 % of these genes are organized in tandem arrays scattered throughout the genome. This feature was observed in five additional plant taxa (three dicots and two monocots), suggesting that tandem duplication has played an important role in the evolution of the PR genes in higher plants. Expression profiling captured the dynamics and complexity of PR genes expression at basal levels and after induction by two cacao pathogens (the oomycete, Phytophthora palmivora, and the fungus, Colletotrichum theobromicola), identifying specific genes within families that are more responsive to pathogen challenge. Subsequent qRT-PCR validated the induction of several PR-1, PR-3, PR-4, and PR-10 family members, with greater than 1000 fold induction detected for specific genes. We describe candidate genes that are likely to be involved in cacao's defense against Phytophthora and Colletotrichum infection and could be potentially useful for marker-assisted selection for breeding of disease resistant cacao varieties. The data presented here, along with existing cacao-omics resources, will enable targeted functional genetic screening of defense genes likely to play critical functions in cacao's defense against its pathogens.

Immunological Response to Single Pathogen Challenge with Agents of the Bovine Respiratory Disease Complex: An RNA-Sequence Analysis of the Bronchial Lymph Node Transcriptome.

PubMed

Tizioto, Polyana C; Kim, JaeWoo; Seabury, Christopher M; Schnabel, Robert D; Gershwin, Laurel J; Van Eenennaam, Alison L; Toaff-Rosenstein, Rachel; Neibergs, Holly L; Taylor, Jeremy F

2015-01-01

Susceptibility to bovine respiratory disease (BRD) is multi-factorial and is influenced by stress in conjunction with infection by both bacterial and viral pathogens. While vaccination is broadly used in an effort to prevent BRD, it is far from being fully protective and cases diagnosed from a combination of observed clinical signs without any attempt at identifying the causal pathogens are usually treated with antibiotics. Dairy and beef cattle losses from BRD are profound worldwide and genetic studies have now been initiated to elucidate host loci which underlie susceptibility with the objective of enabling molecular breeding to reduce disease prevalence. In this study, we employed RNA sequencing to examine the bronchial lymph node transcriptomes of controls and beef cattle which had individually been experimentally challenged with bovine respiratory syncytial virus, infectious bovine rhinotracheitis, bovine viral diarrhea virus, Pasteurella multocida, Mannheimia haemolytica or Mycoplasma bovis to identify the genes that are involved in the bovine immune response to infection. We found that 142 differentially expressed genes were located in previously described quantitative trait locus regions associated with risk of BRD. Mutations affecting the expression or amino acid composition of these genes may affect disease susceptibility and could be incorporated into molecular breeding programs. Genes involved in innate immunity were generally found to be differentially expressed between the control and pathogen-challenged animals suggesting that variation in these genes may lead to a heritability of susceptibility that is pathogen independent. However, we also found pathogen-specific expression profiles which suggest that host genetic variation for BRD susceptibility is pathogen dependent.

Engineered resistance and hypersusceptibility through functional metabolic studies of 100 genes in soybean to its major pathogen, the soybean cyst nematode.

PubMed

Matthews, Benjamin F; Beard, Hunter; MacDonald, Margaret H; Kabir, Sara; Youssef, Reham M; Hosseini, Parsa; Brewer, Eric

2013-05-01

During pathogen attack, the host plant induces genes to ward off the pathogen while the pathogen often produces effector proteins to increase susceptibility of the host. Gene expression studies of syncytia formed in soybean root by soybean cyst nematode (Heterodera glycines) identified many genes altered in expression in resistant and susceptible roots. However, it is difficult to assess the role and impact of these genes on resistance using gene expression patterns alone. We selected 100 soybean genes from published microarray studies and individually overexpressed them in soybean roots to determine their impact on cyst nematode development. Nine genes reduced the number of mature females by more than 50 % when overexpressed, including genes encoding ascorbate peroxidase, β-1,4-endoglucanase, short chain dehydrogenase, lipase, DREPP membrane protein, calmodulin, and three proteins of unknown function. One gene encoding a serine hydroxymethyltransferase decreased the number of mature cyst nematode females by 45 % and is located at the Rhg4 locus. Four genes increased the number of mature cyst nematode females by more than 200 %, while thirteen others increased the number of mature cyst nematode females by more than 150 %. Our data support a role for auxin and ethylene in susceptibility of soybean to cyst nematodes. These studies highlight the contrasting gene sets induced by host and nematode during infection and provide new insights into the interactions between host and pathogen at the molecular level. Overexpression of some of these genes result in a greater decrease in the number of cysts formed than recognized soybean cyst nematode resistance loci.

Transcriptomic and Proteomic Analyses of Resistant Host Responses in Arachis diogoi Challenged with Late Leaf Spot Pathogen, Phaeoisariopsis personata

PubMed Central

Kumar, Dilip; Kirti, Pulugurtha Bharadwaja

2015-01-01

Late leaf spot is a serious disease of peanut caused by the imperfect fungus, Phaeoisariopsis personata. Wild diploid species, Arachis diogoi. is reported to be highly resistant to this disease and asymptomatic. The objective of this study is to investigate the molecular responses of the wild peanut challenged with the late leaf spot pathogen using cDNA-AFLP and 2D proteomic study. A total of 233 reliable, differentially expressed genes were identified in Arachis diogoi. About one third of the TDFs exhibit no significant similarity with the known sequences in the data bases. Expressed sequence tag data showed that the characterized genes are involved in conferring resistance in the wild peanut to the pathogen challenge. Several genes for proteins involved in cell wall strengthening, hypersensitive cell death and resistance related proteins have been identified. Genes identified for other proteins appear to function in metabolism, signal transduction and defence. Nineteen TDFs based on the homology analysis of genes associated with defence, signal transduction and metabolism were further validated by quantitative real time PCR (qRT-PCR) analyses in resistant wild species in comparison with a susceptible peanut genotype in time course experiments. The proteins corresponding to six TDFs were differentially expressed at protein level also. Differentially expressed TDFs and proteins in wild peanut indicate its defence mechanism upon pathogen challenge and provide initial breakthrough of genes possibly involved in recognition events and early signalling responses to combat the pathogen through subsequent development of resistivity. This is the first attempt to elucidate the molecular basis of the response of the resistant genotype to the late leaf spot pathogen, and its defence mechanism. PMID:25646800

Repressor- and Activator-Type Ethylene Response Factors Functioning in Jasmonate Signaling and Disease Resistance Identified via a Genome-Wide Screen of Arabidopsis Transcription Factor Gene Expression[w

PubMed Central

McGrath, Ken C.; Dombrecht, Bruno; Manners, John M.; Schenk, Peer M.; Edgar, Cameron I.; Maclean, Donald J.; Scheible, Wolf-Rüdiger; Udvardi, Michael K.; Kazan, Kemal

2005-01-01

To identify transcription factors (TFs) involved in jasmonate (JA) signaling and plant defense, we screened 1,534 Arabidopsis (Arabidopsis thaliana) TFs by real-time quantitative reverse transcription-PCR for their altered transcript at 6 h following either methyl JA treatment or inoculation with the incompatible pathogen Alternaria brassicicola. We identified 134 TFs that showed a significant change in expression, including many APETALA2/ethylene response factor (AP2/ERF), MYB, WRKY, and NAC TF genes with unknown functions. Twenty TF genes were induced by both the pathogen and methyl JA and these included 10 members of the AP2/ERF TF family, primarily from the B1a and B3 subclusters. Functional analysis of the B1a TF AtERF4 revealed that AtERF4 acts as a novel negative regulator of JA-responsive defense gene expression and resistance to the necrotrophic fungal pathogen Fusarium oxysporum and antagonizes JA inhibition of root elongation. In contrast, functional analysis of the B3 TF AtERF2 showed that AtERF2 is a positive regulator of JA-responsive defense genes and resistance to F. oxysporum and enhances JA inhibition of root elongation. Our results suggest that plants coordinately express multiple repressor- and activator-type AP2/ERFs during pathogen challenge to modulate defense gene expression and disease resistance. PMID:16183832

Gene Expression Profiles of Chlamydophila pneumoniae during the Developmental Cycle and Iron Depletion–Mediated Persistence

PubMed Central

Mäurer, André P; Mehlitz, Adrian; Mollenkopf, Hans J; Meyer, Thomas F

2007-01-01

The obligate intracellular, gram-negative bacterium Chlamydophila pneumoniae (Cpn) has impact as a human pathogen. Little is known about changes in the Cpn transcriptome during its biphasic developmental cycle (the acute infection) and persistence. The latter stage has been linked to chronic diseases. To analyze Cpn CWL029 gene expression, we designed a pathogen-specific oligo microarray and optimized the extraction method for pathogen RNA. Throughout the acute infection, ratio expression profiles for each gene were generated using 48 h post infection as a reference. Based on these profiles, significantly expressed genes were separated into 12 expression clusters using self-organizing map clustering and manual sorting into the “early”, “mid”, “late”, and “tardy” cluster classes. The latter two were differentiated because the “tardy” class showed steadily increasing expression at the end of the cycle. The transcriptome of the Cpn elementary body (EB) and published EB proteomics data were compared to the cluster profile of the acute infection. We found an intriguing association between “late” genes and genes coding for EB proteins, whereas “tardy” genes were mainly associated with genes coding for EB mRNA. It has been published that iron depletion leads to Cpn persistence. We compared the gene expression profiles during iron depletion–mediated persistence with the expression clusters of the acute infection. This led to the finding that establishment of iron depletion–mediated persistence is more likely a mid-cycle arrest in development rather than a completely distinct gene expression pattern. Here, we describe the Cpn transcriptome during the acute infection, differentiating “late” genes, which correlate to EB proteins, and “tardy” genes, which lead to EB mRNA. Expression profiles during iron mediated–persistence led us to propose the hypothesis that the transcriptomic “clock” is arrested during acute mid-cycle. PMID:17590080

Gene expression analysis of whole blood RNA from pigs infected with low and high pathogenic African swine fever viruses

DOE PAGES

Jaing, Crystal; Rowland, Raymond R. R.; Allen, Jonathan E.; ...

2017-08-31

African swine fever virus (ASFV) is a macrophage-tropic virus responsible for ASF, a transboundary disease that threatens swine production world-wide. Since there are no vaccines available to control ASF after an outbreak, obtaining an understanding of the virus-host interaction is important for developing new intervention strategies. In this study, a whole transcriptomic RNA-Seq method was used to characterize differentially expressed genes in pigs infected with a low pathogenic ASFV isolate, OUR T88/3 (OURT), or the highly pathogenic Georgia 2007/1 (GRG). After infection, pigs infected with OURT showed no or few clinical signs; whereas, GRG produced clinical signs consistent with acutemore » ASF. RNA-Seq detected the expression of ASFV genes from the whole blood of the GRG, but not the OURT pigs, consistent with the pathotypes of these strains and the replication of GRG in circulating monocytes. Even though GRG and OURT possess different pathogenic properties, there was significant overlap in the most upregulated host genes. A small number of differentially expressed microRNAs were also detected in GRG and OURT pigs. These data confirm previous studies describing the response of macrophages and lymphocytes to ASFV infection, as well as reveal unique gene pathways upregulated in response to infection with GRG.« less

Gene expression analysis of whole blood RNA from pigs infected with low and high pathogenic African swine fever viruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaing, Crystal; Rowland, Raymond R. R.; Allen, Jonathan E.

African swine fever virus (ASFV) is a macrophage-tropic virus responsible for ASF, a transboundary disease that threatens swine production world-wide. Since there are no vaccines available to control ASF after an outbreak, obtaining an understanding of the virus-host interaction is important for developing new intervention strategies. In this study, a whole transcriptomic RNA-Seq method was used to characterize differentially expressed genes in pigs infected with a low pathogenic ASFV isolate, OUR T88/3 (OURT), or the highly pathogenic Georgia 2007/1 (GRG). After infection, pigs infected with OURT showed no or few clinical signs; whereas, GRG produced clinical signs consistent with acutemore » ASF. RNA-Seq detected the expression of ASFV genes from the whole blood of the GRG, but not the OURT pigs, consistent with the pathotypes of these strains and the replication of GRG in circulating monocytes. Even though GRG and OURT possess different pathogenic properties, there was significant overlap in the most upregulated host genes. A small number of differentially expressed microRNAs were also detected in GRG and OURT pigs. These data confirm previous studies describing the response of macrophages and lymphocytes to ASFV infection, as well as reveal unique gene pathways upregulated in response to infection with GRG.« less

Generation of Reactive Oxygen Species via NOXa Is Important for Development and Pathogenicity of Mycosphaerella graminicola.

PubMed

Choi, Yoon-E; Lee, Changsu; Goodwin, Stephen B

2016-03-01

The ascomycete fungus Mycosphaerella graminicola (synonym Zymoseptoria tritici) is an important pathogen of wheat causing economically significant losses. The primary nutritional mode of this fungus is thought to be hemibiotrophic. This pathogenic lifestyle is associated with an early biotrophic stage of nutrient uptake followed by a necrotrophic stage aided possibly by production of a toxin or reactive oxygen species (ROS). In many other fungi, the genes CREA and AREA are important during the biotrophic stage of infection, while the NOXa gene product is important during necrotrophic growth. To test the hypothesis that these genes are important for pathogenicity of M. graminicola, we employed an over-expression strategy for the selected target genes CREA, AREA, and NOXa, which might function as regulators of nutrient acquisition or ROS generation. Increased expressions of CREA, AREA, and NOXa in M. graminicola were confirmed via quantitative real-time PCR and strains were subsequently assayed for pathogenicity. Among them, the NOXa over-expression strain, NO2, resulted in significantly increased virulence. Moreover, instead of the usual filamentous growth, we observed a predominance of yeast-like growth of NO2 which was correlated with ROS production. Our data indicate that ROS generation via NOXa is important to pathogenicity as well as development in M. graminicola.

Defense Responses to Mycotoxin-Producing Fungi Fusarium proliferatum, F. subglutinans, and Aspergillus flavus in Kernels of Susceptible and Resistant Maize Genotypes.

PubMed

Lanubile, Alessandra; Maschietto, Valentina; De Leonardis, Silvana; Battilani, Paola; Paciolla, Costantino; Marocco, Adriano

2015-05-01

Developing kernels of resistant and susceptible maize genotypes were inoculated with Fusarium proliferatum, F. subglutinans, and Aspergillus flavus. Selected defense systems were investigated using real-time reverse transcription-polymerase chain reaction to monitor the expression of pathogenesis-related (PR) genes (PR1, PR5, PRm3, PRm6) and genes protective from oxidative stress (peroxidase, catalase, superoxide dismutase and ascorbate peroxidase) at 72 h postinoculation. The study was also extended to the analysis of the ascorbate-glutathione cycle and catalase, superoxide dismutase, and cytosolic and wall peroxidases enzymes. Furthermore, the hydrogen peroxide and malondialdehyde contents were studied to evaluate the oxidation level. Higher gene expression and enzymatic activities were observed in uninoculated kernels of resistant line, conferring a major readiness to the pathogen attack. Moreover expression values of PR genes remained higher in the resistant line after inoculation, demonstrating a potentiated response to the pathogen invasions. In contrast, reactive oxygen species-scavenging genes were strongly induced in the susceptible line only after pathogen inoculation, although their enzymatic activity was higher in the resistant line. Our data provide an important basis for further investigation of defense gene functions in developing kernels in order to improve resistance to fungal pathogens. Maize genotypes with overexpressed resistance traits could be profitably utilized in breeding programs focused on resistance to pathogens and grain safety.

Apoptosis, Toll-like, RIG-I-like and NOD-like Receptors Are Pathways Jointly Induced by Diverse Respiratory Bacterial and Viral Pathogens

PubMed Central

Martínez, Isidoro; Oliveros, Juan C.; Cuesta, Isabel; de la Barrera, Jorge; Ausina, Vicente; Casals, Cristina; de Lorenzo, Alba; García, Ernesto; García-Fojeda, Belén; Garmendia, Junkal; González-Nicolau, Mar; Lacoma, Alicia; Menéndez, Margarita; Moranta, David; Nieto, Amelia; Ortín, Juan; Pérez-González, Alicia; Prat, Cristina; Ramos-Sevillano, Elisa; Regueiro, Verónica; Rodriguez-Frandsen, Ariel; Solís, Dolores; Yuste, José; Bengoechea, José A.; Melero, José A.

2017-01-01

Lower respiratory tract infections are among the top five leading causes of human death. Fighting these infections is therefore a world health priority. Searching for induced alterations in host gene expression shared by several relevant respiratory pathogens represents an alternative to identify new targets for wide-range host-oriented therapeutics. With this aim, alveolar macrophages were independently infected with three unrelated bacterial (Streptococcus pneumoniae, Klebsiella pneumoniae, and Staphylococcus aureus) and two dissimilar viral (respiratory syncytial virus and influenza A virus) respiratory pathogens, all of them highly relevant for human health. Cells were also activated with bacterial lipopolysaccharide (LPS) as a prototypical pathogen-associated molecular pattern. Patterns of differentially expressed cellular genes shared by the indicated pathogens were searched by microarray analysis. Most of the commonly up-regulated host genes were related to the innate immune response and/or apoptosis, with Toll-like, RIG-I-like and NOD-like receptors among the top 10 signaling pathways with over-expressed genes. These results identify new potential broad-spectrum targets to fight the important human infections caused by the bacteria and viruses studied here. PMID:28298903

Following Pathogen Development and Gene Expression in a Food Ecosystem: the Case of a Staphylococcus aureus Isolate in Cheese

PubMed Central

Aigle, Marina; Fleurot, Renaud; Darrigo, Claire; Hennekinne, Jacques-Antoine; Gruss, Alexandra; Borezée-Durant, Elise; Delacroix-Buchet, Agnès

2014-01-01

Human intoxication or infection due to bacterial food contamination constitutes an economic challenge and a public health problem. Information on the in situ distribution and expression of pathogens responsible for this risk is to date lacking, largely because of technical bottlenecks in detecting signals from minority bacterial populations within a complex microbial and physicochemical ecosystem. We simulated the contamination of a real high-risk cheese with a natural food isolate of Staphylococcus aureus, an enterotoxin-producing pathogen responsible for food poisoning. To overcome the problem of a detection limit in a solid matrix, we chose to work with a fluorescent reporter (superfolder green fluorescent protein) that would allow spatiotemporal monitoring of S. aureus populations and targeted gene expression. The combination of complementary techniques revealed that S. aureus localizes preferentially on the cheese surface during ripening. Immunochemistry and confocal laser scanning microscopy enabled us to visualize, in a single image, dairy bacteria and pathogen populations, virulence gene expression, and the toxin produced. This procedure is readily applicable to other genes of interest, other bacteria, and different types of food matrices. PMID:24928871

Erwinia amylovora Expresses Fast and Simultaneously hrp/dsp Virulence Genes during Flower Infection on Apple Trees

PubMed Central

Pester, Doris; Milčevičová, Renáta; Schaffer, Johann; Wilhelm, Eva; Blümel, Sylvia

2012-01-01

Background Pathogen entry through host blossoms is the predominant infection pathway of the Gram-negative bacterium Erwinia amylovora leading to manifestation of the disease fire blight. Like in other economically important plant pathogens, E. amylovora pathogenicity depends on a type III secretion system encoded by hrp genes. However, timing and transcriptional order of hrp gene expression during flower infections are unknown. Methodology/Principal Findings Using quantitative real-time PCR analyses, we addressed the questions of how fast, strong and uniform key hrp virulence genes and the effector dspA/E are expressed when bacteria enter flowers provided with the full defense mechanism of the apple plant. In non-invasive bacterial inoculations of apple flowers still attached to the tree, E. amylovora activated expression of key type III secretion genes in a narrow time window, mounting in a single expression peak of all investigated hrp/dspA/E genes around 24–48 h post inoculation (hpi). This single expression peak coincided with a single depression in the plant PR-1 expression at 24 hpi indicating transient manipulation of the salicylic acid pathway as one target of E. amylovora type III effectors. Expression of hrp/dspA/E genes was highly correlated to expression of the regulator hrpL and relative transcript abundances followed the ratio: hrpA>hrpN>hrpL>dspA/E. Acidic conditions (pH 4) in flower infections led to reduced virulence/effector gene expression without the typical expression peak observed under natural conditions (pH 7). Conclusion/Significance The simultaneous expression of hrpL, hrpA, hrpN, and the effector dspA/E during early floral infection indicates that speed and immediate effector transmission is important for successful plant invasion. When this delicate balance is disturbed, e.g., by acidic pH during infection, virulence gene expression is reduced, thus partly explaining the efficacy of acidification in fire blight control on a molecular level. PMID:22412891

Differential gene expression for Curvularia eragrostidis pathogenic incidence in crabgrass (Digitaria sanguinalis) revealed by cDNA-AFLP analysis.

PubMed

Wang, Jianshu; Wang, Xuemin; Yuan, Bohua; Qiang, Sheng

2013-01-01

Gene expression profiles of Digitaria sanguinalis infected by Curvularia eragrostidis strain QZ-2000 at two concentrations of conidia and two dew durations were analyzed by cDNA amplified fragment length polymorphisms (cDNA-AFLP). Inoculum strength was more determinant of gene expression than dew duration. A total of 256 primer combinations were used for selective amplification and 1214 transcript-derived fragments (TDFs) were selected for their differential expression. Of these, 518 up-regulated differentially expressed TDFs were identified. Forty-six differential cDNA fragments were chosen to be cloned and 35 of them were successfully cloned and sequenced, of which 25 were homologous to genes of known function according to the GenBank database. Only 6 genes were up-regulated in Curvularia eragrostidis-inoculated D. sanguinalis, with functions involved in signal transduction, energy metabolism, cell growth and development, stress responses, abscisic acid biosynthesis and response. It appears that a few pathways may be important parts of the pathogenic strategy of C. eragrostidis strain QZ-2000 on D. sanguinalis. Our study provides the fundamentals to further study the pathogenic mechanism, screen for optimal C. eragrostidis strains as potential mycoherbicide and apply this product to control D. sanguinalis.

Expression profiling during arabidopsis/downy mildew interaction reveals a highly-expressed effector that attenuates responses to salicylic acid.

PubMed

Asai, Shuta; Rallapalli, Ghanasyam; Piquerez, Sophie J M; Caillaud, Marie-Cécile; Furzer, Oliver J; Ishaque, Naveed; Wirthmueller, Lennart; Fabro, Georgina; Shirasu, Ken; Jones, Jonathan D G

2014-10-01

Plants have evolved strong innate immunity mechanisms, but successful pathogens evade or suppress plant immunity via effectors delivered into the plant cell. Hyaloperonospora arabidopsidis (Hpa) causes downy mildew on Arabidopsis thaliana, and a genome sequence is available for isolate Emoy2. Here, we exploit the availability of genome sequences for Hpa and Arabidopsis to measure gene-expression changes in both Hpa and Arabidopsis simultaneously during infection. Using a high-throughput cDNA tag sequencing method, we reveal expression patterns of Hpa predicted effectors and Arabidopsis genes in compatible and incompatible interactions, and promoter elements associated with Hpa genes expressed during infection. By resequencing Hpa isolate Waco9, we found it evades Arabidopsis resistance gene RPP1 through deletion of the cognate recognized effector ATR1. Arabidopsis salicylic acid (SA)-responsive genes including PR1 were activated not only at early time points in the incompatible interaction but also at late time points in the compatible interaction. By histochemical analysis, we found that Hpa suppresses SA-inducible PR1 expression, specifically in the haustoriated cells into which host-translocated effectors are delivered, but not in non-haustoriated adjacent cells. Finally, we found a highly-expressed Hpa effector candidate that suppresses responsiveness to SA. As this approach can be easily applied to host-pathogen interactions for which both host and pathogen genome sequences are available, this work opens the door towards transcriptome studies in infection biology that should help unravel pathogen infection strategies and the mechanisms by which host defense responses are overcome.

Genome-wide identification, classification and expression analysis in fungal-plant interactions of cutinase gene family and functional analysis of a putative ClCUT7 in Curvularia lunata.

PubMed

Liu, Tong; Hou, Jumei; Wang, Yuying; Jin, Yazhong; Borth, Wayne; Zhao, Fengzhou; Liu, Zheng; Hu, John; Zuo, Yuhu

2016-06-01

Cutinase is described as playing various roles in fungal-plant pathogen interactions, such as eliciting host-derived signals, fungal spore attachment and carbon acquisition during saprophytic growth. However, the characteristics of the cutinase genes, their expression in compatible interactions and their roles in pathogenesis have not been reported in Curvularia lunata, an important leaf spot pathogen of maize in China. Therefore, a cutinase gene family analysis could have profound significance. In this study, we identified 13 cutinase genes (ClCUT1 to ClCUT13) in the C. lunata genome. Multiple sequence alignment showed that most fungal cutinase proteins had one highly conserved GYSQG motif and a similar DxVCxG[ST]-[LIVMF](3)-x(3)H motif. Gene structure analyses of the cutinases revealed a complex intron-exon pattern with differences in the position and number of introns and exons. Based on phylogenetic relationship analysis, C. lunata cutinases and 78 known cutinase proteins from other fungi were classified into four groups with subgroups, but the C. lunata cutinases clustered in only three of the four groups. Motif analyses showed that each group of cutinases from C. lunata had a common motif. Real-time PCR indicated that transcript levels of the cutinase genes in a compatible interaction between pathogen and host had varied expression patterns. Interestingly, the transcript levels of ClCUT7 gradually increased during early pathogenesis with the most significant up-regulation at 3 h post-inoculation. When ClCUT7 was deleted, pathogenicity of the mutant decreased on unwounded maize (Zea mays) leaves. On wounded maize leaves, however, the mutant caused symptoms similar to the wild-type strain. Moreover, the ClCUT7 mutant had an approximately 10 % reduction in growth rate when cutin was the sole carbon source. In conclusion, we identified and characterized the cutinase family genes of C. lunata, analyzed their expression patterns in a compatible host-pathogen interaction, and explored the role of ClCUT7 in pathogenicity. This work will increase our understanding of cutinase genes in other fungal-plant pathogens.

The LBP Gene and Its Association with Resistance to Aeromonas hydrophila in Tilapia

PubMed Central

Fu, Gui Hong; Liu, Feng; Xia, Jun Hong; Yue, Gen Hua

2014-01-01

Resistance to pathogens is important for the sustainability and profitability of food fish production. In immune-related genes, the lipopolysaccharide-binding protein (LBP) gene is an important mediator of the inflammatory reaction. We analyzed the cDNA and genomic structure of the LBP gene in tilapia. The full-length cDNA (1901 bp) of the gene contained a 1416 bp open reading frame, encoding 471 amino acid residues. Its genomic sequence was 5577 bp, comprising 15 exons and 14 introns. Under normal conditions, the gene was constitutively expressed in all examined tissues. The highest expression was detected in intestine and kidney. We examined the responses of the gene to challenges with two bacterial pathogens Streptcoccus agalactiae and Aeromonas hydrophila. The gene was significantly upregulated in kidney and spleen post-infection with S. agalactiae and A. hydrophila, respectively. However, the expression profiles of the gene after the challenge with the two pathogens were different. Furthermore, we identified three SNPs in the gene. There were significant associations (p < 0.05) of two of the three SNPs with the resistance to A. hydrophila, but not with the resistance to S. agalactiae or growth performance. These results suggest that the LBP gene is involved in the acute-phase immunologic response to the bacterial infections, and the responses to the two bacterial pathogens are different. The two SNPs associated with the resistance to A. hydrophila may be useful in the selection of tilapia resistant to A. hydrophila. PMID:25470022

Comparative evolutionary genomics of Corynebacterium with special reference to codon and amino acid usage diversities.

PubMed

Pal, Shilpee; Sarkar, Indrani; Roy, Ayan; Mohapatra, Pradeep K Das; Mondal, Keshab C; Sen, Arnab

2018-02-01

The present study has been aimed to the comparative analysis of high GC composition containing Corynebacterium genomes and their evolutionary study by exploring codon and amino acid usage patterns. Phylogenetic study by MLSA approach, indel analysis and BLAST matrix differentiated Corynebacterium species in pathogenic and non-pathogenic clusters. Correspondence analysis on synonymous codon usage reveals that, gene length, optimal codon frequencies and tRNA abundance affect the gene expression of Corynebacterium. Most of the optimal codons as well as translationally optimal codons are C ending i.e. RNY (R-purine, N-any nucleotide base, and Y-pyrimidine) and reveal translational selection pressure on codon bias of Corynebacterium. Amino acid usage is affected by hydrophobicity, aromaticity, protein energy cost, etc. Highly expressed genes followed the cost minimization hypothesis and are less diverged at their synonymous positions of codons. Functional analysis of core genes shows significant difference in pathogenic and non-pathogenic Corynebacterium. The study reveals close relationship between non-pathogenic and opportunistic pathogenic Corynebaterium as well as between molecular evolution and survival niches of the organism.

Usage of the Heterologous Expression of the Antimicrobial Gene afp From Aspergillus giganteus for Increasing Fungal Resistance in Olive

PubMed Central

Narvaez, Isabel; Khayreddine, Titouh; Pliego, Clara; Cerezo, Sergio; Jiménez-Díaz, Rafael M.; Trapero-Casas, José L.; López-Herrera, Carlos; Arjona-Girona, Isabel; Martín, Carmen; Mercado, José A.; Pliego-Alfaro, Fernando

2018-01-01

The antifungal protein (AFP) produced by Aspergillus giganteus, encoded by the afp gene, has been used to confer resistance against a broad range of fungal pathogens in several crops. In this research, transgenic olive plants expressing the afp gene under the control of the constitutive promoter CaMV35S were generated and their disease response against two root infecting fungal pathogens, Verticillium dahliae and Rosellinia necatrix, was evaluated. Embryogenic cultures derived from a mature zygotic embryo of cv. ‘Picual’ were used for A. tumefaciens transformation. Five independent transgenic lines were obtained, showing a variable level of afp expression in leaves and roots. None of these transgenic lines showed enhanced resistance to Verticillium wilt. However, some of the lines displayed a degree of incomplete resistance to white root rot caused by R. necatrix compared with disease reaction of non-transformed plants or transgenic plants expressing only the GUS gene. The level of resistance to this pathogen correlated with that of the afp expression in root and leaves. Our results indicate that the afp gene can be useful for enhanced partial resistance to R. necatrix in olive, but this gene does not protect against V. dahliae. PMID:29875785

Heterologous expression of pathogen-specific genes ligA and ligB in the saprophyte Leptospira biflexa confers enhanced adhesion to cultured cells and fibronectin

PubMed Central

2011-01-01

Background In comparison to other bacterial pathogens, our knowledge of the molecular basis of the pathogenesis of leptospirosis is extremely limited. An improved understanding of leptospiral pathogenetic mechanisms requires reliable tools for functional genetic analysis. Leptospiral immunoglobulin-like (Lig) proteins are surface proteins found in pathogenic Leptospira, but not in saprophytes. Here, we describe a system for heterologous expression of the Leptospira interrogans genes ligA and ligB in the saprophyte Leptospira biflexa serovar Patoc. Results The genes encoding LigA and LigB under the control of a constitutive spirochaetal promoter were inserted into the L. biflexa replicative plasmid. We were able to demonstrate expression and surface localization of LigA and LigB in L. biflexa. We found that the expression of the lig genes significantly enhanced the ability of transformed L. biflexa to adhere in vitro to extracellular matrix components and cultured cells, suggesting the involvement of Lig proteins in cell adhesion. Conclusions This work reports a complete description of the system we have developed for heterologous expression of pathogen-specific proteins in the saprophytic L. biflexa. We show that expression of LigA and LigB proteins from the pathogen confers a virulence-associated phenotype on L. biflexa, namely adhesion to eukaryotic cells and fibronectin in vitro. This study indicates that L. biflexa can serve as a surrogate host to characterize the role of key virulence factors of the causative agent of leptospirosis. PMID:21658265

Selection of reference genes for expression analysis in the entomophthoralean fungus Pandora neoaphidis.

PubMed

Chen, Chun; Xie, Tingna; Ye, Sudan; Jensen, Annette Bruun; Eilenberg, Jørgen

2016-01-01

The selection of suitable reference genes is crucial for accurate quantification of gene expression and can add to our understanding of host-pathogen interactions. To identify suitable reference genes in Pandora neoaphidis, an obligate aphid pathogenic fungus, the expression of three traditional candidate genes including 18S rRNA(18S), 28S rRNA(28S) and elongation factor 1 alpha-like protein (EF1), were measured by quantitative polymerase chain reaction at different developmental stages (conidia, conidia with germ tubes, short hyphae and elongated hyphae), and under different nutritional conditions. We calculated the expression stability of candidate reference genes using four algorithms including geNorm, NormFinder, BestKeeper and Delta Ct. The analysis results revealed that the comprehensive ranking of candidate reference genes from the most stable to the least stable was 18S (1.189), 28S (1.414) and EF1 (3). The 18S was, therefore, the most suitable reference gene for real-time RT-PCR analysis of gene expression under all conditions. These results will support further studies on gene expression in P. neoaphidis. Copyright © 2015 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Differential expression of CPKs and cytosolic Ca2+ variation in resistant and susceptible apple cultivars (Malus x domestica) in response to the pathogen Erwinia amylovora and mechanical wounding

PubMed Central

2013-01-01

Background Plant calcium (Ca2+) signals are involved in a wide array of intracellular signalling pathways following pathogen invasion. Ca2+-binding sensory proteins such as Ca2+-dependent protein kinases (CPKs) have been predicted to mediate signalling following Ca2+ influx after pathogen infection. However, to date this prediction has remained elusive. Results We conducted a genome-wide identification of the Malus x domestica CPK (MdCPK) gene family and identified 30 CPK genes. Comparative phylogenetic analysis of Malus CPKs with CPKs of Arabidopsis thaliana (AtCPKs), Oryza sativa (OsCPKs), Populous trichocarpa (PtCPKs) and Zea mays (ZmCPKs) revealed four different groups. From the phylogenetic tree, we found that MdCPKs are closely related to AtCPKs and PtCPKs rather than OsCPKs and ZmCPKs, indicating their dicot-specific origin. Furthermore, comparative quantitative real time PCR and intracellular cytosolic calcium ([Ca2+]cyt) analysis were carried out on fire blight resistant and susceptible M. x domestica apple cultivars following infection with a pathogen (Erwinia amylovora) and/or mechanical damage. Calcium analysis showed an increased [Ca2+]cyt over time in resistant cultivars as compared to susceptible cultivars. Gene expression studies showed that 11 out of the 30 MdCPKs were differentially expressed following pathogen infection. Conclusions We studied the genome-wide analysis of MdCPK gene family in Malus x domestica and analyzed their differential gene expression along with cytosolic calcium variation upon pathogen infection. There was a striking difference in MdCPKs gene expressions and [Ca2+]cyt variations between resistant and susceptible M. x domestica cultivars in response to E. amylovora and mechanical wounding. Our genomic and bioinformatic analysis provided an important insight about the role of MdCPKs in modulating defence responses in susceptible and resistant apple cultivars. It also provided further information on early signalling and downstream signalling cascades in response to pathogenic and mechanical stress. PMID:24192013

Systematic Analysis of Zn2Cys6 Transcription Factors Required for Development and Pathogenicity by High-Throughput Gene Knockout in the Rice Blast Fungus

PubMed Central

Huang, Pengyun; Lin, Fucheng

2014-01-01

Because of great challenges and workload in deleting genes on a large scale, the functions of most genes in pathogenic fungi are still unclear. In this study, we developed a high-throughput gene knockout system using a novel yeast-Escherichia-Agrobacterium shuttle vector, pKO1B, in the rice blast fungus Magnaporthe oryzae. Using this method, we deleted 104 fungal-specific Zn2Cys6 transcription factor (TF) genes in M. oryzae. We then analyzed the phenotypes of these mutants with regard to growth, asexual and infection-related development, pathogenesis, and 9 abiotic stresses. The resulting data provide new insights into how this rice pathogen of global significance regulates important traits in the infection cycle through Zn2Cys6TF genes. A large variation in biological functions of Zn2Cys6TF genes was observed under the conditions tested. Sixty-one of 104 Zn2Cys6 TF genes were found to be required for fungal development. In-depth analysis of TF genes revealed that TF genes involved in pathogenicity frequently tend to function in multiple development stages, and disclosed many highly conserved but unidentified functional TF genes of importance in the fungal kingdom. We further found that the virulence-required TF genes GPF1 and CNF2 have similar regulation mechanisms in the gene expression involved in pathogenicity. These experimental validations clearly demonstrated the value of a high-throughput gene knockout system in understanding the biological functions of genes on a genome scale in fungi, and provided a solid foundation for elucidating the gene expression network that regulates the development and pathogenicity of M. oryzae. PMID:25299517

Genome-Wide Analysis of Germline Signaling Genes Regulating Longevity and Innate Immunity in the Nematode Pristionchus pacificus

PubMed Central

Sommer, Ralf J.

2012-01-01

Removal of the reproductive system of many animals including fish, flies, nematodes, mice and humans can increase lifespan through mechanisms largely unknown. The abrogation of the germline in Caenorhabditis elegans increases longevity by 60% due to a signal emitted from the somatic gonad. Apart from increased longevity, germline-less C. elegans is also resistant to other environmental stressors such as feeding on bacterial pathogens. However, the evolutionary conservation of this pathogen resistance, its genetic basis and an understanding of genes involved in producing this extraordinary survival phenotype are currently unknown. To study these evolutionary aspects we used the necromenic nematode Pristionchus pacificus, which is a genetic model system used in comparison to C. elegans. By ablation of germline precursor cells and subsequent feeding on the pathogen Serratia marcescens we discovered that P. pacificus shows remarkable resistance to bacterial pathogens and that this response is evolutionarily conserved across the Genus Pristionchus. To gain a mechanistic understanding of the increased resistance to bacterial pathogens and longevity in germline-ablated P. pacificus we used whole genome microarrays to profile the transcriptional response comparing germline ablated versus un-ablated animals when fed S. marcescens. We show that lipid metabolism, maintenance of the proteasome, insulin signaling and nuclear pore complexes are essential for germline deficient phenotypes with more than 3,300 genes being differentially expressed. In contrast, gene expression of germline-less P. pacificus on E. coli (longevity) and S. marcescens (immunity) is very similar with only 244 genes differentially expressed indicating that longevity is due to abundant gene expression also involved in immunity. By testing existing mutants of Ppa-DAF-16/FOXO and the nuclear hormone receptor Ppa-DAF-12 we show a conserved function of both genes in resistance to bacterial pathogens and longevity. This is the first study to show that the influence of the reproductive system on extending lifespan and innate immunity is conserved in evolution. PMID:22912581

AlgU controls expression of virulence genes in Pseudomonas syringae pv. tomato DC3000

USDA-ARS?s Scientific Manuscript database

Plant pathogenic bacteria are able to integrate information about their environment and adjust gene expression to provide adaptive functions. AlgU, an ECF sigma factor encoded by Pseudomonas syringae, controls expression of genes for alginate biosynthesis and is active while the bacteria are associa...

De-novo assembly and characterization of the transcriptome of Metschnikowia fructicola reveals differences in gene expression following interaction with Penicillium digitatum and grapefruit peel

PubMed Central

2013-01-01

Background The yeast Metschnikowia fructicola is an antagonist with biological control activity against postharvest diseases of several fruits. We performed a transcriptome analysis, using RNA-Seq technology, to examine the response of M. fructicola with citrus fruit and with the postharvest pathogen, Penicillium digitatum. Results More than 26 million sequencing reads were assembled into 9,674 unigenes. Approximately 50% of the unigenes could be annotated based on homology matches in the NCBI database. Based on homology, sequences were annotated with a gene description, gene ontology (GO term), and clustered into functional groups. An analysis of differential expression when the yeast was interacting with the fruit vs. the pathogen revealed more than 250 genes with specific expression responses. In the antagonist-pathogen interaction, genes related to transmembrane, multidrug transport and to amino acid metabolism were induced. In the antagonist-fruit interaction, expression of genes involved in oxidative stress, iron homeostasis, zinc homeostasis, and lipid metabolism were induced. Patterns of gene expression in the two interactions were examined at the individual transcript level by quantitative real-time PCR analysis (RT-qPCR). Conclusion This study provides new insight into the biology of the tritrophic interactions that occur in a biocontrol system such as the use of the yeast, M. fructicola for the control of green mold on citrus caused by P. digitatum. PMID:23496978

Identification and expression analysis of WRKY transcription factor genes in canola (Brassica napus L.) in response to fungal pathogens and hormone treatments.

PubMed

Yang, Bo; Jiang, Yuanqing; Rahman, Muhammad H; Deyholos, Michael K; Kav, Nat N V

2009-06-03

Members of plant WRKY transcription factor families are widely implicated in defense responses and various other physiological processes. For canola (Brassica napus L.), no WRKY genes have been described in detail. Because of the economic importance of this crop, and its evolutionary relationship to Arabidopsis thaliana, we sought to characterize a subset of canola WRKY genes in the context of pathogen and hormone responses. In this study, we identified 46 WRKY genes from canola by mining the expressed sequence tag (EST) database and cloned cDNA sequences of 38 BnWRKYs. A phylogenetic tree was constructed using the conserved WRKY domain amino acid sequences, which demonstrated that BnWRKYs can be divided into three major groups. We further compared BnWRKYs to the 72 WRKY genes from Arabidopsis and 91 WRKY from rice, and we identified 46 presumptive orthologs of AtWRKY genes. We examined the subcellular localization of four BnWRKY proteins using green fluorescent protein (GFP) and we observed the fluorescent green signals in the nucleus only.The responses of 16 selected BnWRKY genes to two fungal pathogens, Sclerotinia sclerotiorum and Alternaria brassicae, were analyzed by quantitative real time-PCR (qRT-PCR). Transcript abundance of 13 BnWRKY genes changed significantly following pathogen challenge: transcripts of 10 WRKYs increased in abundance, two WRKY transcripts decreased after infection, and one decreased at 12 h post-infection but increased later on (72 h). We also observed that transcript abundance of 13/16 BnWRKY genes was responsive to one or more hormones, including abscisic acid (ABA), and cytokinin (6-benzylaminopurine, BAP) and the defense signaling molecules jasmonic acid (JA), salicylic acid (SA), and ethylene (ET). We compared these transcript expression patterns to those previously described for presumptive orthologs of these genes in Arabidopsis and rice, and observed both similarities and differences in expression patterns. We identified a set of 13 BnWRKY genes from among 16 BnWRKY genes assayed, that are responsive to both fungal pathogens and hormone treatments, suggesting shared signaling mechanisms for these responses. This study suggests that a large number of BnWRKY proteins are involved in the transcriptional regulation of defense-related genes in response to fungal pathogens and hormone stimuli.

Longitudinal analysis of the group A Streptococcus transcriptome in experimental pharyngitis in cynomolgus macaques.

PubMed

Virtaneva, Kimmo; Porcella, Stephen F; Graham, Morag R; Ireland, Robin M; Johnson, Claire A; Ricklefs, Stacy M; Babar, Imran; Parkins, Larye D; Romero, Romina A; Corn, G Judson; Gardner, Don J; Bailey, John R; Parnell, Michael J; Musser, James M

2005-06-21

Identification of the genetic events that contribute to host-pathogen interactions is important for understanding the natural history of infectious diseases and developing therapeutics. Transcriptome studies conducted on pathogens have been central to this goal in recent years. However, most of these investigations have focused on specific end points or disease phases, rather than analysis of the entire time course of infection. To gain a more complete understanding of how bacterial gene expression changes over time in a primate host, the transcriptome of group A Streptococcus (GAS) was analyzed during an 86-day infection protocol in 20 cynomolgus macaques with experimental pharyngitis. The study used 260 custom Affymetrix (Santa Clara, CA) chips, and data were confirmed by TaqMan analysis. Colonization, acute, and asymptomatic phases of disease were identified. Successful colonization and severe inflammation were significantly correlated with an early onset of superantigen gene expression. The differential expression of two-component regulators covR and spy0680 (M1_spy0874) was significantly associated with GAS colony-forming units, inflammation, and phases of disease. Prophage virulence gene expression and prophage induction occurred predominantly during high pathogen cell densities and acute inflammation. We discovered that temporal changes in the GAS transcriptome were integrally linked to the phase of clinical disease and host-defense response. Knowledge of the gene expression patterns characterizing each phase of pathogen-host interaction provides avenues for targeted investigation of proven and putative virulence factors and genes of unknown function and will assist vaccine research.

Expression of the Grape VaSTS19 Gene in Arabidopsis Improves Resistance to Powdery Mildew and Botrytis cinerea but Increases Susceptibility to Pseudomonas syringe pv Tomato DC3000

PubMed Central

Wang, Yaqiong; Wang, Dejun; Wang, Fan; Huang, Li; Tian, Xiaomin; van Nocker, Steve; Gao, Hua; Wang, Xiping

2017-01-01

Stilbene synthase (STS) is a key enzyme that catalyzes the biosynthesis of resveratrol compounds and plays an important role in disease resistance. The molecular pathways linking STS with pathogen responses and their regulation are not known. We isolated an STS gene, VaSTS19, from a Chinese wild grape, Vitis amurensis Rupr. cv. “Tonghua-3”, and transferred this gene to Arabidopsis. We then generated VaSTS19-expressing Arabidopsis lines and evaluated the functions of VaSTS19 in various pathogen stresses, including powdery mildew, B. cinerea and Pseudomonas syringae pv. tomato DC3000 (PstDC3000). VaSTS19 enhanced resistance to powdery mildew and B. cinerea, but increased susceptibility to PstDC3000. Aniline blue staining revealed that VaSTS19 transgenic lines accumulated more callose compared to nontransgenic control plants, and showed smaller stomatal apertures when exposed to pathogen-associated molecular patterns (flagellin fragment (flg22) or lipopolysaccharides (LPS)). Analysis of the expression of several disease-related genes suggested that VaSTS19 expression enhanced defense responses though salicylic acid (SA) and/or jasmonic acid (JA) signaling pathways. These findings provide a deeper insight into the function of STS genes in defense against pathogens, and a better understanding of the regulatory cross talk between SA and JA pathways. PMID:28926983

Expression of the Grape VaSTS19 Gene in Arabidopsis Improves Resistance to Powdery Mildew and Botrytis cinerea but Increases Susceptibility to Pseudomonas syringe pv Tomato DC3000.

PubMed

Wang, Yaqiong; Wang, Dejun; Wang, Fan; Huang, Li; Tian, Xiaomin; van Nocker, Steve; Gao, Hua; Wang, Xiping

2017-09-17

Stilbene synthase (STS) is a key enzyme that catalyzes the biosynthesis of resveratrol compounds and plays an important role in disease resistance. The molecular pathways linking STS with pathogen responses and their regulation are not known. We isolated an STS gene, VaSTS19 , from a Chinese wild grape, Vitis amurensis Rupr. cv. "Tonghua-3", and transferred this gene to Arabidopsis . We then generated VaSTS19 -expressing Arabidopsis lines and evaluated the functions of VaSTS19 in various pathogen stresses, including powdery mildew, B. cinerea and Pseudomonas syringae pv. tomato DC3000 ( Pst DC3000). VaSTS19 enhanced resistance to powdery mildew and B. cinerea , but increased susceptibility to Pst DC3000. Aniline blue staining revealed that VaSTS19 transgenic lines accumulated more callose compared to nontransgenic control plants, and showed smaller stomatal apertures when exposed to pathogen-associated molecular patterns (flagellin fragment (flg22) or lipopolysaccharides (LPS)). Analysis of the expression of several disease-related genes suggested that VaSTS19 expression enhanced defense responses though salicylic acid (SA) and/or jasmonic acid (JA) signaling pathways. These findings provide a deeper insight into the function of STS genes in defense against pathogens, and a better understanding of the regulatory cross talk between SA and JA pathways.

Signaling events in pathogen-induced macrophage foam cell formation.

PubMed

Shaik-Dasthagirisaheb, Yazdani B; Mekasha, Samrawit; He, Xianbao; Gibson, Frank C; Ingalls, Robin R

2016-08-01

Macrophage foam cell formation is a key event in atherosclerosis. Several triggers induce low-density lipoprotein (LDL) uptake by macrophages to create foam cells, including infections with Porphyromonas gingivalis and Chlamydia pneumoniae, two pathogens that have been linked to atherosclerosis. While gene regulation during foam cell formation has been examined, comparative investigations to identify shared and specific pathogen-elicited molecular events relevant to foam cell formation are not well documented. We infected mouse bone marrow-derived macrophages with P. gingivalis or C. pneumoniae in the presence of LDL to induce foam cell formation, and examined gene expression using an atherosclerosis pathway targeted plate array. We found over 30 genes were significantly induced in response to both pathogens, including PPAR family members that are broadly important in atherosclerosis and matrix remodeling genes that may play a role in plaque development and stability. Six genes mainly involved in lipid transport were significantly downregulated. The response overall was remarkably similar and few genes were regulated in a pathogen-specific manner. Despite very divergent lifestyles, P. gingivalis and C. pneumoniae activate similar gene expression profiles during foam cell formation that may ultimately serve as targets for modulating infection-elicited foam cell burden, and progression of atherosclerosis. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Isolation, characterization, and expression analyses of plant elicitor peptides (pep) genes in maize

USDA-ARS?s Scientific Manuscript database

PROPEP1, PROPEP 2, and PROPEP3 genes appear to have roles in a feedback loop that amplifies defense signaling pathways initiated by pathogens. We present evidence to support the role of peptides derived from PROPEP genes as endogenous elicitors that are generated in response to pathogens. The preval...

Molecular genetic techniques for gene manipulation in Candida albicans.

PubMed

Xu, Qiu-Rong; Yan, Lan; Lv, Quan-Zhen; Zhou, Mi; Sui, Xue; Cao, Yong-Bing; Jiang, Yuan-Ying

2014-05-15

Candida albicans is one of the most common fungal pathogen in humans due to its high frequency as an opportunistic and pathogenic fungus causing superficial as well as invasive infections in immunocompromised patients. An understanding of gene function in C. albicans is necessary to study the molecular basis of its pathogenesis, virulence and drug resistance. Several manipulation techniques have been used for investigation of gene function in C. albicans, including gene disruption, controlled gene expression, protein tagging, gene reintegration, and overexpression. In this review, the main cassettes containing selectable markers used for gene manipulation in C. albicans are summarized; the advantages and limitations of these cassettes are discussed concerning the influences on the target gene expression and the virulence of the mutant strains.

Discovery of genes implicated in whirling disease infection and resistance in rainbow trout using genome-wide expression profiling

PubMed Central

Baerwald, Melinda R; Welsh, Amy B; Hedrick, Ronald P; May, Bernie

2008-01-01

Background Whirling disease, caused by the pathogen Myxobolus cerebralis, afflicts several salmonid species. Rainbow trout are particularly susceptible and may suffer high mortality rates. The disease is persistent and spreading in hatcheries and natural waters of several countries, including the U.S.A., and the economic losses attributed to whirling disease are substantial. In this study, genome-wide expression profiling using cDNA microarrays was conducted for resistant Hofer and susceptible Trout Lodge rainbow trout strains following pathogen exposure with the primary objective of identifying specific genes implicated in whirling disease resistance. Results Several genes were significantly up-regulated in skin following pathogen exposure for both the resistant and susceptible rainbow trout strains. For both strains, response to infection appears to be linked with the interferon system. Expression profiles for three genes identified with microarrays were confirmed with qRT-PCR. Ubiquitin-like protein 1 was up-regulated over 100 fold and interferon regulating factor 1 was up-regulated over 15 fold following pathogen exposure for both strains. Expression of metallothionein B, which has known roles in inflammation and immune response, was up-regulated over 5 fold in the resistant Hofer strain but was unchanged in the susceptible Trout Lodge strain following pathogen exposure. Conclusion The present study has provided an initial view into the genetic basis underlying immune response and resistance of rainbow trout to the whirling disease parasite. The identified genes have allowed us to gain insight into the molecular mechanisms implicated in salmonid immune response and resistance to whirling disease infection. PMID:18218127

Insect and wound induced GUS gene expression from a Beta vulgaris proteinase inhibitor gene promoter

USDA-ARS?s Scientific Manuscript database

Inducible gene promoters that are specifically activated by pathogen invasion or insect pest attack are needed for effective expression of resistance genes to control plant diseases. In the present study, a promoter from a serine proteinase inhibitor gene (BvSTI) shown to be up-regulated in resist...

Host and non-host pathogens elicit different jasmonate/ethylene responses in Arabidopsis.

PubMed

Zimmerli, Laurent; Stein, Mónica; Lipka, Volker; Schulze-Lefert, Paul; Somerville, Shauna

2004-12-01

Arabidopsis does not support the growth and asexual reproduction of the barley pathogen, Blumeria graminis f. sp. hordei Bgh). A majority of germlings fail to penetrate the epidermal cell wall and papillae. To gain additional insight into this interaction, we determined whether the salicylic acid (SA) or jasmonate (JA)/ethylene (ET) defence pathways played a role in blocking barley powdery mildew infections. Only the eds1 mutant and NahG transgenics supported a modest increase in penetration success by the barley powdery mildew. We also compared the global gene expression patterns of Arabidopsis inoculated with the non-host barley powdery mildew to those inoculated with a virulent, host powdery mildew, Erysiphe cichoracearum. Genes repressed by inoculations with non-host and host powdery mildews relative to non-inoculated control plants accounted for two-thirds of the differentially expressed genes. A majority of these genes encoded components of photosynthesis and general metabolism. Consistent with this observation, Arabidopsis growth was inhibited following inoculation with Bgh, suggesting a shift in resource allocation from growth to defence. A number of defence-associated genes were induced during both interactions. These genes likely are components of basal defence responses, which do not effectively block host powdery mildew infections. In addition, genes encoding defensins, anti-microbial peptides whose expression is under the control of the JA/ET signalling pathway, were induced exclusively by non-host pathogens. Ectopic activation of JA/ET signalling protected Arabidopsis against two biotrophic host pathogens. Taken together, these data suggest that biotrophic host pathogens must either suppress or fail to elicit the JA/ET signal transduction pathway.

Activation of Pathogenesis-related Genes by the Rhizobacterium, Bacillus sp. JS, Which Induces Systemic Resistance in Tobacco Plants.

PubMed

Kim, Ji-Seong; Lee, Jeongeun; Lee, Chan-Hui; Woo, Su Young; Kang, Hoduck; Seo, Sang-Gyu; Kim, Sun-Hyung

2015-06-01

Plant growth promoting rhizobacteria (PGPR) are known to confer disease resistance to plants. Bacillus sp. JS demonstrated antifungal activities against five fungal pathogens in in vitro assays. To verify whether the volatiles of Bacillus sp. JS confer disease resistance, tobacco leaves pre-treated with the volatiles were damaged by the fungal pathogen, Rhizoctonia solani and oomycete Phytophthora nicotianae. Pre-treated tobacco leaves had smaller lesion than the control plant leaves. In pathogenesis-related (PR) gene expression analysis, volatiles of Bacillus sp. JS caused the up-regulation of PR-2 encoding β-1,3-glucanase and acidic PR-3 encoding chitinase. Expression of acidic PR-4 encoding chitinase and acidic PR-9 encoding peroxidase increased gradually after exposure of the volatiles to Bacillus sp. JS. Basic PR-14 encoding lipid transfer protein was also increased. However, PR-1 genes, as markers of salicylic acid (SA) induced resistance, were not expressed. These results suggested that the volatiles of Bacillus sp. JS confer disease resistance against fungal and oomycete pathogens through PR genes expression.

Comparative transcriptome analysis of a lowly virulent strain of Erwinia amylovora in shoots of two apple cultivars - susceptible and resistant to fire blight.

PubMed

Puławska, Joanna; Kałużna, Monika; Warabieda, Wojciech; Mikiciński, Artur

2017-11-13

Erwinia amylovora is generally considered to be a homogeneous species in terms of phenotypic and genetic features. However, strains show variation in their virulence, particularly on hosts with different susceptibility to fire blight. We applied the RNA-seq technique to elucidate transcriptome-level changes of the lowly virulent E. amylovora 650 strain during infection of shoots of susceptible (Idared) and resistant (Free Redstar) apple cultivars. The highest number of differentially expressed E. amylovora genes between the two apple genotypes was observed at 24 h after inoculation. Six days after inoculation, only a few bacterial genes were differentially expressed in the susceptible and resistant apple cultivars. The analysis of differentially expressed gene functions showed that generally, higher expression of genes related to stress response and defence against toxic compounds was observed in Free Redstar. Also in this cultivar, higher expression of flagellar genes (FlaI), which are recognized as PAMP (pathogen-associated molecular pattern) by the innate immune systems of plants, was noted. Additionally, several genes that have not yet been proven to play a role in the pathogenic abilities of E. amylovora were found to be differentially expressed in the two apple cultivars. This RNA-seq analysis generated a novel dataset describing the transcriptional response of the lowly virulent strain of E. amylovora in susceptible and resistant apple cultivar. Most genes were regulated in the same way in both apple cultivars, but there were also some cultivar-specific responses suggesting that the environment in Free Redstar is more stressful for bacteria what can be the reason of their inability to infect of this cultivar. Among genes with the highest fold change in expression between experimental combinations or with the highest transcript abundance, there are many genes without ascribed functions, which have never been tested for their role in pathogenicity. Overall, this study provides the first transcriptional profile by RNA-seq of E. amylovora during infection of a host plant and insights into the transcriptional response of this pathogen in the environments of susceptible and resistant apple plants.

Identification and validation of reference genes for qRT-PCR studies of the obligate aphid pathogenic fungus Pandora neoaphidis during different developmental stages.

PubMed

Zhang, Shutao; Chen, Chun; Xie, Tingna; Ye, Sudan

2017-01-01

The selection of stable reference genes is a critical step for the accurate quantification of gene expression. To identify and validate the reference genes in Pandora neoaphidis-an obligate aphid pathogenic fungus-the expression of 13classical candidate reference genes were evaluated by quantitative real-time reverse transcriptase polymerase chain reaction(qPCR) at four developmental stages (conidia, conidia with germ tubes, short hyphae and elongated hyphae). Four statistical algorithms, including geNorm, NormFinder, BestKeeper and Delta Ct method were used to rank putative reference genes according to their expression stability and indicate the best reference gene or combination of reference genes for accurate normalization. The analysis of comprehensive ranking revealed that ACT1and 18Swas the most stably expressed genes throughout the developmental stages. To further validate the suitability of the reference genes identified in this study, the expression of cell division control protein 25 (CDC25) and Chitinase 1(CHI1) genes were used to further confirm the validated candidate reference genes. Our study presented the first systematic study of reference gene(s) selection for P. neoaphidis study and provided guidelines to obtain more accurate qPCR results for future developmental efforts.

A Temperature-Responsive Network Links Cell Shape and Virulence Traits in a Primary Fungal Pathogen

PubMed Central

Beyhan, Sinem; Gutierrez, Matias; Voorhies, Mark; Sil, Anita

2013-01-01

Survival at host temperature is a critical trait for pathogenic microbes of humans. Thermally dimorphic fungal pathogens, including Histoplasma capsulatum, are soil fungi that undergo dramatic changes in cell shape and virulence gene expression in response to host temperature. How these organisms link changes in temperature to both morphologic development and expression of virulence traits is unknown. Here we elucidate a temperature-responsive transcriptional network in H. capsulatum, which switches from a filamentous form in the environment to a pathogenic yeast form at body temperature. The circuit is driven by three highly conserved factors, Ryp1, Ryp2, and Ryp3, that are required for yeast-phase growth at 37°C. Ryp factors belong to distinct families of proteins that control developmental transitions in fungi: Ryp1 is a member of the WOPR family of transcription factors, and Ryp2 and Ryp3 are both members of the Velvet family of proteins whose molecular function is unknown. Here we provide the first evidence that these WOPR and Velvet proteins interact, and that Velvet proteins associate with DNA to drive gene expression. Using genome-wide chromatin immunoprecipitation studies, we determine that Ryp1, Ryp2, and Ryp3 associate with a large common set of genomic loci that includes known virulence genes, indicating that the Ryp factors directly control genes required for pathogenicity in addition to their role in regulating cell morphology. We further dissect the Ryp regulatory circuit by determining that a fourth transcription factor, which we name Ryp4, is required for yeast-phase growth and gene expression, associates with DNA, and displays interdependent regulation with Ryp1, Ryp2, and Ryp3. Finally, we define cis-acting motifs that recruit the Ryp factors to their interwoven network of temperature-responsive target genes. Taken together, our results reveal a positive feedback circuit that directs a broad transcriptional switch between environmental and pathogenic states in response to temperature. PMID:23935449

Cladosporium fulvum CfHNNI1 induces hypersensitive necrosis, defence gene expression and disease resistance in both host and nonhost plants.

PubMed

Cai, Xin-Zhong; Zhou, Xin; Xu, You-Ping; Joosten, Matthieu H A J; de Wit, Pierre J G M

2007-05-01

Nonhost resistance as a durable and broad-spectrum defence strategy is of great potential for agricultural applications. We have previously isolated a cDNA showing homology with genes encoding bZIP transcription factors from tomato leaf mould pathogen Cladosporium fulvum. Upon expression, the cDNA results in necrosis in C. fulvum host tomato and nonhost tobacco plants and is thus named CfHNNI1 (for C . f ulvum host and nonhost plant necrosis inducer 1). In the present study we report the induction of necrosis in a variety of nonhost plant species belonging to three families by the transient in planta expression of CfHNNI1 using virus-based vectors. Additionally, transient expression of CfHNNI1 also induced expression of the HR marker gene LeHSR203 and greatly reduced the accumulation of recombinant Potato virus X. Stable CfHNNI1 transgenic tobacco plants were generated in which the expression of CfHNNI1 is under the control of the pathogen-inducible hsr203J promoter. When infected with the oomycetes pathogen Phytophthora parasitica var. nicotianae, these transgenic plants manifested enhanced expression of CfHNNI1 and subsequent accumulation of CfHNNI1 protein, resulting in high expression of the HSR203J and PR genes, and strong resistance to the pathogen. The CfHNNI1 transgenic plants also exhibited induced resistance to Pseudomonas syringae pv. tabaci and Tobacco mosaic virus. Furthermore, CfHNNI1 was highly expressed and the protein was translocated into plant cells during the incompatible interactions between C. fulvum and host and nonhost plants. Our results demonstrate that CfHNNI1 is a potential general elicitor of hypersensitive response and nonhost resistance.

Suppression of polygalacturonase gene expression in the phytopathogenic fungus Ophiostoma novo-ulmi by RNA interference.

PubMed

Carneiro, Joyce S; de la Bastide, Paul Y; Chabot, Meghan; Lerch, Lindsey; Hintz, William E

2010-05-01

The fungal pathogen, Ophiostomo novo-ulmi, has been responsible for the rapid decline of American elm (Ulmus americana) across North America and remains a serious threat to surviving elm populations. The production of pectinolytic polygalacturonase enzymes has been implicated as a virulence factor for many fungal pathogens, including O. novo-ulmi. Previous work has shown that the targeted disruption of the endopolygalacturonase gene locus epg1 of O. novo-ulmi reduced, but did not eliminate pectinase activity. In the present study, we evaluated the use of RNA interference (RNAi) as a method of suppressing expression of the epg1 locus in O. novo-ulmi and compared its efficiency to the gene disruption method. While there was a reduction in epg1-specific mRNA transcripts and in the amount of polygalacturonase enzyme secreted for both methods of gene regulation, neither method completely suppressed the expression of pectinase activity. There was, however, a significantly greater reduction in both transcript levels and secreted enzyme observed for some of the RNAi transformants. As the first demonstration of RNAi in O. novo-ulmi, this method of gene regulation shows promise in future studies of gene expression and pathogenicity. Copyright 2010 Elsevier Inc. All rights reserved.

Expression of the Bovine NK-Lysin Gene Family and Activity against Respiratory Pathogens.

PubMed

Chen, Junfeng; Yang, Chingyuan; Tizioto, Polyana C; Huang, Huan; Lee, Mi O K; Payne, Harold R; Lawhon, Sara D; Schroeder, Friedhelm; Taylor, Jeremy F; Womack, James E

2016-01-01

Unlike the genomes of many mammals that have a single NK-lysin gene, the cattle genome contains a family of four genes, one of which is expressed preferentially in the lung. In this study, we compared the expression of the four bovine NK-lysin genes in healthy animals to animals challenged with pathogens known to be associated with bovine respiratory disease (BRD) using transcriptome sequencing (RNA-seq). The expression of several NK-lysins, especially NK2C, was elevated in challenged relative to control animals. The effects of synthetic peptides corresponding to functional region helices 2 and 3 of each gene product were tested on both model membranes and bio-membranes. Circular dichroism spectroscopy indicated that these peptides adopted a more helical secondary structure upon binding to an anionic model membrane and liposome leakage assays suggested that these peptides disrupt membranes. Bacterial killing assays further confirmed the antimicrobial effects of these peptides on BRD-associated bacteria, including both Pasteurella multocida and Mannhemia haemolytica and an ultrastructural examination of NK-lysin-treated P. multocida cells by transmission electron microscopy revealed the lysis of target membranes. These studies demonstrate that the expanded bovine NK-lysin gene family is potentially important in host defense against pathogens involved in bovine respiratory disease.

Transcriptome and Metabolite Profiling of the Infection Cycle of Zymoseptoria tritici on Wheat Reveals a Biphasic Interaction with Plant Immunity Involving Differential Pathogen Chromosomal Contributions and a Variation on the Hemibiotrophic Lifestyle Definition1[OPEN

PubMed Central

Rudd, Jason J.; Kanyuka, Kostya; Hassani-Pak, Keywan; Derbyshire, Mark; Andongabo, Ambrose; Devonshire, Jean; Lysenko, Artem; Saqi, Mansoor; Desai, Nalini M.; Powers, Stephen J.; Hooper, Juliet; Ambroso, Linda; Bharti, Arvind; Farmer, Andrew; Hammond-Kosack, Kim E.; Dietrich, Robert A.; Courbot, Mikael

2015-01-01

The hemibiotrophic fungus Zymoseptoria tritici causes Septoria tritici blotch disease of wheat (Triticum aestivum). Pathogen reproduction on wheat occurs without cell penetration, suggesting that dynamic and intimate intercellular communication occurs between fungus and plant throughout the disease cycle. We used deep RNA sequencing and metabolomics to investigate the physiology of plant and pathogen throughout an asexual reproductive cycle of Z. tritici on wheat leaves. Over 3,000 pathogen genes, more than 7,000 wheat genes, and more than 300 metabolites were differentially regulated. Intriguingly, individual fungal chromosomes contributed unequally to the overall gene expression changes. Early transcriptional down-regulation of putative host defense genes was detected in inoculated leaves. There was little evidence for fungal nutrient acquisition from the plant throughout symptomless colonization by Z. tritici, which may instead be utilizing lipid and fatty acid stores for growth. However, the fungus then subsequently manipulated specific plant carbohydrates, including fructan metabolites, during the switch to necrotrophic growth and reproduction. This switch coincided with increased expression of jasmonic acid biosynthesis genes and large-scale activation of other plant defense responses. Fungal genes encoding putative secondary metabolite clusters and secreted effector proteins were identified with distinct infection phase-specific expression patterns, although functional analysis suggested that many have overlapping/redundant functions in virulence. The pathogenic lifestyle of Z. tritici on wheat revealed through this study, involving initial defense suppression by a slow-growing extracellular and nutritionally limited pathogen followed by defense (hyper) activation during reproduction, reveals a subtle modification of the conceptual definition of hemibiotrophic plant infection. PMID:25596183

Physiological and molecular responses of Lactuca sativa to colonization by Salmonella enterica serovar Dublin.

PubMed

Klerks, M M; van Gent-Pelzer, M; Franz, E; Zijlstra, C; van Bruggen, A H C

2007-08-01

This paper describes the physiological and molecular interactions between the human-pathogenic organism Salmonella enterica serovar Dublin and the commercially available mini Roman lettuce cv. Tamburo. The association of S. enterica serovar Dublin with lettuce plants was first determined, which indicated the presence of significant populations outside and inside the plants. The latter was evidenced from significant residual concentrations after highly efficient surface disinfection (99.81%) and fluorescence microscopy of S. enterica serovar Dublin in cross sections of lettuce at the root-shoot transition region. The plant biomass was reduced significantly compared to that of noncolonized plants upon colonization with S. enterica serovar Dublin. In addition to the physiological response, transcriptome analysis by cDNA amplified fragment length polymorphism analysis also provided clear differential gene expression profiles between noncolonized and colonized lettuce plants. From these, generally and differentially expressed genes were selected and identified by sequence analysis, followed by reverse transcription-PCR displaying the specific gene expression profiles in time. Functional grouping of the expressed genes indicated a correlation between colonization of the plants and an increase in expressed pathogenicity-related genes. This study indicates that lettuce plants respond to the presence of S. enterica serovar Dublin at physiological and molecular levels, as shown by the reduction in growth and the concurrent expression of pathogenicity-related genes. In addition, it was confirmed that Salmonella spp. can colonize the interior of lettuce plants, thus potentially imposing a human health risk when processed and consumed.

Characterizing the stress/defense transcriptome of Arabidopsis

PubMed Central

Mahalingam, Ramamurthy; Gomez-Buitrago, AnaMaria; Eckardt, Nancy; Shah, Nigam; Guevara-Garcia, Angel; Day, Philip; Raina, Ramesh; Fedoroff, Nina V

2003-01-01

Background To understand the gene networks that underlie plant stress and defense responses, it is necessary to identify and characterize the genes that respond both initially and as the physiological response to the stress or pathogen develops. We used PCR-based suppression subtractive hybridization to identify Arabidopsis genes that are differentially expressed in response to ozone, bacterial and oomycete pathogens and the signaling molecules salicylic acid (SA) and jasmonic acid. Results We identified a total of 1,058 differentially expressed genes from eight stress cDNA libraries. Digital northern analysis revealed that 55% of the stress-inducible genes are rarely transcribed in unstressed plants and 17% of them were not previously represented in Arabidopsis expressed sequence tag databases. More than two-thirds of the genes in the stress cDNA collection have not been identified in previous studies as stress/defense response genes. Several stress-responsive cis-elements showed a statistically significant over-representation in the promoters of the genes in the stress cDNA collection. These include W- and G-boxes, the SA-inducible element, the abscisic acid response element and the TGA motif. Conclusions The stress cDNA collection comprises a broad repertoire of stress-responsive genes encoding proteins that are involved in both the initial and subsequent stages of the physiological response to abiotic stress and pathogens. This set of stress-, pathogen- and hormone-modulated genes is an important resource for understanding the genetic interactions underlying stress signaling and responses and may contribute to the characterization of the stress transcriptome through the construction of standardized specialized arrays. PMID:12620105

Plants, plant pathogens, and microgravity--a deadly trio.

PubMed

Leach, J E; Ryba-White, M; Sun, Q; Wu, C J; Hilaire, E; Gartner, C; Nedukha, O; Kordyum, E; Keck, M; Leung, H; Guikema, J A

2001-06-01

Plants grown in spaceflight conditions are more susceptible to colonization by plant pathogens. The underlying causes for this enhanced susceptibility are not known. Possibly the formation of structural barriers and the activation of plant defense response components are impaired in spaceflight conditions. Either condition would result from altered gene expression of the plant. Because of the tools available, past studies focused on a few physiological responses or biochemical pathways. With recent advances in genomics research, new tools, including microarray technologies, are available to examine the global impact of growth in the spacecraft on the plant's gene expression profile. In ground-based studies, we have developed cDNA subtraction libraries of rice that are enriched for genes induced during pathogen infection and the defense response. Arrays of these genes are being used to dissect plant defense response pathways in a model system involving wild-type rice plants and lesion mimic mutants. The lesion mimic mutants are ideal experimental tools because they erratically develop defense response-like lesions in the absence of pathogens. The gene expression profiles from these ground-based studies will provide the molecular basis for understanding the biochemical and physiological impacts of spaceflight on plant growth, development and disease defense responses. This, in turn, will allow the development of strategies to manage plant disease for life in the space environment.

Plants, plant pathogens, and microgravity--a deadly trio

NASA Technical Reports Server (NTRS)

Leach, J. E.; Ryba-White, M.; Sun, Q.; Wu, C. J.; Hilaire, E.; Gartner, C.; Nedukha, O.; Kordyum, E.; Keck, M.; Leung, H.;

Differential Coexpression Analysis Reveals Extensive Rewiring of Arabidopsis Gene Coexpression in Response to Pseudomonas syringae Infection

PubMed Central

Jiang, Zhenhong; Dong, Xiaobao; Li, Zhi-Gang; He, Fei; Zhang, Ziding

2016-01-01

Plant defense responses to pathogens involve massive transcriptional reprogramming. Recently, differential coexpression analysis has been developed to study the rewiring of gene networks through microarray data, which is becoming an important complement to traditional differential expression analysis. Using time-series microarray data of Arabidopsis thaliana infected with Pseudomonas syringae, we analyzed Arabidopsis defense responses to P. syringae through differential coexpression analysis. Overall, we found that differential coexpression was a common phenomenon of plant immunity. Genes that were frequently involved in differential coexpression tend to be related to plant immune responses. Importantly, many of those genes have similar average expression levels between normal plant growth and pathogen infection but have different coexpression partners. By integrating the Arabidopsis regulatory network into our analysis, we identified several transcription factors that may be regulators of differential coexpression during plant immune responses. We also observed extensive differential coexpression between genes within the same metabolic pathways. Several metabolic pathways, such as photosynthesis light reactions, exhibited significant changes in expression correlation between normal growth and pathogen infection. Taken together, differential coexpression analysis provides a new strategy for analyzing transcriptional data related to plant defense responses and new insights into the understanding of plant-pathogen interactions. PMID:27721457

Genome‐wide gene expression dynamics of the fungal pathogen Dothistroma septosporum throughout its infection cycle of the gymnosperm host Pinus radiata

PubMed Central

Guo, Yanan; Sim, Andre D.; Kabir, M. Shahjahan; Chettri, Pranav; Ozturk, Ibrahim K.; Hunziker, Lukas; Ganley, Rebecca J.; Cox, Murray P.

2015-01-01

Summary We present genome‐wide gene expression patterns as a time series through the infection cycle of the fungal pine needle blight pathogen, Dothistroma septosporum, as it invades its gymnosperm host, Pinus radiata. We determined the molecular changes at three stages of the disease cycle: epiphytic/biotrophic (early), initial necrosis (mid) and mature sporulating lesion (late). Over 1.7 billion combined plant and fungal reads were sequenced to obtain 3.2 million fungal‐specific reads, which comprised as little as 0.1% of the sample reads early in infection. This enriched dataset shows that the initial biotrophic stage is characterized by the up‐regulation of genes encoding fungal cell wall‐modifying enzymes and signalling proteins. Later necrotrophic stages show the up‐regulation of genes for secondary metabolism, putative effectors, oxidoreductases, transporters and starch degradation. This in‐depth through‐time transcriptomic study provides our first snapshot of the gene expression dynamics that characterize infection by this fungal pathogen in its gymnosperm host. PMID:25919703

Effect of external and internal factors on the expression of reporter genes driven by the N resistance gene promoter.

PubMed

Kathiria, Palak; Sidler, Corinne; Woycicki, Rafal; Yao, Youli; Kovalchuk, Igor

2013-07-01

The role of resistance (R) genes in plant pathogen interaction has been studied extensively due to its economical impact on agriculture. Interaction between tobacco mosaic virus (TMV) and the N protein from tobacco is one of the most widely used models to understand various aspects of pathogen resistance. The transcription activity governed by N gene promoter is one of the least understood elements of the model. In this study, the N gene promoter was cloned and fused with two different reporter genes, one encoding β-glucuronidase (N::GUS) and another, luciferase (N::LUC). Tobacco plants transformed with the N::GUS or N::LUC reporter constructs were screened for homozygosity and stable expression. Histochemical analysis of N::GUS tobacco plants revealed that the expression is organ specific and developmentally regulated. Whereas two week old plants expressed GUS in midveins only, 6-wk-old plants also expressed GUS in leaf lamella. Roots did not show GUS expression at any time during development. Experiments to address effects of external stress were performed using N::LUC tobacco plants. These experiments showed that N gene promoter expression was suppressed when plants were exposed to high but not low temperatures. Expression was also upregulated in response to TMV, but no changes were observed in plants treated with SA.

Protein Poly(ADP-ribosyl)ation Regulates Arabidopsis Immune Gene Expression and Defense Responses

PubMed Central

Feng, Baomin; Liu, Chenglong; de Oliveira, Marcos V. V.; Intorne, Aline C.; Li, Bo; Babilonia, Kevin; de Souza Filho, Gonçalo A.; Shan, Libo; He, Ping

2015-01-01

Perception of microbe-associated molecular patterns (MAMPs) elicits transcriptional reprogramming in hosts and activates defense to pathogen attacks. The molecular mechanisms underlying plant pattern-triggered immunity remain elusive. A genetic screen identified Arabidopsis poly(ADP-ribose) glycohydrolase 1 (atparg1) mutant with elevated immune gene expression upon multiple MAMP and pathogen treatments. Poly(ADP-ribose) glycohydrolase (PARG) is predicted to remove poly(ADP-ribose) polymers on acceptor proteins modified by poly(ADP-ribose) polymerases (PARPs) with three PARPs and two PARGs in Arabidopsis genome. AtPARP1 and AtPARP2 possess poly(ADP-ribose) polymerase activity, and the activity of AtPARP2 was enhanced by MAMP treatment. AtPARG1, but not AtPARG2, carries glycohydrolase activity in vivo and in vitro. Importantly, mutation (G450R) in atparg1 blocks its activity and the corresponding residue is highly conserved and essential for human HsPARG activity. Consistently, mutant atparp1atparp2 plants exhibited compromised immune gene activation and enhanced susceptibility to pathogen infections. Our study indicates that protein poly(ADP-ribosyl)ation plays critical roles in plant immune gene expression and defense to pathogen attacks. PMID:25569773

R Script Approach to Infer Toxoplasma Infection Mechanisms From Microarrays and Domain-Domain Protein Interactions

PubMed Central

Arenas, Ailan F; Salcedo, Gladys E; Gomez-Marin, Jorge E

2017-01-01

Pathogen-host protein-protein interaction systems examine the interactions between the protein repertoires of 2 distinct organisms. Some of these pathogen proteins interact with the host protein system and may manipulate it for their own advantages. In this work, we designed an R script by concatenating 2 functions called rowDM and rowCVmed to infer pathogen-host interaction using previously reported microarray data, including host gene enrichment analysis and the crossing of interspecific domain-domain interactions. We applied this script to the Toxoplasma-host system to describe pathogen survival mechanisms from human, mouse, and Toxoplasma Gene Expression Omnibus series. Our outcomes exhibited similar results with previously reported microarray analyses, but we found other important proteins that could contribute to toxoplasma pathogenesis. We observed that Toxoplasma ROP38 is the most differentially expressed protein among toxoplasma strains. Enrichment analysis and KEGG mapping indicated that the human retinal genes most affected by Toxoplasma infections are those related to antiapoptotic mechanisms. We suggest that proteins PIK3R1, PRKCA, PRKCG, PRKCB, HRAS, and c-JUN could be the possible substrates for differentially expressed Toxoplasma kinase ROP38. Likewise, we propose that Toxoplasma causes overexpression of apoptotic suppression human genes. PMID:29317802

Characterization of a tryptophan 2-monooxygenase gene from Puccinia graminis f. sp. tritici involved in auxin biosynthesis and rust pathogenicity.

PubMed

Yin, Chuntao; Park, Jeong-Jin; Gang, David R; Hulbert, Scot H

2014-03-01

The plant hormone indole-3-acetic acid (IAA) is best known as a regulator of plant growth and development but its production can also affect plant-microbe interactions. Microorganisms, including numerous plant-associated bacteria and several fungi, are also capable of producing IAA. The stem rust fungus Puccinia graminis f. sp. tritici induced wheat plants to accumulate auxin in infected leaf tissue. A gene (Pgt-IaaM) encoding a putative tryptophan 2-monooxygenase, which makes the auxin precursor indole-3-acetamide (IAM), was identified in the P. graminis f. sp. tritici genome and found to be expressed in haustoria cells in infected plant tissue. Transient silencing of the gene in infected wheat plants indicated that it was required for full pathogenicity. Expression of Pgt-IaaM in Arabidopsis caused a typical auxin expression phenotype and promoted susceptibility to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000.

Identification of Quantitative Trait Loci Controlling Gene Expression during the Innate Immunity Response of Soybean1[W][OA

PubMed Central

Valdés-López, Oswaldo; Thibivilliers, Sandra; Qiu, Jing; Xu, Wayne Wenzhong; Nguyen, Tran H.N.; Libault, Marc; Le, Brandon H.; Goldberg, Robert B.; Hill, Curtis B.; Hartman, Glen L.; Diers, Brian; Stacey, Gary

2011-01-01

Microbe-associated molecular pattern-triggered immunity (MTI) is an important component of the plant innate immunity response to invading pathogens. However, most of our knowledge of MTI comes from studies of model systems with relatively little work done with crop plants. In this work, we report on variation in both the microbe-associated molecular pattern-triggered oxidative burst and gene expression across four soybean (Glycine max) genotypes. Variation in MTI correlated with the level of pathogen resistance for each genotype. A quantitative trait locus analysis on these traits identified four loci that appeared to regulate gene expression during MTI in soybean. Likewise, we observed that both MTI variation and pathogen resistance were quantitatively inherited. The approach utilized in this study may have utility for identifying key resistance loci useful for developing improved soybean cultivars. PMID:21963820

mRNA-Seq Analysis of the Pseudoperonospora cubensis Transcriptome During Cucumber (Cucumis sativus L.) Infection

PubMed Central

Hamilton, John P.; Vaillancourt, Brieanne; Buell, C. Robin; Day, Brad

2012-01-01

Pseudoperonospora cubensis, an oomycete, is the causal agent of cucurbit downy mildew, and is responsible for significant losses on cucurbit crops worldwide. While other oomycete plant pathogens have been extensively studied at the molecular level, Ps. cubensis and the molecular basis of its interaction with cucurbit hosts has not been well examined. Here, we present the first large-scale global gene expression analysis of Ps. cubensis infection of a susceptible Cucumis sativus cultivar, ‘Vlaspik’, and identification of genes with putative roles in infection, growth, and pathogenicity. Using high throughput whole transcriptome sequencing, we captured differential expression of 2383 Ps. cubensis genes in sporangia and at 1, 2, 3, 4, 6, and 8 days post-inoculation (dpi). Additionally, comparison of Ps. cubensis expression profiles with expression profiles from an infection time course of the oomycete pathogen Phytophthora infestans on Solanum tuberosum revealed similarities in expression patterns of 1,576–6,806 orthologous genes suggesting a substantial degree of overlap in molecular events in virulence between the biotrophic Ps. cubensis and the hemi-biotrophic P. infestans. Co-expression analyses identified distinct modules of Ps. cubensis genes that were representative of early, intermediate, and late infection stages. Collectively, these expression data have advanced our understanding of key molecular and genetic events in the virulence of Ps. cubensis and thus, provides a foundation for identifying mechanism(s) by which to engineer or effect resistance in the host. PMID:22545137

Caste-, sex-, and age-dependent expression of immune-related genes in a Japanese subterranean termite, Reticulitermes speratus

PubMed Central

Kobayashi, Kazuya; Matsuura, Kenji

2017-01-01

Insects protect themselves from microbial infections through innate immune responses, including pathogen recognition, phagocytosis, the activation of proteolytic cascades, and the synthesis of antimicrobial peptides. Termites, eusocial insects inhabiting microbe-rich wood, live in closely-related family groups that are susceptible to shared pathogen infections. To resist pathogenic infection, termite families have evolved diverse immune adaptations at both individual and societal levels, and a strategy of trade-offs between reproduction and immunity has been suggested. Although termite immune-inducible genes have been identified, few studies have investigated the differential expression of these genes between reproductive and neuter castes, and between sexes in each caste. In this study, we compared the expression levels of immune-related genes among castes, sexes, and ages in a Japanese subterranean termite, Reticulitermes speratus. Using RNA-seq, we found 197 immune-related genes, including 40 pattern recognition proteins, 97 signalling proteins, 60 effectors. Among these genes, 174 showed differential expression among castes. Comparing expression levels between males and females in each caste, we found sexually dimorphic expression of immune-related genes not only in reproductive castes, but also in neuter castes. Moreover, we identified age-related differential expression of 162 genes in male and/or female reproductives. In addition, although R. speratus is known to use the antibacterial peptide C-type lysozyme as an egg recognition pheromone, we determined that R. speratus has not only C-type, but also P-type and I-type lysozymes, as well as other termite species. Our transcriptomic analyses revealed immune response plasticity among all castes, and sex-biased expression of immune genes even in neuter castes, suggesting a sexual division of labor in the immune system of R. speratus. This study heightens the understanding of the evolution of antimicrobial strategies in eusocial insects, and of sexual roles in insect societies as a whole. PMID:28410430

Caste-, sex-, and age-dependent expression of immune-related genes in a Japanese subterranean termite, Reticulitermes speratus.

