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.

Utilization of evolutionary model, bioinformatics and heuristics for development of a multiplex Escherichia coli O157:H7 PCR assay

USDA-ARS?s Scientific Manuscript database

Introduction: Escherichia coli O157:H7 is a devastating foodborne pathogen causing many foodborne outbreaks worldwide with significant morbidity and mortality. The plasticity of the E. coli O157:H7 genome, inconsistent expression of surface antigens, and sharing of genetic elements with other non-...

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.

A Bacteriophage Capsid Protein Is an Inhibitor of a Conserved Transcription Terminator of Various Bacterial Pathogens.

PubMed

Ghosh, Gairika; Reddy, Jayavardhana; Sambhare, Susmit; Sen, Ranjan

2018-01-01

Rho is a hexameric molecular motor that functions as a conserved transcription terminator in the majority of bacterial species and is a potential drug target. Psu is a bacteriophage P4 capsid protein that inhibits Escherichia coli Rho by obstructing its ATPase and translocase activities. In this study, we explored the anti-Rho activity of Psu for Rho proteins from different pathogens. Sequence alignment and homology modeling of Rho proteins from pathogenic bacteria revealed the conserved nature of the Psu-interacting regions in all these proteins. We chose Rho proteins from various pathogens, including Mycobacterium smegmatis , Mycobacterium bovis , Mycobacterium tuberculosis , Xanthomonas campestris , Xanthomonas oryzae , Corynebacterium glutamicum , Vibrio cholerae , Salmonella enterica , and Pseudomonas syringae The purified recombinant Rho proteins of these organisms showed variable rates of ATP hydrolysis on poly(rC) as the substrate and were capable of releasing RNA from the E. coli transcription elongation complexes. Psu was capable of inhibiting these two functions of all these Rho proteins. In vivo pulldown assays revealed direct binding of Psu with many of these Rho proteins. In vivo expression of psu induced killing of M. smegmatis , M. bovis , X. campestris , and E. coli expressing S. enterica Rho indicating Psu-induced inhibition of Rho proteins of these strains under physiological conditions. We propose that the "universal" inhibitory function of the Psu protein against the Rho proteins from both Gram-negative and Gram-positive bacteria could be useful for designing peptides with antimicrobial functions and that these peptides could contribute to synergistic antibiotic treatment of the pathogens by compromising the Rho functions. IMPORTANCE Bacteriophage-derived protein factors modulating different bacterial processes could be converted into unique antimicrobial agents. Bacteriophage P4 capsid protein Psu is an inhibitor of the E. coli transcription terminator Rho. Here we show that apart from antagonizing E. coli Rho, Psu is able to inhibit Rho proteins from various phylogenetically unrelated Gram-negative and Gram-positive pathogens. Upon binding to these Rho proteins, Psu inhibited them by affecting their ATPase and RNA release functions. The expression of Psu in vivo kills various pathogens, such as Mycobacterium and Xanthomonas species. Hence, Psu could be useful for identifying peptide sequences with anti-Rho activities and might constitute part of synergistic antibiotic treatment against pathogens. Copyright © 2017 American Society for Microbiology.

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.

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

Regulation of Cell Division, Biofilm Formation, and Virulence by FlhC in Escherichia coli O157:H7 Grown on Meat▿†

PubMed Central

Sule, Preeti; Horne, Shelley M.; Logue, Catherine M.; Prüß, Birgit M.

2011-01-01

To understand the continuous problems that Escherichia coli O157:H7 causes as food pathogen, this study assessed global gene regulation in bacteria growing on meat. Since FlhD/FlhC of E. coli K-12 laboratory strains was previously established as a major control point in transducing signals from the environment to several cellular processes, this study compared the expression pattern of an E. coli O157:H7 parent strain to that of its isogenic flhC mutant. This was done with bacteria that had been grown on meat. Microarray experiments revealed 287 putative targets of FlhC. Real-time PCR was performed as an alternative estimate of transcription and confirmed microarray data for 13 out of 15 genes tested (87%). The confirmed genes are representative of cellular functions, such as central metabolism, cell division, biofilm formation, and pathogenicity. An additional 13 genes from the same cellular functions that had not been hypothesized as being regulated by FlhC by the microarray experiment were tested with real-time PCR and also exhibited higher expression levels in the flhC mutant than in the parent strain. Physiological experiments were performed and confirmed that FlhC reduced the cell division rate, the amount of biofilm biomass, and pathogenicity in a chicken embryo lethality model. Altogether, this study provides valuable insight into the complex regulatory network of the pathogen that enables its survival under various environmental conditions. This information may be used to develop strategies that could be used to reduce the number of cells or pathogenicity of E. coli O157:H7 on meat by interfering with the signal transduction pathways. PMID:21498760

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.

Enterobacteria secrete an inhibitor of Pseudomonas virulence during clinical bacteriuria

PubMed Central

Ohlemacher, Shannon I.; Giblin, Daryl E.; d’Avignon, D. André; Stapleton, Ann E.; Trautner, Barbara W.; Henderson, Jeffrey P.

2017-01-01

Escherichia coli and other Enterobacteriaceae are among the most common pathogens of the human urinary tract. Among the genetic gains of function associated with urinary E. coli isolates is the Yersinia high pathogenicity island (HPI), which directs the biosynthesis of yersiniabactin (Ybt), a virulence-associated metallophore. Using a metabolomics approach, we found that E. coli and other Enterobacteriaceae expressing the Yersinia HPI also secrete escherichelin, a second metallophore whose chemical structure matches a known synthetic inhibitor of the virulence-associated pyochelin siderophore system in Pseudomonas aeruginosa. We detected escherichelin during clinical E. coli urinary tract infection (UTI) and experimental human colonization with a commensal, potentially probiotic E. coli bacteriuria strain. Escherichelin production by colonizing enterobacteria may help human hosts resist opportunistic infections by Pseudomonas and other pyochelin-expressing bacteria. This siderophore-based mechanism of microbial antagonism may be one of many elements contributing to the protective effects of the human microbiome. Future UTI-preventive probiotic strains may benefit by retaining the escherichelin biosynthetic capacity of the Yersinia HPI while eliminating the Ybt biosynthetic capacity. PMID:28945201

[In vitro function of outer membrane protease T of Escherichia coli K1 pathogenic strain].

PubMed

Hui, Changye; Guo, Yan; Wu, Shuchi; Peng, Liang; Cao, Hong; Huang, Shenghe

2010-01-01

Plasminogen activation and antimicrobial peptide hydrolysis contribute to pathogens invasion and survival in vivo. To demonstrate the expression of outer membrane protease T in E. coli K1 pathogenic strain E44, its activity of plasminogen activator and protamine hydrolysis. After Benzamidine Sepharose Fast Flow and SOURCE 30Q chromatography, we got E44 outer membrane mixed fraction, and examined its activity of plasminogen activation with chromogenic substrate S-2251 method. An ompT deletion mutant of E44 was constructed by using the suicide vector pCVD442, termed as E44ompT. We examined 0.1 mg/mL cationic antimicrobial peptide protamine susceptibility of E44, ompT mutant strain E44ompT and E44ompT harboring pUCT, which was constructed by inserting complete ompT open reading frame into pUC13. We got about 37 kDa E44 membrane extract, which could activate plasminogen, and activation was membrane extract dose dependent. This confirmed the expression of outer membrane protease T in the outer membrane of E44. E44ompT was, more susceptible to 0.1 mg/mL protamine than E44, and E440mpT was partially complemented by pUCT. Outer membrane protease T is expressed in E. coli K1 pathogenic strain E44, and can activate plasminogen and hydrolyze protamine.

Microarray analysis and draft genomes of two Escherichia coli 0157:H7 lineage II cattle isolates FRIK966 and FRIK2000 investigating lack of Shiga toxin expression

USDA-ARS?s Scientific Manuscript database

The existence of two separate lineages of Escherichia coli O157:H7 has previously been reported, and research indicates that lineage I might be more pathogenic towards human hosts than lineage II. We have previously shown that lineage I expresses higher levels of Shiga toxin 2 (Stx2). To evaluate w...

Lipocalin 2 regulates intestine bacterial survival by interplaying with siderophore in a weaned piglet model of Escherichia coli infection.

PubMed

Guo, Bing-Xiu; Wang, Qian-Qian; Li, Jia-Hui; Gan, Zhen-Shun; Zhang, Xiao-Feng; Wang, Yi-Zhen; Du, Hua-Hua

2017-09-12

Iron is an essential nutrient that facilitates cell proliferation and growth, which plays a pivotal role in modulating the battle for survival between mammalian hosts and their pathogens. Pathogenic bacteria secrete siderophores to acquire iron from the host. However, lipocalin 2 (Lcn2), a siderophore-binding antimicrobial protein, binds to siderophores to prevent bacterial uptake of iron, which is critical for the control of systemic infection with Escherichia coli ( E. coli ). But few studies focus on the anti-infective response of Lcn2 in the intestines by inhibiting bacterial proliferation based on microbial iron metabolism. In this study, we showed that iron was sequestrated within cells in a piglet model of E. coli K88 infection. Siderophores was produced following E. coli K88 infection and siderophore-related genes expression was upregulated in iron-deficiency environment in vitro . Meanwhile, we found that Lcn2 expression was rapidly and robustly induced in jejunum by E. coli K88 infection and could be stimulated by IL-17 and IL-22. Furthermore, both Lcn2 induced in epithelial cells IPEC-1 and added exogenously as a recombinant protein could inhibit the growth of E. coli . We can conclude that Lcn2 is a crucial component of mucosal immune defense against intestinal infection with E. coli K88.

Heterologous Expression, Purification and Characterization of an Oligopeptidase A from the Pathogen Leptospira interrogans.

PubMed

Anu, Prasannan V; Madanan, Madathiparambil G; Nair, Ananthakrishnan J; Nair, Gangaprasad A; Nair, Govinda Pillai M; Sudhakaran, Perumana R; Satheeshkumar, Padikara K

2018-04-01

Oligopeptidases are enzymes involved in the degradation of short peptides (generally less than 30 amino acids in size) which help pathogens evade the host defence mechanisms. Leptospira is a zoonotic pathogen and causes leptospirosis in mammals. Proteome analysis of Leptospira revealed the presence of oligopeptidase A (OpdA) among other membrane proteins. To study the role of oligopeptidase in leptospirosis, the OpdA of L. interrogans was cloned and expressed in Escherichia coli with a histidine tag (His-tag). The protein showed maximum expression at 37 °C with 0.5 mM of IPTG after 2 h of induction. Recombinant OpdA protein was purified to homogeneity using Ni-affinity chromatography. The purified OpdA showed more than 80% inhibition with a serine protease inhibitor but the activity was reduced to 30% with the cysteine protease inhibitor. The peptidase activity was increased significantly in the presence of Zn 2+ at a neutral pH. Inhibitor assay indicate the presence of more than one active sites for peptidase activity as reported with the OpdA of E. coli and Salmonella. Over-expression of OpdA in E. coli BL21 (DE3) did not cause any negative effects on normal cell growth and viability. The role of OpdA as virulence factor in Leptospira and its potential as a therapeutic and diagnostic target in leptospirosis is yet to be identified.

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.

Postgenomics Characterization of an Essential Genetic Determinant of Mammary Pathogenic Escherichia coli.

PubMed

Blum, Shlomo E; Goldstone, Robert J; Connolly, James P R; Répérant-Ferter, Maryline; Germon, Pierre; Inglis, Neil F; Krifucks, Oleg; Mathur, Shubham; Manson, Erin; Mclean, Kevin; Rainard, Pascal; Roe, Andrew J; Leitner, Gabriel; Smith, David G E

2018-04-03

Escherichia coli are major bacterial pathogens causing bovine mastitis, a disease of great economic impact on dairy production worldwide. This work aimed to study the virulence determinants of mammary pathogenic E. coli (MPEC). By whole-genome sequencing analysis of 40 MPEC and 22 environmental ("dairy-farm" E. coli [DFEC]) strains, we found that only the fec locus ( fecIRABCDE ) for ferric dicitrate uptake was present in the core genome of MPEC and that it was absent in DFEC genomes ( P < 0.05). Expression of the FecA receptor in the outer membrane was shown to be citrate dependent by mass spectrometry. FecA was overexpressed when bacteria were grown in milk. Transcription of the fecA gene and of the inner membrane transport component fecB gene was upregulated in bacteria recovered from experimental intramammary infection. The presence of the fec system was shown to affect the ability of E. coli to grow in milk. While the rate of growth in milk of fec -positive ( fec + ) DFEC was similar to that of MPEC, it was significantly lower in DFEC lacking fec Furthermore, deletion of fec reduced the rate of growth in milk of MPEC strain P4, whereas fec -transformed non-mammary gland-pathogenic DFEC strain K71 gained the phenotype of the level of growth in milk observed in MPEC. The role of fec in E. coli intramammary pathogenicity was investigated in vivo in cows, with results showing that an MPEC P4 mutant lacking fec lost its ability to induce mastitis, whereas the fec + DFEC K71 mutant was able to trigger intramammary inflammation. For the first time, a single molecular locus was shown to be crucial in MPEC pathogenicity. IMPORTANCE Bovine mastitis is the major infectious disease in dairy cows and the leading cause of economic loss to the global dairy industry, directly contributing to the price of dairy products on supermarket shelves and the financial hardships suffered by dairy farmers. Mastitis is also the leading reason for the use of antibiotics in dairy farms. Good farm management practices in many countries have dramatically reduced the incidence of contagious mastitis; however, the problems associated with the incidence of environmental mastitis caused by bacteria such as Escherichia coli have proven intractable. E. coli bacteria cause acute mastitis, which affects the health and welfare of cows and in extreme cases may be fatal. Here we show for the first time that the pathogenicity of E. coli causing mastitis in cows is highly dependent on the fecIRABCDE ferric citrate uptake system that allows the bacterium to capture iron from citrate. The Fec system is highly expressed during infection in the bovine udder and is ubiquitous in and necessary for the E. coli bacteria that cause mammary infections in cattle. These results have far-reaching implications, raising the possibility that mastitis may be controllable by targeting this system. Copyright © 2018 Blum et al.

Postgenomics Characterization of an Essential Genetic Determinant of Mammary Pathogenic Escherichia coli

PubMed Central

2018-01-01

ABSTRACT Escherichia coli are major bacterial pathogens causing bovine mastitis, a disease of great economic impact on dairy production worldwide. This work aimed to study the virulence determinants of mammary pathogenic E. coli (MPEC). By whole-genome sequencing analysis of 40 MPEC and 22 environmental (“dairy-farm” E. coli [DFEC]) strains, we found that only the fec locus (fecIRABCDE) for ferric dicitrate uptake was present in the core genome of MPEC and that it was absent in DFEC genomes (P < 0.05). Expression of the FecA receptor in the outer membrane was shown to be citrate dependent by mass spectrometry. FecA was overexpressed when bacteria were grown in milk. Transcription of the fecA gene and of the inner membrane transport component fecB gene was upregulated in bacteria recovered from experimental intramammary infection. The presence of the fec system was shown to affect the ability of E. coli to grow in milk. While the rate of growth in milk of fec-positive (fec+) DFEC was similar to that of MPEC, it was significantly lower in DFEC lacking fec. Furthermore, deletion of fec reduced the rate of growth in milk of MPEC strain P4, whereas fec-transformed non-mammary gland-pathogenic DFEC strain K71 gained the phenotype of the level of growth in milk observed in MPEC. The role of fec in E. coli intramammary pathogenicity was investigated in vivo in cows, with results showing that an MPEC P4 mutant lacking fec lost its ability to induce mastitis, whereas the fec+ DFEC K71 mutant was able to trigger intramammary inflammation. For the first time, a single molecular locus was shown to be crucial in MPEC pathogenicity. PMID:29615502

Antimicrobial activity of gamma-thionin-like soybean SE60 in E. coli and tobacco plants.

PubMed

Choi, Yeonhee; Choi, Yang Do; Lee, Jong Seob

2008-10-17

The SE60, a low molecular weight, sulfur-rich protein in soybean, is known to be homologous to wheat gamma-purothionin. To elucidate the functional role of SE60, we expressed SE60 cDNA in Escherichia coli and in tobacco plants. A single protein band was detected by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) after anti-FLAG affinity purification of the protein from transformed E. coli. While the control E. coli cells harboring pFLAG-1 showed standard growth with Isopropyl beta-d-1-thiogalactopyranoside (IPTG) induction, E. coli cells expressing the SE60 fusion protein did not grow at all, suggesting that SE60 has toxic effects on E. coli growth. Genomic integration and the expression of transgene in the transgenic tobacco plants were confirmed by Southern and Northern blot analysis, respectively. The transgenic plants demonstrated enhanced resistance against the pathogen Pseudomonas syringae. Taken together, these results strongly suggest that SE60 has antimicrobial activity and play a role in the defense mechanism in soybean plants.

Uropathogenic E. coli Exploit CEA to Promote Colonization of the Urogenital Tract Mucosa

PubMed Central

Muenzner, Petra; Kengmo Tchoupa, Arnaud; Klauser, Benedikt; Brunner, Thomas; Putze, Johannes; Dobrindt, Ulrich; Hauck, Christof R.

2016-01-01

Attachment to the host mucosa is a key step in bacterial pathogenesis. On the apical surface of epithelial cells, members of the human carcinoembryonic antigen (CEA) family are abundant glycoproteins involved in cell-cell adhesion and modulation of cell signaling. Interestingly, several gram-negative bacterial pathogens target these receptors by specialized adhesins. The prototype of a CEACAM-binding pathogen, Neisseria gonorrhoeae, utilizes colony opacity associated (Opa) proteins to engage CEA, as well as the CEA-related cell adhesion molecules CEACAM1 and CEACAM6 on human epithelial cells. By heterologous expression of neisserial Opa proteins in non-pathogenic E. coli we find that the Opa protein-CEA interaction is sufficient to alter gene expression, to increase integrin activity and to promote matrix adhesion of infected cervical carcinoma cells and immortalized vaginal epithelial cells in vitro. These CEA-triggered events translate in suppression of exfoliation and improved colonization of the urogenital tract by Opa protein-expressing E. coli in CEA-transgenic compared to wildtype mice. Interestingly, uropathogenic E. coli expressing an unrelated CEACAM-binding protein of the Afa/Dr adhesin family recapitulate the in vitro and in vivo phenotype. In contrast, an isogenic strain lacking the CEACAM-binding adhesin shows reduced colonization and does not suppress epithelial exfoliation. These results demonstrate that engagement of human CEACAMs by distinct bacterial adhesins is sufficient to blunt exfoliation and to promote host infection. Our findings provide novel insight into mucosal colonization by a common UPEC pathotype and help to explain why human CEACAMs are a preferred epithelial target structure for diverse gram-negative bacteria to establish a foothold on the human mucosa. PMID:27171273

Impact of blaNDM-1 on fitness and pathogenicity of Escherichia coli and Klebsiella pneumoniae.

PubMed

Göttig, Stephan; Riedel-Christ, Sara; Saleh, Ahmad; Kempf, Volkhard A J; Hamprecht, Axel

2016-06-01

The objective of this study was to determine whether acquisition of New Delhi metallo-β-lactamase-1 (NDM-1) has an impact on the fitness and virulence of Escherichia coli and Klebsiella pneumoniae. Growth kinetics and the cost of fitness of NDM-1 plasmid carriage were assessed in isogenic E. coli J53 and K. pneumoniae PRZ in vitro by pairwise competition assays. The pathogenicity of NDM-1-expressing E. coli and K. pneumoniae strains and their isogenic controls was analysed in vivo using a Galleria mellonella infection model. The cytotoxicity of NDM-1 was assessed in A549 human lung epithelial cells using the lactate dehydrogenase (LDH) assay. No differences in growth kinetics were recorded between NDM-1-expressing strains and controls (P = 0.92). A reduction in fitness of NDM-1-carrying strains was observed both for E. coli J53 and K. pneumoniae PRZ [selection rate constant (s) = -1.27 ± 0.27 for E. coli J53 and -0.19 ± 0.14 for K. pneumoniae PRZ; P < 0.0001]. Survival of G. mellonella larvae infected with NDM-1-expressing strains and controls was similar for E. coli J53 and K. pneumoniae PRZ. Cytotoxicity in A549 cells was not affected by NDM-1 expression (P > 0.05). The presence of blaNDM-1 did not increase the virulence or cytotoxicity of isogenic strains. However, there was a considerable cost of fitness incurred by carriage of the pNDM-1 plasmid. Interestingly, the cost of fitness was significantly higher in E. coli J53 compared with K. pneumoniae PRZ. Copyright © 2016 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

The Yersinia pseudotuberculosis and Yersinia pestis toxin complex is active against cultured mammalian cells.

PubMed

Hares, Michelle C; Hinchliffe, Stewart J; Strong, Philippa C R; Eleftherianos, Ioannis; Dowling, Andrea J; ffrench-Constant, Richard H; Waterfield, Nick

2008-11-01

The toxin complex (Tc) genes were first identified in the insect pathogen Photorhabdus luminescens and encode approximately 1 MDa protein complexes which are toxic to insect pests. Subsequent genome sequencing projects have revealed the presence of tc orthologues in a range of bacterial pathogens known to be associated with insects. Interestingly, members of the mammalian-pathogenic yersiniae have also been shown to encode Tc orthologues. Studies in Yersinia enterocolitica have shown that divergent tc loci either encode insect-active toxins or play a role in colonization of the gut in gastroenteritis models of rats. So far little is known about the activity of the Tc proteins in the other mammalian-pathogenic yersiniae. Here we present work to suggest that Tc proteins in Yersinia pseudotuberculosis and Yersinia pestis are not insecticidal toxins but have evolved for mammalian pathogenicity. We show that Tc is secreted by Y. pseudotuberculosis strain IP32953 during growth in media at 28 degrees C and 37 degrees C. We also demonstrate that oral toxicity of strain IP32953 to Manduca sexta larvae is not due to Tc expression and that lysates of Escherichia coli BL21 expressing the Yersinia Tc proteins are not toxic to Sf9 insect cells but are toxic to cultured mammalian cell lines. Cell lysates of E. coli BL21 expressing the Y. pseudotuberculosis Tc proteins caused actin ruffles, vacuoles and multi-nucleation in cultured human gut cells (Caco-2); similar morphology was observed after application of a lysate of E. coli BL21 expressing the Y. pestis Tc proteins to mouse fibroblast NIH3T3 cells, but not Caco-2 cells. Finally, transient expression of the individual Tc proteins in Caco-2 and NIH3T3 cell lines reproduced the actin and nuclear rearrangement observed with the topical applications. Together these results add weight to the growing hypothesis that the Tc proteins in Y. pseudotuberculosis and Y. pestis have been adapted for mammalian pathogenicity. We further conclude that Tc proteins from Y. pseudotuberculosis and Y. pestis display differential mammalian cell specificity in their toxicity.

Injury and mechanism of recombinant E. coli expressing STa on piglets colon.

PubMed

Lv, Yang; Li, Xueni; Zhang, Lin; Shi, Yutao; DU, Linxiao; Ding, Binying; Hou, Yongqing; Gong, Joshua; Wu, Tao

2018-02-09

Enterotoxigenic Escherichia coli (ETEC) is primary pathogenic bacteria of piglet diarrhea, over two thirds of piglets diarrhea caused by ETEC are resulted from STa-producing ETEC strains. This experiment was conducted to construct the recombinant E. coli expressing STa and study the injury and mechanism of recombinant E. coli expressing STa on 7 days old piglets colon. Twenty-four 7 days old piglets were allotted to four treatments: control group, STa group (2 × 10 9 CFU E. coli LMG194-STa), LMG194 group (2 × 10 9 CFU E. coli LMG194) and K88 group (2 × 10 9 CFU E. coli K88). The result showed that E. coli infection significantly increased diarrhea rates; changed DAO activity in plasma and colon; damaged colonic mucosal morphology including crypt depth, number of globet cells, density of lymphocytes and lamina propria cell density; substantially reduced antioxidant capacity by altering activities of GSH-Px, SOD, and TNOS and productions of MDA and H 2 O 2 ; obviously decreased AQP3, AQP4 and KCNJ13 protein expression levels; substantially altered the gene expression levels of inflammatory cytokines. Conclusively, STa group had the biggest effect on these indices in four treatment groups. These results suggested that the recombinant strain expressed STa can induce piglets diarrhea and colonic morphological and funtional damage by altering expression of proteins connect to transportation function and genes associated with intestinal injury and inflammatory cytokines.

Injury and mechanism of recombinant E. coli expressing STa on piglets colon

PubMed Central

LV, Yang; LI, Xueni; ZHANG, Lin; SHI, Yutao; DU, Linxiao; DING, Binying; HOU, Yongqing; GONG, Joshua; WU, Tao

2017-01-01

Enterotoxigenic Escherichia coli (ETEC) is primary pathogenic bacteria of piglet diarrhea, over two thirds of piglets diarrhea caused by ETEC are resulted from STa-producing ETEC strains. This experiment was conducted to construct the recombinant E. coli expressing STa and study the injury and mechanism of recombinant E. coli expressing STa on 7 days old piglets colon. Twenty-four 7 days old piglets were allotted to four treatments: control group, STa group (2 × 109 CFU E. coli LMG194-STa), LMG194 group (2 × 109 CFU E. coli LMG194) and K88 group (2 × 109 CFU E. coli K88). The result showed that E. coli infection significantly increased diarrhea rates; changed DAO activity in plasma and colon; damaged colonic mucosal morphology including crypt depth, number of globet cells, density of lymphocytes and lamina propria cell density; substantially reduced antioxidant capacity by altering activities of GSH-Px, SOD, and TNOS and productions of MDA and H2O2; obviously decreased AQP3, AQP4 and KCNJ13 protein expression levels; substantially altered the gene expression levels of inflammatory cytokines. Conclusively, STa group had the biggest effect on these indices in four treatment groups. These results suggested that the recombinant strain expressed STa can induce piglets diarrhea and colonic morphological and funtional damage by altering expression of proteins connect to transportation function and genes associated with intestinal injury and inflammatory cytokines. PMID:29187713

The CpxRA two-component system is essential for Citrobacter rodentium virulence.

PubMed

Thomassin, Jenny-Lee; Giannakopoulou, Natalia; Zhu, Lei; Gross, Jeremy; Salmon, Kristiana; Leclerc, Jean-Mathieu; Daigle, France; Le Moual, Hervé; Gruenheid, Samantha

2015-05-01

Citrobacter rodentium is a murine intestinal pathogen used as a model for the foodborne human pathogens enterohemorrhagic Escherichia coli and enteropathogenic E. coli. During infection, these pathogens use two-component signal transduction systems to detect and adapt to changing environmental conditions. In E. coli, the CpxRA two-component signal transduction system responds to envelope stress by modulating the expression of a myriad of genes. Quantitative real-time PCR showed that cpxRA was expressed in the colon of C57BL/6J mice infected with C. rodentium. To determine whether CpxRA plays a role during C. rodentium infection, a cpxRA deletion strain was generated and found to have a colonization defect during infection. This defect was independent of an altered growth rate or a defective type III secretion system, and single-copy chromosomal complementation of cpxRA restored virulence. The C. rodentium strains were then tested in C3H/HeJ mice, a lethal intestinal infection model. Mice infected with the ΔcpxRA strain survived infection, whereas mice infected with the wild-type or complemented strains succumbed to infection. Furthermore, we found that the cpxRA expression level was higher during early infection than at a later time point. Taken together, these data demonstrate that the CpxRA two-component signal transduction system is essential for the in vivo virulence of C. rodentium. In addition, these data suggest that fine-tuned cpxRA expression is important for infection. This is the first study that identifies a C. rodentium two-component transduction system required for pathogenesis. This study further indicates that CpxRA is an interesting target for therapeutics against enteric pathogens. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Regulation of nrf operon expression in pathogenic enteric bacteria: sequence divergence reveals new regulatory complexity

PubMed Central

Godfrey, Rita E.; Lee, David J.; Busby, Stephen J. W.

2017-01-01

Summary The Escherichia coli K‐12 nrf operon encodes a periplasmic nitrite reductase, the expression of which is driven from a single promoter, pnrf. Expression from pnrf is activated by the FNR transcription factor in response to anaerobiosis and further increased in response to nitrite by the response regulator proteins, NarL and NarP. FNR‐dependent transcription is suppressed by the binding of two nucleoid associated proteins, IHF and Fis. As Fis levels increase in cells grown in rich medium, the positioning of its binding site, overlapping the promoter −10 element, ensures that pnrf is sharply repressed. Here, we investigate the expression of the nrf operon promoter from various pathogenic enteric bacteria. We show that pnrf from enterohaemorrhagic E. coli is more active than its K‐12 counterpart, exhibits substantial FNR‐independent activity and is insensitive to nutrient quality, due to an improved −10 element. We also demonstrate that the Salmonella enterica serovar Typhimurium core promoter is more active than previously thought, due to differences around the transcription start site, and that its expression is repressed by downstream sequences. We identify the CsrA RNA binding protein as being responsible for this, and show that CsrA differentially regulates the E. coli K‐12 and Salmonella nrf operons. PMID:28211111

FNR Regulates the Expression of Important Virulence Factors Contributing to the Pathogenicity of Avian Pathogenic Escherichia coli

PubMed Central

Barbieri, Nicolle L.; Vande Vorde, Jessica A.; Baker, Alison R.; Horn, Fabiana; Li, Ganwu; Logue, Catherine M.; Nolan, Lisa K.

2017-01-01

Avian pathogenic Escherichia coli (APEC) is the etiologic agent of colibacillosis, an important cause of morbidity and mortality in poultry. Though, many virulence factors associated with APEC pathogenicity are known, their regulation remains unclear. FNR (fumarate and nitrate reduction) is a well-known global regulator that works as an oxygen sensor and has previously been described as a virulence regulator in bacterial pathogens. The goal of this study was to examine the role of FNR in the regulation of APEC virulence factors, such as Type I fimbriae, and processes such as adherence and invasion, type VI secretion, survival during oxidative stress, and growth in iron-restricted environments. To accomplish this goal, APEC O1, a well-characterized, highly virulent, and fully sequenced strain of APEC harboring multiple virulence mechanisms, some of which are plasmid-linked, was compared to its FNR mutant for expression of various virulence traits. Deletion of FNR was found to affect APEC O1's adherence, invasion and expression of ompT, a plasmid-encoded outer membrane protein, type I fimbriae, and aatA, encoding an autotransporter. Indeed, the fnr− mutant showed an 8-fold reduction in expression of type I fimbriae and a highly significant (P < 0.0001) reduction in expression of fimA, ompT (plasmid-borne), and aatA. FNR was also found to regulate expression of the type VI secretion system, affecting the expression of vgrG. Further, FNR was found to be important to APEC O1's growth in iron-deficient media and survival during oxidative stress with the mutant showing a 4-fold decrease in tolerance to oxidative stress, as compared to the wild type. Thus, our results suggest that FNR functions as an important regulator of APEC virulence. PMID:28690981

DNA microarray-mediated transcriptional profiling of avian pathogenic Escherichia coli O2 strain E058 during its infection of chicken.

PubMed

Gao, Qingqing; Xia, Le; Liu, Juanhua; Wang, Xiaobo; Gao, Song; Liu, Xiufan

2016-11-01

Avian pathogenic Escherichia coli (APEC) cause typical extraintestinal infections in poultry, including acute fatal septicemia, subacute pericarditis, and airsacculitis. These bacteria most often infect chickens, turkeys, ducks, and other avian species, and therefore pose a significant economic burden on the poultry industry worldwide. Few studies have analyzed the genome-wide transcriptional profile of APEC during infection in vivo. In this study, we examined the genome-wide transcriptional response of APEC O2 strain E058 in an in vivo chicken infection model to better understand the factors necessary for APEC colonization, growth, and survival in vivo. An Affymetrix multigenome DNA microarray, which contains most of the genomic open reading frames of E. coli K-12 strain MG1655, uropathogenic E. coli strain CFT073, and E. coli O157:H7 strain EDL 933, was used to profile the gene expression in APEC E058. We identified the in vivo transcriptional response of APEC E058 bacteria collected directly from the blood of infected chickens. Significant differences in expression levels were detected between the in vivo expression profile and the in vitro expression profile in LB medium. The genes highly expressed during infection were involved in metabolism, iron acquisition or transport, virulence, response to stress, and biological regulation. The reliability of the microarray data was confirmed by performing quantitative real-time PCR on 12 representative genes. Moreover, several significantly upregulated genes, including yjiY, sodA, phoB and spy, were selected to study their role in APEC pathogenesis. The data will help to better understand the mechanisms of APEC pathogenesis. Copyright © 2016 Elsevier Ltd. All rights reserved.

Differential cytokine expression in Chlamydophila psittaci genotype A-, B- or D-infected chicken macrophages after exposure to Escherichia coli O2:K1 LPS.

PubMed

Beeckman, Delphine Sylvie Anne; Rothwell, Lisa; Kaiser, Pete; Vanrompay, Daisy C G

2010-08-01

Chlamydophila (Cp.) psittaci and avian pathogenic Escherichia (E.) coli infections contribute to the respiratory disease complex observed in turkeys. Secondary infection with E. coli exacerbates Cp. psittaci pathogenicity and augments E. coli excretion. The innate immune response initiated by both pathogens in their avian host is unknown. We therefore determined the cytokine responses following Cp. psittaci infection and E. coli superinfection of avian monocytes/macrophages by examining gene transcripts of IL-1beta, IL-6, CXCLi2 (IL-8), CXCLi1 (K60), IL-10, IL-12alpha/beta, IL-18, TGF-beta4 and CCLi2 at 4h post-inoculation with different Cp. psittaci strains or 4h post-treatment with avian E. coli LPS of Cp. psittaci pre-infected HD11 cells. Cp. psittaci strains used were 84/55 and 92/1293 (highly virulent), CP3 (low virulent) and 84/2334 (phylogenetically intermediate between Cp. psittaci and Chlamydophila abortus). At 4h post chlamydial infection, an increased expression of IL-1beta and IL-6 as well as CXCLi2, CXCLi1 and CCLi2 was observed compared to levels in uninfected HD11 controls. This effect was less pronounced for the milder CP3 strain. The pro-inflammatory response of Cp. psittaci infected cells to E. coli LPS was significantly lowered compared to uninfected controls, especially when the cells were pre-infected with highly virulent Cp. psittaci strains. In both experiments, exceptionally high IL-10 and no TGF-beta4 responses were observed, and we propose that this could induce macrophage deactivation and NF-kappaB suppression. Consequently, pro-inflammatory and Th1-promoting responses to both the primary Cp. psittaci infection and E. coli would be inhibited, thus explaining the observed aggravated in vivo pathology. Copyright 2010 Elsevier Ltd. All rights reserved.

Hha Represses Biofilm Formation in Escherichia coli O157:H7 by Affecting the Expression of Flagella and Curli Fimbriae

USDA-ARS?s Scientific Manuscript database

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a zoonotic pathogen that produces a broad-spectrum of diarrheal illnesses in infected humans. Although the genetic and molecular mechanisms enabling EHEC O157:H7 to produce characteristic adherence on epithelial cells are well characterized, the g...

[Avian Escherichia coli virulence factors associated with coli septicemia in broiler chickens].

PubMed

Ramirez Santoyo, R M; Moreno Sala, A; Almanza Marquez, Y

2001-01-01

In order to detect phenotypic characteristics associated with pathogenicity, 25 strains of Escherichia coli, isolated from clinical cases of colisepticemia in broiler chickens, were examined to determine the following properties: colicinogenicity, colicin V production, type 1 fimbriae, hemolysin expression and motility. Colicinogenicity occurred in 72% of the strains, 56% of all strains produced colicin V, 84% were positive for type 1 fimbriae and 80% were positive for motility. None of the strains had hemolytic activity; however, all of them, expressed at least one of the other characteristics studied. These results suggest that the diversity of phenotypes detected partially explain the multifactorial nature of avian colisepticemia.

A new sulfonamide resistance gene (sul3) in Escherichia coli is widespread in the pig population of Switzerland.

PubMed

Perreten, Vincent; Boerlin, Patrick

2003-03-01

A new gene, sul3, which specifies a 263-amino-acid protein similar to a dihydropteroate synthase encoded by the 54-kb conjugative plasmid pVP440 from Escherichia coli was characterized. Expression of the cloned sul3 gene conferred resistance to sulfamethoxazole on E. coli. Two copies of the insertion element IS15Delta/26 flanked the region containing sul3. The sul3 gene was detected in one-third of the sulfonamide-resistant pathogenic E. coli isolates from pigs in Switzerland.

A New Sulfonamide Resistance Gene (sul3) in Escherichia coli Is Widespread in the Pig Population of Switzerland

PubMed Central

Perreten, Vincent; Boerlin, Patrick

2003-01-01

A new gene, sul3, which specifies a 263-amino-acid protein similar to a dihydropteroate synthase encoded by the 54-kb conjugative plasmid pVP440 from Escherichia coli was characterized. Expression of the cloned sul3 gene conferred resistance to sulfamethoxazole on E. coli. Two copies of the insertion element IS15Δ/26 flanked the region containing sul3. The sul3 gene was detected in one-third of the sulfonamide-resistant pathogenic E. coli isolates from pigs in Switzerland. PMID:12604565

Drosophila Embryos as Model Systems for Monitoring Bacterial Infection in Real Time

PubMed Central

Evans, Iwan R.; Waterfield, Nicholas; ffrench-Constant, Richard H.; Wood, Will

2009-01-01

Drosophila embryos are well studied developmental microcosms that have been used extensively as models for early development and more recently wound repair. Here we extend this work by looking at embryos as model systems for following bacterial infection in real time. We examine the behaviour of injected pathogenic (Photorhabdus asymbiotica) and non-pathogenic (Escherichia coli) bacteria and their interaction with embryonic hemocytes using time-lapse confocal microscopy. We find that embryonic hemocytes both recognise and phagocytose injected wild type, non-pathogenic E. coli in a Dscam independent manner, proving that embryonic hemocytes are phagocytically competent. In contrast, injection of bacterial cells of the insect pathogen Photorhabdus leads to a rapid ‘freezing’ phenotype of the hemocytes associated with significant rearrangement of the actin cytoskeleton. This freezing phenotype can be phenocopied by either injection of the purified insecticidal toxin Makes Caterpillars Floppy 1 (Mcf1) or by recombinant E. coli expressing the mcf1 gene. Mcf1 mediated hemocyte freezing is shibire dependent, suggesting that endocytosis is required for Mcf1 toxicity and can be modulated by dominant negative or constitutively active Rac expression, suggesting early and unexpected effects of Mcf1 on the actin cytoskeleton. Together these data show how Drosophila embryos can be used to track bacterial infection in real time and how mutant analysis can be used to genetically dissect the effects of specific bacterial virulence factors. PMID:19609447

Computational Analysis of Host-Pathogen Protein Interactions between Humans and Different Strains of Enterohemorrhagic Escherichia coli.

PubMed

Bose, Tungadri; Venkatesh, K V; Mande, Sharmila S

2017-01-01

Serotype O157:H7, an enterohemorrhagic Escherichia coli (EHEC), is known to cause gastrointestinal and systemic illnesses ranging from diarrhea and hemorrhagic colitis to potentially fatal hemolytic uremic syndrome. Specific genetic factors like ompA, nsrR , and LEE genes are known to play roles in EHEC pathogenesis. However, these factors are not specific to EHEC and their presence in several non-pathogenic strains indicates that additional factors are involved in pathogenicity. We propose a comprehensive effort to screen for such potential genetic elements, through investigation of biomolecular interactions between E. coli and their host. In this work, an in silico investigation of the protein-protein interactions (PPIs) between human cells and four EHEC strains (viz., EDL933, Sakai, EC4115, and TW14359) was performed in order to understand the virulence and host-colonization strategies of these strains. Potential host-pathogen interactions (HPIs) between human cells and the "non-pathogenic" E. coli strain MG1655 were also probed to evaluate whether and how the variations in the genomes could translate into altered virulence and host-colonization capabilities of the studied bacterial strains. Results indicate that a small subset of HPIs are unique to the studied pathogens and can be implicated in virulence. This subset of interactions involved E. coli proteins like YhdW, ChuT, EivG, and HlyA. These proteins have previously been reported to be involved in bacterial virulence. In addition, clear differences in lineage and clade-specific HPI profiles could be identified. Furthermore, available gene expression profiles of the HPI-proteins were utilized to estimate the proportion of proteins which may be involved in interactions. We hypothesized that a cumulative score of the ratios of bound:unbound proteins (involved in HPIs) would indicate the extent of colonization. Thus, we designed the Host Colonization Index (HCI) measure to determine the host colonization potential of the E. coli strains. Pathogenic strains of E. coli were observed to have higher HCIs as compared to a non-pathogenic laboratory strain. However, no significant differences among the HCIs of the two pathogenic groups were observed. Overall, our findings are expected to provide additional insights into EHEC pathogenesis and are likely to aid in designing alternate preventive and therapeutic strategies.

Impact of a short-term exposure to spaceflight on the phenotype, genome, transcriptome and proteome of Escherichia coli

NASA Astrophysics Data System (ADS)

Li, Tianzhi; Chang, De; Xu, Huiwen; Chen, Jiapeng; Su, Longxiang; Guo, Yinghua; Chen, Zhenhong; Wang, Yajuan; Wang, Li; Wang, Junfeng; Fang, Xiangqun; Liu, Changting

2015-07-01

Escherichia coli (E. coli) is the most widely applied model organism in current biological science. As a widespread opportunistic pathogen, E. coli can survive not only by symbiosis with human, but also outside the host as well, which necessitates the evaluation of its response to the space environment. Therefore, to keep humans safe in space, it is necessary to understand how the bacteria respond to this environment. Despite extensive investigations for a few decades, the response of E. coli to the real space environment is still controversial. To better understand the mechanisms how E. coli overcomes harsh environments such as microgravity in space and to investigate whether these factors may induce pathogenic changes in E. coli that are potentially detrimental to astronauts, we conducted detailed genomics, transcriptomic and proteomic studies on E. coli that experienced 17 days of spaceflight. By comparing two flight strains LCT-EC52 and LCT-EC59 to a control strain LCT-EC106 that was cultured under the same temperature conditions on the ground, we identified metabolism changes, polymorphism changes, differentially expressed genes and proteins in the two flight strains. The flight strains differed from the control in the utilization of more than 30 carbon sources. Two single nucleotide polymorphisms (SNPs) and one deletion were identified in the flight strains. The expression level of more than 1000 genes altered in flight strains. Genes involved in chemotaxis, lipid metabolism and cell motility express differently. Moreover, the two flight strains also differed extensively from each other in terms of metabolism, transcriptome and proteome, indicating the impact of space environment on individual cells is heterogeneous and probably genotype-dependent. This study presents the first systematic profile of E. coli genome, transcriptome and proteome after spaceflight, which helps to elucidate the mechanism that controls the adaptation of microbes to the space environment.

O-acetyltransferase gene neuO is segregated according to phylogenetic background and contributes to environmental desiccation resistance in Escherichia coli K1.

PubMed

Mordhorst, Ines L; Claus, Heike; Ewers, Christa; Lappann, Martin; Schoen, Christoph; Elias, Johannes; Batzilla, Julia; Dobrindt, Ulrich; Wieler, Lothar H; Bergfeld, Anne K; Mühlenhoff, Martina; Vogel, Ulrich

2009-12-01

Escherichia coli K1 causes disease in humans and birds. Its polysialic acid capsule can be O-acetylated via phase-variable expression of the acetyltransferase NeuO encoded by prophage CUS-3. The role of capsule O-acetylation in ecological adaptation or pathogenic invasion of E. coli K1 is largely unclear. A population genetics approach was performed to study the distribution of neuO among E. coli K1 isolates from human and avian sources. Multilocus sequence typing revealed 39 different sequence types (STs) among 183 E. coli K1 strains. The proportion of the ST95 complex (STC95) was 44%. NeuO was found in 98% of the STC95 strains, but only in 24% of other STs. Grouping of STs and prophage genotypes revealed a segregation of prophage types according to STs, suggesting coevolution of CUS-3 and the E. coli K1 host. Within the STC95, which is known to harbour both human and avian pathogenic isolates, CUS-3 genotypes were shared irrespective of the host species. Functional analysis of a variety of strain pairs revealed that NeuO-mediated K1 capsule O-acetylation enhanced desiccation resistance. In contrast, NeuO expression led to a reduced biofilm formation in biofilm positive E. coli K1 isolates. These findings suggest a delicate ecological balance of neuO'on'/'off' switching.

Escherichia coli K-12 pathogenicity in the pea aphid, Acyrthosiphon pisum, reveals reduced antibacterial defense in aphids.

PubMed

Altincicek, Boran; Ter Braak, Bas; Laughton, Alice M; Udekwu, Klas I; Gerardo, Nicole M

2011-10-01

To better understand the molecular basis underlying aphid immune tolerance to beneficial bacteria and immune defense to pathogenic bacteria, we characterized how the pea aphid Acyrthosiphon pisum responds to Escherichia coli K-12 infections. E. coli bacteria, usually cleared in the hemolymph of other insect species, were capable of growing exponentially and killing aphids within a few days. Red fluorescence protein expressing E. coli K-12 laboratory strain multiplied in the aphid hemolymph as well as in the digestive tract, resulting in death of infected aphids. Selected gene deletion mutants of the E. coli K-12 predicted to have reduced virulence during systemic infections showed no difference in either replication or killing rate when compared to the wild type E. coli strain. Of note, however, the XL1-Blue E. coli K-12 strain exhibited a significant lag phase before multiplying and killing aphids. This bacterial strain has recently been shown to be more sensitive to oxidative stress than other E. coli K-12 strains, revealing a potential role for reactive oxygen species-mediated defenses in the otherwise reduced aphid immune system. Copyright © 2011 Elsevier Ltd. All rights reserved.

HlSRB, a Class B Scavenger Receptor, Is Key to the Granulocyte-Mediated Microbial Phagocytosis in Ticks

PubMed Central

Aung, Kyaw Min; Boldbaatar, Damdinsuren; Umemiya-Shirafuji, Rika; Liao, Min; Tsuji, Naotoshi; Xuenan, Xuan; Suzuki, Hiroshi; Kume, Aiko; Galay, Remil Linggatong; Tanaka, Tetsuya; Fujisaki, Kozo

2012-01-01

Ixodid ticks transmit various pathogens of deadly diseases to humans and animals. However, the specific molecule that functions in the recognition and control of pathogens inside ticks is not yet to be identified. Class B scavenger receptor CD36 (SRB) participates in internalization of apoptotic cells, certain bacterial and fungal pathogens, and modified low-density lipoproteins. Recently, we have reported on recombinant HlSRB, a 50-kDa protein with one hydrophobic SRB domain from the hard tick, Haemaphysalis longicornis. Here, we show that HlSRB plays vital roles in granulocyte-mediated phagocytosis to invading Escherichia coli and contributes to the first-line host defense against various pathogens. Data clearly revealed that granulocytes that up-regulated the expression of cell surface HlSRB are almost exclusively involved in hemocyte-mediated phagocytosis for E. coli in ticks, and post-transcriptional silencing of the HlSRB-specific gene ablated the granulocytes' ability to phagocytose E. coli and resulted in the mortality of ticks due to high bacteremia. This is the first report demonstrating that a scavenger receptor molecule contributes to hemocyte-mediated phagocytosis against exogenous pathogens, isolated and characterized from hematophagous arthropods. PMID:22479406

Tracking contamination through ground beef production and identifying points of recontamination using a novel green fluorescent protein (GFP) expressing, E. coli O103, non-pathogenic surrogate

USDA-ARS?s Scientific Manuscript database

Introduction: Commonly, ground beef processors conduct studies to model contaminant flow through their production systems using surrogate organisms. Typical surrogate organisms may not behave as Escherichia coli O157:H7 during grinding and are not easy to detect at very low levels. Purpose: Develop...

Epithelial and Mesenchymal Cells in the Bovine Colonic Mucosa Differ in Their Responsiveness to Escherichia coli Shiga Toxin 1

USDA-ARS?s Scientific Manuscript database

Cells in the depth of the crypts in the bovine colon express CD77 molecules that potentially act as receptors for Shiga toxins (Stx). The implication of this finding for the intestinal colonization 25 of cattle with human pathogenic Stx-producing Escherichia coli (STEC) remains undefined. We used f...

Epigenetic Regulation of the Nitrosative Stress Response and Intracellular Macrophage Survival by Extraintestinal Pathogenic Escherichia coli

PubMed Central

Bateman, Stacey L.; Seed, Patrick C.

2013-01-01

Summary Extraintestinal pathogenic Escherichia coli (ExPEC) reside in the enteric tract as a commensal reservoir, but can transition to a pathogenic state by invading normally sterile niches, establishing infection, and disseminating to invasive sites like the bloodstream. Macrophages are required for ExPEC dissemination, suggesting the pathogen has developed mechanisms to persist within professional phagocytes. Here, we report that FimX, an ExPEC-associated DNA invertase that regulates the major virulence factor type 1 pili (T1P), is also an epigenetic regulator of a LuxR-like response regulator HyxR. FimX regulated hyxR expression through bidirectional phase inversion of its promoter region at sites different from the type 1 pili promoter and independent of integration host factor IHF. In vitro, transition from high to low HyxR expression produced enhanced tolerance of reactive nitrogen intermediates (RNI), primarily through de-repression of hmpA, encoding a nitric oxide detoxifying flavohemoglobin. However, in the macrophage, HyxR produced large effects on intracellular survival in the presence and absence of RNI and independent of Hmp. Collectively, we have shown that the ability of ExPEC to survive in macrophages is contingent upon the proper transition from high to low HyxR expression through epigenetic regulatory control by FimX. PMID:22221182

Engineering Escherichia coli into a protein delivery system for mammalian cells.

PubMed

Reeves, Analise Z; Spears, William E; Du, Juan; Tan, Kah Yong; Wagers, Amy J; Lesser, Cammie F

2015-05-15

Many Gram-negative pathogens encode type 3 secretion systems, sophisticated nanomachines that deliver proteins directly into the cytoplasm of mammalian cells. These systems present attractive opportunities for therapeutic protein delivery applications; however, their utility has been limited by their inherent pathogenicity. Here, we report the reengineering of a laboratory strain of Escherichia coli with a tunable type 3 secretion system that can efficiently deliver heterologous proteins into mammalian cells, thereby circumventing the need for virulence attenuation. We first introduced a 31 kB region of Shigella flexneri DNA that encodes all of the information needed to form the secretion nanomachine onto a plasmid that can be directly propagated within E. coli or integrated into the E. coli chromosome. To provide flexible control over type 3 secretion and protein delivery, we generated plasmids expressing master regulators of the type 3 system from either constitutive or inducible promoters. We then constructed a Gateway-compatible plasmid library of type 3 secretion sequences to enable rapid screening and identification of sequences that do not perturb function when fused to heterologous protein substrates and optimized their delivery into mammalian cells. Combining these elements, we found that coordinated expression of the type 3 secretion system and modified target protein substrates produces a nonpathogenic strain that expresses, secretes, and delivers heterologous proteins into mammalian cells. This reengineered system thus provides a highly flexible protein delivery platform with potential for future therapeutic applications.

Development of genetic techniques for the psychrotrophic fish pathogen Flavobacterium psychrophilum.

PubMed

Alvarez, B; Secades, P; McBride, M J; Guijarro, J A

2004-01-01

Flavobacterium psychrophilum, a member of the Cytophaga-Flavobacterium-Bacteroides group, is an important pathogen of salmonid fish. Previous attempts to develop genetic techniques for this fastidious, psychrotrophic bacterium have met with failure. Here we describe the development of techniques for the genetic manipulation of F. psychrophilum and the identification of plasmids, selectable markers, a reporter system, and a transposon that function in several isolates of this fish pathogen. The antibiotic resistance genes ermF, cfxA, and tetQ function in F. psychrophilum. Cloning vectors based on the F. psychrophilum cryptic plasmid pCP1 which carried these selectable markers were introduced by conjugation from E. coli, resulting in antibiotic-resistant colonies of F. psychrophilum. Conjugative transfer of DNA into F. psychrophilum was strain dependent. Efficient transfer was observed for two of the seven strains tested (THC02-90 and THC04-90). E. coli lacZY functioned in F. psychrophilum when expressed from a pCP1 promoter, allowing its development as a reporter for studies of gene expression. Plasmids isolated from F. psychrophilum were efficiently introduced into F. psychrophilum by electroporation, but plasmids isolated from E. coli were not suitable for transfer by this route, suggesting the presence of a restriction barrier. DNA isolated from F. psychrophilum was resistant to digestion by Sau3AI and BamHI, indicating that a Sau3AI-like restriction modification system may constitute part of this barrier. Tn4351 was introduced into F. psychrophilum from E. coli and transposed with apparent randomness, resulting in erythromycin-resistant colonies. The techniques developed in this study allow for genetic manipulation and analysis of this important fish pathogen.

Colonization by non-pathogenic bacteria alters mRNA expression of cytochromes P450 in originally germ-free mice.

PubMed

Jourová, L; Anzenbacher, P; Lišková, B; Matušková, Z; Hermanová, P; Hudcovic, T; Kozáková, H; Hrnčířová, L; Anzenbacherová, E

2017-11-01

Gut microbiota provides a wide range of beneficial function for the host and has an immense effect on the host's health state. It has also been shown that gut microbiome is often involved in the biotransformation of xenobiotics; however, the molecular mechanisms of the interaction between the gut bacteria and the metabolism of drugs by the host are still unclear. To investigate the effect of microbial colonization on messenger RNA (mRNA) expression of liver cytochromes P450 (CYPs), the main drug-metabolizing enzymes, we used germ-free (GF) mice, lacking the intestinal flora and mice monocolonized by non-pathogenic bacteria Lactobacillus plantarum NIZO2877 or probiotic bacteria Escherichia coli Nissle 1917 compared to specific pathogen-free (SPF) mice. Our results show that the mRNA expression of Cyp1a2 and Cyp2e1 was significantly increased, while the expression of Cyp3a11 mRNA was decreased under GF conditions compared to the SPF mice. The both bacteria L. plantarum NIZO2877 and E. coli Nissle 1917 given to the GF mice decreased the level of Cyp1a2 mRNA and normalized it to the control level. On the other hand, the colonization by these bacteria had no effect on the expression of Cyp3a11 mRNA in the liver of the GF mice (which remained decreased). Surprisingly, monocolonization with chosen bacterial strains has shown a different effect on the expression of Cyp2e1 mRNA in GF mice. Increased level of Cyp2e1 expression observed in the GF mice was found also in mice colonized by L. plantarum NIZO2877 ; however, the colonization with probiotic E. coli Nissle 1917 caused a decrease in Cyp2e1 expression and partially restored the SPF mice conditions.

Characterization of Stg Fimbriae from an Avian Pathogenic Escherichia coli O78:K80 Strain and Assessment of Their Contribution to Colonization of the Chicken Respiratory Tract

PubMed Central

Lymberopoulos, Maria H.; Houle, Sébastien; Daigle, France; Léveillé, Simon; Brée, Annie; Moulin-Schouleur, Maryvonne; Johnson, James R.; Dozois, Charles M.

2006-01-01

In a previous study, ecs-3, a sequence from avian pathogenic Escherichia coli (APEC) O78:K80 strain χ7122, was found to be expressed in vivo in infected chicken tissues. The region encompassing ecs-3 carries a fimbrial gene cluster that is a putative ortholog of the stg fimbrial gene cluster of Salmonella enterica serovar Typhi. This APEC fimbrial gene cluster, which we have termed stg, is a member of a distinct group of related fimbriae that are located in the glmS-pstS intergenic region of certain E. coli and S. enterica strains. Under the control of the pBAD promoter, the production of Stg fimbriae was demonstrated by Western blotting and immunogold electron microscopy with E. coli K-12. Transcriptional fusions suggest that stg expression is influenced by the carbohydrate source and decreased by the addition of iron and that Fur plays a role in the regulation of stg expression. stg sequences were associated with APEC O78 isolates, and stg was phylogenetically distributed among E. coli reference strains and clinical isolates from human urinary tract infections. Stg fimbriae contributed to the adherence of a nonfimbriated E. coli K-12 strain to avian lung sections and human epithelial cells in vitro. Coinfection experiments with APEC strain χ7122 and an isogenic Δstg mutant demonstrated that compared to the wild-type parent, the Δstg mutant was less able to colonize air sacs, equally able to colonize lungs, and able to more effectively colonize tracheas of infected chickens. Stg fimbriae, together with other adhesins, may therefore contribute to the colonization of avian respiratory tissues by certain APEC strains. PMID:16952934

Some Like It Hot: Heat Resistance of Escherichia coli in Food

PubMed Central

Li, Hui; Gänzle, Michael

2016-01-01

Heat treatment and cooking are common interventions for reducing the numbers of vegetative cells and eliminating pathogenic microorganisms in food. Current cooking method requires the internal temperature of beef patties to reach 71°C. However, some pathogenic Escherichia coli such as the beef isolate E. coli AW 1.7 are extremely heat resistant, questioning its inactivation by current heat interventions in beef processing. To optimize the conditions of heat treatment for effective decontaminations of pathogenic E. coli strains, sufficient estimations, and explanations are necessary on mechanisms of heat resistance of target strains. The heat resistance of E. coli depends on the variability of strains and properties of food formulations including salt and water activity. Heat induces alterations of E. coli cells including membrane, cytoplasm, ribosome and DNA, particularly on proteins including protein misfolding and aggregations. Resistant systems of E. coli act against these alterations, mainly through gene regulations of heat response including EvgA, heat shock proteins, σE and σS, to re-fold of misfolded proteins, and achieve antagonism to heat stress. Heat resistance can also be increased by expression of key proteins of membrane and stabilization of membrane fluidity. In addition to the contributions of the outer membrane porin NmpC and overcome of osmotic stress from compatible solutes, the new identified genomic island locus of heat resistant performs a critical role to these highly heat resistant strains. This review aims to provide an overview of current knowledge on heat resistance of E. coli, to better understand its related mechanisms and explore more effective applications of heat interventions in food industry. PMID:27857712

Secreted Gaussia princeps luciferase as a reporter of Escherichia coli replication in a mouse tissue cage model of infection.

PubMed

Liu, Mingyu; Blinn, Christina; McLeod, Sarah M; Wiseman, John W; Newman, Joseph V; Fisher, Stewart L; Walkup, Grant K

2014-01-01

Measurement of bacterial burden in animal infection models is a key component for both bacterial pathogenesis studies and therapeutic agent research. The traditional quantification means for in vivo bacterial burden requires frequent animal sacrifice and enumerating colony forming units (CFU) recovered from infection loci. To address these issues, researchers have developed a variety of luciferase-expressing bacterial reporter strains to enable bacterial detection in living animals. To date, all such luciferase-based bacterial reporters are in cell-associated form. Production of luciferase-secreting recombinant bacteria could provide the advantage of reporting CFU from both infection loci themselves and remote sampling (eg. body fluid and plasma). Toward this end, we have genetically manipulated a pathogenic Escherichia coli (E. coli) strain, ATCC25922, to secrete the marine copepod Gaussia princeps luciferase (Gluc), and assessed the use of Gluc as both an in situ and ex situ reporter for bacterial burden in mouse tissue cage infections. The E. coli expressing Gluc demonstrates in vivo imaging of bacteria in a tissue cage model of infection. Furthermore, secreted Gluc activity and bacterial CFUs recovered from tissue cage fluid (TCF) are correlated along 18 days of infection. Importantly, secreted Gluc can also be detected in plasma samples and serve as an ex situ indicator for the established tissue cage infection, once high bacterial burdens are achieved. We have demonstrated that Gluc from marine eukaryotes can be stably expressed and secreted by pathogenic E. coli in vivo to enable a facile tool for longitudinal evaluation of persistent bacterial infection.

Characterization of extraintestinal pathogenic Escherichia coli isolated from captive wild felids with bacteremia.

PubMed

Carvalho, Vania M; Osugui, Lika; Setzer, Ariela P; Lopez, Rodrigo P G; Pestana de Castro, Antonio F; Irino, Kinue; Catão-Dias, José L

2012-09-01

Diseases caused by extraintestinal pathogenic Escherichia coli (ExPEC) in wild felids are rarely reported. Although urinary tract infections are infrequently reported in domestic cats, such infections when present are commonly caused by ExPEC. The present work characterized ExPEC strains isolated from 2 adult felines, a snow leopard (Panthera uncia) and a black leopard (Panthera pardus melas), that died from secondary bacteremia associated with urinary tract infections. Isolates from both animals were classified into the B2 phylogenetic group and expressed virulence genotypes that allowed them to cause severe disease. In addition, strains from the black leopard showed multidrug resistance.

Chromosomal features of Escherichia coli serotype O2:K2, an avian pathogenic E. coli.

PubMed

Jørgensen, Steffen L; Kudirkiene, Egle; Li, Lili; Christensen, Jens P; Olsen, John E; Nolan, Lisa; Olsen, Rikke H

2017-01-01

Escherichia coli causing infection outside the gastrointestinal system are referred to as extra-intestinal pathogenic E. coli. Avian pathogenic E. coli is a subgroup of extra-intestinal pathogenic E. coli and infections due to avian pathogenic E. coli have major impact on poultry production economy and welfare worldwide. An almost defining characteristic of avian pathogenic E. coli is the carriage of plasmids, which may encode virulence factors and antibiotic resistance determinates. For the same reason, plasmids of avian pathogenic E. coli have been intensively studied. However, genes encoded by the chromosome may also be important for disease manifestation and antimicrobial resistance. For the E. coli strain APEC_O2 the plasmids have been sequenced and analyzed in several studies, and E. coli APEC_O2 may therefore serve as a reference strain in future studies. Here we describe the chromosomal features of E. coli APEC_O2. E. coli APEC_O2 is a sequence type ST135, has a chromosome of 4,908,820 bp (plasmid removed), comprising 4672 protein-coding genes, 110 RNA genes, and 156 pseudogenes, with an average G + C content of 50.69%. We identified 82 insertion sequences as well as 4672 protein coding sequences, 12 predicated genomic islands, three prophage-related sequences, and two clustered regularly interspaced short palindromic repeats regions on the chromosome, suggesting the possible occurrence of horizontal gene transfer in this strain. The wildtype strain of E. coli APEC_O2 is resistant towards multiple antimicrobials, however, no (complete) antibiotic resistance genes were present on the chromosome, but a number of genes associated with extra-intestinal disease were identified. Together, the information provided here on E. coli APEC_O2 will assist in future studies of avian pathogenic E. coli strains, in particular regarding strain of E. coli APEC_O2, and aid in the general understanding of the pathogenesis of avian pathogenic E. coli .

Role of CD14 in responses to clinical isolates of Escherichia coli: effects of K1 capsule expression.

PubMed

Metkar, Shalaka; Awasthi, Shanjana; Denamur, Erick; Kim, Kwang Sik; Gangloff, Sophie C; Teichberg, Saul; Haziot, Alain; Silver, Jack; Goyert, Sanna M

2007-11-01

Severe bacterial infections leading to sepsis or septic shock can be induced by bacteria that utilize different factors to drive pathogenicity and/or virulence, leading to disease in the host. One major factor expressed by all clinical isolates of gram-negative bacteria is lipopolysaccharide (LPS); a second factor expressed by some Escherichia coli strains is a K1 polysaccharide capsule. To determine the role of the CD14 LPS receptor in the pathogenic effects of naturally occurring E. coli, the responses of CD14-/- and CD14+/+ mice to three different isolates of E. coli obtained from sepsis patients were compared; two isolates express both smooth LPS and the K1 antigen, while the third isolate expresses only LPS and is negative for K1. An additional K1-positive isolate obtained from a newborn with meningitis and a K1-negative isogenic mutant of this strain were also used for these studies. CD14-/- mice were resistant to the lethal effects of the K1-negative isolates. This resistance was accompanied by significantly lower levels of systemic tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6) in these mice than in CD14+/+ mice, enhanced clearance of the bacteria, and significantly fewer additional gross symptoms. In contrast, CD14-/- mice were as sensitive as CD14+/+ mice to the lethal effects of the K1-positive isolates, even though they had significantly lower levels of TNF-alpha and IL-6 than CD14+/+ mice. These studies show that different bacterial isolates can use distinctly different mechanisms to cause disease and suggest that new, nonantibiotic therapeutics need to be directed against multiple targets.

Frequent combination of antimicrobial multiresistance and extraintestinal pathogenicity in Escherichia coli isolates from urban rats (Rattus norvegicus) in Berlin, Germany.

PubMed

Guenther, Sebastian; Bethe, Astrid; Fruth, Angelika; Semmler, Torsten; Ulrich, Rainer G; Wieler, Lothar H; Ewers, Christa

2012-01-01

Urban rats present a global public health concern as they are considered a reservoir and vector of zoonotic pathogens, including Escherichia coli. In view of the increasing emergence of antimicrobial resistant E. coli strains and the on-going discussion about environmental reservoirs, we intended to analyse whether urban rats might be a potential source of putatively zoonotic E. coli combining resistance and virulence. For that, we took fecal samples from 87 brown rats (Rattus norvegicus) and tested at least three E. coli colonies from each animal. Thirty two of these E. coli strains were pre-selected from a total of 211 non-duplicate isolates based on their phenotypic resistance to at least three antimicrobial classes, thus fulfilling the definition of multiresistance. As determined by multilocus sequence typing (MLST), these 32 strains belonged to 24 different sequence types (STs), indicating a high phylogenetic diversity. We identified STs, which frequently occur among extraintestinal pathogenic E. coli (ExPEC), such as STs 95, 131, 70, 428, and 127. Also, the detection of a number of typical virulence genes confirmed that the rats tested carried ExPEC-like strains. In particular, the finding of an Extended-spectrum beta-lactamase (ESBL)-producing strain which belongs to a highly virulent, so far mainly human- and avian-restricted ExPEC lineage (ST95), which expresses a serogroup linked with invasive strains (O18:NM:K1), and finally, which produces an ESBL-type frequently identified among human strains (CTX-M-9), pointed towards the important role, urban rats might play in the transmission of multiresistant and virulent E. coli strains. Indeed, using a chicken infection model, this strain showed a high in vivo pathogenicity. Imagining the high numbers of urban rats living worldwide, the way to the transmission of putatively zoonotic, multiresistant, and virulent strains might not be far ahead. The unforeseeable consequences of such an emerging public health threat need careful consideration in the future.

Connexin 26 facilitates gastrointestinal bacterial infection in vitro.

PubMed

Simpson, Charlotte; Kelsell, David P; Marchès, Olivier

2013-01-01

Escherichia coli, including enteropathogenic E. coli (EPEC), represents the most common cause of diarrhoea worldwide and is therefore a serious public health burden. Treatment for gastrointestinal pathogens is hindered by the emergence of multiple antibiotic resistance, leading to the requirement for the development of new therapies. A variety of mechanisms act in combination to mediate gastrointestinal-bacterial-associated diarrhoea development. For example, EPEC infection of enterocytes induces attaching and effacing lesion formation and the disruption of tight junctions. An alternative enteric pathogen, Shigella flexneri, manipulates the expression of Connexin 26 (Cx26), a gap junction protein. S. flexneri can open Cx26 hemichannels allowing the release of ATP, whereas HeLa cells expressing mutant gap-junction-associated Cx26 are less susceptible to cellular invasion by S. flexneri than cells expressing wild-type (WT) Cx26. We have investigated further the link between Cx26 expression and gastrointestinal infection by using EPEC and S. flexneri as in vitro models of infection. In this study, a significant reduction in EPEC adherence was observed in cells expressing mutant Cx26 compared with WT Cx26. Furthermore, a significant reduction in both cellular invasion by S. flexneri and adherence by EPEC was demonstrated in human intestinal cell lines following treatment with Cx26 short interfering RNA. These in vitro results suggest that the loss of functional Cx26 expression provides improved protection against gastrointestinal bacterial pathogens. Thus, Cx26 represents a potential therapeutic target for gastrointestinal bacterial infection.

Structural Modification of Lipopolysaccharide Conferred by mcr-1 in Gram-Negative ESKAPE Pathogens.

PubMed

Liu, Yi-Yun; Chandler, Courtney E; Leung, Lisa M; McElheny, Christi L; Mettus, Roberta T; Shanks, Robert M Q; Liu, Jian-Hua; Goodlett, David R; Ernst, Robert K; Doi, Yohei

2017-06-01

mcr-1 was initially reported as the first plasmid-mediated colistin resistance gene in clinical isolates of Escherichia coli and Klebsiella pneumoniae in China and has subsequently been identified worldwide in various species of the family Enterobacteriaceae mcr-1 encodes a phosphoethanolamine transferase, and its expression has been shown to generate phosphoethanolamine-modified bis-phosphorylated hexa-acylated lipid A in E. coli Here, we investigated the effects of mcr-1 on colistin susceptibility and on lipopolysaccharide structures in laboratory and clinical strains of the Gram-negative ESKAPE ( Enterococcus faecium , Staphylococcus aureus , K. pneumoniae , Acinetobacter baumannii , Pseudomonas aeruginosa , and Enterobacter species) pathogens, which are often treated clinically by colistin. The effects of mcr-1 on colistin resistance were determined using MIC assays of laboratory and clinical strains of E. coli , K. pneumoniae , A. baumannii , and P. aeruginosa Lipid A structural changes resulting from MCR-1 were analyzed by mass spectrometry. The introduction of mcr-1 led to colistin resistance in E. coli , K. pneumoniae , and A. baumannii but only moderately reduced susceptibility in P. aeruginosa Phosphoethanolamine modification of lipid A was observed consistently for all four species. These findings highlight the risk of colistin resistance as a consequence of mcr-1 expression among ESKAPE pathogens, especially in K. pneumoniae and A. baumannii Furthermore, the observation that lipid A structures were modified despite only modest increases in colistin MICs in some instances suggests more sophisticated surveillance methods may need to be developed to track the dissemination of mcr-1 or plasmid-mediated phosphoethanolamine transferases in general. Copyright © 2017 American Society for Microbiology.

Structural Modification of Lipopolysaccharide Conferred by mcr-1 in Gram-Negative ESKAPE Pathogens

PubMed Central

Liu, Yi-Yun; Chandler, Courtney E.; Leung, Lisa M.; McElheny, Christi L.; Mettus, Roberta T.; Liu, Jian-Hua; Goodlett, David R.

2017-01-01

ABSTRACT mcr-1 was initially reported as the first plasmid-mediated colistin resistance gene in clinical isolates of Escherichia coli and Klebsiella pneumoniae in China and has subsequently been identified worldwide in various species of the family Enterobacteriaceae. mcr-1 encodes a phosphoethanolamine transferase, and its expression has been shown to generate phosphoethanolamine-modified bis-phosphorylated hexa-acylated lipid A in E. coli. Here, we investigated the effects of mcr-1 on colistin susceptibility and on lipopolysaccharide structures in laboratory and clinical strains of the Gram-negative ESKAPE (Enterococcus faecium, Staphylococcus aureus, K. pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogens, which are often treated clinically by colistin. The effects of mcr-1 on colistin resistance were determined using MIC assays of laboratory and clinical strains of E. coli, K. pneumoniae, A. baumannii, and P. aeruginosa. Lipid A structural changes resulting from MCR-1 were analyzed by mass spectrometry. The introduction of mcr-1 led to colistin resistance in E. coli, K. pneumoniae, and A. baumannii but only moderately reduced susceptibility in P. aeruginosa. Phosphoethanolamine modification of lipid A was observed consistently for all four species. These findings highlight the risk of colistin resistance as a consequence of mcr-1 expression among ESKAPE pathogens, especially in K. pneumoniae and A. baumannii. Furthermore, the observation that lipid A structures were modified despite only modest increases in colistin MICs in some instances suggests more sophisticated surveillance methods may need to be developed to track the dissemination of mcr-1 or plasmid-mediated phosphoethanolamine transferases in general. PMID:28373195

Bacillus subtilis and yeast cell wall improve the intestinal health of broilers challenged by Clostridium perfringens.

PubMed

Li, Z; Wang, W; Lv, Z; Liu, D; Guo, Y

2017-12-01

1. The objective was to investigate the effects of Bacillus subtilis, yeast cell wall (YCW) and their combination on intestinal health of broilers challenged by Clostridium perfringens over a 21-d period. 2. Using a 5 × 2 factorial arrangement of treatments, 800 1-d-old male Cobb 500 broilers were used to study the effects of feed additives (without additive or with zinc bacitracin, B. subtilis, YCW, and the combination of B. subtilis and YCW), pathogen challenge (without or with Clostridium perfringens challenge), and their interactive effects. 3. C. perfringens infection increased intestinal lesions scores, damaged intestinal histomorphology, increased serum endotoxin concentration, cytokine mRNA expression and intestinal population of C. perfringens and Escherichia coli and decreased ileal bifidobacteria numbers. The 4 additives decreased serum endotoxin. Zinc bacitracin tended to decrease cytokine mRNA expression and the intestinal number of C. perfringens and E. coli. B. subtilis, YCW and their combination increased cytokine mRNA expression. B. subtilis and YCW decreased the number of C. perfringens and E. coli in the ileum, and their combination decreased pathogens numbers in the ileum and caecum. 4. In conclusion, B. subtilis, YCW and their combination improved the intestinal health of NE-infected broilers, and could be potential alternatives to antibiotics.

Survival of pathogenic Escherichia coli on basil, lettuce, and spinach

USDA-ARS?s Scientific Manuscript database

The contamination of lettuce, spinach and basil with pathogenic E. coli has caused numerous illnesses over the past decade. E. coli O157:H7, E. coli O104:H4 and avian pathogenic E. coli (APECstx- and APECstx+) were inoculated on basil plants and in promix soiless substrate using drip and overhead ir...

Protective effects of indigenous Escherichia coli against a pathogenic E. coli challenge strain in pigs.

PubMed

Vahjen, W; Cuisiniere, T; Zentek, J

2017-10-13

To investigate the inhibitory effect of indigenous enterobacteria on pathogenic Escherichia coli, a challenge trial with postweaning pigs was conducted. A pathogenic E. coli strain was administered to all animals and their health was closely monitored thereafter. Faecal samples were taken from three healthy and three diarrhoeic animals. Samples were cultivated on MacConkey agar and isolates were subcultured. A soft agar overlay assay was used to determine the inhibitory activity of the isolates. A total of 1,173 enterobacterial isolates were screened for their ability to inhibit the E. coli challenge strain. Colony forming units of enterobacteria on MacConkey agar were not different between healthy and diarrhoeic animals in the original samples. Furthermore, numbers of isolates per animal were also not significantly different between healthy (482 isolates) and diarrhoeic animals (691 isolates). A total of 43 isolates (3.7%) with inhibitory activity against the pathogenic E. coli challenge strain were detected. All inhibitory isolates were identified as E. coli via MALDI-TOF. The isolates belonged to the phylotypes A, C and E. Many isolates (67.4%) were commensal E. coli without relevant porcine pathogenic factors, but toxin- and fimbrial genes (stx2e, fae, estIb, elt1a, fas, fan) were detected in 14 inhibitory isolates. Healthy animals showed significantly (P=0.003) more inhibitory isolates (36 of 482 isolates; 7.5%) than diseased animals (7 of 691 isolates; 1.0%). There were no significant correlations regarding phylotype or pathogenic factors between healthy and diseased animals. This study has shown that a small proportion of indigenous E. coli is able to inhibit in vitro growth of a pathogenic E. coli strain in pigs. Furthermore, healthy animals possess significantly more inhibitory E. coli strains than diarrhoeic animals. The inhibition of pathogenic E. coli by specific indigenous E. coli strains may be an underlying principle for the containment of pathogenic E. coli in pigs.

A secretome view of colonisation factors in Shiga toxin-encoding Escherichia coli (STEC): from enterohaemorrhagic E. coli (EHEC) to related enteropathotypes.

PubMed

Monteiro, Ricardo; Ageorges, Valentin; Rojas-Lopez, Maricarmen; Schmidt, Herbert; Weiss, Agnes; Bertin, Yolande; Forano, Evelyne; Jubelin, Grégory; Henderson, Ian R; Livrelli, Valérie; Gobert, Alain P; Rosini, Roberto; Soriani, Marco; Desvaux, Mickaël

2016-08-01

Shiga toxin-encoding Escherichia coli (STEC) regroup strains that carry genes encoding Shiga toxin (Stx). Among intestinal pathogenic E. coli, enterohaemorrhagic E. coli (EHEC) constitute the major subgroup of virulent STEC. EHEC cause serious human disease such as haemorrhagic colitis and haemolytic-uremic syndrome. While EHEC have evolved from enteropathogenic E. coli, hybrids with enteroaggregative E. coli have recently emerged. Of note, some enteroinvasive E. coli also belong to the STEC group. While the LEE (locus of enterocyte effacement) is a key and prominent molecular determinant in the pathogenicity, neither all EHEC nor STEC contain the LEE, suggesting that they possess additional virulence and colonisation factors. Currently, nine protein secretion systems have been described in diderm-lipopolysaccharide bacteria (archetypal Gram-negative) and can be involved in the secretion of extracellular effectors, cell-surface proteins or assembly of cell-surface organelles, such as flagella or pili. In this review, we focus on the secretome of STEC and related enteropathotypes, which are relevant to the colonisation of biotic and abiotic surfaces. Considering the wealth of potential protein trafficking mechanisms, the different combinations of colonisation factors and modulation of their expression is further emphasised with regard to the ecophysiology of STEC. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Recombinant production of a chimeric antimicrobial peptide in E. coli and assessment of its activity against some avian clinically isolated pathogens.

PubMed

Tanhaiean, Abass; Azghandi, Marjan; Razmyar, Jamshid; Mohammadi, Elyas; Sekhavati, Mohammad Hadi

2018-06-08

Over the last decades, poultry industry faced to the rapid emergence of multidrug-resistant bacteria as a global concern. Antimicrobial peptide (AMPs) known as potential antibiotic alternative and were considered as a new antimicrobial agent. Current methods of production and purification of AMPs have several limitations such as: costly, time-consuming and killing the producing host cells in recombinant form. In the present study, a chimeric peptide derived from camel lactoferrin was produced in Escherichia coli periplasmic space using a pET-based expression system and its antibacterial activity was determined on some avian pathogens in vitro. A carboxy-terminal polyhistidine tag was used for purification by Ni 2+ affinity chromatography with an average yield of 0.42 g/L. The His-tagged chimeric peptide showed different range of antimicrobial activity against clinically isolated avian pathogens with low chicken blood hemolysis activity and high serum stability. Overall, the results of this investigation showed the recombinant chimeric peptide was successfully expressed in pET-based expression system and could be considered as a proper alternative for some currently used antibiotics in poultry industry and drugs veterinary medicine. Copyright © 2018 Elsevier Ltd. All rights reserved.

Comparison of expression of monomeric and multimeric adenoregulin genes in Escherichia coli and Pichia pastorias.

PubMed

Zhou, Yuxun; Cao, Wei; Wang, Jinzhi; Ma, Yushu; Wei, Dongzhi

2005-05-01

Adenoregulin is a 33 amino acid antibiotic peptide who belongs to dermaseptin family which is the first vertebrate family to show lethal effects against filamentous fungi, as well as a broad spectrum of pathogenic microorganisms. Synthetic adenoregulin gene was cloned in 2, 4 and 6 tandem repeats and subcloned in pET32a and pET22b vectors. Recombinant plasmids were transformed into E. coli BL21(DE3), Fusion proteins of Trx-ADR1, Trx-ADR2 and Trx-ADR4 could be expressed after the hosts were induced by IPTG, but the expression level decreased dramatically with the number of tandem repeats increased. ADR1, ADR4 and ADR6 could not be expressed by E. coli without carrier proteins. But for Pichia pastoris GS115, ADR1 and ADR6 in the fermentation broth of the hosts could be detected by ELISA, and the bactericidal activities could also be observed.

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.

Risk of Transmission of Antimicrobial Resistant Escherichia coli from Commercial Broiler and Free-Range Retail Chicken in India

PubMed Central

Hussain, Arif; Shaik, Sabiha; Ranjan, Amit; Nandanwar, Nishant; Tiwari, Sumeet K.; Majid, Mohammad; Baddam, Ramani; Qureshi, Insaf A.; Semmler, Torsten; Wieler, Lothar H.; Islam, Mohammad A.; Chakravortty, Dipshikha; Ahmed, Niyaz

2017-01-01

Multidrug-resistant Escherichia coli infections are a growing public health concern. This study analyzed the possibility of contamination of commercial poultry meat (broiler and free-range) with pathogenic and or multi-resistant E. coli in retail chain poultry meat markets in India. We analyzed 168 E. coli isolates from broiler and free-range retail poultry (meat/ceca) sampled over a wide geographical area, for their antimicrobial sensitivity, phylogenetic groupings, virulence determinants, extended-spectrum-β-lactamase (ESBL) genotypes, fingerprinting by Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR and genetic relatedness to human pathogenic E. coli using whole genome sequencing (WGS). The prevalence rates of ESBL producing E. coli among broiler chicken were: meat 46%; ceca 40%. Whereas, those for free range chicken were: meat 15%; ceca 30%. E. coli from broiler and free-range chicken exhibited varied prevalence rates for multi-drug resistance (meat 68%; ceca 64% and meat 8%; ceca 26%, respectively) and extraintestinal pathogenic E. coli (ExPEC) contamination (5 and 0%, respectively). WGS analysis confirmed two globally emergent human pathogenic lineages of E. coli, namely the ST131 (H30-Rx subclone) and ST117 among our poultry E. coli isolates. These results suggest that commercial poultry meat is not only an indirect public health risk by being a possible carrier of non-pathogenic multi-drug resistant (MDR)-E. coli, but could as well be the carrier of human E. coli pathotypes. Further, the free-range chicken appears to carry low risk of contamination with antimicrobial resistant and extraintestinal pathogenic E. coli (ExPEC). Overall, these observations reinforce the understanding that poultry meat in the retail chain could possibly be contaminated by MDR and/or pathogenic E. coli. PMID:29180984

Risk of Transmission of Antimicrobial Resistant Escherichia coli from Commercial Broiler and Free-Range Retail Chicken in India.

PubMed

Hussain, Arif; Shaik, Sabiha; Ranjan, Amit; Nandanwar, Nishant; Tiwari, Sumeet K; Majid, Mohammad; Baddam, Ramani; Qureshi, Insaf A; Semmler, Torsten; Wieler, Lothar H; Islam, Mohammad A; Chakravortty, Dipshikha; Ahmed, Niyaz

2017-01-01

Multidrug-resistant Escherichia coli infections are a growing public health concern. This study analyzed the possibility of contamination of commercial poultry meat (broiler and free-range) with pathogenic and or multi-resistant E. coli in retail chain poultry meat markets in India. We analyzed 168 E. coli isolates from broiler and free-range retail poultry (meat/ceca) sampled over a wide geographical area, for their antimicrobial sensitivity, phylogenetic groupings, virulence determinants, extended-spectrum-β-lactamase (ESBL) genotypes, fingerprinting by Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR and genetic relatedness to human pathogenic E. coli using whole genome sequencing (WGS). The prevalence rates of ESBL producing E. coli among broiler chicken were: meat 46%; ceca 40%. Whereas, those for free range chicken were: meat 15%; ceca 30%. E. coli from broiler and free-range chicken exhibited varied prevalence rates for multi-drug resistance (meat 68%; ceca 64% and meat 8%; ceca 26%, respectively) and extraintestinal pathogenic E. coli (ExPEC) contamination (5 and 0%, respectively). WGS analysis confirmed two globally emergent human pathogenic lineages of E. coli , namely the ST131 ( H 30-Rx subclone) and ST117 among our poultry E. coli isolates. These results suggest that commercial poultry meat is not only an indirect public health risk by being a possible carrier of non-pathogenic multi-drug resistant (MDR)- E. coli , but could as well be the carrier of human E. coli pathotypes. Further, the free-range chicken appears to carry low risk of contamination with antimicrobial resistant and extraintestinal pathogenic E. coli (ExPEC). Overall, these observations reinforce the understanding that poultry meat in the retail chain could possibly be contaminated by MDR and/or pathogenic E. coli.

Characterization and Expression Analysis of Receptor for Activated C Kinase from Silk-producing Insect Antheraea pernyi.

PubMed

Zhu, Bao-Jian; Yu, Hao; Tian, Sen; Dai, Li-Shang; Sun, Yu; Liu, Chao-Liang

2016-01-01

The receptor for activated C kinase (RACK) is an important scaffold protein with regulatory functions in cells. However, its role in the immune response of Antheraea pernyi to pathogen challenge remains unclear. To investigate the biological functions of RACK in the wild silkworm A. pernyi, cloning was performed and the expression patterns of the RACK gene were analyzed. Sequence analysis revealed that the RACK gene was 1120 bp containing a 960-bp open reading frame. The deduced RACK protein sequence reveals the higher identity with its homologs from other insects. SDS-PAGE and western blot analysis demonstrated successful expression of a 36-kDa recombinant RACK protein in Escherichia coli. The titer of a rabbit-raised antibody against recombinant RACK protein was about 1: 20000, determined by ELISA. Real-time PCR analysis showed that RACK expression was higher in fat bodies than in other examined A. pernyi tissues. The expression of RACK mRNA in fat bodies of fifth larvae of A. pernyi was obviously induced after nucleopolyhedrovirus, E. coli or Beauveria bassiana challenge. However, the expression patterns of RACK were different in response to these pathogens. Our data suggest that RACK may play a role in the innate immune responses of A. pernyi.