PubMed

Mitaka, Yuki; Kobayashi, Kazuya; Matsuura, Kenji

2017-01-01

Insects protect themselves from microbial infections through innate immune responses, including pathogen recognition, phagocytosis, the activation of proteolytic cascades, and the synthesis of antimicrobial peptides. Termites, eusocial insects inhabiting microbe-rich wood, live in closely-related family groups that are susceptible to shared pathogen infections. To resist pathogenic infection, termite families have evolved diverse immune adaptations at both individual and societal levels, and a strategy of trade-offs between reproduction and immunity has been suggested. Although termite immune-inducible genes have been identified, few studies have investigated the differential expression of these genes between reproductive and neuter castes, and between sexes in each caste. In this study, we compared the expression levels of immune-related genes among castes, sexes, and ages in a Japanese subterranean termite, Reticulitermes speratus. Using RNA-seq, we found 197 immune-related genes, including 40 pattern recognition proteins, 97 signalling proteins, 60 effectors. Among these genes, 174 showed differential expression among castes. Comparing expression levels between males and females in each caste, we found sexually dimorphic expression of immune-related genes not only in reproductive castes, but also in neuter castes. Moreover, we identified age-related differential expression of 162 genes in male and/or female reproductives. In addition, although R. speratus is known to use the antibacterial peptide C-type lysozyme as an egg recognition pheromone, we determined that R. speratus has not only C-type, but also P-type and I-type lysozymes, as well as other termite species. Our transcriptomic analyses revealed immune response plasticity among all castes, and sex-biased expression of immune genes even in neuter castes, suggesting a sexual division of labor in the immune system of R. speratus. This study heightens the understanding of the evolution of antimicrobial strategies in eusocial insects, and of sexual roles in insect societies as a whole.

The Transcription Factor Rbf1 Is the Master Regulator for b-Mating Type Controlled Pathogenic Development in Ustilago maydis

PubMed Central

Vranes, Miroslav; Wahl, Ramon; Pothiratana, Chetsada; Schuler, David; Vincon, Volker; Finkernagel, Florian; Flor-Parra, Ignacio; Kämper, Jörg

2010-01-01

In the phytopathogenic basidiomycete Ustilago maydis, sexual and pathogenic development are tightly connected and controlled by the heterodimeric bE/bW transcription factor complex encoded by the b-mating type locus. The formation of the active bE/bW heterodimer leads to the formation of filaments, induces a G2 cell cycle arrest, and triggers pathogenicity. Here, we identify a set of 345 bE/bW responsive genes which show altered expression during these developmental changes; several of these genes are associated with cell cycle coordination, morphogenesis and pathogenicity. 90% of the genes that show altered expression upon bE/bW-activation require the zinc finger transcription factor Rbf1, one of the few factors directly regulated by the bE/bW heterodimer. Rbf1 is a novel master regulator in a multilayered network of transcription factors that facilitates the complex regulatory traits of sexual and pathogenic development. PMID:20700446

Identification of promising host-induced silencing targets among genes preferentially transcribed in haustoria of Puccinia

USDA-ARS?s Scientific Manuscript database

Expression of dsRNA fragments of rust pathogen genes in wheat seedlings through the barley stripe mosaic virus (BSMV) based host-induced gene silencing (HIGS) system can reduce the expression of the corresponding genes in the rust fungus. The highest levels of suppression have generally been observe...

Overexpression of SOS genes in ciprofloxacin resistant Escherichia coli mutants.

PubMed

Pourahmad Jaktaji, Razieh; Pasand, Shirin

2016-01-15

Fluoroquinolones are important antibiotics for the treatment of urinary tract infections caused by Escherichia coli. Mutational studies have shown that ciprofloxacin, a member of fluoroquinolones induces SOS response and mutagenesis in pathogenic bacteria which in turn develop antibiotic resistance. However, inhibition of SOS response can increase recombination activity which in turn leads to genetic variation. The aim of this study was to measure 5 SOS genes expressions in nine E. coli mutants with different MICs for ciprofloxacin following exposure to ciprofloxacin. Gene expression was assessed by quantitative real time PCR. Gene alteration assessment was conducted by PCR amplification and DNA sequencing. Results showed that the expression of recA was increased in 5 mutants. This overexpression is not related to gene alteration, and enhances the expression of polB and umuCD genes encoding nonmutagenic and mutagenic polymerases, respectively. The direct relationship between the level of SOS expression and the level of resistance to ciprofloxacin was also indicated. It was concluded that novel therapeutic strategy that inhibits RecA activity would enhance the efficiency of common antibiotics against pathogenic bacteria. Copyright © 2015 Elsevier B.V. All rights reserved.

Response of Vitis vinifera cell cultures to Eutypa lata and Trichoderma atroviride culture filtrates: expression of defence-related genes and phenotypes.

PubMed

Mutawila, C; Stander, C; Halleen, F; Vivier, M A; Mostert, L

2017-03-01

Cell suspension cultures of Vitis vinifera cv. Dauphine berries were used to study the response to the vascular pathogen, Eutypa lata, in comparison with a biological control agent, Trichoderma atroviride, that was previously shown to be effective in pruning wound protection. The expression of genes coding for enzymes of the phenylpropanoid pathway and pathogenesis-related (PR) proteins was profiled over a 48-h period using quantitative reverse transcriptase PCR. The cell cultures responded to elicitors of both fungi with a hypersensitive-like response that lead to a decrease in cell viability. Similar genes were triggered by both the pathogen and biocontrol agent, but the timing patterns and magnitude of expression was dependent on the specific fungal elicitor. Culture filtrates of both fungi caused upregulation of phenylalanine ammonia-lyase (PAL), 4-coumaroyl Co-A ligase (CCo-A) and stilbene synthase (STS), and a downregulation of chalcone synthase (CHS) genes. The pathogen filtrate caused a biphasic pattern in the upregulation of PAL and STS genes which was not observed in cells treated with filtrates of the biocontrol agent. Analytical assays showed significantly higher total phenolic content and chitinolytic enzyme activity in the cell cultures treated with the T. atroviride filtrate compared to the pathogen filtrate. These results corresponded well to the higher expression of PAL and chitinase class IV genes. The response of the cell cultures to T. atroviride filtrate provides support for the notion that the wound protection by the biocontrol agent at least partially relies on the induction of grapevine resistance mechanisms.

Global Genome and Transcriptome Analyses of Magnaporthe oryzae Epidemic Isolate 98-06 Uncover Novel Effectors and Pathogenicity-Related Genes, Revealing Gene Gain and Lose Dynamics in Genome Evolution

PubMed Central

Dong, Yanhan; Li, Ying; Zhao, Miaomiao; Jing, Maofeng; Liu, Xinyu; Liu, Muxing; Guo, Xianxian; Zhang, Xing; Chen, Yue; Liu, Yongfeng; Liu, Yanhong; Ye, Wenwu; Zhang, Haifeng; Wang, Yuanchao; Zheng, Xiaobo; Wang, Ping; Zhang, Zhengguang

2015-01-01

Genome dynamics of pathogenic organisms are driven by pathogen and host co-evolution, in which pathogen genomes are shaped to overcome stresses imposed by hosts with various genetic backgrounds through generation of a variety of isolates. This same principle applies to the rice blast pathogen Magnaporthe oryzae and the rice host; however, genetic variations among different isolates of M. oryzae remain largely unknown, particularly at genome and transcriptome levels. Here, we applied genomic and transcriptomic analytical tools to investigate M. oryzae isolate 98-06 that is the most aggressive in infection of susceptible rice cultivars. A unique 1.4 Mb of genomic sequences was found in isolate 98-06 in comparison to reference strain 70-15. Genome-wide expression profiling revealed the presence of two critical expression patterns of M. oryzae based on 64 known pathogenicity-related (PaR) genes. In addition, 134 candidate effectors with various segregation patterns were identified. Five tested proteins could suppress BAX-mediated programmed cell death in Nicotiana benthamiana leaves. Characterization of isolate-specific effector candidates Iug6 and Iug9 and PaR candidate Iug18 revealed that they have a role in fungal propagation and pathogenicity. Moreover, Iug6 and Iug9 are located exclusively in the biotrophic interfacial complex (BIC) and their overexpression leads to suppression of defense-related gene expression in rice, suggesting that they might participate in biotrophy by inhibiting the SA and ET pathways within the host. Thus, our studies identify novel effector and PaR proteins involved in pathogenicity of the highly aggressive M. oryzae field isolate 98-06, and reveal molecular and genomic dynamics in the evolution of M. oryzae and rice host interactions. PMID:25837042

Comparative Transcriptomic Analysis of Race 1 and Race 4 of Fusarium oxysporum f. sp. cubense Induced with Different Carbon Sources

PubMed Central

Qin, Shiwen; Ji, Chunyan; Li, Yunfeng; Wang, Zhenzhong

2017-01-01

The fungal pathogen Fusarium oxysporum f. sp. cubense causes Fusarium wilt, one of the most destructive diseases in banana and plantain cultivars. Pathogenic race 1 attacks the “Gros Michel” banana cultivar, and race 4 is pathogenic to the Cavendish banana cultivar and those cultivars that are susceptible to Foc1. To understand the divergence in gene expression modules between the two races during degradation of the host cell wall, we performed RNA sequencing to compare the genome-wide transcriptional profiles of the two races grown in media containing banana cell wall, pectin, or glucose as the sole carbon source. Overall, the gene expression profiles of Foc1 and Foc4 in response to host cell wall or pectin appeared remarkably different. When grown with host cell wall, a much larger number of genes showed altered levels of expression in Foc4 in comparison with Foc1, including genes encoding carbohydrate-active enzymes (CAZymes) and other virulence-related genes. Additionally, the levels of gene expression were higher in Foc4 than in Foc1 when grown with host cell wall or pectin. Furthermore, a great majority of genes were differentially expressed in a variety-specific manner when induced by host cell wall or pectin. More specific CAZymes and other pathogenesis-related genes were expressed in Foc4 than in Foc1 when grown with host cell wall. The first transcriptome profiles obtained for Foc during degradation of the host cell wall may provide new insights into the mechanism of banana cell wall polysaccharide decomposition and the genetic basis of Foc host specificity. PMID:28468818

Comparative Transcriptomic Analysis of Race 1 and Race 4 of Fusarium oxysporum f. sp. cubense Induced with Different Carbon Sources.

PubMed

Qin, Shiwen; Ji, Chunyan; Li, Yunfeng; Wang, Zhenzhong

2017-07-05

The fungal pathogen Fusarium oxysporum f. sp. cubense causes Fusarium wilt, one of the most destructive diseases in banana and plantain cultivars. Pathogenic race 1 attacks the "Gros Michel" banana cultivar, and race 4 is pathogenic to the Cavendish banana cultivar and those cultivars that are susceptible to Foc1. To understand the divergence in gene expression modules between the two races during degradation of the host cell wall, we performed RNA sequencing to compare the genome-wide transcriptional profiles of the two races grown in media containing banana cell wall, pectin, or glucose as the sole carbon source. Overall, the gene expression profiles of Foc1 and Foc4 in response to host cell wall or pectin appeared remarkably different. When grown with host cell wall, a much larger number of genes showed altered levels of expression in Foc4 in comparison with Foc1, including genes encoding carbohydrate-active enzymes (CAZymes) and other virulence-related genes. Additionally, the levels of gene expression were higher in Foc4 than in Foc1 when grown with host cell wall or pectin. Furthermore, a great majority of genes were differentially expressed in a variety-specific manner when induced by host cell wall or pectin. More specific CAZymes and other pathogenesis-related genes were expressed in Foc4 than in Foc1 when grown with host cell wall. The first transcriptome profiles obtained for Foc during degradation of the host cell wall may provide new insights into the mechanism of banana cell wall polysaccharide decomposition and the genetic basis of Foc host specificity. Copyright © 2017 Qin et al.

The Quantitative Basis of the Arabidopsis Innate Immune System to Endemic Pathogens Depends on Pathogen Genetics

PubMed Central

Corwin, Jason A.; Copeland, Daniel; Feusier, Julie; Subedy, Anushriya; Eshbaugh, Robert; Palmer, Christine; Maloof, Julin; Kliebenstein, Daniel J.

2016-01-01

The most established model of the eukaryotic innate immune system is derived from examples of large effect monogenic quantitative resistance to pathogens. However, many host-pathogen interactions involve many genes of small to medium effect and exhibit quantitative resistance. We used the Arabidopsis-Botrytis pathosystem to explore the quantitative genetic architecture underlying host innate immune system in a population of Arabidopsis thaliana. By infecting a diverse panel of Arabidopsis accessions with four phenotypically and genotypically distinct isolates of the fungal necrotroph B. cinerea, we identified a total of 2,982 genes associated with quantitative resistance using lesion area and 3,354 genes associated with camalexin production as measures of the interaction. Most genes were associated with resistance to a specific Botrytis isolate, which demonstrates the influence of pathogen genetic variation in analyzing host quantitative resistance. While known resistance genes, such as receptor-like kinases (RLKs) and nucleotide-binding site leucine-rich repeat proteins (NLRs), were found to be enriched among associated genes, they only account for a small fraction of the total genes associated with quantitative resistance. Using publically available co-expression data, we condensed the quantitative resistance associated genes into co-expressed gene networks. GO analysis of these networks implicated several biological processes commonly connected to disease resistance, including defense hormone signaling and ROS production, as well as novel processes, such as leaf development. Validation of single gene T-DNA knockouts in a Col-0 background demonstrate a high success rate (60%) when accounting for differences in environmental and Botrytis genetic variation. This study shows that the genetic architecture underlying host innate immune system is extremely complex and is likely able to sense and respond to differential virulence among pathogen genotypes. PMID:26866607

The Quantitative Basis of the Arabidopsis Innate Immune System to Endemic Pathogens Depends on Pathogen Genetics.

PubMed

Corwin, Jason A; Copeland, Daniel; Feusier, Julie; Subedy, Anushriya; Eshbaugh, Robert; Palmer, Christine; Maloof, Julin; Kliebenstein, Daniel J

2016-02-01

The most established model of the eukaryotic innate immune system is derived from examples of large effect monogenic quantitative resistance to pathogens. However, many host-pathogen interactions involve many genes of small to medium effect and exhibit quantitative resistance. We used the Arabidopsis-Botrytis pathosystem to explore the quantitative genetic architecture underlying host innate immune system in a population of Arabidopsis thaliana. By infecting a diverse panel of Arabidopsis accessions with four phenotypically and genotypically distinct isolates of the fungal necrotroph B. cinerea, we identified a total of 2,982 genes associated with quantitative resistance using lesion area and 3,354 genes associated with camalexin production as measures of the interaction. Most genes were associated with resistance to a specific Botrytis isolate, which demonstrates the influence of pathogen genetic variation in analyzing host quantitative resistance. While known resistance genes, such as receptor-like kinases (RLKs) and nucleotide-binding site leucine-rich repeat proteins (NLRs), were found to be enriched among associated genes, they only account for a small fraction of the total genes associated with quantitative resistance. Using publically available co-expression data, we condensed the quantitative resistance associated genes into co-expressed gene networks. GO analysis of these networks implicated several biological processes commonly connected to disease resistance, including defense hormone signaling and ROS production, as well as novel processes, such as leaf development. Validation of single gene T-DNA knockouts in a Col-0 background demonstrate a high success rate (60%) when accounting for differences in environmental and Botrytis genetic variation. This study shows that the genetic architecture underlying host innate immune system is extremely complex and is likely able to sense and respond to differential virulence among pathogen genotypes.

Plant-pathogen interactions: what microarray tells about it?

PubMed

Lodha, T D; Basak, J

2012-01-01

Plant defense responses are mediated by elementary regulatory proteins that affect expression of thousands of genes. Over the last decade, microarray technology has played a key role in deciphering the underlying networks of gene regulation in plants that lead to a wide variety of defence responses. Microarray is an important tool to quantify and profile the expression of thousands of genes simultaneously, with two main aims: (1) gene discovery and (2) global expression profiling. Several microarray technologies are currently in use; most include a glass slide platform with spotted cDNA or oligonucleotides. Till date, microarray technology has been used in the identification of regulatory genes, end-point defence genes, to understand the signal transduction processes underlying disease resistance and its intimate links to other physiological pathways. Microarray technology can be used for in-depth, simultaneous profiling of host/pathogen genes as the disease progresses from infection to resistance/susceptibility at different developmental stages of the host, which can be done in different environments, for clearer understanding of the processes involved. A thorough knowledge of plant disease resistance using successful combination of microarray and other high throughput techniques, as well as biochemical, genetic, and cell biological experiments is needed for practical application to secure and stabilize yield of many crop plants. This review starts with a brief introduction to microarray technology, followed by the basics of plant-pathogen interaction, the use of DNA microarrays over the last decade to unravel the mysteries of plant-pathogen interaction, and ends with the future prospects of this technology.

A Network Approach of Gene Co-expression in the Zea mays/Aspergillus flavus Pathosystem to Map Host/Pathogen Interaction Pathways.

PubMed

Musungu, Bryan M; Bhatnagar, Deepak; Brown, Robert L; Payne, Gary A; OBrian, Greg; Fakhoury, Ahmad M; Geisler, Matt

2016-01-01

A gene co-expression network (GEN) was generated using a dual RNA-seq study with the fungal pathogen Aspergillus flavus and its plant host Zea mays during the initial 3 days of infection. The analysis deciphered novel pathways and mapped genes of interest in both organisms during the infection. This network revealed a high degree of connectivity in many of the previously recognized pathways in Z. mays such as jasmonic acid, ethylene, and reactive oxygen species (ROS). For the pathogen A. flavus , a link between aflatoxin production and vesicular transport was identified within the network. There was significant interspecies correlation of expression between Z. mays and A. flavus for a subset of 104 Z. mays , and 1942 A. flavus genes. This resulted in an interspecies subnetwork enriched in multiple Z. mays genes involved in the production of ROS. In addition to the ROS from Z. mays , there was enrichment in the vesicular transport pathways and the aflatoxin pathway for A. flavus . Included in these genes, a key aflatoxin cluster regulator, AflS, was found to be co-regulated with multiple Z. mays ROS producing genes within the network, suggesting AflS may be monitoring host ROS levels. The entire GEN for both host and pathogen, and the subset of interspecies correlations, is presented as a tool for hypothesis generation and discovery for events in the early stages of fungal infection of Z. mays by A. flavus .

A Network Approach of Gene Co-expression in the Zea mays/Aspergillus flavus Pathosystem to Map Host/Pathogen Interaction Pathways

PubMed Central

Musungu, Bryan M.; Bhatnagar, Deepak; Brown, Robert L.; Payne, Gary A.; OBrian, Greg; Fakhoury, Ahmad M.; Geisler, Matt

2016-01-01

A gene co-expression network (GEN) was generated using a dual RNA-seq study with the fungal pathogen Aspergillus flavus and its plant host Zea mays during the initial 3 days of infection. The analysis deciphered novel pathways and mapped genes of interest in both organisms during the infection. This network revealed a high degree of connectivity in many of the previously recognized pathways in Z. mays such as jasmonic acid, ethylene, and reactive oxygen species (ROS). For the pathogen A. flavus, a link between aflatoxin production and vesicular transport was identified within the network. There was significant interspecies correlation of expression between Z. mays and A. flavus for a subset of 104 Z. mays, and 1942 A. flavus genes. This resulted in an interspecies subnetwork enriched in multiple Z. mays genes involved in the production of ROS. In addition to the ROS from Z. mays, there was enrichment in the vesicular transport pathways and the aflatoxin pathway for A. flavus. Included in these genes, a key aflatoxin cluster regulator, AflS, was found to be co-regulated with multiple Z. mays ROS producing genes within the network, suggesting AflS may be monitoring host ROS levels. The entire GEN for both host and pathogen, and the subset of interspecies correlations, is presented as a tool for hypothesis generation and discovery for events in the early stages of fungal infection of Z. mays by A. flavus. PMID:27917194

The FUN of identifying gene function in bacterial pathogens; insights from Salmonella functional genomics.

PubMed

Hammarlöf, Disa L; Canals, Rocío; Hinton, Jay C D

2013-10-01

The availability of thousands of genome sequences of bacterial pathogens poses a particular challenge because each genome contains hundreds of genes of unknown function (FUN). How can we easily discover which FUN genes encode important virulence factors? One solution is to combine two different functional genomic approaches. First, transcriptomics identifies bacterial FUN genes that show differential expression during the process of mammalian infection. Second, global mutagenesis identifies individual FUN genes that the pathogen requires to cause disease. The intersection of these datasets can reveal a small set of candidate genes most likely to encode novel virulence attributes. We demonstrate this approach with the Salmonella infection model, and propose that a similar strategy could be used for other bacterial pathogens. Copyright © 2013 Elsevier Ltd. All rights reserved.

Identification and functional characterization of the soybean GmaPPO12 promoter conferring Phytophthora sojae induced expression.

PubMed

Chai, Chunyue; Lin, Yanling; Shen, Danyu; Wu, Yuren; Li, Hongjuan; Dou, Daolong

2013-01-01

Identification of pathogen-inducible promoters largely lags behind cloning of the genes for disease resistance. Here, we cloned the soybean GmaPPO12 gene and found that it was rapidly and strongly induced by Phytophthorasojae infection. Computational analysis revealed that its promoter contained many known cis-elements, including several defense related transcriptional factor-binding boxes. We showed that the promoter could mediate induction of GUS expression upon infection in both transient expression assays in Nicotianabenthamiana and stable transgenic soybean hairy roots. Importantly, we demonstrated that pathogen-induced expression of the GmaPPO12 promoter was higher than that of the soybean GmaPR1a promoter. A progressive 5' and 3' deletion analysis revealed two fragments that were essential for promoter activity. Thus, the cloned promoter could be used in transgenic plants to enhance resistance to phytophthora pathogens, and the identified fragment could serve as a candidate to produce synthetic pathogen-induced promoters.

Gene expression platform for synthetic biology in the human pathogen Streptococcus pneumoniae.

PubMed

Sorg, Robin A; Kuipers, Oscar P; Veening, Jan-Willem

2015-03-20

The human pathogen Streptococcus pneumoniae (pneumococcus) is a bacterium that owes its success to complex gene expression regulation patterns on both the cellular and the population level. Expression of virulence factors enables a mostly hazard-free presence of the commensal, in balance with the host and niche competitors. Under specific circumstances, changes in this expression can result in a more aggressive behavior and the reversion to the invasive form as pathogen. These triggering conditions are very difficult to study due to the fact that environmental cues are often unknown or barely possible to simulate outside the host (in vitro). An alternative way of investigating expression patterns is found in synthetic biology approaches of reconstructing regulatory networks that mimic an observed behavior with orthogonal components. Here, we created a genetic platform suitable for synthetic biology approaches in S. pneumoniae and characterized a set of standardized promoters and reporters. We show that our system allows for fast and easy cloning with the BglBrick system and that reliable and robust gene expression after integration into the S. pneumoniae genome is achieved. In addition, the cloning system was extended to allow for direct linker-based assembly of ribosome binding sites, peptide tags, and fusion proteins, and we called this new generally applicable standard "BglFusion". The gene expression platform and the methods described in this study pave the way for employing synthetic biology approaches in S. pneumoniae.

Cyt toxin expression reveals an inverse regulation of insect and plant virulence factors of Dickeya dadantii.

PubMed

Costechareyre, Denis; Dridi, Bedis; Rahbé, Yvan; Condemine, Guy

2010-12-01

The plant pathogenic bacteria Dickeya dadantii is also a pathogen of the pea aphid Acyrthosiphon pisum. The genome of the bacteria contains four cyt genes, encoding homologues of Bacillus thuringiensis Cyt toxins, which are involved in its pathogenicity to insects. We show here that these genes are transcribed as an operon, and we determined the conditions necessary for their expression. Their expression is induced at high temperature and at an osmolarity equivalent to that found in the plant phloem sap. The regulators of cyt genes have also been identified: their expression is repressed by H-NS and VfmE and activated by PecS. These genes are already known to regulate plant virulence factors, but in an opposite way. When tested in a virulence assay by ingestion, the pecS mutant was almost non-pathogenic while hns and vfmE mutants behaved in the same way as the wild-type strain. Mutants of other regulators of plant virulence, GacA, OmpR and PhoP, that do not control Cyt toxin production, also showed reduced pathogenicity. In an assay by injection of bacteria, the gacA strain was less pathogenic but, surprisingly, the pecS mutant was slightly more virulent. These results show that Cyt toxins are not the only virulence factors required to kill aphids, and that these factors act at different stages of the infection. Moreover, their production is controlled by general virulence regulators known for their role in plant virulence. This integration could indicate that virulence towards insects is a normal mode of life for D. dadantii. © 2010 Society for Applied Microbiology and Blackwell Publishing Ltd.

A chromosomally encoded virulence factor protects the Lyme disease pathogen against host-adaptive immunity.

PubMed

Yang, Xiuli; Coleman, Adam S; Anguita, Juan; Pal, Utpal

2009-03-01

Borrelia burgdorferi, the bacterial pathogen of Lyme borreliosis, differentially expresses select genes in vivo, likely contributing to microbial persistence and disease. Expression analysis of spirochete genes encoding potential membrane proteins showed that surface-located membrane protein 1 (lmp1) transcripts were expressed at high levels in the infected murine heart, especially during early stages of infection. Mice and humans with diagnosed Lyme borreliosis also developed antibodies against Lmp1. Deletion of lmp1 severely impaired the pathogen's ability to persist in diverse murine tissues including the heart, and to induce disease, which was restored upon chromosomal complementation of the mutant with the lmp1 gene. Lmp1 performs an immune-related rather than a metabolic function, as its deletion did not affect microbial persistence in immunodeficient mice, but significantly decreased spirochete resistance to the borreliacidal effects of anti-B. burgdorferi sera in a complement-independent manner. These data demonstrate the existence of a virulence factor that helps the pathogen evade host-acquired immune defense and establish persistent infection in mammals.

Mitigating effects of pollen during paraquat exposure on gene expression and pathogen prevalence in Apis mellifera L.

PubMed

de Mattos, Igor Medici; Soares, Ademilson E E; Tarpy, David R

2018-01-01

Honey bee (Apis mellifera L.) populations have been experiencing notable mortality in Europe and North America. No single cause has been identified for these dramatic losses, but rather multiple interacting factors are likely responsible (such as pesticides, malnutrition, habitat loss, and pathogens). Paraquat is one of the most widely used non-selective herbicides, especially in developing countries. This herbicide is considered slightly toxic to honey bees, despite being reported as a highly effective inducer of oxidative stress in a wide range of living systems. Here, we test the effects of paraquat on the expression of detoxification and antioxidant-related genes, as well as on the dynamics of pathogen titers. Moreover, we tested the effects of pollen as mitigating factor to paraquat exposure. Our results show significant changes in the expression of several antioxidant-related and detoxification-related genes in the presence of paraquat, as well as an increase of pathogens titers. Finally, we demonstrate a mitigating effect of pollen through the up-regulation of specific genes and improvement of survival of bees exposed to paraquat. The presence of pollen in the diet was also correlated with a reduced prevalence of Nosema and viral pathogens. We discuss the importance of honey bees' nutrition, especially the availability of pollen, on colony losses chronically reported in the USA and Europe.

The Arabidopsis leaf transcriptome reveals distinct but also overlapping responses to colonization by phyllosphere commensals and pathogen infection with impact on plant health.

PubMed

Vogel, Christine; Bodenhausen, Natacha; Gruissem, Wilhelm; Vorholt, Julia A

2016-10-01

Plants are colonized by a variety of bacteria, most of which are not pathogenic. Currently, the plant responses to phyllosphere commensals or to pathogen infection in the presence of commensals are not well understood. Here, we examined the transcriptional response of Arabidopsis thaliana leaves to colonization by common commensal bacteria in a gnotobiotic system using RNA sequencing and conducted plant mutant assays. Arabidopsis responded differently to the model bacteria Sphingomonas melonis Fr1 (S.Fr1) and Methylobacterium extorquens PA1 (M.PA1). Whereas M.PA1 only marginally affected the expression of plant genes (< 10), S.Fr1 colonization changed the expression of almost 400 genes. For the latter, genes related to defense responses were activated and partly overlapped with those elicited by the pathogen Pseudomonas syringae DC3000 (Pst). As S.Fr1 is able to mediate plant protective activity against Pst, we tested plant immunity mutants and found that the pattern-recognition co-receptor mutant bak1/bkk1 showed attenuated S.Fr1-dependent plant protection. The experiments demonstrate that the plant responds differently to members of its natural phyllosphere microbiota. A subset of commensals trigger expression of defense-related genes and thereby may contribute to plant health upon pathogen encounter. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

Constitutive expression of a grapevine polygalacturonase-inhibiting protein affects gene expression and cell wall properties in uninfected tobacco

PubMed Central

2011-01-01

Background Polygalacturonase-inhibiting proteins (PGIPs) directly limit the effective ingress of fungal pathogens by inhibiting cell wall-degrading endopolygalacturonases (ePGs). Transgenic tobacco plants over-expressing grapevine (Vitis vinifera) Vvpgip1 have previously been shown to be resistant to Botrytis infection. In this study we characterized two of these PGIP over-expressing lines with known resistance phenotypes by gene expression and hormone profiling in the absence of pathogen infection. Results Global gene expression was performed by a cross-species microarray approach using a potato cDNA microarray. The degree of potential cross-hybridization between probes was modeled by a novel computational workflow designed in-house. Probe annotations were updated by predicting probe-to-transcript hybridizations and combining information derived from other plant species. Comparing uninfected Vvpgip1-overexpressing lines to wild-type (WT), 318 probes showed significant change in expression. Functional groups of genes involved in metabolism and associated to the cell wall were identified and consequent cell wall analysis revealed increased lignin-levels in the transgenic lines, but no major differences in cell wall-derived polysaccharides. GO enrichment analysis also identified genes responsive to auxin, which was supported by elevated indole-acetic acid (IAA) levels in the transgenic lines. Finally, a down-regulation of xyloglucan endotransglycosylase/hydrolases (XTHs), which are important in cell wall remodeling, was linked to a decrease in total XTH activity. Conclusions This evaluation of PGIP over-expressing plants performed under pathogen-free conditions to exclude the classical PGIP-ePG inhibition interaction indicates additional roles for PGIPs beyond the inhibition of ePGs. PMID:22078230

From deep-sea volcanoes to human pathogens: a conserved quorum-sensing signal in Epsilonproteobacteria.

PubMed

Pérez-Rodríguez, Ileana; Bolognini, Marie; Ricci, Jessica; Bini, Elisabetta; Vetriani, Costantino

2015-05-01

Chemosynthetic Epsilonproteobacteria from deep-sea hydrothermal vents colonize substrates exposed to steep thermal and redox gradients. In many bacteria, substrate attachment, biofilm formation, expression of virulence genes and host colonization are partly controlled via a cell density-dependent mechanism involving signal molecules, known as quorum sensing. Within the Epsilonproteobacteria, quorum sensing has been investigated only in human pathogens that use the luxS/autoinducer-2 (AI-2) mechanism to control the expression of some of these functions. In this study we showed that luxS is conserved in Epsilonproteobacteria and that pathogenic and mesophilic members of this class inherited this gene from a thermophilic ancestor. Furthermore, we provide evidence that the luxS gene is expressed--and a quorum-sensing signal is produced--during growth of Sulfurovum lithotrophicum and Caminibacter mediatlanticus, two Epsilonproteobacteria from deep-sea hydrothermal vents. Finally, we detected luxS transcripts in Epsilonproteobacteria-dominated biofilm communities collected from deep-sea hydrothermal vents. Taken together, our findings indicate that the epsiloproteobacterial lineage of the LuxS enzyme originated in high-temperature geothermal environments and that, in vent Epsilonproteobacteria, luxS expression is linked to the production of AI-2 signals, which are likely produced in situ at deep-sea vents. We conclude that the luxS gene is part of the ancestral epsilonproteobacterial genome and represents an evolutionary link that connects thermophiles to human pathogens.

Quantitative gene expression deregulation in mantle-cell lymphoma: correlation with clinical and biologic factors.

PubMed

Kienle, Dirk; Katzenberger, Tiemo; Ott, German; Saupe, Doreen; Benner, Axel; Kohlhammer, Holger; Barth, Thomas F E; Höller, Sylvia; Kalla, Jörg; Rosenwald, Andreas; Müller-Hermelink, Hans Konrad; Möller, Peter; Lichter, Peter; Döhner, Hartmut; Stilgenbauer, Stephan

2007-07-01

There is evidence for a direct role of quantitative gene expression deregulation in mantle-cell lymphoma (MCL) pathogenesis. Our aim was to investigate gene expression associations with other pathogenic factors and the significance of gene expression in a multivariate survival analysis. Quantitative expression of 20 genes of potential relevance for MCL prognosis and pathogenesis were analyzed using real-time reverse transcriptase polymerase chain reaction and correlated with clinical and genetic factors, tumor morphology, and Ki-67 index in 65 MCL samples. Genomic losses at the loci of TP53, RB1, and P16 were associated with reduced transcript levels of the respective genes, indicating a gene-dosage effect as the pathomechanism. Analysis of gene expression correlations between the candidate genes revealed a separation into two clusters, one dominated by proliferation activators, another by proliferation inhibitors and regulators of apoptosis. Whereas only weak associations were identified between gene expression and clinical parameters or blastoid morphology, several genes were correlated closely with the Ki-67 index, including the short CCND1 variant (positive correlation) and RB1, ATM, P27, and BMI (negative correlation). In multivariate survival analysis, expression levels of MYC, MDM2, EZH2, and CCND1 were the strongest prognostic factors independently of tumor proliferation and clinical factors. These results indicate a pathogenic contribution of several gene transcript levels to the biology and clinical course of MCL. Genes can be differentiated into factors contributing to proliferation deregulation, either by enhancement or loss of inhibition, and proliferation-independent factors potentially contributing to MCL pathogenesis by apoptosis impairment.

Transcriptional analysis of sweet orange trees co-infected with 'Candidatus Liberibacter asiaticus' and mild or severe strains of Citrus tristeza virus.

PubMed

Fu, Shimin; Shao, Jonathan; Paul, Cristina; Zhou, Changyong; Hartung, John S

2017-10-31

Citrus worldwide is threatened by huanglongbing (HLB) and tristeza diseases caused by 'Candidatus Liberibacter asiaticus' (CaLas) and Citrus tristeza virus (CTV). Although the pathogens are members of the α-proteobacteria and Closteroviridae, respectively, both are restricted to phloem cells in infected citrus and are transmitted by insect vectors. The response of sweet orange to single infection by either of these two pathogens has been characterized previously by global gene expression analysis. But because of the ubiquity of these pathogens where the diseases occur, co-infection by both pathogens is very common and could lead to increased disease severity based on synergism. We therefore co-inoculated sweet orange trees with CaLas and either a mild or a severe strain of CTV, and measured changes of gene expression in host plants. In plants infected with CaLas-B232, the overall alteration in gene expression was much greater in plants co-inoculated with the severe strain of CTV, B6, than when co-infected with the mild strain of CTV, B2. Plants co-infected with CaLas-B232 and either strain of CTV died but trees co-infected with CTV-B2 survived much longer than those co-infected with CTV-B6. Many important pathways were perturbed by both CTV-B2/CaLas-B232 and/or CTV-B6/CaLas-B232, but always more severely by CTV-B6/CaLas-B232. Genes related to cell wall modification and metal transport responded differently to infection by the pathogens in combination than by the same pathogens singly. The expressions of genes encoding phloem proteins and sucrose loading proteins were also differentially altered in response to CTV-B2 or CTV-B6 in combination with CaLas-B232, leading to different phloem environments in plants co-infected by CaLas and mild or severe CTV. Many host genes were expressed differently in response to dual infection as compared to single infections with the same pathogens. Interactions of the pathogens within the host may lead to a better or worse result for the host plant. CTV-B6 may exert a synergistic effect with CaLas-B232 in weakening the plant; on the other hand, the responses activated by the mild strain CTV-B2 may provide some beneficial effects against CaLas-B232 by increasing the defense response of the host.

Transcriptome Analysis of Bombyx mori Larval Midgut during Persistent and Pathogenic Cytoplasmic Polyhedrosis Virus Infection

PubMed Central

Kolliopoulou, Anna; Van Nieuwerburgh, Filip; Stravopodis, Dimitrios J.; Deforce, Dieter; Swevers, Luc; Smagghe, Guy

2015-01-01

Many insects can be persistently infected with viruses but do not show any obvious adverse effects with respect to physiology, development or reproduction. Here, Bombyx mori strain Daizo, persistently infected with cytoplasmic polyhedrosis virus (BmCPV), was used to study the host’s transcriptional response after pathogenic infection with the same virus in midgut tissue of larvae persistently and pathogenically infected as 2nd and 4th instars. Next generation sequencing revealed that from 13,769 expressed genes, 167 were upregulated and 141 downregulated in both larval instars following pathogenic infection. Several genes that could possibly be involved in B. mori immune response against BmCPV or that may be induced by the virus in order to increase infectivity were identified, whereas classification of differentially expressed transcripts (confirmed by qRT-PCR) resulted in gene categories related to physical barriers, immune responses, proteolytic / metabolic enzymes, heat-shock proteins, hormonal signaling and uncharacterized proteins. Comparison of our data with the available literature (pathogenic infection of persistently vs. non-persistently infected larvae) unveiled various similarities of response in both cases, which suggests that pre-existing persistent infection does not affect in a major way the transcriptome response against pathogenic infection. To investigate the possible host’s RNAi response against BmCPV challenge, the differential expression of RNAi-related genes and the accumulation of viral small RNAs (vsRNAs) were studied. During pathogenic infection, siRNA-like traces like the 2-fold up-regulation of the core RNAi genes Ago-2 and Dcr-2 as well as a peak of 20 nt small RNAs were observed. Interestingly, vsRNAs of the same size were detected at lower rates in persistently infected larvae. Collectively, our data provide an initial assessment of the relative significance of persistent infection of silkworm larvae on the host response following pathogenic infection with CPV, while they also highlight the relative importance of RNAi as an antiviral mechanism. PMID:25816294

Genes involved in Beauveria bassiana infection to Galleria mellonella.

PubMed

Chen, Anhui; Wang, Yulong; Shao, Ying; Zhou, Qiumei; Chen, Shanglong; Wu, Yonghua; Chen, Hongwei; Liu, Enqi

2018-05-01

The ascomycete fungus Beauveria bassiana is a natural pathogen of hundreds of insect species and is commercially produced as an environmentally friendly mycoinsecticide. Many genes involved in fungal insecticide infection have been identified but few have been further explored. In this study, we constructed three transcriptomes of B. bassiana at 24, 48 and 72 h post infection of insect pests (BbI) or control (BbC). There were 3148, 3613 and 4922 genes differentially expressed at 24, 48 and 72 h post BbI/BbC infection, respectively. A large number of genes and pathways involved in infection were identified. To further analyze those genes, expression patterns across different infection stages (0, 12, 24, 36, 48, 60, 72 and 84 h) were studied using quantitative RT-PCR. This analysis showed that the infection-related genes could be divided into four patterns: highly expressed throughout the whole infection process (thioredoxin 1); highly expressed during early stages of infection but lowly expressed after the insect death (adhesin protein Mad1); lowly expressed during early infection but highly expressed after insect death (cation transporter, OpS13); or lowly expressed across the entire infection process (catalase protein). The data provide novel insights into the insect-pathogen interaction and help to uncover the molecular mechanisms involved in fungal infection of insect pests.

Evaluation of reference genes in Vibrio parahaemolyticus for gene expression analysis using quantitative RT-PCR

USDA-ARS?s Scientific Manuscript database

Vibrio parahaemolyticus is a significant human pathogen capable of causing foodborne gastroenteritis associated with the consumption of contaminated raw or undercooked seafood. Quantitative RT-PCR (qRT-PCR) is a useful tool for studying gene expression in V. parahaemolyticus to characterize the viru...

RNA-seq comparative analysis of Peking ducks spleen gene expression 24 h post-infected with duck plague virulent or attenuated virus.

PubMed

Liu, Tian; Cheng, Anchun; Wang, Mingshu; Jia, Renyong; Yang, Qiao; Wu, Ying; Sun, Kunfeng; Zhu, Dekang; Chen, Shun; Liu, Mafeng; Zhao, XinXin; Chen, Xiaoyue

2017-09-13

Duck plague virus (DPV), a member of alphaherpesvirus sub-family, can cause significant economic losses on duck farms in China. DPV Chinese virulent strain (CHv) is highly pathogenic and could induce massive ducks death. Attenuated DPV vaccines (CHa) have been put into service against duck plague with billions of doses in China each year. Researches on DPV have been development for many years, however, a comprehensive understanding of molecular mechanisms underlying pathogenicity of CHv strain and protection of CHa strain to ducks is still blank. In present study, we performed RNA-seq technology to analyze transcriptome profiling of duck spleens for the first time to identify differentially expressed genes (DEGs) associated with the infection of CHv and CHa at 24 h. Comparison of gene expression with mock ducks revealed 748 DEGs and 484 DEGs after CHv and CHa infection, respectively. Gene pathway analysis of DEGs highlighted valuable biological processes involved in host immune response, cell apoptosis and viral invasion. Genes expressed in those pathways were different in CHv infected duck spleens and CHa vaccinated duck spleens. The results may provide valuable information for us to explore the reasons of pathogenicity caused by CHv strain and protection activated by CHa strain.

Analysis of the grape (Vitis vinifera L.) thaumatin-like protein (TLP) gene family and demonstration that TLP29 contributes to disease resistance.

PubMed

Yan, Xiaoxiao; Qiao, Hengbo; Zhang, Xiuming; Guo, Chunlei; Wang, Mengnan; Wang, Yuejin; Wang, Xiping

2017-06-27

Thaumatin-like protein (TLP) is present as a large family in plants, and individual members play different roles in various responses to biotic and abiotic stresses. Here we studied the role of 33 putative grape (Vitis vinifera L.) TLP genes (VvTLP) in grape disease resistance. Heat maps analysis compared the expression profiles of 33 genes in disease resistant and susceptible grape species infected with anthracnose (Elsinoe ampelina), powdery mildew (Erysiphe necator) or Botrytis cinerea. Among these 33 genes, the expression level of TLP29 increased following the three pathogens inoculations, and its homolog from the disease resistant Chinese wild grape V. quinquangularis cv. 'Shang-24', was focused for functional studies. Over-expression of TLP29 from grape 'Shang-24' (VqTLP29) in Arabidopsis thaliana enhanced its resistance to powdery mildew and the bacterium Pseudomonas syringae pv. tomato DC3000, but decreased resistance to B. cinerea. Moreover, the stomatal closure immunity response to pathogen associated molecular patterns was strengthened in the transgenic lines. A comparison of the expression profiles of various resistance-related genes after infection with different pathogens indicated that VqTLP29 may be involved in the salicylic acid and jasmonic acid/ethylene signaling pathways.

Identification and characterization of NF-YB family genes in tung tree.

PubMed

Yang, Susu; Wang, Yangdong; Yin, Hengfu; Guo, Haobo; Gao, Ming; Zhu, Huiping; Chen, Yicun

2015-12-01

The NF-YB transcription factor gene family encodes a subunit of the CCAAT box-binding factor (CBF), a highly conserved trimeric activator that strongly binds to the CCAAT box promoter element. Studies on model plants have shown that NF-YB proteins participate in important developmental and physiological processes, but little is known about NF-YB proteins in trees. Here, we identified seven NF-YB transcription factor-encoding genes in Vernicia fordii, an important oilseed tree in China. A phylogenetic analysis separated the genes into two groups; non-LEC1 type (VfNF-YB1, 5, 7, 9, 11, 13) and LEC1-type (VfNF-YB 14). A gene structure analysis showed that VfNF-YB 5 has three introns and the other genes have no introns. The seven VfNF-YB sequences contain highly conserved domains, a disordered region at the N terminus, and two long helix structures at the C terminus. Phylogenetic analyses showed that VfNF-YB family genes are highly homologous to GmNF-YB genes, and many of them are closely related to functionally characterized NF-YBs. In expression analyses of various tissues (root, stem, leaf, and kernel) and the root during pathogen infection, VfNF-YB1, 5, and 11 were dominantly expressed in kernels, and VfNF-YB7 and 9 were expressed only in the root. Different VfNF-YB family genes showed different responses to pathogen infection, suggesting that they play different roles in the pathogen response. Together, these findings represent the first extensive evaluation of the NF-YB family in tung tree and provide a foundation for dissecting the functions of VfNF-YB genes in seed development, stress adaption, fatty acid synthesis, and pathogen response.

Investigating Conservation of the Cell-Cycle-Regulated Transcriptional Program in the Fungal Pathogen, Cryptococcus neoformans

PubMed Central

Sierra, Crystal S.; Haase, Steven B.

2016-01-01

The pathogenic yeast Cryptococcus neoformans causes fungal meningitis in immune-compromised patients. Cell proliferation in the budding yeast form is required for C. neoformans to infect human hosts, and virulence factors such as capsule formation and melanin production are affected by cell-cycle perturbation. Thus, understanding cell-cycle regulation is critical for a full understanding of virulence factors for disease. Our group and others have demonstrated that a large fraction of genes in Saccharomyces cerevisiae is expressed periodically during the cell cycle, and that proper regulation of this transcriptional program is important for proper cell division. Despite the evolutionary divergence of the two budding yeasts, we found that a similar percentage of all genes (~20%) is periodically expressed during the cell cycle in both yeasts. However, the temporal ordering of periodic expression has diverged for some orthologous cell-cycle genes, especially those related to bud emergence and bud growth. Genes regulating DNA replication and mitosis exhibited a conserved ordering in both yeasts, suggesting that essential cell-cycle processes are conserved in periodicity and in timing of expression (i.e. duplication before division). In S. cerevisiae cells, we have proposed that an interconnected network of periodic transcription factors (TFs) controls the bulk of the cell-cycle transcriptional program. We found that temporal ordering of orthologous network TFs was not always maintained; however, the TF network topology at cell-cycle commitment appears to be conserved in C. neoformans. During the C. neoformans cell cycle, DNA replication genes, mitosis genes, and 40 genes involved in virulence are periodically expressed. Future work toward understanding the gene regulatory network that controls cell-cycle genes is critical for developing novel antifungals to inhibit pathogen proliferation. PMID:27918582

Identification of Powdery Mildew Responsive Genes in Hevea brasiliensis through mRNA Differential Display

PubMed Central

Li, Xiang; Bi, Zhenghong; Di, Rong; Liang, Peng; He, Qiguang; Liu, Wenbo; Miao, Weiguo; Zheng, Fucong

2016-01-01

Powdery mildew is an important disease of rubber trees caused by Oidium heveae B. A. Steinmann. As far as we know, none of the resistance genes related to powdery mildew have been isolated from the rubber tree. There is little information available at the molecular level regarding how a rubber tree develops defense mechanisms against this pathogen. We have studied rubber tree mRNA transcripts from the resistant RRIC52 cultivar by differential display analysis. Leaves inoculated with the spores of O. heveae were collected from 0 to 120 hpi in order to identify pathogen-regulated genes at different infection stages. We identified 78 rubber tree genes that were differentially expressed during the plant–pathogen interaction. BLAST analysis for these 78 ESTs classified them into seven functional groups: cell wall and membrane pathways, transcription factor and regulatory proteins, transporters, signal transduction, phytoalexin biosynthesis, other metabolism functions, and unknown functions. The gene expression for eight of these genes was validated by qRT-PCR in both RRIC52 and the partially susceptible Reyan 7-33-97 cultivars, revealing the similar or differential changes of gene expressions between these two cultivars. This study has improved our overall understanding of the molecular mechanisms of rubber tree resistance to powdery mildew. PMID:26840302

Regulation of disease-responsive genes mediated by epigenetic factors: interaction of Arabidopsis-Pseudomonas.

PubMed

De-La-Peña, Clelia; Rangel-Cano, Alicia; Alvarez-Venegas, Raúl

2012-05-01

Genes in eukaryotic organisms function within the context of chromatin, and the mechanisms that modulate the structure of chromatin are defined as epigenetic. In Arabidopsis, pathogen infection induces the expression of at least one histone deacetylase, suggesting that histone acetylation/deacetylation has an important role in the pathogenic response in plants. How/whether histone methylation affects gene response to pathogen infection is unknown. To gain a better understanding of the epigenetic mechanisms regulating the interaction between Pseudomonas syringae and Arabidopsis thaliana, we analysed three different Arabidopsis ash1-related (absent, small or homeotic discs 1) mutants. We found that the loss of function of ASHH2 and ASHR1 resulted in faster hypersensitive responses (HRs) to both mutant (hrpA) and pathogenic (DC3000) strains of P. syringae, whereas control (Col-0) and ashr3 mutants appeared to be more resistant to the infection after 2 days. Furthermore, we showed that, in the ashr3 background, the PR1 gene (PATHOGENESIS-RELATED GENE 1) displayed the highest expression levels on infection with DC3000, correlating with increased resistance against this pathogen. Our results show that, in both the ashr1 and ashh2 backgrounds, the histone H3 lysine 4 dimethylation (H3K4me2) levels decreased at the promoter region of PR1 on infection with the DC3000 strain, suggesting that an epigenetically regulated PR1 expression is involved in the plant defence. Our results suggest that histone methylation in response to pathogen infection may be a critical component in the signalling and defence processes occurring between plants and microbes. © 2011 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2011 BSPP AND BLACKWELL PUBLISHING LTD.

Use of Gnotobiotic Zebrafish to Study Vibrio anguillarum Pathogenicity

PubMed Central

Oyarbide, Usua; Iturria, Iñaki; Rainieri, Sandra

2015-01-01

Abstract We evaluated the use of the gnotobiotic zebrafish system to study the effects of bacterial infection, and analyzed expression of genes involved in zebrafish innate immunity. Using a GFP-labeled strain of Vibrio anguillarum, we fluorescently monitored colonization of the zebrafish intestinal tract and used gene expression analysis to compare changes in genes involved in innate immunity between nongnotobiotic and gnotobiotic larvae. The experiments performed with the gnotobiotic zebrafish reveal new insights into V. anguillarum pathogenesis. Specifically, an alteration of the host immune system was detected through the suppression of a number of innate immune genes (NFKB, IL1B, TLR4, MPX, and TRF) during the first 3 h post infection. This immunomodulation can be indicative of a “stealth mechanism” of mucus invasion in which the pathogen found a sheltered niche, a typical trait of intracellular pathogens. PMID:25548877

Transcription analysis of stx1, marA, and eaeA genes in Escherichia coli O157:H7 treated with sodium benzoate.

PubMed

Critzer, Faith J; Dsouza, Doris H; Golden, David A

2008-07-01

Expression of the multiple antibiotic resistance (mar) operon causes increased antimicrobial resistance in bacterial pathogens. The activator of this operon, MarA, can alter expression of >60 genes in Escherichia coli K-12. However, data on the expression of virulence and resistance genes when foodborne pathogens are exposed to antimicrobial agents are lacking. This study was conducted to determine transcription of marA (mar activator), stx1 (Shiga toxin 1), and eaeA (intimin) genes of E. coli O157:H7 EDL933 as affected by sodium benzoate. E. coli O157:H7 was grown in Luria-Bertani broth containing 0 (control) and 1% sodium benzoate at 37 degrees C for 24 h, and total RNA was extracted. Primers were designed for hemX (209 bp; housekeeping gene), marA (261 bp), and eaeA (223 bp) genes; previously reported primers were used for stx1. Tenfold dilutions of RNA were used in a real-time one-step reverse transcriptase PCR to determine transcription levels. All experiments were conducted in triplicate, and product detection was validated by gel electrophoresis. For marA and stx1, real-time one-step reverse transcriptase PCR products were detected at a 1-log-greater dilution in sodium benzoate-treated cells than in control cells, although cell numbers for each were similar (7.28 and 7.57 log CFU/ml, respectively). This indicates a greater (albeit slight) level of their transcription in treated cells than in control cells. No difference in expression of eaeA was observed. HemX is a putative uroporphyrinogen III methylase. The hemX gene was expressed at the same level in control and treated cells, validating hemX as an appropriate housekeeping marker. These data indicate that stx1 and marA genes could play a role in pathogen virulence and survival when treated with sodium benzoate, whereas eaeA expression is not altered. Understanding adaptations of E. coli O157:H7 during antimicrobial exposure is essential to better understand and implement methods to inhibit or control survival of this pathogen in foods.

An emerging cyberinfrastructure for biodefense pathogen and pathogen-host data.

PubMed

Zhang, C; Crasta, O; Cammer, S; Will, R; Kenyon, R; Sullivan, D; Yu, Q; Sun, W; Jha, R; Liu, D; Xue, T; Zhang, Y; Moore, M; McGarvey, P; Huang, H; Chen, Y; Zhang, J; Mazumder, R; Wu, C; Sobral, B

2008-01-01

The NIAID-funded Biodefense Proteomics Resource Center (RC) provides storage, dissemination, visualization and analysis capabilities for the experimental data deposited by seven Proteomics Research Centers (PRCs). The data and its publication is to support researchers working to discover candidates for the next generation of vaccines, therapeutics and diagnostics against NIAID's Category A, B and C priority pathogens. The data includes transcriptional profiles, protein profiles, protein structural data and host-pathogen protein interactions, in the context of the pathogen life cycle in vivo and in vitro. The database has stored and supported host or pathogen data derived from Bacillus, Brucella, Cryptosporidium, Salmonella, SARS, Toxoplasma, Vibrio and Yersinia, human tissue libraries, and mouse macrophages. These publicly available data cover diverse data types such as mass spectrometry, yeast two-hybrid (Y2H), gene expression profiles, X-ray and NMR determined protein structures and protein expression clones. The growing database covers over 23 000 unique genes/proteins from different experiments and organisms. All of the genes/proteins are annotated and integrated across experiments using UniProt Knowledgebase (UniProtKB) accession numbers. The web-interface for the database enables searching, querying and downloading at the level of experiment, group and individual gene(s)/protein(s) via UniProtKB accession numbers or protein function keywords. The system is accessible at http://www.proteomicsresource.org/.

The Rhizoctonia solani AG1-IB (isolate 7/3/14) transcriptome during interaction with the host plant lettuce (Lactuca sativa L.).

PubMed

Verwaaijen, Bart; Wibberg, Daniel; Kröber, Magdalena; Winkler, Anika; Zrenner, Rita; Bednarz, Hanna; Niehaus, Karsten; Grosch, Rita; Pühler, Alfred; Schlüter, Andreas

2017-01-01

The necrotrophic pathogen Rhizoctonia solani is one of the most economically important soil-borne pathogens of crop plants. Isolates of R. solani AG1-IB are the major pathogens responsible for bottom-rot of lettuce (Lactuca sativa L.) and are also responsible for diseases in other plant species. Currently, there is lack of information regarding the molecular responses in R. solani during the pathogenic interaction between the necrotrophic soil-borne pathogen and its host plant. The genome of R. solani AG1-IB (isolate 7/3/14) was recently established to obtain insights into its putative pathogenicity determinants. In this study, the transcriptional activity of R. solani AG1-IB was followed during the course of its pathogenic interaction with the host plant lettuce under controlled conditions. Based on visual observations, three distinct pathogen-host interaction zones on lettuce leaves were defined which covered different phases of disease progression on tissue inoculated with the AG1-IB (isolate 7/3/14). The zones were defined as: Zone 1-symptomless, Zone 2-light brown discoloration, and Zone 3-dark brown, necrotic lesions. Differences in R. solani hyphae structure in these three zones were investigated by microscopic observation. Transcriptional activity within these three interaction zones was used to represent the course of R. solani disease progression applying high-throughput RNA sequencing (RNA-Seq) analysis of samples collected from each Zone. The resulting three transcriptome data sets were analyzed for their highest expressed genes and for differentially transcribed genes between the respective interaction zones. Among the highest expressed genes was a group of not previously described genes which were transcribed exclusively during early stages of interaction, in Zones 1 and 2. Previously described importance of up-regulation in R. solani agglutinin genes during disease progression could be further confirmed; here, the corresponding genes exhibited extremely high transcription levels. Most differentially higher expressed transcripts were found within Zone 2. In Zone 3, the zone with the strongest degree of interaction, gene transcripts indicative of apoptotic activity were highly abundant. The transcriptome data presented in this work support previous models of the disease and interaction cycle of R. solani and lettuce and may influence effective techniques for control of this pathogen.

Catabolite and Oxygen Regulation of Enterohemorrhagic Escherichia coli Virulence.

PubMed

Carlson-Banning, Kimberly M; Sperandio, Vanessa

2016-11-22

The biogeography of the gut is diverse in its longitudinal axis, as well as within specific microenvironments. Differential oxygenation and nutrient composition drive the membership of microbial communities in these habitats. Moreover, enteric pathogens can orchestrate further modifications to gain a competitive advantage toward host colonization. These pathogens are versatile and adept when exploiting the human colon. They expertly navigate complex environmental cues and interkingdom signaling to colonize and infect their hosts. Here we demonstrate how enterohemorrhagic Escherichia coli (EHEC) uses three sugar-sensing transcription factors, Cra, KdpE, and FusR, to exquisitely regulate the expression of virulence factors associated with its type III secretion system (T3SS) when exposed to various oxygen concentrations. We also explored the effect of mucin-derived nonpreferred carbon sources on EHEC growth and expression of virulence genes. Taken together, the results show that EHEC represses the expression of its T3SS when oxygen is absent, mimicking the largely anaerobic lumen, and activates its T3SS when oxygen is available through Cra. In addition, when EHEC senses mucin-derived sugars heavily present in the O-linked and N-linked glycans of the large intestine, virulence gene expression is initiated. Sugars derived from pectin, a complex plant polysaccharide digested in the large intestine, also increased virulence gene expression. Not only does EHEC sense host- and microbiota-derived interkingdom signals, it also uses oxygen availability and mucin-derived sugars liberated by the microbiota to stimulate expression of the T3SS. This precision in gene regulation allows EHEC to be an efficient pathogen with an extremely low infectious dose. Enteric pathogens have to be crafty when interpreting multiple environmental cues to successfully establish themselves within complex and diverse gut microenvironments. Differences in oxygen tension and nutrient composition determine the biogeography of the gut microbiota and provide unique niches that can be exploited by enteric pathogens. EHEC is an enteric pathogen that colonizes the colon and causes outbreaks of bloody diarrhea and hemolytic-uremic syndrome worldwide. It has a very low infectious dose, which requires it to be an extremely effective pathogen. Hence, here we show that EHEC senses multiple sugar sources and oxygen levels to optimally control the expression of its virulence repertoire. This exquisite regulatory control equips EHEC to sense different intestinal compartments to colonize the host. Copyright © 2016 Carlson-Banning and Sperandio.

Infection by Rhodococcus fascians maintains cotyledons as a sink tissue for the pathogen

PubMed Central

Dhandapani, Pragatheswari; Song, Jiancheng; Novak, Ondrej

2017-01-01

Background and Aims Pisum sativum L. (pea) seed is a source of carbohydrate and protein for the developing plant. By studying pea seeds inoculated by the cytokinin-producing bacterium, Rhodococcus fascians, we sought to determine the impact of both an epiphytic (avirulent) strain and a pathogenic strain on source–sink activity within the cotyledons during and following germination. Methods Bacterial spread was monitored microscopically, and real-time reverse transcription–quantitative PCR was used to determine the expression of cytokinin biosynthesis, degradation and response regulator gene family members, along with expression of family members of SWEET, SUT, CWINV and AAP genes – gene families identified initially in pea by transcriptomic analysis. The endogenous cytokinin content was also determined. Key Results The cotyledons infected by the virulent strain remained intact and turned green, while multiple shoots were formed and root growth was reduced. The epiphytic strain had no such marked impact. Isopentenyl adenine was elevated in the cotyledons infected by the virulent strain. Strong expression of RfIPT, RfLOG and RfCKX was detected in the cotyledons infected by the virulent strain throughout the experiment, with elevated expression also observed for PsSWEET, PsSUT and PsINV gene family members. The epiphytic strain had some impact on the expression of these genes, especially at the later stages of reserve mobilization from the cotyledons. Conclusions The pathogenic strain retained the cotyledons as a sink tissue for the pathogen rather than the cotyledon converting completely to a source tissue for the germinating plant. We suggest that the interaction of cytokinins, CWINVs and SWEETs may lead to the loss of apical dominance and the appearance of multiple shoots. PMID:27864224

Functional modulation of human intestinal epithelial cell responses by Bifidobacterium infantis and Lactobacillus salivarius

PubMed Central

O'Hara, Ann M; O'Regan, Padraig; Fanning, Áine; O'Mahony, Caitlin; MacSharry, John; Lyons, Anne; Bienenstock, John; O'Mahony, Liam; Shanahan, Fergus

2006-01-01

Intestinal epithelial cells (IECs) and dendritic cells (DCs) play a pivotal role in antigen sampling and the maintenance of gut homeostasis. However, the interaction of commensal bacteria with the intestinal surface remains incompletely understood. Here we investigated immune cell responses to commensal and pathogenic bacteria. HT-29 human IECs were incubated with Bifidobacterium infantis 35624, Lactobacillus salivarius UCC118 or Salmonella typhimurium UK1 for varying times, or were pretreated with a probiotic for 2 hr prior to stimulation with S. typhimurium or flagellin. Gene arrays were used to examine inflammatory gene expression. Nuclear factor (NF)-κB activation, interleukin (IL)-8 secretion, pathogen adherence to IECs, and mucin-3 (MUC3) and E-cadherin gene expression were assayed by TransAM assay, enzyme-linked immunosorbent assay (ELISA), fluorescence, and real-time reverse transcriptase–polymerase chain reaction (RT-PCR), respectively. IL-10 and tumour necrosis factor (TNF)-α secretion by bacteria-treated peripheral blood-derived DCs were measured using ELISA. S. typhimurium increased expression of 36 of the 847 immune-related genes assayed, including NF-κB and IL-8. The commensal bacteria did not alter expression levels of any of the 847 genes. However, B. infantis and L. salivarius attenuated both IL-8 secretion at baseline and S. typhimurium-induced pro-inflammatory responses. B. infantis also limited flagellin-induced IL-8 protein secretion. The commensal bacteria did not increase MUC3 or E-cadherin expression, or interfere with pathogen binding to HT-29 cells, but they did stimulate IL-10 and TNF-α secretion by DCs. The data demonstrate that, although the intestinal epithelium is immunologically quiescent when it encounters B. infantis or L. salivarius, these commensal bacteria exert immunomodulatory effects on intestinal immune cells that mediate host responses to flagellin and enteric pathogens. PMID:16771855

Gene expression profiling of the plant pathogenic basidiomycetous fungus Rhizoctonia solani AG 4 reveals putative virulence factors

USDA-ARS?s Scientific Manuscript database

Rhizoctonia solani is a ubiquitous basidiomycetous soilborne fungal pathogen causing damping off of seedlings, aerial blights and postharvest diseases. To gain insight into the molecular mechanisms of pathogenesis a global approach based on analysis of expressed sequence tags (ESTs) was undertaken. ...

Transgenic expression of medicago truncatula PR10 and PR5 promoters in alfalfa shows pathogen-induced up-regulation of transgene expression

USDA-ARS?s Scientific Manuscript database

Genetic modification of alfalfa to introduce novel traits requires promoters for controlling gene expression. Promoters that are constitutively activated for expression of genes that enhance disease resistance pose a great energy load on the plant and exert a strong selective pressure on the pathoge...

Genomic mechanisms accounting for the adaptation to parasitism in nematode-trapping fungi.

PubMed

Meerupati, Tejashwari; Andersson, Karl-Magnus; Friman, Eva; Kumar, Dharmendra; Tunlid, Anders; Ahrén, Dag

2013-11-01

Orbiliomycetes is one of the earliest diverging branches of the filamentous ascomycetes. The class contains nematode-trapping fungi that form unique infection structures, called traps, to capture and kill free-living nematodes. The traps have evolved differently along several lineages and include adhesive traps (knobs, nets or branches) and constricting rings. We show, by genome sequencing of the knob-forming species Monacrosporium haptotylum and comparison with the net-forming species Arthrobotrys oligospora, that two genomic mechanisms are likely to have been important for the adaptation to parasitism in these fungi. Firstly, the expansion of protein domain families and the large number of species-specific genes indicated that gene duplication followed by functional diversification had a major role in the evolution of the nematode-trapping fungi. Gene expression indicated that many of these genes are important for pathogenicity. Secondly, gene expression of orthologs between the two fungi during infection indicated that differential regulation was an important mechanism for the evolution of parasitism in nematode-trapping fungi. Many of the highly expressed and highly upregulated M. haptotylum transcripts during the early stages of nematode infection were species-specific and encoded small secreted proteins (SSPs) that were affected by repeat-induced point mutations (RIP). An active RIP mechanism was revealed by lack of repeats, dinucleotide bias in repeats and genes, low proportion of recent gene duplicates, and reduction of recent gene family expansions. The high expression and rapid divergence of SSPs indicate a striking similarity in the infection mechanisms of nematode-trapping fungi and plant and insect pathogens from the crown groups of the filamentous ascomycetes (Pezizomycotina). The patterns of gene family expansions in the nematode-trapping fungi were more similar to plant pathogens than to insect and animal pathogens. The observation of RIP activity in the Orbiliomycetes suggested that this mechanism was present early in the evolution of the filamentous ascomycetes.

Genomic Mechanisms Accounting for the Adaptation to Parasitism in Nematode-Trapping Fungi

PubMed Central

Meerupati, Tejashwari; Andersson, Karl-Magnus; Friman, Eva; Kumar, Dharmendra; Tunlid, Anders; Ahrén, Dag

2013-01-01

Orbiliomycetes is one of the earliest diverging branches of the filamentous ascomycetes. The class contains nematode-trapping fungi that form unique infection structures, called traps, to capture and kill free-living nematodes. The traps have evolved differently along several lineages and include adhesive traps (knobs, nets or branches) and constricting rings. We show, by genome sequencing of the knob-forming species Monacrosporium haptotylum and comparison with the net-forming species Arthrobotrys oligospora, that two genomic mechanisms are likely to have been important for the adaptation to parasitism in these fungi. Firstly, the expansion of protein domain families and the large number of species-specific genes indicated that gene duplication followed by functional diversification had a major role in the evolution of the nematode-trapping fungi. Gene expression indicated that many of these genes are important for pathogenicity. Secondly, gene expression of orthologs between the two fungi during infection indicated that differential regulation was an important mechanism for the evolution of parasitism in nematode-trapping fungi. Many of the highly expressed and highly upregulated M. haptotylum transcripts during the early stages of nematode infection were species-specific and encoded small secreted proteins (SSPs) that were affected by repeat-induced point mutations (RIP). An active RIP mechanism was revealed by lack of repeats, dinucleotide bias in repeats and genes, low proportion of recent gene duplicates, and reduction of recent gene family expansions. The high expression and rapid divergence of SSPs indicate a striking similarity in the infection mechanisms of nematode-trapping fungi and plant and insect pathogens from the crown groups of the filamentous ascomycetes (Pezizomycotina). The patterns of gene family expansions in the nematode-trapping fungi were more similar to plant pathogens than to insect and animal pathogens. The observation of RIP activity in the Orbiliomycetes suggested that this mechanism was present early in the evolution of the filamentous ascomycetes. PMID:24244185

Tissue-specific and pathogen-inducible expression of a fusion protein containing a Fusarium-specific antibody and a fungal chitinase protects wheat against Fusarium pathogens and mycotoxins.

PubMed

Cheng, Wei; Li, He-Ping; Zhang, Jing-Bo; Du, Hong-Jie; Wei, Qi-Yong; Huang, Tao; Yang, Peng; Kong, Xian-Wei; Liao, Yu-Cai

2015-06-01

Fusarium head blight (FHB) in wheat and other small grain cereals is a globally devastating disease caused by toxigenic Fusarium pathogens. Controlling FHB is a challenge because germplasm that is naturally resistant against these pathogens is inadequate. Current control measures rely on fungicides. Here, an antibody fusion comprised of the Fusarium spp.-specific recombinant antibody gene CWP2 derived from chicken, and the endochitinase gene Ech42 from the biocontrol fungus Trichoderma atroviride was introduced into the elite wheat cultivar Zhengmai9023 by particle bombardment. Expression of this fusion gene was regulated by the lemma/palea-specific promoter Lem2 derived from barley; its expression was confirmed as lemma/palea-specific in transgenic wheat. Single-floret inoculation of independent transgenic wheat lines of the T3 to T6 generations revealed significant resistance (type II) to fungal spreading, and natural infection assays in the field showed significant resistance (type I) to initial infection. Gas chromatography-mass spectrometry analysis revealed marked reduction of mycotoxins in the grains of the transgenic wheat lines. Progenies of crosses between the transgenic lines and the FHB-susceptible cultivar Huamai13 also showed significantly enhanced FHB resistance. Quantitative real-time PCR analysis revealed that the tissue-specific expression of the antibody fusion was induced by salicylic acid drenching and induced to a greater extent by F. graminearum infection. Histochemical analysis showed substantial restriction of mycelial growth in the lemma tissues of the transgenic plants. Thus, the combined tissue-specific and pathogen-inducible expression of this Fusarium-specific antibody fusion can effectively protect wheat against Fusarium pathogens and reduce mycotoxin content in grain. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Antioxidant genes of plants and fungal pathogens are distinctly regulated during disease development in different Rhizoctonia solani pathosystems.

PubMed

Samsatly, Jamil; Copley, Tanya R; Jabaji, Suha H

2018-01-01

Biotic stress, as a result of plant-pathogen interactions, induces the accumulation of reactive oxygen species in the cells, causing severe oxidative damage to plants and pathogens. To overcome this damage, both the host and pathogen have developed antioxidant systems to quench excess ROS and keep ROS production and scavenging systems under control. Data on ROS-scavenging systems in the necrotrophic plant pathogen Rhizoctonia solani are just emerging. We formerly identified vitamin B6 biosynthetic machinery of R. solani AG3 as a powerful antioxidant exhibiting a high ability to quench ROS, similar to CATALASE (CAT) and GLUTATHIONE S-TRANSFERASE (GST). Here, we provide evidence on the involvement of R. solani vitamin B6 biosynthetic pathway genes; RsolPDX1 (KF620111.1), RsolPDX2 (KF620112.1), and RsolPLR (KJ395592.1) in vitamin B6 de novo biosynthesis by yeast complementation assays. Since gene expression studies focusing on oxidative stress responses of both the plant and the pathogen following R. solani infection are very limited, this study is the first coexpression analysis of genes encoding vitamin B6, CAT and GST in plant and fungal tissues of three pathosystems during interaction of different AG groups of R. solani with their respective hosts. The findings indicate that distinct expression patterns of fungal and host antioxidant genes were correlated in necrotic tissues and their surrounding areas in each of the three R. solani pathosystems: potato sprout-R. solani AG3; soybean hypocotyl-R. solani AG4 and soybean leaves-R. solani AG1-IA interactions. Levels of ROS increased in all types of potato and soybean tissues, and in fungal hyphae following infection of R. solani AGs as determined by non-fluorescence and fluorescence methods using H2DCF-DA and DAB, respectively. Overall, we demonstrate that the co-expression and accumulation of certain plant and pathogen ROS-antioxidant related genes in each pathosystem are highlighted and might be critical during disease development from the plant's point of view, and in pathogenicity and developing of infection structures from the fungal point of view.

Antioxidant genes of plants and fungal pathogens are distinctly regulated during disease development in different Rhizoctonia solani pathosystems

PubMed Central

Samsatly, Jamil; Copley, Tanya R.

2018-01-01

Biotic stress, as a result of plant-pathogen interactions, induces the accumulation of reactive oxygen species in the cells, causing severe oxidative damage to plants and pathogens. To overcome this damage, both the host and pathogen have developed antioxidant systems to quench excess ROS and keep ROS production and scavenging systems under control. Data on ROS-scavenging systems in the necrotrophic plant pathogen Rhizoctonia solani are just emerging. We formerly identified vitamin B6 biosynthetic machinery of R. solani AG3 as a powerful antioxidant exhibiting a high ability to quench ROS, similar to CATALASE (CAT) and GLUTATHIONE S-TRANSFERASE (GST). Here, we provide evidence on the involvement of R. solani vitamin B6 biosynthetic pathway genes; RsolPDX1 (KF620111.1), RsolPDX2 (KF620112.1), and RsolPLR (KJ395592.1) in vitamin B6 de novo biosynthesis by yeast complementation assays. Since gene expression studies focusing on oxidative stress responses of both the plant and the pathogen following R. solani infection are very limited, this study is the first coexpression analysis of genes encoding vitamin B6, CAT and GST in plant and fungal tissues of three pathosystems during interaction of different AG groups of R. solani with their respective hosts. The findings indicate that distinct expression patterns of fungal and host antioxidant genes were correlated in necrotic tissues and their surrounding areas in each of the three R. solani pathosystems: potato sprout-R. solani AG3; soybean hypocotyl-R. solani AG4 and soybean leaves-R. solani AG1-IA interactions. Levels of ROS increased in all types of potato and soybean tissues, and in fungal hyphae following infection of R. solani AGs as determined by non-fluorescence and fluorescence methods using H2DCF-DA and DAB, respectively. Overall, we demonstrate that the co-expression and accumulation of certain plant and pathogen ROS-antioxidant related genes in each pathosystem are highlighted and might be critical during disease development from the plant’s point of view, and in pathogenicity and developing of infection structures from the fungal point of view. PMID:29466404

Root defense analysis against Fusarium oxysporum reveals new regulators to confer resistance

PubMed Central

Chen, Yi Chung; Wong, Chin Lin; Muzzi, Frederico; Vlaardingerbroek, Ido; Kidd, Brendan N.; Schenk, Peer M.

2014-01-01

Fusarium oxysporum is a root-infecting fungal pathogen that causes wilt disease on a broad range of plant species, including Arabidopsis thaliana. Investigation of the defense response against this pathogen had primarily been conducted using leaf tissue and little was known about the root defense response. In this study, we profiled the expression of root genes after infection with F. oxysporum by microarray analysis. In contrast to the leaf response, root tissue did not show a strong induction of defense-associated gene expression and instead showed a greater proportion of repressed genes. Screening insertion mutants from differentially expressed genes in the microarray uncovered a role for the transcription factor ETHYLENE RESPONSE FACTOR72 (ERF72) in susceptibility to F. oxysporum. Due to the role of ERF72 in suppressing programmed cell death and detoxifying reactive oxygen species (ROS), we examined the pub22/pub23/pub24 U-box type E3 ubiquitin ligase triple mutant which is known to possess enhanced ROS production in response to pathogen challenge. We found that the pub22/23/24 mutant is more resistant to F. oxysporum infection, suggesting that a heightened innate immune response provides protection against F. oxysporum. We conclude that root-mediated defenses against soil-borne pathogens can be provided at multiple levels. PMID:24998294

RNA Editing of the GP Gene of Ebola Virus is an Important Pathogenicity Factor.

PubMed

Volchkova, Valentina A; Dolnik, Olga; Martinez, Mikel J; Reynard, Olivier; Volchkov, Viktor E

2015-10-01

Synthesis of the surface glycoprotein GP of Ebola virus (EBOV) is dependent on transcriptional RNA editing, whereas direct expression of the GP gene results in synthesis of nonstructural secreted glycoprotein sGP. In this study, we investigate the role of RNA editing in the pathogenicity of EBOV using a guinea pig model and recombinant guinea pig-adapted EBOV containing mutations at the editing site, allowing expression of surface GP without the need for RNA editing, and also preventing synthesis of sGP. We demonstrate that the elimination of the editing site leads to EBOV attenuation in vivo, explained by lower virus spread caused by the higher virus cytotoxicity and, most likely, by an increased ability of the host defense systems to recognize and eliminate virus-infected cells. We also demonstrate that expression of sGP does not affect pathogenicity of EBOV in guinea pigs. In conclusion, data obtained indicate that downregulation of the level of surface GP expression through a mechanism of GP gene RNA editing plays an important role in the high pathogenicity of EBOV. © The Author 2015. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Early host response in the mammary gland after experimental Streptococcus uberis challenge in heifers.

PubMed

de Greeff, Astrid; Zadoks, Ruth; Ruuls, Lisette; Toussaint, Mathilda; Nguyen, Thi Kim Anh; Downing, Alison; Rebel, Johanna; Stockhofe-Zurwieden, Norbert; Smith, Hilde

2013-06-01

Streptococcus uberis is a highly prevalent causative agent of bovine mastitis, which leads to large economic losses in the dairy industry. The aim of this study was to examine the host response during acute inflammation after experimental challenge with capsulated Strep. uberis. Gene expression in response to Strep. uberis was compared between infected and control quarters in 3 animals. All quarters (n=16) were sampled at 16 different locations. Microarray data showed that 239 genes were differentially expressed between infected and control quarters. No differences in gene expression were observed between the different locations. Microarray data were confirmed for several genes using quantitative PCR analysis. Genes differentially expressed due to early Strep. uberis mastitis represented several stages of the process of infection: (1) pathogen recognition; (2) chemoattraction of neutrophils; (3) tissue repair mechanisms; and (4) bactericidal activity. Three different pathogen recognition genes were induced: ficolins, lipopolysaccharide binding protein, and toll-like receptor 2. Calgranulins were found to be the most strongly upregulated genes during early inflammation. By histology and immunohistochemistry, we demonstrated that changes in gene expression in response to Strep. uberis were induced both in infiltrating somatic milk cells and in mammary epithelial cells, demonstrating that the latter cell type plays a role in milk production as well as immune responsiveness. Given the rapid development of inflammation or mastitis after infection, early diagnosis of (Strep. uberis) mastitis is required for prevention of disease and spread of the pathogen. Insight into host responses could help to design immunomodulatory therapies to dampen inflammation after (early) diagnosis of Strep. uberis mastitis. Future research should focus on development of these early diagnostics and immunomodulatory components for mastitis treatment. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Characterization of the Carbohydrate Binding Module 18 gene family in the amphibian pathogen Batrachochytrium dendrobatidis.

PubMed

Liu, Peng; Stajich, Jason E

2015-04-01

Batrachochytrium dendrobatidis (Bd) is the causative agent of chytridiomycosis responsible for worldwide decline in amphibian populations. Previous analysis of the Bd genome revealed a unique expansion of the carbohydrate-binding module family 18 (CBM18) predicted to be a sub-class of chitin recognition domains. CBM expansions have been linked to the evolution of pathogenicity in a variety of fungal species by protecting the fungus from the host. Based on phylogenetic analysis and presence of additional protein domains, the gene family can be classified into 3 classes: Tyrosinase-, Deacetylase-, and Lectin-like. Examination of the mRNA expression levels from sporangia and zoospores of nine of the cbm18 genes found that the Lectin-like genes had the highest expression while the Tyrosinase-like genes showed little expression, especially in zoospores. Heterologous expression of GFP-tagged copies of four CBM18 genes in Saccharomyces cerevisiae demonstrated that two copies containing secretion signal peptides are trafficked to the cell boundary. The Lectin-like genes cbm18-ll1 and cbm18-ll2 co-localized with the chitinous cell boundaries visualized by staining with calcofluor white. In vitro assays of the full length and single domain copies from CBM18-LL1 demonstrated chitin binding and no binding to cellulose or xylan. Expressed CBM18 domain proteins were demonstrated to protect the fungus, Trichoderma reeseii, in vitro against hydrolysis from exogenously added chitinase, likely by binding and limiting exposure of fungal chitin. These results demonstrate that cbm18 genes can play a role in fungal defense and expansion of their copy number may be an important pathogenicity factor of this emerging infectious disease of amphibians. Copyright © 2015 Elsevier Inc. All rights reserved.

Concurrent Host-Pathogen Transcriptional Responses in a Clostridium perfringens Murine Myonecrosis Infection

PubMed Central

2018-01-01

ABSTRACT To obtain an insight into host-pathogen interactions in clostridial myonecrosis, we carried out comparative transcriptome analysis of both the bacterium and the host in a murine Clostridium perfringens infection model, which is the first time that such an investigation has been conducted. Analysis of the host transcriptome from infected muscle tissues indicated that many genes were upregulated compared to the results seen with mock-infected mice. These genes were enriched for host defense pathways, including Toll-like receptor (TLR) and Nod-like receptor (NLR) signaling components. Real-time PCR confirmed that host TLR2 and NLRP3 inflammasome genes were induced in response to C. perfringens infection. Comparison of the transcriptome of C. perfringens cells from the infected tissues with that from broth cultures showed that host selective pressure induced a global change in C. perfringens gene expression. A total of 33% (923) of C. perfringens genes were differentially regulated, including 10 potential virulence genes that were upregulated relative to their expression in vitro. These genes encoded putative proteins that may be involved in the synthesis of cell wall-associated macromolecules, in adhesion to host cells, or in protection from host cationic antimicrobial peptides. This report presents the first successful expression profiling of coregulated transcriptomes of bacterial and host genes during a clostridial myonecrosis infection and provides new insights into disease pathogenesis and host-pathogen interactions. PMID:29588405

Genetic anaylsis of a disease resistance gene from loblolly pine

Treesearch

Yinghua Huang; Nili Jin; Alex Diner; Chuck Tauer; Yan Zhang; John Damicone

2003-01-01

Rapid advances in molecular genetics provide great opportunities for studies of host defense mechanisms. Examination of plant responses to disease at the cellular and molecular level permits both discovery of changes in gene expression in the tissues attacked by pathogens, and identification of genetic components involved in the interaction between host and pathogens....

Horizontal Acquisition and Transcriptional Integration of Novel Genes in Mosquito-Associated Spiroplasma.

PubMed

Lo, Wen-Sui; Kuo, Chih-Horng

2017-12-01

Genetic differentiation among symbiotic bacteria is important in shaping biodiversity. The genus Spiroplasma contains species occupying diverse niches and is a model system for symbiont evolution. Previous studies have established that two mosquito-associated species have diverged extensively in their carbohydrate metabolism genes despite having a close phylogenetic relationship. Notably, although the commensal Spiroplasma diminutum lacks identifiable pathogenicity factors, the pathogenic Spiroplasma taiwanense was found to have acquired a virulence factor glpO and its associated genes through horizontal transfer. However, it is unclear if these acquired genes have been integrated into the regulatory network. In this study, we inferred the gene content evolution in these bacteria, as well as examined their transcriptomes in response to glucose availability. The results indicated that both species have many more gene acquisitions from the Mycoides-Entomoplasmataceae clade, which contains several important pathogens of ruminants, than previously thought. Moreover, several acquired genes have higher expression levels than the vertically inherited homologs, indicating possible functional replacement. Finally, the virulence factor and its functionally linked genes in S. taiwanense were up-regulated in response to glucose starvation, suggesting that these acquired genes are under expression regulation and the pathogenicity may be a stress response. In summary, although differential gene losses are a major process for symbiont divergence, gene gains are critical in counteracting genome degradation and driving diversification among facultative symbionts. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Moraxella osloensis gene expression in the slug host Deroceras reticulatum.