Escherichia coli pathotypes

USDA-ARS?s Scientific Manuscript database

Escherichia coli strains are important commensals of the intestinal tract of humans and animals; however, pathogenic strains, including diarrhea-inducing E. coli and extraintestinal pathogenic E. coli. Intestinal E. coli pathotypes may cause a dehydrating watery diarrhea, or more severe diseases su...

Targeting virulence: salmochelin modification tunes the antibacterial activity spectrum of β-lactams for pathogen-selective killing of Escherichia coli † †Electronic supplementary information (ESI) available: Tables of bacterial strains employed in this study, iron content of the antimicrobial activity medium, characterization of GlcEnt–Amp/Amx 7–10, GlcEnt-PEG3-N3 12–13, and BLAST search for iroN sequence. Figures of HPLC traces of MceC- and IroB-catalyzed glucosylation of Ent-PEG3-N3 11, optical absorption spectra of GlcEnt–Amp/Amx 7–10, additional antimicrobial activity assays, time-kill kinetics, competition assays for FepA and IroN recognition, mixed-species antimicrobial activity assays, Lcn2 effect on antibacterial activity of GlcEnt–Amp/Amx 7–10, and cytotoxicity assays against T84 cells. See DOI: 10.1039/c5sc00962f Click here for additional data file.

PubMed Central

Chairatana, Phoom; Zheng, Tengfei

2015-01-01

New antibiotics are required to treat bacterial infections and counteract the emergence of antibiotic resistance. Pathogen-specific antibiotics have several advantages over broad-spectrum drugs, which include minimal perturbation to the commensal microbiota. We present a strategy for targeting antibiotics to bacterial pathogens that utilises the salmochelin-mediated iron uptake machinery of Gram-negative Escherichia coli. Salmochelins are C-glucosylated derivatives of the siderophore enterobactin. The biosynthesis and utilisation of salmochelins are important for virulence because these siderophores allow pathogens to acquire iron and evade the enterobactin-scavenging host-defense protein lipocalin-2. Inspired by the salmochelins, we report the design and chemoenzymatic preparation of glucosylated enterobactin–β-lactam conjugates that harbour the antibiotics ampicillin (Amp) and amoxicillin (Amx), hereafter GlcEnt–Amp/Amx. The GlcEnt scaffolds are based on mono- and diglucosylated Ent where one catechol moiety is functionalized at the C5 position for antibiotic attachment. We demonstrate that GlcEnt–Amp/Amx provide up to 1000-fold enhanced antimicrobial activity against uropathogenic E. coli relative to the parent β-lactams. Moreover, GlcEnt–Amp/Amx based on a diglucosylated Ent (DGE) platform selectively kill uropathogenic E. coli that express the salmochelin receptor IroN in the presence of non-pathogenic E. coli and other bacterial strains that include the commensal microbe Lactobacillus rhamnosus GG. Moreover, GlcEnt–Amp/Amx evade the host-defense protein lipocalin-2, and exhibit low toxicity to mammalian cells. Our work establishes that siderophore–antibiotic conjugates provide a strategy for targeting virulence, narrowing the activity spectrum of antibiotics in clinical use, and achieving selective delivery of antibacterial cargos to pathogenic bacteria on the basis of siderophore receptor expression. PMID:28717471

Toll-like receptor prestimulation increases phagocytosis of Escherichia coli DH5alpha and Escherichia coli K1 strains by murine microglial cells.

PubMed

Ribes, Sandra; Ebert, Sandra; Czesnik, Dirk; Regen, Tommy; Zeug, Andre; Bukowski, Stephanie; Mildner, Alexander; Eiffert, Helmut; Hanisch, Uwe-Karsten; Hammerschmidt, Sven; Nau, Roland

2009-01-01

Meningitis and meningoencephalitis caused by Escherichia coli are associated with high rates of mortality. When an infection occurs, Toll-like receptors (TLRs) expressed by microglial cells can recognize pathogen-associated molecular patterns and activate multiple steps in the inflammatory response that coordinate the brain's local defense, such as phagocytosis of invading pathogens. An upregulation of the phagocytic ability of reactive microglia could improve the host defense in immunocompromised patients against pathogens such as E. coli. Here, murine microglial cultures were stimulated with the TLR agonists Pam(3)CSK(4) (TLR1/TLR2), lipopolysaccharide (TLR4), and CpG oligodeoxynucleotide (TLR9) for 24 h. Upon stimulation, levels of tumor necrosis factor alpha and the neutrophil chemoattractant CXCL1 were increased, indicating microglial activation. Phagocytic activity was studied after adding either E. coli DH5alpha or E. coli K1 strains. After 60 and 90 min of bacterial exposure, the number of ingested bacteria was significantly higher in cells prestimulated with TLR agonists than in unstimulated controls (P < 0.01). Addition of cytochalasin D, an inhibitor of actin polymerization, blocked >90% of phagocytosis. We also analyzed the ability of microglia to kill the ingested E. coli strains. Intracellularly surviving bacteria were quantified at different time points (90, 150, 240, and 360 min) after 90 min of phagocytosis. The number of bacteria killed intracellularly after 6 h was higher in cells primed with the different TLR agonists than in unstimulated microglia. Our data suggest that microglial stimulation by the TLR system can increase bacterial phagocytosis and killing. This approach could improve central nervous system resistance to infections in immunocompromised patients.

Inhibition of Inducible Nitric Oxide Controls Pathogen Load and Brain Damage by Enhancing Phagocytosis of Escherichia coli K1 in Neonatal Meningitis

PubMed Central

Mittal, Rahul; Gonzalez-Gomez, Ignacio; Goth, Kerstin A.; Prasadarao, Nemani V.

2010-01-01

Escherichia coli K1 is a leading cause of neonatal meningitis in humans. In this study, we sought to determine the pathophysiologic relevance of inducible nitric oxide (iNOS) in experimental E. coli K1 meningitis. By using a newborn mouse model of meningitis, we demonstrate that E. coli infection triggered the expression of iNOS in the brains of mice. Additionally, iNOS−/− mice were resistant to E. coli K1 infection, displaying normal brain histology, no bacteremia, no disruption of the blood–brain barrier, and reduced inflammatory response. Treatment with an iNOS specific inhibitor, aminoguanidine (AG), of wild-type animals before infection prevented the development of bacteremia and the occurrence of meningitis. The infected animals treated with AG after the development of bacteremia also completely cleared the pathogen from circulation and prevented brain damage. Histopathological and micro-CT analysis of brains revealed significant damage in E. coli K1–infected mice, which was completely abrogated by AG administration. Peritoneal macrophages and polymorphonuclear leukocytes isolated from iNOS−/− mice or pretreated with AG demonstrated enhanced uptake and killing of the bacteria compared with macrophages and polymorphonuclear leukocytes from wild-type mice in which E. coli K1 survive and multiply. Thus, NO produced by iNOS may be beneficial for E. coli to survive inside the macrophages, and prevention of iNOS could be a therapeutic strategy to treat neonatal E. coli meningitis. PMID:20093483

Inhibition of inducible nitric oxide controls pathogen load and brain damage by enhancing phagocytosis of Escherichia coli K1 in neonatal meningitis.

PubMed

Mittal, Rahul; Gonzalez-Gomez, Ignacio; Goth, Kerstin A; Prasadarao, Nemani V

2010-03-01

Escherichia coli K1 is a leading cause of neonatal meningitis in humans. In this study, we sought to determine the pathophysiologic relevance of inducible nitric oxide (iNOS) in experimental E. coli K1 meningitis. By using a newborn mouse model of meningitis, we demonstrate that E. coli infection triggered the expression of iNOS in the brains of mice. Additionally, iNOS-/- mice were resistant to E. coli K1 infection, displaying normal brain histology, no bacteremia, no disruption of the blood-brain barrier, and reduced inflammatory response. Treatment with an iNOS specific inhibitor, aminoguanidine (AG), of wild-type animals before infection prevented the development of bacteremia and the occurrence of meningitis. The infected animals treated with AG after the development of bacteremia also completely cleared the pathogen from circulation and prevented brain damage. Histopathological and micro-CT analysis of brains revealed significant damage in E. coli K1-infected mice, which was completely abrogated by AG administration. Peritoneal macrophages and polymorphonuclear leukocytes isolated from iNOS-/- mice or pretreated with AG demonstrated enhanced uptake and killing of the bacteria compared with macrophages and polymorphonuclear leukocytes from wild-type mice in which E. coli K1 survive and multiply. Thus, NO produced by iNOS may be beneficial for E. coli to survive inside the macrophages, and prevention of iNOS could be a therapeutic strategy to treat neonatal E. coli meningitis.

Uterine diseases in cattle after parturition

PubMed Central

Sheldon, I. Martin; Williams, Erin J.; Miller, Aleisha N.A.; Nash, Deborah M.; Herath, Shan

2008-01-01

Bacterial contamination of the uterine lumen is common in cattle after parturition, often leading to infection and uterine disease. Clinical disease can be diagnosed and scored by examination of the vaginal mucus, which reflects the presence of pathogenic bacteria such as Escherichia coli and Arcanobacterium pyogenes. Viruses may also cause uterine disease and bovine herpesvirus 4 (BoHV-4) is tropic for endometrial cells, causing a rapid cytopathic effect. The elimination of pathogens by the innate immune system is dependent on pattern recognition receptors binding pathogen-associated molecules. Uterine epithelial and stromal cells express receptors such as Toll-like Receptor 4 that binds E. coli lipopolysaccharide. The infertility associated with uterine disease is caused by damage to the endometrium and disruption of ovarian cyclic activity. Bacteria modulate endometrial prostaglandin secretion, and perturb ovarian follicle growth and function. Understanding the molecular basis of uterine disease will lead to novel approaches to treating infertility. PMID:18329302

Inactivation of conserved genes induces microbial aversion, drug detoxification, and innate immunity in C.elegans

PubMed Central

Melo, Justine A.; Ruvkun, Gary

2012-01-01

Summary The nematode C. elegans consumes benign bacteria such as E. coli and is repelled by pathogens and toxins. Here we show that RNAi and toxin-mediated disruption of core cellular activities, including translation, respiration, and protein turnover, stimulates behavioral avoidance of attractive E. coli. RNAi of such essential processes also induces expression of detoxification and innate immune response genes in the absence of toxins or pathogens. Disruption of core processes in non-neuronal tissues can stimulate aversion behavior, revealing a neuroendocrine axis of control. Microbial avoidance requires serotonergic and Jnk kinase signaling. We propose that surveillance pathways oversee critical cellular activities to detect pathogens, many of which deploy toxins and virulence factors to disrupt these same host pathways. Variation in cellular surveillance and endocrine pathways controlling behavior, detoxification and immunity selected by past toxin or microbial interactions could underlie aberrant responses to foods, medicines, and microbes. PMID:22500807

Development of a dual vaccine for prevention of Brucella abortus infection and Escherichia coli O157:H7 intestinal colonization.

PubMed

Iannino, Florencia; Herrmann, Claudia K; Roset, Mara S; Briones, Gabriel

2015-05-05

Zoonoses that affect human and animal health have an important economic impact. In the study now presented, a bivalent vaccine has been developed that has the potential for preventing the transmission from cattle to humans of two bacterial pathogens: Brucella abortus and Shiga toxin-producing Escherichia coli (STEC). A 66kDa chimeric antigen, composed by EspA, Intimin, Tir, and H7 flagellin (EITH7) from STEC, was constructed and expressed in B. abortus Δpgm vaccine strain (BabΔpgm). Mice orally immunized with BabΔpgm(EITH7) elicited an immune response with the induction of anti-EITH7 antibodies (IgA) that clears an intestinal infection of E. coli O157:H7 three times faster (t=4 days) than mice immunized with BabΔpgm carrier strain (t=12 days). As expected, mice immunized with BabΔpgm(EITH7) strain also elicited a protective immune response against B. abortus infection. A Brucella-based vaccine platform is described capable of eliciting a combined protective immune response against two bacterial pathogens with diverse lifestyles-the intracellular pathogen B. abortus and the intestinal extracellular pathogen STEC. Copyright © 2015 Elsevier Ltd. All rights reserved.

The Gene tia, Harbored by the Subtilase-Encoding Pathogenicity Island, Is Involved in the Ability of Locus of Enterocyte Effacement-Negative Shiga Toxin-Producing Escherichia coli Strains To Invade Monolayers of Epithelial Cells

PubMed Central

Chiani, Paola; Michelacci, Valeria; Minelli, Fabio; Caprioli, Alfredo; Morabito, Stefano

2017-01-01

ABSTRACT Locus of enterocyte effacement (LEE)-negative Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are human pathogens that lack the LEE locus, a pathogenicity island (PAI) involved in the intimate adhesion of LEE-positive strains to the host gut epithelium. The mechanism used by LEE-negative STEC strains to colonize the host intestinal mucosa is still not clear. The cell invasion determinant tia, previously described in enterotoxigenic E. coli strains, has been identified in LEE-negative STEC strains that possess the subtilase-encoding pathogenicity island (SE-PAI). We evaluated the role of the gene tia, present in these LEE-negative STEC strains, in the invasion of monolayers of cultured cells. We observed that these strains were able to invade Caco-2 and HEp-2 cell monolayers and compared their invasion ability with that of a mutant strain in which the gene tia had been inactivated. Mutation of the gene tia resulted in a strong reduction of the invasive phenotype, and complementation of the tia mutation with a functional copy of the gene restored the invasion activity. Moreover, we show that the gene tia is overexpressed in bacteria actively invading cell monolayers, demonstrating that tia is involved in the ability to invade cultured monolayers of epithelial cells shown by SE-PAI-positive E. coli, including STEC, strains. However, the expression of the tia gene in the E. coli K-12 strain JM109 was not sufficient, in its own right, to confer to this strain the ability to invade cell monolayers, suggesting that at least another factor must be involved in the invasion ability displayed by the SE-PAI-positive strains. PMID:28893912

The Gene tia, Harbored by the Subtilase-Encoding Pathogenicity Island, Is Involved in the Ability of Locus of Enterocyte Effacement-Negative Shiga Toxin-Producing Escherichia coli Strains To Invade Monolayers of Epithelial Cells.

PubMed

Bondì, Roslen; Chiani, Paola; Michelacci, Valeria; Minelli, Fabio; Caprioli, Alfredo; Morabito, Stefano

2017-12-01

Locus of enterocyte effacement (LEE)-negative Shiga toxin (Stx)-producing Escherichia coli (STEC) strains are human pathogens that lack the LEE locus, a pathogenicity island (PAI) involved in the intimate adhesion of LEE-positive strains to the host gut epithelium. The mechanism used by LEE-negative STEC strains to colonize the host intestinal mucosa is still not clear. The cell invasion determinant tia , previously described in enterotoxigenic E. coli strains, has been identified in LEE-negative STEC strains that possess the subtilase-encoding pathogenicity island (SE-PAI). We evaluated the role of the gene tia , present in these LEE-negative STEC strains, in the invasion of monolayers of cultured cells. We observed that these strains were able to invade Caco-2 and HEp-2 cell monolayers and compared their invasion ability with that of a mutant strain in which the gene tia had been inactivated. Mutation of the gene tia resulted in a strong reduction of the invasive phenotype, and complementation of the tia mutation with a functional copy of the gene restored the invasion activity. Moreover, we show that the gene tia is overexpressed in bacteria actively invading cell monolayers, demonstrating that tia is involved in the ability to invade cultured monolayers of epithelial cells shown by SE-PAI-positive E. coli , including STEC, strains. However, the expression of the tia gene in the E. coli K-12 strain JM109 was not sufficient, in its own right, to confer to this strain the ability to invade cell monolayers, suggesting that at least another factor must be involved in the invasion ability displayed by the SE-PAI-positive strains. Copyright © 2017 American Society for Microbiology.

Insights into the evolution of pathogenicity of Escherichia coli from genomic analysis of intestinal E. coli of Marmota himalayana in Qinghai–Tibet plateau of China

PubMed Central

Lu, Shan; Jin, Dong; Wu, Shusheng; Yang, Jing; Lan, Ruiting; Bai, Xiangning; Liu, Sha; Meng, Qiong; Yuan, Xuejiao; Zhou, Juan; Pu, Ji; Chen, Qiang; Dai, Hang; Hu, Yuanyuan; Xiong, Yanwen; Ye, Changyun; Xu, Jianguo

2016-01-01

Escherichia coli is both of a widespread harmless gut commensal and a versatile pathogen of humans. Domestic animals are a well-known reservoir for pathogenic E. coli. However, studies of E. coli populations from wild animals that have been separated from human activities had been very limited. Here we obtained 580 isolates from intestinal contents of 116 wild Marmot Marmota himalayana from Qinghai–Tibet plateau, China, with five isolates per animal. We selected 125 (hereinafter referred to as strains) from the 580 isolates for genome sequencing, based on unique pulse field gel electrophoresis patterns and at least one isolate per animal. Whole genome sequence analysis revealed that all 125 strains carried at least one and the majority (79.2%) carried multiple virulence genes based on the analysis of 22 selected virulence genes. In particular, the majority of the strains carried virulence genes from different pathovars as potential 'hybrid pathogens'. The alleles of eight virulence genes from the Marmot E. coli were found to have diverged earlier than all known alleles from human and other animal E. coli. Phylogenetic analysis of the 125 Marmot E. coli genomes and 355 genomes selected from 1622 human and other E. coli strains identified two new phylogroups, G and H, both of which diverged earlier than the other phylogroups. Eight of the 12 well-known pathogenic E. coli lineages were found to share a most recent common ancestor with one or more Marmot E. coli strains. Our results suggested that the intestinal E. coli of the Marmots contained a diverse virulence gene pool and is potentially pathogenic to humans. These findings provided a new understanding of the evolutionary origin of pathogenic E. coli. PMID:27924811

Insights into the evolution of pathogenicity of Escherichia coli from genomic analysis of intestinal E. coli of Marmota himalayana in Qinghai-Tibet plateau of China.

PubMed

Lu, Shan; Jin, Dong; Wu, Shusheng; Yang, Jing; Lan, Ruiting; Bai, Xiangning; Liu, Sha; Meng, Qiong; Yuan, Xuejiao; Zhou, Juan; Pu, Ji; Chen, Qiang; Dai, Hang; Hu, Yuanyuan; Xiong, Yanwen; Ye, Changyun; Xu, Jianguo

2016-12-07

Escherichia coli is both of a widespread harmless gut commensal and a versatile pathogen of humans. Domestic animals are a well-known reservoir for pathogenic E. coli. However, studies of E. coli populations from wild animals that have been separated from human activities had been very limited. Here we obtained 580 isolates from intestinal contents of 116 wild Marmot Marmota himalayana from Qinghai-Tibet plateau, China, with five isolates per animal. We selected 125 (hereinafter referred to as strains) from the 580 isolates for genome sequencing, based on unique pulse field gel electrophoresis patterns and at least one isolate per animal. Whole genome sequence analysis revealed that all 125 strains carried at least one and the majority (79.2%) carried multiple virulence genes based on the analysis of 22 selected virulence genes. In particular, the majority of the strains carried virulence genes from different pathovars as potential 'hybrid pathogens'. The alleles of eight virulence genes from the Marmot E. coli were found to have diverged earlier than all known alleles from human and other animal E. coli. Phylogenetic analysis of the 125 Marmot E. coli genomes and 355 genomes selected from 1622 human and other E. coli strains identified two new phylogroups, G and H, both of which diverged earlier than the other phylogroups. Eight of the 12 well-known pathogenic E. coli lineages were found to share a most recent common ancestor with one or more Marmot E. coli strains. Our results suggested that the intestinal E. coli of the Marmots contained a diverse virulence gene pool and is potentially pathogenic to humans. These findings provided a new understanding of the evolutionary origin of pathogenic E. coli.

Pertussis Toxin Exploits Specific Host Cell Signaling Pathways for Promoting Invasion and Translocation of Escherichia coli K1 RS218 in Human Brain-derived Microvascular Endothelial Cells*

PubMed Central

Karassek, Sascha; Starost, Laura; Solbach, Johanna; Greune, Lilo; Sano, Yasuteru; Kanda, Takashi; Kim, KwangSik; Schmidt, M. Alexander

2015-01-01

Pertussis toxin (PTx), an AB5 toxin and major virulence factor of the whooping cough-causing pathogen Bordetella pertussis, has been shown to affect the blood-brain barrier. Dysfunction of the blood-brain barrier may facilitate penetration of bacterial pathogens into the brain, such as Escherichia coli K1 (RS218). In this study, we investigated the influence of PTx on blood-brain barrier permissiveness to E. coli infection using human brain-derived endothelial HBMEC and TY10 cells as in vitro models. Our results indicate that PTx acts at several key points of host cell intracellular signaling pathways, which are also affected by E. coli K1 RS218 infection. Application of PTx increased the expression of the pathogen binding receptor gp96. Further, we found an activation of STAT3 and of the small GTPase Rac1, which have been described as being essential for bacterial invasion involving host cell actin cytoskeleton rearrangements at the bacterial entry site. In addition, we showed that PTx induces a remarkable relocation of VE-cadherin and β-catenin from intercellular junctions. The observed changes in host cell signaling molecules were accompanied by differences in intracellular calcium levels, which might act as a second messenger system for PTx. In summary, PTx not only facilitates invasion of E. coli K1 RS218 by activating essential signaling cascades; it also affects intercellular barriers to increase paracellular translocation. PMID:26324705

Study on E. coli and Salmonella biofilms from fresh fruits and vegetables.

PubMed

Amrutha, Balagopal; Sundar, Kothandapani; Shetty, Prathapkumar Halady

2017-04-01

Foodborne outbreaks associated with fresh fruits and vegetables are on the rise worldwide. Biofilm formation is one of the important traits of pathogens making them strongly attached to substrates as well as express virulence phenotypes. Present study investigates the biofilm forming ability of E. coli and Salmonella sp. isolated from fresh fruits and vegetables. A total of 53 strains, including 35 E. coli and 18 Salmonella sp. isolated from different fruit and vegetable samples were taken into account for the study. Initial screening for biofilm formation was done using Congo Red agar plate test. Results revealed that 22.8% E. coli and 22.2% Salmonella sp. were potential biofilm formers. However, the MTP (Micro-Titre Plate) assay suggested more isolates of both E. coli and Salmonella sp. were moderate to strong biofilm producers. Agar plate diffusion assay with Agrobacterium tumefaciens NTL-4 showed the production of quorum signaling molecules (AHLs) by three isolates of E. coli and one Salmonella sp. Two E. coli isolates showed a significant amount of EPS production indicating higher biofilm forming potential. The Presence of LUX R homologue gene ( sdi A) in two of the Salmonella isolates were confirmed by PCR which demonstrated their potential pathogenicity. Results of the work underline the biofilm forming and potentially virulent capacities of isolates from the surface of fruits and vegetables.

Comparative Study on Different Expression Hosts for Alkaline Phytase Engineered in Escherichia coli.

PubMed

Chen, Weiwei; Yu, Hongwei; Ye, Lidan

2016-07-01

The application of alkaline phytase as a feed additive is restricted by the poor specific activity. Escherichia coli is a frequently used host for directed evolution of proteins including alkaline phytase towards improved activity. However, it is not suitable for production of food-grade products due to potential pathogenicity. To combine the advantages of different expression systems, mutants of the alkaline phytase originated from Bacillus subtilis 168 (phy168) were first generated via directed evolution in E. coli and then transformed to food-grade hosts B. subtilis and Pichia pastoris for secretory expression. In order to investigate the suitability of different expression systems, the phy168 mutants expressed in different hosts were characterized and compared in terms of specific activity, pH profile, pH stability, temperature profile, and thermostability. The specific activity of B. subtilis-expressed D24G/K70R/K111E/N121S mutant at pH 7.0 and 60 °C was 30.4 U/mg, obviously higher than those in P. pastoris (22.7 U/mg) and E. coli (19.7 U/mg). Moreover, after 10 min incubation at 80 °C, the B. subtilis-expressed D24G/K70R/K111E/N121S retained about 70 % of the activity at pH 7.0 and 37 °C, whereas the values were only about 25 and 50 % when expressed in P. pastoris and E. coli, respectively. These results suggested B. subtilis as an appropriate host for expression of phy168 mutants and that the strategy of creating mutants in one host and expressing them in another might be a new solution to industrial production of proteins with desired properties.

Mixed biofilm formation by Shiga toxin-producing Escherichia coli and Salmonella enterica serovar Typhimurium enhanced bacterial resistance to sanitization due to extracellular polymeric substances.

PubMed

Wang, Rong; Kalchayanand, Norasak; Schmidt, John W; Harhay, Dayna M

2013-09-01

Shiga toxin-producing Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium are important foodborne pathogens capable of forming single-species biofilms or coexisting in multispecies biofilm communities. Bacterial biofilm cells are usually more resistant to sanitization than their planktonic counterparts, so these foodborne pathogens in biofilms pose a serious food safety concern. We investigated how the coexistence of E. coli O157:H7 and Salmonella Typhimurium strains would affect bacterial planktonic growth competition and mixed biofilm composition. Furthermore, we also investigated how mixed biofilm formation would affect bacterial resistance to common sanitizers. Salmonella Typhimurium strains were able to outcompete E. coli strains in the planktonic growth phase; however, mixed biofilm development was highly dependent upon companion strain properties in terms of the expression of bacterial extracellular polymeric substances (EPS), including curli fimbriae and exopolysaccharide cellulose. The EPS-producing strains with higher biofilm-forming abilities were able to establish themselves in mixed biofilms more efficiently. In comparison to single-strain biofilms, Salmonella or E. coli strains with negative EPS expression obtained significantly enhanced resistance to sanitization by forming mixed biofilms with an EPS-producing companion strain of the other species. These observations indicate that the bacterial EPS components not only enhance the sanitizer resistance of the EPS-producing strains but also render protections to their companion strains, regardless of species, in mixed biofilms. Our study highlights the potential risk of cross-contamination by multispecies biofilms in food safety and the need for increased attention to proper sanitization practices in food processing facilities.

Epigenetic Mechanisms Regulate Innate Immunity against Uropathogenic and Commensal-Like Escherichia coli in the Surrogate Insect Model Galleria mellonella.

PubMed

Heitmueller, Miriam; Billion, André; Dobrindt, Ulrich; Vilcinskas, Andreas; Mukherjee, Krishnendu

2017-10-01

Innate-immunity-related genes in humans are activated during urinary tract infections (UTIs) caused by pathogenic strains of Escherichia coli but are suppressed by commensals. Epigenetic mechanisms play a pivotal role in the regulation of gene expression in response to environmental stimuli. To determine whether epigenetic mechanisms can explain the different behaviors of pathogenic and commensal bacteria, we infected larvae of the greater wax moth, Galleria mellonella , a widely used model insect host, with a uropathogenic E. coli (UPEC) strain that causes symptomatic UTIs in humans or a commensal-like strain that causes asymptomatic bacteriuria (ABU). Infection with the UPEC strain (CFT073) was more lethal to larvae than infection with the attenuated ABU strain (83972) due to the recognition of each strain by different Toll-like receptors, ultimately leading to differential DNA/RNA methylation and histone acetylation. We used next-generation sequencing and reverse transcription (RT)-PCR to correlate epigenetic changes with the induction of innate-immunity-related genes. Transcriptomic analysis of G. mellonella larvae infected with E. coli strains CFT073 and 83972 revealed strain-specific variations in the class and expression levels of genes encoding antimicrobial peptides, cytokines, and enzymes controlling DNA methylation and histone acetylation. Our results provide evidence for the differential epigenetic regulation of transcriptional reprogramming by UPEC and ABU strains of E. coli in G. mellonella larvae, which may be relevant to understanding the different behaviors of these bacterial strains in the human urinary tract. Copyright © 2017 American Society for Microbiology.

Diversity, Replication, Pathogenicity and Cell Biology of Crimean Congo Hemorrhagic Fever Virus

DTIC Science & Technology

2006-10-01

recombinant protein produced and purified in E . coli was able to bind to ssRNA in a dot blot filter binding assay. In order to identify domains in the N...Fig. 1. RNA binding domains of the N protein of CCHFV. The depicted GST-fusion proteins were expressed in E . coli and purified using a...detected and NSm protein produced after cleavage of the glycoprotein precursor in virus infected cells. The NSm is stable and transported to the Golgi

Analysis of Genome Plasticity in Pathogenic and Commensal Escherichia coli Isolates by Use of DNA Arrays

PubMed Central

Dobrindt, Ulrich; Agerer, Franziska; Michaelis, Kai; Janka, Andreas; Buchrieser, Carmen; Samuelson, Martin; Svanborg, Catharina; Gottschalk, Gerhard; Karch, Helge; Hacker, Jörg

2003-01-01

Genomes of prokaryotes differ significantly in size and DNA composition. Escherichia coli is considered a model organism to analyze the processes involved in bacterial genome evolution, as the species comprises numerous pathogenic and commensal variants. Pathogenic and nonpathogenic E. coli strains differ in the presence and absence of additional DNA elements contributing to specific virulence traits and also in the presence and absence of additional genetic information. To analyze the genetic diversity of pathogenic and commensal E. coli isolates, a whole-genome approach was applied. Using DNA arrays, the presence of all translatable open reading frames (ORFs) of nonpathogenic E. coli K-12 strain MG1655 was investigated in 26 E. coli isolates, including various extraintestinal and intestinal pathogenic E. coli isolates, 3 pathogenicity island deletion mutants, and commensal and laboratory strains. Additionally, the presence of virulence-associated genes of E. coli was determined using a DNA “pathoarray” developed in our laboratory. The frequency and distributional pattern of genomic variations vary widely in different E. coli strains. Up to 10% of the E. coli K-12-specific ORFs were not detectable in the genomes of the different strains. DNA sequences described for extraintestinal or intestinal pathogenic E. coli are more frequently detectable in isolates of the same origin than in other pathotypes. Several genes coding for virulence or fitness factors are also present in commensal E. coli isolates. Based on these results, the conserved E. coli core genome is estimated to consist of at least 3,100 translatable ORFs. The absence of K-12-specific ORFs was detectable in all chromosomal regions. These data demonstrate the great genome heterogeneity and genetic diversity among E. coli strains and underline the fact that both the acquisition and deletion of DNA elements are important processes involved in the evolution of prokaryotes. PMID:12618447

Pathogenic Escherichia coli producing Extended-Spectrum β-Lactamases isolated from surface water and wastewater.

PubMed

Franz, Eelco; Veenman, Christiaan; van Hoek, Angela H A M; de Roda Husman, Ana; Blaak, Hetty

2015-09-24

To assess public health risks from environmental exposure to Extended-Spectrum β-Lactamases (ESBL)-producing bacteria, it is necessary to have insight in the proportion of relative harmless commensal variants and potentially pathogenic ones (which may directly cause disease). In the current study, 170 ESBL-producing E. coli from Dutch wastewater (n = 82) and surface water (n = 88) were characterized with respect to ESBL-genotype, phylogenetic group, resistance phenotype and virulence markers associated with enteroaggregative E. coli (EAEC), enteroinvasive E. coli (EIEC), enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), extraintesinal E. coli (ExPEC), and Shiga toxin-producing E. coli (STEC). Overall, 17.1% of all ESBL-producing E. coli were suspected pathogenic variants. Suspected ExPECs constituted 8.8% of all ESBL-producing variants and 8.3% were potential gastrointestinal pathogens (4.1% EAEC, 1.8% EPEC, 1.2% EIEC, 1.2% ETEC, no STEC). Suspected pathogens were significantly associated with ESBL-genotype CTX-M-15 (X(2) = 14.7, P < 0.001) and phylogenetic group B2 (X(2) = 23.5, P < 0.001). Finally, 84% of the pathogenic ESBL-producing E. coli isolates were resistant to three or more different classes of antibiotics. In conclusion, this study demonstrates that the aquatic environment is a potential reservoir of E. coli variants that combine ESBL-genes, a high level of multi-drug resistance and virulence factors, and therewith pose a health risk to humans upon exposure.

Expression of Clostridium perfringens epsilon-beta fusion toxin gene in E. coli and its immunologic studies in mouse.

PubMed

Pilehchian Langroudi, Reza; Shamsara, Mehdi; Aghaiypour, Khosrow

2013-07-11

Clostridium perfringens is an anaerobic spore-forming, pathogenic bacterium that is responsible for severe diseases in humans and livestock. In the present study, an epsilon-beta fusion toxin was expressed as a soluble protein in E. coli and the recombinant cell lysate was used for immunization studies in mouse. Potency of the toxin (as an antigen) induced 6 and 10IU/ml of epsilon and beta anti-toxin in rabbit, respectively. These titers were higher than the minimum level required by the European Pharmacopoeia for epsilon and beta toxins. Experimental challenge with the recombinant fusion toxoid revealed that it could protect mice against C. perfringens epsilon and beta toxins. Toxicity of the fusion toxin was studied by histopathological findings, which were the same as the native toxins. In conclusion, E. coli is a suitable expression host for immunogenic epsilon-beta fusion toxin of C. perfringens. Copyright © 2013 Elsevier Ltd. All rights reserved.

Significance of Viable but Nonculturable Escherichia coli: Induction, Detection, and Control.

PubMed

Ding, Tian; Suo, Yuanjie; Xiang, Qisen; Zhao, Xihong; Chen, Shiguo; Ye, Xingqian; Liu, Donghong

2017-03-28

Diseases caused by foodborne or waterborne pathogens are emerging. Many pathogens can enter into the viable but nonculturable (VBNC) state, which is a survival strategy when exposed to harsh environmental stresses. Pathogens in the VBNC state have the ability to evade conventional microbiological detection methods, posing a significant and potential health risk. Therefore, controlling VBNC bacteria in food processing and the environment is of great importance. As the typical one of the gram-negatives, Escherichia coli ( E. coli ) is a widespread foodborne and waterborne pathogenic bacterium and is able to enter into a VBNC state in extreme conditions (similar to the other gram-negative bacteria), including inducing factors and resuscitation stimulus. VBNC E. coli has the ability to recover both culturability and pathogenicity, which may bring potential health risk. This review describes the concrete factors (nonthermal treatment, chemical agents, and environmental factors) that induce E. coli into the VBNC state, the condition or stimulus required for resuscitation of VBNC E. coli , and the methods for detecting VBNC E. coli . Furthermore, the mechanism of genes and proteins involved in the VBNC E. coli is also discussed in this review.

RNA-Mediated Thermoregulation of Iron-Acquisition Genes in Shigella dysenteriae and Pathogenic Escherichia coli

PubMed Central

Kouse, Andrew B.; Righetti, Francesco; Kortmann, Jens; Narberhaus, Franz; Murphy, Erin R.

2013-01-01

The initiation, progression and transmission of most bacterial infections is dependent upon the ability of the invading pathogen to acquire iron from each of the varied environments encountered during the course of a natural infection. In total, 95% of iron within the human body is complexed within heme, making heme a potentially rich source of host-associated nutrient iron for invading bacteria. As heme is encountered only within the host, pathogenic bacteria often regulate synthesis of heme utilization factors such that production is maximal under host-associated environmental conditions. This study examines the regulated production of ShuA, an outer-membrane receptor required for the utilization of heme as a source of nutrient iron by Shigella dysenteriae, a pathogenic bacterium that causes severe diarrheal diseases in humans. Specifically, the impact of the distinct environmental temperatures encountered during infection within a host (37°C) and transmission between hosts (25°C) on shuA expression is investigated. We show that shuA expression is subject to temperature-dependent post-transcriptional regulation resulting in increased ShuA production at 37°C. The observed thermoregulation is mediated by nucleic acid sequences within the 5′ untranslated region. In addition, we have identified similar nucleotide sequences within the 5′ untranslated region of the orthologous chuA transcript of enteropathogenic E. coli and have demonstrated that it also functions to confer temperature-dependent post-transcriptional regulation. In both function and predicted structure, the regulatory element within the shuA and chuA 5′ untranslated regions closely resembles a FourU RNA thermometer, a zipper-like RNA structure that occludes the Shine-Dalgarno sequence at low temperatures. Increased production of ShuA and ChuA in response to the host body temperature allows for maximal production of these heme acquisition factors within the environment where S. dysenteriae and pathogenic E. coli strains would encounter heme, a host-specific iron source. PMID:23704938

Sex and virulence in Escherichia coli: an evolutionary perspective

PubMed Central

Wirth, Thierry; Falush, Daniel; Lan, Ruiting; Colles, Frances; Mensa, Patience; Wieler, Lothar H; Karch, Helge; Reeves, Peter R; Maiden, Martin CJ; Ochman, Howard; Achtman, Mark

2006-01-01

Pathogenic Escherichia coli cause over 160 million cases of dysentery and one million deaths per year, whereas non-pathogenic E. coli constitute part of the normal intestinal flora of healthy mammals and birds. The evolutionary pathways underlying this dichotomy in bacterial lifestyle were investigated by multilocus sequence typing of a global collection of isolates. Specific pathogen types [enterohaemorrhagic E. coli, enteropathogenic E. coli, enteroinvasive E. coli, K1 and Shigella] have arisen independently and repeatedly in several lineages, whereas other lineages contain only few pathogens. Rates of evolution have accelerated in pathogenic lineages, culminating in highly virulent organisms whose genomic contents are altered frequently by increased rates of homologous recombination; thus, the evolution of virulence is linked to bacterial sex. This long-term pattern of evolution was observed in genes distributed throughout the genome, and thereby is the likely result of episodic selection for strains that can escape the host immune response. PMID:16689791

Comparative genomics of transport proteins in probiotic and pathogenic Escherichia coli and Salmonella enterica strains.

PubMed

Do, Jimmy; Zafar, Hassan; Saier, Milton H

2017-06-01

Escherichia coli is a genetically diverse species that can be pathogenic, probiotic, commensal, or a harmless laboratory strain. Pathogenic strains of E. coli cause urinary tract infections, diarrhea, hemorrhagic colitis, and pyelonephritis, while the two known probiotic E. coli strains combat inflammatory bowel disease and play a role in immunomodulation. Salmonella enterica, a close relative of E. coli, includes two important pathogenic serovars, Typhi and Typhimurium, causing typhoid fever and enterocolitis in humans, respectively, with the latter strain also causing a lethal typhoid fever-like disease in mice. In this study, we identify the transport systems and their substrates within seven E. coli strains: two probiotic strains, two extracellular pathogens, two intracellular pathogens, and K-12, as well as the two intracellular pathogenic S. enterica strains noted above. Transport systems characteristic of each probiotic or pathogenic species were thus identified, and the tabulated results obtained with all of these strains were compared. We found that the probiotic and pathogenic strains generally contain more iron-siderophore and sugar transporters than E. coli K-12. Pathogens have increased numbers of pore-forming toxins, protein secretion systems, decarboxylation-driven Na + exporters, electron flow-driven monovalent cation exporters, and putative transporters of unknown function compared to the probiotic strains. Both pathogens and probiotic strains encode metabolite transporters that reflect their intracellular versus extracellular environments. The results indicate that the probiotic strains live extracellularly. It seems that relatively few virulence factors can convert a beneficial or commensal microorganism into a pathogen. Taken together, the results reveal the distinguishing features of these strains and provide a starting point for future engineering of beneficial enteric bacteria. Copyright © 2017 Elsevier Ltd. All rights reserved.

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.

Translation inhibition of the developmental cycle protein HctA by the small RNA IhtA is conserved across Chlamydia.

PubMed

Tattersall, Jeremiah; Rao, Geeta Vittal; Runac, Justin; Hackstadt, Ted; Grieshaber, Scott S; Grieshaber, Nicole A

2012-01-01

The developmental cycle of the obligate intracellular pathogen Chlamydia trachomatis serovar L2 is controlled in part by the small non-coding RNA (sRNA), IhtA. All Chlamydia alternate in a regulated fashion between the infectious elementary body (EB) and the replicative reticulate body (RB) which asynchronously re-differentiates back to the terminal EB form at the end of the cycle. The histone like protein HctA is central to RB:EB differentiation late in the cycle as it binds to and occludes the genome, thereby repressing transcription and translation. The sRNA IhtA is a critical component of this regulatory loop as it represses translation of hctA until late in infection at which point IhtA transcription decreases, allowing HctA expression to occur and RB to EB differentiation to proceed. It has been reported that IhtA is expressed during infection by the human pathogens C. trachomatis serovars L2, D and L2b and C. pneumoniae. We show in this work that IhtA is also expressed by the animal pathogens C. caviae and C. muridarum. Expression of HctA in E. coli is lethal and co-expression of IhtA relieves this phenotype. To determine if regulation of HctA by IhtA is a conserved mechanism across pathogenic chlamydial species, we cloned hctA and ihtA from C. trachomatis serovar D, C. muridarum, C. caviae and C. pneumoniae and assayed for rescue of growth repression in E. coli co-expression studies. In each case, co-expression of ihtA with the cognate hctA resulted in relief of growth repression. In addition, expression of each chlamydial species IhtA rescued the lethal phenotype of C. trachomatis serovar L2 HctA expression. As biolayer interferometry studies indicate that IhtA interacts directly with hctA message for all species tested, we predict that conserved sequences of IhtA are necessary for function and/or binding.

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.

Pertussis Toxin Exploits Specific Host Cell Signaling Pathways for Promoting Invasion and Translocation of Escherichia coli K1 RS218 in Human Brain-derived Microvascular Endothelial Cells.

PubMed

Karassek, Sascha; Starost, Laura; Solbach, Johanna; Greune, Lilo; Sano, Yasuteru; Kanda, Takashi; Kim, KwangSik; Schmidt, M Alexander

2015-10-09

Pertussis toxin (PTx), an AB5 toxin and major virulence factor of the whooping cough-causing pathogen Bordetella pertussis, has been shown to affect the blood-brain barrier. Dysfunction of the blood-brain barrier may facilitate penetration of bacterial pathogens into the brain, such as Escherichia coli K1 (RS218). In this study, we investigated the influence of PTx on blood-brain barrier permissiveness to E. coli infection using human brain-derived endothelial HBMEC and TY10 cells as in vitro models. Our results indicate that PTx acts at several key points of host cell intracellular signaling pathways, which are also affected by E. coli K1 RS218 infection. Application of PTx increased the expression of the pathogen binding receptor gp96. Further, we found an activation of STAT3 and of the small GTPase Rac1, which have been described as being essential for bacterial invasion involving host cell actin cytoskeleton rearrangements at the bacterial entry site. In addition, we showed that PTx induces a remarkable relocation of VE-cadherin and β-catenin from intercellular junctions. The observed changes in host cell signaling molecules were accompanied by differences in intracellular calcium levels, which might act as a second messenger system for PTx. In summary, PTx not only facilitates invasion of E. coli K1 RS218 by activating essential signaling cascades; it also affects intercellular barriers to increase paracellular translocation. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Identification of novel bacteriophage peptides using a combination of gene sequence LC-MS-MS analysis and BLASTP

USDA-ARS?s Scientific Manuscript database

Introduction: In an effort to characterize novel bacteriophage with lytic activity against pathogenic E.coli associated with foodborne illness, gene sequencing and mass spectrometry have been used to identify expressed peptides which differentiate isolated bacteriophage from other known phage. Here,...

Sensitivity of pathogenic and attenuated E. coli O157:H7 strains to ultraviolet-C light as assessed by conventional plating methods and ethidium monoazide-PCR

USDA-ARS?s Scientific Manuscript database

In this study, the UV-C sensitivity of six pathogenic E. coli O157:H7 strains associated with recent outbreaks of foodborne illnesses and four attenuated E. coli O157:H7 strains was investigated. Futhermore, the mechanism of UV-C impact on two pathogenic E. coli strains with different UV-C sensitiv...

Escherichia coli, Salmonella, and Mycobacterium avium subsp. paratuberculosis in wild European starlings at a Kansas cattle feedlot.

PubMed

Gaukler, Shannon M; Linz, George M; Sherwood, Julie S; Dyer, Neil W; Bleier, William J; Wannemuehler, Yvonne M; Nolan, Lisa K; Logue, Catherine M

2009-12-01

The prevalence of Escherichia coli, Salmonella spp., and Mycobacterium avium subsp. paratuberculosis isolated from the feces of wild European starlings (Sturnus vulgaris) humanely trapped at a feedlot in central Kansas was assessed. All E. coli and Salmonella isolates recovered were tested for antimicrobial susceptibility using National Antimicrobial Resistance Monitoring System panels and the E. coli isolates were classified as to their content of genes associated with pathogenic E. coli of birds and cattle, including cvaC, iroN2, ompTp, hlyF2, eitC, iss, iutA, ireA, papC, stxI, stxII, sta, K99, F41, and eae. Escherichia coli O157:H7 and Mycobacterium avium subsp. paratuberculosis were not detected and Salmonella was isolated from only three samples, two of which displayed antimicrobial resistance. Approximately half of the E. coli isolates were resistant to antimicrobial agents with 96% showing resistance to tetracycline. Only one isolate was positive for a single gene associated with bovine pathogenic E. coli. An interesting finding of this study was that 5% of the E. coli isolates tested met the criteria established for identification as avian pathogenic E. coli (APEC). Thus these findings suggest that starlings are not a significant source of Salmonella spp., Mycobacterium avium subsp. paratuberculosis, E. coli O157, or other shiga toxin-producing E. coli in this feedlot. However, they may have the potential to spread APEC, an important pathogen of poultry and a potential pathogen to human beings.

Expression in Escherichia coli, purification, refolding and antifungal activity of an osmotin from Solanum nigrum

PubMed Central

Campos, Magnólia de A; Silva, Marilia S; Magalhães, Cláudio P; Ribeiro, Simone G; Sarto, Rafael PD; Vieira, Eduardo A; Grossi de Sá, Maria F

2008-01-01

Background Heterologous protein expression in microorganisms may contribute to identify and demonstrate antifungal activity of novel proteins. The Solanum nigrum osmotin-like protein (SnOLP) gene encodes a member of pathogenesis-related (PR) proteins, from the PR-5 sub-group, the last comprising several proteins with different functions, including antifungal activity. Based on deduced amino acid sequence of SnOLP, computer modeling produced a tertiary structure which is indicative of antifungal activity. Results To validate the potential antifungal activity of SnOLP, a hexahistidine-tagged mature SnOLP form was overexpressed in Escherichia coli M15 strain carried out by a pQE30 vector construction. The urea solubilized His6-tagged mature SnOLP protein was affinity-purified by immobilized-metal (Ni2+) affinity column chromatography. As SnOLP requires the correct formation of eight disulfide bonds, not correctly formed in bacterial cells, we adapted an in vitro method to refold the E. coli expressed SnOLP by using reduced:oxidized gluthatione redox buffer. This method generated biologically active conformations of the recombinant mature SnOLP, which exerted antifungal action towards plant pathogenic fungi (Fusarium solani f. sp.glycines, Colletotrichum spp., Macrophomina phaseolina) and oomycete (Phytophthora nicotiana var. parasitica) under in vitro conditions. Conclusion Since SnOLP displays activity against economically important plant pathogenic fungi and oomycete, it represents a novel PR-5 protein with promising utility for biotechnological applications. PMID:18334031

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

Influence of medium components on the expression of recombinant lipoproteins in Escherichia coli.

PubMed

Tseng, Chi-Ling; Leng, Chih-Hsiang

2012-02-01

Bacterial lipoproteins are crucial antigens for protective immunity against bacterial pathogens. Expression of exogenous lipoproteins in Escherichia coli at high levels is thought to be an extremely difficult endeavor because it frequently results in incomplete or absent lipid modification. Previously, we identified a fusion sequence (D1) from a Neisseria meningitidis lipoprotein that induced a non-lipidated protein, E3 (the domain III of the dengue virus envelope protein), to become lipidated. However, without optimizing the growth conditions, some of the D1-fusion proteins were not lipidated. Here, we report the influence of medium components on the expression of recombinant lipoproteins in E. coli. For high-level expression of mature lipoproteins in the C43 (DE3) strain, M9 medium was better than M63 and the rich medium. Furthermore, we analyzed the influence of other media factors (including nitrogen and carbon sources, phosphate, ferrous ions, calcium, magnesium, and pH) on the levels of lipoprotein expression. The results showed that excess nitrogen sources and phosphate in M9 medium could increase the amount of immature lipoproteins, and glucose was a better carbon source than glycerol for expressing mature lipoproteins. We also found that lipoproteins tended to be completely processed in the alkaline environment, even in the nutrient-rich medium. Additional constructs expressing different immunogens or lipid signal peptides as targets were also utilized, demonstrating that these targets could be expressed as completely mature lipoproteins in the M9 medium but not in the rich medium. Our results provide the useful information for expressing mature exogenous lipoproteins in E. coli.

Outbreak of extended-spectrum β-lactamase-producing Escherichia coli transmitted through breast milk sharing in a neonatal intensive care unit.

PubMed

Nakamura, K; Kaneko, M; Abe, Y; Yamamoto, N; Mori, H; Yoshida, A; Ohashi, K; Miura, S; Yang, T T; Momoi, N; Kanemitsu, K

2016-01-01

Routine surveillance in a neonatal intensive care unit (NICU) showed an increased detection of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli (ESBL-E. coli) in August 2012, following nearly a year without detection. To describe the investigation and interventions by a hospital infection control team of an outbreak of ESBL-E. coli in a NICU. Six neonates with positive cultures of ESBL-E. coli (five with respiratory colonization, one with a urinary tract infection), control infants who were negative for ESBL-E. coli during the study period, and mothers who donated their breast milk were included. A case-control study was performed to identify possible risk factors for positive ESBL-E. coli cultures and molecular typing of isolated strains by pulsed-field gel electrophoresis. The odds ratio for ESBL-E. coli infection after receiving shared unpasteurized breast milk during the study period was 49.17 (95% confidence interval: 6.02-354.68; P < 0.05). The pulsed-field gel electrophoresis pattern showed that all strains were identical, and the same pathogen was detected in freshly expressed milk of a particular donor. After ceasing the breast milk sharing, the outbreak was successfully terminated. This outbreak indicates that contamination of milk packs can result in transmission of a drug-resistant pathogen to newborn infants. Providers of human breast milk need to be aware of the necessity for low-temperature pasteurization and bacterial cultures, which should be conducted before and after freezing, before prescribing to infants. Copyright © 2015 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

A novel hemagglutinin protein produced in bacteria protects chickens against H5N1 highly pathogenic avian influenza viruses by inducing H5 subtype-specific neutralizing antibodies

PubMed Central

Sączyńska, Violetta; Romanik, Agnieszka; Florys, Katarzyna; Cecuda-Adamczewska, Violetta; Kęsik-Brodacka, Małgorzata; Śmietanka, Krzysztof; Olszewska, Monika; Domańska-Blicharz, Katarzyna; Minta, Zenon; Szewczyk, Bogusław; Płucienniczak, Grażyna; Płucienniczak, Andrzej

2017-01-01

The highly pathogenic (HP) H5N1 avian influenza viruses (AIVs) cause a mortality rate of up to 100% in infected chickens and pose a permanent pandemic threat. Attempts to obtain effective vaccines against H5N1 HPAIVs have focused on hemagglutinin (HA), an immunodominant viral antigen capable of eliciting neutralizing antibodies. The vast majority of vaccine projects have been performed using eukaryotic expression systems. In contrast, we used a bacterial expression system to produce vaccine HA protein (bacterial HA) according to our own design. The HA protein with the sequence of the H5N1 HPAIV strain was efficiently expressed in Escherichia coli, recovered in the form of inclusion bodies and refolded by dilution between two chromatographic purification steps. Antigenicity studies showed that the resulting antigen, referred to as rH5-E. coli, preserves conformational epitopes targeted by antibodies specific for H5-subtype HAs, inhibiting hemagglutination and/or neutralizing influenza viruses in vitro. The proper conformation of this protein and its ability to form functional oligomers were confirmed by a hemagglutination test. Consistent with the biochemical characteristics, prime-boost immunizations with adjuvanted rH5-E. coli protected 100% and 70% of specific pathogen-free, layer-type chickens against challenge with homologous and heterologous H5N1 HPAIVs, respectively. The observed protection was related to the positivity in the FluAC H5 test (IDVet) but not to hemagglutination-inhibiting antibody titers. Due to full protection, the effective contact transmission of the homologous challenge virus did not occur. Survivors from both challenges did not or only transiently shed the viruses, as established by viral RNA detection in oropharyngeal and cloacal swabs. Our results demonstrate that vaccination with rH5-E. coli could confer control of H5N1 HPAIV infection and transmission rates in chicken flocks, accompanied by reduced virus shedding. Moreover, the role of H5 subtype-specific neutralizing antibodies in anti-influenza immunity and a novel correlate of protection are indicated. PMID:28212428

Proteomic Analysis to Elucidate the Antibacterial Action of Silver Ions Against Bovine Mastitis Pathogens.

PubMed

Kang, Seog Jin; Cho, Yong Il; Kim, Ki Hyun; Cho, Eun Seok

2016-05-01

Silver ions act as a powerful, broad-spectrum antimicrobial agent and are known to kill over 650 different kinds of pathogens. We investigated the protein expression pattern and identity after silver ion treatment in Escherichia coli and Staphylococcus aureus, which are primarily responsible for the majority of bovine mastitis cases using proteomics. Two-dimensional electrophoresis showed that silver ion treatment significantly reduced 5 spot's density in E. coli and S. aureus, respectively. We identified 10 proteins (alkyl hydroperoxide reductase C22 subunit, phosphoglucomutase, fructose-1-phosphate kinase, putative carbamoyl transferase, alpha-galactosidase, carbamate kinase, ornithine transcarbamoylase, fumarate hydratase class II, alcohol dehydrogenase, and conserved hypothetical protein) by matrix-assisted laser desorption ionization time of flight (MALDI-TOF). These results demonstrated that silver ions have bactericidal effects through energy deprivation, inhibition of DNA replication, and accumulation of oxidants in bovine mastitis pathogens and suggested that silver ions can be applied for the treatment of bovine mastitis.

Diarrheagenic Escherichia coli

PubMed Central

Nataro, James P.; Kaper, James B.

1998-01-01

Escherichia coli is the predominant nonpathogenic facultative flora of the human intestine. Some E. coli strains, however, have developed the ability to cause disease of the gastrointestinal, urinary, or central nervous system in even the most robust human hosts. Diarrheagenic strains of E. coli can be divided into at least six different categories with corresponding distinct pathogenic schemes. Taken together, these organisms probably represent the most common cause of pediatric diarrhea worldwide. Several distinct clinical syndromes accompany infection with diarrheagenic E. coli categories, including traveler’s diarrhea (enterotoxigenic E. coli), hemorrhagic colitis and hemolytic-uremic syndrome (enterohemorrhagic E. coli), persistent diarrhea (enteroaggregative E. coli), and watery diarrhea of infants (enteropathogenic E. coli). This review discusses the current level of understanding of the pathogenesis of the diarrheagenic E. coli strains and describes how their pathogenic schemes underlie the clinical manifestations, diagnostic approach, and epidemiologic investigation of these important pathogens. PMID:9457432

Droplet microfluidics for amplification-free genetic detection of single cells.

PubMed

Rane, Tushar D; Zec, Helena C; Puleo, Chris; Lee, Abraham P; Wang, Tza-Huei

2012-09-21

In this article we present a novel droplet microfluidic chip enabling amplification-free detection of single pathogenic cells. The device streamlines multiple functionalities to carry out sample digitization, cell lysis, probe-target hybridization for subsequent fluorescent detection. A peptide nucleic acid fluorescence resonance energy transfer probe (PNA beacon) is used to detect 16S rRNA present in pathogenic cells. Initially the sensitivity and quantification abilities of the platform are tested using a synthetic target mimicking the actual expression level of 16S rRNA in single cells. The capability of the device to perform "sample-to-answer" pathogen detection of single cells is demonstrated using E. coli as a model pathogen.

Role of curli and plant cultivation conditions on Escherichia coli O157:H7 internalization into spinach grown on hydroponics and in soil.

PubMed

Macarisin, Dumitru; Patel, Jitendra; Sharma, Vijay K

2014-03-03

Contamination of fresh produce could represent a public health concern because no terminal kill step is applied during harvest or at the processing facility to kill pathogens. In addition, once contaminated, pathogens may internalize into produce and be protected from disinfectants during the postharvest processing step. The objective of the current study was to determine the potential internalization of Escherichia coli O157:H7 into spinach roots and subsequent transfer to the edible parts. Because curli are involved in biofilm formation, we investigated whether their presence influence the internalization of E. coli O157:H7 into spinach. Further, the effect of the spinach cultivar on E. coli O157:H7 internalization was evaluated. Spinach plants were grown in contaminated soil as well as hydroponically to prevent mechanical wounding of the roots and inadvertent transfer of pathogens from the contamination source to the non-exposed plant surfaces. Results showed that E. coli O157:H7 could internalize into hydroponically grown intact spinach plants through the root system and move to the stem and leaf level. The incidence of internalization was significantly higher in hydroponically grown plants when roots were exposed to 7 log CFU/mL compared to those exposed to 5 log CFU/mL. The effect of cultivar on E. coli O157:H7 internalization was not significant (P>0.05) for the analyzed spinach varieties, internalization incidences showing almost equal distribution between Space and Waitiki, 49.06% and 50.94% respectively. Wounding of the root system in hydroponically grown spinach increased the incidence of E. coli O157:H7 internalization and translocation to the edible portions of the plant. Experimental contamination of the plants grown in soil resulted in a greater number of internalization events then in those grown hydroponically, suggesting that E. coli O157:H7 internalization is dependent on root damage, which is more likely to occur when plants are grown in soil. Curli expression by E. coli O157:H7 had no significant effect on its root uptake by spinach plants. Published by Elsevier B.V.

β-1,3-1,4-glucanase gene from Bacillus velezensis ZJ20 exerts antifungal effect on plant pathogenic fungi.

PubMed

Xu, Ting; Zhu, Tianhui; Li, Shujiang

2016-02-01

Bacillus velezensis is a known antifungal bacteria. To understand the role of β-1, 3-1, 4-glucanase played on B. velezensis about the mechanism which exerts effect on fungi, we isolated and cloned the β-1, 3-1, 4-glucanase gene (Bglu1) from B. velezensis ZJ20. The Bglu1 open reading frame was 732 bp that encoded a protein with 243 amino acids and a calculated molecular weight of 27.3 kDa. The same gene without the signal peptide, termed Bglu2, was also cloned and expressed in E. coli BL21. Among the two variants, only Bglu2 protein was expressed. Purified Bglu2 could be eluted with imidazole solution at concentrations ranging from 100 to 500 mM although the highest expression was observed at 150 and 200 mM and the purest was at 500 mM. In addition, activity of the crude enzyme was 1527 U ml(-1) and the highest activity of the purified enzyme was 1706 U ml(-1). The purified β-1, 3-1, 4-glucanase had activity on a wide range of pH and temperatures and displayed optimal activity at pH 5.0 and 35 °C. More importantly, the mycelial morphology of three pathogenic fungi was destroyed by the purified β-1, 3-1, 4-glucanase. In conclusion, β-1, 3-1, 4-glucanase from B. velezensis ZJ20 can be highly expressed in E. coli BL21 and the recombinant protein is pathogenic to fungi.

Genome-Based Comparison of Cyclic Di-GMP Signaling in Pathogenic and Commensal Escherichia coli Strains

PubMed Central

Povolotsky, Tatyana L.

2015-01-01

ABSTRACT The ubiquitous bacterial second messenger cyclic di-GMP (c-di-GMP) has recently become prominent as a trigger for biofilm formation in many bacteria. It is generated by diguanylate cyclases (DGCs; with GGDEF domains) and degraded by specific phosphodiesterases (PDEs; containing either EAL or HD-GYP domains). Most bacterial species contain multiples of these proteins with some having specific functions that are based on direct molecular interactions in addition to their enzymatic activities. Escherichia coli K-12 laboratory strains feature 29 genes encoding GGDEF and/or EAL domains, resulting in a set of 12 DGCs, 13 PDEs, and four enzymatically inactive “degenerate” proteins that act by direct macromolecular interactions. We present here a comparative analysis of GGDEF/EAL domain-encoding genes in 61 genomes of pathogenic, commensal, and probiotic E. coli strains (including enteric pathogens such as enteroaggregative, enterohemorrhagic, enteropathogenic, enterotoxigenic, and adherent and invasive Escherichia coli and the 2011 German outbreak O104:H4 strain, as well as extraintestinal pathogenic E. coli, such as uropathogenic and meningitis-associated E. coli). We describe additional genes for two membrane-associated DGCs (DgcX and DgcY) and four PDEs (the membrane-associated PdeT, as well as the EAL domain-only proteins PdeW, PdeX, and PdeY), thus showing the pangenome of E. coli to contain at least 35 GGDEF/EAL domain proteins. A core set of only eight proteins is absolutely conserved in all 61 strains: DgcC (YaiC), DgcI (YliF), PdeB (YlaB), PdeH (YhjH), PdeK (YhjK), PdeN (Rtn), and the degenerate proteins CsrD and CdgI (YeaI). In all other GGDEF/EAL domain genes, diverse point and frameshift mutations, as well as small or large deletions, were discovered in various strains. IMPORTANCE Our analysis reveals interesting trends in pathogenic Escherichia coli that could reflect different host cell adherence mechanisms. These may either benefit from or be counteracted by the c-di-GMP-stimulated production of amyloid curli fibers and cellulose. Thus, EAEC, which adhere in a “stacked brick” biofilm mode, have a potential for high c-di-GMP accumulation due to DgcX, a strongly expressed additional DGC. In contrast, EHEC and UPEC, which use alternative adherence mechanisms, tend to have extra PDEs, suggesting that low cellular c-di-GMP levels are crucial for these strains under specific conditions. Overall, our study also indicates that GGDEF/EAL domain proteins evolve rapidly and thereby contribute to adaptation to host-specific and environmental niches of various types of E. coli. PMID:26303830

Genome-Based Comparison of Cyclic Di-GMP Signaling in Pathogenic and Commensal Escherichia coli Strains.

PubMed

Povolotsky, Tatyana L; Hengge, Regine

2016-01-01

The ubiquitous bacterial second messenger cyclic di-GMP (c-di-GMP) has recently become prominent as a trigger for biofilm formation in many bacteria. It is generated by diguanylate cyclases (DGCs; with GGDEF domains) and degraded by specific phosphodiesterases (PDEs; containing either EAL or HD-GYP domains). Most bacterial species contain multiples of these proteins with some having specific functions that are based on direct molecular interactions in addition to their enzymatic activities. Escherichia coli K-12 laboratory strains feature 29 genes encoding GGDEF and/or EAL domains, resulting in a set of 12 DGCs, 13 PDEs, and four enzymatically inactive "degenerate" proteins that act by direct macromolecular interactions. We present here a comparative analysis of GGDEF/EAL domain-encoding genes in 61 genomes of pathogenic, commensal, and probiotic E. coli strains (including enteric pathogens such as enteroaggregative, enterohemorrhagic, enteropathogenic, enterotoxigenic, and adherent and invasive Escherichia coli and the 2011 German outbreak O104:H4 strain, as well as extraintestinal pathogenic E. coli, such as uropathogenic and meningitis-associated E. coli). We describe additional genes for two membrane-associated DGCs (DgcX and DgcY) and four PDEs (the membrane-associated PdeT, as well as the EAL domain-only proteins PdeW, PdeX, and PdeY), thus showing the pangenome of E. coli to contain at least 35 GGDEF/EAL domain proteins. A core set of only eight proteins is absolutely conserved in all 61 strains: DgcC (YaiC), DgcI (YliF), PdeB (YlaB), PdeH (YhjH), PdeK (YhjK), PdeN (Rtn), and the degenerate proteins CsrD and CdgI (YeaI). In all other GGDEF/EAL domain genes, diverse point and frameshift mutations, as well as small or large deletions, were discovered in various strains. Our analysis reveals interesting trends in pathogenic Escherichia coli that could reflect different host cell adherence mechanisms. These may either benefit from or be counteracted by the c-di-GMP-stimulated production of amyloid curli fibers and cellulose. Thus, EAEC, which adhere in a "stacked brick" biofilm mode, have a potential for high c-di-GMP accumulation due to DgcX, a strongly expressed additional DGC. In contrast, EHEC and UPEC, which use alternative adherence mechanisms, tend to have extra PDEs, suggesting that low cellular c-di-GMP levels are crucial for these strains under specific conditions. Overall, our study also indicates that GGDEF/EAL domain proteins evolve rapidly and thereby contribute to adaptation to host-specific and environmental niches of various types of E. coli. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Fimbria-Encoding Gene yadC Has a Pleiotropic Effect on Several Biological Characteristics and Plays a Role in Avian Pathogenic Escherichia coli Pathogenicity.

PubMed

Verma, Renu; Rojas, Thaís Cabrera Galvão; Maluta, Renato Pariz; Leite, Janaína Luisa; da Silva, Livia Pilatti Mendes; Nakazato, Gerson; Dias da Silveira, Wanderley

2016-01-01

The extraintestinal pathogen termed avian pathogenic Escherichia coli (APEC) is known to cause colibacillosis in chickens. The molecular basis of APEC pathogenesis is not fully elucidated yet. In this work, we deleted a component of the Yad gene cluster (yadC) in order to understand the role of Yad in the pathogenicity of the APEC strain SCI-07. In vitro, the transcription level of yadC was upregulated at 41°C and downregulated at 22°C. The yadC expression in vivo was more pronounced in lungs than in spleen, suggesting a role in the early steps of the infection. Chicks infected with the wild-type and mutant strains presented, respectively, 80% and 50% mortality rates. The ΔyadC strain presented a slightly decreased ability to adhere to HeLa cells with or without the d-mannose analog compared with the wild type. Real-time PCR (RT-PCR) assays showed that fimH was downregulated (P < 0.05) and csgA and ecpA were slightly upregulated in the mutant strain, showing that yadC modulates expression of other fimbriae. Bacterial internalization studies showed that the ΔyadC strain had a lower number of intracellular bacteria recovered from Hep-2 cells and HD11 cells than the wild-type strain (P < 0.05). Motility assays in soft agar demonstrated that the ΔyadC strain was less motile than the wild type (P < 0.01). Curiously, flagellum-associated genes were not dramatically downregulated in the ΔyadC strain. Taken together, the results show that the fimbrial adhesin Yad contributes to the pathogenicity and modulates different biological characteristics of the APEC strain SCI-07. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Bioinformatics comparisons of RNA-binding proteins of pathogenic and non-pathogenic Escherichia coli strains reveal novel virulence factors.

PubMed

Ghosh, Pritha; Sowdhamini, Ramanathan

2017-08-24

Pathogenic bacteria have evolved various strategies to counteract host defences. They are also exposed to environments that are undergoing constant changes. Hence, in order to survive, bacteria must adapt themselves to the changing environmental conditions by performing regulations at the transcriptional and/or post-transcriptional levels. Roles of RNA-binding proteins (RBPs) as virulence factors have been very well studied. Here, we have used a sequence search-based method to compare and contrast the proteomes of 16 pathogenic and three non-pathogenic E. coli strains as well as to obtain a global picture of the RBP landscape (RBPome) in E. coli. Our results show that there are no significant differences in the percentage of RBPs encoded by the pathogenic and the non-pathogenic E. coli strains. The differences in the types of Pfam domains as well as Pfam RNA-binding domains, encoded by these two classes of E. coli strains, are also insignificant. The complete and distinct RBPome of E. coli has been established by studying all known E. coli strains till date. We have also identified RBPs that are exclusive to pathogenic strains, and most of them can be exploited as drug targets since they appear to be non-homologous to their human host proteins. Many of these pathogen-specific proteins were uncharacterised and their identities could be resolved on the basis of sequence homology searches with known proteins. Detailed structural modelling, molecular dynamics simulations and sequence comparisons have been pursued for selected examples to understand differences in stability and RNA-binding. The approach used in this paper to cross-compare proteomes of pathogenic and non-pathogenic strains may also be extended to other bacterial or even eukaryotic proteomes to understand interesting differences in their RBPomes. The pathogen-specific RBPs reported in this study, may also be taken up further for clinical trials and/or experimental validations.

Analysis of the Genome Structure of the Nonpathogenic Probiotic Escherichia coli Strain Nissle 1917

PubMed Central

Grozdanov, Lubomir; Raasch, Carsten; Schulze, Jürgen; Sonnenborn, Ulrich; Gottschalk, Gerhard; Hacker, Jörg; Dobrindt, Ulrich

2004-01-01

Nonpathogenic Escherichia coli strain Nissle 1917 (O6:K5:H1) is used as a probiotic agent in medicine, mainly for the treatment of various gastroenterological diseases. To gain insight on the genetic level into its properties of colonization and commensalism, this strain's genome structure has been analyzed by three approaches: (i) sequence context screening of tRNA genes as a potential indication of chromosomal integration of horizontally acquired DNA, (ii) sequence analysis of 280 kb of genomic islands (GEIs) coding for important fitness factors, and (iii) comparison of Nissle 1917 genome content with that of other E. coli strains by DNA-DNA hybridization. PCR-based screening of 324 nonpathogenic and pathogenic E. coli isolates of different origins revealed that some chromosomal regions are frequently detectable in nonpathogenic E. coli and also among extraintestinal and intestinal pathogenic strains. Many known fitness factor determinants of strain Nissle 1917 are localized on four GEIs which have been partially sequenced and analyzed. Comparison of these data with the available knowledge of the genome structure of E. coli K-12 strain MG1655 and of uropathogenic E. coli O6 strains CFT073 and 536 revealed structural similarities on the genomic level, especially between the E. coli O6 strains. The lack of defined virulence factors (i.e., alpha-hemolysin, P-fimbrial adhesins, and the semirough lipopolysaccharide phenotype) combined with the expression of fitness factors such as microcins, different iron uptake systems, adhesins, and proteases, which may support its survival and successful colonization of the human gut, most likely contributes to the probiotic character of E. coli strain Nissle 1917. PMID:15292145

Occurrence and distribution of fecal indicator bacteria and gene markers of pathogenic bacteria in Great Lakes tributaries, March-October 2011

USGS Publications Warehouse

Brennan, Angela K.; Johnson, Heather E.; Totten, Alexander R.; Duris, Joseph W.

2015-01-01

Water samples were analyzed for the presence of FIB concentrations (FIB; fecal coliform bacteria, Escherichia coli [E. coli], and enterococci) by using membrane filtration and serial dilution methods. The resulting enrichments from standard culturing of the samples were then analyzed by using polymerase chain reaction (PCR) to determine the occurrence of pathogen gene markers for Shigella species, Campylobacter jejuni and coli, Salmonellaspecies, and pathogenic E. coli, including Shiga toxin-producing E. coli (STEC).

Deciphering the roles of outer membrane protein A extracellular loops in the pathogenesis of Escherichia coli K1 meningitis.

PubMed

Mittal, Rahul; Krishnan, Subramanian; Gonzalez-Gomez, Ignacio; Prasadarao, Nemani V

2011-01-21

Outer membrane protein A (OmpA) has been implicated as an important virulence factor in several gram-negative bacterial infections such as Escherichia coli K1, a leading cause of neonatal meningitis associated with significant mortality and morbidity. In this study, we generated E. coli K1 mutants that express OmpA in which three or four amino acids from various extracellular loops were changed to alanines, and we examined their ability to survive in several immune cells. We observed that loop regions 1 and 2 play an important role in the survival of E. coli K1 inside neutrophils and dendritic cells, and loop regions 1 and 3 are needed for survival in macrophages. Concomitantly, E. coli K1 mutants expressing loop 1 and 2 mutations were unable to cause meningitis in a newborn mouse model. Of note, mutations in loop 4 of OmpA enhance the severity of the pathogenesis by allowing the pathogen to survive better in circulation and to produce high bacteremia levels. These results demonstrate, for the first time, the roles played by different regions of extracellular loops of OmpA of E. coli K1 in the pathogenesis of meningitis and may help in designing effective preventive strategies against this deadly disease.

Biological and Physicochemical Wastewater Treatment Processes Reduce the Prevalence of Virulent Escherichia coli

PubMed Central

Biswal, Basanta Kumar; Mazza, Alberto; Masson, Luke; Gehr, Ronald

2013-01-01

Effluents discharged from wastewater treatment plants are possible sources of pathogenic bacteria, including Escherichia coli, in the freshwater environment, and determining the possible selection of pathogens is important. This study evaluated the impact of activated sludge and physicochemical wastewater treatment processes on the prevalence of potentially virulent E. coli. A total of 719 E. coli isolates collected from four municipal plants in Québec before and after treatment were characterized by using a customized DNA microarray to determine the impact of treatment processes on the frequency of specific pathotypes and virulence genes. The percentages of potentially pathogenic E. coli isolates in the plant influents varied between 26 and 51%, and in the effluents, the percentages were 14 to 31%, for a reduction observed at all plants ranging between 14 and 45%. Pathotypes associated with extraintestinal pathogenic E. coli (ExPEC) were the most abundant at three of the four plants and represented 24% of all isolates, while intestinal pathogenic E. coli pathotypes (IPEC) represented 10% of the isolates. At the plant where ExPEC isolates were not the most abundant, a large number of isolates were classified as both ExPEC and IPEC; overall, 6% of the isolates were classified in both groups, with the majority being from the same plant. The reduction of the proportion of pathogenic E. coli could not be explained by the preferential loss of one virulence gene or one type of virulence factor; however, the quinolone resistance gene (qnrS) appears to enhance the loss of virulence genes, suggesting a mechanism involving the loss of pathogenicity islands. PMID:23160132

Captive and free-living urban pigeons (Columba livia) from Brazil as carriers of multidrug-resistant pathogenic Escherichia coli.

PubMed

Borges, Clarissa A; Maluta, Renato P; Beraldo, Lívia G; Cardozo, Marita V; Guastalli, Elisabete A L; Kariyawasam, Subhashinie; DebRoy, Chitrita; Ávila, Fernando A

2017-01-01

Thirty Escherichia coli isolates from captive and free-living pigeons in Brazil were characterised. Virulence-associated genes identified in pigeons included those which occur relatively frequently in avian pathogenic E. coli (APEC) from commercial poultry worldwide. Eleven of 30 E. coli isolates from pigeons, belonging mainly to B1 and B2 phylogenetic groups, had high or intermediate pathogenicity for 1-day-old chicks. The frequency of multi-drug resistant (MDR) E. coli in captive pigeons was relatively high and included one isolate positive for the extended-spectrum β-lactamase (ESBL) gene bla CTX-M-8 . Pulsed field gel electrophoresis (PFGE) showed high heterogeneity among isolates. There is potential for pigeons to transmit antibiotic resistant pathogenic E. coli to other species through environmental contamination or direct contact. Copyright © 2017 Elsevier Ltd. All rights reserved.

Genome sequence of E. coli O104:H4 leads to rapid development of a targeted antimicrobial agent against this emerging pathogen.

USDA-ARS?s Scientific Manuscript database

A recent widespread outbreak of Escherichia coli O104:H4 in Germany demonstrates the dynamic nature of emerging and re-emerging food-borne pathogens, particularly STECs and related pathogenic E. coli. Rapid genomic sequencing and public availability of these data from the German outbreak strain allo...

Mass spectrometry-based method of detecting and distinguishing type 1 and type 2 Shiga-like toxins in human serum

USDA-ARS?s Scientific Manuscript database

Shiga-like toxins (verotoxins) are a class of AB5 holotoxins that are responsible for the virulence associated with bacterial pathogens such as Shigella dysenteriae, shigatoxigenic and enterohemorrhagic strains of Escherichia coli (STEC and EHEC), and some Enterobacter strains. The actual expression...

Procurement and persistence of GFP-expressing Escherichia coli and Salmonella Typhimurium in male and female house flies exposed to cattle manure

USDA-ARS?s Scientific Manuscript database

Introduction: Adult house flies, Musca domestica L., are associated with animal manure and other microbe-rich substrates. Consequently, both sexes can acquire and potentially disseminate pathogenic bacteria to surrounding environments, including residential areas, via contaminated body parts and/or ...

Immunomodulation of Host Chitinase 3-Like 1 During a Mammary Pathogenic Escherichia coli Infection

PubMed Central

Breyne, Koen; Steenbrugge, Jonas; Demeyere, Kristel; Lee, Chun Geun; Elias, Jack A.; Petzl, Wolfram; Smith, David G. E.; Germon, Pierre; Meyer, Evelyne

2018-01-01

Chitin is a N-acetyl-d-glucosamine biopolymer that can be recognized by chitin-binding proteins. Although mammals lack chitin synthase, they induce proteins responsible for detecting chitin in response to bacterial infections. Our aim was to investigate whether chitinase 3-like 1 (CHI3L1) has a potential role in the innate immunity of the Escherichia coli (E. coli) infected mammary gland. CHI3L1 protein was found to be secreted in whey of naturally coliform-affected quarters compared to whey samples isolated from healthy udders. In addition, gene expression of CHI3L1 was confirmed in udder tissue of cows experimentally infected with a mammary pathogenic E. coli (MPEC) strain. Despite the known anatomical differences, the bovine udders’ innate immune response was mimicked by applying an experimental mouse model using MPEC or non-MPEC isolates. The effect of CHI3L1 expression in the murine mammary gland in response to coliform bacteria was investigated through the use of CHI3L1−/− mice as well as through treatment with either a pan-caspase inhibitor or chitin particles in wild-type mice. The local induction of CHI3L1 postinfection with different E. coli strains was demonstrated to be independent of both bacterial growth and mammary interleukin (IL)-8 levels. Indeed, CHI3L1 emerged as a regulator impacting on the transcytosis of Ly6G-positive cells from the interstitial space into the alveolar lumen of the mammary tissue. Furthermore, CHI3L1 was found to be upstream regulated by caspase activity and had a major downstream effect on the local pro-inflammatory cytokine profile, including IL-1beta, IL-6, and RANTES/CCL5. In conclusion, CHI3L1 was demonstrated to play a key role in the cytokine and caspase signaling during E. coli triggered inflammation of the mammary gland. PMID:29892291

Suppression subtractive hybridization identifies an autotransporter adhesin gene of E. coli IMT5155 specifically associated with avian pathogenic Escherichia coli (APEC).

PubMed

Dai, Jianjun; Wang, Shaohui; Guerlebeck, Doreen; Laturnus, Claudia; Guenther, Sebastian; Shi, Zhenyu; Lu, Chengping; Ewers, Christa

2010-09-09

Extraintestinal pathogenic E. coli (ExPEC) represent a phylogenetically diverse group of bacteria which are implicated in a large range of infections in humans and animals. Although subgroups of different ExPEC pathotypes, including uropathogenic, newborn meningitis causing, and avian pathogenic E. coli (APEC) share a number of virulence features, there still might be factors specifically contributing to the pathogenesis of a certain subset of strains or a distinct pathotype. Thus, we made use of suppression subtractive hybridization and compared APEC strain IMT5155 (O2:K1:H5; sequence type complex 95) with human uropathogenic E. coli strain CFT073 (O6:K2:H5; sequence type complex 73) to identify factors which may complete the currently existing model of APEC pathogenicity and further elucidate the position of this avian pathotype within the whole ExPEC group. Twenty-eight different genomic loci were identified, which are present in IMT5155 but not in CFT073. One of these loci contained a gene encoding a putative autotransporter adhesin. The open reading frame of the gene spans a 3,498 bp region leading to a putative 124-kDa adhesive protein. A specific antibody was raised against this protein and expression of the adhesin was shown under laboratory conditions. Adherence and adherence inhibition assays demonstrated a role for the corresponding protein in adhesion to DF-1 chicken fibroblasts. Sequence analyses revealed that the flanking regions of the chromosomally located gene contained sequences of mobile genetic elements, indicating a probable spread among different strains by horizontal gene transfer. In accordance with this hypothesis, the adhesin was found to be present not only in different phylogenetic groups of extraintestinal pathogenic but also of commensal E. coli strains, yielding a significant association with strains of avian origin. We identified a chromosomally located autotransporter gene in a highly virulent APEC strain which confers increased adherence of a non-fimbriated E. coli K-12 strain to a chicken fibroblast cell line. Even though flanked by mobile genetic elements and three different genetic regions upstream of the gene, most probably indicating horizontal gene transfer events, the adhesin gene was significantly linked with strains of avian origin. Due to the nucleotide sequence similarity of 98% to a recently published adhesin-related gene, located on plasmid pAPEC-O1-ColBM, the name aatA (APEC autotransporter adhesin A) was adopted from that study.Our data substantiate that AatA might not only be of relevance in APEC pathogenicity but also in facilitating their reservoir life style in the chicken intestine, which might pave the way for future intestinal preventive strategies.

Evaluation of Animal Genetic and Physiological Factors That Affect the Prevalence of Escherichia coli O157 in Cattle

PubMed Central

Jeon, Soo Jin; Elzo, Mauricio; DiLorenzo, Nicolas; Lamb, G. Cliff; Jeong, Kwang Cheol

2013-01-01

Controlling the prevalence of Escherichia coli O157 in cattle at the pre-harvest level is critical to reduce outbreaks of this pathogen in humans. Multilayers of factors including the environmental and bacterial factors modulate the colonization and persistence of E. coli O157 in cattle that serve as a reservoir of this pathogen. Here, we report animal factors contributing to the prevalence of E. coli O157 in cattle. We observe the lowest number of E. coli O157 in Brahman breed when compared with other crosses in an Angus-Brahman multibreed herd, and bulls excrete more E. coli O157 than steers in the pens where cattle were housed together. The presence of super-shedders, cattle excreting >105 CFU/rectal anal swab, increases the concentration of E. coli O157 in the pens; thereby super-shedders enhance transmission of this pathogen among cattle. Molecular subtyping analysis reveal only one subtype of E. coli O157 in the multibreed herd, indicating the variance in the levels of E. coli O157 in cattle is influenced by animal factors. Furthermore, strain tracking after relocation of the cattle to a commercial feedlot reveals farm-to-farm transmission of E. coli O157, likely via super-shedders. Our results reveal high risk factors in the prevalence of E. coli O157 in cattle whereby animal genetic and physiological factors influence whether this pathogen can persist in cattle at high concentration, providing insights to intervene this pathogen at the pre-harvest level. PMID:23405204

A systematic review and meta-analysis of the epidemiology of pathogenic Escherichia coli of calves and the role of calves as reservoirs for human pathogenic E. coli.

PubMed

Kolenda, Rafał; Burdukiewicz, Michał; Schierack, Peter

2015-01-01

Escherichia coli bacteria are the most common causes of diarrhea and septicemia in calves. Moreover, calves form a major reservoir for transmission of pathogenic E. coli to humans. Systematic reviews and meta-analyses of publications on E. coli as calf pathogens and the role of calves as reservoir have not been done so far. We reviewed studies between 1951 and 2013 reporting the presence of virulence associated factors (VAFs) in calf E. coli and extracted the following information: year(s) and country of sampling, animal number, health status, isolate number, VAF prevalence, serotypes, diagnostic methods, and biological assays. The prevalence of VAFs or E. coli pathotypes was compared between healthy and diarrheic animals and was analyzed for time courses. Together, 106 papers with 25,982 E. coli isolates from 27 countries tested for VAFs were included. F5, F17, and F41 fimbriae and heat-stable enterotoxin (ST) - VAFs of enterotoxigenic E. coli (ETEC) were significantly associated with calf diarrhea. On the contrary, ETEC VAF F4 fimbriae and heat-labile enterotoxin as well as enteropathogenic (EPEC), Shiga toxin-producing (STEC), and enterohemorrhagic E. coli (EHEC) were not associated with diarrhea. The prevalence increased overtime for ST-positive isolates, but decreased for F5- and STEC-positive isolates. Our study provides useful information about the history of scientific investigations performed in this domain so far, and helps to define etiological agents of calf disease, and to evaluate calves as reservoir hosts for human pathogenic E. coli.

The NAG Sensor NagC Regulates LEE Gene Expression and Contributes to Gut Colonization by Escherichia coli O157:H7.

PubMed

Le Bihan, Guillaume; Sicard, Jean-Félix; Garneau, Philippe; Bernalier-Donadille, Annick; Gobert, Alain P; Garrivier, Annie; Martin, Christine; Hay, Anthony G; Beaudry, Francis; Harel, Josée; Jubelin, Grégory

2017-01-01

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 are human pathogens responsible for bloody diarrhea and renal failures. EHEC employ a type 3 secretion system to attach directly to the human colonic epithelium. This structure is encoded by the locus of enterocyte effacement (LEE) whose expression is regulated in response to specific nutrients. In this study, we show that the mucin-derived sugars N-acetylglucosamine (NAG) and N-acetylneuraminic acid (NANA) inhibit EHEC adhesion to epithelial cells through down-regulation of LEE expression. The effect of NAG and NANA is dependent on NagC, a transcriptional repressor of the NAG catabolism in E. coli . We show that NagC is an activator of the LEE1 operon and a critical regulator for the colonization of mice intestine by EHEC. Finally, we demonstrate that NAG and NANA as well as the metabolic activity of Bacteroides thetaiotaomicron affect the in vivo fitness of EHEC in a NagC-dependent manner. This study highlights the role of NagC in coordinating metabolism and LEE expression in EHEC and in promoting EHEC colonization in vivo .