PubMed

An, Ruisheng; Sreevatsan, Srinand; Grewal, Parwinder S

2008-01-28

The bacterium Moraxella osloensis is a mutualistic symbiont of the slug-parasitic nematode Phasmarhabditis hermaphrodita. In nature, P. hermaphrodita vectors M. osloensis into the shell cavity of the slug host Deroceras reticulatum in which the bacteria multiply and kill the slug. As M. osloensis is the main killing agent, genes expressed by M. osloensis in the slug are likely to play important roles in virulence. Studies on pathogenic interactions between bacteria and lower order hosts are few, but such studies have the potential to shed light on the evolution of bacterial virulence. Therefore, we investigated such an interaction by determining gene expression of M. osloensis in its slug host D. reticulatum by selectively capturing transcribed sequences. Thirteen M. osloensis genes were identified to be up-regulated post infection in D. reticulatum. Compared to the in vitro expressed genes in the stationary phase, we found that genes of ubiquinone synthetase (ubiS) and acyl-coA synthetase (acs) were up-regulated in both D. reticulatum and stationary phase in vitro cultures, but the remaining 11 genes were exclusively expressed in D. reticulatum and are hence infection specific. Mutational analysis on genes of protein-disulfide isomerase (dsbC) and ubiS showed that the virulence of both mutants to slugs was markedly reduced and could be complemented. Further, compared to the growth rate of wild-type M. osloensis, the dsbC and ubiS mutants showed normal and reduced growth rate in vitro, respectively. We conclude that 11 out of the 13 up-regulated M. osloensis genes are infection specific. Distribution of these identified genes in various bacterial pathogens indicates that the virulence genes are conserved among different pathogen-host interactions. Mutagenesis, growth rate and virulence bioassays further confirmed that ubiS and dsbC genes play important roles in M. osloensis survival and virulence, respectively in D. reticulatum.

Moraxella osloensis Gene Expression in the Slug Host Deroceras reticulatum

PubMed Central

An, Ruisheng; Sreevatsan, Srinand; Grewal, Parwinder S

2008-01-01

Background The bacterium Moraxella osloensis is a mutualistic symbiont of the slug-parasitic nematode Phasmarhabditis hermaphrodita. In nature, P. hermaphrodita vectors M. osloensis into the shell cavity of the slug host Deroceras reticulatum in which the bacteria multiply and kill the slug. As M. osloensis is the main killing agent, genes expressed by M. osloensis in the slug are likely to play important roles in virulence. Studies on pathogenic interactions between bacteria and lower order hosts are few, but such studies have the potential to shed light on the evolution of bacterial virulence. Therefore, we investigated such an interaction by determining gene expression of M. osloensis in its slug host D. reticulatum by selectively capturing transcribed sequences. Results Thirteen M. osloensis genes were identified to be up-regulated post infection in D. reticulatum. Compared to the in vitro expressed genes in the stationary phase, we found that genes of ubiquinone synthetase (ubiS) and acyl-coA synthetase (acs) were up-regulated in both D. reticulatum and stationary phase in vitro cultures, but the remaining 11 genes were exclusively expressed in D. reticulatum and are hence infection specific. Mutational analysis on genes of protein-disulfide isomerase (dsbC) and ubiS showed that the virulence of both mutants to slugs was markedly reduced and could be complemented. Further, compared to the growth rate of wild-type M. osloensis, the dsbC and ubiS mutants showed normal and reduced growth rate in vitro, respectively. Conclusion We conclude that 11 out of the 13 up-regulated M. osloensis genes are infection specific. Distribution of these identified genes in various bacterial pathogens indicates that the virulence genes are conserved among different pathogen-host interactions. Mutagenesis, growth rate and virulence bioassays further confirmed that ubiS and dsbC genes play important roles in M. osloensis survival and virulence, respectively in D. reticulatum. PMID:18226222

Genome-wide characterization of pectin methyl esterase genes reveals members differentially expressed in tolerant and susceptible wheats in response to Fusarium graminearum.

PubMed

Zega, Alessandra; D'Ovidio, Renato

2016-11-01

Pectin methyl esterase (PME) genes code for enzymes that are involved in structural modifications of the plant cell wall during plant growth and development. They are also involved in plant-pathogen interaction. PME genes belong to a multigene family and in this study we report the first comprehensive analysis of the PME gene family in bread wheat (Triticum aestivum L.). Like in other species, the members of the TaPME family are dispersed throughout the genome and their encoded products retain the typical structural features of PMEs. qRT-PCR analysis showed variation in the expression pattern of TaPME genes in different tissues and revealed that these genes are mainly expressed in flowering spikes. In our attempt to identify putative TaPME genes involved in wheat defense, we revealed a strong variation in the expression of the TaPME following Fusarium graminearum infection, the causal agent of Fusarium head blight (FHB). Particularly interesting was the finding that the expression profile of some PME genes was markedly different between the FHB-resistant wheat cultivar Sumai3 and the FHB-susceptible cultivar Bobwhite, suggesting a possible involvement of these PME genes in FHB resistance. Moreover, the expression analysis of the TaPME genes during F. graminearum progression within the spike revealed those genes that responded more promptly to pathogen invasion. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Pathogen recognition in the innate immune response.

PubMed

Kumar, Himanshu; Kawai, Taro; Akira, Shizuo

2009-04-28

Immunity against microbial pathogens primarily depends on the recognition of pathogen components by innate receptors expressed on immune and non-immune cells. Innate receptors are evolutionarily conserved germ-line-encoded proteins and include TLRs (Toll-like receptors), RLRs [RIG-I (retinoic acid-inducible gene-I)-like receptors] and NLRs (Nod-like receptors). These receptors recognize pathogens or pathogen-derived products in different cellular compartments, such as the plasma membrane, the endosomes or the cytoplasm, and induce the expression of cytokines, chemokines and co-stimulatory molecules to eliminate pathogens and instruct pathogen-specific adaptive immune responses. In the present review, we will discuss the recent progress in the study of pathogen recognition by TLRs, RLRs and NLRs and their signalling pathways.

The Genome Biology of Effector Gene Evolution in Filamentous Plant Pathogens.

PubMed

Sánchez-Vallet, Andrea; Fouché, Simone; Fudal, Isabelle; Hartmann, Fanny E; Soyer, Jessica L; Tellier, Aurélien; Croll, Daniel

2018-05-16

Filamentous pathogens, including fungi and oomycetes, pose major threats to global food security. Crop pathogens cause damage by secreting effectors that manipulate the host to the pathogen's advantage. Genes encoding such effectors are among the most rapidly evolving genes in pathogen genomes. Here, we review how the major characteristics of the emergence, function, and regulation of effector genes are tightly linked to the genomic compartments where these genes are located in pathogen genomes. The presence of repetitive elements in these compartments is associated with elevated rates of point mutations and sequence rearrangements with a major impact on effector diversification. The expression of many effectors converges on an epigenetic control mediated by the presence of repetitive elements. Population genomics analyses showed that rapidly evolving pathogens show high rates of turnover at effector loci and display a mosaic in effector presence-absence polymorphism among strains. We conclude that effective pathogen containment strategies require a thorough understanding of the effector genome biology and the pathogen's potential for rapid adaptation. Expected final online publication date for the Annual Review of Phytopathology Volume 56 is August 25, 2018. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

Pathogenic Germline Variants in 10,389 Adult Cancers.

PubMed

Huang, Kuan-Lin; Mashl, R Jay; Wu, Yige; Ritter, Deborah I; Wang, Jiayin; Oh, Clara; Paczkowska, Marta; Reynolds, Sheila; Wyczalkowski, Matthew A; Oak, Ninad; Scott, Adam D; Krassowski, Michal; Cherniack, Andrew D; Houlahan, Kathleen E; Jayasinghe, Reyka; Wang, Liang-Bo; Zhou, Daniel Cui; Liu, Di; Cao, Song; Kim, Young Won; Koire, Amanda; McMichael, Joshua F; Hucthagowder, Vishwanathan; Kim, Tae-Beom; Hahn, Abigail; Wang, Chen; McLellan, Michael D; Al-Mulla, Fahd; Johnson, Kimberly J; Lichtarge, Olivier; Boutros, Paul C; Raphael, Benjamin; Lazar, Alexander J; Zhang, Wei; Wendl, Michael C; Govindan, Ramaswamy; Jain, Sanjay; Wheeler, David; Kulkarni, Shashikant; Dipersio, John F; Reimand, Jüri; Meric-Bernstam, Funda; Chen, Ken; Shmulevich, Ilya; Plon, Sharon E; Chen, Feng; Ding, Li

2018-04-05

We conducted the largest investigation of predisposition variants in cancer to date, discovering 853 pathogenic or likely pathogenic variants in 8% of 10,389 cases from 33 cancer types. Twenty-one genes showed single or cross-cancer associations, including novel associations of SDHA in melanoma and PALB2 in stomach adenocarcinoma. The 659 predisposition variants and 18 additional large deletions in tumor suppressors, including ATM, BRCA1, and NF1, showed low gene expression and frequent (43%) loss of heterozygosity or biallelic two-hit events. We also discovered 33 such variants in oncogenes, including missenses in MET, RET, and PTPN11 associated with high gene expression. We nominated 47 additional predisposition variants from prioritized VUSs supported by multiple evidences involving case-control frequency, loss of heterozygosity, expression effect, and co-localization with mutations and modified residues. Our integrative approach links rare predisposition variants to functional consequences, informing future guidelines of variant classification and germline genetic testing in cancer. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

The platelet-activating factor acetylhydrolase gene derived from Trichoderma harzianum induces maize resistance to Curvularia lunata through the jasmonic acid signaling pathway.

PubMed

Yu, Chuanjin; Fan, Lili; Gao, Jinxin; Wang, Meng; Wu, Qiong; Tang, Jun; Li, Yaqian; Chen, Jie

2015-01-01

Platelet-activating factor acetylhydrolase (PAF-AH) derived from Trichoderma harzianum was upregulated by the interaction of T. harzianum with maize roots or the foliar pathogen Curvularia lunata. PAF-AH was associated with chitinase and cellulase expressions, but especially with chitinase, because its activity in the KO40 transformant (PAF-AH disruption transformant) was lower, compared with the wild-type strain T28. The result demonstrated that the colonization of maize roots by T. harzianum induced systemic protection of leaves inoculated with C. lunata. Such protection was associated with the expression of inducible jasmonic acid pathway-related genes. Moreover, the data from liquid chromatography-mass spectrometry confirmed that the concentration of jasmonic acid in maize leaves was associated with the expression level of defense-related genes, suggesting that PAF-AH induced resistance to the foliar pathogen. Our findings showed that PAF-AH had an important function in inducing systemic resistance to maize leaf spot pathogen.

BIOSYNTHESIS AND ACTION OF JASMONATES IN PLANTS.

PubMed

Creelman, Robert A.; Mullet, John E.

1997-06-01

Jasmonic acid and its derivatives can modulate aspects of fruit ripening, production of viable pollen, root growth, tendril coiling, and plant resistance to insects and pathogens. Jasmonate activates genes involved in pathogen and insect resistance, and genes encoding vegetative storage proteins, but represses genes encoding proteins involved in photosynthesis. Jasmonic acid is derived from linolenic acid, and most of the enzymes in the biosynthetic pathway have been extensively characterized. Modulation of lipoxygenase and allene oxide synthase gene expression in transgenic plants raises new questions about the compartmentation of the biosynthetic pathway and its regulation. The activation of jasmonic acid biosynthesis by cell wall elicitors, the peptide systemin, and other compounds will be related to the function of jasmonates in plants. Jasmonate modulates gene expression at the level of translation, RNA processing, and transcription. Promoter elements that mediate responses to jasmonate have been isolated. This review covers recent advances in our understanding of how jasmonate biosynthesis is regulated and relates this information to knowledge of jasmonate modulated gene expression.

Lynch syndrome associated with two MLH1 promoter variants and allelic imbalance of MLH1 expression.

PubMed

Hesson, Luke B; Packham, Deborah; Kwok, Chau-To; Nunez, Andrea C; Ng, Benedict; Schmidt, Christa; Fields, Michael; Wong, Jason W H; Sloane, Mathew A; Ward, Robyn L

2015-06-01

Lynch syndrome is a hereditary cancer syndrome caused by a constitutional mutation in one of the mismatch repair genes. The implementation of predictive testing and targeted preventative surveillance is hindered by the frequent finding of sequence variants of uncertain significance in these genes. We aimed to determine the pathogenicity of previously reported variants (c.-28A>G and c.-7C>T) within the MLH1 5'untranslated region (UTR) in two individuals from unrelated suspected Lynch syndrome families. We investigated whether these variants were associated with other pathogenic alterations using targeted high-throughput sequencing of the MLH1 locus. We also determined their relationship to gene expression and epigenetic alterations at the promoter. Sequencing revealed that the c.-28A>G and c.-7C>T variants were the only potentially pathogenic alterations within the MLH1 gene. In both individuals, the levels of transcription from the variant allele were reduced to 50% compared with the wild-type allele. Partial loss of expression occurred in the absence of constitutional epigenetic alterations within the MLH1 promoter. We propose that these variants may be pathogenic due to constitutional partial loss of MLH1 expression, and that this may be associated with intermediate penetrance of a Lynch syndrome phenotype. Our findings provide further evidence of the potential importance of noncoding variants in the MLH1 5'UTR in the pathogenesis of Lynch syndrome. © 2015 The Authors. **Human Mutation published by Wiley Periodicals, Inc.

Lynch Syndrome Associated with Two MLH1 Promoter Variants and Allelic Imbalance of MLH1 Expression

PubMed Central

Hesson, Luke B; Packham, Deborah; Kwok, Chau-To; Nunez, Andrea C; Ng, Benedict; Schmidt, Christa; Fields, Michael; Wong, Jason WH; Sloane, Mathew A; Ward, Robyn L

2015-01-01

Lynch syndrome is a hereditary cancer syndrome caused by a constitutional mutation in one of the mismatch repair genes. The implementation of predictive testing and targeted preventative surveillance is hindered by the frequent finding of sequence variants of uncertain significance in these genes. We aimed to determine the pathogenicity of previously reported variants (c.-28A>G and c.-7C>T) within the MLH1 5′untranslated region (UTR) in two individuals from unrelated suspected Lynch syndrome families. We investigated whether these variants were associated with other pathogenic alterations using targeted high-throughput sequencing of the MLH1 locus. We also determined their relationship to gene expression and epigenetic alterations at the promoter. Sequencing revealed that the c.-28A>G and c.-7C>T variants were the only potentially pathogenic alterations within the MLH1 gene. In both individuals, the levels of transcription from the variant allele were reduced to 50% compared with the wild-type allele. Partial loss of expression occurred in the absence of constitutional epigenetic alterations within the MLH1 promoter. We propose that these variants may be pathogenic due to constitutional partial loss of MLH1 expression, and that this may be associated with intermediate penetrance of a Lynch syndrome phenotype. Our findings provide further evidence of the potential importance of noncoding variants in the MLH1 5′UTR in the pathogenesis of Lynch syndrome. PMID:25762362

Pseudogymnoascus destructans transcriptome changes during white-nose syndrome infections

PubMed Central

Reeder, Sophia M.; Palmer, Jonathan M.; Prokkola, Jenni M.; Lilley, Thomas M.; Reeder, DeeAnn M.

2017-01-01

ABSTRACT White nose syndrome (WNS) is caused by the psychrophilic fungus Pseudogymnoascus destructans that can grow in the environment saprotrophically or parasitically by infecting hibernating bats. Infections are pathological in many species of North American bats, disrupting hibernation and causing mortality. To determine what fungal pathways are involved in infection of living tissue, we examined fungal gene expression using RNA-Seq. We compared P. destructans gene expression when grown in culture to that during infection of a North American bat species, Myotis lucifugus, that shows high WNS mortality. Cultured P. destructans was grown at 10 to 14 C and P. destructans growing in vivo was presumably exposed to temperatures ranging from 4 to 8 C during torpor and up to 37 C during periodic arousals. We found that when P. destructans is causing WNS, the most significant differentially expressed genes were involved in heat shock responses, cell wall remodeling, and micronutrient acquisition. These results indicate that this fungal pathogen responds to host-pathogen interactions by regulating gene expression in ways that may contribute to evasion of host responses. Alterations in fungal cell wall structures could allow P. destructans to avoid detection by host pattern recognition receptors and antibody responses. This study has also identified several fungal pathways upregulated during WNS infection that may be candidates for mitigating infection pathology. By identifying host-specific pathogen responses, these observations have important implications for host-pathogen evolutionary relationships in WNS and other fungal diseases. PMID:28614673

Pseudogymnoascus destructans transcriptome changes during white-nose syndrome infections.

PubMed

Reeder, Sophia M; Palmer, Jonathan M; Prokkola, Jenni M; Lilley, Thomas M; Reeder, DeeAnn M; Field, Kenneth A

2017-11-17

White nose syndrome (WNS) is caused by the psychrophilic fungus Pseudogymnoascus destructans that can grow in the environment saprotrophically or parasitically by infecting hibernating bats. Infections are pathological in many species of North American bats, disrupting hibernation and causing mortality. To determine what fungal pathways are involved in infection of living tissue, we examined fungal gene expression using RNA-Seq. We compared P. destructans gene expression when grown in culture to that during infection of a North American bat species, Myotis lucifugus, that shows high WNS mortality. Cultured P. destructans was grown at 10 to 14 C and P. destructans growing in vivo was presumably exposed to temperatures ranging from 4 to 8 C during torpor and up to 37 C during periodic arousals. We found that when P. destructans is causing WNS, the most significant differentially expressed genes were involved in heat shock responses, cell wall remodeling, and micronutrient acquisition. These results indicate that this fungal pathogen responds to host-pathogen interactions by regulating gene expression in ways that may contribute to evasion of host responses. Alterations in fungal cell wall structures could allow P. destructans to avoid detection by host pattern recognition receptors and antibody responses. This study has also identified several fungal pathways upregulated during WNS infection that may be candidates for mitigating infection pathology. By identifying host-specific pathogen responses, these observations have important implications for host-pathogen evolutionary relationships in WNS and other fungal diseases.

Mechanisms of pathogenesis of emerging adenoviruses.

PubMed

Cook, James; Radke, Jay

2017-01-01

Periodic outbreaks of human adenovirus infections can cause severe illness in people with no known predisposing conditions. The reasons for this increased viral pathogenicity are uncertain. Adenoviruses are constantly undergoing mutation during circulation in the human population, but related phenotypic changes of the viruses are rarely detected because of the infrequency of such outbreaks and the limited biological studies of the emergent strains. Mutations and genetic recombinations have been identified in these new strains. However, the linkage between these genetic changes and increased pathogenicity is poorly understood. It has been observed recently that differences in virus-induced immunopathogenesis can be associated with altered expression of non-mutant viral genes associated with changes in viral modulation of the host innate immune response. Initial small animal studies indicate that these changes in viral gene expression can be associated with enhanced immunopathogenesis in vivo . Available evidence suggests the hypothesis that there is a critical threshold of expression of certain viral genes that determines both the sustainability of viral transmission in the human population and the enhancement of immunopathogenesis. Studies of this possibility will require extension of the analysis of outbreak viral strains from a sequencing-based focus to biological studies of relationships between viral gene expression and pathogenic responses. Advances in this area will require increased coordination among public health organizations, diagnostic microbiology laboratories, and research laboratories to identify, catalog, and systematically study differences between prototype and emergent viral strains that explain the increased pathogenicity that can occur during clinical outbreaks.

Dynamic changes in host gene expression associated with H5N8 avian influenza virus infection in mice.

PubMed

Park, Su-Jin; Kumar, Mukesh; Kwon, Hyeok-il; Seong, Rak-Kyun; Han, Kyudong; Song, Jae-min; Kim, Chul-Joong; Choi, Young-Ki; Shin, Ok Sarah

2015-11-18

Emerging outbreaks of newly found, highly pathogenic avian influenza (HPAI) A(H5N8) viruses have been reported globally. Previous studies have indicated that H5N8 pathogenicity in mice is relatively moderate compared with H5N1 pathogenicity. However, detailed mechanisms underlying avian influenza pathogenicity are still undetermined. We used a high-throughput RNA-seq method to analyse host and pathogen transcriptomes in the lungs of mice infected with A/MD/Korea/W452/2014 (H5N8) and A/EM/Korea/W149/2006 (H5N1) viruses. Sequenced numbers of viral transcripts and expression levels of host immune-related genes at 1 day post infection (dpi) were higher in H5N8-infected than H5N1-infected mice. Dual sequencing of viral transcripts revealed that in contrast to the observations at 1 dpi, higher number of H5N1 genes than H5N8 genes was sequenced at 3 and 7 dpi, which is consistent with higher viral titres and virulence observed in infected lungs in vivo. Ingenuity pathway analysis revealed a more significant upregulation of death receptor signalling, driven by H5N1 than with H5N8 infection at 3 and 7 dpi. Early induction of immune response-related genes may elicit protection in H5N8-infected mice, which correlates with moderate pathogenicity in vivo. Collectively, our data provide new insight into the underlying mechanisms of the differential pathogenicity of avian influenza viruses.

Dynamic changes in host gene expression associated with H5N8 avian influenza virus infection in mice

PubMed Central

Park, Su-Jin; Kumar, Mukesh; Kwon, Hyeok-il; Seong, Rak-Kyun; Han, Kyudong; Song, Jae-min; Kim, Chul-Joong; Choi, Young-Ki; Shin, Ok Sarah

2015-01-01

Emerging outbreaks of newly found, highly pathogenic avian influenza (HPAI) A(H5N8) viruses have been reported globally. Previous studies have indicated that H5N8 pathogenicity in mice is relatively moderate compared with H5N1 pathogenicity. However, detailed mechanisms underlying avian influenza pathogenicity are still undetermined. We used a high-throughput RNA-seq method to analyse host and pathogen transcriptomes in the lungs of mice infected with A/MD/Korea/W452/2014 (H5N8) and A/EM/Korea/W149/2006 (H5N1) viruses. Sequenced numbers of viral transcripts and expression levels of host immune-related genes at 1 day post infection (dpi) were higher in H5N8-infected than H5N1-infected mice. Dual sequencing of viral transcripts revealed that in contrast to the observations at 1 dpi, higher number of H5N1 genes than H5N8 genes was sequenced at 3 and 7 dpi, which is consistent with higher viral titres and virulence observed in infected lungs in vivo. Ingenuity pathway analysis revealed a more significant upregulation of death receptor signalling, driven by H5N1 than with H5N8 infection at 3 and 7 dpi. Early induction of immune response-related genes may elicit protection in H5N8-infected mice, which correlates with moderate pathogenicity in vivo. Collectively, our data provide new insight into the underlying mechanisms of the differential pathogenicity of avian influenza viruses. PMID:26576844

The expression of propionicin PLG-1 gene (plg-1) by lactic starters.

PubMed

Mohamed, Sameh E; Tahoun, Mahmoud K

2015-05-01

Propionicin PLG-1 is a bacteriocin produced by Propionibacterium thoenii P127. Such bacteriocin inhibits wide range of food-borne pathogens such as pathogenic Escherichia coli, Pseudomonas aeruginosa, Vibrio parahaemolyticus, Yersinia enterocolitica and a strain of Corynebacterium sp. In the present study, plg-1 gene expressing propionicin PLG-1 was isolated, sequenced for the first time and the resulting sequence was analysed using several web-based bioinformatics programs. The PCR product containing plg-1 gene was transferred to different lactic acid bacterial (LAB) strains using pLEB590 as a cloning vector to give the modified vector pLEBPLG-1. LAB transformants showed an antimicrobial activity against Esch. coli DH5α (most affected strain), Listeria monocytogenes 18116, and Salmonella enterica 25566 as model pathogenic strains. Such LAB transformants can be used in dairy industry to control the food-borne pathogens that are largely distributed worldwide and to feed schoolchildren in the poor countries where dangerous epidemic diseases and diarrhoea prevail.

From genes to genomes: a new paradigm for studying fungal pathogenesis in Magnaporthe oryzae.

PubMed

Xu, Jin-Rong; Zhao, Xinhua; Dean, Ralph A

2007-01-01

Magnaporthe oryzae is the most destructive fungal pathogen of rice worldwide and because of its amenability to classical and molecular genetic manipulation, availability of a genome sequence, and other resources it has emerged as a leading model system to study host-pathogen interactions. This chapter reviews recent progress toward elucidation of the molecular basis of infection-related morphogenesis, host penetration, invasive growth, and host-pathogen interactions. Related information on genome analysis and genomic studies of plant infection processes is summarized under specific topics where appropriate. Particular emphasis is placed on the role of MAP kinase and cAMP signal transduction pathways and unique features in the genome such as repetitive sequences and expanded gene families. Emerging developments in functional genome analysis through large-scale insertional mutagenesis and gene expression profiling are detailed. The chapter concludes with new prospects in the area of systems biology, such as protein expression profiling, and highlighting remaining crucial information needed to fully appreciate host-pathogen interactions.

Arabidopsis serotonin N-acetyltransferase knockout mutant plants exhibit decreased melatonin and salicylic acid levels resulting in susceptibility to an avirulent pathogen.

PubMed

Lee, Hyoung Yool; Byeon, Yeong; Tan, Dun-Xian; Reiter, Russel J; Back, Kyoungwhan

2015-04-01

Serotonin N-acetyltransferase (SNAT) is the penultimate enzyme in the melatonin biosynthesis pathway in plants. We examined the effects of SNAT gene inactivation in two Arabidopsis T-DNA insertion mutant lines. After inoculation with the avirulent pathogen Pseudomonas syringe pv. tomato DC3000 harboring the elicitor avrRpt2 (Pst-avrRpt2), melatonin levels in the snat knockout mutant lines were 50% less than in wild-type Arabidopsis Col-0 plants. The snat knockout mutant lines exhibited susceptibility to pathogen infection that coincided with decreased induction of defense genes including PR1, ICS1, and PDF1.2. Because melatonin acts upstream of salicylic acid (SA) synthesis, the reduced melatonin levels in the snat mutant lines led to decreased SA levels compared to wild-type, suggesting that the increased pathogen susceptibility of the snat mutant lines could be attributed to decreased SA levels and subsequent attenuation of defense gene induction. Exogenous melatonin treatment failed to induce defense gene expression in nahG Arabidopsis plants, but restored the induction of defense gene expression in the snat mutant lines. In addition, melatonin caused translocation of NPR1 (nonexpressor of PR1) protein from the cytoplasm into the nucleus indicating that melatonin-elicited pathogen resistance in response to avirulent pathogen attack is SA-dependent in Arabidopsis. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Interchromosomal Transfer of Immune Regulation During Infection of Barley with the Powdery Mildew Pathogen

PubMed Central

Surana, Priyanka; Xu, Ruo; Fuerst, Gregory; Chapman, Antony V. E.; Nettleton, Dan; Wise, Roger P.

2017-01-01

Powdery mildew pathogens colonize over 9500 plant species, causing critical yield loss. The Ascomycete fungus, Blumeria graminis f. sp. hordei (Bgh), causes powdery mildew disease in barley (Hordeum vulgare L.). Successful infection begins with penetration of host epidermal cells, culminating in haustorial feeding structures, facilitating delivery of fungal effectors to the plant and exchange of nutrients from host to pathogen. We used expression Quantitative Trait Locus (eQTL) analysis to dissect the temporal control of immunity-associated gene expression in a doubled haploid barley population challenged with Bgh. Two highly significant regions possessing trans eQTL were identified near the telomeric ends of chromosomes (Chr) 2HL and 1HS. Within these regions reside diverse resistance loci derived from barley landrace H. laevigatum (MlLa) and H. vulgare cv. Algerian (Mla1), which associate with the altered expression of 961 and 3296 genes during fungal penetration of the host and haustorial development, respectively. Regulatory control of transcript levels for 299 of the 961 genes is reprioritized from MlLa on 2HL to Mla1 on 1HS as infection progresses, with 292 of the 299 alternating the allele responsible for higher expression, including Adaptin Protein-2 subunit μ AP2M and Vesicle Associated Membrane Protein VAMP72 subfamily members VAMP721/722. AP2M mediates effector-triggered immunity (ETI) via endocytosis of plasma membrane receptor components. VAMP721/722 and SNAP33 form a Soluble N-ethylmaleimide-sensitive factor Attachment Protein REceptor (SNARE) complex with SYP121 (PEN1), which is engaged in pathogen associated molecular pattern (PAMP)-triggered immunity via exocytosis. We postulate that genes regulated by alternate chromosomal positions are repurposed as part of a conserved immune complex to respond to different pathogen attack scenarios. PMID:28790145

Overwintering Is Associated with Reduced Expression of Immune Genes and Higher Susceptibility to Virus Infection in Honey Bees

PubMed Central

Steinmann, Nadja; Corona, Miguel; Neumann, Peter; Dainat, Benjamin

2015-01-01

The eusocial honey bee, Apis mellifera, has evolved remarkable abilities to survive extreme seasonal differences in temperature and availability of resources by dividing the worker caste into two groups that differ in physiology and lifespan: summer and winter bees. Most of the recent major losses of managed honey bee colonies occur during the winter, suggesting that winter bees may have compromised immune function and higher susceptibility to diseases. We tested this hypothesis by comparing the expression of eight immune genes and naturally occurring infection levels of deformed wing virus (DWV), one of the most widespread viruses in A. mellifera populations, between summer and winter bees. Possible interactions between immune response and physiological activity were tested by measuring the expression of vitellogenin and methyl farnesoate epoxidase, a gene coding for the last enzyme involved in juvenile hormone biosynthesis. Our data show that high DWV loads in winter bees correlate with reduced expression of genes involved in the cellular immune response and physiological activity and high expression of humoral immune genes involved in antibacterial defense compared with summer bees. This expression pattern could reflect evolutionary adaptations to resist bacterial pathogens and economize energy during the winter under a pathogen landscape with reduced risk of pathogenic viral infections. The outbreak of Varroa destructor infestation could have overcome these adaptations by promoting the transmission of viruses. Our results suggest that reduced cellular immune function during the winter may have increased honey bee’s susceptibility to DWV. These results contribute to our understanding of honey bee colony losses in temperate regions. PMID:26121358

Expression of H5 hemagglutinin vaccine antigen in common duckweed (Lemna minor) protects against H5N1 high pathogenicity avian influenza virus challenge in immunized chickens

USDA-ARS?s Scientific Manuscript database

A synthetic hemagglutinin (HA) gene from the highly pathogenic avian influenza (HPAI) virus A/chicken/Indonesia/7/2003 (H5N1) (Indo/03) was expressed in aquatic plant Lemna minor (rLemna-HA). In Experiment 1, efficacy of rLemna-HA was tested on specific pathogen free (SPF) birds immunized with 0.2 ...

Avian Paramyxovirus Type-3 as a Vaccine Vector: Identification of a Genome Location for High Level Expression of a Foreign Gene

PubMed Central

Yoshida, Asuka; Samal, Siba K.

2017-01-01

Avian paramyxovirus serotype 3 (APMV-3) causes infection in a wide variety of avian species, but it does not cause apparent diseases in chickens. On the contrary, APMV-1, also known as Newcastle disease virus (NDV), can cause severe disease in chickens. Currently, natural low virulence strains of NDV are used as live-attenuated vaccines throughout the world. NDV is also being evaluated as a vaccine vector against poultry pathogens. However, due to routine vaccination programs, chickens often possess pre-existing antibodies against NDV, which may cause the chickens to be less sensitive to recombinant NDV vaccines expressing antigens of other avian pathogens. Therefore, it may be possible for an APMV-3 vector vaccine to circumvent this issue. In this study, we determined the optimal insertion site in the genome of APMV-3 for high level expression of a foreign gene. We generated recombinant APMV-3 viruses expressing the green fluorescent protein (GFP) by inserting the GFP gene at five different intergenic regions in the genome. The levels of GFP transcription and translation were evaluated. Interestingly, the levels of GFP transcription and translation did not follow the 3′-to-5′ attenuation mechanism of non-segmented, negative-sense RNA viruses. The insertion of GFP gene into the P-M gene junction resulted in higher level of expression of GFP than when the gene was inserted into the upstream N-P gene junction. Unlike NDV, insertion of GFP did not attenuate the growth efficiency of AMPV-3. Thus, APMV-3 could be a more useful vaccine vector for avian pathogens than NDV. PMID:28473820

Avian Paramyxovirus Type-3 as a Vaccine Vector: Identification of a Genome Location for High Level Expression of a Foreign Gene.

PubMed

Yoshida, Asuka; Samal, Siba K

2017-01-01

Avian paramyxovirus serotype 3 (APMV-3) causes infection in a wide variety of avian species, but it does not cause apparent diseases in chickens. On the contrary, APMV-1, also known as Newcastle disease virus (NDV), can cause severe disease in chickens. Currently, natural low virulence strains of NDV are used as live-attenuated vaccines throughout the world. NDV is also being evaluated as a vaccine vector against poultry pathogens. However, due to routine vaccination programs, chickens often possess pre-existing antibodies against NDV, which may cause the chickens to be less sensitive to recombinant NDV vaccines expressing antigens of other avian pathogens. Therefore, it may be possible for an APMV-3 vector vaccine to circumvent this issue. In this study, we determined the optimal insertion site in the genome of APMV-3 for high level expression of a foreign gene. We generated recombinant APMV-3 viruses expressing the green fluorescent protein (GFP) by inserting the GFP gene at five different intergenic regions in the genome. The levels of GFP transcription and translation were evaluated. Interestingly, the levels of GFP transcription and translation did not follow the 3'-to-5' attenuation mechanism of non-segmented, negative-sense RNA viruses. The insertion of GFP gene into the P-M gene junction resulted in higher level of expression of GFP than when the gene was inserted into the upstream N-P gene junction. Unlike NDV, insertion of GFP did not attenuate the growth efficiency of AMPV-3. Thus, APMV-3 could be a more useful vaccine vector for avian pathogens than NDV.

Pathogenic effects of Rift Valley fever virus NSs gene are alleviated in cultured cells by expressed antiviral short hairpin RNAs.

PubMed

Scott, Tristan; Paweska, Janusz T; Arbuthnot, Patrick; Weinberg, Marc S

2012-01-01

Rift Valley fever virus (RVFV), a member of the Bunyaviridae family, may cause severe hepatitis, encephalitis and haemorrhagic fever in humans. There are currently no available licensed vaccines or therapies to treat the viral infection in humans. RNA interference (RNAi)-based viral gene silencing offers a promising approach to inhibiting replication of this highly pathogenic virus. The small (S) segment of the RVFV tripartite genome carries the genetic determinates for pathogenicity during infection. This segment encodes the non-structural S (NSs) and essential nucleocapsid (N) genes. To advance RNAi-based inhibition of RVFV replication, we designed several Pol III short hairpin RNA (shRNA) expression cassettes against the NSs and N genes, including a multimerized plasmid vector that included four shRNA expression cassettes. Effective target silencing was demonstrated using full- and partial-length target reporter assays, and confirmed by western blot analysis of exogenous N and NSs expression. Small RNA northern blots showed detectable RNAi guide strand formation from single and multimerized shRNA constructs. Using a cell culture model of RVFV replication, shRNAs targeting the N gene decreased intracellular nucleocapsid protein concentration and viral replication. The shRNAs directed against the NSs gene reduced NSs protein concentrations and alleviated NSs-mediated cytotoxicity, which may be caused by host transcription suppression. These data are the first demonstration that RNAi activators have a potential therapeutic benefit for countering RVFV infection.

Gene expression profiling during asexual development of the late blight pathogen Phytophthora infestans reveals a highly dynamic transcriptome.

PubMed

Judelson, Howard S; Ah-Fong, Audrey M V; Aux, George; Avrova, Anna O; Bruce, Catherine; Cakir, Cahid; da Cunha, Luis; Grenville-Briggs, Laura; Latijnhouwers, Maita; Ligterink, Wilco; Meijer, Harold J G; Roberts, Samuel; Thurber, Carrie S; Whisson, Stephen C; Birch, Paul R J; Govers, Francine; Kamoun, Sophien; van West, Pieter; Windass, John

2008-04-01

Much of the pathogenic success of Phytophthora infestans, the potato and tomato late blight agent, relies on its ability to generate from mycelia large amounts of sporangia, which release zoospores that encyst and form infection structures. To better understand these stages, Affymetrix GeneChips based on 15,650 unigenes were designed and used to profile the life cycle. Approximately half of P. infestans genes were found to exhibit significant differential expression between developmental transitions, with approximately (1)/(10) being stage-specific and most changes occurring during zoosporogenesis. Quantitative reverse-transcription polymerase chain reaction assays confirmed the robustness of the array results and showed that similar patterns of differential expression were obtained regardless of whether hyphae were from laboratory media or infected tomato. Differentially expressed genes encode potential cellular regulators, especially protein kinases; metabolic enzymes such as those involved in glycolysis, gluconeogenesis, or the biosynthesis of amino acids or lipids; regulators of DNA synthesis; structural proteins, including predicted flagellar proteins; and pathogenicity factors, including cell-wall-degrading enzymes, RXLR effector proteins, and enzymes protecting against plant defense responses. Curiously, some stage-specific transcripts do not appear to encode functional proteins. These findings reveal many new aspects of oomycete biology, as well as potential targets for crop protection chemicals.

Differential role of gpaB and sidA gene expressions in relation to virulence in Aspergillus species from patients with invasive aspergillosis.

PubMed

Ghods, Nayereh; Falahati, Mehraban; Roudbary, Maryam; Farahyar, Shirin; Shamaei, Masoud; Pourabdollah, Mahin; Seif, Farhad

2018-02-03

The virulence genes in invasive aspergillosis (IA) have not been analyzed adequately. The present study was designed to evaluate the expression of gpaB and sidA genes, which are important virulence genes in Aspergillus spp. from bronchoalveolar lavage (BAL) samples. Direct examination and culture on Czapek Agar and Sabouraud Dextrose Agar media were performed for 600 BAL specimens isolated from patients with possible aspergillosis. A Galactomannan ELISA assay was also carried out. The expression levels of the gpaB and sidA genes in isolates were analyzed using quantitative real-time PCR (qRT-PCR). We identified 2 species, including Aspergillus flavus (A. flavus) and Aspergillus fumigatus (A. fumigatus) in 25 positive samples for invasive aspergillosis as validated using GM-ELISA. A. flavus is the main pathogen threatening transplant recipients and cancer patients worldwide. In this study, A. flavus had low levels of the gpaB gene expression compared to A. fumigatus (p=0.006). The highest sidA expression was detected in transplant recipients (p=0.05). There was no significant correlation between sidA expression and underlying disease (p=0.15). The sidA and gpaB gene expression patterns may provide evidence that these virulence genes play important roles in the pathogenicity of Aspergillus isolates; however, there are several regulatory genes responsible for the unexpressed sidA and gpaB genes in the isolates. Copyright © 2018 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

Small-molecule inhibitors suppress the expression of both type III secretion and amylovoran biosynthesis genes in Erwinia amylovora.

PubMed

Yang, Fan; Korban, Schuyler S; Pusey, P Lawrence; Elofsson, Michael; Sundin, George W; Zhao, Youfu

2014-01-01

The type III secretion system (T3SS) and exopolysaccharide (EPS) amylovoran are two essential pathogenicity factors in Erwinia amylovora, the causal agent of the serious bacterial disease fire blight. In this study, small molecules that inhibit T3SS gene expression in E. amylovora under hrp (hypersensitive response and pathogenicity)-inducing conditions were identified and characterized using green fluorescent protein (GFP) as a reporter. These compounds belong to salicylidene acylhydrazides and also inhibit amylovoran production. Microarray analysis of E. amylovora treated with compounds 3 and 9 identified a total of 588 significantly differentially expressed genes. Among them, 95 and 78 genes were activated and suppressed by both compounds, respectively, when compared with the dimethylsulphoxide (DMSO) control. The expression of the majority of T3SS genes in E. amylovora, including hrpL and the avrRpt2 effector gene, was suppressed by both compounds. Compound 3 also suppressed the expression of amylovoran precursor and biosynthesis genes. However, both compounds induced significantly the expression of glycogen biosynthesis genes and siderophore biosynthesis, regulatory and transport genes. Furthermore, many membrane, lipoprotein and exported protein-encoding genes were also activated by both compounds. Similar expression patterns were observed for compounds 1, 2 and 4. Using crab apple flower as a model, compound 3 was capable of reducing disease development in pistils. These results suggest a common inhibition mechanism shared by salicylidene acylhydrazides and indicate that small-molecule inhibitors that disable T3SS function could be explored to control fire blight disease. © 2013 BSPP AND JOHN WILEY & SONS LTD.

Host-Induced Gene Silencing of Rice Blast Fungus Magnaporthe oryzae Pathogenicity Genes Mediated by the Brome Mosaic Virus.

PubMed

Zhu, Lin; Zhu, Jian; Liu, Zhixue; Wang, Zhengyi; Zhou, Cheng; Wang, Hong

2017-09-26

Magnaporthe oryzae is a devastating plant pathogen, which has a detrimental impact on rice production worldwide. Despite its agronomical importance, some newly-emerging pathotypes often overcome race-specific disease resistance rapidly. It is thus desirable to develop a novel strategy for the long-lasting resistance of rice plants to ever-changing fungal pathogens. Brome mosaic virus (BMV)-induced RNA interference (RNAi) has emerged as a useful tool to study host-resistance genes for rice blast protection. Planta-generated silencing of targeted genes inside biotrophic pathogens can be achieved by expression of M. oryzae -derived gene fragments in the BMV-mediated gene silencing system, a technique termed host-induced gene silencing (HIGS). In this study, the effectiveness of BMV-mediated HIGS in M. oryzae was examined by targeting three predicted pathogenicity genes, MoABC1, MoMAC1 and MoPMK1 . Systemic generation of fungal gene-specific small interfering RNA (siRNA) molecules induced by inoculation of BMV viral vectors inhibited disease development and reduced the transcription of targeted fungal genes after subsequent M. oryzae inoculation. Combined introduction of fungal gene sequences in sense and antisense orientation mediated by the BMV silencing vectors significantly enhanced the efficiency of this host-generated trans-specific RNAi, implying that these fungal genes played crucial roles in pathogenicity. Collectively, our results indicated that BMV-HIGS system was a great strategy for protecting host plants against the invasion of pathogenic fungi.

The wheat Lr34 multipathogen resistance gene confers resistance to anthracnose and rust in sorghum.

PubMed

Schnippenkoetter, Wendelin; Lo, Clive; Liu, Guoquan; Dibley, Katherine; Chan, Wai Lung; White, Jodie; Milne, Ricky; Zwart, Alexander; Kwong, Eunjung; Keller, Beat; Godwin, Ian; Krattinger, Simon G; Lagudah, Evans

2017-11-01

The ability of the wheat Lr34 multipathogen resistance gene (Lr34res) to function across a wide taxonomic boundary was investigated in transgenic Sorghum bicolor. Increased resistance to sorghum rust and anthracnose disease symptoms following infection with the biotrophic pathogen Puccinia purpurea and the hemibiotroph Colletotrichum sublineolum, respectively, occurred in transgenic plants expressing the Lr34res ABC transporter. Transgenic sorghum lines that highly expressed the wheat Lr34res gene exhibited immunity to sorghum rust compared to the low-expressing single copy Lr34res genotype that conferred partial resistance. Pathogen-induced pigmentation mediated by flavonoid phytoalexins was evident on transgenic sorghum leaves following P. purpurea infection within 24-72 h, which paralleled Lr34res gene expression. Elevated expression of flavone synthase II, flavanone 4-reductase and dihydroflavonol reductase genes which control the biosynthesis of flavonoid phytoalexins characterized the highly expressing Lr34res transgenic lines 24-h post-inoculation with P. purpurea. Metabolite analysis of mesocotyls infected with C. sublineolum showed increased levels of 3-deoxyanthocyanidin metabolites were associated with Lr34res expression, concomitant with reduced symptoms of anthracnose. © 2017 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Comprehensive analysis of microRNA-Seq and target mRNAs of rice sheath blight pathogen provides new insights into pathogenic regulatory mechanisms.

PubMed

Lin, Runmao; He, Liye; He, Jiayu; Qin, Peigang; Wang, Yanran; Deng, Qiming; Yang, Xiaoting; Li, Shuangcheng; Wang, Shiquan; Wang, Wenming; Liu, Huainian; Li, Ping; Zheng, Aiping

2016-07-03

MicroRNAs (miRNAs) are ∼22 nucleotide non-coding RNAs that regulate gene expression by targeting mRNAs for degradation or inhibiting protein translation. To investigate whether miRNAs regulate the pathogenesis in necrotrophic fungus Rhizoctonia solani AG1 IA, which causes significant yield loss in main economically important crops, and to determine the regulatory mechanism occurring during pathogenesis, we constructed hyphal small RNA libraries from six different infection periods of the rice leaf. Through sequencing and analysis, 177 miRNA-like small RNAs (milRNAs) were identified, including 15 candidate pathogenic novel milRNAs predicted by functional annotations of their target mRNAs and expression patterns of milRNAs and mRNAs during infection. Reverse transcription-quantitative polymerase chain reaction results for randomly selected milRNAs demonstrated that our novel comprehensive predictions had a high level of accuracy. In our predicted pathogenic protein-protein interaction network of R. solani, we added the related regulatory milRNAs of these core coding genes into the network, and could understand the relationships among these regulatory factors more clearly at the systems level. Furthermore, the putative pathogenic Rhi-milR-16, which negatively regulates target gene expression, was experimentally validated to have regulatory functions by a dual-luciferase reporter assay. Additionally, 23 candidate rice miRNAs that may involve in plant immunity against R. solani were discovered. This first study on novel pathogenic milRNAs of R. solani AG1 IA and the recognition of target genes involved in pathogenicity, as well as rice miRNAs, participated in defence against R. solani could provide new insights into revealing the pathogenic mechanisms of the severe rice sheath blight disease. © The Author 2016. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

Investigation of Reference Genes in Vibrio parahaemolyticus for Gene Expression Analysis Using Quantitative RT-PCR.

PubMed

Ma, Yue-Jiao; Sun, Xiao-Hong; Xu, Xiao-Yan; Zhao, Yong; Pan, Ying-Jie; Hwang, Cheng-An; Wu, Vivian C H

2015-01-01

Vibrio parahaemolyticus is a significant human pathogen capable of causing foodborne gastroenteritis associated with the consumption of contaminated raw or undercooked seafood. Quantitative RT-PCR (qRT-PCR) is a useful tool for studying gene expression in V. parahaemolyticus to characterize its virulence factors and understand the effect of environmental conditions on its pathogenicity. However, there is not a stable gene in V. parahaemolyticus that has been identified for use as a reference gene for qRT-PCR. This study evaluated the stability of 6 reference genes (16S rRNA, recA, rpoS, pvsA, pvuA, and gapdh) in 5 V. parahaemolyticus strains (O3:K6-clinical strain-tdh+, ATCC33846-tdh+, ATCC33847-tdh+, ATCC17802-trh+, and F13-environmental strain-tdh+) cultured at 4 different temperatures (15, 25, 37 and 42°C). Stability values were calculated using GeNorm, NormFinder, BestKeeper, and Delta CT algorithms. The results indicated that recA was the most stably expressed gene in the V. parahaemolyticus strains cultured at different temperatures. This study examined multiple V. parahaemolyticus strains and growth temperatures, hence the finding provided stronger evidence that recA can be used as a reference gene for gene expression studies in V. parahaemolyticus.

Genome-wide characterization and expression profiling of immune genes in the diamondback moth, Plutella xylostella (L.)

PubMed Central

Xia, Xiaofeng; Yu, Liying; Xue, Minqian; Yu, Xiaoqiang; Vasseur, Liette; Gurr, Geoff M.; Baxter, Simon W.; Lin, Hailan; Lin, Junhan; You, Minsheng

2015-01-01

The diamondback moth, Plutella xylostella (L.), is a destructive pest that attacks cruciferous crops worldwide. Immune responses are important for interactions between insects and pathogens and information on these underpins the development of strategies for biocontrol-based pest management. Little, however, is known about immune genes and their regulation patterns in P. xylostella. A total of 149 immune-related genes in 20 gene families were identified through comparison of P. xylostella genome with the genomes of other insects. Complete and conserved Toll, IMD and JAK-STAT signaling pathways were found in P. xylostella. Genes involved in pathogen recognition were expanded and more diversified than genes associated with intracellular signal transduction. Gene expression profiles showed that the IMD pathway may regulate expression of antimicrobial peptide (AMP) genes in the midgut, and be related to an observed down-regulation of AMPs in experimental lines of insecticide-resistant P. xylostella. A bacterial feeding study demonstrated that P. xylostella could activate different AMPs in response to bacterial infection. This study has established a framework of comprehensive expression profiles that highlight cues for immune regulation in a major pest. Our work provides a foundation for further studies on the functions of P. xylostella immune genes and mechanisms of innate immunity. PMID:25943446

Genome-wide characterization and expression profiling of immune genes in the diamondback moth, Plutella xylostella (L.).

PubMed

Xia, Xiaofeng; Yu, Liying; Xue, Minqian; Yu, Xiaoqiang; Vasseur, Liette; Gurr, Geoff M; Baxter, Simon W; Lin, Hailan; Lin, Junhan; You, Minsheng

2015-05-06

The diamondback moth, Plutella xylostella (L.), is a destructive pest that attacks cruciferous crops worldwide. Immune responses are important for interactions between insects and pathogens and information on these underpins the development of strategies for biocontrol-based pest management. Little, however, is known about immune genes and their regulation patterns in P. xylostella. A total of 149 immune-related genes in 20 gene families were identified through comparison of P. xylostella genome with the genomes of other insects. Complete and conserved Toll, IMD and JAK-STAT signaling pathways were found in P. xylostella. Genes involved in pathogen recognition were expanded and more diversified than genes associated with intracellular signal transduction. Gene expression profiles showed that the IMD pathway may regulate expression of antimicrobial peptide (AMP) genes in the midgut, and be related to an observed down-regulation of AMPs in experimental lines of insecticide-resistant P. xylostella. A bacterial feeding study demonstrated that P. xylostella could activate different AMPs in response to bacterial infection. This study has established a framework of comprehensive expression profiles that highlight cues for immune regulation in a major pest. Our work provides a foundation for further studies on the functions of P. xylostella immune genes and mechanisms of innate immunity.

The PAPI-1 pathogenicity island-encoded small RNA PesA influences Pseudomonas aeruginosa virulence and modulates pyocin S3 production

PubMed Central

Ferrara, Silvia; Falcone, Marilena; Macchi, Raffaella; Bragonzi, Alessandra; Girelli, Daniela; Cariani, Lisa; Cigana, Cristina

2017-01-01

Small non-coding RNAs (sRNAs) are post-transcriptional regulators of gene expression that have been recognized as key contributors to bacterial virulence and pathogenic mechanisms. In this study, we characterized the sRNA PesA of the opportunistic human pathogen Pseudomonas aeruginosa. We show that PesA, which is transcribed within the pathogenicity island PAPI-1 of P. aeruginosa strain PA14, contributes to P. aeruginosa PA14 virulence. In fact, pesA gene deletion resulted in a less pathogenic strain, showing higher survival of cystic fibrosis human bronchial epithelial cells after infection. Moreover, we show that PesA influences positively the expression of pyocin S3 whose genetic locus comprises two structural genes, pyoS3A and pyoS3I, encoding the killing S3A and the immunity S3I proteins, respectively. Interestingly, the deletion of pesA gene results in increased sensitivity to UV irradiation and to the fluoroquinolone antibiotic ciprofloxacin. The degree of UV sensitivity displayed by the PA14 strain lacking PesA is comparable to that of a strain deleted for pyoS3A-I. These results suggest an involvement of pyocin S3 in DNA damage repair and a regulatory role of PesA on this function. PMID:28665976

Effects of immunostimulation on social behavior, chemical communication and genome-wide gene expression in honey bee workers (Apis mellifera)

PubMed Central

2012-01-01

Background Social insects, such as honey bees, use molecular, physiological and behavioral responses to combat pathogens and parasites. The honey bee genome contains all of the canonical insect immune response pathways, and several studies have demonstrated that pathogens can activate expression of immune effectors. Honey bees also use behavioral responses, termed social immunity, to collectively defend their hives from pathogens and parasites. These responses include hygienic behavior (where workers remove diseased brood) and allo-grooming (where workers remove ectoparasites from nestmates). We have previously demonstrated that immunostimulation causes changes in the cuticular hydrocarbon profiles of workers, which results in altered worker-worker social interactions. Thus, cuticular hydrocarbons may enable workers to identify sick nestmates, and adjust their behavior in response. Here, we test the specificity of behavioral, chemical and genomic responses to immunostimulation by challenging workers with a panel of different immune stimulants (saline, Sephadex beads and Gram-negative bacteria E. coli). Results While only bacteria-injected bees elicited altered behavioral responses from healthy nestmates compared to controls, all treatments resulted in significant changes in cuticular hydrocarbon profiles. Immunostimulation caused significant changes in expression of hundreds of genes, the majority of which have not been identified as members of the canonical immune response pathways. Furthermore, several new candidate genes that may play a role in cuticular hydrocarbon biosynthesis were identified. Effects of immune challenge expression of several genes involved in immune response, cuticular hydrocarbon biosynthesis, and the Notch signaling pathway were confirmed using quantitative real-time PCR. Finally, we identified common genes regulated by pathogen challenge in honey bees and other insects. Conclusions These results demonstrate that honey bee genomic responses to immunostimulation are substantially broader than the previously identified canonical immune response pathways, and may mediate the behavioral changes associated with social immunity by orchestrating changes in chemical signaling. These studies lay the groundwork for future research into the genomic responses of honey bees to native honey bee parasites and pathogens. PMID:23072398

Effects of immunostimulation on social behavior, chemical communication and genome-wide gene expression in honey bee workers (Apis mellifera).

PubMed

Richard, Freddie-Jeanne; Holt, Holly L; Grozinger, Christina M

2012-10-16

Social insects, such as honey bees, use molecular, physiological and behavioral responses to combat pathogens and parasites. The honey bee genome contains all of the canonical insect immune response pathways, and several studies have demonstrated that pathogens can activate expression of immune effectors. Honey bees also use behavioral responses, termed social immunity, to collectively defend their hives from pathogens and parasites. These responses include hygienic behavior (where workers remove diseased brood) and allo-grooming (where workers remove ectoparasites from nestmates). We have previously demonstrated that immunostimulation causes changes in the cuticular hydrocarbon profiles of workers, which results in altered worker-worker social interactions. Thus, cuticular hydrocarbons may enable workers to identify sick nestmates, and adjust their behavior in response. Here, we test the specificity of behavioral, chemical and genomic responses to immunostimulation by challenging workers with a panel of different immune stimulants (saline, Sephadex beads and Gram-negative bacteria E. coli). While only bacteria-injected bees elicited altered behavioral responses from healthy nestmates compared to controls, all treatments resulted in significant changes in cuticular hydrocarbon profiles. Immunostimulation caused significant changes in expression of hundreds of genes, the majority of which have not been identified as members of the canonical immune response pathways. Furthermore, several new candidate genes that may play a role in cuticular hydrocarbon biosynthesis were identified. Effects of immune challenge expression of several genes involved in immune response, cuticular hydrocarbon biosynthesis, and the Notch signaling pathway were confirmed using quantitative real-time PCR. Finally, we identified common genes regulated by pathogen challenge in honey bees and other insects. These results demonstrate that honey bee genomic responses to immunostimulation are substantially broader than the previously identified canonical immune response pathways, and may mediate the behavioral changes associated with social immunity by orchestrating changes in chemical signaling. These studies lay the groundwork for future research into the genomic responses of honey bees to native honey bee parasites and pathogens.

The Wor1-like protein Fgp1 regulates pathogenicity, toxin synthesis and reproduction in the phytopathogenic fungus Fusarium graminearum

USDA-ARS?s Scientific Manuscript database

WOR1 is a gene for a conserved fungal regulatory protein controlling the dimorphic switch and pathogenicity in Candida albicans and its ortholog in the plant pathogen Fusarium oxysporum, called SGE1, is also required for pathogenicity and expression of plant effector proteins. F. graminearum, an imp...

From deep-sea volcanoes to human pathogens: a conserved quorum-sensing signal in Epsilonproteobacteria

PubMed Central

Pérez-Rodríguez, Ileana; Bolognini, Marie; Ricci, Jessica; Bini, Elisabetta; Vetriani, Costantino

2015-01-01

Chemosynthetic Epsilonproteobacteria from deep-sea hydrothermal vents colonize substrates exposed to steep thermal and redox gradients. In many bacteria, substrate attachment, biofilm formation, expression of virulence genes and host colonization are partly controlled via a cell density-dependent mechanism involving signal molecules, known as quorum sensing. Within the Epsilonproteobacteria, quorum sensing has been investigated only in human pathogens that use the luxS/autoinducer-2 (AI-2) mechanism to control the expression of some of these functions. In this study we showed that luxS is conserved in Epsilonproteobacteria and that pathogenic and mesophilic members of this class inherited this gene from a thermophilic ancestor. Furthermore, we provide evidence that the luxS gene is expressed—and a quorum-sensing signal is produced—during growth of Sulfurovum lithotrophicum and Caminibacter mediatlanticus, two Epsilonproteobacteria from deep-sea hydrothermal vents. Finally, we detected luxS transcripts in Epsilonproteobacteria-dominated biofilm communities collected from deep-sea hydrothermal vents. Taken together, our findings indicate that the epsiloproteobacterial lineage of the LuxS enzyme originated in high-temperature geothermal environments and that, in vent Epsilonproteobacteria, luxS expression is linked to the production of AI-2 signals, which are likely produced in situ at deep-sea vents. We conclude that the luxS gene is part of the ancestral epsilonproteobacterial genome and represents an evolutionary link that connects thermophiles to human pathogens. PMID:25397946

Mechanisms of Surface Antigenic Variation in the Human Pathogenic Fungus Pneumocystis jirovecii.

PubMed

Schmid-Siegert, Emanuel; Richard, Sophie; Luraschi, Amanda; Mühlethaler, Konrad; Pagni, Marco; Hauser, Philippe M

2017-11-07

Microbial pathogens commonly escape the human immune system by varying surface proteins. We investigated the mechanisms used for that purpose by Pneumocystis jirovecii This uncultivable fungus is an obligate pulmonary pathogen that in immunocompromised individuals causes pneumonia, a major life-threatening infection. Long-read PacBio sequencing was used to assemble a core of subtelomeres of a single P. jirovecii strain from a bronchoalveolar lavage fluid specimen from a single patient. A total of 113 genes encoding surface proteins were identified, including 28 pseudogenes. These genes formed a subtelomeric gene superfamily, which included five families encoding adhesive glycosylphosphatidylinositol (GPI)-anchored glycoproteins and one family encoding excreted glycoproteins. Numerical analyses suggested that diversification of the glycoproteins relies on mosaic genes created by ectopic recombination and occurs only within each family. DNA motifs suggested that all genes are expressed independently, except those of the family encoding the most abundant surface glycoproteins, which are subject to mutually exclusive expression. PCR analyses showed that exchange of the expressed gene of the latter family occurs frequently, possibly favored by the location of the genes proximal to the telomere because this allows concomitant telomere exchange. Our observations suggest that (i) the P. jirovecii cell surface is made of a complex mixture of different surface proteins, with a majority of a single isoform of the most abundant glycoprotein, (ii) genetic mosaicism within each family ensures variation of the glycoproteins, and (iii) the strategy of the fungus consists of the continuous production of new subpopulations composed of cells that are antigenically different. IMPORTANCE Pneumocystis jirovecii is a fungus causing severe pneumonia in immunocompromised individuals. It is the second most frequent life-threatening invasive fungal infection. We have studied the mechanisms of antigenic variation used by this pathogen to escape the human immune system, a strategy commonly used by pathogenic microorganisms. Using a new DNA sequencing technology generating long reads, we could characterize the highly repetitive gene families encoding the proteins that are present on the cellular surface of this pest. These gene families are localized in the regions close to the ends of all chromosomes, the subtelomeres. Such chromosomal localization was found to favor genetic recombinations between members of each gene family and to allow diversification of these proteins continuously over time. This pathogen seems to use a strategy of antigenic variation consisting of the continuous production of new subpopulations composed of cells that are antigenically different. Such a strategy is unique among human pathogens. Copyright © 2017 Schmid-Siegert et al.

Genome-wide identification of sweet orange (Citrus sinensis) histone modification gene families and their expression analysis during the fruit development and fruit-blue mold infection process.

PubMed

Xu, Jidi; Xu, Haidan; Liu, Yuanlong; Wang, Xia; Xu, Qiang; Deng, Xiuxin

2015-01-01

In eukaryotes, histone acetylation and methylation have been known to be involved in regulating diverse developmental processes and plant defense. These histone modification events are controlled by a series of histone modification gene families. To date, there is no study regarding genome-wide characterization of histone modification related genes in citrus species. Based on the two recent sequenced sweet orange genome databases, a total of 136 CsHMs (Citrus sinensis histone modification genes), including 47 CsHMTs (histone methyltransferase genes), 23 CsHDMs (histone demethylase genes), 50 CsHATs (histone acetyltransferase genes), and 16 CsHDACs (histone deacetylase genes) were identified. These genes were categorized to 11 gene families. A comprehensive analysis of these 11 gene families was performed with chromosome locations, phylogenetic comparison, gene structures, and conserved domain compositions of proteins. In order to gain an insight into the potential roles of these genes in citrus fruit development, 42 CsHMs with high mRNA abundance in fruit tissues were selected to further analyze their expression profiles at six stages of fruit development. Interestingly, a numbers of genes were expressed highly in flesh of ripening fruit and some of them showed the increasing expression levels along with the fruit development. Furthermore, we analyzed the expression patterns of all 136 CsHMs response to the infection of blue mold (Penicillium digitatum), which is the most devastating pathogen in citrus post-harvest process. The results indicated that 20 of them showed the strong alterations of their expression levels during the fruit-pathogen infection. In conclusion, this study presents a comprehensive analysis of the histone modification gene families in sweet orange and further elucidates their behaviors during the fruit development and the blue mold infection responses.

Efficient transformation and expression of gfp gene in Valsa mali var. mali.

PubMed

Chen, Liang; Sun, Gengwu; Wu, Shujing; Liu, Huixiang; Wang, Hongkai

2015-01-01

Valsa mali var. mali, the causal agent of valsa canker of apple, causes great loss of apple production in apple producing regions. The pathogenic mechanism of the pathogen has not been studied extensively, thus a suitable gene marker for pathogenic invasion analysis and a random insertion of T-DNA for mutants are desirable. In this paper, we reported the construction of a binary vector pKO1-HPH containing a positive selective gene hygromycin phosphotransferase (hph), a reporter gene gfp conferring green fluorescent protein, and an efficient protocol for V. mali var. mali transformation mediated by Agrobacterium tumefaciens. A transformation efficiency up to about 75 transformants per 10(5) conidia was achieved when co-cultivation of V. mali var. mali and A. tumefaciens for 48 h in A. tumefaciens inductive medium agar plates. The insertions of hph gene and gfp gene into V. mali var. mali genome verified by polymerase chain reaction and southern blot analysis showed that 10 randomly-selected transformants exhibited a single, unique hybridization pattern. This is the first report of A. tumefaciens-mediated transformation of V. mali var mali carrying a 'reporter' gfp gene that stably and efficiently expressed in the transformed V. mali var. mali species.

Preponderance of toxigenic Escherichia coli in stool pathogens correlates with toxin detection in accessible drinking-water sources.

PubMed

Igbokwe, H; Bhattacharyya, S; Gradus, S; Khubbar, M; Griswold, D; Navidad, J; Igwilo, C; Masson-Meyers, D; Azenabor, A A

2015-02-01

Since early detection of pathogens and their virulence factors contribute to intervention and control strategies, we assessed the enteropathogens in diarrhoea disease and investigated the link between toxigenic strains of Escherichia coli from stool and drinking-water sources; and determined the expression of toxin genes by antibiotic-resistant E. coli in Lagos, Nigeria. This was compared with isolates from diarrhoeal stool and water from Wisconsin, USA. The new Luminex xTAG GPP (Gastroplex) technique and conventional real-time PCR were used to profile enteric pathogens and E. coli toxin gene isolates, respectively. Results showed the pathogen profile of stool and indicated a relationship between E. coli toxin genes in water and stool from Lagos which was absent in Wisconsin isolates. The Gastroplex technique was efficient for multiple enteric pathogens and toxin gene detection. The co-existence of antibiotic resistance with enteroinvasive E. coli toxin genes suggests an additional prognostic burden on patients.

Expression of tomato prosystemin gene in Arabidopsis reveals systemic translocation of its mRNA and confers necrotrophic fungal resistance.

PubMed

Zhang, Haiyan; Yu, Pengli; Zhao, Jiuhai; Jiang, Hongling; Wang, Haiyang; Zhu, Yingfang; Botella, Miguel A; Šamaj, Jozef; Li, Chuanyou; Lin, Jinxing

2018-01-01

Systemin (SYS), an octadecapeptide hormone processed from a 200-amino-acid precursor (prosystemin, PS), plays a central role in the systemic activation of defense genes in tomato in response to herbivore and pathogen attacks. However, whether PS mRNA is transferable and its role in systemic defense responses remain unknown. We created the transgenic tomato PS gene tagged with the green fluorescent protein (PS-GFP) using a shoot- or root-specific promoter, and the constitutive 35S promoter in Arabidopsis. Subcellular localization of PS-/SYS-GFP was observed using confocal laser scanning microscopy and gene transcripts were determined using quantitative real-time PCR. In Arabidopsis, PS protein can be processed and SYS is secreted. Shoot-/root-specific expression of PS-GFP in Arabidopsis, and grafting experiments, revealed that the PS mRNA moves in a bi-directional manner. We also found that ectopic expression of PS improves Arabidopsis resistance to the necrotrophic fungus Botrytis cinerea, consistent with substantial upregulation of the transcript levels of specific pathogen-responsive genes. Our results provide novel insights into the multifaceted mechanism of SYS signaling transport and its potential application in genetic engineering for increasing pathogen resistance across diverse plant families. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Large-scale transcriptome analysis reveals arabidopsis metabolic pathways are frequently influenced by different pathogens.

PubMed

Jiang, Zhenhong; He, Fei; Zhang, Ziding

2017-07-01

Through large-scale transcriptional data analyses, we highlighted the importance of plant metabolism in plant immunity and identified 26 metabolic pathways that were frequently influenced by the infection of 14 different pathogens. Reprogramming of plant metabolism is a common phenomenon in plant defense responses. Currently, a large number of transcriptional profiles of infected tissues in Arabidopsis (Arabidopsis thaliana) have been deposited in public databases, which provides a great opportunity to understand the expression patterns of metabolic pathways during plant defense responses at the systems level. Here, we performed a large-scale transcriptome analysis based on 135 previously published expression samples, including 14 different pathogens, to explore the expression pattern of Arabidopsis metabolic pathways. Overall, metabolic genes are significantly changed in expression during plant defense responses. Upregulated metabolic genes are enriched on defense responses, and downregulated genes are enriched on photosynthesis, fatty acid and lipid metabolic processes. Gene set enrichment analysis (GSEA) identifies 26 frequently differentially expressed metabolic pathways (FreDE_Paths) that are differentially expressed in more than 60% of infected samples. These pathways are involved in the generation of energy, fatty acid and lipid metabolism as well as secondary metabolite biosynthesis. Clustering analysis based on the expression levels of these 26 metabolic pathways clearly distinguishes infected and control samples, further suggesting the importance of these metabolic pathways in plant defense responses. By comparing with FreDE_Paths from abiotic stresses, we find that the expression patterns of 26 FreDE_Paths from biotic stresses are more consistent across different infected samples. By investigating the expression correlation between transcriptional factors (TFs) and FreDE_Paths, we identify several notable relationships. Collectively, the current study will deepen our understanding of plant metabolism in plant immunity and provide new insights into disease-resistant crop improvement.

Cloning and characterization of a novel chitinase gene (chi46) from Chaetomium globosum and identification of its biological activity.

PubMed

Liu, Z H; Yang, Q; Hu, S; Zhang, J D; Ma, J

2008-08-01

Chitinases play a major role in the defensive strategies of plants against fungal pathogens. In the current study, the gene for a 46-kDa endochitinase (chi46) was cloned from Chaetomium globosum, an important biocontrol fungus. The corresponding complementary deoxyribonucleic acid sequence was 1,350 bp in length, encoding 449 amino acid residues. The temporal expression of chi46, in response to the treatments of cell walls of six pathogens and confrontation against two fungal pathogens, was measured in C. globosum using real-time reverse transcription polymerase chain reaction. The expression of chi46 can be highly induced by exposure to the cell walls of plant pathogens and living pathogens, suggesting a role in plant disease resistance. The chi46 gene was inserted into the pPIC9 vector and transferred into the cells of Pichia pastoris GS115 for heterologous expression. The optimal reaction conditions for chitinase CHI46 activity were: 45 degrees C, pH of 5.0, and 5 mmol l(-1) of Cu2+. The maximum enzyme activity was 1.42 U ml(-1) following exposure to the cell wall chitin of Septoria tritici. The CHI46 enzyme can efficiently degrade cell walls of the phytopathogenic Rhizoctonia solani, Fusarium oxysporum, Sclerotinia sclerotiorum, Valsa sordida, S. tritici, and Phytophthora sojae, demonstrating that it may be involved in the biocontrol mechanism of C. globosum.

Global Expression Studies of Yersinia Pestis Pathogenicity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Garcia, E; Motin, V; Brubaker, R

2002-10-15

The aim of these studies continues to be the investigation into the molecular mechanisms that underlie the virulence process in Yersinia pestis. In particular, the focus of this work centers on the identification of novel genes and pathways responsible for the pathogenic properties of this organism. In spite of more than four decades of intense investigation in this field, the dilemma as to what makes Y. pestis such a virulent and lethal pathogen remains unanswered. The method being employed makes use microarray technology (DNA chip) that enables the examination of the global activities of the whole complement of genes inmore » this pathogen. Two primary resources available to the investigators (one directly obtained from a separate CBNP-funded project) make these studies possible: (1) Whole genome comparisons of the genes in Y. pestis and its near neighbors with attenuated or non pathogenic characteristics, and (2) the ability to duplicate in vitro, conditions that mimic the infection process of this pathogen. This year we have extended our studies from the original work of characterizing the global transcriptional regulation in Y. pestis triggered during temperature transition from 26 C to 37 C (roughly conditions found in the flea vector and the mammalian host, respectively) to studies of regulation encountered during shift between growth from conditions of neutral pH to acidic pH (the latter conditions, those mimic the environment found inside macrophages, a likely environment found by these cells during infection.). For this work, DNA arrays containing some 5,000 genes (the entire genome of Y. pestis plus those genes found uniquely in the enteropathogen, and near neighbor, Y. pseudotuberculosis) are used to monitor the simultaneous expression levels of each gene of known and unknown function in Y. pestis. Those genes that are up-regulate under the experimental conditions represent genes potentially involved in the pathogenic process. The ultimate role in pathogenicity of those candidate genes uncovered from these studies will be further ascertained by direct knock outs (gene inactivation) and by in vivo studies using an animal model. Discovery of new virulence factors in Y. pestis will directly impact the development of new signatures for detection and geo-location since it will help us to understand and identify those genes that are essential in making the organism pathogenic. These are genes that cannot be altered or removed from the pathogen and as such constitute the best type of signature that we can utilize in their detection and identification. Applications such as this will also enable the utilization of similar technologies to study other pathogens such as Francisella and Brucella, for which we know substantially less in terms of their modality of virulence.« less

Genome-wide identification of the SWEET gene family in wheat.

PubMed

Gao, Yue; Wang, Zi Yuan; Kumar, Vikranth; Xu, Xiao Feng; Yuan, De Peng; Zhu, Xiao Feng; Li, Tian Ya; Jia, Baolei; Xuan, Yuan Hu

2018-02-05

The SWEET (sugars will eventually be exported transporter) family is a newly characterized group of sugar transporters. In plants, the key roles of SWEETs in phloem transport, nectar secretion, pollen nutrition, stress tolerance, and plant-pathogen interactions have been identified. SWEET family genes have been characterized in many plant species, but a comprehensive analysis of SWEET members has not yet been performed in wheat. Here, 59 wheat SWEETs (hereafter TaSWEETs) were identified through homology searches. Analyses of phylogenetic relationships, numbers of transmembrane helices (TMHs), gene structures, and motifs showed that TaSWEETs carrying 3-7 TMHs could be classified into four clades with 10 different types of motifs. Examination of the expression patterns of 18 SWEET genes revealed that a few are tissue-specific while most are ubiquitously expressed. In addition, the stem rust-mediated expression patterns of SWEET genes were monitored using a stem rust-susceptible cultivar, 'Little Club' (LC). The resulting data showed that the expression of five out of the 18 SWEETs tested was induced following inoculation. In conclusion, we provide the first comprehensive analysis of the wheat SWEET gene family. Information regarding the phylogenetic relationships, gene structures, and expression profiles of SWEET genes in different tissues and following stem rust disease inoculation will be useful in identifying the potential roles of SWEETs in specific developmental and pathogenic processes. Copyright © 2017 Elsevier B.V. All rights reserved.

Selection of suitable reference genes for gene expression studies in Staphylococcus capitis during growth under erythromycin stress.

PubMed

Cui, Bintao; Smooker, Peter M; Rouch, Duncan A; Deighton, Margaret A

2016-08-01

Accurate and reproducible measurement of gene transcription requires appropriate reference genes, which are stably expressed under different experimental conditions to provide normalization. Staphylococcus capitis is a human pathogen that produces biofilm under stress, such as imposed by antimicrobial agents. In this study, a set of five commonly used staphylococcal reference genes (gyrB, sodA, recA, tuf and rpoB) were systematically evaluated in two clinical isolates of Staphylococcus capitis (S. capitis subspecies urealyticus and capitis, respectively) under erythromycin stress in mid-log and stationary phases. Two public software programs (geNorm and NormFinder) and two manual calculation methods, reference residue normalization (RRN) and relative quantitative (RQ), were applied. The potential reference genes selected by the four algorithms were further validated by comparing the expression of a well-studied biofilm gene (icaA) with phenotypic biofilm formation in S. capitis under four different experimental conditions. The four methods differed considerably in their ability to predict the most suitable reference gene or gene combination for comparing icaA expression under different conditions. Under the conditions used here, the RQ method provided better selection of reference genes than the other three algorithms; however, this finding needs to be confirmed with a larger number of isolates. This study reinforces the need to assess the stability of reference genes for analysis of target gene expression under different conditions and the use of more than one algorithm in such studies. Although this work was conducted using a specific human pathogen, it emphasizes the importance of selecting suitable reference genes for accurate normalization of gene expression more generally.

Pathogenic Leptospira species express surface-exposed proteins belonging to the bacterial immunoglobulin superfamily

PubMed Central

Matsunaga, James; Barocchi, Michele A.; Croda, Julio; Young, Tracy A.; Sanchez, Yolanda; Siqueira, Isadora; Bolin, Carole A.; Reis, Mitermayer G.; Riley, Lee W.; Haake, David A.; Ko, Albert I.

2005-01-01

Summary Proteins with bacterial immunoglobulin-like (Big) domains, such as the Yersinia pseudotuberculosis invasin and Escherichia coli intimin, are surface-expressed proteins that mediate host mammalian cell invasion or attachment. Here, we report the identification and characterization of a new family of Big domain proteins, referred to as Lig (leptospiral Ig-like) proteins, in pathogenic Leptospira. Screening of L. interrogans and L. kirschneri expression libraries with sera from leptospirosis patients identified 13 lambda phage clones that encode tandem repeats of the 90 amino acid Big domain. Two lig genes, designated ligA and ligB, and one pseudo-gene, ligC, were identified. The ligA and ligB genes encode amino-terminal lipoprotein signal peptides followed by 10 or 11 Big domain repeats and, in the case of ligB, a unique carboxy-terminal non-repeat domain. The organization of ligC is similar to that of ligB but contains mutations that disrupt the reading frame. The lig sequences are present in pathogenic but not saprophytic Leptospira species. LigA and LigB are expressed by a variety of virulent leptospiral strains. Loss of Lig protein and RNA transcript expression is correlated with the observed loss of virulence during culture attenuation of pathogenic strains. High-pressure freeze substitution followed by immunocytochemical electron microscopy confirmed that the Lig proteins were localized to the bacterial surface. Immunoblot studies with patient sera found that the Lig proteins are a major antigen recognized during the acute host infection. These observations demonstrate that the Lig proteins are a newly identified surface protein of pathogenic Leptospira, which by analogy to other bacterial immunoglobulin superfamily virulence factors, may play a role in host cell attachment and invasion during leptospiral pathogenesis. PMID:12890019

Nicotiana plumbaginifolia plants silenced for the ATP-binding cassette transporter gene NpPDR1 show increased susceptibility to a group of fungal and oomycete pathogens.

PubMed

Bultreys, Alain; Trombik, Tomasz; Drozak, Anna; Boutry, Marc

2009-09-01

SUMMARY The behaviour of Nicotiana plumbaginifolia plants silenced for the ATP-binding cassette transporter gene NpPDR1 was investigated in response to fungal and oomycete infections. The importance of NpPDR1 in plant defence was demonstrated for two organs in which NpPDR1 is constitutively expressed: the roots and the petal epidermis. The roots of the plantlets of two lines silenced for NpPDR1 expression were clearly more sensitive than those of controls to the fungal pathogens Botrytis cinerea, Fusarium oxysporum sp., F. oxysporum f. sp. nicotianae, F. oxysporum f. sp. melonis and Rhizoctonia solani, as well as to the oomycete pathogen Phytophthora nicotianae race 0. The Ph gene-linked resistance of N. plumbaginifolia to P. nicotianae race 0 was totally ineffective in NpPDR1-silenced lines. In addition, the petals of the NpPDR1-silenced lines were spotted 15%-20% more rapidly by B. cinerea than were the controls. The rapid induction (after 2-4 days) of NpPDR1 expression in N. plumbaginifolia and N. tabacum mature leaves in response to pathogen presence was demonstrated for the first time with fungi and one oomycete: R. solani, F. oxysporum and P. nicotianae. With B. cinerea, such rapid expression was not observed in healthy mature leaves. NpPDR1 expression was not observed during latent infections of B. cinerea in N. plumbaginifolia and N. tabacum, but was induced when conditions facilitated B. cinerea development in leaves, such as leaf ageing or an initial root infection. This work demonstrates the increased sensitivity of NpPDR1-silenced N. plumbaginifolia plants to all of the fungal and oomycete pathogens investigated.

Transcriptome dynamics of a susceptible wheat upon Fusarium head blight reveals that molecular responses to Fusarium graminearum infection fit over the grain development processes.

PubMed

Chetouhi, Cherif; Bonhomme, Ludovic; Lasserre-Zuber, Pauline; Cambon, Florence; Pelletier, Sandra; Renou, Jean-Pierre; Langin, Thierry

2016-03-01

In many plant/pathogen interactions, host susceptibility factors are key determinants of disease development promoting pathogen growth and spreading in plant tissues. In the Fusarium head blight (FHB) disease, the molecular basis of wheat susceptibility is still poorly understood while it could provide new insights into the understanding of the wheat/Fusarium graminearum (Fg) interaction and guide future breeding programs to produce cultivars with sustainable resistance. To identify the wheat grain candidate genes, a genome-wide gene expression profiling was performed in the French susceptible wheat cultivar, Recital. Gene-specific two-way ANOVA of about 40 K transcripts at five grain developmental stages identified 1309 differentially expressed genes. Out of these, 536 were impacted by the Fg effect alone. Most of these Fg-responsive genes belonged to biological and molecular functions related to biotic and abiotic stresses indicating the activation of common stress pathways during susceptibility response of wheat grain to FHB. This analysis revealed also 773 other genes displaying either specific Fg-responsive profiles along with grain development stages or synergistic adjustments with the grain development effect. These genes were involved in various molecular pathways including primary metabolism, cell death, and gene expression reprogramming. An increasingly complex host response was revealed, as was the impact of both Fg infection and grain ontogeny on the transcription of wheat genes. This analysis provides a wealth of candidate genes and pathways involved in susceptibility responses to FHB and depicts new clues to the understanding of the susceptibility determinism in plant/pathogen interactions.

A remarkable synergistic effect at the transcriptomic level in peach fruits doubly infected by prunus necrotic ringspot virus and peach latent mosaic viroid.

PubMed

Herranz, Mari Carmen; Niehl, Annette; Rosales, Marlene; Fiore, Nicola; Zamorano, Alan; Granell, Antonio; Pallas, Vicente

2013-05-28

Microarray profiling is a powerful technique to investigate expression changes of large amounts of genes in response to specific environmental conditions. The majority of the studies investigating gene expression changes in virus-infected plants are limited to interactions between a virus and a model host plant, which usually is Arabidopsis thaliana or Nicotiana benthamiana. In the present work, we performed microarray profiling to explore changes in the expression profile of field-grown Prunus persica (peach) originating from Chile upon single and double infection with Prunus necrotic ringspot virus (PNRSV) and Peach latent mosaic viroid (PLMVd), worldwide natural pathogens of peach trees. Upon single PLMVd or PNRSV infection, the number of statistically significant gene expression changes was relatively low. By contrast, doubly-infected fruits presented a high number of differentially regulated genes. Among these, down-regulated genes were prevalent. Functional categorization of the gene expression changes upon double PLMVd and PNRSV infection revealed protein modification and degradation as the functional category with the highest percentage of repressed genes whereas induced genes encoded mainly proteins related to phosphate, C-compound and carbohydrate metabolism and also protein modification. Overrepresentation analysis upon double infection with PLMVd and PNRSV revealed specific functional categories over- and underrepresented among the repressed genes indicating active counter-defense mechanisms of the pathogens during infection. Our results identify a novel synergistic effect of PLMVd and PNRSV on the transcriptome of peach fruits. We demonstrate that mixed infections, which occur frequently in field conditions, result in a more complex transcriptional response than that observed in single infections. Thus, our data demonstrate for the first time that the simultaneous infection of a viroid and a plant virus synergistically affect the host transcriptome in infected peach fruits. These field studies can help to fully understand plant-pathogen interactions and to develop appropriate crop protection strategies.

A remarkable synergistic effect at the transcriptomic level in peach fruits doubly infected by prunus necrotic ringspot virus and peach latent mosaic viroid

PubMed Central

2013-01-01

Background Microarray profiling is a powerful technique to investigate expression changes of large amounts of genes in response to specific environmental conditions. The majority of the studies investigating gene expression changes in virus-infected plants are limited to interactions between a virus and a model host plant, which usually is Arabidopsis thaliana or Nicotiana benthamiana. In the present work, we performed microarray profiling to explore changes in the expression profile of field-grown Prunus persica (peach) originating from Chile upon single and double infection with Prunus necrotic ringspot virus (PNRSV) and Peach latent mosaic viroid (PLMVd), worldwide natural pathogens of peach trees. Results Upon single PLMVd or PNRSV infection, the number of statistically significant gene expression changes was relatively low. By contrast, doubly-infected fruits presented a high number of differentially regulated genes. Among these, down-regulated genes were prevalent. Functional categorization of the gene expression changes upon double PLMVd and PNRSV infection revealed protein modification and degradation as the functional category with the highest percentage of repressed genes whereas induced genes encoded mainly proteins related to phosphate, C-compound and carbohydrate metabolism and also protein modification. Overrepresentation analysis upon double infection with PLMVd and PNRSV revealed specific functional categories over- and underrepresented among the repressed genes indicating active counter-defense mechanisms of the pathogens during infection. Conclusions Our results identify a novel synergistic effect of PLMVd and PNRSV on the transcriptome of peach fruits. We demonstrate that mixed infections, which occur frequently in field conditions, result in a more complex transcriptional response than that observed in single infections. Thus, our data demonstrate for the first time that the simultaneous infection of a viroid and a plant virus synergistically affect the host transcriptome in infected peach fruits. These field studies can help to fully understand plant-pathogen interactions and to develop appropriate crop protection strategies. PMID:23710752

DNA-Binding Protein HU Coordinates Pathogenicity in Vibrio parahaemolyticus.

PubMed

Phan, Ngoc Quang; Uebanso, Takashi; Shimohata, Takaaki; Nakahashi, Mutsumi; Mawatari, Kazuaki; Takahashi, Akira

2015-09-01

HU is one of the most abundant nucleoid-associated proteins in bacterial cells and regulates the expression of many genes involved in growth, motility, metabolism, and virulence. It is known that Vibrio parahaemolyticus pathogenicity is related to its characteristic rapid growth and that type III secretion system 1 (T3SS1) contributes to its cytotoxicity. However, it is not known if HU plays a role in the pathogenicity of V. parahaemolyticus. In the present study, we investigated the effect of HU proteins HU-2 (HUα) (V. parahaemolyticus 2911 [vp2911]) and HUβ (vp0920) on the pathogenicity of V. parahaemolyticus. We found that a deletion of both HU subunits (yielding the ΔHUs [Δvp0920 Δvp2911] strain), but not single deletions, led to a reduction of the growth rate. In addition, expression levels of T3SS1-related genes, including exsA (positive regulator), exsD (negative regulator), vp1680 (cytotoxic effector), and vp1671 (T3SS1 apparatus), were reduced in the ΔHUs strain compared to the wild type (WT). As a result, cytotoxicity to HeLa cells was decreased in the ΔHUs strain. The additional deletion of exsD in the ΔHUs strain restored T3SS1-related gene expression levels and cytotoxicity but not the growth rate. These results suggest that the HU protein regulates the levels of T3SS1 gene expression and cytotoxicity in a growth rate-independent manner. Nucleoid-binding protein HU regulates cellular behaviors, including nucleoid structuring, general recombination, transposition, growth, replication, motility, metabolism, and virulence. It is thought that both the number of bacteria and the number of virulence factors may affect the pathogenicity of bacteria. In the present study, we investigated which factor(s) has a dominant role during infection in one of the most rapidly growing bacterial species, Vibrio parahaemolyticus. We found that V. parahaemolyticus cytotoxicity is regulated, in a growth rate-independent manner, by the HU proteins through regulation of a number of virulence factors, including T3SS1 gene expression. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

A De Novo-Assembly Based Data Analysis Pipeline for Plant Obligate Parasite Metatranscriptomic Studies.

PubMed

Guo, Li; Allen, Kelly S; Deiulio, Greg; Zhang, Yong; Madeiras, Angela M; Wick, Robert L; Ma, Li-Jun

2016-01-01

Current and emerging plant diseases caused by obligate parasitic microbes such as rusts, downy mildews, and powdery mildews threaten worldwide crop production and food safety. These obligate parasites are typically unculturable in the laboratory, posing technical challenges to characterize them at the genetic and genomic level. Here we have developed a data analysis pipeline integrating several bioinformatic software programs. This pipeline facilitates rapid gene discovery and expression analysis of a plant host and its obligate parasite simultaneously by next generation sequencing of mixed host and pathogen RNA (i.e., metatranscriptomics). We applied this pipeline to metatranscriptomic sequencing data of sweet basil (Ocimum basilicum) and its obligate downy mildew parasite Peronospora belbahrii, both lacking a sequenced genome. Even with a single data point, we were able to identify both candidate host defense genes and pathogen virulence genes that are highly expressed during infection. This demonstrates the power of this pipeline for identifying genes important in host-pathogen interactions without prior genomic information for either the plant host or the obligate biotrophic pathogen. The simplicity of this pipeline makes it accessible to researchers with limited computational skills and applicable to metatranscriptomic data analysis in a wide range of plant-obligate-parasite systems.

InvA protein is a Nudix hydrolase required for infection by pathogenic Leptospira in cell lines and animals.

PubMed

Luo, Yihui; Liu, Yan; Sun, Dexter; Ojcius, David M; Zhao, Jinfang; Lin, Xuai; Wu, Dong; Zhang, Rongguang; Chen, Ming; Li, Lanjuan; Yan, Jie

2011-10-21

Leptospirosis caused by pathogenic species of the genus Leptospira is a re-emerging zoonotic disease, which affects a wide variety of host species and is transmitted by contaminated water. The genomes of several pathogenic Leptospira species contain a gene named invA, which contains a Nudix domain. However, the function of this gene has never been characterized. Here, we demonstrated that the invA gene was highly conserved in protein sequence and present in all tested pathogenic Leptospira species. The recombinant InvA protein of pathogenic L. interrogans strain Lai hydrolyzed several specific dinucleoside oligophosphate substrates, reflecting the enzymatic activity of Nudix in Leptospira species. Pathogenic leptospires did not express this protein in media but temporarily expressed it at early stages (within 60 min) of infection of macrophages and nephric epithelial cells. Comparing with the wild type, the invA-deficient mutant displayed much lower infectivity and a significantly reduced survival rate in macrophages and nephric epithelial cells. Moreover, the invA-deficient leptospires presented an attenuated virulence in hamsters, caused mild histopathological damage, and were transmitted in lower numbers in the urine, compared with the wild-type strain. The invA revertant, made by complementing the invA-deficient mutant with the invA gene, reacquired virulence similar to the wild type in vitro and in vivo. The LD(50) in hamsters was 1000-fold higher for the invA-deficient mutant than for the invA revertant and wild type. These results demonstrate that the InvA protein is a Nudix hydrolase, and the invA gene is essential for virulence in pathogenic Leptospira species.

Comparison of immune transcriptome response following infection with PRRSV, PCV2 and SIV

USDA-ARS?s Scientific Manuscript database

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major pathogen of swine. Here we identified and compared gene expression changes in tracheobronchial lymph nodes (TBLN) following viral infection using Digital Gene Expression Tag Profiling (DGETP). Pigs were infected with 1 x 10**5 ce...

Comparative Evolution of Morphological Regulatory Functions in Candida Species

PubMed Central

Lackey, Erika; Vipulanandan, Geethanjali; Childers, Delma S.

2013-01-01

Morphological transitions play an important role in virulence and virulence-related processes in a wide variety of pathogenic fungi, including the most commonly isolated human fungal pathogen Candida albicans. While environmental signals, transcriptional regulators, and target genes associated with C. albicans morphogenesis are well-characterized, considerably little is known about morphological regulatory mechanisms and the extent to which they are evolutionarily conserved in less pathogenic and less filamentous non-albicans Candida species (NACS). We have identified specific optimal filament-inducing conditions for three NACS (C. tropicalis, C. parapsilosis, and C. guilliermondii), which are very limited, suggesting that these species may be adapted for niche-specific filamentation in the host. Only a subset of evolutionarily conserved C. albicans filament-specific target genes were induced upon filamentation in C. tropicalis, C. parapsilosis, and C. guilliermondii. One of the genes showing conserved expression was UME6, a key filament-specific regulator of C. albicans hyphal development. Constitutive high-level expression of UME6 was sufficient to drive increased filamentation as well as biofilm formation and partly restore conserved filament-specific gene expression in both C. tropicalis and C. parapsilosis, suggesting that evolutionary differences in filamentation ability among pathogenic Candida species may be partially attributed to alterations in the expression level of a conserved filamentous growth machinery. In contrast to UME6, NRG1, an important repressor of C. albicans filamentation, showed only a partly conserved role in controlling NACS filamentation. Overall, our results suggest that C. albicans morphological regulatory functions are partially conserved in NACS and have evolved to respond to more specific sets of host environmental cues. PMID:23913541

Escherichia coli global gene expression in urine from women with urinary tract infection.

PubMed

Hagan, Erin C; Lloyd, Amanda L; Rasko, David A; Faerber, Gary J; Mobley, Harry L T

2010-11-11

Murine models of urinary tract infection (UTI) have provided substantial data identifying uropathogenic E. coli (UPEC) virulence factors and assessing their expression in vivo. However, it is unclear how gene expression in these animal models compares to UPEC gene expression during UTI in humans. To address this, we used a UPEC strain CFT073-specific microarray to measure global gene expression in eight E. coli isolates monitored directly from the urine of eight women presenting at a clinic with bacteriuria. The resulting gene expression profiles were compared to those of the same E. coli isolates cultured statically to exponential phase in pooled, sterilized human urine ex vivo. Known fitness factors, including iron acquisition and peptide transport systems, were highly expressed during human UTI and support a model in which UPEC replicates rapidly in vivo. While these findings were often consistent with previous data obtained from the murine UTI model, host-specific differences were observed. Most strikingly, expression of type 1 fimbrial genes, which are among the most highly expressed genes during murine experimental UTI and encode an essential virulence factor for this experimental model, was undetectable in six of the eight E. coli strains from women with UTI. Despite the lack of type 1 fimbrial expression in the urine samples, these E. coli isolates were generally capable of expressing type 1 fimbriae in vitro and highly upregulated fimA upon experimental murine infection. The findings presented here provide insight into the metabolic and pathogenic profile of UPEC in urine from women with UTI and represent the first transcriptome analysis for any pathogenic E. coli during a naturally occurring infection in humans.

Transcriptomic analysis reveals tomato genes whose expression is induced specifically during effector-triggered immunity and identifies the Epk1 protein kinase which is required for the host response to three bacterial effector proteins.

PubMed

Pombo, Marina A; Zheng, Yi; Fernandez-Pozo, Noe; Dunham, Diane M; Fei, Zhangjun; Martin, Gregory B

2014-01-01

Plants have two related immune systems to defend themselves against pathogen attack. Initially,pattern-triggered immunity is activated upon recognition of microbe-associated molecular patterns by pattern recognition receptors. Pathogenic bacteria deliver effector proteins into the plant cell that interfere with this immune response and promote disease. However, some plants express resistance proteins that detect the presence of specific effectors leading to a robust defense response referred to as effector-triggered immunity. The interaction of tomato with Pseudomonas syringae pv. tomato is an established model system for understanding the molecular basis of these plant immune responses. We apply high-throughput RNA sequencing to this pathosystem to identify genes whose expression changes specifically during pattern-triggered or effector-triggered immunity. We then develop reporter genes for each of these responses that will enable characterization of the host response to the large collection of P. s. pv. tomato strains that express different combinations of effectors. Virus-induced gene silencing of 30 of the effector-triggered immunity-specific genes identifies Epk1 which encodes a predicted protein kinase from a family previously unknown to be involved in immunity. Knocked-down expression of Epk1 compromises effector-triggered immunity triggered by three bacterial effectors but not by effectors from non-bacterial pathogens. Epistasis experiments indicate that Epk1 acts upstream of effector-triggered immunity-associated MAP kinase signaling. Using RNA-seq technology we identify genes involved in specific immune responses. A functional genomics screen led to the discovery of Epk1, a novel predicted protein kinase required for plant defense activation upon recognition of three different bacterial effectors.

Can-miRn37a mediated suppression of ethylene response factors enhances the resistance of chilli against anthracnose pathogen Colletotrichum truncatum L.

PubMed

Mishra, Rukmini; Mohanty, Jatindra Nath; Chand, Subodh Kumar; Joshi, Raj Kumar

2018-02-01

Pepper anthracnose, caused by Colletotrichum species complex is the most destructive disease of chilli (Capsicum annuum L.). miRNAs are key modulators of transcriptional and post- transcriptional expression of genes during defense responses. In the present study, we performed a comparative miRNA profiling of susceptible (Arka Lohit-AL) and resistant (Punjab Lal-PL) chilli cultivars to identify 35 differentially expressed miRNAs that could be classified as positive, negative or basal regulators of defense against C. truncatum, the most potent anthracnose pathogen. Interestingly, a novel microRNA can-miRn37a was significantly induced in PL but largely repressed in AL genotype post pathogen attack. Subsequent over-expression of can-miRn37a in AL showed enhanced resistance to anthracnose, as evidenced by decreased fungal growth and induced expression of defense-related genes. Consequently, the expression of its three target genes encoding the ethylene response factors (ERFs) was down-regulated in PL as well as in the over-expression lines of AL genotypes. The ability of these targets to be regulated by can-miRn37a was further confirmed by transient co-expression in Nicotiana benthamiana. Additionally, the virus-induced silencing of the three targets in the susceptible AL cultivar revealed their role in fungal colonization and induction of C. truncatum pathogenicity in chilli. Taken together, our study suggests that can-miRn37a provides a potential miRNA mediated approach of engineering anthracnose resistance in chilli by repressing ERFs and preventing fungal colonization. Copyright © 2017 Elsevier B.V. All rights reserved.

PtrWRKY73, a salicylic acid-inducible poplar WRKY transcription factor, is involved in disease resistance in Arabidopsis thaliana.

PubMed

Duan, Yanjiao; Jiang, Yuanzhong; Ye, Shenglong; Karim, Abdul; Ling, Zhengyi; He, Yunqiu; Yang, Siqi; Luo, Keming

2015-05-01

A salicylic acid-inducible WRKY gene, PtrWRKY73, from Populus trichocarpa , was isolated and characterized. Overexpression of PtrWRKY73 in Arabidopsis thaliana increased resistance to biotrophic pathogens but reduced resistance against necrotrophic pathogens. WRKY transcription factors are commonly involved in plant defense responses. However, limited information is available about the roles of the WRKY genes in poplar defense. In this study, we isolated a salicylic acid (SA)-inducible WRKY gene, PtrWRKY73, from Populus trichocarpa, belonging to group I family and containing two WRKY domains, a D domain and an SP cluster. PtrWRKY73 was expressed predominantly in roots, old leaves, sprouts and stems, especially in phloem and its expression was induced in response to treatment with exogenous SA. PtrWRKY73 was localized to the nucleus of plant cells and exhibited transcriptional activation. Overexpression of PtrWRKY73 in Arabidopsis thaliana resulted in increased resistance to a virulent strain of the bacterial pathogen Pseudomonas syringae (PstDC3000), but more sensitivity to the necrotrophic fungal pathogen Botrytis cinerea. The SA-mediated defense-associated genes, such as PR1, PR2 and PAD4, were markedly up-regulated in transgenic plants overexpressing PtrWRKY73. Arabidopsis non-expressor of PR1 (NPR1) was not affected, whereas a defense-related gene PAL4 had reduced in PtrWRKY73 overexpressor plants. Together, these results indicated that PtrWRKY73 plays a positive role in plant resistance to biotrophic pathogens but a negative effect on resistance against necrotrophic pathogens.

Epigenetic control of effector gene expression in the plant pathogenic fungus Leptosphaeria maculans.

PubMed

Soyer, Jessica L; El Ghalid, Mennat; Glaser, Nicolas; Ollivier, Bénédicte; Linglin, Juliette; Grandaubert, Jonathan; Balesdent, Marie-Hélène; Connolly, Lanelle R; Freitag, Michael; Rouxel, Thierry; Fudal, Isabelle

2014-03-01

Plant pathogens secrete an arsenal of small secreted proteins (SSPs) acting as effectors that modulate host immunity to facilitate infection. SSP-encoding genes are often located in particular genomic environments and show waves of concerted expression at diverse stages of plant infection. To date, little is known about the regulation of their expression. The genome of the Ascomycete Leptosphaeria maculans comprises alternating gene-rich GC-isochores and gene-poor AT-isochores. The AT-isochores harbor mosaics of transposable elements, encompassing one-third of the genome, and are enriched in putative effector genes that present similar expression patterns, namely no expression or low-level expression during axenic cultures compared to strong induction of expression during primary infection of oilseed rape (Brassica napus). Here, we investigated the involvement of one specific histone modification, histone H3 lysine 9 methylation (H3K9me3), in epigenetic regulation of concerted effector gene expression in L. maculans. For this purpose, we silenced the expression of two key players in heterochromatin assembly and maintenance, HP1 and DIM-5 by RNAi. By using HP1-GFP as a heterochromatin marker, we observed that almost no chromatin condensation is visible in strains in which LmDIM5 was silenced by RNAi. By whole genome oligoarrays we observed overexpression of 369 or 390 genes, respectively, in the silenced-LmHP1 and -LmDIM5 transformants during growth in axenic culture, clearly favouring expression of SSP-encoding genes within AT-isochores. The ectopic integration of four effector genes in GC-isochores led to their overexpression during growth in axenic culture. These data strongly suggest that epigenetic control, mediated by HP1 and DIM-5, represses the expression of at least part of the effector genes located in AT-isochores during growth in axenic culture. Our hypothesis is that changes of lifestyle and a switch toward pathogenesis lift chromatin-mediated repression, allowing a rapid response to new environmental conditions.

Network Analysis Reveals a Common Host-Pathogen Interaction Pattern in Arabidopsis Immune Responses.

PubMed

Li, Hong; Zhou, Yuan; Zhang, Ziding

2017-01-01

Many plant pathogens secrete virulence effectors into host cells to target important proteins in host cellular network. However, the dynamic interactions between effectors and host cellular network have not been fully understood. Here, an integrative network analysis was conducted by combining Arabidopsis thaliana protein-protein interaction network, known targets of Pseudomonas syringae and Hyaloperonospora arabidopsidis effectors, and gene expression profiles in the immune response. In particular, we focused on the characteristic network topology of the effector targets and differentially expressed genes (DEGs). We found that effectors tended to manipulate key network positions with higher betweenness centrality. The effector targets, especially those that are common targets of an individual effector, tended to be clustered together in the network. Moreover, the distances between the effector targets and DEGs increased over time during infection. In line with this observation, pathogen-susceptible mutants tended to have more DEGs surrounding the effector targets compared with resistant mutants. Our results suggest a common plant-pathogen interaction pattern at the cellular network level, where pathogens employ potent local impact mode to interfere with key positions in the host network, and plant organizes an in-depth defense by sequentially activating genes distal to the effector targets.

Gene expression systems in corynebacteria.

PubMed

Srivastava, Preeti; Deb, J K

2005-04-01

Corynebacterium belongs to a group of gram-positive bacteria having moderate to high G+C content, the other members being Mycobacterium, Nocardia, and Rhodococcus. Considerable information is now available on the plasmids, gene regulatory elements, and gene expression in corynebacteria, especially in soil corynebacteria such as Corynebacterium glutamicum. These bacteria are non-pathogenic and, unlike Bacillus and Streptomyces, are low in proteolytic activity and thus have the potential of becoming attractive systems for expression of heterologous proteins. This review discusses recent advances in our understanding of the organization of various regulatory elements, such as promoters, transcription terminators, and development of vectors for cloning and gene expression.

In vivo expression of Salmonella enterica serotype Typhi genes in the blood of patients with typhoid fever in Bangladesh.

PubMed

Sheikh, Alaullah; Charles, Richelle C; Sharmeen, Nusrat; Rollins, Sean M; Harris, Jason B; Bhuiyan, Md Saruar; Arifuzzaman, Mohammad; Khanam, Farhana; Bukka, Archana; Kalsy, Anuj; Porwollik, Steffen; Leung, Daniel T; Brooks, W Abdullah; LaRocque, Regina C; Hohmann, Elizabeth L; Cravioto, Alejandro; Logvinenko, Tanya; Calderwood, Stephen B; McClelland, Michael; Graham, James E; Qadri, Firdausi; Ryan, Edward T

2011-12-01

Salmonella enterica serotype Typhi is the cause of typhoid fever. It is a human-restricted pathogen, and few data exist on S. Typhi gene expression in humans. We applied an RNA capture and amplification technique, Selective Capture of Transcribed Sequences (SCOTS), and microarray hybridization to identify S. Typhi transcripts expressed in the blood of five humans infected with S. Typhi in Bangladesh. In total, we detected the expression of mRNAs for 2,046 S. Typhi genes (44% of the S. Typhi genome) in human blood; expression of 912 genes was detected in all 5 patients, and expression of 1,100 genes was detected in 4 or more patients. Identified transcripts were associated with the virulence-associated PhoP regulon, Salmonella pathogenicity islands, the use of alternative carbon and energy sources, synthesis and transport of iron, thiamine, and biotin, and resistance to antimicrobial peptides and oxidative stress. The most highly represented group were genes currently annotated as encoding proteins designated as hypothetical, unknown, or unclassified. Of the 2,046 detected transcripts, 1,320 (29% of the S. Typhi genome) had significantly different levels of detection in human blood compared to in vitro cultures; detection of 141 transcripts was significantly different in all 5 patients, and detection of 331 transcripts varied in at least 4 patients. These mRNAs encode proteins of unknown function, those involved in energy metabolism, transport and binding, cell envelope, cellular processes, and pathogenesis. We confirmed increased expression of a subset of identified mRNAs by quantitative-PCR. We report the first characterization of bacterial transcriptional profiles in the blood of patients with typhoid fever. S. Typhi is an important global pathogen whose restricted host range has greatly inhibited laboratory studies. Our results suggest that S. Typhi uses a largely uncharacterized genetic repertoire to survive within cells and utilize alternate energy sources during infection.

Insights into organ-specific pathogen defense responses in plants: RNA-seq analysis of potato tuber-Phytophthora infestans interactions.

PubMed

Gao, Liangliang; Tu, Zheng Jin; Millett, Benjamin P; Bradeen, James M

2013-05-23

The late blight pathogen Phytophthora infestans can attack both potato foliage and tubers. Although interaction transcriptome dynamics between potato foliage and various pathogens have been reported, no transcriptome study has focused specifically upon how potato tubers respond to pathogen infection. When inoculated with P. infestans, tubers of nontransformed 'Russet Burbank' (WT) potato develop late blight disease while those of transgenic 'Russet Burbank' line SP2211 (+RB), which expresses the potato late blight resistance gene RB (Rpi-blb1), do not. We compared transcriptome responses to P. infestans inoculation in tubers of these two lines. We demonstrated the practicality of RNA-seq to study tetraploid potato and present the first RNA-seq study of potato tuber diseases. A total of 483 million paired end Illumina RNA-seq reads were generated, representing the transcription of around 30,000 potato genes. Differentially expressed genes, gene groups and ontology bins that exhibited differences between the WT and +RB lines were identified. P. infestans transcripts, including those of known effectors, were also identified. Faster and stronger activation of defense related genes, gene groups and ontology bins correlate with successful tuber resistance against P. infestans. Our results suggest that the hypersensitive response is likely a general form of resistance against the hemibiotrophic P. infestans-even in potato tubers, organs that develop below ground.

Insights into organ-specific pathogen defense responses in plants: RNA-seq analysis of potato tuber-Phytophthora infestans interactions

PubMed Central

2013-01-01

Background The late blight pathogen Phytophthora infestans can attack both potato foliage and tubers. Although interaction transcriptome dynamics between potato foliage and various pathogens have been reported, no transcriptome study has focused specifically upon how potato tubers respond to pathogen infection. When inoculated with P. infestans, tubers of nontransformed ‘Russet Burbank’ (WT) potato develop late blight disease while those of transgenic ‘Russet Burbank’ line SP2211 (+RB), which expresses the potato late blight resistance gene RB (Rpi-blb1), do not. We compared transcriptome responses to P. infestans inoculation in tubers of these two lines. Results We demonstrated the practicality of RNA-seq to study tetraploid potato and present the first RNA-seq study of potato tuber diseases. A total of 483 million paired end Illumina RNA-seq reads were generated, representing the transcription of around 30,000 potato genes. Differentially expressed genes, gene groups and ontology bins that exhibited differences between the WT and +RB lines were identified. P. infestans transcripts, including those of known effectors, were also identified. Conclusion Faster and stronger activation of defense related genes, gene groups and ontology bins correlate with successful tuber resistance against P. infestans. Our results suggest that the hypersensitive response is likely a general form of resistance against the hemibiotrophic P. infestans—even in potato tubers, organs that develop below ground. PMID:23702331

FvSNF1, the sucrose non-fermenting protein kinase gene of Fusarium virguliforme, is required for cell-wall-degrading enzymes expression and sudden death syndrome development in soybean.

PubMed

Islam, Kazi T; Bond, Jason P; Fakhoury, Ahmad M

2017-08-01

Fusarium virguliforme is a soil-borne pathogenic fungus that causes sudden death syndrome (SDS) in soybean. Its pathogenicity is believed to require the activity of cell-wall-degrading enzymes (CWDEs). The sucrose non-fermenting protein kinase 1 gene (SNF1) is a key component of the glucose de-repression pathway in yeast, and a regulator of gene expression for CWDEs in some plant pathogenic fungi. To elucidate the functional role of the SNF1 homolog in F. virguliforme, FvSNF1 was disrupted using a split-marker strategy. Disruption of FvSNF1 in F. virguliforme abolishes galactose utilization and causes poor growth on xylose, arabinose and sucrose. However, the resulting Fvsnf1 mutant grew similar to wild-type and ectopic transformants on glucose, fructose, maltose, or pectin as the main source of carbon. The Fvsnf1 mutant displayed no expression of the gene-encoding galactose oxidase (GAO), a secretory enzyme that catalyzes oxidation of D-galactose. It also exhibited a significant reduction in the expression of several CWDE-coding genes in contrast to the wild-type strain. Greenhouse pathogenicity assays revealed that the Fvsnf1 mutant was severely impaired in its ability to cause SDS on challenged soybean plants. Microscopy and microtome studies on infected roots showed that the Fvsnf1 mutant was defective in colonizing vascular tissue of infected plants. Cross and longitudinal sections of infected roots stained with fluorescein-labeled wheat germ agglutinin and Congo red showed that the Fvsnf1 mutant failed to colonize the xylem vessels and phloem tissue at later stages of infection. Quantification of the fungal biomass in inoculated roots further confirmed a reduced colonization of roots by the Fvsnf1 mutant when compared to the wild type. These findings suggest that FvSNF1 regulates the expression of CWDEs in F. virguliforme, thus affecting the virulence of the fungus on soybean.

Human macrophage gamma interferon decreases gene expression but not replication of Mycobacterium tuberculosis: analysis of the host-pathogen reciprocal influence on transcription in a comparison of strains H37Rv and CMT97.

PubMed

Cappelli, G; Volpe, P; Sanduzzi, A; Sacchi, A; Colizzi, V; Mariani, F

2001-12-01

Mycobacterium tuberculosis is an intracellular pathogen that readily survives and replicates in human macrophages (MPhi). Host cells have developed different mycobactericidal mechanisms, including the production of inflammatory cytokines. The aim of this study was to compare the MPhi response, in terms of cytokine gene expression, to infection with the M. tuberculosis laboratory strain H37Rv and the clinical M. tuberculosis isolate CMT97. Both strains induce the production of interleukin-12 (IL-12) and IL-16 at comparable levels. However, the clinical isolate induces a significantly higher and more prolonged MPhi activation, as shown by reverse transcription-PCR analysis of IL-1beta, IL-6, IL-10, transforming growth factor beta, tumor necrosis factor alpha, and gamma interferon (IFN-gamma) transcripts. Interestingly, when IFN-gamma transcription is high, the number of M. tuberculosis genes expressed decreases and vice versa, whereas no mycobactericidal effect was observed in terms of bacterial growth. Expression of 11 genes was also studied in the two M. tuberculosis strains by infecting resting or activated MPhi and compared to bacterial intracellular survival. In both cases, a peculiar inverse correlation between expression of these genes and multiplication was observed. The number and type of genes expressed by the two strains differed significantly.

Transcriptomic profiling of microbe-microbe interactions reveals the specific response of the biocontrol strain P. fluorescens In5 to the phytopathogen Rhizoctonia solani.

PubMed

Hennessy, Rosanna C; Glaring, Mikkel A; Olsson, Stefan; Stougaard, Peter

2017-08-10

Few studies to date report the transcriptional response of biocontrol bacteria toward phytopathogens. In order to gain insights into the potential mechanism underlying the antagonism of the antimicrobial producing strain P. fluorescens In5 against the phytopathogens Rhizoctonia solani and Pythium aphanidermatum, global RNA sequencing was performed. Differential gene expression profiling of P. fluorescens In5 in response to either R. solani or P. aphanidermatum was investigated using transcriptome sequencing (RNA-seq). Total RNA was isolated from single bacterial cultures of P. fluorescens In5 or bacterial cultures in dual-culture for 48 h with each pathogen in biological triplicates. RNA-seq libraries were constructed following a default Illumina stranded RNA protocol including rRNA depletion and were sequenced 2 × 100 bases on Illumina HiSeq generating approximately 10 million reads per sample. No significant changes in global gene expression were recorded during dual-culture of P. fluorescens In5 with any of the two pathogens but rather each pathogen appeared to induce expression of a specific set of genes. A particularly strong transcriptional response to R. solani was observed and notably several genes possibly associated with secondary metabolite detoxification and metabolism were highly upregulated in response to the fungus. A total of 23 genes were significantly upregulated and seven genes were significantly downregulated with at least respectively a threefold change in expression level in response to R. solani compared to the no fungus control. In contrast, only one gene was significantly upregulated over threefold and three transcripts were significantly downregulated over threefold in response to P. aphanidermatum. Genes known to be involved in synthesis of secondary metabolites, e.g. non-ribosomal synthetases and hydrogen cyanide were not differentially expressed at the time points studied. This study demonstrates that genes possibly involved in metabolite detoxification are highly upregulated in P. fluorescens In5 when co-cultured with plant pathogens and in particular the fungus R. solani. This highlights the importance of studying microbe-microbe interactions to gain a better understanding of how different systems function in vitro and ultimately in natural systems where biocontrol agents can be used for the sustainable management of plant diseases.

GhMAP3K65, a Cotton Raf-Like MAP3K Gene, Enhances Susceptibility to Pathogen Infection and Heat Stress by Negatively Modulating Growth and Development in Transgenic Nicotiana benthamiana.

PubMed

Zhai, Na; Jia, Haihong; Liu, Dongdong; Liu, Shuchang; Ma, Manli; Guo, Xingqi; Li, Han

2017-11-21

Mitogen-activated protein kinase kinase kinases (MAP3Ks), the top components of MAPK cascades, modulate many biological processes, such as growth, development and various environmental stresses. Nevertheless, the roles of MAP3Ks remain poorly understood in cotton. In this study, GhMAP3K65 was identified in cotton, and its transcription was inducible by pathogen infection, heat stress, and multiple signalling molecules. Silencing of GhMAP3K65 enhanced resistance to pathogen infection and heat stress in cotton. In contrast, overexpression of GhMAP3K65 enhanced susceptibility to pathogen infection and heat stress in transgenic Nicotiana benthamiana . The expression of defence-associated genes was activated in transgenic N. benthamiana plants after pathogen infection and heat stress, indicating that GhMAP3K65 positively regulates plant defence responses. Nevertheless, transgenic N. benthamiana plants impaired lignin biosynthesis and stomatal immunity in their leaves and repressed vitality of their root systems. In addition, the expression of lignin biosynthesis genes and lignin content were inhibited after pathogen infection and heat stress. Collectively, these results demonstrate that GhMAP3K65 enhances susceptibility to pathogen infection and heat stress by negatively modulating growth and development in transgenic N. benthamiana plants.

Effect of wheat roots infected with the pathogenic fungus Gaeumannomyces graminis var. tritici on gene expression of the biocontrol bacterium Pseudomonas fluorescens Pf29Arp.

PubMed

Barret, Matthieu; Frey-Klett, Pascale; Guillerm-Erckelboudt, Anne-Yvonne; Boutin, Morgane; Guernec, Gregory; Sarniguet, Alain

2009-12-01

Traits contributing to the competence of biocontrol bacteria to colonize plant roots are often induced in the rhizosphere in response to plant components. These interactions have been studied using the two partners in gnotobiotic systems. However, in nature, beneficial or pathogenic fungi often colonize roots. Influence of these plant-fungus interactions on bacterial behavior remains to be investigated. Here, we have examined the influence of colonization of wheat roots by the take-all fungus Gaeumannomyces graminis var. tritici on gene expression of the biocontrol bacterium Pseudomonas fluorescens Pf29Arp. Bacteria were inoculated onto healthy, early G. graminis var. tritici-colonized and necrotic roots and transcriptomes were compared by shotgun DNA microarray. Pf29Arp decreased disease severity when inoculated before the onset of necrosis. Necrotic roots exerted a broader effect on gene expression compared with early G. graminis var. tritici-colonized and healthy roots. A gene encoding a putative type VI secretion system effector was only induced in necrotic conditions. A common pool of Pf29Arp genes differentially expressed on G. graminis var. tritici-colonized roots was related to carbon metabolism and oxidative stress, with a highest fold-change with necrosis. Overall, the data showed that the association of the pathogenic fungus with the roots strongly altered Pf29Arp adaptation with differences between early and late G. graminis var. tritici infection steps.

Transcriptome analysis of the fungal pathogen Fusarium oxysporum f. sp. medicaginis during colonisation of resistant and susceptible Medicago truncatula hosts identifies differential pathogenicity profiles and novel candidate effectors.

PubMed

Thatcher, Louise F; Williams, Angela H; Garg, Gagan; Buck, Sally-Anne G; Singh, Karam B

2016-11-03

Pathogenic members of the Fusarium oxysporum species complex are responsible for vascular wilt disease on many important crops including legumes, where they can be one of the most destructive disease causing necrotrophic fungi. We previously developed a model legume-infecting pathosystem based on the reference legume Medicago truncatula and a pathogenic F. oxysporum forma specialis (f. sp.) medicaginis (Fom). To dissect the molecular pathogenicity arsenal used by this root-infecting pathogen, we sequenced its transcriptome during infection of a susceptible and resistant host accession. High coverage RNA-Seq of Fom infected root samples harvested from susceptible (DZA315) or resistant (A17) M. truncatula seedlings at early or later stages of infection (2 or 7 days post infection (dpi)) and from vegetative (in vitro) samples facilitated the identification of unique and overlapping sets of in planta differentially expressed genes. This included enrichment, particularly in DZA315 in planta up-regulated datasets, for proteins associated with sugar, protein and plant cell wall metabolism, membrane transport, nutrient uptake and oxidative processes. Genes encoding effector-like proteins were identified, including homologues of the F. oxysporum f. sp. lycopersici Secreted In Xylem (SIX) proteins, and several novel candidate effectors based on predicted secretion, small protein size and high in-planta induced expression. The majority of the effector candidates contain no known protein domains but do share high similarity to predicted proteins predominantly from other F. oxysporum ff. spp. as well as other Fusaria (F. solani, F. fujikori, F. verticilloides, F. graminearum and F. pseudograminearum), and from another wilt pathogen of the same class, a Verticillium species. Overall, this suggests these novel effector candidates may play important roles in Fusaria and wilt pathogen virulence. Combining high coverage in planta RNA-Seq with knowledge of fungal pathogenicity protein features facilitated the identification of differentially expressed pathogenicity associated genes and novel effector candidates expressed during infection of a resistant or susceptible M. truncatula host. The knowledge from this first in depth in planta transcriptome sequencing of any F. oxysporum ff. spp. pathogenic on legumes will facilitate the dissection of Fusarium wilt pathogenicity mechanisms on many important legume crops.

Dual-species transcriptional profiling during systemic candidiasis reveals organ-specific host-pathogen interactions.

PubMed

Hebecker, Betty; Vlaic, Sebastian; Conrad, Theresia; Bauer, Michael; Brunke, Sascha; Kapitan, Mario; Linde, Jörg; Hube, Bernhard; Jacobsen, Ilse D

2016-11-03

Candida albicans is a common cause of life-threatening fungal bloodstream infections. In the murine model of systemic candidiasis, the kidney is the primary target organ while the fungal load declines over time in liver and spleen. To better understand these organ-specific differences in host-pathogen interaction, we performed gene expression profiling of murine kidney, liver and spleen and determined the fungal transcriptome in liver and kidney. We observed a delayed transcriptional immune response accompanied by late induction of fungal stress response genes in the kidneys. In contrast, early upregulation of the proinflammatory response in the liver was associated with a fungal transcriptome resembling response to phagocytosis, suggesting that phagocytes contribute significantly to fungal control in the liver. Notably, C. albicans hypha-associated genes were upregulated in the absence of visible filamentation in the liver, indicating an uncoupling of gene expression and morphology and a morphology-independent effect by hypha-associated genes in this organ. Consistently, integration of host and pathogen transcriptional data in an inter-species gene regulatory network indicated connections of C. albicans cell wall remodelling and metabolism to the organ-specific immune responses.

Dual-species transcriptional profiling during systemic candidiasis reveals organ-specific host-pathogen interactions

PubMed Central

Hebecker, Betty; Vlaic, Sebastian; Conrad, Theresia; Bauer, Michael; Brunke, Sascha; Kapitan, Mario; Linde, Jörg; Hube, Bernhard; Jacobsen, Ilse D.

2016-01-01

Candida albicans is a common cause of life-threatening fungal bloodstream infections. In the murine model of systemic candidiasis, the kidney is the primary target organ while the fungal load declines over time in liver and spleen. To better understand these organ-specific differences in host-pathogen interaction, we performed gene expression profiling of murine kidney, liver and spleen and determined the fungal transcriptome in liver and kidney. We observed a delayed transcriptional immune response accompanied by late induction of fungal stress response genes in the kidneys. In contrast, early upregulation of the proinflammatory response in the liver was associated with a fungal transcriptome resembling response to phagocytosis, suggesting that phagocytes contribute significantly to fungal control in the liver. Notably, C. albicans hypha-associated genes were upregulated in the absence of visible filamentation in the liver, indicating an uncoupling of gene expression and morphology and a morphology-independent effect by hypha-associated genes in this organ. Consistently, integration of host and pathogen transcriptional data in an inter-species gene regulatory network indicated connections of C. albicans cell wall remodelling and metabolism to the organ-specific immune responses. PMID:27808111

Lemna (duckweed) expressed hemagglutinin from avian influenza H5N1 protects chickens against H5N1 high pathogenicity avian influenza virus challenge

USDA-ARS?s Scientific Manuscript database

In the last two decades, transgenic plants have been explored as safe and cost effective alternative expression platforms for producing recombinant proteins. In this study, a synthetic hemagglutinin (HA) gene from the high pathogenicity avian influenza (HPAI) virus A/chicken/Indonesia/7/2003 (H5N1)...

Intercellular and intracellular signalling systems that globally control the expression of virulence genes in plant pathogenic bacteria.

PubMed

Ham, Jong Hyun

2013-04-01

Plant pathogenic bacteria utilize complex signalling systems to control the expression of virulence genes at the cellular level and within populations. Quorum sensing (QS), an important intercellular communication mechanism, is mediated by different types of small molecules, including N-acyl homoserine lactones (AHLs), fatty acids and small proteins. AHL-mediated signalling systems dependent on the LuxI and LuxR family proteins play critical roles in the virulence of a wide range of Gram-negative plant pathogenic bacteria belonging to the Alphaproteobacteria, Betaproteobacteria and Gammaproteobacteria. Xanthomonas spp. and Xylella fastidiosa, members of the Gammaproteobacteria, however, possess QS systems that are mediated by fatty acid-type diffusible signal factors (DSFs). Recent studies have demonstrated that Ax21, a 194-amino-acid protein in Xanthomonas oryzae pv. oryzae, plays dual functions in activating a rice innate immune pathway through binding to the rice XA21 pattern recognition receptor and in regulating bacterial virulence and biofilm formation as a QS signal molecule. In xanthomonads, DSF-mediated QS systems are connected with the signalling pathways mediated by cyclic diguanosine monophosphate (c-di-GMP), which functions as a second messenger for the control of virulence gene expression in these bacterial pathogens. © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

Expression profiling of marker genes responsive to the defence-associated phytohormones salicylic acid, jasmonic acid and ethylene in Brachypodium distachyon.

PubMed

Kouzai, Yusuke; Kimura, Mamiko; Yamanaka, Yurie; Watanabe, Megumi; Matsui, Hidenori; Yamamoto, Mikihiro; Ichinose, Yuki; Toyoda, Kazuhiro; Onda, Yoshihiko; Mochida, Keiichi; Noutoshi, Yoshiteru

2016-03-02

Brachypodium distachyon is a promising model plants for grasses. Infections of Brachypodium by various pathogens that severely impair crop production have been reported, and the species accordingly provides an alternative platform for investigating molecular mechanisms of pathogen virulence and plant disease resistance. To date, we have a broad picture of plant immunity only in Arabidopsis and rice; therefore, Brachypodium may constitute a counterpart that displays the commonality and uniqueness of defence systems among plant species. Phytohormones play key roles in plant biotic stress responses, and hormone-responsive genes are used to qualitatively and quantitatively evaluate disease resistance responses during pathogen infection. For these purposes, defence-related phytohormone marker genes expressed at time points suitable for defence-response monitoring are needed. Information about their expression profiles over time as well as their response specificity is also helpful. However, useful marker genes are still rare in Brachypodium. We selected 34 candidates for Brachypodium marker genes on the basis of protein-sequence similarity to known marker genes used in Arabidopsis and rice. Brachypodium plants were treated with the defence-related phytohormones salicylic acid, jasmonic acid and ethylene, and their transcription levels were measured 24 and 48 h after treatment. Two genes for salicylic acid, 7 for jasmonic acid and 2 for ethylene were significantly induced at either or both time points. We then focused on 11 genes encoding pathogenesis-related (PR) 1 protein and compared their expression patterns with those of Arabidopsis and rice. Phylogenetic analysis suggested that Brachypodium contains several PR1-family genes similar to rice genes. Our expression profiling revealed that regulation patterns of some PR1 genes as well as of markers identified for defence-related phytohormones are closely related to those in rice. We propose that the Brachypodium immune hormone marker genes identified in this study will be useful to plant pathologists who use Brachypodium as a model pathosystem, because the timing of their transcriptional activation matches that of the disease resistance response. Our results using Brachypodium also suggest that monocots share a characteristic immune system, defined as the common defence system, that is different from that of dicots.

Ca2+/Calmodulin-Dependent AtSR1/CAMTA3 Plays Critical Roles in Balancing Plant Growth and Immunity.

PubMed

Yuan, Peiguo; Du, Liqun; Poovaiah, B W

2018-06-14

During plant-pathogen interactions, plants have to relocate their resources including energy to defend invading organisms; as a result, plant growth and development are usually reduced. Arabidopsis signal responsive1 (AtSR1) has been documented as a negative regulator of plant immune responses and could serve as a positive regulator of plant growth and development. However, the mechanism by which AtSR1 balances plant growth and immunity is poorly understood. Here, we performed a global gene expression profiling using Affymetrix microarrays to study how AtSR1 regulates defense- and growth-related genes in plants with and without bacterial pathogen infection. Results revealed that AtSR1 negatively regulates most of the immune-related genes involved in molecular pattern-triggered immunity (PTI), effector-triggered immunity (ETI), and in salicylic acid (SA)- and jasmonate (JA)-mediated signaling pathways. AtSR1 may rigidly regulate several steps of the SA-mediated pathway, from the activation of SA synthesis to the perception of SA signal. Furthermore, AtSR1 may also regulate plant growth through its involvement in regulating auxin- and BRs-related pathways. Although microarray data revealed that expression levels of defense-related genes induced by pathogens are higher in wild-type (WT) plants than that in atsr1 mutant plants, WT plants are more susceptible to the infection of virulent pathogen as compared to atsr1 mutant plants. These observations indicate that the AtSR1 functions in suppressing the expression of genes induced by pathogen attack and contributes to the rapid establishment of resistance in WT background. Results of electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP)-PCR assays suggest that AtSR1 acts as transcription factor in balancing plant growth and immunity, through interaction with the “CGCG” containing CG-box in the promotors of its target genes.

Retransformation of marker-free potato for enhanced resistance against fungal pathogens by pyramiding chitinase and wasabi defensin genes.

PubMed

Khan, Raham Sher; Darwish, Nader Ahmed; Khattak, Bushra; Ntui, Valentine Otang; Kong, Kynet; Shimomae, Kazuki; Nakamura, Ikuo; Mii, Masahiro

2014-09-01

Multi-auto-transformation vector system has been one of the strategies to produce marker-free transgenic plants without using selective chemicals and plant growth regulators and thus facilitating transgene stacking. In the study reported here, retransformation was carried out in marker-free transgenic potato CV. May Queen containing ChiC gene (isolated from Streptomyces griseus strain HUT 6037) with wasabi defensin (WD) gene (isolated from Wasabia japonica) to pyramid the two disease resistant genes. Molecular analyses of the developed shoots confirmed the existence of both the genes of interest (ChiC and WD) in transgenic plants. Co-expression of the genes was confirmed by RT-PCR, northern blot, and western blot analyses. Disease resistance assay of in vitro plants showed that the transgenic lines co-expressing both the ChiC and WD genes had higher resistance against the fungal pathogens, Fusarium oxysporum (Fusarium wilt) and Alternaria solani (early blight) compared to the non-transformed control and the transgenic lines expressing either of the ChiC or WD genes. The disease resistance potential of the transgenic plants could be increased by transgene stacking or multiple transformations.

Identification of host genes leading to West Nile virus encephalitis in mice brain using RNA-seq analysis

PubMed Central

Kumar, Mukesh; Belcaid, Mahdi; Nerurkar, Vivek R.

2016-01-01

Differential host responses may be critical determinants of distinct pathologies of West Nile virus (WNV) NY99 (pathogenic) and WNV Eg101 (non-pathogenic) strains. We employed RNA-seq technology to analyze global differential gene expression in WNV-infected mice brain and to identify the host cellular factors leading to lethal encephalitis. We identified 1,400 and 278 transcripts, which were differentially expressed after WNV NY99 and WNV Eg101 infections, respectively, and 147 genes were common to infection with both the viruses. Genes that were up-regulated in infection with both the viruses were mainly associated with interferon signaling. Genes associated with inflammation and cell death/apoptosis were only expressed after WNV NY99 infection. We demonstrate that differences in the activation of key pattern recognition receptors resulted in the induction of unique innate immune profiles, which corresponded with the induction of interferon and inflammatory responses. Pathway analysis of differentially expressed genes indicated that after WNV NY99 infection, TREM-1 mediated activation of toll-like receptors leads to the high inflammatory response. In conclusion, we have identified both common and specific responses to WNV NY99 and WNV Eg101 infections as well as genes linked to potential resistance to infection that may be targets for therapeutics. PMID:27211830

Identification of differentially expressed genes associated with changes in the morphology of Pichia fermentans on apple and peach fruit.

PubMed

Fiori, Stefano; Scherm, Barbara; Liu, Jia; Farrell, Robert; Mannazzu, Ilaria; Budroni, Marilena; Maserti, Bianca E; Wisniewski, Michael E; Migheli, Quirico

2012-11-01

Pichia fermentans (strain DISAABA 726) is an effective biocontrol agent against Monilinia fructicola and Botrytis cinerea when inoculated in artificially wounded apple fruit but is an aggressive pathogen when inoculated on wounded peach fruit, causing severe fruit decay. Pichia fermentans grows as budding yeast on apple tissue and exhibits pseudohyphal growth on peach tissue, suggesting that dimorphism may be associated with pathogenicity. Two complementary suppressive subtractive hybridization (SSH) strategies, that is, rapid subtraction hybridization (RaSH) and PCR-based subtraction, were performed to identify genes differentially expressed by P. fermentans after 24-h growth on apple vs. peach fruit. Gene products that were more highly expressed on peach than on apple tissue, or vice versa, were sequenced and compared with available yeast genome sequence databases. Several of the genes more highly expressed, when P. fermentans was grown on peach, were related to stress response, glycolysis, amino acid metabolism, and alcoholic fermentation but surprisingly not to cell wall degrading enzymes such as pectinases or cellulases. The dual activity of P. fermentans as both a biocontrol agent and a pathogen emphasizes the need for a thorough risk analysis of potential antagonists to avoid unpredictable results that could negatively impact the safe use of postharvest biocontrol strategies. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Spleen transcriptome response to infection with avian pathogenic Escherichia coli in broiler chickens

PubMed Central

2011-01-01

Background Avian pathogenic Escherichia coli (APEC) is detrimental to poultry health and its zoonotic potential is a food safety concern. Regulation of antimicrobials in food-production animals has put greater focus on enhancing host resistance to bacterial infections through genetics. To better define effective mechanism of host resistance, global gene expression in the spleen of chickens, harvested at two times post-infection (PI) with APEC, was measured using microarray technology, in a design that will enable investigation of effects of vaccination, challenge, and pathology level. Results There were 1,101 genes significantly differentially expressed between severely infected and non-infected groups on day 1 PI and 1,723 on day 5 PI. Very little difference was seen between mildly infected and non-infected groups on either time point. Between birds exhibiting mild and severe pathology, there were 2 significantly differentially expressed genes on day 1 PI and 799 on day 5 PI. Groups with greater pathology had more genes with increased expression than decreased expression levels. Several predominate immune pathways, Toll-like receptor, Jak-STAT, and cytokine signaling, were represented between challenged and non-challenged groups. Vaccination had, surprisingly, no detectible effect on gene expression, although it significantly protected the birds from observable gross lesions. Functional characterization of significantly expressed genes revealed unique gene ontology classifications during each time point, with many unique to a particular treatment or class contrast. Conclusions More severe pathology caused by APEC infection was associated with a high level of gene expression differences and increase in gene expression levels. Many of the significantly differentially expressed genes were unique to a particular treatment, pathology level or time point. The present study not only investigates the transcriptomic regulations of APEC infection, but also the degree of pathology associated with that infection. This study will allow for greater discovery into host mechanisms for disease resistance, providing targets for marker assisted selection and advanced drug development. PMID:21951686

Spleen transcriptome response to infection with avian pathogenic Escherichia coli in broiler chickens.

PubMed

Sandford, Erin E; Orr, Megan; Balfanz, Emma; Bowerman, Nate; Li, Xianyao; Zhou, Huaijun; Johnson, Timothy J; Kariyawasam, Subhashinie; Liu, Peng; Nolan, Lisa K; Lamont, Susan J

2011-09-27

Avian pathogenic Escherichia coli (APEC) is detrimental to poultry health and its zoonotic potential is a food safety concern. Regulation of antimicrobials in food-production animals has put greater focus on enhancing host resistance to bacterial infections through genetics. To better define effective mechanism of host resistance, global gene expression in the spleen of chickens, harvested at two times post-infection (PI) with APEC, was measured using microarray technology, in a design that will enable investigation of effects of vaccination, challenge, and pathology level. There were 1,101 genes significantly differentially expressed between severely infected and non-infected groups on day 1 PI and 1,723 on day 5 PI. Very little difference was seen between mildly infected and non-infected groups on either time point. Between birds exhibiting mild and severe pathology, there were 2 significantly differentially expressed genes on day 1 PI and 799 on day 5 PI. Groups with greater pathology had more genes with increased expression than decreased expression levels. Several predominate immune pathways, Toll-like receptor, Jak-STAT, and cytokine signaling, were represented between challenged and non-challenged groups. Vaccination had, surprisingly, no detectible effect on gene expression, although it significantly protected the birds from observable gross lesions. Functional characterization of significantly expressed genes revealed unique gene ontology classifications during each time point, with many unique to a particular treatment or class contrast. More severe pathology caused by APEC infection was associated with a high level of gene expression differences and increase in gene expression levels. Many of the significantly differentially expressed genes were unique to a particular treatment, pathology level or time point. The present study not only investigates the transcriptomic regulations of APEC infection, but also the degree of pathology associated with that infection. This study will allow for greater discovery into host mechanisms for disease resistance, providing targets for marker assisted selection and advanced drug development.

Competitive Performance of Transgenic Wheat Resistant to Powdery Mildew

PubMed Central

Kalinina, Olena; Zeller, Simon L.; Schmid, Bernhard

2011-01-01

Genetically modified (GM) plants offer an ideal model system to study the influence of single genes that confer constitutive resistance to pathogens on the ecological behaviour of plants. We used phytometers to study competitive interactions between GM lines of spring wheat Triticum aestivum carrying such genes and control lines. We hypothesized that competitive performance of GM lines would be reduced due to enhanced transgene expression under pathogen levels typically encountered in the field. The transgenes pm3b from wheat (resistance against powdery mildew Blumeria graminis) or chitinase and glucanase genes from barley (resistance against fungi in general) were introduced with the ubiquitin promoter from maize (pm3b and chitinase genes) or the actin promoter from rice (glucanase gene). Phytometers of 15 transgenic and non-transgenic wheat lines were transplanted as seedlings into plots sown with the same 15 lines as competitive environments and subject to two soil nutrient levels. Pm3b lines had reduced mildew incidence compared with control lines. Chitinase and chitinase/glucanase lines showed the same high resistance to mildew as their control in low-nutrient treatment and slightly lower mildew rates than the control in high-nutrient environment. Pm3b lines were weaker competitors than control lines. This resulted in reduced yield and seed number. The Pm3b line with the highest transgene expression had 53.2% lower yield than the control whereas the Pm3b line which segregated in resistance and had higher mildew rates showed only minor costs under competition. The line expressing both chitinase and glucanase genes also showed reduced yield and seed number under competition compared with its control. Our results suggest that single transgenes conferring constitutive resistance to pathogens can have ecological costs and can weaken plant competitiveness even in the presence of the pathogen. The magnitude of these costs appears related to the degree of expression of the transgenes. PMID:22132219

The Genome of Tolypocladium inflatum: Evolution, Organization, and Expression of the Cyclosporin Biosynthetic Gene Cluster

PubMed Central

Bushley, Kathryn E.; Raja, Rajani; Jaiswal, Pankaj; Cumbie, Jason S.; Nonogaki, Mariko; Boyd, Alexander E.; Owensby, C. Alisha; Knaus, Brian J.; Elser, Justin; Miller, Daniel; Di, Yanming; McPhail, Kerry L.; Spatafora, Joseph W.

2013-01-01

The ascomycete fungus Tolypocladium inflatum, a pathogen of beetle larvae, is best known as the producer of the immunosuppressant drug cyclosporin. The draft genome of T. inflatum strain NRRL 8044 (ATCC 34921), the isolate from which cyclosporin was first isolated, is presented along with comparative analyses of the biosynthesis of cyclosporin and other secondary metabolites in T. inflatum and related taxa. Phylogenomic analyses reveal previously undetected and complex patterns of homology between the nonribosomal peptide synthetase (NRPS) that encodes for cyclosporin synthetase (simA) and those of other secondary metabolites with activities against insects (e.g., beauvericin, destruxins, etc.), and demonstrate the roles of module duplication and gene fusion in diversification of NRPSs. The secondary metabolite gene cluster responsible for cyclosporin biosynthesis is described. In addition to genes necessary for cyclosporin biosynthesis, it harbors a gene for a cyclophilin, which is a member of a family of immunophilins known to bind cyclosporin. Comparative analyses support a lineage specific origin of the cyclosporin gene cluster rather than horizontal gene transfer from bacteria or other fungi. RNA-Seq transcriptome analyses in a cyclosporin-inducing medium delineate the boundaries of the cyclosporin cluster and reveal high levels of expression of the gene cluster cyclophilin. In medium containing insect hemolymph, weaker but significant upregulation of several genes within the cyclosporin cluster, including the highly expressed cyclophilin gene, was observed. T. inflatum also represents the first reference draft genome of Ophiocordycipitaceae, a third family of insect pathogenic fungi within the fungal order Hypocreales, and supports parallel and qualitatively distinct radiations of insect pathogens. The T. inflatum genome provides additional insight into the evolution and biosynthesis of cyclosporin and lays a foundation for further investigations of the role of secondary metabolite gene clusters and their metabolites in fungal biology. PMID:23818858

Transcriptome-wide analysis of WRKY transcription factors in wheat and their leaf rust responsive expression profiling.

PubMed

Satapathy, Lopamudra; Singh, Dharmendra; Ranjan, Prashant; Kumar, Dhananjay; Kumar, Manish; Prabhu, Kumble Vinod; Mukhopadhyay, Kunal

2014-12-01

WRKY, a plant-specific transcription factor family, has important roles in pathogen defense, abiotic cues and phytohormone signaling, yet little is known about their roles and molecular mechanism of function in response to rust diseases in wheat. We identified 100 TaWRKY sequences using wheat Expressed Sequence Tag database of which 22 WRKY sequences were novel. Identified proteins were characterized based on their zinc finger motifs and phylogenetic analysis clustered them into six clades consisting of class IIc and class III WRKY proteins. Functional annotation revealed major functions in metabolic and cellular processes in control plants; whereas response to stimuli, signaling and defense in pathogen inoculated plants, their major molecular function being binding to DNA. Tag-based expression analysis of the identified genes revealed differential expression between mock and Puccinia triticina inoculated wheat near isogenic lines. Gene expression was also performed with six rust-related microarray experiments at Gene Expression Omnibus database. TaWRKY10, 15, 17 and 56 were common in both tag-based and microarray-based differential expression analysis and could be representing rust specific WRKY genes. The obtained results will bestow insight into the functional characterization of WRKY transcription factors responsive to leaf rust pathogenesis that can be used as candidate genes in molecular breeding programs to improve biotic stress tolerance in wheat.

Genome sequence of the necrotrophic plant pathogen Pythium ultimum reveals original pathogenicity mechanisms and effector repertoire.

USDA-ARS?s Scientific Manuscript database

The P. ultimum DAOM BR144 (=CBS 805.95 = ATCC200006) genome (42.8 Mb) encodes 15,290 genes, and has extensive sequence similarity and synteny with related Phytophthora spp., including the potato late blight pathogen Phytophthora infestans. Whole transcriptome sequencing revealed expression of 86 % o...

The Wor1-like Protein Fgp1 Regulates Pathogenicity, Toxin Synthesis and Reproduction in the Phytopathogenic Fungus Fusarium graminearum

PubMed Central

Jonkers, Wilfried; Dong, Yanhong; Broz, Karen; Corby Kistler, H.

2012-01-01

WOR1 is a gene for a conserved fungal regulatory protein controlling the dimorphic switch and pathogenicity determents in Candida albicans and its ortholog in the plant pathogen Fusarium oxysporum, called SGE1, is required for pathogenicity and expression of key plant effector proteins. F. graminearum, an important pathogen of cereals, is not known to employ switching and no effector proteins from F. graminearum have been found to date that are required for infection. In this study, the potential role of the WOR1-like gene in pathogenesis was tested in this toxigenic fungus. Deletion of the WOR1 ortholog (called FGP1) in F. graminearum results in greatly reduced pathogenicity and loss of trichothecene toxin accumulation in infected wheat plants and in vitro. The loss of toxin accumulation alone may be sufficient to explain the loss of pathogenicity to wheat. Under toxin-inducing conditions, expression of genes for trichothecene biosynthesis and many other genes are not detected or detected at lower levels in Δfgp1 strains. FGP1 is also involved in the developmental processes of conidium formation and sexual reproduction and modulates a morphological change that accompanies mycotoxin production in vitro. The Wor1-like proteins in Fusarium species have highly conserved N-terminal regions and remarkably divergent C-termini. Interchanging the N- and C- terminal portions of proteins from F. oxysporum and F. graminearum resulted in partial to complete loss of function. Wor1-like proteins are conserved but have evolved to regulate pathogenicity in a range of fungi, likely by adaptations to the C-terminal portion of the protein. PMID:22693448

Defects in the C. elegans acyl-CoA Synthase, acs-3, and Nuclear Hormone Receptor, nhr-25, Cause Sensitivity to Distinct, but Overlapping Stresses

PubMed Central

Ward, Jordan D.; Mullaney, Brendan; Schiller, Benjamin J.; He, Le D.; Petnic, Sarah E.; Couillault, Carole; Pujol, Nathalie; Bernal, Teresita U.; Van Gilst, Marc R.; Ashrafi, Kaveh; Ewbank, Jonathan J.; Yamamoto, Keith R.

2014-01-01

Metazoan transcription factors control distinct networks of genes in specific tissues, yet understanding how these networks are integrated into physiology, development, and homeostasis remains challenging. Inactivation of the nuclear hormone receptor nhr-25 ameliorates developmental and metabolic phenotypes associated with loss of function of an acyl-CoA synthetase gene, acs-3. ACS-3 activity prevents aberrantly high NHR-25 activity. Here, we investigated this relationship further by examining gene expression patterns following acs-3 and nhr-25 inactivation. Unexpectedly, we found that the acs-3 mutation or nhr-25 RNAi resulted in similar transcriptomes with enrichment in innate immunity and stress response gene expression. Mutants of either gene exhibited distinct sensitivities to pathogens and environmental stresses. Only nhr-25 was required for wild-type levels of resistance to the bacterial pathogen P. aeruginosa and only acs-3 was required for wild-type levels of resistance to osmotic stress and the oxidative stress generator, juglone. Inactivation of either acs-3 or nhr-25 compromised lifespan and resistance to the fungal pathogen D. coniospora. Double mutants exhibited more severe defects in the lifespan and P. aeruginosa assays, but were similar to the single mutants in other assays. Finally, acs-3 mutants displayed defects in their epidermal surface barrier, potentially accounting for the observed sensitivities. Together, these data indicate that inactivation of either acs-3 or nhr-25 causes stress sensitivity and increased expression of innate immunity/stress genes, most likely by different mechanisms. Elevated expression of these immune/stress genes appears to abrogate the transcriptional signatures relevant to metabolism and development. PMID:24651852

Expression of the insect metalloproteinase inhibitor IMPI in the fat body of Galleria mellonella exposed to infection with Beauveria bassiana.

PubMed

Vertyporokh, Lidiia; Wojda, Iwona

2017-01-01

The inducible metalloproteinase inhibitor (IMPI) discovered in Galleria mellonella is currently the only specific inhibitor of metalloproteinases found in animals. Its role is to inhibit the activity of metalloproteinases secreted by pathogenic organisms as virulence factors to degrade immune-relevant polypeptides of the infected host. This is a good example of an evolutionary arms race between the insect hosts and their natural pathogens. In this report, we analyze the expression of a gene encoding an inducible metalloproteinase inhibitor (IMPI) in fat bodies of the greater wax moth larvae Galleria mellonella infected with an entomopathogenic fungus Beauveria bassiana. We have used a natural infection, i.e. covering larval integument with fungal aerospores, as well as injection of fungal blastospores directly into the larval hemocel. We compare the expression of IMPI with the expression of genes encoding proteins with fungicidal activity, gallerimycin and galiomycin, whose expression reflects the stimulation of Galleria mellonella defense mechanisms. Also, gene expression is analyzed in the light of survival of animals after spore injection.

An emerging cyberinfrastructure for biodefense pathogen and pathogen–host data

PubMed Central

Zhang, C.; Crasta, O.; Cammer, S.; Will, R.; Kenyon, R.; Sullivan, D.; Yu, Q.; Sun, W.; Jha, R.; Liu, D.; Xue, T.; Zhang, Y.; Moore, M.; McGarvey, P.; Huang, H.; Chen, Y.; Zhang, J.; Mazumder, R.; Wu, C.; Sobral, B.

2008-01-01

The NIAID-funded Biodefense Proteomics Resource Center (RC) provides storage, dissemination, visualization and analysis capabilities for the experimental data deposited by seven Proteomics Research Centers (PRCs). The data and its publication is to support researchers working to discover candidates for the next generation of vaccines, therapeutics and diagnostics against NIAID's Category A, B and C priority pathogens. The data includes transcriptional profiles, protein profiles, protein structural data and host–pathogen protein interactions, in the context of the pathogen life cycle in vivo and in vitro. The database has stored and supported host or pathogen data derived from Bacillus, Brucella, Cryptosporidium, Salmonella, SARS, Toxoplasma, Vibrio and Yersinia, human tissue libraries, and mouse macrophages. These publicly available data cover diverse data types such as mass spectrometry, yeast two-hybrid (Y2H), gene expression profiles, X-ray and NMR determined protein structures and protein expression clones. The growing database covers over 23 000 unique genes/proteins from different experiments and organisms. All of the genes/proteins are annotated and integrated across experiments using UniProt Knowledgebase (UniProtKB) accession numbers. The web-interface for the database enables searching, querying and downloading at the level of experiment, group and individual gene(s)/protein(s) via UniProtKB accession numbers or protein function keywords. The system is accessible at http://www.proteomicsresource.org/. PMID:17984082

Temperature-responsive genetic loci in the plant pathogen Pseudomonas syringae pv. glycinea.

PubMed

Ullrich, M S; Schergaut, M; Boch, J; Ullrich, B

2000-10-01

Plant-pathogenic bacteria may sense variations in environmental factors, such as temperature, to adapt to plant-associated habitats during pathogenesis or epiphytic growth. The bacterial blight pathogen of soybean, Pseudomonas syringae pv. glycinea PG4180, preferentially produces the phytotoxin coronatine at 18 degrees C and infects the host plant under conditions of low temperature and high humidity. A miniTn5-based promoterless glucuronidase (uidA) reporter gene was used to identify genetic loci of PG4180 preferentially expressed at 18 or 28 degrees C. Out of 7500 transposon mutants, 61 showed thermoregulated uidA expression as determined by a three-step screening procedure. Two-thirds of these mutants showed an increased reporter gene expression at 18 degrees C whilst the remainder exhibited higher uidA expression at 28 degrees C. MiniTn5-uidA insertion loci from these mutants were subcloned and their nucleotide sequences were determined. Several of the mutants induced at 18 degrees C contained the miniTn5-uidA insertion within the 32.8 kb coronatine biosynthetic gene cluster. Among the other mutants with increased uidA expression at 18 degrees C, insertions were found in genes encoding formaldehyde dehydrogenase, short-chain dehydrogenase and mannuronan C-5-epimerase, in a plasmid-borne replication protein, and in the hrpT locus, involved in pathogenicity of P. syringae. Among the mutants induced at 28 degrees C, insertions disrupted loci with similarities to a repressor of conjugal plasmid transfer, UV resistance determinants, an isoflavanoid-degrading enzyme, a HU-like DNA-binding protein, two additional regulatory proteins, a homologue of bacterial adhesins, transport proteins, LPS synthesis enzymes and two proteases. Genetic loci from 13 mutants did not show significant similarities to any database entries. Results of plant inoculations showed that three of the mutants tested were inhibited in symptom development and in planta multiplication rates. Temperature-shift experiments suggested that all of the identified loci showed a rather slow induction of expression upon change of temperature.

Gene expression profiling of a pressure-tolerant Listeria monocytogenes Scott A CtsR deletion mutant

USDA-ARS?s Scientific Manuscript database

Listeria monocytogenes is a food-borne pathogen of significant threat to public health. High hydrostatic pressure (HPP) treatment can be used to control Listeria monocytogenes in food. The CtsR (class three stress gene repressor) protein negatively regulates the expression of class III heat shock ...

Fowl adenovirus serotype 9 vectored vaccine for protection of avian influenza virus

USDA-ARS?s Scientific Manuscript database

A fowl adenovirus serotype 9, a non-pathogenic large double stranded DNA virus, was developed as a viral vector to express influenza genes as a potential vaccine. Two separate constructs were developed that expressed either the hemagglutinin gene of A/Chicken/Jalisco/2012 (H7) or A/ Chicken/Iowa/20...

Cloning and expression of Pectobacterium carotovorum endo-polygalacturonase gene in Pichia pastoris for production of oligogalacturonates

USDA-ARS?s Scientific Manuscript database

A bacterial endo-polygalacturonase (endo-PGase) gene from the plant pathogen Pectobacterium carotovorum was cloned into pGAPZaA vector and constitutively expressed in Pichia pastoris. The recombinant endo-PGase secreted by the Pichia clone showed a 1.7 fold increase when the culture medium included ...

Identification of Early Response Genes in Human Peripheral Leukocytes Infected with Orientia tsutsugamushi: The Emergent of a Unique Gene Expression Profile for Diagnosis of O. tsutsugamush Infection

DTIC Science & Technology

2010-01-01

dynein to move from the cell periphery to the microtubule organizing center [22]. Therefore, the initial interactions between host and intracellular...used to study host-pathogen interactions , mainly by identifying genes from pathogens that may be involved in pathogenecity and by surveying the scope...toward understanding the host-Orientia tsutsugamushi interaction at the molecular level, we used human cDNA microarray technology to examine in detail

A Rapid Spin Column-Based Method to Enrich Pathogen Transcripts from Eukaryotic Host Cells Prior to Sequencing

DOE PAGES

Bent, Zachary W.; Poorey, Kunal; LaBauve, Annette E.; ...

2016-12-21

When analyzing pathogen transcriptomes during the infection of host cells, the signal-to-background (pathogen-to-host) ratio of nucleic acids (NA) in infected samples is very small. Despite the advancements in next-generation sequencing, the minute amount of pathogen NA makes standard RNA-seq library preps inadequate for effective gene-level analysis of the pathogen in cases with low bacterial loads. In order to provide a more complete picture of the pathogen transcriptome during an infection, we developed a novel pathogen enrichment technique, which can enrich for transcripts from any cultivable bacteria or virus, using common, readily available laboratory equipment and reagents. To evenly enrich formore » pathogen transcripts, we generate biotinylated pathogen-targeted capture probes in an enzymatic process using the entire genome of the pathogen as a template. The capture probes are hybridized to a strand-specific cDNA library generated from an RNA sample. The biotinylated probes are captured on a monomeric avidin resin in a miniature spin column, and enriched pathogen-specific cDNA is eluted following a series of washes. To test this method, we performed an in vitro time-course infection using Klebsiella pneumoniae to infect murine macrophage cells. K. pneumoniae transcript enrichment efficiency was evaluated using RNA-seq. Bacterial transcripts were enriched up to ~400-fold, and allowed the recovery of transcripts from ~2000–3600 genes not observed in untreated control samples. These additional transcripts revealed interesting aspects of K. pneumoniae biology including the expression of putative virulence factors and the expression of several genes responsible for antibiotic resistance even in the absence of drugs.« less

A Rapid Spin Column-Based Method to Enrich Pathogen Transcripts from Eukaryotic Host Cells Prior to Sequencing

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bent, Zachary W.; Poorey, Kunal; LaBauve, Annette E.

When analyzing pathogen transcriptomes during the infection of host cells, the signal-to-background (pathogen-to-host) ratio of nucleic acids (NA) in infected samples is very small. Despite the advancements in next-generation sequencing, the minute amount of pathogen NA makes standard RNA-seq library preps inadequate for effective gene-level analysis of the pathogen in cases with low bacterial loads. In order to provide a more complete picture of the pathogen transcriptome during an infection, we developed a novel pathogen enrichment technique, which can enrich for transcripts from any cultivable bacteria or virus, using common, readily available laboratory equipment and reagents. To evenly enrich formore » pathogen transcripts, we generate biotinylated pathogen-targeted capture probes in an enzymatic process using the entire genome of the pathogen as a template. The capture probes are hybridized to a strand-specific cDNA library generated from an RNA sample. The biotinylated probes are captured on a monomeric avidin resin in a miniature spin column, and enriched pathogen-specific cDNA is eluted following a series of washes. To test this method, we performed an in vitro time-course infection using Klebsiella pneumoniae to infect murine macrophage cells. K. pneumoniae transcript enrichment efficiency was evaluated using RNA-seq. Bacterial transcripts were enriched up to ~400-fold, and allowed the recovery of transcripts from ~2000–3600 genes not observed in untreated control samples. These additional transcripts revealed interesting aspects of K. pneumoniae biology including the expression of putative virulence factors and the expression of several genes responsible for antibiotic resistance even in the absence of drugs.« less

Human pathogenic Mycoplasma species induced cytokine gene expression in Epstein-Barr virus (EBV)-positive lymphoblastoid cell lines.

PubMed

Schäffner, E; Opitz, O; Pietsch, K; Bauer, G; Ehlers, S; Jacobs, E

1998-04-01

We addressed the question whether the in vitro interaction of two Epstein-Barr virus (EBV)-genome-positive B cell lines (EB-3 and HilB-gamma) with either Mycoplasma pneumoniae or M. hominis, with the mycoplasma species (M. fermentans, M. fermentans subsp. incognitus, M. penetrans, M. genitalium) or with mycoplasma species known to be mere commensals of the respiratory tract (M. orale and M. salivarium) would result in expression of mRNAs for IL-2, IL-2R, IL-4 and IL-6 as determined by reverse transcriptase (RT)-PCR after 4 and 24 h of cocultivation. The pattern of cytokine gene expression observed depended on (i) the origin of the transformed cell line, (ii) the pathogenicity of the Mycoplasma species, and (iii) the length of cocultivation. The EBV-immortalized lymphoblastoid cell line HilB-gamma showed mRNA expression for IL-2, IL-2-receptor, IL-4 and IL-6 peaking 24 h after stimulation with M. pneumoniae and all AIDS-related mycoplasma species tested. The Burkitt lymphoma cell line EB-3 showed a distinct and isolated strong II-2/IL-2 R-mRNA expression within 4 h after contact with the pathogenic and all of the AIDS related mycoplasma species. In neither EBV-containing cell line cytokine was gene expression detectable after stimulation with the commensal mycoplasma species, M. orale and M. salivarium, indicating species differences in the ability of mycoplasmas to interact with and stimulate B-cell lines. Our data suggest that some mcyoplasma species may act as immunomodulatory cofactors by eliciting inappropriate cytokine gene expression in B cells latently infected with EBV. Therefore, this cultivation model may prove useful in evaluating the pathogenetic potential of novel isolated mycoplasma species. Copyright 1998 Academic Press Limited.

Differentially Expressed Genes in Resistant and Susceptible Common Bean (Phaseolus vulgaris L.) Genotypes in Response to Fusarium oxysporum f. sp. phaseoli

PubMed Central

Xue, Renfeng; Wu, Jing; Zhu, Zhendong; Wang, Lanfen; Wang, Xiaoming; Wang, Shumin; Blair, Matthew W.

2015-01-01

Fusarium wilt of common bean (Phaseolus vulgaris L.), caused by Fusarium oxysporum Schlechtend.:Fr. f.sp. phaseoli (Fop), is one of the most important diseases of common beans worldwide. Few natural sources of resistance to Fop exist and provide only moderate or partial levels of protection. Despite the economic importance of the disease across multiple crops, only a few of Fop induced genes have been analyzed in legumes. Therefore, our goal was to identify transcriptionally regulated genes during an incompatible interaction between common bean and the Fop pathogen using the cDNA amplified fragment length polymorphism (cDNA-AFLP) technique. We generated a total of 8,730 transcript-derived fragments (TDFs) with 768 primer pairs based on the comparison of a moderately resistant and a susceptible genotype. In total, 423 TDFs (4.9%) displayed altered expression patterns after inoculation with Fop inoculum. We obtained full amplicon sequences for 122 selected TDFs, of which 98 were identified as annotated known genes in different functional categories based on their putative functions, 10 were predicted but non-annotated genes and 14 were not homologous to any known genes. The 98 TDFs encoding genes of known putative function were classified as related to metabolism (22), signal transduction (21), protein synthesis and processing (20), development and cytoskeletal organization (12), transport of proteins (7), gene expression and RNA metabolism (4), redox reactions (4), defense and stress responses (3), energy metabolism (3), and hormone responses (2). Based on the analyses of homology, 19 TDFs from different functional categories were chosen for expression analysis using quantitative RT-PCR. The genes found to be important here were implicated at various steps of pathogen infection and will allow a better understanding of the mechanisms of defense and resistance to Fop and similar pathogens. The differential response genes discovered here could also be used as molecular markers in association mapping or QTL analysis. PMID:26030070

Identification of DNA Methyltransferase Genes in Human Pathogenic Bacteria by Comparative Genomics.

PubMed

Brambila-Tapia, Aniel Jessica Leticia; Poot-Hernández, Augusto Cesar; Perez-Rueda, Ernesto; Rodríguez-Vázquez, Katya

2016-06-01

DNA methylation plays an important role in gene expression and virulence in some pathogenic bacteria. In this report, we describe DNA methyltransferases (MTases) present in human pathogenic bacteria and compared them with related species, which are not pathogenic or less pathogenic, based in comparative genomics. We performed a search in the KEGG database of the KEGG database orthology groups associated with adenine and cytosine DNA MTase activities (EC: 2.1.1.37, EC: 2.1.1.113 and EC: 2.1.1.72) in 37 human pathogenic species and 18 non/less pathogenic relatives and performed comparisons of the number of these MTases sequences according to their genome size, the DNA MTase type and with their non-less pathogenic relatives. We observed that Helicobacter pylori and Neisseria spp. presented the highest number of MTases while ten different species did not present a predicted DNA MTase. We also detected a significant increase of adenine MTases over cytosine MTases (2.19 vs. 1.06, respectively, p < 0.001). Adenine MTases were the only MTases associated with restriction modification systems and DNA MTases associated with type I restriction modification systems were more numerous than those associated with type III restriction modification systems (0.84 vs. 0.17, p < 0.001); additionally, there was no correlation with the genome size and the total number of DNA MTases, indicating that the number of DNA MTases is related to the particular evolution and lifestyle of specific species, regulating the expression of virulence genes in some pathogenic bacteria.

Intracellular Growth Is Dependent on Tyrosine Catabolism in the Dimorphic Fungal Pathogen Penicillium marneffei

PubMed Central

Boyce, Kylie J.; McLauchlan, Alisha; Schreider, Lena; Andrianopoulos, Alex

2015-01-01

During infection, pathogens must utilise the available nutrient sources in order to grow while simultaneously evading or tolerating the host’s defence systems. Amino acids are an important nutritional source for pathogenic fungi and can be assimilated from host proteins to provide both carbon and nitrogen. The hpdA gene of the dimorphic fungus Penicillium marneffei, which encodes an enzyme which catalyses the second step of tyrosine catabolism, was identified as up-regulated in pathogenic yeast cells. As well as enabling the fungus to acquire carbon and nitrogen, tyrosine is also a precursor in the formation of two types of protective melanin; DOPA melanin and pyomelanin. Chemical inhibition of HpdA in P. marneffei inhibits ex vivo yeast cell production suggesting that tyrosine is a key nutrient source during infectious growth. The genes required for tyrosine catabolism, including hpdA, are located in a gene cluster and the expression of these genes is induced in the presence of tyrosine. A gene (hmgR) encoding a Zn(II)2-Cys6 binuclear cluster transcription factor is present within the cluster and is required for tyrosine induced expression and repression in the presence of a preferred nitrogen source. AreA, the GATA-type transcription factor which regulates the global response to limiting nitrogen conditions negatively regulates expression of cluster genes in the absence of tyrosine and is required for nitrogen metabolite repression. Deletion of the tyrosine catabolic genes in the cluster affects growth on tyrosine as either a nitrogen or carbon source and affects pyomelanin, but not DOPA melanin, production. In contrast to other genes of the tyrosine catabolic cluster, deletion of hpdA results in no growth within macrophages. This suggests that the ability to catabolise tyrosine is not required for macrophage infection and that HpdA has an additional novel role to that of tyrosine catabolism and pyomelanin production during growth in host cells. PMID:25812137

Carbohydrate Availability Regulates Virulence Gene Expression in Streptococcus suis

PubMed Central

Ferrando, M. Laura; van Baarlen, Peter; Orrù, Germano; Piga, Rosaria; Bongers, Roger S.; Wels, Michiel; De Greeff, Astrid; Smith, Hilde E.; Wells, Jerry M.

2014-01-01

Streptococcus suis is a major bacterial pathogen of young pigs causing worldwide economic problems for the pig industry. S. suis is also an emerging pathogen of humans. Colonization of porcine oropharynx by S. suis is considered to be a high risk factor for invasive disease. In the oropharyngeal cavity, where glucose is rapidly absorbed but dietary α-glucans persist, there is a profound effect of carbohydrate availability on the expression of virulence genes. Nineteen predicted or confirmed S. suis virulence genes that promote adhesion to and invasion of epithelial cells were expressed at higher levels when S. suis was supplied with the α-glucan starch/pullulan compared to glucose as the single carbon source. Additionally the production of suilysin, a toxin that damages epithelial cells, was increased more than ten-fold when glucose levels were low and S. suis was growing on pullulan. Based on biochemical, bioinformatics and in vitro and in vivo gene expression studies, we developed a biological model that postulates the effect of carbon catabolite repression on expression of virulence genes in the mucosa, organs and blood. This research increases our understanding of S. suis virulence mechanisms and has important implications for the design of future control strategies including the development of anti-infective strategies by modulating animal feed composition. PMID:24642967

cDNA Microarray Analysis of Host-Pathogen Interactions in a Porcine In Vitro Model for Toxoplasma gondii Infection†

PubMed Central

Okomo-Adhiambo, Margaret; Beattie, Craig; Rink, Anette

2006-01-01

Toxoplasma gondii induces the expression of proinflammatory cytokines, reorganizes organelles, scavenges nutrients, and inhibits apoptosis in infected host cells. We used a cDNA microarray of 420 annotated porcine expressed sequence tags to analyze the molecular basis of these changes at eight time points over a 72-hour period in porcine kidney epithelial (PK13) cells infected with T. gondii. A total of 401 genes with Cy3 and Cy5 spot intensities of ≥500 were selected for analysis, of which 263 (65.6%) were induced ≥2-fold (expression ratio, ≥2.0; P ≤ 0.05 [t test]) over at least one time point and 48 (12%) were significantly down-regulated. At least 12 functional categories of genes were modulated (up- or down-regulated) by T. gondii. The majority of induced genes were clustered as transcription, signal transduction, host immune response, nutrient metabolism, and apoptosis related. The expression of selected genes altered by T. gondii was validated by quantitative real-time reverse transcription-PCR. These results suggest that significant changes in gene expression occur in response to T. gondii infection in PK13 cells, facilitating further analysis of host-pathogen interactions in toxoplasmosis in a secondary host. PMID:16790800

Generation and analysis of expression sequence tags from haustoria of the wheat stripe rust fungus Puccinia striiformis f. sp. Tritici

PubMed Central

2009-01-01

Background Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat (Triticum aestivum L.) worldwide. In spite of its agricultural importance, the genomics and genetics of the pathogen are poorly characterized. Pst transcripts from urediniospores and germinated urediniospores have been examined previously, but little is known about genes expressed during host infection. Some genes involved in virulence in other rust fungi have been found to be specifically expressed in haustoria. Therefore, the objective of this study was to generate a cDNA library to characterize genes expressed in haustoria of Pst. Results A total of 5,126 EST sequences of high quality were generated from haustoria of Pst, from which 287 contigs and 847 singletons were derived. Approximately 10% and 26% of the 1,134 unique sequences were homologous to proteins with known functions and hypothetical proteins, respectively. The remaining 64% of the unique sequences had no significant similarities in GenBank. Fifteen genes were predicted to be proteins secreted from Pst haustoria. Analysis of ten genes, including six secreted protein genes, using quantitative RT-PCR revealed changes in transcript levels in different developmental and infection stages of the pathogen. Conclusions The haustorial cDNA library was useful in identifying genes of the stripe rust fungus expressed during the infection process. From the library, we identified 15 genes encoding putative secreted proteins and six genes induced during the infection process. These genes are candidates for further studies to determine their functions in wheat-Pst interactions. PMID:20028560

Expression of the 12-oxophytodienoic acid 10,11-reductase gene in the compatible interaction between pea and fungal pathogen.

PubMed

Ishiga, Yasuhiro; Funato, Akiko; Tachiki, Tomoyuki; Toyoda, Kazuhiro; Shiraishi, Tomonori; Yamada, Tetsuji; Ichinose, Yuki

2002-10-01

Suppressors produced by Mycosphaerella pinodes are glycopeptides to block pea defense responses induced by elicitors. A clone, S64, was isolated as cDNA for suppressor-inducible gene from pea epicotyls. The treatment of pea epicotyls with suppressor alone induced an increase of S64 mRNA within 1 h, and it reached a maximum level at 3 h after treatment. The induction was not affected by application of the elicitor, indicating that the suppressor has a dominant action to regulate S64 gene expression. S64 was also induced by inoculation with a virulent pathogen, M. pinodes, but not by inoculation with a non-pathogen, Ascochyta rabiei, nor by treatment with fungal elicitor. The deduced structure of S64 showed high homology to 12-oxophytodienoic acid reductase (OPR) in Arabidopsis thaliana. A recombinant protein derived from S64 had OPR activity, suggesting compatibility-specific activation of the octadecanoid pathway in plants. Treatment with jasmonic acid (JA) or methyl jasmonic acid, end products of the octadecanoid pathway, inhibited the elicitor-induced accumulation of PAL mRNA in pea. These results indicate that the suppressor-induced S64 gene expression leads to the production of JA or related compounds, which might contribute to the establishment of compatibility by inhibiting the phenylpropanoid biosynthetic pathway.

Plant viral synergism: the potyviral genome encodes a broad-range pathogenicity enhancer that transactivates replication of heterologous viruses.