The NAG Sensor NagC Regulates LEE Gene Expression and Contributes to Gut Colonization by Escherichia coli O157:H7

PubMed Central

Le Bihan, Guillaume; Sicard, Jean-Félix; Garneau, Philippe; Bernalier-Donadille, Annick; Gobert, Alain P.; Garrivier, Annie; Martin, Christine; Hay, Anthony G.; Beaudry, Francis; Harel, Josée; Jubelin, Grégory

2017-01-01

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 are human pathogens responsible for bloody diarrhea and renal failures. EHEC employ a type 3 secretion system to attach directly to the human colonic epithelium. This structure is encoded by the locus of enterocyte effacement (LEE) whose expression is regulated in response to specific nutrients. In this study, we show that the mucin-derived sugars N-acetylglucosamine (NAG) and N-acetylneuraminic acid (NANA) inhibit EHEC adhesion to epithelial cells through down-regulation of LEE expression. The effect of NAG and NANA is dependent on NagC, a transcriptional repressor of the NAG catabolism in E. coli. We show that NagC is an activator of the LEE1 operon and a critical regulator for the colonization of mice intestine by EHEC. Finally, we demonstrate that NAG and NANA as well as the metabolic activity of Bacteroides thetaiotaomicron affect the in vivo fitness of EHEC in a NagC-dependent manner. This study highlights the role of NagC in coordinating metabolism and LEE expression in EHEC and in promoting EHEC colonization in vivo. PMID:28484684

Complete Genome Sequence of the Avian Pathogenic Escherichia coli Strain APEC O78

PubMed Central

Mangiamele, Paul; Nicholson, Bryon; Wannemuehler, Yvonne; Seemann, Torsten; Logue, Catherine M.; Li, Ganwu; Tivendale, Kelly A.

2013-01-01

Colibacillosis, caused by avian pathogenic Escherichia coli (APEC), is a significant disease, causing extensive animal and financial losses globally. Because of the significance of this disease, more knowledge is needed regarding APEC's mechanisms of virulence. Here, we present the fully closed genome sequence of a typical avian pathogenic E. coli strain belonging to the serogroup O78. PMID:23516182

Enterohaemorrhagic Escherichia coli gains a competitive advantage by using ethanolamine as a nitrogen source in the bovine intestinal content.

PubMed

Bertin, Yolande; Girardeau, J P; Chaucheyras-Durand, F; Lyan, Bernard; Pujos-Guillot, Estelle; Harel, Josée; Martin, Christine

2011-02-01

The bovine gastrointestinal tract is the main reservoir for enterohaemorrhagic Escherichia coli (EHEC) responsible for food-borne infections. Characterization of nutrients that promote the carriage of these pathogens by the ruminant would help to develop ecological strategies to reduce their survival in the bovine gastrointestinal tract. In this study, we show for the first time that free ethanolamine (EA) constitutes a nitrogen source for the O157:H7 EHEC strain EDL933 in the bovine intestinal content because of induction of the eut (ethanolamine utilization) gene cluster. In contrast, the eut gene cluster is absent in the genome of most species constituting the mammalian gut microbiota. Furthermore, the eutB gene (encoding a subunit of the enzyme that catalyses the release of ammonia from EA) is poorly expressed in non-pathogenic E. coli. Accordingly, EA is consumed by EHEC but is poorly metabolized by endogenous microbiota of the bovine small intestine, including commensal E. coli. Interestingly, the capacity to utilize EA as a nitrogen source confers a growth advantage to E. coli O157:H7 when the bacteria enter the stationary growth phase. These data demonstrate that EHEC strains take advantage of a nitrogen source that is not consumed by the resident microbiota, and suggest that EA represents an ecological niche favouring EHEC persistence in the bovine intestine.

Demonstration of the IgG antibody repertoire against the bacteria Escherichia coli in Chinese intravenous immunoglobulins.

PubMed

Ye, Shengliang; Lei, Min; Jiang, Peng; Liu, Fengjuan; Wang, Zongkui; Cao, Haijun; Du, Xi; Yuan, Jing; Chen, Yunhua; Ma, Li; Li, Changqing

2017-01-30

Intravenous immunoglobulin (IVIg) is produced by pooling plasma from thousands of healthy blood donors, and the diversity of the antibody is critical for the clinical efficacy of IVIg. This study investigated the antibody diversity of Chinese IVIg. Firstly, 2-dimensional gel electrophoresis and immunoblotting with protein extracts of Escherichia coli (E. coli) O157:H7 were used to study IgG antibody repertoire of 8 IVIg preparations from different Chinese manufacturers. This was followed by the identification of the antibody-reactive proteins of E. coli by mass spectrometry and the sequence similarity of the proteins was aligned by bioinformatics analysis. The results showed that all IVIg preparations expressed a large range of antibody reactivities against E. coli proteins. 94-238 antigens were recognized by the 8 IVIg preparations. 33 interesting target antigens were selected and identified as 29 different proteins, mainly including membrane proteins, molecular chaperones, metabolism enzymes, and proteins involved in cell cycle processes. Additionally, these antigens were highly conserved proteins which were found extensively in a variety of other pathogenic microorganisms. Our study indicated that Chinese IVIg preparations recognized a large range of high conserved proteins which play key roles in pathogenic microorganisms, and showed each IVIg had its own distinct antibody repertoire. Copyright © 2016 Elsevier B.V. All rights reserved.

Shiga toxin-converting phages and the emergence of new pathogenic Escherichia coli: a world in motion

PubMed Central

Tozzoli, Rosangela; Grande, Laura; Michelacci, Valeria; Ranieri, Paola; Maugliani, Antonella; Caprioli, Alfredo; Morabito, Stefano

2014-01-01

Shiga toxin (Stx)-producing Escherichia coli (STEC) are pathogenic E. coli causing diarrhea, hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). STEC are characterized by a constellation of virulence factors additional to Stx and have long been regarded as capable to cause HC and HUS when possessing the ability of inducing the attaching and effacing (A/E) lesion to the enterocyte, although strains isolated from such severe infections sometimes lack this virulence feature. Interestingly, the capability to cause the A/E lesion is shared with another E. coli pathogroup, the Enteropathogenic E. coli (EPEC). In the very recent times, a different type of STEC broke the scene causing a shift in the paradigm for HUS-associated STEC. In 2011, a STEC O104:H4 caused a large outbreak with more than 800 HUS and 50 deaths. Such a strain presented the adhesion determinants of Enteroaggregative E. coli (EAggEC). We investigated the possibility that, besides STEC and EAggEC, other pathogenic E. coli could be susceptible to infection with stx-phages. A panel of stx2-phages obtained from STEC isolated from human disease was used to infect experimentally E. coli strains representing all the known pathogenic types, including both diarrheagenic E. coli (DEC) and extra-intestinal pathogenic E. coli (ExPEC). We observed that all the E. coli pathogroups used in the infection experiments were susceptible to the infection. Our results suggest that the stx2-phages used may not have specificity for E. coli adapted to the intestinal environment, at least in the conditions used. Additionally, we could only observe transient lysogens suggesting that the event of stable stx2-phage acquisition occurs rarely. PMID:24999453

Current pathogenic Escherichia coli foodborne outbreak cases and therapy development.

PubMed

Yang, Shih-Chun; Lin, Chih-Hung; Aljuffali, Ibrahim A; Fang, Jia-You

2017-08-01

Food contamination by pathogenic microorganisms has been a serious public health problem and a cause of huge economic losses worldwide. Foodborne pathogenic Escherichia coli (E. coli) contamination, such as that with E. coli O157 and O104, is very common, even in developed countries. Bacterial contamination may occur during any of the steps in the farm-to-table continuum from environmental, animal, or human sources and cause foodborne illness. To understand the causes of the foodborne outbreaks by E. coli and food-contamination prevention measures, we collected and investigated the past 10 years' worldwide reports of foodborne E. coli contamination cases. In the first half of this review article, we introduce the infection and symptoms of five major foodborne diarrheagenic E. coli pathotypes: enteropathogenic E. coli (EPEC), Shiga toxin-producing E. coli/enterohemorrhagic E. coli (STEC/EHEC), Shigella/enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), and enterotoxigenic E. coli (ETEC). In the second half of this review article, we introduce the foodborne outbreak cases caused by E. coli in natural foods and food products. Finally, we discuss current developments that can be applied to control and prevent bacterial food contamination.

Recombinant expression of Streptococcus pneumoniae capsular polysaccharides in Escherichia coli

PubMed Central

Kay, Emily J.; Yates, Laura E.; Terra, Vanessa S.; Cuccui, Jon; Wren, Brendan W.

2016-01-01

Currently, Streptococcus pneumoniae is responsible for over 14 million cases of pneumonia worldwide annually, and over 1 million deaths, the majority of them children. The major determinant for pathogenesis is a polysaccharide capsule that is variable and is used to distinguish strains based on their serotype. The capsule forms the basis of the pneumococcal polysaccharide vaccine (PPV23) that contains purified capsular polysaccharide from 23 serotypes, and the pneumococcal conjugate vaccine (PCV13), containing 13 common serotypes conjugated to CRM197 (mutant diphtheria toxin). Purified capsule from S. pneumoniae is required for pneumococcal conjugate vaccine production, and costs can be prohibitively high, limiting accessibility of the vaccine in low-income countries. In this study, we demonstrate the recombinant expression of the capsule-encoding locus from four different serotypes of S. pneumoniae within Escherichia coli. Furthermore, we attempt to identify the minimum set of genes necessary to reliably and efficiently express these capsules heterologously. These E. coli strains could be used to produce a supply of S. pneumoniae serotype-specific capsules without the need to culture pathogenic bacteria. Additionally, these strains could be applied to synthetic glycobiological applications: recombinant vaccine production using E. coli outer membrane vesicles or coupling to proteins using protein glycan coupling technology. PMID:27110302

Enteric Pathogen Survival Varies Substantially in Irrigation Water from Belgian Lettuce Producers

PubMed Central

Van Der Linden, Inge; Cottyn, Bart; Uyttendaele, Mieke; Berkvens, Nick; Vlaemynck, Geertrui; Heyndrickx, Marc; Maes, Martine

2014-01-01

It is accepted that irrigation water is a potential carrier of enteric pathogens, such as Salmonella and E. coli O157:H7 and, therefore, a source for contamination of fresh produce. We tested this by comparing irrigation water samples taken from five different greenhouses in Belgium. The water samples were inoculated with four zoonotic strains, two Salmonella and two E. coli O157:H7 strains, and pathogen survival and growth in the water were monitored up till 14 days. The influence of water temperature and chemical water quality was evaluated, and the survival tests were also performed in water samples from which the resident aquatic microbiota had previously been eliminated by filter sterilization. The pathogen’s survival differed greatly in the different irrigation waters. Three water samples contained nutrients to support important growth of the pathogens, and another enabled weaker growth. However, for all, growth was only observed in the samples that did not contain the resident aquatic microbiota. In the original waters with their specific water biota, pathogen levels declined. The same survival tendencies existed in water of 4 °C and 20 °C, although always more expressed at 20 °C. Low water temperatures resulted in longer pathogen survival. Remarkably, the survival capacity of two E. coli 0157:H7 strains differed, while Salmonella Thompson and Salmonella Typhimurium behaved similarly. The pathogens were also transferred to detached lettuce leaves, while suspended in two of the water samples or in a buffer. The effect of the water sample on the pathogen’s fitness was also reproduced on the leaves when stored at 100% relative humidity. Inoculation of the suspension in buffer or in one of the water samples enabled epiphytic growth and survival, while the pathogen level in the other water sample decreased once loaded on the leaves. Our results show that irrigation waters from different origin may have a different capacity to transmit enteric pathogens and an important impact on the fitness of the pathogens to sustain and even grow on the leaf surface. PMID:25268508

Controlled progressive innate immune stimulation regimen prevents the induction of sickness behavior in the open field test

PubMed Central

Chen, Qun; Tarr, Andrew J; Liu, Xiaoyu; Wang, Yufen; Reed, Nathaniel S; DeMarsh, Cameron P; Sheridan, John F; Quan, Ning

2013-01-01

Peripheral immune activation by bacterial mimics or live replicating pathogens is well known to induce central nervous system activation. Sickness behavior alterations are often associated with inflammation-induced increases in peripheral proinflammatory cytokines (eg, interleukin [IL]-1β and IL-6). However, most researchers have used acute high dose endotoxin/bacterial challenges to observe these outcomes. Using this methodology may pose inherent risks in the translational interpretation of the experimental data in these studies. Studies using Escherichia coli have yet to establish the full kinetics of repeated E. coli peripheral injections. Therefore, we sought to examine the effects of repeated low dose E. coli on sickness behavior and local peripheral inflammation in the open field test. Results from the current experiments showed a behavioral dose response, where increased amounts of E. coli resulted in correspondingly increased sickness behavior. Furthermore, animals that received a subthreshold dose (ie, one that did not cause sickness behavior) of E. coli 24 hours prior were able to withstand a larger dose of E. coli on the second day (a dose that would normally cause sickness behavior in mice without prior exposure) without inducing sickness behavior. In addition, animals that received escalating subthreshold doses of E. coli on days 1 and 2 behaviorally tolerated a dose of E. coli 25 times higher than what would normally cause sickness behavior if given acutely. Lastly, increased levels of E. coli caused increased IL-6 and IL-1β protein expression in the peritoneal cavity, and this increase was blocked by administering a subthreshold dose of E. coli 24 hours prior. These data show that progressive challenges with subthreshold levels of E. coli may obviate the induction of sickness behavior and proinflammatory cytokine expression. PMID:23950656

Controlled progressive innate immune stimulation regimen prevents the induction of sickness behavior in the open field test.

PubMed

Chen, Qun; Tarr, Andrew J; Liu, Xiaoyu; Wang, Yufen; Reed, Nathaniel S; Demarsh, Cameron P; Sheridan, John F; Quan, Ning

2013-01-01

Peripheral immune activation by bacterial mimics or live replicating pathogens is well known to induce central nervous system activation. Sickness behavior alterations are often associated with inflammation-induced increases in peripheral proinflammatory cytokines (eg, interleukin [IL]-1β and IL-6). However, most researchers have used acute high dose endotoxin/bacterial challenges to observe these outcomes. Using this methodology may pose inherent risks in the translational interpretation of the experimental data in these studies. Studies using Escherichia coli have yet to establish the full kinetics of repeated E. coli peripheral injections. Therefore, we sought to examine the effects of repeated low dose E. coli on sickness behavior and local peripheral inflammation in the open field test. Results from the current experiments showed a behavioral dose response, where increased amounts of E. coli resulted in correspondingly increased sickness behavior. Furthermore, animals that received a subthreshold dose (ie, one that did not cause sickness behavior) of E. coli 24 hours prior were able to withstand a larger dose of E. coli on the second day (a dose that would normally cause sickness behavior in mice without prior exposure) without inducing sickness behavior. In addition, animals that received escalating subthreshold doses of E. coli on days 1 and 2 behaviorally tolerated a dose of E. coli 25 times higher than what would normally cause sickness behavior if given acutely. Lastly, increased levels of E. coli caused increased IL-6 and IL-1β protein expression in the peritoneal cavity, and this increase was blocked by administering a subthreshold dose of E. coli 24 hours prior. These data show that progressive challenges with subthreshold levels of E. coli may obviate the induction of sickness behavior and proinflammatory cytokine expression.

Medical Surveillance Monthly Report (MSMR). Volume 4, Number 6, September 1998

DTIC Science & Technology

1998-09-01

bacterial pathogens (from both cases and noncases) were Salmonella enteritidis , entero-adherent E. coli (EAEC), entero- toxigenic E. coli (ETEC), and...Isolation rate (% of cultures + for pathogen) No. of cultures + for pathogen Isolation rate (% of cultures + for pathogen) Salmonella enteritidis 32...infections ............. 8 ARD surveillance update .................................................. 13 Foodborne outbreak, Salmonella

Identification of Bacterial Factors Involved in Type 1 Fimbria Expression using an Escherichia coli K12 Proteome Chip*

PubMed Central

Chen, Yi-Wen; Teng, Ching-Hao; Ho, Yu-Hsuan; Jessica Ho, Tien Yu; Huang, Wen-Chun; Hashimoto, Masayuki; Chiang, I-Yuan; Chen, Chien-Sheng

2014-01-01

Type 1 fimbriae are filamentous structures on Escherichia coli. These structures are important adherence factors. Because binding to the host cells is the first step of infection, type 1 fimbria is an important virulence factor of pathogenic E. coli. Expression of type 1 fimbria is regulated by a phase variation in which each individual bacterium can alternate between fimbriated (phase-ON) and nonfimbriated (phase-OFF) states. The phase variation is regulated by the flipping of the 314-bp fimS fragment, which contains the promoter driving the expression of the genes required for the synthesis of type 1 fimbria. Thus, the bacterial proteins able to interact with fimS are likely to be involved in regulating the expression of type 1 fimbria. To identify novel type 1 fimbria-regulating factors, we used an E. coli K12 proteome chip to screen for the bacterial factors able to interact with a 602-bp DNA fragment containing fimS and its adjacent regions. The Spr protein was identified by the proteome chip-based screening and further confirmed to be able to interact with fimS by electrophoretic mobility shift assay. Deletion of spr in the neonatal meningitis E. coli strain RS218 significantly increased the ratio of the bacterial colonies that contained the type 1 fimbria phase-ON cells on agar plates. In addition, Spr interfered with the interactions of fimS with the site-specific recombinases, FimB and FimE, which are responsible for mediating the flipping of fimS. These results suggest that Spr is involved in the regulation of type 1 fimbria expression through direct interaction with the invertible element fimS. These findings facilitate our understanding of the regulation of type 1 fimbria. PMID:24692643

Crystallization and preliminary crystallographic analysis of recombinant immunoglobulin G-binding protein from Streptococcus suis

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

Khan, Abdul Hamid; Chu, Fuliang; Feng, Youjun

2008-08-01

Crystallization of recombinant IgG-binding protein expressed in Escherichia coli using the hanging-drop vapour-diffusion method is described. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 38.98, b = 43.94, c = 78.17 Å. Streptococcus suis, an important zoonotic pathogen, expresses immunoglobulin G-binding protein, which is thought to be helpful to the organism in eluding the host defence system. Recombinant IgG-binding protein expressed in Escherichia coli has been crystallized using the hanging-drop vapour-diffusion method. The crystals belonged to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 38.98, b = 43.94, c =more » 78.17 Å and one molecule in the asymmetric unit. Diffraction data were collected to 2.60 Å resolution.« less

Growth modeling of uropathogenic Escherichia coli in ground chicken meat

USDA-ARS?s Scientific Manuscript database

Extraintestinal Pathogenic Escherichia coli (ExPEC), including Uropathogenic E. coli (UPEC), are common contaminants in poultry meat, and are a major pathogen associated with inflammatory bowel disease, ulcerative colitis, sepsis, and urinary tract infections. The purpose of this study was to determ...

In silico prediction of escherichia coli proteins targeting the host cell nucleus, with special reference to their role in colon cancer etiology.

PubMed

Khan, Abdul Arif

2014-06-01

The potential role of Escherichia coli in the development of colorectal carcinoma (CRC) has been investigated in many studies. Although the exact mechanism is not clear, chronic inflammation caused by E. coli and other related events are suggested as possible causes behind E. coli-induced colon cancer. It has been found that CRC cells, but not normal cells, are colonized by an intracellular form of E. coli. We predicted nuclear targeting of bacterial proteins in the host cell through computational tools nuclear localization signal (NLS) mapper and balanced subcellular localization predictor (BaCeILo). During intracellular E. coli residence, such targeting is highly likely and may have a possible role in colon cancer etiology. We observed that several gene expression-associated proteins of E. coli can migrate to the host nucleus during intracellular infections. This situation provides an opportunity for competitive interaction of host and pathogen proteins with similar cellular substrates, thereby increasing the chances of development of colon cancer. Moreover, the results indicated that proteins localized in the membrane of E. coli mostly act as secretary proteins in host cells. No exact correlation was observed between NLS prediction and nuclear localization prediction by BaCeILo. This is partly because of a number of reasons, including that only 30% of nuclear proteins carry NLS and that proteins <40 kDa molecular weight can passively target the host nucleus. This study concludes that detection of gene expression-specific E. coli proteins and their targeting of the nucleus may have a profound impact on CRC etiology.

Recent Advances in Understanding Enteric Pathogenic Escherichia coli

PubMed Central

Croxen, Matthew A.; Law, Robyn J.; Scholz, Roland; Keeney, Kristie M.; Wlodarska, Marta

2013-01-01

SUMMARY Although Escherichia coli can be an innocuous resident of the gastrointestinal tract, it also has the pathogenic capacity to cause significant diarrheal and extraintestinal diseases. Pathogenic variants of E. coli (pathovars or pathotypes) cause much morbidity and mortality worldwide. Consequently, pathogenic E. coli is widely studied in humans, animals, food, and the environment. While there are many common features that these pathotypes employ to colonize the intestinal mucosa and cause disease, the course, onset, and complications vary significantly. Outbreaks are common in developed and developing countries, and they sometimes have fatal consequences. Many of these pathotypes are a major public health concern as they have low infectious doses and are transmitted through ubiquitous mediums, including food and water. The seriousness of pathogenic E. coli is exemplified by dedicated national and international surveillance programs that monitor and track outbreaks; unfortunately, this surveillance is often lacking in developing countries. While not all pathotypes carry the same public health profile, they all carry an enormous potential to cause disease and continue to present challenges to human health. This comprehensive review highlights recent advances in our understanding of the intestinal pathotypes of E. coli. PMID:24092857

Electrochemical recognition and quantification of cytochrome c expression in Bacillus subtilis and aerobe/anaerobe Escherichia coli using N,N,N′,N′-tetramethyl-para-phenylene-diamine (TMPD)† †Electronic supplementary information (ESI) available. See DOI: 10.1039/c7sc03498a

PubMed Central

Kuss, S.; Tanner, E. E. L.; Ordovas-Montanes, M.

2017-01-01

The colorimetric identification of pathogenic and non-pathogenic bacteria in cell culture is commonly performed using the redox mediator N,N,N′,N′-tetramethyl-para-phenylene-diamine (TMPD) in the so-called oxidase test, which indicates the presence of bacterial cytochrome c oxidases. The presented study demonstrates the ability of electrochemistry to employ TMPD to detect bacteria and quantify the activity of bacterial cytochrome c oxidases. Cyclic voltammetry studies and chronoamperometry measurements performed on the model organism Bacillus subtilis result in a turnover number, calculated for single bacteria. Furthermore, trace amounts of cytochrome c oxidases were revealed in aerobically cultured Escherichia coli, which to our knowledge no other technique is currently able to quantify in molecular biology. The reported technique could be applied to a variety of pathogenic bacteria and has the potential to be employed in future biosensing technology. PMID:29568431

Phage-protease-peptide: a novel trifecta enabling multiplex detection of viable bacterial pathogens.

PubMed

Alcaine, S D; Tilton, L; Serrano, M A C; Wang, M; Vachet, R W; Nugen, S R

2015-10-01

Bacteriophages represent rapid, readily targeted, and easily produced molecular probes for the detection of bacterial pathogens. Molecular biology techniques have allowed researchers to make significant advances in the bioengineering of bacteriophage to further improve speed and sensitivity of detection. Despite their host specificity, bacteriophages have not been meaningfully leveraged in multiplex detection of bacterial pathogens. We propose a proof-of-principal phage-based scheme to enable multiplex detection. Our scheme involves bioengineering bacteriophage to carry a gene for a specific protease, which is expressed during infection of the target cell. Upon lysis, the protease is released to cleave a reporter peptide, and the signal detected. Here we demonstrate the successful (i) modification of T7 bacteriophage to carry tobacco etch virus (TEV) protease; (ii) expression of TEV protease by Escherichia coli following infection by our modified T7, an average of 2000 units of protease per phage are produced during infection; and (iii) proof-of-principle detection of E. coli in 3 h after a primary enrichment via TEV protease activity using a fluorescent peptide and using a designed target peptide for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis (MALDI-TOF MS) analysis. This proof-of-principle can be translated to other phage-protease-peptide combinations to enable multiplex bacterial detection and readily adopted on multiple platforms, like MALDI-TOF MS or fluorescent readers, commonly found in labs.

Molecular Analysis of Escherichia coli from retail meats (2002-2004) from the United States National Antimicrobial Resistance Monitoring System.

PubMed

Johnson, James R; McCabe, James S; White, David G; Johnston, Brian; Kuskowski, Michael A; McDermott, Patrick

2009-07-15

The origins and virulence potential of meat product-associated Escherichia coli are undefined. Two hundred eighty-seven E. coli isolates (145 resistant and 142 susceptible to trimethoprim-sulfamethoxazole, nalidixic acid, and/or ceftiofur), recovered by the United States National Antimicrobial Monitoring System from retail beef, pork, chicken, and turkey products (from Oregon, Tennessee, Georgia, and Maryland, 2002-2004) underwent polymerase chain reaction testing for phylogenetic groupings and 59 virulence-associated genes. However analyzed, resistant and susceptible isolates differed minimally according to the assessed characteristics. In contrast, the 4 meat types differed greatly for multiple individual traits and aggregate virulence scores. Poultry isolates exhibited virulence genes associated with avian pathogenic E. coli; beef isolates exhibited traits associated with E. coli from diseased cattle. Overall, 20% of isolates qualified as extraintestinal pathogenic E. coli, with poultry isolates exhibiting significantly higher virulence scores than beef and pork isolates (P < .001). Within this systematically collected, geographically distributed sample of recent retail meat isolates, the carriage of extraintestinal pathogenic E. coli virulence genes in antimicrobial-resistant and antimicrobial-susceptible E. coli appeared similar, whereas isolates from different types of meat differed, consistent with on-farm acquisition of resistance within host species-specific E. coli populations. A substantial minority of meat-source E. coli (whether susceptible or resistant) may represent potential human pathogens.

Effect of nanoliposomes containing Zataria multiflora Boiss. essential oil on gene expression of Shiga toxin 2 in Escherichia coli O157:H7.

PubMed

Khatibi, S A; Misaghi, A; Moosavy, M H; Akhondzadeh Basti, A; Mohamadian, S; Khanjari, A

2018-02-01

Enterohaemorrhagic Escherichia coli serotype O157:H7 as a major human pathogen is responsible for food borne outbreaks, bloody diarrhoea, haemorrhagic colitis and haemolytic uraemic syndrome and even death. In this study, the antibacterial activity of the Zataria multiflora essential oil (ZMEO) and nanoliposome-encapsulated ZMEO was evaluated on the pathogenicity of E. coli O157:H7. The minimum inhibitory concentrations (MIC) of essential oil (EO) were determined against the bacterium before and after encapsulation into nanoliposome. Then, the effect of subinhibitory concentrations was evaluated on Shiga toxin 2 (Stx2) production. The effect of free and nanoliposomal EO was also studied on the gene expression of Stx2 by real-time PCR. It was found that inhibitory activity of EO was improved after incorporation into nanoliposomes (P < 0·05). The MIC of free EO against E. coli O157:H7 was 0·03% (v/v), while this value decreased to 0·015%, after encapsulation of EO into nanoliposomes. Furthermore, subinhibitory concentrations of liposomal EO (50 and 75% MIC) had significantly higher inhibitory effect on Stx2 titre than its free form (P < 0·05). Sub-MICs of nanoencapsulated EO also showed a better activity in reduction of Stx2A gene expression than free EO. Using 75% MIC of nanoliposomal EO, the relative transcriptional level of Stx2A gene was decreased from 0·721 to 0·646. The findings of present study suggest that application of nanoliposomes can improve the antibacterial effect of EOs like ZMEO. Due to the enhancement of antimicrobial activity, nanoencapsulation of plant EOs and extracts may increase their commercial application not only in food area but also in the pharmaceutics, cosmetics and health products. © 2017 The Society for Applied Microbiology.

CdiA Effectors from Uropathogenic Escherichia coli Use Heterotrimeric Osmoporins as Receptors to Recognize Target Bacteria

PubMed Central

Beck, Christina M.; Willett, Julia L. E.; Kim, Jeff J.; Low, David A.; Hayes, Christopher S.

2016-01-01

Many Gram-negative bacterial pathogens express contact-dependent growth inhibition (CDI) systems that promote cell-cell interaction. CDI+ bacteria express surface CdiA effector proteins, which transfer their C-terminal toxin domains into susceptible target cells upon binding to specific receptors. CDI+ cells also produce immunity proteins that neutralize the toxin domains delivered from neighboring siblings. Here, we show that CdiAEC536 from uropathogenic Escherichia coli 536 (EC536) uses OmpC and OmpF as receptors to recognize target bacteria. E. coli mutants lacking either ompF or ompC are resistant to CDIEC536-mediated growth inhibition, and both porins are required for target-cell adhesion to inhibitors that express CdiAEC536. Experiments with single-chain OmpF fusions indicate that the CdiAEC536 receptor is heterotrimeric OmpC-OmpF. Because the OmpC and OmpF porins are under selective pressure from bacteriophages and host immune systems, their surface-exposed loops vary between E. coli isolates. OmpC polymorphism has a significant impact on CDIEC536 mediated competition, with many E. coli isolates expressing alleles that are not recognized by CdiAEC536. Analyses of recombinant OmpC chimeras suggest that extracellular loops L4 and L5 are important recognition epitopes for CdiAEC536. Loops L4 and L5 also account for much of the sequence variability between E. coli OmpC proteins, raising the possibility that CDI contributes to the selective pressure driving OmpC diversification. We find that the most efficient CdiAEC536 receptors are encoded by isolates that carry the same cdi gene cluster as E. coli 536. Thus, it appears that CdiA effectors often bind preferentially to "self" receptors, thereby promoting interactions between sibling cells. As a consequence, these effector proteins cannot recognize nor suppress the growth of many potential competitors. These findings suggest that self-recognition and kin selection are important functions of CDI. PMID:27723824

An in silico model for identification of small RNAs in whole bacterial genomes: characterization of antisense RNAs in pathogenic Escherichia coli and Streptococcus agalactiae strains.

PubMed

Pichon, Christophe; du Merle, Laurence; Caliot, Marie Elise; Trieu-Cuot, Patrick; Le Bouguénec, Chantal

2012-04-01

Characterization of small non-coding ribonucleic acids (sRNA) among the large volume of data generated by high-throughput RNA-seq or tiling microarray analyses remains a challenge. Thus, there is still a need for accurate in silico prediction methods to identify sRNAs within a given bacterial species. After years of effort, dedicated software were developed based on comparative genomic analyses or mathematical/statistical models. Although these genomic analyses enabled sRNAs in intergenic regions to be efficiently identified, they all failed to predict antisense sRNA genes (asRNA), i.e. RNA genes located on the DNA strand complementary to that which encodes the protein. The statistical models enabled any genomic region to be analyzed theorically but not efficiently. We present a new model for in silico identification of sRNA and asRNA candidates within an entire bacterial genome. This model was successfully used to analyze the Gram-negative Escherichia coli and Gram-positive Streptococcus agalactiae. In both bacteria, numerous asRNAs are transcribed from the complementary strand of genes located in pathogenicity islands, strongly suggesting that these asRNAs are regulators of the virulence expression. In particular, we characterized an asRNA that acted as an enhancer-like regulator of the type 1 fimbriae production involved in the virulence of extra-intestinal pathogenic E. coli.

An in silico model for identification of small RNAs in whole bacterial genomes: characterization of antisense RNAs in pathogenic Escherichia coli and Streptococcus agalactiae strains

PubMed Central

Pichon, Christophe; du Merle, Laurence; Caliot, Marie Elise; Trieu-Cuot, Patrick; Le Bouguénec, Chantal

2012-01-01

Characterization of small non-coding ribonucleic acids (sRNA) among the large volume of data generated by high-throughput RNA-seq or tiling microarray analyses remains a challenge. Thus, there is still a need for accurate in silico prediction methods to identify sRNAs within a given bacterial species. After years of effort, dedicated software were developed based on comparative genomic analyses or mathematical/statistical models. Although these genomic analyses enabled sRNAs in intergenic regions to be efficiently identified, they all failed to predict antisense sRNA genes (asRNA), i.e. RNA genes located on the DNA strand complementary to that which encodes the protein. The statistical models enabled any genomic region to be analyzed theorically but not efficiently. We present a new model for in silico identification of sRNA and asRNA candidates within an entire bacterial genome. This model was successfully used to analyze the Gram-negative Escherichia coli and Gram-positive Streptococcus agalactiae. In both bacteria, numerous asRNAs are transcribed from the complementary strand of genes located in pathogenicity islands, strongly suggesting that these asRNAs are regulators of the virulence expression. In particular, we characterized an asRNA that acted as an enhancer-like regulator of the type 1 fimbriae production involved in the virulence of extra-intestinal pathogenic E. coli. PMID:22139924

Prevalence of Avian-Pathogenic Escherichia coli Strain O1 Genomic Islands among Extraintestinal and Commensal E. coli Isolates

PubMed Central

Johnson, Timothy J.; Wannemuehler, Yvonne; Kariyawasam, Subhashinie; Johnson, James R.; Logue, Catherine M.

2012-01-01

Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include pathogens of humans and animals. Previously, the genome of avian-pathogenic E. coli (APEC) O1:K1:H7 strain O1, from ST95, was sequenced and compared to those of several other E. coli strains, identifying 43 genomic islands. Here, the genomic islands of APEC O1 were compared to those of other sequenced E. coli strains, and the distribution of 81 genes belonging to 12 APEC O1 genomic islands among 828 human and avian ExPEC and commensal E. coli isolates was determined. Multiple islands were highly prevalent among isolates belonging to the O1 and O18 serogroups within phylogenetic group B2, which are implicated in human neonatal meningitis. Because of the extensive genomic similarities between APEC O1 and other human ExPEC strains belonging to the ST95 phylogenetic lineage, its ability to cause disease in a rat model of sepsis and meningitis was assessed. Unlike other ST95 lineage strains, APEC O1 was unable to cause bacteremia or meningitis in the neonatal rat model and was significantly less virulent than uropathogenic E. coli (UPEC) CFT073 in a mouse sepsis model, despite carrying multiple neonatal meningitis E. coli (NMEC) virulence factors and belonging to the ST95 phylogenetic lineage. These results suggest that host adaptation or genome modifications have occurred either in APEC O1 or in highly virulent ExPEC isolates, resulting in differences in pathogenicity. Overall, the genomic islands examined provide targets for further discrimination of the different ExPEC subpathotypes, serogroups, phylogenetic types, and sequence types. PMID:22467781

Prevalence of avian-pathogenic Escherichia coli strain O1 genomic islands among extraintestinal and commensal E. coli isolates.

PubMed

Johnson, Timothy J; Wannemuehler, Yvonne; Kariyawasam, Subhashinie; Johnson, James R; Logue, Catherine M; Nolan, Lisa K

2012-06-01

Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include pathogens of humans and animals. Previously, the genome of avian-pathogenic E. coli (APEC) O1:K1:H7 strain O1, from ST95, was sequenced and compared to those of several other E. coli strains, identifying 43 genomic islands. Here, the genomic islands of APEC O1 were compared to those of other sequenced E. coli strains, and the distribution of 81 genes belonging to 12 APEC O1 genomic islands among 828 human and avian ExPEC and commensal E. coli isolates was determined. Multiple islands were highly prevalent among isolates belonging to the O1 and O18 serogroups within phylogenetic group B2, which are implicated in human neonatal meningitis. Because of the extensive genomic similarities between APEC O1 and other human ExPEC strains belonging to the ST95 phylogenetic lineage, its ability to cause disease in a rat model of sepsis and meningitis was assessed. Unlike other ST95 lineage strains, APEC O1 was unable to cause bacteremia or meningitis in the neonatal rat model and was significantly less virulent than uropathogenic E. coli (UPEC) CFT073 in a mouse sepsis model, despite carrying multiple neonatal meningitis E. coli (NMEC) virulence factors and belonging to the ST95 phylogenetic lineage. These results suggest that host adaptation or genome modifications have occurred either in APEC O1 or in highly virulent ExPEC isolates, resulting in differences in pathogenicity. Overall, the genomic islands examined provide targets for further discrimination of the different ExPEC subpathotypes, serogroups, phylogenetic types, and sequence types.

Influence of surface polysaccharides of Escherichia coli O157:H7 on plant defense response and survival of the human enteric pathogen on Arabidopsis thaliana and lettuce (Lactuca sativa).

PubMed

Jang, Hyein; Matthews, Karl R

2018-04-01

This study aimed to determine the influence of bacterial surface polysaccharides (cellulose, colanic acid, and lipopolysaccharide; LPS) on the colonization or survival of Escherichia coli O157:H7 on plants and the plant defense response. Survival of E. coli O157:H7 were evaluated on Arabidopsis thaliana and romaine lettuce as a model plant and an edible crop (leafy vegetable), respectively. The population of the wild-type strain of E. coli O157:H7 on Arabidopsis plants and lettuce was significantly (P < 0.05) greater compared with the colanic acid-deficient and LPS-truncated mutants on day 1 and day 5 post-inoculation. This result indicates that colanic acid and LPS structures may contribute to the ability of bacterial survival or persistence on plants. The wild-type strain of E. coli O157:H7 produced approximately twice the amount (P < 0.05) of capsular polysaccharide (CPS) than the colanic acid and LPS-truncated mutants. The significantly lower production of CPS was associated with significantly greater (2-fold) expression of pathogenesis-related gene (PR1) compared with the wild-type and cellulose-deficient mutant (P < 0.05). Collectively, the results of this study may suggest that specific surface polysaccharides of E. coli O157:H7 differentially induce the plant defense response, consequently affecting the survival of the human pathogen on plants. The survival and persistence of E. coli O157:H7 was similar on Arabidopsis and lettuce regardless of day post-inoculation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Escherichia coli Contamination across Multiple Environmental Compartments (Soil, Hands, Drinking Water, and Handwashing Water) in Urban Harare: Correlations and Risk Factors.

PubMed

Navab-Daneshmand, Tala; Friedrich, Max N D; Gächter, Marja; Montealegre, Maria Camila; Mlambo, Linn S; Nhiwatiwa, Tamuka; Mosler, Hans-Joachim; Julian, Timothy R

2018-03-01

Escherichia coli pathotypes (i.e., enteropathogenic and enterotoxigenic) have been identified among the pathogens most responsible for moderate-to-severe diarrhea in low- and middle-income countries (LMICs). Pathogenic E. coli are transmitted from infected human or animal feces to new susceptible hosts via environmental reservoirs such as hands, water, and soil. Commensal E. coli , which includes nonpathogenic E. coli strains, are widely used as fecal bacteria indicator, with their presence associated with increased likelihood of enteric pathogens and/or diarrheal disease. In this study, we investigated E. coli contamination in environmental reservoirs within households ( N = 142) in high-population density communities of Harare, Zimbabwe. We further assessed the interconnectedness of the environmental compartments by investigating associations between, and household-level risk factors for, E. coli contamination. From the data we collected, the source and risk factors for E. coli contamination are not readily apparent. One notable exception is the presence of running tap water on the household plot, which is associated with significantly less E. coli contamination of drinking water, handwashing water, and hands after handwashing. In addition, E. coli levels on hands after washing are significantly associated with handwashing water contamination, hand contamination before washing, and diarrhea incidence. Finally, we observed that animal ownership increases E. coli contamination in soil, and E. coli in soil are correlated with contamination on hands before washing. This study highlights the complexity of E. coli contamination in household environments within LMICs. More, larger, studies are needed to better identify sources and exposure pathways of E. coli -and enteric pathogens generally-to identify effective interventions.

Outer membrane vesicles displaying engineered glycotopes elicit protective antibodies

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

Chen, Linxiao; Valentine, Jenny L.; Huang, Chung-Jr

The O-antigen polysaccharide (O-PS) component of lipopolysaccharides on the surface of gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies elicited by O-PS often confer protection against infection; therefore, O-PS glycoconjugate vaccines have proven useful against a number of different pathogenic bacteria. However, conventional methods for natural extraction or chemical synthesis of O-PS are technically demanding, inefficient, and expensive. In this paper, we describe an alternative methodology for producing glycoconjugate vaccines whereby recombinant O-PS biosynthesis is coordinated with vesiculation in laboratory strains of Escherichia coli to yield glycosylated outer membrane vesicles (glycOMVs) decorated with pathogen-mimetic glycotopes. Usingmore » this approach, glycOMVs corresponding to eight different pathogenic bacteria were generated. For example, expression of a 17-kb O-PS gene cluster from the highly virulent Francisella tularensis subsp. tularensis (type A) strain Schu S4 in hypervesiculating E. coli cells yielded glycOMVs that displayed F. tularensis O-PS. Immunization of BALB/c mice with glycOMVs elicited significant titers of O-PS–specific serum IgG antibodies as well as vaginal and bronchoalveolar IgA antibodies. Importantly, glycOMVs significantly prolonged survival upon subsequent challenge with F. tularensis Schu S4 and provided complete protection against challenge with two different F. tularensis subsp. holarctica (type B) live vaccine strains, thereby demonstrating the vaccine potential of glycOMVs. Finally, given the ease with which recombinant glycotopes can be expressed on OMVs, the strategy described here could be readily adapted for developing vaccines against many other bacterial pathogens.« less

Outer membrane vesicles displaying engineered glycotopes elicit protective antibodies

DOE PAGES

Chen, Linxiao; Valentine, Jenny L.; Huang, Chung-Jr; ...

2016-06-06

The O-antigen polysaccharide (O-PS) component of lipopolysaccharides on the surface of gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies elicited by O-PS often confer protection against infection; therefore, O-PS glycoconjugate vaccines have proven useful against a number of different pathogenic bacteria. However, conventional methods for natural extraction or chemical synthesis of O-PS are technically demanding, inefficient, and expensive. In this paper, we describe an alternative methodology for producing glycoconjugate vaccines whereby recombinant O-PS biosynthesis is coordinated with vesiculation in laboratory strains of Escherichia coli to yield glycosylated outer membrane vesicles (glycOMVs) decorated with pathogen-mimetic glycotopes. Usingmore » this approach, glycOMVs corresponding to eight different pathogenic bacteria were generated. For example, expression of a 17-kb O-PS gene cluster from the highly virulent Francisella tularensis subsp. tularensis (type A) strain Schu S4 in hypervesiculating E. coli cells yielded glycOMVs that displayed F. tularensis O-PS. Immunization of BALB/c mice with glycOMVs elicited significant titers of O-PS–specific serum IgG antibodies as well as vaginal and bronchoalveolar IgA antibodies. Importantly, glycOMVs significantly prolonged survival upon subsequent challenge with F. tularensis Schu S4 and provided complete protection against challenge with two different F. tularensis subsp. holarctica (type B) live vaccine strains, thereby demonstrating the vaccine potential of glycOMVs. Finally, given the ease with which recombinant glycotopes can be expressed on OMVs, the strategy described here could be readily adapted for developing vaccines against many other bacterial pathogens.« less

Comparative genomics and transcriptomics of Escherichia coli isolates carrying virulence factors of both enteropathogenic and enterotoxigenic E. coli.

PubMed

Hazen, Tracy H; Michalski, Jane; Luo, Qingwei; Shetty, Amol C; Daugherty, Sean C; Fleckenstein, James M; Rasko, David A

2017-06-14

Escherichia coli that are capable of causing human disease are often classified into pathogenic variants (pathovars) based on their virulence gene content. However, disease-associated hybrid E. coli, containing unique combinations of multiple canonical virulence factors have also been described. Such was the case of the E. coli O104:H4 outbreak in 2011, which caused significant morbidity and mortality. Among the pathovars of diarrheagenic E. coli that cause significant human disease are the enteropathogenic E. coli (EPEC) and enterotoxigenic E. coli (ETEC). In the current study we use comparative genomics, transcriptomics, and functional studies to characterize isolates that contain virulence factors of both EPEC and ETEC. Based on phylogenomic analysis, these hybrid isolates are more genomically-related to EPEC, but appear to have acquired ETEC virulence genes. Global transcriptional analysis using RNA sequencing, demonstrated that the EPEC and ETEC virulence genes of these hybrid isolates were differentially-expressed under virulence-inducing laboratory conditions, similar to reference isolates. Immunoblot assays further verified that the virulence gene products were produced and that the T3SS effector EspB of EPEC, and heat-labile toxin of ETEC were secreted. These findings document the existence and virulence potential of an E. coli pathovar hybrid that blurs the distinction between E. coli pathovars.

Genetic diversity and features analysis of type VI secretion systems loci in avian pathogenic Escherichia coli by wide genomic scanning.

PubMed

Ma, Jiale; Sun, Min; Bao, Yinli; Pan, Zihao; Zhang, Wei; Lu, Chengping; Yao, Huochun

2013-12-01

Avian pathogenic Escherichia coli (APEC) strains frequently cause extra-intestinal infections and significant economic losses. Recent studies revealed that the type VI secretion system (T6SS) is involved in APEC pathogenesis. Here we provide the first evidence of three distinguishable and conserved T6SS loci in APEC genomes. In addition, we present the prevalence and comparative genomic analysis of these three T6SS loci in 472 APEC isolates. The prevalence of T6SS1, T6SS2 and T6SS3 loci were 14.62% (69/472), 2.33% (11/472) and 0.85% (4/472) positive in the APEC collections, respectively, and revealed that >85% of the strains contained T6SS loci which consisted of the virulent phylogenetic groups D and B2. Comprehensive analysis showed prominent characteristics of T6SS1 locus, including wildly prevalence, rich sequence diversity, versatile VgrG islands and excellent expression competence in various E. coli pathotypes. Whereas the T6SS2 locus infatuated with ECOR groups B2 and sequence conservation, of which are only expressed in meningitis E. coli. Regrettably, the T6SS3 locus was encoded in negligible APEC isolates and lacked several key genes. An in-depth analysis about VgrG proteins indicated that their COG4253 and gp27 domain were involved in the transport of putative effector islands and recognition of host cells respectively, which revealed that VgrG proteins played an important role in functions formation of T6SS. Copyright © 2013 The Authors. Published by Elsevier B.V. All rights reserved.

Chromogenic agar medium for detection and isolation of Escherichia coli serogroups O26,O45,O103,O111,O121, and O145 from fresh beef and cattle feces

USDA-ARS?s Scientific Manuscript database

Non-O157 Shiga toxin–producing Escherichia coli (STEC) strains are clinically important foodborne pathogens. Unlike E. coli O157:H7, these foodborne pathogens have no unique biochemical characteristics to readily distinguish them from other E. coli strains growing on plating media. In this study, a ...

Influence of poultry litter and dairy manure on persistence of non-pathogenic E. coli and E. coli O157:H7 in two geographically distinct mid-Atlantic fields

USDA-ARS?s Scientific Manuscript database

A study was established to investigate how long non-pathogenic E. coli and E. coli O157:H7 persist in an agricultural environment after application of raw manure to fields intended to grow produce. The study is currently being conducted at two geographically distinct sites: Princess Anne, MD (Unive...

Colonization with Extraintestinal Pathogenic Escherichia coli among Nursing Home Residents and Its Relationship to Fluoroquinolone Resistance

PubMed Central

Maslow, Joel N.; Lautenbach, Ebbing; Glaze, Thomas; Bilker, Warren; Johnson, James R.

2004-01-01

In a cross-sectional fecal prevalence survey involving 49 residents of a Veterans Affairs nursing home, 59% of subjects were colonized with extraintestinal pathogenic Escherichia coli (ExPEC), 22% were colonized with adhesin-positive E. coli, and 51% were colonized with fluoroquinolone-resistant E. coli. Among 80 unique isolates, adhesins correlated negatively and aerobactin correlated positively with fluoroquinolone resistance. PMID:15328142

Multidrug-resistant pathogenic Escherichia coli isolated from wild birds in a veterinary hospital.

PubMed

Borges, C A; Beraldo, L G; Maluta, R P; Cardozo, M V; Barboza, K B; Guastalli, E A L; Kariyawasam, S; DebRoy, C; Ávila, F A

2017-02-01

Wild birds are carriers of Escherichia coli. However, little is known about their role as reservoirs for extra-intestinal pathogenic E. coli (ExPEC). In this work we investigated E. coli strains carrying virulence genes related to human and animal ExPEC isolated from free-living wild birds treated in a veterinary hospital. Multidrug resistance was found in 47.4% of the strains, but none of them were extended-spectrum beta-lactamase producers. Not only the virulence genes, but also the serogroups (e.g. O1 and O2) detected in the isolates of E. coli have already been implicated in human and bird diseases. The sequence types detected were also found in wild, companion and food animals, environmental and human clinical isolates in different countries. Furthermore, from the 19 isolates, 17 (89.5%) showed a degree of pathogenicity on an in vivo infection model. The isolates showed high heterogeneity by pulsed-field gel electrophoresis indicating that E. coli from these birds are clonally diverse. Overall, the results showed that wild birds can be reservoirs and/or vectors of highly pathogenic and multidrug-resistant E. coli that have the potential to cause disease in humans and poultry.

Heterologous Expression of Der Homologs in an Escherichia coli der Mutant and Their Functional Complementation

PubMed Central

Choi, Eunsil; Kang, Nalae; Jeon, Young; Pai, Hyun-Sook

2016-01-01

ABSTRACT The unique Escherichia coli GTPase Der (double Era-like GTPase), which contains tandemly repeated GTP-binding domains, has been shown to play an essential role in 50S ribosomal subunit biogenesis. The depletion of Der results in the accumulation of precursors of 50S ribosomal subunits that are structurally unstable at low Mg2+ concentrations. Der homologs are ubiquitously found in eubacteria. Conversely, very few are conserved in eukaryotes, and none is conserved in archaea. In the present study, to verify their conserved role in bacterial 50S ribosomal subunit biogenesis, we cloned Der homologs from two gammaproteobacteria, Klebsiella pneumoniae and Salmonella enterica serovar Typhimurium; two pathogenic bacteria, Staphylococcus aureus and Neisseria gonorrhoeae; and the extremophile Deinococcus radiodurans and then evaluated whether they could functionally complement the E. coli der-null phenotype. Only K. pneumoniae and S. Typhimurium Der proteins enabled the E. coli der-null strain to grow under nonpermissive conditions. Sucrose density gradient experiments revealed that the expression of K. pneumoniae and S. Typhimurium Der proteins rescued the structural instability of 50S ribosomal subunits, which was caused by E. coli Der depletion. To determine what allows their complementation, we constructed Der chimeras. We found that only Der chimeras harboring both the linker and long C-terminal regions could reverse the growth defects of the der-null strain. Our findings suggest that ubiquitously conserved essential GTPase Der is involved in 50S ribosomal subunit biosynthesis in various bacteria and that the linker and C-terminal regions may participate in species-specific recognition or interaction with the 50S ribosomal subunit. IMPORTANCE In Escherichia coli, Der (double Era-like GTPase) is an essential GTPase that is important for the production of mature 50S ribosomal subunits. However, to date, its precise role in ribosome biogenesis has not been clarified. In this study, we used five Der homologs from gammaproteobacteria, pathogenic bacteria, and an extremophile to elucidate their conserved function in 50S ribosomal subunit biogenesis. Among them, Klebsiella pneumoniae and Salmonella enterica serovar Typhimurium Der homologs implicated the participation of Der in ribosome assembly in E. coli. Our results show that the linker and C-terminal regions of Der homologs are correlated with its functional complementation in E. coli der mutants, suggesting that they are involved in species-specific recognition or interaction with 50S ribosomal subunits. PMID:27297882

Rapid detection of E. coli on goat meat by electronic nose

USDA-ARS?s Scientific Manuscript database

Much attention has been paid on the foodborne illness of food, which is easily contaminated with bacterial or pathogens. Escherichia coli (E. coli) is one of these bacterial that commonly live in the contaminated animal meat. There is a growing need in the food industry for pathogen detection syst...

Draft genomic sequencing of six potential extraintestinal pathogenic Escherichia coli isolates from retail chicken meat.

USDA-ARS?s Scientific Manuscript database

Potential Extraintestinal pathogenic Escherichia coli isolates DP254, WH333, WH398, F356, FEX675 and FEX725 were isolated from retail chicken meat products. Here, we report the draft genome sequences for these six E. coli isolates, which are currently being used in food safety research....

Escherichia coli Contamination across Multiple Environmental Compartments (Soil, Hands, Drinking Water, and Handwashing Water) in Urban Harare: Correlations and Risk Factors

PubMed Central

Navab-Daneshmand, Tala; Friedrich, Max N. D.; Gächter, Marja; Montealegre, Maria Camila; Mlambo, Linn S.; Nhiwatiwa, Tamuka; Mosler, Hans-Joachim; Julian, Timothy R.

2018-01-01

Abstract. Escherichia coli pathotypes (i.e., enteropathogenic and enterotoxigenic) have been identified among the pathogens most responsible for moderate-to-severe diarrhea in low- and middle-income countries (LMICs). Pathogenic E. coli are transmitted from infected human or animal feces to new susceptible hosts via environmental reservoirs such as hands, water, and soil. Commensal E. coli, which includes nonpathogenic E. coli strains, are widely used as fecal bacteria indicator, with their presence associated with increased likelihood of enteric pathogens and/or diarrheal disease. In this study, we investigated E. coli contamination in environmental reservoirs within households (N = 142) in high-population density communities of Harare, Zimbabwe. We further assessed the interconnectedness of the environmental compartments by investigating associations between, and household-level risk factors for, E. coli contamination. From the data we collected, the source and risk factors for E. coli contamination are not readily apparent. One notable exception is the presence of running tap water on the household plot, which is associated with significantly less E. coli contamination of drinking water, handwashing water, and hands after handwashing. In addition, E. coli levels on hands after washing are significantly associated with handwashing water contamination, hand contamination before washing, and diarrhea incidence. Finally, we observed that animal ownership increases E. coli contamination in soil, and E. coli in soil are correlated with contamination on hands before washing. This study highlights the complexity of E. coli contamination in household environments within LMICs. More, larger, studies are needed to better identify sources and exposure pathways of E. coli—and enteric pathogens generally—to identify effective interventions. PMID:29363444

“Black holes” and bacterial pathogenicity: A large genomic deletion that enhances the virulence of Shigella spp. and enteroinvasive Escherichia coli

PubMed Central

Maurelli, Anthony T.; Fernández, Reinaldo E.; Bloch, Craig A.; Rode, Christopher K.; Fasano, Alessio

1998-01-01

Plasmids, bacteriophages, and pathogenicity islands are genomic additions that contribute to the evolution of bacterial pathogens. For example, Shigella spp., the causative agents of bacillary dysentery, differ from the closely related commensal Escherichia coli in the presence of a plasmid in Shigella that encodes virulence functions. However, pathogenic bacteria also may lack properties that are characteristic of nonpathogens. Lysine decarboxylase (LDC) activity is present in ≈90% of E. coli strains but is uniformly absent in Shigella strains. When the gene for LDC, cadA, was introduced into Shigella flexneri 2a, virulence became attenuated, and enterotoxin activity was inhibited greatly. The enterotoxin inhibitor was identified as cadaverine, a product of the reaction catalyzed by LDC. Comparison of the S. flexneri 2a and laboratory E. coli K-12 genomes in the region of cadA revealed a large deletion in Shigella. Representative strains of Shigella spp. and enteroinvasive E. coli displayed similar deletions of cadA. Our results suggest that, as Shigella spp. evolved from E. coli to become pathogens, they not only acquired virulence genes on a plasmid but also shed genes via deletions. The formation of these “black holes,” deletions of genes that are detrimental to a pathogenic lifestyle, provides an evolutionary pathway that enables a pathogen to enhance virulence. Furthermore, the demonstration that cadaverine can inhibit enterotoxin activity may lead to more general models about toxin activity or entry into cells and suggests an avenue for antitoxin therapy. Thus, understanding the role of black holes in pathogen evolution may yield clues to new treatments of infectious diseases. PMID:9520472

Occurrence of waterborne pathogens and Escherichia coli at offshore drinking water intakes in lake Ontario.

PubMed

Edge, T A; Khan, I U H; Bouchard, R; Guo, J; Hill, S; Locas, A; Moore, L; Neumann, N; Nowak, E; Payment, P; Yang, R; Yerubandi, R; Watson, S

2013-10-01

The occurrence of waterborne pathogens was investigated at three drinking water intakes located about 2 km offshore in Lake Ontario. Water sampling was conducted over 3 years for Campylobacter spp., Cryptosporidium spp., Giardia spp., cultivable enteric viruses, and water quality parameters. All pathogens were detected in the offshore source water for each water treatment plant (WTP1 to WTP3), although at relatively low frequencies and concentrations. Giardia was the most common pathogen, occurring in 36% of water samples from the influent of WTP1 (n = 46), and with a maximum concentration of 0.70 cysts/liter in this influent. Cryptosporidium occurred as frequently as 15% in the WTP2 influent (n = 35), with a maximum concentration of 0.40 oocysts/liter in the WTP1 influent. The human Bacteroidales HF183 DNA marker was most common in the WTP1 influent (19%), and this was the only WTP where the Cryptosporidium hominis genotype was detected. No water quality parameter was predictive of pathogen occurrence across all three WTP influents. Escherichia coli was often below detection when pathogens were detected, and spikes in E. coli concentrations often did not coincide with pathogen occurrence. After summer rain events, river plumes had E. coli concentrations as high as 222 CFU/100 ml in surface waters 2 km offshore, without impacting drinking water intakes below the thermocline on the lake bottom. At times, prechlorination to control mussels at offshore intake cribs compromised the use of E. coli for "raw" water quality assessment, particularly for chlorine-resistant Cryptosporidium. E. coli measured by standard methods did not reliably predict pathogen occurrence at drinking water intakes in offshore ecosystems.

Occurrence of Waterborne Pathogens and Escherichia coli at Offshore Drinking Water Intakes in Lake Ontario

PubMed Central

Khan, I. U. H.; Bouchard, R.; Guo, J.; Hill, S.; Locas, A.; Moore, L.; Neumann, N.; Nowak, E.; Payment, P.; Yang, R.; Yerubandi, R.; Watson, S.

2013-01-01

The occurrence of waterborne pathogens was investigated at three drinking water intakes located about 2 km offshore in Lake Ontario. Water sampling was conducted over 3 years for Campylobacter spp., Cryptosporidium spp., Giardia spp., cultivable enteric viruses, and water quality parameters. All pathogens were detected in the offshore source water for each water treatment plant (WTP1 to WTP3), although at relatively low frequencies and concentrations. Giardia was the most common pathogen, occurring in 36% of water samples from the influent of WTP1 (n = 46), and with a maximum concentration of 0.70 cysts/liter in this influent. Cryptosporidium occurred as frequently as 15% in the WTP2 influent (n = 35), with a maximum concentration of 0.40 oocysts/liter in the WTP1 influent. The human Bacteroidales HF183 DNA marker was most common in the WTP1 influent (19%), and this was the only WTP where the Cryptosporidium hominis genotype was detected. No water quality parameter was predictive of pathogen occurrence across all three WTP influents. Escherichia coli was often below detection when pathogens were detected, and spikes in E. coli concentrations often did not coincide with pathogen occurrence. After summer rain events, river plumes had E. coli concentrations as high as 222 CFU/100 ml in surface waters 2 km offshore, without impacting drinking water intakes below the thermocline on the lake bottom. At times, prechlorination to control mussels at offshore intake cribs compromised the use of E. coli for “raw” water quality assessment, particularly for chlorine-resistant Cryptosporidium. E. coli measured by standard methods did not reliably predict pathogen occurrence at drinking water intakes in offshore ecosystems. PMID:23835181

``Black Holes" and Bacterial Pathogenicity: A Large Genomic Deletion that Enhances the Virulence of Shigella spp. and Enteroinvasive Escherichia coli

NASA Astrophysics Data System (ADS)

Maurelli, Anthony T.; Fernandez, Reinaldo E.; Bloch, Craig A.; Rode, Christopher K.; Fasano, Alessio

1998-03-01

Plasmids, bacteriophages, and pathogenicity islands are genomic additions that contribute to the evolution of bacterial pathogens. For example, Shigella spp., the causative agents of bacillary dysentery, differ from the closely related commensal Escherichia coli in the presence of a plasmid in Shigella that encodes virulence functions. However, pathogenic bacteria also may lack properties that are characteristic of nonpathogens. Lysine decarboxylate (LDC) activity is present in ≈ 90% of E. coli strains but is uniformly absent in Shigella strains. When the gene for LDC, cadA, was introduced into Shigella flexneri 2a, virulence became attenuated, and enterotoxin activity was inhibited greatly. The enterotoxin inhibitor was identified as cadaverine, a product of the reaction catalyzed by LDC. Comparison of the S. flexneri 2a and laboratory E. coli K-12 genomes in the region of cadA revealed a large deletion in Shigella. Representative strains of Shigella spp. and enteroinvasive E. coli displayed similar deletions of cadA. Our results suggest that, as Shigella spp. evolved from E. coli to become pathogens, they not only acquired virulence genes on a plasmid but also shed genes via deletions. The formation of these ``black holes,'' deletions of genes that are detrimental to a pathogenic lifestyle, provides an evolutionary pathway that enables a pathogen to enhance virulence. Furthermore, the demonstration that cadaverine can inhibit enterotoxin activity may lead to more general models about toxin activity or entry into cells and suggests an avenue for antitoxin therapy. Thus, understanding the role of black holes in pathogen evolution may yield clues to new treatments of infectious diseases.

Consequences of reductive evolution for gene expression in an obligate endosymbiont.

PubMed

Wilcox, Jennifer L; Dunbar, Helen E; Wolfinger, Russell D; Moran, Nancy A

2003-06-01

The smallest cellular genomes are found in obligate symbiotic and pathogenic bacteria living within eukaryotic hosts. In comparison with large genomes of free-living relatives, these reduced genomes are rearranged and have lost most regulatory elements. To test whether reduced bacterial genomes incur reduced regulatory capacities, we used full-genome microarrays to evaluate transcriptional response to environmental stress in Buchnera aphidicola, the obligate endosymbiont of aphids. The 580 genes of the B. aphidicola genome represent a subset of the 4500 genes known from the related organism, Escherichia coli. Although over 20 orthologues of E. coli heat stress (HS) genes are retained by B. aphidicola, only five were differentially expressed after near-lethal heat stress treatments, and only modest shifts were observed. Analyses of upstream regulatory regions revealed loss or degradation of most HS (sigma32) promoters. Genomic rearrangements downstream of an intact HS promoter yielded upregulation of a functionally unrelated and an inactivated gene. Reanalyses of comparable experimental array data for E. coli and Bacillus subtilis revealed that genome-wide differential expression was significantly lower in B. aphidicola. Our demonstration of a diminished stress response validates reports of temperature sensitivity in B. aphidicola and suggests that this reduced bacterial genome exhibits transcriptional inflexibility.

Apa is a trimeric autotransporter adhesin of Actinobacillus pleuropneumoniae responsible for autoagglutination and host cell adherence.

PubMed

Xiao, Longwen; Zhou, Liang; Sun, Changjiang; Feng, Xin; Du, ChongTao; Gao, Yu; Ji, Qun; Yang, Shuxin; Wang, Yu; Han, Wenyu; Langford, P R; Lei, Liancheng

2012-10-01

Actinobacillus pleuropneumoniae is the causative agent of porcine pleuropneumonia, and adherence to host cells is a key step in the pathogenic process. Although trimeric autotransporter adhesins (TAAs) were identified in many pathogenic bacteria in recent years, none in A. pleuropneumoniae have been characterized. In this study, we identified a TAA from A. pleuropneumoniae, Apa, and characterized the contribution of its amino acid residues to the adhesion process. Sequence analysis of the C-terminal amino acid residues of Apa revealed the presence of a putative translocator domain and six conserved HsfBD1-like or HsfBD2-like binding domains. Western blot analysis revealed that the 126 C-terminal amino acids of Apa could form trimeric molecules. By confocal laser scanning microscopy, one of these six domains (ApaBD3) was determined to mediate adherence to epithelial cells. Adherence assays and adherence inhibition assays using a recombinant E. coli- ApaBD3 strain which expressed ApaBD3 on the surface of E. coli confirmed that this domain was responsible for the adhesion activity. Moreover, cellular enzyme-linked immunosorbent assays demonstrated that ApaBD3 mediated high-level adherence to epithelial cell lines. Intriguingly, autoagglutination was observed with the E. coli- ApaBD3 strain, and this phenomenon was dependent upon the association of the expressed ApaBD3 with the C-terminal translocator domain. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

E. coli O124 K72 alters the intestinal barrier and the tight junctions proteins of guinea pig intestine.

PubMed

Ren, Xiaomeng; Zhu, Yanyan; Gamallat, Yaser; Ma, Shenhao; Chiwala, Gift; Meyiah, Abdo; Xin, Yi

2017-10-01

Our research group previously isolated and identified a strain of pathogenic Escherichia coli from clinical samples called E. coli O124 K72. The present study was aimed at determining the potential effects of E. coli O124 K72 on intestinal barrier functions and structural proteins integrity in guinea pig. Guinea pigs were grouped into three groups; control (CG); E. coli O124 K72 (E. coli); and probiotics Lactobacillus rhamnosus (LGG). Initially, we create intestinal dysbiosis by giving all animals Levofloxacin for 10days, but the control group (CG) received the same volume of saline. Then, the animals received either E. coli O124 K72 (E. coli) or Lactobacillus rhamnosus (LGG) according to their assigned group. E. coli O124 K72 treatment significantly affected colon morphology and distorted intestinal barrier function by up-regulating Claudin2 and down-regulating Occludin. In addition, E. coli upregulated the mRNA expression of MUC1, MUC2, MUC13 and MUC15. Furthermore, suspected tumor was found in the E. coli treated animals. Our results suggested that E. coli O124 K72 strain has adverse effects on intestinal barrier functions and is capable of altering integrity of structural proteins in guinea pig model while at same time it may have a role in colon carcinogenesis. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

The phagocytic capacity and immunological potency of human dendritic cells is improved by α2,6‐sialic acid deficiency

PubMed Central

Cabral, M. Guadalupe; Silva, Zélia; Ligeiro, Dário; Seixas, Elsa; Crespo, Hélio; Carrascal, Mylène A.; Silva, Mariana; Piteira, Ana R.; Paixão, Paulo; Lau, Joseph T.; Videira, Paula A.

2013-01-01

Summary Dendritic cells (DCs) play an essential role in immunity against bacteria by phagocytosis and by eliciting adaptive immune responses. Previously, we demonstrated that human monocyte‐derived DCs (MDDCs) express a high content of cell surface α2,6‐sialylated glycans. However, the relative role of these sialylated structures in phagocytosis of bacteria has not been reported. Here, we show that treatment with a sialidase significantly improved the capacity of both immature and mature MDDCs to phagocytose Escherichia coli. Desialylated MDDCs had a significantly more mature phenotype, with higher expression of MHC molecules and interleukin (IL)‐12, tumour necrosis factor‐α, IL‐6 and IL‐10 cytokines, and nuclear factor‐κB activation. T lymphocytes primed by desialylated MDDCs expressed more interferon‐γ when compared with priming by sialylated MDDCs. Improved phagocytosis required E. coli sialic acids, indicating a mechanism of host–pathogen interaction dependent on sialic acid moieties. The DCs harvested from mice deficient in the ST6Gal.1 sialyltransferase showed improved phagocytosis capacity, demonstrating that the observed sialidase effect was a result of the removal of α2,6‐sialic acid. The phagocytosis of different pathogenic E. coli isolates was also enhanced by sialidase, which suggests that modifications on MDDC sialic acids may be considered in the development of MDDC‐based antibacterial therapies. Physiologically, our findings shed new light on mechanisms that modulate the function of both immature and mature MDDCs, in the context of host–bacteria interaction. Hence, with particular relevance to DC‐based therapies, the engineering of α2,6‐sialic acid cell surface is a novel possibility to fine tune DC phagocytosis and immunological potency. PMID:23113614

Surface display of Salmonella epitopes in Escherichia coli and Staphylococcus carnosus.

PubMed

Nhan, Nguyen Thanh; Gonzalez de Valdivia, Ernesto; Gustavsson, Martin; Hai, Truong Nam; Larsson, Gen

2011-04-11

Salmonella enterica serotype Enteritidis (SE) is considered to be one of the most potent pathogenic Salmonella serotypes causing food-borne disease in humans. Since a live bacterial vaccine based on surface display of antigens has many advantages over traditional vaccines, we have studied the surface display of the SE antigenic proteins, H:gm and SefA in Escherichia coli by the β-autotransporter system, AIDA. This procedure was compared to protein translocation in Staphylococcus carnosus, using a staphylococci hybrid vector earlier developed for surface display of other vaccine epitopes. Both SefA and H:gm were translocated to the outer membrane in Escherichia coli. SefA was expressed to full length but H:gm was shorter than expected, probably due to a proteolytic cleavage of the N-terminal during passage either through the periplasm or over the membrane. FACS analysis confirmed that SefA was facing the extracellular environment, but this could not be conclusively established for H:gm since the N-terminal detection tag (His6) was cleaved off. Polyclonal salmonella antibodies confirmed the sustained antibody-antigen binding towards both proteins. The surface expression data from Staphylococcus carnosus suggested that the H:gm and SefA proteins were transported to the cell wall since the detection marker was displayed by FACS analysis. Apart from the accumulated knowledge and the existence of a wealth of equipment and techniques, the results indicate the selection of E. coli for further studies for surface expression of salmonella antigens. Surface expression of the full length protein facing the cell environment was positively proven by standard analysis, and the FACS signal comparison to expression in Staphylococcus carnosus shows that the distribution of the surface protein on each cell was comparatively very narrow in E. coli, the E. coli outer membrane molecules can serve as an adjuvant for the surface antigenic proteins and multimeric forms of the SefA protein were detected which would probably be positive for the realisation of a strong antigenic property. The detection of specific and similar proteolytic cleavage patterns for both the proteins provides a further starting point for the investigation and development of the Escherichia coli AIDA autotransporter efficiency.

Meat Science and Muscle Biology Symposium: Escherichia coli O157:H7, diet, and fecal microbiome in beef cattle.

PubMed

Wells, J E; Kim, M; Bono, J L; Kuehn, L A; Benson, A K

2014-04-01

Shiga-toxigenic Escherichia coli, such as E. coli O157:H7, are foodborne zoonotic pathogens that can cause severe illness and death in humans. The gastrointestinal tract of ruminant animals has been identified as a primary habitat for E. coli O157:H7 and, in cattle, the hindgut tract appears to be a primary site for colonization. This pathogen has been found in cattle feces, on cattle hides, and in the production environment, and transmission to humans has occurred as a result of consumption of contaminated ground beef, water, and produce. Interventions to reduce the pathogen at beef harvest have significantly reduced the occurrence of the pathogen, but outbreaks and recalls due to the pathogen still occur for beef products. Interventions in the feedyard before harvest have had little success, but critical control points for implementing interventions are limited compared with the beef abattoir. The percentage of animals shedding E. coli O157:H7 in the feces can be highly variable from pen to pen, and the levels in the feces can vary from animal to animal. Animals colonized and shedding E. coli O157:H7 at high levels are a small fraction of animals in a pen but are important source for transferring the pathogen amongst the penmates. Recent research has indicated that diet may greatly influence the shedding of E. coli O157:H7. In addition, diet can influence the microbiota composition of the feces. However, little is known about the interaction between the indigenous microbiota and fecal shedding of E. coli O157:H7. Understanding the influence of indigenous microbiota on the colonization and shedding of E. coli O157:H7 will provide a potential avenue for intervention in the preharvest production environment not yet exploited.

Use of a novel antigen expressing system to study the Salmonella enterica serovar Typhi protein recognition by T cells

PubMed Central

Lou, David; Steiner, Stephanie; Rezwanul, Tasmia; Guo, Qin; Picking, William D.; Nene, Vishvanath; Sztein, Marcelo B.

2017-01-01

Salmonella enterica serovar Typhi (S. Typhi), the causative agent of the typhoid fever, is a pathogen of great public health importance. Typhoid vaccines have the potential to be cost-effective measures towards combating this disease, yet the antigens triggering host protective immune responses are largely unknown. Given the key role of cellular-mediated immunity in S. Typhi protection, it is crucial to identify S. Typhi proteins involved in T-cell responses. Here, cells from individuals immunized with Ty21a typhoid vaccine were collected before and after immunization and used as effectors. We also used an innovative antigen expressing system based on the infection of B-cells with recombinant Escherichia coli (E. coli) expressing one of four S. Typhi gene products (i.e., SifA, OmpC, FliC, GroEL) as targets. Using flow cytometry, we found that the pattern of response to specific S. Typhi proteins was variable. Some individuals responded to all four proteins while others responded to only one or two proteins. We next evaluated whether T-cells responding to recombinant E. coli also possess the ability to respond to purified proteins. We observed that CD4+ cell responses, but not CD8+ cell responses, to recombinant E. coli were significantly associated with the responses to purified proteins. Thus, our results demonstrate the feasibility of using an E. coli expressing system to uncover the antigen specificity of T-cells and highlight its applicability to vaccine studies. These results also emphasize the importance of selecting the stimuli appropriately when evaluating CD4+ and CD8+ cell responses. PMID:28873442

[Cloning of Clostridium perfringens alpha-toxin gene and extracellular expression in Escherichia coli].

PubMed

Inoue, Masaharu; Kikuchi, Maho; Komoriya, Tomoe; Watanabe, Kunitomo; Kouno, Hideki

2007-01-01

Clostridium perfringens (C. perfringens) is a Gram-positive bacterial pathogen that widely propagets in the soil and the gastrointestinal tract of human and animals. This bacteria causes food poisoning, gas gangrene and other various range of infectious diseases. But there is no standard diagnosis method of C. perfringens. In order to develop a new type of immunoassay for clinical purpose, we studied expression and extracellular secretion of recombinant alpha-toxin having enzyme activity in E. coli expression system. Cloning was carried out after PCR amplification from C. perfringens GAI 94074 which was clinical isolate. Three kinds of fragment were cloned using pET100/D-TOPO vector. These fragments coded for ribosome binding site, signal peptide, and alpha-toxin gene respectively. Recombinant pET100 plasmid transformed into TOP 10 cells and the obtained plasmids were transformed into BL21 (DE3) cells. Then, the transformants were induced expression with IPTG. In conclusion, we successfully cloned, expressed and exteracellular secreted C. perfringens alpha-toxin containing signal peptide. Biologically, the obtained recombinant protein was positive for phospholipase C activity.

Molecular cloning, sequencing, and expression of Eimeria tenella HSP70 partial gene.

PubMed

Bogado, A L G; Martins, G F; Sasse, J P; Guimarães, J da S; Garcia, J L

2017-03-15

Members of the Eimeria genus are protozoan parasites of the subphylum Apicomplexa (Eimeriidae family), and belong to the coccidia group. Eimeria tenella is one of the most pathogenic species owing to its ability to penetrate the mucosa, and cause inflammation and damage. It is an obligate intracellular parasite that causes disease by destroying the host cells during multiplication. Heat shock protein 70 (HSP70) is a molecular chaperone that prevents cellular stress. The objective of this study was to clone, sequence, and express E. tenella HSP70 protein. After selecting the region of highest hydrophilicity in the hsp70 gene, we cloned complementary DNA (cDNA) into a pTrcHis2-TOPO vector and transformed it into TOP10 Escherichia coli cells; after induction, the bacteria expressed a 23-kDa protein with insoluble expression levels of approximately 5 mg/L. In summary, the partial hsp70 gene was successfully expressed in E. coli, producing a 23-kDa protein under insoluble conditions, and the antigen characteristics predicted by hydrophilicity analysis suggest the development of a vaccine for use in avian coccidiosis.

Extra-corporeal membrane oxygenation-associated infections: implication of extra-intestinal pathogenic Escherichia coli clones.

PubMed

Messika, Jonathan; Clermont, Olivier; Landraud, Luce; Schmidt, Matthieu; Aubry, Alexandra; Sougakoff, Wladimir; Fernandes, Romain; Combes, Alain; Denamur, Erick; Ricard, Jean-Damien

2017-08-01

Extra-corporeal membrane oxygenation (ECMO) is a promising life-saving technique for critically ill patients. Bacterial infection is a frequent complication, and Escherichia coli the predominant causative pathogen, but little is known about the characteristics of E. coli strains in these infections. We therefore conducted a retrospective study of 33 E. coli strains responsible for 33 ECMO-related infections, in 30 subjects. Antimicrobial susceptibility, phylotyping, O-typing, clonal relatedness determination and the screening for four virulence factor genes were conducted. Polymicrobial infections were evidenced in 61.6 % of episodes, irrespective of E. coli characteristics. Extra-intestinal pathogenic strains represented the large majority (69.7 %) of all E. coli isolates. Their advantageous genetic background may explain their predominance in this context. The potential for targeted digestive decontamination should be investigated in these patients for whom infectious complications are a heavy burden.

Correlation between E. coli levels and the presence of foodborne pathogens in surface irrigation water: Establishment of a sampling program.

PubMed

Truchado, Pilar; Hernandez, Natalia; Gil, Maria I; Ivanek, Renata; Allende, Ana

2018-01-01

To establish the association between microbial indicators and the presence of foodborne pathogens in irrigation water, Escherichia coli was enumerated using two quantification methods (plate counts and PMA-qPCR) and presence/absence of pathogenic microorganisms, including five strains from the Shiga toxigenic E. coli (O157:H7, O26, O103, O111 and O145) and Salmonella spp. were evaluated. The results confirmed that surface water can be considered a microbial hazard when used for irrigation. The levels of viable E. coli were very similar to those of cultivable E. coli, except for irrigation water obtained from water reservoirs. Comparison between the E. coli counts in samples positive and negative for the presence of pathogenic bacteria for the evaluated water sources identified E. coli level of 2.35 log cfu/100 mL as a cut-off able to correctly predict positive and negative samples with 93% sensitivity and 66% specificity, respectively. Thus, for the samples with levels of E. coli under 2.35 log cfu/100 mL (e.g., 2.24 log cfu/100 mL) there was a 90% probability that the samples were not contaminated with pathogenic microorganism in locations with similar prevalence. E. coli levels in irrigation water were affected by the ambient temperature confirming that water source and climate conditions should be taken into account by growers when designing a sampling program and the frequency of the monitoring to make a better and more efficient use of their resources. Copyright © 2017 Elsevier Ltd. All rights reserved.

Escherichia coli O157:H7 induces stronger plant immunity than Salmonella enterica Typhimurium SL1344.

PubMed

Roy, Debanjana; Panchal, Shweta; Rosa, Bruce A; Melotto, Maeli

2013-04-01

Consumption of fresh produce contaminated with bacterial human pathogens has resulted in various, sometimes deadly, disease outbreaks. In this study, we assessed plant defense responses induced by the fully pathogenic bacteria Escherichia coli O157:H7 and Salmonella enterica serovar Typhimurium SL1344 in both Arabidopsis thaliana and lettuce (Lactuca sativa). Unlike SL1344, O157:H7 induced strong plant immunity at both pre-invasion and post-invasion steps of infection. For instance, O157:H7 triggered stomatal closure even under high relative humidity, an environmental condition that generally weakens plant defenses against bacteria in the field and laboratory conditions. SL1344 instead induced a transient stomatal immunity. We also observed that PR1 gene expression was significantly higher in Arabidopsis leaves infected with O157:H7 compared with SL1344. These results suggest that plants may recognize and respond to some human pathogens more effectively than others. Furthermore, stomatal immunity can diminish the penetration of human pathogens through the leaf epidermis, resulting in low bacterial titers in the plant apoplast and suggesting that additional control measures can be employed to prevent food contamination. The understanding of how plant responses can diminish bacterial contamination is paramount in preventing outbreaks and improving the safety of food supplies.

Escherichia coli and selected veterinary and zoonotic pathogens isolated from environmental sites in companion animal veterinary hospitals in southern Ontario.

PubMed

Murphy, Colleen P; Reid-Smith, Richard J; Boerlin, Patrick; Weese, J Scott; Prescott, John F; Janecko, Nicol; Hassard, Lori; McEwen, Scott A

2010-09-01

Hospital-based infection control in veterinary medicine is emerging and the role of the environment in hospital-acquired infections (HAI) in veterinary hospitals is largely unknown. This study was initiated to determine the recovery of Escherichia coli and selected veterinary and zoonotic pathogens from the environments of 101 community veterinary hospitals. The proportion of hospitals with positive environmental swabs were: E. coli--92%, Clostridium difficile--58%, methicillin-resistant Staphylococcus aureus (MRSA)--9%, CMY-2 producing E. coli--9%, methicillin-resistant Staphylococcus pseudintermedius--7%, and Salmonella--2%. Vancomycin-resistant Enterococcus spp., canine parvovirus, and feline calicivirus were not isolated. Prevalence of antimicrobial resistance in E. coli isolates was low. Important potential veterinary and human pathogens were recovered including Canadian epidemic strains MRSA-2 and MRSA-5, and C. difficile ribotype 027. There is an environmental reservoir of pathogens in veterinary hospitals; therefore, additional studies are required to characterize risk factors associated with HAI in companion animals, including the role of the environment.

The pap Operon of Avian Pathogenic Escherichia coli Strain O1:K1 Is Located on a Novel Pathogenicity Island

PubMed Central

Kariyawasam, Subhashinie; Johnson, Timothy J.; Nolan, Lisa K.

2006-01-01

We have identified a 56-kb pathogenicity island (PAI) in avian pathogenic Escherichia coli strain O1:K1 (APEC-O1). This PAI, termed PAI IAPEC-O1, is integrated adjacent to the 3′ end of the pheV tRNA gene. It carries putative virulence genes of APEC (pap operon), other E. coli genes (tia and ireA), and a 1.5-kb region unique to APEC-O1. The kps gene cluster required for the biosynthesis of polysialic acid capsule was mapped to a location immediately downstream of this PAI. PMID:16369033

Draft Genomic Sequencing of Six Potential Extraintestinal Pathogenic Escherichia coli Isolates from Retail Chicken Meat

PubMed Central

Xu, Aixia; Johnson, James R.; Sheen, Shiowshuh; Needleman, David S.

2018-01-01

ABSTRACT Potential extraintestinal pathogenic Escherichia coli strains DP254, WH333, WH398, F356, FEX675, and FEX725 were isolated from retail chicken meat products. Here, we report the draft genome sequences for these six E. coli isolates, which are currently being used in food safety research. PMID:29798928

Agrochemicals indirectly increase survival of E. coli O157:H7 and indicator bacteria by reducing ecosystem services.

PubMed

Staley, Zachery R; Rohr, Jason R; Senkbeil, Jacob K; Harwood, Valerie J

Storm water and agricultural runoff frequently contain agrochemicals, fecal indicator bacteria (FIB), and zoonotic pathogens. Entry of such contaminants into aquatic ecosystems may affect ecology and human health. This study tested the hypothesis that the herbicide atrazine and the fungicide chlorothalonil indirectly affect the survival of FIB (Escherichia coli and Enterococcus faecalis) and a pathogen (E. coli O157:H7) by altering densities of protozoan predators or by altering competition from autochthonous bacteria. Streptomycin-resistant E. coli, En. faecalis, and E. coli O157:H7 were added to microcosms composed of Florida river water containing natural protozoan and bacterial populations. FIB, pathogen, and protozoan densities were monitored over six days. Known metabolic inhibitors, cycloheximide and streptomycin, were used to inhibit autochthonous protozoa or bacteria, respectively. The inhibitors made it possible to isolate the effects of predation or competition on survival of allochthonous bacteria, and each treatment increased the survival of FIB and pathogens. Chlorothalonil's effect was similar to that of cycloheximide, significantly reducing protozoan densities and elevating densities of FIB and pathogens relative to the control. Atrazine treatment did not affect protozoan densities, but, through an effect on competition, resulted in significantly greater densities of En. faecalis and E. coli O157:H7. Hence, by reducing predaceous protozoa and bacterial competitors that facilitate purifying water bodies of FIBs and human pathogens, chlorothalonil and atrazine indirectly diminished an ecosystem service of fresh water.

Label-Free 3D Ag Nanoflower-Based Electrochemical Immunosensor for the Detection of Escherichia coli O157:H7 Pathogens

NASA Astrophysics Data System (ADS)

Huang, He; Liu, Minghuan; Wang, Xiangsheng; Zhang, Wenjie; Yang, Da-Peng; Cui, Lianhua; Wang, Xiansong

2016-11-01

It is highly desirable to develop a rapid and simple method to detect pathogens. Combining nanomaterials with electrochemical techniques is an efficient way for pathogen detection. Herein, a novel 3D Ag nanoflower was prepared via a biomineralization method by using bovine serum albumin (BSA) as a template. It was adopted as a sensing interface to construct an electrochemical bacteria immunosensor for the rapid detection of foodborne pathogens Escherichia coli ( E. coli) O157:H7. Bacterial antibody was immobilized onto the surface of Ag nanoflowers through covalent conjugation. Electrochemical impedance spectroscopy (EIS) was used to detect and validate the resistance changes, where [Fe(CN)6]3-/4- acted as the redox probe. A linear relation between R et and E. coli concentration was obtained in the E. coli concentration range of 3.0 × 102-3.0 × 108 cfu mL-1. The as-prepared biosensor gave rise to an obvious response to E. coli but had no distinct response to Cronobacter sakazakii, methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus albus, Lactobacillus easei, and Shigella flexneri, revealing a high selectivity for the detection of the pathogens down to 100 cfu mL-1 in a short time. We believe that this BSA-conjugated 3D Ag nanoflowers could be used as a powerful interface material with good conductivity and biocompatibility for improving pathogen detection and treatment in the field of medicine, environment, and food safety.