PubMed Central

Pruss, G; Ge, X; Shi, X M; Carrington, J C; Bowman Vance, V

1997-01-01

Synergistic viral diseases of higher plants are caused by the interaction of two independent viruses in the same host and are characterized by dramatic increases in symptoms and in accumulation of one of the coinfecting viruses. In potato virus X (PVX)/potyviral synergism, increased pathogenicity and accumulation of PVX are mediated by the expression of potyviral 5' proximal sequences encoding P1, the helper component proteinase (HC-Pro), and a fraction of P3. Here, we report that the same potyviral sequence (termed P1/HC-Pro) enhances the pathogenicity and accumulation of two other heterologous viruses: cucumber mosaic virus and tobacco mosaic virus. In the case of PVX-potyviral synergism, we show that the expression of the HC-Pro gene product, but not the RNA sequence itself, is sufficient to induce the increase in PVX pathogenicity and that both P1 and P3 coding sequences are dispensable for this aspect of the synergistic interaction. In protoplasts, expression of the potyviral P1/HC-Pro region prolongs the accumulation of PVX (-) strand RNA and transactivates expression of a reporter gene from a PVX subgenomic promoter. Unlike the synergistic enhancement of PVX pathogenicity, which requires only expression of HC-Pro, the enhancement of PVX (-) strand RNA accumulation in protoplasts is significantly greater when the entire P1/HC-Pro sequence is expressed. These results indicate that the potyviral P1/HC-Pro region affects a step in disease development that is common to a broad range of virus infections and suggest a mechanism involving transactivation of viral replication. PMID:9212462

Differential expression of oil palm pathology genes during interactions with Ganoderma boninense and Trichoderma harzianum.

PubMed

Alizadeh, Fahimeh; Abdullah, Siti Nor Akmar; Khodavandi, Alireza; Abdullah, Faridah; Yusuf, Umi Kalsom; Chong, Pei Pei

2011-07-01

The expression profiles of Δ9 stearoyl-acyl carrier protein desaturase (SAD1 and SAD2) and type 3 metallothionein (MT3-A and MT3-B) were investigated in seedlings of oil palm (Elaeis guineensis) artificially inoculated with the pathogenic fungus Ganoderma boninense and the symbiotic fungus Trichoderma harzianum. Expression of SAD1 and MT3-A in roots and SAD2 in leaves were significantly up-regulated in G. boninense inoculated seedlings at 21 d after treatment when physical symptoms had not yet appeared and thereafter decreased to basal levels when symptoms became visible. Our finding demonstrated that the SAD1 expression in leaves was significantly down-regulated to negligible levels at 42 and 63 d after treatment. The transcripts of MT3 genes were synthesized in G. boninense inoculated leaves at 42 d after treatment, and the analyses did not show detectable expression of these genes before 42 d after treatment. In T. harzianum inoculated seedlings, the expression levels of SAD1 and SAD2 increased gradually and were stronger in roots than leaves, while for MT3-A and MT3-B, the expression levels were induced in leaves at 3d after treatment and subsequently maintained at same levels until 63d after treatment. The MT3-A expression was significantly up-regulated in roots at 3d after treatment and thereafter were maintained at this level. Both SAD and MT3 expression were maintained at maximum levels or at levels higher than basal. This study demonstrates that oil palm was able to distinguish between pathogenic and symbiotic fungal interactions, thus resulting in different transcriptional activation profiles of SAD and MT3 genes. Increases in expression levels of SAD and MT3 would lead to enhanced resistance against G. boninense and down-regulation of genes confer potential for invasive growth of the pathogen. Differences in expression profiles of SAD and MT3 relate to plant resistance mechanisms while supporting growth enhancing effects of symbiotic T. harzianum. Copyright © 2011 Elsevier GmbH. All rights reserved.

Genome Analyses of an Aggressive and Invasive Lineage of the Irish Potato Famine Pathogen

PubMed Central

Raffaele, Sylvain; Bain, Ruairidh A.; Cooke, Louise R.; Etherington, Graham J.; Deahl, Kenneth L.; Farrer, Rhys A.; Gilroy, Eleanor M.; Goss, Erica M.; Grünwald, Niklaus J.; Hein, Ingo; MacLean, Daniel; McNicol, James W.; Randall, Eva; Oliva, Ricardo F.; Pel, Mathieu A.; Shaw, David S.; Squires, Julie N.; Taylor, Moray C.; Vleeshouwers, Vivianne G. A. A.; Birch, Paul R. J.; Lees, Alison K.; Kamoun, Sophien

2012-01-01

Pest and pathogen losses jeopardise global food security and ever since the 19th century Irish famine, potato late blight has exemplified this threat. The causal oomycete pathogen, Phytophthora infestans, undergoes major population shifts in agricultural systems via the successive emergence and migration of asexual lineages. The phenotypic and genotypic bases of these selective sweeps are largely unknown but management strategies need to adapt to reflect the changing pathogen population. Here, we used molecular markers to document the emergence of a lineage, termed 13_A2, in the European P. infestans population, and its rapid displacement of other lineages to exceed 75% of the pathogen population across Great Britain in less than three years. We show that isolates of the 13_A2 lineage are among the most aggressive on cultivated potatoes, outcompete other aggressive lineages in the field, and overcome previously effective forms of plant host resistance. Genome analyses of a 13_A2 isolate revealed extensive genetic and expression polymorphisms particularly in effector genes. Copy number variations, gene gains and losses, amino-acid replacements and changes in expression patterns of disease effector genes within the 13_A2 isolate likely contribute to enhanced virulence and aggressiveness to drive this population displacement. Importantly, 13_A2 isolates carry intact and in planta induced Avrblb1, Avrblb2 and Avrvnt1 effector genes that trigger resistance in potato lines carrying the corresponding R immune receptor genes Rpi-blb1, Rpi-blb2, and Rpi-vnt1.1. These findings point towards a strategy for deploying genetic resistance to mitigate the impact of the 13_A2 lineage and illustrate how pathogen population monitoring, combined with genome analysis, informs the management of devastating disease epidemics. PMID:23055926

Expressed sequence tag (EST) analysis of two subspecies of Metarhizium anisopliae reveals a plethora of secreted proteins with potential activity in insect hosts.

PubMed

Freimoser, Florian M; Screen, Steven; Bagga, Savita; Hu, Gang; St Leger, Raymond J

2003-01-01

Expressed sequence tag (EST) libraries for Metarhizium anisopliae, the causative agent of green muscardine disease, were developed from the broad host-range pathogen Metarhizium anisopliae sf. anisopliae and the specific grasshopper pathogen, M. anisopliae sf. acridum. Approximately 1,700 5' end sequences from each subspecies were generated from cDNA libraries representing fungi grown under conditions that maximize secretion of cuticle-degrading enzymes. Both subspecies had ESTs for virtually all pathogenicity-related genes cloned to date from M. anisopliae, but many novel genes encoding potential virulence factors were also tagged. Enzymes with potential targets in the insect host included proteases, chitinases, phospholipases, lipases, esterases, phosphatases and enzymes producing toxic secondary metabolites. A diverse array of proteases composed 36 % of all M. anisopliae sf. anisopliae ESTs. Eighty percent of the ESTs that could be clustered into functional groups had significant matches (E<10(-5)) in other ascomycete fungi. These included genes reported to have specific roles in pathogens with plant or vertebrate hosts. Many of the remaining ESTs had their best BLAST match among animal, plant and bacterial sequences. These include genes with plant and microbial counterparts that produce potent antimicrobials. The abundance of transcripts discovered for different functional groups varied between the two subspecies of M. anisopliae in a manner consistent with ecological adaptations of the two pathogens. By hastening gene discovery this project has enhanced development of improved mycoinsecticides. In addition, the M. anisopliae ESTs represent a significant contribution to the extensive database of sequences from ascomycetes that are saprophytes or plant and vertebrate pathogens. Comparative analyses of these sequences is providing important information about the biology and evolutionary history of this clade.

Gene cloning and prokaryotic expression of recombinant outer membrane protein from Vibrio parahaemolyticus

NASA Astrophysics Data System (ADS)

Yuan, Ye; Wang, Xiuli; Guo, Sheping; Qiu, Xuemei

2011-06-01

Gram-negative Vibrio parahaemolyticus is a common pathogen in humans and marine animals. The outer membrane protein of bacteria plays an important role in the infection and pathogenicity to the host. Thus, the outer membrane proteins are an ideal target for vaccines. We amplified a complete outer membrane protein gene (ompW) from V. parahaemolyticus ATCC 17802. We then cloned and expressed the gene into Escherichia coli BL21 (DE3) cells. The gene coded for a protein that was 42.78 kDa. We purified the protein using Ni-NTA affinity chromatography and Anti-His antibody Western blotting, respectively. Our results provide a basis for future application of the OmpW protein as a vaccine candidate against infection by V. parahaemolyticus. In addition, the purified OmpW protein can be used for further functional and structural studies.

The Trophic Life Cycle Stage of the Opportunistic Fungal Pathogen Pneumocystis murina Hinders the Ability of Dendritic Cells To Stimulate CD4+ T Cell Responses

PubMed Central

Evans, Heather M.; Simpson, Andrew; Shen, Shu; Stromberg, Arnold J.; Pickett, Carol L.

2017-01-01

ABSTRACT The life cycle of the opportunistic fungal pathogen Pneumocystis murina consists of a trophic stage and an ascus-like cystic stage. Infection with the cyst stage induces proinflammatory immune responses, while trophic forms suppress the cytokine response to multiple pathogen-associated molecular patterns (PAMPs), including β-glucan. A targeted gene expression assay was used to evaluate the dendritic cell response following stimulation with trophic forms alone, with a normal mixture of trophic forms and cysts, or with β-glucan. We demonstrate that stimulation with trophic forms downregulated the expression of multiple genes normally associated with the response to infection, including genes encoding transcription factors. Trophic forms also suppressed the expression of genes related to antigen processing and presentation, including the gene encoding the major histocompatibility complex (MHC) class II transactivator, CIITA. Stimulation of dendritic cells with trophic forms, but not a mixture of trophic forms and cysts, reduced the expression of MHC class II and the costimulatory molecule CD40 on the surface of the cells. These defects in the expression of MHC class II and costimulatory molecules corresponded with a reduced capacity for trophic form-loaded dendritic cells to stimulate CD4+ T cell proliferation and polarization. These data are consistent with the delayed innate and adaptive responses previously observed in immunocompetent mice inoculated with trophic forms compared to responses in mice inoculated with a mixture of trophic forms and cysts. We propose that trophic forms broadly inhibit the ability of dendritic cells to fulfill their role as antigen-presenting cells. PMID:28694293

Searching for resistance genes to Bursaphelenchus xylophilus using high throughput screening.

PubMed

Santos, Carla S; Pinheiro, Miguel; Silva, Ana I; Egas, Conceição; Vasconcelos, Marta W

2012-11-07

Pine wilt disease (PWD), caused by the pinewood nematode (PWN; Bursaphelenchus xylophilus), damages and kills pine trees and is causing serious economic damage worldwide. Although the ecological mechanism of infestation is well described, the plant's molecular response to the pathogen is not well known. This is due mainly to the lack of genomic information and the complexity of the disease. High throughput sequencing is now an efficient approach for detecting the expression of genes in non-model organisms, thus providing valuable information in spite of the lack of the genome sequence. In an attempt to unravel genes potentially involved in the pine defense against the pathogen, we hereby report the high throughput comparative sequence analysis of infested and non-infested stems of Pinus pinaster (very susceptible to PWN) and Pinus pinea (less susceptible to PWN). Four cDNA libraries from infested and non-infested stems of P. pinaster and P. pinea were sequenced in a full 454 GS FLX run, producing a total of 2,083,698 reads. The putative amino acid sequences encoded by the assembled transcripts were annotated according to Gene Ontology, to assign Pinus contigs into Biological Processes, Cellular Components and Molecular Functions categories. Most of the annotated transcripts corresponded to Picea genes-25.4-39.7%, whereas a smaller percentage, matched Pinus genes, 1.8-12.8%, probably a consequence of more public genomic information available for Picea than for Pinus. The comparative transcriptome analysis showed that when P. pinaster was infested with PWN, the genes malate dehydrogenase, ABA, water deficit stress related genes and PAR1 were highly expressed, while in PWN-infested P. pinea, the highly expressed genes were ricin B-related lectin, and genes belonging to the SNARE and high mobility group families. Quantitative PCR experiments confirmed the differential gene expression between the two pine species. Defense-related genes triggered by nematode infestation were detected in both P. pinaster and P. pinea transcriptomes utilizing 454 pyrosequencing technology. P. pinaster showed higher abundance of genes related to transcriptional regulation, terpenoid secondary metabolism (including some with nematicidal activity) and pathogen attack. P. pinea showed higher abundance of genes related to oxidative stress and higher levels of expression in general of stress responsive genes. This study provides essential information about the molecular defense mechanisms utilized by P. pinaster and P. pinea against PWN infestation and contributes to a better understanding of PWD.

Gene expression of indoor fungal communities under damp building conditions: Implications for human health.

PubMed

Hegarty, B; Dannemiller, K C; Peccia, J

2018-03-03

Dampness and visible mold growth in homes are associated with negative human health outcomes, but causal relationships between fungal exposure and health are not well established. The purpose of this study was to determine whether dampness in buildings impacts fungal community gene expression and how, in turn, gene expression may modulate human health impacts. A metatranscriptomic study was performed on house dust fungal communities to investigate the expression of genes and metabolic processes in chamber experiments at water activity levels of 0.5, 0.85, and 1.0. Fungi at water activities as low as 0.5 were metabolically active, focusing their transcriptional resources on primary processes essential for cell maintenance. Metabolic complexity increased with water activity where communities at 1.0 displayed more diverse secondary metabolic processes. Greater gene expression at increasing water activity has important implications for human health: Fungal communities at 1.0 a w upregulated a greater number of allergen-, mycotoxin-, and pathogenicity-encoding genes versus communities at 0.85 and 0.5 a w . In damp buildings, fungi may display increases in secondary metabolic processes with the potential for greater per-cell production of allergens, toxins, and pathogenicity. Assessments in wet versus dry buildings that do not account for this elevated health impact may not accurately reflect exposure. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Enhanced resistance to blister blight in transgenic tea (Camellia sinensis [L.] O. Kuntze) by overexpression of class I chitinase gene from potato (Solanum tuberosum).

PubMed

Singh, H Ranjit; Deka, Manab; Das, Sudripta

2015-07-01

Tea is the second most consumed beverage in the world. A crop loss of up to 43 % has been reported due to blister blight disease of tea caused by a fungus, Exobasidium vexans. Thus, it directly affects the tea industry qualitatively and quantitatively. Solanum tuberosum class I chitinase gene (AF153195) is a plant pathogenesis-related gene. It was introduced into tea genome via Agrobacterium-mediated transformation with hygromycin phosphotransferase (hpt) gene conferring hygromycin resistance as plant selectable marker. A total of 41 hygromycin resistant plantlets were obtained, and PCR analysis established 12 plantlets confirming about the stable integration of transgene in the plant genome. Real-time PCR detected transgene expression in four transgenic plantlets (T28, C57, C9, and T31). Resistance to biotrophic fungal pathogen, E. vexans, was tested by detached leaf infection assay of greenhouse acclimated plantlets. An inhibitory activity against the fungal pathogen was evident from the detached leaves from the transformants compared with the control. Fungal lesion formed on control plantlet whereas the transgenic plantlets showed resistance to inoculated fungal pathogen by the formation of hypersensitivity reaction area. This result suggests that constitutive expression of the potato class I chitinase gene can be exploited to improve resistance to fungal pathogen, E. vexans, in economical perennial plantation crop like tea.

Silencing of an α-dioxygenase gene, Ca-DOX, retards growth and suppresses basal disease resistance responses in Capsicum annum.

PubMed

Hong, Chi Eun; Ha, Young-Im; Choi, Hyoju; Moon, Ju Yeon; Lee, Jiyoung; Shin, Ah-Young; Park, Chang Jin; Yoon, Gyeong Mee; Kwon, Suk-Yoon; Jo, Ick-Hyun; Park, Jeong Mee

2017-03-01

Alpha-dioxygenases (α-DOX) catalyzing the primary oxygenation of fatty acids to oxylipins were recently found in plants. Here, the biological roles of the pepper α-DOX (Ca-DOX) gene, which is strongly induced during non-host pathogen infection in chili pepper, were examined. Virus-induced gene silencing demonstrated that down-regulation of Ca-DOX enhanced susceptibility to bacterial pathogens and suppressed the hypersensitive response via the suppression of pathogenesis-related genes such as PR4, proteinase inhibitor II and lipid transfer protein (PR14). Ca-DOX-silenced pepper plants also exhibited more retarded growth with lower epidermal cell numbers and reduced cell wall thickness than control plants. To better understand regulation of Ca-DOX, transgenic Arabidopsis plants harboring the β-glucuronidase (GUS) reporter gene driven from a putative Ca-DOX promoter were generated. GUS expression was significantly induced upon avirulent pathogen infection in transgenic Arabidopsis leaves, whereas GUS induction was relatively weak upon virulent pathogen treatment. After treatment with plant hormones, early and strong GUS expression was seen after treatment of salicylic acid, whereas ethylene and methyl jasmonate treatments produced relatively weak and late GUS signals. These results will enable us to further understand the role of α-DOX, which is important in lipid metabolism, defense responses, and growth development in plants.

Genetic reprogramming of host cells by bacterial pathogens.

PubMed

Tran Van Nhieu, Guy; Arbibe, Laurence

2009-10-29

During the course of infection, pathogens often induce changes in gene expression in host cells and these changes can be long lasting and global or transient and of limited amplitude. Defining how, when, and why bacterial pathogens reprogram host cells represents an exciting challenge that opens up the opportunity to grasp the essence of pathogenesis and its molecular details.

Pathogenomic Inference of Virulence-Associated Genes in Leptospira interrogans

PubMed Central

Lehmann, Jason S.; Fouts, Derrick E.; Haft, Daniel H.; Cannella, Anthony P.; Ricaldi, Jessica N.; Brinkac, Lauren; Harkins, Derek; Durkin, Scott; Sanka, Ravi; Sutton, Granger; Moreno, Angelo; Vinetz, Joseph M.; Matthias, Michael A.

2013-01-01

Leptospirosis is a globally important, neglected zoonotic infection caused by spirochetes of the genus Leptospira. Since genetic transformation remains technically limited for pathogenic Leptospira, a systems biology pathogenomic approach was used to infer leptospiral virulence genes by whole genome comparison of culture-attenuated Leptospira interrogans serovar Lai with its virulent, isogenic parent. Among the 11 pathogen-specific protein-coding genes in which non-synonymous mutations were found, a putative soluble adenylate cyclase with host cell cAMP-elevating activity, and two members of a previously unstudied ∼15 member paralogous gene family of unknown function were identified. This gene family was also uniquely found in the alpha-proteobacteria Bartonella bacilliformis and Bartonella australis that are geographically restricted to the Andes and Australia, respectively. How the pathogenic Leptospira and these two Bartonella species came to share this expanded gene family remains an evolutionary mystery. In vivo expression analyses demonstrated up-regulation of 10/11 Leptospira genes identified in the attenuation screen, and profound in vivo, tissue-specific up-regulation by members of the paralogous gene family, suggesting a direct role in virulence and host-pathogen interactions. The pathogenomic experimental design here is generalizable as a functional systems biology approach to studying bacterial pathogenesis and virulence and should encourage similar experimental studies of other pathogens. PMID:24098822

Pathogenomic inference of virulence-associated genes in Leptospira interrogans.

PubMed

Lehmann, Jason S; Fouts, Derrick E; Haft, Daniel H; Cannella, Anthony P; Ricaldi, Jessica N; Brinkac, Lauren; Harkins, Derek; Durkin, Scott; Sanka, Ravi; Sutton, Granger; Moreno, Angelo; Vinetz, Joseph M; Matthias, Michael A

2013-01-01

Leptospirosis is a globally important, neglected zoonotic infection caused by spirochetes of the genus Leptospira. Since genetic transformation remains technically limited for pathogenic Leptospira, a systems biology pathogenomic approach was used to infer leptospiral virulence genes by whole genome comparison of culture-attenuated Leptospira interrogans serovar Lai with its virulent, isogenic parent. Among the 11 pathogen-specific protein-coding genes in which non-synonymous mutations were found, a putative soluble adenylate cyclase with host cell cAMP-elevating activity, and two members of a previously unstudied ∼15 member paralogous gene family of unknown function were identified. This gene family was also uniquely found in the alpha-proteobacteria Bartonella bacilliformis and Bartonella australis that are geographically restricted to the Andes and Australia, respectively. How the pathogenic Leptospira and these two Bartonella species came to share this expanded gene family remains an evolutionary mystery. In vivo expression analyses demonstrated up-regulation of 10/11 Leptospira genes identified in the attenuation screen, and profound in vivo, tissue-specific up-regulation by members of the paralogous gene family, suggesting a direct role in virulence and host-pathogen interactions. The pathogenomic experimental design here is generalizable as a functional systems biology approach to studying bacterial pathogenesis and virulence and should encourage similar experimental studies of other pathogens.

Single sea urchin phagocytes express messages of a single sequence from the diverse Sp185/333 gene family in response to bacterial challenge.

PubMed

Majeske, Audrey J; Oren, Matan; Sacchi, Sandro; Smith, L Courtney

2014-12-01

Immune systems in animals rely on fast and efficient responses to a wide variety of pathogens. The Sp185/333 gene family in the purple sea urchin, Strongylocentrotus purpuratus, consists of an estimated 50 (±10) members per genome that share a basic gene structure but show high sequence diversity, primarily due to the mosaic appearance of short blocks of sequence called elements. The genes show significantly elevated expression in three subpopulations of phagocytes responding to marine bacteria. The encoded Sp185/333 proteins are highly diverse and have central effector functions in the immune system. In this study we report the Sp185/333 gene expression in single sea urchin phagocytes. Sea urchins challenged with heat-killed marine bacteria resulted in a typical increase in coelomocyte concentration within 24 h, which included an increased proportion of phagocytes expressing Sp185/333 proteins. Phagocyte fractions enriched from coelomocytes were used in limiting dilutions to obtain samples of single cells that were evaluated for Sp185/333 gene expression by nested RT-PCR. Amplicon sequences showed identical or nearly identical Sp185/333 amplicon sequences in single phagocytes with matches to six known Sp185/333 element patterns, including both common and rare element patterns. This suggested that single phagocytes show restricted expression from the Sp185/333 gene family and infers a diverse, flexible, and efficient response to pathogens. This type of expression pattern from a family of immune response genes in single cells has not been identified previously in other invertebrates. Copyright © 2014 by The American Association of Immunologists, Inc.

Pepper pectin methylesterase inhibitor protein CaPMEI1 is required for antifungal activity, basal disease resistance and abiotic stress tolerance.

PubMed

An, Soo Hyun; Sohn, Kee Hoon; Choi, Hyong Woo; Hwang, In Sun; Lee, Sung Chul; Hwang, Byung Kook

2008-06-01

Pectin is one of the main components of the plant cell wall that functions as the primary barrier against pathogens. Among the extracellular pectinolytic enzymes, pectin methylesterase (PME) demethylesterifies pectin, which is secreted into the cell wall in a highly methylesterified form. Here, we isolated and functionally characterized the pepper (Capsicum annuum L.) gene CaPMEI1, which encodes a pectin methylesterase inhibitor protein (PMEI), in pepper leaves infected by Xanthomonas campestris pv. vesicatoria (Xcv). CaPMEI1 transcripts are localized in the xylem of vascular bundles in leaf tissues, and pathogens and abiotic stresses can induce differential expression of this gene. Purified recombinant CaPMEI1 protein not only inhibits PME, but also exhibits antifungal activity against some plant pathogenic fungi. Virus-induced gene silencing of CaPMEI1 in pepper confers enhanced susceptibility to Xcv, accompanied by suppressed expression of some defense-related genes. Transgenic Arabidopsis CaPMEI1-overexpression lines exhibit enhanced resistance to Pseudomonas syringae pv. tomato, mannitol and methyl viologen, but not to the biotrophic pathogen Hyaloperonospora parasitica. Together, these results suggest that CaPMEI1, an antifungal protein, may be involved in basal disease resistance, as well as in drought and oxidative stress tolerance in plants.

A De Novo-Assembly Based Data Analysis Pipeline for Plant Obligate Parasite Metatranscriptomic Studies

PubMed Central

Guo, Li; Allen, Kelly S.; Deiulio, Greg; Zhang, Yong; Madeiras, Angela M.; Wick, Robert L.; Ma, Li-Jun

2016-01-01

Current and emerging plant diseases caused by obligate parasitic microbes such as rusts, downy mildews, and powdery mildews threaten worldwide crop production and food safety. These obligate parasites are typically unculturable in the laboratory, posing technical challenges to characterize them at the genetic and genomic level. Here we have developed a data analysis pipeline integrating several bioinformatic software programs. This pipeline facilitates rapid gene discovery and expression analysis of a plant host and its obligate parasite simultaneously by next generation sequencing of mixed host and pathogen RNA (i.e., metatranscriptomics). We applied this pipeline to metatranscriptomic sequencing data of sweet basil (Ocimum basilicum) and its obligate downy mildew parasite Peronospora belbahrii, both lacking a sequenced genome. Even with a single data point, we were able to identify both candidate host defense genes and pathogen virulence genes that are highly expressed during infection. This demonstrates the power of this pipeline for identifying genes important in host–pathogen interactions without prior genomic information for either the plant host or the obligate biotrophic pathogen. The simplicity of this pipeline makes it accessible to researchers with limited computational skills and applicable to metatranscriptomic data analysis in a wide range of plant-obligate-parasite systems. PMID:27462318

Identification of pathogenicity-related genes in Fusarium oxysporum f. sp. cepae.

PubMed

Taylor, Andrew; Vágány, Viktória; Jackson, Alison C; Harrison, Richard J; Rainoni, Alessandro; Clarkson, John P

2016-09-01

Pathogenic isolates of Fusarium oxysporum, distinguished as formae speciales (f. spp.) on the basis of their host specificity, cause crown rots, root rots and vascular wilts on many important crops worldwide. Fusarium oxysporum f. sp. cepae (FOC) is particularly problematic to onion growers worldwide and is increasing in prevalence in the UK. We characterized 31 F. oxysporum isolates collected from UK onions using pathogenicity tests, sequencing of housekeeping genes and identification of effectors. In onion seedling and bulb tests, 21 isolates were pathogenic and 10 were non-pathogenic. The molecular characterization of these isolates, and 21 additional isolates comprising other f. spp. and different Fusarium species, was carried out by sequencing three housekeeping genes. A concatenated tree separated the F. oxysporum isolates into six clades, but did not distinguish between pathogenic and non-pathogenic isolates. Ten putative effectors were identified within FOC, including seven Secreted In Xylem (SIX) genes first reported in F. oxysporum f. sp. lycopersici. Two highly homologous proteins with signal peptides and RxLR motifs (CRX1/CRX2) and a gene with no previously characterized domains (C5) were also identified. The presence/absence of nine of these genes was strongly related to pathogenicity against onion and all were shown to be expressed in planta. Different SIX gene complements were identified in other f. spp., but none were identified in three other Fusarium species from onion. Although the FOC SIX genes had a high level of homology with other f. spp., there were clear differences in sequences which were unique to FOC, whereas CRX1 and C5 genes appear to be largely FOC specific. © 2015 The Authors Molecular Plant Pathology Published by British Society for Plant Pathology and John Wiley & Sons Ltd.

Natural Variation in the Pto Pathogen Resistance Gene Within Species of Wild Tomato (Lycopersicon). I. Functional Analysis of Pto Alleles

PubMed Central

Rose, Laura E.; Langley, Charles H.; Bernal, Adriana J.; Michelmore, Richard W.

2005-01-01

Disease resistance to the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) in the cultivated tomato, Lycopersicon esculentum, and the closely related L. pimpinellifolium is triggered by the physical interaction between plant disease resistance protein, Pto, and the pathogen avirulence protein, AvrPto. To investigate the extent to which variation in the Pto gene is responsible for naturally occurring variation in resistance to Pst, we determined the resistance phenotype of 51 accessions from seven species of Lycopersicon to isogenic strains of Pst differing in the presence of avrPto. One-third of the plants displayed resistance specifically when the pathogen expressed AvrPto, consistent with a gene-for-gene interaction. To test whether this resistance in these species was conferred specifically by the Pto gene, alleles of Pto were amplified and sequenced from 49 individuals and a subset (16) of these alleles was tested in planta using Agrobacterium-mediated transient assays. Eleven alleles conferred a hypersensitive resistance response (HR) in the presence of AvrPto, while 5 did not. Ten amino acid substitutions associated with the absence of AvrPto recognition and HR were identified, none of which had been identified in previous structure-function studies. Additionally, 3 alleles encoding putative pseudogenes of Pto were isolated from two species of Lycopersicon. Therefore, a large proportion, but not all, of the natural variation in the reaction to strains of Pst expressing AvrPto can be attributed to sequence variation in the Pto gene. PMID:15944360

The rice endophyte Harpophora oryzae genome reveals evolution from a pathogen to a mutualistic endophyte

PubMed Central

Xu, Xi-Hui; Su, Zhen-Zhu; Wang, Chen; Kubicek, Christian P.; Feng, Xiao-Xiao; Mao, Li-Juan; Wang, Jia-Ying; Chen, Chen; Lin, Fu-Cheng; Zhang, Chu-Long

2014-01-01

The fungus Harpophora oryzae is a close relative of the pathogen Magnaporthe oryzae and a beneficial endosymbiont of wild rice. Here, we show that H. oryzae evolved from a pathogenic ancestor. The overall genomic structures of H. and M. oryzae were found to be similar. However, during interactions with rice, the expression of 11.7% of all genes showed opposing trends in the two fungi, suggesting differences in gene regulation. Moreover, infection patterns, triggering of host defense responses, signal transduction and nutritional preferences exhibited remarkable differentiation between the two fungi. In addition, the H. oryzae genome was found to contain thousands of loci of transposon-like elements, which led to the disruption of 929 genes. Our results indicate that the gain or loss of orphan genes, DNA duplications, gene family expansions and the frequent translocation of transposon-like elements have been important factors in the evolution of this endosymbiont from a pathogenic ancestor. PMID:25048173

The Lr34 adult plant rust resistance gene provides seedling resistance in durum wheat without senescence.

PubMed

Rinaldo, Amy; Gilbert, Brian; Boni, Rainer; Krattinger, Simon G; Singh, Davinder; Park, Robert F; Lagudah, Evans; Ayliffe, Michael

2017-07-01

The hexaploid wheat (Triticum aestivum) adult plant resistance gene, Lr34/Yr18/Sr57/Pm38/Ltn1, provides broad-spectrum resistance to wheat leaf rust (Lr34), stripe rust (Yr18), stem rust (Sr57) and powdery mildew (Pm38) pathogens, and has remained effective in wheat crops for many decades. The partial resistance provided by this gene is only apparent in adult plants and not effective in field-grown seedlings. Lr34 also causes leaf tip necrosis (Ltn1) in mature adult plant leaves when grown under field conditions. This D genome-encoded bread wheat gene was transferred to tetraploid durum wheat (T. turgidum) cultivar Stewart by transformation. Transgenic durum lines were produced with elevated gene expression levels when compared with the endogenous hexaploid gene. Unlike nontransgenic hexaploid and durum control lines, these transgenic plants showed robust seedling resistance to pathogens causing wheat leaf rust, stripe rust and powdery mildew disease. The effectiveness of seedling resistance against each pathogen correlated with the level of transgene expression. No evidence of accelerated leaf necrosis or up-regulation of senescence gene markers was apparent in these seedlings, suggesting senescence is not required for Lr34 resistance, although leaf tip necrosis occurred in mature plant flag leaves. Several abiotic stress-response genes were up-regulated in these seedlings in the absence of rust infection as previously observed in adult plant flag leaves of hexaploid wheat. Increasing day length significantly increased Lr34 seedling resistance. These data demonstrate that expression of a highly durable, broad-spectrum adult plant resistance gene can be modified to provide seedling resistance in durum wheat. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Regulation of Chlamydia Gene Expression by Tandem Promoters with Different Temporal Patterns.

PubMed

Rosario, Christopher J; Tan, Ming

2016-01-15

Chlamydia is a genus of pathogenic bacteria with an unusual intracellular developmental cycle marked by temporal waves of gene expression. The three main temporal groups of chlamydial genes are proposed to be controlled by separate mechanisms of transcriptional regulation. However, we have noted genes with discrepancies, such as the early gene dnaK and the midcycle genes bioY and pgk, which have promoters controlled by the late transcriptional regulators EUO and σ(28). To resolve this issue, we analyzed the promoters of these three genes in vitro and in Chlamydia trachomatis bacteria grown in cell culture. Transcripts from the σ(28)-dependent promoter of each gene were detected only at late times in the intracellular infection, bolstering the role of σ(28) RNA polymerase in late gene expression. In each case, however, expression prior to late times was due to a second promoter that was transcribed by σ(66) RNA polymerase, which is the major form of chlamydial polymerase. These results demonstrate that chlamydial genes can be transcribed from tandem promoters with different temporal profiles, leading to a composite expression pattern that differs from the expression profile of a single promoter. In addition, tandem promoters allow a gene to be regulated by multiple mechanisms of transcriptional regulation, such as DNA supercoiling or late regulation by EUO and σ(28). We discuss how tandem promoters broaden the repertoire of temporal gene expression patterns in the chlamydial developmental cycle and can be used to fine-tune the expression of specific genes. Chlamydia is a pathogenic bacterium that is responsible for the majority of infectious disease cases reported to the CDC each year. It causes an intracellular infection that is characterized by coordinated expression of chlamydial genes in temporal waves. Chlamydial transcription has been shown to be regulated by DNA supercoiling, alternative forms of RNA polymerase, and transcription factors, but the number of transcription factors found in Chlamydia is far fewer than the number found in most bacteria. This report describes the use of tandem promoters that allow the temporal expression of a gene or operon to be controlled by more than one regulatory mechanism. This combinatorial strategy expands the range of expression patterns that are available to regulate chlamydial genes. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Genome-wide identification of sweet orange (Citrus sinensis) histone modification gene families and their expression analysis during the fruit development and fruit-blue mold infection process

PubMed Central

Xu, Jidi; Xu, Haidan; Liu, Yuanlong; Wang, Xia; Xu, Qiang; Deng, Xiuxin

2015-01-01

In eukaryotes, histone acetylation and methylation have been known to be involved in regulating diverse developmental processes and plant defense. These histone modification events are controlled by a series of histone modification gene families. To date, there is no study regarding genome-wide characterization of histone modification related genes in citrus species. Based on the two recent sequenced sweet orange genome databases, a total of 136 CsHMs (Citrus sinensis histone modification genes), including 47 CsHMTs (histone methyltransferase genes), 23 CsHDMs (histone demethylase genes), 50 CsHATs (histone acetyltransferase genes), and 16 CsHDACs (histone deacetylase genes) were identified. These genes were categorized to 11 gene families. A comprehensive analysis of these 11 gene families was performed with chromosome locations, phylogenetic comparison, gene structures, and conserved domain compositions of proteins. In order to gain an insight into the potential roles of these genes in citrus fruit development, 42 CsHMs with high mRNA abundance in fruit tissues were selected to further analyze their expression profiles at six stages of fruit development. Interestingly, a numbers of genes were expressed highly in flesh of ripening fruit and some of them showed the increasing expression levels along with the fruit development. Furthermore, we analyzed the expression patterns of all 136 CsHMs response to the infection of blue mold (Penicillium digitatum), which is the most devastating pathogen in citrus post-harvest process. The results indicated that 20 of them showed the strong alterations of their expression levels during the fruit-pathogen infection. In conclusion, this study presents a comprehensive analysis of the histone modification gene families in sweet orange and further elucidates their behaviors during the fruit development and the blue mold infection responses. PMID:26300904

Pathogen Proliferation Governs the Magnitude but Compromises the Function of CD8 T Cells1

PubMed Central

Sad, Subash; Dudani, Renu; Gurnani, Komal; Russell, Marsha; van Faassen, Henk; Finlay, Brett; Krishnan, Lakshmi

2014-01-01

CD8+ T cell memory is critical for protection against many intracellular pathogens. However, it is not clear how pathogen virulence influences the development and function of CD8+ T cells. Salmonella typhimurium (ST) is an intracellular bacterium that causes rapid fatality in susceptible mice and chronic infection in resistant strains. We have constructed recombinant mutants of ST, expressing the same immunodominant Ag OVA, but defective in various key virulence genes. We show that the magnitude of CD8+ T cell response correlates directly to the intracellular proliferation of ST. Wild-type ST displayed efficient intracellular proliferation and induced increased numbers of OVA-specific CD8+ T cells upon infection in mice. In contrast, mutants with defective Salmonella pathogenicity island II genes displayed poor intracellular proliferation and induced reduced numbers of OVA-specific CD8+ T cells. However, when functionality of the CD8+ T cell response was measured, mutants of ST induced a more functional response compared with the wild-type ST. Infection with wild-type ST, in contrast to mutants defective in pathogenicity island II genes, induced the generation of mainly effector-memory CD8+ T cells that expressed little IL-2, failed to mediate efficient cytotoxicity, and proliferated poorly in response to Ag challenge in vivo. Taken together, these results indicate that pathogens that proliferate rapidly and chronically in vivo may evoke functionally inferior memory CD8+ T cells which may promote the survival of the pathogen. PMID:18424704

Constitutive activation of jasmonate signaling in an Arabidopsis mutant correlates with enhanced resistance to Erysiphe cichoracearum, Pseudomonas syringae, and Myzus persicae.

PubMed

Ellis, Christine; Karafyllidis, Ioannis; Turner, John G

2002-10-01

In Arabidopsis spp., the jasmonate (JA) response pathway generally is required for defenses against necrotrophic pathogens and chewing insects, while the salicylic acid (SA) response pathway is generally required for specific, resistance (R) gene-mediated defenses against both biotrophic and necrotrophic pathogens. For example, SA-dependent defenses are required for resistance to the biotrophic fungal pathogen Erysiphe cichoracearum UCSC1 and the bacterial pathogen Pseudomonas syringae pv. maculicola, and also are expressed during response to the green peach aphid Myzus persicae. However, recent evidence indicates that the expression of JA-dependent defenses also may confer resistance to E. cichoracearum. To confirm and to extend this observation, we have compared the disease and pest resistance of wild-type Arabidopsis plants with that of the mutants coil, which is insensitive to JA, and cev1, which has constitutive JA signaling. Measurements of the colonization of these plants by E. cichoracearum, P. syringae pv. maculicola, and M. persicae indicated that activation of the JA signal pathway enhanced resistance, and was associated with the activation of JA-dependent defense genes and the suppression of SA-dependent defense genes. We conclude that JA and SA induce alternative defense pathways that can confer resistance to the same pathogens and pests.

Analysis of gene expression in two growth states of Xylella fastidiosa and its relationship with pathogenicity.

PubMed

de Souza, Alessandra A; Takita, Marco A; Coletta-Filho, Helvécio D; Caldana, Camila; Goldman, Gustavo H; Yanai, Giane M; Muto, Nair H; de Oliveira, Regina C; Nunes, Luiz R; Machado, Marcos A

2003-10-01

Xylella fastidiosa is a plant pathogen responsible for diseases of economically important crops. Although there is considerable disagreement about its mechanism of pathogenicity, blockage of the vessels is one of the most accepted hypotheses. Loss of virulence by this bacterium was observed after serial passages in axenic culture. To confirm the loss of pathogenicity of X. fastidiosa, the causing agent of citrus variegated chlorosis (CVC), freshly-isolated bacteria (first passage [FP] condition) as well as bacteria obtained after 46 passages in axenic culture (several passage [SP] condition) were inoculated into sweet orange and periwinkle plants. Using real time quantitative polymerase chain reaction, we verified that the colonization of FP cells was more efficient for both hosts. The sequence of the complete X. fastidiosa genome allowed the construction of a DNA microarray that was used to investigate the total changes in gene expression associated with the FP condition. Most genes found to be induced in the FP condition were associated with adhesion and probably with adaptation to the host environment. This report represents the first study of the transcriptome of this pathogen, which has recently gained more importance, since the genome of several strains has been either partially or entirely sequenced.

A review of the immune molecules in the sea cucumber.

PubMed

Xue, Zhuang; Li, Hui; Wang, Xiuli; Li, Xia; Liu, Yang; Sun, Jing; Liu, Cenjie

2015-05-01

It is very important to identify and characterize the immune-related genes that respond to pathogens. Until recently, only some of the immune-related genes in sea cucumbers had been characterized. Their expression patterns after pathogen challenges have been analyzed via expressed sequence tag libraries, microarray studies and proteomic approaches. These genes include lectins, antimicrobial peptides, lysozyme, enzymes, clotting protein, pattern recognition proteins, Toll receptors, complement C3 and other humoral factors that might participate in the innate immune system of sea cucumbers. Although the participation of some of these immune molecules in the sea cucumber's innate immune defense against invading pathogens has been demonstrated, the functions of many of the molecules remain unclear. This review focuses on the discovery and functional characterization of the immune-related molecules from the sea cucumber for the first time and provides new insights into the immune mechanisms of the sea cucumber, which opens new possibilities for developing drugs for novel anti-bacterial and antiviral applications in fisheries. Copyright © 2015 Elsevier Ltd. All rights reserved.

Clustering, haplotype diversity and locations of MIC-3: a unique root-specific defense-related gene family in upland cotton (Gossypium hirsutum L.)

USDA-ARS?s Scientific Manuscript database

MIC-3-related genes of cotton (Gossypium spp.) were identified and shown to have root-specific expression, associated with pathogen defense-related function and specifically increased expression in root-knot nematode (RKN) resistant plants after nematode infection. Here we cloned and sequenced MIC-...

Immune response of the Caribbean sea fan, Gorgonia ventalina, exposed to an Aplanochytrium parasite as revealed by transcriptome sequencing

PubMed Central

Burge, Colleen A.; Mouchka, Morgan E.; Harvell, C. Drew; Roberts, Steven

2013-01-01

Coral reef communities are undergoing marked declines due to a variety of stressors including disease. The sea fan coral, Gorgonia ventalina, is a tractable study system to investigate mechanisms of immunity to a naturally occurring pathogen. Functional studies in Gorgonia ventalina immunity indicate that several key pathways and cellular components are involved in response to natural microbial invaders, although to date the functional and regulatory pathways remain largely un-described. This study used short-read sequencing (Illumina GAIIx) to identify genes involved in the response of G. ventalina to a naturally occurring Aplanochytrium spp. parasite. De novo assembly of the G. ventalina transcriptome yielded 90,230 contigs of which 40,142 were annotated. RNA-Seq analysis revealed 210 differentially expressed genes in sea fans exposed to the Aplanochytrium parasite. Differentially expressed genes involved in immunity include pattern recognition molecules, anti-microbial peptides, and genes involved in wound repair and reactive oxygen species formation. Gene enrichment analysis indicated eight biological processes were enriched representing 36 genes, largely involved with protein translation and energy production. This is the first report using high-throughput sequencing to characterize the host response of a coral to a natural pathogen. Furthermore, we have generated the first transcriptome for a soft (octocoral or non-scleractinian) coral species. Expression analysis revealed genes important in invertebrate innate immune pathways, as well as those whose role is previously un-described in cnidarians. This resource will be valuable in characterizing G. ventalina immune response to infection and co-infection of pathogens in the context of environmental change. PMID:23898300

Fungal-specific transcription factor AbPf2 activates pathogenicity in Alternaria brassicicola

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cho, Yangrae; Ohm, Robin A.; Grigoriev, Igor V.

Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. To identify molecular determinants of pathogenicity, we created non-pathogenic mutants of a transcription factor-encoding gene, AbPf2. The frequency and timing of germination and appressorium formation on host plants were similar between the non-pathogenic abpf2 mutants and wild-type A. brassicicola. The mutants were also similar in vitro to wild-type A. brassicicola in terms of vegetative growth, conidium production, and responses to a phytoalexin, reactive oxygen species and osmolites. The hyphae of the mutants grew slowly but did not cause disease symptoms on the surface of host plants. Transcripts of the AbPf2more » gene increased exponentially soon after wild-type conidia contacted their host plants . A small amount of AbPf2 protein, as monitored using GFP fusions, was present in young, mature conidia. The protein level decreased during saprophytic growth, but increased and was located primarily in fungal nuclei during pathogenesis. Levels of the proteins and transcripts sharply decreased following colonization of host tissues beyond the initial infection site. When expression of the transcription factor was induced in the wild-type during early pathogenesis, 106 fungal genes were also induced in the wild-type but not in the abpf2 mutants. Notably, 33 of the 106 genes encoded secreted proteins, including eight putative effector proteins. Plants inoculated with abpf2 mutants expressed higher levels of genes associated with photosynthesis, the pentose phosphate pathway and primary metabolism, but lower levels of defense-related genes. Our results suggest that AbPf2 is an important regulator of pathogenesis, but does not affect other cellular processes in A. brassicicola.« less

The global regulator of pathogenesis PnCon7 positively regulates Tox3 effector gene expression through direct interaction in the wheat pathogen Parastagonospora nodorum.

PubMed

Lin, Shao-Yu; Chooi, Yit-Heng; Solomon, Peter S

2018-05-03

To investigate effector gene regulation in the wheat pathogenic fungus Parastagonospora nodorum, the promoter and expression of Tox3 was characterised through a series of complementary approaches. Promoter deletion and DNase I footprinting experiments identified a 25 bp region in the Tox3 promoter as being required for transcription. Subsequent yeast one-hybrid analysis using the DNA sequence as bait identified that interacting partner as the C2H2 zinc finger transcription factor PnCon7, a putative master regulator of pathogenesis. Silencing of PnCon7 resulted in the down-regulation of Tox3 demonstrating that the transcription factor has a positive regulatory role on gene expression. Analysis of Tox3 expression in the PnCon7 silenced strains revealed a strong correlation with PnCon7 transcript levels, supportive of a direct regulatory role. Subsequent pathogenicity assays using PnCon7-silenced isolates revealed that the transcription factor was required for Tox3-mediated disease. The expression of two other necrotrophic effectors (ToxA and Tox1) was also affected but in a non-dose dependent manner suggesting that the regulatory role of PnCon7 on these genes was indirect. Collectively, these data have advanced our fundamental understanding of the Con7 master regulator of pathogenesis by demonstrating its positive regulatory role on the Tox3 effector in P. nodorum through direct interaction. This article is protected by copyright. All rights reserved. © 2018 John Wiley & Sons Ltd.

Whole-transcriptome, high-throughput RNA sequence analysis of the bovine macrophage response to Mycobacterium bovis infection in vitro.

PubMed

Nalpas, Nicolas C; Park, Stephen D E; Magee, David A; Taraktsoglou, Maria; Browne, John A; Conlon, Kevin M; Rue-Albrecht, Kévin; Killick, Kate E; Hokamp, Karsten; Lohan, Amanda J; Loftus, Brendan J; Gormley, Eamonn; Gordon, Stephen V; MacHugh, David E

2013-04-08

Mycobacterium bovis, the causative agent of bovine tuberculosis, is an intracellular pathogen that can persist inside host macrophages during infection via a diverse range of mechanisms that subvert the host immune response. In the current study, we have analysed and compared the transcriptomes of M. bovis-infected monocyte-derived macrophages (MDM) purified from six Holstein-Friesian females with the transcriptomes of non-infected control MDM from the same animals over a 24 h period using strand-specific RNA sequencing (RNA-seq). In addition, we compare gene expression profiles generated using RNA-seq with those previously generated by us using the high-density Affymetrix® GeneChip® Bovine Genome Array platform from the same MDM-extracted RNA. A mean of 7.2 million reads from each MDM sample mapped uniquely and unambiguously to single Bos taurus reference genome locations. Analysis of these mapped reads showed 2,584 genes (1,392 upregulated; 1,192 downregulated) and 757 putative natural antisense transcripts (558 upregulated; 119 downregulated) that were differentially expressed based on sense and antisense strand data, respectively (adjusted P-value ≤ 0.05). Of the differentially expressed genes, 694 were common to both the sense and antisense data sets, with the direction of expression (i.e. up- or downregulation) positively correlated for 693 genes and negatively correlated for the remaining gene. Gene ontology analysis of the differentially expressed genes revealed an enrichment of immune, apoptotic and cell signalling genes. Notably, the number of differentially expressed genes identified from RNA-seq sense strand analysis was greater than the number of differentially expressed genes detected from microarray analysis (2,584 genes versus 2,015 genes). Furthermore, our data reveal a greater dynamic range in the detection and quantification of gene transcripts for RNA-seq compared to microarray technology. This study highlights the value of RNA-seq in identifying novel immunomodulatory mechanisms that underlie host-mycobacterial pathogen interactions during infection, including possible complex post-transcriptional regulation of host gene expression involving antisense RNA.

A Genetic Screen for Pathogenicity Genes in the Hemibiotrophic Fungus Colletotrichum higginsianum Identifies the Plasma Membrane Proton Pump Pma2 Required for Host Penetration

PubMed Central

Dahl, Marlis; Müller, Susanne; Voll, Lars M.; Koch, Christian

2015-01-01

We used insertional mutagenesis by Agrobacterium tumefaciens mediated transformation (ATMT) to isolate pathogenicity mutants of Colletotrichum higginsianum. From a collection of 7200 insertion mutants we isolated 75 mutants with reduced symptoms. 19 of these were affected in host penetration, while 17 were affected in later stages of infection, like switching to necrotrophic growth. For 16 mutants the location of T-DNA insertions could be identified by PCR. A potential plasma membrane H+-ATPase Pma2 was targeted in five independent insertion mutants. We genetically inactivated the Ku80 component of the non-homologous end-joining pathway in C. higginsianum to establish an efficient gene knockout protocol. Chpma2 deletion mutants generated by homologous recombination in the ΔChku80 background form fully melanized appressoria but entirely fail to penetrate the host tissue and are non-pathogenic. The ChPMA2 gene is induced upon appressoria formation and infection of A. thaliana. Pma2 activity is not important for vegetative growth of saprophytically growing mycelium, since the mutant shows no growth penalty under these conditions. Colletotrichum higginsianum codes for a closely related gene (ChPMA1), which is highly expressed under most growth conditions. ChPMA1 is more similar to the homologous yeast genes for plasma membrane pumps. We propose that expression of a specific proton pump early during infection may be common to many appressoria forming fungal pathogens as we found ChPMA2 orthologs in several plant pathogenic fungi. PMID:25992547

Functional Analysis of Genes Comprising the Locus of Heat Resistance in Escherichia coli.

PubMed

Mercer, Ryan; Nguyen, Oanh; Ou, Qixing; McMullen, Lynn; Gänzle, Michael G

2017-10-15

The locus of heat resistance (LHR) is a 15- to 19-kb genomic island conferring exceptional heat resistance to organisms in the family Enterobacteriaceae , including pathogenic strains of Salmonella enterica and Escherichia coli The complement of LHR-comprising genes that is necessary for heat resistance and the stress-induced or growth-phase-induced expression of LHR-comprising genes are unknown. This study determined the contribution of the seven LHR-comprising genes yfdX1 GI , yfdX2 , hdeD GI , orf11 , trx GI , kefB , and psiE GI by comparing the heat resistances of E. coli strains harboring plasmid-encoded derivatives of the different LHRs in these genes. (Genes carry a subscript "GI" [genomic island] if an ortholog of the same gene is present in genomes of E. coli ) LHR-encoded heat shock proteins sHSP20, ClpK GI , and sHSP GI are not sufficient for the heat resistance phenotype; YfdX1, YfdX2, and HdeD are necessary to complement the LHR heat shock proteins and to impart a high level of resistance. Deletion of trx GI , kefB , and psiE GI from plasmid-encoded copies of the LHR did not significantly affect heat resistance. The effect of the growth phase and the NaCl concentration on expression from the putative LHR promoter p2 was determined by quantitative reverse transcription-PCR and by a plasmid-encoded p2:GFP promoter fusion. The expression levels of exponential- and stationary-phase E. coli cells were not significantly different, but the addition of 1% NaCl significantly increased LHR expression. Remarkably, LHR expression in E. coli was dependent on a chromosomal copy of evgA In conclusion, this study improved our understanding of the genes required for exceptional heat resistance in E. coli and factors that increase their expression in food. IMPORTANCE The locus of heat resistance (LHR) is a genomic island conferring exceptional heat resistance to several foodborne pathogens. The exceptional level of heat resistance provided by the LHR questions the control of pathogens by current food processing and preparation techniques. The function of LHR-comprising genes and their regulation, however, remain largely unknown. This study defines a core complement of LHR-encoded proteins that are necessary for heat resistance and demonstrates that regulation of the LHR in E. coli requires a chromosomal copy of the gene encoding EvgA. This study provides insight into the function of a transmissible genomic island that allows otherwise heat-sensitive enteric bacteria, including pathogens, to lead a thermoduric lifestyle and thus contributes to the detection and control of heat-resistant enteric bacteria in food. Copyright © 2017 American Society for Microbiology.

Functional Analysis of Genes Comprising the Locus of Heat Resistance in Escherichia coli

PubMed Central

Mercer, Ryan; Nguyen, Oanh; Ou, Qixing; McMullen, Lynn

2017-01-01

ABSTRACT The locus of heat resistance (LHR) is a 15- to 19-kb genomic island conferring exceptional heat resistance to organisms in the family Enterobacteriaceae, including pathogenic strains of Salmonella enterica and Escherichia coli. The complement of LHR-comprising genes that is necessary for heat resistance and the stress-induced or growth-phase-induced expression of LHR-comprising genes are unknown. This study determined the contribution of the seven LHR-comprising genes yfdX1GI, yfdX2, hdeDGI, orf11, trxGI, kefB, and psiEGI by comparing the heat resistances of E. coli strains harboring plasmid-encoded derivatives of the different LHRs in these genes. (Genes carry a subscript “GI” [genomic island] if an ortholog of the same gene is present in genomes of E. coli.) LHR-encoded heat shock proteins sHSP20, ClpKGI, and sHSPGI are not sufficient for the heat resistance phenotype; YfdX1, YfdX2, and HdeD are necessary to complement the LHR heat shock proteins and to impart a high level of resistance. Deletion of trxGI, kefB, and psiEGI from plasmid-encoded copies of the LHR did not significantly affect heat resistance. The effect of the growth phase and the NaCl concentration on expression from the putative LHR promoter p2 was determined by quantitative reverse transcription-PCR and by a plasmid-encoded p2:GFP promoter fusion. The expression levels of exponential- and stationary-phase E. coli cells were not significantly different, but the addition of 1% NaCl significantly increased LHR expression. Remarkably, LHR expression in E. coli was dependent on a chromosomal copy of evgA. In conclusion, this study improved our understanding of the genes required for exceptional heat resistance in E. coli and factors that increase their expression in food. IMPORTANCE The locus of heat resistance (LHR) is a genomic island conferring exceptional heat resistance to several foodborne pathogens. The exceptional level of heat resistance provided by the LHR questions the control of pathogens by current food processing and preparation techniques. The function of LHR-comprising genes and their regulation, however, remain largely unknown. This study defines a core complement of LHR-encoded proteins that are necessary for heat resistance and demonstrates that regulation of the LHR in E. coli requires a chromosomal copy of the gene encoding EvgA. This study provides insight into the function of a transmissible genomic island that allows otherwise heat-sensitive enteric bacteria, including pathogens, to lead a thermoduric lifestyle and thus contributes to the detection and control of heat-resistant enteric bacteria in food. PMID:28802266

L-glutamine Induces Expression of Listeria monocytogenes Virulence Genes

PubMed Central

Lobel, Lior; Burg-Golani, Tamar; Sigal, Nadejda; Rose, Jessica; Livnat-Levanon, Nurit; Lewinson, Oded; Herskovits, Anat A.

2017-01-01

The high environmental adaptability of bacteria is contingent upon their ability to sense changes in their surroundings. Bacterial pathogen entry into host poses an abrupt and dramatic environmental change, during which successful pathogens gauge multiple parameters that signal host localization. The facultative human pathogen Listeria monocytogenes flourishes in soil, water and food, and in ~50 different animals, and serves as a model for intracellular infection. L. monocytogenes identifies host entry by sensing both physical (e.g., temperature) and chemical (e.g., metabolite concentrations) factors. We report here that L-glutamine, an abundant nitrogen source in host serum and cells, serves as an environmental indicator and inducer of virulence gene expression. In contrast, ammonia, which is the most abundant nitrogen source in soil and water, fully supports growth, but fails to activate virulence gene transcription. We demonstrate that induction of virulence genes only occurs when the Listerial intracellular concentration of L-glutamine crosses a certain threshold, acting as an on/off switch: off when L-glutamine concentrations are below the threshold, and fully on when the threshold is crossed. To turn on the switch, L-glutamine must be present, and the L-glutamine high affinity ABC transporter, GlnPQ, must be active. Inactivation of GlnPQ led to complete arrest of L-glutamine uptake, reduced type I interferon response in infected macrophages, dramatic reduction in expression of virulence genes, and attenuated virulence in a mouse infection model. These results may explain observations made with other pathogens correlating nitrogen metabolism and virulence, and suggest that gauging of L-glutamine as a means of ascertaining host localization may be a general mechanism. PMID:28114430

Transcriptomic analysis of two Beauveria bassiana strains grown on cuticle extracts of the silkworm uncovers their different metabolic response at early infection stage.

PubMed

Wang, Jing-Jie; Bai, Wen-Wen; Zhou, Wei; Liu, Jing; Chen, Jie; Liu, Xiao-Yuan; Xiang, Ting-Ting; Liu, Ren-Hua; Wang, Wen-Hui; Zhang, Bao-Ling; Wan, Yong-Ji

2017-05-01

Beauveria bassiana is an important entomopathogenic fungus which not only widely distributes in the environment but also shows phenotypic diversity. However, the mechanism of pathogenic differences among natural B. bassiana strains has not been revealed at transcriptome-wide level. In the present study, in order to explore the mechanism, two B. bassiana strains with different pathogenicity were isolated from silkworms (Bombyx mori L.) and selected to analyze the gene expression of early stage by culturing on cuticle extracts of the silkworm and using RNA-sequencing technique. A total of 2108 up-regulated and 1115 down-regulated genes were identified in B. bassiana strain GXsk1011 (hyper-virulent strain) compared with B. bassiana strain GXtr1009 (hypo-virulent strain), respectively. The function categorization of differential expressed genes (DEGs) showed that most of them involved in metabolic process, biosynthesis of secondary metabolites, catalytic activity, and some involved in nutrition uptake, adhesion and host defense were also noted. Based on our data, distinct pathogenicity among different strains of B. bassiana may largely attribute to unique gene expression pattern which differed at very early infection process. Most of the genes involved in conidia adhesion, cuticle degradation and fungal growth were up-regulated in hyper-virulent B. bassiana strain GXsk1011. Furthermore, in combination with fungal growth analysis, our research provided a clue that fungal growth may also play an important role during early infection process. The results will help to explain why different B. bassiana strains show distinct pathogenicity on the same host even under same condition. Moreover, the transcriptome data were also useful for screening potential virulence factors. Copyright © 2017 Elsevier Inc. All rights reserved.

Gasmin (BV2-5), a polydnaviral-acquired gene in Spodoptera exigua. Trade-off in the defense against bacterial and viral infections.

PubMed

Gasmi, Laila; Jakubowska, Agata K; Herrero, Salvador

2016-03-01

Thousands of Hymenopteran endoparasitoids have developed a unique symbiotic relationship with viruses named polydnavirus (PDVs). These viruses immunocompromise the lepidopteran host allowing the survival of the wasp eggs. In a previous work, we have shown the horizontal transfer of some polydnaviral genes into the genome of the Lepidoptera, Spodoptera exigua. One of these genes, BV2-5 (named gasmin) interferes with actin polymerization, negatively affecting the multiplication of baculovirus in cell culture. In this work, we have focused in the study of the effect of Gasmin expression on different aspects of the baculovirus production. In addition, and since actin polymerization is crucial for phagocytosis, we have studied the effect of Gasmin expression on the larval interaction with bacterial pathogens. Over-expression of Gasmin on hemocytes significantly reduces their capacity to phagocytize the pathogenic bacteria Bacillus thuringiensis. According to these results, gasmin domestication negatively affects baculovirus replication, but increases larvae susceptibility to bacterial infections as pay off. Although the effect of Gasmin on the insect interaction with other pathogens or parasitoids remain unknown, the opposite effects described here could shape the biological history of this species based on the abundance of certain type of pathogens as suggested by the presence of truncated forms of this protein in several regions of the world. Copyright © 2015 Elsevier Ltd. All rights reserved.

Role and regulation of the orphan AphA protein of quorum sensing in pathogenic Vibrios.

PubMed

Lu, Renfei; Osei-Adjei, George; Huang, Xinxiang; Zhang, Yiquan

2018-03-01

Quorum sensing (QS), a cell-to-cell communication process, is widely distributed in the bacterial kingdom. Bacteria use QS to control gene expression in response to cell density by detecting the signal molecules called autoinducers. AphA protein is the master QS regulator of vibrios operating at low cell density. It regulates the expression of a variety of genes, especially those encoding virulence factors, flagella/motility and biofilm formation. The role and regulation of AphA in vibrios, especially in human pathogenic vibrios, are summarized in this review. Clarification of the roles of AphA will help us to understand the pathogenesis of vibrios.

Lessons learned: Optimization of a murine small bowel resection model

PubMed Central

Taylor, Janice A.; Martin, Colin A.; Nair, Rajalakshmi; Guo, Jun; Erwin, Christopher R.; Warner, Brad W.

2008-01-01

Background/Purpose Central to the use of murine models of disease is the ability to derive reproducible data. The purpose of this study was to determine factors contributing to variability in our murine model of small bowel resection (SBR). Methods Male C57Bl/6 mice were randomized to sham or 50% SBR. The effect of housing type (pathogen-free versus standard housing), nutrition (reconstituted powder versus tube feeding formulation), and correlates of intestinal morphology with gene expression changes were investigated Multiple linear regression modeling or one-way ANOVA was used for data analysis. Results Pathogen-free mice had significantly shorter ileal villi at baseline and demonstrated greater villus growth after SBR compared to mice housed in standard rooms. Food type did not affect adaptation. Gene expression changes were more consistent and significant in isolated crypt cells that demonstrated adaptive growth when compared with crypts that did not deepen after SBR. Conclusion Maintenance of mice in pathogen-free conditions and restricting gene expression analysis to individual animals exhibiting morphologic adaptation enhances sensitivity and specificity of data derived from this model. These refinements will minimize experimental variability and lead to improved understanding of the complex process of intestinal adaptation. PMID:18558176

Arabidopsis Defense against Botrytis cinerea: Chronology and Regulation Deciphered by High-Resolution Temporal Transcriptomic Analysis[C][W

PubMed Central

Windram, Oliver; Madhou, Priyadharshini; McHattie, Stuart; Hill, Claire; Hickman, Richard; Cooke, Emma; Jenkins, Dafyd J.; Penfold, Christopher A.; Baxter, Laura; Breeze, Emily; Kiddle, Steven J.; Rhodes, Johanna; Atwell, Susanna; Kliebenstein, Daniel J.; Kim, Youn-sung; Stegle, Oliver; Borgwardt, Karsten; Zhang, Cunjin; Tabrett, Alex; Legaie, Roxane; Moore, Jonathan; Finkenstadt, Bärbel; Wild, David L.; Mead, Andrew; Rand, David; Beynon, Jim; Ott, Sascha; Buchanan-Wollaston, Vicky; Denby, Katherine J.

2012-01-01

Transcriptional reprogramming forms a major part of a plant’s response to pathogen infection. Many individual components and pathways operating during plant defense have been identified, but our knowledge of how these different components interact is still rudimentary. We generated a high-resolution time series of gene expression profiles from a single Arabidopsis thaliana leaf during infection by the necrotrophic fungal pathogen Botrytis cinerea. Approximately one-third of the Arabidopsis genome is differentially expressed during the first 48 h after infection, with the majority of changes in gene expression occurring before significant lesion development. We used computational tools to obtain a detailed chronology of the defense response against B. cinerea, highlighting the times at which signaling and metabolic processes change, and identify transcription factor families operating at different times after infection. Motif enrichment and network inference predicted regulatory interactions, and testing of one such prediction identified a role for TGA3 in defense against necrotrophic pathogens. These data provide an unprecedented level of detail about transcriptional changes during a defense response and are suited to systems biology analyses to generate predictive models of the gene regulatory networks mediating the Arabidopsis response to B. cinerea. PMID:23023172

Expressed sequences tags of the anther smut fungus, Microbotryum violaceum, identify mating and pathogenicity genes

PubMed Central

Yockteng, Roxana; Marthey, Sylvain; Chiapello, Hélène; Gendrault, Annie; Hood, Michael E; Rodolphe, François; Devier, Benjamin; Wincker, Patrick; Dossat, Carole; Giraud, Tatiana

2007-01-01

Background The basidiomycete fungus Microbotryum violaceum is responsible for the anther-smut disease in many plants of the Caryophyllaceae family and is a model in genetics and evolutionary biology. Infection is initiated by dikaryotic hyphae produced after the conjugation of two haploid sporidia of opposite mating type. This study describes M. violaceum ESTs corresponding to nuclear genes expressed during conjugation and early hyphal production. Results A normalized cDNA library generated 24,128 sequences, which were assembled into 7,765 unique genes; 25.2% of them displayed significant similarity to annotated proteins from other organisms, 74.3% a weak similarity to the same set of known proteins, and 0.5% were orphans. We identified putative pheromone receptors and genes that in other fungi are involved in the mating process. We also identified many sequences similar to genes known to be involved in pathogenicity in other fungi. The M. violaceum EST database, MICROBASE, is available on the Web and provides access to the sequences, assembled contigs, annotations and programs to compare similarities against MICROBASE. Conclusion This study provides a basis for cloning the mating type locus, for further investigation of pathogenicity genes in the anther smut fungi, and for comparative genomics. PMID:17692127

Global gene expression analysis of peripheral blood mononuclear cells in rhesus monkey infants with CA16 infection-induced HFMD.

PubMed

Song, Jie; Hu, Yajie; Hu, Yunguang; Wang, Jingjing; Zhang, Xiaolong; Wang, Lichun; Guo, Lei; Wang, Yancui; Ning, Ruotong; Liao, Yun; Zhang, Ying; Zheng, Huiwen; Shi, Haijing; He, Zhanlong; Li, Qihan; Liu, Longding

2016-03-02

Coxsackievirus A16 (CA16) is a dominant pathogen that results in hand, foot, and mouth disease and causes outbreaks worldwide, particularly in the Asia-Pacific region. However, the underlying molecular mechanisms remain unclear. Our previous study has demonstrated that the basic CA16 pathogenic process was successfully mimicked in rhesus monkey infant. The present study focused on the global gene expression changes in peripheral blood mononuclear cells of rhesus monkey infants with hand, foot, and mouth disease induced by CA16 infection at different time points. Genome-wide expression analysis was performed with Agilent whole-genome microarrays and established bioinformatics tools. Nine hundred and forty-eight significant differentially expressed genes that were associated with 5 gene ontology categories, including cell communication, cell cycle, immune system process, regulation of transcription and metabolic process were identified. Subsequently, the mapping of genes related to the immune system process by PANTHER pathway analysis revealed the predominance of inflammation mediated by chemokine and cytokine signaling pathways and the interleukin signaling pathway. Ultimately, co-expressed genes and their networks were analyzed. The results revealed the gene expression profile of the immune system in response to CA16 in rhesus monkey infants and suggested that such an immune response was generated as a result of the positive mobilization of the immune system. This initial microarray study will provide insights into the molecular mechanism of CA16 infection and will facilitate the identification of biomarkers for the evaluation of vaccines against this virus. Copyright © 2016 Elsevier B.V. All rights reserved.

Gene expression in honey bee (Apis mellifera) larvae exposed to pesticides and Varroa mites (Varroa destructor).

PubMed

Gregorc, Aleš; Evans, Jay D; Scharf, Mike; Ellis, James D

2012-08-01

Honey bee (Apis mellifera) larvae reared in vitro were exposed to one of nine pesticides and/or were challenged with the parasitic mite, Varroa destructor. Total RNA was extracted from individual larvae and first strand cDNAs were generated. Gene-expression changes in larvae were measured using quantitative PCR (qPCR) targeting transcripts for pathogens and genes involved in physiological processes, bee health, immunity, and/or xenobiotic detoxification. Transcript levels for Peptidoglycan Recognition Protein (PGRPSC), a pathogen recognition gene, increased in larvae exposed to Varroa mites (P<0.001) and were not changed in pesticide treated larvae. As expected, Varroa-parasitized brood had higher transcripts of Deformed Wing Virus than did control larvae (P<0.001). Varroa parasitism, arguably coupled with virus infection, resulted in significantly higher transcript abundances for the antimicrobial peptides abaecin, hymenoptaecin, and defensin1. Transcript levels for Prophenoloxidase-activating enzyme (PPOact), an immune end product, were elevated in larvae treated with myclobutanil and chlorothalonil (both are fungicides) (P<0.001). Transcript levels for Hexameric storage protein (Hsp70) were significantly upregulated in imidacloprid, fluvalinate, coumaphos, myclobutanil, and amitraz treated larvae. Definitive impacts of pesticides and Varroa parasitism on honey bee larval gene expression were demonstrated. Interactions between larval treatments and gene expression for the targeted genes are discussed. Copyright © 2012 Elsevier Ltd. All rights reserved.

Pathogenic mechanisms of intracellular bacteria.

PubMed

Niller, Hans Helmut; Masa, Roland; Venkei, Annamária; Mészáros, Sándor; Minarovits, Janos

2017-06-01

We wished to overview recent data on a subset of epigenetic changes elicited by intracellular bacteria in human cells. Reprogramming the gene expression pattern of various host cells may facilitate bacterial growth, survival, and spread. DNA-(cytosine C5)-methyltransferases of Mycoplasma hyorhinis targeting cytosine-phosphate-guanine (CpG) dinucleotides and a Mycobacterium tuberculosis methyltransferase targeting non-CpG sites methylated the host cell DNA and altered the pattern of gene expression. Gene silencing by CpG methylation and histone deacetylation, mediated by cellular enzymes, also occurred in M. tuberculosis-infected macrophages. M. tuberculosis elicited cell type-specific epigenetic changes: it caused increased DNA methylation in macrophages, but induced demethylation, deposition of euchromatic histone marks and activation of immune-related genes in dendritic cells. A secreted transposase of Acinetobacter baumannii silenced a cellular gene, whereas Mycobacterium leprae altered the epigenotype, phenotype, and fate of infected Schwann cells. The 'keystone pathogen' oral bacterium Porphyromonas gingivalis induced local DNA methylation and increased the level of histone acetylation in host cells. These epigenetic changes at the biofilm-gingiva interface may contribute to the development of periodontitis. Epigenetic regulators produced by intracellular bacteria alter the epigenotype and gene expression pattern of host cells and play an important role in pathogenesis.

Changes in transcript abundance relating to colony collapse disorder in honey bees (Apis mellifera).

PubMed

Johnson, Reed M; Evans, Jay D; Robinson, Gene E; Berenbaum, May R

2009-09-01

Colony collapse disorder (CCD) is a mysterious disappearance of honey bees that has beset beekeepers in the United States since late 2006. Pathogens and other environmental stresses, including pesticides, have been linked to CCD, but a causal relationship has not yet been demonstrated. Because the gut acts as a primary interface between the honey bee and its environment as a site of entry for pathogens and toxins, we used whole-genome microarrays to compare gene expression between guts of bees from CCD colonies originating on both the east and west coasts of the United States and guts of bees from healthy colonies sampled before the emergence of CCD. Considerable variation in gene expression was associated with the geographical origin of bees, but a consensus list of 65 transcripts was identified as potential markers for CCD status. Overall, elevated expression of pesticide response genes was not observed. Genes involved in immune response showed no clear trend in expression pattern despite the increased prevalence of viruses and other pathogens in CCD colonies. Microarray analysis revealed unusual ribosomal RNA fragments that were conspicuously more abundant in the guts of CCD bees. The presence of these fragments may be a possible consequence of picorna-like viral infection, including deformed wing virus and Israeli acute paralysis virus, and may be related to arrested translation. Ribosomal fragment abundance and presence of multiple viruses may prove to be useful diagnostic markers for colonies afflicted with CCD.

Changes in transcript abundance relating to colony collapse disorder in honey bees (Apis mellifera)

PubMed Central

Johnson, Reed M.; Evans, Jay D.; Robinson, Gene E.; Berenbaum, May R.

2009-01-01

Colony collapse disorder (CCD) is a mysterious disappearance of honey bees that has beset beekeepers in the United States since late 2006. Pathogens and other environmental stresses, including pesticides, have been linked to CCD, but a causal relationship has not yet been demonstrated. Because the gut acts as a primary interface between the honey bee and its environment as a site of entry for pathogens and toxins, we used whole-genome microarrays to compare gene expression between guts of bees from CCD colonies originating on both the east and west coasts of the United States and guts of bees from healthy colonies sampled before the emergence of CCD. Considerable variation in gene expression was associated with the geographical origin of bees, but a consensus list of 65 transcripts was identified as potential markers for CCD status. Overall, elevated expression of pesticide response genes was not observed. Genes involved in immune response showed no clear trend in expression pattern despite the increased prevalence of viruses and other pathogens in CCD colonies. Microarray analysis revealed unusual ribosomal RNA fragments that were conspicuously more abundant in the guts of CCD bees. The presence of these fragments may be a possible consequence of picorna-like viral infection, including deformed wing virus and Israeli acute paralysis virus, and may be related to arrested translation. Ribosomal fragment abundance and presence of multiple viruses may prove to be useful diagnostic markers for colonies afflicted with CCD. PMID:19706391

Expanding the genetic toolbox for Leptospira species by generation of fluorescent bacteria.

PubMed

Aviat, Florence; Slamti, Leyla; Cerqueira, Gustavo M; Lourdault, Kristel; Picardeau, Mathieu

2010-12-01

Our knowledge of the genetics and molecular basis of the pathogenesis associated with Leptospira, in comparison to those of other bacterial species, is very limited. An improved understanding of pathogenic mechanisms requires reliable genetic tools for functional genetic analysis. Here, we report the expression of gfp and mRFP1 genes under the control of constitutive spirochetal promoters in both saprophytic and pathogenic Leptospira strains. We were able to reliably measure the fluorescence of Leptospira by fluorescence microscopy and a fluorometric microplate reader-based assay. We showed that the expression of the gfp gene had no significant effects on growth in vivo and pathogenicity in L. interrogans. We constructed an expression vector for L. biflexa that contains the lacI repressor, an inducible lac promoter, and gfp as the reporter, demonstrating that the lac system is functional in Leptospira. Green fluorescent protein (GFP) expression was induced by the addition of isopropyl-β-d-thiogalactopyranoside (IPTG) in L. biflexa transformants harboring the expression vector. Finally, we showed that GFP can be used as a reporter to assess promoter activity in different environmental conditions. These results may facilitate further advances for studying the genetics of Leptospira spp.

Gene Expression by Mouse Inner Ear Hair Cells during Development

PubMed Central

Scheffer, Déborah I.; Shen, Jun

2015-01-01

Hair cells of the inner ear are essential for hearing and balance. As a consequence, pathogenic variants in genes specifically expressed in hair cells often cause hereditary deafness. Hair cells are few in number and not easily isolated from the adjacent supporting cells, so the biochemistry and molecular biology of hair cells can be difficult to study. To study gene expression in hair cells, we developed a protocol for hair cell isolation by FACS. With nearly pure hair cells and surrounding cells, from cochlea and utricle and from E16 to P7, we performed a comprehensive cell type-specific RNA-Seq study of gene expression during mouse inner ear development. Expression profiling revealed new hair cell genes with distinct expression patterns: some are specific for vestibular hair cells, others for cochlear hair cells, and some are expressed just before or after maturation of mechanosensitivity. We found that many of the known hereditary deafness genes are much more highly expressed in hair cells than surrounding cells, suggesting that genes preferentially expressed in hair cells are good candidates for unknown deafness genes. PMID:25904789

Characterization of recombinant Raccoonpox Vaccine Vectors in Chickens

USGS Publications Warehouse

Hwa, S.-H.; Iams, Keith P.; Hall, Jeffrey S.; Kingstad, B.A.; Osorio, Jorge E.

2010-01-01

Raccoonpox virus (RCN) has been used as a recombinant vector against several mammalian pathogens but has not been tested in birds. The replication of RCN in chick embryo fibroblasts (CEFs) and chickens was studied with the use of highly pathogenic avian influenza virus H5N1 hemagglutinin (HA) as a model antigen and luciferase (luc) as a reporter gene. Although RCN replicated to low levels in CEFs, it efficiently expressed recombinant proteins and, in vivo, elicited anti-HA immunoglobulin yolk (IgY) antibody responses comparable to inactivated influenza virus. Biophotonic in vivo imaging of 1-wk-old chicks with RCN-luc showed strong expression of the luc reporter gene lasting up to 3 days postinfection. These studies demonstrate the potential of RCN as a vaccine vector for avian influenza and other poultry pathogens. ?? American Association of Avian Pathologists 2010.

Guava Leaf Extract Inhibits Quorum-Sensing and Chromobacterium violaceum Induced Lysis of Human Hepatoma Cells: Whole Transcriptome Analysis Reveals Differential Gene Expression

PubMed Central

Tiwary, Bipransh Kumar; Kumar, Anoop

2014-01-01

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value≤0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum. PMID:25229331

Guava leaf extract inhibits quorum-sensing and Chromobacterium violaceum induced lysis of human hepatoma cells: whole transcriptome analysis reveals differential gene expression.

PubMed

Ghosh, Runu; Tiwary, Bipransh Kumar; Kumar, Anoop; Chakraborty, Ranadhir

2014-01-01

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value ≤ 0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum.

Comparison of virulence factors and expression of specific genes between uropathogenic Escherichia coli and avian pathogenic E. coli in a murine urinary tract infection model and a chicken challenge model.

PubMed

Zhao, Lixiang; Gao, Song; Huan, Haixia; Xu, Xiaojing; Zhu, Xiaoping; Yang, Weixia; Gao, Qingqing; Liu, Xiufan

2009-05-01

Avian pathogenic Escherichia coli (APEC) and uropathogenic E. coli (UPEC) establish infections in extraintestinal habitats of different hosts. As the diversity, epidemiological sources and evolutionary origins of extraintestinal pathogenic E. coli (ExPEC) are so far only partially defined, in the present study,100 APEC isolates and 202 UPEC isolates were compared by their content of virulence genes and phylogenetic groups. The two groups showed substantial overlap in terms of their serogroups, phylogenetic groups and virulence genotypes, including their possession of certain genes associated with large transmissible plasmids of APEC. In a chicken challenge model, both UPEC U17 and APEC E058 had similar LD(50), demonstrating that UPEC U17 had the potential to cause significant disease in poultry. To gain further information about the similarities between UPEC and APEC, the in vivo expression of 152 specific genes of UPEC U17 and APEC E058 in both a murine urinary tract infection (UTI) model and a chicken challenge model was compared with that of these strains grown statically to exponential phase in rich medium. It was found that in the same model (murine UTI or chicken challenge), various genes of UPEC U17 and APEC E058 showed a similar tendency of expression. Several iron-related genes were upregulated in the UTI model and/or chicken challenge model, indicating that iron acquisition is important for E. coli to survive in blood or the urinary tract. Based on these results, the potential for APEC to act as human UPEC or as a reservoir of virulence genes for UPEC should be considered. Further, this study compared the transcriptional profile of virulence genes among APEC and UPEC in vivo.

Urbanization Increases Pathogen Pressure on Feral and Managed Honey Bees.

PubMed

Youngsteadt, Elsa; Appler, R Holden; López-Uribe, Margarita M; Tarpy, David R; Frank, Steven D

2015-01-01

Given the role of infectious disease in global pollinator decline, there is a need to understand factors that shape pathogen susceptibility and transmission in bees. Here we ask how urbanization affects the immune response and pathogen load of feral and managed colonies of honey bees (Apis mellifera Linnaeus), the predominant economically important pollinator worldwide. Using quantitative real-time PCR, we measured expression of 4 immune genes and relative abundance of 10 honey bee pathogens. We also measured worker survival in a laboratory bioassay. We found that pathogen pressure on honey bees increased with urbanization and management, and the probability of worker survival declined 3-fold along our urbanization gradient. The effect of management on pathogens appears to be mediated by immunity, with feral bees expressing immune genes at nearly twice the levels of managed bees following an immune challenge. The effect of urbanization, however, was not linked with immunity; instead, urbanization may favor viability and transmission of some disease agents. Feral colonies, with lower disease burdens and stronger immune responses, may illuminate ways to improve honey bee management. The previously unexamined effects of urbanization on honey-bee disease are concerning, suggesting that urban areas may favor problematic diseases of pollinators.

Urbanization Increases Pathogen Pressure on Feral and Managed Honey Bees

PubMed Central

López-Uribe, Margarita M.; Tarpy, David R.; Frank, Steven D.

2015-01-01

Given the role of infectious disease in global pollinator decline, there is a need to understand factors that shape pathogen susceptibility and transmission in bees. Here we ask how urbanization affects the immune response and pathogen load of feral and managed colonies of honey bees (Apis mellifera Linnaeus), the predominant economically important pollinator worldwide. Using quantitative real-time PCR, we measured expression of 4 immune genes and relative abundance of 10 honey bee pathogens. We also measured worker survival in a laboratory bioassay. We found that pathogen pressure on honey bees increased with urbanization and management, and the probability of worker survival declined 3-fold along our urbanization gradient. The effect of management on pathogens appears to be mediated by immunity, with feral bees expressing immune genes at nearly twice the levels of managed bees following an immune challenge. The effect of urbanization, however, was not linked with immunity; instead, urbanization may favor viability and transmission of some disease agents. Feral colonies, with lower disease burdens and stronger immune responses, may illuminate ways to improve honey bee management. The previously unexamined effects of urbanization on honey-bee disease are concerning, suggesting that urban areas may favor problematic diseases of pollinators. PMID:26536606

Genomic analyses and expression evaluation of thaumatin-like gene family in the cacao fungal pathogen Moniliophthora perniciosa.

PubMed

Franco, Sulamita de Freitas; Baroni, Renata Moro; Carazzolle, Marcelo Falsarella; Teixeira, Paulo José Pereira Lima; Reis, Osvaldo; Pereira, Gonçalo Amarante Guimarães; Mondego, Jorge Maurício Costa

2015-10-30

Thaumatin-like proteins (TLPs) are found in diverse eukaryotes. Plant TLPs, known as Pathogenicity Related Protein (PR-5), are considered fungal inhibitors. However, genes encoding TLPs are frequently found in fungal genomes. In this work, we have identified that Moniliophthora perniciosa, a basidiomycete pathogen that causes the Witches' Broom Disease (WBD) of cacao, presents thirteen putative TLPs from which four are expressed during WBD progression. One of them is similar to small TLPs, which are present in phytopathogenic basidiomycete, such as wheat stem rust fungus Puccinia graminis. Fungi genomes annotation and phylogenetic data revealed a larger number of TLPs in basidiomycetes when comparing with ascomycetes, suggesting that these proteins could be involved in specific traits of mushroom-forming species. Based on the present data, we discuss the contribution of TLPs in the combat against fungal competitors and hypothesize a role of these proteins in M. perniciosa pathogenicity. Copyright © 2015 Elsevier Inc. All rights reserved.

Living in two worlds: the plant and insect lifestyles of Xylella fastidiosa.

PubMed

Chatterjee, Subhadeep; Almeida, Rodrigo P P; Lindow, Steven

2008-01-01

Diseases caused by Xylella fastidiosa have attained great importance worldwide as the pathogen and its insect vectors have been disseminated. Since this is the first plant pathogenic bacterium for which a complete genome sequence was determined, much progress has been made in understanding the process by which it spreads within the xylem vessels of susceptible plants as well as the traits that contribute to its acquisition and transmission by sharpshooter vectors. Although this pathogen shares many similarities with Xanthomonas species, such as its use of a small fatty acid signal molecule to coordinate virulence gene expression, the traits that it utilizes to cause disease and the manner in which they are regulated differ substantially from those of related plant pathogens. Its complex lifestyle as both a plant and insect colonist involves traits that are in conflict with these stages, thus apparently necessitating the use of a gene regulatory scheme that allows cells expressing different traits to co-occur in the plant.