Thymus transcriptome reveals novel pathways in response to avian pathogenic Escherichia coli infection

PubMed Central

Sun, H.; Liu, P.; Nolan, L. K.; Lamont, S. J.

2016-01-01

Avian pathogenic Escherichia coli (APEC) can cause significant morbidity in chickens. The thymus provides the essential environment for T cell development; however, the thymus transcriptome has not been examined for gene expression in response to APEC infection. An improved understanding of the host genomic response to APEC infection could inform future breeding programs for disease resistance and APEC control. We therefore analyzed the transcriptome of the thymus of birds challenged with APEC, contrasting susceptible and resistant phenotypes. Thousands of genes were differentially expressed in birds of the 5-day post infection (dpi) challenged-susceptible group vs. 5 dpi non-challenged, in 5 dpi challenged-susceptible vs. 5 dpi challenged-resistant birds, as well as in 5 dpi vs. one dpi challenged-susceptible birds. The Toll-like receptor signaling pathway was the major innate immune response for birds to respond to APEC infection. Moreover, lysosome and cell adhesion molecules pathways were common mechanisms for chicken response to APEC infection. The T-cell receptor signaling pathway, cell cycle, and p53 signaling pathways were significantly activated in resistant birds to resist APEC infection. These results provide a comprehensive assessment of global gene networks and biological functionalities of differentially expressed genes in the thymus under APEC infection. These findings provide novel insights into key molecular genetic mechanisms that differentiate host resistance from susceptibility in this primary lymphoid tissue, the thymus. PMID:27466434

Genomic characterisation of an endometrial pathogenic Escherichia coli strain reveals the acquisition of genetic elements associated with extra-intestinal pathogenicity.

PubMed

Goldstone, Robert J; Popat, Roman; Schuberth, Hans-Joachim; Sandra, Olivier; Sheldon, I Martin; Smith, David G E

2014-12-06

Strains of Escherichia coli cause a wide variety of intestinal and extra-intestinal diseases in both humans and animals, and are also often found in healthy individuals or the environment. Broadly, a strong phylogenetic relationship exists that distinguishes most E. coli causing intestinal disease from those that cause extra-intestinal disease, however, isolates within a recently described subclass of Extra-Intestinal Pathogenic E. coli (ExPEC), termed endometrial pathogenic E. coli, tend to be phylogenetically distant from the vast majority of characterised ExPECs, and more closely related to human intestinal pathogens. In this work, we investigate the genetic basis for ExPEC infection in the prototypic endometrial pathogenic E. coli strain MS499. By investigating the genome of MS499 in comparison with a range of other E. coli sequences, we have discovered that this bacterium has acquired substantial lengths of DNA which encode factors more usually associated with ExPECs and less frequently found in the phylogroup relatives of MS499. Many of these acquired factors, including several iron acquisition systems and a virulence plasmid similar to that found in several ExPECs such as APEC O1 and the neonatal meningitis E. coli S88, play characterised roles in a variety of typical ExPEC infections and appear to have been acquired recently by the evolutionary lineage leading to MS499. Taking advantage of the phylogenetic relationship we describe between MS499 and several other closely related E. coli isolates from across the globe, we propose a step-wise evolution of a novel clade of sequence type 453 ExPECs within phylogroup B1, involving the recruitment of ExPEC virulence factors into the genome of an ancestrally non-extraintestinal E. coli, which has repurposed this lineage with the capacity to cause extraintestinal disease. These data reveal the genetic components which may be involved in this phenotype switching, and argue that horizontal gene exchange may be a key factor in the emergence of novel lineages of ExPECs.

Outsider to insider: resetting the natural host niche of commensal E. coli K-12.

PubMed

Sahu, Upasana; Kar, Sudeshna

2012-01-01

The status of E. coli K-12 as an exclusively non-invasive, non-pathogenic bacterium has almost been incontrovertible. Our recent finding that a mutation in one of its main architectural protein, HU, converts E. coli K-12 to an actively invasive form suggests that gaining host cell entry might be an expedient survival tactic for traditional commensals during certain altered host conditions. The mutant E. coli (SK3842) exhibits properties usually associated with pathogenic bacteria: host cell invasion, phagosomal disruption and intracellular replication. However, unlike the situation with some pathogens, internalized SK3842 imparts anti-apoptotic and cyto-protective effects rather than lethality on the host cell, both in vitro and in vivo. Here, we show that SK3842 also provides colonization resistance against other invasive pathogens--a trait not shared by the parental commensal strain. Thus, the altered lifestyle of SK3842 encompasses characteristics both from traditional pathogens as well as beneficial probiotic strains.

A neighborhood statistics model for predicting stream pathogen indicator levels.

PubMed

Pandey, Pramod K; Pasternack, Gregory B; Majumder, Mahbubul; Soupir, Michelle L; Kaiser, Mark S

2015-03-01

Because elevated levels of water-borne Escherichia coli in streams are a leading cause of water quality impairments in the U.S., water-quality managers need tools for predicting aqueous E. coli levels. Presently, E. coli levels may be predicted using complex mechanistic models that have a high degree of unchecked uncertainty or simpler statistical models. To assess spatio-temporal patterns of instream E. coli levels, herein we measured E. coli, a pathogen indicator, at 16 sites (at four different times) within the Squaw Creek watershed, Iowa, and subsequently, the Markov Random Field model was exploited to develop a neighborhood statistics model for predicting instream E. coli levels. Two observed covariates, local water temperature (degrees Celsius) and mean cross-sectional depth (meters), were used as inputs to the model. Predictions of E. coli levels in the water column were compared with independent observational data collected from 16 in-stream locations. The results revealed that spatio-temporal averages of predicted and observed E. coli levels were extremely close. Approximately 66 % of individual predicted E. coli concentrations were within a factor of 2 of the observed values. In only one event, the difference between prediction and observation was beyond one order of magnitude. The mean of all predicted values at 16 locations was approximately 1 % higher than the mean of the observed values. The approach presented here will be useful while assessing instream contaminations such as pathogen/pathogen indicator levels at the watershed scale.

Vaccines for viral and bacterial pathogens causing acute gastroenteritis: Part II: Vaccines for Shigella, Salmonella, enterotoxigenic E. coli (ETEC) enterohemorragic E. coli (EHEC) and Campylobacter jejuni

PubMed Central

O’Ryan, Miguel; Vidal, Roberto; del Canto, Felipe; Carlos Salazar, Juan; Montero, David

2015-01-01

In Part II we discuss the following bacterial pathogens: Shigella, Salmonella (non-typhoidal), diarrheogenic E. coli (enterotoxigenic and enterohemorragic) and Campylobacter jejuni. In contrast to the enteric viruses and Vibrio cholerae discussed in Part I of this series, for the bacterial pathogens described here there is only one licensed vaccine, developed primarily for Vibrio cholerae and which provides moderate protection against enterotoxigenic E. coli (ETEC) (Dukoral®), as well as a few additional candidates in advanced stages of development for ETEC and one candidate for Shigella spp. Numerous vaccine candidates in earlier stages of development are discussed. PMID:25715096

Genome dynamics and its impact on evolution of Escherichia coli.

PubMed

Dobrindt, Ulrich; Chowdary, M Geddam; Krumbholz, G; Hacker, J

2010-08-01

The Escherichia coli genome consists of a conserved part, the so-called core genome, which encodes essential cellular functions and of a flexible, strain-specific part. Genes that belong to the flexible genome code for factors involved in bacterial fitness and adaptation to different environments. Adaptation includes increase in fitness and colonization capacity. Pathogenic as well as non-pathogenic bacteria carry mobile and accessory genetic elements such as plasmids, bacteriophages, genomic islands and others, which code for functions required for proper adaptation. Escherichia coli is a very good example to study the interdependency of genome architecture and lifestyle of bacteria. Thus, these species include pathogenic variants as well as commensal bacteria adapted to different host organisms. In Escherichia coli, various genetic elements encode for pathogenicity factors as well as factors, which increase the fitness of non-pathogenic bacteria. The processes of genome dynamics, such as gene transfer, genome reduction, rearrangements as well as point mutations contribute to the adaptation of the bacteria into particular environments. Using Escherichia coli model organisms, such as uropathogenic strain 536 or commensal strain Nissle 1917, we studied mechanisms of genome dynamics and discuss these processes in the light of the evolution of microbes.

[The detection of strains of Esherichia coll producing Shiga toxin in populations of normal intestinal microbiota in children with functional disorders of gastrointestinal tract].

PubMed

Ivanova, E I; Popkova, S M; Dzhioev, Iu P; Rakova, E B; Nemchenko, U M; Rychkova, L V

2014-11-01

In intestinal ecosystem, interchange of genetic material between different types of bacteria and other representatives of family Enterobacteriaceae results in development of types of normal colibacillus with genetic characteristics of pathogenicity. This occurrence can be considered as a theoretical substantiation for labeling such strains as pathobionts. The polymerase chain reaction was implemented to analyze 96 strains of different types of Escherichia coli (with normal and weak zymogenic activity and hemolytic activity) isolated from children with functional disorders of gastrointestinal tract. The purpose was to detect presence of gens coding capacity of toxin production (six1, stx2). In intestinal biotope of children, circulation of strains of Escherichia coli producing shiga toxin having no relation to pathogenic group being representatives of normal indigenous microbiota. The presence of gens stx1 and stx2 in various biochemical types of Escherichia coli permits establishing fact of forming of reservoir of potential pathogenicity in non-pathogenic forms of Escherichia coli. The presence of gen (verotoxin 1) in genome of various types of Escherichia coli isolated from one single biotope testifies possible horizontal transmission of factors of pathogenicity in intestinal biotope.

Released products of pathogenic bacteria stimulate biofilm formation by Escherichia coli K-12 strains.

PubMed

Vacheva, Anna; Ivanova, Radka; Paunova-Krasteva, Tsvetelina; Stoitsova, Stoyanka

2012-06-01

It has recently been shown that pathogens with a limited capacity for sessile growth (like some Escherichia coli O157 strains) can benefit from the presence of other bacteria and form mixed biofilms with companion strains. This study addresses the question whether pathogens may influence attached growth of E. coli non-pathogenic strains via secreted factors. We compared the biofilm-modulating effects of sterile stationary-phase culture media of a biofilm non-producing strain of E. coli O157:H, a laboratory biofilm-producing E. coli K-12 strain and a biofilm-forming strain of the pathogen Yersina enterocolitica O:3. Sessile growth was monitored as biomass (crystal violet assay), exopolysaccharide (ELLA) and morphology (scanning electron and confocal laser microscopy). With two of the E. coli K-12 strains stimulation of biofilm formation by all supernatants was achieved, but only the pathogens' secreted products induced biomass increase in some 'biofilm-deficient' K-12 strains. Lectin-peroxidase labeling indicated changes in colanic acid and poly-N-acetylglucosamine amounts in extracellular matrices. The contribution of indole, protein and polysaccharide to the biofilm-modulating activities of the supernatants was compared. Indole, in concentrations equal to those established in the supernatants, suppressed sessile growth in one K-12 strain. Proteinase K significantly reduced the stimulatory effects of all supernatants, indicating a prominent role of protein/peptide factor(s) in biofilm promotion. The amount of released polysaccharides (rPS) in the supernatants was quantitated then comparable quantities of isolated rPS were applied during biofilm growth. The three rPS had notable strain-specific effects with regard to both the strain-source of the rPS and the E. coli K-12 target strain.

Fecal contamination of drinking water in Kericho District, Western Kenya: role of source and household water handling and hygiene practices.

PubMed

Too, Johana Kiplagat; Kipkemboi Sang, Willy; Ng'ang'a, Zipporah; Ngayo, Musa Otieno

2016-08-01

Inadequate protection of water sources, and poor household hygienic and handling practices have exacerbated fecal water contamination in Kenya. This study evaluated the rate and correlates of thermotolerant coliform (TTC) household water contamination in Kericho District, Western Kenya. Culture and multiplex polymerase chain reaction (PCR) techniques were used to characterize TTCs. The disk diffusion method was used for antibiotic susceptibility profiling of pathogenic Escherichia coli. Out of the 103 households surveyed, 48 (46.6%) had TTC contaminated drinking water (TTC levels of >10 cfu/100 mL). Five of these households were contaminated with pathogenic E. coli, including 40% enteroaggregative E. coli, 40% enterotoxigenic E. coli, and 20% enteropathogenic E. coli. All these pathogenic E. coli strains were multidrug resistant to sulfamethoxazole/trimethoprim, ampicillin, tetracycline and ampicillin/sulbactam. Rural household locality, drinking water hand contact, water storage container cleaning practice, hand washing before water withdrawal, water source total coliforms <10 cfu/100 mL, temperature, and free chlorine levels were associated with TTC contamination of household drinking water. Significant proportions of household drinking water in Kericho District are contaminated with TTCs including with pathogenic multidrug-resistant E. coli. Source and household hygiene and practices contribute significantly to drinking water contamination.

Spondylodiscitis in a healthy 12-year-old girl with Extraintestinal pathogenic Escherichia coli (ExPEC) bacteraemia.

PubMed

Gaschignard, J; Geslain, G; Mallet, C; Lorrot, M; Blot, N; Alison, M; Bonacorsi, S

2017-05-31

Escherichia coli (E. coli) is rarely implicated in bone or joint infections in children. We discuss the case of a healthy 12-year-old girl with an E. coli bacteraemia and a T11-T12 spondylodiscitis revealed by magnetic resonance imaging. The strain harboured serogroup O1:K1 and virulence factors common to highly virulent extra intestinal pathogenic E. coli (ExPEC). Immunological work-up was normal. The identification of E. coli in a spondylodiscitis should lead to the search for immunosuppression of the host and virulence factors of the strain, particularly those of ExPEC.

Detection of Pathogenic Escherichia coli in Samples Collected at an Abattoir in Zaria, Nigeria and at Different Points in the Surrounding Environment

PubMed Central

Kabiru, Lawan Mohammed; Bello, Mohammed; Kabir, Junaid; Grande, Laura; Morabito, Stefano

2015-01-01

Pathogenic Escherichia coli can be released with the wastes coming from slaughterhouses into the environment, where they can persist. We investigated the presence of diarrheagenic E. coli in specimens taken at an abattoir located in the Zaria region, Nigeria, in samples of water from the river Koreye, where the effluent from the abattoir spills in, and vegetable specimens taken at a nearby farm. All the isolated E. coli were assayed for the production of Shiga toxins (Stx) by using the Ridascreen verotoxin Immunoassay and by PCR amplification of genes associated with the diarrheagenic E. coli. Three strains from the rectal content of two slaughtered animals and a cabbage were positive for the presence of the Stx-coding genes. Additionally we have isolated one Enteroaggregative E. coli (EAggEC) from the abattoir effluent and two Subtilase-producing E. coli from the slaughterhouse’s effluent and a sample of carrots. Our results provide evidence that pathogenic E. coli can contaminate the environment as a result of the discharge into the environment of untreated abattoir effluent, representing a reservoir for STEC and other diarrheagenic E. coli favouring their spread to crops. PMID:25590145

Coculture of Escherichia coli O157:H7 with a Nonpathogenic E. coli Strain Increases Toxin Production and Virulence in a Germfree Mouse Model

PubMed Central

Goswami, Kakolie; Chen, Chun; Xiaoli, Lingzi; Eaton, Kathryn A.

2015-01-01

Escherichia coli O157:H7 is a notorious foodborne pathogen due to its low infectious dose and the disease symptoms it causes, which include bloody diarrhea and severe abdominal cramps. In some cases, the disease progresses to hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS), due to the expression of one or more Shiga toxins (Stx). Isoforms of Stx, including Stx2a, are encoded within temperate prophages. In the presence of certain antibiotics, phage induction occurs, which also increases the expression of toxin genes. Additionally, increased Stx2 accumulation has been reported when O157:H7 was cocultured with phage-susceptible nonpathogenic E. coli. This study characterized an E. coli O157:H7 strain, designated PA2, that belongs to the hypervirulent clade 8 cluster. Stx2a levels after ciprofloxacin induction were lower for PA2 than for the prototypical outbreak strains Sakai and EDL933. However, during coculture with the nonpathogenic strain E. coli C600, PA2 produced Stx2a levels that were 2- to 12-fold higher than those observed during coculture with EDL933 and Sakai, respectively. Germfree mice cocolonized by PA2 and C600 showed greater kidney damage, increased Stx2a accumulation in feces, and more visible signs of disease than mice given PA2 or C600 alone. These data suggest one mechanism by which microorganisms associated with the colonic microbiota could enhance the virulence of E. coli O157:H7, particularly a subset of clade 8 strains. PMID:26259815

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

On the Transport of Viable but Non-Culturable (VBNC) E.coli O157:H7 in Soil and Groundwater

NASA Astrophysics Data System (ADS)

Kartz, C. R.; Kachanoski, G.; Dyck, M. F.

2010-12-01

The influence of the viable but non-culturable (VBNC) state on the expression of specific phenotypic traits of Enterohemorrhagic Escherichia coli O157:H7 as well as its transport behaviour in porous media has been examined in this study. E.coli O157:H7 is a human pathogen capable of entering a viable but non-culturable (VBNC) state following exposure to sublethal stress. In the VBNC state, E.coli O157:H7 is not detectable by standard culture techniques, yet is able to retain its virulence and ability to cause illness in humans. To date there is no in-depth information regarding the transport of VBNC E.coli species in soil or groundwater. Due to the public health risk, it becomes important to examine whether discrepancies exist between the transport behaviors of culturable and VBNC E.coli O157:H7 to help decide if current protocols for detecting this pathogen are accurate. This study identifies and contrasts transport-related properties of the two cell stages including hydrophobicity, extracellular polymeric substance (EPS) composition, and cell widths/lengths. Transport behaviors of the two cellular states are quantified and compared using column transport assays. Our results show that when E.coli O157:H7 cells enter into the VBNC state, there is an accompanied decrease in the hydrophobicity of the cells, shrinking of the cell profile from rod-shaped to coccoid, as well as a significant increase in tightly-bound surface proteins and sugars. Transport assays revealed a notable increase in mass flux when cells were in the VBNC state versus the culturable state. This research will contribute to the current knowledge-base describing E.coli O157:H7 cells in the VBNC state, spark dialogue concerning the accuracy of currently-used identification protocols, as well as add further evidence to the notion that bacteria transport in the subsurface is a truly dynamic process.

Role of F1C fimbriae, flagella, and secreted bacterial components in the inhibitory effect of probiotic Escherichia coli Nissle 1917 on atypical enteropathogenic E. coli infection.

PubMed

Kleta, Sylvia; Nordhoff, Marcel; Tedin, Karsten; Wieler, Lothar H; Kolenda, Rafal; Oswald, Sibylle; Oelschlaeger, Tobias A; Bleiss, Wilfried; Schierack, Peter

2014-05-01

Enteropathogenic Escherichia coli (EPEC) is recognized as an important intestinal pathogen that frequently causes acute and persistent diarrhea in humans and animals. The use of probiotic bacteria to prevent diarrhea is gaining increasing interest. The probiotic E. coli strain Nissle 1917 (EcN) is known to be effective in the treatment of several gastrointestinal disorders. While both in vitro and in vivo studies have described strong inhibitory effects of EcN on enteropathogenic bacteria, including pathogenic E. coli, the underlying molecular mechanisms remain largely unknown. In this study, we examined the inhibitory effect of EcN on infections of porcine intestinal epithelial cells with atypical enteropathogenic E. coli (aEPEC) with respect to single infection steps, including adhesion, microcolony formation, and the attaching and effacing phenotype. We show that EcN drastically reduced the infection efficiencies of aEPEC by inhibiting bacterial adhesion and growth of microcolonies, but not the attaching and effacing of adherent bacteria. The inhibitory effect correlated with EcN adhesion capacities and was predominantly mediated by F1C fimbriae, but also by H1 flagella, which served as bridges between EcN cells. Furthermore, EcN seemed to interfere with the initial adhesion of aEPEC to host cells by secretion of inhibitory components. These components do not appear to be specific to EcN, but we propose that the strong adhesion capacities enable EcN to secrete sufficient local concentrations of the inhibitory factors. The results of this study are consistent with a mode of action whereby EcN inhibits secretion of virulence-associated proteins of EPEC, but not their expression.

Broad-host-range plasmids for red fluorescent protein labeling of gram-negative bacteria for use in the zebrafish model system.

PubMed

Singer, John T; Phennicie, Ryan T; Sullivan, Matthew J; Porter, Laura A; Shaffer, Valerie J; Kim, Carol H

2010-06-01

To observe real-time interactions between green fluorescent protein-labeled immune cells and invading bacteria in the zebrafish (Danio rerio), a series of plasmids was constructed for the red fluorescent protein (RFP) labeling of a variety of fish and human pathogens. The aim of this study was to create a collection of plasmids that would express RFP pigments both constitutively and under tac promoter regulation and that would be nontoxic and broadly transmissible to a variety of Gram-negative bacteria. DNA fragments encoding the RFP dimeric (d), monomeric (m), and tandem dimeric (td) derivatives d-Tomato, td-Tomato, m-Orange, and m-Cherry were cloned into the IncQ-based vector pMMB66EH in Escherichia coli. Plasmids were mobilized into recipient strains by conjugal mating. Pigment production was inducible in Escherichia coli, Pseudomonas aeruginosa, Edwardsiella tarda, and Vibrio (Listonella) anguillarum strains by isopropyl-beta-d-thiogalactopyranoside (IPTG) treatment. A spontaneous mutant exconjugant of P. aeruginosa PA14 was isolated that expressed td-Tomato constitutively. Complementation analysis revealed that the constitutive phenotype likely was due to a mutation in lacI(q) carried on pMMB66EH. DNA sequence analysis confirmed the presence of five transitions, four transversions, and a 2-bp addition within a 14-bp region of lacI. Vector DNA was purified from this constitutive mutant, and structural DNA sequences for RFP pigments were cloned into the constitutive vector. Exconjugants of P. aeruginosa, E. tarda, and V. anguillarum expressed all pigments in an IPTG-independent fashion. Results from zebrafish infectivity studies indicate that RFP-labeled pathogens will be useful for the study of real-time interactions between host cells of the innate immune system and the infecting pathogen.

Screening host proteins required for bacterial adherence after H9N2 virus infection.

PubMed

Ma, Li-Li; Sun, Zhen-Hong; Xu, Yu-Lin; Wang, Shu-Juan; Wang, Hui-Ning; Zhang, Hao; Hu, Li-Ping; Sun, Xiao-Mei; Zhu, Lin; Shang, Hong-Qi; Zhu, Rui-Liang; Wei, Kai

2018-01-01

H9N2 subtype low pathogenic avian influenza virus (LPAIV) is distributed worldwide and causes great economic losses in the poultry industry, especially when complicated with other bacterial infections. Tissue damages caused by virus infection provide an opportunity for bacteria invasion, but this mechanism is not sufficient for low pathogenic strains. Moreover, although H9N2 virus infection was demonstrated to promote bacterial infection in several studies, its mechanism remained unclear. In this study, infection experiments in vivo and in vitro demonstrated that the adhesion of Escherichia coli (E. coli) to host cells significantly increased after H9N2 virus infection, and this increase was not caused by pathological damages. Subsequently, we constructed a late chicken embryo infection model and used proteomics techniques to analyze the expression of proteins associated with bacterial adhesion after H9N2 virus infection. A total of 279 significantly differential expressed proteins were detected through isobaric tags for relative and absolute quantitation (iTRAQ) coupled with nano-liquid chromatography-tandem mass spectrometry (nano-LC-MS/MS) analysis. The results of Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis showed that differentially expressed proteins were enriched in host innate immunity; cell proliferation, differentiation, and apoptosis; and pathogenicity-related signaling pathways. Finally, we screened out several proteins, such as TGF-β1, integrins, cortactin, E-cadherin, vinculin, and fibromodulin, which were probably associated with bacterial adhesion. The study analyzed the mechanism of secondary bacterial infection induced by H9N2 virus infection from a novel perspective, which provided theoretical and data support for investigating the synergistic infection mechanism between the H9N2 virus and bacteria. Copyright © 2017 Elsevier B.V. All rights reserved.

Pathotyping of Escherichia coli isolated from community toilet wastewater and stored drinking water in a slum in Bangladesh.

PubMed

Harada, H; Fujimori, Y; Gomi, R; Ahsan, Md N; Fujii, S; Sakai, A; Matsuda, T

2018-06-01

This study investigated the occurrence of Escherichia coli pathotypes in sanitary wastewater and drinking water in a Bangladeshi urban slum and the potential associations between these sources. We examined 621 E. coli isolates from sanitary wastewater and stored drinking water by multiplex PCR and dual-index sequencing, classifying them into eight pathotypes based on 14 virulence genes and additionally evaluating the possession of the human-specific E. coli genetic biomarker H8. The proportions of pathogenic E. coli were significantly different (P < 0·001) between wastewater (18·6%) and drinking water (1·7%). StIb-positive enterotoxigenic E. coli (ETEC) were predominant in wastewater, indicating that people in the site carried ETEC. In contrast, no ETEC was present in drinking water and the proportion of H8-positive isolates was significantly smaller (7·8%) than that in wastewater (16·3%) (P = 0·001). Our findings indicate that sanitary wastewater from the slum was heavily contaminated with pathogenic E. coli, posing a great health risk. Furthermore, E. coli contamination of drinking water could be derived from not only human but also other sources. Sanitary wastewater from an urban slum was heavily contaminated with pathogenic Escherichia coli. It is worth noting a great health risk of accidental exposure to pathogenically contaminated wastewater improperly discharged in and around urban slums. The distinct difference in pathotypes between wastewater and drinking water and the significantly smaller positive proportion of the human-specific E. coli genetic biomarker (H8) in drinking water indicate that drinking water contamination could be derived from not only human but also other sources. This highlights that pathotyping in association with the H8 marker provides an indication of pathogen contamination sources of environmental transmission media. © 2018 The Society for Applied Microbiology.

Epidemic potential of Escherichia coli ST131 and Klebsiella pneumoniae ST258: a systematic review and meta-analysis

PubMed Central

Dautzenberg, M J D; Haverkate, M R; Bonten, M J M; Bootsma, M C J

2016-01-01

Objectives Observational studies have suggested that Escherichia coli sequence type (ST) 131 and Klebsiella pneumoniae ST258 have hyperendemic properties. This would be obvious from continuously high incidence and/or prevalence of carriage or infection with these bacteria in specific patient populations. Hyperendemicity could result from increased transmissibility, longer duration of infectiousness, and/or higher pathogenic potential as compared with other lineages of the same species. The aim of our research is to quantitatively estimate these critical parameters for E. coli ST131 and K. pneumoniae ST258, in order to investigate whether E. coli ST131 and K. pneumoniae ST258 are truly hyperendemic clones. Primary outcome measures A systematic literature search was performed to assess the evidence of transmissibility, duration of infectiousness, and pathogenicity for E. coli ST131 and K. pneumoniae ST258. Meta-regression was performed to quantify these characteristics. Results The systematic literature search yielded 639 articles, of which 19 data sources provided information on transmissibility (E. coli ST131 n=9; K. pneumoniae ST258 n=10)), 2 on duration of infectiousness (E. coli ST131 n=2), and 324 on pathogenicity (E. coli ST131 n=285; K. pneumoniae ST258 n=39). Available data on duration of carriage and on transmissibility were insufficient for quantitative assessment. In multivariable meta-regression E. coli isolates causing infection were associated with ST131, compared to isolates only causing colonisation, suggesting that E. coli ST131 can be considered more pathogenic than non-ST131 isolates. Date of isolation, location and resistance mechanism also influenced the prevalence of ST131. E. coli ST131 was 3.2 (95% CI 2.0 to 5.0) times more pathogenic than non-ST131. For K. pneumoniae ST258 there were not enough data for meta-regression assessing the influence of colonisation versus infection on ST258 prevalence. Conclusions With the currently available data, it cannot be confirmed nor rejected, that E. coli ST131 or K. pneumoniae ST258 are hyperendemic clones. PMID:26988349

A Family of Indoles Regulate Virulence and Shiga Toxin Production in Pathogenic E. coli

PubMed Central

Izrayelit, Yevgeniy; Bhatt, Shantanu; Cartwright, Emily; Wang, Wei; Swimm, Alyson I.; Benian, Guy M.; Schroeder, Frank C.; Kalman, Daniel

2013-01-01

Enteropathogenic Escherichia coli (EPEC), enterohemorrhagic E. coli (EHEC) and enteroaggregative E. coli (EAEC) are intestinal pathogens that cause food and water-borne disease in humans. Using biochemical methods and NMR-based comparative metabolomics in conjunction with the nematode Caenorhabditis elegans, we developed a bioassay to identify secreted small molecules produced by these pathogens. We identified indole, indole-3-carboxaldehyde (ICA), and indole-3-acetic acid (IAA), as factors that only in combination are sufficient to kill C. elegans. Importantly, although lethal to C. elegans, these molecules downregulate several bacterial processes important for pathogenesis in mammals. These include motility, biofilm formation and production of Shiga toxins. Some pathogenic E. coli strains are known to contain a Locus of Enterocyte Effacement (LEE), which encodes virulence factors that cause “attaching and effacing” (A/E) lesions in mammals, including formation of actin pedestals. We found that these indole derivatives also downregulate production of LEE virulence factors and inhibit pedestal formation on mammalian cells. Finally, upon oral administration, ICA inhibited virulence and promoted survival in a lethal mouse infection model. In summary, the C. elegans model in conjunction with metabolomics has facilitated identification of a family of indole derivatives that broadly regulate physiology in E. coli, and virulence in pathogenic strains. These molecules may enable development of new therapeutics that interfere with bacterial small-molecule signaling. PMID:23372726

Using the agricultural environment to select better surrogates for foodborne pathogens associated with fresh produce.

PubMed

Cook, Kimberly L; Givan, Ethan C; Mayton, Holly M; Parekh, Rohan R; Taylor, Ritchie; Walker, Sharon L

2017-12-04

Despite continuing efforts to reduce foodborne pathogen contamination of fresh produce, significant outbreaks continue to occur. Identification of appropriate surrogates for foodborne pathogens facilitates relevant research to identify reservoirs and amplifiers of these contaminants in production and processing environments. Therefore, the objective of this study was to identify environmental Escherichia coli isolates from manures (poultry, swine and dairy) and surface water sources with properties similar to those of the produce associated foodborne pathogens E. coli O157:H7 and Salmonella enterica serotype Typhimurium. The most similar environmental E. coli isolates were from poultry (n=3) and surface water (n=1) sources. The best environmental E. coli surrogates had cell surface characteristics (zeta potential, hydrophobicity and exopolysaccharide composition) that were similar (i.e., within 15%) to those of S. Typhimurium and/or formed biofilms more often when grown in low nutrient media prepared from lettuce lysates (24%) than when grown on high nutrient broth (7%). The rate of attachment of environmental isolates to lettuce leaves was also similar to that of S. Typhimurium. In contrast, E. coli O157:H7, a commonly used E. coli quality control strain and swine isolates behaved similarly; all were in the lowest 10% of isolates for biofilm formation and leaf attachment. These data suggest that the environment may provide a valuable resource for selection of surrogates for foodborne pathogens. Published by Elsevier B.V.

A multi-pathogen selective enrichment broth for simultaneous growth of Salmonella enteria, Escherichia coli O157:H7 and Shigella flexneri

USDA-ARS?s Scientific Manuscript database

Salmonella, Shigella, and Escherichia coli O157:H7 contaminate similar types of food and all three can cause foodborne disease. Traditional microbiological enrichment broths to detect these pathogens are different in terms of their composition, which limits the application of multi-pathogen detectio...

Effects of co-occurring Wolbachia and Spiroplasma endosymbionts on the Drosophila immune response against insect pathogenic and non-pathogenic bacteria.

PubMed

Shokal, Upasana; Yadav, Shruti; Atri, Jaishri; Accetta, Julia; Kenney, Eric; Banks, Katherine; Katakam, Akash; Jaenike, John; Eleftherianos, Ioannis

2016-02-09

Symbiotic interactions between microbes and animals are common in nature. Symbiotic organisms are particularly common in insects and, in some cases, they may protect their hosts from pathogenic infections. Wolbachia and Spiroplasma endosymbionts naturally inhabit various insects including Drosophila melanogaster fruit flies. Therefore, this symbiotic association is considered an excellent model to investigate whether endosymbiotic bacteria participate in host immune processes against certain pathogens. Here we have investigated whether the presence of Wolbachia alone or together with Spiroplasma endosymbionts in D. melanogaster adult flies affects the immune response against the virulent insect pathogen Photorhabdus luminescens and against non-pathogenic Escherichia coli bacteria. We found that D. melanogaster flies carrying no endosymbionts, those carrying both Wolbachia and Spiroplasma, and those containing Wolbachia only had similar survival rates after infection with P. luminescens or Escherichia coli bacteria. However, flies carrying both endosymbionts or Wolbachia only contained higher numbers of E. coli cells at early time-points post infection than flies without endosymbiotic bacteria. Interestingly, flies containing Wolbachia only had lower titers of this endosymbiont upon infection with the pathogen P. luminescens than uninfected flies of the same strain. We further found that the presence of Wolbachia and Spiroplasma in D. melanogaster up-regulated certain immune-related genes upon infection with P. luminescens or E. coli bacteria, but it failed to alter the phagocytic ability of the flies toward E. coli inactive bioparticles. Our results suggest that the presence of Wolbachia and Spiroplasma in D. melanogaster can modulate immune signaling against infection by certain insect pathogenic and non-pathogenic bacteria. Results from such studies are important for understanding the molecular basis of the interactions between endosymbiotic bacteria of insects and exogenous microbes.

Probiotic E. coli Nissle 1917 biofilms on silicone substrates for bacterial interference against pathogen colonization.

PubMed

Chen, Quan; Zhu, Zhiling; Wang, Jun; Lopez, Analette I; Li, Siheng; Kumar, Amit; Yu, Fei; Chen, Haoqing; Cai, Chengzhi; Zhang, Lijuan

2017-03-01

Bacterial interference is an alternative strategy to fight against device-associated bacterial infections. Pursuing this strategy, a non-pathogenic bacterial biofilm is used as a live, protective barrier to fence off pathogen colonization. In this work, biofilms formed by probiotic Escherichia coli strain Nissle 1917 (EcN) are investigated for their potential for long-term bacterial interference against infections associated with silicone-based urinary catheters and indwelling catheters used in the digestive system, such as feeding tubes and voice prostheses. We have shown that EcN can form stable biofilms on silicone substrates, particularly those modified with a biphenyl mannoside derivative. These biofilms greatly reduced the colonization by pathogenic Enterococcus faecalis in Lysogeny broth (LB) for 11days. Bacterial interference is an alternative strategy to fight against device-associated bacterial infections. Pursuing this strategy, we use non-pathogenic bacteria to form a biofilm that serves as a live, protective barrier against pathogen colonization. Herein, we report the first use of preformed probiotic E. coli Nissle 1917 biofilms on the mannoside-presenting silicone substrates to prevent pathogen colonization. The biofilms serve as a live, protective barrier to fence off the pathogens, whereas current antimicrobial/antifouling coatings are subjected to gradual coverage by the biomass from the rapidly growing pathogens in a high-nutrient environment. It should be noted that E. coli Nissle 1917 is commercially available and has been used in many clinical trials. We also demonstrated that this probiotic strain performed significantly better than the non-commercial, genetically modified E. coli strain that we previously reported. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Fecal-indicator bacteria in surface waters of the Santee River Basin and coastal drainages, North and South Carolina, 1995-98

USGS Publications Warehouse

Wilhelm, Lance J.; Maluk, Terry L.

1998-01-01

IntroductionHigh levels of fecal-indicator bacteria in rivers and streams can indicate the possible presence of pathogenic (disease-causing) microorganisms. Cholera, typhoid fever, bacterial dysentery, infectious hepatitis, and cryptosporidiosis are some of the well known waterborne diseases that spread through water contaminated and fecal matter. Eye, ear, nose, and throat infections also can result from contact with contaminated water. In general, methods are not routinely used to detect pathogens in water. Instead, bacteria such as total coliforms, fecal coliforms, fecal streptococci, Escherichia coli (E coli), and enterococci are used as indicators of sanitary water quality, because they are present in high numbers in fecal material and have been shown to be associated with some waterborne disease-causing organisms. Indicator bacteria usually are harmless, more plentiful, and easier to detect than pathogens. The concentration of bacteria in a sample of water is usually expressed as the number of bacterial colonies per 100 milliliters of water sample.As part of the U.S. Geological Survey National Water-Quality Assessment Program, 145 samples were collected and analyzed for selected water-quality constituents, fecal coliforms, and fecal streptococci at 17 sites in North and South Carolina from October 1995 through September 1996. Of the original 17 sites, 4 in South Carolina were sampled for E. coli and total coliforms from April through September 1997. At two sites, this sampling continued from October 1997 through April 1998.

The effects of upaB deletion and the double/triple deletion of upaB, aatA, and aatB genes on pathogenicity of avian pathogenic Escherichia coli.

PubMed

Zhu-Ge, Xiang-Kai; Pan, Zi-Hao; Tang, Fang; Mao, Xiang; Hu, Lin; Wang, Shao-Hui; Xu, Bin; Lu, Cheng-Ping; Fan, Hong-Jie; Dai, Jian-Jun

2015-12-01

Autotransporters (ATs) are associated with pathogenesis of Avian Pathogenic Escherichia coli (APEC). The molecular characterization of APEC ATs can provide insights about their relevance to APEC pathogenesis. Here, we characterized a conventional autotransporter UpaB in APEC DE205B genome. The upaB existed in 41.9 % of 236 APEC isolates and was predominantly associated with ECOR B2 and D. Our studies showed that UpaB mediates the DE205B adhesion in DF-1 cells, and enhances autoaggregation and biofilm formation of fimbria-negative E. coli AAEC189 (MG1655Δfim) in vitro. Deletion of upaB of DE205B attenuates the virulence in duck model and early colonization in the duck lungs during APEC systemic infection. Furthermore, double and triple deletion of upaB, aatA, and aatB genes cumulatively attenuated DE205B adhesion in DF-1 cells, accompanying with decreased 50 % lethal dose (LD50) in duck model and the early colonization in the duck lungs. However, DE205BΔupaB/ΔaatA/ΔaatB might "compensate" the influence of gene deletion by upregulating the expression of fimbrial adhesin genes yqiL, yadN, and vacuolating autotransporter vat during early colonization of APEC. Finally, we demonstrated that vaccination with recombinant UpaB, AatA, and AatB proteins conferred protection against colisepticemia caused by DE205B infection in duck model.

Raman spectroscopy for the microbiological characterization and identification of medically relevant bacteria

NASA Astrophysics Data System (ADS)

Hamasha, Khozima Mahmoud

The detection and identification of pathogenic bacteria has become more important than ever due to the increase of potential bioterrorism threats and the high mortality rate of bacterial infections worldwide. Raman spectroscopy has recently gained popularity as an attractive robust approach for the molecular characterization, rapid identification, and accurate classification of a wide range of bacteria. In this dissertation, Raman spectroscopy utilizing advanced statistical techniques was used to identify and discriminate between different pathogenic and non-pathogenic bacterial strains of E. coli and Staphylococcus aureus bacterial species by probing the molecular compositions of the cells. The five-carbon sugar xylitol, which cannot be metabolized by the oral and nasopharyngeal bacteria, had been recognized by clinicians as a preventive agents for dental caries and many studies have demonstrated that xylitol causes a reduction in otitis media (chronic inner ear infections) and other nasopharyngeal infections. Raman spectroscopy was used to characterize the uptake and metabolic activity of xylitol in pathogenic (viridans group Streptococcus) and nonpathogenic (E. coli) bacteria by taking their Raman spectra before xylitol exposure and after growing with xylitol and quantifying the significant differences in the molecular vibrational modes due to this exposure. The results of this study showed significant stable spectral changes in the S. viridians bacteria induced by xylitol and those changes were not the same as in some E. coli strains. Finally, Raman spectroscopy experiments were conducted to provide important information about the function of a certain protein (wag31) of Mycobacterium tuberculosis using a relative non-pathogenic bacterium called Mycobacterium smegmatis. Raman spectra of conditional mutants of bacteria expressing three different phosphorylation forms of wag31 were collected and analyzed. The results show that that the phosphorylation of wag31 causes significant differences in the molecular structure, namely the quantity of amino acids associated with peptidoglycan precursor proteins and lipid II as observed in the Raman spectra of these cells. Raman spectra were also acquired from the isolated cell envelope fraction of the cells expressing different forms of wag31 and the results showed that a significant number of the molecular vibrational differences observed in the cells were also observed in the cell envelope fraction, indicating that these differences are localized in the cell envelope.

Role of the SRRz/Rz1 lambdoid lysis cassette in the pathoadaptive evolution of Shigella.

PubMed

Leuzzi, Adriano; Grossi, Milena; Di Martino, Maria Letizia; Pasqua, Martina; Micheli, Gioacchino; Colonna, Bianca; Prosseda, Gianni

2017-06-01

Shigella, the etiological agent of bacillary dysentery (shigellosis), is a highly adapted human pathogen. It evolved from an innocuous ancestor resembling the Escherichia coli strain by gain and loss of genes and functions. While the gain process concerns the acquisition of the genetic determinants of virulence, the loss is related to the adaptation of the genome to the new pathogenic status and occurs by pathoadaptive mutation of antivirulence genes. In this study, we highlight that the SRRz/Rz 1 lambdoid lysis cassette, even though stably adopted in E. coli K12 by virtue of its beneficial effect on cell physiology, has undergone a significant decay in Shigella. Moreover, we show the antivirulence nature of the SRRz/Rz 1 lysis cassette in Shigella. In fact, by restoring the SRRz/Rz 1 expression in this pathogen, we observe an increased release of peptidoglycan fragments, causing an unbalance in the fine control exerted by Shigella on host innate immunity and a mitigation of its virulence. This strongly affects the virulence of Shigella and allows to consider the loss of SRRz/Rz 1 lysis cassette as another pathoadaptive event in the life of Shigella. Copyright © 2017 Elsevier GmbH. All rights reserved.

A long-term epigenetic memory switch controls bacterial virulence bimodality

PubMed Central

Ronin, Irine; Katsowich, Naama; Rosenshine, Ilan; Balaban, Nathalie Q

2017-01-01

When pathogens enter the host, sensing of environmental cues activates the expression of virulence genes. Opposite transition of pathogens from activating to non-activating conditions is poorly understood. Interestingly, variability in the expression of virulence genes upon infection enhances colonization. In order to systematically detect the role of phenotypic variability in enteropathogenic E. coli (EPEC), an important human pathogen, both in virulence activating and non-activating conditions, we employed the ScanLag methodology. The analysis revealed a bimodal growth rate. Mathematical modeling combined with experimental analysis showed that this bimodality is mediated by a hysteretic memory-switch that results in the stable co-existence of non-virulent and hyper-virulent subpopulations, even after many generations of growth in non-activating conditions. We identified the per operon as the key component of the hysteretic switch. This unique hysteretic memory switch may result in persistent infection and enhanced host-to-host spreading. DOI: http://dx.doi.org/10.7554/eLife.19599.001 PMID:28178445

The Intriguing Evolutionary Journey of Enteroinvasive E. coli (EIEC) toward Pathogenicity

PubMed Central

Pasqua, Martina; Michelacci, Valeria; Di Martino, Maria Letizia; Tozzoli, Rosangela; Grossi, Milena; Colonna, Bianca; Morabito, Stefano; Prosseda, Gianni

2017-01-01

Among the intestinal pathogenic Escherichia coli, enteroinvasive E. coli (EIEC) are a group of intracellular pathogens able to enter epithelial cells of colon, multiplicate within them, and move between adjacent cells with a mechanism similar to Shigella, the ethiological agent of bacillary dysentery. Despite EIEC belong to the same pathotype of Shigella, they neither have the full set of traits that define Shigella nor have undergone the extensive gene decay observed in Shigella. Molecular analysis confirms that EIEC are widely distributed among E. coli phylogenetic groups and correspond to bioserotypes found in many E. coli serogroups. Like Shigella, also in EIEC the critical event toward a pathogenic life-style consisted in the acquisition by horizontal gene transfer of a large F-type plasmid (pINV) containing the genes required for invasion, intracellular survival, and spreading through the intestinal mucosa. In Shigella, the ample gain in virulence determinants has been counteracted by a substantial loss of functions that, although important for the survival in the environment, are redundant or deleterious for the life inside the host. The pathoadaptation process that has led Shigella to modify its metabolic profile and increase its pathogenic potential is still in infancy in EIEC, although maintenance of some features typical of E. coli might favor their emerging relevance as intestinal pathogens worldwide, as documented by recent outbreaks in industrialized countries. In this review, we will discuss the evolution of EIEC toward Shigella-like invasive forms going through the epidemiology, including the emergence of new virulent strains, their genome organization, and the complex interactions they establish with the host. PMID:29259590

Graphene-interfaced electrical biosensor for label-free and sensitive detection of foodborne pathogenic E. coli O157:H7.

PubMed

Pandey, Ashish; Gurbuz, Yasar; Ozguz, Volkan; Niazi, Javed H; Qureshi, Anjum

2017-05-15

E. coli O157:H7 is an enterohemorrhagic bacteria responsible for serious foodborne outbreaks that causes diarrhoea, fever and vomiting in humans. Recent foodborne E. coli outbreaks has left a serious concern to public health. Therefore, there is an increasing demand for a simple, rapid and sensitive method for pathogen detection in contaminated foods. In this study, we developed a label-free electrical biosensor interfaced with graphene for sensitive detection of pathogenic bacteria. This biosensor was fabricated by interfacing graphene with interdigitated microelectrodes of capacitors that were biofunctionalized with E. coli O157:H7 specific antibodies for sensitive pathogenic bacteria detection. Here, graphene nanostructures on the sensor surface provided superior chemical properties such as high carrier mobility and biocompatibility with antibodies and bacteria. The sensors transduced the signal based on changes in dielectric properties (capacitance) through (i) polarization of captured cell-surface charges, (ii) cells' internal bioactivity, (iii) cell-wall's electronegativity or dipole moment and their relaxation and (iv) charge carrier mobility of graphene that modulated the electrical properties once the pathogenic E. coli O157:H7 captured on the sensor surface. Sensitive capacitance changes thus observed with graphene based capacitors were specific to E. coli O157:H7 strain with a sensitivity as low as 10-100 cells/ml. The proposed graphene based electrical biosensor provided advantages of speed, sensitivity, specificity and in-situ bacterial detection with no chemical mediators, represents a versatile approach for detection of a wide variety of other pathogens. Copyright © 2016 Elsevier B.V. All rights reserved.

The Intriguing Evolutionary Journey of Enteroinvasive E. coli (EIEC) toward Pathogenicity.

PubMed

Pasqua, Martina; Michelacci, Valeria; Di Martino, Maria Letizia; Tozzoli, Rosangela; Grossi, Milena; Colonna, Bianca; Morabito, Stefano; Prosseda, Gianni

2017-01-01

Among the intestinal pathogenic Escherichia coli , enteroinvasive E. coli (EIEC) are a group of intracellular pathogens able to enter epithelial cells of colon, multiplicate within them, and move between adjacent cells with a mechanism similar to Shigella , the ethiological agent of bacillary dysentery. Despite EIEC belong to the same pathotype of Shigella , they neither have the full set of traits that define Shigella nor have undergone the extensive gene decay observed in Shigella . Molecular analysis confirms that EIEC are widely distributed among E. coli phylogenetic groups and correspond to bioserotypes found in many E. coli serogroups. Like Shigella , also in EIEC the critical event toward a pathogenic life-style consisted in the acquisition by horizontal gene transfer of a large F-type plasmid (pINV) containing the genes required for invasion, intracellular survival, and spreading through the intestinal mucosa. In Shigella , the ample gain in virulence determinants has been counteracted by a substantial loss of functions that, although important for the survival in the environment, are redundant or deleterious for the life inside the host. The pathoadaptation process that has led Shigella to modify its metabolic profile and increase its pathogenic potential is still in infancy in EIEC, although maintenance of some features typical of E. coli might favor their emerging relevance as intestinal pathogens worldwide, as documented by recent outbreaks in industrialized countries. In this review, we will discuss the evolution of EIEC toward Shigella -like invasive forms going through the epidemiology, including the emergence of new virulent strains, their genome organization, and the complex interactions they establish with the host.

Antibacterial activity and mechanism of action of ε-poly-L-lysine

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

Ye, Ruosong; Xu, Hengyi; Wan, Cuixiang

Highlights: •Antibacterial activity and mechanism of ε-PL against E. coli O157:H7 was investigated. •Critical inhibitory factors toward the growth of E. coli O157:H7 by ε-PL was analyzed. •Cell membrane integrity and cell morphology of E. coli O157:H7 was affected by ε-PL. •A positive correlation between reactive oxygen species levels and ε-PL concentration in E. coli O157:H7 cells. •ε-PL induced the expression of different genes related to oxidative/redox stress, SOS response, virulence. -- Abstract: ε-Poly-L-lysine (ε-PL) is widely used as an antibacterial agent because of its broad antimicrobial spectrum. However, the mechanism of ε-PL against pathogens at the molecular level hasmore » not been elucidated. This study investigated the antibacterial activity and mechanism of ε-PL against Escherichia coli O157:H7 CMCC44828. Propidium monoazide-PCR test results indicated that the threshold condition of ε-PL for complete membrane lysis of E. coli O157:H7 was 10 μg/mL (90% mortality for 5 μg/mL). Further verification of the destructive effect of ε-PL on cell structure was performed by atomic force microscopy and transmission electron microscopy. Results showed a positive correlation between reactive oxygen species (ROS) levels and ε-PL concentration in E. coli O157:H7 cells. Moreover, the mortality of E. coli O157:H7 was reduced when antioxidant N-acetylcysteine was added. Results from real-time quantitative PCR (RT-qPCR) indicated that the expression levels of oxidative stress genes sodA and oxyR were up-regulated 4- and 16-fold, respectively, whereas virulence genes eaeA and espA were down-regulated after ε-PL treatment. Expression of DNA damage response (SOS response) regulon genes recA and lexA were also affected by ε-PL. In conclusion, the antibacterial mechanism of ε-PL against E. coli O157:H7 may be attributed to disturbance on membrane integrity, oxidative stress by ROS, and effects on various gene expressions, such as regulation of oxidative stress, SOS response, and changes in virulence.« less

Coating of silicone with mannoside-PAMAM dendrimers to enhance formation of non-pathogenic Escherichia coli biofilms against colonization of uropathogens.

PubMed

Zhu, Zhiling; Yu, Fei; Chen, Haoqing; Wang, Jun; Lopez, Analette I; Chen, Quan; Li, Siheng; Long, Yuyu; Darouiche, Rabih O; Hull, Richard A; Zhang, Lijuan; Cai, Chengzhi

2017-12-01

Bacterial interference using non-pathogenic Escherichia coli 83972 is a novel strategy for preventing catheter-associated urinary tract infection (CAUTI). Crucial to the success of this strategy is to establish a high coverage and stable biofilm of the non-pathogenic bacteria on the catheter surface. However, this non-pathogenic strain is sluggish to form biofilms on silicone as the most widely used material for urinary catheters. We have addressed this issue by modifying the silicone catheter surfaces with mannosides that promote the biofilm formation, but the stability of the non-pathogenic biofilms challenged by uropathogens over long-term remains a concern. Herein, we report our study on the stability of the non-pathogenic biofilms grown on propynylphenyl mannoside-modified silicone. The result shows that 94% non-pathogenic bacteria were retained on the modified silicone under >0.5 Pa shear stress. After being challenged by three multidrug-resistant uropathogenic isolates in artificial urine for 11 days, large amounts (>4 × 10 6  CFU cm -2 ) of the non-pathogenic bacteria remained on the surfaces. These non-pathogenic biofilms reduced the colonization of the uropathogens by >3.2-log. In bacterial interference, the non-pathogenic Escherichia coli strains are sluggish to form biofilms on the catheter surfaces, due to rapid removal by urine flow. We have demonstrated a solution to this bottleneck by pre-functionalization of mannosides on the silicone surfaces to promote E. coli biofilm formation. A pre-conjugated high affinity propynylphenyl mannoside ligand tethered to the nanometric amino-terminated poly(amido amine) (PAMAM) dendrimer is used for binding to a major E. coli adhesin FimH. It greatly improves the efficiency for the catheter modification, the non-pathogenic biofilm coverage, as well as the (long-term) stability for prevention of uropathogen infections. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Occurrence and antimicrobial resistance of pathogenic Escherichia coli and Salmonella spp. in retail raw table eggs sold for human consumption in Enugu state, Nigeria

PubMed Central

Okorie-Kanu, O. Josephine; Ezenduka, E. Vivienne; Okorie-Kanu, C. Onwuchokwe; Ugwu, L. Chinweokwu; Nnamani, U. John

2016-01-01

Aim: This study was conducted to investigate the occurrence of pathogenic Escherichia coli and Salmonella species in retail raw table eggs sold for human consumption in Enugu State and to determine the resistance of these pathogens to antimicrobials commonly used in human and veterinary practices in Nigeria. Materials and Methods: A total of 340 raw table eggs comprising 68 composite samples (5 eggs per composite sample) were collected from five selected farms (13 composite samples from the farms) and 10 retail outlets (55 composite samples from the retail outlets) in the study area over a period of 4-month (March-June, 2014). The eggs were screened for pathogenic E. coli and Salmonella species following standard procedures within 24 h of sample collection. Isolates obtained were subjected to in-vitro antimicrobial susceptibility test with 15 commonly used antimicrobials using the disk diffusion method. Results: About 37 (54.4%) and 7 (10.3%) of the 68 composite samples were positive for pathogenic E. coli and Salmonella species, respectively. The shells showed significantly higher (p<0.05) contaminations than the contents for both microorganisms. The eggs from the farms showed higher contamination with pathogenic E. coli than eggs from the retail outlets while the reverse was the case for Salmonella species even though they were not significant (p>0.05). The organisms obtained showed a multiple drug resistance. They were completely resistant to nitrofurantoin, sulfamethoxazole/trimethoprim, penicillin G and oxacillin. In addition to these, Salmonella spp. also showed 100% resistance to tetracycline. The pathogenic E. coli isolates obtained were 100% susceptible to gentamicin, neomycin, ciprofloxacin, and amoxicillin-clavulanic acid while Salmonella spp. showed 100% susceptibility to erythromycin, neomycin, and rifampicin. Both organisms showed varying degrees of resistance to streptomycin, amoxicillin, vancomycin, and doxycycline. Conclusion: From the results of the study, it can be concluded that the raw table eggs marketed for human consumption in Enugu State, Nigeria is contaminated with pathogenic E. coli and Salmonella species that showed multiple drug resistance to antimicrobial agents commonly used in veterinary and human practice. PMID:27956787

Occurrence and antimicrobial resistance of pathogenic Escherichia coli and Salmonella spp. in retail raw table eggs sold for human consumption in Enugu state, Nigeria.

PubMed

Okorie-Kanu, O Josephine; Ezenduka, E Vivienne; Okorie-Kanu, C Onwuchokwe; Ugwu, L Chinweokwu; Nnamani, U John

2016-11-01

This study was conducted to investigate the occurrence of pathogenic Escherichia coli and Salmonella species in retail raw table eggs sold for human consumption in Enugu State and to determine the resistance of these pathogens to antimicrobials commonly used in human and veterinary practices in Nigeria. A total of 340 raw table eggs comprising 68 composite samples (5 eggs per composite sample) were collected from five selected farms (13 composite samples from the farms) and 10 retail outlets (55 composite samples from the retail outlets) in the study area over a period of 4-month (March-June, 2014). The eggs were screened for pathogenic E. coli and Salmonella species following standard procedures within 24 h of sample collection. Isolates obtained were subjected to in-vitro antimicrobial susceptibility test with 15 commonly used antimicrobials using the disk diffusion method. About 37 (54.4%) and 7 (10.3%) of the 68 composite samples were positive for pathogenic E. coli and Salmonella species, respectively. The shells showed significantly higher (p<0.05) contaminations than the contents for both microorganisms. The eggs from the farms showed higher contamination with pathogenic E. coli than eggs from the retail outlets while the reverse was the case for Salmonella species even though they were not significant (p>0.05). The organisms obtained showed a multiple drug resistance. They were completely resistant to nitrofurantoin, sulfamethoxazole/trimethoprim, penicillin G and oxacillin. In addition to these, Salmonella spp. also showed 100% resistance to tetracycline. The pathogenic E. coli isolates obtained were 100% susceptible to gentamicin, neomycin, ciprofloxacin, and amoxicillin-clavulanic acid while Salmonella spp. showed 100% susceptibility to erythromycin, neomycin, and rifampicin. Both organisms showed varying degrees of resistance to streptomycin, amoxicillin, vancomycin, and doxycycline. From the results of the study, it can be concluded that the raw table eggs marketed for human consumption in Enugu State, Nigeria is contaminated with pathogenic E. coli and Salmonella species that showed multiple drug resistance to antimicrobial agents commonly used in veterinary and human practice.

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.

Medical and economic impact of extraintestinal infections due to Escherichia coli: focus on an increasingly important endemic problem.

PubMed

Russo, Thomas A; Johnson, James R

2003-04-01

Escherichia coli is probably the best-known bacterial species and one of the most frequently isolated organisms from clinical specimens. Despite this, underappreciation and misunderstandings exist among medical professionals and the lay public alike regarding E. coli as an extraintestinal pathogen. Underappreciated features include (i) the wide variety of extraintestinal infections E. coli can cause, (ii) the high incidence and associated morbidity, mortality, and costs of these diverse clinical syndromes, (iii) the pathogenic potential of different groups of E. coli strains for causing intestinal versus extraintestinal disease, and (iv) increasing antimicrobial resistance. In this era in which health news often sensationalizes uncommon infection syndromes or pathogens, the strains of E. coli that cause extraintestinal infection are an increasingly important endemic problem and underappreciated "killers". Billions of health care dollars, millions of work days, and hundreds of thousands of lives are lost each year to extraintestinal infections due to E. coli. New treatments and prevention measures will be needed for improved outcomes and a diminished disease burden.

[Haemorrhagic septicaemia in a pig caused by extraintestinal pathogenic Escherichia coli (ExPEC) as a differential diagnosis in classical swine fever--case report and review of the literature].

PubMed

Reiner, Gerald; von Berg, Stephan; Hillen, Sonja; Clemens, Nina; Huisinger, Maike; Burkhardt, Eberhard; Weiss, Reinhardt; Reinacher, Manfred

2010-01-01

Domestic pig herds in some regions of Germany are permanently threatened by Classical Swine Fever. In the case of suspicion, a series of infectious and non infectious causes has to be excluded. The present paper describes a case of Escherichia coli septicaemia, with clinical and pathological symptoms that could not be differentiated from European or African Swine Fever. The E. coli strain could not be classified by standard serotyping. Virulence factors common for ETEC (enterotoxic E. coli) or EDEC (edema-disease E. coli) were not detected. Instead, we found P-fimbriae and aerobactin, thus characterising this strain as an extraintestinal pathogenic strain. Such strains have sporadicly been reported as the cause of septicaemia in piglets or weaners, but the present case is the first report of an E. coli-associated septicaemia in an adult pig. This case shows that extraintestinal pathogenic E. coli can be the cause of severe septicaemia and haemorrhagia. They thus have to be considered as a further differential diagnosis in swine fever.

Escherichia coli and selected veterinary and zoonotic pathogens isolated from environmental sites in companion animal veterinary hospitals in southern Ontario

PubMed Central

Murphy, Colleen P.; Reid-Smith, Richard J.; Boerlin, Patrick; Weese, J. Scott; Prescott, John F.; Janecko, Nicol; Hassard, Lori; McEwen, Scott A.

2010-01-01

Hospital-based infection control in veterinary medicine is emerging and the role of the environment in hospital-acquired infections (HAI) in veterinary hospitals is largely unknown. This study was initiated to determine the recovery of Escherichia coli and selected veterinary and zoonotic pathogens from the environments of 101 community veterinary hospitals. The proportion of hospitals with positive environmental swabs were: E. coli — 92%, Clostridium difficile — 58%, methicillin-resistant Staphylococcus aureus (MRSA) — 9%, CMY-2 producing E. coli — 9%, methicillin-resistant Staphylococcus pseudintermedius — 7%, and Salmonella — 2%. Vancomycin-resistant Enterococcus spp., canine parvovirus, and feline calicivirus were not isolated. Prevalence of antimicrobial resistance in E. coli isolates was low. Important potential veterinary and human pathogens were recovered including Canadian epidemic strains MRSA-2 and MRSA-5, and C. difficile ribotype 027. There is an environmental reservoir of pathogens in veterinary hospitals; therefore, additional studies are required to characterize risk factors associated with HAI in companion animals, including the role of the environment. PMID:21119862

Toxin-Antitoxin Systems Are Important for Niche-Specific Colonization and Stress Resistance of Uropathogenic Escherichia coli

PubMed Central

Norton, J. Paul; Mulvey, Matthew A.

2012-01-01

Toxin-antitoxin (TA) systems are prevalent in many bacterial genomes and have been implicated in biofilm and persister cell formation, but the contribution of individual chromosomally encoded TA systems during bacterial pathogenesis is not well understood. Of the known TA systems encoded by Escherichia coli, only a subset is associated with strains of extraintestinal pathogenic E. coli (ExPEC). These pathogens colonize diverse niches and are a major cause of sepsis, meningitis, and urinary tract infections. Using a murine infection model, we show that two TA systems (YefM-YoeB and YbaJ-Hha) independently promote colonization of the bladder by the reference uropathogenic ExPEC isolate CFT073, while a third TA system comprised of the toxin PasT and the antitoxin PasI is critical to ExPEC survival within the kidneys. The PasTI TA system also enhances ExPEC persister cell formation in the presence of antibiotics and markedly increases pathogen resistance to nutrient limitation as well as oxidative and nitrosative stresses. On its own, low-level expression of PasT protects ExPEC from these stresses, whereas overexpression of PasT is toxic and causes bacterial stasis. PasT-induced stasis can be rescued by overexpression of PasI, indicating that PasTI is a bona fide TA system. By mutagenesis, we find that the stress resistance and toxic effects of PasT can be uncoupled and mapped to distinct domains. Toxicity was specifically linked to sequences within the N-terminus of PasT, a region that also promotes the development of persister cells. These results indicate discrete, multipurpose functions for a TA-associated toxin and demonstrate that individual TA systems can provide bacteria with pronounced fitness advantages dependent on toxin expression levels and the specific environmental niche occupied. PMID:23055930

Fate of naturally occurring Escherichia coli O157:H7 and other zoonotic pathogens during minimally managed bovine feedlot manure composting processes

USDA-ARS?s Scientific Manuscript database

Reducing Escherichia coli O157:H7 in livestock manures before application to cropland is critical for reducing the risk of foodborne illness associated with produce. Our objective was to determine the fate of naturally occurring E. coli O157:H7 and other pathogens during minimally managed on-farm bo...

Complete Genome Sequence of Enteroinvasive Escherichia coli O96:H19 Associated with a Severe Foodborne Outbreak

PubMed Central

Pettengill, Emily A.; Hoffmann, Maria; Roberts, Richard J.; Payne, Justin; Allard, Marc; Michelacci, Valeria; Minelli, Fabio; Morabito, Stefano

2015-01-01

We present here the complete genome sequence of a strain of enteroinvasive Escherichia coli O96:H19 from a severe foodborne outbreak in a canteen in Italy in 2014. The complete genome may provide important information about the acquired pathogenicity of this strain and the transition between commensal and pathogenic E. coli. PMID:26251502

Genomic and Phenomic Study of Mammary Pathogenic Escherichia coli

PubMed Central

Blum, Shlomo E.; Heller, Elimelech D.; Sela, Shlomo; Elad, Daniel; Edery, Nir; Leitner, Gabriel

2015-01-01

Escherichia coli is a major etiological agent of intra-mammary infections (IMI) in cows, leading to acute mastitis and causing great economic losses in dairy production worldwide. Particular strains cause persistent IMI, leading to recurrent mastitis. Virulence factors of mammary pathogenic E. coli (MPEC) involved pathogenesis of mastitis as well as those differentiating strains causing acute or persistent mastitis are largely unknown. This study aimed to identify virulence markers in MPEC through whole genome and phenome comparative analysis. MPEC strains causing acute (VL2874 and P4) or persistent (VL2732) mastitis were compared to an environmental strain (K71) and to the genomes of strains representing different E. coli pathotypes. Intra-mammary challenge in mice confirmed experimentally that the strains studied here have different pathogenic potential, and that the environmental strain K71 is non-pathogenic in the mammary gland. Analysis of whole genome sequences and predicted proteomes revealed high similarity among MPEC, whereas MPEC significantly differed from the non-mammary pathogenic strain K71, and from E. coli genomes from other pathotypes. Functional features identified in MPEC genomes and lacking in the non-mammary pathogenic strain were associated with synthesis of lipopolysaccharide and other membrane antigens, ferric-dicitrate iron acquisition and sugars metabolism. Features associated with cytotoxicity or intra-cellular survival were found specifically in the genomes of strains from severe and acute (VL2874) or persistent (VL2732) mastitis, respectively. MPEC genomes were relatively similar to strain K-12, which was subsequently shown here to be possibly pathogenic in the mammary gland. Phenome analysis showed that the persistent MPEC was the most versatile in terms of nutrients metabolized and acute MPEC the least. Among phenotypes unique to MPEC compared to the non-mammary pathogenic strain were uric acid and D-serine metabolism. This study reveals virulence factors and phenotypic characteristics of MPEC that may play a role in pathogenesis of E. coli mastitis. PMID:26327312

Growth and survival of foodborne pathogens in beer.

PubMed

Menz, Garry; Aldred, Peter; Vriesekoop, Frank

2011-10-01

This work aimed to assess the growth and survival of four foodborne pathogens (Escherichia coli O157:H7, Salmonella Typhimurium, Listeria monocytogenes, and Staphylococcus aureus) in beer. The effects of ethanol, pH, and storage temperature were investigated for the gram-negative pathogens (E. coli O157:H7 and Salmonella Typhimurium), whereas the presence of hops ensured that the gram-positive pathogens (L. monocytogenes and S. aureus) were rapidly inactivated in alcohol-free beer. The pathogens E. coli O157:H7 and Salmonella Typhimurium could not grow in the mid-strength or full-strength beers, although they could survive for more than 30 days in the mid-strength beer when held at 4°C. These pathogens grew rapidly in the alcohol-free beer; however, growth was prevented when the pH of the alcohol-free beer was lowered from the "as received" value of 4.3 to 4.0. Pathogen survival in all beers was prolonged at lowered storage temperatures.

Zoonotic Potential of Escherichia coli Isolates from Retail Chicken Meat Products and Eggs

PubMed Central

Mitchell, Natalie M.; Johnson, James R.; Johnston, Brian; Curtiss, Roy

2014-01-01

Chicken products are suspected as a source of extraintestinal pathogenic Escherichia coli (ExPEC), which causes diseases in humans. The zoonotic risk to humans from chicken-source E. coli is not fully elucidated. To clarify the zoonotic risk posed by ExPEC in chicken products and to fill existing knowledge gaps regarding ExPEC zoonosis, we evaluated the prevalence of ExPEC on shell eggs and compared virulence-associated phenotypes between ExPEC and non-ExPEC isolates from both chicken meat and eggs. The prevalence of ExPEC among egg-source isolates was low, i.e., 5/108 (4.7%). Based on combined genotypic and phenotypic screening results, multiple human and avian pathotypes were represented among the chicken-source ExPEC isolates, including avian-pathogenic E. coli (APEC), uropathogenic E. coli (UPEC), neonatal meningitis E. coli (NMEC), and sepsis-associated E. coli (SEPEC), as well as an undefined ExPEC group, which included isolates with fewer virulence factors than the APEC, UPEC, and NMEC isolates. These findings document a substantial prevalence of human-pathogenic ExPEC-associated genes and phenotypes among E. coli isolates from retail chicken products and identify key virulence traits that could be used for screening. PMID:25480753

Age-related susceptibility to infection with diarrheagenic E. coli in infants from peri-urban areas of Lima, Peru

PubMed Central

Ochoa, Theresa J.; Ecker, Lucie; Barletta, Francesca; Mispireta, Mónica L.; Gil, Ana I.; Contreras, Carmen; Molina, Margarita; Amemiya, Isabel; Verastegui, Hector; Hall, Eric R.; Cleary, Thomas G.; Lanata, Claudio F.

2009-01-01

Background Diarrheagenic E. coli are being recognized as important pediatric enteropathogens worldwide. However, it is unclear whether there are differences in age-related susceptibility to specific agents, especially among infants. Methods We conducted a passive surveillance diarrhea cohort study of 1034 children from 2 to 12 months of age in Lima, Perú. Control stool samples were collected from randomly selected children without diarrhea. All samples were analyzed for common enteric pathogens and for the diarrheagenic E. coli by a multiplex real-time PCR. Results The most commonly isolated pathogens from 1065 diarrheal episodes were the diarrheagenic E. coli (31%), including enteroaggregative (15.1%) and enteropathogenic E. coli (EPEC) (7.6%). Diarrheagenic E. coli, Campylobacter and rotavirus were more frequently isolated from infants ≥ 6m. Diffusely adherent E. coli and enterotoxigenic E. coli (ETEC) were more frequently isolated in diarrheal samples than in controls in older infants (p<0.05). Children ≥ 6m infected with ETEC had a 4.56-fold increased risk for diarrhea (95% CI, 1.20 to 17.28). Persistent diarrhea was more frequent in infants < 6m (13.5% vs. 3.6%, p<0.001). Among diarrheagenic E. coli positive samples, co-infections with other pathogens were more common in diarrhea than in controls (40.1% vs. 15.6%, p<0.001). Conclusions Diarrheagenic E. coli were more frequently isolated in older infants. In this setting with high frequency of pathogen exposure and high frequency of breastfeeding, we hypothesize that the major age-related differences result from decreased exposure to milk protective factors and with increased exposure to contaminated food and water. PMID:19857163

[Pathogenic characteristics of Escherichia coli strains in cases of asymptomatic bacteriuria and symptomatic urinary tract infections].

PubMed

Misiewicz, I A; Galiński, J

1989-01-01

The aim of this study was to examine if E. coli isolated from asymptomatic bacteriuria differed in pathogenic features from strains isolated from symptomatic infections of urinary tract. In this study 130 strains of E. coli isolated from women having asymptomatic bacteriuria and 112 strains isolated from patients with symptoms of urinary tract infection were examined. It was shown that E. coli isolated from patients with symptomatic urinary tract infection showed the more frequently ability to cause mannose-resistant haemagglutination of human erythrocytes, resistance to bactericidal activity of serum and haemolytic properties than those isolated from asymptomatic bacteriuria. These strains showed also the higher ability to adhere to Vero cells in tissue culture. Among E. coli strains isolated from persons with asymptomatic bacteriuria the pathogenic features were most frequently found in strains from healthy women and the most rarely in isolated from diabetic women.

A Novel Protective Vaccine Antigen from the Core Escherichia coli Genome

PubMed Central

Moriel, Danilo G.; Tan, Lendl; Goh, Kelvin G. K.; Ipe, Deepak S.; Lo, Alvin W.; Peters, Kate M.

2016-01-01

ABSTRACT Escherichia coli is a versatile pathogen capable of causing intestinal and extraintestinal infections that result in a huge burden of global human disease. The diversity of E. coli is reflected by its multiple different pathotypes and mosaic genome composition. E. coli strains are also a major driver of antibiotic resistance, emphasizing the urgent need for new treatment and prevention measures. Here, we used a large data set comprising 1,700 draft and complete genomes to define the core and accessory genome of E. coli and demonstrated the overlapping relationship between strains from different pathotypes. In combination with proteomic investigation, this analysis revealed core genes that encode surface-exposed or secreted proteins that represent potential broad-coverage vaccine antigens. One of these antigens, YncE, was characterized as a conserved immunogenic antigen able to protect against acute systemic infection in mice after vaccination. Overall, this work provides a genomic blueprint for future analyses of conserved and accessory E. coli genes. The work also identified YncE as a novel antigen that could be exploited in the development of a vaccine against all pathogenic E. coli strains—an important direction given the high global incidence of infections caused by multidrug-resistant strains for which there are few effective antibiotics. IMPORTANCE E. coli is a multifaceted pathogen of major significance to global human health and an important contributor to increasing antibiotic resistance. Given the paucity of therapies still effective against multidrug-resistant pathogenic E. coli strains, novel treatment and prevention strategies are urgently required. In this study, we defined the core and accessory components of the E. coli genome by examining a large collection of draft and completely sequenced strains available from public databases. This data set was mined by employing a reverse-vaccinology approach in combination with proteomics to identify putative broadly protective vaccine antigens. One such antigen was identified that was highly immunogenic and induced protection in a mouse model of bacteremia. Overall, our study provides a genomic and proteomic framework for the selection of novel vaccine antigens that could mediate broad protection against pathogenic E. coli. PMID:27904885

Construction and expression of immunogenic hybrid enterotoxigenic Escherichia coli CFA/I and CS2 colonization fimbriae for use in vaccines.

PubMed

Tobias, Joshua; Svennerholm, Ann-Mari; Holmgren, Jan; Lebens, Michael

2010-07-01

Enterotoxigenic Escherichia coli (ETEC) are an important cause of diarrheal morbidity in developing countries, especially in children and also of traveler's diarrhea. Colonization factors (CFs) of ETEC, like CFA/I and CS2 which are genetically and structurally related, play a substantial role in pathogenicity, and since intestinal-mucosal immune responses against CFs appear to be protective, much effort has focused on the development of a CF-based ETEC vaccine. We have constructed hybrid operons in which the major CS2 subunit-encoding cotA gene was inserted into the CFA/I operon, either replacing (hybrid I) or being added to the major CFA/I subunit-encoding cfaB gene (hybrid II). Using specific monoclonal antibodies against the major subunits of CFA/I and CS2, high levels of surface expression of both fimbrial subunits were shown in E. coli carrying the hybrid II operon. Oral immunization of mice with formalin-killed bacteria expressing hybrid II fimbriae induced strong CFA/I- and CS2-specific serum IgG + IgM and fecal IgA antibody responses, which were higher than those achieved by similar immunization with the reference strains. Bacteria expressing hybrid fimbriae are potential candidate strains in an oral-killed CF-ETEC vaccine, and the approach represents an attractive and novel means of producing a broad-spectrum ETEC vaccine.

Gene expression profiling of Escherichia coli in response to interactions with the lettuce rhizosphere.

PubMed

Hou, Z; Fink, R C; Black, E P; Sugawara, M; Zhang, Z; Diez-Gonzalez, F; Sadowsky, M J

2012-11-01

The objective of this study was to examine transcriptional changes in Escherichia coli when the bacterium was growing in the lettuce rhizoshpere. A combination of microarray analyses, colonization assays and confocal microscopy was used to gain a more complete understanding of bacterial genes involved in the colonization and growth of E. coli K12 in the lettuce root rhizosphere using a novel hydroponic assay system. After 3 days of interaction with lettuce roots, E. coli genes involved in protein synthesis, stress responses and attachment were up-regulated. Mutants in curli production (crl, csgA) and flagella synthesis (fliN) had a reduced capacity to attach to roots as determined by bacterial counts and by confocal laser scanning microscopy. This study indicates that E. coli K12 has the capability to colonize lettuce roots by using attachment genes and can readily adapt to the rhizosphere of lettuce plants. Results of this study show curli production and biofilm modulation genes are important for rhizosphere colonization and may provide useful targets to disrupt this process. Further studies using pathogenic strains will provide additional information about lettuce-E. coli interactions. © 2012 The Authors Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.

Abundance of Pathogenic Escherichia coli Virulence-Associated Genes in Well and Borehole Water Used for Domestic Purposes in a Peri-Urban Community of South Africa

PubMed Central

Abia, Akebe Luther King; Schaefer, Lisa; Ubomba-Jaswa, Eunice; Le Roux, Wouter

2017-01-01

In the absence of pipe-borne water, many people in Africa, especially in rural communities, depend on alternative water sources such as wells, boreholes and rivers for household and personal hygiene. Poor maintenance and nearby pit latrines, however, lead to microbial pollution of these sources. We evaluated the abundance of Escherichia coli and the prevalence of pathogenic E. coli virulence genes in water from wells, boreholes and a river in a South African peri-urban community. Monthly samples were collected between August 2015 and November 2016. In all, 144 water samples were analysed for E. coli using the Colilert 18 system. Virulence genes (eagg, eaeA, stx1, stx2, flichH7, ST, ipaH, ibeA) were investigated using real-time polymerase chain reaction. Mean E. coli counts ranged between 0 and 443.1 Most Probable Number (MPN)/100 mL of water sample. Overall, 99.3% of samples were positive for at least one virulence gene studied, with flicH7 being the most detected gene (81/140; 57.6%) and the stx2 gene the least detected gene (8/140; 5.7%). Both intestinal and extraintestinal pathogenic E. coli genes were detected. The detection of virulence genes in these water sources suggests the presence of potentially pathogenic E. coli strains and is a public health concern. PMID:28335539

Abundance of Pathogenic Escherichia coli Virulence-Associated Genes in Well and Borehole Water Used for Domestic Purposes in a Peri-Urban Community of South Africa.

PubMed

Abia, Akebe Luther King; Schaefer, Lisa; Ubomba-Jaswa, Eunice; Le Roux, Wouter

2017-03-20

In the absence of pipe-borne water, many people in Africa, especially in rural communities, depend on alternative water sources such as wells, boreholes and rivers for household and personal hygiene. Poor maintenance and nearby pit latrines, however, lead to microbial pollution of these sources. We evaluated the abundance of Escherichia coli and the prevalence of pathogenic E. coli virulence genes in water from wells, boreholes and a river in a South African peri-urban community. Monthly samples were collected between August 2015 and November 2016. In all, 144 water samples were analysed for E. coli using the Colilert 18 system. Virulence genes ( eagg , eaeA , stx1 , stx2 , flichH7 , ST , ipaH , ibeA ) were investigated using real-time polymerase chain reaction. Mean E. coli counts ranged between 0 and 443.1 Most Probable Number (MPN)/100 mL of water sample. Overall, 99.3% of samples were positive for at least one virulence gene studied, with flicH7 being the most detected gene (81/140; 57.6%) and the stx2 gene the least detected gene (8/140; 5.7%). Both intestinal and extraintestinal pathogenic E. coli genes were detected. The detection of virulence genes in these water sources suggests the presence of potentially pathogenic E. coli strains and is a public health concern.

Occurrence of Virulence Genes Associated with Diarrheagenic Escherichia coli Isolated from Raw Cow’s Milk from Two Commercial Dairy Farms in the Eastern Cape Province, South Africa

PubMed Central

Caine, Lesley-Anne; Nwodo, Uchechukwu U.; Okoh, Anthony I.; Ndip, Roland N.; Green, Ezekiel

2014-01-01

Escherichia coli remains a public health concern worldwide as an organism that causes diarrhea and its reservoir in raw milk may play an important role in the survival and transport of pathogenic strains. Diarrheagenic E. coli strains are diverse food-borne pathogens and causes diarrhea with varying virulence in humans. We investigated the prevalence of pathogenic E. coli in raw milk from two commercial dairy farms. Four hundred raw milk samples, 200 from each dairy farm, were screened for the presence of fliCH7, eagR, ial, eagg, lt, and papC genes. In dairy farm A, 100 E. coli were identified based on culture, oxidase and Gram staining, while 88 isolates from dairy farm B were identified in the same manner. Gene detection showed fliCH7 27 (54%) to be the highest gene detected from farm A and lt 2 (4%) to be the lowest. The highest gene detected in dairy farm B was fliCH7 16 (43.2%) and papC 1 (2.7%) was the least. The amplification of pathogenic genes associated with diarrheagenic E. coli from cows’ raw milk demonstrates that potentially virulent E. coli strains are widely distributed in raw milk and may be a cause of concern for human health. PMID:25411727

A new protocol to detect multiple foodborne pathogens with PCR dipstick DNA chromatography after a six-hour enrichment culture in a broad-range food pathogen enrichment broth.

PubMed

Hayashi, Masahiro; Natori, Tatsuya; Kubota-Hayashi, Sayoko; Miyata, Machiko; Ohkusu, Kiyofumi; Kawamoto, Keiko; Kurazono, Hisao; Makino, Souichi; Ezaki, Takayuki

2013-01-01

A quick foodborne pathogen screening method after six-hour enrichment culture with a broad-range food pathogen enrichment broth is described. Pathogenic factors of Salmonella enterica, Shigella spp., enteroinvasive Escherichia coli, and enterohemorrhagic E. coli are amplified with a cocktail primer and rapid polymerase chain reaction (PCR), which finishes amplification in 30 min. The PCR amplicon was differentiated with a dipstick DNA chromatography assay in 5-10 min. Starting from a four- to six-hour enrichment culture, this assay was finished within 45 min. Detection sensitivity of this protocol was less than 2.5 CFU/25 g for S. enterica and 3.3 CFU/25 g for enterohemorrhagic E. coli in spiked ground meat experiments.

Physiology and pathogenicity of cpdB deleted mutant of avian pathogenic Escherichia coli.

PubMed

Liu, Huifang; Chen, Liping; Si, Wei; Wang, Chunlai; Zhu, Fangna; Li, Guangxing; Liu, Siguo

2017-04-01

Avian colibacillosis is one of the most common infectious diseases caused partially or entirely by avian pathogenic Escherichia coli (APEC) in birds. In addition to spontaneous infection, APEC can also cause secondary infections that result in greater severity of illness and greater losses to the poultry industry. In order to assess the role of 2', 3'-cyclic phosphodiesterase (cpdB) in APEC on disease physiology and pathogenicity, an avian pathogenic Escherichia coli-34 (APEC-34) cpdB mutant was obtained using the Red system. The cpdB mutant grew at a slower rate than the natural strain APEC-34. Scanning electron microscopy (SEM) indicated that the bacteria of the cpdB mutant were significantly longer than the bacteria observed in the natural strain (P<0.01), and that the width of the cpdB mutant was significantly smaller than its natural counterpart (P<0.01). In order to evaluate the role of cpdB in APEC in the colonization of internal organs (lung, liver and spleen) in poultry, seven-day-old SPF chicks were infected with 10 9 CFU/chick of the cpdB mutant or the natural strain. No colonizations of cpdB mutants were observed in the internal organs 10days following the infection, though numerous natural strains were observed at 20days following infection. Additionally, the relative expression of division protein ftsZ, outer membrane protein A ompA, ferric uptake regulator fur and tryptophanase tnaA genes in the mutant strain were all significantly lower than in the natural strain (P<0.05 or P<0.01). These results suggested that cpdB is involved in the long-term colonization of APEC in the internal organs of the test subjects. The deletion of the cpdB gene also significantly affected the APEC growth and morphology. Copyright © 2016. Published by Elsevier Ltd.

[Study on national active monitoring for food borne pathogens and antimicrobial resistance in China 2001].

PubMed

Wang, Maoqi; Ran, Lu; Wang, Zhutian; Li, Zhigang

2004-01-01

To survey food borne pathogens and antimicrobial resistance in China. A total of 4034 samples of foods (raw meats, raw milk, cooked meats, ice cream, yoghurt, aquatic product and vegetable) were examined for the presence of Escherichia coli O157:H7 Salmonella spp and Listeria monocytogens by national active foodborne pathogens surveillance system. The samples were obtained from 11 provinces in 2001. Approximate 5.50% of the all samples yielded 3 pathogenes, whereas Escherichia coli O157:H7 (0.82%), Salmonella servoars (3.32%) and Listeria monocytogens (1.29%). The most heavy contamination by three food borne pathogens are in raw meat (12.96%). Top seven serotype of the 137 Salmonella isolates are S. derby, S. agona, S. enteritidis, S. reading, S. anatum. S. muenster, S. typhimurium. Serotypes and antibiotic resistance patterns of Salmonella isolates are different in 11 provinces. E. coli O157:H7 strains that are isolated from raw meat and cooked meat have VT2, eae, Hly genes. Salmonella and E. coli strains of multidrug resistance were isolated and identified.

Virulence factors in Escherichia coli urinary tract infection.

PubMed Central

Johnson, J R

1991-01-01

Uropathogenic strains of Escherichia coli are characterized by the expression of distinctive bacterial properties, products, or structures referred to as virulence factors because they help the organism overcome host defenses and colonize or invade the urinary tract. Virulence factors of recognized importance in the pathogenesis of urinary tract infection (UTI) include adhesins (P fimbriae, certain other mannose-resistant adhesins, and type 1 fimbriae), the aerobactin system, hemolysin, K capsule, and resistance to serum killing. This review summarizes the virtual explosion of information regarding the epidemiology, biochemistry, mechanisms of action, and genetic basis of these urovirulence factors that has occurred in the past decade and identifies areas in need of further study. Virulence factor expression is more common among certain genetically related groups of E. coli which constitute virulent clones within the larger E. coli population. In general, the more virulence factors a strain expresses, the more severe an infection it is able to cause. Certain virulence factors specifically favor the development of pyelonephritis, others favor cystitis, and others favor asymptomatic bacteriuria. The currently defined virulence factors clearly contribute to the virulence of wild-type strains but are usually insufficient in themselves to transform an avirulent organism into a pathogen, demonstrating that other as-yet-undefined virulence properties await discovery. Virulence factor testing is a useful epidemiological and research tool but as yet has no defined clinical role. Immunological and biochemical anti-virulence factor interventions are effective in animal models of UTI and hold promise for the prevention of UTI in humans. Images PMID:1672263

SIGIRR, a negative regulator of TLR/IL-1R signalling promotes Microbiota dependent resistance to colonization by enteric bacterial pathogens.

PubMed

Sham, Ho Pan; Yu, Emily Yi Shan; Gulen, Muhammet F; Bhinder, Ganive; Stahl, Martin; Chan, Justin M; Brewster, Lara; Morampudi, Vijay; Gibson, Deanna L; Hughes, Michael R; McNagny, Kelly M; Li, Xiaoxia; Vallance, Bruce A

2013-01-01

Enteric bacterial pathogens such as enterohemorrhagic E. coli (EHEC) and Salmonella Typhimurium target the intestinal epithelial cells (IEC) lining the mammalian gastrointestinal tract. Despite expressing innate Toll-like receptors (TLRs), IEC are innately hypo-responsive to most bacterial products. This is thought to prevent maladaptive inflammatory responses against commensal bacteria, but it also limits antimicrobial responses by IEC to invading bacterial pathogens, potentially increasing host susceptibility to infection. One reason for the innate hypo-responsiveness of IEC is their expression of Single Ig IL-1 Related Receptor (SIGIRR), a negative regulator of interleukin (IL)-1 and TLR signaling. To address whether SIGIRR expression and the innate hypo-responsiveness of IEC impacts on enteric host defense, Sigirr deficient (-/-) mice were infected with the EHEC related pathogen Citrobacter rodentium. Sigirr -/- mice responded with accelerated IEC proliferation and strong pro-inflammatory and antimicrobial responses but surprisingly, Sigirr -/- mice proved dramatically more susceptible to infection than wildtype mice. Through haematopoietic transplantation studies, it was determined that SIGIRR expression by non-haematopoietic cells (putative IEC) regulated these responses. Moreover, the exaggerated responses were found to be primarily dependent on IL-1R signaling. Whilst exploring the basis for their susceptibility, Sigirr -/- mice were found to be unusually susceptible to intestinal Salmonella Typhimurium colonization, developing enterocolitis without the typical requirement for antibiotic based removal of competing commensal microbes. Strikingly, the exaggerated antimicrobial responses seen in Sigirr -/- mice were found to cause a rapid and dramatic loss of commensal microbes from the infected intestine. This depletion appears to reduce the ability of the microbiota to compete for space and nutrients (colonization resistance) with the invading pathogens, leaving the intestine highly susceptible to pathogen colonization. Thus, SIGIRR expression by IEC reflects a strategy that sacrifices maximal innate responsiveness by IEC in order to promote commensal microbe based colonization resistance against bacterial pathogens.

Evaluating the pathogenic potential of environmental Escherichia coli by using the Caenorhabditis elegans infection model.

PubMed

Merkx-Jacques, Alexandra; Coors, Anja; Brousseau, Roland; Masson, Luke; Mazza, Alberto; Tien, Yuan-Ching; Topp, Edward

2013-04-01

The detection and abundance of Escherichia coli in water is used to monitor and mandate the quality of drinking and recreational water. Distinguishing commensal waterborne E. coli isolates from those that cause diarrhea or extraintestinal disease in humans is important for quantifying human health risk. A DNA microarray was used to evaluate the distribution of virulence genes in 148 E. coli environmental isolates from a watershed in eastern Ontario, Canada, and in eight clinical isolates. Their pathogenic potential was evaluated with Caenorhabditis elegans, and the concordance between the bioassay result and the pathotype deduced by genotyping was explored. Isolates identified as potentially pathogenic on the basis of their complement of virulence genes were significantly more likely to be pathogenic to C. elegans than those determined to be potentially nonpathogenic. A number of isolates that were identified as nonpathogenic on the basis of genotyping were pathogenic in the infection assay, suggesting that genotyping did not capture all potentially pathogenic types. The detection of the adhesin-encoding genes sfaD, focA, and focG, which encode adhesins; of iroN2, which encodes a siderophore receptor; of pic, which encodes an autotransporter protein; and of b1432, which encodes a putative transposase, was significantly associated with pathogenicity in the infection assay. Overall, E. coli isolates predicted to be pathogenic on the basis of genotyping were indeed so in the C. elegans infection assay. Furthermore, the detection of C. elegans-infective environmental isolates predicted to be nonpathogenic on the basis of genotyping suggests that there are hitherto-unrecognized virulence factors or combinations thereof that are important in the establishment of infection.

Comparison of fecal indicators with pathogenic bacteria and rotavirus in groundwater.

PubMed

Ferguson, Andrew S; Layton, Alice C; Mailloux, Brian J; Culligan, Patricia J; Williams, Daniel E; Smartt, Abby E; Sayler, Gary S; Feighery, John; McKay, Larry D; Knappett, Peter S K; Alexandrova, Ekaterina; Arbit, Talia; Emch, Michael; Escamilla, Veronica; Ahmed, Kazi Matin; Alam, Md Jahangir; Streatfield, P Kim; Yunus, Mohammad; van Geen, Alexander

2012-08-01

Groundwater is routinely analyzed for fecal indicators but direct comparisons of fecal indicators to the presence of bacterial and viral pathogens are rare. This study was conducted in rural Bangladesh where the human population density is high, sanitation is poor, and groundwater pumped from shallow tubewells is often contaminated with fecal bacteria. Five indicator microorganisms (E. coli, total coliform, F+RNA coliphage, Bacteroides and human-associated Bacteroides) and various environmental parameters were compared to the direct detection of waterborne pathogens by quantitative PCR in groundwater pumped from 50 tubewells. Rotavirus was detected in groundwater filtrate from the largest proportion of tubewells (40%), followed by Shigella (10%), Vibrio (10%), and pathogenic E. coli (8%). Spearman rank correlations and sensitivity-specificity calculations indicate that some, but not all, combinations of indicators and environmental parameters can predict the presence of pathogens. Culture-dependent fecal indicator bacteria measured on a single date did not predict total bacterial pathogens, but annually averaged monthly measurements of culturable E. coli did improve prediction for total bacterial pathogens. A qPCR-based E. coli assay was the best indicator for the bacterial pathogens. F+RNA coliphage were neither correlated nor sufficiently sensitive towards rotavirus, but were predictive of bacterial pathogens. Since groundwater cannot be excluded as a significant source of diarrheal disease in Bangladesh and neighboring countries with similar characteristics, the need to develop more effective methods for screening tubewells with respect to microbial contamination is necessary. Copyright © 2012 Elsevier B.V. All rights reserved.

Comparison of fecal indicators with pathogenic bacteria and rotavirus in groundwater

PubMed Central

Ferguson, Andrew S.; Layton, Alice C.; Mailloux, Brian J; Culligan, Patricia J.; Williams, Daniel E.; Smartt, Abby E.; Sayler, Gary S.; Feighery, John; McKay, Larry; Knappett, Peter S.K.; Alexandrova, Ekaterina; Arbit, Talia; Emch, Michael; Escamilla, Veronica; Ahmed, Kazi Matin; Alam, Md. Jahangir; Streatfield, P. Kim; Yunus, Mohammad; van Geen, Alexander

2012-01-01

Groundwater is routinely analyzed for fecal indicators but direct comparisons of fecal indicators to the presence of bacterial and viral pathogens are rare. This study was conducted in rural Bangladesh where the human population density is high, sanitation is poor, and groundwater pumped from shallow tubewells is often contaminated with fecal bacteria. Five indicator microorganisms (E. coli, total coliform, F+RNA coliphage, Bacteroides and human-associated Bacteroides) and various environmental parameters were compared to the direct detection of waterborne pathogens by quantitative PCR in groundwater pumped from 50 tubewells. Rotavirus was detected in groundwater filtrate from the largest proportion of tubewells (40%), followed by Shigella (10%), Vibrio (10%), and pathogenic E. coli (8%). Spearman rank correlations and sensitivity-specificity calculations indicate that some, but not all, combinations of indicators and environmental parameters can predict the presence of pathogens. Culture-dependent fecal indicator bacteria measured on a single date did not predict total bacterial pathogens, but annually averaged monthly measurements of culturable E. coli did improve prediction for total bacterial pathogens. A qPCR-based E. coli assay was the best indicator for the bacterial pathogens. F+RNA coliphage were neither correlated nor sufficiently sensitive towards rotavirus, but were predictive of bacterial pathogens. Since groundwater cannot be excluded as a significant source of diarrheal disease in Bangladesh and neighboring countries with similar characteristics, the need to develop more effective methods for screening tubewells with respect to microbial contamination is necessary. PMID:22705866

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.

Expression and purification of functional Clostridium perfringens alpha and epsilon toxins in Escherichia coli.

PubMed

Zhao, Yao; Kang, Lin; Gao, Shan; Zhou, Yang; Su, Libo; Xin, Wenwen; Su, Yuxin; Wang, Jinglin

2011-06-01

The alpha and epsilon toxins are 2 of the 4 major lethal toxins of the pathogen Clostridium perfringens. In this study, the expression of the epsilon toxin (etx) gene of C. perfringens was optimized by replacing rare codons with high-frequency codons, and the optimized gene was synthesized using overlapping PCR. Then, the etx gene or the alpha-toxin gene (cpa) was individually inserted into the pTIG-Trx expression vector with a hexahistidine tag and a thioredoxin (Trx) to facilitate their purification and induce the expression of soluble proteins. The recombinant alpha toxin (rCPA) and epsilon toxin (rETX) were highly expressed as soluble forms in the recipient Escherichia coli BL21 strain, respectively. The rCPA and rETX were purified using Ni(2+)-chelating chromatography and size-exclusion chromatography. And the entire purification process recovered about 40% of each target protein from the starting materials. The purified target toxins formed single band at about 42kDa (rCPA) or 31kDa (rETX) in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and their functional activity was confirmed by bioactivity assays. We have shown that the production of large amounts of soluble and functional proteins by using the pTIG-Trx vector in E. coli is a good alternative for the production of native alpha and epsilon toxins and could also be useful for the production of other toxic proteins with soluble forms. Copyright © 2011 Elsevier Inc. All rights reserved.

Escherichia coli and other Enterobacteriaceae: Food poisoning and health effects

USDA-ARS?s Scientific Manuscript database

The family Enterobactericeae consists of rod-shaped, Gram-negative, facultatively anaerobic, non-spore forming bacteria and also includes the food-borne pathogens, Cronobacter spp., Escherichia coli, Salmonella enterica, Shigella spp., and Yersinia spp. Illness caused by these pathogens is acquired...

Salmonella enterica suppresses Pectobacterium carotovorum subsp. carotovorum population and soft rot progression by acidifying the microaerophilic environment.

PubMed

Kwan, Grace; Charkowski, Amy O; Barak, Jeri D

2013-02-12

Although enteric human pathogens are usually studied in the context of their animal hosts, a significant portion of their life cycle occurs on plants. Plant disease alters the phyllosphere, leading to enhanced growth of human pathogens; however, the impact of human pathogens on phytopathogen biology and plant health is largely unknown. To characterize the interaction between human pathogens and phytobacterial pathogens in the phyllosphere, we examined the interactions between Pectobacterium carotovorum subsp. carotovorum and Salmonella enterica or Escherichia coli O157:H7 with regard to bacterial populations, soft rot progression, and changes in local pH. The presence of P. carotovorum subsp. carotovorum enhanced the growth of both S. enterica and E. coli O157:H7 on leaves. However, in a microaerophilic environment, S. enterica reduced P. carotovorum subsp. carotovorum populations and soft rot progression by moderating local environmental pH. Reduced soft rot was not due to S. enterica proteolytic activity. Limitations on P. carotovorum subsp. carotovorum growth, disease progression, and pH elevation were not observed on leaves coinoculated with E. coli O157:H7 or when leaves were coinoculated with S. enterica in an aerobic environment. S. enterica also severely undermined the relationship between the phytobacterial population and disease progression of a P. carotovorum subsp. carotovorum budB mutant defective in the 2,3-butanediol pathway for acid neutralization. Our results show that S. enterica and E. coli O157:H7 interact differently with the enteric phytobacterial pathogen P. carotovorum subsp. carotovorum. S. enterica inhibition of soft rot progression may conceal a rapidly growing human pathogen population. Whereas soft rotted produce can alert consumers to the possibility of food-borne pathogens, healthy-looking produce may entice consumption of contaminated vegetables. Salmonella enterica and Escherichia coli O157:H7 may use plants to move between animal and human hosts. Their populations are higher on plants cocolonized with the common bacterial soft rot pathogen Pectobacterium carotovorum subsp. carotovorum, turning edible plants into a risk factor for human disease. We inoculated leaves with P. carotovorum subsp. carotovorum and S. enterica or E. coli O157:H7 to study the interactions between these bacteria. While P. carotovorum subsp. carotovorum enhanced the growth of both S. enterica and E. coli O157:H7, these human pathogens affected P. carotovorum subsp. carotovorum fundamentally differently. S. enterica reduced P. carotovorum subsp. carotovorum growth and acidified the environment, leading to less soft rot on leaves; E. coli O157:H7 had no such effects. As soft rot signals a food safety risk, the reduction of soft rot symptoms in the presence of S. enterica may lead consumers to eat healthy-looking but S. enterica-contaminated produce.

Identification and differentiation of Campylobacter species by high-resolution melting curve analysis.

PubMed

Hoseinpour, Fatemeh; Foroughi, Azadeh; Nomanpour, Bizhan; Nasab, Rezvan Sobhani

2017-07-01

Campylobacter jejuni and Campylobacter coli are the important food-born zoonotic pathogen, also are leading causes of human food borne illnesses worldwide. cadF gene is expressed in all C. jejuni and C. coli strains and mediates cell binding to the cell matrix protein, Fibronectin. High-resolution melting (HRM) analysis is emerging as an efficient and robust method for discriminating DNA sequence variants. The goal of this study was to apply HRM analysis to identification of C. jejuni and C. coli. A total of 100 samples were obtained from chicken in Kermanshah, Iran. HRM analysis based on cadF gene and Eva Green was developed to the identification of Campylobacter to the species level. Fifty-five of 100 samples were positive as campylobacter (7 C. jejuni, 29 C. coli, 16 mixes and 3 unknown). Minor variations were observed in melting point temperatures of C. coli or C. jejuni isolates and this variation can be used to the differentiation between C. coli or C. jejuni isolates. The results of this study indicated that HRM curve analysis can be a suitable technique and rapid technique for distinguishing between C. jejuni and C. coli isolates. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Survey for Escherichia coli Virulence Factors in Asymptomatic Free-Ranging Parrots

PubMed Central

Becker Saidenberg, André; Robaldo Guedes, Neiva Maria; Fernandes Seixas, Gláucia Helena; da Costa Allgayer, Mariangela; Pacífico de Assis, Erica; Fabio Silveira, Luis; Anne Melville, Priscilla; Benites, Nilson Roberti

2012-01-01

Parrots in captivity are frequently affected by Escherichia coli (E. coli) infections. The objective of this study was to collect information on the carrier state for E. coli pathotypes in asymptomatic free-ranging parrots. Cloacal swabs were collected from nestlings of Hyacinth, Lear's macaws and Blue-fronted Amazon parrots and tested by polymerase chain reaction (PCR) for virulence factors commonly found in enteropathogenic, avian pathogenic, and uropathogenic E. coli strains. In total, 44 samples were cultured and E. coli isolates were yielded, from which DNA was extracted and processed by PCR. Genes commonly found in APEC isolates from Blue-fronted Amazon parrots and Hyacinth macaws were expressed in 14 of these 44 samples. One atypical EPEC isolate was obtained from a sample from Lear's macaw. The most commonly found gene was the increased serum survival (iss) gene. This is the first report, that describes such pathotypes in asymptomatic free-living parrots. The findings of this study suggest the presence of a stable host/parasite relationship at the time of the sampling brings a new understanding to the role that E. coli plays in captive and wild parrots. Such information can be used to improve husbandry protocols as well as help conservation efforts of free-living populations. PMID:23738135

A Microbiological Water Quality Evaluation of Ganges River Deltaic Aquifers

NASA Astrophysics Data System (ADS)

Yerby, C. J.; Gragg, S. E.; Page, J.; Leavens, J.; Bhattacharya, P.; Harrington, J.; Datta, S.

2014-12-01

Substantial natural contamination from trace elements (like arsenic) and pathogens make Ganges Deltaic aquifers an area of utmost concern. Following millions of cases of chronic arsenic poisoning from the groundwaters of the region, numerous residents are still knowingly ingesting water from shallow to intermediate accessible depth drinking water wells. Added to the calamity of arsenic is the prevalence of pathogenic bacteria in these waters. The increasing frequency of gastroenteritis signifies the need to quantify the magnitude and extensiveness of health degrading agents--bacterial pathogens (i.e. Salmonella) and non-pathogens (i.e. Enterobacteriaceae) --within the water supply in accessible Gangetic aquifers. To assess the dissolved microbiological quality in the region, present study sampling locations are along defined piezometer nests in an area in SE Asia (Bangladesh). Every nest contains samples from wells at varying depths covering shallow to deep aquifers. To date, 17 of the 76 water samples were analyzed for Salmonella, generic Escherichia coli (E. coli) and coliforms. Briefly, samples were plated in duplicate onto E. coli/Coliform petrifilm and incubated at 370C for 48 hours. Next, each sample was enriched in buffered peptone water and incubated at 370C for 18 hours. Bacterial DNA was extracted and amplified using a qPCR machine. Amplification plots were analyzed to determine presence/absence of microorganisms. All water samples (n=~76) are analyzed for Salmonella, Escherichia coli O157:H7, Listeria spp. and Shigella. Pathogen populations of PCR-positive water samples are enumerated using the agar direct plate method. Non-pathogenic bacterial indicator organisms (i.e. Enterobacteriaceae) will also be enumerated. Over the course of the experiment, we hypothesize that shallower wells will 1)have a higher pathogen prevalence and 2)harbor pathogens and nonpathogens at higher concentrations. While the 17 samples analyzed to date were negative for Salmonella, and E. coli, we anticipate subsequent sample analyses may reveal, E. coli or pathogenic (i.e. Salmonella) contamination due to livestock and anthropogenic wastes in the area. With farmers using shallow depth aquifers to irrigate crops, there is a very real threat of foodborne illness and the risk to public health is great.

An overview of molecular stress response mechanisms in Escherichia coli contributing to survival of Shiga toxin-producing Escherichia coli during raw milk cheese production.

PubMed

Peng, Silvio; Tasara, Taurai; Hummerjohann, Jörg; Stephan, Roger

2011-05-01

The ability of foodborne pathogens to survive in certain foods mainly depends on stress response mechanisms. Insight into molecular properties enabling pathogenic bacteria to survive in food is valuable for improvement of the control of pathogens during food processing. Raw milk cheeses are a potential source for human infections with Shiga toxin-producing Escherichia coli (STEC). In this review, we focused on the stress response mechanisms important for allowing STEC to survive raw milk cheese production processes. The major components and regulation pathways for general, acid, osmotic, and heat shock stress responses in E. coli and the implications of these responses for the survival of STEC in raw milk cheeses are discussed.

Immune function of a Rab-related protein by modulating the JAK-STAT signaling pathway in the silkworm, Bombyx mori.

PubMed

Chen, Chen; Eldein, Salah; Zhou, Xiaosan; Sun, Yu; Gao, Jin; Sun, Yuxuan; Liu, Chaoliang; Wang, Lei

2018-01-01

The Rab-family GTPases mainly regulate intracellular vesicle transport, and play important roles in the innate immune response in invertebrates. However, the function and signal transduction of Rab proteins in immune reactions remain unclear in silkworms. In this study, we analyzed a Rab-related protein of silkworm Bombyx mori (BmRABRP) by raising antibodies against its bacterially expressed recombinant form. Tissue distribution analysis showed that BmRABRP mRNA and protein were high expressed in the Malpighian tubule and fat body, respectively. However, among the different stages, only the fourth instar larvae and pupae showed significant BmRABRP levels. After challenge with four pathogenic microorganisms (Escherichia coli, BmNPV, Beauveria bassiana, Micrococcus luteus), the expression of BmRABRP mRNA in the fat body was significantly upregulated. In contrast, the BmRABRP protein was significantly upregulated after infection with BmNPV, while it was downregulated by E. coli, B. bassiana, and M. luteus. A specific dsRNA was used to explore the immune function and relationship between BmRABRP and the JAK-STAT signaling pathway. After BmRABRP gene interference, significant reduction in the number of nodules and increased mortality suggested that BmRABRP plays an important role in silkworm's response to bacterial challenge. In addition, four key genes (BmHOP, BmSTAT, BmSOCS2, and BmSOCS6) of the JAK-STAT signaling pathway showed significantly altered expressions after BmRABRP silencing. BmHOP and BmSOCS6 expressions were significantly decreased, while BmSTAT and BmSOCS2 were significantly upregulated. Our results suggested that BmRABRP is involved in the innate immune response against pathogenic microorganisms through the JAK-STAT signaling pathway in silkworm. © 2017 Wiley Periodicals, Inc.

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

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.

Characterization of urinary tract infection-associated Shiga toxin-producing Escherichia coli.

PubMed

Toval, Francisco; Schiller, Roswitha; Meisen, Iris; Putze, Johannes; Kouzel, Ivan U; Zhang, Wenlan; Karch, Helge; Bielaszewska, Martina; Mormann, Michael; Müthing, Johannes; Dobrindt, Ulrich

2014-11-01

Enterohemorrhagic Escherichia coli (EHEC), a subgroup of Shiga toxin (Stx)-producing E. coli (STEC), is a leading cause of diarrhea and hemolytic-uremic syndrome (HUS) in humans. However, urinary tract infections (UTIs) caused by this microorganism but not associated with diarrhea have occasionally been reported. We geno- and phenotypically characterized three EHEC isolates obtained from the urine of hospitalized patients suffering from UTIs. These isolates carried typical EHEC virulence markers and belonged to HUS-associated E. coli (HUSEC) clones, but they lacked virulence markers typical of uropathogenic E. coli. One isolate exhibited a localized adherence (LA)-like pattern on T24 urinary bladder epithelial cells. Since the glycosphingolipids (GSLs) globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer) are well-known receptors for Stx but also for P fimbriae, a major virulence factor of extraintestinal pathogenic E. coli (ExPEC), the expression of Gb3Cer and Gb4Cer by T24 cells and in murine urinary bladder tissue was examined by thin-layer chromatography and mass spectrometry. We provide data indicating that Stxs released by the EHEC isolates bind to Gb3Cer and Gb4Cer isolated from T24 cells, which were susceptible to Stx. All three EHEC isolates expressed stx genes upon growth in urine. Two strains were able to cause UTI in a murine infection model and could not be outcompeted in urine in vitro by typical uropathogenic E. coli isolates. Our results indicate that despite the lack of ExPEC virulence markers, EHEC variants may exhibit in certain suitable hosts, e.g., in hospital patients, a uropathogenic potential. The contribution of EHEC virulence factors to uropathogenesis remains to be further investigated. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Coculture of Escherichia coli O157:H7 with a Nonpathogenic E. coli Strain Increases Toxin Production and Virulence in a Germfree Mouse Model.

PubMed

Goswami, Kakolie; Chen, Chun; Xiaoli, Lingzi; Eaton, Kathryn A; Dudley, Edward G

2015-11-01

Escherichia coli O157:H7 is a notorious foodborne pathogen due to its low infectious dose and the disease symptoms it causes, which include bloody diarrhea and severe abdominal cramps. In some cases, the disease progresses to hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS), due to the expression of one or more Shiga toxins (Stx). Isoforms of Stx, including Stx2a, are encoded within temperate prophages. In the presence of certain antibiotics, phage induction occurs, which also increases the expression of toxin genes. Additionally, increased Stx2 accumulation has been reported when O157:H7 was cocultured with phage-susceptible nonpathogenic E. coli. This study characterized an E. coli O157:H7 strain, designated PA2, that belongs to the hypervirulent clade 8 cluster. Stx2a levels after ciprofloxacin induction were lower for PA2 than for the prototypical outbreak strains Sakai and EDL933. However, during coculture with the nonpathogenic strain E. coli C600, PA2 produced Stx2a levels that were 2- to 12-fold higher than those observed during coculture with EDL933 and Sakai, respectively. Germfree mice cocolonized by PA2 and C600 showed greater kidney damage, increased Stx2a accumulation in feces, and more visible signs of disease than mice given PA2 or C600 alone. These data suggest one mechanism by which microorganisms associated with the colonic microbiota could enhance the virulence of E. coli O157:H7, particularly a subset of clade 8 strains. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

IscR Regulates Synthesis of Colonization Factor Antigen I Fimbriae in Response to Iron Starvation in Enterotoxigenic Escherichia coli

PubMed Central

Arnaud-Barbe, Nadège; Poncet, David; Reverchon, Sylvie; Wawrzyniak, Julien; Nasser, William

2015-01-01

ABSTRACT Iron availability functions as an environmental cue for enteropathogenic bacteria, signaling arrival within the human host. As enterotoxigenic Escherichia coli (ETEC) is a major cause of human diarrhea, the effect of iron on ETEC virulence factors was evaluated here. ETEC pathogenicity is directly linked to production of fimbrial colonization factors and secretion of heat-labile enterotoxin (LT) and/or heat-stable enterotoxin (ST). Efficient colonization of the small intestine further requires at least the flagellin binding adhesin EtpA. Under iron starvation, production of the CFA/I fimbriae was increased in the ETEC H10407 prototype strain. In contrast, LT secretion was inhibited. Furthermore, under iron starvation, gene expression of the cfa (CFA/I) and etp (EtpBAC) operons was induced, whereas transcription of toxin genes was either unchanged or repressed. Transcriptional reporter fusion experiments focusing on the cfa operon further showed that iron starvation stimulated cfaA promoter activity in ETEC, indicating that the impact of iron on CFA/I production was mediated by transcriptional regulation. Evaluation of cfaA promoter activity in heterologous E. coli single mutant knockout strains identified IscR as the regulator responsible for inducing cfa fimbrial gene expression in response to iron starvation, and this was confirmed in an ETEC ΔiscR strain. The global iron response regulator, Fur, was not implicated. IscR binding sites were identified in silico within the cfaA promoter and fixation confirmed by DNase I footprinting, indicating that IscR directly binds the promoter region to induce CFA/I. IMPORTANCE Pathogenic enterobacteria modulate expression of virulence genes in response to iron availability. Although the Fur transcription factor represents the global regulator of iron homeostasis in Escherichia coli, we show that several ETEC virulence factors are modulated by iron, with expression of the major fimbriae under the control of the iron-sulfur cluster regulator, IscR. Furthermore, we demonstrate that the apo form of IscR, lacking an Fe-S cluster, is able to directly fix the corresponding promoter region. These results provide further evidence implicating IscR in bacterial virulence and suggest that IscR may represent a more general regulator mediating the iron response in enteropathogens. PMID:26124243

The Evolution of Campylobacter jejuni and Campylobacter coli

PubMed Central

Sheppard, Samuel K.; Maiden, Martin C.J.

2015-01-01

The global significance of Campylobacter jejuni and Campylobacter coli as gastrointestinal human pathogens has motivated numerous studies to characterize their population biology and evolution. These bacteria are a common component of the intestinal microbiota of numerous bird and mammal species and cause disease in humans, typically via consumption of contaminated meat products, especially poultry meat. Sequence-based molecular typing methods, such as multilocus sequence typing (MLST) and whole genome sequencing (WGS), have been instructive for understanding the epidemiology and evolution of these bacteria and how phenotypic variation relates to the high degree of genetic structuring in C. coli and C. jejuni populations. Here, we describe aspects of the relatively short history of coevolution between humans and pathogenic Campylobacter, by reviewing research investigating how mutation and lateral or horizontal gene transfer (LGT or HGT, respectively) interact to create the observed population structure. These genetic changes occur in a complex fitness landscape with divergent ecologies, including multiple host species, which can lead to rapid adaptation, for example, through frame-shift mutations that alter gene expression or the acquisition of novel genetic elements by HGT. Recombination is a particularly strong evolutionary force in Campylobacter, leading to the emergence of new lineages and even large-scale genome-wide interspecies introgression between C. jejuni and C. coli. The increasing availability of large genome datasets is enhancing understanding of Campylobacter evolution through the application of methods, such as genome-wide association studies, but MLST-derived clonal complex designations remain a useful method for describing population structure. PMID:26101080

Antibacterial activity and mechanism of action of ε-poly-L-lysine.

PubMed

Ye, Ruosong; Xu, Hengyi; Wan, Cuixiang; Peng, Shanshan; Wang, Lijun; Xu, Hong; Aguilar, Zoraida P; Xiong, Yonghua; Zeng, Zheling; Wei, Hua

2013-09-13

ε-Poly-L-lysine (ε-PL)(2) is widely used as an antibacterial agent because of its broad antimicrobial spectrum. However, the mechanism of ε-PL against pathogens at the molecular level has not been elucidated. This study investigated the antibacterial activity and mechanism of ε-PL against Escherichia coli O157:H7 CMCC44828. Propidium monoazide-PCR test results indicated that the threshold condition of ε-PL for complete membrane lysis of E. coli O157:H7 was 10 μg/mL (90% mortality for 5 μg/mL). Further verification of the destructive effect of ε-PL on cell structure was performed by atomic force microscopy and transmission electron microscopy. Results showed a positive correlation between reactive oxygen species (ROS)(3) levels and ε-PL concentration in E. coli O157:H7 cells. Moreover, the mortality of E. coli O157:H7 was reduced when antioxidant N-acetylcysteine was added. Results from real-time quantitative PCR (RT-qPCR)(4) indicated that the expression levels of oxidative stress genes sodA and oxyR were up-regulated 4- and 16-fold, respectively, whereas virulence genes eaeA and espA were down-regulated after ε-PL treatment. Expression of DNA damage response (SOS response)(5) regulon genes recA and lexA were also affected by ε-PL. In conclusion, the antibacterial mechanism of ε-PL against E. coli O157:H7 may be attributed to disturbance on membrane integrity, oxidative stress by ROS, and effects on various gene expressions, such as regulation of oxidative stress, SOS response, and changes in virulence. Copyright © 2013. Published by Elsevier Inc.

Biofiltration for stormwater harvesting: Comparison of Campylobacter spp. and Escherichia coli removal under normal and challenging operational conditions

NASA Astrophysics Data System (ADS)

Chandrasena, G. I.; Deletic, A.; McCarthy, D. T.

2016-06-01

Knowledge of pathogen removal in stormwater biofilters (also known as stormwater bioretention systems or rain gardens) has predominately been determined using bacterial indicators, and the removal of reference pathogens in these systems has rarely been investigated. Furthermore, current understanding of indicator bacteria removal in these systems is largely built upon laboratory-scale work. This paper examines whether indicator organism removal from urban stormwater using biofilters in laboratory settings are representative of the removal of pathogens in field conditions, by studying the removal of Escherichia coli (a typical indicator microorganism) and Campylobacter spp. (a typical reference pathogen) from urban stormwater by two established field-scale biofilters. It was found that E. coli log reduction was higher than that of Campylobacter spp. in both biofilters, and that there was no correlation between E. coli and Campylobacter spp. log removal performance. This confirms that E. coli behaves significantly differently to this reference pathogen, reinforcing that single organisms should not be employed to understand faecal microorganism removal in urban stormwater treatment systems. The average reduction in E. coli from only one of the tested biofilters was able to meet the log reduction targets suggested in the current Australian stormwater harvesting guidelines for irrigating sports fields and golf courses. The difference in the performance of the two biofilters is likely a result of a number of design and operational factors; the most important being that the biofilter that did not meet the guidelines was tested using extremely high influent volumes and microbial concentrations, and long antecedent dry weather periods. As such, the E. coli removal performances identified in this study confirmed laboratory findings that inflow concentration and antecedent dry period impact overall microbial removal. In general, this paper emphasizes the need for the validation of stormwater harvesting systems, namely, the testing of treatment systems under challenging operational conditions using multiple indicators and reference pathogens.

Effect of chlorine treatment on inhibition of E. coli serogroup O2 incorporation into 7-day-old biofilm on polyvinylchloride surface.

PubMed

Maharjan, P; Dey, S; Huff, G; Zhang, W; Phillips, G K; Watkins, S

2017-08-01

Poultry waterlines are constructed using polyvinylchloride (PVC) material on which bacterial biofilm can easily form. Biofilm can harbor pathogens including avian pathogenic E. coli (APEC) strains. An in vitro evaluation was performed to determine if E. coli sero group O2 (avian pathogenic) could attach on a PVC surface that had pre-formed biofilm and if this phenomenon could be affected when water was treated with chlorine. Initially, biofilm growth was induced in PVC test coupons (15.16 cm2) for a 7-day period mimicking the waterline scenario in the first wk of poultry brooding; and then this biofilm was challenged with E. coli O2 seeded water in presence/absence of chlorine treatment. After rinsing, test coupons were sampled for bacterial (APC) and E. coli O2 enumeration at various occasions post seeding the pathogen and chlorine treatment. Day 7 APC recovered from coupons was 4.35 log10 cfu/cm2 in trial 1 and 3.66 log10 cfu/cm2 in trial 2. E. coli O2 was not recovered from chlorine treated test coupons (P < 0.05), whereas it was retrieved from untreated coupons (untreated contained > 3 log10 cfu/cm2 in trial 1 and > 2 log10 cfu/cm2 in trial 2). This study suggests that E. coli O2 can incorporate into pre-formed biofilm on a PVC surface within 24 h if water sanitation is not present, and the attachment time of the pathogen can prolong in the absence of already formed biofilm. © 2017 Poultry Science Association Inc.

Dynamics of E.coli virulence factors in dairy cow herds

USDA-ARS?s Scientific Manuscript database

Background. Dairy farms are known reservoirs of entero-pathogenic E. coli (EPEC). EPEC, or the virulence factors associated with pathogenicity, have been detected in manure, milk, and the farm environment. However, it is unclear which farm compartments are reservoirs contributing to EPEC persistence...

Virulence gene content in Escherichia coli isolates from poultry flocks with clinical signs of colibacillosis in Brazil.

PubMed

De Carli, Silvia; Ikuta, Nilo; Lehmann, Fernanda Kieling Moreira; da Silveira, Vinicius Proença; de Melo Predebon, Gabriela; Fonseca, André Salvador Kazantzi; Lunge, Vagner Ricardo

2015-11-01

Escherichia coli is a commensal bacterium of the bird's intestinal tract, but it can invade different tissues resulting in systemic symptoms (colibacillosis). This disease occurs only when the E. coli infecting strain presents virulence factors (encoded by specific genes) that enable the adhesion and proliferation in the host organism. Thus, it is important to differentiate pathogenic (APEC, avian pathogenic E. coli) and non-pathogenic or fecal (AFEC, avian fecal E. coli) isolates. Previous studies analyzed the occurrence of virulence factors in E. coli strains isolated from birds with colibacillosis, demonstrating a high frequency of the bacterial genes cvaC, iroN, iss, iutA, sitA, tsh, fyuA, irp-2, ompT and hlyF in pathogenic strains. The aim of the present study was to evaluate the occurrence and frequency of these virulence genes in E. coli isolated from poultry flocks in Brazil. A total of 138 isolates of E. coli was obtained from samples of different tissues and/or organs (spleen, liver, kidney, trachea, lungs, skin, ovary, oviduct, intestine, cloaca) and environmental swabs collected from chicken and turkey flocks suspected to have colibacillosis in farms from the main Brazilian producing regions. Total DNA was extracted and the 10 virulence genes were detected by traditional and/or real-time PCR. At least 11 samples of each gene were sequenced and compared to reference strains. All 10 virulence factors were detected in Brazilian E. coli isolates, with frequencies ranging from 39.9% (irp-2) to 68.8% (hlyF and sitA). Moreover, a high nucleotide similarity (over 99%) was observed between gene sequences of Brazilian isolates and reference strains. Seventy-nine isolates were defined as pathogenic (APEC) and 59 as fecal (AFEC) based on previously described criteria. In conclusion, the main virulence genes of the reference E. coli strains are also present in isolates associated with colibacillosis in Brazil. The analysis of this set of virulence factors can be used to differentiate between APEC and AFEC isolates in Brazil. © 2015 Poultry Science Association Inc.

[Genetic mechanisms of Salmonella enteritidis biodiversity and clinical features of salmonellosis].

PubMed

Mavziutov, A R; Murzabaeva, R T; Nazmutdinova, R G; Mirsaiapova, I A

2010-01-01

To assess prevalence of fragments of Escherichia coli pathogenicity islands in Salmonella enteritidis strains as well as to study clinical signs of disease caused by these strains in adults. Ninety-six patients with salmonellosis were studied. Ninety strains of S. enteritidis were isolated and tested by PCR for the presence of genes associated with pathogenicity islands of E. coli: hlyA, hlyB, sfaG, and sfaA. It was determined that DNA fragments homologous to pathogenicity islands of E. coli were present in 87 (96.7%) of S. enteritidis clinical isolates. Disease caused by Salmonella strains which possess only sfaG was mostly mild--7 (33.3%), whereas strains which had sfaG with fragments of hlyA and/or hlyB caused severe disease--7 (50%). sfaA fragments were found mostly in combination with other genes. In such cases the disease was mostly severe--6 (42.8%). Correlation between presence of E. coli pathogenicity islands in Salmonella spp., their antibiotic resistance and severity of infection was established.

Escherichia coli LF82 differentially regulates ROS production and mucin expression in intestinal epithelial T84 cells: implication of NOX1.

PubMed

Elatrech, Imen; Marzaioli, Viviana; Boukemara, Hanane; Bournier, Odile; Neut, Christel; Darfeuille-Michaud, Arlette; Luis, José; Dubuquoy, Laurent; El-Benna, Jamel; My-Chan Dang, Pham; Marie, Jean-Claude

2015-05-01

Increased reactive oxygen species (ROS) production is associated with inflamed ileal lesions in Crohn's disease colonized by pathogenic adherent-invasive Escherichia coli LF82. We investigated whether such ileal bacteria can modulate ROS production by epithelial cells, thus impacting on inflammation and mucin expression. Ileal bacteria from patients with Crohn's disease were incubated with cultured epithelial T84 cells, and ROS production was assayed using the luminol-amplified chemiluminescence method. The gentamicin protection assay was used for bacterial invasion of T84 cell. The expression of NADPH oxidase (NOX) subunits, mucin, and IL-8 was analyzed by quantitative real-time PCR and Western blots. Involvement of NOX and ROS was analyzed using diphenyleneiodonium (DPI) and N-acetylcysteine (NAC). Among different bacteria tested, only LF82 induced an increase of ROS production by T84 cells in a dose-dependent manner. This response was inhibited by DPI and NAC. Heat- or ethanol-attenuated LF82 bacteria and the mutant LF82ΔFimA, which does not express pili type 1 and poorly adheres to epithelial cells, did not induce the oxidative response. The LF82-induced oxidative response coincides with its invasion in T84 cells, and both processes were inhibited by DPI. Also, we observed an increased expression of NOX1 and NOXO1 in response to LF82 bacteria versus the mutant LF82ΔFimA. Furthermore, LF82 inhibited mucin gene expression (MUC2 and MUC5AC) in T84 cells while increasing the chemotactic IL-8 expression, both in a DPI-sensitive manner. Adherent-invasive E. coli LF82 induced ROS production by intestinal NADPH oxidase and altered mucin and IL-8 expression, leading to perpetuation of inflammatory lesions in Crohn's disease.

A New Protocol to Detect Multiple Foodborne Pathogens with PCR Dipstick DNA Chromatography after a Six-Hour Enrichment Culture in a Broad-Range Food Pathogen Enrichment Broth

PubMed Central

Hayashi, Masahiro; Natori, Tatsuya; Kubota-Hayashi, Sayoko; Miyata, Machiko; Ohkusu, Kiyofumi; Kurazono, Hisao; Makino, Souichi; Ezaki, Takayuki

2013-01-01

A quick foodborne pathogen screening method after six-hour enrichment culture with a broad-range food pathogen enrichment broth is described. Pathogenic factors of Salmonella enterica, Shigella spp., enteroinvasive Escherichia coli, and enterohemorrhagic E. coli are amplified with a cocktail primer and rapid polymerase chain reaction (PCR), which finishes amplification in 30 min. The PCR amplicon was differentiated with a dipstick DNA chromatography assay in 5–10 min. Starting from a four- to six-hour enrichment culture, this assay was finished within 45 min. Detection sensitivity of this protocol was less than 2.5 CFU/25 g for S. enterica and 3.3 CFU/25 g for enterohemorrhagic E. coli in spiked ground meat experiments. PMID:24364031

Thymus transcriptome reveals novel pathways in response to avian pathogenic Escherichia coli infection.

PubMed

Sun, H; Liu, P; Nolan, L K; Lamont, S J

2016-12-01

Avian pathogenic Escherichia coli (APEC) can cause significant morbidity in chickens. The thymus provides the essential environment for T cell development; however, the thymus transcriptome has not been examined for gene expression in response to APEC infection. An improved understanding of the host genomic response to APEC infection could inform future breeding programs for disease resistance and APEC control. We therefore analyzed the transcriptome of the thymus of birds challenged with APEC, contrasting susceptible and resistant phenotypes. Thousands of genes were differentially expressed in birds of the 5-day post infection (dpi) challenged-susceptible group vs. 5 dpi non-challenged, in 5 dpi challenged-susceptible vs. 5 dpi challenged-resistant birds, as well as in 5 dpi vs. one dpi challenged-susceptible birds. The Toll-like receptor signaling pathway was the major innate immune response for birds to respond to APEC infection. Moreover, lysosome and cell adhesion molecules pathways were common mechanisms for chicken response to APEC infection. The T-cell receptor signaling pathway, cell cycle, and p53 signaling pathways were significantly activated in resistant birds to resist APEC infection. These results provide a comprehensive assessment of global gene networks and biological functionalities of differentially expressed genes in the thymus under APEC infection. These findings provide novel insights into key molecular genetic mechanisms that differentiate host resistance from susceptibility in this primary lymphoid tissue, the thymus. © The Author 2016. Published by Oxford University Press on behalf of Poultry Science Association.

The alkaloid compound harmane increases the lifespan of Caenorhabditis elegans during bacterial infection, by modulating the nematode's innate immune response.

PubMed

Jakobsen, Henrik; Bojer, Martin S; Marinus, Martin G; Xu, Tao; Struve, Carsten; Krogfelt, Karen A; Løbner-Olesen, Anders

2013-01-01

The nematode Caenorhabditis elegans has in recent years been proven to be a powerful in vivo model for testing antimicrobial compounds. We report here that the alkaloid compound Harmane (2-methyl-β-carboline) increases the lifespan of nematodes infected with a human pathogen, the Shiga toxin-producing Escherichia coli O157:H7 strain EDL933 and several other bacterial pathogens. This was shown to be unrelated to the weak antibiotic effect of Harmane. Using GFP-expressing E. coli EDL933, we showed that Harmane does not lower the colonization burden in the nematodes. We also found that the expression of the putative immune effector gene F35E12.5 was up-regulated in response to Harmane treatment. This indicates that Harmane stimulates the innate immune response of the nematode; thereby increasing its lifespan during bacterial infection. Expression of F35E12.5 is predominantly regulated through the p38 MAPK pathway; however, intriguingly the lifespan extension resulting from Harmane was higher in p38 MAPK-deficient nematodes. This indicates that Harmane has a complex effect on the innate immune system of C. elegans. Harmane could therefore be a useful tool in the further research into C. elegans immunity. Since the innate immunity of C. elegans has a high degree of evolutionary conservation, drugs such as Harmane could also be possible alternatives to classic antibiotics. The C. elegans model could prove to be useful for selection and development of such drugs.

Impedance based detection of pathogenic E. coli O157:H7 using a ferrocene-antimicrobial peptide modified biosensor.

PubMed

Li, Yongxin; Afrasiabi, Rouzbeh; Fathi, Farkhondeh; Wang, Nan; Xiang, Cuili; Love, Ryan; She, Zhe; Kraatz, Heinz-Bernhard

2014-08-15

Escherichia coli O157:H7 can cause life-threatening gastrointestinal diseases and has been a severe public health problem worldwide in recent years. A novel biosensor for the detection of E. coli O157:H7 is described here using a film composed of ferrocene-peptide conjugates, in which the antimicrobial peptide magainin I has been incorporated as the biorecognition element. Electrochemical impedance spectroscopy was employed to investigate the surface characteristics of the newly developed biosensor and to monitor the interactions between the peptide film and the pathogenic bacteria. X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry (ToF-SIMS) were employed to confirm the immobilization of ferrocene-conjugate onto the gold surface. Non-pathogenic E. coli K12, Staphylococcus epidermidis and Bacillus subtilis were used in this study to evaluate the selectivity of the proposed biosensor. The results have shown the order of the preferential selectivity of the method is E. coli O157:H7>non-pathogenic E. coli>gram positive species. The detection of E. coli O157:H7 with a sensitivity of 10(3)cfu/mL is enabled by the biosensor. The experimental conditions have been optimized and the plot of changes of charge transfer resistance (ΔRCT) and the logarithm of the cell concentration of E. coli O157:H7 shows a linear correlation in the range of 10(3)-10(7)cfu/mL with a correlation coefficient of 0.983. Copyright © 2014 Elsevier B.V. All rights reserved.

Evaluation of Escherichia coli isolates from healthy chickens to determine their potential risk to poultry and human health.

PubMed

Stromberg, Zachary R; Johnson, James R; Fairbrother, John M; Kilbourne, Jacquelyn; Van Goor, Angelica; Curtiss, Roy; Mellata, Melha

2017-01-01

Extraintestinal pathogenic Escherichia coli (ExPEC) strains are important pathogens that cause diverse diseases in humans and poultry. Some E. coli isolates from chicken feces contain ExPEC-associated virulence genes, so appear potentially pathogenic; they conceivably could be transmitted to humans through handling and/or consumption of contaminated meat. However, the actual extraintestinal virulence potential of chicken-source fecal E. coli is poorly understood. Here, we assessed whether fecal E. coli isolates from healthy production chickens could cause diseases in a chicken model of avian colibacillosis and three rodent models of ExPEC-associated human infections. From 304 E. coli isolates from chicken fecal samples, 175 E. coli isolates were screened by PCR for virulence genes associated with human-source ExPEC or avian pathogenic E. coli (APEC), an ExPEC subset that causes extraintestinal infections in poultry. Selected isolates genetically identified as ExPEC and non-ExPEC isolates were assessed in vitro for virulence-associated phenotypes, and in vivo for disease-causing ability in animal models of colibacillosis, sepsis, meningitis, and urinary tract infection. Among the study isolates, 13% (40/304) were identified as ExPEC; the majority of these were classified as APEC and uropathogenic E. coli, but none as neonatal meningitis E. coli. Multiple chicken-source fecal ExPEC isolates resembled avian and human clinical ExPEC isolates in causing one or more ExPEC-associated illnesses in experimental animal infection models. Additionally, some isolates that were classified as non-ExPEC were able to cause ExPEC-associated illnesses in animal models, and thus future studies are needed to elucidate their mechanisms of virulence. These findings show that E. coli isolates from chicken feces contain ExPEC-associated genes, exhibit ExPEC-associated in vitro phenotypes, and can cause ExPEC-associated infections in animal models, and thus may pose a health threat to poultry and consumers.

Evaluation of Escherichia coli isolates from healthy chickens to determine their potential risk to poultry and human health

PubMed Central

Johnson, James R.; Fairbrother, John M.; Kilbourne, Jacquelyn; Van Goor, Angelica; Curtiss, Roy; Mellata, Melha

2017-01-01

Extraintestinal pathogenic Escherichia coli (ExPEC) strains are important pathogens that cause diverse diseases in humans and poultry. Some E. coli isolates from chicken feces contain ExPEC-associated virulence genes, so appear potentially pathogenic; they conceivably could be transmitted to humans through handling and/or consumption of contaminated meat. However, the actual extraintestinal virulence potential of chicken-source fecal E. coli is poorly understood. Here, we assessed whether fecal E. coli isolates from healthy production chickens could cause diseases in a chicken model of avian colibacillosis and three rodent models of ExPEC-associated human infections. From 304 E. coli isolates from chicken fecal samples, 175 E. coli isolates were screened by PCR for virulence genes associated with human-source ExPEC or avian pathogenic E. coli (APEC), an ExPEC subset that causes extraintestinal infections in poultry. Selected isolates genetically identified as ExPEC and non-ExPEC isolates were assessed in vitro for virulence-associated phenotypes, and in vivo for disease-causing ability in animal models of colibacillosis, sepsis, meningitis, and urinary tract infection. Among the study isolates, 13% (40/304) were identified as ExPEC; the majority of these were classified as APEC and uropathogenic E. coli, but none as neonatal meningitis E. coli. Multiple chicken-source fecal ExPEC isolates resembled avian and human clinical ExPEC isolates in causing one or more ExPEC-associated illnesses in experimental animal infection models. Additionally, some isolates that were classified as non-ExPEC were able to cause ExPEC-associated illnesses in animal models, and thus future studies are needed to elucidate their mechanisms of virulence. These findings show that E. coli isolates from chicken feces contain ExPEC-associated genes, exhibit ExPEC-associated in vitro phenotypes, and can cause ExPEC-associated infections in animal models, and thus may pose a health threat to poultry and consumers. PMID:28671990

A conserved virulence plasmidic region contributes to the virulence of the multiresistant Escherichia coli meningitis strain S286 belonging to phylogenetic group C.

PubMed

Lemaître, Chloé; Mahjoub-Messai, Farah; Dupont, Damien; Caro, Valérie; Diancourt, Laure; Bingen, Edouard; Bidet, Philippe; Bonacorsi, Stéphane

2013-01-01

Recent isolation of the non-K1 Escherichia coli neonatal meningitis strain S286, belonging to phylogroup C, which is closely related to major group B1, and producing an extended-spectrum beta-lactamase, encouraged us to seek the genetic determinants responsible for its virulence. We show that S286 belongs to the sequence O type ST23O78 and harbors 4 large plasmids. The largest one, pS286colV (~120 kb), not related to resistance, contains genes characteristic of a Conserved Virulence Plasmidic (CVP) region initially identified in B2 extra-intestinal avian pathogenic E. coli (APEC) strains and in the B2 neonatal meningitis E. coli strain S88. The sequence of this CVP region has a strong homology (98%) with that of the recently sequenced plasmid pChi7122-1 of the O78 APEC strain Chi7122. A CVP plasmid-cured variant of S286 was less virulent than the wild type strain in a neonatal rat sepsis model with a significant lower level of bacteremia at 24 h (4.1 ± 1.41 versus 2.60 ± 0.16 log CFU/ml, p = 0.001) and mortality. However, the mortality in the model of adult mice was comparable between wild type and variant indicating that pS286colV is not sufficient by itself to fully explain the virulence of S286. Gene expression analysis of pS286colV in iron depleted environment was very close to that of pS88, suggesting that genes of CVP region may be expressed similarly in two very different genetic backgrounds (group C versus group B2). Screening a collection of 178 human A/B1 extraintestinal pathogenic E. coli (ExPEC) strains revealed that the CVP region is highly prevalent (23%) and MLST analysis indicated that these CVP positive strains belong to several clusters and mostly to phylogroup C. The virulence of S286 is explained in part by the presence of CVP region and this region has spread in different clusters of human A/B1 ExPEC, especially in group C.

Occurrence of pathogenicity island I(APEC-O1) genes among Escherichia coli implicated in avian colibacillosis.

PubMed

Kariyawasam, Subhashinie; Johnson, Timothy J; Debroy, Chitrita; Nolan, Lisa K

2006-09-01

Colibacillosis caused by avian pathogenic Escherichia coli (APEC) is a leading cause of economic loss to the poultry industry worldwide. The ability of APEC to cause disease is determined by certain virulence markers, some of which are located on pathogenicity islands (PAls). We recently described one such PAI in an APEC O1:K1 strain (APEC-O1). This PAI, termed PAI I(APEC-O1), carries the genes of the pap operon, a region similar to the tia invasion determinant of enterotoxigenic E coli; ireA, a gene that encodes an iron-responsive element; and a novel 1.5-kb region, ORF 54. Here, the occurrence of six selected loci of PAI I(APEC-O1) (papA, papC, papG, ireA, tia, and ORF 54) among APEC and fecal E. coli strains from apparently healthy chickens (avian commensal E. coli) was determined using polymerase chain reaction (PCR) techniques. None of the commensal E. coli was positive for all six traits, whereas 7.2% of the APEC isolates were positive for all the traits. Although there was no significant difference in the occurrence of ORF 54 among APEC and commensal E. coli, tia, ireA, papC, and papG genes were predominantly present in APEC rather than in avian commensal E. coli. papA was detected in only 6.3% of APEC, perhaps because of the presence of allelic variants of the gene. Additionally, the presence of all six traits was tested with PCR in APEC isolates collected in the 1980s, and these results were compared with those obtained with the APEC isolated in the 1990s. There was no significant difference in the occurrence of tia, ireA, papC, papG, and ORF 54 between APEC isolates collected during the different decades. However, papA was more frequently present in APEC from the 1980s than it was in APEC from the 1990s. Phylogenetic group of an isolate did not correlate with pathogenicity or the presence of PAI traits, except that more APEC of the low-pathogenicity group belonged to the phylogenetic group B1. However, PAI traits occurred more frequently in isolates belonging to the intermediate- and high-pathogenicity groups than in isolates of low pathogenicity.

Engineering the Controlled Assembly of Filamentous Injectisomes in E. coli K-12 for Protein Translocation into Mammalian Cells.

PubMed

Ruano-Gallego, David; Álvarez, Beatriz; Fernández, Luis Ángel

2015-09-18

Bacterial pathogens containing type III protein secretion systems (T3SS) assemble large needle-like protein complexes in the bacterial envelope, called injectisomes, for translocation of protein effectors into host cells. The application of these "molecular syringes" for the injection of proteins into mammalian cells is hindered by their structural and genomic complexity, requiring multiple polypeptides encoded along with effectors in various transcriptional units (TUs) with intricate regulation. In this work, we have rationally designed the controlled expression of the filamentous injectisomes found in enteropathogenic Escherichia coli (EPEC) in the nonpathogenic strain E. coli K-12. All structural components of EPEC injectisomes, encoded in a genomic island called the locus of enterocyte effacement (LEE), were engineered in five TUs (eLEEs) excluding effectors, promoters and transcriptional regulators. These eLEEs were placed under the control of the IPTG-inducible promoter Ptac and integrated into specific chromosomal sites of E. coli K-12 using a marker-less strategy. The resulting strain, named synthetic injector E. coli (SIEC), assembles filamentous injectisomes similar to those in EPEC. SIEC injectisomes form pores in the host plasma membrane and are able to translocate T3-substrate proteins (e.g., translocated intimin receptor, Tir) into the cytoplasm of HeLa cells reproducing the phenotypes of intimate attachment and polymerization of actin-pedestals elicited by EPEC bacteria. Hence, SIEC strain allows the controlled expression of functional filamentous injectisomes for efficient translocation of proteins with T3S-signals into mammalian cells.

The Genotoxin Colibactin Is a Determinant of Virulence in Escherichia coli K1 Experimental Neonatal Systemic Infection.

PubMed

McCarthy, Alex J; Martin, Patricia; Cloup, Emilie; Stabler, Richard A; Oswald, Eric; Taylor, Peter W

2015-09-01

Escherichia coli strains expressing the K1 capsule are a major cause of sepsis and meningitis in human neonates. The development of these diseases is dependent on the expression of a range of virulence factors, many of which remain uncharacterized. Here, we show that all but 1 of 34 E. coli K1 neonatal isolates carried clbA and clbP, genes contained within the pks pathogenicity island and required for the synthesis of colibactin, a polyketide-peptide genotoxin that causes genomic instability in eukaryotic cells by induction of double-strand breaks in DNA. Inactivation of clbA and clbP in E. coli A192PP, a virulent strain of serotype O18:K1 that colonizes the gastrointestinal tract and translocates to the blood compartment with very high frequency in experimental infection of the neonatal rat, significantly reduced the capacity of A192PP to colonize the gut, engender double-strand breaks in DNA, and cause invasive, lethal disease. Mutation of clbA, which encodes a pleiotropic enzyme also involved in siderophore synthesis, impacted virulence to a greater extent than mutation of clbP, encoding an enzyme specific to colibactin synthesis. Restoration of colibactin gene function by complementation reestablished the fully virulent phenotype. We conclude that colibactin contributes to the capacity of E. coli K1 to colonize the neonatal gastrointestinal tract and to cause invasive disease in the susceptible neonate. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

The Genotoxin Colibactin Is a Determinant of Virulence in Escherichia coli K1 Experimental Neonatal Systemic Infection

PubMed Central

McCarthy, Alex J.; Martin, Patricia; Cloup, Emilie; Stabler, Richard A.

2015-01-01

Escherichia coli strains expressing the K1 capsule are a major cause of sepsis and meningitis in human neonates. The development of these diseases is dependent on the expression of a range of virulence factors, many of which remain uncharacterized. Here, we show that all but 1 of 34 E. coli K1 neonatal isolates carried clbA and clbP, genes contained within the pks pathogenicity island and required for the synthesis of colibactin, a polyketide-peptide genotoxin that causes genomic instability in eukaryotic cells by induction of double-strand breaks in DNA. Inactivation of clbA and clbP in E. coli A192PP, a virulent strain of serotype O18:K1 that colonizes the gastrointestinal tract and translocates to the blood compartment with very high frequency in experimental infection of the neonatal rat, significantly reduced the capacity of A192PP to colonize the gut, engender double-strand breaks in DNA, and cause invasive, lethal disease. Mutation of clbA, which encodes a pleiotropic enzyme also involved in siderophore synthesis, impacted virulence to a greater extent than mutation of clbP, encoding an enzyme specific to colibactin synthesis. Restoration of colibactin gene function by complementation reestablished the fully virulent phenotype. We conclude that colibactin contributes to the capacity of E. coli K1 to colonize the neonatal gastrointestinal tract and to cause invasive disease in the susceptible neonate. PMID:26150540

Antibiotic-Resistant Pathogenic Escherichia Coli Isolated from Rooftop Rainwater-Harvesting Tanks in the Eastern Cape, South Africa.

PubMed

Malema, Mokaba Shirley; Abia, Akebe Luther King; Tandlich, Roman; Zuma, Bonga; Mwenge Kahinda, Jean-Marc; Ubomba-Jaswa, Eunice

2018-05-01

Although many developing countries use harvested rainwater (HRW) for drinking and other household purposes, its quality is seldom monitored. Continuous assessment of the microbial quality of HRW would ensure the safety of users of such water. The current study investigated the prevalence of pathogenic Escherichia coli strains and their antimicrobial resistance patterns in HRW tanks in the Eastern Cape, South Africa. Rainwater samples were collected weekly between June and September 2016 from 11 tanks in various areas of the province. Enumeration of E. coli was performed using the Colilert ® 18/Quanti-Tray ® 2000 method. E. coli isolates were obtained and screened for their virulence potentials using polymerase chain reaction (PCR), and subsequently tested for antibiotic resistance using the disc-diffusion method against 11 antibiotics. The pathotype most detected was the neonatal meningitis E. coli (NMEC) ( ibeA 28%) while pathotype enteroaggregative E. coli (EAEC) was not detected. The highest resistance of the E. coli isolates was observed against Cephalothin (76%). All tested pathotypes were susceptible to Gentamicin, and 52% demonstrated multiple-antibiotic resistance (MAR). The results of the current study are of public health concern since the use of untreated harvested rainwater for potable purposes may pose a risk of transmission of pathogenic and antimicrobial-resistant E. coli.

Antibiotic-Resistant Pathogenic Escherichia Coli Isolated from Rooftop Rainwater-Harvesting Tanks in the Eastern Cape, South Africa

PubMed Central

Malema, Mokaba Shirley; Tandlich, Roman; Zuma, Bonga; Mwenge Kahinda, Jean-Marc

2018-01-01

Although many developing countries use harvested rainwater (HRW) for drinking and other household purposes, its quality is seldom monitored. Continuous assessment of the microbial quality of HRW would ensure the safety of users of such water. The current study investigated the prevalence of pathogenic Escherichia coli strains and their antimicrobial resistance patterns in HRW tanks in the Eastern Cape, South Africa. Rainwater samples were collected weekly between June and September 2016 from 11 tanks in various areas of the province. Enumeration of E. coli was performed using the Colilert®18/Quanti-Tray® 2000 method. E. coli isolates were obtained and screened for their virulence potentials using polymerase chain reaction (PCR), and subsequently tested for antibiotic resistance using the disc-diffusion method against 11 antibiotics. The pathotype most detected was the neonatal meningitis E. coli (NMEC) (ibeA 28%) while pathotype enteroaggregative E. coli (EAEC) was not detected. The highest resistance of the E. coli isolates was observed against Cephalothin (76%). All tested pathotypes were susceptible to Gentamicin, and 52% demonstrated multiple-antibiotic resistance (MAR). The results of the current study are of public health concern since the use of untreated harvested rainwater for potable purposes may pose a risk of transmission of pathogenic and antimicrobial-resistant E. coli. PMID:29723970

Zoonotic potential of Escherichia coli isolates from retail chicken meat products and eggs.

PubMed

Mitchell, Natalie M; Johnson, James R; Johnston, Brian; Curtiss, Roy; Mellata, Melha

2015-02-01

Chicken products are suspected as a source of extraintestinal pathogenic Escherichia coli (ExPEC), which causes diseases in humans. The zoonotic risk to humans from chicken-source E. coli is not fully elucidated. To clarify the zoonotic risk posed by ExPEC in chicken products and to fill existing knowledge gaps regarding ExPEC zoonosis, we evaluated the prevalence of ExPEC on shell eggs and compared virulence-associated phenotypes between ExPEC and non-ExPEC isolates from both chicken meat and eggs. The prevalence of ExPEC among egg-source isolates was low, i.e., 5/108 (4.7%). Based on combined genotypic and phenotypic screening results, multiple human and avian pathotypes were represented among the chicken-source ExPEC isolates, including avian-pathogenic E. coli (APEC), uropathogenic E. coli (UPEC), neonatal meningitis E. coli (NMEC), and sepsis-associated E. coli (SEPEC), as well as an undefined ExPEC group, which included isolates with fewer virulence factors than the APEC, UPEC, and NMEC isolates. These findings document a substantial prevalence of human-pathogenic ExPEC-associated genes and phenotypes among E. coli isolates from retail chicken products and identify key virulence traits that could be used for screening. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Risk Factors for Salmonella, Shiga Toxin-Producing Escherichia coli and Campylobacter Occurrence in Primary Production of Leafy Greens and Strawberries

PubMed Central

Ceuppens, Siele; Johannessen, Gro S.; Allende, Ana; Tondo, Eduardo César; El-Tahan, Fouad; Sampers, Imca; Jacxsens, Liesbeth; Uyttendaele, Mieke

2015-01-01

The microbiological sanitary quality and safety of leafy greens and strawberries were assessed in the primary production in Belgium, Brazil, Egypt, Norway and Spain by enumeration of Escherichia coli and detection of Salmonella, Shiga toxin-producing E. coli (STEC) and Campylobacter. Water samples were more prone to containing pathogens (54 positives out of 950 analyses) than soil (16/1186) and produce on the field (18/977 for leafy greens and 5/402 for strawberries). The prevalence of pathogens also varied markedly according to the sampling region. Flooding of fields increased the risk considerably, with odds ratio (OR) 10.9 for Salmonella and 7.0 for STEC. A significant association between elevated numbers of generic E. coli and detection of pathogens (OR of 2.3 for STEC and 2.7 for Salmonella) was established. Generic E. coli was found to be a suitable index organism for Salmonella and STEC, but to a lesser extent for Campylobacter. Guidelines on frequency of sampling and threshold values for E. coli in irrigation water may differ from region to region. PMID:26295251

IraL Is an RssB Anti-adaptor That Stabilizes RpoS during Logarithmic Phase Growth in Escherichia coli and Shigella

PubMed Central

Hryckowian, Andrew J.; Battesti, Aurelia; Lemke, Justin J.; Meyer, Zachary C.

2014-01-01

ABSTRACT RpoS (σS), the general stress response sigma factor, directs the expression of genes under a variety of stressful conditions. Control of the cellular σS concentration is critical for appropriately scaled σS-dependent gene expression. One way to maintain appropriate levels of σS is to regulate its stability. Indeed, σS degradation is catalyzed by the ClpXP protease and the recognition of σS by ClpXP depends on the adaptor protein RssB. Three anti-adaptors (IraD, IraM, and IraP) exist in Escherichia coli K-12; each interacts with RssB and inhibits RssB activity under different stress conditions, thereby stabilizing σS. Unlike K-12, some E. coli isolates, including uropathogenic E. coli strain CFT073, show comparable cellular levels of σS during the logarithmic and stationary growth phases, suggesting that there are differences in the regulation of σS levels among E. coli strains. Here, we describe IraL, an RssB anti-adaptor that stabilizes σS during logarithmic phase growth in CFT073 and other E. coli and Shigella strains. By immunoblot analyses, we show that IraL affects the levels and stability of σS during logarithmic phase growth. By computational and PCR-based analyses, we reveal that iraL is found in many E. coli pathotypes but not in laboratory-adapted strains. Finally, by bacterial two-hybrid and copurification analyses, we demonstrate that IraL interacts with RssB by a mechanism distinct from that used by other characterized anti-adaptors. We introduce a fourth RssB anti-adaptor found in E. coli species and suggest that differences in the regulation of σS levels may contribute to host and niche specificity in pathogenic and nonpathogenic E. coli strains. PMID:24865554

The first 30 years of Shiga toxin-producing Escherichia coli in cattle production: Incidence, preharvest ecology, and management

USDA-ARS?s Scientific Manuscript database

Of the 700 serotypes of Escherichia coli, most are commensal; however, some range from mildly to highly pathogenic and can cause death. The disease-causing enterovirulent E. coli are classified as: Enterotoxigenic E. coli (ETEC), Enteropathogenic E. coli (EPEC), Enteroinvasive E. coli (EIEC), and ...

MICROBIAL CONTAMINATION OF STREET VENDED FOODS FROM A UNIVERSITY CAMPUS IN BANGLADESH.

PubMed

Islam, Sufia; Nasrin, Nishat; Rizwan, Farhana; Nahar, Lutfun; Bhowmik, Adity; Esha, Sayma Afrin; Talukder, Kaisar Ali; Akter, Mahmuda; Roy, Ajoy; Ahmed, Muniruddin

2015-05-01

The microbiological quality of street vended food samples from Dhaka, Bangladesh was evaluated. The objective of the study was to identify the presence of common pathogens (Escherichia coli, Shigella spp, Salmonella and Vibrio spp) and to describe the molecular characterization of E coli, a commonly found pathogen in various street foods. Fifty food samples were collected from fixed and mobile vendors from two sampling locations (Mohakhali and Aftabnagar) in Dhaka city, Bangladesh. The tested samples included deep fried and fried snacks; quick lunch items; pickles; fruit chutney; baked items; spicy, sour and hot snacks etc: Juices, tamarind water and plain drinking water were also tested. Sterile polythene bags were used for collecting 200 g of each category of samples. They were tested for the presence of microorganisms following conventional microbiological processes. Biochemical tests followed by serology were done for the confirmation of Shigella and Salmonella. Serological reaction was carried out for confirmation of Vibrio spp. DNA was isolated for the molecular characterization to detect the pathogenic E. coli by polymerase chain reaction (PCR). Out of 50 food samples, six (12%) were confirmed to contain different species of E. coli and Shigella. Molecular characterization of E. coli revealed that three samples were contaminated with enteroaggregative E. coli (EAEC) and one was contaminated with enterotoxigenic E. coli (ETEC). Shigellaflexneri X variant was detected in one food item and Shigella flexneri 2a was found in drinking water. All these enteric pathogens could be the potential cause for foodborne illnesses.

Multiplex quantification of Escherichia coli, Salmonella typhi and Vibrio cholera with three DNA targets in single reaction assay.

PubMed

Jangampalli Adi, Pradeepkiran; Naidu, Jagadish R; Matcha, Bhaskar

2017-09-01

Escherichia coli (E. coli), Salmonella typhi and Vibrio cholera harmful pathogens, which causes various diseases in humans. Rapid diagnosis of bacterial infection is an important for patient management and appropriate therapy during the early phase of the bacterial infected diseases. Among the existing techniques for identifying pathogens were less sensitive and time-consuming processes. In the present study total, 48 clinical 31 blood and 17 urine samples of patients suspected with the infections were collected from SVRR Hospital and used to detect the pathogens. Multiplex polymerase chain reaction (PCR) assay was set to design for the identification of Escherichia coli, Salmonella typhi and Vibrio cholera from the different clinical samples. Rapid diagnosis of Escherichia coli (E. coli), Salmonella and Vibrio cholera pathogens can be done with simultaneously in a single multiplex PCR assay by using specific primers with adjusted PCR conditions. Through this approach, the results represented with out of 31 blood samples 1-15 shows the positive with E. coli and remaining 14 only 11 were correlated with multiplex results of Vibrio cholera, remaining the urine samples all are positive with 17 samples correlate with the Salmonella typhi. Through the high specificity benefits of excellent sensitivity, with high resolution and reproducibility. This method of results proved and illustrates the best potential system for diagnosing the infectious disease with modern trendy. Copyright © 2017 Elsevier Ltd. All rights reserved.

Sensitive detection of viable Escherichia coli O157:H7 from foods using a luciferase-reporter phage phiV10lux.

PubMed

Kim, Jinwoo; Kim, Minsik; Kim, Seongmi; Ryu, Sangryeol

2017-08-02

Escherichia coli O157:H7, a major foodborne pathogen, is a major public health concern associated with life-threatening diseases such as hemolytic uremic syndrome. To alleviate this burden, a sensitive and rapid system is required to detect this pathogen in various kinds of foods. Herein, we propose a phage-based pathogen detection method to replace laborious and time-consuming conventional methods. We engineered an E. coli O157:H7-specific phage phiV10 to rapidly and sensitively detect this notorious pathogen. The luxCDABE operon was introduced into the phiV10 genome and allowed the engineered phage phiV10lux to generate bioluminescence proportional to the number of viable E. coli O157:H7 cells without any substrate addition. The phage phiV10lux was able to detect at least 1CFU/ml of E. coli O157:H7 in a pure culture within 40min after 5h of pre-incubation. In artificially contaminated romaine lettuce, apple juice (pH3.51), and ground beef, the reporter phage could detect approximately 10CFU/cm 2 , 13CFU/ml, and 17CFU/g of E. coli O157:H7, respectively. Taken together, the constructed reporter phage phiV10lux could be applied as a powerful tool for rapid and sensitive detection of live E. coli O157:H7 in foods. Copyright © 2017 Elsevier B.V. All rights reserved.

Escherichia coli as other Enterobacteriaceae: food poisoning and health effects

USDA-ARS?s Scientific Manuscript database

Many Escherichia coli strains are harmless, and they are an important commensal in the intestinal microflora; however, pathogenic strains also exist. The pathogenic strains can be divided into diarrhea-inducing strains and strains that reside in the intestines but only cause disease in bodily sites...

The seaweed fly (Coelopidae) can facilitate environmental survival and transmission of E. coli O157 at sandy beaches.

PubMed

Swinscoe, Isobel; Oliver, David M; Gilburn, Andre S; Quilliam, Richard S

2018-06-19

The sustainable management of recreational beaches is essential for minimising risk of human exposure to microbial pathogens whilst simultaneously maintaining valuable ecosystem services. Decaying seaweed on public beaches is gaining recognition as a substrate for microbial contamination, and is a potentially significant reservoir for human pathogens in close proximity to beach users. Closely associated with beds of decaying seaweed are dense populations of the seaweed fly (Coelopidae), which could influence the spatio-temporal fate of seaweed-associated human pathogens within beach environments. Replicated mesocosms containing seaweed inoculated with a bioluminescent strain of the zoonotic pathogen E. coli O157:H7, were used to determine the effects of two seaweed flies, Coelopa frigida and C. pilipes, on E. coli O157:H7 survival dynamics. Multiple generations of seaweed flies and their larvae significantly enhanced persistence of E. coli O157:H7 in simulated wrack habitats, demonstrating that both female and male C. frigida flies are capable of transferring E. coli O157:H7 between individual wrack beds and into the sand. Adult fly faeces can contain significant concentrations of E. coli O157:H7, which suggests they are capable of acting as biological vectors and bridge hosts between wrack habitats and other seaweed fly populations, and facilitate the persistence and dispersal of E. coli O157:H7 in sandy beach environments. This study provides the first evidence that seaweed fly populations inhabiting natural wrack beds contaminated with the human pathogen E. coli O157:H7 have the capacity to amplify the hazard source, and therefore potential transmission risk, to beach users exposed to seaweed and sand in the intertidal zone. The risk to public health from seaweed flies and decaying wrack beds is usually limited by human avoidance behaviour; however, seaweed fly migration and nuisance inland plagues in urban areas could increase human exposure routes beyond the beach environment. Copyright © 2018 Elsevier Ltd. All rights reserved.

The Intimin-Like Protein FdeC Is Regulated by H-NS and Temperature in Enterohemorrhagic Escherichia coli

PubMed Central

Easton, Donna M.; Allsopp, Luke P.; Phan, Minh-Duy; Moriel, Danilo Gomes; Goh, Guan Kai; Beatson, Scott A.; Mahony, Timothy J.; Cobbold, Rowland N.

2014-01-01

Enterohemorrhagic Escherichia coli (EHEC) is a Shiga-toxigenic pathogen capable of inducing severe forms of enteritis (e.g., hemorrhagic colitis) and extraintestinal sequelae (e.g., hemolytic-uremic syndrome). The molecular basis of colonization of human and animal hosts by EHEC is not yet completely understood, and an improved understanding of EHEC mucosal adherence may lead to the development of interventions that could disrupt host colonization. FdeC, also referred to by its IHE3034 locus tag ECOK1_0290, is an intimin-like protein that was recently shown to contribute to kidney colonization in a mouse urinary tract infection model. The expression of FdeC is tightly regulated in vitro, and FdeC shows promise as a vaccine candidate against extraintestinal E. coli strains. In this study, we characterized the prevalence, regulation, and function of fdeC in EHEC. We showed that the fdeC gene is conserved in both O157 and non-O157 EHEC and encodes a protein that is expressed at the cell surface and promotes biofilm formation under continuous-flow conditions in a recombinant E. coli strain background. We also identified culture conditions under which FdeC is expressed and showed that minor alterations of these conditions, such as changes in temperature, can significantly alter the level of FdeC expression. Additionally, we demonstrated that the transcription of the fdeC gene is repressed by the global regulator H-NS. Taken together, our data suggest a role for FdeC in EHEC when it grows at temperatures above 37°C, a condition relevant to its specialized niche at the rectoanal junctions of cattle. PMID:25239893

Multiepitope fusion antigen induces broadly protective antibodies that prevent adherence of Escherichia coli strains expressing colonization factor antigen I (CFA/I), CFA/II, and CFA/IV.

PubMed

Ruan, Xiaosai; Knudsen, David E; Wollenberg, Katie M; Sack, David A; Zhang, Weiping

2014-02-01

Diarrhea is the second leading cause of death in children younger than 5 years and continues to be a major threat to global health. Enterotoxigenic Escherichia coli (ETEC) strains are the most common bacteria causing diarrhea in developing countries. ETEC strains are able to attach to host small intestinal epithelial cells by using bacterial colonization factor antigen (CFA) adhesins. This attachment helps to initiate the diarrheal disease. Vaccines that induce antiadhesin immunity to block adherence of ETEC strains that express immunologically heterogeneous CFA adhesins are expected to protect against ETEC diarrhea. In this study, we created a CFA multiepitope fusion antigen (MEFA) carrying representative epitopes of CFA/I, CFA/II (CS1, CS2, and CS3), and CFA/IV (CS4, CS5, and CS6), examined its immunogenicity in mice, and assessed the potential of this MEFA as an antiadhesin vaccine against ETEC. Mice intraperitoneally immunized with this CFA MEFA exhibited no adverse effects and developed immune responses to CFA/I, CFA/II, and CFA/IV adhesins. Moreover, after incubation with serum of the immunized mice, ETEC or E. coli strains expressing CFA/I, CFA/II, or CFA/IV adhesins were significantly inhibited in adherence to Caco-2 cells. Our results indicated this CFA MEFA elicited antibodies that not only cross-reacted to CFA/I, CFA/II and CFA/IV adhesins but also broadly inhibited adherence of E. coli strains expressing these seven adhesins and suggested that this CFA MEFA could be a candidate to induce broad-spectrum antiadhesin protection against ETEC diarrhea. Additionally, this antigen construction approach (creating an MEFA) may be generally used in vaccine development against heterogenic pathogens.

Occurrence and characterization of Shiga toxin-producing Escherichia coli O157:H7 and other non-sorbitol-fermenting E. coli in cattle and humans in urban areas of Morogoro, Tanzania.

PubMed

Lupindu, Athumani M; Olsen, John E; Ngowi, Helena A; Msoffe, Peter L M; Mtambo, Madundo M; Scheutz, Flemming; Dalsgaard, Anders

2014-07-01

Escherichia coli strains such as Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli, enterotoxigenic, attaching, and effacing E. coli, and enteroinvasive E. coli cause diarrhea in humans. Although other serotypes exist, the most commonly reported STEC in outbreaks is O157:H7. A cross-sectional study was conducted to isolate and characterize non-sorbitol-fermenting (NSF) E. coli O157:H7 from urban and periurban livestock settings of Morogoro, Tanzania. Human stool, cattle feces, and soil and water samples were collected. Observations and questionnaire interview studies were used to gather information about cattle and manure management practices in the study area. E. coli were isolated on sorbitol MacConkey agar and characterized by conventional biochemical tests. Out of 1049 samples, 143 (13.7%) yielded NSF E. coli. Serological and antimicrobial tests and molecular typing were performed to NSF E. coli isolates. These procedures detected 10 (7%) pathogenic E. coli including STEC (n=7), enteropathogenic E. coli (EPEC) (n=2), and attaching and effacing E. coli (A/EEC) (n=1) strains. The STEC strains had the ability to produce VT1 and different VT2 toxin subtypes that caused cytopathic effects on Vero cells. The prevalence of STEC in cattle was 1.6%, out of which 0.9% was serotype O157:H7 and the overall prevalence of diarrheagenic E. coli in cattle was 2.2%. The serotypes O157:H7, O142:H34, O113:H21, O+:H-, O+:H16, and O25:H4 were identified. One ESBL-producing isolate showed the MLST type ST131. To our knowledge, this is the first finding in Tanzania of this recently emerged worldwide pandemic clonal group, causing widespread antimicrobial-resistant infections, and adds knowledge of the geographical distribution of ST131. Cattle manure was indiscriminately deposited within residential areas, and there was direct contact between humans and cattle feces during manure handling. Cattle and manure management practices expose humans, animals, and the environment to pathogenic E. coli and other manure-borne pathogens. Therefore, there is a need to improve manure management practices in urban and periurban areas to prevent pathogen spread and associated human health risks.

Development and evaluation of a decision-supporting model for identifying the source location of microbial intrusions in real gravity sewer systems.

PubMed

Kim, Minyoung; Choi, Christopher Y; Gerba, Charles P

2013-09-01

Assuming a scenario of a hypothetical pathogenic outbreak, we aimed this study at developing a decision-support model for identifying the location of the pathogenic intrusion as a means of facilitating rapid detection and efficient containment. The developed model was applied to a real sewer system (the Campbell wash basin in Tucson, AZ) in order to validate its feasibility. The basin under investigation was divided into 14 sub-basins. The geometric information associated with the sewer network was digitized using GIS (Geological Information System) and imported into an urban sewer network simulation model to generate microbial breakthrough curves at the outlet. A pre-defined amount of Escherichia coli (E. coli), which is an indicator of fecal coliform bacteria, was hypothetically introduced into 56 manholes (four in each sub-basin, chosen at random), and a total of 56 breakthrough curves of E. coli were generated using the simulation model at the outlet. Transport patterns were classified depending upon the location of the injection site (manhole), various known characteristics (peak concentration and time, pipe length, travel time, etc.) extracted from each E. coli breakthrough curve and the layout of sewer network. Using this information, we back-predicted the injection location once an E. coli intrusion was detected at a monitoring site using Artificial Neural Networks (ANNs). The results showed that ANNs identified the location of the injection sites with 57% accuracy; ANNs correctly recognized eight out of fourteen expressions with relying on data from a single detection sensor. Increasing the available sensors within the basin significantly improved the accuracy of the simulation results (from 57% to 100%). Copyright © 2013 Elsevier Ltd. All rights reserved.

Pathogen transport and fate modeling in the Upper Salem River Watershed using SWAT model.

PubMed

Niazi, Mehran; Obropta, Christopher; Miskewitz, Robert

2015-03-15

Simulation of the fate and transport of pathogen contamination was conducted with SWAT for the Upper Salem River Watershed, located in Salem County, New Jersey. This watershed is 37 km(2) and land uses are predominantly agricultural. The watershed drains to a 32 km stretch of the Salem River upstream of the head of tide. This strech is identified on the 303(d) list as impaired for pathogens. The overall goal of this research was to use SWAT as a tool to help to better understand how two pathogen indicators (Escherichia coli and fecal coliform) are transported throughout the watershed, by determining the model parameters that control the fate and transport of these two indicator species. This effort was the first watershed modeling attempt with SWAT to successfully simulate E. coli and fecal coliform simultaneously. Sensitivity analysis has been performed for flow as well as fecal coliform and E. coli. Hydrologic calibration at six sampling locations indicate that the model provides a "good" prediction of watershed outlet flow (E = 0.69) while at certain upstream calibration locations predictions are less representative (0.32 < E < 0.70). Monthly calibration and validation of the pathogen transport and fate model was conducted for both fecal coliform (0.07 < E < 0.47 and -0.94 < E < 0.33) and E. coli (0.03 < E < 0.39 and -0.81 < E < 0.31) for the six sampling points. The fit of the model compared favorably with many similar pathogen modeling efforts. The research contributes new knowledge in E. coli and fecal coliform modeling and will help increase the understanding of sensitivity analysis and pathogen modeling with SWAT at the watershed scale. Copyright © 2014 Elsevier Ltd. All rights reserved.

Main Concerns of Pathogenic Microorganisms in Meat

NASA Astrophysics Data System (ADS)

Nørrung, Birgit; Andersen, Jens Kirk; Buncic, Sava

Although various foods can serve as sources of foodborne illness, meat and meat products are important sources of human infections with a variety of foodborne pathogens, i.e. Salmonella spp., Campylobacter jejuni/coli, Yersinia enterocolitica, Verotoxigenic E. coli and, to some extent, Listeria monocytogenes. All these may be harboured in the gastrointestinal tract of food-producing animals. The most frequent chain of events leading to meat-borne illness involves food animals, which are healthy carriers of the pathogens that are subsequently transferred to humans through production, handling and consumption of meat and meat products. Occurrences of Salmonella spp., C. jejuni/coli, Y. enterocolitica and Verotoxigenic E. coli in fresh red meat vary relatively widely, although most often are between 1 and 10%, depending on a range of factors including the organism, geographical factors, farming and/or meat production practices.

An Escherichia coli O157-specific engineered pyocin prevents and ameliorates infection by E. coli O157:H7 in an animal model of diarrheal disease.

PubMed

Ritchie, Jennifer M; Greenwich, Jennifer L; Davis, Brigid M; Bronson, Roderick T; Gebhart, Dana; Williams, Steven R; Martin, David; Scholl, Dean; Waldor, Matthew K

2011-12-01

AvR2-V10.3 is an engineered R-type pyocin that specifically kills Escherichia coli O157, an enteric pathogen that is a major cause of food-borne diarrheal disease. New therapeutics to counteract E. coli O157 are needed, as currently available antibiotics can exacerbate the consequences of infection. We show here that orogastric administration of AvR2-V10.3 can prevent or ameliorate E. coli O157:H7-induced diarrhea and intestinal inflammation in an infant rabbit model of infection when the compound is administered either in a postexposure prophylactic regimen or after the onset of symptoms. Notably, administration of AvR2-V10.3 also reduces bacterial carriage and fecal shedding of this pathogen. Our findings support the further development of pathogen-specific R-type pyocins as a way to treat enteric infections.

Prevalence and Genomic Characterization of Escherichia coli O157:H7 in Cow-Calf Herds throughout California.