Divergent and convergent modes of interaction between wheat and Puccinia graminis f. sp. tritici isolates revealed by the comparative gene co-expression network and genome analyses.

PubMed

Rutter, William B; Salcedo, Andres; Akhunova, Alina; He, Fei; Wang, Shichen; Liang, Hanquan; Bowden, Robert L; Akhunov, Eduard

2017-04-12

Two opposing evolutionary constraints exert pressure on plant pathogens: one to diversify virulence factors in order to evade plant defenses, and the other to retain virulence factors critical for maintaining a compatible interaction with the plant host. To better understand how the diversified arsenals of fungal genes promote interaction with the same compatible wheat line, we performed a comparative genomic analysis of two North American isolates of Puccinia graminis f. sp. tritici (Pgt). The patterns of inter-isolate divergence in the secreted candidate effector genes were compared with the levels of conservation and divergence of plant-pathogen gene co-expression networks (GCN) developed for each isolate. Comprative genomic analyses revealed substantial level of interisolate divergence in effector gene complement and sequence divergence. Gene Ontology (GO) analyses of the conserved and unique parts of the isolate-specific GCNs identified a number of conserved host pathways targeted by both isolates. Interestingly, the degree of inter-isolate sub-network conservation varied widely for the different host pathways and was positively associated with the proportion of conserved effector candidates associated with each sub-network. While different Pgt isolates tended to exploit similar wheat pathways for infection, the mode of plant-pathogen interaction varied for different pathways with some pathways being associated with the conserved set of effectors and others being linked with the diverged or isolate-specific effectors. Our data suggest that at the intra-species level pathogen populations likely maintain divergent sets of effectors capable of targeting the same plant host pathways. This functional redundancy may play an important role in the dynamic of the "arms-race" between host and pathogen serving as the basis for diverse virulence strategies and creating conditions where mutations in certain effector groups will not have a major effect on the pathogen's ability to infect the host.

Identification of the Regulon of AphB and Its Essential Roles in LuxR and Exotoxin Asp Expression in the Pathogen Vibrio alginolyticus.

PubMed

Gao, Xiating; Liu, Yang; Liu, Huan; Yang, Zhen; Liu, Qin; Zhang, Yuanxing; Wang, Qiyao

2017-10-15

In Vibrio species, AphB is essential to activate virulence cascades by sensing low-pH and anaerobiosis signals; however, its regulon remains largely unknown. Here, AphB is found to be a key virulence regulator in Vibrio alginolyticus , a pathogen for marine animals and humans. Chromatin immunoprecipitation followed by high-throughput DNA sequencing (ChIP-seq) enabled the detection of 20 loci in the V. alginolyticus genome that contained AphB-binding peaks. An AphB-specific binding consensus was confirmed by electrophoretic mobility shift assays (EMSAs), and the regulation of genes flanking such binding sites was demonstrated using quantitative real-time PCR analysis. AphB binds directly to its own promoter and positively controls its own expression in later growth stages. AphB also activates the expression of the exotoxin Asp by binding directly to the promoter regions of asp and the master quorum-sensing (QS) regulator luxR DNase I footprinting analysis uncovered distinct AphB-binding sites (BBS) in these promoters. Furthermore, a BBS in the luxR promoter region overlaps that of LuxR-binding site I, which mediates the positive control of luxR promoter activity by AphB. This study provides new insights into the AphB regulon and reveals the mechanisms underlying AphB regulation of physiological adaptation and QS-controlled virulence in V. alginolyticus IMPORTANCE In this work, AphB is determined to play essential roles in the expression of genes associated with QS, physiology, and virulence in V. alginolyticus , a pathogen for marine animals and humans. AphB was found to bind directly to 20 genes and control their expression by a 17-bp consensus binding sequence. Among the 20 genes, the aphB gene itself was identified to be positively autoregulated, and AphB also positively controlled asp and luxR expression. Taken together, these findings improve our understanding of the roles of AphB in controlling physiological adaptation and QS-controlled virulence gene expression. Copyright © 2017 American Society for Microbiology.

Indications for distinct pathogenic mechanisms of asbestos and silica through gene expression profiling of the response of lung epithelial cells

PubMed Central

Perkins, Timothy N.; Peeters, Paul M.; Shukla, Arti; Arijs, Ingrid; Dragon, Julie; Wouters, Emiel F.M.; Reynaert, Niki L.; Mossman, Brooke T.

2015-01-01

Occupational and environmental exposures to airborne asbestos and silica are associated with the development of lung fibrosis in the forms of asbestosis and silicosis, respectively. However, both diseases display distinct pathologic presentations, likely associated with differences in gene expression induced by different mineral structures, composition and bio-persistent properties. We hypothesized that effects of mineral exposure in the airway epithelium may dictate deviating molecular events that may explain the different pathologies of asbestosis versus silicosis. Using robust gene expression-profiling in conjunction with in-depth pathway analysis, we assessed early (24 h) alterations in gene expression associated with crocidolite asbestos or cristobalite silica exposures in primary human bronchial epithelial cells (NHBEs). Observations were confirmed in an immortalized line (BEAS-2B) by QRT-PCR and protein assays. Utilization of overall gene expression, unsupervised hierarchical cluster analysis and integrated pathway analysis revealed gene alterations that were common to both minerals or unique to either mineral. Our findings reveal that both minerals had potent effects on genes governing cell adhesion/migration, inflammation, and cellular stress, key features of fibrosis. Asbestos exposure was most specifically associated with aberrant cell proliferation and carcinogenesis, whereas silica exposure was highly associated with additional inflammatory responses, as well as pattern recognition, and fibrogenesis. These findings illustrate the use of gene-profiling as a means to determine early molecular events that may dictate pathological processes induced by exogenous cellular insults. In addition, it is a useful approach for predicting the pathogenicity of potentially harmful materials. PMID:25351596

Changing the Game: Using Integrative Genomics to Probe Virulence Mechanisms of the Stem Rust Pathogen Puccinia graminis f. sp. tritici.

PubMed

Figueroa, Melania; Upadhyaya, Narayana M; Sperschneider, Jana; Park, Robert F; Szabo, Les J; Steffenson, Brian; Ellis, Jeff G; Dodds, Peter N

2016-01-01

The recent resurgence of wheat stem rust caused by new virulent races of Puccinia graminis f. sp. tritici (Pgt) poses a threat to food security. These concerns have catalyzed an extensive global effort toward controlling this disease. Substantial research and breeding programs target the identification and introduction of new stem rust resistance (Sr) genes in cultivars for genetic protection against the disease. Such resistance genes typically encode immune receptor proteins that recognize specific components of the pathogen, known as avirulence (Avr) proteins. A significant drawback to deploying cultivars with single Sr genes is that they are often overcome by evolution of the pathogen to escape recognition through alterations in Avr genes. Thus, a key element in achieving durable rust control is the deployment of multiple effective Sr genes in combination, either through conventional breeding or transgenic approaches, to minimize the risk of resistance breakdown. In this situation, evolution of pathogen virulence would require changes in multiple Avr genes in order to bypass recognition. However, choosing the optimal Sr gene combinations to deploy is a challenge that requires detailed knowledge of the pathogen Avr genes with which they interact and the virulence phenotypes of Pgt existing in nature. Identifying specific Avr genes from Pgt will provide screening tools to enhance pathogen virulence monitoring, assess heterozygosity and propensity for mutation in pathogen populations, and confirm individual Sr gene functions in crop varieties carrying multiple effective resistance genes. Toward this goal, much progress has been made in assembling a high quality reference genome sequence for Pgt, as well as a Pan-genome encompassing variation between multiple field isolates with diverse virulence spectra. In turn this has allowed prediction of Pgt effector gene candidates based on known features of Avr genes in other plant pathogens, including the related flax rust fungus. Upregulation of gene expression in haustoria and evidence for diversifying selection are two useful parameters to identify candidate Avr genes. Recently, we have also applied machine learning approaches to agnostically predict candidate effectors. Here, we review progress in stem rust pathogenomics and approaches currently underway to identify Avr genes recognized by wheat Sr genes.

Unique morphological changes in plant pathogenic phytoplasma-infected petunia flowers are related to transcriptional regulation of floral homeotic genes in an organ-specific manner.

PubMed

Himeno, Misako; Neriya, Yutaro; Minato, Nami; Miura, Chihiro; Sugawara, Kyoko; Ishii, Yoshiko; Yamaji, Yasuyuki; Kakizawa, Shigeyuki; Oshima, Kenro; Namba, Shigetou

2011-09-01

Abnormal flowers are often induced by infection of certain plant pathogens, e.g. phytoplasma, but the molecular mechanisms underlying these malformations have remained poorly understood. Here, we show that infection with OY-W phytoplasma (Candidatus Phytoplasma asteris, onion yellows phytoplasma strain, line OY-W) affects the expression of the floral homeotic genes of petunia plants in an organ-specific manner. Upon infection with OY-W phytoplasma, floral morphological changes, including conversion to leaf-like structures, were observed in sepals, petals and pistils, but not in stamens. As the expression levels of homeotic genes differ greatly between floral organs, we examined the expression levels of homeotic genes in each floral organ infected by OY-W phytoplasma, compared with healthy plants. The expression levels of several homeotic genes required for organ development, such as PFG, PhGLO1 and FBP7, were significantly downregulated by the phytoplasma infection in floral organs, except the stamens, suggesting that the unique morphological changes caused by the phytoplasma infection might result from the significant decrease in expression of some crucial homeotic genes. Moreover, the expression levels of TER, ALF and DOT genes, which are known to participate in floral meristem identity, were significantly downregulated in the phytoplasma-infected petunia meristems, implying that phytoplasma would affect an upstream signaling pathway of floral meristem identity. Our results suggest that phytoplasma infection may have complex effects on floral development, resulting in the unique phenotypes that were clearly distinct from the mutant flower phenotypes produced by the knock-out or the overexpression of certain homeotic genes. © 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.

Molecular cloning, characterization and expression of immunoglobulin D on pathogen challenge and pathogen associated molecular patterns stimulation in freshwater carp, Catla catla.

PubMed

Banerjee, Rajanya; Patel, Bhakti; Basu, Madhubanti; Lenka, Saswati S; Paicha, Mahismita; Samanta, Mrinal; Das, Surajit

2017-10-01

The primordial immunoglobulin class, IgD, was the first non-IgM isotype discovered in teleosts. The crucial roles of IgM and IgZ in imparting systemic and mucosal immunity, respectively, in various fish species have been widely established. However, the putative function of a unique IgD isotype during pathogenic invasions has not been well explored. The present study reports the existence of an IgD ortholog in freshwater carp, Catla catla, and further evaluates its differential expression profile in response to bacterial, parasitic and viral antigenic exposure and pathogen associated molecular patterns (PAMPs) stimulation. The IgD of C. catla (CcIgD) cDNA sequence was found to encode 226 amino acids and confirmed homology with heavy chain delta region of Cyprinidae family members. Phylogenetic analysis of CcIgD exhibited greatest similarity with Ctenopharyngodon idella. qRT-PCR analysis revealed significant upregulation (P < 0.001) of IgD gene expression in kidney with respect to other tissues at 24 hr post-Aeromonas hydrophila challenge. CcIgD gene expression in skin was enhanced following Streptococcus uberis infection and in blood following Argulus infection and inactivated rhabdoviral antigen stimulation. Further, the treatment of bacterial and viral products (PAMPs) also triggered significant (P < 0.05) increases in CcIgD mRNA expression in kidney. These findings indicate the functional importance of teleost IgD in orchestrating tissue specific neutralization of antigens on stimulation with different pathogens and PAMPs. © 2017 The Societies and John Wiley & Sons Australia, Ltd.

Field tolerance to fungal pathogens of Brassica napus constitutively expressing a chimeric chitinase gene

DOE Office of Scientific and Technical Information (OSTI.GOV)

Grison, R.; Grezes-Besset, B.; Lucante, N.

1996-05-01

Constitutive overexpression of a protein involved in plant defense mechanisms to disease is one of the strategies proposed to increase plant tolerance to fungal pathogens. A hybrid endochitinase gene under a constitutive promoter was introduced by Agrobacterium-mediated transformation into a winter-type oilseed rape (Brassica napus var. oleifera) inbred line. Progeny from transformed plants was challenged using three different fungal pathogens (Cylindrosporium concentricum, Phoma lingam, Sclerotinia sclerotiorum) in field trials at two different geographical locations. These plants exhibited an increased tolerance to disease as compared with the nontransgenic parental plants. 31 refs., 1 fig., 2 tabs.

Dual Transcriptome Profiling of Leishmania-Infected Human Macrophages Reveals Distinct Reprogramming Signatures.

PubMed

Fernandes, Maria Cecilia; Dillon, Laura A L; Belew, Ashton Trey; Bravo, Hector Corrada; Mosser, David M; El-Sayed, Najib M

2016-05-10

Macrophages are mononuclear phagocytes that constitute a first line of defense against pathogens. While lethal to many microbes, they are the primary host cells of Leishmania spp. parasites, the obligate intracellular pathogens that cause leishmaniasis. We conducted transcriptomic profiling of two Leishmania species and the human macrophage over the course of intracellular infection by using high-throughput RNA sequencing to characterize the global gene expression changes and reprogramming events that underlie the interactions between the pathogen and its host. A systematic exclusion of the generic effects of large-particle phagocytosis revealed a vigorous, parasite-specific response of the human macrophage early in the infection that was greatly tempered at later time points. An analogous temporal expression pattern was observed with the parasite, suggesting that much of the reprogramming that occurs as parasites transform into intracellular forms generally stabilizes shortly after entry. Following that, the parasite establishes an intracellular niche within macrophages, with minimal communication between the parasite and the host cell later during the infection. No significant difference was observed between parasite species transcriptomes or in the transcriptional response of macrophages infected with each species. Our comparative analysis of gene expression changes that occur as mouse and human macrophages are infected by Leishmania spp. points toward a general signature of the Leishmania-macrophage infectome. Little is known about the transcriptional changes that occur within mammalian cells harboring intracellular pathogens. This study characterizes the gene expression signatures of Leishmania spp. parasites and the coordinated response of infected human macrophages as the pathogen enters and persists within them. After accounting for the generic effects of large-particle phagocytosis, we observed a parasite-specific response of the human macrophages early in infection that was reduced at later time points. A similar expression pattern was observed in the parasites. Our analyses provide specific insights into the interplay between human macrophages and Leishmania parasites and constitute an important general resource for the study of how pathogens evade host defenses and modulate the functions of the cell to survive intracellularly. Copyright © 2016 Fernandes et al.

Helicobacter pylori potentiates epithelial:mesenchymal transition in gastric cancer: links to soluble HB-EGF, gastrin and matrix metalloproteinase-7

PubMed Central

Yin, Yinfei; Grabowska, Anna M; Clarke, Philip A; Whelband, Elisabeth; Robinson, Karen; Argent, Richard H; Tobias, Amanda; Kumari, Rajendra; Atherton, John C

2010-01-01

Background and aims Helicobacter pylori (H pylori) infection is a major risk factor in the development of distal gastric adenocarcinoma. Development of the invasive phenotype is associated with the phenomenon of epithelial:mesenchymal transition (EMT). Soluble heparin-binding epidermal growth factor (HB-EGF) has been implicated in this process. A study was undertaken to investigate the possibility that matrix metalloproteinase (MMP)-7 is upregulated in H pylori infection as a result of hypergastrinaemia, which may enhance shedding of HB-EGF and contribute towards EMT in gastric adenocarcinoma cell lines. Methods Three gastric epithelial cell lines (AGS, MGLVA1 and ST16) were co-cultured with the pathogenic H pylori strain 60190 and non-pathogenic strain Tx30a in an in vitro infection model. Gene expression was quantified by real-time PCR, HB-EGF shedding by ELISA and protein expression by immunofluorescence or immunohistochemistry. The INS-GAS mouse, a transgenic mouse model of gastric carcinogenesis which overexpresses amidated gastrin, was used to investigate the in vivo relationship between HB-EGF, MMP-7, gastrin and EMT. Results The pathogenic strain of H pylori significantly upregulated EMT-associated genes Snail, Slug and vimentin in all three gastric cell lines to a greater degree than the non-pathogenic strain. Pathogenic H pylori also upregulated HB-EGF shedding, a factor implicated in EMT, which was partially dependent on both gastrin and MMP-7 expression. Gastrin and MMP-7 siRNAs and MMP-7 neutralising antibody significantly reduced upregulation of HB-EGF shedding in H pylori infected gastric cell lines and reduced EMT gene expression. The effect of H pylori on EMT was also reversed by gastrin siRNA. Neutralisation of gastrin in the INS-GAS mouse model reduced expression of MMP-7, HB-EGF and key EMT proteins. Conclusion The upregulation of MMP-7 by pathogenic H pylori is partially dependent on gastrin and may have a role in the development of gastric cancer, potentially through EMT, by indirectly increasing levels of soluble HB-EGF. PMID:20584780

Genetic transformation of novel isolates of chicken Lactobacillus bearing probiotic features for expression of heterologous proteins: a tool to develop live oral vaccines

PubMed Central

Mota, Rodrigo M; Moreira, João Luiz S; Souza, Marcelo R; Fátima Horta, M; Teixeira, Santuza MR; Neumann, Elisabeth; Nicoli, Jacques R; Nunes, Álvaro C

2006-01-01

Background The use of lactic acid bacteria as vehicles to delivery antigens to immunize animals is a promising issue. When genetically modified, these bacteria can induce a specific local and systemic immune response against selected pathogens. Gastric acid and bile salts tolerance, production of antagonistic substances against pathogenic microorganisms, and adhesive ability to gut epithelium are other important characteristics that make these bacteria useful for oral immunization. Results Bacteria isolated on de Man, Rogosa and Sharpe medium (MRS) from different gastrointestinal portions of broiler chicks were evaluated for their resistance to artificial gastric acid and bile salts, production of hydrogen peroxide, and cell surface hydrophobicity. Thirty-eight isolates were first typed at species level by PCR amplification of 16S-23S rRNA intergenic spacers using universal primers that anneal within 16S and 23S genes, followed by restriction digestion analyses of PCR amplicons (PCR-ARDRA). An expression cassette was assembled onto the pCR2.1-Topo vector by cloning the promoter, leader peptide, cell wall anchor and terminator sequences derived from the laminin binding S-layer protein gene of L. crispatus strain F5.7 (lbs gene). A sequence encoding the green fluorescent protein (GFP) was inserted as reporter gene, and an erythromycin resistance gene was added as selective marker. All constructs were able to express GFP in the cloning host E. coli XL1-Blue and different Lactobacillus strains as verified by FACS and laser scanning confocal microscopy. Conclusion Lactobacillus isolated from gastrointestinal tract of broiler chickens and selected for probiotic characteristics can be genetically modified by introducing an expression cassette into the lbs locus. The transformed bacteria expressed on its cell wall surface different fluorescent proteins used as reporters of promoter function. It is possible then that similar bacterial model expressing pathogen antigens can be used as live oral vaccines to immunize broilers against infectious diseases. PMID:16396687

Response of the goat mammary gland to infection with Staphylococcus aureus revealed by gene expression profiling in milk somatic and white blood cells

PubMed Central

2012-01-01

Background S. aureus is one of the main pathogens responsible for the intra-mammary infection in dairy ruminants. Although much work has been carried out to understand the complex physiological and cellular events that occur in the mammary gland in response to S. aureus, the protective mechanisms are still poorly understood. The objectives of the present study were to investigate gene expression during the early response of the goat mammary gland to an experimental challenge with S. aureus, in order to better understand the local and systemic response and to compare them in two divergent lines of goat selected for high and low milk somatic cell scores. Results No differences in gene expression were found between high and low SCS (Somatic Cells Score) selection lines. Analysing the two groups together, an expression of 300 genes were found to change from T0 before infection, and T4 at 24 hours and T5 at 30 hours following challenge. In blood derived white blood cells 8 genes showed increased expression between T0 and T5 and 1 gene has reduced expression. The genes showing the greatest increase in expression following challenge (5.65 to 3.16 fold change) play an important role in (i) immune and inflammatory response (NFKB1, TNFAIP6, BASP1, IRF1, PLEK, BATF3); (ii) the regulation of innate resistance to pathogens (PTX3); and (iii) the regulation of cell metabolism (CYTH4, SLC2A6, ARG2). The genes with reduced expression (−1.5 to −2.5 fold) included genes involved in (i) lipid metabolism (ABCG2, FASN), (ii) chemokine, cytokine and intracellular signalling (SPPI), and (iii) cell cytoskeleton and extracellular matrix (KRT19). Conclusions Analysis of genes with differential expression following infection showed an inverse relationship between immune response and lipid metabolism in the early response of the mammary gland to the S. aureus challenge. PTX3 showed a large change in expression in both milk and blood, and is therefore a candidate for further studies on immune response associated with mastitis. PMID:23046560

Trichodiene production in a Trichoderma harzianum erg1-silenced strain provides evidence of the importance of the sterol biosynthetic pathway in inducing plant defense-related gene expression

USDA-ARS?s Scientific Manuscript database

Trichoderma species are often used as biocontrol agents against plant-pathogenic fungi. A complex molecular interaction occurs among the biocontrol agent, the antagonistic fungus, and the plant. Terpenes and sterols produced by the biocontrol fungus have been found to affect gene expression in both ...

Effects of Light and Temperature on Daily Activity and Clock Gene Expression in Two Mosquito Disease Vectors.

PubMed

Rivas, Gustavo B S; Teles-de-Freitas, Rayane; Pavan, Márcio G; Lima, José B P; Peixoto, Alexandre A; Bruno, Rafaela Vieira

2018-06-01

Most organisms feature an endogenous circadian clock capable of synchronization with their environment. The most well-known synchronizing agents are light and temperature. The circadian clock of mosquitoes, vectors of many pathogens, drives important behaviors related to vectoral capacity, including oviposition, host seeking, and hematophagy. Main clock gene expression, as well as locomotor activity patterns, has been identified in Aedes aegypti and Culex quinquefasciatus under artificial light-dark cycles. Given that these mosquito species thrive in tropical areas, it is reasonable to speculate that temperature plays an important role in the circadian clock. Here, we provide data supporting a different hierarchy of light and temperature as zeitgebers of two mosquito species. We recorded their locomotor activity and quantified mRNA expression of the main clock genes in several combinations of light and temperature cycles. We observed that A. aegypti is more sensitive to temperature, while C. quinquefasciatus is more responsive to light. These variations in clock gene expression and locomotor activity may have affected the mosquito species' metabolism, energy expenditure, fitness cost, and pathogen transmission efficiency. Our findings are relevant to chronobiology studies and also have epidemiological implications.

An efficient cDNA-AFLP-based strategy for the identification of putative pathogenicity factors from the potato cyst nematode Globodera rostochiensis.

PubMed

Qin, L; Overmars, H; Helder, J; Popeijus, H; van der Voort, J R; Groenink, W; van Koert, P; Schots, A; Bakker, J; Smant, G

2000-08-01

A new strategy has been designed to identify putative pathogenicity factors from the dorsal or subventral esophageal glands of the potato cyst nematode Globodera rostochiensis. Three independent criteria were used for selection. First, genes of interest should predominantly be expressed in infective second-stage juveniles, and not, or to a far lesser extent, in younger developmental stages. For this, gene expression profiles from five different developmental stages were generated with cDNA-AFLP (amplified fragment length polymorphism). Secondly, the mRNA corresponding to such a putative pathogenicity factor should predominantly be present in the esophageal glands of pre-parasitic juveniles. This was checked by in situ hybridization. As a third criterion, these proteinaceous factors should be preceded by a signal peptide for secretion. Expression profiles of more than 4,000 genes were generated and three up-regulated, dorsal gland-specific proteins preceded by signal peptide for secretion were identified. No dorsal gland genes have been cloned before from plant-parasitic nematodes. The partial sequence of these three factors, A4, A18, and A41, showed no significant homology to any known gene. Their presence in the dorsal glands of infective juveniles suggests that these proteins could be involved in feeding cell initiation, and not in migration in the plant root or in protection against plant defense responses. Finally, the applicability of this new strategy in other plant-microbe interactions is discussed.

Identification of in vivo regulators of the Vibrio cholerae xds gene using a high-throughput genetic selection

PubMed Central

McDonough, EmilyKate; Lazinski, David W.; Camilli, Andrew

2014-01-01

Summary Vibrio cholerae, the causative agent of cholera, remains a threat to public health in areas with inadequate sanitation. As a waterborne pathogen, V. cholerae moves between two dissimilar environments, aquatic reservoirs and the intestinal tract of humans. Accordingly, this pathogen undergoes adaptive shifts in gene expression throughout the different stages of its lifecycle. One particular gene, xds, encodes a secreted exonuclease that was previously identified as being induced during infection. Here we sought to identify regulators responsible for the in vivo-specific induction of xds. A transcriptional fusion of xds to two consecutive antibiotic resistance genes was used to select transposon mutants that had inserted within or adjacent to regulatory genes and thereby caused increased expression of the xds fusion under non-inducing conditions. Large pools of selected insertion sites were sequenced in a high throughput manner using Tn-seq to identify potential mechanisms of xds regulation. Our selection identified the two-component system PhoB/R as the dominant activator of xds expression. In vitro validation confirmed that PhoB, a protein which is only active during phosphate limitation, was responsible for xds activation. Using xds expression as a biosensor of the extracellular phosphate level, we observed that the mouse small intestine is a phosphate-limited environment. PMID:24673931

Ubiquitin-conjugating enzyme E2-like gene associated to pathogen response in Concholepas concholepas: SNP identification and transcription expression.

PubMed

Núñez-Acuña, Gustavo; Aguilar-Espinoza, Andrea; Chávez-Mardones, Jacqueline; Gallardo-Escárate, Cristian

2012-10-01

Ubiquitin-conjugated E2 enzyme (UBE2) is one of the main components of the proteasome degradation cascade. Previous studies have shown an increase of expression levels in individuals challenged to some pathogen organism such as virus and bacteria. The study was to characterize the immune response of UBE2 gene in the gastropod Concholepas concholepas through expression analysis and single nucleotide polymorphisms (SNP) discovery. Hence, UBE2 was identified from a cDNA library by 454 pyrosequencing, while SNP identification and validation were performed using De novo assembly and high resolution melting analysis. Challenge trials with Vibrio anguillarum was carried out to evaluate the relative transcript abundance of UBE2 gene from two to thirty-three hours post-treatment. The results showed a partial UBE2 sequence of 889 base pair (bp) with a partial coding region of 291 bp. SNP variation (A/C) was observed at the 546th position. Individuals challenged by V. anguillarum showed an overexpression of the UBE2 gene, the expression being significantly higher in homozygous individuals (AA) than (CC) or heterozygous individuals (A/C). This study contributes useful information relating to the UBE2 gene and its association with innate immune response in marine invertebrates. Copyright © 2012 Elsevier Ltd. All rights reserved.

Identification of pathogenicity‐related genes in Fusarium oxysporum f. sp. cepae

PubMed Central

Vágány, Viktória; Jackson, Alison C.; Harrison, Richard J.; Rainoni, Alessandro; Clarkson, John P.

2016-01-01

Summary Pathogenic isolates of Fusarium oxysporum, distinguished as formae speciales (f. spp.) on the basis of their host specificity, cause crown rots, root rots and vascular wilts on many important crops worldwide. Fusarium oxysporum f. sp. cepae (FOC) is particularly problematic to onion growers worldwide and is increasing in prevalence in the UK. We characterized 31 F. oxysporum isolates collected from UK onions using pathogenicity tests, sequencing of housekeeping genes and identification of effectors. In onion seedling and bulb tests, 21 isolates were pathogenic and 10 were non‐pathogenic. The molecular characterization of these isolates, and 21 additional isolates comprising other f. spp. and different Fusarium species, was carried out by sequencing three housekeeping genes. A concatenated tree separated the F. oxysporum isolates into six clades, but did not distinguish between pathogenic and non‐pathogenic isolates. Ten putative effectors were identified within FOC, including seven Secreted In Xylem (SIX) genes first reported in F. oxysporum f. sp. lycopersici. Two highly homologous proteins with signal peptides and RxLR motifs (CRX1/CRX2) and a gene with no previously characterized domains (C5) were also identified. The presence/absence of nine of these genes was strongly related to pathogenicity against onion and all were shown to be expressed in planta. Different SIX gene complements were identified in other f. spp., but none were identified in three other Fusarium species from onion. Although the FOC SIX genes had a high level of homology with other f. spp., there were clear differences in sequences which were unique to FOC, whereas CRX1 and C5 genes appear to be largely FOC specific. PMID:26609905

Pathogenic Escherichia coli and food handlers in luxury hotels in Nairobi, Kenya.

PubMed

Onyango, Abel O; Kenya, Eucharia U; Mbithi, John J N; Ng'ayo, Musa O

2009-11-01

The epidemiology and virulence properties of pathogenic Escherichia coli among food handlers in tourist destination hotels in Kenya are largely uncharacterized. This cross-sectional study among consenting 885 food handlers working in nine luxurious tourist hotels in Nairobi, Kenya determined the epidemiology, virulence properties, antibiotics susceptibility profiles and conjugation abilities of pathogenic Escherichia coli. Pathogenic Escherichia coli was detected among 39 (4.4%) subjects, including 1.8% enteroaggregative Escherichia coli (EAEC) harboring aggR genes, 1.2% enterotoxigenic Escherichia coli (ETEC) expressing both LT and STp toxins, 1.1% enteropathogenic Escherichia coli (EPEC) and 0.2% Shiga-like Escherichia coli (EHEC) both harboring eaeA and stx2 genes respectively. All the pathotypes had increased surface hydrophobicity. Using multivariate analyses, food handlers with loose stools were more likely to be infected with pathogenic Escherichia coli. Majority 53.8% of the pathotypes were resistant to tetracycline with 40.2% being multi-drug resistant. About 85.7% pathotypes trans-conjugated with Escherichia coli K12 F(-) NA(r) LA. The carriage of multi-drug resistant, toxin expressing pathogenic Escherichia coli by this population is of public health concern because exposure to low doses can result in infection. Screening food handlers and implementing public awareness programs is recommended as an intervention to control transmission of enteric pathogens.

Virulence control in group A Streptococcus by a two-component gene regulatory system: global expression profiling and in vivo infection modeling.

PubMed

Graham, Morag R; Smoot, Laura M; Migliaccio, Cristi A Lux; Virtaneva, Kimmo; Sturdevant, Daniel E; Porcella, Stephen F; Federle, Michael J; Adams, Gerald J; Scott, June R; Musser, James M

2002-10-15

Two-component gene regulatory systems composed of a membrane-bound sensor and cytoplasmic response regulator are important mechanisms used by bacteria to sense and respond to environmental stimuli. Group A Streptococcus, the causative agent of mild infections and life-threatening invasive diseases, produces many virulence factors that promote survival in humans. A two-component regulatory system, designated covRS (cov, control of virulence; csrRS), negatively controls expression of five proven or putative virulence factors (capsule, cysteine protease, streptokinase, streptolysin S, and streptodornase). Inactivation of covRS results in enhanced virulence in mouse models of invasive disease. Using DNA microarrays and quantitative RT-PCR, we found that CovR influences transcription of 15% (n = 271) of all chromosomal genes, including many that encode surface and secreted proteins mediating host-pathogen interactions. CovR also plays a central role in gene regulatory networks by influencing expression of genes encoding transcriptional regulators, including other two-component systems. Differential transcription of genes influenced by covR also was identified in mouse soft-tissue infection. This analysis provides a genome-scale overview of a virulence gene network in an important human pathogen and adds insight into the molecular mechanisms used by group A Streptococcus to interact with the host, promote survival, and cause disease.

Tissue-specific and pathogen-induced regulation of a Nicotiana plumbaginifolia beta-1,3-glucanase gene.

PubMed Central

Castresana, C; de Carvalho, F; Gheysen, G; Habets, M; Inzé, D; Van Montagu, M

1990-01-01

The Nicotiana plumbaginifolia gn1 gene encoding a beta-1,3-glucanase isoform has been characterized. The gn1 product represents an isoform distinct from the previously identified tobacco beta-1,3-glucanases. By expressing gn1 in Escherichia coli, we have determined directly that the encoded protein does, indeed, correspond to a beta-1,3-glucanase. In N. plumbaginifolia, gn1 was found to be expressed in roots and older leaves. Transgenic tobacco plants containing the 5'-noncoding region of gn1 fused to the beta-glucuronidase (GUS) reporter gene also showed maximum levels of GUS activity in roots and older leaves. No detectable activity was present in the upper part of the transgenic plants with the exception of stem cells at the bases of emerging shoots. The expression conferred by the gn1 promoter was differentially induced in response to specific plant stress treatments. Studies of three plant-bacteria interactions showed high levels of GUS activity when infection resulted in a hypersensitive reaction. Increased gene expression was confined to cells surrounding the necrotic lesions. The observed expression pattern suggests that the characterized beta-1,3-glucanase plays a role both in plant development and in the defense response against pathogen infection. PMID:2152158

A HIGHLY SELECTIVE PCR PROTOCOL FOR DETECTING 16S RRNA GENES OF THE GENUS PSEUDOMONAS (SENSU STRICTO) IN ENVIRONMENTAL SAMPLES

EPA Science Inventory

Pseudomonas species are plant, animal, and human pathogens; exhibit plant pathogen-suppressing properties useful in biological control; or express metabolic versatilities valued in biotechnology and bioremediation. Specific detection of Pseudomonas species in the environment may ...

The silencing suppressor (NSs) protein of the plant virus Tomato spotted wilt virus enhances heterologous protein expression and baculovirus pathogenicity in cells and lepidopteran insects.

PubMed

de Oliveira, Virgínia Carla; da Silva Morgado, Fabricio; Ardisson-Araújo, Daniel Mendes Pereira; Resende, Renato Oliveira; Ribeiro, Bergmann Morais

2015-11-01

In this work, we showed that cell death induced by a recombinant (vAcNSs) Autographa californica multiple nucleopolyhedrovirus (AcMNPV) expressing the silencing suppressor (NSs) protein of Tomato spotted wilt virus (TSWV) was enhanced on permissive and semipermissive cell lines. The expression of a heterologous gene (firefly luciferase) during co-infection of insect cells with vAcNSs and a second recombinant baculovirus (vAgppolhfluc) was shown to increase when compared to single vAgppolhfluc infections. Furthermore, the vAcNSs mean time-to-death values were significantly lower than those for wild-type AcMNPV on larvae of Spodoptera frugiperda and Anticarsia gemmatalis. These results showed that the TSWV-NSs protein could efficiently increase heterologous protein expression in insect cells as well as baculovirus pathogenicity and virulence, probably by suppressing the gene-silencing machinery in insects.

Analysis of a polygalacturonase gene of Ustilago maydis and characterization of the encoded enzyme.

PubMed

Castruita-Domínguez, José P; González-Hernández, Sandra E; Polaina, Julio; Flores-Villavicencio, Lérida L; Alvarez-Vargas, Aurelio; Flores-Martínez, Alberto; Ponce-Noyola, Patricia; Leal-Morales, Carlos A

2014-05-01

Ustilago maydis is a pathogenic fungus that produces the corn smut. It is a biotrophic parasite that depends on living plant tissues for its proliferation and development. Polygalacturonases are secreted by pathogens to solubilize the plant cell-wall and are required for pathogen virulence. In this paper, we report the isolation of a U. maydis polygalacturonase gene (Pgu1) and the functional and structural characterization of the encoded enzyme. The U. maydis Pgu1 gene is expressed when the fungus is grown in liquid culture media containing different carbon sources. In plant tissue, the expression increased as a function of incubation time. Pgu1 gene expression was detected during plant infection around 10 days post-infection with U. maydis FB-D12 strain in combination with teliospore formation. Synthesis and secretion of active recombinant PGU1 were achieved using Pichia pastoris, the purified enzyme had a optimum temperature of 34 °C, optimum pH of 4.5, a Km of 57.84 g/L for polygalacturonic acid, and a Vmax of 28.9 µg/min mg. Structural models of PGU1 based on homologous enzymes yielded a typical right-handed β-helix fold of pectinolytic enzymes classified in the glycosyl hydrolases family 28, and the U. maydis PGU1 is related with endo rather than exo polygalacturonases. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Virulent and pathogenic features on the Cronobacter sakazakii polymyxin resistant pmr mutant strain s-3.

PubMed

Bao, Xuerui; Yang, Ling; Chen, Lequn; Li, Bing; Li, Lin; Li, Yanyan; Xu, Zhenbo

2017-09-01

Cronobacter sakazakii is a well-known opportunistic pathogen responsible for necrotizing enterocolitis, meningitis and septicaemia in the premature, immunocompromised infants and neonates. This pathogen possesses various virulence factors and regulatory systems, and pmrA/pmrB regulatory system has been identified in a variety of bacterial species. The current study aims to investigate role of pmrA gene in the pathogenicity and virulence characteristics of Cronobacter sakazakii using whole genome sequencing and RNA-seq. Results demonstrated that the absence of pmrA has the potential to affect Cronobacter sakazakii on its pathogenicity, virulence and resistance abilities by regulating expression of numerous related genes, including CusB, CusC, CusR and ESA_pESA3p05434. Copyright © 2017 Elsevier Ltd. All rights reserved.

Human microbiomes and their roles in dysbiosis, common diseases, and novel therapeutic approaches.

PubMed

Belizário, José E; Napolitano, Mauro

2015-01-01

The human body is the residence of a large number of commensal (non-pathogenic) and pathogenic microbial species that have co-evolved with the human genome, adaptive immune system, and diet. With recent advances in DNA-based technologies, we initiated the exploration of bacterial gene functions and their role in human health. The main goal of the human microbiome project is to characterize the abundance, diversity and functionality of the genes present in all microorganisms that permanently live in different sites of the human body. The gut microbiota expresses over 3.3 million bacterial genes, while the human genome expresses only 20 thousand genes. Microbe gene-products exert pivotal functions via the regulation of food digestion and immune system development. Studies are confirming that manipulation of non-pathogenic bacterial strains in the host can stimulate the recovery of the immune response to pathogenic bacteria causing diseases. Different approaches, including the use of nutraceutics (prebiotics and probiotics) as well as phages engineered with CRISPR/Cas systems and quorum sensing systems have been developed as new therapies for controlling dysbiosis (alterations in microbial community) and common diseases (e.g., diabetes and obesity). The designing and production of pharmaceuticals based on our own body's microbiome is an emerging field and is rapidly growing to be fully explored in the near future. This review provides an outlook on recent findings on the human microbiomes, their impact on health and diseases, and on the development of targeted therapies.

Human microbiomes and their roles in dysbiosis, common diseases, and novel therapeutic approaches

PubMed Central

Belizário, José E.; Napolitano, Mauro

2015-01-01

The human body is the residence of a large number of commensal (non-pathogenic) and pathogenic microbial species that have co-evolved with the human genome, adaptive immune system, and diet. With recent advances in DNA-based technologies, we initiated the exploration of bacterial gene functions and their role in human health. The main goal of the human microbiome project is to characterize the abundance, diversity and functionality of the genes present in all microorganisms that permanently live in different sites of the human body. The gut microbiota expresses over 3.3 million bacterial genes, while the human genome expresses only 20 thousand genes. Microbe gene-products exert pivotal functions via the regulation of food digestion and immune system development. Studies are confirming that manipulation of non-pathogenic bacterial strains in the host can stimulate the recovery of the immune response to pathogenic bacteria causing diseases. Different approaches, including the use of nutraceutics (prebiotics and probiotics) as well as phages engineered with CRISPR/Cas systems and quorum sensing systems have been developed as new therapies for controlling dysbiosis (alterations in microbial community) and common diseases (e.g., diabetes and obesity). The designing and production of pharmaceuticals based on our own body’s microbiome is an emerging field and is rapidly growing to be fully explored in the near future. This review provides an outlook on recent findings on the human microbiomes, their impact on health and diseases, and on the development of targeted therapies. PMID:26500616

Comparative study of CXC chemokines modulation in brown trout (Salmo trutta) following infection with a bacterial or viral pathogen.

PubMed

Gorgoglione, Bartolomeo; Zahran, Eman; Taylor, Nick G H; Feist, Stephen W; Zou, Jun; Secombes, Christopher J

2016-03-01

Chemokine modulation in response to pathogens still needs to be fully characterised in fish, in view of the recently described novel chemokines present. This paper reports the first comparative study of CXC chemokine genes transcription in salmonids (brown trout), with a particular focus on the fish specific CXC chemokines (CXCL_F). Adopting new primer sets, optimised to specifically target mRNA, a RT-qPCR gene screening was carried out. Constitutive gene expression was assessed first in six tissues from SPF brown trout. Transcription modulation was next investigated in kidney and spleen during septicaemic infection induced by a RNA virus (Viral Haemorrhagic Septicaemia virus, genotype Ia) or by a Gram negative bacterium (Yersinia ruckeri, ser. O1/biot. 2). From each target organ specific pathogen burden, measured detecting VHSV-glycoprotein or Y. ruckeri 16S rRNA, and IFN-γ gene expression were analysed for their correlation to chemokine transcription. Both pathogens modulated CXC chemokine gene transcript levels, with marked up-regulation seen in some cases, and with both temporal and tissue specific effects apparent. For example, Y. ruckeri strongly induced chemokine transcription in spleen within 24h, whilst VHS generally induced the largest increases at 3d.p.i. in both tissues. This study gives clues to the role of the novel CXC chemokines, in comparison to the other known CXC chemokines in salmonids. Copyright © 2016 Elsevier Ltd. All rights reserved.

Transcriptional profile of P. syringae pv. phaseolicola NPS3121 at low temperature: Physiology of phytopathogenic bacteria

PubMed Central

2013-01-01

Background Low temperatures play key roles in the development of most plant diseases, mainly because of their influence on the expression of various virulence factors in phytopathogenic bacteria. Thus far, studies regarding this environmental parameter have focused on specific themes and little is known about phytopathogenic bacteria physiology under these conditions. To obtain a global view regarding phytopathogenic bacteria strategies in response to physiologically relevant temperature changes, we used DNA microarray technology to compare the gene expression profile of the model bacterial pathogen P. syringae pv. phaseolicola NPS3121 grown at 18°C and 28°C. Results A total of 236 differentially regulated genes were identified, of which 133 were up-regulated and 103 were down-regulated at 18°C compared to 28°C. The majority of these genes are involved in pathogenicity and virulence processes. In general, the results of this study suggest that the expression profile obtained may be related to the fact that low temperatures induce oxidative stress in bacterial cells, which in turn influences the expression of iron metabolism genes. The expression also appears to be correlated with the profile expression obtained in genes related to motility, biofilm production, and the type III secretion system. Conclusions From the data obtained in this study, we can begin to understand the strategies used by this phytopathogen during low temperature growth, which can occur in host interactions and disease development. PMID:23587016

Genomic evidence for genes encoding leucine-rich repeat receptors linked to resistance against the eukaryotic extra- and intracellular Brassica napus pathogens Leptosphaeria maculans and Plasmodiophora brassicae.

PubMed

Stotz, Henrik U; Harvey, Pascoe J; Haddadi, Parham; Mashanova, Alla; Kukol, Andreas; Larkan, Nicholas J; Borhan, M Hossein; Fitt, Bruce D L

2018-01-01

Genes coding for nucleotide-binding leucine-rich repeat (LRR) receptors (NLRs) control resistance against intracellular (cell-penetrating) pathogens. However, evidence for a role of genes coding for proteins with LRR domains in resistance against extracellular (apoplastic) fungal pathogens is limited. Here, the distribution of genes coding for proteins with eLRR domains but lacking kinase domains was determined for the Brassica napus genome. Predictions of signal peptide and transmembrane regions divided these genes into 184 coding for receptor-like proteins (RLPs) and 121 coding for secreted proteins (SPs). Together with previously annotated NLRs, a total of 720 LRR genes were found. Leptosphaeria maculans-induced expression during a compatible interaction with cultivar Topas differed between RLP, SP and NLR gene families; NLR genes were induced relatively late, during the necrotrophic phase of pathogen colonization. Seven RLP, one SP and two NLR genes were found in Rlm1 and Rlm3/Rlm4/Rlm7/Rlm9 loci for resistance against L. maculans on chromosome A07 of B. napus. One NLR gene at the Rlm9 locus was positively selected, as was the RLP gene on chromosome A10 with LepR3 and Rlm2 alleles conferring resistance against L. maculans races with corresponding effectors AvrLm1 and AvrLm2, respectively. Known loci for resistance against L. maculans (extracellular hemi-biotrophic fungus), Sclerotinia sclerotiorum (necrotrophic fungus) and Plasmodiophora brassicae (intracellular, obligate biotrophic protist) were examined for presence of RLPs, SPs and NLRs in these regions. Whereas loci for resistance against P. brassicae were enriched for NLRs, no such signature was observed for the other pathogens. These findings demonstrate involvement of (i) NLR genes in resistance against the intracellular pathogen P. brassicae and a putative NLR gene in Rlm9-mediated resistance against the extracellular pathogen L. maculans.

Polygalacturonase gene pgxB in Aspergillus niger is a virulence factor in apple fruit.

PubMed

Liu, Cheng-Qian; Hu, Kang-Di; Li, Ting-Ting; Yang, Ying; Yang, Feng; Li, Yan-Hong; Liu, He-Ping; Chen, Xiao-Yan; Zhang, Hua

2017-01-01

Aspergillus niger, a saprophytic fungus, is widely distributed in soil, air and cereals, and can cause postharvest diseases in fruit. Polygalacturonase (PG) is one of the main enzymes in fungal pathogens to degrade plant cell wall. To evaluate whether the deletion of an exo-polygalacturonase gene pgxB would influence fungal pathogenicity to fruit, pgxB gene was deleted in Aspergillus niger MA 70.15 (wild type) via homologous recombination. The ΔpgxB mutant showed similar growth behavior compared with the wild type. Pectin medium induced significant higher expression of all pectinase genes in both wild type and ΔpgxB in comparison to potato dextrose agar medium. However, the ΔpgxB mutant was less virulent on apple fruits as the necrosis diameter caused by ΔpgxB mutant was significantly smaller than that of wild type. Results of quantitive-PCR showed that, in the process of infection in apple fruit, gene expressions of polygalacturonase genes pgaI, pgaII, pgaA, pgaC, pgaD and pgaE were enhanced in ΔpgxB mutant in comparison to wild type. These results prove that, despite the increased gene expression of other polygalacturonase genes in ΔpgxB mutant, the lack of pgxB gene significantly reduced the virulence of A. niger on apple fruit, suggesting that pgxB plays an important role in the infection process on the apple fruit.

Electrotransformation and expression of bacterial genes encoding hygromycin phosphotransferase and beta-galactosidase in the pathogenic fungus Histoplasma capsulatum.

PubMed

Woods, J P; Heinecke, E L; Goldman, W E

1998-04-01

We developed an efficient electrotransformation system for the pathogenic fungus Histoplasma capsulatum and used it to examine the effects of features of the transforming DNA on transformation efficiency and fate of the transforming DNA and to demonstrate fungal expression of two recombinant Escherichia coli genes, hph and lacZ. Linearized DNA and plasmids containing Histoplasma telomeric sequences showed the greatest transformation efficiencies, while the plasmid vector had no significant effect, nor did the derivation of the selectable URA5 marker (native Histoplasma gene or a heterologous Podospora anserina gene). Electrotransformation resulted in more frequent multimerization, other modification, or possibly chromosomal integration of transforming telomeric plasmids when saturating amounts of DNA were used, but this effect was not observed with smaller amounts of transforming DNA. We developed another selection system using a hygromycin B resistance marker from plasmid pAN7-1, consisting of the E. coli hph gene flanked by Aspergillus nidulans promoter and terminator sequences. Much of the heterologous fungal sequences could be removed without compromising function in H. capsulatum, allowing construction of a substantially smaller effective marker fragment. Transformation efficiency increased when nonselective conditions were maintained for a time after electrotransformation before selection with the protein synthesis inhibitor hygromycin B was imposed. Finally, we constructed a readily detectable and quantifiable reporter gene by fusing Histoplasma URA5 with E. coli lacZ, resulting in expression of functional beta-galactosidase in H. capsulatum. Demonstration of expression of bacterial genes as effective selectable markers and reporters, together with a highly efficient electrotransformation system, provide valuable approaches for molecular genetic analysis and manipulation of H. capsulatum, which have proven useful for examination of targeted gene disruption, regulated gene expression, and potential virulence determinants in this fungus.

The role of thionins in rice defence against root pathogens.

PubMed

Ji, Hongli; Gheysen, Godelieve; Ullah, Chhana; Verbeek, Ruben; Shang, Chenjing; De Vleesschauwer, David; Höfte, Monica; Kyndt, Tina

2015-10-01

Thionins are antimicrobial peptides that are involved in plant defence. Here, we present an in-depth analysis of the role of rice thionin genes in defence responses against two root pathogens: the root-knot nematode Meloidogyne graminicola and the oomycete Pythium graminicola. The expression of rice thionin genes was observed to be differentially regulated by defence-related hormones, whereas all analysed genes were consistently down-regulated in M. graminicola-induced galls, at least until 7 days post-inoculation (dpi). Transgenic lines of Oryza sativa cv. Nipponbare overproducing OsTHI7 revealed decreased susceptibility to M. graminicola infection and P. graminicola colonization. Taken together, these results demonstrate the role of rice thionin genes in defence against two of the most damaging root pathogens attacking rice. © 2015 BSPP AND JOHN WILEY & SONS LTD.

[FANCA gene mutation analysis in Fanconi anemia patients].

PubMed

Chen, Fei; Peng, Guang-Jie; Zhang, Kejian; Hu, Qun; Zhang, Liu-Qing; Liu, Ai-Guo

2005-10-01

To screen the FANCA gene mutation and explore the FANCA protein function in Fanconi anemia (FA) patients. FANCA protein expression and its interaction with FANCF were analyzed using Western blot and immunoprecipitation in 3 cases of FA-A. Genomic DNA was used for MLPA analysis followed by sequencing. FANCA protein was undetectable and FANCA and FANCF protein interaction was impaired in these 3 cases of FA-A. Each case of FA-A contained biallelic pathogenic mutations in FANCA gene. No functional FANCA protein was found in these 3 cases of FA-A, and intragenic deletion, frame shift and splice site mutation were the major pathogenic mutations found in FANCA gene.

Oligogalacturonide-mediated induction of a gene involved in jasmonic acid synthesis in response to the cell-wall-degrading enzymes of the plant pathogen Erwinia carotovora.

PubMed

Norman, C; Vidal, S; Palva, E T

1999-07-01

Identification of Arabidopsis thaliana genes responsive to plant cell-wall-degrading enzymes of Erwinia carotovora subsp. carotovora led to the isolation of a cDNA clone with high sequence homology to the gene for allene oxide synthase, an enzyme involved in the biosynthesis of jasmonates. Expression of the corresponding gene was induced by the extracellular enzymes from this pathogen as well as by treatment with methyl jasmonate and short oligogalacturonides (OGAs). This suggests that OGAs are involved in the induction of the jasmonate pathway during plant defense response to E. carotovora subsp. carotovora attack.

Comparative transcriptome and gene co-expression network analysis reveal genes and signaling pathways adaptively responsive to varied adverse stresses in the insect fungal pathogen, Beauveria bassiana.

PubMed

He, Zhangjiang; Zhao, Xin; Lu, Zhuoyue; Wang, Huifang; Liu, Pengfei; Zeng, Fanqin; Zhang, Yongjun

2018-01-01

Sensing, responding, and adapting to the surrounding environment are crucial for all living organisms to survive, proliferate, and differentiate in their biological niches. Beauveria bassiana is an economically important insect-pathogenic fungus which is widely used as a biocontrol agent to control a variety of insect pests. The fungal pathogen unavoidably encounters a variety of adverse environmental stresses and defense response from the host insects during application of the fungal agents. However, few are known about the transcription response of the fungus to respond or adapt varied adverse stresses. Here, we comparatively analyzed the transcriptome of B. bassiana in globe genome under the varied stationary-phase stresses including osmotic agent (0.8 M NaCl), high temperature (32 °C), cell wall-perturbing agent (Congo red), and oxidative agents (H 2 O 2 or menadione). Total of 12,412 reads were obtained, and mapped to the 6767 genes of the B. bassiana. All of these stresses caused transcription responses involved in basal metabolism, cell wall construction, stress response or cell rescue/detoxification, signaling transduction and gene transcription regulation, and likely other cellular processes. An array of genes displayed similar transcription patterns in response to at least two of the five stresses, suggesting a shared transcription response to varied adverse stresses. Gene co-expression network analysis revealed that mTOR signaling pathway, but not HOG1 MAP kinase pathway, played a central role in regulation the varied adverse stress responses, which was verified by RNAi-mediated knockdown of TOR1. Our findings provided an insight of transcription response and gene co-expression network of B. bassiana in adaptation to varied environments. Copyright © 2017 Elsevier Inc. All rights reserved.

Two novel mutations in the PPIB gene cause a rare pedigree of osteogenesis imperfecta type IX.

PubMed

Jiang, Yu; Pan, Jingxin; Guo, Dongwei; Zhang, Wei; Xie, Jie; Fang, Zishui; Guo, Chunmiao; Fang, Qun; Jiang, Weiying; Guo, Yibin

2017-06-01

Osteogenesis imperfecta (OI) is a rare genetic skeletal disorder characterized by increased bone fragility and vulnerability to fractures. PPIB is identified as a candidate gene for OI-IX, here we detect two pathogenic mutations in PPIB and analyze the genotype-phenotype correlation in a Chinese family with OI. Next-generation sequencing (NGS) was used to screen the whole exome of the parents of proband. Screening of variation frequency, evolutionary conservation comparisons, pathogenicity evaluation, and protein structure prediction were conducted to assess the pathogenicity of the novel mutations. Sanger sequencing was used to confirm the candidate variants. RTQ-PCR was used to analyze the PPIB gene expression. All mutant genes screened out by NGS were excluded except PPIB. Two novel heterozygous PPIB mutations (father, c.25A>G; mother, c.509G>A) were identified in relation to osteogenesis imperfecta type IX. Both mutations were predicted to be pathogenic by bioinformatics analysis and RTQ-PCR analysis revealed downregulated PPIB expression in the two carriers. We report a rare pedigree with an autosomal recessive osteogenesis imperfecta type IX (OI-IX) caused by two novel PPIB mutations identified for the first time in China. The current study expands our knowledge of PPIB mutations and their associated phenotypes, and provides new information on the genetic defects associated with this disease for clinical diagnosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of the significance of cell wall polymers in flax infected with a pathogenic strain of Fusarium oxysporum.

PubMed

Wojtasik, Wioleta; Kulma, Anna; Dymińska, Lucyna; Hanuza, Jerzy; Czemplik, Magdalena; Szopa, Jan

2016-03-22

Fusarium oxysporum infection leads to Fusarium-derived wilt, which is responsible for the greatest losses in flax (Linum usitatissimum) crop yield. Plants infected by Fusarium oxysporum show severe symptoms of dehydration due to the growth of the fungus in vascular tissues. As the disease develops, vascular browning and leaf yellowing can be observed. In the case of more virulent strains, plants die. The pathogen's attack starts with secretion of enzymes degrading the host cell wall. The main aim of the study was to evaluate the role of the cell wall polymers in the flax plant response to the infection in order to better understand the process of resistance and develop new ways to protect plants against infection. For this purpose, the expression of genes involved in cell wall polymer metabolism and corresponding polymer levels were investigated in flax seedlings after incubation with Fusarium oxysporum. This analysis was facilitated by selecting two groups of genes responding differently to the infection. The first group comprised genes strongly affected by the infection and activated later (phenylalanine ammonia lyase and glucosyltransferase). The second group comprised genes which are slightly affected (up to five times) and their expression vary as the infection progresses. Fusarium oxysporum infection did not affect the contents of cell wall polymers, but changed their structure. The results suggest that the role of the cell wall polymers in the plant response to Fusarium oxysporum infection is manifested through changes in expression of their genes and rearrangement of the cell wall polymers. Our studies provided new information about the role of cellulose and hemicelluloses in the infection process, the change of their structure and the expression of genes participating in their metabolism during the pathogen infection. We also confirmed the role of pectin and lignin in this process, indicating the major changes at the mRNA level of lignin metabolism genes and the loosening of the pectin structure.

Expression of glycine-rich protein genes, AtGRP5 and AtGRP23, induced by the cutin monomer 16-hydroxypalmitic acid in Arabidopsis thaliana.

PubMed

Park, Jong Ho; Suh, Mi Chung; Kim, Tae Hyun; Kim, Moon Chul; Cho, Sung Ho

2008-11-01

Glycine-rich proteins (GRPs) belong to a large family of heterogenous proteins that are enriched in glycine residues. The expression of two GRP genes of Arabidopsis thaliana, AtGRP5 and AtGRP23, was induced by 16-hydroxypalmitic acid (HPA), a major component of cutin. The expression of AtGRP3, which encodes a GRP protein that is structurally different from AtGRP5 and AtGRP23, was not responsive to HPA application. Treatment with HPA also induced expression of the pathogen-related PR-1 and PR-4 genes. Abscisic acid and salicylic acid treatments enhanced the transcript levels of AtGRP5 and AtGRP23 as well as those of AtGRP3. It was also demonstrated that HPA effectively elicited the accumulation of H2O2 in rosette leaves of Arabidopsis. Results suggest the possible role of some species of GRPs, such as AtGRP5 and AtGRP23, in response to the pathogenic invasion mediated by cutin monomers in plants.

Adaptation Mechanisms in the Evolution of Moss Defenses to Microbes

PubMed Central

Ponce de León, Inés; Montesano, Marcos

2017-01-01

Bryophytes, including mosses, liverworts and hornworts are early land plants that have evolved key adaptation mechanisms to cope with abiotic stresses and microorganisms. Microbial symbioses facilitated plant colonization of land by enhancing nutrient uptake leading to improved plant growth and fitness. In addition, early land plants acquired novel defense mechanisms to protect plant tissues from pre-existing microbial pathogens. Due to its evolutionary stage linking unicellular green algae to vascular plants, the non-vascular moss Physcomitrella patens is an interesting organism to explore the adaptation mechanisms developed in the evolution of plant defenses to microbes. Cellular and biochemical approaches, gene expression profiles, and functional analysis of genes by targeted gene disruption have revealed that several defense mechanisms against microbial pathogens are conserved between mosses and flowering plants. P. patens perceives pathogen associated molecular patterns by plasma membrane receptor(s) and transduces the signal through a MAP kinase (MAPK) cascade leading to the activation of cell wall associated defenses and expression of genes that encode proteins with different roles in plant resistance. After pathogen assault, P. patens also activates the production of ROS, induces a HR-like reaction and increases levels of some hormones. Furthermore, alternative metabolic pathways are present in P. patens leading to the production of a distinct metabolic scenario than flowering plants that could contribute to defense. P. patens has acquired genes by horizontal transfer from prokaryotes and fungi, and some of them could represent adaptive benefits for resistance to biotic stress. In this review, the current knowledge related to the evolution of plant defense responses against pathogens will be discussed, focusing on the latest advances made in the model plant P. patens. PMID:28360923

Expression of the β-1,3-glucanase gene bgn13.1 from Trichoderma harzianum in strawberry increases tolerance to crown rot diseases but interferes with plant growth.

PubMed

Mercado, José A; Barceló, Marta; Pliego, Clara; Rey, Manuel; Caballero, José L; Muñoz-Blanco, Juan; Ruano-Rosa, David; López-Herrera, Carlos; de Los Santos, Berta; Romero-Muñoz, Fernando; Pliego-Alfaro, Fernando

2015-12-01

The expression of antifungal genes from Trichoderma harzianum, mainly chitinases, has been used to confer plant resistance to fungal diseases. However, the biotechnological potential of glucanase genes from Trichoderma has been scarcely assessed. In this research, transgenic strawberry plants expressing the β-1,3-glucanase gene bgn13.1 from T. harzianum, under the control of the CaMV35S promoter, have been generated. After acclimatization, five out of 12 independent lines analysed showed a stunted phenotype when growing in the greenhouse. Moreover, most of the lines displayed a reduced yield due to both a reduction in the number of fruit per plant and a lower fruit size. Several transgenic lines showing higher glucanase activity in leaves than control plants were selected for pathogenicity tests. When inoculated with Colletotrichum acutatum, one of the most important strawberry pathogens, transgenic lines showed lower anthracnose symptoms in leaf and crown than control. In the three lines selected, the percentage of plants showing anthracnose symptoms in crown decreased from 61 % to a mean value of 16.5 %, in control and transgenic lines, respectively. Some transgenic lines also showed an enhanced resistance to Rosellinia necatrix, a soil-borne pathogen causing root and crown rot in strawberry. These results indicate that bgn13.1 from T. harzianum can be used to increase strawberry tolerance to crown rot diseases, although its constitutive expression affects plant growth and fruit yield. Alternative strategies such as the use of tissue specific promoters might avoid the negative effects of bgn13.1 expression in plant performance.

Lactobacillus rhamnosus GG and its SpaC pilus adhesin modulate inflammatory responsiveness and TLR-related gene expression in the fetal human gut

PubMed Central

Ganguli, Kriston; Collado, Maria Carmen; Rautava, Jaana; Lu, Lei; Satokari, Reetta; von Ossowski, Ingemar; Reunanen, Justus; de Vos, Willem M.; Palva, Airi; Isolauri, Erika; Salminen, Seppo; Walker, W. Allan; Rautava, Samuli

2015-01-01

Background Bacterial contact in utero modulates fetal and neonatal immune responses. Maternal probiotic supplementation reduces the risk of immune-mediated disease in the infant. We investigated the immunomodulatory properties of live Lactobacillus rhamnosus GG and its SpaC pilus adhesin in human fetal intestinal models. Methods TNF-α mRNA expression was measured by qPCR in a human fetal intestinal organ culture model exposed to live L. rhamnosus GG and proinflammatory stimuli. Binding of recombinant SpaC pilus protein to intestinal epithelial cells was assessed in human fetal intestinal organ culture and the human fetal intestinal epithelial cell line H4 by immunohistochemistry and immunofluorescence, respectively. TLR-related gene expression in fetal ileal organ culture after exposure to recombinant SpaC was assessed by qPCR. Results Live L. rhamnosus GG significantly attenuates pathogen-induced TNF-α mRNA expression in the human fetal gut. Recombinant SpaC protein was found to adhere to the fetal gut and to modulate varying levels of TLR-related gene expression. Conclusion The human fetal gut is responsive to luminal microbes. L. rhamnosus GG significantly attenuates fetal intestinal inflammatory responses to pathogenic bacteria. The L. rhamnosus GG pilus adhesin SpaC binds to immature human intestinal epithelial cells and directly modulates intestinal epithelial cell innate immune gene expression. PMID:25580735

Cutin monomer induces expression of the rice OsLTP5 lipid transfer protein gene.

PubMed

Kim, Tae Hyun; Park, Jong Ho; Kim, Moon Chul; Cho, Sung Ho

2008-01-01

Treatment with the cutin monomer 16-hydroxypalmitic acid (HPA), a major component of cutin, elicited the synthesis of hydrogen peroxide (H2O2) in rice leaves and induced the expression of the lipid transfer protein gene OsLTP5. Treatment with HPA also induced expression of OsLTP1, OsLTP2, and the pathogen-related PR-10 genes to a lesser extent. The OsLTP5 transcript was expressed prominently in stems and flowers, but was barely detectable in leaves. Expression of OsLTP5 was induced in shoots in response to ABA and salicylic acid. It is proposed that HPA is perceived by rice as a signal, inducing defense reactions.

Searching for resistance genes to Bursaphelenchus xylophilus using high throughput screening

PubMed Central

2012-01-01

Background Pine wilt disease (PWD), caused by the pinewood nematode (PWN; Bursaphelenchus xylophilus), damages and kills pine trees and is causing serious economic damage worldwide. Although the ecological mechanism of infestation is well described, the plant’s molecular response to the pathogen is not well known. This is due mainly to the lack of genomic information and the complexity of the disease. High throughput sequencing is now an efficient approach for detecting the expression of genes in non-model organisms, thus providing valuable information in spite of the lack of the genome sequence. In an attempt to unravel genes potentially involved in the pine defense against the pathogen, we hereby report the high throughput comparative sequence analysis of infested and non-infested stems of Pinus pinaster (very susceptible to PWN) and Pinus pinea (less susceptible to PWN). Results Four cDNA libraries from infested and non-infested stems of P. pinaster and P. pinea were sequenced in a full 454 GS FLX run, producing a total of 2,083,698 reads. The putative amino acid sequences encoded by the assembled transcripts were annotated according to Gene Ontology, to assign Pinus contigs into Biological Processes, Cellular Components and Molecular Functions categories. Most of the annotated transcripts corresponded to Picea genes-25.4-39.7%, whereas a smaller percentage, matched Pinus genes, 1.8-12.8%, probably a consequence of more public genomic information available for Picea than for Pinus. The comparative transcriptome analysis showed that when P. pinaster was infested with PWN, the genes malate dehydrogenase, ABA, water deficit stress related genes and PAR1 were highly expressed, while in PWN-infested P. pinea, the highly expressed genes were ricin B-related lectin, and genes belonging to the SNARE and high mobility group families. Quantitative PCR experiments confirmed the differential gene expression between the two pine species. Conclusions Defense-related genes triggered by nematode infestation were detected in both P. pinaster and P. pinea transcriptomes utilizing 454 pyrosequencing technology. P. pinaster showed higher abundance of genes related to transcriptional regulation, terpenoid secondary metabolism (including some with nematicidal activity) and pathogen attack. P. pinea showed higher abundance of genes related to oxidative stress and higher levels of expression in general of stress responsive genes. This study provides essential information about the molecular defense mechanisms utilized by P. pinaster and P. pinea against PWN infestation and contributes to a better understanding of PWD. PMID:23134679

Trichostatin A effects on gene expression in the protozoan parasite Entamoeba histolytica

PubMed Central

Ehrenkaufer, Gretchen M; Eichinger, Daniel J; Singh, Upinder

2007-01-01

Background Histone modification regulates chromatin structure and influences gene expression associated with diverse biological functions including cellular differentiation, cancer, maintenance of genome architecture, and pathogen virulence. In Entamoeba, a deep-branching eukaryote, short chain fatty acids (SCFA) affect histone acetylation and parasite development. Additionally, a number of active histone modifying enzymes have been identified in the parasite genome. However, the overall extent of gene regulation tied to histone acetylation is not known. Results In order to identify the genome-wide effects of histone acetylation in regulating E. histolytica gene expression, we used whole-genome expression profiling of parasites treated with SCFA and Trichostatin A (TSA). Despite significant changes in histone acetylation patterns, exposure of parasites to SCFA resulted in minimal transcriptional changes (11 out of 9,435 genes transcriptionally regulated). In contrast, exposure to TSA, a more specific inhibitor of histone deacetylases, significantly affected transcription of 163 genes (122 genes upregulated and 41 genes downregulated). Genes modulated by TSA were not regulated by treatment with 5-Azacytidine, an inhibitor of DNA-methyltransferase, indicating that in E. histolytica the crosstalk between DNA methylation and histone modification is not substantial. However, the set of genes regulated by TSA overlapped substantially with genes regulated during parasite development: 73/122 genes upregulated by TSA exposure were upregulated in E. histolytica cysts (p-value = 6 × 10-53) and 15/41 genes downregulated by TSA exposure were downregulated in E. histolytica cysts (p-value = 3 × 10-7). Conclusion This work represents the first genome-wide analysis of histone acetylation and its effects on gene expression in E. histolytica. The data indicate that SCFAs, despite their ability to influence histone acetylation, have minimal effects on gene transcription in cultured parasites. In contrast, the effect of TSA on E. histolytica gene expression is more substantial and includes genes involved in the encystation pathway. These observations will allow further dissection of the effects of histone acetylation and the genetic pathways regulating stage conversion in this pathogenic parasite. PMID:17612405

Adaptation to the Host Environment by Plant-Pathogenic Fungi.

PubMed

van der Does, H Charlotte; Rep, Martijn

2017-08-04

Many fungi can live both saprophytically and as endophyte or pathogen inside a living plant. In both environments, complex organic polymers are used as sources of nutrients. Propagation inside a living host also requires the ability to respond to immune responses of the host. We review current knowledge of how plant-pathogenic fungi do this. First, we look at how fungi change their global gene expression upon recognition of the host environment, leading to secretion of effectors, enzymes, and secondary metabolites; changes in metabolism; and defense against toxic compounds. Second, we look at what is known about the various cues that enable fungi to sense the presence of living plant cells. Finally, we review literature on transcription factors that participate in gene expression in planta or are suspected to be involved in that process because they are required for the ability to cause disease.

Increased filamentous growth of Candida albicans in simulated microgravity.

PubMed

Altenburg, Sara D; Nielsen-Preiss, Sheila M; Hyman, Linda E

2008-03-01

Knowledge of simulated microgravity (SMG)-induced changes in the pathogenicity of microorganisms is important for success of long-term spaceflight. In a previous study using the high aspect ratio vessel bioreactor, we showed that the yeast species Saccharomyces cerevisiae underwent a significant phenotypic response when grown in modeled microgravity, which was reflected in the analysis of gene expression profiles. In this study, we establish that Candida albicans responds to SMG in a similar fashion, demonstrating that there is a conserved response among yeast to this environmental stress. We also report that the growth of C. albicans in SMG results in a morphogenic switch that is consistent with enhanced pathogenicity. Specifically, we observed an increase in filamentous forms of the organism and accompanying changes in the expression of two genes associated with the yeast-hyphal transition. The morphological response may have significant implications for astronauts' safety, as the fungal pathogen may become more virulent during spaceflight.

Activation and manipulation of host responses by a Gram-positive bacterium

PubMed Central

Balaji, Vasudevan

2008-01-01

The interaction between tomato plants and Clavibacter michiganensis subsp. michiganensis (Cmm) represents a model pathosystem to study the interplay between the virulence determinants of a Gram-positive bacterium and the attempt of a crop plant to counteract pathogen invasion. To investigate plant responses activated during this compatible interaction, we recently analyzed gene expression profiles of tomato stems infected with Cmm. This analysis revealed activation of basal defense responses that are typically observed upon plant perception of pathogen-associated molecular patterns. In addition, Cmm infection upregulated the expression of host genes related to ethylene synthesis and response. Further analysis of tomato plants impaired in ethylene perception and production demonstrated an important role for ethylene in the development of disease symptoms. Here we discuss possible molecular strategies used by the plant to recognize Cmm infection and possible mechanisms employed by the pathogen to interfere with the activation of plant defense responses and promote disease. PMID:19704516

Foliar aphid feeding recruits rhizosphere bacteria and primes plant immunity against pathogenic and non-pathogenic bacteria in pepper.

PubMed

Lee, Boyoung; Lee, Soohyun; Ryu, Choong-Min

2012-07-01

Plants modulate defence signalling networks in response to different biotic stresses. The present study evaluated the effect of a phloem-sucking aphid on plant defence mechanisms in pepper (Capsicum annuum) during subsequent pathogen attacks on leaves and rhizosphere bacteria on roots. Plants were pretreated with aphids and/or the chemical trigger benzothiadiazol (BTH) 7 d before being challenged with two pathogenic bacteria, Xanthomonas axonopodis pv. vesicatoria (Xav) as a compatible pathogen and X. axonopodis pv. glycines (Xag) as an incompatible (non-host) pathogen. Disease severity was noticeably lower in aphid- and BTH + aphid-treated plants than in controls. Although treatment with BTH or aphids alone did not affect the hypersensitive response (HR) against Xag strain 8ra, the combination treatment had a synergistic effect on the HR. The aphid population was reduced by BTH pretreatment and by combination treatment with BTH and bacterial pathogens in a synergistic manner. Analysis of the expression of the defence-related genes Capsicum annum pathogenesis-related gene 9 (CaPR9), chitinase 2 (CaCHI2), SAR8·2 and Lipoxygenase1 (CaLOX1) revealed that aphid infestation resulted in the priming of the systemic defence responses against compatible and incompatible pathogens. Conversely, pre-challenge with the compatible pathogen Xav on pepper leaves significantly reduced aphid numbers. Aphid infestation increased the population of the beneficial Bacillus subtilis GB03 but reduced that of the pathogenic Ralstonia solanacearum SL1931. The expression of defence-related genes in the root and leaf after aphid feeding indicated that the above-ground aphid infestation elicited salicylic acid and jasmonic acid signalling throughout the whole plant. The findings of this study show that aphid feeding elicits plant resistance responses and attracts beneficial bacterial populations to help the plant cope with subsequent pathogen attacks.

Foliar aphid feeding recruits rhizosphere bacteria and primes plant immunity against pathogenic and non-pathogenic bacteria in pepper

PubMed Central

Lee, Boyoung; Lee, Soohyun; Ryu, Choong-Min

2012-01-01

Background and Aims Plants modulate defence signalling networks in response to different biotic stresses. The present study evaluated the effect of a phloem-sucking aphid on plant defence mechanisms in pepper (Capsicum annuum) during subsequent pathogen attacks on leaves and rhizosphere bacteria on roots. Methods Plants were pretreated with aphids and/or the chemical trigger benzothiadiazol (BTH) 7 d before being challenged with two pathogenic bacteria, Xanthomonas axonopodis pv. vesicatoria (Xav) as a compatible pathogen and X. axonopodis pv. glycines (Xag) as an incompatible (non-host) pathogen. Key Results Disease severity was noticeably lower in aphid- and BTH + aphid-treated plants than in controls. Although treatment with BTH or aphids alone did not affect the hypersensitive response (HR) against Xag strain 8ra, the combination treatment had a synergistic effect on the HR. The aphid population was reduced by BTH pretreatment and by combination treatment with BTH and bacterial pathogens in a synergistic manner. Analysis of the expression of the defence-related genes Capsicum annum pathogenesis-related gene 9 (CaPR9), chitinase 2 (CaCHI2), SAR8·2 and Lipoxygenase1 (CaLOX1) revealed that aphid infestation resulted in the priming of the systemic defence responses against compatible and incompatible pathogens. Conversely, pre-challenge with the compatible pathogen Xav on pepper leaves significantly reduced aphid numbers. Aphid infestation increased the population of the beneficial Bacillus subtilis GB03 but reduced that of the pathogenic Ralstonia solanacearum SL1931. The expression of defence-related genes in the root and leaf after aphid feeding indicated that the above-ground aphid infestation elicited salicylic acid and jasmonic acid signalling throughout the whole plant. Conclusions The findings of this study show that aphid feeding elicits plant resistance responses and attracts beneficial bacterial populations to help the plant cope with subsequent pathogen attacks. PMID:22437662

Potato stem cuttings to study Verticillium dahliae infection for resistance breeding and ‘omics’ studies

USDA-ARS?s Scientific Manuscript database

The use of consistent and effective methods for early discrimination of resistance to pathogens and selection of appropriate times for tissue sampling are important for experiments focused on global gene expression and metabolomics. Assays for resistance to the vascular pathogen Verticillium dahliae...

High-throughput SuperSAGE for gene expression analysis of Nicotiana tabacum - Rhizoctonia solani interaction

USDA-ARS?s Scientific Manuscript database

Plants are under continuous threat of infection by pathogens endowed with diverse strategies to colonize their host. Knowledge of plant susceptibility factors and the molecular processes involved in the infection process are critical for understanding plant-pathogen interactions. We used SuperSAGE t...

Identifying differentially expressed genes in leaves of Glycine tomentella in the presence of the fungal pathogen Phakopsora pachyrhizi

USDA-ARS?s Scientific Manuscript database

Transcription profiles of Glycine tomentella genotypes having different responses to soybean rust, caused by the fungal pathogen Phakopsora pachyrhizi, were compared using suppression subtractive hybridization (SSH). Four cDNA libraries were constructed from infected and non-infected leaves of resis...

Analysis and functional annotation of expressed sequence tags from the fall armyworm Spodoptera frugiperda

PubMed Central

Deng, Youping; Dong, Yinghua; Thodima, Venkata; Clem, Rollie J; Passarelli, A Lorena

2006-01-01

Background Little is known about the genome sequences of lepidopteran insects, although this group of insects has been studied extensively in the fields of endocrinology, development, immunity, and pathogen-host interactions. In addition, cell lines derived from Spodoptera frugiperda and other lepidopteran insects are routinely used for baculovirus foreign gene expression. This study reports the results of an expressed sequence tag (EST) sequencing project in cells from the lepidopteran insect S. frugiperda, the fall armyworm. Results We have constructed an EST database using two cDNA libraries from the S. frugiperda-derived cell line, SF-21. The database consists of 2,367 ESTs which were assembled into 244 contigs and 951 singlets for a total of 1,195 unique sequences. Conclusion S. frugiperda is an agriculturally important pest insect and genomic information will be instrumental for establishing initial transcriptional profiling and gene function studies, and for obtaining information about genes manipulated during infections by insect pathogens such as baculoviruses. PMID:17052344

Expression of genes associated with stress conditions by Listeria monocytogenes in interaction with nisin producer Lactococcus lactis.

PubMed

Miranda, Rodrigo Otávio; Campos-Galvão, Maria Emilene Martino; Nero, Luís Augusto

2018-03-01

The use of nisin producers in foods is considered a mitigation strategy to control foodborne pathogens growth, such as Listeria monocytogenes, due to the production of this bacteriocin in situ. However, when the bacteriocin does not reach an adequate concentration, the target bacteria can develop a cross-response to different stress conditions in food, such as acid, thermal and osmotic. This study aimed to evaluate the interaction of a nisin-producing strain of Lactococcus lactis DY-13 and L. monocytogenes in BHI and skim milk, and its influence on general (sigB), acid (gadD2), thermal (groEL) and osmotic (gbu) stress-related genes of the pathogen. L. monocytogenes populations decreased approximately 2log in BHI and 1log in milk after 24h in co-culture with the nisin producer L. lactis, coherent with the increasing expression of nisK. Expression of stress-related genes by L. monocytogenes presented lower oscillation in BHI than in milk, indicating its better ability to survive in milk, despite the higher nisin production. Stress-related genes presented a varied expression by L. monocytogenes in the tested conditions: sigB expression remained stable or reduced over time; gadD2 presented high expression in milk; groEL presented low expression in BHI when compared to milk, trending to decrease overtime; gbu expression in milk after 24h was lower than in BHI. The presented study demonstrated the growth of a nisin producer L. lactis can affect the expression of stress-related genes by L. monocytogenes, and understating these mechanisms is crucial to enhance the conservation methods employed in foods. Copyright © 2017 Elsevier Ltd. All rights reserved.

microRNAs and Their Targets in Apple (Malus domestica cv. "Fuji") Involved in Response to Infection of Pathogen Valsa mali.

PubMed

Feng, Hao; Xu, Ming; Zheng, Xiang; Zhu, Tongyi; Gao, Xiaoning; Huang, Lili

2017-01-01

miRNAs are important regulators involving in plant-pathogen interactions. However, their roles in apple tree response to Valsa canker pathogen ( Valsa mali, Vm ) infection were poorly understood. In this study, we constructed two miRNA libraries using the twig bark tissues of apple tree ( Malus domestica Borkh. cv. "Fuji") inoculated with Vm (IVm) and PDA medium (control, BMd). Among all detected miRNAs, 23 miRNAs were specifically isolated from BMd and 39 miRNAs were specifically isolated from IVm. Meanwhile, the expression of 294 miRNAs decreased; and another 172 miRNAs showed an increased expression trend in IVm compared with that in BMd. Furthermore, two degradome sequencing libraries were also constructed to identify the target genes of these miRNAs. In total, 353 differentially expressed miRNAs between IVm and BMd were detected to be able to target 1,077 unigenes with 2,251 cleavage sites. Based on GO and KEGG analysis, these genes were found to be mainly related to transcription regulation and signal transduction. In addition, we selected 17 miRNAs and 22 corresponding target genes to screen the expression profiles when apple twigs were infected by Vm . The expression trends of most miRNAs/target genes were consist with the results of deep sequencing. Many of them may involve in the apple twig- Vm interaction by inducing/reducing their expression. What's more, miRNAs and their target genes regulate the apple twig- Vm interaction by forming many complicated regulation networks rather than one to one model. It is worth that a conserved miRNAs mdm-miR482b, which was down regulated in IVm compared with BMd, has 14 potential target genes, most of which are disease resistance related genes. This indicates that mdm-miR482b may play important roles in apple twig response to Vm . More important, the feedback regulation of sRNA pathway in apple twig is also very complex, and play critical role in the interaction between apple twig and Vm based on the results of expression analysis. In all, the results will provide insights into the crucial functions of miRNAs in the woody plant, apple tree- Vm interaction.

microRNAs and Their Targets in Apple (Malus domestica cv. “Fuji”) Involved in Response to Infection of Pathogen Valsa mali

PubMed Central

Feng, Hao; Xu, Ming; Zheng, Xiang; Zhu, Tongyi; Gao, Xiaoning; Huang, Lili

2017-01-01

miRNAs are important regulators involving in plant-pathogen interactions. However, their roles in apple tree response to Valsa canker pathogen (Valsa mali, Vm) infection were poorly understood. In this study, we constructed two miRNA libraries using the twig bark tissues of apple tree (Malus domestica Borkh. cv. “Fuji”) inoculated with Vm (IVm) and PDA medium (control, BMd). Among all detected miRNAs, 23 miRNAs were specifically isolated from BMd and 39 miRNAs were specifically isolated from IVm. Meanwhile, the expression of 294 miRNAs decreased; and another 172 miRNAs showed an increased expression trend in IVm compared with that in BMd. Furthermore, two degradome sequencing libraries were also constructed to identify the target genes of these miRNAs. In total, 353 differentially expressed miRNAs between IVm and BMd were detected to be able to target 1,077 unigenes with 2,251 cleavage sites. Based on GO and KEGG analysis, these genes were found to be mainly related to transcription regulation and signal transduction. In addition, we selected 17 miRNAs and 22 corresponding target genes to screen the expression profiles when apple twigs were infected by Vm. The expression trends of most miRNAs/target genes were consist with the results of deep sequencing. Many of them may involve in the apple twig-Vm interaction by inducing/reducing their expression. What's more, miRNAs and their target genes regulate the apple twig-Vm interaction by forming many complicated regulation networks rather than one to one model. It is worth that a conserved miRNAs mdm-miR482b, which was down regulated in IVm compared with BMd, has 14 potential target genes, most of which are disease resistance related genes. This indicates that mdm-miR482b may play important roles in apple twig response to Vm. More important, the feedback regulation of sRNA pathway in apple twig is also very complex, and play critical role in the interaction between apple twig and Vm based on the results of expression analysis. In all, the results will provide insights into the crucial functions of miRNAs in the woody plant, apple tree-Vm interaction. PMID:29270184

Diversifying selection in the wheat stem rust fungus acts predominantly on pathogen-associated gene families and reveals candidate effectors

PubMed Central

Sperschneider, Jana; Ying, Hua; Dodds, Peter N.; Gardiner, Donald M.; Upadhyaya, Narayana M.; Singh, Karam B.; Manners, John M.; Taylor, Jennifer M.