PubMed

Worley, Jay N; Flores, Kristopher A; Yang, Xun; Chase, Jennifer A; Cao, Guojie; Tang, Shuai; Meng, Jianghong; Atwill, Edward R

2017-08-15

Escherichia coli serotype O157:H7 is a zoonotic food- and waterborne bacterial pathogen that causes a high hospitalization rate and can cause life-threatening complications. Increasingly, E. coli O157:H7 infections appear to originate from fresh produce. Ruminants, such as cattle, are a prominent reservoir of E. coli O157:H7 in the United States. California is one of the most agriculturally productive regions in the world for fresh produce, beef, and milk. The close proximity of fresh produce and cattle presents food safety challenges on a uniquely large scale. We performed a survey of E. coli O157:H7 on 20 farms in California to observe the regional diversity and prevalence of E. coli O157:H7. Isolates were obtained from enrichment cultures of cow feces. Some farms were sampled on two dates. Genomes from isolates were sequenced to determine their relatedness and pathogenic potential. E. coli O157:H7 was isolated from approximately half of the farms. The point prevalence of E. coli O157:H7 on farms was highly variable, ranging from zero to nearly 90%. Within farms, generally one or a few lineages were found, even when the rate of isolation was high. On farms with high isolation rates, a single clonal lineage accounted for most of the isolates. Farms that were visited months after the first visit might have had the same lineages of E. coli O157:H7. Strains of E. coli O157:H7 may be persistent for months on farms. IMPORTANCE This survey of 20 cow-calf operations from different regions of California provides an in depth look at resident Escherichia coli O157:H7 populations at the molecular level. E. coli O157:H7 is found to have a highly variable prevalence, and with whole-genome sequencing, high prevalences in herds were found to be due to a single lineage shed from multiple cows. Few repeat lineages were found between farms in this area; therefore, we predict that E. coli O157:H7 has significant diversity in this area beyond what is detected in this survey. All isolates from this study were found to have pathogenic potential based on the presence of key virulence gene sequences. This represents a novel insight into pathogen diversity within a single subtype and will inform future attempts to survey regional pathogen populations. Copyright © 2017 American Society for Microbiology.

Prevalence and Genomic Characterization of Escherichia coli O157:H7 in Cow-Calf Herds throughout California

PubMed Central

Worley, Jay N.; Flores, Kristopher A.; Yang, Xun; Chase, Jennifer A.; Cao, Guojie; Tang, Shuai; Meng, Jianghong

2017-01-01

ABSTRACT Escherichia coli serotype O157:H7 is a zoonotic food- and waterborne bacterial pathogen that causes a high hospitalization rate and can cause life-threatening complications. Increasingly, E. coli O157:H7 infections appear to originate from fresh produce. Ruminants, such as cattle, are a prominent reservoir of E. coli O157:H7 in the United States. California is one of the most agriculturally productive regions in the world for fresh produce, beef, and milk. The close proximity of fresh produce and cattle presents food safety challenges on a uniquely large scale. We performed a survey of E. coli O157:H7 on 20 farms in California to observe the regional diversity and prevalence of E. coli O157:H7. Isolates were obtained from enrichment cultures of cow feces. Some farms were sampled on two dates. Genomes from isolates were sequenced to determine their relatedness and pathogenic potential. E. coli O157:H7 was isolated from approximately half of the farms. The point prevalence of E. coli O157:H7 on farms was highly variable, ranging from zero to nearly 90%. Within farms, generally one or a few lineages were found, even when the rate of isolation was high. On farms with high isolation rates, a single clonal lineage accounted for most of the isolates. Farms that were visited months after the first visit might have had the same lineages of E. coli O157:H7. Strains of E. coli O157:H7 may be persistent for months on farms. IMPORTANCE This survey of 20 cow-calf operations from different regions of California provides an in depth look at resident Escherichia coli O157:H7 populations at the molecular level. E. coli O157:H7 is found to have a highly variable prevalence, and with whole-genome sequencing, high prevalences in herds were found to be due to a single lineage shed from multiple cows. Few repeat lineages were found between farms in this area; therefore, we predict that E. coli O157:H7 has significant diversity in this area beyond what is detected in this survey. All isolates from this study were found to have pathogenic potential based on the presence of key virulence gene sequences. This represents a novel insight into pathogen diversity within a single subtype and will inform future attempts to survey regional pathogen populations. PMID:28550057

The first 30 years of Shiga toxin-producing Escherichia coli in cattle production: Preharvest intervention strategies

USDA-ARS?s Scientific Manuscript database

Cattle are naturally colonized with foodborne pathogenic bacteria, including Shiga toxin-producing E. coli (STEC). While these foodborne pathogens threaten food safety, they also cause human illnesses via cross contamination of other foods, the water supply, and via direct animal contact. Pre-harv...

Potential Pathogenicity and Host Range of Extended-Spectrum β-Lactamase-Producing Escherichia coli Isolates from Healthy Poultry ▿

PubMed Central

Bortolaia, Valeria; Larsen, Jesper; Damborg, Peter; Guardabassi, Luca

2011-01-01

Thirty of 33 epidemiologically unrelated extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates from healthy poultry lacked the virulence genes commonly associated with human-pathogenic strains. The main zoonotic risk is associated with the broad host range of avian E. coli belonging to sequence type complex 10 and of IncN and IncI1 plasmids carrying blaCTX-M or blaSHV. PMID:21705531

Fate of Escherichia coli O26 in Corn Silage Experimentally Contaminated at Ensiling, at Silo Opening, or after Aerobic Exposure, and Protective Effect of Various Bacterial Inoculants▿

PubMed Central

Dunière, Lysiane; Gleizal, Audrey; Chaucheyras-Durand, Frédérique; Chevallier, Isabelle; Thévenot-Sergentet, Delphine

2011-01-01

Shiga toxin-producing Escherichia coli (STEC) strains are responsible for human illness. Ruminants are recognized as a major reservoir of STEC, and animal feeds, such as silages, have been pointed out as a possible vehicle for the spread of STEC. The present study aimed to monitor the fate of pathogenic E. coli O26 strains in corn material experimentally inoculated (105 CFU/g) during ensiling, just after silo opening, and after several days of aerobic exposure. The addition of 3 bacterial inoculants, Propionibacterium sp., Lactobacillus buchneri, and Leuconostoc mesenteroides (106 CFU/g), was evaluated for their abilities to control these pathogens. The results showed that E. coli O26 could not survive in corn silage 5 days postensiling, and the 3 inoculants tested did not modify the fate of pathogen survival during ensiling. In the case of direct contamination at silo opening, E. coli O26 could be totally eradicated from corn silage previously inoculated with Leuconostoc mesenteroides. The combination of proper ensiling techniques and the utilization of selected bacterial inoculants appears to represent a good strategy to guarantee nutritional qualities of cattle feed while at the same time limiting the entry of pathogenic E. coli into the epidemiological cycle to improve the microbial safety of the food chain. PMID:21984243

[Analysis of Pathogenic Bacteria in Reclaimed Water and Impact of UV Disinfection on the Removal of Pathogenic Bacteria].

PubMed

Jing, Ming; Wang, Lei

2016-02-15

In the study, 454-pyrosequencing technology was employed to investigate the species of pathogenic bacteria and the proportion of each pathogen in secondary effluent. Culture-based, qPCR and Q-RT-PCR methods were employed to analyze the removal of indicator (E. coli) and pathogen (Salmonella and Mycobacterium) by ultraviolet (UV) disinfection at a dose of 60 mJ x Cm(-2). The results showed that 11 kinds of pathogenic bacteria were found and the most abundant potentially pathogenic bacteria in the secondary effluent were affiliated with the genera of Clostridium (2.96%), Arcobacter (0.82%) and Mycobacterium (0.36%). 99.9% of culturable E. coli and Salmonella were removed by UV disinfection (60 mJ x cm(-2), however, less than 90% of culturable Mycobacterium were removed. The removal efficiencies of viable E. coli, Salmonella and Mycobacterium were low. Q-RT-PCR seemed to be a promising method for evaluating viable microorganisms in samples. Besides, pathogenic bacteria entered into VBNC state at a UV dose of 60 mJ x cm(-2). Other advanced treatment processes were needed to ensure safe utilization of reclaimed water.

Optimization of multi-epitopic HIV-1 recombinant protein expression in prokaryote system and conjugation to mouse DEC-205 monoclonal antibody: implication for in-vivo targeted delivery of dendritic cells

PubMed Central

Rahimi, Roghayeh; Ebtekar, Massoumeh; Moazzeni, Seyed Mohammad; Mostafaie, Ali; Mahdavi, Mehdi

2015-01-01

Objective(s): Multi-epitopic protein vaccines and direction of vaccine delivery to dendritic cells (DCs) are promising approaches for enhancing immune responses against mutable pathogens. Escherichia coli is current host for expression of recombinant proteins, and it is important to optimize expression condition. The aim of this study was the optimization of multi-epitopic HIV-1 tat/pol/gag/env recombinant protein (HIVtop4) expression by E. coli and conjugation of purified protein to anti DEC-205 monoclonal antibody as candidate vaccine. Materials and Methods: In this study, expression was induced in BL21 (DE3) E. coli cells by optimization of induction condition, post induction incubation time, temperature and culture medium formula. Some culture mediums were used for cell culture, and isopropyl-beta-D-thiogalactopyranoside was used for induction of expression. Protein was purified by Ni-NTA column chromatography and confirmed against anti-His antibody in western-blotting. To exploit DCs properties for immunization purposes, recombinant protein chemically coupled to αDEC-205 monoclonal antibody and confirmed against anti-His antibody in western-blotting. Results: The optimum condition for expression was 1 mM IPTG during 4 hr cultures in 2XYT medium, and final protein produced in soluble form. Conjugation of purified protein to αDEC-205 antibody resulted in smears of protein: antibodies conjugate in different molecular weights. Conclusion: The best cultivation condition for production of HIVtop4 protein is induction by 1 mM IPTG during 4 hr in 2XYT medium. The final concentration of purified protein was 500 µg/ml. PMID:25810888

[Plasticity of bacterial genomes: pathogenicity islands and the locus of enterocyte effacement (LEE)].

PubMed

Kirsch, Petra; Jores, Jörg; Wieler, Lothar H

2004-01-01

Many bacterial virulence attributes, like toxins, adhesins, invasins, iron uptake systems, are encoded within specific regions of the bacterial genome. These in size varying regions are termed pathogenicity islands (PAIs) since they confer pathogenic properties to the respective micro-organism. Per definition PAIs are exclusively found in pathogenic strains and are often inserted near transfer-RNA genes. Nevertheless, non-pathogenic bacteria also possess foreign DNA elements that confer advantageous features, leading to improved fitness. These additional DNA elements as well as PAIs are termed genomic islands and were acquired during bacterial evolution. Significant G+C content deviation in pathogenicity islands with respect to the rest of the genome, the presence of direct repeat sequences at the flanking regions, the presence of integrase gene determinants as other mobility features,the particular insertion site (tRNA gene) as well as the observed genetic instability suggests that pathogenicity islands were acquired by horizontal gene transfer. PAIs are the fascinating proof of the plasticity of bacterial genomes. PAIs were originally described in human pathogenic Escherichia (E.) coli strains. In the meantime PAIs have been found in various pathogenic bacteria of humans, animals and even plants. The Locus of Enterocyte Effacement (LEE) is one particular widely distributed PAI of E coli. In addition, it also confers pathogenicity to the related species Citrobacter (C.) rodentium and Escherichia (E.) alvei. The LEE is an important virulence feature of several animal pathogens. It is an obligate PAI of all animal and human enteropathogenic E. coli (EPEC), and most enterohaemorrhegic E. coli (EHEC) also harbor the LEE. The LEE encodes a type III secretion system, an adhesion (intimin) that mediates the intimate contact between the bacterium and the epithelial cell, as well as various proteins which are secreted via the type III secretion system. The LEE encoded virulence features are responsible for the formation of so called attaching and effacing (AE) lesions in the intestinal epithelium. Due to its wide distribution in animal pathogens, LEE encoded antigens are suitable vaccine antigens. Acquisition and structure of the LEE pathogenicity island is the crucial point of numerous investigations. However, the evolution of the LEE, its origin and further spread in E. coli, are far from being resolved.

A rapid two dot filter assay for the detection of E. coli O157 in water samples.

PubMed

Kamma, Sujatha; Tang, Lily; Leung, Kelvin; Ashton, Edie; Newman, Norman; Suresh, Mavanur R

2008-07-31

E. coli O157:H7 is an enterohemorrhagic bacteria that cause deadly water-borne infections implicated in outbreaks of a wide spectrum of human gastrointestinal diseases. It is therefore important to have a rapid convenient, simple and sensitive range of detection of E. coli O157:H7. A new E. coli O157 MAb designated P124 was developed for ultrasensitive detection of E. coli O157 in water, apple juice and beef for routine use. A prototype filter dot assay was designed with anti-E. coli O157 MAb bound to 0.2 microm nitrocellulose filter disk as the capture antibody. A 100 ml water sample spiked with 1-50 CFU of E. coli O157 either in the presence or absence of other non-specific bacteria were filtered for capture of the pathogen on the antibody coated nitrocellulose disk. The detection of the pathogen was successfully accomplished by the same antibody both as a capture and detecting antibody as a homosandwich. In a non-enriched format, detection of E. coli was possible with a sensitivity of 2500 CFU/100 ml. Ultrasensitive detection of ~1 CFU/100 ml sample could be achieved by a prior pathogen enrichment step before the addition of the labeled antibody. The design of this diagnostic test is based on the common architecture of all bacteria, viruses and spores, namely the manifestation of repeat lipopolysaccharide epitopes on the surface. We have developed an easy-to-use two dot visual filter assay for translation into current water testing in public health laboratories to detect E. coli O157:H7. In a 5 h assay approximately 1 CFU and approximately 5 CFU of E. coli O157 could be detected in 100 ml of water or juice and lake samples respectively. This simple homosandwich enrichment strategy can also be used to detect low levels of other water-borne pathogens.

Interactions of the Hindgut Mucosa-Associated Microbiome with Its Host Regulate Shedding of Escherichia coli O157:H7 by Cattle.

PubMed

Wang, Ou; McAllister, Tim A; Plastow, Graham; Stanford, Kim; Selinger, Brent; Guan, Le Luo

2018-01-01

Cattle are the primary carrier of Escherichia coli O157:H7, a foodborne human pathogen, and those shedding >10 4 CFU/gram of feces of E. coli O157:H7 are defined as supershedders (SS). This study investigated the rectoanal junction (RAJ) mucosa-associated microbiota and its relationship with host gene expression in SS and in cattle from which E. coli O157:H7 was not detected (nonshedders [NS]), aiming to elucidate the mechanisms involved in supershedding. In total, 14 phyla, 66 families, and 101 genera of RAJ mucosa-associated bacteria were identified and Firmicutes (61.5 ± 7.5%), Bacteroidetes (27.9 ± 6.4%), and Proteobacteria (5.5 ± 2.1%) were the predominant phyla. Differential abundance analysis of operational taxonomic units (OTUs) identified 2 OTUs unique to SS which were members of Bacteroides and Clostridium and 7 OTUs unique to NS which were members of Coprococcus , Prevotella , Clostridium , and Paludibacter Differential abundance analysis of predicted microbial functions (using PICRUSt [phylogenetic investigation of communities by reconstruction of unobserved states]) revealed that 3 pathways had higher abundance (log 2 fold change, 0.10 to 0.23) whereas 12 pathways had lower abundance (log 2 fold change, -0.36 to -0.20) in SS. In addition, we identified significant correlations between expression of 19 differentially expressed genes and the relative abundance of predicted microbial functions, including nucleic acid polymerization and carbohydrate and amino acid metabolism. Our findings suggest that differences in RAJ microbiota at both the compositional and functional levels may be associated with E. coli O157:H7 supershedding and that certain microbial groups and microbial functions may influence RAJ physiology of SS by affecting host gene expression. IMPORTANCE Cattle with fecal E. coli O157:H7 at >10 4 CFU per gram of feces have been defined as the supershedders, and they are responsible for the most of the E. coli O157:H7 spread into farm environment. Currently, no method is available for beef producers to eliminate shedding of E. coli O157:H7 in cattle, and the lack of information about the mechanisms of supershedding greatly impedes the development of effective methods. This study investigated the role of the rectoanal junction (RAJ) mucosa-associated microbiome in E. coli O157:H7 shedding, and our results indicated that the compositions and functions of RAJ microbiota differed between supershedders and nonshedders. The identified relationship between the differentially abundant microbes and 19 previously identified differentially expressed genes suggests the role of host-microbial interactions involved in E. coli O157:H7 supershedding. Our findings provide a fundamental understanding of the supershedding phenomenon which is essential for the development of strategies, such as the use of directly fed microbials, to reduce E. coli O157:H7 shedding in cattle. Copyright © 2017 American Society for Microbiology.

Virulence characteristics and genetic affinities of multiple drug resistant uropathogenic Escherichia coli from a semi urban locality in India.

PubMed

Jadhav, Savita; Hussain, Arif; Devi, Savita; Kumar, Ashutosh; Parveen, Sana; Gandham, Nageshwari; Wieler, Lothar H; Ewers, Christa; Ahmed, Niyaz

2011-03-25

Extraintestinal pathogenic Escherichia coli (ExPEC) are of significant health concern. The emergence of drug resistant E. coli with high virulence potential is alarming. Lack of sufficient data on transmission dynamics, virulence spectrum and antimicrobial resistance of certain pathogens such as the uropathogenic E. coli (UPEC) from countries with high infection burden, such as India, hinders the infection control and management efforts. In this study, we extensively genotyped and phenotyped a collection of 150 UPEC obtained from patients belonging to a semi-urban, industrialized setting near Pune, India. The isolates representing different clinical categories were analyzed in comparison with 50 commensal E. coli isolates from India as well as 50 ExPEC strains from Germany. Virulent strains were identified based on hemolysis, haemagglutination, cell surface hydrophobicity, serum bactericidal activity as well as with the help of O serotyping. We generated antimicrobial resistance profiles for all the clinical isolates and carried out phylogenetic analysis based on repetitive extragenic palindromic (rep)-PCR. E. coli from urinary tract infection cases expressed higher percentages of type I (45%) and P fimbriae (40%) when compared to fecal isolates (25% and 8% respectively). Hemolytic group comprised of 60% of UPEC and only 2% of E. coli from feces. Additionally, we found that serum resistance and cell surface hydrophobicity were not significantly (p = 0.16/p = 0.51) associated with UPEC from clinical cases. Moreover, clinical isolates exhibited highest resistance against amoxicillin (67.3%) and least against nitrofurantoin (57.3%). We also observed that 31.3% of UPEC were extended-spectrum beta-lactamase (ESBL) producers belonging to serotype O25, of which four were also positive for O25b subgroup that is linked to B2-O25b-ST131-CTX-M-15 virulent/multiresistant type. Furthermore, isolates from India and Germany (as well as global sources) were found to be genetically distinct with no evidence to espouse expansion of E. coli from India to the west or vice-versa.

Genome-Wide Mapping of Cystitis Due to Streptococcus agalactiae and Escherichia coli in Mice Identifies a Unique Bladder Transcriptome That Signifies Pathogen-Specific Antimicrobial Defense against Urinary Tract Infection

PubMed Central

Tan, Chee K.; Carey, Alison J.; Cui, Xiangqin; Webb, Richard I.; Ipe, Deepak; Crowley, Michael; Cripps, Allan W.; Benjamin, William H.; Ulett, Kimberly B.; Schembri, Mark A.

2012-01-01

The most common causes of urinary tract infections (UTIs) are Gram-negative pathogens such as Escherichia coli; however, Gram-positive organisms, including Streptococcus agalactiae, or group B streptococcus (GBS), also cause UTI. In GBS infection, UTI progresses to cystitis once the bacteria colonize the bladder, but the host responses triggered in the bladder immediately following infection are largely unknown. Here, we used genome-wide expression profiling to map the bladder transcriptome of GBS UTI in mice infected transurethrally with uropathogenic GBS that was cultured from a 35-year-old women with cystitis. RNA from bladders was applied to Affymetrix Gene-1.0ST microarrays; quantitative reverse transcriptase PCR (qRT-PCR) was used to analyze selected gene responses identified in array data sets. A surprisingly small significant-gene list of 172 genes was identified at 24 h; this compared to 2,507 genes identified in a side-by-side comparison with uropathogenic E. coli (UPEC). No genes exhibited significantly altered expression at 2 h in GBS-infected mice according to arrays despite high bladder bacterial loads at this early time point. The absence of a marked early host response to GBS juxtaposed with broad-based bladder responses activated by UPEC at 2 h. Bioinformatics analyses, including integrative system-level network mapping, revealed multiple activated biological pathways in the GBS bladder transcriptome that regulate leukocyte activation, inflammation, apoptosis, and cytokine-chemokine biosynthesis. These findings define a novel, minimalistic type of bladder host response triggered by GBS UTI, which comprises collective antimicrobial pathways that differ dramatically from those activated by UPEC. Overall, this study emphasizes the unique nature of bladder immune activation mechanisms triggered by distinct uropathogens. PMID:22733575

Survival of pathogenic bacteria in compost with special reference to Escherichia coli.

PubMed

Gong, Chun-ming; Koichi, Inoue; Shunji, Inanaga; Takashi, Someya

2005-01-01

Application of compost in agricultural practice could potentially cause contamination of foodstuffs with pathogenic bacteria such as Escherichia coli O157:H7 (E. Coli O157). We investigated pathogenic bacteria in compost collected from the compost facilities, and evaluated the survival of E. coli K12 and O157 in laboratory experiments. Out of 19 compost product samples, coliform bacteria and salmonella were detected in 7 and 3 samples respectively. The number of coliform bacteria was 1.8 x 10(2) to 2.5 x 10(6) CFU/g dw and that of salmonella was 4.2 x 10(1) to 6.0 x 10(3) CFU/g dw. Moreover, coliform bacteria, fecal coliform, E. coli and salmonella were detected during composting at 54 degrees C to 67 degrees C. The results indicated that moisture content was a very important factor to the heat sensitivity of pathogenic bacteria in compost, E. coil in compost of high moisture content was more sensitive than that in compost of low moisture content, cells harvested in logarithmic phase was more sensitive than these in stationary phase, and E. coli K12 was more sensitive than E. coli O157. Based on the D values, the lethal time of E. coli K12 and O157 from l0(8) to 10(0) CFU/g dw were 16.3 and 28.8 min, respectively, at 60 degrees C in compost with 40% moisture content. However, some E. coil cells survived in composting process at 54 degrees C to 67 degrees C. Water potential (low moisture content) and physiological aspects of bacteria (stationary phase) could explain only in part of the prolonged survival of E. coil in compost, and there should be some other factors that are conducive to bacterial survival in compost.

PREVALENCE AND CHARACTERISTICS OF ESCHERICHIA COLI AND SALMONELLA SPP. IN THE FECES OF WILD URBAN NORWAY AND BLACK RATS (RATTUS NORVEGICUS AND RATTUS RATTUS) FROM AN INNER-CITY NEIGHBORHOOD OF VANCOUVER, CANADA.

PubMed

Himsworth, Chelsea G; Zabek, Erin; Desruisseau, Andrea; Parmley, E Jane; Reid-Smith, Richard; Jardine, Claire M; Tang, Patrick; Patrick, David M

2015-07-01

Although rat feces are widely suspected to be a source of pathogenic bacteria, few investigators have studied fecal pathogens in rats. We investigated the prevalence and characteristics of Escherichia coli and Salmonella spp. in Norway and black rats (Rattus norvegicus and Rattus rattus, respectively) from an urban neighborhood of Vancouver, Canada, collected September 2011-August 2012. Colon content was cultured for E. coli and Salmonella spp. and screened for the seven most-common enteropathogenic Shiga toxin-producing E. coli (STEC) serotypes by PCR. Isolates were tested for antimicrobial resistance and Salmonella isolates were serotyped. We detected E. coli in 397/633 (62.7%) urban rats. Forty-one of 397 (6.5%) E. coli isolates were resistant to ≥ 1 antimicrobial while 17 (4.3%) were multidrug resistant (including two isolates demonstrating extended-spectrum β-lactamase resistance). Ten of 633 (1.6%) urban rats were carrying STEC serotypes including O145, O103, O26, and O45. Norway rats were more likely to be carrying E. coli compared to black rats, and there was geographic clustering of specific resistance patterns and STEC serotypes. Salmonella spp. were detected in 3/633 (0.5%) rats including serotypes Derby, Indiana, and Enteritidis. In contrast to zoonotic pathogens for which rats are the natural reservoir (e.g., Leptospira interrogans, Rickettsia typhi, Seoul virus), rats likely acquired E. coli and Salmonella spp. from their environment. The ability of rats to be a 'sponge' for environmental pathogens has received little consideration, and the ecology and public health significance of these organisms in rats requires further investigation.

Temporospatial fate of bacteria and immune effector expression in house flies (Musca domestica L.) fed GFP-E. coli O157:H7

PubMed Central

Fleming, A.; Kumar, H.V.; Joyner, C.; Reynolds, A.; Nayduch, D.

2014-01-01

House flies (Diptera: Muscidae; Musca domestica L.) harbor and transmit a variety of human enteropathogens including E. coli O157:H7. Interactions between ingested bacteria and the fly gut directly impact bacterial persistence, survival and ultimately fly vector competence. We assessed the temporospatial fate of GFP-E. coli O157:H7 (GFP-ECO157) in house flies along with fly antimicrobial responses for 12 h post-ingestion. In flies fed GFP-ECO157, culture and microscopy revealed a steady decrease in bacterial load over 12 h, which was likely attributable to the combined effects of immobilization within the peritrophic matrix, lysis and peristaltic excretion. However, flies can putatively transmit this pathogen in excreta because intact bacteria were observed in the crop and rectum. qRT-PCR analysis of antimicrobial peptides (AMP) and lysozyme gene expression showed minimal upregulation in both the gut and carcass of house flies fed GFP-ECO157. However, these genes were upregulated in fly heads and salivary glands, and effector proteins were detected in the gut of some flies. Collectively, these data indicate that house flies can serve as reservoirs of E. coli O157:H7 for up to 12 h, and factors in addition to AMPs and lysozyme may contribute to bacteria destruction in the gut. PMID:24712451

Effect of RNase E deficiency on translocon protein synthesis in an RNase E-inducible strain of enterohemorrhagic Escherichia coli O157:H7.

PubMed

Lodato, Patricia B; Thuraisamy, Thujitha; Richards, Jamie; Belasco, Joel G

2017-07-06

Enterohemorrhagic Escherichia coli (EHEC) is a food-borne pathogen that assembles a type III secretion system (T3SS) on its surface. The last portion of the T3SS, called the 'translocon', is composed of a filament and a pore complex that is inserted into the membrane of intestinal epithelial cells. The genes encoding the translocon (espADB) are part of the LEE4 operon. Their expression is regulated by a complex post-transcriptional mechanism that involves the processing of LEE4 mRNA by the essential endoribonuclease RNase E. Here, we report the construction of an EHEC strain (TEA028-rne) in which RNase E can be induced by adding IPTG to the culture medium. EHEC cells deficient in RNase E displayed an abnormal morphology and slower growth, in agreement with published observations in E. coli K-12. Under those conditions, EspA and EspB were produced at higher concentrations, and protein secretion still occurred. These results indicate that RNase E negatively regulates translocon protein synthesis and demonstrate the utility of E. coli strain TEA028-rne as a tool for investigating the influence of this ribonuclease on EHEC gene expression in vitro. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Engineering new mycobacterial vaccine design for HIV–TB pediatric vaccine vectored by lysine auxotroph of BCG

PubMed Central

Saubi, Narcís; Gea-Mallorquí, Ester; Ferrer, Pau; Hurtado, Carmen; Sánchez-Úbeda, Sara; Eto, Yoshiki; Gatell, Josep M; Hanke, Tomáš; Joseph, Joan

2014-01-01

In this study, we have engineered a new mycobacterial vaccine design by using an antibiotic-free plasmid selection system. We assembled a novel Escherichia coli (E. coli)–mycobacterial shuttle plasmid p2auxo.HIVA, expressing the HIV-1 clade A immunogen HIVA. This shuttle vector employs an antibiotic resistance-free mechanism for plasmid selection and maintenance based on glycine complementation in E. coli and lysine complementation in mycobacteria. This plasmid was first transformed into glycine auxotroph of E. coli strain and subsequently transformed into lysine auxotroph of Mycobacterium bovis BCG strain to generate vaccine BCG.HIVA2auxo. We demonstrated that the episomal plasmid p2auxo.HIVA was stable in vivo over a 7-week period and genetically and phenotypically characterized the BCG.HIVA2auxo vaccine strain. The BCG.HIVA2auxo vaccine in combination with modified vaccinia virus Ankara (MVA). HIVA was safe and induced HIV-1 and Mycobacterium tuberculosis-specific interferon-γ-producing T-cell responses in adult BALB/c mice. Polyfunctional HIV-1-specific CD8+ T cells, which produce interferon-γ and tumor necrosis factor-α and express the degranulation marker CD107a, were induced. Thus, we engineered a novel, safer, good laboratory practice–compatible BCG-vectored vaccine using prototype immunogen HIVA. This antibiotic-free plasmid selection system based on “double” auxotrophic complementation might be a new mycobacterial vaccine platform to develop not only recombinant BCG-based vaccines expressing second generation of HIV-1 immunogens but also other major pediatric pathogens to prime protective response soon after birth. PMID:26015961

Rapid detection of foodborne microorganisms on food surface using Fourier transform Raman spectroscopy

NASA Astrophysics Data System (ADS)

Yang, Hong; Irudayaraj, Joseph

2003-02-01

Fourier transform (FT) Raman spectroscopy was used for non-destructive characterization and differentiation of six different microorganisms including the pathogen Escherichia coli O157:H7 on whole apples. Mahalanobis distance metric was used to evaluate and quantify the statistical differences between the spectra of six different microorganisms. The same procedure was extended to discriminate six different strains of E. coli. The FT-Raman procedure was not only successful in discriminating the different E. coli strain but also accurately differentiated the pathogen from non-pathogens. Results demonstrate that FT-Raman spectroscopy can be an excellent tool for rapid examination of food surfaces for microorganism contamination and for the classification of microbial cultures.

Functional characterization of the recombinant antimicrobial peptide Trx-Ace-AMP1 and its application on the control of tomato early blight disease.

PubMed

Wu, Yin; He, Yue; Ge, Xiaochun

2011-05-01

Ace-AMP1 is a potent antifungal peptide found in onion (Allium cepa) seeds with sequence similarity to plant lipid transfer proteins. Transgenic plants over-expressing Ace-AMP1 gene have enhanced disease resistance to some fungal pathogens. However, mass production in heterologous systems and in vitro application of this peptide have not been reported. In this study, Ace-AMP1 was highly expressed in a prokaryotic Escherichia coli system as a fusion protein. The purified protein inhibited the growth of many plant fungal pathogens, especially Alternaria solani, Fusarium oxysporum f. sp. vasinfectum, and Verticillium dahliae. The inhibitory effect was accompanied by hyphal hyperbranching for V. dahliae but not for F. oxysporum f. sp. vasinfectum and A. solani, suggesting that the mode of action of Ace-AMP1 on different fungi might be different. Application of Ace-AMP1 on tomato leaves showed that the recombinant protein conferred strong resistance to the tomato pathogen A. solani and could be used as an effective fungicide.

Uropathogenic E.coli (UPEC) Infection Induces Proliferation through Enhancer of Zeste Homologue 2 (EZH2)

PubMed Central

Penna, Frank; Samiei, Alaleh Najdi; Sidler, Martin; Jiang, Jia-Xin; Ibrahim, Fadi; Tolg, Cornelia; Delgado-Olguin, Paul; Rosenblum, Norman; Bägli, Darius J.

2016-01-01

Host-pathogen interactions can induce epigenetic changes in the host directly, as well as indirectly through secreted factors. Previously, uropathogenic Escherichia coli (UPEC) was shown to increase DNA methyltransferase activity and expression, which was associated with methylation-dependent alterations in the urothelial expression of CDKN2A. Here, we showed that paracrine factors from infected cells alter expression of another epigenetic writer, EZH2, coordinate with proliferation. Urothelial cells were inoculated with UPEC, UPEC derivatives, or vehicle (mock infection) at low moi, washed, then maintained in media with Gentamycin. Urothelial conditioned media (CM) and extracellular vesicles (EV) were isolated after the inoculations and used to treat naïve urothelial cells. EZH2 increased with UPEC infection, inoculation-induced CM, and inoculation-induced EV vs. parallel stimulation derived from mock-inoculated urothelial cells. We found that infection also increased proliferation at one day post-infection, which was blocked by the EZH2 inhibitor UNC1999. Inhibition of demethylation at H3K27me3 had the opposite effect and augmented proliferation. CONCLUSION: Uropathogen-induced paracrine factors act epigenetically by altering expression of EZH2, which plays a key role in early host cell proliferative responses to infection. PMID:26964089

Probiotics inhibit enteropathogenic E. coli adherence in vitro by inducing intestinal mucin gene expression.

PubMed

Mack, D R; Michail, S; Wei, S; McDougall, L; Hollingsworth, M A

1999-04-01

Probiotic agents, live microorganisms with beneficial effects for the host, may offer an alternative to conventional antimicrobials in the treatment and prevention of enteric infections. The probiotic agents Lactobacillus plantarum 299v and Lactobacillus rhamnosus GG quantitatively inhibited the adherence of an attaching and effacing pathogenic Escherichia coli to HT-29 intestinal epithelial cells but did not inhibit adherence to nonintestinal HEp-2 cells. HT-29 cells were grown under conditions that induced high levels of either MUC2 or MUC3 mRNA, but HEp-2 cells expressed only minimal levels of MUC2 and no MUC3 mRNA. Media enriched for MUC2 and MUC3 mucin were added exogenously to binding assays and were shown to be capable of inhibiting enteropathogen adherence to HEp-2 cells. Incubation of L. plantarum 299v with HT-29 cells increased MUC2 and MUC3 mRNA expression levels. From these in vitro studies, we propose the hypothesis that the ability of probiotic agents to inhibit adherence of attaching and effacing organisms to intestinal epithelial cells is mediated through their ability to increase expression of MUC2 and MUC3 intestinal mucins.

[Etiological surveillance and analysis of infectious diarrhea in Beijing in year 2010].

PubMed

Huang, Fang; Deng, Ying; Qu, Mei; Liu, Gui-Rong; Liu, Yuan; Zhang, Xin; Li, Jie; Yan, Han-Qiu; Gao, Zhi-Yong; Liu, Bai-Wei; Li, Xi-Tai; Li, Xin-Yu

2011-09-01

To explore the pathogenic form, epidemic features and serotype distribution of the pathogenic bacteria causing infectious diarrhea in Beijing. A total of 2118 samples of rectal swabs and stool specimens of diarrheal patients were collected from 6 surveillant intestinal tract clinics during the period between April and October, 2010. Enteric multiple pathogens including Vibrio cholerae, Vibrio parahaemolyticus, Salmonella, Shigella and diarrheagenic Escherichia coli were detected by the isolation culture, biochemical identification and serotyping methods. The population distribution, temporal distribution and serotype distribution of the above pathogenic bacteria were analyzed by descriptive statistical methods. 478 strains isolated from the total 2118 specimens were positive for pathogen detection, accounting to 22.6%. Among the 478 strains of pathogenic bacteria, Shigella accounting for 40.8% (195/478) was the most frequent pathogen, followed by Vibrio parahaemolyticus accouting for 23.8% (114/478), Salmonella accounting for 19.0% (91/478) and diarrheagenic Escherichia coli accounting for 4.8% (23/478). Enteric pathogenic bacteria spread mainly among adults aging between 20 and 39; and the distribution was different among different age groups, while the highest detected rate was in 30 - 39 age group, accounting for 27.2% (92/338). The detected rate of pathogenic bacteria showed evident seasonal variations, with a peak from July to October, whose detected rates were 23.5% (114/486), 32.8% (176/536), 36.1% (90/249) and 25.9% (29/112) respectively. The detected rates in other months were all under 16.0%. Shigella Sonnei was the dominant serotype, accounting for 83.1% (162/195). O3:K6 was the dominant serotype among Vibrio parahaemolyticus, accounting for 63.2% (72/114). Salmonella Enteritidis and Salmonella Typhimurium were dominant serotypes among Salmonella, accounting for 13.2% (12/91) and 12.1% (11/91) separately. Enterpathogenic Escherichia coli and enterotoxigenic Escherichia coli were the dominant serotypes among Diarrheagenic Escherichia coli, accounting for 69.6% (16/23) and 30.4% (7/23) respectively. The three main pathogenic bacteria causing infectious diarrhea in Beijing are Shigella, Vibrio parahaemolyticus, Salmonella; and there are obvious changes in the serotype distribution of Shigella and Samonella compared to previous years.

Pathogenic bacteria induce colonic PepT1 expression: an implication in host defense response

PubMed Central

Nguyen, Hang Thi Thu; Dalmasso, Guillaume; Powell, Kimberly R.; Yan, Yutao; Bhatt, Shantanu; Kalman, Daniel; Sitaraman, Shanthi; Merlin, Didier

2009-01-01

Background & Aims Expression of the di/tripeptide transporter PepT1 has been observed in the colon under inflammatory conditions, however, the inducing factors and underlying mechanisms remain unknown. Here, we address the effects of pathogenic bacteria on colonic PepT1 expression together with its functional consequences. Methods Human colonic HT29-Cl.19A cells were infected with the attaching and effacing (A/E) enteropathogenic E. coli (EPEC). Wild-type and PepT1 transgenic mice or cultured colonic tissues derived from these mice were infected with Citrobacter rodentium, a murine A/E pathogen related to EPEC. Results EPEC induced PepT1 expression and activity in HT29-Cl.19A cells by intimately attaching to host cells through lipid rafts. Induction of PepT1 expression by EPEC required the transcription factor Cdx2. PepT1 expression reduced binding of EPEC to lipid rafts, as well as activation of NF-κB and MAP kinase and production of IL-8. Accordingly, ex vivo and in vivo experiments revealed that C. rodentium induced colonic PepT1 expression and that, compared to their wild-type counterparts, PepT1 transgenic mice infected with C. rodentium exhibited decreased bacterial colonization, production of pro-inflammatory cytokines, and neutrophil infiltration into the colon. Conclusions Our findings demonstrate a molecular mechanism underlying the regulation of colonic PepT1 expression under pathological conditions and reveal a novel role for PepT1 in host defense via its capacity to modulate bacterial-epithelial interactions and intestinal inflammation. PMID:19549526

The chicken as a natural model for extraintestinal infections caused by avian pathogenic Escherichia coli (APEC).

PubMed

Antão, Esther-Maria; Glodde, Susanne; Li, Ganwu; Sharifi, Reza; Homeier, Timo; Laturnus, Claudia; Diehl, Ines; Bethe, Astrid; Philipp, Hans-C; Preisinger, Rudolf; Wieler, Lothar H; Ewers, Christa

2008-01-01

E. coli infections in avian species have become an economic threat to the poultry industry worldwide. Several factors have been associated with the virulence of E. coli in avian hosts, but no specific virulence gene has been identified as being entirely responsible for the pathogenicity of avian pathogenic E. coli (APEC). Needless to say, the chicken would serve as the best model organism for unravelling the pathogenic mechanisms of APEC, an extraintestinal pathogen. Five-week-old white leghorn SPF chickens were infected intra-tracheally with a well characterized APEC field strain IMT5155 (O2:K1:H5) using different doses corresponding to the respective models of infection established, that is, the lung colonization model allowing re-isolation of bacteria only from the lung but not from other internal organs, and the systemic infection model. These two models represent the crucial steps in the pathogenesis of APEC infections, including the colonization of the lung epithelium and the spread of bacteria throughout the bloodstream. The read-out system includes a clinical score, pathomorphological changes and bacterial load determination. The lung colonization model has been established and described for the first time in this study, in addition to a comprehensive account of a systemic infection model which enables the study of severe extraintestinal pathogenic E. coli (ExPEC) infections. These in vivo models enable the application of various molecular approaches to study host-pathogen interactions more closely. The most important application of such genetic manipulation techniques is the identification of genes required for extraintestinal virulence, as well as host genes involved in immunity in vivo. The knowledge obtained from these studies serves the dual purpose of shedding light on the nature of virulence itself, as well as providing a route for rational attenuation of the pathogen for vaccine construction, a measure by which extraintestinal infections, including those caused by APEC, could eventually be controlled and prevented in the field.

Isolation and evaluation of cocktail phages for the control of multidrug-resistant Escherichia coli serotype O104: H4 and E. coli O157: H7 isolates causing diarrhea.

PubMed

Safwat Mohamed, Doaa; Farouk Ahmed, Eman; Mohamed Mahmoud, Abobakr; Abd El-Baky, Rehab Mahmoud; John, James

2018-02-01

Escherichia coli serotype O157: H7 and E. coli O104: H4 are well known foodborne pathogens causing sever enteric illness. Using bacteriophages as biocontrol agents of some foodborne pathogens and multidrug-resistant (MDR) bacteria has a great attention nowadays. This study aims to test the effect of cocktail phages on the growth of some foodborne pathogens and MDR E. coli. Routine conventional PCR was used to confirm the identification of E. coli isolates. Double-layered culture technique was used to isolate phages from sewage water. Morphology of bacteriophage was described using transmission electron microscopy, and spot test was performed to determine host range of the phage cocktail. Phage cocktail of Siphoviridae and Podoviridae family infecting E. coli O157: H7, E. coli O104: H4 and untypeable E. coli (neither O157 nor O104) has been isolated from sewage water. Phage cocktail showed both lytic and lysogenic activity. Lytic activity was observed against E. coli O157: H7, E. coli O104: H4 isolates, Staphylococcus. aureus ATCC6538 and Pseudomonas aeruginosa ATCC 10145, while the lysogenic activity was observed against the untypeable strain. The tested phage cocktail showed a promising inhibitory action on E. coli O157: H7 and O104: H4, S. aureus ATCC6538 and P. aeruginosa ATCC 10145, suggesting the possibility of its use as a biocontrol tool or as natural food preservatives for many food products. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

The target of daptomycin is absent from Escherichia coli and other gram-negative pathogens.

PubMed

Randall, Christopher P; Mariner, Katherine R; Chopra, Ian; O'Neill, Alex J

2013-01-01

Antistaphylococcal agents commonly lack activity against Gram-negative bacteria like Escherichia coli owing to the permeability barrier presented by the outer membrane and/or the action of efflux transporters. When these intrinsic resistance mechanisms are artificially compromised, such agents almost invariably demonstrate antibacterial activity against Gram negatives. Here we show that this is not the case for the antibiotic daptomycin, whose target appears to be absent from E. coli and other Gram-negative pathogens.

The role of unusual pathogens in prostatitis syndrome.

PubMed

Skerk, Visnja; Krhen, Ivan; Schonwald, Slavko; Cajic, Vjeran; Markovinovic, Leo; Roglic, Srdan; Zekan, Sime; Andracevic, Arjana Tambic; Kruzic, Vladimira

2004-09-01

A total of 1442 patients with symptoms of chronic prostatitis were examined over a 4-year period at the Outpatient Department for Urogenital Infections, University Hospital for Infectious Diseases "Dr. Fran Mihaljevic", Zagreb, Croatia. An infectious aetiology was determined in 1070 (74.2%) patients. In 561 of 1070 (52.4%) patients the inflammatory finding (>10 WBC/hpf) was found in expressed prostatic secretions (EPS) or voided bladder urine (VB(3)). Normal, <10 WBCs/hpf was found in 362 of 536 (67.5%) patients with symptoms of chronic prostatitis in whom Chlamydia trachomatis was detected in EPS or VB(3), in 51 of 151 (33.8%) patients with isolated Trichomonas vaginalis and in 40 of 72 (55.6%) patients with isolated Ureaplasma urealyticum. Escherichia coli was the causative pathogen in 95, Enterococcus in 68, Proteus mirabilis in 37, Klebsiella pneumoniae in 16, Streptococcus agalactiae in 19, and Pseudomonas aeruginosa in 3 patients with chronic prostatitis. Other patients had a mixed infection. In patients with chronic bacterial prostatitis (CBP) caused by E. coli, P. mirabilis, K. pneumoniae, E. or S. agalactiae, an inflammatory finding was regularly found in EPS or VB(3).

[Outbreaks caused by diarrheagenic Escherichia coli].

PubMed

Vila Estapé, Jordi; Zboromyrska, Yuliya

2012-02-01

Escherichia coli are ubiquitous bacteria from a wide variety of ecosystems including the gastrointestinal tract of humans and warm-blooded animals. E. coli can play a role as an opportunistic bacteria causing a variety of infectious diseases including, among many others, sepsis, urinary tract infections, meningitis, and wound infections. Moreover, these bacteria can also act as primary pathogens in the intestinal tract. There are several pathotypes of E. coli that cause enteritis, and both sporadic cases and outbreaks have been reported. In this article, we review the pathogenicity and epidemiology of enteritis caused by these E. coli pathotypes, and provide some examples of outbreaks described in the scientific literature and the measures required to prevent them. Copyright © 2011 Elsevier España, S.L. All rights reserved.

Prevention and cure of systemic Escherichia coli K1 infection by modification of the bacterial phenotype.

PubMed

Mushtaq, Naseem; Redpath, Maria B; Luzio, J Paul; Taylor, Peter W

2004-05-01

Escherichia coli is a common cause of meningitis and sepsis in the newborn infant, and the large majority of isolates from these infections produce a polysialic acid (PSA) capsular polysaccharide, the K1 antigen, that protects the bacterial cell from immune attack. We determined whether a capsule-depolymerizing enzyme, by removing this protective barrier, could alter the outcome of systemic infection in an animal model. Bacteriophage-derived endosialidase E (endoE) selectively degrades the PSA capsule on the surface of E. coli K1 strains. Intraperitoneal administration of small quantities of recombinant endoE (20 micro g) to 3-day-old rats, colonized with a virulent strain of K1, prevented bacteremia and death from systemic infection. The enzyme had no effect on the viability of E. coli strains but sensitized strains expressing PSA to killing by the complement system. This study demonstrates the potential therapeutic efficacy of agents that cure infections by modification of the bacterial phenotype rather than by killing or inhibition of growth of the pathogen.

Applications of immunomagnetic capture and time-resolved fluorescence to detect outbreak Escherichia coli O157 and Salmonella in alfalfa sprouts

NASA Astrophysics Data System (ADS)

Tu, Shu-I.; Gordon, Marsha; Fett, William F.; Gehring, Andrew G.; Irwin, Peter L.

2004-03-01

Commercially available alfalfa seeds were inoculated with low levels (~ 4 CFU/g) of pathogenic bacteria. The inoculated seeds were then allowed to sprout in sterile tap water at 22°C. After 48 hours, the irrigation water and sprouts were separately transferred to bovine heart infusion (BHI) media. The microbes in the BHI samples were allowed to grow for 4 hours at 37°C and 160 rpm. Specific immunomagnetic beads (IMB) were then applied to capture the E.coli O157 and/or Salmonella in the growth media. Separation and concentration of IMB-captured pathogens were achieved using magnetic separators. The captured E. coli O157:H7 and Salmonella spp were further tagged with europium (Eu) labeled anti-E. coli O157 antibodies and samarium (Sm) labeled anti-Salmonella antibodies, respectively. After washing, the lanthanide labels were extracted out from the complexes by specific chelators to form strongly fluorescent chelates. The specific time-resolved fluorescence (TRF) associated with Eu or Sm was measured to estimate the extent of capture of the E. coli O157 and Salmonella, respectively. The results indicated that the approach could detect E. coli O157 and Salmonella enterica from the seeds inoculated with ~ 4 CFU/g of the pathogens. Non-targeted bacteria, e.g., Aeromonas and Citrobacter exhibited essentially no cross reactivity. Since the pathogen detection from the sprouts was achieved within 6 hours, the developed methodology could be use as a rapid, sensitive and specific screening process for E. coli O157 and Salmonella enterica in this popular salad food.

tir- and stx-Positive Escherichia coli in Stream Waters in a Metropolitan Area

PubMed Central

Higgins, James A.; Belt, Kenneth T.; Karns, Jeffrey S.; Russell-Anelli, Jonathan; Shelton, Daniel R.

2005-01-01

Diarrheagenic Escherichia coli, which may include the enteropathogenic E. coli and the enterohemorrhagic E. coli, are a significant cause of diarrheal disease among infants and children in both developing and developed areas. Disease outbreaks related to freshwater exposure have been documented, but the presence of these organisms in the urban aquatic environment is not well characterized. From April 2002 through April 2004 we conducted weekly surveys of streams in the metropolitan Baltimore, Md., area for the prevalence of potentially pathogenic E. coli by using PCR assays targeting the tir and stx1 and stx2 genes. Coliforms testing positive for the presence of the tir gene were cultured from 653 of 1,218 samples (53%), with a greater prevalence associated with urban, polluted streams than in suburban and forested watershed streams. Polluted urban streams were also more likely to test positive for the presence of one of the stx genes. Sequence analysis of the tir amplicon, as well as the entire tir gene from three isolates, indicated that the pathogenic E. coli present in the stream waters has a high degree of sequence homology with the E. coli O157:H7 serotype. Our data indicate that pathogenic E. coli are continually deposited into a variety of stream habitats and suggest that this organism may be a permanent member of the gastrointestinal microflora of humans and animals in the metropolitan Baltimore area. PMID:15870341

Survival and interaction of Escherichia coli O104:H4 on Arabidopsis thaliana and lettuce (Lactuca sativa) in comparison to E. coli O157:H7: Influence of plant defense response and bacterial capsular polysaccharide.

PubMed

Jang, Hyein; Matthews, Karl R

2018-06-01

Shiga toxin-producing Escherichia coli (STEC) has been associated with illnesses and outbreaks linked to fresh vegetables, prompting a growing public health concern. Most studies regarding interactions of STEC on fresh produce focused on E. coli O157:H7. Limited information is available about survival or fitness of E. coli O104:H4, non-O157 pathogen that was linked to one of the largest outbreaks of hemolytic uremic syndrome in 2011. In this study, survival of E. coli O104:H4 was evaluated on Arabidopsis thaliana plant and lettuce for 5 days compared with E. coli O157:H7, and expression of pathogenesis-realted gene (PR1; induction of plant defense response) was examined by reverse transcription quantitative PCR, and potential influence of capsular polysaccharide (CPS) on the bacterial fitness on plant was investigated. Populations of E. coli O104:H4 strains (RG1, C3493, and LpfA) on Arabidopsis and lettuce were significantly (P < 0.05) greater than those of E. coli O157:H7 strains (7386 and sakai) at day 5 post-inoculation, indicating E. coli O104:H4 may have better survival ability on the plants. In addition, the E. coli O104:H4 strains produced significantly (P < 0.05) higher amounts of CPS compared with the E. coli O157:H7 strains. RG1 strain (1.5-fold) initiated significantly (P < 0.05) lower expression of PR1 gene indicating induction of plant defense response compared with E. coli O157:H7 strains 7386 (2.9-fold) and sakai (2.7-fold). Collectively, the results in this study suggests that different level of CPS production and plant defense response initiated by each STEC strain might influence the bacterial survival or persistence on plants. The present study provides better understanding of survival behavior of STEC, particularly E. coli O104:H4, using a model plant and vegetable under pre-harvest conditions with plant defense response. Copyright © 2018 Elsevier Ltd. All rights reserved.

Survival of generic E. coli and surrogate E. coli O157:H7 in manure-amended soils

USDA-ARS?s Scientific Manuscript database

Introduction: Recently released U.S. FDA standards state that untreated biological soil amendments must be applied to soil 9 months before produce crop harvest to reduce pathogen contamination risk on produce. Manure and soil type may impact survival of bacterial pathogens. Purpose: Determine survi...

Bacterial delivery of RNAi effectors: transkingdom RNAi.

PubMed

Lage, Hermann; Krühn, Andrea

2010-08-18

RNA interference (RNAi) represents a high effective mechanism for specific inhibition of mRNA expression. Besides its potential as a powerful laboratory tool, the RNAi pathway appears to be promising for therapeutic utilization. For development of RNA interference (RNAi)-based therapies, delivery of RNAi-mediating agents to target cells is one of the major obstacles. A novel strategy to overcome this hurdle is transkingdom RNAi (tkRNAi). This technology uses non-pathogenic bacteria, e.g. Escherichia coli, to produce and deliver therapeutic short hairpin RNA (shRNA) into target cells to induce RNAi. A first-generation tkRNAi-mediating vector, TRIP, contains the bacteriophage T7 promoter for expression regulation of a therapeutic shRNA of interest. Furthermore, TRIP has the Inv locus from Yersinia pseudotuberculosis that encodes invasin, which permits natural noninvasive bacteria to enter beta1-integrin-positive mammalian cells and the HlyA gene from Listeria monocytogenes, which produces listeriolysin O. This enzyme allows the therapeutic shRNA to escape from entry vesicles within the cytoplasm of the target cell. TRIP constructs are introduced into a competent non-pathogenic Escherichia coli strain, which encodes T7 RNA polymerase necessary for the T7 promoter-driven synthesis of shRNAs. A well-characterized cancer-associated target molecule for different RNAi strategies is ABCB1 (MDR1/P-glycoprotein, MDR1/P-gp). This ABC-transporter acts as a drug extrusion pump and mediates the "classical" ABCB1-mediated multidrug resistance (MDR) phenotype of human cancer cells which is characterized by a specific cross resistance pattern. Different ABCB1-expressing MDR cancer cells were treated with anti-ABCB1 shRNA expression vector bearing E. coli. This procedure resulted in activation of the RNAi pathways within the cancer cells and a considerable down regulation of the ABCB1 encoding mRNA as well as the corresponding drug extrusion pump. Accordingly, drug accumulation was enhanced in the pristine drug-resistant cancer cells and the MDR phenotype was reversed. By means of this model the data provide the proof-of-concept that tkRNAi is suitable for modulation of cancer-associated factors, e.g. ABCB1, in human cancer cells.

Identification and Characterization of T5-Like Bacteriophages Representing Two Novel Subgroups from Food Products

PubMed Central

Sváb, Domonkos; Falgenhauer, Linda; Rohde, Manfred; Szabó, Judit; Chakraborty, Trinad; Tóth, István

2018-01-01

During recent years, interest in the use of bacteriophages as biocontrol agents against foodborne pathogens has increased, particularly for members of the family Enterobacteriaceae, with pathogenic Escherichia coli, Shigella, and Salmonella strains among them. Here, we report the isolation and characterisation of 12 novel T5-like bacteriophages from confiscated food samples. All bacterophages effectively lysed E. coli K-12 strains and were able to infect pathogenic E. coli strains representing enterohaemorrhagic (EHEC), enteropathogenic (EPEC), enterotoxigenic (ETEC), and enteroinvasive (EIEC) pathotypes, Shigella dysenteriae, S. sonnei strains, as well as multidrug-resistant (MDR) E. coli and multiple strains representing different Salmonella enterica serovars. All the bacteriophages exhibited Siphoviridae morphology. Whole genome sequencing of the novel T5-like bacteriophages showed that they represent two distinct groups, with the genome-based grouping correlating to the different host spectra. As these bacteriophages are of food origin, their stability and lack of any virulence genes, as well as their broad and mutually complementary host spectrum makes these new T5-like bacteriophages valuable candidates for use as biocontrol agents against foodborne pathogenic enterobacteria. PMID:29487585

Community-acquired urinary tract infection in hospitalized children: etiology and antimicrobial resistance. A comparison between first episode and recurrent infection.

PubMed

Sakran, Waheeb; Smolkin, Vladislav; Odetalla, Ahmad; Halevy, Raphael; Koren, Ariel

2015-05-01

Urinary tract infection (UTI) is common in infants and children, and Escherichia coli is the leading pathogen. The aims of this study were to compare first episode of UTI with recurrent infection, reveal organisms that cause UTI, uropathogen resistance, and presence of bacteria producing extended-spectrum β-lactamase (ESBL). The first-UTI group included 456 children. E coli was the leading pathogen (80.5%), and Pseudomonas aeruginosa was found in 1.5%. The uropathogens were resistant to gentamicin (3.41%) and cefuroxime (5.71%), and highly resistant to cefamezin (37.39%). The recurrent-infection group included 106 children. E coli was also the leading pathogen, but 7.5% of the isolates were P aeruginosa (P = .002 compared with first-episode group); 6.6% were ESBL-producing bacteria compared with 1.1% in the first-episode group (P = .002). E coli is the leading pathogen in both groups. P aeruginosa and ESBL-producing bacteria were more common in the recurrent infection group. © The Author(s) 2014.

[The detection of occurrence rate of genes coding capability to form pili binding in auto-strains of Escherichia coli].

PubMed

Ivanova, E I; Popkova, S M; Dzhioev, Iu P; Rakova, E B; Dolgikh, V V; Savel'kaeva, M V; Nemchenko, U M; Bukharova, E V; Serdiuk, L V

2015-01-01

E. coli is a commensal of intestine of the vertebrata. The exchange of genetic material of different types of bacteria between themselves and with other representatives of family of Enterobacteriaceae in intestinal ecosystem results in development of types of normal colibacillus with genetic characteristics of pathogenicity that can serve as a theoretical substantiation to attribute such strains to pathobionts. The entero-pathogenic colibacillus continues be an important cause of diarrhea in children in developing countries. The gene responsible for formation of pili binding is a necessary condition for virulence of entero-pathogenic colibacillus. The polymerase chain reaction was applied to examine 316 strains of different types of E. coli (normal, with weak enzyme activity and hemolytic activity) isolated from healthy children and children with functional disorders of gastro-intestinal tract for presence of genes coding capability to form pill binding. The presence of this gene in different biochemical types of E. coli permits to establish the fact of formation of reservoir of pathogenicity in indigent microbiota of intestinal biocenosis.

Microbiological analysis of pre-packed sweet basil (Ocimum basilicum) and coriander (Coriandrum sativum) leaves for the presence of Salmonella spp. and Shiga toxin-producing E. coli.

PubMed

Delbeke, Stefanie; Ceuppens, Siele; Jacxsens, Liesbeth; Uyttendaele, Mieke

2015-09-02

Enteric pathogens, such as Salmonella spp. and pathogenic Escherichia coli, have been detected and associated with food borne outbreaks from (imported) fresh leafy herbs. Screening on imported herbs from South East Asian countries has been described. However, limited information on prevalence of these pathogens is available from other sourcing regions. Therefore, fresh pre-packed basil and coriander leaves from a Belgian trading company were investigated for the presence of Salmonella spp., Shiga toxin-producing E. coli (STEC), generic E. coli and coliforms. In total 592 samples were collected originating from Belgium, Israel and Cyprus during 2013-2014. Multiplex PCR followed by further culture confirmation was used for the detection of Salmonella spp. and STEC, whereas the Petrifilm Select E. coli and VRBL-agar were used, respectively, for the enumeration of E. coli and coliforms. Salmonella was detected in 10 out of 592 samples (25g) (1.7%; 5 from basil and 5 from coriander), of which two samples were sourced from Israel and eight from Cyprus. The presence of STEC was suspected in 11 out of 592 samples (25g) (1.9%; 3 basil and 8 coriander), due to the detection of stx and eae genes, of which one sample originated from Belgium, four from Israel and six from Cyprus. No STEC was isolated by culture techniques, but in three samples a serotype (O26, O103 or O111) with its most likely associated eae-variant (β or θ) was detected by PCR. Generic E. coli was enumerated in 108 out of 592 samples, whereby 55, 32 and 13 samples respectively between 10-100, 100-1000 and 1000-10,000cfu/g and 8 samples exceeding 10,000cfu/g. Coliforms were enumerated in all herb samples at variable levels ranging from 1.6 to 7.5logcfu/g. Further statistics indicate that the E. coli class (categorized by level) was significantly correlated with the presence of Salmonella (p<0.001) or STEC (p=0.019), while coliform counts were significant correlated with Salmonella (p<0.001), but not with STEC (p=0.405). Generic E. coli class is a better indicator for the presence of enteric pathogens than coliforms on fresh herbs, but the relationship between E. coli and Salmonella or STEC was not strong enough to provide a threshold value for E. coli to assure food safety (i.e. no pathogens present). Results indicate that fresh leafy herbs like basil and coriander sourced from different cultivation regions, may contain enteric pathogens and potentially pose a risk for human health. Copyright © 2015 Elsevier B.V. All rights reserved.

Diarrheagenic Escherichia coli strains recovered from urban pigeons (Columba livia) in Brazil and their antimicrobial susceptibility patterns.

PubMed

Silva, Vânia L; Nicoli, Jacques R; Nascimento, Thiago C; Diniz, Cláudio G

2009-09-01

Urban pigeons (Columba livia) come into close contact with humans and animals, and may contribute to the spread of infectious agents. These may include human pathogens such as diarrheagenic Escherichia coli strains, which are able to survive in pigeon feces, thus creating potential for human exposure and infection. Our objectives were to determine the occurrence of diarrheagenic E. coli strains in fresh feces from urban pigeons and their drug susceptibility patterns. E. coli strains were isolated from 100 fresh feces samples and presumptive phenotypic species identification was carried out, confirmed by amplification of specific 16S ribosomal RNA encoding DNA. Multiplex PCR was performed to characterize pathogenic strains. Drug susceptibility patterns were determined by the agar dilution method. Enteroinvasive E. coli, Shiga toxin-producing E. coli, enteropathogenic E. coli, and enterotoxigenic E. coli were detected at an overall rate of 12.1%. Among the isolated E. coli strains, 62.1% were susceptible to all tested drugs, whereas 37.9% were resistant to at least one of the antimicrobials tested. Amikacin was the less effective drug (36.8% resistance), followed by ampicillin (7.8%). No resistance was detected to gentamicin, ceftriaxone, and ceftazidime and almost all the isolates were susceptible to ampicillin-sulbactam (98.4%), levofloxacin (97.8%), and trimethoprim-sulfamethoxazole (96.1%). Since these pigeons may harbor multidrug-resistant pathogens, their presence in an urban environment could be an important component of infection spread, with impact on public health.

Microbiological quality and safety assessment of lettuce production in Brazil.

PubMed

Ceuppens, Siele; Hessel, Claudia Titze; de Quadros Rodrigues, Rochele; Bartz, Sabrina; Tondo, Eduardo César; Uyttendaele, Mieke

2014-07-02

The microbiological quality and safety of lettuce during primary production in Brazil were determined by enumeration of hygiene indicators Escherichia coli, coliforms and enterococci and detection of enteric pathogens Salmonella and E. coli O157:H7 in organic fertilizers, soil, irrigation water, lettuce crops, harvest boxes and worker's hands taken from six different lettuce farms throughout the crop growth cycle. Generic E. coli was a suitable indicator for the presence of Salmonella and E. coli O157:H7, while coliforms and enterococci were not. Few pathogens were detected: 5 salmonellae and 2 E. coli O157:H7 from 260 samples, of which only one was lettuce and the others were manure, soil and water. Most (5/7) pathogens were isolated from the same farm and all were from organic production. Statistical analysis revealed the following environmental and agro-technical risk factors for increased microbial load and pathogen prevalence in lettuce production: high temperature, flooding of lettuce fields, application of contaminated organic fertilizer, irrigation with water of inferior quality and large distances between the field and toilets. Control of the composting process of organic fertilizers and the irrigation water quality appear most crucial to improve and/or maintain the microbiological quality and safety during the primary production of lettuce. Copyright © 2014 Elsevier B.V. All rights reserved.

Change in the Structure of Escherichia coli Population and the Pattern of Virulence Genes along a Rural Aquatic Continuum

PubMed Central

Petit, Fabienne; Clermont, Olivier; Delannoy, Sabine; Servais, Pierre; Gourmelon, Michèle; Fach, Patrick; Oberlé, Kenny; Fournier, Matthieu; Denamur, Erick; Berthe, Thierry

2017-01-01

The aim of this study was to investigate the diversity of the Escherichia coli population, focusing on the occurrence of pathogenic E. coli, in surface water draining a rural catchment. Two sampling campaigns were carried out in similar hydrological conditions (wet period, low flow) along a river continuum, characterized by two opposite density gradients of animals (cattle and wild animals) and human populations. While the abundance of E. coli slightly increased along the river continuum, the abundance of both human and ruminant-associated Bacteroidales markers, as well as the number of E. coli multi-resistant to antibiotics, evidenced a fecal contamination originating from animals at upstream rural sites, and from humans at downstream urban sites. A strong spatial modification of the structure of the E. coli population was observed. At the upstream site close to a forest, a higher abundance of the B2 phylogroup and Escherichia clade strains were observed. At the pasture upstream site, a greater proportion of both E and B1 phylogroups was detected, therefore suggesting a fecal contamination of mainly bovine origin. Conversely, in downstream urban sites, A, D, and F phylogroups were more abundant. To assess the occurrence of intestinal pathogenic strains, virulence factors [afaD, stx1, stx2, eltB (LT), estA (ST), ipaH, bfpA, eae, aaiC and aatA] were screened among 651 E. coli isolates. Intestinal pathogenic strains STEC O174:H21 (stx2) and EHEC O26:H11 (eae, stx1) were isolated in water and sediments close to the pasture site. In contrast, in the downstream urban site aEPEC/EAEC and DAEC of human origin, as well as extra-intestinal pathogenic E. coli belonging to clonal group A of D phylogroup, were sampled. Even if the estimated input of STEC (Shiga toxin-producing E. coli) – released in water at the upstream pasture site – at the downstream site was low, we show that STEC could persist in sediment. These results show that, the run-off of small cattle farms contributed, as much as the wastewater effluent, in the dissemination of pathogenic E. coli in both water and sediments, even if the microbiological quality of the water was good or to average quality according to the French water index. PMID:28458656

MOLECULAR CHARACTERISATION AND ANTIMICROBIAL RESISTANCE PATTERNS OF ESCHERICHIA COLI ISOLATES FROM GOATS SLAUGHTERED IN PARTS OF KENYA.

PubMed

Njoroge, S; Muigai, A W T; Njiruh, P N; Kariuki, S

2013-03-01

To determine the antibiotic resistance patterns of pathogenic Escherichia coli on goat meat carcass at Huruma and Kiserian abattoirs in Kenya. Laboratory based study. Huruma and Kiserian abattoirs in Kenya, 400 slaughtered goats inspected by veterinary health officers and approved for human consumption. A Total of 400 slaughtered goats which were inspected by veterinary health officers and approved for human consumption were sampled from Huruma and Kiserian abattoir. Goat carcass swabs were collected by passing each swab tissue on four parts of the carcass mainly neck, right and left forelimbs, right and left hind limbs, and brisket. A total of 54 E. coli isolates were isolated and confirmed to be pathogenic. The percentage of isolates resistant to various microbial agents was recorded as follows: ampicillin (26 %), amoxycillin-clavulanic acid (17%), tetracycline (15%), chroramphenicol (4%), and ceftrixone (2% each). All Escherichia coli isolates were susceptible to gentamicin sulphamethaxazole-trimethomprin, kanamycin, cetriazididine (CAZ, 30pg), ciproxacin, nalidixic acid and chloramphenicol. Isolates were resistant to one or more of the antibiotics tested. Among the drugs tested, resistance was more frequently observed against ampicillin, amoxycillin-clavulanic acid, tetracycline, ceftrixone and chroramphenicol antibiotics. Among the isolates 26(48%) were positive for the stx1 gene, 19(35%) had the eae gene, 10(19%) possessed est gene,while 8(15%) harboured elt gene. Overall five isolates (10%) possessed aspu gene and two (4%) had aggR gene. No isolate possessed ipah gene. This study demonstrated that there is a significant level of antimicrobial resistance in pathogenic E. coli isolated from goat meat from Huruma and Kiserian abattoir. This indicates that goat meat from abattoirs could pose a risk of transmission of pathogenic antibiotic resistant strains to human. Poor hygienic standards and indiscriminate use of antimicrobials are the two main reasons for the presence of resistant pathogens in goat carcasses. Implemention of appropriate hygiene measures to control contamination of meat with pathogenic E. coli.

The Interplay between Entamoeba and Enteropathogenic Bacteria Modulates Epithelial Cell Damage

PubMed Central

Galván-Moroyoqui, José Manuel; Domínguez-Robles, M. del Carmen; Franco, Elizabeth; Meza, Isaura

2008-01-01

Background Mixed intestinal infections with Entamoeba histolytica, Entamoeba dispar and bacteria with exacerbated manifestations of disease are common in regions where amoebiasis is endemic. However, amoeba–bacteria interactions remain largely unexamined. Methodology Trophozoites of E. histolytica and E. dispar were co-cultured with enteropathogenic bacteria strains Escherichia coli (EPEC), Shigella dysenteriae and a commensal Escherichia coli. Amoebae that phagocytosed bacteria were tested for a cytopathic effect on epithelial cell monolayers. Cysteine proteinase activity, adhesion and cell surface concentration of Gal/GalNAc lectin were analyzed in amoebae showing increased virulence. Structural and functional changes and induction of IL-8 expression were determined in epithelial cells before and after exposure to bacteria. Chemotaxis of amoebae and neutrophils to human IL-8 and conditioned culture media from epithelial cells exposed to bacteria was quantified. Principal Findings E. histolytica digested phagocytosed bacteria, although S. dysenteriae retained 70% viability after ingestion. Phagocytosis of pathogenic bacteria augmented the cytopathic effect of E. histolytica and increased expression of Gal/GalNAc lectin on the amoebic surface and increased cysteine proteinase activity. E. dispar remained avirulent. Adhesion of amoebae and damage to cells exposed to bacteria were increased. Additional increases were observed if amoebae had phagocytosed bacteria. Co-culture of epithelial cells with enteropathogenic bacteria disrupted monolayer permeability and induced expression of IL-8. Media from these co-cultures and human recombinant IL-8 were similarly chemotactic for neutrophils and E. histolytica. Conclusions Epithelial monolayers exposed to enteropathogenic bacteria become more susceptible to E. histolytica damage. At the same time, phagocytosis of pathogenic bacteria by amoebae further increased epithelial cell damage. Significance The in vitro system presented here provides evidence that the Entamoeba/enteropathogenic bacteria interplay modulates epithelial cell responses to the pathogens. In mixed intestinal infections, where such interactions are possible, they could influence the outcome of disease. The results offer insights to continue research on this phenomenon. PMID:18648517

Antibacterial Activities of Wasabi against Escherichia coli O157:H7 and Staphylococcus aureus

PubMed Central

Lu, Zhongjing; Dockery, Christopher R.; Crosby, Michael; Chavarria, Katherine; Patterson, Brett; Giedd, Matthew

2016-01-01

Escherichia coli O157:H7 and Staphylococcus aureus are two of the major pathogens frequently involved in foodborne outbreaks. Control of these pathogens in foods is essential to food safety. It is of great interest in the use of natural antimicrobial compounds present in edible plants to control foodborne pathogens as consumers prefer more natural “green” foods. Allyl isothiocyanate (AITC) is an antimicrobial compound naturally present in wasabi (Japanese horseradish) and several other edible plants. Although the antibacterial effects of pure AITC and wasabi extract (essential oil) against several bacteria have been reported, the antibacterial property of natural wasabi has not been well studied. This study investigated the antibacterial activities of wasabi as well as AITC against E. coli O157:H7 and S. aureus. Chemical analysis showed that AITC is the major isothiocyanate in wasabi. The AITC concentration in the wasabi powder used in this study was 5.91 ± 0.59 mg/g. The minimum inhibitory concentration (MIC) of wasabi against E. coli O157:H7 or S. aureus was 1% (or 10 mg/ml). Wasabi at 4% displayed higher bactericidal activity against S. aureus than against E. coli O157:H7. The MIC of AITC against either pathogen was between 10 and 100 μg/ml. AITC at 500 μg/ml was bactericidal against both pathogens while AITC at 1000 μg/ml eliminated E. coli O157:H7 much faster than S. aureus. The results from this study showed that wasabi has strong antibacterial property and has high potential to effectively control E. coli O157:H7 and S. aureus in foods. The antibacterial property along with its natural green color, unique flavor, and advantage to safeguard foods at the point of ingestion makes wasabi a promising natural edible antibacterial plant. The results from this study may be of significant interest to the food industry as they develop new and safe foods. These results may also stimulate more research to evaluate the antibacterial effect of wasabi against other foodborne pathogens and to explore other edible plants for their antimicrobial properties. To our knowledge, this is the first report on the antibacterial activity of wasabi in its natural form of consumption against E. coli O157:H7 and S. aureus. PMID:27708622

Antibacterial Activities of Wasabi against Escherichia coli O157:H7 and Staphylococcus aureus.

PubMed

Lu, Zhongjing; Dockery, Christopher R; Crosby, Michael; Chavarria, Katherine; Patterson, Brett; Giedd, Matthew

2016-01-01

Escherichia coli O157:H7 and Staphylococcus aureus are two of the major pathogens frequently involved in foodborne outbreaks. Control of these pathogens in foods is essential to food safety. It is of great interest in the use of natural antimicrobial compounds present in edible plants to control foodborne pathogens as consumers prefer more natural "green" foods. Allyl isothiocyanate (AITC) is an antimicrobial compound naturally present in wasabi (Japanese horseradish) and several other edible plants. Although the antibacterial effects of pure AITC and wasabi extract (essential oil) against several bacteria have been reported, the antibacterial property of natural wasabi has not been well studied. This study investigated the antibacterial activities of wasabi as well as AITC against E . coli O157:H7 and S . aureus . Chemical analysis showed that AITC is the major isothiocyanate in wasabi. The AITC concentration in the wasabi powder used in this study was 5.91 ± 0.59 mg/g. The minimum inhibitory concentration (MIC) of wasabi against E. coli O157:H7 or S. aureus was 1% (or 10 mg/ml). Wasabi at 4% displayed higher bactericidal activity against S. aureus than against E. coli O157:H7. The MIC of AITC against either pathogen was between 10 and 100 μg/ml. AITC at 500 μg/ml was bactericidal against both pathogens while AITC at 1000 μg/ml eliminated E. coli O157:H7 much faster than S. aureus . The results from this study showed that wasabi has strong antibacterial property and has high potential to effectively control E. coli O157:H7 and S. aureus in foods. The antibacterial property along with its natural green color, unique flavor, and advantage to safeguard foods at the point of ingestion makes wasabi a promising natural edible antibacterial plant. The results from this study may be of significant interest to the food industry as they develop new and safe foods. These results may also stimulate more research to evaluate the antibacterial effect of wasabi against other foodborne pathogens and to explore other edible plants for their antimicrobial properties. To our knowledge, this is the first report on the antibacterial activity of wasabi in its natural form of consumption against E. coli O157:H7 and S. aureus .

Molecular Profiling of Shiga Toxin-Producing Escherichia coli and Enteropathogenic E. coli Strains Isolated from French Coastal Environments.

PubMed

Balière, C; Rincé, A; Delannoy, S; Fach, P; Gourmelon, M

2016-07-01

Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) strains may be responsible for food-borne infections in humans. Twenty-eight STEC and 75 EPEC strains previously isolated from French shellfish-harvesting areas and their watersheds and belonging to 68 distinguishable serotypes were characterized in this study. High-throughput real-time PCR was used to search for the presence of 75 E. coli virulence-associated gene targets, and genes encoding Shiga toxin (stx) and intimin (eae) were subtyped using PCR tests and DNA sequencing, respectively. The results showed a high level of diversity between strains, with 17 unique virulence gene profiles for STEC and 56 for EPEC. Seven STEC and 15 EPEC strains were found to display a large number or a particular combination of genetic markers of virulence and the presence of stx and/or eae variants, suggesting their potential pathogenicity for humans. Among these, an O26:H11 stx1a eae-β1 strain was associated with a large number of virulence-associated genes (n = 47), including genes carried on the locus of enterocyte effacement (LEE) or other pathogenicity islands, such as OI-122, OI-71, OI-43/48, OI-50, OI-57, and the high-pathogenicity island (HPI). One O91:H21 STEC strain containing 4 stx variants (stx1a, stx2a, stx2c, and stx2d) was found to possess genes associated with pathogenicity islands OI-122, OI-43/48, and OI-15. Among EPEC strains harboring a large number of virulence genes (n, 34 to 50), eight belonged to serotype O26:H11, O103:H2, O103:H25, O145:H28, O157:H7, or O153:H2. The species E. coli includes a wide variety of strains, some of which may be responsible for severe infections. This study, a molecular risk assessment study of E. coli strains isolated from the coastal environment, was conducted to evaluate the potential risk for shellfish consumers. This report describes the characterization of virulence gene profiles and stx/eae polymorphisms of E. coli isolates and clearly highlights the finding that the majority of strains isolated from coastal environment are potentially weakly pathogenic, while some are likely to be more pathogenic. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Rapid fluorescence detection of pathogenic bacteria using magnetic enrichment technique combined with magnetophoretic chromatography.

PubMed

Che, Yulan; Xu, Yi; Wang, Renjie; Chen, Li

2017-08-01

A rapid and sensitive analytical method was developed to detect pathogenic bacteria which combined magnetic enrichment, fluorescence labeling with polyethylene glycol (PEG) magnetophoretic chromatography. As pathogenic bacteria usually exist in complex matrixes at low concentration, an efficient enrichment is essential for diagnosis. In order to capture series types of pathogenic bacteria in samples, amino-modified magnetic nanoparticles (Fe 3 O 4 @SiO 2 -NH 2 ) were prepared for efficient enrichment by the electrostatic interaction with pathogenic bacteria. It was shown that the capture efficiency reached up to 95.4% for Escherichia coli (E. coli). Furthermore, quantitative analysis of the bacteria was achieved by using acridine orange (AO) as a fluorescence probe for the captured E. coli due to its ability of staining series types of bacteria and rapid labeling. In order to remove the free magnetic nanoparticles and redundant fluorescent reagent, the labeled suspension was poured into a PEG separation column and was separated by applying an external magnetic field. The presence of 100 cfu mL -1 E. coli could be detected for semi-quantitative analysis by observing the separation column with the naked eye, and the concentration could be further evaluated by fluorescence detection. All the above processes were finished within 80 min. It was demonstrated that a good linear relationship existed between the fluorescence intensity and the concentration of E. coli ranging from 10 2 to 10 6  cfu mL -1 , with a detection limit of 100 cfu mL -1 when E. coli acted as target bacteria. The recovery rate of E. coli was 93.6∼102.0% in tap water and cooked meat samples, and the RSD was lower than 7% (n = 6); the result coincided with the conventional plate count method. Graphical abstract ᅟ.

E. coli survival in waters: applicability of the Arrhenius equation

USDA-ARS?s Scientific Manuscript database

E. coli is an important microorganism indicator used to show the presence of pathogens and fecal contamination in waters. Knowing E. coli survival rates is important for assessing the severity of contamination that has occurred and making appropriate management evaluations. E. coli survival rates ...

Preliminary survey of antibiotic-resistant fecal indicator bacteria and pathogenic Escherichia coli from river-water samples collected in Oakland County, Michigan, 2003

USGS Publications Warehouse

Fogarty, Lisa R.; Duris, Joseph W.; Aichele, Stephen S.

2005-01-01

A preliminary study was done in Oakland County, Michigan, to determine the concentration of fecal indicator bacteria (fecal coliform bacteria and enterococci), antibiotic resistance patterns of these two groups, and the presence of potentially pathogenic Escherichia coli (E. coli). For selected sites, specific members of these groups [E. coli, Enterococcus faecium (E. faecium) and Enterococcus faecalis (E. faecalis)] were isolated and tested for levels of resistance to specific antibiotics used to treat human infections by pathogens in these groups and for their potential to transfer these resistances. In addition, water samples from all sites were tested for indicators of potentially pathogenic E. coli by three assays: a growth-based assay for sorbitol-negative E. coli, an immunological assay for E. coli O157, and a molecular assay for three virulence and two serotype genes. Samples were also collected from two non-urbanized sites outside of Oakland County. Results from the urbanized Oakland County area were compared to those from these two non-urbanized sites. Fecal indicator bacteria concentrations exceeded State of Michigan recreational water-quality standards and (or) recommended U.S. Environmental Protection Agency (USEPA) standards in samples from all but two Oakland County sites. Multiple-antibiotic-resistant fecal coliform bacteria were found at all sites, including two reference sites from outside the county. Two sites (Stony Creek and Paint Creek) yielded fecal coliform isolates resistant to all tested antibiotics. Patterns indicative of extended-spectrum-β-lactamase (ESBL)- producing fecal coliform bacteria were found at eight sites in Oakland County and E. coli resistant to clinically significant antibiotics were recovered from the River Rouge, Clinton River, and Paint Creek. Vancomycin-resistant presumptive enterococci were found at six sites in Oakland County and were not found at the reference sites. Evidence of acquired antibiotic resistances was detected in bacteria from multiple sites in Oakland County but not detected in bacteria from the reference sites. Integrons capable of transferring resistance were detected in isolates from the River Rouge and Clinton River. E. faecium and E. faecalis identified in samples collected from Kearsley Creek and Evans Ditch were resistant to high levels of vancomycin and carried transferable genes responsible for resistance. Several sites in Oakland County had indicators of pathogenic E. coli in August and (or) September 2003. Two samples from the Clinton River in August tested positive for all three E. coli O157 tests. Both the August and September samples from one River Rouge site were positive for the immunological and molecular assay for E. coli O157. A combination of virulence genes commonly associated with human illness was detected at five sites in August and seven sites in September. Antibiotic-resistance profiles of clinical concern along with genes capable of transferring the resistance were found at several sites throughout Oakland County; samples from many of these sites also contained potentially pathogenic E. coli.

Widespread Fosfomycin Resistance in Gram-Negative Bacteria Attributable to the Chromosomal fosA Gene

PubMed Central

Ito, Ryota; Tomich, Adam D.; Callaghan, Jake D.; McElheny, Christi L.; Mettus, Roberta T.; Sluis-Cremer, Nicolas

2017-01-01

ABSTRACT Fosfomycin is a decades-old antibiotic which is being revisited because of its perceived activity against many extensively drug-resistant Gram-negative pathogens. FosA proteins are Mn2+ and K+-dependent glutathione S-transferases which confer fosfomycin resistance in Gram-negative bacteria by conjugation of glutathione to the antibiotic. Plasmid-borne fosA variants have been reported in fosfomycin-resistant Escherichia coli strains. However, the prevalence and distribution of fosA in other Gram-negative bacteria are not known. We systematically surveyed the presence of fosA in Gram-negative bacteria in over 18,000 published genomes from 18 Gram-negative species and investigated their contribution to fosfomycin resistance. We show that FosA homologues are present in the majority of genomes in some species (e.g., Klebsiella spp., Enterobacter spp., Serratia marcescens, and Pseudomonas aeruginosa), whereas they are largely absent in others (e.g., E. coli, Acinetobacter baumannii, and Burkholderia cepacia). FosA proteins in different bacterial pathogens are highly divergent, but key amino acid residues in the active site are conserved. Chromosomal fosA genes conferred high-level fosfomycin resistance when expressed in E. coli, and deletion of chromosomal fosA in S. marcescens eliminated fosfomycin resistance. Our results indicate that FosA is encoded by clinically relevant Gram-negative species and contributes to intrinsic fosfomycin resistance. PMID:28851843

Prevalence of foodborne pathogens in grilled chicken from street vendors and retail outlets in Reynosa, Tamaulipas, Mexico.

PubMed

Díaz-López, A; Cantú-Ramírez, R C; Garza-González, E; Ruiz-Tolentino, L; Tellez-Luis, S J; Rivera, G; Bocanegra-García, V

2011-08-01

We analyzed a total of 70 grilled chicken samples bought randomly from street vendors and retail outlets in the city of Reynosa, Mexico, to determine the prevalence of Escherichia coli (Shiga toxin producing and enterotoxin producing), Salmonella spp., Staphylococcus aureus, Listeria spp., and Campylobacter spp. using microbiological methods and PCR detection of bacterial sequences. Of the 70 samples, 27 (38.5%) were from retail outlets and 43 (61.4%) from street vendors. All specimens were negative by both microbiological and molecular methods for Listeria monocytogenes, Shiga toxin 2 of Shiga toxin-producing E. coli, lt of enterotoxin-producing E. coli, and st enterotoxin, and all were negative for Salmonella spp. and Campylobacter jejuni by PCR. Of the samples studied, 49 (70%) had undetectable levels of the foodborne pathogens studied with the methods used. In the remaining 21 (30%) specimens, at least one pathogen was isolated or detected, with E. coli being the pathogen most frequently isolated and with two samples bearing the hlyA gene. We found no statistical difference in bacterial prevalence between retail and street vendor samples. The presence of pathogens in grilled chicken is an important public health risk because of the great demand for and daily consumption of this product in this region.

Microbiological quality of water from the rivers of Curitiba, Paraná State, Brazil, and the susceptibility to antimicrobial drugs and pathogenicity of Escherichia coli.