2014-01-01

Plant pathogens cause severe losses to crop plants and threaten global food production. One striking example is the wheat stem rust fungus, Puccinia graminis f. sp. tritici, which can rapidly evolve new virulent pathotypes in response to resistant host lines. Like several other filamentous fungal and oomycete plant pathogens, its genome features expanded gene families that have been implicated in host-pathogen interactions, possibly encoding effector proteins that interact directly with target host defense proteins. Previous efforts to understand virulence largely relied on the prediction of secreted, small and cysteine-rich proteins as candidate effectors and thus delivered an overwhelming number of candidates. Here, we implement an alternative analysis strategy that uses the signal of adaptive evolution as a line of evidence for effector function, combined with comparative information and expression data. We demonstrate that in planta up-regulated genes that are rapidly evolving are found almost exclusively in pathogen-associated gene families, affirming the impact of host-pathogen co-evolution on genome structure and the adaptive diversification of specialized gene families. In particular, we predict 42 effector candidates that are conserved only across pathogens, induced during infection and rapidly evolving. One of our top candidates has recently been shown to induce genotype-specific hypersensitive cell death in wheat. This shows that comparative genomics incorporating the evolutionary signal of adaptation is powerful for predicting effector candidates for laboratory verification. Our system can be applied to a wide range of pathogens and will give insight into host-pathogen dynamics, ultimately leading to progress in strategies for disease control. PMID:25225496

Signature-tagged mutagenesis in a chicken infection model leads to the identification of a novel avian pathogenic Escherichia coli fimbrial adhesin.

PubMed

Antão, Esther-Maria; Ewers, Christa; Gürlebeck, Doreen; Preisinger, Rudolf; Homeier, Timo; Li, Ganwu; Wieler, Lothar H

2009-11-12

The extraintestinal pathogen, avian pathogenic E. coli (APEC), known to cause systemic infections in chickens, is responsible for large economic losses in the poultry industry worldwide. In order to identify genes involved in the early essential stages of pathogenesis, namely adhesion and colonization, Signature-tagged mutagenesis (STM) was applied to a previously established lung colonization model of infection by generating and screening a total of 1,800 mutants of an APEC strain IMT5155 (O2:K1:H5; Sequence type complex 95). The study led to the identification of new genes of interest, including two adhesins, one of which coded for a novel APEC fimbrial adhesin (Yqi) not described for its role in APEC pathogenesis to date. Its gene product has been temporarily designated ExPEC Adhesin I (EA/I) until the adhesin-specific receptor is identified. Deletion of the ExPEC adhesin I gene resulted in reduced colonization ability by APEC strain IMT5155 both in vitro and in vivo. Furthermore, complementation of the adhesin gene restored its ability to colonize epithelial cells in vitro. The ExPEC adhesin I protein was successfully expressed in vitro. Electron microscopy of an afimbriate strain E. coli AAEC189 over-expressed with the putative EA/I gene cluster revealed short fimbrial-like appendages protruding out of the bacterial outer membrane. We observed that this adhesin coding gene yqi is prevalent among extraintestinal pathogenic E. coli (ExPEC) isolates, including APEC (54.4%), uropathogenic E. coli (UPEC) (65.9%) and newborn meningitic E. coli (NMEC) (60.0%), and absent in all of the 153 intestinal pathogenic E. coli strains tested, thereby validating the designation of the adhesin as ExPEC Adhesin I. In addition, prevalence of EA/I was most frequently associated with the B2 group of the EcoR classification and ST95 complex of the multi locus sequence typing (MLST) scheme, with evidence of a positive selection within this highly pathogenic complex. This is the first report of the newly identified and functionally characterized ExPEC adhesin I and its significant role during APEC infection in chickens.

MicroRNA396a-5p and -3p induce tomato disease susceptibility by suppressing target genes and upregulating salicylic acid.

PubMed

Chen, Lei; Meng, Jun; Zhai, Junmiao; Xu, Pinsan; Luan, Yushi

2017-12-01

Plants have evolved a variety of mechanisms to perceive and resist the assault of pathogens. The biotrophs, necrotrophs and hemibiotrophs are types of plant pathogens that activate diverse salicylic acid (SA) and jasmonic acid (JA) signaling pathways. In this study we showed that the expressions of miR396a-5p and -3p in Solanum lycopersicum (S. lycopersicum) were both down-regulated after infection by hemibiotroph Phytophthora infestans (P. infestans) and necrotroph Botrytis cinerea (B. cinerea) infection. Overexpression of miR396a-5p and -3p in transgenic tomato enhanced the susceptibility of S. lycopersicum to P. infestans and B. cinerea infection and the tendency to produce reactive oxygen species (ROS) under pathogen-related biotic stress. Additionally, miR396a regulated growth-regulating factor1 (GRF1), salicylic acid carboxyl methyltransferase (SAMT), glycosyl hydrolases (GH) and nucleotide-binding site-leucine-rich repeat (NBS-LRR) and down-regulated their levels. This ultimately led to inhibition of the expression of pathogenesis-related 1 (PR1), TGA transcription factors1 and 2 (TGA1 and TGA2) and JA-dependent proteinase inhibitors I and II (PI I and II), but enhanced the endogenous SA content and nonexpressor of pathogenesis-related genes 1 (NPR1) expression. Taken together, our results showed that negative regulation of target genes and their downstream genes expressions by miR396a-5p and -3p are critical for tomato abiotic stresses via affecting SA or JA signaling pathways. Copyright © 2017 Elsevier B.V. All rights reserved.

Control of Gene Expression in Leptospira spp. by Transcription Activator-Like Effectors Demonstrates a Potential Role for LigA and LigB in Leptospira interrogans Virulence

PubMed Central

Pappas, Christopher J.

2015-01-01

Leptospirosis is a zoonotic disease that affects ∼1 million people annually, with a mortality rate of >10%. Currently, there is an absence of effective genetic manipulation tools for targeted mutagenesis in pathogenic leptospires. Transcription activator-like effectors (TALEs) are a recently described group of repressors that modify transcriptional activity in prokaryotic and eukaryotic cells by directly binding to a targeted sequence within the host genome. To determine the applicability of TALEs within Leptospira spp., two TALE constructs were designed. First, a constitutively expressed TALE gene specific for the lacO-like region upstream of bgaL was trans inserted in the saprophyte Leptospira biflexa (the TALEβgal strain). Reverse transcriptase PCR (RT-PCR) analysis and enzymatic assays demonstrated that BgaL was not expressed in the TALEβgal strain. Second, to study the role of LigA and LigB in pathogenesis, a constitutively expressed TALE gene with specificity for the homologous promoter regions of ligA and ligB was cis inserted into the pathogen Leptospira interrogans (TALElig). LigA and LigB expression was studied by using three independent clones: TALElig1, TALElig2, and TALElig3. Immunoblot analysis of osmotically induced TALElig clones demonstrated 2- to 9-fold reductions in the expression levels of LigA and LigB, with the highest reductions being noted for TALElig1 and TALElig2, which were avirulent in vivo and nonrecoverable from animal tissues. This study reconfirms galactosidase activity in the saprophyte and suggests a role for LigA and LigB in pathogenesis. Collectively, this study demonstrates that TALEs are effective at reducing the expression of targeted genes within saprophytic and pathogenic strains of Leptospira spp., providing an additional genetic manipulation tool for this genus. PMID:26341206

Control of Gene Expression in Leptospira spp. by Transcription Activator-Like Effectors Demonstrates a Potential Role for LigA and LigB in Leptospira interrogans Virulence.

PubMed

Pappas, Christopher J; Picardeau, Mathieu

2015-11-01

Leptospirosis is a zoonotic disease that affects ∼1 million people annually, with a mortality rate of >10%. Currently, there is an absence of effective genetic manipulation tools for targeted mutagenesis in pathogenic leptospires. Transcription activator-like effectors (TALEs) are a recently described group of repressors that modify transcriptional activity in prokaryotic and eukaryotic cells by directly binding to a targeted sequence within the host genome. To determine the applicability of TALEs within Leptospira spp., two TALE constructs were designed. First, a constitutively expressed TALE gene specific for the lacO-like region upstream of bgaL was trans inserted in the saprophyte Leptospira biflexa (the TALEβgal strain). Reverse transcriptase PCR (RT-PCR) analysis and enzymatic assays demonstrated that BgaL was not expressed in the TALEβgal strain. Second, to study the role of LigA and LigB in pathogenesis, a constitutively expressed TALE gene with specificity for the homologous promoter regions of ligA and ligB was cis inserted into the pathogen Leptospira interrogans (TALElig). LigA and LigB expression was studied by using three independent clones: TALElig1, TALElig2, and TALElig3. Immunoblot analysis of osmotically induced TALElig clones demonstrated 2- to 9-fold reductions in the expression levels of LigA and LigB, with the highest reductions being noted for TALElig1 and TALElig2, which were avirulent in vivo and nonrecoverable from animal tissues. This study reconfirms galactosidase activity in the saprophyte and suggests a role for LigA and LigB in pathogenesis. Collectively, this study demonstrates that TALEs are effective at reducing the expression of targeted genes within saprophytic and pathogenic strains of Leptospira spp., providing an additional genetic manipulation tool for this genus. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Evidence for a Common Toolbox Based on Necrotrophy in a Fungal Lineage Spanning Necrotrophs, Biotrophs, Endophytes, Host Generalists and Specialists

PubMed Central

Andrew, Marion; Barua, Reeta; Short, Steven M.; Kohn, Linda M.

2012-01-01

The Sclerotiniaceae (Ascomycotina, Leotiomycetes) is a relatively recently evolved lineage of necrotrophic host generalists, and necrotrophic or biotrophic host specialists, some latent or symptomless. We hypothesized that they inherited a basic toolbox of genes for plant symbiosis from their common ancestor. Maintenance and evolutionary diversification of symbiosis could require selection on toolbox genes or on timing and magnitude of gene expression. The genes studied were chosen because their products have been previously investigated as pathogenicity factors in the Sclerotiniaceae. They encode proteins associated with cell wall degradation: acid protease 1 (acp1), aspartyl protease (asps), and polygalacturonases (pg1, pg3, pg5, pg6), and the oxalic acid (OA) pathway: a zinc finger transcription factor (pac1), and oxaloacetate acetylhydrolase (oah), catalyst in OA production, essential for full symptom production in Sclerotinia sclerotiorum. Site-specific likelihood analyses provided evidence for purifying selection in all 8 pathogenicity-related genes. Consistent with an evolutionary arms race model, positive selection was detected in 5 of 8 genes. Only generalists produced large, proliferating disease lesions on excised Arabidopsis thaliana leaves and oxalic acid by 72 hours in vitro. In planta expression of oah was 10–300 times greater among the necrotrophic host generalists than necrotrophic and biotrophic host specialists; pac1 was not differentially expressed. Ability to amplify 6/8 pathogenicity related genes and produce oxalic acid in all genera are consistent with the common toolbox hypothesis for this gene sample. That our data did not distinguish biotrophs from necrotrophs is consistent with 1) a common toolbox based on necrotrophy and 2) the most conservative interpretation of the 3-locus housekeeping gene phylogeny – a baseline of necrotrophy from which forms of biotrophy emerged at least twice. Early oah overexpression likely expands the host range of necrotrophic generalists in the Sclerotiniaceae, while specialists and biotrophs deploy oah, or other as-yet-unknown toolbox genes, differently. PMID:22253834

RNA-Seq Analysis Reveals Candidate Genes for Ontogenic Resistance in Malus-Venturia Pathosystem

PubMed Central

Gusberti, Michele; Gessler, Cesare; Broggini, Giovanni A. L.

2013-01-01

Ontogenic scab resistance in apple leaves and fruits is a horizontal resistance against the plant pathogen Venturia inaequalis and is expressed as a decrease in disease symptoms and incidence with the ageing of the leaves. Several studies at the biochemical level tried to unveil the nature of this resistance; however, no conclusive results were reported. We decided therefore to investigate the genetic origin of this phenomenon by performing a full quantitative transcriptome sequencing and comparison of young (susceptible) and old (ontogenic resistant) leaves, infected or not with the pathogen. Two time points at 72 and 96 hours post-inoculation were chosen for RNA sampling and sequencing. Comparison between the different conditions (young and old leaves, inoculated or not) should allow the identification of differentially expressed genes which may represent different induced plant defence reactions leading to ontogenic resistance or may be the cause of a constitutive (uninoculated with the pathogen) shift toward resistance in old leaves. Differentially expressed genes were then characterised for their function by homology to A. thaliana and other plant genes, particularly looking for genes involved in pathways already suspected of appertaining to ontogenic resistance in apple or other hosts, or to plant defence mechanisms in general. In this work, five candidate genes putatively involved in the ontogenic resistance of apple were identified: a gene encoding an “enhanced disease susceptibility 1 protein” was found to be down-regulated in both uninoculated and inoculated old leaves at 96 hpi, while the other four genes encoding proteins (metallothionein3-like protein, lipoxygenase, lipid transfer protein, and a peroxidase 3) were found to be constitutively up-regulated in inoculated and uninoculated old leaves. The modulation of the five candidate genes has been validated using the real-time quantitative PCR. Thus, ontogenic resistance may be the result of the corresponding up- and down-regulation of these genes. PMID:24223809

Linkage of cold acclimation and disease resistance through plant-pathogen interaction pathway in Vitis amurensis grapevine.

PubMed

Wu, Jiao; Zhang, Yali; Yin, Ling; Qu, Junjie; Lu, Jiang

2014-12-01

Low temperatures cause severe damage to none cold hardy grapevines. A preliminary survey with Solexa sequencing technology was used to analyze gene expression profiles of cold hardy Vitis amurensis 'Zuoshan-1' after cold acclimation at 4 °C for 48 h. A total of 16,750 and 18,068 putative genes were annotated for 4 °C-treated and control library, respectively. Among them, 393 genes were upregulated for at least 20-fold, while 69 genes were downregulated for at least 20-fold under the 4 °C treatment for 48 h. A subset of 101 genes from this survey was investigated further using reverse transcription polymerase chain reaction (RT-PCR). Genes associated with signaling events in pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI), including generation of calcium signals (CNGC, CMLs), jasmonic acid signal (JAZ1), oxidative burst (Rboh), and phosphorylation (FLS2, BAK, MEKK1, MKKs) cascades, were upregulated after cold acclimation. Disease resistance genes (RPM1, RPS5, RIN4, PBS1) in the process of effector-triggered immunity (ETI) were also upregulated in the current condition. Defense-related genes (WRKYs, PR1, MIN7) involved in both PTI and ETI processes were abundantly expressed after cold acclimation. Our results indicated that plant-pathogen interaction pathways were linked to the cold acclimation in V. amurensis grapevine. Other biotic- and abiotic-related genes, such as defense (protein phosphatase 2C, U-box domain proteins, NCED1, stilbene synthase), transcription (DREBs, MYBs, ERFs, ZFPs), signal transduction (kinase, calcium, and auxin signaling), transport (ATP-binding cassette (ABC) transporters, auxin:hydrogen symporter), and various metabolism, were also abundantly expressed in the cold acclimation of V. Amurensis 'Zuoshan-1' grapevine. This study revealed a series of critical genes and pathways to delineate important biological processes affected by low temperature in 'Zuoshan-1'.

Molecular and functional analyses of novel anti-lipopolysaccharide factors in giant river prawn (Macrobrachium rosenbergii, De Man) and their expression responses under pathogen and temperature exposure.

PubMed

Srisapoome, Prapansak; Klongklaew, Nawanith; Areechon, Nontawith; Wongpanya, Ratree

2018-06-15

Anti-lipopolysaccharide factor (ALF) is an immune-related protein that is crucially involved in immune defense mechanisms against invading pathogens in crustaceans. In the current study, three different ALFs of giant river prawn (Mr-ALF3, Mr-ALF8 and Mr-ALF9) were discovered. Based on sequence analysis, Mr-ALF3 and Mr-ALF9 were identified as new members of ALFs in crustaceans (groups F and G, respectively). Structurally, each newly identified Mr-ALF contained three α-helices packed against a four-stranded β-sheet bearing the LPS-binding motif, which usually binds to the cell wall components of bacteria. Tissue expression analysis using quantitative real-time RT-PCR (qRT-PCR) demonstrated that Mr-ALF3 was expressed in most tissues, and the highest expression was in the heart and hemocytes. The Mr-ALF8 gene was highly expressed in the heart, hemocytes, midgut, hepatopacreas and hindgut, respectively, while the Mr-ALF9 gene was modestly expressed in the heart and hemocytes, respectively. The transcriptional responses of the Mr-ALFs to Aeromonas hydrophila and hot/cold temperatures were investigated by qRT-PCR in the gills, hepatopancreas and hemocytes. We found that all Mr-ALFs were clearly suppressed in all tested tissues when the experimental prawns were exposed to extreme temperatures (25 and 35 °C). Moreover, the expression levels of these genes were significantly induced in all examined tissues by 2 different concentrations of A. hydrophila (1 × 10 6 and 1 × 10 9  CFU/ml), particularly 12 and 96 h after the injection. Finally, binding activity analysis of LPS-motif peptides of each Mr-ALF revealed that the LPS peptide of Mr-ALF3 exhibited the strongest adhesion to two pathogenic Gram-negative bacteria, A. hydrophila and Vibrio harveyi, and the non-pathogenic Gram-positive Bacillus megaterium. The results also showed that the Mr-ALF8 and Mr-ALF9 peptides had mild antimicrobial effects against similar tested bacteria. Based on information obtained in this study, novel ALF genes were clearly identified. Analyses of their responses under pathogenic and temperature stresses demonstrated the binding and antimicrobial activities of these ALFs and the consequent physiological effects, indicating their crucial functional roles in the prawn immune system. Copyright © 2018 Elsevier Ltd. All rights reserved.

Identification, characterisation and expression analysis of natural killer receptor genes in Chlamydia pecorum infected koalas (Phascolarctos cinereus).

PubMed

Morris, Katrina M; Mathew, Marina; Waugh, Courtney; Ujvari, Beata; Timms, Peter; Polkinghorne, Adam; Belov, Katherine

2015-10-15

Koalas (Phascolarctos cinereus), an iconic Australian marsupial, are being heavily impacted by the spread of Chlamydia pecorum, an obligate intracellular bacterial pathogen. Koalas vary in their response to this pathogen, with some showing no symptoms, while others suffer severe symptoms leading to infertility, blindness or death. Little is known about the pathology of this disease and the immune response against it in this host. Studies have demonstrated that natural killer (NK) cells, key components of the innate immune system, are involved in the immune response to chlamydial infections in humans. These cells can directly lyse cells infected by intracellular pathogens and their ability to recognise these infected cells is mediated through NK receptors on their surface. These are encoded in two regions of the genome, the leukocyte receptor complex (LRC) and the natural killer complex (NKC). These two families evolve rapidly and different repertoires of genes, which have evolved by gene duplication, are seen in different species. In this study we aimed to characterise genes belonging to the NK receptor clusters in the koala by searching available koala transcriptomes using a combination of search methods. We developed a qPCR assay to quantify relative expression of four genes, two encoded within the NK receptor cluster (CLEC1B, CLEC4E) and two known to play a role in NK response to Chalmydia in humans (NCR3, PRF1). We found that the NK receptor repertoire of the koala closely resembles that of the Tasmanian devil, with minimal genes in the NKC, but with lineage specific expansions in the LRC. Additional genes important for NK cell activity, NCR3 and PRF1, were also identified and characterised. In a preliminary study to investigate whether these genes are involved in the koala immune response to infection by its chlamydial pathogen, C. pecorum, we investigated the expression of four genes in koalas with active chlamydia infection, those with past infection and those without infection using qPCR. This analysis revealed that one of these four, CLEC4E, may be upregulated in response to chlamydia infection. We have characterised genes of the NKC and LRC in koalas and have discovered evidence that one of these genes may be upregulated in koalas with chlamydia, suggesting that these receptors may play a role in the immune response of koalas to chlamydia infection.

Silencing of genes involved in Anaplasma marginale-tick interactions affects the pathogen developmental cycle in Dermacentor variabilis.

PubMed

Kocan, Katherine M; Zivkovic, Zorica; Blouin, Edmour F; Naranjo, Victoria; Almazán, Consuelo; Mitra, Ruchira; de la Fuente, José

2009-07-16

The cattle pathogen, Anaplasma marginale, undergoes a developmental cycle in ticks that begins in gut cells. Transmission to cattle occurs from salivary glands during a second tick feeding. At each site of development two forms of A. marginale (reticulated and dense) occur within a parasitophorous vacuole in the host cell cytoplasm. However, the role of tick genes in pathogen development is unknown. Four genes, found in previous studies to be differentially expressed in Dermacentor variabilis ticks in response to infection with A. marginale, were silenced by RNA interference (RNAi) to determine the effect of silencing on the A. marginale developmental cycle. These four genes encoded for putative glutathione S-transferase (GST), salivary selenoprotein M (SelM), H+ transporting lysosomal vacuolar proton pump (vATPase) and subolesin. The impact of gene knockdown on A. marginale tick infections, both after acquiring infection and after a second transmission feeding, was determined and studied by light microscopy. Silencing of these genes had a different impact on A. marginale development in different tick tissues by affecting infection levels, the densities of colonies containing reticulated or dense forms and tissue morphology. Salivary gland infections were not seen in any of the gene-silenced ticks, raising the question of whether these ticks were able to transmit the pathogen. The results of this RNAi and light microscopic analyses of tick tissues infected with A. marginale after the silencing of genes functionally important for pathogen development suggest a role for these molecules during pathogen life cycle in ticks.

Survival, gene and metabolite responses of Litoria verreauxii alpina frogs to fungal disease chytridiomycosis

NASA Astrophysics Data System (ADS)

Grogan, Laura F.; Mulvenna, Jason; Gummer, Joel P. A.; Scheele, Ben C.; Berger, Lee; Cashins, Scott D.; McFadden, Michael S.; Harlow, Peter; Hunter, David A.; Trengove, Robert D.; Skerratt, Lee F.

2018-03-01

The fungal skin disease chytridiomycosis has caused the devastating decline and extinction of hundreds of amphibian species globally, yet the potential for evolving resistance, and the underlying pathophysiological mechanisms remain poorly understood. We exposed 406 naïve, captive-raised alpine tree frogs (Litoria verreauxii alpina) from multiple populations (one evolutionarily naïve to chytridiomycosis) to the aetiological agent Batrachochytrium dendrobatidis in two concurrent and controlled infection experiments. We investigated (A) survival outcomes and clinical pathogen burdens between populations and clutches, and (B) individual host tissue responses to chytridiomycosis. Here we present multiple interrelated datasets associated with these exposure experiments, including animal signalment, survival and pathogen burden of 355 animals from Experiment A, and the following datasets related to 61 animals from Experiment B: animal signalment and pathogen burden; raw RNA-Seq reads from skin, liver and spleen tissues; de novo assembled transcriptomes for each tissue type; raw gene expression data; annotation data for each gene; and raw metabolite expression data from skin and liver tissues. These data provide an extensive baseline for future analyses.

The fire blight pathogen Erwinia amylovora requires the rpoN gene for pathogenicity in apple.

PubMed

Ramos, Laura S; Lehman, Brian L; Sinn, Judith P; Pfeufer, Emily E; Halbrendt, Noemi O; McNellis, Timothy W

2013-10-01

RpoN is a σ(54) factor regulating essential virulence gene expression in several plant pathogenic bacteria, including Pseudomonas syringae and Pectobacterium carotovorum. In this study, we found that mutation of rpoN in the fire blight pathogen Erwinia amylovora caused a nonpathogenic phenotype. The E. amylovora rpoN Tn5 transposon mutant rpoN1250::Tn5 did not cause fire blight disease symptoms on shoots of mature apple trees. In detached immature apple fruits, the rpoN1250::Tn5 mutant failed to cause fire blight disease symptoms and grew to population levels 12 orders of magnitude lower than the wild-type. In addition, the rpoN1250::Tn5 mutant failed to elicit a hypersensitive response when infiltrated into nonhost tobacco plant leaves, and rpoN1250::Tn5 cells failed to express HrpN protein when grown in hrp (hypersensitive response and pathogenicity)-inducing liquid medium. A plasmid-borne copy of the wild-type rpoN gene complemented all the rpoN1250::Tn5 mutant phenotypes tested. The rpoN1250::Tn5 mutant was prototrophic on minimal solid and liquid media, indicating that the rpoN1250::Tn5 nonpathogenic phenotype was not caused by a defect in basic metabolism or growth. This study provides clear genetic evidence that rpoN is an essential virulence gene of E. amylovora, suggesting that rpoN has the same function in E. amylovora as in P. syringae and Pe. carotovorum. 2013 BSPP and JOHN WILEY & SONS LTD

Disarming Bacterial Virulence through Chemical Inhibition of the DNA Binding Domain of an AraC-like Transcriptional Activator Protein*

PubMed Central

Yang, Ji; Hocking, Dianna M.; Cheng, Catherine; Dogovski, Con; Perugini, Matthew A.; Holien, Jessica K.; Parker, Michael W.; Hartland, Elizabeth L.; Tauschek, Marija; Robins-Browne, Roy M.

2013-01-01

The misuse of antibiotics during past decades has led to pervasive antibiotic resistance in bacteria. Hence, there is an urgent need for the development of new and alternative approaches to combat bacterial infections. In most bacterial pathogens the expression of virulence is tightly regulated at the transcriptional level. Therefore, targeting pathogens with drugs that interfere with virulence gene expression offers an effective alternative to conventional antimicrobial chemotherapy. Many Gram-negative intestinal pathogens produce AraC-like proteins that control the expression of genes required for infection. In this study we investigated the prototypical AraC-like virulence regulator, RegA, from the mouse attaching and effacing pathogen, Citrobacter rodentium, as a potential drug target. By screening a small molecule chemical library and chemical optimization, we identified two compounds that specifically inhibited the ability of RegA to activate its target promoters and thus reduced expression of a number of proteins required for virulence. Biophysical, biochemical, genetic, and computational analyses indicated that the more potent of these two compounds, which we named regacin, disrupts the DNA binding capacity of RegA by interacting with amino acid residues within a conserved region of the DNA binding domain. Oral administration of regacin to mice, commencing 15 min before or 12 h after oral inoculation with C. rodentium, caused highly significant attenuation of intestinal colonization by the mouse pathogen comparable to that of an isogenic regA-deletion mutant. These findings demonstrate that chemical inhibition of the DNA binding domains of transcriptional regulators is a viable strategy for the development of antimicrobial agents that target bacterial pathogens. PMID:24019519

Gene Expression Analysis of Plum pox virus (Sharka) Susceptibility/Resistance in Apricot (Prunus armeniaca L.).

PubMed

Rubio, Manuel; Ballester, Ana Rosa; Olivares, Pedro Manuel; Castro de Moura, Manuel; Dicenta, Federico; Martínez-Gómez, Pedro

2015-01-01

RNA-Seq has proven to be a very powerful tool in the analysis of the Plum pox virus (PPV, sharka disease)/Prunus interaction. This technique is an important complementary tool to other means of studying genomics. In this work an analysis of gene expression of resistance/susceptibility to PPV in apricot is performed. RNA-Seq has been applied to analyse the gene expression changes induced by PPV infection in leaves from two full-sib apricot genotypes, "Rojo Pasión" and "Z506-7", resistant and susceptible to PPV, respectively. Transcriptomic analyses revealed the existence of more than 2,000 genes related to the pathogen response and resistance to PPV in apricot. These results showed that the response to infection by the virus in the susceptible genotype is associated with an induction of genes involved in pathogen resistance such as the allene oxide synthase, S-adenosylmethionine synthetase 2 and the major MLP-like protein 423. Over-expression of the Dicer protein 2a may indicate the suppression of a gene silencing mechanism of the plant by PPV HCPro and P1 PPV proteins. On the other hand, there were 164 genes involved in resistance mechanisms that have been identified in apricot, 49 of which are located in the PPVres region (scaffold 1 positions from 8,050,804 to 8,244,925), which is responsible for PPV resistance in apricot. Among these genes in apricot there are several MATH domain-containing genes, although other genes inside (Pleiotropic drug resistance 9 gene) or outside (CAP, Cysteine-rich secretory proteins, Antigen 5 and Pathogenesis-related 1 protein; and LEA, Late embryogenesis abundant protein) PPVres region could also be involved in the resistance.

Gene Expression Analysis of Plum pox virus (Sharka) Susceptibility/Resistance in Apricot (Prunus armeniaca L.)

PubMed Central

Rubio, Manuel; Ballester, Ana Rosa; Olivares, Pedro Manuel; Castro de Moura, Manuel; Dicenta, Federico; Martínez-Gómez, Pedro

2015-01-01

RNA-Seq has proven to be a very powerful tool in the analysis of the Plum pox virus (PPV, sharka disease)/Prunus interaction. This technique is an important complementary tool to other means of studying genomics. In this work an analysis of gene expression of resistance/susceptibility to PPV in apricot is performed. RNA-Seq has been applied to analyse the gene expression changes induced by PPV infection in leaves from two full-sib apricot genotypes, “Rojo Pasión” and “Z506-7”, resistant and susceptible to PPV, respectively. Transcriptomic analyses revealed the existence of more than 2,000 genes related to the pathogen response and resistance to PPV in apricot. These results showed that the response to infection by the virus in the susceptible genotype is associated with an induction of genes involved in pathogen resistance such as the allene oxide synthase, S-adenosylmethionine synthetase 2 and the major MLP-like protein 423. Over-expression of the Dicer protein 2a may indicate the suppression of a gene silencing mechanism of the plant by PPV HCPro and P1 PPV proteins. On the other hand, there were 164 genes involved in resistance mechanisms that have been identified in apricot, 49 of which are located in the PPVres region (scaffold 1 positions from 8,050,804 to 8,244,925), which is responsible for PPV resistance in apricot. Among these genes in apricot there are several MATH domain-containing genes, although other genes inside (Pleiotropic drug resistance 9 gene) or outside (CAP, Cysteine-rich secretory proteins, Antigen 5 and Pathogenesis-related 1 protein; and LEA, Late embryogenesis abundant protein) PPVres region could also be involved in the resistance. PMID:26658051

Investigation of the Fusarium virguliforme Transcriptomes Induced during Infection of Soybean Roots Suggests that Enzymes with Hydrolytic Activities Could Play a Major Role in Root Necrosis

PubMed Central

Sahu, Binod B.; Baumbach, Jordan L.; Singh, Prashant; Srivastava, Subodh K.; Yi, Xiaoping

2017-01-01

Sudden death syndrome (SDS) is caused by the fungal pathogen, Fusarium virguliforme, and is a major threat to soybean production in North America. There are two major components of this disease: (i) root necrosis and (ii) foliar SDS. Root symptoms consist of root necrosis with vascular discoloration. Foliar SDS is characterized by interveinal chlorosis and leaf necrosis, and in severe cases by flower and pod abscission. A major toxin involved in initiating foliar SDS has been identified. Nothing is known about how root necrosis develops. In order to unravel the mechanisms used by the pathogen to cause root necrosis, the transcriptome of the pathogen in infected soybean root tissues of a susceptible cultivar, ‘Essex’, was investigated. The transcriptomes of the germinating conidia and mycelia were also examined. Of the 14,845 predicted F. virguliforme genes, we observed that 12,017 (81%) were expressed in germinating conidia and 12,208 (82%) in mycelia and 10,626 (72%) in infected soybean roots. Of the 10,626 genes induced in infected roots, 224 were transcribed only following infection. Expression of several infection-induced genes encoding enzymes with oxidation-reduction properties suggests that degradation of antimicrobial compounds such as the phytoalexin, glyceollin, could be important in early stages of the root tissue infection. Enzymes with hydrolytic and catalytic activities could play an important role in establishing the necrotrophic phase. The expression of a large number of genes encoding enzymes with catalytic and hydrolytic activities during the late infection stages suggests that cell wall degradation could be involved in root necrosis and the establishment of the necrotrophic phase in this pathogen. PMID:28095498

Investigation of the Fusarium virguliforme Transcriptomes Induced during Infection of Soybean Roots Suggests that Enzymes with Hydrolytic Activities Could Play a Major Role in Root Necrosis.

PubMed

Sahu, Binod B; Baumbach, Jordan L; Singh, Prashant; Srivastava, Subodh K; Yi, Xiaoping; Bhattacharyya, Madan K

2017-01-01

Sudden death syndrome (SDS) is caused by the fungal pathogen, Fusarium virguliforme, and is a major threat to soybean production in North America. There are two major components of this disease: (i) root necrosis and (ii) foliar SDS. Root symptoms consist of root necrosis with vascular discoloration. Foliar SDS is characterized by interveinal chlorosis and leaf necrosis, and in severe cases by flower and pod abscission. A major toxin involved in initiating foliar SDS has been identified. Nothing is known about how root necrosis develops. In order to unravel the mechanisms used by the pathogen to cause root necrosis, the transcriptome of the pathogen in infected soybean root tissues of a susceptible cultivar, 'Essex', was investigated. The transcriptomes of the germinating conidia and mycelia were also examined. Of the 14,845 predicted F. virguliforme genes, we observed that 12,017 (81%) were expressed in germinating conidia and 12,208 (82%) in mycelia and 10,626 (72%) in infected soybean roots. Of the 10,626 genes induced in infected roots, 224 were transcribed only following infection. Expression of several infection-induced genes encoding enzymes with oxidation-reduction properties suggests that degradation of antimicrobial compounds such as the phytoalexin, glyceollin, could be important in early stages of the root tissue infection. Enzymes with hydrolytic and catalytic activities could play an important role in establishing the necrotrophic phase. The expression of a large number of genes encoding enzymes with catalytic and hydrolytic activities during the late infection stages suggests that cell wall degradation could be involved in root necrosis and the establishment of the necrotrophic phase in this pathogen.

The bZIP Transcription Factor Fgap1 Mediates Oxidative Stress Response and Trichothecene Biosynthesis But Not Virulence in Fusarium graminearum

PubMed Central

Montibus, Mathilde; Ducos, Christine; Bonnin-Verdal, Marie-Noelle; Bormann, Jorg; Ponts, Nadia; Richard-Forget, Florence; Barreau, Christian

2013-01-01

Redox sensing is of primary importance for fungi to cope with oxidant compounds found in their environment. Plant pathogens are particularly subject to the oxidative burst during the primary steps of infection. In the budding yeast Saccharomyces cerevisiae, it is the transcription factor Yap1 that mediates the response to oxidative stress via activation of genes coding for detoxification enzymes. In the cereal pathogen Fusarium graminearum, Fgap1 a homologue of Yap1 was identified and its role was investigated. During infection, this pathogen produces mycotoxins belonging to the trichothecenes family that accumulate in the grains. The global regulation of toxin biosynthesis is not completely understood. However, it is now clearly established that an oxidative stress activates the production of toxins by F. graminearum. The involvement of Fgap1 in this activation was investigated. A deleted mutant and a strain expressing a truncated constitutive form of Fgap1 were constructed. None of the mutants was affected in pathogenicity. The deleted mutant showed higher level of trichothecenes production associated with overexpression of Tri genes. Moreover activation of toxin accumulation in response to oxidative stress was no longer observed. Regarding the mutant with the truncated constitutive form of Fgap1, toxin production was strongly reduced. Expression of oxidative stress response genes was not activated in the deleted mutant and expression of the gene encoding the mitochondrial superoxide dismutase MnSOD1 was up-regulated in the mutant with the truncated constitutive form of Fgap1. Our results demonstrate that Fgap1 plays a key role in the link between oxidative stress response and F. graminearum secondary metabolism. PMID:24349499

Dissection of the cis-2-decenoic acid signaling network in Pseudomonas aeruginosa using microarray technique

PubMed Central

Rahmani-Badi, Azadeh; Sepehr, Shayesteh; Fallahi, Hossein; Heidari-Keshel, Saeed

2015-01-01

Many bacterial pathogens use quorum-sensing (QS) signaling to regulate the expression of factors contributing to virulence and persistence. Bacteria produce signals of different chemical classes. The signal molecule, known as diffusible signal factor (DSF), is a cis-unsaturated fatty acid that was first described in the plant pathogen Xanthomonas campestris. Previous works have shown that human pathogen, Pseudomonas aeruginosa, also synthesizes a structurally related molecule, characterized as cis-2-decenoic acid (C10: Δ2, CDA) that induces biofilm dispersal by multiple types of bacteria. Furthermore, CDA has been shown to be involved in inter-kingdom signaling that modulates fungal behavior. Therefore, an understanding of its signaling mechanism could suggest strategies for interference, with consequences for disease control. To identify the components of CDA signaling pathway in this pathogen, a comparative transcritpome analysis was conducted, in the presence and absence of CDA. A protein-protein interaction (PPI) network for differentially expressed (DE) genes with known function was then constructed by STRING and Cytoscape. In addition, the effects of CDA in combination with antimicrobial agents on the biofilm surface area and bacteria viability were evaluated using fluorescence microscopy and digital image analysis. Microarray analysis identified 666 differentially expressed genes in the presence of CDA and gene ontology (GO) analysis revealed that in P. aeruginosa, CDA mediates dispersion of biofilms through signaling pathways, including enhanced motility, metabolic activity, virulence as well as persistence at different temperatures. PPI data suggested that a cluster of five genes (PA4978, PA4979, PA4980, PA4982, PA4983) is involved in the CDA synthesis and perception. Combined treatments using both CDA and antimicrobial agents showed that following exposure of the biofilms to CDA, remaining cells on the surface were easily removed and killed by antimicrobials. PMID:25972860

Genome-Wide Analysis of Host Responses to Four Different Types of Microorganisms in Bombyx Mori (Lepidoptera: Bombycidae).

PubMed

Cheng, Tingcai; Lin, Ping; Huang, Lulin; Wu, Yuqian; Jin, Shengkai; Liu, Chun; Xia, Qingyou

2016-01-01

Several pathogenic microorganisms have been used to investigate the genome-wide transcriptional responses of Bombyx mori to infection. However, studies have so far each focused on one microorganism, and systematic genome-wide comparison of transcriptional responses to different pathogenic microorganisms has not been undertaken. Here, we surveyed transcriptional responses of B. mori to its natural bacterial, viral, and fungal pathogens, Bacillus bombyseptieus, B. mori nucleopolyhedrovirus (BmNPV), and Beauveria bassiana, respectively, and to nonpathogenic Escherichia coli, by microarray analysis. In total, the expression of 2,436, 1,804, 1,743, and 912 B. mori genes was modulated by infection with B. bombyseptieus, BmNPV, B. bassiana, and E. coli, respectively. Notably, the expression of 620, 400, 177, or 165 of these genes was only modulated by infection with B. bombyseptieus, BmNPV, B. bassiana, or E. coli, respectively. In contrast to the expression of genes related to juvenile hormone synthesis and metabolism, that of genes encoding juvenile hormone binding proteins was microorganism-specific. Three basal metabolic pathways were modulated by infection with any of the four microorganisms, and 3, 14, 5, and 2 metabolic pathways were specifically modulated by infection with B. bombyseptieus, BmNPV, B. bassiana, and E. coli, respectively. Interestingly, BmNPV infection modulated the JAK/STAT signaling pathway, whereas both the Imd and Toll signaling pathways were modulated by infection with B. bombyseptieus, B. bassiana, or E. coli These results elucidate potential molecular mechanisms of the host response to different microorganisms, and provide a foundation for further work on host-pathogen interaction. © The Author 2016. Published by Oxford University Press on behalf of the Entomological Society of America.

The veA gene of the pine needle pathogen Dothistroma septosporum regulates sporulation and secondary metabolism

USDA-ARS?s Scientific Manuscript database

Fungi possess genetic systems to regulate the expression of genes involved in complex processes such as development and secondary metabolite biosynthesis. The product of the velvet gene veA, first identified and characterized in Aspergillus nidulans, is a key player in the regulation of both of thes...

What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira

PubMed Central

Fouts, Derrick E.; Matthias, Michael A.; Adhikarla, Haritha; Adler, Ben; Amorim-Santos, Luciane; Berg, Douglas E.; Bulach, Dieter; Buschiazzo, Alejandro; Chang, Yung-Fu; Galloway, Renee L.; Haake, David A.; Haft, Daniel H.; Hartskeerl, Rudy; Ko, Albert I.; Levett, Paul N.; Matsunaga, James; Mechaly, Ariel E.; Monk, Jonathan M.; Nascimento, Ana L. T.; Nelson, Karen E.; Palsson, Bernhard; Peacock, Sharon J.; Picardeau, Mathieu; Ricaldi, Jessica N.; Thaipandungpanit, Janjira; Wunder, Elsio A.; Yang, X. Frank; Zhang, Jun-Jie; Vinetz, Joseph M.

2016-01-01

Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade’s refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic) vs. non-infectious Leptospira, this work provides new insights into the evolution of a genus of bacterial pathogens. This work will be a comprehensive roadmap for understanding leptospirosis pathogenesis. More generally, it provides new insights into mechanisms by which bacterial pathogens adapt to mammalian hosts. PMID:26890609

What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira.

PubMed

Fouts, Derrick E; Matthias, Michael A; Adhikarla, Haritha; Adler, Ben; Amorim-Santos, Luciane; Berg, Douglas E; Bulach, Dieter; Buschiazzo, Alejandro; Chang, Yung-Fu; Galloway, Renee L; Haake, David A; Haft, Daniel H; Hartskeerl, Rudy; Ko, Albert I; Levett, Paul N; Matsunaga, James; Mechaly, Ariel E; Monk, Jonathan M; Nascimento, Ana L T; Nelson, Karen E; Palsson, Bernhard; Peacock, Sharon J; Picardeau, Mathieu; Ricaldi, Jessica N; Thaipandungpanit, Janjira; Wunder, Elsio A; Yang, X Frank; Zhang, Jun-Jie; Vinetz, Joseph M

2016-02-01

Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade's refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic) vs. non-infectious Leptospira, this work provides new insights into the evolution of a genus of bacterial pathogens. This work will be a comprehensive roadmap for understanding leptospirosis pathogenesis. More generally, it provides new insights into mechanisms by which bacterial pathogens adapt to mammalian hosts.

Involvement of Abscisic Acid in the Coordinated Regulation of a Stress-Inducible Hexose Transporter (VvHT5) and a Cell Wall Invertase in Grapevine in Response to Biotrophic Fungal Infection[W

PubMed Central

Hayes, Matthew A.; Feechan, Angela; Dry, Ian B.

2010-01-01

Biotrophic fungal and oomycete pathogens alter carbohydrate metabolism in infected host tissues. Symptoms such as elevated soluble carbohydrate concentrations and increased invertase activity suggest that a pathogen-induced carbohydrate sink is established. To identify pathogen-induced regulators of carbohydrate sink strength, quantitative real-time polymerase chain reaction was used to measure transcript levels of invertase and hexose transporter genes in biotrophic pathogen-infected grapevine (Vitis vinifera) leaves. The hexose transporter VvHT5 was highly induced in coordination with the cell wall invertase gene VvcwINV by powdery and downy mildew infection. However, similar responses were also observed in response to wounding, suggesting that this is a generalized response to stress. Analysis of the VvHT5 promoter region indicated the presence of multiple abscisic acid (ABA) response elements, suggesting a role for ABA in the transition from source to sink under stress conditions. ABA treatment of grape leaves was found to reproduce the same gene-specific transcriptional changes as observed under biotic and abiotic stress conditions. Furthermore, the key regulatory ABA biosynthetic gene, VvNCED1, was activated under these same stress conditions. VvHT5 promoter::β-glucuronidase-directed expression in transgenic Arabidopsis (Arabidopsis thaliana) was activated by infection with powdery mildew and by ABA treatment, and the expression was closely associated with vascular tissue adjacent to infected regions. Unlike VvHT1 and VvHT3, which appear to be predominantly involved in hexose transport in developing leaves and berries, VvHT5 appears to have a specific role in enhancing sink strength under stress conditions, and this is controlled through ABA. Our data suggest a central role for ABA in the regulation of VvcwINV and VvHT5 expression during the transition from source to sink in response to infection by biotrophic pathogens. PMID:20348211

Pathogenic Leptospira species express surface-exposed proteins belonging to the bacterial immunoglobulin superfamily.

PubMed

Matsunaga, James; Barocchi, Michele A; Croda, Julio; Young, Tracy A; Sanchez, Yolanda; Siqueira, Isadora; Bolin, Carole A; Reis, Mitermayer G; Riley, Lee W; Haake, David A; Ko, Albert I

2003-08-01

Proteins with bacterial immunoglobulin-like (Big) domains, such as the Yersinia pseudotuberculosis invasin and Escherichia coli intimin, are surface-expressed proteins that mediate host mammalian cell invasion or attachment. Here, we report the identification and characterization of a new family of Big domain proteins, referred to as Lig (leptospiral Ig-like) proteins, in pathogenic Leptospira. Screening of L. interrogans and L. kirschneri expression libraries with sera from leptospirosis patients identified 13 lambda phage clones that encode tandem repeats of the 90 amino acid Big domain. Two lig genes, designated ligA and ligB, and one pseudogene, ligC, were identified. The ligA and ligB genes encode amino-terminal lipoprotein signal peptides followed by 10 or 11 Big domain repeats and, in the case of ligB, a unique carboxy-terminal non-repeat domain. The organization of ligC is similar to that of ligB but contains mutations that disrupt the reading frame. The lig sequences are present in pathogenic but not saprophytic Leptospira species. LigA and LigB are expressed by a variety of virulent leptospiral strains. Loss of Lig protein and RNA transcript expression is correlated with the observed loss of virulence during culture attenuation of pathogenic strains. High-pressure freeze substitution followed by immunocytochemical electron microscopy confirmed that the Lig proteins were localized to the bacterial surface. Immunoblot studies with patient sera found that the Lig proteins are a major antigen recognized during the acute host infection. These observations demonstrate that the Lig proteins are a newly identified surface protein of pathogenic Leptospira, which by analogy to other bacterial immunoglobulin superfamily virulence factors, may play a role in host cell attachment and invasion during leptospiral pathogenesis.

microRNA-mediated R gene regulation: molecular scabbards for double-edged swords.

PubMed

Deng, Yingtian; Liu, Minglei; Li, Xiaofei; Li, Feng

2018-02-01

Plant resistance (R) proteins are immune receptors that recognize pathogen effectors and trigger rapid defense responses, namely effector-triggered immunity. R protein-mediated pathogen resistance is usually race specific. During plant-pathogen coevolution, plant genomes accumulated large numbers of R genes. Even though plant R genes provide important natural resources for breeding disease-resistant crops, their presence in the plant genome comes at a cost. Misregulation of R genes leads to developmental defects, such as stunted growth and reduced fertility. In the past decade, many microRNAs (miRNAs) have been identified to target various R genes in plant genomes. miRNAs reduce R gene levels under normal conditions and allow induction of R gene expression under various stresses. For these reasons, we consider R genes to be double-edged "swords" and miRNAs as molecular "scabbards". In the present review, we summarize the contributions and potential problems of these "swords" and discuss the features and production of the "scabbards", as well as the mechanisms used to pull the "sword" from the "scabbard" when needed.

Genetic characterization of the HrpL regulon of the fire blight pathogen Erwinia amylovora reveals novel virulence factors.

PubMed

McNally, R Ryan; Toth, Ian K; Cock, Peter J A; Pritchard, Leighton; Hedley, Pete E; Morris, Jenny A; Zhao, Youfu; Sundin, George W

2012-02-01

The bacterial pathogen Erwinia amylovora is the causal agent of fire blight, an economically significant disease of apple and pear. Disease initiation by E. amylovora requires the translocation of effector proteins into host cells via the hypersensitive response and pathogenicity (hrp) type III secretion system (T3SS). The alternative sigma factor HrpL positively regulates the transcription of structural and translocated components of the T3SS via hrp promoter elements. To characterize genome-wide HrpL-dependent gene expression in E. amylovora Ea1189, wild-type and Ea1189ΔhrpL strains were cultured in hrp-inducing minimal medium, and total RNA was compared using a custom microarray designed to represent the annotated genes of E. amylovora ATCC 49946. The results revealed 24 genes differentially regulated in Ea1189ΔhrpL relative to Ea1189 with fold-change expression ratios greater than 1.5; of these, 19 genes exhibited decreased transcript abundance and five genes showed increased transcript abundance relative to Ea1189. To expand our understanding of the HrpL regulon and to elucidate direct versus indirect HrpL-mediated effects on gene expression, the genome of E. amylovora ATCC 49946 was examined in silico using a hidden Markov model assembled from known Erwinia spp. hrp promoters. This technique identified 15 putative type III novel hrp promoters, seven of which were validated with quantitative polymerase chain reaction based on expression analyses. It was found that HrpL-regulated genes encode all known components of the hrp T3SS, as well as five putative type III effectors. Eight genes displayed apparent indirect HrpL regulation, suggesting that the HrpL regulon is connected to downstream signalling networks. The construction of deletion mutants of three novel HrpL-regulated genes resulted in the identification of additional virulence factors as well as mutants displaying abnormal motility and biofilm phenotypes. © 2011 The Authors. Molecular Plant Pathology © 2011 BSPP and Blackwell Publishing Ltd.

Uncovering leaf rust responsive miRNAs in wheat (Triticum aestivum L.) using high-throughput sequencing and prediction of their targets through degradome analysis.

PubMed

Kumar, Dhananjay; Dutta, Summi; Singh, Dharmendra; Prabhu, Kumble Vinod; Kumar, Manish; Mukhopadhyay, Kunal

2017-01-01

Deep sequencing identified 497 conserved and 559 novel miRNAs in wheat, while degradome analysis revealed 701 targets genes. QRT-PCR demonstrated differential expression of miRNAs during stages of leaf rust progression. Bread wheat (Triticum aestivum L.) is an important cereal food crop feeding 30 % of the world population. Major threat to wheat production is the rust epidemics. This study was targeted towards identification and functional characterizations of micro(mi)RNAs and their target genes in wheat in response to leaf rust ingression. High-throughput sequencing was used for transcriptome-wide identification of miRNAs and their expression profiling in retort to leaf rust using mock and pathogen-inoculated resistant and susceptible near-isogenic wheat plants. A total of 1056 mature miRNAs were identified, of which 497 miRNAs were conserved and 559 miRNAs were novel. The pathogen-inoculated resistant plants manifested more miRNAs compared with the pathogen infected susceptible plants. The miRNA counts increased in susceptible isoline due to leaf rust, conversely, the counts decreased in the resistant isoline in response to pathogenesis illustrating precise spatial tuning of miRNAs during compatible and incompatible interaction. Stem-loop quantitative real-time PCR was used to profile 10 highly differentially expressed miRNAs obtained from high-throughput sequencing data. The spatio-temporal profiling validated the differential expression of miRNAs between the isolines as well as in retort to pathogen infection. Degradome analysis provided 701 predicted target genes associated with defense response, signal transduction, development, metabolism, and transcriptional regulation. The obtained results indicate that wheat isolines employ diverse arrays of miRNAs that modulate their target genes during compatible and incompatible interaction. Our findings contribute to increase knowledge on roles of microRNA in wheat-leaf rust interactions and could help in rust resistance breeding programs.

Differential Expression of Inflammatory Cytokines and Stress Genes in Male and Female Mice in Response to a Lipopolysaccharide Challenge

PubMed Central

Everhardt Queen, Ashleigh; Moerdyk-Schauwecker, Megan; McKee, Leslie M.; Leamy, Larry J.

2016-01-01

Background Sex plays a key role in an individual’s immune response against pathogenic challenges such that females fare better when infected with certain pathogens. It is thought that sex hormones impact gene expression in immune cells and lead to sexually dimorphic responses to pathogens. We predicted that, in the presence of E. coli gram-negative lipopolysaccharide (LPS), there would be a sexually dimorphic response in proinflammatory cytokine production and acute phase stress gene expression and that these responses might vary among different mouse strains and times in a pattern opposite to that of body temperature associated with LPS-induced shock. Materials and Methods Interleukin-6 (IL-6), macrophage inflammatory protein-Iβ (MIP-1β), tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β) as well as beta-fibrinogen (Fgb) and metallothionein-1 (Mt-1) mRNA expression were measured at four time points (0, 2, 4 and 7 hours) after injection of E. coli LPS in mice from three inbred strains. Results Statistical analysis using analyses of variance (ANOVAs) showed that the levels of the all six traits changed over time, generally peaking at 2 hours after LPS injection. Mt-1, Fgb, and IL-6 showed differences among strains, although these were time-specific. Sexual dimorphism was seen for Fgb and IL6, and was most pronounced at the latest time period (7 hours) where male levels exceeded those for females. Trends for all six cytokine/gene expression traits were negatively correlated with those for body temperatures. Discussion The higher levels of expression of Fgb and IL6 in males compared with females are consistent with the greater vulnerability of males to infection and subsequent inflammation. Temperature appears to be a useful proxy for mortality in endotoxic shock, but sexual dimorphism in cytokine and stress gene expression levels may persist after an LPS challenge even if temperatures in the two sexes are similar and have begun to stabilize. PMID:27120355

Innovative approach for transcriptomic analysis of obligate intracellular pathogen: selective capture of transcribed sequences of Ehrlichia ruminantium

PubMed Central

2009-01-01

Background Whole genome transcriptomic analysis is a powerful approach to elucidate the molecular mechanisms controlling the pathogenesis of obligate intracellular bacteria. However, the major hurdle resides in the low quantity of prokaryotic mRNAs extracted from host cells. Our model Ehrlichia ruminantium (ER), the causative agent of heartwater, is transmitted by tick Amblyomma variegatum. This bacterium affects wild and domestic ruminants and is present in Sub-Saharan Africa and the Caribbean islands. Because of its strictly intracellular location, which constitutes a limitation for its extensive study, the molecular mechanisms involved in its pathogenicity are still poorly understood. Results We successfully adapted the SCOTS method (Selective Capture of Transcribed Sequences) on the model Rickettsiales ER to capture mRNAs. Southern Blots and RT-PCR revealed an enrichment of ER's cDNAs and a diminution of ribosomal contaminants after three rounds of capture. qRT-PCR and whole-genome ER microarrays hybridizations demonstrated that SCOTS method introduced only a limited bias on gene expression. Indeed, we confirmed the differential gene expression between poorly and highly expressed genes before and after SCOTS captures. The comparative gene expression obtained from ER microarrays data, on samples before and after SCOTS at 96 hpi was significantly correlated (R2 = 0.7). Moreover, SCOTS method is crucial for microarrays analysis of ER, especially for early time points post-infection. There was low detection of transcripts for untreated samples whereas 24% and 70.7% were revealed for SCOTS samples at 24 and 96 hpi respectively. Conclusions We conclude that this SCOTS method has a key importance for the transcriptomic analysis of ER and can be potentially used for other Rickettsiales. This study constitutes the first step for further gene expression analyses that will lead to a better understanding of both ER pathogenicity and the adaptation of obligate intracellular bacteria to their environment. PMID:20034374

Regulatory principles governing Salmonella and Yersinia virulence

PubMed Central

Erhardt, Marc; Dersch, Petra

2015-01-01

Enteric pathogens such as Salmonella and Yersinia evolved numerous strategies to survive and proliferate in different environmental reservoirs and mammalian hosts. Deciphering common and pathogen-specific principles for how these bacteria adjust and coordinate spatiotemporal expression of virulence determinants, stress adaptation, and metabolic functions is fundamental to understand microbial pathogenesis. In order to manage sudden environmental changes, attacks by the host immune systems and microbial competition, the pathogens employ a plethora of transcriptional and post-transcriptional control elements, including transcription factors, sensory and regulatory RNAs, RNAses, and proteases, to fine-tune and control complex gene regulatory networks. Many of the contributing global regulators and the molecular mechanisms of regulation are frequently conserved between Yersinia and Salmonella. However, the interplay, arrangement, and composition of the control elements vary between these closely related enteric pathogens, which generate phenotypic differences leading to distinct pathogenic properties. In this overview we present common and different regulatory networks used by Salmonella and Yersinia to coordinate the expression of crucial motility, cell adhesion and invasion determinants, immune defense strategies, and metabolic adaptation processes. We highlight evolutionary changes of the gene regulatory circuits that result in different properties of the regulatory elements and how this influences the overall outcome of the infection process. PMID:26441883

Burkholderia mallei and Burkholderia pseudomallei Cluster 1 Type VI Secretion System Gene Expression Is Negatively Regulated by Iron and Zinc

PubMed Central

Burtnick, Mary N.; Brett, Paul J.

2013-01-01

Burkholderia mallei is a facultative intracellular pathogen that causes glanders in humans and animals. Previous studies have demonstrated that the cluster 1 type VI secretion system (T6SS-1) expressed by this organism is essential for virulence in hamsters and is positively regulated by the VirAG two-component system. Recently, we have shown that T6SS-1 gene expression is up-regulated following internalization of this pathogen into phagocytic cells and that this system promotes multinucleated giant cell formation in infected tissue culture monolayers. In the present study, we further investigated the complex regulation of this important virulence factor. To assess T6SS-1 expression, B. mallei strains were cultured in various media conditions and Hcp1 production was analyzed by Western immunoblotting. Transcript levels of several VirAG-regulated genes (bimA, tssA, hcp1 and tssM) were also determined using quantitative real time PCR. Consistent with previous observations, T6SS-1 was not expressed during growth of B. mallei in rich media. Curiously, growth of the organism in minimal media (M9G) or minimal media plus casamino acids (M9CG) facilitated robust expression of T6SS-1 genes whereas growth in minimal media plus tryptone (M9TG) did not. Investigation of this phenomenon confirmed a regulatory role for VirAG in this process. Additionally, T6SS-1 gene expression was significantly down-regulated by the addition of iron and zinc to M9CG. Other genes under the control of VirAG did not appear to be as tightly regulated by these divalent metals. Similar results were observed for B. pseudomallei, but not for B. thailandensis. Collectively, our findings indicate that in addition to being positively regulated by VirAG, B. mallei and B. pseudomallei T6SS-1 gene expression is negatively regulated by iron and zinc. PMID:24146925

Burkholderia mallei and Burkholderia pseudomallei cluster 1 type VI secretion system gene expression is negatively regulated by iron and zinc.

PubMed

Burtnick, Mary N; Brett, Paul J

2013-01-01

Burkholderia mallei is a facultative intracellular pathogen that causes glanders in humans and animals. Previous studies have demonstrated that the cluster 1 type VI secretion system (T6SS-1) expressed by this organism is essential for virulence in hamsters and is positively regulated by the VirAG two-component system. Recently, we have shown that T6SS-1 gene expression is up-regulated following internalization of this pathogen into phagocytic cells and that this system promotes multinucleated giant cell formation in infected tissue culture monolayers. In the present study, we further investigated the complex regulation of this important virulence factor. To assess T6SS-1 expression, B. mallei strains were cultured in various media conditions and Hcp1 production was analyzed by Western immunoblotting. Transcript levels of several VirAG-regulated genes (bimA, tssA, hcp1 and tssM) were also determined using quantitative real time PCR. Consistent with previous observations, T6SS-1 was not expressed during growth of B. mallei in rich media. Curiously, growth of the organism in minimal media (M9G) or minimal media plus casamino acids (M9CG) facilitated robust expression of T6SS-1 genes whereas growth in minimal media plus tryptone (M9TG) did not. Investigation of this phenomenon confirmed a regulatory role for VirAG in this process. Additionally, T6SS-1 gene expression was significantly down-regulated by the addition of iron and zinc to M9CG. Other genes under the control of VirAG did not appear to be as tightly regulated by these divalent metals. Similar results were observed for B. pseudomallei, but not for B. thailandensis. Collectively, our findings indicate that in addition to being positively regulated by VirAG, B. mallei and B. pseudomallei T6SS-1 gene expression is negatively regulated by iron and zinc.

Chromatin versus pathogens: the function of epigenetics in plant immunity.

PubMed

Ding, Bo; Wang, Guo-Liang

2015-01-01

To defend against pathogens, plants have developed a sophisticated innate immunity that includes effector recognition, signal transduction, and rapid defense responses. Recent evidence has demonstrated that plants utilize the epigenetic control of gene expression to fine-tune their defense when challenged by pathogens. In this review, we highlight the current understanding of the molecular mechanisms of histone modifications (i.e., methylation, acetylation, and ubiquitination) and chromatin remodeling that contribute to plant immunity against pathogens. Functions of key histone-modifying and chromatin remodeling enzymes are discussed.

Expression of the Theobroma cacao Bax-inhibitor-1 gene in tomato reduces infection by the hemibiotrophic pathogen Moniliophthora perniciosa.

PubMed

Scotton, Danielle Camargo; Azevedo, Mariana Da Silva; Sestari, Ivan; Da Silva, Jamille Santos; Souza, Lucas Anjos; Peres, Lázaro Eustáquio Pereira; Leal, Gildemberg Amorim; Figueira, Antonio

2017-10-01

Programmed cell death (PCD) plays a key role in plant responses to pathogens, determining the success of infection depending on the pathogen lifestyle and on which participant of the interaction triggers cell death. The hemibiotrophic basidiomycete Moniliophthora perniciosa is the causal agent of witches' broom disease of Theobroma cacao L. (cacao), a serious constraint for production in South America and the Caribbean. It has been hypothesized that M. perniciosa pathogenesis involves PCD, initially as a plant defence mechanism, which is diverted by the fungus to induce necrosis during the dikaryotic phase of the mycelia. Here, we evaluated whether the expression of a cacao anti-apoptotic gene would affect the incidence and severity of M. perniciosa infection using the 'Micro-Tom' (MT) tomato as a model. The cacao Bax-inhibitor-1 (TcBI-1) gene, encoding a putative basal attenuator of PCD, was constitutively expressed in MT to evaluate function. Transformants expressing TcBI-1, when treated with tunicamycin, an inducer of endoplasmic reticulum stress, showed a decrease in cell peroxidation. When the same transformants were inoculated with the necrotrophic fungal pathogens Sclerotinia sclerotiorum, Sclerotium rolfsii and Botrytis cinerea, a significant reduction in infection severity was observed, confirming TcBI-1 function. After inoculation with M. perniciosa, TcBI-1 transformant lines showed a significant reduction in disease incidence compared with MT. The overexpression of TcBI-1 appears to affect the ability of germinating spores to penetrate susceptible tissues, restoring part of the non-host resistance in MT against the S-biotype of M. perniciosa. © 2016 BSPP AND JOHN WILEY & SONS LTD.

Enhanced Transcription of the Arabidopsis Disease Resistance Genes RPW8.1 and RPW8.2 via a Salicylic Acid–Dependent Amplification Circuit Is Required for Hypersensitive Cell Death

PubMed Central

Xiao, Shunyuan; Brown, Samantha; Patrick, Elaine; Brearley, Charles; Turner, John G.

2003-01-01

The Arabidopsis disease resistance (R) genes RPW8.1 and RPW8.2 couple the recognition of powdery mildew pathogens of this plant with the subsequent induction of a localized necrosis, or hypersensitive response (HR). The HR restricts the spread of the infection and renders the plant resistant. One-third of Arabidopsis plants transformed with a genomic fragment containing RPW8.1 and RPW8.2 developed spontaneous HR-like lesions (SHL) in the absence of pathogens. We demonstrate that SHL occurs in transgenic lines that contain multiple copies of the transgene and express RPW8.1 and RPW8.2 at high levels. SHL is associated with salicylic acid (SA) accumulation, and at the site of the lesion, there is increased expression of RPW8.1, increased production of H2O2, and increased expression of pathogenesis-related genes. These lesions are physiologically similar to the pathogen-induced HR mediated by RPW8.1 and RPW8.2. Significantly, environmental conditions that suppress SHL suppress the transcription of RPW8.1 and RPW8.2 and also suppress resistance to powdery mildews, even in transgenic lines containing RPW8.1 and RPW8.2 that normally do not express SHL. Furthermore, treatment with SA increases the transcription of RPW8.1 and RPW8.2, induces SHL, and enhances resistance to powdery mildews. We conclude that HR requires the transcription of RPW8.1 and RPW8.2, which is regulated independently of the pathogen by SA-dependent feedback amplification. PMID:12509520

Towards the elements of successful insect Ribonucleic acid interference (RNAi)

USDA-ARS?s Scientific Manuscript database

Ribonucleic acid interference (RNAi), the sequence-specific suppression of gene expression, offers great opportunities for insect science, especially to analyze gene function, manage pest populations, and reduce disease pathogens. The accumulating body of literature on insect RNAi has revealed that ...

The plant nitrogen mobilization promoted by Colletotrichum lindemuthianum in Phaseolus leaves depends on fungus pathogenicity.

PubMed

Tavernier, Virginie; Cadiou, Sandrine; Pageau, Karine; Laugé, Richard; Reisdorf-Cren, Michèle; Langin, Thierry; Masclaux-Daubresse, Céline

2007-01-01

Nitrogen plays an essential role in the nutrient relationship between plants and pathogens. Some studies report that the nitrogen-mobilizing plant metabolism that occurs during abiotic and biotic stress could be a 'slash-and-burn' defence strategy. In order to study nitrogen recycling and mobilization in host plants during pathogen attack and invasion, the Colletotrichum lindemuthianum/Phaseolus vulgaris interaction was used as a model. C. lindemuthianum is a hemibiotroph that causes anthracnose disease on P. vulgaris. Non-pathogenic mutants and the pathogenic wild-type strain were used to compare their effects on plant metabolism. The deleterious effects of infection were monitored by measuring changes in chlorophyll, protein, and amino acid concentrations. It was shown that amino acid composition changed depending on the plant-fungus interaction and that glutamine accumulated mainly in the leaves infected by the pathogenic strain. Glutamine accumulation correlated with the accumulation of cytosolic glutamine synthetase (GS1 alpha) mRNA. The most striking result was that the GS1 alpha gene was induced in all the fungus-infected leaves, independent of the strain used for inoculation, and that GS1 alpha expression paralleled the PAL3 and CHS defence gene expression. It is concluded that a role of GS1 alpha in plant defence has to be considered.

Flow cytometric purification of Colletotrichum higginsianum biotrophic hyphae from Arabidopsis leaves for stage-specific transcriptome analysis.

PubMed

Takahara, Hiroyuki; Dolf, Andreas; Endl, Elmar; O'Connell, Richard

2009-08-01

Generation of stage-specific cDNA libraries is a powerful approach to identify pathogen genes that are differentially expressed during plant infection. Biotrophic pathogens develop specialized infection structures inside living plant cells, but sampling the transcriptome of these structures is problematic due to the low ratio of fungal to plant RNA, and the lack of efficient methods to isolate them from infected plants. Here we established a method, based on fluorescence-activated cell sorting (FACS), to purify the intracellular biotrophic hyphae of Colletotrichum higginsianum from homogenates of infected Arabidopsis leaves. Specific selection of viable hyphae using a fluorescent vital marker provided intact RNA for cDNA library construction. Pilot-scale sequencing showed that the library was enriched with plant-induced and pathogenicity-related fungal genes, including some encoding small, soluble secreted proteins that represent candidate fungal effectors. The high purity of the hyphae (94%) prevented contamination of the library by sequences derived from host cells or other fungal cell types. RT-PCR confirmed that genes identified in the FACS-purified hyphae were also expressed in planta. The method has wide applicability for isolating the infection structures of other plant pathogens, and will facilitate cell-specific transcriptome analysis via deep sequencing and microarray hybridization, as well as proteomic analyses.

Predicting gene expression levels from codon biases in alpha-proteobacterial genomes.

PubMed

Karlin, Samuel; Barnett, Melanie J; Campbell, Allan M; Fisher, Robert F; Mrazek, Jan

2003-06-10

Predicted highly expressed (PHX) genes in five currently available high G+C complete alpha-proteobacterial genomes are analyzed. These include: the nitrogen-fixing plant symbionts Sinorhizobium meliloti (SINME) and Mesorhizobium loti (MESLO), the nonpathogenic aquatic bacterium Caulobacter crescentus (CAUCR), the plant pathogen Agrobacterium tumefaciens (AGRTU), and the mammalian pathogen Brucella melitensis (BRUME). Three of these genomes, SINME, AGRTU, and BRUME, contain multiple chromosomes or megaplasmids (>1 Mb length). PHX genes in these genomes are concentrated mainly in the major (largest) chromosome with few PHX genes found in the secondary chromosomes and megaplasmids. Tricarboxylic acid cycle and aerobic respiration genes are strongly PHX in all five genomes, whereas anaerobic pathways of glycolysis and fermentation are mostly not PHX. Only in MESLO (but not SINME) and BRUME are most glycolysis genes PHX. Many flagellar genes are PHX in MESLO and CAUCR, but mostly are not PHX in SINME and AGRTU. The nonmotile BRUME also carries many flagellar genes but these are generally not PHX and all but one are located in the second chromosome. CAUCR stands out among available prokaryotic genomes with 25 PHX TonB-dependent receptors. These are putatively involved in uptake of iron ions and other nonsoluble compounds.

Quantitative real-time PCR normalization for gene expression studies in the plant pathogenic fungi Lasiodiplodia theobromae.

PubMed

Paolinelli-Alfonso, Marcos; Galindo-Sánchez, Clara Elizabeth; Hernandez-Martinez, Rufina

2016-08-01

Lasiodiplodia theobromae is a highly virulent plant pathogen. It has been suggested that heat stress increases its virulence. The aim of this work was to evaluate, compare, and recommend normalization strategies for gene expression analysis of the fungus growing with grapevine wood under heat stress. Using RT-qPCR-derived data, reference gene stability was evaluated through geNorm, NormFinder and Bestkeeper applications. Based on the geometric mean using the ranking position obtained for each independent analysis, genes were ranked from least to most stable as follows: glyceraldehyde-3-phosphate dehydrogenase (GAPDH), actin (ACT), β-tubulin (TUB) and elongation factor-1α (EF1α). Using RNAseq-derived data based on the calculated tagwise dispersion these genes were ordered by increasing stability as follows: GAPDH, ACT, TUB, and EF1α. The correlation between RNAseq and RTqPCR results was used as criteria to identify the best RT-qPCR normalization approach. The gene TUB is recommended as the best option for normalization among the commonly used reference genes, but alternative fungal reference genes are also suggested. Copyright © 2016 Elsevier B.V. All rights reserved.

Regulation of gene expression in protozoa parasites.

PubMed

Gomez, Consuelo; Esther Ramirez, M; Calixto-Galvez, Mercedes; Medel, Olivia; Rodríguez, Mario A

2010-01-01

Infections with protozoa parasites are associated with high burdens of morbidity and mortality across the developing world. Despite extensive efforts to control the transmission of these parasites, the spread of populations resistant to drugs and the lack of effective vaccines against them contribute to their persistence as major public health problems. Parasites should perform a strict control on the expression of genes involved in their pathogenicity, differentiation, immune evasion, or drug resistance, and the comprehension of the mechanisms implicated in that control could help to develop novel therapeutic strategies. However, until now these mechanisms are poorly understood in protozoa. Recent investigations into gene expression in protozoa parasites suggest that they possess many of the canonical machineries employed by higher eukaryotes for the control of gene expression at transcriptional, posttranscriptional, and epigenetic levels, but they also contain exclusive mechanisms. Here, we review the current understanding about the regulation of gene expression in Plasmodium sp., Trypanosomatids, Entamoeba histolytica and Trichomonas vaginalis.

Green tissue-specific co-expression of chitinase and oxalate oxidase 4 genes in rice for enhanced resistance against sheath blight.

PubMed

Karmakar, Subhasis; Molla, Kutubuddin Ali; Chanda, Palas K; Sarkar, Sailendra Nath; Datta, Swapan K; Datta, Karabi

2016-01-01

Green tissue-specific simultaneous overexpression of two defense-related genes ( OsCHI11 & OsOXO4 ) in rice leads to significant resistance against sheath blight pathogen ( R. solani ) without distressing any agronomically important traits. Overexpressing two defense-related genes (OsOXO4 and OsCHI11) cloned from rice is effective at enhancing resistance against sheath blight caused by Rhizoctonia solani. These genes were expressed under the control of two different green tissue-specific promoters, viz. maize phosphoenolpyruvate carboxylase gene promoter, PEPC, and rice cis-acting 544-bp DNA element, immediately upstream of the D54O translational start site, P D54O-544 . Putative T0 transgenic rice plants were screened by PCR and integration of genes was confirmed by Southern hybridization of progeny (T1) rice plants. Successful expression of OsOXO4 and OsCHI11 in all tested plants was confirmed. Expression of PR genes increased significantly following pathogen infection in overexpressing transgenic plants. Following infection, transgenic plants exhibited elevated hydrogen peroxide levels, significant changes in activity of ROS scavenging enzymes and reduced membrane damage when compared to their wild-type counterpart. In a Rhizoctonia solani toxin assay, a detached leaf inoculation test and an in vivo plant bioassay, transgenic plants showed a significant reduction in disease symptoms in comparison to non-transgenic control plants. This is the first report of overexpression of two different PR genes driven by two green tissue-specific promoters providing enhanced sheath blight resistance in transgenic rice.

Analysis of apolipoprotein genes and their involvement in disease response of channel catfish after bacterial infection.

PubMed

Yang, Yujia; Fu, Qiang; Zhou, Tao; Li, Yun; Liu, Shikai; Zeng, Qifan; Wang, Xiaozhu; Jin, Yulin; Tian, Changxu; Qin, Zhenkui; Dunham, Rex; Liu, Zhanjiang

2017-02-01

Apolipoproteins are protein component of plasma lipoproteins. They exert crucial roles in lipoprotein metabolism and serve as enzyme cofactors, receptor ligands, and lipid transfer carriers in mammals. In teleosts, apolipoproteins are also involved in diverse processes including embryonic and ontogenic development, liver and digestive system organogenesis, and innate immunity. In this study, we identified a set of 19 apolipoprotein genes in channel catfish (Ictalurus punctatus). Phylogenetic analysis and syntenic analysis were conducted to determine their identities and evolutionary relationships. The expression signatures of apolipoproteins in channel catfish were determined in healthy tissues and after infections with two major bacterial pathogens, Edwardsiella ictaluri and Flavobacterium columnare. In healthy channel catfish, most apolipoprotein genes exhibited tissue-specific expression patterns in channel catfish. After ESC and columnaris infections, 5 and 7 apolipoprotein genes were differentially expressed respectively, which presented a pathogen-specific and time-dependent pattern of regulation. After ESC infection, three exchangeable apolipoproteins (apoA-IB, apoC-I, and apoE-B) were suppressed in catfish intestine, while two nonexchangeable apolipoproteins (apoB-A and apoB-B) were slightly up-regulated. After columnaris infection, apoB-B, apoD-B, and apoE-A were significantly down-regulated in catfish gill, while apoF, apoL-IV, apoO-like, and apo-14 kDa showed significantly up-regulation. Taken together, these results suggested that apolipoprotein genes may play significant roles in innate immune responses to bacterial pathogens in channel catfish. Copyright © 2016 Elsevier Ltd. All rights reserved.

The promoter of the pepper pathogen-induced membrane protein gene CaPIMP1 mediates environmental stress responses in plants.

PubMed

Hong, Jeum Kyu; Hwang, Byung Kook

2009-01-01

The promoter of the pepper pathogen-induced membrane protein gene CaPIMP1 was analyzed by an Agrobacterium-mediated transient expression assay in tobacco leaves. Several stress-related cis-acting elements (GT-1, W-box and ABRE) are located within the CaPIMP1 promoter. In tobacco leaf tissues transiently transformed with a CaPIMP1 promoter-beta-glucuronidase (GUS) gene fusion, serially 5'-deleted CaPIMP1 promoters were differentially activated by Pseudomonas syringae pv. tabaci, ethylene, methyl jasmonate, abscisic acid, and nitric oxide. The -1,193 bp region of the CaPIMP1 gene promoter sequence exhibited full promoter activity. The -417- and -593 bp promoter regions were sufficient for GUS gene activation by ethylene and methyl jasmonate treatments, respectively. However, CaPIMP1 promoter sequences longer than -793 bp were required for promoter activation by abscisic acid and sodium nitroprusside treatments. CaPIMP1 expression was activated in pepper leaves by treatment with ethylene, methyl jasmonate, abscisic acid, beta-amino-n-butyric acid, NaCl, mechanical wounding, and low temperature, but not with salicylic acid. Overexpression of CaPIMP1 in Arabidopsis conferred hypersensitivity to mannitol, NaCl, and ABA during seed germination but not during seedling development. In contrast, transgenic plants overexpressing CaPIMP1 exhibited enhanced tolerance to oxidative stress induced by methyl viologen during germination and early seedling stages. These results suggest that CaPIMP1 expression may alter responsiveness to environmental stress, as well as to pathogen infection.

The Genomes of the Fungal Plant Pathogens Cladosporium fulvum and Dothistroma septosporum Reveal Adaptation to Different Hosts and Lifestyles But Also Signatures of Common Ancestry

DOE Office of Scientific and Technical Information (OSTI.GOV)

de Wit, Pierre J. G. M.; van der Burgt, Ate; Okmen, Bilal

2012-05-04

We sequenced and compared the genomes of the Dothideomycete fungal plant pathogens Cladosporium fulvum (Cfu) (syn. Passalora fulva) and Dothistroma septosporum (Dse) that are closely related phylogenetically, but have different lifestyles and hosts. Although both fungi grow extracellularly in close contact with host mesophyll cells, Cfu is a biotroph infecting tomato, while Dse is a hemibiotroph infecting pine. The genomes of these fungi have a similar set of genes (70percent of gene content in both genomes are homologs), but differ significantly in size (Cfu >61.1-Mb; Dse 31.2-Mb), which is mainly due to the difference in repeat content (47.2percent in Cfumore » versus 3.2percent in Dse). Recent adaptation to different lifestyles and hosts is suggested by diverged sets of genes. Cfu contains an tomatinase gene that we predict might be required for detoxification of tomatine, while this gene is absent in Dse. Many genes encoding secreted proteins are unique to each species and the repeat-rich areas in Cfu are enriched for these species-specific genes. In contrast, conserved genes suggest common host ancestry. Homologs of Cfu effector genes, including Ecp2 and Avr4, are present in Dse and induce a Cf-Ecp2- and Cf-4-mediated hypersensitive response, respectively. Strikingly, genes involved in production of the toxin dothistromin, a likely virulence factor for Dse, are conserved in Cfu, but their expression differs markedly with essentially no expression by Cfu in planta. Likewise, Cfu has a carbohydrate-degrading enzyme catalog that is more similar to that of necrotrophs or hemibiotrophs and a larger pectinolytic gene arsenal than Dse, but many of these genes are not expressed in planta or are pseudogenized. Overall, comparison of their genomes suggests that these closely related plant pathogens had a common ancestral host but since adapted to different hosts and lifestyles by a combination of differentiated gene content, pseudogenization, and gene regulation.« less

Large-Scale Gene Disruption in Magnaporthe oryzae Identifies MC69, a Secreted Protein Required for Infection by Monocot and Dicot Fungal Pathogens

PubMed Central

Saitoh, Hiromasa; Fujisawa, Shizuko; Mitsuoka, Chikako; Ito, Akiko; Hirabuchi, Akiko; Ikeda, Kyoko; Irieda, Hiroki; Yoshino, Kae; Yoshida, Kentaro; Matsumura, Hideo; Tosa, Yukio; Win, Joe; Kamoun, Sophien; Takano, Yoshitaka; Terauchi, Ryohei

2012-01-01

To search for virulence effector genes of the rice blast fungus, Magnaporthe oryzae, we carried out a large-scale targeted disruption of genes for 78 putative secreted proteins that are expressed during the early stages of infection of M. oryzae. Disruption of the majority of genes did not affect growth, conidiation, or pathogenicity of M. oryzae. One exception was the gene MC69. The mc69 mutant showed a severe reduction in blast symptoms on rice and barley, indicating the importance of MC69 for pathogenicity of M. oryzae. The mc69 mutant did not exhibit changes in saprophytic growth and conidiation. Microscopic analysis of infection behavior in the mc69 mutant revealed that MC69 is dispensable for appressorium formation. However, mc69 mutant failed to develop invasive hyphae after appressorium formation in rice leaf sheath, indicating a critical role of MC69 in interaction with host plants. MC69 encodes a hypothetical 54 amino acids protein with a signal peptide. Live-cell imaging suggested that fluorescently labeled MC69 was not translocated into rice cytoplasm. Site-directed mutagenesis of two conserved cysteine residues (Cys36 and Cys46) in the mature MC69 impaired function of MC69 without affecting its secretion, suggesting the importance of the disulfide bond in MC69 pathogenicity function. Furthermore, deletion of the MC69 orthologous gene reduced pathogenicity of the cucumber anthracnose fungus Colletotrichum orbiculare on both cucumber and Nicotiana benthamiana leaves. We conclude that MC69 is a secreted pathogenicity protein commonly required for infection of two different plant pathogenic fungi, M. oryzae and C. orbiculare pathogenic on monocot and dicot plants, respectively. PMID:22589729

Comparative transcript profiling by SuperSAGE identifies novel candidate genes for controlling potato quantitative resistance to late blight not compromised by late maturity.

PubMed

Draffehn, Astrid M; Li, Li; Krezdorn, Nicolas; Ding, Jia; Lübeck, Jens; Strahwald, Josef; Muktar, Meki S; Walkemeier, Birgit; Rotter, Björn; Gebhardt, Christiane

2013-01-01

Resistance to pathogens is essential for survival of wild and cultivated plants. Pathogen susceptibility causes major losses of crop yield and quality. Durable field resistance combined with high yield and other superior agronomic characters are therefore, important objectives in every crop breeding program. Precision and efficacy of resistance breeding can be enhanced by molecular diagnostic tools, which result from knowledge of the molecular basis of resistance and susceptibility. Breeding uses resistance conferred by single R genes and polygenic quantitative resistance. The latter is partial but considered more durable. Molecular mechanisms of plant pathogen interactions are elucidated mainly in experimental systems involving single R genes, whereas most genes important for quantitative resistance in crops like potato are unknown. Quantitative resistance of potato to Phytophthora infestans causing late blight is often compromised by late plant maturity, a negative agronomic character. Our objective was to identify candidate genes for quantitative resistance to late blight not compromised by late plant maturity. We used diagnostic DNA-markers to select plants with different field levels of maturity corrected resistance (MCR) to late blight and compared their leaf transcriptomes before and after infection with P. infestans using SuperSAGE (serial analysis of gene expression) technology and next generation sequencing. We identified 2034 transcripts up or down regulated upon infection, including a homolog of the kiwi fruit allergen kiwellin. 806 transcripts showed differential expression between groups of genotypes with contrasting MCR levels. The observed expression patterns suggest that MCR is in part controlled by differential transcript levels in uninfected plants. Functional annotation suggests that, besides biotic and abiotic stress responses, general cellular processes such as photosynthesis, protein biosynthesis, and degradation play a role in MCR.

Inducible expression of Bs2 R gene from Capsicum chacoense in sweet orange (Citrus sinensis L. Osbeck) confers enhanced resistance to citrus canker disease.

PubMed

Sendín, Lorena Noelia; Orce, Ingrid Georgina; Gómez, Rocío Liliana; Enrique, Ramón; Grellet Bournonville, Carlos Froilán; Noguera, Aldo Sergio; Vojnov, Adrián Alberto; Marano, María Rosa; Castagnaro, Atilio Pedro; Filippone, María Paula

2017-04-01

Transgenic expression of the pepper Bs2 gene confers resistance to Xanthomonas campestris pv. vesicatoria (Xcv) pathogenic strains which contain the avrBs2 avirulence gene in susceptible pepper and tomato varieties. The avrBs2 gene is highly conserved among members of the Xanthomonas genus, and the avrBs2 of Xcv shares 96% homology with the avrBs2 of Xanthomonas citri subsp. citri (Xcc), the causal agent of citrus canker disease. A previous study showed that the transient expression of pepper Bs2 in lemon leaves reduced canker formation and induced plant defence mechanisms. In this work, the effect of the stable expression of Bs2 gene on citrus canker resistance was evaluated in transgenic plants of Citrus sinensis cv. Pineapple. Interestingly, Agrobacterium-mediated transformation of epicotyls was unsuccessful when a constitutive promoter (2× CaMV 35S) was used in the plasmid construction, but seven transgenic lines were obtained with a genetic construction harbouring Bs2 under the control of a pathogen-inducible promoter, from glutathione S-transferase gene from potato. A reduction of disease symptoms of up to 70% was observed in transgenic lines expressing Bs2 with respect to non-transformed control plants. This reduction was directly dependent on the Xcc avrBs2 gene since no effect was observed when a mutant strain of Xcc with a disruption in avrBs2 gene was used for inoculations. Additionally, a canker symptom reduction was correlated with levels of the Bs2 expression in transgenic plants, as assessed by real-time qPCR, and accompanied by the production of reactive oxygen species. These results indicate that the pepper Bs2 resistance gene is also functional in a family other than the Solanaceae, and could be considered for canker control.

Heat shock and prolonged heat stress attenuate neurotoxin and sporulation gene expression in group I Clostridium botulinum strain ATCC 3502

PubMed Central

Selby, Katja; Mascher, Gerald; Somervuo, Panu; Korkeala, Hannu

2017-01-01

Foodborne pathogenic bacteria are exposed to a number of environmental stresses during food processing, storage, and preparation, and in the human body. In order to improve the safety of food, the understanding of molecular stress response mechanisms foodborne pathogens employ is essential. Many response mechanisms that are activated during heat shock may cross-protect bacteria against other environmental stresses. To better understand the molecular mechanisms Clostridium botulinum, the causative agent of botulism, utilizes during acute heat stress and during adaptation to stressfully high temperature, the C. botulinum Group I strain ATCC 3502 was grown in continuous culture at 39°C and exposed to heat shock at 45°C, followed by prolonged heat stress at 45°C to allow adaptation of the culture to the high temperature. Growth in continuous culture was performed to exclude secondary growth phase effects or other environmental impacts on bacterial gene transcription. Changes in global gene expression profiles were studied using DNA microarray hybridization. During acute heat stress, Class I and III heat shock genes as well as members of the SOS regulon were activated. The neurotoxin gene botA and genes encoding the neurotoxin-associated proteins were suppressed throughout the study. Prolonged heat stress led to suppression of the sporulation machinery whereas genes related to chemotaxis and motility were activated. Induced expression of a large proportion of prophage genes was detected, suggesting an important role of acquired genes in the stress resistance of C. botulinum. Finally, changes in the expression of a large number of genes related to carbohydrate and amino acid metabolism indicated remodeling of the cellular metabolism. PMID:28464023

Heat shock and prolonged heat stress attenuate neurotoxin and sporulation gene expression in group I Clostridium botulinum strain ATCC 3502.

PubMed

Selby, Katja; Mascher, Gerald; Somervuo, Panu; Lindström, Miia; Korkeala, Hannu

2017-01-01

Foodborne pathogenic bacteria are exposed to a number of environmental stresses during food processing, storage, and preparation, and in the human body. In order to improve the safety of food, the understanding of molecular stress response mechanisms foodborne pathogens employ is essential. Many response mechanisms that are activated during heat shock may cross-protect bacteria against other environmental stresses. To better understand the molecular mechanisms Clostridium botulinum, the causative agent of botulism, utilizes during acute heat stress and during adaptation to stressfully high temperature, the C. botulinum Group I strain ATCC 3502 was grown in continuous culture at 39°C and exposed to heat shock at 45°C, followed by prolonged heat stress at 45°C to allow adaptation of the culture to the high temperature. Growth in continuous culture was performed to exclude secondary growth phase effects or other environmental impacts on bacterial gene transcription. Changes in global gene expression profiles were studied using DNA microarray hybridization. During acute heat stress, Class I and III heat shock genes as well as members of the SOS regulon were activated. The neurotoxin gene botA and genes encoding the neurotoxin-associated proteins were suppressed throughout the study. Prolonged heat stress led to suppression of the sporulation machinery whereas genes related to chemotaxis and motility were activated. Induced expression of a large proportion of prophage genes was detected, suggesting an important role of acquired genes in the stress resistance of C. botulinum. Finally, changes in the expression of a large number of genes related to carbohydrate and amino acid metabolism indicated remodeling of the cellular metabolism.

Identification of differentially expressed genes from Trichoderma harzianum during growth on cell wall of Fusarium solani as a tool for biotechnological application

PubMed Central

2013-01-01

Background The species of T. harzianum are well known for their biocontrol activity against many plant pathogens. However, there is a lack of studies concerning its use as a biological control agent against F. solani, a pathogen involved in several crop diseases. In this study, we have used subtractive library hybridization (SSH) and quantitative real-time PCR (RT-qPCR) techniques in order to explore changes in T. harzianum genes expression during growth on cell wall of F. solani (FSCW) or glucose. RT-qPCR was also used to examine the regulation of 18 genes, potentially involved in biocontrol, during confrontation between T. harzianum and F. solani. Results Data obtained from two subtractive libraries were compared after annotation using the Blast2GO suite. A total of 417 and 78 readable EST sequence were annotated in the FSCW and glucose libraries, respectively. Functional annotation of these genes identified diverse biological processes and molecular functions required during T. harzianum growth on FSCW or glucose. We identified various genes of biotechnological value encoding to proteins which function such as transporters, hydrolytic activity, adherence, appressorium development and pathogenesis. Fifteen genes were up-regulated and sixteen were down-regulated at least at one-time point during growth of T. harzianum in FSCW. During the confrontation assay most of the genes were up-regulated, mainly after contact, when the interaction has been established. Conclusions This study demonstrates that T. harzianum expressed different genes when grown on FSCW compared to glucose. It provides insights into the mechanisms of gene expression involved in mycoparasitism of T. harzianum against F. solani. The identification and evaluation of these genes may contribute to the development of an efficient biological control agent. PMID:23497274

Identification of differentially expressed genes from Trichoderma harzianum during growth on cell wall of Fusarium solani as a tool for biotechnological application.

PubMed

Vieira, Pabline Marinho; Coelho, Alexandre Siqueira Guedes; Steindorff, Andrei Stecca; de Siqueira, Saulo José Linhares; Silva, Roberto do Nascimento; Ulhoa, Cirano José

2013-03-15

The species of T. harzianum are well known for their biocontrol activity against many plant pathogens. However, there is a lack of studies concerning its use as a biological control agent against F. solani, a pathogen involved in several crop diseases. In this study, we have used subtractive library hybridization (SSH) and quantitative real-time PCR (RT-qPCR) techniques in order to explore changes in T. harzianum genes expression during growth on cell wall of F. solani (FSCW) or glucose. RT-qPCR was also used to examine the regulation of 18 genes, potentially involved in biocontrol, during confrontation between T. harzianum and F. solani. Data obtained from two subtractive libraries were compared after annotation using the Blast2GO suite. A total of 417 and 78 readable EST sequence were annotated in the FSCW and glucose libraries, respectively. Functional annotation of these genes identified diverse biological processes and molecular functions required during T. harzianum growth on FSCW or glucose. We identified various genes of biotechnological value encoding to proteins which function such as transporters, hydrolytic activity, adherence, appressorium development and pathogenesis. Fifteen genes were up-regulated and sixteen were down-regulated at least at one-time point during growth of T. harzianum in FSCW. During the confrontation assay most of the genes were up-regulated, mainly after contact, when the interaction has been established. This study demonstrates that T. harzianum expressed different genes when grown on FSCW compared to glucose. It provides insights into the mechanisms of gene expression involved in mycoparasitism of T. harzianum against F. solani. The identification and evaluation of these genes may contribute to the development of an efficient biological control agent.

The virulence factor ychO has a pleiotropic action in an Avian Pathogenic Escherichia coli (APEC) strain.

PubMed

Pilatti, Livia; Boldrin de Paiva, Jacqueline; Rojas, Thaís Cabrera Galvão; Leite, Janaína Luisa; Conceição, Rogério Arcuri; Nakazato, Gerson; Dias da Silveira, Wanderley

2016-03-10

Avian pathogenic Escherichia coli strains cause extraintestinal diseases in birds, leading to substantial economic losses to the poultry industry worldwide. Bacteria that invade cells can overcome the host humoral immune response, resulting in a higher pathogenicity potential. Invasins are members of a large family of outer membrane proteins that allow pathogen invasion into host cells by interacting with specific receptors on the cell surface. An in silico analysis of the genome of a septicemic APEC strain (SEPT362) demonstrated the presence of a putative invasin homologous to the ychO gene from E. coli str. K-12 substr. MG1655. In vitro and in vivo assays comparing a mutant strain carrying a null mutation of this gene, a complemented strain, and its counterpart wild-type strain showed that ychO plays a role in the pathogenicity of APEC strain SEPT362. In vitro assays demonstrated that the mutant strain exhibited significant decreases in bacterial adhesiveness and invasiveness in chicken cells and biofilm formation. In vivo assay indicated a decrease in pathogenicity of the mutant strain. Moreover, transcriptome analysis demonstrated that the ychO deletion affected the expression of 426 genes. Among the altered genes, 93.66% were downregulated in the mutant, including membrane proteins and metabolism genes. The results led us to propose that gene ychO contributes to the pathogenicity of APEC strain SEPT362 influencing, in a pleiotropic manner, many biological characteristics, such as adhesion and invasion of in vitro cultured cells, biofilm formation and motility, which could be due to the possible membrane location of this protein. All of these results suggest that the absence of gene ychO would influence the virulence of the APEC strain herein studied.

Identification of in vivo regulators of the Vibrio cholerae xds gene using a high-throughput genetic selection.

PubMed

McDonough, Emilykate; Lazinski, David W; Camilli, Andrew

2014-04-01

Vibrio cholerae, the causative agent of cholera, remains a threat to public health in areas with inadequate sanitation. As a waterborne pathogen, V. cholerae moves between two dissimilar environments, aquatic reservoirs and the intestinal tract of humans. Accordingly, this pathogen undergoes adaptive shifts in gene expression throughout the different stages of its lifecycle. One particular gene, xds, encodes a secreted exonuclease that was previously identified as being induced during infection. Here we sought to identify regulators responsible for the in vivo-specific induction of xds. A transcriptional fusion of xds to two consecutive antibiotic resistance genes was used to select transposon mutants that had inserted within or adjacent to regulatory genes and thereby caused increased expression of the xds fusion under non-inducing conditions. Large pools of selected insertion sites were sequenced in a high throughput manner using Tn-seq to identify potential mechanisms of xds regulation. Our selection identified the two-component system PhoB/R as the dominant activator of xds expression. In vitro validation confirmed that PhoB, a protein which is only active during phosphate limitation, was responsible for xds activation. Using xds expression as a biosensor of the extracellular phosphate level, we observed that the mouse small intestine is a phosphate-limited environment. © 2014 John Wiley & Sons Ltd.

Similarities and differences in global gene expression profiles between herbicide- and pathogen-induced PSII inhibition

USDA-ARS?s Scientific Manuscript database

Plant pathogens, and photosynthesis inhibiting herbicides, can both damage photosystem II (PSII), causing it to be highly sensitive to damage by light energy, and to release high levels of reactive oxygen species (ROS). This photoinhibition of PSII could possibly be the source of the strong oxidativ...

Evolution of a zoonotic pathogen: investigating prophage diversity in enterohaemorrhagic Escherichia coli O157 by long-read sequencing

USDA-ARS?s Scientific Manuscript database

Enterohaemorrhagic E. coli 0157 is a zoonotic pathogen for which colonisation of cattle and virulence in humans is associated with the expression of multiple horizontally acquired genes, the majority present in active or cryptic prophages. Our understanding of the evolution and phylogeny of E. coli ...

AutA and AutR, Two Novel Global Transcriptional Regulators, Facilitate Avian Pathogenic Escherichia coli Infection.

PubMed

Zhuge, Xiangkai; Tang, Fang; Zhu, Hongfei; Mao, Xiang; Wang, Shaohui; Wu, Zongfu; Lu, Chengping; Dai, Jianjun; Fan, Hongjie

2016-04-26

Bacteria can change its lifestyle during inhabiting in host niches where they survive and replicate by rapidly altering gene expression pattern to accommodate the new environment. In this study, two novel regulators in avian pathogenic Escherichia coli (APEC) were identified and designated as AutA and AutR. RT-PCR and β-galactosidase assay results showed that AutA and AutR co-regulated the expression of adhesin UpaB in APEC strain DE205B. Electrophoretic mobility shift assay showed that AutA and AutR could directly bind the upaB promoter DNA. In vitro transcription assay indicated that AutA could activate the upaB transcription, while AutR inhibited the upaB transcription due to directly suppressing the activating effect of AutA on UpaB expression. Transcriptome analysis showed that AutA and AutR coherently affected the expression of hundreds of genes. Our study confirmed that AutA and AutR co-regulated the expression of DE205B K1 capsule and acid resistance systems in E. coli acid fitness island (AFI). Moreover, phenotypic heterogeneity in expression of K1 capsule and acid resistance systems in AFI during host-pathogen interaction was associated with the regulation of AutA and AutR. Collectively speaking, our studies presented that AutA and AutR are involved in APEC adaptive lifestyle change to facilitate its infection.

Distinctive Expansion of Potential Virulence Genes in the Genome of the Oomycete Fish Pathogen Saprolegnia parasitica

PubMed Central

Belmonte, Rodrigo; Löbach, Lars; Christie, James; van den Ackerveken, Guido; Bottin, Arnaud; Bulone, Vincent; Díaz-Moreno, Sara M.; Dumas, Bernard; Fan, Lin; Gaulin, Elodie; Govers, Francine; Grenville-Briggs, Laura J.; Horner, Neil R.; Levin, Joshua Z.; Mammella, Marco; Meijer, Harold J. G.; Morris, Paul; Nusbaum, Chad; Oome, Stan; Phillips, Andrew J.; van Rooyen, David; Rzeszutek, Elzbieta; Saraiva, Marcia; Secombes, Chris J.; Seidl, Michael F.; Snel, Berend; Stassen, Joost H. M.; Sykes, Sean; Tripathy, Sucheta; van den Berg, Herbert; Vega-Arreguin, Julio C.; Wawra, Stephan; Young, Sarah K.; Zeng, Qiandong; Dieguez-Uribeondo, Javier; Russ, Carsten; Tyler, Brett M.; van West, Pieter

2013-01-01

Oomycetes in the class Saprolegniomycetidae of the Eukaryotic kingdom Stramenopila have evolved as severe pathogens of amphibians, crustaceans, fish and insects, resulting in major losses in aquaculture and damage to aquatic ecosystems. We have sequenced the 63 Mb genome of the fresh water fish pathogen, Saprolegnia parasitica. Approximately 1/3 of the assembled genome exhibits loss of heterozygosity, indicating an efficient mechanism for revealing new variation. Comparison of S. parasitica with plant pathogenic oomycetes suggests that during evolution the host cellular environment has driven distinct patterns of gene expansion and loss in the genomes of plant and animal pathogens. S. parasitica possesses one of the largest repertoires of proteases (270) among eukaryotes that are deployed in waves at different points during infection as determined from RNA-Seq data. In contrast, despite being capable of living saprotrophically, parasitism has led to loss of inorganic nitrogen and sulfur assimilation pathways, strikingly similar to losses in obligate plant pathogenic oomycetes and fungi. The large gene families that are hallmarks of plant pathogenic oomycetes such as Phytophthora appear to be lacking in S. parasitica, including those encoding RXLR effectors, Crinkler's, and Necrosis Inducing-Like Proteins (NLP). S. parasitica also has a very large kinome of 543 kinases, 10% of which is induced upon infection. Moreover, S. parasitica encodes several genes typical of animals or animal-pathogens and lacking from other oomycetes, including disintegrins and galactose-binding lectins, whose expression and evolutionary origins implicate horizontal gene transfer in the evolution of animal pathogenesis in S. parasitica. PMID:23785293

Antigenic variation in malaria: in situ switching, relaxed and mutually exclusive transcription of var genes during intra-erythrocytic development in Plasmodium falciparum.

PubMed Central

Scherf, A; Hernandez-Rivas, R; Buffet, P; Bottius, E; Benatar, C; Pouvelle, B; Gysin, J; Lanzer, M

1998-01-01

Members of the Plasmodium falciparum var gene family encode clonally variant adhesins, which play an important role in the pathogenicity of tropical malaria. Here we employ a selective panning protocol to generate isogenic P.falciparum populations with defined adhesive phenotypes for CD36, ICAM-1 and CSA, expressing single and distinct var gene variants. This technique has established the framework for examining var gene expression, its regulation and switching. It was found that var gene switching occurs in situ. Ubiquitous transcription of all var gene variants appears to occur in early ring stages. However, var gene expression is tightly regulated in trophozoites and is exerted through a silencing mechanism. Transcriptional control is mutually exclusive in parasites that express defined adhesive phenotypes. In situ var gene switching is apparently mediated at the level of transcriptional initiation, as demonstrated by nuclear run-on analyses. Our results suggest that an epigenetic mechanism(s) is involved in var gene regulation. PMID:9736619

The Fusarium crown rot pathogen Fusarium pseudograminearum triggers a suite of transcriptional and metabolic changes in bread wheat (Triticum aestivum L.)

PubMed Central

Carere, Jason; Fitzgerald, Timothy L.; Stiller, Jiri; Covarelli, Lorenzo; Xu, Qian; Gubler, Frank; Colgrave, Michelle L.; Gardiner, Donald M.; Manners, John M.; Henry, Robert J.; Kazan, Kemal

2017-01-01

Abstract Background and Aims Fusarium crown rot caused by the fungal pathogen Fusarium pseudograminearum is a disease of wheat and barley, bearing significant economic cost. Efforts to develop effective resistance to this disease have been hampered by the quantitative nature of resistance and a lack of understanding of the factors associated with resistance and susceptibility. Here, we aimed to dissect transcriptional responses triggered in wheat by F. pseudograminearum infection. Methods We used an RNA-seq approach to analyse host responses during a compatible interaction and identified >2700 wheat genes differentially regulated after inoculation with F. pseudograminearum. The production of a few key metabolites and plant hormones in the host during the interaction was also analysed. Key Results Analysis of gene ontology enrichment showed that a disproportionate number of genes involved in primary and secondary metabolism, signalling and transport were differentially expressed in infected seedlings. A number of genes encoding pathogen-responsive uridine-diphosphate glycosyltransferases (UGTs) potentially involved in detoxification of the Fusarium mycotoxin deoxynivalenol (DON) were differentially expressed. Using a F. pseudograminearum DON-non-producing mutant, DON was shown to play an important role in virulence during Fusarium crown rot. An over-representation of genes involved in the phenylalanine, tryptophan and tyrosine biosynthesis pathways was observed. This was confirmed through metabolite analyses that demonstrated tryptamine and serotonin levels are induced after F. pseudograminearum inoculation. Conclusions Overall, the observed host response in bread wheat to F. pseudograminearum during early infection exhibited enrichment of processes related to pathogen perception, defence signalling, transport and metabolism and deployment of chemical and enzymatic defences. Additional functional analyses of candidate genes should reveal their roles in disease resistance or susceptibility. Better understanding of host responses contributing to resistance and/or susceptibility will aid the development of future disease improvement strategies against this important plant pathogen. PMID:27941094

The Fusarium crown rot pathogen Fusarium pseudograminearum triggers a suite of transcriptional and metabolic changes in bread wheat (Triticum aestivum L.).

PubMed

Powell, Jonathan J; Carere, Jason; Fitzgerald, Timothy L; Stiller, Jiri; Covarelli, Lorenzo; Xu, Qian; Gubler, Frank; Colgrave, Michelle L; Gardiner, Donald M; Manners, John M; Henry, Robert J; Kazan, Kemal

2017-03-01

Fusarium crown rot caused by the fungal pathogen Fusarium pseudograminearum is a disease of wheat and barley, bearing significant economic cost. Efforts to develop effective resistance to this disease have been hampered by the quantitative nature of resistance and a lack of understanding of the factors associated with resistance and susceptibility. Here, we aimed to dissect transcriptional responses triggered in wheat by F. pseudograminearum infection. We used an RNA-seq approach to analyse host responses during a compatible interaction and identified >2700 wheat genes differentially regulated after inoculation with F. pseudograminearum . The production of a few key metabolites and plant hormones in the host during the interaction was also analysed. Analysis of gene ontology enrichment showed that a disproportionate number of genes involved in primary and secondary metabolism, signalling and transport were differentially expressed in infected seedlings. A number of genes encoding pathogen-responsive uridine-diphosphate glycosyltransferases (UGTs) potentially involved in detoxification of the Fusarium mycotoxin deoxynivalenol (DON) were differentially expressed. Using a F. pseudograminearum DON-non-producing mutant, DON was shown to play an important role in virulence during Fusarium crown rot. An over-representation of genes involved in the phenylalanine, tryptophan and tyrosine biosynthesis pathways was observed. This was confirmed through metabolite analyses that demonstrated tryptamine and serotonin levels are induced after F. pseudograminearum inoculation. Overall, the observed host response in bread wheat to F. pseudograminearum during early infection exhibited enrichment of processes related to pathogen perception, defence signalling, transport and metabolism and deployment of chemical and enzymatic defences. Additional functional analyses of candidate genes should reveal their roles in disease resistance or susceptibility. Better understanding of host responses contributing to resistance and/or susceptibility will aid the development of future disease improvement strategies against this important plant pathogen. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company.

Basic leucine zipper domain transcription factors: the vanguards in plant immunity.

PubMed

Noman, Ali; Liu, Zhiqin; Aqeel, Muhammad; Zainab, Madiha; Khan, Muhammad Ifnan; Hussain, Ansar; Ashraf, Muhammad Furqan; Li, Xia; Weng, Yahong; He, Shuilin

2017-12-01

Regulation of spatio-temporal expression patterns of stress tolerance associated plant genes is an essential component of the stress responses. Eukaryotes assign a large amount of their genome to transcription with multiple transcription factors (TFs). Often, these transcription factors fit into outsized gene groups which, in several cases, exclusively belong to plants. Basic leucine zipper domain (bZIP) transcription factors regulate vital processes in plants and animals. In plants, bZIPs are implicated in numerous fundamental processes like seed development, energy balance, and responses to abiotic or biotic stresses. Systematic analysis of the information obtained over the last two decades disclosed a constitutive role of bZIPs against biotic stress. bZIP TFs are vital players in plant innate immunity due to their ability to regulate genes associated with PAMP-triggered immunity, effector-triggered immunity, and hormonal signaling networks. Expression analysis of studied bZIP genes suggests that exploration and functional characterization of novel bZIP TFs in planta is helpful in improving crop resistance against pathogens and environmental stresses. Our review focuses on major advancements in bZIP TFs and plant responses against different pathogens. The integration of genomics information with the functional studies provides new insights into the regulation of plant defense mechanisms and engineering crops with improved resistance to invading pathogens. Conclusively, succinct functions of bZIPs as positive or negative regulator mediate resistance to the plant pathogens and lay a foundation for understanding associated genes and TFs regulating different pathways. Moreover, bZIP TFs may offer a comprehensive transgenic gizmo for engineering disease resistance in plant breeding programs.

A Virulence Essential CRN Effector of Phytophthora capsici Suppresses Host Defense and Induces Cell Death in Plant Nucleus.

PubMed

Mafurah, Joseph Juma; Ma, Huifei; Zhang, Meixiang; Xu, Jing; He, Feng; Ye, Tingyue; Shen, Danyu; Chen, Yanyu; Rajput, Nasir Ahmed; Dou, Daolong

2015-01-01

Phytophthora capsici is a soil-borne plant pathogen with a wide range of hosts. The pathogen secretes a large array of effectors during infection of host plants, including Crinkler (CRN) effectors. However, it remains largely unknown on the roles of these effectors in virulence especially in P. capsici. In this study, we identified a cell death-inducing CRN effector PcCRN4 using agroinfiltration approach. Transient expression of PcCRN4 gene induced cell death in N. benthamiana, N. tabacum and Solanum lycopersicum. Overexpression of the gene in N. benthamiana enhanced susceptibility to P. capsici. Subcellular localization results showed that PcCRN4 localized to the plant nucleus, and the localization was required for both of its cell death-inducing activity and virulent function. Silencing PcCRN4 gene in P. capsici significantly reduced pathogen virulence. The expression of the pathogenesis-related gene PR1b in N. benthamiana was significantly induced when plants were inoculated with PcCRN4-silenced P. capsici transformant compared to the wilt-type. Callose deposits were also abundant at sites inoculated with PcCRN4-silenced transformant, indicating that silencing of PcCRN4 in P. capsici reduced the ability of the pathogen to suppress plant defenses. Transcriptions of cell death-related genes were affected when PcCRN4-silenced line were inoculated on Arabidopsis thaliana, suggesting that PcCRN4 may induce cell death by manipulating cell death-related genes. Overall, our results demonstrate that PcCRN4 is a virulence essential effector and it needs target to the plant nucleus to suppress plant immune responses.

Transcriptomic insight into pathogenicity-associated factors of Conidiobolus obscurus, an obligate aphid-pathogenic fungus belonging to Entomopthoromycota.

PubMed

Wang, Jianghong; Zhou, Xiang; Guo, Kai; Zhang, Xinqi; Lin, Haiping; Montalva, Cristian

2018-01-16

Conidiobolus obscurus is a widespread fungal entomopathogen with aphid biocontrol potential. This study focused on a de novo transcriptomic analysis of C. obscurus. A number of pathogenicity-associated factors were annotated for the first time from the assembled 17 231 fungal unigenes, including those encoding subtilisin-like proteolytic enzymes (Pr1s), trypsin-like proteases, metalloproteases, carboxypeptidases and endochitinases. Many of these genes were transcriptionally up-regulated by at least twofold in mycotized cadavers compared with the in vitro fungal cultures. The resultant transcriptomic database was validated by the transcript levels of three selected pathogenicity-related genes quantified from different in vivo and in vitro material in real-time quantitative polymerase chain reaction (PCR). The involvement of multiple Pr1 proteases in the first stage of fungal infection was also suggested. Interestingly, a unique cytolytic (Cyt)-like δ-endotoxin gene was highly expressed in both mycotized cadavers and fungal cultures, and was more or less distinct from its homologues in bacteria and other fungi. Our findings provide the first global insight into various pathogenicity-related genes in this obligate aphid pathogen and may help to develop novel biocontrol strategy against aphid pests. © 2018 Society of Chemical Industry. © 2018 Society of Chemical Industry.

STING-Dependent 2'-5' Oligoadenylate Synthetase-Like Production Is Required for Intracellular Mycobacterium leprae Survival.

PubMed

de Toledo-Pinto, Thiago Gomes; Ferreira, Anna Beatriz Robottom; Ribeiro-Alves, Marcelo; Rodrigues, Luciana Silva; Batista-Silva, Leonardo Ribeiro; Silva, Bruno Jorge de Andrade; Lemes, Robertha Mariana Rodrigues; Martinez, Alejandra Nóbrega; Sandoval, Felipe Galvan; Alvarado-Arnez, Lucia Elena; Rosa, Patrícia Sammarco; Shannon, Edward Joseph; Pessolani, Maria Cristina Vidal; Pinheiro, Roberta Olmo; Antunes, Sérgio Luís Gomes; Sarno, Euzenir Nunes; Lara, Flávio Alves; Williams, Diana Lynn; Ozório Moraes, Milton

2016-07-15

Cytosolic detection of nucleic acids elicits a type I interferon (IFN) response and plays a critical role in host defense against intracellular pathogens. Herein, a global gene expression profile of Mycobacterium leprae-infected primary human Schwann cells identified the genes differentially expressed in the type I IFN pathway. Among them, the gene encoding 2'-5' oligoadenylate synthetase-like (OASL) underwent the greatest upregulation and was also shown to be upregulated in M. leprae-infected human macrophage cell lineages, primary monocytes, and skin lesion specimens from patients with a disseminated form of leprosy. OASL knock down was associated with decreased viability of M. leprae that was concomitant with upregulation of either antimicrobial peptide expression or autophagy levels. Downregulation of MCP-1/CCL2 release was also observed during OASL knock down. M. leprae-mediated OASL expression was dependent on cytosolic DNA sensing mediated by stimulator of IFN genes signaling. The addition of M. leprae DNA enhanced nonpathogenic Mycobacterium bovis bacillus Calmette-Guerin intracellular survival, downregulated antimicrobial peptide expression, and increased MCP-1/CCL2 secretion. Thus, our data uncover a promycobacterial role for OASL during M. leprae infection that directs the host immune response toward a niche that permits survival of the pathogen. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Cross-talk of the biotrophic pathogen Claviceps purpurea and its host Secale cereale.

PubMed

Oeser, Birgitt; Kind, Sabine; Schurack, Selma; Schmutzer, Thomas; Tudzynski, Paul; Hinsch, Janine

2017-04-04

The economically important Ergot fungus Claviceps purpurea is an interesting biotrophic model system because of its strict organ specificity (grass ovaries) and the lack of any detectable plant defense reactions. Though several virulence factors were identified, the exact infection mechanisms are unknown, e.g. how the fungus masks its attack and if the host detects the infection at all. We present a first dual transcriptome analysis using an RNA-Seq approach. We studied both, fungal and plant gene expression in young ovaries infected by the wild-type and two virulence-attenuated mutants. We can show that the plant recognizes the fungus, since defense related genes are upregulated, especially several phytohormone genes. We present a survey of in planta expressed fungal genes, among them several confirmed virulence genes. Interestingly, the set of most highly expressed genes includes a high proportion of genes encoding putative effectors, small secreted proteins which might be involved in masking the fungal attack or interfering with host defense reactions. As known from several other phytopathogens, the C. purpurea genome contains more than 400 of such genes, many of them clustered and probably highly redundant. Since the lack of effective defense reactions in spite of recognition of the fungus could very well be achieved by effectors, we started a functional analysis of some of the most highly expressed candidates. However, the redundancy of the system made the identification of a drastic effect of a single gene most unlikely. We can show that at least one candidate accumulates in the plant apoplast. Deletion of some candidates led to a reduced virulence of C. purpurea on rye, indicating a role of the respective proteins during the infection process. We show for the first time that- despite the absence of effective plant defense reactions- the biotrophic pathogen C. purpurea is detected by its host. This points to a role of effectors in modulation of the effective plant response. Indeed, several putative effector genes are among the highest expressed genes in planta.

Signal signature and transcriptome changes of Arabidopsis during pathogen and insect attack.

PubMed

De Vos, Martin; Van Oosten, Vivian R; Van Poecke, Remco M P; Van Pelt, Johan A; Pozo, Maria J; Mueller, Martin J; Buchala, Antony J; Métraux, Jean-Pierre; Van Loon, L C; Dicke, Marcel; Pieterse, Corné M J

2005-09-01

Plant defenses against pathogens and insects are regulated differentially by cross-communicating signaling pathways in which salicylic acid (SA), jasmonic acid (JA), and ethylene (ET) play key roles. To understand how plants integrate pathogen- and insect-induced signals into specific defense responses, we monitored the dynamics of SA, JA, and ET signaling in Arabidopsis after attack by a set of microbial pathogens and herbivorous insects with different modes of attack. Arabidopsis plants were exposed to a pathogenic leaf bacterium (Pseudomonas syringae pv. tomato), a pathogenic leaf fungus (Alternaria brassicicola), tissue-chewing caterpillars (Pieris rapae), cell-content-feeding thrips (Frankliniella occidentalis), or phloem-feeding aphids (Myzus persicae). Monitoring the signal signature in each plant-attacker combination showed that the kinetics of SA, JA, and ET production varies greatly in both quantity and timing. Analysis of global gene expression profiles demonstrated that the signal signature characteristic of each Arabidopsis-attacker combination is orchestrated into a surprisingly complex set of transcriptional alterations in which, in all cases, stress-related genes are overrepresented. Comparison of the transcript profiles revealed that consistent changes induced by pathogens and insects with very different modes of attack can show considerable overlap. Of all consistent changes induced by A. brassicicola, Pieris rapae, and E occidentalis, more than 50% also were induced consistently by P. syringae. Notably, although these four attackers all stimulated JA biosynthesis, the majority of the changes in JA-responsive gene expression were attacker specific. All together, our study shows that SA, JA, and ET play a primary role in the orchestration of the plant's defense response, but other regulatory mechanisms, such as pathway cross-talk or additional attacker-induced signals, eventually shape the highly complex attacker-specific defense response.

Polygalacturonase gene pgxB in Aspergillus niger is a virulence factor in apple fruit

PubMed Central

Yang, Ying; Yang, Feng; Li, Yan-Hong; Liu, He-Ping; Chen, Xiao-Yan

2017-01-01

Aspergillus niger, a saprophytic fungus, is widely distributed in soil, air and cereals, and can cause postharvest diseases in fruit. Polygalacturonase (PG) is one of the main enzymes in fungal pathogens to degrade plant cell wall. To evaluate whether the deletion of an exo-polygalacturonase gene pgxB would influence fungal pathogenicity to fruit, pgxB gene was deleted in Aspergillus niger MA 70.15 (wild type) via homologous recombination. The ΔpgxB mutant showed similar growth behavior compared with the wild type. Pectin medium induced significant higher expression of all pectinase genes in both wild type and ΔpgxB in comparison to potato dextrose agar medium. However, the ΔpgxB mutant was less virulent on apple fruits as the necrosis diameter caused by ΔpgxB mutant was significantly smaller than that of wild type. Results of quantitive-PCR showed that, in the process of infection in apple fruit, gene expressions of polygalacturonase genes pgaI, pgaII, pgaA, pgaC, pgaD and pgaE were enhanced in ΔpgxB mutant in comparison to wild type. These results prove that, despite the increased gene expression of other polygalacturonase genes in ΔpgxB mutant, the lack of pgxB gene significantly reduced the virulence of A. niger on apple fruit, suggesting that pgxB plays an important role in the infection process on the apple fruit. PMID:28257463

The Roles of Heat Shock Proteins 70 and 90 in Exopalaemon carinicauda After WSSV and Vibrio anguillarum Challenges

NASA Astrophysics Data System (ADS)

Li, Jitao; Li, Jian; Duan, Yafei; Chen, Ping; Liu, Ping

2018-04-01

Heat shock proteins (HSPs), such as HSP70 and HSP90, are a suite of highly conserved proteins produced in all cellular organisms when they are exposed to stresses. In aquatic animals, they have been proved to play important roles in response to environmental pollutants and particularly in the non-specific immune responses to pathogen infections. In the present study, the expression profiles of HSP70 and HSP90 genes in hemocytes and hepatopancreas from the ridgetail white prawn Exopalaemon carinicauda infected with WSSV and Vibrio anguillarum were detected using reverse transcription polymerase chain reaction (RT-PCR). After WSSV challenge, the expression level of HSP 70 gene transcripts in the hemocytes and hepatopancreas increased to peak level at 6 h and 48 h, respectively. HSP90 gene transcripts in hemocytes and hepatopancreas were up-regulated significantly at 12 h and 6 h, respectively. During V. anguillarum challenge, the mRNA content of HSP70 gene in hemocytes and hepatopancreas increased significantly at 12 h and 6 h post-infection, respectively. The expression level of HSP90 gene both in hemocytes and hepatopancreas were up-regulated in the first 3 h. The expression patterns of HSP70 and HSP90 genes in hemocytes and hepatopancreas showed temporal and spatial differences after challenged with WSSV and V. anguillarum. The results suggested that HSPs might be involved in immune responses to pathogens challenge in E. carinicauda.

The roles of heat shock proteins 70 and 90 in Exopalaemon carinicauda after WSSV and Vibrio anguillarum challenges

NASA Astrophysics Data System (ADS)

Li, Jitao; Li, Jian; Duan, Yafei; Chen, Ping; Liu, Ping

2017-12-01

Heat shock proteins (HSPs), such as HSP70 and HSP90, are a suite of highly conserved proteins produced in all cellular organisms when they are exposed to stresses. In aquatic animals, they have been proved to play important roles in response to environmental pollutants and particularly in the non-specific immune responses to pathogen infections. In the present study, the expression profiles of HSP70 and HSP90 genes in hemocytes and hepatopancreas from the ridgetail white prawn Exopalaemon carinicauda infected with WSSV and Vibrio anguillarum were detected using reverse transcription polymerase chain reaction (RT-PCR). After WSSV challenge, the expression level of HSP 70 gene transcripts in the hemocytes and hepatopancreas increased to peak level at 6 h and 48 h, respectively. HSP90 gene transcripts in hemocytes and hepatopancreas were up-regulated significantly at 12 h and 6 h, respectively. During V. anguillarum challenge, the mRNA content of HSP70 gene in hemocytes and hepatopancreas increased significantly at 12 h and 6 h post-infection, respectively. The expression level of HSP90 gene both in hemocytes and hepatopancreas were up-regulated in the first 3 h. The expression patterns of HSP70 and HSP90 genes in hemocytes and hepatopancreas showed temporal and spatial differences after challenged with WSSV and V. anguillarum. The results suggested that HSPs might be involved in immune responses to pathogens challenge in E. carinicauda.

Genome sequencing of the lizard parasite Leishmania tarentolae reveals loss of genes associated to the intracellular stage of human pathogenic species

PubMed Central

Raymond, Frédéric; Boisvert, Sébastien; Roy, Gaétan; Ritt, Jean-François; Légaré, Danielle; Isnard, Amandine; Stanke, Mario; Olivier, Martin; Tremblay, Michel J.; Papadopoulou, Barbara; Ouellette, Marc; Corbeil, Jacques

2012-01-01

The Leishmania tarentolae Parrot-TarII strain genome sequence was resolved to an average 16-fold mean coverage by next-generation DNA sequencing technologies. This is the first non-pathogenic to humans kinetoplastid protozoan genome to be described thus providing an opportunity for comparison with the completed genomes of pathogenic Leishmania species. A high synteny was observed between all sequenced Leishmania species. A limited number of chromosomal regions diverged between L. tarentolae and L. infantum, while remaining syntenic to L. major. Globally, >90% of the L. tarentolae gene content was shared with the other Leishmania species. We identified 95 predicted coding sequences unique to L. tarentolae and 250 genes that were absent from L. tarentolae. Interestingly, many of the latter genes were expressed in the intracellular amastigote stage of pathogenic species. In addition, genes coding for products involved in antioxidant defence or participating in vesicular-mediated protein transport were underrepresented in L. tarentolae. In contrast to other Leishmania genomes, two gene families were expanded in L. tarentolae, namely the zinc metallo-peptidase surface glycoprotein GP63 and the promastigote surface antigen PSA31C. Overall, L. tarentolae's gene content appears better adapted to the promastigote insect stage rather than the amastigote mammalian stage. PMID:21998295

The in planta transcriptome of Ralstonia solanacearum: conserved physiological and virulence strategies during bacterial wilt of tomato.

PubMed

Jacobs, Jonathan M; Babujee, Lavanya; Meng, Fanhong; Milling, Annett; Allen, Caitilyn

2012-01-01

Plant xylem fluid is considered a nutrient-poor environment, but the bacterial wilt pathogen Ralstonia solanacearum is well adapted to it, growing to 10(8) to 10(9) CFU/g tomato stem. To better understand how R. solanacearum succeeds in this habitat, we analyzed the transcriptomes of two phylogenetically distinct R. solanacearum strains that both wilt tomato, strains UW551 (phylotype II) and GMI1000 (phylotype I). We profiled bacterial gene expression at ~6 × 10(8) CFU/ml in culture or in plant xylem during early tomato bacterial wilt pathogenesis. Despite phylogenetic differences, these two strains expressed their 3,477 common orthologous genes in generally similar patterns, with about 12% of their transcriptomes significantly altered in planta versus in rich medium. Several primary metabolic pathways were highly expressed during pathogenesis. These pathways included sucrose uptake and catabolism, and components of these pathways were encoded by genes in the scrABY cluster. A UW551 scrA mutant was significantly reduced in virulence on resistant and susceptible tomato as well as on potato and the epidemiologically important weed host Solanum dulcamara. Functional scrA contributed to pathogen competitive fitness during colonization of tomato xylem, which contained ~300 µM sucrose. scrA expression was induced by sucrose, but to a much greater degree by growth in planta. Unexpectedly, 45% of the genes directly regulated by HrpB, the transcriptional activator of the type 3 secretion system (T3SS), were upregulated in planta at high cell densities. This result modifies a regulatory model based on bacterial behavior in culture, where this key virulence factor is repressed at high cell densities. The active transcription of these genes in wilting plants suggests that T3SS has a biological role throughout the disease cycle. IMPORTANCE Ralstonia solanacearum is a widespread plant pathogen that causes bacterial wilt disease. It inflicts serious crop losses on tropical farmers, with major economic and human consequences. It is also a model for the many destructive microbes that colonize the water-conducting plant xylem tissue, which is low in nutrients and oxygen. We extracted bacteria from infected tomato plants and globally identified the biological functions that R. solanacearum expresses during plant pathogenesis. This revealed the unexpected presence of sucrose in tomato xylem fluid and the pathogen's dependence on host sucrose for virulence on tomato, potato, and the common weed bittersweet nightshade. Further, R. solanacearum was highly responsive to the plant environment, expressing several metabolic and virulence functions quite differently in the plant than in pure culture. These results reinforce the utility of studying pathogens in interaction with hosts and suggest that selecting for reduced sucrose levels could generate wilt-resistant crops.

GMC oxidoreductase, a highly expressed protein in a potent biocontrol agent Fusarium oxysporum Cong:1-2, is dispensable for biocontrol activity.

PubMed

Kawabe, Masato; Okabe Onokubo, Akiko; Arimoto, Yutaka; Yoshida, Takanobu; Azegami, Koji; Teraoka, Tohru; Arie, Tsutomu

2011-01-01

A spontaneous non-pathogenic variant (Cong:1-2) derived from Fusarium oxysporum f. sp. conglutinans (Cong: 1-1), a causal agent of cabbage yellows, carries biocontrol activity for cabbage yellows. We found a GMC oxidoreductase (ODX1) among the proteins expressed much more in Cong:1-2 than Cong:1-1 by 2D-DIGE comparison. GMC oxidoreductases have been reported to be involved in biocontrol activity of several plant pathogenic fungi. The gene encoding ODX1 in Cong:1-2 was cloned, and targeted disruption of the gene in Cong:1-2 did not affect its biocontrol activity, suggesting that GMC oxidoreductase is dispensable for biocontrol activity in the fungal biocontrol agent.

Tick salivary compounds: their role in modulation of host defences and pathogen transmission

PubMed Central

Kazimírová, Mária; Štibrániová, Iveta

2013-01-01

Ticks require blood meal to complete development and reproduction. Multifunctional tick salivary glands play a pivotal role in tick feeding and transmission of pathogens. Tick salivary molecules injected into the host modulate host defence responses to the benefit of the feeding ticks. To colonize tick organs, tick-borne microorganisms must overcome several barriers, i.e., tick gut membrane, tick immunity, and moulting. Tick-borne pathogens co-evolved with their vectors and hosts and developed molecular adaptations to avoid adverse effects of tick and host defences. Large gaps exist in the knowledge of survival strategies of tick-borne microorganisms and on the molecular mechanisms of tick-host-pathogen interactions. Prior to transmission to a host, the microorganisms penetrate and multiply in tick salivary glands. As soon as the tick is attached to a host, gene expression and production of salivary molecules is upregulated, primarily to facilitate feeding and avoid tick rejection by the host. Pathogens exploit tick salivary molecules for their survival and multiplication in the vector and transmission to and establishment in the hosts. Promotion of pathogen transmission by bioactive molecules in tick saliva was described as saliva-assisted transmission (SAT). SAT candidates comprise compounds with anti-haemostatic, anti-inflammatory and immunomodulatory functions, but the molecular mechanisms by which they mediate pathogen transmission are largely unknown. To date only a few tick salivary molecules associated with specific pathogen transmission have been identified and their functions partially elucidated. Advanced molecular techniques are applied in studying tick-host-pathogen interactions and provide information on expression of vector and pathogen genes during pathogen acquisition, establishment and transmission. Understanding the molecular events on the tick-host-pathogen interface may lead to development of new strategies to control tick-borne diseases. PMID:23971008

Use of Heme Compounds as Iron Sources by Pathogenic Neisseriae Requires the Product of the hemO Gene

PubMed Central

Zhu, Wenming; Hunt, Desiree J.; Richardson, Anthony R.; Stojiljkovic, Igor

2000-01-01

Heme compounds are an important source of iron for neisseriae. We have identified a neisserial gene, hemO, that is essential for heme, hemoglobin (Hb), and haptoglobin-Hb utilization. The hemO gene is located 178 bp upstream of the hmbR Hb receptor gene in Neisseria meningitidis isolates. The product of the hemO gene is homologous to enzymes that degrade heme; 21% of its amino acid residues are identical, and 44% are similar, to those of the human heme oxygenase-1. DNA sequences homologous to hemO were ubiquitous in commensal and pathogenic neisseriae. HemO genetic knockout strains of Neisseria gonorrhoeae and N. meningitidis were unable to use any heme source, while the assimilation of transferrin-iron and iron-citrate complexes was unaffected. A phenotypic characterization of a conditional hemO mutant, constructed by inserting an isopropyl-β-d-thiogalactopyranoside (IPTG)-regulated promoter upstream of the ribosomal binding site of hemO, confirmed the indispensability of the HemO protein in heme utilization. The expression of HemO also protected N. meningitidis cells against heme toxicity. hemO mutants were still able to transport heme into the cell, since both heme and Hb could complement an N. meningitidis hemA hemO double mutant for growth. The expression of the HmbR receptor was reduced significantly by the inactivation of the hemO gene, suggesting that hemO and hmbR are transcriptionally linked. The expression of the unlinked Hb receptor, HpuAB, was not altered. Comparison of the polypeptide patterns of the wild type and the hemO mutant led to detection of six protein spots with an altered expression pattern, suggesting a more general role of HemO in the regulation of gene expression in Neisseriae. PMID:10629191

RNA- and protein-mediated control of Listeria monocytogenes virulence gene expression

PubMed Central

Lebreton, Alice; Cossart, Pascale

2017-01-01

ABSTRACT The model opportunistic pathogen Listeria monocytogenes has been the object of extensive research, aiming at understanding its ability to colonize diverse environmental niches and animal hosts. Bacterial transcriptomes in various conditions reflect this efficient adaptability. We review here our current knowledge of the mechanisms allowing L. monocytogenes to respond to environmental changes and trigger pathogenicity, with a special focus on RNA-mediated control of gene expression. We highlight how these studies have brought novel concepts in prokaryotic gene regulation, such as the ‘excludon’ where the 5′-UTR of a messenger also acts as an antisense regulator of an operon transcribed in opposite orientation, or the notion that riboswitches can regulate non-coding RNAs to integrate complex metabolic stimuli into regulatory networks. Overall, the Listeria model exemplifies that fine RNA tuners act together with master regulatory proteins to orchestrate appropriate transcriptional programmes. PMID:27217337

A comparative analysis of host responses to avian influenza infection in ducks and chickens highlights a role for the interferon-induced transmembrane proteins in viral resistance.

PubMed

Smith, Jacqueline; Smith, Nikki; Yu, Le; Paton, Ian R; Gutowska, Maria Weronika; Forrest, Heather L; Danner, Angela F; Seiler, J Patrick; Digard, Paul; Webster, Robert G; Burt, David W

2015-08-04

Chickens are susceptible to infection with a limited number of Influenza A viruses and are a potential source of a human influenza pandemic. In particular, H5 and H7 haemagglutinin subtypes can evolve from low to highly pathogenic strains in gallinaceous poultry. Ducks on the other hand are a natural reservoir for these viruses and are able to withstand most avian influenza strains. Transcriptomic sequencing of lung and ileum tissue samples from birds infected with high (H5N1) and low (H5N2) pathogenic influenza viruses has allowed us to compare the early host response to these infections in both these species. Chickens (but not ducks) lack the intracellular receptor for viral ssRNA, RIG-I and the gene for an important RIG-I binding protein, RNF135. These differences in gene content partly explain the differences in host responses to low pathogenic and highly pathogenic avian influenza virus in chicken and ducks. We reveal very different patterns of expression of members of the interferon-induced transmembrane protein (IFITM) gene family in ducks and chickens. In ducks, IFITM1, 2 and 3 are strongly up regulated in response to highly pathogenic avian influenza, where little response is seen in chickens. Clustering of gene expression profiles suggests IFITM1 and 2 have an anti-viral response and IFITM3 may restrict avian influenza virus through cell membrane fusion. We also show, through molecular phylogenetic analyses, that avian IFITM1 and IFITM3 genes have been subject to both episodic and pervasive positive selection at specific codons. In particular, avian IFITM1 showed evidence of positive selection in the duck lineage at sites known to restrict influenza virus infection. Taken together these results support a model where the IFITM123 protein family and RIG-I all play a crucial role in the tolerance of ducks to highly pathogenic and low pathogenic strains of avian influenza viruses when compared to the chicken.

Gene Rearrangement Attenuates Expression and Lethality of a Nonsegmented Negative Strand RNA Virus

NASA Astrophysics Data System (ADS)

Williams Wertz, Gail; Perepelitsa, Victoria P.; Ball, L. Andrew

1998-03-01

The nonsegmented negative strand RNA viruses comprise hundreds of human, animal, insect, and plant pathogens. Gene expression of these viruses is controlled by the highly conserved order of genes relative to the single transcriptional promoter. We utilized this regulatory mechanism to alter gene expression levels of vesicular stomatitis virus by rearranging the gene order. This report documents that gene expression levels and the viral phenotype can be manipulated in a predictable manner. Translocation of the promoter-proximal nucleocapsid protein gene N, whose product is required stoichiometrically for genome replication, to successive positions down the genome reduced N mRNA and protein expression in a stepwise manner. The reduction in N gene expression resulted in a stepwise decrease in genomic RNA replication. Translocation of the N gene also attenuated the viruses to increasing extents for replication in cultured cells and for lethality in mice, without compromising their ability to elicit protective immunity. Because monopartite negative strand RNA viruses have not been reported to undergo homologous recombination, gene rearrangement should be irreversible and may provide a rational strategy for developing stably attenuated live vaccines against this type of virus.

Identifying a gene expression signature of cluster headache in blood

PubMed Central

Eising, Else; Pelzer, Nadine; Vijfhuizen, Lisanne S.; Vries, Boukje de; Ferrari, Michel D.; ‘t Hoen, Peter A. C.; Terwindt, Gisela M.; van den Maagdenberg, Arn M. J. M.

2017-01-01

Cluster headache is a relatively rare headache disorder, typically characterized by multiple daily, short-lasting attacks of excruciating, unilateral (peri-)orbital or temporal pain associated with autonomic symptoms and restlessness. To better understand the pathophysiology of cluster headache, we used RNA sequencing to identify differentially expressed genes and pathways in whole blood of patients with episodic (n = 19) or chronic (n = 20) cluster headache in comparison with headache-free controls (n = 20). Gene expression data were analysed by gene and by module of co-expressed genes with particular attention to previously implicated disease pathways including hypocretin dysregulation. Only moderate gene expression differences were identified and no associations were found with previously reported pathogenic mechanisms. At the level of functional gene sets, associations were observed for genes involved in several brain-related mechanisms such as GABA receptor function and voltage-gated channels. In addition, genes and modules of co-expressed genes showed a role for intracellular signalling cascades, mitochondria and inflammation. Although larger study samples may be required to identify the full range of involved pathways, these results indicate a role for mitochondria, intracellular signalling and inflammation in cluster headache. PMID:28074859

Sequence variations and protein expression levels of the two immune evasion proteins Gpm1 and Pra1 influence virulence of clinical Candida albicans isolates.

PubMed

Luo, Shanshan; Hipler, Uta-Christina; Münzberg, Christin; Skerka, Christine; Zipfel, Peter F

2015-01-01

Candida albicans, the important human fungal pathogen uses multiple evasion strategies to control, modulate and inhibit host complement and innate immune attack. Clinical C. albicans strains vary in pathogenicity and in serum resistance, in this work we analyzed sequence polymorphisms and variations in the expression levels of two central fungal complement evasion proteins, Gpm1 (phosphoglycerate mutase 1) and Pra1 (pH-regulated antigen 1) in thirteen clinical C. albicans isolates. Four nucleotide (nt) exchanges, all representing synonymous exchanges, were identified within the 747-nt long GPM1 gene. For the 900-nt long PRA1 gene, sixteen nucleotide exchanges were identified, which represented synonymous, as well as non-synonymous exchanges. All thirteen clinical isolates had a homozygous exchange (A to G) at position 73 of the PRA1 gene. Surface levels of Gpm1 varied by 8.2, and Pra1 levels by 3.3 fold in thirteen tested isolates and these differences influenced fungal immune fitness. The high Gpm1/Pra1 expressing candida strains bound the three human immune regulators more efficiently, than the low expression strains. The difference was 44% for Factor H binding, 51% for C4BP binding and 23% for plasminogen binding. This higher Gpm1/Pra1 expressing strains result in enhanced survival upon challenge with complement active, Factor H depleted human serum (difference 40%). In addition adhesion to and infection of human endothelial cells was increased (difference 60%), and C3b surface deposition was less effective (difference 27%). Thus, variable expression levels of central immune evasion protein influences immune fitness of the human fungal pathogen C. albicans and thus contribute to fungal virulence.

The expression of heterologous MAM-7 in Lactobacillus rhamnosus reduces its intrinsic capacity to inhibit colonization of pathogen Vibrio parahaemolyticus in vitro.

PubMed

Beltran, Sebastian; Munoz-Bergmann, Cristian A; Elola-Lopez, Ana; Quintana, Javiera; Segovia, Cristopher; Trombert, Annette N

2016-01-07

Vibrio parahaemolyticus (V. parahaemolyticus) is a Gram-negative, halophilic bacterium recognized as one of the most important foodborne pathogen. When ingested, V. parahaemolyticus causes a self-limiting illness (Vibriosis), characterized mainly by watery diarrhoea. Treatment is usually oral rehydration and/or antibiotics in complicated cases. Since 1996, the pathogenic and pandemic V. parahaemolyticus O3:K6 serotype has spread worldwide, increasing the reported number of vibriosis cases. Thus, the design of new strategies for pathogen control and illness prevention is necessary. Lactobacillus sp. grouped Gram positive innocuous bacteria, part of normal intestinal microbiota and usually used as oral vaccines for several diarrheic diseases. Recombinants strains of Lactobacillus (RL) expressing pathogen antigens can be used as part of an anti-adhesion strategy where RL block the pathogen union sites in host cells. Thus, we aimed to express MAM-7 V. parahaemolyticus adhesion protein in Lactobacillus sp. to generate an RL that prevents pathogen colonization. We cloned the MAM-7 gene from V. parahaemolyticus RIMD 2210633 in Lactobacillus expression vectors. Recombinant strains (Lactobacillus rhamnosus pSEC-MAM7 and L. rhamnosus pCWA-MAM7) adhered to CaCo-2 cells and competed with the pathogen. However, the L. rhamnosus wild type strain showed the best capacity to inhibit pathogen colonization in vitro. In addition, LDH-assay showed that recombinant strains were cytotoxic compared with the wild type isogenic strain. MAM-7 expression in lactobacilli reduces the intrinsic inhibitory capacity of L. rhamnosus against V. parahaemolyticus.

Comparative Digital Gene Expression Analysis of Tissue-Cultured Plantlets of Highly Resistant and Susceptible Banana Cultivars in Response to Fusarium oxysporum

PubMed Central

Niu, Yuqing; Hu, Bei; Li, Xiaoquan; Chen, Houbin; Šamaj, Jozef; Xu, Chunxiang

2018-01-01

Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc) is one of the most destructive soil-borne diseases. In this study, young tissue-cultured plantlets of banana (Musa spp. AAA) cultivars differing in Foc susceptibility were used to reveal their differential responses to this pathogen using digital gene expression (DGE). Data were evaluated by various bioinformatic tools (Venn diagrams, gene ontology (GO) annotation and Kyoto encyclopedia of genes and genomes (KEGG) pathway analyses) and immunofluorescence labelling method to support the identification of gene candidates determining the resistance of banana against Foc. Interestingly, we have identified MaWRKY50 as an important gene involved in both constitutive and induced resistance. We also identified new genes involved in the resistance of banana to Foc, including several other transcription factors (TFs), pathogenesis-related (PR) genes and some genes related to the plant cell wall biosynthesis or degradation (e.g., pectinesterases, β-glucosidases, xyloglucan endotransglucosylase/hydrolase and endoglucanase). The resistant banana cultivar shows activation of PR-3 and PR-4 genes as well as formation of different constitutive cell barriers to restrict spreading of the pathogen. These data suggest new mechanisms of banana resistance to Foc. PMID:29364855

Gall midges (Hessian flies) as plant pathogens.

PubMed

Stuart, Jeff J; Chen, Ming-Shun; Shukle, Richard; Harris, Marion O

2012-01-01

Gall midges constitute an important group of plant-parasitic insects. The Hessian fly (HF; Mayetiola destructor), the most investigated gall midge, was the first insect hypothesized to have a gene-for-gene interaction with its host plant, wheat (Triticum spp.). Recent investigations support that hypothesis. The minute larval mandibles appear to act in a manner that is analogous to nematode stylets and the haustoria of filamentous plant pathogens. Putative effector proteins are encoded by hundreds of genes and expressed in the HF larval salivary gland. Cultivar-specific resistance (R) genes mediate a highly localized plant reaction that prevents the survival of avirulent HF larvae. Fine-scale mapping of HF avirulence (Avr) genes provides further evidence of effector-triggered immunity (ETI) against HF in wheat. Taken together, these discoveries suggest that the HF, and other gall midges, may be considered biotrophic, or hemibiotrophic, plant pathogens, and they demonstrate the potential that the wheat-HF interaction has in the study of insect-induced plant gall formation.

The Verticillium-specific protein VdSCP7 localizes to the plant nucleus and modulates immunity to fungal infections.

PubMed

Zhang, Lisha; Ni, Hao; Du, Xuan; Wang, Sheng; Ma, Xiao-Wei; Nürnberger, Thorsten; Guo, Hui-Shan; Hua, Chenlei

2017-07-01

Fungal pathogens secrete effector proteins to suppress plant basal defense for successful colonization. Resistant plants, however, can recognize effectors by cognate R proteins to induce effector-triggered immunity (ETI). By analyzing secretomes of the vascular fungal pathogen Verticillium dahliae, we identified a novel secreted protein VdSCP7 that targets the plant nucleus. The green fluorescent protein (GFP)-tagged VdSCP7 gene with either a mutated nuclear localization signal motif or with additional nuclear export signal was transiently expressed in Nicotiana benthamiana, and investigated for induction of plant immunity. The role of VdSCP7 in V. dahliae pathogenicity was characterized by gene knockout and complementation, and GFP labeling. Expression of the VdSCP7 gene in N. benthamiana activated both salicylic acid and jasmonate signaling, and altered the plant's susceptibility to the pathogens Botrytis cinerea and Phytophthora capsici. The immune response activated by VdSCP7 was highly dependent on its initial extracellular secretion and subsequent nuclear localization in plants. Knockout of the VdSCP7 gene significantly enhanced V. dahliae aggressiveness on cotton. GFP-labeled VdSCP7 is secreted by V. dahliae and accumulates in the plant nucleus. We conclude that VdSCP7 is a novel effector protein that targets the host nucleus to modulate plant immunity, and suggest that plants can recognize VdSCP7 to activate ETI during fungal infection. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Genes that are involved in high hydrostatic pressure treatments in a Listeria monocytogenes Scott A ctsR deletion mutant

USDA-ARS?s Scientific Manuscript database

Listeria monocytogenes is a food-borne pathogen of significant threat to public health. High Hydrostatic Pressure (HHP) treatment can be used to control L. monocytogenes in food. The CtsR (class three stress gene repressor) protein negatively regulates the expression of class III heat shock genes....

Transgenic expression of a maize geranyl geranyl transferase gene sequence in maize callus increases resistance to ear rot pathogens

USDA-ARS?s Scientific Manuscript database

Determining the genes responsible for pest resistance in maize can allow breeders to develop varieties with lower losses and less contamination with undesirable toxins. A gene sequence coding for a geranyl geranyl transferase-like protein located in a fungal ear rot resistance quantitative trait loc...

Transcriptome profiling analysis of Vibrio vulnificus during human infection.

PubMed

Bisharat, Naiel; Bronstein, Michal; Korner, Mira; Schnitzer, Temima; Koton, Yael

2013-09-01

Vibrio vulnificus is a waterborne pathogen that was responsible for an outbreak of severe soft-tissue infections among fish farmers and fish consumers in Israel. Several factors have been shown to be associated with virulence. However, the transcriptome profile of the pathogen during human infection has not been determined yet. We compared the transcriptome profile, using RNA sequencing, of a human-pathogenic strain harvested directly from tissue of a patient suffering from severe soft-tissue infection with necrotizing fasciitis, with the same strain and three other environmental strains grown in vitro. The five sequenced libraries were aligned to the reference genomes of V. vulnificus strains CMCP6 and YJ016. Approximately 47.8 to 62.3 million paired-end raw reads were generated from the five runs. Nearly 84 % of the genome was covered by reads from at least one of the five runs, suggesting that nearly 16 % of the genome is not transcribed or is transcribed at low levels. We identified 123 genes that were differentially expressed during the acute phase of infection. Sixty-three genes were mapped to the large chromosome, 47 genes mapped to the small chromosome and 13 genes mapped to the YJ016 plasmid. The 123 genes fell into a variety of functional categories including transcription, signal transduction, cell motility, carbohydrate metabolism, intracellular trafficking and cell envelope biogenesis. Among the genes differentially expressed during human infection we identified genes encoding bacterial toxin (RtxA1) and genes involved in flagellar components, Flp-coding region, GGDEF family protein, iron acquisition system and sialic acid metabolism.

Transcriptome profiling of resistant and susceptible Cavendish banana roots following inoculation with Fusarium oxysporum f. sp. cubense tropical race 4

PubMed Central

2012-01-01

Background Fusarium wilt, caused by the fungal pathogen Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4), is considered the most lethal disease of Cavendish bananas in the world. The disease can be managed in the field by planting resistant Cavendish plants generated by somaclonal variation. However, little information is available on the genetic basis of plant resistance to Foc TR4. To a better understand the defense response of resistant banana plants to the Fusarium wilt pathogen, the transcriptome profiles in roots of resistant and susceptible Cavendish banana challenged with Foc TR4 were compared. Results RNA-seq analysis generated more than 103 million 90-bp clean pair end (PE) reads, which were assembled into 88,161 unigenes (mean size = 554 bp). Based on sequence similarity searches, 61,706 (69.99%) genes were identified, among which 21,273 and 50,410 unigenes were assigned to gene ontology (GO) categories and clusters of orthologous groups (COG), respectively. Searches in the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG) mapped 33,243 (37.71%) unigenes to 119 KEGG pathways. A total of 5,008 genes were assigned to plant-pathogen interactions, including disease defense and signal transduction. Digital gene expression (DGE) analysis revealed large differences in the transcriptome profiles of the Foc TR4-resistant somaclonal variant and its susceptible wild-type. Expression patterns of genes involved in pathogen-associated molecular pattern (PAMP) recognition, activation of effector-triggered immunity (ETI), ion influx, and biosynthesis of hormones as well as pathogenesis-related (PR) genes, transcription factors, signaling/regulatory genes, cell wall modification genes and genes with other functions were analyzed and compared. The results indicated that basal defense mechanisms are involved in the recognition of PAMPs, and that high levels of defense-related transcripts may contribute to Foc TR4 resistance in banana. Conclusions This study generated a substantial amount of banana transcript sequences and compared the defense responses against Foc TR4 between resistant and susceptible Cavendish bananas. The results contribute to the identification of candidate genes related to plant resistance in a non-model organism, banana, and help to improve the current understanding of host-pathogen interactions. PMID:22863187

HDT701, a histone H4 deacetylase, negatively regulates plant innate immunity by modulating histone H4 acetylation of defense-related genes in rice.

PubMed

Ding, Bo; Bellizzi, Maria del Rosario; Ning, Yuese; Meyers, Blake C; Wang, Guo-Liang

2012-09-01

Histone acetylation and deacetylation play an important role in the modification of chromatin structure and regulation of gene expression in eukaryotes. Chromatin acetylation status is modulated antagonistically by histone acetyltransferases and histone deacetylases (HDACs). In this study, we characterized the function of histone deacetylase701 (HDT701), a member of the plant-specific HD2 subfamily of HDACs, in rice (Oryza sativa) innate immunity. Transcription of HDT701 is increased in the compatible reaction and decreased in the incompatible reaction after infection by the fungal pathogen Magnaporthe oryzae. Overexpression of HDT701 in transgenic rice leads to decreased levels of histone H4 acetylation and enhanced susceptibility to the rice pathogens M. oryzae and Xanthomonas oryzae pv oryzae (Xoo). By contrast, silencing of HDT701 in transgenic rice causes elevated levels of histone H4 acetylation and elevated transcription of pattern recognition receptor (PRR) and defense-related genes, increased generation of reactive oxygen species after pathogen-associated molecular pattern elicitor treatment, as well as enhanced resistance to both M. oryzae and Xoo. We also found that HDT701 can bind to defense-related genes to regulate their expression. Taken together, these results demonstrate that HDT701 negatively regulates innate immunity by modulating the levels of histone H4 acetylation of PRR and defense-related genes in rice.

Phylogenetic analysis of PR genes in some pome fruit species with the emphasis on transcriptional analysis and ROS response under Erwinia amylovora inoculation in apple.

PubMed

Hassani, Maryam; Salami, Seyed Alireza; Nasiri, Jaber; Abdollahi, Hamid; Ghahremani, Zahra

2016-02-01

Attempts were made to identify eight pathogenesis related (PR) genes (i.e., PR-1a, PR3-ch1, PR3-Ch2, PR3-Ch3, PR3-Ch4, PR3-Ch5, PR-5 and PR-8) from 27 genotypes of apple, quince and pear, which are induced in response to inoculation with the pathogen Erwinia amylovora, the causal agent of fire blight. Totally, 32 PR genes of different families were obtained, excepting PR3-Ch2 (amplified only in apple) and PR3-Ch4 (amplified only in apple and pear), the others were successfully amplified in all the genotypes of apple, quince and pear. Evolutionary, the genes of each family exhibited significant homology with each other, as the corresponded phylogenetic neighbor-joining-based dendrograms were taken into consideration. Meanwhile, according to the expression assay, it was deduced that the pathogen activity can significantly affect the expression levels of some selected PR genes of PR3-Ch2, PR3-Ch4, PR3-Ch5 and particularly Cat I in both resistant (MM-111) and semi-susceptible (MM-106) apple rootstocks. Lastly, it was concluded that the pathogen E. amylovora is able to stimulate ROS response, particularly using generation of hydrogen peroxide (H2O2) in both aforementioned apple rootstock.

Comparative Genomic Analysis of Drechmeria coniospora Reveals Core and Specific Genetic Requirements for Fungal Endoparasitism of Nematodes

PubMed Central

Thakur, Nishant; Arguel, Marie-Jeanne; Polanowska, Jolanta; Henrissat, Bernard; Record, Eric; Magdelenat, Ghislaine; Barbe, Valérie; Raffaele, Sylvain; Barbry, Pascal

2016-01-01

Drechmeria coniospora is an obligate fungal pathogen that infects nematodes via the adhesion of specialized spores to the host cuticle. D. coniospora is frequently found associated with Caenorhabditis elegans in environmental samples. It is used in the study of the nematode’s response to fungal infection. Full understanding of this bi-partite interaction requires knowledge of the pathogen’s genome, analysis of its gene expression program and a capacity for genetic engineering. The acquisition of all three is reported here. A phylogenetic analysis placed D. coniospora close to the truffle parasite Tolypocladium ophioglossoides, and Hirsutella minnesotensis, another nematophagous fungus. Ascomycete nematopathogenicity is polyphyletic; D. coniospora represents a branch that has not been molecularly characterized. A detailed in silico functional analysis, comparing D. coniospora to 11 fungal species, revealed genes and gene families potentially involved in virulence and showed it to be a highly specialized pathogen. A targeted comparison with nematophagous fungi highlighted D. coniospora-specific genes and a core set of genes associated with nematode parasitism. A comparative gene expression analysis of samples from fungal spores and mycelia, and infected C. elegans, gave a molecular view of the different stages of the D. coniospora lifecycle. Transformation of D. coniospora allowed targeted gene knock-out and the production of fungus that expresses fluorescent reporter genes. It also permitted the initial characterisation of a potential fungal counter-defensive strategy, involving interference with a host antimicrobial mechanism. This high-quality annotated genome for D. coniospora gives insights into the evolution and virulence of nematode-destroying fungi. Coupled with genetic transformation, it opens the way for molecular dissection of D. coniospora physiology, and will allow both sides of the interaction between D. coniospora and C. elegans, as well as the evolutionary arms race that exists between pathogen and host, to be studied. PMID:27153332

Bifidobacterium breve IPLA20005 affects in vitro the expression of hly and luxS genes, related to the virulence of Listeria monocytogenes Lm23.

PubMed

Rios-Covian, David; Nogacka, Alicja; Salazar, Nuria; Hernández-Barranco, A M; Cuesta, Isabel; Gueimonde, Miguel; de Los Reyes Gavilán, Clara G

2018-03-01

Mechanistic features that characterize the interaction and inhibition of the food-borne pathogen Listeria monocytogenes by members of the genus Bifidobacterium still remain unclear. In the present work, we tried to shed light on the influence that co-cultivation of L. monocytogenes with Bifidobacterium breve may exert on both microorganisms and on virulence of the pathogen. Production of acetate and lactate was measured by gas chromatography and high-performance liquid chromatography, respectively; bacterial counts were obtained by plate count; gene expression was determined by RT-qPCR; and haemolytic activity was analyzed against goat erythrocytes. We found slightly but significantly lower final counts of Listeria and Bifidobacterium (p < 0.05) and lower haemolytic efficiency in L. monocytogenes cells from cocultures than in those from monocultures. In contrast, the hly and luxS genes, which code for the cytolysin listeriolysin O and participate in biofilm formation, respectively, were overexpressed when L. monocytogenes was grown in coculture. This indicates that the presence of Bifidobacterium is able to modify the gene expression and haemolytic activity of L. monocytogenes when both microorganisms grow together.

Screening, Expression, Purification and Functional Characterization of Novel Antimicrobial Peptide Genes from Hermetia illucens (L.).

PubMed

Elhag, Osama; Zhou, Dingzhong; Song, Qi; Soomro, Abdul Aziz; Cai, Minmin; Zheng, Longyu; Yu, Ziniu; Zhang, Jibin

2017-01-01

Antimicrobial peptides from a wide spectrum of insects possess potent microbicidal properties against microbial-related diseases. In this study, seven new gene fragments of three types of antimicrobial peptides were obtained from Hermetia illucens (L), and were named cecropinZ1, sarcotoxin1, sarcotoxin (2a), sarcotoxin (2b), sarcotoxin3, stomoxynZH1, and stomoxynZH1(a). Among these genes, a 189-basepair gene (stomoxynZH1) was cloned into the pET32a expression vector and expressed in the Escherichia coli as a fusion protein with thioredoxin. Results show that Trx-stomoxynZH1 exhibits diverse inhibitory activity on various pathogens, including Gram-positive bacterium Staphylococcus aureus, Gram-negative bacterium Escherichia coli, fungus Rhizoctonia solani Khün (rice)-10, and fungus Sclerotinia sclerotiorum (Lib.) de Bary-14. The minimum inhibitory concentration of Trx-stomoxynZH1 is higher against Gram-positive bacteria than against Gram-negative bacteria but similar between the fungal strains. These results indicate that H. illucens (L.) could provide a rich source for the discovery of novel antimicrobial peptides. Importantly, stomoxynZH1 displays a potential benefit in controlling antibiotic-resistant pathogens.

Screening, Expression, Purification and Functional Characterization of Novel Antimicrobial Peptide Genes from Hermetia illucens (L.)

PubMed Central

Elhag, Osama; Zhou, Dingzhong; Song, Qi; Soomro, Abdul Aziz; Cai, Minmin; Zheng, Longyu; Yu, Ziniu; Zhang, Jibin

2017-01-01

Antimicrobial peptides from a wide spectrum of insects possess potent microbicidal properties against microbial-related diseases. In this study, seven new gene fragments of three types of antimicrobial peptides were obtained from Hermetia illucens (L), and were named cecropinZ1, sarcotoxin1, sarcotoxin (2a), sarcotoxin (2b), sarcotoxin3, stomoxynZH1, and stomoxynZH1(a). Among these genes, a 189-basepair gene (stomoxynZH1) was cloned into the pET32a expression vector and expressed in the Escherichia coli as a fusion protein with thioredoxin. Results show that Trx-stomoxynZH1 exhibits diverse inhibitory activity on various pathogens, including Gram-positive bacterium Staphylococcus aureus, Gram-negative bacterium Escherichia coli, fungus Rhizoctonia solani Khün (rice)-10, and fungus Sclerotinia sclerotiorum (Lib.) de Bary-14. The minimum inhibitory concentration of Trx-stomoxynZH1 is higher against Gram-positive bacteria than against Gram-negative bacteria but similar between the fungal strains. These results indicate that H. illucens (L.) could provide a rich source for the discovery of novel antimicrobial peptides. Importantly, stomoxynZH1 displays a potential benefit in controlling antibiotic-resistant pathogens. PMID:28056070

Metaproteomics reveals abundant transposase expression in mutualistic endosymbionts

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kleiner, Manuel; Young, Jacque C; Shah, Manesh B

2013-01-01

Transposases, enzymes that catalyze the movement of mobile genetic elements, are the most abundant genes in nature. While many bacteria encode an abundance of transposases in their genomes, the current paradigm is that transposase gene expression is tightly regulated and generally low due to its severe mutagenic effects. In the current study, we detected the highest number of transposase proteins ever reported in bacteria, in symbionts of the gutless marine worm Olavius algarvensis using metaproteomics. At least 26 different transposases from 12 different families were detected and genomic and proteomic analyses suggest many of these are active. This high expressionmore » of transposases indicates that the mechanisms for their tight regulation have been disabled or destroyed. Based on recent studies on other symbionts and pathogens that showed high transposase transcription, we speculate that abundant transposase expression might be common in symbionts and pathogens.« less

Fungal and host transcriptome analysis of pH-regulated genes during colonization of apple fruits by Penicillium expansum.

PubMed

Barad, Shiri; Sela, Noa; Kumar, Dilip; Kumar-Dubey, Amit; Glam-Matana, Nofar; Sherman, Amir; Prusky, Dov

2016-05-04

Penicillium expansum is a destructive phytopathogen that causes decay in deciduous fruits during postharvest handling and storage. During colonization the fungus secretes D-gluconic acid (GLA), which modulates environmental pH and regulates mycotoxin accumulation in colonized tissue. Till now no transcriptomic analysis has addressed the specific contribution of the pathogen's pH regulation to the P. expansum colonization process. For this purpose total RNA from the leading edge of P. expansum-colonized apple tissue of cv. 'Golden Delicious' and from fungal cultures grown under pH 4 or 7 were sequenced and their gene expression patterns were compared. We present a large-scale analysis of the transcriptome data of P. expansum and apple response to fungal colonization. The fungal analysis revealed nine different clusters of gene expression patterns that were divided among three major groups in which the colonized tissue showed, respectively: (i) differing transcript expression patterns between mycelial growth at pH 4 and pH 7; (ii) similar transcript expression patterns of mycelial growth at pH 4; and (iii) similar transcript expression patterns of mycelial growth at pH 7. Each group was functionally characterized in order to decipher genes that are important for pH regulation and also for colonization of apple fruits by Penicillium. Furthermore, comparison of gene expression of healthy apple tissue with that of colonized tissue showed that differentially expressed genes revealed up-regulation of the jasmonic acid and mevalonate pathways, and also down-regulation of the glycogen and starch biosynthesis pathways. Overall, we identified important genes and functionalities of P. expansum that were controlled by the environmental pH. Differential expression patterns of genes belonging to the same gene family suggest that genes were selectively activated according to their optimal environmental conditions (pH, in vitro or in vivo) to enable the fungus to cope with varying conditions and to make optimal use of available enzymes. Comparison between the activation of the colonized host's gene responses by alkalizing Colletotrichum gloeosporioides and acidifying P. expansum pathogens indicated similar gene response patterns, but stronger responses to P. expansum, suggesting the importance of acidification by P. expansum as a factor in its increased aggressiveness.

Identification of Scedosporium boydii catalase A1 gene, a reactive oxygen species detoxification factor highly expressed in response to oxidative stress and phagocytic cells.

PubMed

Mina, Sara; Staerck, Cindy; d'Almeida, Sènan M; Marot, Agnès; Delneste, Yves; Calenda, Alphonse; Tabiasco, Julie; Bouchara, Jean-Philippe; Fleury, Maxime J J

2015-12-01

Scedosporium boydii is an opportunistic filamentous fungus which may be responsible for a large variety of infections in both immunocompetent and immunocompromised individuals. This fungus belongs to the Scedosporium apiospermum species complex which usually ranks second among the filamentous fungi colonizing the airways of patients with cystic fibrosis (CF). Species of the S. apiospermum complex are able to chronically colonize the CF airways suggesting pathogenic mechanisms allowing persistence and growth of these fungi in the respiratory tract. Few putative virulence factors have been purified and characterized so far in the S. apiospermum complex including a cytosolic Cu,Zn-superoxide dismutase (SOD) and a monofunctional catalase (catalase A1). Upon microbial infection, host phagocytes release reactive oxygen species (ROS), such as hydrogen peroxide, as part of the antimicrobial response. Catalases are known to protect pathogens against ROS by degradation of the hydrogen peroxide. Here, we identified the S. boydii catalase A1 gene (CATA1) and investigated its expression in response to the environmental conditions encountered in the CF airways and to the oxidative stress. Results showed that S. boydii CATA1 gene expression is not affected by hypoxia, hypercapnia or pH changes. In contrast, CATA1 gene was overexpressed in response to a chemically induced oxidative stress with a relative gene expression 37-fold higher in the presence of 250 μM H(2)O(2), 20-fold higher with 250 μM menadione and 5-fold higher with 2 mM paraquat. Moreover, S. boydii CATA1 gene expression progressively increased upon exposure to activated THP-1-derived macrophages, reaching a maximum after 12 h (26 fold). Activated HL60-derived neutrophils and activated human peripheral blood neutrophils more rapidly induced S. boydii CATA1 gene overexpression, a maximum gene expression level being reached at 75 min (17 fold) and 60 min (15 fold), respectively. In contrast expression of the gene encoding the Cu,Zn-SOD (SODC gene) was not affected by H(2)O(2), menadione, paraquat or in co-culture with phagocytic cells. These results suggest that S. boydii CATA1 gene is highly stimulated by the oxidative burst response whereas SODC gene is constitutively expressed. Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Transmembrane transporter expression regulated by the glucosylceramide pathway in Cryptococcus neoformans.

PubMed

Singh, Arpita; Rella, Antonella; Schwacke, John; Vacchi-Suzzi, Caterina; Luberto, Chiara; Del Poeta, Maurizio

2015-11-16

The sphingolipid glucosylceramide (GlcCer) and factors involved in the fungal GlcCer pathways were shown earlier to be an integral part of fungal virulence, especially in fungal replication at 37 °C, in neutral/alkaline pH and 5 % CO2 environments (e.g. alveolar spaces). Two mutants, ∆gcs 1 lacking glucosylceramide synthase 1 gene (GCS1) which catalyzes the formation of sphingolipid GlcCer from the C9-methyl ceramide and ∆smt1 lacking sphingolipid C9 methyltransferase gene (SMT1), which adds a methyl group to position nine of the sphingosine backbone of ceramide, of this pathway were attenuated in virulence and have a growth defect at the above-mentioned conditions. These mutants with either no or structurally modified GlcCer located on the cell-membrane have reduced membrane rigidity, which may have altered not only the physical location of membrane proteins but also their expression, as the pathogen's mode of adaptation to changing need. Importantly, pathogens are known to adapt themselves to the changing host environments by altering their patterns of gene expression. By transcriptional analysis of gene expression, we identified six genes whose expression was changed from their wild-type counterpart grown in the same conditions, i.e. they became either down regulated or up regulated in these two mutants. The microarray data was validated by real-time PCR, which confirmed their fold change in gene expression. All the six genes we identified, viz siderochrome-iron transporter (CNAG_02083), monosaccharide transporter (CNAG_05340), glucose transporter (CNAG_03772), membrane protein (CNAG_03912), membrane transport protein (CNAG_00539), and sugar transporter (CNAG_06963), are membrane-localized and have significantly altered gene expression levels. Therefore, we hypothesize that these genes function either independently or in tandem with a structurally modified cell wall/plasma membrane resulting from the modifications of the GlcCer pathway and thus possibly disrupt transmembrane signaling complex, which in turn contributes to cryptococcal osmotic, pH, ion homeostasis and its pathobiology. Six genes identified from gene expression microarrays by gene set enrichment analysis and validated by RT-PCR, are membrane located and associated with the growth defect at neutral-alkaline pH due to the absence and or presence of a structurally modified GlcCer. They may be involved in the transmembrane signaling network in Cryptococcus neoformans, and therefore the pathobiology of the fungus in these conditions.

Mucin2 is Required for Probiotic Agents-Mediated Blocking Effects on Meningitic E. coli-Induced Pathogenicities.

PubMed

Yu, Jing-Yi; He, Xiao-Long; Puthiyakunnon, Santhosh; Peng, Liang; Li, Yan; Wu, Li-Sha; Peng, Wen-Ling; Zhang, Ya; Gao, Jie; Zhang, Yao-Yuan; Boddu, Swapna; Long, Min; Cao, Hong; Huang, Sheng-He

2015-10-01

Mucin2 (MUC2), an important regulatory factor in the immune system, plays an important role in the host defense system against bacterial translocation. Probiotics known to regulate MUC2 gene expression have been widely studied, but the interactions among probiotic, pathogens, and mucin gene are still not fully understood. The aim of this study was to investigate the role of MUC2 in blocking effects of probiotics on meningitic E. coli-induced pathogenicities. In this study, live combined probiotic tablets containing living Bifidobacterium, Lactobacillus bulgaricus, and Streptococcus thermophilus were used. MUC2 expression was knocked down in Caco-2 cells by RNA interference. 5-Aza-2'-deoxycytidine (5-Aza-CdR), which enhances mucin-promoted probiotic effects through inducing production of Sadenosyl- L-methionine (SAMe), was used to up-regulate MUC2 expression in Caco-2 cells. The adhesion to and invasion of meningitic E. coli were detected by competition assays. Our studies showed that probiotic agents could block E. coli-caused intestinal colonization, bacteremia, and meningitis in a neonatal sepsis and meningitis rat model. MUC2 gene expression in the neonatal rats given probiotic agents was obviously higher than that of the infected and uninfected control groups without probiotic treatment. The prohibitive effects of probiotic agents on MUC2-knockdown Caco-2 cells infected with E44 were significantly reduced compared with nontransfected Caco-2 cells. Moreover, the results also showed that 5- Aza-CdR, a drug enhancing the production of SAMe that is a protective agent of probiotics, was able to significantly suppress adhesion and invasion of E44 to Caco-2 cells by upregulation of MUC2 expression. Taken together, our data suggest that probiotic agents can efficiently block meningitic E. coli-induced pathogenicities in a manner dependent on MUC2.

Identification of pathogenic genes and upstream regulators in age-related macular degeneration.

PubMed

Zhao, Bin; Wang, Mengya; Xu, Jing; Li, Min; Yu, Yuhui

2017-06-26

Age-related macular degeneration (AMD) is the leading cause of irreversible blindness in older individuals. Our study aims to identify the key genes and upstream regulators in AMD. To screen pathogenic genes of AMD, an integrated analysis was performed by using the microarray datasets in AMD derived from the Gene Expression Omnibus (GEO) database. The functional annotation and potential pathways of differentially expressed genes (DEGs) were further discovered by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. We constructed the AMD-specific transcriptional regulatory network to find the crucial transcriptional factors (TFs) which target the DEGs in AMD. Quantitative real time polymerase chain reaction (qRT-PCR) was performed to verify the DEGs and TFs obtained by integrated analysis. From two GEO datasets obtained, we identified 1280 DEGs (730 up-regulated and 550 down-regulated genes) between AMD and normal control (NC). After KEGG analysis, steroid biosynthesis is a significantly enriched pathway for DEGs. The expression of 8 genes (TNC, GRP, TRAF6, ADAMTS5, GPX3, FAP, DHCR7 and FDFT1) was detected. Except for TNC and GPX3, the other 6 genes in qRT-PCR played the same pattern with that in our integrated analysis. The dysregulation of these eight genes may involve with the process of AMD. Two crucial transcription factors (c-rel and myogenin) were concluded to play a role in AMD. Especially, myogenin was associated with AMD by regulating TNC, GRP and FAP. Our finding can contribute to developing new potential biomarkers, revealing the underlying pathogenesis, and further raising new therapeutic targets for AMD.