PubMed

Giowanella, Melissa; Bozza, Angela; do Rocio Dalzoto, Patricia; Dionísio, Jair Alves; Andraus, Sumaia; Guimarães, Edson Luiz Gomes; Pimentel, Ida Chapaval

2015-11-01

Water safety is determined by several markers, and Escherichia coli is one of the most important indicators of water quality. The objective of this study was to evaluate the microbiological parameters in environmental samples of fresh water from rivers of Curitiba and its metropolitan area in Paraná State, Brazil. In addition, we evaluated the pathogenicity and susceptibility to antimicrobial drugs in E. coli. These evaluations were performed by quantitative and qualitative methods employing selective media for isolating thermotolerant coliforms and biochemical tests for identifying E. coli. Pathogenic strains of E. coli were detected by PCR multiplex using specific primers. From the water samples, 494 thermotolerant coliforms were obtained, of which 96 (19.43%) isolates were characterized as E. coli. Three isolates were identified as enteroaggregative E. coli, one as enterotoxigenic E. coli, one as enteropathogenic E. coli, and two carried the Eae virulence gene. E. coli susceptibility to commonly employed antimicrobial drugs was analyzed by the disc diffusion method. The results showed 49 (51.04%) isolates resistant to all the drugs assayed, 16 (16.67%) with an intermediate resistance to all drugs, and 31 (32.29%) intermediately or fully resistant to one or more drugs tested. The highest rate of resistance was observed for tetracycline 30 μg, streptomycin 10 μg, and ceftazidime 30 μg. Detection of E. coli is associated with water contamination by fecal material from humans and warm-blooded animals. The occurrence of resistant strains can be the result of the indiscriminate use of antimicrobial drugs and poor sanitation in the areas assayed.

Biofilm Formation and Dispersal under the Influence of the Global Regulator CsrA of Escherichia coli

PubMed Central

Jackson, Debra W.; Suzuki, Kazushi; Oakford, Lawrence; Simecka, Jerry W.; Hart, Mark E.; Romeo, Tony

2002-01-01

The predominant mode of growth of bacteria in the environment is within sessile, matrix-enclosed communities known as biofilms. Biofilms often complicate chronic and difficult-to-treat infections by protecting bacteria from the immune system, decreasing antibiotic efficacy, and dispersing planktonic cells to distant body sites. While the biology of bacterial biofilms has become a major focus of microbial research, the regulatory mechanisms of biofilm development remain poorly defined and those of dispersal are unknown. Here we establish that the RNA binding global regulatory protein CsrA (carbon storage regulator) of Escherichia coli K-12 serves as both a repressor of biofilm formation and an activator of biofilm dispersal under a variety of culture conditions. Ectopic expression of the E. coli K-12 csrA gene repressed biofilm formation by related bacterial pathogens. A csrA knockout mutation enhanced biofilm formation in E. coli strains that were defective for extracellular, surface, or regulatory factors previously implicated in biofilm formation. In contrast, this csrA mutation did not affect biofilm formation by a glgA (glycogen synthase) knockout mutant. Complementation studies with glg genes provided further genetic evidence that the effects of CsrA on biofilm formation are mediated largely through the regulation of intracellular glycogen biosynthesis and catabolism. Finally, the expression of a chromosomally encoded csrA′-′lacZ translational fusion was dynamically regulated during biofilm formation in a pattern consistent with its role as a repressor. We propose that global regulation of central carbon flux by CsrA is an extremely important feature of E. coli biofilm development. PMID:11741870

Cloacael Carriage and Multidrug Resistance Escherichia coli O157:H7 from Poultry Farms, Eastern Ethiopia

PubMed Central

Shecho, Mude; Muktar, Yimer

2017-01-01

A cross-sectional study was carried out to determine antimicrobial drug resistance patterns of E. coli O157:H7 isolates and estimate the level of the pathogen. A total of 194 cloacae swab samples were collected randomly in two poultry farms. Standard cultural, biochemical, and serological (latex agglutination) methods were used to isolate E. coli O157:H7. The isolates were subjected to antimicrobial susceptibility testing using disc diffusion method. Out of 194 cloacae samples examined, 13.4% (n = 26) were found to be positive for E. coli O157:H7. The finding indicated differences in E. coli O157:H7 infection among the different risk factors. Chicken from Adele Poultry Farm showed higher E. coli O157:H7 infection (OR = 3.89) than Haramaya University poultry farm and young birds had more infection (OR = 4.62) than adult birds. Of the total 14 antimicrobials included in the panel of study, the susceptibility results were varied with 96.15% and 0% E. coli O157:H7 isolates expressing resistance to erythromycin, clindamycin, spectinomycin, and ciprofloxacin, respectively. Multidrug resistance to more than two antimicrobial agents was detected in 24 (92.30%) of the isolates. The study showed high presence of antimicrobial resistant isolates of E. coli O157:H7. Further study is required to better understand the ecology and evolution of bacterial resistance to antimicrobial agents. PMID:28349121

Cloacael Carriage and Multidrug Resistance Escherichia coli O157:H7 from Poultry Farms, Eastern Ethiopia.

PubMed

Shecho, Mude; Thomas, Naod; Kemal, Jelalu; Muktar, Yimer

2017-01-01

A cross-sectional study was carried out to determine antimicrobial drug resistance patterns of E. coli O157:H7 isolates and estimate the level of the pathogen. A total of 194 cloacae swab samples were collected randomly in two poultry farms. Standard cultural, biochemical, and serological (latex agglutination) methods were used to isolate E. coli O157:H7. The isolates were subjected to antimicrobial susceptibility testing using disc diffusion method. Out of 194 cloacae samples examined, 13.4% ( n = 26) were found to be positive for E. coli O157:H7. The finding indicated differences in E. coli O157:H7 infection among the different risk factors. Chicken from Adele Poultry Farm showed higher E. coli O157:H7 infection (OR = 3.89) than Haramaya University poultry farm and young birds had more infection (OR = 4.62) than adult birds. Of the total 14 antimicrobials included in the panel of study, the susceptibility results were varied with 96.15% and 0% E. coli O157:H7 isolates expressing resistance to erythromycin, clindamycin, spectinomycin, and ciprofloxacin, respectively. Multidrug resistance to more than two antimicrobial agents was detected in 24 (92.30%) of the isolates. The study showed high presence of antimicrobial resistant isolates of E. coli O157:H7. Further study is required to better understand the ecology and evolution of bacterial resistance to antimicrobial agents.

The Alkaloid Compound Harmane Increases the Lifespan of Caenorhabditis elegans during Bacterial Infection, by Modulating the Nematode’s Innate Immune Response

PubMed Central

Marinus, Martin G.; Xu, Tao; Struve, Carsten; Krogfelt, Karen A.; Løbner-Olesen, Anders

2013-01-01

The nematode Caenorhabditis elegans has in recent years been proven to be a powerful in vivo model for testing antimicrobial compounds. We report here that the alkaloid compound Harmane (2-methyl-β-carboline) increases the lifespan of nematodes infected with a human pathogen, the Shiga toxin-producing Escherichia coli O157:H7 strain EDL933 and several other bacterial pathogens. This was shown to be unrelated to the weak antibiotic effect of Harmane. Using GFP-expressing E. coli EDL933, we showed that Harmane does not lower the colonization burden in the nematodes. We also found that the expression of the putative immune effector gene F35E12.5 was up-regulated in response to Harmane treatment. This indicates that Harmane stimulates the innate immune response of the nematode; thereby increasing its lifespan during bacterial infection. Expression of F35E12.5 is predominantly regulated through the p38 MAPK pathway; however, intriguingly the lifespan extension resulting from Harmane was higher in p38 MAPK-deficient nematodes. This indicates that Harmane has a complex effect on the innate immune system of C. elegans. Harmane could therefore be a useful tool in the further research into C. elegans immunity. Since the innate immunity of C. elegans has a high degree of evolutionary conservation, drugs such as Harmane could also be possible alternatives to classic antibiotics. The C. elegans model could prove to be useful for selection and development of such drugs. PMID:23544153

The dps gene of symbiotic "Candidatus Legionella jeonii" in Amoeba proteus responds to hydrogen peroxide and phagocytosis.

PubMed

Park, Miey; Yun, Seong Tae; Hwang, Sue-Yun; Chun, Choong-Ill; Ahn, Tae In

2006-11-01

To survive in host cells, intracellular pathogens or symbiotic bacteria require protective mechanisms to overcome the oxidative stress generated by phagocytic activities of the host. By genomic library tagging, we cloned a dps (stands for DNA-binding protein from starved cells) gene of the symbiotic "Candidatus Legionella jeonii" organism (called the X bacterium) (dps(X)) that grows in Amoeba proteus. The gene encodes a 17-kDa protein (pI 5.19) with 91% homology to Dps and DNA-binding ferritin-like proteins of other organisms. The cloned gene complemented the dps mutant of Escherichia coli and conferred resistance to hydrogen peroxide. Dps(X) proteins purified from E. coli transformed with the dps(X) gene were in oligomeric form, formed a complex with pBlueskript SKII DNA, and protected the DNA from DNase I digestion and H(2)O(2)-mediated damage. The expression of the dps(X) gene in "Candidatus Legionella jeonii" was enhanced when the host amoeba was treated with 2 mM H(2)O(2) and by phagocytic activities of the host cell. These results suggested that the Dps protein has a function protective of the bacterial DNA and that its gene expression responds to oxidative stress generated by phagocytic activities of the host cell. With regard to the fact that invasion of Legionella sp. into respiratory phagocytic cells causes pneumonia in mammals, further characterization of dps(X) expression in the Legionella sp. that multiplies in a protozoan host in the natural environment may provide valuable information toward understanding the protective mechanisms of intracellular pathogens.

Nutritional basis for colonization resistance by human commensal Escherichia coli strains HS and Nissle 1917 against E. coli O157:H7 in the mouse intestine.

PubMed

Maltby, Rosalie; Leatham-Jensen, Mary P; Gibson, Terri; Cohen, Paul S; Conway, Tyrrell

2013-01-01

Escherichia coli is a single species consisting of many biotypes, some of which are commensal colonizers of mammals and others that cause disease. Humans are colonized on average with five commensal biotypes, and it is widely thought that the commensals serve as a barrier to infection by pathogens. Previous studies showed that a combination of three pre-colonized commensal E. coli strains prevents colonization of E. coli O157:H7 in a mouse model (Leatham, et al., 2010, Infect Immun 77: 2876-7886). The commensal biotypes included E. coli HS, which is known to successfully colonize humans at high doses with no adverse effects, and E. coli Nissle 1917, a human commensal strain that is used in Europe as a preventative of traveler's diarrhea. We hypothesized that commensal biotypes could exert colonization resistance by consuming nutrients needed by E. coli O157:H7 to colonize, thus preventing this first step in infection. Here we report that to colonize streptomycin-treated mice E. coli HS consumes six of the twelve sugars tested and E. coli Nissle 1917 uses a complementary yet divergent set of seven sugars to colonize, thus establishing a nutritional basis for the ability of E. coli HS and Nissle 1917 to occupy distinct niches in the mouse intestine. Together these two commensals use the five sugars previously determined to be most important for colonization of E. coli EDL933, an O157:H7 strain. As predicted, the two commensals prevented E. coli EDL933 colonization. The results support a model in which invading pathogenic E. coli must compete with the gut microbiota to obtain the nutrients needed to colonize and establish infection; accordingly, the outcome of the challenge is determined by the aggregate capacity of the native microbiota to consume the nutrients required by the pathogen.

Molecular epidemiological view on Shiga toxin-producing Escherichia coli causing human disease in Germany: Diversity, prevalence, and outbreaks.

PubMed

Fruth, Angelika; Prager, Rita; Tietze, Erhard; Rabsch, Wolfgang; Flieger, Antje

2015-10-01

Infections by intestinal pathogenic Escherichia coli (E. coli) are among those causing a high mortality and morbidity due to diarrheal disease and post infection sequelae worldwide. Since introduction of the Infection Protection Act in Germany 2001, these pathogens rank third among bacterial infections of the gastrointestinal tract. As a major pathovar Shiga toxin-producing E. coli (STEC) which include enterohemorrhagic E. coli (EHEC) play a leading role in occurrence of sporadic cases and disease outbreaks. An outstanding example is the large outbreak in spring 2011 caused by EHEC/EAEC O104:H4. To monitor and trace back STEC infections, national surveillance programs have been implemented including activities of the German National Reference Centre for Salmonella and other Enteric Bacterial Pathogens (NRC). This review highlights advances in our understanding of STEC in the last 20 years of STEC surveillance by the NRC. Here important characteristics of STEC strains from human infections and outbreaks in Germany between 1997 and 2013 are summarized. Copyright © 2015. Published by Elsevier GmbH.

ISOLATION AND MOLECULAR IDENTIFICATION OF POTENTIALLY PATHOGENIC Escherichia coli AND Campylobacter jejuni IN FERAL PIGEONS FROM AN URBAN AREA IN THE CITY OF LIMA, PERU

PubMed Central

CABALLERO, Moisés; RIVERA, Isabel; JARA, Luis M.; ULLOA-STANOJLOVIC, Francisco M.; SHIVA, Carlos

2015-01-01

SUMMARY Feral pigeons (Columbia livia) live in close contact with humans and other animals. They can transmit potentially pathogenic and zoonotic agents. The objective of this study was to isolate and detect strains of diarrheagenic Escherichia coli and Campylobacter jejuni of urban feral pigeons from an area of Lima, Peru. Fresh dropping samples from urban parks were collected for microbiological isolation of E. coli strains in selective agar, and Campylobacter by filtration method. Molecular identification of diarrheagenic pathotypes of E.coli and Campylobacter jejuni was performed by PCR. Twenty-two parks were sampled and 16 colonies of Campylobacter spp. were isolated. The 100% of isolates were identified as Campylobacter jejuni. Furthermore, 102 colonies of E. coliwere isolated and the 5.88% resulted as Enteropathogenic (EPEC) type and 0.98% as Shiga toxin-producing E. coli (STEC). The urban feral pigeons of Lima in Peru can act as a reservoir or carriers of zoonotic potentially pathogenic enteric agents. PMID:26603225

Enteric pathogen-plant interactions: molecular connections leading to colonization and growth and implications for food safety.

PubMed

Martínez-Vaz, Betsy M; Fink, Ryan C; Diez-Gonzalez, Francisco; Sadowsky, Michael J

2014-01-01

Leafy green vegetables have been identified as a source of foodborne illnesses worldwide over the past decade. Human enteric pathogens, such as Escherichia coli O157:H7 and Salmonella, have been implicated in numerous food poisoning outbreaks associated with the consumption of fresh produce. An understanding of the mechanisms responsible for the establishment of pathogenic bacteria in or on vegetable plants is critical for understanding and ameliorating this problem as well as ensuring the safety of our food supply. While previous studies have described the growth and survival of enteric pathogens in the environment and also the risk factors associated with the contamination of vegetables, the molecular events involved in the colonization of fresh produce by enteric pathogens are just beginning to be elucidated. This review summarizes recent findings on the interactions of several bacterial pathogens with leafy green vegetables. Changes in gene expression linked to the bacterial attachment and colonization of plant structures are discussed in light of their relevance to plant-microbe interactions. We propose a mechanism for the establishment and association of enteric pathogens with plants and discuss potential strategies to address the problem of foodborne illness linked to the consumption of leafy green vegetables.

Different Cellular Origins and Functions of Extracellular Proteins from Escherichia coli O157:H7 and O104:H4 as Determined by Comparative Proteomic Analysis

PubMed Central

Islam, Nazrul; Nagy, Attila; Garrett, Wesley M.; Shelton, Dan

2016-01-01

ABSTRACT Extracellular proteins play important roles in bacterial interactions with the environmental matrices. In this study, we examined the extracellular proteins from Escherichia coli O157:H7 and O104:H4 by tandem mass spectrometry. We identified 500 and 859 proteins from the growth media of E. coli O157:H7 and O104:H4, respectively, including 371 proteins common to both strains. Among proteins that were considered specific to E. coli O157:H7 or present at higher relative abundances in O157:H7 medium, most (57 of 65) had secretion signal sequences in their encoding genes. Noticeably, the proteins included locus of enterocyte effacement (LEE) virulence factors, proteins required for peptidyl-lipoprotein accumulation, and proteins involved in iron scavenging. In contrast, a much smaller proportion of proteins (37 of 150) that were considered specific to O104:H4 or presented at higher relative abundances in O104:H4 medium had signals targeting them for secretion. These proteins included Shiga toxin 2 subunit B and O104:H4 signature proteins, including AAF/1 major fimbrial subunit and serine protease autotransporters. Most of the abundant proteins from the growth medium of E. coli O104:H4 were annotated as having functions in the cytoplasm. We provide evidence that the extensive presence of cytoplasmic proteins in E. coli O104:H4 growth medium was due to biological processes independent of cell lysis, indicating alternative mechanisms for this potent pathogen releasing cytoplasmic contents into the growth milieu, which could play a role in interaction with the environmental matrices, such as pathogenesis and biofilm formation. IMPORTANCE In this study, we compared the extracellular proteins from two of the most prominent foodborne pathogenic E. coli organisms that have caused severe outbreaks in the United States and in Europe. E. coli O157:H7 is a well-studied Shiga toxigenic foodborne pathogen of the enterohemorrhagic pathotype that has caused numerous outbreaks associated with various contaminated foods worldwide. E. coli O104:H4 is a newly emerged Shiga toxigenic foodborne pathogen of the enteroaggregative pathotype that gained notoriety for causing one of the most deadly foodborne outbreaks in Europe in 2011. Comparison of proteins in the growth medium revealed significant differences in the compositions of the extracellular proteins for these two pathogens. These differences may provide valuable information regarding the cellular responses of these pathogens to their environment, including cell survival and pathogenesis. PMID:27208096

A small diffusible signal molecule is responsible for the global control of virulence and exoenzyme production in the plant pathogen Erwinia carotovora.

PubMed Central

Pirhonen, M; Flego, D; Heikinheimo, R; Palva, E T

1993-01-01

Virulence of the plant pathogen Erwinia carotovora subsp. carotovora is dependent on the production and secretion of a complex arsenal of plant cell wall-degrading enzymes. Production of these exoenzymes is controlled by a global regulatory mechanism. A virulent mutants in one of the regulatory loci, expI, show a pleiotropic defect in the growth phase-dependent transcriptional activation of exoenzyme gene expression. The expI gene encodes a 26 kDa polypeptide that is structurally and functionally related to the luxI gene product of Vibrio fischeri. Functional similarity of expI and luxI has been demonstrated by reciprocal genetic complementation experiments. LuxI controls bioluminescence in V.fischeri in a growth phase-dependent manner by directing the synthesis of the diffusible autoinducer, N-(3-oxohexanoyl) homoserine lactone. E.c. subsp. carotovora expI+ strains or Escherichia coli harboring the cloned expI gene excrete a small diffusible signal molecule that complements the expI mutation of Erwinia as well as a luxI mutation of V.fischeri. This extracellular complementation can also be achieved by E.coli harboring the luxI gene from V.fischeri or by adding the synthetic V.fischeri autoinducer. Both the production of the plant tissue-macerating exoenzymes and the ability of the bacteria to propagate in planta are restored in expI mutants by autoinducer addition. These data suggest that the same signal molecule is employed in control of such diverse processes as virulence in a plant pathogen and bioluminescence in a marine bacterium, and may represent a general mechanism by which bacteria modulate gene expression in response to changing environmental conditions. Images PMID:8508772

A small diffusible signal molecule is responsible for the global control of virulence and exoenzyme production in the plant pathogen Erwinia carotovora.

PubMed

Pirhonen, M; Flego, D; Heikinheimo, R; Palva, E T

1993-06-01

Virulence of the plant pathogen Erwinia carotovora subsp. carotovora is dependent on the production and secretion of a complex arsenal of plant cell wall-degrading enzymes. Production of these exoenzymes is controlled by a global regulatory mechanism. A virulent mutants in one of the regulatory loci, expI, show a pleiotropic defect in the growth phase-dependent transcriptional activation of exoenzyme gene expression. The expI gene encodes a 26 kDa polypeptide that is structurally and functionally related to the luxI gene product of Vibrio fischeri. Functional similarity of expI and luxI has been demonstrated by reciprocal genetic complementation experiments. LuxI controls bioluminescence in V.fischeri in a growth phase-dependent manner by directing the synthesis of the diffusible autoinducer, N-(3-oxohexanoyl) homoserine lactone. E.c. subsp. carotovora expI+ strains or Escherichia coli harboring the cloned expI gene excrete a small diffusible signal molecule that complements the expI mutation of Erwinia as well as a luxI mutation of V.fischeri. This extracellular complementation can also be achieved by E.coli harboring the luxI gene from V.fischeri or by adding the synthetic V.fischeri autoinducer. Both the production of the plant tissue-macerating exoenzymes and the ability of the bacteria to propagate in planta are restored in expI mutants by autoinducer addition. These data suggest that the same signal molecule is employed in control of such diverse processes as virulence in a plant pathogen and bioluminescence in a marine bacterium, and may represent a general mechanism by which bacteria modulate gene expression in response to changing environmental conditions.

Pathogenesis of human diffusely adhering Escherichia coli expressing Afa/Dr adhesins (Afa/Dr DAEC): current insights and future challenges.

PubMed

Servin, Alain L

2014-10-01

The pathogenicity and clinical pertinence of diffusely adhering Escherichia coli expressing the Afa/Dr adhesins (Afa/Dr DAEC) in urinary tract infections (UTIs) and pregnancy complications are well established. In contrast, the implication of intestinal Afa/Dr DAEC in diarrhea is still under debate. These strains are age dependently involved in diarrhea in children, are apparently not involved in diarrhea in adults, and can also be asymptomatic intestinal microbiota strains in children and adult. This comprehensive review analyzes the epidemiology and diagnosis and highlights recent progress which has improved the understanding of Afa/Dr DAEC pathogenesis. Here, I summarize the roles of Afa/Dr DAEC virulence factors, including Afa/Dr adhesins, flagella, Sat toxin, and pks island products, in the development of specific mechanisms of pathogenicity. In intestinal epithelial polarized cells, the Afa/Dr adhesins trigger cell membrane receptor clustering and activation of the linked cell signaling pathways, promote structural and functional cell lesions and injuries in intestinal barrier, induce proinflammatory responses, create angiogenesis, instigate epithelial-mesenchymal transition-like events, and lead to pks-dependent DNA damage. UTI-associated Afa/Dr DAEC strains, following adhesin-membrane receptor cell interactions and activation of associated lipid raft-dependent cell signaling pathways, internalize in a microtubule-dependent manner within urinary tract epithelial cells, develop a particular intracellular lifestyle, and trigger a toxin-dependent cell detachment. In response to Afa/Dr DAEC infection, the host epithelial cells generate antibacterial defense responses. Finally, I discuss a hypothetical role of intestinal Afa/Dr DAEC strains that can act as "silent pathogens" with the capacity to emerge as "pathobionts" for the development of inflammatory bowel disease and intestinal carcinogenesis. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Transcriptome Analysis of Escherichia coli O157:H7 Exposed to Lysates of Lettuce Leaves ▿

PubMed Central

Kyle, Jennifer L.; Parker, Craig T.; Goudeau, Danielle; Brandl, Maria T.

2010-01-01

Harvesting and processing of leafy greens inherently cause plant tissue damage, creating niches on leaves that human pathogens can exploit. We previously demonstrated that Escherichia coli O157:H7 (EcO157) multiplies more rapidly on shredded leaves than on intact leaves (M. T. Brandl, Appl. Environ. Microbiol. 74:5285-5289, 2008). To investigate how EcO157 cells adapt to physicochemical conditions in injured lettuce tissue, we used microarray-based whole-genome transcriptional profiling to characterize gene expression patterns in EcO157 after 15- and 30-min exposures to romaine lettuce lysates. Multiple carbohydrate transport systems that have a role in the utilization of substrates known to be prevalent in plant cells were activated in EcO157. This indicates the availability to the human pathogen of a variety of carbohydrates released from injured plant cells that may promote its extensive growth in leaf lysates and, thus, in wounded leaf tissue. In addition, microarray analysis revealed the upregulation of numerous genes associated with EcO157 attachment and virulence, with oxidative stress and antimicrobial resistance (including the OxyR and Mar regulons), with detoxification of noxious compounds, and with DNA repair. Upregulation of oxidative stress and antimicrobial resistance genes in EcO157 was confirmed on shredded lettuce by quantitative reverse transcription-PCR. We further demonstrate that this adaptation to stress conditions imparts the pathogen with increased resistance to hydrogen peroxide and calcium hypochlorite. This enhanced resistance to chlorinated sanitizers combined with increased expression of virulence determinants and multiplication at sites of injury on the leaves may help explain the association of processed leafy greens with outbreaks of EcO157. PMID:20061451

Complete Genome Sequence of the Avian-Pathogenic Escherichia coli Strain APEC O18

PubMed Central

Nicholson, Bryon A.; Wannemuehler, Yvonne M.; Logue, Catherine M.; Li, Ganwu

2016-01-01

Avian-pathogenic Escherichia coli (APEC) is the causative agent of colibacillosis, a disease that affects all facets of poultry production worldwide, resulting in multimillion dollar losses annually. Here, we report the genome sequence of an APEC O18 sequence type 95 (ST95) strain associated with disease in a chicken. PMID:27811098

Comparison of U.S. Environmental Protection Agency and U.S. Composting Council... Escherichia coli O157:H7 in finished compost

USDA-ARS?s Scientific Manuscript database

Composting management or conditions that result in inadequate exposure of the compostable materials to destructive time-temperature regimens can result in survival of enteric human pathogens. Bacterial pathogens, such as Escherichia coli O157:H7 and Salmonella spp., can regrow in finished compost. ...

Automated immunomagnetic separation for the detection of Escherichia coli O157:H7 from spinach

USDA-ARS?s Scientific Manuscript database

Escherichia coli O157:H7 is a major cause of foodborne illness and methods for rapid and sensitive detection of this deadly pathogen are needed to protect consumers. The use of immunomagnetic separation (IMS) for the capture and concentration of foodborne pathogens has been gaining popularity, in p...

Evolution of a zoonotic pathogen: investigating prophage diversity in enterohaemorrhagic E. coli O157 by long-read sequencing

USDA-ARS?s Scientific Manuscript database

Enterohaemorrhagic Escherichia Coli (EHEC) is a zoonotic pathogen known to be potentially lethal in humans. Its main animal reservoir is ruminants, specifically cattle, and yearly outbreaks occur worldwide with the most prevalent serotype being EHEC O157:H7. Most virulence factors of EHEC O157, incl...

Of woods and webs: possible alternatives to the tree of life for studying genomic fluidity in E. coli.

PubMed

Beauregard-Racine, Julie; Bicep, Cédric; Schliep, Klaus; Lopez, Philippe; Lapointe, François-Joseph; Bapteste, Eric

2011-07-20

We introduce several forest-based and network-based methods for exploring microbial evolution, and apply them to the study of thousands of genes from 30 strains of E. coli. This case study illustrates how additional analyses could offer fast heuristic alternatives to standard tree of life (TOL) approaches. We use gene networks to identify genes with atypical modes of evolution, and genome networks to characterize the evolution of genetic partnerships between E. coli and mobile genetic elements. We develop a novel polychromatic quartet method to capture patterns of recombination within E. coli, to update the clanistic toolkit, and to search for the impact of lateral gene transfer and of pathogenicity on gene evolution in two large forests of trees bearing E. coli. We unravel high rates of lateral gene transfer involving E. coli (about 40% of the trees under study), and show that both core genes and shell genes of E. coli are affected by non-tree-like evolutionary processes. We show that pathogenic lifestyle impacted the structure of 30% of the gene trees, and that pathogenic strains are more likely to transfer genes with one another than with non-pathogenic strains. In addition, we propose five groups of genes as candidate mobile modules of pathogenicity. We also present strong evidence for recent lateral gene transfer between E. coli and mobile genetic elements. Depending on which evolutionary questions biologists want to address (i.e. the identification of modules, genetic partnerships, recombination, lateral gene transfer, or genes with atypical evolutionary modes, etc.), forest-based and network-based methods are preferable to the reconstruction of a single tree, because they provide insights and produce hypotheses about the dynamics of genome evolution, rather than the relative branching order of species and lineages. Such a methodological pluralism - the use of woods and webs - is to be encouraged to analyse the evolutionary processes at play in microbial evolution.This manuscript was reviewed by: Ford Doolittle, Tal Pupko, Richard Burian, James McInerney, Didier Raoult, and Yan Boucher.

The arrhenius equation as means to simulate E. Coli survival in waters

USDA-ARS?s Scientific Manuscript database

E. coli is an important microorganism indicator used to show the presence of pathogens and fecal contamination in waters. Knowing E. coli survival rates is important for assessing the severity of contamination that has occurred and making appropriate management evaluations. E. coli survival rates ...

Editorial: Emerging approaches for typing, detection, characterization, and traceback of Escherichia coli

USDA-ARS?s Scientific Manuscript database

Commensal E. coli inhabit the large intestines of humans and animals and are important in maintaining normal intestinal homeostasis. There are also many groups of disease-causing E. coli, including diarrheagenic and extra-intestinal pathogenic E. coli (ExPEC). There are approximately O188 somatic O...

Characteristics of volatile organic compounds produced from five pathogenic bacteria by headspace-solid phase micro-extraction/gas chromatography-mass spectrometry.

PubMed

Chen, Juan; Tang, Junni; Shi, Hui; Tang, Cheng; Zhang, Rong

2017-03-01

The characteristics of volatile compounds from five different bacterial species, Escherichia coli O157:H7, Salmonella Enteritidis, Shigella flexneri, Staphylococcus aureus, and Listeria monocytogenes, growing, respectively, in trypticase soy broth were monitored by headspace solid-phase micro-extraction/gas chromatography-mass spectrometry. The results showed that most volatile organic compounds (VOCs) of five pathogens started to increase after the sixth to tenth hour. Methyl ketones and long chain alcohols were representative volatiles for three Gram-negative bacteria. The especially high production of indole was characterized to E. coli O157:H7. The production of 3-hydroxy-2-butanone was indicative of the presence of two Gram-positive bacteria. Both 3-methyl-butanoic acid and 3-methyl-butanal were unique biomarkers for S. aureus. The population dynamics of individual pathogen could be monitored using the accumulation of VOCs correlated with its growth. And these five pathogens could be distinguishable though principle component analysis of 18 volatile metabolites. Moreover, the mixed culture of S. aureus and E. coli O157:H7 was also investigated. The levels of 3-methyl-butanal and 3-methyl-butanoic acid were largely reduced; while the level of indole almost unchanged and correlated with E. coli O157:H7 growth very well. The characteristics of volatiles from the five foodborne pathogens could lay a fundamental basis for further research into pathogen contamination control by detecting volatile signatures of pathogens. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Occurrence and antibacterial susceptibility pattern of bacterial pathogens isolated from diarrheal patients in Pakistan.

PubMed

Rasool, Muhammad H; Siddique, Abu B; Saqalein, Muhammad; Asghar, Muhammad J; Zahoor, Muhammad A; Aslam, Bilal; Shafiq, Humerah B; Nisar, Muhammad A

2016-03-01

To determine the occurrence of bacterial pathogens responsible for diarrhea and to engender information regarding the effectiveness of commonly used antibiotic against diarrhea. This cross-sectional study was conducted between April and July 2014. Samples were collected from the Divisional Headquarter and Allied Hospital, Faisalabad, Pakistan. The differential and selective media were used to isolate bacterial pathogens, which were identified through cultural characteristics, microscopy, and biochemical tests. Disc diffusion assay was carried out using Muller Hinton agar medium, and minimum inhibitory concentration was determined using broth dilution method against isolated pathogens. One hundred and forty-one (100%) samples were positive for some bacteria. Frequency of occurrence was Bacillus cereus (B. cereus) (66%), Escherichia coli (E.coli) (48.5%), Salmonella typhi (S. Typhi) (27.7%), Pseudomonas aeruginosa (P. aeruginosa) (8.5%), and Staphylococcus aureus (S. aureus) (4.3%). Single pathogen was detected in 20 (14.2%) samples whereas combinations were found in 121 (85.8%) samples. Bacillus cereus and E.coli were the most frequently detected pathogens followed by the S. Typhi, P. aeruginosa, and Staph. aureus. The percentage occurrence of isolated pathogens was 31% in B. cereus, 31% in E. coli, 18% in S. Typhi, 5% in P. aeruginosa, and 3% in Staph. aureus. Pseudomonas aeruginosa showed resistance against Amoxicillin and Cefotaxime, whereas S. aureus was found resistant against Cefotaxime. Statistical analysis using one way Analysis of Variance revealed that Ofloxacin and Gentamicin had significant (p less than 0.05) differences against all isolates as compared with other antibiotics used in this study.

Microbial Copper-binding Siderophores at the Host-Pathogen Interface*

PubMed Central

Koh, Eun-Ik; Henderson, Jeffrey P.

2015-01-01

Numerous pathogenic microorganisms secrete small molecule chelators called siderophores defined by their ability to bind extracellular ferric iron, making it bioavailable to microbes. Recently, a siderophore produced by uropathogenic Escherichia coli, yersiniabactin, was found to also bind copper ions during human infections. The ability of yersiniabactin to protect E. coli from copper toxicity and redox-based phagocyte defenses distinguishes it from other E. coli siderophores. Here we compare yersiniabactin to other extracellular copper-binding molecules and review how copper-binding siderophores may confer virulence-associated gains of function during infection pathogenesis. PMID:26055720

Novel Compounds from Shark and Stingray Epidermal Mucus With Antimicrobial Activity Against Wound Infection Pathogens

DTIC Science & Technology

2013-03-01

530-E6 MSSA, MRSA, E. coli, VRE, E. faecalis, B. subtilis 530-E7 MSSA, MRSA, E. coli, VRE, E. faecalis, B. subtilis B. anthracis, Micrococcus sp...530-E10 MSSA, MRSA, E. coli, VRE, E. faecalis, B. subtilis Micrococcus sp. 530-A5 VRE 530-B12 VRE 530-C12 VRE B. anthracis 530-D12 VRE 530-E12...VRE Micrococcus sp. 530-F11 VRE Micrococcus sp. 530-F12 VRE Enterococcus. faecium Figure 9. Pathogen overlay assay plates showing

Modeling the effect of seasonal variation in ambient temperature on the transmission dynamics of a pathogen with a free-living stage: example of Escherichia coli O157:H7 in a dairy herd.

PubMed

Gautam, Raju; Bani-Yaghoub, Majid; Neill, William H; Döpfer, Dörte; Kaspar, Charles; Ivanek, Renata

2011-10-01

To explore the potential role of ambient temperature on infection transmission dynamics for pathogens, we used Escherichia coli O157:H7 in a dairy herd and the surrounding farm environment as a model system. For this system, we developed a mathematical model in which a Susceptible-Infectious-Susceptible (SIS) model of infection spread through the host population is coupled with a metapopulation model of E. coli O157:H7 free-living stage in the environment allowing bacterial growth to be influenced by ambient temperature. Model results indicate that seasonal variation in ambient temperature could have a considerable impact on pathogen populations in the environment, specifically on barn surfaces and in water troughs, and consequently on the prevalence of infection in the host population. Based on model assumptions, contaminated drinking water was the most important pathway of E. coli O157:H7 transmission to cattle. Sensitivity analysis indicated that water-borne transmission is amplified during the warmer months if the amount of standing drinking water available to the cattle herd is high. This is because warmer ambient temperature favors faster pathogen replication which when combined with slower water replacement-rate due to high amount of available standing water leads to a greater pathogen load in drinking water. These results offer a possible explanation of the seasonal variation in E. coli O157:H7 prevalence in cattle and suggest that improved drinking-water management could be used for control of this infection in cattle. Our study demonstrates how consideration of ambient temperature in transmission cycles of pathogens able to survive and grow in the environment outside the host could offer novel perspectives on the spread and control of infections caused by such pathogens. Copyright © 2011 Elsevier B.V. All rights reserved.

GIS-based analysis of the fate of waste-related pathogens Cryptosporidium parvum, Giardia lamblia and Escherichia coli in a tropical canal network.

PubMed

Diallo, Mamadou B C; Anceno, Alfredo J; Tawatsupa, Benjawan; Tripathi, Nitin K; Wangsuphachart, Voranuch; Shipin, Oleg V

2009-03-01

Urban canals play a major socio-economic role in many tropical countries and, particularly, Thailand. One of the overlooked functions that they perform is a significant attenuation of waste-related pathogens posing considerable health risk, as well as pollution attenuation in general. The study dealt with a comparison of three canals receiving: (i) municipal, (ii) mainly industrial and (iii) mainly agricultural wastewater, listed in order of progressively decreasing organic loading. The occurrence and fate of waterborne Cryptosporidium parvum, Giardia lamblia and Escherichia coli were monitored in the canals by both real-time PCR and conventionally for 12 months. The pathogens are etiological agents of an estimated 38% and 47% of diarrhea cases worldwide and in Thailand, respectively. The geographic information system (GIS) was used to evaluate and map point and, particularly, non-point pollution sources which allowed differentiating the canal sections in terms of predominant pathogen sources. The flowthrough canals, which can be viewed as waste stabilization ponds, were found to be efficiently removing the pathogens at the following generalized specific rates: 0.3 (C. parvum), 1.2 (G. lamblia), 1.8 (E. coli) log10/km.d in the dry season. The rates decreased in the rainy season for E. coli and G. lamblia, but increased for C. parvum which indicated different removal mechanisms. Data suggest that E. coli and G. lamblia were mainly removed through sedimentation and sunlight (UV) irradiation, while the likely mechanism for C. parvum was predation. Overall, the specific pathogen removal rates positively correlated with the canal organic loading rates in the rainy season. As an important result, an estimate of the municipal pollution mitigation by over 2280 km canals in the Greater Bangkok suggests that concomitant to the pathogens at least 36-95 tons of BOD5 is being removed daily, thereby saving the receiving Chao Phraya River and Bight of Bangkok, by far exceeding current, from major eutrophication problems.

Assessment of bacterial pathogens in fresh rainwater and airborne particulate matter using Real-Time PCR

NASA Astrophysics Data System (ADS)

Kaushik, Rajni; Balasubramanian, Rajasekhar

2012-01-01

Bacterial pathogens in airborne particulate matter (PM) and in rainwater (RW) were detected using a robust and sensitive Real-Time PCR method. Both RW and PM were collected simultaneously in the tropical atmosphere of Singapore, which were then subjected to analysis for the presence of selected bacterial pathogens and potential pathogen of health concern ( Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa and Aeromonas hydrophila). These pathogens were found to be prevalent in both PM and RW samples with E. coli being the most prevalent potential pathogen in both types of samples. The temporal distribution of these pathogens in PM and RW was found to be similar to each other. Using the proposed microbiological technique, the atmospheric deposition (dry and wet deposition) of bacterial pathogens to lakes and reservoirs can be studied in view of growing concerns about the outbreak of waterborne diseases.

Bacterial Prostatitis: Bacterial Virulence, Clinical Outcomes, and New Directions.

PubMed

Krieger, John N; Thumbikat, Praveen

2016-02-01

Four prostatitis syndromes are recognized clinically: acute bacterial prostatitis, chronic bacterial prostatitis, chronic prostatitis/chronic pelvic pain syndrome, and asymptomatic prostatitis. Because Escherichia coli represents the most common cause of bacterial prostatitis, we investigated the importance of bacterial virulence factors and antimicrobial resistance in E. coli strains causing prostatitis and the potential association of these characteristics with clinical outcomes. A structured literature review revealed that we have limited understanding of the virulence-associated characteristics of E. coli causing acute prostatitis. Therefore, we completed a comprehensive microbiological and molecular investigation of a unique strain collection isolated from healthy young men. We also considered new data from an animal model system suggesting certain E. coli might prove important in the etiology of chronic prostatitis/chronic pelvic pain syndrome. Our human data suggest that E. coli needs multiple pathogenicity-associated traits to overcome anatomic and immune responses in healthy young men without urological risk factors. The phylogenetic background and accumulation of an exceptional repertoire of extraintestinal pathogenic virulence-associated genes indicate that these E. coli strains belong to a highly virulent subset of uropathogenic variants. In contrast, antibiotic resistance confers little added advantage to E. coli strains in these healthy outpatients. Our animal model data also suggest that certain pathogenic E. coli may be important in the etiology of chronic prostatitis/chronic pelvic pain syndrome through mechanisms that are dependent on the host genetic background and the virulence of the bacterial strain.

Escherichia coli transfer from simulated wildlife feces to lettuce during foliar irrigation: A field study in the Northeastern United States.

PubMed

Weller, Daniel L; Kovac, Jasna; Kent, David J; Roof, Sherry; Tokman, Jeffrey I; Mudrak, Erika; Kowalcyk, Barbara; Oryang, David; Aceituno, Anna; Wiedmann, Martin

2017-12-01

Wildlife intrusion has been associated with pathogen contamination of produce. However, few studies have examined pathogen transfer from wildlife feces to pre-harvest produce. This study was performed to calculate transfer coefficients for Escherichia coli from simulated wildlife feces to field-grown lettuce during irrigation. Rabbit feces inoculated with a 3-strain cocktail of non-pathogenic E. coli were placed in a lettuce field 2.5-72 h before irrigation. Following irrigation, the E. coli concentration on the lettuce was determined. After exclusion of an outlier with high E. coli levels (Most Probable Number = 5.94*10 8 ), the average percent of E. coli in the feces that transferred to intact lettuce heads was 0.0267% (Standard Error [SE] = 0.0172). Log-linear regression showed that significantly more E. coli transferred to outer leaves compared to inner leaves (Effect = 1.3; 95% Confidence Interval = 0.4, 2.1). Additionally, the percent of E. coli that transferred from the feces to the lettuce decreased significantly with time after fecal placement, and as the distance between the lettuce and the feces, and the lettuce and the sprinklers increased. These findings provide key data that may be used in future quantitative risk assessments to identify potential intervention strategies for reducing food safety risks associated with fresh produce. Copyright © 2017 Elsevier Ltd. All rights reserved.

A Recombinant Horseshoe Crab Plasma Lectin Recognizes Specific Pathogen-Associated Molecular Patterns of Bacteria through Rhamnose

PubMed Central

Ng, Sim-Kun; Huang, Yu-Tsyr; Lee, Yuan-Chuan; Low, Ee-Ling; Chiu, Cheng-Hsun; Chen, Shiu-Ling; Mao, Liang-Chi; Chang, Margaret Dah-Tsyr

2014-01-01

Horseshoe crab is an ancient marine arthropod that, in the absence of a vertebrate-like immune system, relies solely on innate immune responses by defense molecules found in hemolymph plasma and granular hemocytes for host defense. A plasma lectin isolated from the hemolymph of Taiwanese Tachypleus tridentatus recognizes bacteria and lipopolysaccharides (LPSs), yet its structure and mechanism of action remain unclear, largely because of limited availability of horseshoe crabs and the lack of a heterogeneous expression system. In this study, we have successfully expressed and purified a soluble and functional recombinant horseshoe crab plasma lectin (rHPL) in an Escherichia coli system. Interestingly, rHPL bound not only to bacteria and LPSs like the native HPL but also to selective medically important pathogens isolated from clinical specimens, such as Gram-negative Pseudomonas aeruginosa and Klebsiella pneumoniae and Gram-positive Streptococcus pneumoniae serotypes. The binding was demonstrated to occur through a specific molecular interaction with rhamnose in pathogen-associated molecular patterns (PAMPs) on the bacterial surface. Additionally, rHPL inhibited the growth of P. aeruginosa PAO1 in a concentration-dependent manner. The results suggest that a specific protein-glycan interaction between rHPL and rhamnosyl residue may further facilitate development of novel diagnostic and therapeutic strategies for microbial pathogens. PMID:25541995

Targeting Cattle-Borne Zoonoses and Cattle Pathogens Using a Novel Trypanosomatid-Based Delivery System

PubMed Central

Mott, G. Adam; Wilson, Raymond; Fernando, Anuruddika; Robinson, Ailie; MacGregor, Paula; Kennedy, David; Schaap, Dick; Matthews, Jacqueline B.; Matthews, Keith R.

2011-01-01

Trypanosomatid parasites are notorious for the human diseases they cause throughout Africa and South America. However, non-pathogenic trypanosomatids are also found worldwide, infecting a wide range of hosts. One example is Trypanosoma (Megatrypanum) theileri, a ubiquitous protozoan commensal of bovids, which is distributed globally. Exploiting knowledge of pathogenic trypanosomatids, we have developed Trypanosoma theileri as a novel vehicle to deliver vaccine antigens and other proteins to cattle. Conditions for the growth and transfection of T. theileri have been optimised and expressed heterologous proteins targeted for secretion or specific localisation at the cell interior or surface using trafficking signals from Trypanosoma brucei. In cattle, the engineered vehicle could establish in the context of a pre-existing natural T. theileri population, was maintained long-term and generated specific immune responses to an expressed Babesia antigen at protective levels. Building on several decades of basic research into trypanosomatid pathogens, Trypanosoma theileri offers significant potential to target multiple infections, including major cattle-borne zoonoses such as Escherichia coli, Salmonella spp., Brucella abortus and Mycobacterium spp. It also has the potential to deliver therapeutics to cattle, including the lytic factor that protects humans from cattle trypanosomiasis. This could alleviate poverty by protecting indigenous African cattle from African trypanosomiasis. PMID:22046137

Strand-specific transcriptomes of Enterohemorrhagic Escherichia coli in response to interactions with ground beef microbiota: interactions between microorganisms in raw meat.

PubMed

Galia, Wessam; Leriche, Francoise; Cruveiller, Stéphane; Garnier, Cindy; Navratil, Vincent; Dubost, Audrey; Blanquet-Diot, Stéphanie; Thevenot-Sergentet, Delphine

2017-08-03

Enterohemorrhagic Escherichia coli (EHEC) are zoonotic agents associated with outbreaks worldwide. Growth of EHEC strains in ground beef could be inhibited by background microbiota that is present initially at levels greater than that of the pathogen E. coli. However, how the microbiota outcompetes the pathogenic bacteria is unknown. Our objective was to identify metabolic pathways of EHEC that were altered by natural microbiota in order to improve our understanding of the mechanisms controlling the growth and survival of EHECs in ground beef. Based on 16S metagenomics analysis, we identified the microbial community structure in our beef samples which was an essential preliminary for subtractively analyzing the gene expression of the EHEC strains. Then, we applied strand-specific RNA-seq to investigate the effects of this microbiota on the global gene expression of EHEC O26 21765 and O157 EDL933 strains by comparison with their behavior in beef meat without microbiota. In strain O26 21765 , the expression of genes connected with nitrate metabolism and nitrite detoxification, DNA repair, iron and nickel acquisition and carbohydrate metabolism, and numerous genes involved in amino acid metabolism were down-regulated. Further, the observed repression of ftsL and murF, involved respectively in building the cytokinetic ring apparatus and in synthesizing the cytoplasmic precursor of cell wall peptidoglycan, might help to explain the microbiota's inhibitory effect on EHECs. For strain O157 EDL933 , the induced expression of the genes implicated in detoxification and the general stress response and the repressed expression of the peR gene, a gene negatively associated with the virulence phenotype, might be linked to the survival and virulence of O157:H7 in ground beef with microbiota. In the present study, we show how RNA-Seq coupled with a 16S metagenomics analysis can be used to identify the effects of a complex microbial community on relevant functions of an individual microbe within it. These findings add to our understanding of the behavior of EHECs in ground beef. By measuring transcriptional responses of EHEC, we could identify putative targets which may be useful to develop new strategies to limit their shedding in ground meat thus reducing the risk of human illnesses.

Neuropathogenic Escherichia coli K1 does not exhibit proteolytic activities to exert its pathogenicity.

PubMed

Iqbal, Junaid; Rajani, Mehak; Siddiqui, Ruqaiyyah; Khan, Naveed Ahmed

2013-05-01

Proteases are well-known virulence factors that promote survival, pathogenesis and immune evasion of many pathogens. Several lines of evidence suggest that the blood-brain barrier permeability is a prerequisite in microbial invasion of the central nervous system. Because proteases are frequently associated with vascular permeability by targeting junctional proteins, here it is hypothesized that neuropathogenic Escherichia coli K1 exhibit proteolytic activities to exert its pathogenicity. Zymographic assays were performed using collagen and gelatin as substrates. The lysates of whole E. coli K1 strain E44, or E. coli K-12 strain HB101 were tested for proteolytic activities. The conditioned media were prepared by incubating bacteria in RPMI-1640 in the presence or absence of serum. The cell-free supernatants were collected and tested for proteases in zymography as mentioned above. Additionally, proteolytic degradation of host immune factors was determined by co-incubating conditioned media with albumin/immunoglobulins using protease assays. When collagen or gelatin were used as substrates in zymographic assays, neither whole bacteria nor conditioned media exhibited proteolytic activities. The conditioned media of neuropathogenic E. coli K1 strain E44, or E. coli K-12 strain HB101 did not affect degradation of albumin and immunoglobulins using protease assays. Neither zymographic assays nor protease assays detected proteolytic activities in either the whole bacteria or conditioned media of E. coli K1 strain E44 and E. coli K-12 strain HB101. These findings suggest that host cell monolayer disruptions and immune evasion strategies are likely independent of proteolytic activities of neuropathogenic E. coli K1.

Suppression of Escherichia coli O157:H7 by Dung Beetles (Coleoptera: Scarabaeidae) Using the Lowbush Blueberry Agroecosystem as a Model System

PubMed Central

Jones, Matthew S.; Tadepalli, Shravani; Bridges, David F.; Wu, Vivian C. H.; Drummond, Frank

2015-01-01

Wildlife as a source of microbial contamination is a food safety concern. Deer feces (scat) have been determined as a point source for Escherichia coli O157:H7 contamination of fresh produce. The ecological role of the scooped scarab (Onthophagus hecate (Panzer)), a generalist dung beetle species common in Maine blueberry fields, was explored as a biological control agent and alternatively as a pathogen vector between deer scat and food. A large-scale field survey of wildlife scat indicated that pathogenic E. coli O157:H7 was present, albeit at a low prevalence (1.9% of samples, n = 318), in the Maine lowbush blueberry agroecosystem. A manipulative field experiment verified that, should contact occur between deer scat and blueberry plants and fruit during the summer, contamination with E. coli O157:H7 can occur and persist for more than 72 h. For both the positive control and an experimental scat inoculation treatment, the levels of the bacterial population decreased over time, but at different rates (treatment x time interaction: F (1.9,18.8) = 358.486, P < 0.0001). The positive control inoculation, which resulted in a higher initial E. coli level on fruit, decayed at a faster rate than inoculation of fruit via scat in the experimental treatment. We conducted 2 laboratory studies to elucidate aspects of dung beetle feeding ecology as it relates to suppression of E. coli O157:H7 from deer scat to lowbush blueberry fruit. In both experiments, dung beetles buried the same amount of scat whether or not the scat was inoculated with the pathogen (F (1,6) = 0.001; P = 0.999 and (F (2,17) = 4.10, P = 0.147). Beetles feeding on E. coli inoculated deer scat were not found to vector the pathogen to fruit. In two studies, beetles lowered the amount of pathogenic E. coli persisting in soils compared to soils without beetles (F (2,9) = 7.757; P = 0.05 and F (2,17) = 8.0621, P = 0.004). Our study suggests that the dung beetle species, Onthophagus hecate, has the potential to contribute to the suppression of E. coli O157:H7 in agricultural landscapes. PMID:25849430

Suppression of Escherichia coli O157:H7 by Dung Beetles (Coleoptera: Scarabaeidae) using the lowbush blueberry agroecosystem as a model system.

PubMed

Jones, Matthew S; Tadepalli, Shravani; Bridges, David F; Wu, Vivian C H; Drummond, Frank

2015-01-01

Wildlife as a source of microbial contamination is a food safety concern. Deer feces (scat) have been determined as a point source for Escherichia coli O157:H7 contamination of fresh produce. The ecological role of the scooped scarab (Onthophagus hecate (Panzer)), a generalist dung beetle species common in Maine blueberry fields, was explored as a biological control agent and alternatively as a pathogen vector between deer scat and food. A large-scale field survey of wildlife scat indicated that pathogenic E. coli O157:H7 was present, albeit at a low prevalence (1.9% of samples, n = 318), in the Maine lowbush blueberry agroecosystem. A manipulative field experiment verified that, should contact occur between deer scat and blueberry plants and fruit during the summer, contamination with E. coli O157:H7 can occur and persist for more than 72 h. For both the positive control and an experimental scat inoculation treatment, the levels of the bacterial population decreased over time, but at different rates (treatment x time interaction: F (1.9,18.8) = 358.486, P < 0.0001). The positive control inoculation, which resulted in a higher initial E. coli level on fruit, decayed at a faster rate than inoculation of fruit via scat in the experimental treatment. We conducted 2 laboratory studies to elucidate aspects of dung beetle feeding ecology as it relates to suppression of E. coli O157:H7 from deer scat to lowbush blueberry fruit. In both experiments, dung beetles buried the same amount of scat whether or not the scat was inoculated with the pathogen (F(1,6) = 0.001; P = 0.999 and (F (2,17) = 4.10, P = 0.147). Beetles feeding on E. coli inoculated deer scat were not found to vector the pathogen to fruit. In two studies, beetles lowered the amount of pathogenic E. coli persisting in soils compared to soils without beetles (F (2,9) = 7.757; P = 0.05 and F (2,17) = 8.0621, P = 0.004). Our study suggests that the dung beetle species, Onthophagus hecate, has the potential to contribute to the suppression of E. coli O157:H7 in agricultural landscapes.

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

Linking microbial community structure to function in representative simulated systems.

PubMed

Marcus, Ian M; Wilder, Hailey A; Quazi, Shanin J; Walker, Sharon L

2013-04-01

Pathogenic bacteria are generally studied as a single strain under ideal growing conditions, although these conditions are not the norm in the environments in which pathogens typically proliferate. In this investigation, a representative microbial community along with Escherichia coli O157:H7, a model pathogen, was studied in three environments in which such a pathogen could be found: a human colon, a septic tank, and groundwater. Each of these systems was built in the lab in order to retain the physical/chemical and microbial complexity of the environments while maintaining control of the feed into the models. The microbial community in the colon was found to have a high percentage of bacteriodetes and firmicutes, while the septic tank and groundwater systems were composed mostly of proteobacteria. The introduction of E. coli O157:H7 into the simulated systems elicited a shift in the structures and phenotypic cell characteristics of the microbial communities. The fate and transport of the microbial community with E. coli O157:H7 were found to be significantly different from those of E. coli O157:H7 studied as a single isolate, suggesting that the behavior of the organism in the environment was different from that previously conceived. The findings in this study clearly suggest that to gain insight into the fate of pathogens, cells should be grown and analyzed under conditions simulating those of the environment in which the pathogens are present.

Linking Microbial Community Structure to Function in Representative Simulated Systems

PubMed Central

Marcus, Ian M.; Wilder, Hailey A.; Quazi, Shanin J.

2013-01-01

Pathogenic bacteria are generally studied as a single strain under ideal growing conditions, although these conditions are not the norm in the environments in which pathogens typically proliferate. In this investigation, a representative microbial community along with Escherichia coli O157:H7, a model pathogen, was studied in three environments in which such a pathogen could be found: a human colon, a septic tank, and groundwater. Each of these systems was built in the lab in order to retain the physical/chemical and microbial complexity of the environments while maintaining control of the feed into the models. The microbial community in the colon was found to have a high percentage of bacteriodetes and firmicutes, while the septic tank and groundwater systems were composed mostly of proteobacteria. The introduction of E. coli O157:H7 into the simulated systems elicited a shift in the structures and phenotypic cell characteristics of the microbial communities. The fate and transport of the microbial community with E. coli O157:H7 were found to be significantly different from those of E. coli O157:H7 studied as a single isolate, suggesting that the behavior of the organism in the environment was different from that previously conceived. The findings in this study clearly suggest that to gain insight into the fate of pathogens, cells should be grown and analyzed under conditions simulating those of the environment in which the pathogens are present. PMID:23396331

Biofilm formation by enteric pathogens and its role in plant colonization and persistence

PubMed Central

Yaron, Sima; Römling, Ute

2014-01-01

The significant increase in foodborne outbreaks caused by contaminated fresh produce, such as alfalfa sprouts, lettuce, melons, tomatoes and spinach, during the last 30 years stimulated investigation of the mechanisms of persistence of human pathogens on plants. Emerging evidence suggests that Salmonella enterica and Escherichia coli, which cause the vast majority of fresh produce outbreaks, are able to adhere to and to form biofilms on plants leading to persistence and resistance to disinfection treatments, which subsequently can cause human infections and major outbreaks. In this review, we present the current knowledge about host, bacterial and environmental factors that affect the attachment to plant tissue and the process of biofilm formation by S. enterica and E. coli, and discuss how biofilm formation assists in persistence of pathogens on the plants. Mechanisms used by S. enterica and E. coli to adhere and persist on abiotic surfaces and mammalian cells are partially similar and also used by plant pathogens and symbionts. For example, amyloid curli fimbriae, part of the extracellular matrix of biofilms, frequently contribute to adherence and are upregulated upon adherence and colonization of plant material. Also the major exopolysaccharide of the biofilm matrix, cellulose, is an adherence factor not only of S. enterica and E. coli, but also of plant symbionts and pathogens. Plants, on the other hand, respond to colonization by enteric pathogens with a variety of defence mechanisms, some of which can effectively inhibit biofilm formation. Consequently, plant compounds might be investigated for promising novel antibiofilm strategies. PMID:25351039

PATHOGENIC ESCHERICHIA COLI

EPA Science Inventory

Escherichia coli is a bacterial species which inhabits the gastrointestinal tract of man and warm-blooded animals. Because of the ubiquity of this bacterium in the intestinal flora, it serves as an important indicator organism of fecal contamination. E. coli, aside from serving a...

Inactivation of Shiga toxin-producing Escherichia coli O104:H4 using cold atmospheric pressure plasma.

PubMed

Baier, Matthias; Janssen, Traute; Wieler, Lothar H; Ehlbeck, Jörg; Knorr, Dietrich; Schlüter, Oliver

2015-09-01

From cultivation to the end of the post-harvest chain, heat-sensitive fresh produce is exposed to a variety of sources of pathogenic microorganisms. If contaminated, effective gentle means of sanitation are necessary to reduce bacterial pathogen load below their infective dose. The occurrence of rare or new serotypes raises the question of their tenacity to inactivation processes. In this study the antibacterial efficiency of cold plasma by an atmospheric pressure plasma-jet was examined against the Shiga toxin-producing outbreak strain Escherichia coli O104:H4. Argon was transformed into non-thermal plasma at a power input of 8 W and a gas flow of 5 L min(-1). Basic tests were performed on polysaccharide gel discs, including the more common E. coli O157:H7 and non-pathogenic E. coli DSM 1116. At 5 mm treatment distance and 10(5) cfu cm(-2) initial bacterial count, plasma reduced E. coli O104:H4 after 60 s by 4.6 ± 0.6 log, E. coli O157:H7 after 45 s by 4.5 ± 0.6 log, and E. coli DSM 1116 after 30 s by 4.4 ± 1.1 log. On the surface of corn salad leaves, gentle plasma application at 17 mm reduced 10(4) cfu cm(-2) of E. coli O104:H4 by 3.3 ± 1.1 log after 2 min, whereas E. coli O157:H7 was inactivated by 3.2 ± 1.1 log after 60 s. In conclusion, plasma treatment has the potential to reduce pathogens such as E. coli O104:H4 on the surface of fresh produce. However, a serotype-specific adaptation of the process parameters is required. Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Sequencing a piece of history: complete genome sequence of the original Escherichia coli strain

PubMed Central

Dunne, Karl A; Chaudhuri, Roy R; Rossiter, Amanda E; Beriotto, Irene; Browning, Douglas F; Squire, Derrick; Cunningham, Adam F; Cole, Jeffrey A; Loman, Nicholas

2017-01-01

In 1885, Theodor Escherich first described the Bacillus coli commune, which was subsequently renamed Escherichia coli. We report the complete genome sequence of this original strain (NCTC 86). The 5 144 392 bp circular chromosome encodes the genes for 4805 proteins, which include antigens, virulence factors, antimicrobial-resistance factors and secretion systems, of a commensal organism from the pre-antibiotic era. It is located in the E. coli A subgroup and is closely related to E. coli K-12 MG1655. E. coli strain NCTC 86 and the non-pathogenic K-12, C, B and HS strains share a common backbone that is largely co-linear. The exception is a large 2 803 932 bp inversion that spans the replication terminus from gmhB to clpB. Comparison with E. coli K-12 reveals 41 regions of difference (577 351 bp) distributed across the chromosome. For example, and contrary to current dogma, E. coli NCTC 86 includes a nine gene sil locus that encodes a silver-resistance efflux pump acquired before the current widespread use of silver nanoparticles as an antibacterial agent, possibly resulting from the widespread use of silver utensils and currency in Germany in the 1800s. In summary, phylogenetic comparisons with other E. coli strains confirmed that the original strain isolated by Escherich is most closely related to the non-pathogenic commensal strains. It is more distant from the root than the pathogenic organisms E. coli 042 and O157 : H7; therefore, it is not an ancestral state for the species. PMID:28663823

Cloning and sequencing of Staphylococcus aureus murC, a gene essential for cell wall biosynthesis.

PubMed

Lowe, A M; Deresiewicz, R L

1999-01-01

Staphylococcus aureus is a major human pathogen that is increasingly resistant to clinically useful antimicrobial agents. While screening for S. aureus genes expressed during mammalian infection, we isolated murC. This gene encodes UDP-N-acetylmuramoyl-L-alanine synthetase, an enzyme essential for cell wall biosynthesis in a number of bacteria. S. aureus MurC has a predicted mass 49,182 Da and complements the temperature-sensitive murC mutation of E. coli ST222. Sequence data on the DNA flanking staphylococcal murC suggests that the local gene organization there parallels that found in B. subtilis, but differs from that found in gram-negative bacterial pathogens. MurC proteins represent promising targets for broad spectrum antimicrobial drug development.

An improved method to simultaneously detect Salmonella enteritidis, Escherichia coli O157 and Listeria monocytogenes in ground black pepper using multiplex real-time PCR

USDA-ARS?s Scientific Manuscript database

Introduction: The three common foodborne pathogens implicated in foodborne outbreaks are Salmonella spp., Escherichia coli O157:H7 and Listeria monocytogenes. Hence, it is important to identify these pathogens in contaminated foods so that they can be eliminated from the marketplace. At present, the...

Escherichia coli O157:H7 biofilm formation and internalization on lettuce and spinach leaf surfaces reduces efficacy of irradiation and sodium hypochlorite washes

USDA-ARS?s Scientific Manuscript database

Escherichia coli O157:H7 contamination of leafy green vegetables is an ongoing concern for consumers. Biofilm-associated and internalized pathogens are relatively resistant to chemical treatments, but little is known about the response of these protected pathogens to irradiation. Leaves of Romaine l...

Evaluation of commercial ß-agonists, dietary protein, and shade on fecal shedding of Escherichia coli O157:H7 from feedlot cattle

USDA-ARS?s Scientific Manuscript database

Escherichia coli O157:H7 is a foodborne pathogen commonly associated with cattle feces. Diet, including dietary supplements such as b-agonists, may impact fecal shedding of this pathogen. A series of three experiments were conducted to determine if the b- agonists ractopamine hydrochloride (RAC) or ...

Role of major surface structures of Escherichia coli O157:H7 in initial attachment to biotic and abiotic surfaces

USDA-ARS?s Scientific Manuscript database

Infection by human pathogens through fresh, minimally processed produce and solid plant-derived foods is a major concern of U.S. and global food industry and public health services. The enterohemorrhagic Escherichia coli O157:H7 is a frequent and potent food borne pathogen that causes severe disease...

Biofilm formation and sanitizer resistance contributes to “High Event” meat contamination by E. coli O157:H7

USDA-ARS?s Scientific Manuscript database

Background E. coli O157:H7 is an important foodborne pathogen responsible for numerous outbreaks worldwide. FSIS regulates this pathogen as an adulterant in meat products, and a “High Event Period” is defined as a time period in which commercial meat processing plants experience a higher than usual ...

Inactivation of Salmonella, Escherichia coli O157:H7 and non-O157 STEC by hypochlorite solutions with high organic loads

USDA-ARS?s Scientific Manuscript database

Introduction: Salmonella, E. coli O157:H7 and Non-O157 STEC have been recognized as foodborne pathogen concerns for fresh produce. Although chlorinated water (CW) is widely used in fresh produce processing to reduce pathogens and prevent cross-contamination, limited information is available on effic...

Future perspectives, applications and challenges of genomic epidemiology studies for food-borne pathogens: A case study of Enterohemorrhagic Escherichia coli (EHEC) of the O157:H7 serotype

PubMed Central

Eppinger, Mark; Cebula, Thomas A

2015-01-01

The shiga-toxin (Stx)-producing human pathogen Escherichia coli serotype O157:H7 is a highly pathogenic subgroup of Stx-producing E. coli (STEC) with food-borne etiology and bovine reservoir. Each year in the U. S., approximately 100,000 patients are infected with enterohemorrhagic E. coli (EHEC) of the O157:H7 serotype. This food-borne pathogen is a global public health threat responsible for widespread outbreaks of human disease. Since its initial discovery in 1982, O157:H7 has rapidly become the dominant EHEC serotype in North America. Hospitalization rates among patients as high as 50% have been reported for severe outbreaks of human disease. Symptoms of disease can rapidly deteriorate and progress to life-threatening complications such as Hemolytic Uremic Syndrome (HUS), the leading cause of kidney failure in children, or Hemorrhagic Colitis. In depth understanding of the genomic diversity that exists among currently circulating EHEC populations has broad applications for improved molecular-guided biosurveillance, outbreak preparedness, diagnostic risk assessment, and development of alternative toxin-suppressing therapeutics. PMID:25483335

[Pathogenic activity modulation of Escherichia coli TL+ toxin with an isolated protein of Giardia intestinalis and a synthetic peptide].

PubMed

Jiménez-Cardoso, E; Eligio-García, L; Jiménez-Cardoso, J M; Angeles-Anguiano, E; Tobilla-Mercado, J M; Castañeda, G

2001-01-01

It is know that a protein from Giardia intestinalis works as a substrate for V. cholerae and Escherichia coli. The toxic activity of both activates protein G form intestinal mucosa with a pathogenic activity results. In the present study, the pathogenic activity of subunit A of Vibrio cholerae toxin (ADP-ribosyltranferase) using isolated fragments from: Giardia intestinalis and a synthetic peptide were used as modulators in vivo. Adult Neo Zealand males rabbits with ileal loop were prepared and different mixtures of heat labile enterotoxin obtained from Escherichia coli H10407 and ARF protein isolated by electrofocusing from Giardia intestinalis Portland I were inoculated in the loops. The toxin activity was evaluated by luminal liquid secretion and cyclic AMP concentration in tissues (each loop). ADP ribosyltranferase activity was modulated, due to a decreased of luminal secretion and cAMP in tissues. Such results were seen when synthetic peptide and subunit A from Vibrio cholerae were used. The ADP ribosyltranferase activity of heat labile Escherichia coli and Vibrio cholerae toxins were modified by in vitro and in vivo interaction with ARF protein, which modified pathogenic effect over rabbits intestinal epithelium.

Fecal-indicator bacteria and Escherichia coli pathogen data collected near a novel sub-irrigation water-treatment system in Lenawee County, Michigan, June-November 2007

USGS Publications Warehouse

Duris, Joseph W.; Beeler, Stephanie

2008-01-01

The U.S. Geological Survey, in cooperation with the Lenawee County Conservation District in Lenawee County, Mich., conducted a sampling effort over a single growing season (June to November 2007) to evaluate the microbiological water quality around a novel livestock reservoir wetland sub-irrigation system. Samples were collected and analyzed for fecal coliform bacteria, Escherichia coli (E. coli) bacteria, and six genes from pathogenic strains of E. coli.A total of 73 water-quality samples were collected on nine occasions from June to November 2007. These samples were collected within the surface water, shallow ground water, and the manure-treatment system near Bakerlads Farm near Clayton in Lenawee County, Mich. Fecal coliform bacteria concentrations ranged from 10 to 1.26 million colony forming units per 100 milliliters (CFU/100 mL). E. coli bacteria concentrations ranged from 8 to 540,000 CFU/100 mL. Data from the E. coli pathogen analysis showed that 73 percent of samples contained the eaeA gene, 1 percent of samples contained the stx2 gene, 37 percent of samples contained the stx1 gene, 21 percent of samples contained the rfbO157 gene, and 64 percent of samples contained the LTIIa gene.

SURVIVAL OF ESCHERICHIA COLI 0157:H7 IN DAIRY CATTLE FEED WATER

EPA Science Inventory

Cattle feed waters from two dairy farms were used in a study to determine the survival characteristics of the bacterial pathogen Escherichia coli )157:H7 and wild-type E. coli. The E. coli 0157:H7 inoculum consisted of a consortium of isolates obtained from dairy cattle. Fresh ma...

The β-Defensin Gallinacin-6 Is Expressed in the Chicken Digestive Tract and Has Antimicrobial Activity against Food-Borne Pathogens▿

PubMed Central

van Dijk, Albert; Veldhuizen, Edwin J. A.; Kalkhove, Stefanie I. C.; Tjeerdsma-van Bokhoven, Johanna L. M.; Romijn, Roland A.; Haagsman, Henk P.

2007-01-01

Food-borne pathogens are responsible for most cases of food poisoning in developed countries and are often associated with poultry products, including chicken. Little is known about the role of β-defensins in the chicken digestive tract and their efficacy. In this study, the expression of chicken β-defensin gallinacin-6 (Gal-6) and its antimicrobial activity against food-borne pathogens were investigated. Reverse transcription-PCR analysis showed high expression of Gal-6 mRNA in the esophagus and crop, moderate expression in the glandular stomach, and low expression throughout the intestinal tract. Putative transcription factor binding sites for nuclear factor kappa beta, activator protein 1, and nuclear factor interleukin-6 were found in the Gal-6 gene upstream region, which suggests a possible inducible nature of the Gal-6 gene. In colony-counting assays, strong bactericidal and fungicidal activity was observed, including bactericidal activity against food-borne pathogens Campylobacter jejuni, Salmonella enterica serovar Typhimurium, Clostridium perfringens, and Escherichia coli. Treatment with 16 μg/ml synthetic Gal-6 resulted in a 3 log unit reduction in Clostridium perfringens survival within 60 min, indicating fast killing kinetics. Transmission electron microscopy examination of synthetic-Gal-6-treated Clostridium perfringens cells showed dose-dependent changes in morphology after 30 min, including intracellular granulation, cytoplasm retraction, irregular septum formation in dividing cells, and cell lysis. The high expression in the proximal digestive tract and broad antimicrobial activity suggest that chicken β-defensin gallinacin-6 plays an important role in chicken innate host defense. PMID:17194828

Slugs: potential novel vectors of Escherichia coli O157.

PubMed

Sproston, Emma L; Macrae, M; Ogden, Iain D; Wilson, Michael J; Strachan, Norval J C

2006-01-01

Field and laboratory studies were performed to determine whether slugs could act as novel vectors for pathogen (e.g., Escherichia coli O157) transfer from animal feces to salad vegetables. Escherichia coli O157 was isolated from 0.21% of field slugs from an Aberdeenshire sheep farm. These isolates carried the verocytotoxin genes (vt1 and vt2) and the attaching and effacing gene (eae), suggesting that they are potentially pathogenic to humans. Strain typing using multilocus variable number tandem repeats analysis showed that slug and sheep isolates were indistinguishable. Laboratory experiments using an E. coli mutant resistant to nalidixic acid showed that the ubiquitous slug species Deroceras reticulatum could carry viable E. coli on its external surface for up to 14 days. Slugs that had been fed E. coli shed viable bacteria in their feces with numbers showing a short but statistically significant linear log decline. Further, it was found that E. coli persisted for up to 3 weeks in excreted slug feces, and hence, we conclude that slugs have the potential to act as novel vectors of E. coli O157.

Slugs: Potential Novel Vectors of Escherichia coli O157

PubMed Central

Sproston, Emma L.; Macrae, M.; Ogden, Iain D.; Wilson, Michael J.; Strachan, Norval J. C.

2006-01-01

Field and laboratory studies were performed to determine whether slugs could act as novel vectors for pathogen (e.g., Escherichia coli O157) transfer from animal feces to salad vegetables. Escherichia coli O157 was isolated from 0.21% of field slugs from an Aberdeenshire sheep farm. These isolates carried the verocytotoxin genes (vt1 and vt2) and the attaching and effacing gene (eae), suggesting that they are potentially pathogenic to humans. Strain typing using multilocus variable number tandem repeats analysis showed that slug and sheep isolates were indistinguishable. Laboratory experiments using an E. coli mutant resistant to nalidixic acid showed that the ubiquitous slug species Deroceras reticulatum could carry viable E. coli on its external surface for up to 14 days. Slugs that had been fed E. coli shed viable bacteria in their feces with numbers showing a short but statistically significant linear log decline. Further, it was found that E. coli persisted for up to 3 weeks in excreted slug feces, and hence, we conclude that slugs have the potential to act as novel vectors of E. coli O157. PMID:16391036

Differential inhibition of adenylylated and deadenylylated forms of M. tuberculosis glutamine synthetase as a drug discovery platform

PubMed Central

Theron, A.; Roth, R. L.; Hoppe, H.; Parkinson, C.; van der Westhuyzen, C. W.; Stoychev, S.; Wiid, I.; Pietersen, R. D.; Baker, B.

2017-01-01

Glutamine synthetase is a ubiquitous central enzyme in nitrogen metabolism that is controlled by up to four regulatory mechanisms, including adenylylation of some or all of the twelve subunits by adenylyl transferase. It is considered a potential therapeutic target for the treatment of tuberculosis, being essential for the growth of Mycobacterium tuberculosis, and is found extracellularly only in the pathogenic Mycobacterium strains. Human glutamine synthetase is not regulated by the adenylylation mechanism, so the adenylylated form of bacterial glutamine synthetase is of particular interest. Previously published reports show that, when M. tuberculosis glutamine synthetase is expressed in Escherichia coli, the E. coli adenylyl transferase does not optimally adenylylate the M. tuberculosis glutamine synthetase. Here, we demonstrate the production of soluble adenylylated M. tuberulosis glutamine synthetase in E. coli by the co-expression of M. tuberculosis glutamine synthetase and M. tuberculosis adenylyl transferase. The differential inhibition of adenylylated M. tuberulosis glutamine synthetase and deadenylylated M. tuberulosis glutamine synthetase by ATP based scaffold inhibitors are reported. Compounds selected on the basis of their enzyme inhibition were also shown to inhibit M. tuberculosis in the BACTEC 460TB™ assay as well as the intracellular inhibition of M. tuberculosis in a mouse bone-marrow derived macrophage assay. PMID:28972974

Organization and control of genes encoding catabolic enzymes in Rhizobiaceae. Progress report, March 1993

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

Parke, D.; Ornston, L.N.

1993-03-01

Rhizobiaceae, a diverse bacterial group comprising rhizobia and agrobacteria, symbiotic partnership with plants form nitrogen-fixing nodules on plant roots or are plant pathogens. Phenolic compounds produced by plants serve as inducers of rhizobial nodulation genes and agrobacterial virulence genes reflect their capacity to utilize numerous aromatics, including phenolics, as a source of carbon and energy. In many microbes the aerobic degradation of numerous aromatic compounds to tricarboxylic acid cycle intermediates is achieved by the {beta}-ketoadipate pathway. Our initial studies focused on the organization and regulation of the ketoadipate pathway in Agrobacterium tumefaciens. We have cloned, identified and characterized a novelmore » regulatory gene that modulates expression of an adjacent pca (protocatechuate) structural gene, pcaD. Regulation of pcaD is mediated by the regulatory gene, termed pcaQ, in concert with the intermediate {beta}-carboxy-cis,cis-muconate. {beta}-carboxy-cis,cismuconate is an unstable chemical, not marketed commercially, and it is unlikely to permeate Escherichia coli cells if supplied in media. Because of these factors, characterization of pcaQ in E. coli required an in vivo delivery system for {beta}-carboxycis,cis-muconate. This was accomplished by designing an E. coli strain that expressed an Acinetobacter calcoaceticus pcaA gene for conversion of protocatechuate to {beta}-carboxy-cis,cis-muconate.« less

Organization and control of genes encoding catabolic enzymes in Rhizobiaceae

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

Parke, D.; Ornston, L.N.

1993-03-01

Rhizobiaceae, a diverse bacterial group comprising rhizobia and agrobacteria, symbiotic partnership with plants form nitrogen-fixing nodules on plant roots or are plant pathogens. Phenolic compounds produced by plants serve as inducers of rhizobial nodulation genes and agrobacterial virulence genes reflect their capacity to utilize numerous aromatics, including phenolics, as a source of carbon and energy. In many microbes the aerobic degradation of numerous aromatic compounds to tricarboxylic acid cycle intermediates is achieved by the [beta]-ketoadipate pathway. Our initial studies focused on the organization and regulation of the ketoadipate pathway in Agrobacterium tumefaciens. We have cloned, identified and characterized a novelmore » regulatory gene that modulates expression of an adjacent pca (protocatechuate) structural gene, pcaD. Regulation of pcaD is mediated by the regulatory gene, termed pcaQ, in concert with the intermediate [beta]-carboxy-cis,cis-muconate. [beta]-carboxy-cis,cismuconate is an unstable chemical, not marketed commercially, and it is unlikely to permeate Escherichia coli cells if supplied in media. Because of these factors, characterization of pcaQ in E. coli required an in vivo delivery system for [beta]-carboxycis,cis-muconate. This was accomplished by designing an E. coli strain that expressed an Acinetobacter calcoaceticus pcaA gene for conversion of protocatechuate to [beta]-carboxy-cis,cis-muconate.« less

Growth of Escherichia coli O157:H7, Non-O157 Shiga Toxin-Producing Escherichia coli , and Salmonella in Water and Hydroponic Fertilizer Solutions.

PubMed

Shaw, Angela; Helterbran, Kara; Evans, Michael R; Currey, Christopher

2016-12-01

The desire for local, fresh produce year round is driving the growth of hydroponic growing systems in the United States. Many food crops, such as leafy greens and culinary herbs, grown within hydroponics systems have their root systems submerged in recirculating nutrient-dense fertilizer solutions from planting through harvest. If a foodborne pathogen were introduced into this water system, the risk of contamination to the entire crop would be high. Hence, this study was designed to determine whether Escherichia coli O157:H7, non-O157 Shiga toxin-producing E. coli , and Salmonella were able to survive and reproduce in two common hydroponic fertilizer solutions and in water or whether the bacteria would be killed or suppressed by the fertilizer solutions. All the pathogens grew by 1 to 6 log CFU/ml over a 24-h period, depending on the solution. E. coli O157:H7 reached higher levels in the fertilizer solution with plants (3.12 log CFU/ml), whereas non-O157 Shiga toxin-producing E. coli and Salmonella reached higher levels in the fertilizer solution without plants (1.36 to 3.77 log CFU/ml). The foodborne pathogens evaluated here survived for 24 h in the fertilizer solution, and populations grew more rapidly in these solutions than in plain water. Therefore, human pathogens entering the fertilizer solution tanks in hydroponic systems would be expected to rapidly propagate and spread throughout the system and potentially contaminate the entire crop.

Genome Dynamics of Escherichia coli during Antibiotic Treatment: Transfer, Loss, and Persistence of Genetic Elements In situ of the Infant Gut.

PubMed

Porse, Andreas; Gumpert, Heidi; Kubicek-Sutherland, Jessica Z; Karami, Nahid; Adlerberth, Ingegerd; Wold, Agnes E; Andersson, Dan I; Sommer, Morten O A

2017-01-01

Elucidating the adaptive strategies and plasticity of bacterial genomes in situ is crucial for understanding the epidemiology and evolution of pathogens threatening human health. While much is known about the evolution of Escherichia coli in controlled laboratory environments, less effort has been made to elucidate the genome dynamics of E. coli in its native settings. Here, we follow the genome dynamics of co-existing E. coli lineages in situ of the infant gut during the first year of life. One E. coli lineage causes a urinary tract infection (UTI) and experiences several alterations of its genomic content during subsequent antibiotic treatment. Interestingly, all isolates of this uropathogenic E. coli strain carried a highly stable plasmid implicated in virulence of diverse pathogenic strains from all over the world. While virulence elements are certainly beneficial during infection scenarios, their role in gut colonization and pathogen persistence is poorly understood. We performed in vivo competitive fitness experiments to assess the role of this highly disseminated virulence plasmid in gut colonization, but found no evidence for a direct benefit of plasmid carriage. Through plasmid stability assays, we demonstrate that this plasmid is maintained in a parasitic manner, by strong first-line inheritance mechanisms, acting on the single-cell level, rather than providing a direct survival advantage in the gut. Investigating the ecology of endemic accessory genetic elements, in their pathogenic hosts and native environment, is of vital importance if we want to understand the evolution and persistence of highly virulent and drug resistant bacterial isolates.

The genome sequence of avian pathogenic Escherichia coli strain O1:K1:H7 shares strong similarities with human extraintestinal pathogenic E. coli genomes.

PubMed

Johnson, Timothy J; Kariyawasam, Subhashinie; Wannemuehler, Yvonne; Mangiamele, Paul; Johnson, Sara J; Doetkott, Curt; Skyberg, Jerod A; Lynne, Aaron M; Johnson, James R; Nolan, Lisa K

2007-04-01

Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include human uropathogenic E. coli (UPEC) and avian pathogenic E. coli (APEC). Regardless of host of origin, ExPEC strains share many traits. It has been suggested that these commonalities may enable APEC to cause disease in humans. Here, we begin to test the hypothesis that certain APEC strains possess potential to cause human urinary tract infection through virulence genotyping of 1,000 APEC and UPEC strains, generation of the first complete genomic sequence of an APEC (APEC O1:K1:H7) strain, and comparison of this genome to all available human ExPEC genomic sequences. The genomes of APEC O1 and three human UPEC strains were found to be remarkably similar, with only 4.5% of APEC O1's genome not found in other sequenced ExPEC genomes. Also, use of multilocus sequence typing showed that some of the sequenced human ExPEC strains were more like APEC O1 than other human ExPEC strains. This work provides evidence that at least some human and avian ExPEC strains are highly similar to one another, and it supports the possibility that a food-borne link between some APEC and UPEC strains exists. Future studies are necessary to assess the ability of APEC to overcome the hurdles necessary for such a food-borne transmission, and epidemiological studies are required to confirm that such a phenomenon actually occurs.

Influence of Detection Methods in Characterizing Escherichia coli O157:H7 in Raw Goat Meat Using Conventional and Molecular Methods.

PubMed

Tabashsum, Zajeba; Nazneen, Mafruha; Ahsan, C R; Bari, M L; Yasmin, M

2016-01-01

　Presence of Escherichia coli O157:H7 on fresh goat meat samples (n= 40) of Dhaka city was analyzed using conventional and molecular methods. A total of 86 presumptive E. coli O157:H7 colonies were isolated from 60% of the samples using selective agar plating method. After conventional biochemical assay followed by API 20E assay, only 11 isolates were found to be E. coli O157:H7. Further serological test identified only four isolates that has strong agglutination reaction against anti-H7 sensitized latex. The biochemically and serologically confirmed isolates were then screened for major virulence factors include eaeA, rfbE, fliC, stx1 and stx2 genes by PCR. PCR analysis of positive isolates showed, 10 isolates were eaeA and rfbE genes positive but fliC gene was only in six, indicating that these isolates were H7 positive with flagellum antigens which might not expressed or detected in serotyping tests. Multiplex PCR against eaeA, stx1 and stx2 genes of the isolates showed similar results as when done individually. These results revealed that only 7% of the primary presumptive E. coli O157:H7 was found to be stx producing E. coli O157:H7 and thus greatly influenced the detection of the pathogen in meat samples.

Systems metabolic engineering of Escherichia coli for production of the antitumor drugs violacein and deoxyviolacein.

PubMed

Rodrigues, André L; Trachtmann, Nathalie; Becker, Judith; Lohanatha, Ananta F; Blotenberg, Jana; Bolten, Christoph J; Korneli, Claudia; de Souza Lima, André O; Porto, Luismar M; Sprenger, Georg A; Wittmann, Christoph

2013-11-01

Violacein and deoxyviolacein are interesting therapeutics against pathogenic bacteria and viruses as well as tumor cells. In the present work, systems-wide metabolic engineering was applied to target Escherichia coli, a widely accepted recombinant host in pharmaceutical biotechnology, for production of these high-value products. The basic producer, E. coli dVio-1, that expressed the vioABCE cluster from Chromobacterium violaceum under control of the inducible araC system, accumulated 180 mg L(-1) of deoxyviolacein. Targeted intracellular metabolite analysis then identified bottlenecks in tryptophan supporting pathways, the major product building block. This was used for comprehensive engineering of serine, chorismate and tryptophan biosynthesis and the non-oxidative pentose-phosphate pathway. The final strain, E. coli dVio-6, accumulated 320 mg L(-1) deoxyviolacein in shake flask cultures. The created chassis of a high-flux tryptophan pathway was complemented by genomic integration of the vioD gene of Janthinobacterium lividum, which enabled exclusive production of violacein. In a fed-batch process, the resulting producer E. coli Vio-4 accumulated 710 mg L(-1) of the desired product. With straightforward broth extraction and subsequent crystallization, violacein could be obtained with 99.8% purity. This demonstrates the potential of E. coli as a platform for production of tryptophan based therapeutics. Copyright © 2013 Elsevier Inc. All rights reserved.

Pathogen inactivation in liquid dairy manure during anaerobic and aerobic digestions

NASA Astrophysics Data System (ADS)

Biswas, S.; Pandey, P.; Castillo, A. R.; Vaddella, V. K.

2014-12-01

Controlling manure-borne pathogens such as E. coli O157:H7, Salmonella spp. and Listeria monocytogenes are crucial for protecting surface and ground water as well as mitigating risks to human health. In California dairy farms, flushing of dairy manure (mainly animal feces and urine) from freestall barns and subsequent liquid-solid manure separation is a common practice for handling animal waste. The liquid manure fraction is generally pumped into the settling ponds and it goes into aerobic and/or anaerobic lagoons for extended period of time. Considering the importance of controlling pathogens in animal waste, the objective of the study was to understand the effects of anaerobic and aerobic digestions on the survival of three human pathogens in animal waste. The pathogen inactivation was assessed at four temperatures (30, 35, 42, and 50 °C), and the relationships between temperature and pathogen decay were estimated. Results showed a steady decrease of E. coli levels in aerobic and anaerobic digestion processes over the time; however, the decay rates varied with pathogens. The effect of temperature on Salmonella spp. and Listeria monocytogenes survival was different than the E. coli survival. In thermophilic temperatures (42 and 50 °C), decay rate was considerable greater compared to the mesophilic temperatures (30 and 35°C). The E. coli log reductions at 50 °C were 2.1 in both aerobic and anaerobic digestions after 13 days of incubation. The Salmonella spp. log reductions at 50 °C were 5.5 in aerobic digestion, and 5.9 in anaerobic digestion. The Listeria monocytogenes log reductions at 50 °C were 5.0 in aerobic digestion, and 5.6 in anaerobic digestion. The log reduction of E. coli, Salmonella spp., and Listeria monocytogens at 30 °C in aerobic environment were 0.1, 4.7, and 5.6, respectively. In anaerobic environment, the corresponding reductions were 0.4, 4.3, and 5.6, respectively. We anticipate that the outcomes of the study will help improving the existing animal waste management processes to control manure-borne pathogens.

Vimentin, a Novel NF-κB Regulator, Is Required for Meningitic Escherichia coli K1-Induced Pathogen Invasion and PMN Transmigration across the Blood-Brain Barrier.

PubMed

Huang, Sheng-He; Chi, Feng; Peng, Liang; Bo, Tao; Zhang, Bao; Liu, Li-Qun; Wu, Xuedong; Mor-Vaknin, Nirit; Markovitz, David M; Cao, Hong; Zhou, Yan-Hong

NF-κB activation, pathogen invasion, polymorphonuclear leukocytes (PMN) transmigration (PMNT) across the blood-brain barrier (BBB) are the pathogenic triad hallmark features of bacterial meningitis, but the mechanisms underlying these events remain largely unknown. Vimentin, which is a novel NF-κB regulator, is the primary receptor for the major Escherichia coli K1 virulence factor IbeA that contributes to the pathogenesis of neonatal bacterial sepsis and meningitis (NSM). We have previously shown that IbeA-induced NF-κB signaling through its primary receptor vimentin as well as its co-receptor PTB-associated splicing factor (PSF) is required for pathogen penetration and leukocyte transmigration across the BBB. This is the first in vivo study to demonstrate how vimentin and related factors contributed to the pathogenic triad of bacterial meningitis. The role of vimentin in IbeA+ E. coli K1-induced NF-κB activation, pathogen invasion, leukocyte transmigration across the BBB has now been demonstrated by using vimentin knockout (KO) mice. In the in vivo studies presented here, IbeA-induced NF-κB activation, E. coli K1 invasion and polymorphonuclear neutrophil (PMN) transmigration across the BBB were significantly reduced in Vim-/- mice. Decreased neuronal injury in the hippocampal dentate gyrus was observed in Vim-/- mice with meningitis. The major inflammatory regulator α7 nAChR and several signaling molecules contributing to NF-κB activation (p65 and p-CamKII) were significantly reduced in the brain tissues of the Vim-/- mice with E. coli meningitis. Furthermore, Vim KO resulted in significant reduction in neuronal injury and in α7 nAChR-mediated calcium signaling. Vimentin, a novel NF-κB regulator, plays a detrimental role in the host defense against meningitic infection by modulating the NF-κB signaling pathway to increase pathogen invasion, PMN recruitment, BBB permeability and neuronal inflammation. Our findings provide the first evidence for Vim-dependent mechanisms underlying the pathogenic triad of bacterial meningitis.

Vimentin, a Novel NF-κB Regulator, Is Required for Meningitic Escherichia coli K1-Induced Pathogen Invasion and PMN Transmigration across the Blood-Brain Barrier

PubMed Central

Zhang, Bao; Liu, Li-Qun; Wu, Xuedong; Mor-Vaknin, Nirit; Markovitz, David M.; Cao, Hong; Zhou, Yan-Hong

2016-01-01

Background NF-κB activation, pathogen invasion, polymorphonuclear leukocytes (PMN) transmigration (PMNT) across the blood-brain barrier (BBB) are the pathogenic triad hallmark features of bacterial meningitis, but the mechanisms underlying these events remain largely unknown. Vimentin, which is a novel NF-κB regulator, is the primary receptor for the major Escherichia coli K1 virulence factor IbeA that contributes to the pathogenesis of neonatal bacterial sepsis and meningitis (NSM). We have previously shown that IbeA-induced NF-κB signaling through its primary receptor vimentin as well as its co-receptor PTB-associated splicing factor (PSF) is required for pathogen penetration and leukocyte transmigration across the BBB. This is the first in vivo study to demonstrate how vimentin and related factors contributed to the pathogenic triad of bacterial meningitis. Methodology/Principal Findings The role of vimentin in IbeA+ E. coli K1-induced NF-κB activation, pathogen invasion, leukocyte transmigration across the BBB has now been demonstrated by using vimentin knockout (KO) mice. In the in vivo studies presented here, IbeA-induced NF-κB activation, E. coli K1 invasion and polymorphonuclear neutrophil (PMN) transmigration across the BBB were significantly reduced in Vim-/- mice. Decreased neuronal injury in the hippocampal dentate gyrus was observed in Vim-/- mice with meningitis. The major inflammatory regulator α7 nAChR and several signaling molecules contributing to NF-κB activation (p65 and p-CamKII) were significantly reduced in the brain tissues of the Vim-/- mice with E. coli meningitis. Furthermore, Vim KO resulted in significant reduction in neuronal injury and in α7 nAChR-mediated calcium signaling. Conclusion/Significance Vimentin, a novel NF-κB regulator, plays a detrimental role in the host defense against meningitic infection by modulating the NF-κB signaling pathway to increase pathogen invasion, PMN recruitment, BBB permeability and neuronal inflammation. Our findings provide the first evidence for Vim-dependent mechanisms underlying the pathogenic triad of bacterial meningitis. PMID:27657497

Modeling the inactivatin of Escherichia coli 0157:H7 and uropathogenic E. coli in ground beef by high pressure processing and citral

USDA-ARS?s Scientific Manuscript database

Disease causing Escherichia coli commonly found in meat and poultry include intestinal pathogenic E. coli (iPEC) as well as extraintestinal types such as the Uropathogenic E. coli (UPEC). In this study we compared the resistance of iPEC (O157:H7) to UPEC in ground beef using High Pressure Processing...

Modeling the inactivation of Escherichia coli 0157:H7 and uropathogenic E.coli in ground chicken by high pressure processing and thymol

USDA-ARS?s Scientific Manuscript database

Disease causing Escherichia coli commonly found in meat and poultry include intestinal pathogenic E. coli (iPEC) as well as extraintestinal types such as the Uropathogenic E. coli (UPEC). In this study we compare the resistance of iPEC (O157:H7) to UPEC in chicken meat using High Pressure Processing...

Lethality prediction for Escherichia coli 0157:H7 and Uropathogenic E. coli in ground chicken treated with high pressure processing and trans-cinnamaldehyde

USDA-ARS?s Scientific Manuscript database

Pathogenic Escherichia coli, intestinal (O157:H7) as well as extraintestinal types (Uropathogenic E. coli (UPEC)) are commonly found in many foods including chicken meat. In this study we compared the resistance of E. coli O157:H7 to UPEC in chicken meat under the stresses of high hydrostatic pressu...

Secretome analysis of diarrhea-inducing strains of Escherichia coli

PubMed Central

Nirujogi, Raja Sekhar; Muthusamy, Babylakshmi; Kim, Min-Sik; Sathe, Gajanan J.; Lakshmi, P.T.V.; Kovbasnjuk, Olga N.; Prasad, T.S. Keshava; Wade, Mary; Jabbour, Rabih E.

2017-01-01

Secreted proteins constitute a major part of virulence factors that are responsible for pathogenesis caused by Gram-negative bacteria. Enterohemorrhagic Escherichia coli, O157:H7, is the major pathogen often causing outbreaks. However, studies have reported that the significant outbreaks caused by non-O157:H7 E. coli strains, also known as “Big-Six” serogroup strains, are increasing. There is no systematic study describing differential secreted proteins from these non-O157:H7 E. coli strains. In this study, we carried out MS-based differential secretome analysis using tandem mass tags labeling strategy of non-O157:H7 E. coli strains, O103, O111, O121, O145, O26, and O45. We identified 1241 proteins, of which 565 proteins were predicted to be secreted. We also found that 68 proteins were enriched in type III secretion system and several of them were differentially expressed across the strains. Additionally, we identified several strain-specific secreted proteins that could be used for developing potential markers for the identification and strain-level differentiation. To our knowledge, this study is the first comparative proteomic study on secretome of E. coli Big-Six serogroup and the several of these strain-specific secreted proteins can be further studied to develop potential markers for identification and strain-level differentiation. Moreover, the results of this study can be utilized in several applications, including food safety, diagnostics of E. coli outbreaks, and detection and identification of bio threats in biodefense. PMID:28070933

Shared and organism-specific host responses to childhood diarrheal diseases revealed by whole blood transcript profiling.

PubMed

DeBerg, Hannah A; Zaidi, Mussaret B; Altman, Matthew C; Khaenam, Prasong; Gersuk, Vivian H; Campos, Freddy D; Perez-Martinez, Iza; Meza-Segura, Mario; Chaussabel, Damien; Banchereau, Jacques; Estrada-Garcia, Teresa; Linsley, Peter S

2018-01-01

Globally, diarrheal diseases are a leading cause of death in children under five and disproportionately affect children in developing countries. Children who contract diarrheal diseases are rarely screened to identify the etiologic agent due to time and cost considerations associated with pathogen-specific screening and hence pathogen-directed therapy is uncommon. The development of biomarkers to rapidly identify underlying pathogens could improve treatment options and clinical outcomes in childhood diarrheal diseases. Here, we perform RNA sequencing on blood samples collected from children evaluated in an emergency room setting with diarrheal disease where the pathogen(s) present are known. We determine host response gene signatures specific to Salmonella, Shigella and rotavirus, but not E. coli, infections that distinguish them from each other and from healthy controls. Specifically, we observed differential expression of genes related to chemokine receptors or inflammasome signaling in Shigella cases, such as CCR3, CXCR8, and NLRC4, and interferon response genes, such as IFI44 and OASL, in rotavirus cases. Our findings add insight into the host peripheral immune response to these pathogens, and suggest strategies and limitations for the use host response transcript signatures for diagnosing the etiologic agent of childhood diarrheal diseases.

Enteric Pathogens Associated with Childhood Diarrhea in Tripoli-Libya

PubMed Central

Rahouma, Amal; Klena, John D.; Krema, Zaineb; Abobker, Abdalwahed A.; Treesh, Khalid; Franka, Ezzedin; Abusnena, Omar; Shaheen, Hind I.; El Mohammady, Hanan; Abudher, Abdulhafid; Ghenghesh, Khalifa Sifaw

2011-01-01

Stool samples from children < 5 years of age with diarrhea (N = 239) were examined for enteric pathogens using a combination of culture, enzyme-immunoassay, and polymerase chain reaction methods. Pathogens were detected in 122 (51%) stool samples; single pathogens were detected in 37.2% and co-pathogens in 13.8% of samples. Norovirus, rotavirus, and diarrheagenic Escherichia coli (DEC) were the most frequently detected pathogens (15.5%, 13.4%, and 11.2%, respectively); Salmonella, adenovirus, and Aeromonas were detected less frequently (7.9%, 7.1%, and 4.2%). The most commonly detected DEC was enteroaggregative E. coli (5.4%). Resistance to ≥ 3 antimicrobials was observed in 60% (18/30) of the bacterial pathogens. Salmonella resistance to ciprofloxacin (63.1%) has become a concern. Enteric viral pathogens were the most significant causative agents of childhood diarrhea in Tripoli. Bacterial pathogens were also important contributors to pediatric diarrhea. The emergence of ciprofloxacin-resistant Salmonella represents a serious health problem that must be addressed by Libyan health authorities PMID:21633024

Evolution of Bacterial Global Modulators: Role of a Novel H-NS Paralogue in the Enteroaggregative Escherichia coli Strain 042

PubMed Central

2018-01-01

ABSTRACT Bacterial genomes sometimes contain genes that code for homologues of global regulators, the function of which is unclear. In members of the family Enterobacteriaceae, cells express the global regulator H-NS and its paralogue StpA. In Escherichia coli, out of providing a molecular backup for H-NS, the role of StpA is poorly characterized. The enteroaggregative E. coli strain 042 carries, in addition to the hns and stpA genes, a third gene encoding an hns paralogue (hns2). We present in this paper information about its biological function. Transcriptomic analysis has shown that the H-NS2 protein targets a subset of the genes targeted by H-NS. Genes targeted by H-NS2 correspond mainly with horizontally transferred (HGT) genes and are also targeted by the Hha protein, a fine-tuner of H-NS activity. Compared with H-NS, H-NS2 expression levels are lower. In addition, H-NS2 expression exhibits specific features: it is sensitive to the growth temperature and to the nature of the culture medium. This novel H-NS paralogue is widespread within the Enterobacteriaceae. IMPORTANCE Global regulators such as H-NS play key relevant roles enabling bacterial cells to adapt to a changing environment. H-NS modulates both core and horizontally transferred (HGT) genes, but the mechanism by which H-NS can differentially regulate these genes remains to be elucidated. There are several instances of bacterial cells carrying genes that encode homologues of the global regulators. The question is what the roles of these proteins are. We noticed that the enteroaggregative E. coli strain 042 carries a new hitherto uncharacterized copy of the hns gene. We decided to investigate why this pathogenic E. coli strain requires an extra H-NS paralogue, termed H-NS2. In our work, we show that H-NS2 displays specific expression and regulatory properties. H-NS2 targets a subset of H-NS-specific genes and may help to differentially modulate core and HGT genes by the H-NS cellular pool. PMID:29577085

Evolution of Bacterial Global Modulators: Role of a Novel H-NS Paralogue in the Enteroaggregative Escherichia coli Strain 042.

PubMed

Prieto, A; Bernabeu, M; Aznar, S; Ruiz-Cruz, S; Bravo, A; Queiroz, M H; Juárez, A

2018-01-01

Bacterial genomes sometimes contain genes that code for homologues of global regulators, the function of which is unclear. In members of the family Enterobacteriaceae , cells express the global regulator H-NS and its paralogue StpA. In Escherichia coli , out of providing a molecular backup for H-NS, the role of StpA is poorly characterized. The enteroaggregative E. coli strain 042 carries, in addition to the hns and stpA genes, a third gene encoding an hns paralogue ( hns2 ). We present in this paper information about its biological function. Transcriptomic analysis has shown that the H-NS2 protein targets a subset of the genes targeted by H-NS. Genes targeted by H-NS2 correspond mainly with horizontally transferred (HGT) genes and are also targeted by the Hha protein, a fine-tuner of H-NS activity. Compared with H-NS, H-NS2 expression levels are lower. In addition, H-NS2 expression exhibits specific features: it is sensitive to the growth temperature and to the nature of the culture medium. This novel H-NS paralogue is widespread within the Enterobacteriaceae . IMPORTANCE Global regulators such as H-NS play key relevant roles enabling bacterial cells to adapt to a changing environment. H-NS modulates both core and horizontally transferred (HGT) genes, but the mechanism by which H-NS can differentially regulate these genes remains to be elucidated. There are several instances of bacterial cells carrying genes that encode homologues of the global regulators. The question is what the roles of these proteins are. We noticed that the enteroaggregative E. coli strain 042 carries a new hitherto uncharacterized copy of the hns gene. We decided to investigate why this pathogenic E. coli strain requires an extra H-NS paralogue, termed H-NS2. In our work, we show that H-NS2 displays specific expression and regulatory properties. H-NS2 targets a subset of H-NS-specific genes and may help to differentially modulate core and HGT genes by the H-NS cellular pool.

Contribution of Urease to Colonization by Shiga Toxin-Producing Escherichia coli

PubMed Central

Steyert, Susan R.

2012-01-01

Shiga toxin-producing Escherichia coli (STEC) is a food-borne pathogen with a low infectious dose that colonizes the colon in humans and can cause severe clinical manifestations such as hemolytic-uremic syndrome. The urease enzyme, encoded in the STEC chromosome, has been demonstrated to act as a virulence factor in other bacterial pathogens. The NH3 produced as urease hydrolyzes urea can aid in buffering bacteria in acidic environments as well as provide an easily assimilated source of nitrogen that bacteria can use to gain a metabolic advantage over intact microflora. Here, we explore the role of urease in STEC pathogenicity. The STEC urease enzyme exhibited maximum activity near neutral pH and during the stationary-growth phase. Experiments altering growth conditions performed with three phylogenetically distinct urease-positive strains demonstrated that the STEC ure gene cluster is inducible by neither urea nor pH but does respond to nitrogen availability. Quantitative reverse transcription-PCR (qRT-PCR) data indicate that nitrogen inhibits the transcriptional response. The deletion of the ure gene locus was constructed in STEC strain 88-0643, and the ure mutant was used with the wild-type strain in competition experiments in mouse models to examine the contribution of urease. The wild-type strain was twice as likely to survive passage through the acidic stomach and demonstrated an enhanced ability to colonize the intestinal tract compared to the ure mutant strain. These in vivo experiments reveal that, although the benefit STEC gains from urease expression is modest and not absolutely required for colonization, urease can contribute to the pathogenicity of STEC. PMID:22665380

Expression and purification of antimicrobial peptide adenoregulin with C-amidated terminus in Escherichia coli.

PubMed

Cao, Wei; Zhou, Yuxun; Ma, Yushu; Luo, Qingping; Wei, Dongzhi

2005-04-01

Adenoregulin is a 33 amino acid antimicrobial peptide isolated from the skin of the arboreal frog Phyllomedusa bicolor. Natural adenoregulin is synthesized with an amidated valine residue at C-terminus and shows lethal effects against filamentous fungi, as well as a broad spectrum of pathogenic microorganisms. A synthetic gene for adenoregulin (ADR) with an additional amino acid glutamine at C-terminus was cloned into pET32a vector to allow expression of ADR as a Trx fusion protein in Escherichia coli BL21(DE3). The resulting expression level of the fusion protein could reach up to 20% of the total cell proteins. The fusion protein could be purified effectively by Ni2+-chelating chromatography. Released from the fusion protein by enterokinase cleavage and purified to homogeneity, the recombinant ADR displayed antimicrobial activity similar to that of the synthetic ADR reported earlier. Comparing the antimicrobial activities of the recombinant adenoregulin with C-amidated terminus to that without an amidated C-terminus, we found that the amide of glutamine at C-terminus of ADR improved its potency on certain microorganisms such as Tritirachium album and Saccharomyces cerevisiae.

Seeing red; the development of pON.mCherry, a broad-host range constitutive expression plasmid for Gram-negative bacteria.

PubMed

Gebhardt, Michael J; Jacobson, Rachael K; Shuman, Howard A

2017-01-01

The development of plasmid-mediated gene expression control in bacteria revolutionized the field of bacteriology. Many of these expression control systems rely on the addition of small molecules, generally metabolites or non-metabolized analogs thereof, to the growth medium to induce expression of the genes of interest. The paradigmatic example of an expression control system is the lac system from Escherichia coli, which typically relies on the Ptac promoter and the Lac repressor, LacI. In many cases, however, constitutive gene expression is desired, and other experimental approaches require the coordinated control of multiple genes. While multiple systems have been developed for use in E. coli and its close relatives, the utility and/or functionality of these tools does not always translate to other species. For example, for the Gram-negative pathogen, Legionella pneumophila, a causative agent of Legionnaires' Disease, the aforementioned Ptac system represents the only well-established expression control system. In order to enhance the tools available to study bacterial gene expression in L. pneumophila, we developed a plasmid, pON.mCherry, which confers constitutive gene expression from a mutagenized LacI binding site. We demonstrate that pON.mCherry neither interferes with other plasmids harboring an intact LacI-Ptac expression system nor alters the growth of Legionella species during intracellular growth. Furthermore, the broad-host range plasmid backbone of pON.mCherry allows constitutive gene expression in a wide variety of Gram-negative bacterial species, making pON.mCherry a useful tool for the greater research community.

Inactivation of Escherichia coli O157:H7 and Salmonella deposited on gloves in a liquid state and subjected to drying conditions.

PubMed

Erickson, Marilyn C; Liao, Jye-Yin; Webb, Cathy C; Habteselassie, Mussie Y; Cannon, Jennifer L

2018-02-02

Gloves are worn by workers harvesting ready-to-eat produce as a deterrent for contaminating the produce with enteric pathogens that may reside on their hands. As fields are not sterile environments, the probability for gloves to become contaminated still exists and therefore it is critical to understand the conditions that affect the survival of pathogens on gloves. Both Escherichia coli O157:H7 and Salmonella deposited on glove surfaces in a liquid state survived longer when the pathogen had been suspended in lettuce sap than when suspended in water. Despite this protection, pathogens deposited on clean single-use gloves were more likely to survive during drying than pathogens deposited on dirty gloves (a film of lettuce sap had been applied to the surface prior to pathogen application and soil had been ground into the gloves). Survival of both E. coli O157:H7 and Salmonella was biphasic with the greatest losses occurring during the first hour of drying followed by much slower losses in the ensuing hours. Pathogens grown in rich media (tryptic soy broth) versus minimal media (M9) as well as those cultured on solid agar versus liquid broth were also more likely to be resistant to desiccation when deposited onto gloves. Although survival of E. coli O157:H7 on nitrile gloves was in general greater than it was on latex gloves, the relative survival of Salmonella on the two glove types was inconsistent. Due to these inconsistencies, no one glove type is considered better than another in reducing the risk for contamination with enteric pathogens. In addition, the extended survival of what are generally referred to as stress-resistant pathogens suggests that gloves either be changed frequently during the day or washed in a disinfectant to reduce the risk of glove contamination that could otherwise contaminate product handled with the contaminated gloves. Copyright © 2017 Elsevier B.V. All rights reserved.

Conversion of commensal Escherichia coli K-12 to an invasive form via expression of a mutant histone-like protein.

PubMed

Koli, Preeti; Sudan, Sudhanshu; Fitzgerald, David; Adhya, Sankar; Kar, Sudeshna

2011-01-01

The HUα(E38K, V42L) mutant of the bacterial histone-like protein HU causes a major change in the transcription profile of the commensal organism Escherichia coli K-12 (Kar S, Edgar R, Adhya S, Proc. Natl. Acad. Sci. U. S. A. 102:16397-16402, 2005). Among the upregulated genes are several related to pathogenic interactions with mammalian cells, as evidenced by the expression of curli fibers, Ivy, and hemolysin E. When E. coli K-12/ HUα(E38K, V42L) was added to Int-407 cells, there was host cell invasion, phagosomal disruption, and intracellular replication. The invasive trait was also retained in a murine ileal loop model and intestinal explant assays. In addition to invasion, the internalized bacteria caused a novel subversion of host cell apoptosis through modification and regulation of the BH3-only proteins Bim(EL) and Puma. Changes in the transcription profile were attributed to positive supercoiling of DNA leading to the altered availability of relevant promoters. Using the E. coli K-12/HUα(E38K, V42L) variant as a model, we propose that traditional commensal E. coli can adopt an invasive lifestyle through reprogramming its cellular transcription, without gross genetic changes. Escherichia coli K-12 is well established as a benign laboratory strain and a human intestinal commensal. Recent evidences, however, indicate that the typical noninvasive nature of resident E. coli can be reversed under specific circumstances even in the absence of any major genomic flux. We previously engineered an E. coli strain with a mutant histone-like protein, HU, which exhibited significant changes in nucleoid organization and global transcription. Here we showed that the changes induced by the mutant HU have critical functional consequences: from a strict extracellular existence, the mutant E. coli adopts an almost obligate intracellular lifestyle. The internalized E. coli exhibits many of the prototypical characteristics of traditional intracellular bacteria, like phagosomal escape, intracellular replication, and subversion of host cell apoptosis. We suggest that E. coli K-12 can switch between widely divergent lifestyles in relation to mammalian host cells by reprogramming its cellular transcription program and without gross changes in its genomic content.

The enemy within us: lessons from the 2011 European Escherichia coli O104:H4 outbreak.

PubMed

Karch, Helge; Denamur, Erick; Dobrindt, Ulrich; Finlay, B Brett; Hengge, Regine; Johannes, Ludgers; Ron, Eliora Z; Tønjum, Tone; Sansonetti, Philippe J; Vicente, Miguel

2012-09-01

In response to the 2011 European health alert caused by a pathogenic Escherichia coli O104:H4 outbreak, the European Academy of Microbiology (EAM), established by the Federation of European Microbiological Societies (FEMS), convened a meeting in Paris on November 30th, 2011 on 'EHEC infection and control' attended by world renowned experts in pathogenic E. coli. The major aims of this group were to review the scientific issues raised by the outbreak, to assess the handling of the crisis at the scientific and political levels, and to propose future actions. Several conclusions, which will have impact on future potential E. coli outbreaks, are outlined here. Copyright © 2012 EMBO Molecular Medicine.

Plant extracts, spices, and essential oils inactivate E. coli O157:H7 pathogens and reduce formation of potentially carcinogenic heterocyclic amines in grilled beef patties

USDA-ARS?s Scientific Manuscript database

Meats need to be sufficiently heated to inactivate foodborne pathogens such as Escherichia coli O157:H7. High-temperature heat treatment used to prepare well-done meats could, however, increase the formation of potentially carcinogenic heterocyclic amines (HCAs). The objective of this study was to ...

Survival and persistence of non-pathogenic Escherichia coli and attenuated Escherichia coli O157:H7 in soils amended with animal manure in a greenhouse environment

USDA-ARS?s Scientific Manuscript database

Biological soil amendments (BSA's), including dairy cattle, poultry litter, and horse manure, play an important role in agriculture but may contain pathogens that can contaminate raw or ready-to-eat fruit and vegetable crops that are consumed raw. Proposed FDA standards include a 90- or 120-day inte...

Comparative genomic analysis shows that avian pathogenic Escherichia coli isolate IMT5155 (O2:K1:H5; ST complex 95, ST140) shares close relationship with ST95 APEC O1:K1 and human ExPEC O18:K1 strains.

PubMed

Zhu Ge, Xiangkai; Jiang, Jingwei; Pan, Zihao; Hu, Lin; Wang, Shaohui; Wang, Haojin; Leung, Frederick C; Dai, Jianjun; Fan, Hongjie

2014-01-01

Avian pathogenic E. coli and human extraintestinal pathogenic E. coli serotypes O1, O2 and O18 strains isolated from different hosts are generally located in phylogroup B2 and ST complex 95, and they share similar genetic characteristics and pathogenicity, with no or minimal host specificity. They are popular objects for the study of ExPEC genetic characteristics and pathogenesis in recent years. Here, we investigated the evolution and genetic blueprint of APEC pathotype by performing phylogenetic and comparative genome analysis of avian pathogenic E. coli strain IMT5155 (O2:K1:H5; ST complex 95, ST140) with other E. coli pathotypes. Phylogeny analyses indicated that IMT5155 has closest evolutionary relationship with APEC O1, IHE3034, and UTI89. Comparative genomic analysis showed that IMT5155 and APEC O1 shared significant genetic overlap/similarities with human ExPEC dominant O18:K1 strains (IHE3034 and UTI89). Furthermore, the unique PAI I5155 (GI-12) was identified and found to be conserved in APEC O2 serotype isolates. GI-7 and GI-16 encoding two typical T6SSs in IMT5155 might be useful markers for the identification of ExPEC dominant serotypes (O1, O2, and O18) strains. IMT5155 contained a ColV plasmid p1ColV5155, which defined the APEC pathotype. The distribution analysis of 10 sequenced ExPEC pan-genome virulence factors among 47 sequenced E. coli strains provided meaningful information for B2 APEC/ExPEC-specific virulence factors, including several adhesins, invasins, toxins, iron acquisition systems, and so on. The pathogenicity tests of IMT5155 and other APEC O1:K1 and O2:K1 serotypes strains (isolated in China) through four animal models showed that they were highly virulent for avian colisepticemia and able to cause septicemia and meningitis in neonatal rats, suggesting zoonotic potential of these APEC O1:K1 and O2:K1 isolates.

Different Cellular Origins and Functions of Extracellular Proteins from Escherichia coli O157:H7 and O104:H4 as Determined by Comparative Proteomic Analysis.

PubMed

Islam, Nazrul; Nagy, Attila; Garrett, Wesley M; Shelton, Dan; Cooper, Bret; Nou, Xiangwu

2016-07-15

Extracellular proteins play important roles in bacterial interactions with the environmental matrices. In this study, we examined the extracellular proteins from Escherichia coli O157:H7 and O104:H4 by tandem mass spectrometry. We identified 500 and 859 proteins from the growth media of E. coli O157:H7 and O104:H4, respectively, including 371 proteins common to both strains. Among proteins that were considered specific to E. coli O157:H7 or present at higher relative abundances in O157:H7 medium, most (57 of 65) had secretion signal sequences in their encoding genes. Noticeably, the proteins included locus of enterocyte effacement (LEE) virulence factors, proteins required for peptidyl-lipoprotein accumulation, and proteins involved in iron scavenging. In contrast, a much smaller proportion of proteins (37 of 150) that were considered specific to O104:H4 or presented at higher relative abundances in O104:H4 medium had signals targeting them for secretion. These proteins included Shiga toxin 2 subunit B and O104:H4 signature proteins, including AAF/1 major fimbrial subunit and serine protease autotransporters. Most of the abundant proteins from the growth medium of E. coli O104:H4 were annotated as having functions in the cytoplasm. We provide evidence that the extensive presence of cytoplasmic proteins in E. coli O104:H4 growth medium was due to biological processes independent of cell lysis, indicating alternative mechanisms for this potent pathogen releasing cytoplasmic contents into the growth milieu, which could play a role in interaction with the environmental matrices, such as pathogenesis and biofilm formation. In this study, we compared the extracellular proteins from two of the most prominent foodborne pathogenic E. coli organisms that have caused severe outbreaks in the United States and in Europe. E. coli O157:H7 is a well-studied Shiga toxigenic foodborne pathogen of the enterohemorrhagic pathotype that has caused numerous outbreaks associated with various contaminated foods worldwide. E. coli O104:H4 is a newly emerged Shiga toxigenic foodborne pathogen of the enteroaggregative pathotype that gained notoriety for causing one of the most deadly foodborne outbreaks in Europe in 2011. Comparison of proteins in the growth medium revealed significant differences in the compositions of the extracellular proteins for these two pathogens. These differences may provide valuable information regarding the cellular responses of these pathogens to their environment, including cell survival and pathogenesis. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Effect of storage temperature and duration on the behavior of Escherichia coli O157:H7 on packaged fresh-cut salad containing romaine and iceberg lettuce.

PubMed

Luo, Yaguang; He, Qiang; McEvoy, James L

2010-09-01

This study investigated the impact of storage temperature and duration on the fate of Escherichia coli O157:H7 on commercially packaged lettuce salads, and on product quality. Fresh-cut Romaine and Iceberg lettuce salads of different commercial brands were obtained from both retail and wholesale stores. The packages were cut open at one end, the lettuce salad inoculated with E. coli O157:H7 via a fine mist spray, and resealed with or without an initial N(2) flush to match the original package atmospheric levels. The products were stored at 5 and 12 °C until their labeled "Best If Used By" dates, and the microbial counts and product quality were monitored periodically. The results indicate that storage at 5 °C allowed E. coli O157:H7 to survive, but limited its growth, whereas storage at 12 °C facilitated the proliferation of E. coli O157:H7. There was more than 2.0 log CFU/g increase in E. coli O157:H7 populations on lettuce when held at 12 °C for 3 d, followed by additional growth during the remainder of the storage period. Although there was eventually a significant decline in visual quality of lettuce held at 12 °C, the quality of this lettuce was still fully acceptable when E. coli O157:H7 growth reached a statistically significant level. Therefore, maintaining fresh-cut products at 5 °C or below is critical for reducing the food safety risks as E. coli O157:H7 grows at a rapid, temperature-dependent rate prior to significant quality deterioration. Specific information regarding the effect of temperature on pathogen growth on leafy greens is needed to develop science-based food safety guidelines and practices by the regulatory agencies and produce industry. Temperature control is commonly thought to promote quality of leafy greens, not safety, based at least partially on a theory that product quality deterioration precedes pathogen growth at elevated temperatures. This prevalent attitude results in temperature abuse incidents being frequently overlooked in the supply chain. This study demonstrates that human pathogens, such as E. coli O157:H7, can grow significantly on commercially packaged lettuce salads while the product's visual quality is fully acceptable. Packaged fresh-cut salads are marketed as "ready-to-eat" while lacking an effective pathogen kill step during their preparation. Thus, maintaining storage temperature at 5 °C or below is critical to prevent pathogen proliferation and mitigate food safety risks should pathogen contamination inadvertently occur during crop growth or postharvest fresh-cut processing.

Expression and crystallization of SeDsbA, SeDsbL and SeSrgA from Salmonella enterica serovar Typhimurium.

PubMed

Jarrott, R; Shouldice, S R; Guncar, G; Totsika, M; Schembri, M A; Heras, B

2010-05-01

Pathogens require protein-folding enzymes to produce functional virulence determinants. These foldases include the Dsb family of proteins, which catalyze oxidative folding in bacteria. Bacterial disulfide catalytic processes have been well characterized in Escherichia coli K-12 and these mechanisms have been extrapolated to other organisms. However, recent research indicates that the K-12 complement of Dsb proteins is not common to all bacteria. Importantly, many pathogenic bacteria have an extended arsenal of Dsb catalysts that is linked to their virulence. To help to elucidate the process of oxidative folding in pathogens containing a wide repertoire of Dsb proteins, Salmonella enterica serovar Typhimurium has been focused on. This Gram-negative bacterium contains three DsbA proteins: SeDsbA, SeDsbL and SeSrgA. Here, the expression, purification, crystallization and preliminary diffraction analysis of these three proteins are reported. SeDsbA, SeDsbL and SeSrgA crystals diffracted to resolution limits of 1.55, 1.57 and 2.6 A and belonged to space groups P2(1), P2(1)2(1)2 and C2, respectively.

Examination of the Source and Extended Virulence Genotypes of Escherichia coli Contaminating Retail Poultry Meat

PubMed Central

Johnson, Timothy J.; Logue, Catherine M.; Wannemuehler, Yvonne; Kariyawasam, Subhashinie; Doetkott, Curt; DebRoy, Chitrita; White, David G.

2009-01-01

Abstract Extraintestinal pathogenic Escherichia coli (ExPEC) are major players in human urinary tract infections, neonatal bacterial meningitis, and sepsis. Recently, it has been suggested that there might be a zoonotic component to these infections. To determine whether the E. coli contaminating retail poultry are possible extraintestinal pathogens, and to ascertain the source of these contaminants, they were assessed for their genetic similarities to E. coli incriminated in colibacillosis (avian pathogenic E. coli [APEC]), E. coli isolated from multiple locations of apparently healthy birds at slaughter, and human ExPEC. It was anticipated that the retail poultry isolates would most closely resemble avian fecal E. coli since only apparently healthy birds are slaughtered, and fecal contamination of carcasses is the presumed source of meat contamination. Surprisingly, this supposition proved incorrect, as the retail poultry isolates exhibited gene profiles more similar to APEC than to fecal isolates. These isolates contained a number of ExPEC-associated genes, including those associated with ColV virulence plasmids, and many belonged to the B2 phylogenetic group, known to be virulent in human hosts. Additionally, E. coli isolated from the crops and gizzards of apparently healthy birds at slaughter also contained a higher proportion of ExPEC-associated genes than did the avian fecal isolates examined. Such similarities suggest that the widely held beliefs about the sources of poultry contamination may need to be reassessed. Also, the presence of ExPEC-like clones on retail poultry meat means that we cannot yet rule out poultry as a source of ExPEC human disease. PMID:19580453

Biocontrol activity of an alkaline serine protease from Aureobasidium pullulans expressed in Pichia pastoris against four postharvest pathogens on apple.

PubMed

Banani, Houda; Spadaro, Davide; Zhang, Dianpeng; Matic, Slavica; Garibaldi, Angelo; Gullino, Maria Lodovica

2014-07-16

The yeast-like fungus Aureobasidium pullulans PL5 is a microbial antagonist against postharvest pathogens of fruits. The strain is able to produce hydrolases, including glucanases, chitinases and proteases. The alkaline serine protease gene ALP5 from A. pullulans was cloned, inserted into the vector pPIC9 to construct pPIC9/ALP5, and then expressed in Pichia pastoris strain KM71. ALP5 had a molecular mass of 42.9kDa after 5days growth with 1% methanol induction at 28°C. The recombinant protease expressed in P. pastoris showed its highest activity under alkaline conditions (at pH10) and a temperature of 50°C. The antifungal activity of the recombinant protease was investigated against Penicillium expansum, Botrytis cinerea, Monilinia fructicola and Alternaria alternata in vitro and on apple. The recombinant protease reduced significantly the spore germination and the germ tube length of the tested pathogens in PDB medium. The highest level of protease efficacy was observed against M. fructicola and B. cinerea, whereas a lower efficacy was observed against P. expansum and A. alternata indicating a possible effect of the pathogen cell wall composition on the proteolytic activity of the recombinant protease. The presence of protease was able to cause the swelling of the hyphae of B. cinerea, under an optical microscope. The recombinant protease expressed in P. pastoris was more active against the pathogens in vitro than the same enzyme expressed in E. coli in previous studies. The efficacy of ALP5 was also evaluated against the pathogens in vivo on cv Golden Delicious apples. The protease was more efficient in controlling M. fructicola, B. cinerea and P. expansum than A. alternata. However, the extent of the activity was dependent on the enzyme concentration and the length of fruit storage. This study demonstrated the capacity of the alkaline serine protease to keep its enzymatic activity for some days in the unfavorable environment of the fruit wounds. The alkaline serine protease could be developed as a postharvest treatment with antimicrobial activity for fruit undergoing a short storage period. Copyright © 2014 Elsevier B.V. All rights reserved.

iTRAQ-Based Quantitative Proteomic Analysis of the Antimicrobial Mechanism of Peptide F1 against Escherichia coli.

PubMed

Miao, Jianyin; Chen, Feilong; Duan, Shan; Gao, Xiangyang; Liu, Guo; Chen, Yunjiao; Dixon, William; Xiao, Hang; Cao, Yong

2015-08-19

Antimicrobial peptides have received increasing attention in the agricultural and food industries due to their potential to control pathogens. However, to facilitate the development of novel peptide-based antimicrobial agents, details regarding the molecular mechanisms of these peptides need to be elucidated. The aim of this study was to investigate the antimicrobial mechanism of peptide F1, a bacteriocin found in Tibetan kefir, against Escherichia coli at protein levels using iTRAQ-based quantitative proteomic analysis. In response to treatment with peptide F1, 31 of the 280 identified proteins in E. coli showed alterations in their expression, including 10 down-regulated proteins and 21 up-regulated proteins. These 31 proteins all possess different molecular functions and are involved in different molecular pathways, as is evident in referencing the Kyoto Encyclopedia of Genes and Genomes pathways. Specifically, pathways that were significantly altered in E. coli in response to peptide F1 treatment include the tricarboxylic acid cycle, oxidative phosphorylation, glycerophospholipid metabolism, and the cell cycle-caulobacter pathways, which was also associated with inhibition of the cell growth, induction of morphological changes, and cell death. The results provide novel insights into the molecular mechanisms of antimicrobial peptides.

Quantitative Metabolomics Reveals an Epigenetic Blueprint for Iron Acquisition in Uropathogenic Escherichia coli

PubMed Central

Henderson, Jeffrey P.; Crowley, Jan R.; Pinkner, Jerome S.; Walker, Jennifer N.; Tsukayama, Pablo; Stamm, Walter E.; Hooton, Thomas M.; Hultgren, Scott J.

2009-01-01

Bacterial pathogens are frequently distinguished by the presence of acquired genes associated with iron acquisition. The presence of specific siderophore receptor genes, however, does not reliably predict activity of the complex protein assemblies involved in synthesis and transport of these secondary metabolites. Here, we have developed a novel quantitative metabolomic approach based on stable isotope dilution to compare the complement of siderophores produced by Escherichia coli strains associated with intestinal colonization or urinary tract disease. Because uropathogenic E. coli are believed to reside in the gut microbiome prior to infection, we compared siderophore production between urinary and rectal isolates within individual patients with recurrent UTI. While all strains produced enterobactin, strong preferential expression of the siderophores yersiniabactin and salmochelin was observed among urinary strains. Conventional PCR genotyping of siderophore receptors was often insensitive to these differences. A linearized enterobactin siderophore was also identified as a product of strains with an active salmochelin gene cluster. These findings argue that qualitative and quantitative epi-genetic optimization occurs in the E. coli secondary metabolome among human uropathogens. Because the virulence-associated biosynthetic pathways are distinct from those associated with rectal colonization, these results suggest strategies for virulence-targeted therapies. PMID:19229321

Production of transgenic-cloned pigs expressing large quantities of recombinant human lysozyme in milk.

PubMed

Lu, Dan; Liu, Shen; Shang, Shengzhe; Wu, Fangfang; Wen, Xiao; Li, Zhiyuan; Li, Yan; Hu, Xiaoxiang; Zhao, Yaofeng; Li, Qiuyan; Li, Ning

2015-01-01

Human lysozyme is a natural non-specific immune factor in human milk that plays an important role in the defense of breastfed infants against pathogen infection. Although lysozyme is abundant in human milk, there is only trace quantities in pig milk. Here, we successfully generated transgenic cloned pigs with the expression vector pBAC-hLF-hLZ-Neo and their first generation hybrids (F1). The highest concentration of recombinant human lysozyme (rhLZ) with in vitro bioactivity was 2759.6 ± 265.0 mg/L in the milk of F0 sows. Compared with wild-type milk, rhLZ milk inhibited growth of Escherichia coli K88 during the exponential growth phase. Moreover, rhLZ in milk from transgenic sows was directly absorbed by the intestine of piglets with no observable anaphylactic reaction. Our strategy may provide a powerful tool for large-scale production of this important human protein in pigs to improve resistance to pathogen infection.

Bacteria, viruses, and parasites in an intermittent stream protected from and exposed to pasturing cattle: prevalence, densities, and quantitative microbial risk assessment.

PubMed

Wilkes, G; Brassard, J; Edge, T A; Gannon, V; Jokinen, C C; Jones, T H; Neumann, N; Pintar, K D M; Ruecker, N; Schmidt, P J; Sunohara, M; Topp, E; Lapen, D R

2013-10-15

Over 3500 individual water samples, for 131 sampling times, targeting waterborne pathogens/fecal indicator bacteria were collected during a 7-year period from 4 sites along an intermittent stream running through a small livestock pasture system with and without cattle access-to-stream restriction measures. The study assessed the impact of cattle pasturing/riparian zone protection on: pathogen (bacterial, viral, parasite) occurrence, concentrations of fecal indicators, and quantitative microbial risk assessments (QMRA) of the risk of Cryptosporidium, Giardia and Escherichia coli O157:H7 infection in humans. Methodologies were developed to compute QMRA mean risks on the basis of water samples exhibiting potentially human infectious Cryptosporidium and E. coli based on genotyping Crytosporidium, and E. coli O157:H7 presence/absence information paired with enumerated E. coli. All Giardia spp. were considered infectious. No significant pasturing treatment effects were observed among pathogens, with the exception of Campylobacter spp. and E. coli O157:H7. Campylobacter spp. prevalence significantly decreased downstream through pasture treatments and E. coli O157:H7 was observed in a few instances in the middle of the unrestricted pasture. Densities of total coliform, fecal coliform, and E. coli reduced significantly downstream in the restricted pasture system, but not in the unrestricted system. Seasonal and flow conditions were associated with greater indicator bacteria densities, especially in the summer. Norovirus GII was detected at rates of 7-22% of samples for all monitoring sites, and rotavirus in 0-7% of samples for all monitoring sites; pasture treatment trends were not evident, however. Seasonal and stream flow variables (and their interactions) were relatively more important than pasture treatments for initially stratifying pathogen occurrence and higher fecal indicator bacteria densities. Significant positive associations among fecal indicator bacteria and Campylobacter spp. detection were observed. For QMRA, adjusting for the proportion of Cryptosporidium spp. detected that are infectious for humans reduces downstream risk estimates by roughly one order of magnitude. Using QMRA in this manner provides a more refined estimate of beneficial management practice effects on pathogen exposure risks to humans. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

Norwegian Sheep Are an Important Reservoir for Human-Pathogenic Escherichia coli O26:H11

PubMed Central

Sekse, Camilla; Lindstedt, Bjørn-Arne; Sunde, Marianne; Løbersli, Inger; Urdahl, Anne Margrete; Kapperud, Georg

2012-01-01

A previous national survey of Escherichia coli in Norwegian sheep detected eae-positive (eae+) E. coli O26:H11 isolates in 16.3% (80/491) of the flocks. The purpose of the present study was to evaluate the human-pathogenic potential of these ovine isolates by comparing them with E. coli O26 isolates from humans infected in Norway. All human E. coli O26 isolates studied carried the eae gene and shared flagellar type H11. Two-thirds of the sheep flocks and 95.1% of the patients harbored isolates containing arcA allele type 2 and espK and were classified as enterohemorrhagic E. coli (EHEC) (stx positive) or EHEC-like (stx negative). These isolates were further divided into group A (EspK2 positive), associated with stx2-EDL933 and stcEO103, and group B (EspK1 positive), associated with stx1a. Although the stx genes were more frequently present in isolates from patients (46.3%) than in those from sheep flocks (5%), more than half of the ovine isolates in the EHEC/EHEC-like group had multiple-locus variable number of tandem repeat analysis (MLVA) profiles that were identical to those seen in stx-positive human O26:H11 isolates. This indicates that EHEC-like ovine isolates may be able to acquire stx-carrying bacteriophages and thereby have the possibility to cause serious illness in humans. The remaining one-third of the sheep flocks and two of the patients had isolates fulfilling the criteria for atypical enteropathogenic E. coli (aEPEC): arcA allele type 1 and espK negative (group C). The majority of these ovine isolates showed MLVA profiles not previously seen in E. coli O26:H11 isolates from humans. However, according to their virulence gene profile, the aEPEC ovine isolates should be considered potentially pathogenic for humans. In conclusion, sheep are an important reservoir of human-pathogenic E. coli O26:H11 isolates in Norway. PMID:22492457

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

Prevalence and Antibiogram Profiling of Escherichia coli Pathotypes Isolated from the Kat River and the Fort Beaufort Abstraction Water

PubMed Central

Nontongana, Nolonwabo; Sibanda, Timothy; Ngwenya, Elvis; Okoh, Anthony I.

2014-01-01

Escherichia coli is a widespread bacterium encompassing a variety of strains, ranging from highly pathogenic strains, causing worldwide outbreaks of severe diseases to avirulent, well characterized safe laboratory strains. This study evaluated the prevalence and antibiogram profiles of E. coli pathotypes isolated from the Kat River and Fort Beaufort abstraction water. A total of 171 out of 278 confirmed E. coli isolates were positive for at least one pathogenic determinant and these included enteropathogenic E. coli (6%), enterotoxigenic E. coli (47%), uropathogenic E. coli (2%), neonatal meningitis E. coli (5%), diffusely adherent E. coli (1%) and enterohaemorrhagic E. coli (1%). Interestingly, enteroinvasive and enteroaggregative E. coli were not detected. The phenotypic antibiogram profiles of the isolates revealed that all were resistant to penicillin G, while 98% and 38% of the pathotypes were resistant to ampicillin and trimethoprim-sulphamethoxazole, respectively. About 8% of the isolates were resistant to streptomycin. More than half of the isolates exhibited multiple antibiotic resistance with 44% being resistant to three antibiotics and 8% resistant to four antibiotics. We conclude that the Kat River is a reservoir of potentially virulent antibiotic resistant E. coli strains that can cause serious health risks to humans who drink raw water from this river, or in the case that consumption of treated drinking water coincides with failed drinking water processes. PMID:25119699

Potential pathogens, antimicrobial patterns and genotypic diversity of Escherichia coli isolates in constructed wetlands treating swine wastewater.

PubMed

Ibekwe, A M; Murinda, Shelton E; DebRoy, Chitrita; Reddy, Gudigopura B

2016-02-01

Escherichia coli populations originating from swine houses through constructed wetlands were analyzed for potential pathogens, antimicrobial susceptibility patterns, and genotypic diversity. Escherichia coli isolates (n = 493) were screened for the presence of the following virulence genes: stx1, stx2 and eae (Shiga toxin-producing E. coli [STEC]), heat-labile enterotoxin (LT) genes and heat stable toxin STa and STb (enterotoxigenic E. coli (ETEC), cytotoxin necrotizing factors 1 and 2 (cnf1 and cnf2 [necrotoxigenic E. coli- NTEC]), as well as O and H antigens, and the presence of the antibiotic resistance genes blaTEM, blaSHV, blaCMY-2, tet A, tet B, tet C, mph(A), aadA, StrA/B, sul1, sul2 and sul3. The commensal strains were further screened for 16 antimicrobials and characterized by BOX AIR-1 PCR for unique genotypes. The highest antibiotic resistance prevalence was for tetracycline, followed by erythromycin, ampicillin, streptomycin, sulfisoxazole and kanamycin. Our data showed that most of the isolates had high distribution of single or multidrug-resistant (MDR) genotypes. Therefore, the occurrence of MDR E. coli in the wetland is a matter of great concern due to possible transfer of resistance genes from nonpathogenic to pathogenic strains or vice versa in the environment. Published by Oxford University Press on behalf of FEMS 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Significance of Fecal Coliform-Positive Klebsiella1

PubMed Central

Bagley, Susan T.; Seidler, Ramon J.

1977-01-01

A total of 191 Klebsiella pneumoniae isolates of human clinical, bovine mastitis, and a wide variety of environmental sources were tested for fecal coliform (FC) response with the membrane filtration and most probable number techniques. Twenty-seven Escherichia coli cultures of human clinical and environmental origins were also tested. Eighty-five percent (49/58) of known pathogenic K. pneumoniae were FC positive, compared with 16% (19/120) of the environmental strains. E. coli results indicated 93% (13/14) of the clinical and 85% (11/13) of the environmental strains as FC positive. There was no significant difference in the incidence of FC-positive cultures between pathogenic Klebsiella and E. coli. pH measurements of K. pneumoniae and E. coli cultures growing in m-FC broth at 44.5°C revealed three distinct pH ranges correlating with colony morphology. β-Galactosidase assays of Klebsiella and E. coli cultures at 44.5°C indicated all were able to hydrolyze lactose, even if they were FC negative by the membrane filtration or most probable number techniques. The FC response pattern appears stable in K. pneumoniae. Three pathogenic cultures showed no change in FC responses after 270 generations of growth in sterile pulp mill effluent. Since K. pneumoniae is carried in the gastrointestinal tract of humans and animals and 85% of the tested pathogenic strains were FC positive, the isolation of FC-positive Klebsiella organisms from the environment would indicate their fecal or clinical origin or both. The added fact that K. pneumoniae is an opportunistic pathogen of increasing importance makes the occurrence of FC-positive environmental Klebsiella, particularly in large numbers, a potential human and animal health hazard. PMID:18086

No evidence for a bovine mastitis Escherichia coli pathotype.

PubMed

Leimbach, Andreas; Poehlein, Anja; Vollmers, John; Görlich, Dennis; Daniel, Rolf; Dobrindt, Ulrich

2017-05-08

Escherichia coli bovine mastitis is a disease of significant economic importance in the dairy industry. Molecular characterization of mastitis-associated E. coli (MAEC) did not result in the identification of common traits. Nevertheless, a mammary pathogenic E. coli (MPEC) pathotype has been proposed suggesting virulence traits that differentiate MAEC from commensal E. coli. The present study was designed to investigate the MPEC pathotype hypothesis by comparing the genomes of MAEC and commensal bovine E. coli. We sequenced the genomes of eight E. coli isolated from bovine mastitis cases and six fecal commensal isolates from udder-healthy cows. We analyzed the phylogenetic history of bovine E. coli genomes by supplementing this strain panel with eleven bovine-associated E. coli from public databases. The majority of the isolates originate from phylogroups A and B1, but neither MAEC nor commensal strains could be unambiguously distinguished by phylogenetic lineage. The gene content of both MAEC and commensal strains is highly diverse and dominated by their phylogenetic background. Although individual strains carry some typical E. coli virulence-associated genes, no traits important for pathogenicity could be specifically attributed to MAEC. Instead, both commensal strains and MAEC have very few gene families enriched in either pathotype. Only the aerobactin siderophore gene cluster was enriched in commensal E. coli within our strain panel. This is the first characterization of a phylogenetically diverse strain panel including several MAEC and commensal isolates. With our comparative genomics approach we could not confirm previous studies that argue for a positive selection of specific traits enabling MAEC to elicit bovine mastitis. Instead, MAEC are facultative and opportunistic pathogens recruited from the highly diverse bovine gastrointestinal microbiota. Virulence-associated genes implicated in mastitis are a by-product of commensalism with the primary function to enhance fitness in the bovine gastrointestinal tract. Therefore, we put the definition of the MPEC pathotype into question and suggest to designate corresponding isolates as MAEC.

The Arrhenius Equation As Means to Simulate E. Coli Survival in Waters

EPA Science Inventory

E. coli is an important microorganism indicator used to show the presence of pathogens and fecal contamination in waters. Knowing E. coli survival rates is important for assessing the severity of contamination that has occurred and making appropriate management evaluations. E. ...

Pathogenic Escherichia coli

USDA-ARS?s Scientific Manuscript database

Escherichia coli, a member of the Enterobacteriaceae family, is a part of the normal flora of the intestinal tract of humans and a variety of animals. E. coli strains are classified on the basis of antigenic differences in two surface components (serotyping), the somatic antigen (O) of the lipopoly...

Results of a 6-month survey of stool cultures for Escherichia coli O157:H7.

PubMed

Marshall, W F; McLimans, C A; Yu, P K; Allerberger, F J; Van Scoy, R E; Anhalt, J P

1990-06-01

Escherichia coli O157:H7 is a recently recognized enteric pathogen that causes acute hemorrhagic colitis. Although the infection is usually self-limited, it may be complicated by hemolytic uremic syndrome and thrombotic thrombocytopenic purpura. At our institution, stool specimens are now routinely cultured for this organism. To determine the prevalence of E. coli O157:H7-associated diarrhea in our patient population, we surveyed all submitted stool cultures for 6 months for this organism. Specimens were screened for non-sorbitol fermenting E. coli and confirmed by slide-agglutination and immobilization testing. Of 2,164 specimens, 10 yielded E. coli O157:H7. It was the fourth most common bacterial stool pathogen found. Bloody diarrhea and abdominal pain were the most common symptoms of the infected patients. E. coli O157:H7 causes sporadic infections in our patient population and should be considered in the differential diagnosis of acute hemorrhagic colitis.

Structural characterization of CFA/III and Longus type IVb pili from enterotoxigenic Escherichia coli.

PubMed

Kolappan, Subramaniapillai; Roos, Justin; Yuen, Alex S W; Pierce, Owen M; Craig, Lisa

2012-05-01

The type IV pili are helical filaments found on many Gram-negative pathogenic bacteria, with multiple diverse roles in pathogenesis, including microcolony formation, adhesion, and twitching motility. Many pathogenic enterotoxigenic Escherichia coli (ETEC) isolates express one of two type IV pili belonging to the type IVb subclass: CFA/III or Longus. Here we show a direct correlation between CFA/III expression and ETEC aggregation, suggesting that these pili, like the Vibrio cholerae toxin-coregulated pili (TCP), mediate microcolony formation. We report a 1.26-Å resolution crystal structure of CofA, the major pilin subunit from CFA/III. CofA is very similar in structure to V. cholerae TcpA but possesses a 10-amino-acid insertion that replaces part of the α2-helix with an irregular loop containing a 3(10)-helix. Homology modeling suggests a very similar structure for the Longus LngA pilin. A model for the CFA/III pilus filament was generated using the TCP electron microscopy reconstruction as a template. The unique 3(10)-helix insert fits perfectly within the gap between CofA globular domains. This insert, together with differences in surface-exposed residues, produces a filament that is smoother and more negatively charged than TCP. To explore the specificity of the type IV pilus assembly apparatus, CofA was expressed heterologously in V. cholerae by replacing the tcpA gene with that of cofA within the tcp operon. Although CofA was synthesized and processed by V. cholerae, no CFA/III filaments were detected, suggesting that the components of the type IVb pilus assembly system are highly specific to their pilin substrates.

The dps Gene of Symbiotic “Candidatus Legionella jeonii” in Amoeba proteus Responds to Hydrogen Peroxide and Phagocytosis▿

PubMed Central

Park, Miey; Yun, Seong Tae; Hwang, Sue-Yun; Chun, Choong-Ill; Ahn, Tae In

2006-01-01

To survive in host cells, intracellular pathogens or symbiotic bacteria require protective mechanisms to overcome the oxidative stress generated by phagocytic activities of the host. By genomic library tagging, we cloned a dps (stands for DNA-binding protein from starved cells) gene of the symbiotic “Candidatus Legionella jeonii” organism (called the X bacterium) (dpsX) that grows in Amoeba proteus. The gene encodes a 17-kDa protein (pI 5.19) with 91% homology to Dps and DNA-binding ferritin-like proteins of other organisms. The cloned gene complemented the dps mutant of Escherichia coli and conferred resistance to hydrogen peroxide. DpsX proteins purified from E. coli transformed with the dpsX gene were in oligomeric form, formed a complex with pBlueskript SKII DNA, and protected the DNA from DNase I digestion and H2O2-mediated damage. The expression of the dpsX gene in “Candidatus Legionella jeonii” was enhanced when the host amoeba was treated with 2 mM H2O2 and by phagocytic activities of the host cell. These results suggested that the Dps protein has a function protective of the bacterial DNA and that its gene expression responds to oxidative stress generated by phagocytic activities of the host cell. With regard to the fact that invasion of Legionella sp. into respiratory phagocytic cells causes pneumonia in mammals, further characterization of dpsX expression in the Legionella sp. that multiplies in a protozoan host in the natural environment may provide valuable information toward understanding the protective mechanisms of intracellular pathogens. PMID:16950918

Location-specific expression of chemokines, TNF-α and S100 proteins in a teat explant model.

PubMed

Lind, Monique; Sipka, Anja S; Schuberth, Hans-Joachim; Blutke, Andreas; Wanke, Rüdiger; Sauter-Louis, Carola; Duda, Katarzyna A; Holst, Otto; Rainard, Pascal; Germon, Pierre; Zerbe, Holm; Petzl, Wolfram

2015-04-01

The distal compartments of the udder are the first to interact with invading pathogens. The regulatory and effector functions of two major teat regions [Fürstenberg's rosette (FR); teat cistern (TC)] are largely unknown. The objective of this study was to establish an in vitro model with explants of the FR and the TC to analyse their response towards Escherichia coli LPS and Staphylococcus aureus lipoteichoic acid (LTA). Quantitative stereological analysis confirmed differences in the cellular composition of FR and TC explants. Chemokine (CXCL8, CCL5, CCL20) and TNF-α mRNA were expressed at low levels in both locations. Explant stimulation with LPS increased the mRNA abundance of all tested chemokines and TNF-α. Stimulation with LTA only induced CCL20 and CXCL8. LPS- and LTA-stimulated explant supernatants contained CXCL8 and CXCL3. Supernatants significantly attracted neutrophils in vitro. Compared with TC, the FR showed high constitutive mRNA expression of S100 proteins (A8, A9, A12). In the TC, both LPS and LTA significantly induced S100A8, whereas S100A9 and S100A12 expression was only induced by LPS. The novel model system underpins the role of the teat for recognising pathogens and shaping a pathogen- and location-specific immune response. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Synthetic Promoters Functional in Francisella novicida and Escherichia coli

PubMed Central

McWhinnie, Ralph L.

2014-01-01

In this work, we describe the identification of synthetic, controllable promoters that function in the bacterial pathogen Francisella novicida, a model facultative intracellular pathogen. Synthetic DNA fragments consisting of the tetracycline operator (tetO) flanked by a random nucleotide sequence were inserted into a Francisella novicida shuttle plasmid upstream of a promoterless artificial operon containing the reporter genes cat and lacZ. Fragments able to promote transcription were selected for based on their ability to drive expression of the cat gene, conferring chloramphenicol resistance. Promoters of various strengths were found, many of which were repressed in the presence of the tetracycline repressor (TetR) and promoted transcription only in the presence of the TetR inducer anhydrotetracycline. A subset of both constitutive and inducible synthetic promoters were characterized to find their induction ratios and to identify their transcription start sites. In cases where tetO was located between or downstream of the −10 and −35 regions of the promoter, control by TetR was observed. If the tetO region was upstream of the −35 region by more than 9 bp, it did not confer TetR control. We found that three of three promoters isolated in F. novicida functioned at a comparable level in E. coli; however, none of the 10 promoters isolated in E. coli functioned at a significant level in F. novicida. Our results allowed us to isolate minimal F. novicida promoters of 47 and 48 bp in length. PMID:24141126

Involvement of multiple stressors induced by non-thermal plasma-charged aerosols during inactivation of airborne bacteria

PubMed Central

Vaze, Nachiket D.; Park, Sin; Brooks, Ari D.; Fridman, Alexander; Joshi, Suresh G.

2017-01-01

A lab-scale, tunable, single-filament, point-to-point nonthermal dieletric-barrier discharge (DBD) plasma device was built to study the mechanisms of inactivation of aerosolized bacterial pathogens. The system inactivates airborne antibiotic-resistant pathogens efficiently. Nebulization mediated pre-optimized (4 log and 7 log) bacterial loads were challenged to plasma-charged aerosols, and lethal and sublethal doses determined using colony assay, and cell viability assay; and the loss of membrane potential and cellular respiration were determined using cell membrane potential assay and XTT assay. Using the strategies of Escherichia coli wildtype, over-expression mutant, deletion mutants, and peroxide and heat stress scavenging, we analyzed activation of intracellular reactive oxygen species (ROS) and heat shock protein (hsp) chaperons. Superoxide dismutase deletion mutants (ΔsodA, ΔsodB, ΔsodAΔsodB) and catalase mutants ΔkatG and ΔkatEΔkatG did not show significant difference from wildtype strain, and ΔkatE and ΔahpC was found significantly more susceptible to cell death than wildtype. The oxyR regulon was found to mediate plasma-charged aerosol-induced oxidative stress in bacteria. Hsp deficient E. coli (ΔhtpG, ΔgroEL, ΔclpX, ΔgrpE) showed complete inactivation of cells at ambient temperature, and the treatment at cold temperature (4°C) significantly protected hsp deletion mutants and wildtype cells, and indicate a direct involvement of hsp in plasma-charged aerosol mediated E. coli cell death. PMID:28166240

Inactivation of Escherichia coli O157:H7 and Salmonella Typhimurium in black pepper and red pepper by gamma irradiation.

PubMed

Song, Won-Jae; Sung, Hye-Jung; Kim, Sung-Youn; Kim, Kwang-Pyo; Ryu, Sangryeol; Kang, Dong-Hyun

2014-02-17

This study evaluated the efficacy of gamma irradiation to inactivate foodborne pathogens in black pepper (Piper nigrum) and red pepper (dried Capsicum annuum). Black pepper and red pepper inoculated with Escherichia coli O157:H7 and Salmonella Typhimurium were subjected to gamma irradiation in the range of 0, 1, 2, 3 and 5 kGy, and color change was evaluated after treatment. Pathogen populations decreased with increasing treatment doses. A gamma irradiation dose of 5 kGy decreased E. coli O157:H7 and S. Typhimurium populations >4.4 to >5.2 log CFU/g in black pepper without causing color change. Similarly, 5 kGy of gamma irradiation yielded reduction of 3.8 to >5.2 log CFU/g for E. coli O157:H7 and S. Typhimurium in red pepper. During gamma irradiation treatment, L*, a* and b* values of red pepper were not significantly changed except for 297 μm to 420 μm size red pepper treated with 5 kGy of gamma irradiation. Based on the D-value of pathogens in black pepper and red pepper, S. Typhimurium showed more resistant to gamma irradiation than did E. coli O157:H7. These results show that gamma irradiation has potential as a non-thermal process for inactivating foodborne pathogens in spices with minimal color changes. Copyright © 2013 Elsevier B.V. All rights reserved.

Lactobacillus rhamnosus GG Suppresses Meningitic E. coli K1 Penetration across Human Intestinal Epithelial Cells In Vitro and Protects Neonatal Rats against Experimental Hematogenous Meningitis

PubMed Central

Huang, Sheng-He; He, Lina; Zhou, Yanhong; Wu, Chun-Hua; Jong, Ambrose

2009-01-01

The purpose of this study was to examine prophylactic efficacy of probiotics in neonatal sepsis and meningitis caused by E. coli K1. The potential inhibitory effect of Lactobacillus rhamnosus GG (LGG) on meningitic E. coli K1 infection was examined by using (i) in vitro inhibition assays with E44 (a CSF isolate from a newborn baby with E. coli meningitis), and (ii) the neonatal rat model of E. coli sepsis and meningitis. The in vitro studies demonstrated that LGG blocked E44 adhesion, invasion, and transcytosis in a dose-dependent manner. A significant reduction in the levels of pathogen colonization, E. coli bacteremia, and meningitis was observed in the LGG-treated neonatal rats, as assessed by viable cultures, compared to the levels in the control group. In conclusion, probiotic LGG strongly suppresses meningitic E. coli pathogens in vitro and in vivo. The results support the use of probiotic strains such as LGG for prophylaxis of neonatal sepsis and meningitis. PMID:20016677

In vitro effect of vaginal lactobacilli on the growth and adhesion abilities of uropathogenic Escherichia coli.

PubMed

Leccese Terraf, María Cecilia; Juarez Tomás, María Silvina; Rault, Lucie; Le Loir, Yves; Even, Sergine; Nader-Macías, María Elena Fátima

2017-07-01

Escherichia coli is one of the main causes of uncomplicated urinary tract infections and responsible of vaginal infections. Lactobacilli can inhibit this pathogen by the production of antimicrobial substances as organic acids, hydrogen peroxide and/or bacteriocins. The aim of this work was to study the effects of beneficial vaginal lactobacilli on E. coli through in vitro experiments. The inhibitory activity of three vaginal Lactobacillus strains against E. coli was assessed using the agar plate diffusion. Moreover, the effect of Lactobacillus reuteri CRL (Centro de Referencia para Lactobacilos Culture Collection) 1324 on the adhesion and internalization capabilities of E. coli was studied on HeLa cells. Two Lactobacillus strains inhibited the growth of the pathogens by production of organic acids. L. reuteri CRL 1324 reduced the adhesion and internalization of E. coli 275 into HeLa cells. The results obtained suggest that L. reuteri CRL 1324 can be considered as a probiotic candidate for further in vivo studies for the prevention or treatment of urinary tract infections caused by E. coli.

Lactobacillus rhamnosus GG Suppresses Meningitic E. coli K1 Penetration across Human Intestinal Epithelial Cells In Vitro and Protects Neonatal Rats against Experimental Hematogenous Meningitis.

PubMed

Huang, Sheng-He; He, Lina; Zhou, Yanhong; Wu, Chun-Hua; Jong, Ambrose

2009-01-01

The purpose of this study was to examine prophylactic efficacy of probiotics in neonatal sepsis and meningitis caused by E. coli K1. The potential inhibitory effect of Lactobacillus rhamnosus GG (LGG) on meningitic E. coli K1 infection was examined by using (i) in vitro inhibition assays with E44 (a CSF isolate from a newborn baby with E. coli meningitis), and (ii) the neonatal rat model of E. coli sepsis and meningitis. The in vitro studies demonstrated that LGG blocked E44 adhesion, invasion, and transcytosis in a dose-dependent manner. A significant reduction in the levels of pathogen colonization, E. coli bacteremia, and meningitis was observed in the LGG-treated neonatal rats, as assessed by viable cultures, compared to the levels in the control group. In conclusion, probiotic LGG strongly suppresses meningitic E. coli pathogens in vitro and in vivo. The results support the use of probiotic strains such as LGG for prophylaxis of neonatal sepsis and meningitis.

Evidence for Transfer of CMY-2 AmpC β-Lactamase Plasmids between Escherichia coli and Salmonella Isolates from Food Animals and Humans

PubMed Central

Winokur, P. L.; Vonstein, D. L.; Hoffman, L. J.; Uhlenhopp, E. K.; Doern, G. V.

2001-01-01

Escherichia coli is an important pathogen that shows increasing antimicrobial resistance in isolates from both animals and humans. Our laboratory recently described Salmonella isolates from food animals and humans that expressed an identical plasmid-mediated, AmpC-like β-lactamase, CMY-2. In the present study, 59 of 377 E. coli isolates from cattle and swine (15.6%) and 6 of 1,017 (0.6%) isolates of human E. coli from the same geographic region were resistant to both cephamycins and extended-spectrum cephalosporins. An ampC gene could be amplified with CMY-2 primers in 94.8% of animal and 33% of human isolates. Molecular epidemiological studies of chromosomal DNA revealed little clonal relatedness among the animal and human E. coli isolates harboring the CMY-2 gene. The ampC genes from 10 animal and human E. coli isolates were sequenced, and all carried an identical CMY-2 gene. Additionally, all were able to transfer a plasmid containing the CMY-2 gene to a laboratory strain of E. coli. CMY-2 plasmids demonstrated two different plasmid patterns that each showed strong similarities to previously described Salmonella CMY-2 plasmids. Additionally, Southern blot analyses using a CMY-2 probe demonstrated conserved fragments among many of the CMY-2 plasmids identified in Salmonella and E. coli isolates from food animals and humans. These data demonstrate that common plasmids have been transferred between animal-associated Salmonella and E. coli, and identical CMY-2 genes carried by similar plasmids have been identified in humans, suggesting that the CMY-2 plasmid has undergone transfer between different bacterial species and may have been transmitted between food animals and humans. PMID:11557460

Enteric Pathogen-Plant Interactions: Molecular Connections Leading to Colonization and Growth and Implications for Food Safety

PubMed Central

Martínez-Vaz, Betsy M.; Fink, Ryan C.; Diez-Gonzalez, Francisco; Sadowsky, Michael J.

2014-01-01

Leafy green vegetables have been identified as a source of foodborne illnesses worldwide over the past decade. Human enteric pathogens, such as Escherichia coli O157:H7 and Salmonella, have been implicated in numerous food poisoning outbreaks associated with the consumption of fresh produce. An understanding of the mechanisms responsible for the establishment of pathogenic bacteria in or on vegetable plants is critical for understanding and ameliorating this problem as well as ensuring the safety of our food supply. While previous studies have described the growth and survival of enteric pathogens in the environment and also the risk factors associated with the contamination of vegetables, the molecular events involved in the colonization of fresh produce by enteric pathogens are just beginning to be elucidated. This review summarizes recent findings on the interactions of several bacterial pathogens with leafy green vegetables. Changes in gene expression linked to the bacterial attachment and colonization of plant structures are discussed in light of their relevance to plant-microbe interactions. We propose a mechanism for the establishment and association of enteric pathogens with plants and discuss potential strategies to address the problem of foodborne illness linked to the consumption of leafy green vegetables. PMID:24859308

Fecal Leukocytes in Children Infected with Diarrheagenic Escherichia coli▿

PubMed Central

Mercado, Erik H.; Ochoa, Theresa J.; Ecker, Lucie; Cabello, Martin; Durand, David; Barletta, Francesca; Molina, Margarita; Gil, Ana I.; Huicho, Luis; Lanata, Claudio F.; Cleary, Thomas G.

2011-01-01

The purpose of this study was to determine the presence and quantity of fecal leukocytes in children infected with diarrheagenic Escherichia coli and to compare these levels between diarrhea and control cases. We analyzed 1,474 stool samples from 935 diarrhea episodes and 539 from healthy controls of a cohort study of children younger than 2 years of age in Lima, Peru. Stools were analyzed for common enteric pathogens, and diarrheagenic E. coli isolates were studied by a multiplex real-time PCR. Stool smears were stained with methylene blue and read by a blinded observer to determine the number of polymorphonuclear leukocytes per high-power field (L/hpf). Fecal leukocytes at >10 L/hpf were present in 11.8% (110/935) of all diarrheal episodes versus 1.1% (6/539) in controls (P < 0.001). Among stool samples with diarrheagenic E. coli as the only pathogen isolated (excluding coinfection), fecal leukocytes at >10 L/hpf were present in 8.5% (18/212) of diarrhea versus 1.3% (2/157) of control samples (P < 0.01). Ninety-five percent of 99 diarrheagenic E. coli diarrhea samples were positive for fecal lactoferrin. Adjusting for the presence of blood in stools, age, sex, undernutrition, and breastfeeding, enterotoxigenic E. coli (ETEC) isolation as a single pathogen, excluding coinfections, was highly associated with the presence of fecal leukocytes (>10 L/hpf) with an odds ratio (OR) of 4.1 (95% confidence interval [CI], 1.08 to 15.51; P < 0.05). Although diarrheagenic E. coli was isolated with similar frequencies in diarrhea and control samples, clearly it was associated with a more inflammatory response during symptomatic infection; however, in general, these pathogens elicited a mild inflammatory response. PMID:21325554

Antimicrobial resistance profiles of common mastitis pathogens on Canadian dairy farms.

PubMed

Saini, V; McClure, J T; Léger, D; Keefe, G P; Scholl, D T; Morck, D W; Barkema, H W

2012-08-01

Monitoring of antimicrobial resistance (AMR) in bacteria has clinical and public health significance. The present study determined prevalence of AMR in common mastitis pathogens Staphylococcus aureus, including methicillin-resistant Staph. aureus (MRSA; n=1,810), Escherichia coli (n=394), and Klebsiella species (n=139), including extended-spectrum β-lactamase (ESBL)-producing E. coli and Klebsiella species, isolated from milk samples on 89 dairy farms in 6 Canadian provinces. Minimum inhibitory concentrations (MIC) were determined using the Sensititer bovine mastitis plate (Trek Diagnostic Systems Inc., Cleveland, OH) and a National Antimicrobial Resistance Monitoring System gram-negative panel containing antimicrobials commonly used for mastitis treatment and control. Denim blue chromogenic agar and real-time PCR were used to screen and confirm MRSA, respectively. Resistance proportion estimates ranged from 0% for cephalothin and oxacillin to 8.8% for penicillin in Staph. aureus isolates, and 15% of the resistant Staph. aureus isolates were multidrug resistant. One MRSA isolate was confirmed (prevalence: 0.05%). Resistance proportion estimates ranged from 0% for ceftriaxone and ciprofloxacin to 14.8% for tetracycline in E. coli, and 0% for amikacin, ceftiofur, ciprofloxacin, and nalidixic acid to 18.6% for tetracycline in Klebsiella species isolates. Further, 62.8 and 55% of the resistant E. coli and Klebsiella species isolates were multidrug resistant, respectively. Resistance to >5 and >2 antimicrobials was most common in E. coli and Klebsiella species isolates, respectively, and no ESBL producers were found. Prevalence of AMR in bovine mastitis pathogens was low. Most gram-negative udder pathogens were multidrug resistant; MRSA was rarely found, and ESBL E. coli and Klebsiella species isolates were absent in Canadian milk samples. Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Soil Conditions That Can Alter Natural Suppression of Escherichia coli O157:H7 in Ohio Specialty Crop Soils

PubMed Central

Williams, Michele L.; LeJeune, Jeffrey T.

2015-01-01

Food-borne pathogen persistence in soil fundamentally affects the production of safe vegetables and small fruits. Interventions that reduce pathogen survival in soil would have positive impacts on food safety by minimizing preharvest contamination entering the food chain. Laboratory-controlled studies determined the effects of soil pH, moisture content, and soil organic matter (SOM) on the survivability of this pathogen through the creation of single-parameter gradients. Longitudinal field-based studies were conducted in Ohio to quantify the extent to which field soils suppressed Escherichia coli O157:H7 survival. In all experiments, heat-sensitive microorganisms were responsible for the suppression of E. coli O157 in soil regardless of the chemical composition of the soil. In laboratory-based studies, soil pH and moisture content were primary drivers of E. coli O157 survival, with increases in pH after 48 h (P = 0.02) and decreases in moisture content after 48 h (P = 0.007) significantly increasing the log reduction of E. coli O157 numbers. In field-based experiments, E. coli O157 counts from both heated and unheated samples were sensitive to both season (P = 0.004 for heated samples and P = 0.001 for unheated samples) and region (P = 0.002 for heated samples and P = 0.001 for unheated samples). SOM was observed to be a more significant driver of pathogen suppression than the other two factors after 48 h at both planting and harvest (P = 0.002 at planting and P = 0.058 at harvest). This research reinforces the need for both laboratory-controlled experiments and longitudinal field-based experiments to unravel the complex relationships controlling the survival of introduced organisms in soil. PMID:25934621

Candida krusei isolated from fruit juices ultrafiltration membranes promotes colonization of Escherichia coli O157:H7 and Salmonella enterica on stainless steel surfaces.

PubMed

Tarifa, María Clara; Lozano, Jorge Enrique; Brugnoni, Lorena Inés

2017-02-01

To clarify the interactions between a common food spoilage yeast and two pathogenic bacteria involved in outbreaks associated with fruit juices, the present paper studies the effect of the interplay of Candida krusei, collected from UF membranes, with Escherichia coli O157:H7 and Salmonella enterica in the overall process of adhesion and colonization of abiotic surfaces. Two different cases were tested: a) co-adhesion by pathogenic bacteria and yeasts, and b) incorporation of bacteria to pre-adhered C. krusei cells. Cultures were made on stainless steel at 25°C using apple juice as culture medium. After 24 h of co-adhesion with C. krusei, both E. coli O157:H7 and S. enterica increased their counts 1.05 and 1.11 log CFU cm 2 , respectively. Similar increases were obtained when incorporating bacteria to pre-adhered cells of Candida. Nevertheless C. krusei counts decreased in both experimental conditions, in a) 0.40 log CFU cm 2 and 0.55 log CFU cm 2 when exposed to E. coli O157:H7 and S. enterica and in b) 0.18 and 0.68 log CFU cm 2 , respectively. This suggests that C. krusei, E. coli O157:H7, and S. enterica have a complex relationship involving physical and chemical interactions on food contact surfaces. This study supports the possibility that pathogen interactions with members of spoilage microbiota, such as C. krusei, might play an important role for the survival and dissemination of E. coli O157:H7 and Salmonella enterica in food-processing environments. Based on the data obtained from the present study, much more attention should be given to prevent the contamination of these pathogens in acidic drinks.

Epidemiology of bacterial pathogens associated with infectious diarrhea in Djibouti.

PubMed Central

Mikhail, I A; Fox, E; Haberberger, R L; Ahmed, M H; Abbatte, E A

1990-01-01

During a survey examining the causes of diarrhea in the East African country of Djibouti, 140 bacterial pathogens were recovered from 209 diarrheal and 100 control stools. The following pathogens were isolated at comparable frequencies from both diarrheal and control stools: enteroadherent Escherichia coli (EAEC) (10.6 versus 13%), enterotoxigenic E. coli (ETEC) (11 versus 10%), enteropathogenic E. coli (EPEC) (7.7 versus 12%), Salmonella spp. (2.9 versus 3%), and Campylobacter jejuni-C. coli (3.3 versus 5%). Surprisingly, the EAEC strains isolated did not correspond to well-recognized EPEC serogroups. No Yersinia spp., enteroinvasive E. coli, or enterohemorrhagic E. coli were isolated during the course of this study. Only the following two genera were recovered from diarrheal stools exclusively: Shigella spp. (7.7%) and Aeromonas hydrophila group organisms (3.3%). Shigella flexneri was the most common Shigella species isolated. Patients with Shigella species were of a higher average age than were controls (27 versus 13 years), while subjects with Campylobacter or Salmonella species belonged to younger age groups (2.6 and 1.6 years, respectively). Salmonella cases were more often in females. Shigella diarrhea was associated with fecal blood or mucus and leukocytes. ETEC was not associated with nausea or vomiting. Anorexia, weight loss, and fever were associated with the isolation of Salmonella and Aeromonas species. EAEC, ETEC, EPEC, and Shigella species were resistant to most drugs used for treating diarrhea in Africa, while the antibiotic most active against all bacteria tested was norfloxacin. We conclude that in Djibouti in 1989, Shigella and Aeromonas species must be considered as potential pathogens whenever they are isolated from diarrheal stools and that norfloxacin should be considered the drug of choice in adults for treating severe shigellosis and for diarrhea prophylaxis in travelers. PMID:2351738

Avian-pathogenic Escherichia coli strains are similar to neonatal meningitis E. coli strains and are able to cause meningitis in the rat model of human disease.

PubMed

Tivendale, Kelly A; Logue, Catherine M; Kariyawasam, Subhashinie; Jordan, Dianna; Hussein, Ashraf; Li, Ganwu; Wannemuehler, Yvonne; Nolan, Lisa K

2010-08-01

Escherichia coli strains causing avian colibacillosis and human neonatal meningitis, urinary tract infections, and septicemia are collectively known as extraintestinal pathogenic E. coli (ExPEC). Characterization of ExPEC strains using various typing techniques has shown that they harbor many similarities, despite their isolation from different host species, leading to the hypothesis that ExPEC may have zoonotic potential. The present study examined a subset of ExPEC strains: neonatal meningitis E. coli (NMEC) strains and avian-pathogenic E. coli (APEC) strains belonging to the O18 serogroup. The study found that they were not easily differentiated on the basis of multilocus sequence typing, phylogenetic typing, or carriage of large virulence plasmids. Among the APEC strains examined, one strain was found to be an outlier, based on the results of these typing methods, and demonstrated reduced virulence in murine and avian pathogenicity models. Some of the APEC strains tested in a rat model of human neonatal meningitis were able to cause meningitis, demonstrating APEC's ability to cause disease in mammals, lending support to the hypothesis that APEC strains have zoonotic potential. In addition, some NMEC strains were able to cause avian colisepticemia, providing further support for this hypothesis. However, not all of the NMEC and APEC strains tested were able to cause disease in avian and murine hosts, despite the apparent similarities in their known virulence attributes. Thus, it appears that a subset of NMEC and APEC strains harbors zoonotic potential, while other strains do not, suggesting that unknown mechanisms underlie host specificity in some ExPEC strains.

Behavior of Escherichia coli O157:H7 on damaged leaves of spinach, lettuce, cilantro, and parsley stored at abusive temperatures.

PubMed

Khalil, Rowaida K; Frank, Joseph F

2010-02-01

Recent foodborne illness outbreaks associated with the consumption of leafy green produce indicates a need for additional information on the behavior of pathogenic bacteria on these products. Previous research indicates that pathogen growth and survival is enhanced by leaf damage. The objective of this study was to compare the behavior of Escherichia coli O157:H7 on damaged leaves of baby Romaine lettuce, spinach, cilantro, and parsley stored at three abusive temperatures (8, 12, and 15 degrees C). The damaged portions of leaves were inoculated with approximately 10(5) CFU E. coli O157:H7 per leaf. The pathogen grew on damaged spinach leaves held for 3 days at 8 and 12 degrees C (P < 0.05), with the population increasing by 1.18 and 2.08 log CFU per leaf, respectively. E. coli O157:H7 did not grow on damaged Romaine leaves at 8 or 12 degrees C, but growth was observed after 8 h of storage at 15 degrees C, with an increase of less than 1.0 log. Growth of E. coli O157:H7 on Romaine lettuce held at 8 or 12 degrees C was enhanced when inocula were suspended in 0.05% ascorbic acid, indicating the possibility of inhibition by oxidation reactions associated with tissue damage. Damaged cilantro and Italian parsley leaves held at 8 degrees C for 4 days did not support the growth of E. coli O157:H7. Behavior of the pathogen in leaf extracts differed from behavior on the damaged tissue. This study provides evidence that the damaged portion of a leafy green is a distinct growth niche that elicits different microbial responses in the various types of leafy greens.

Agrobacterium tumefaciens supports DNA replication of diverse geminivirus types.

PubMed

Selth, Luke A; Randles, John W; Rezaian, M Ali

2002-04-10

We have previously shown that the soil-borne plant pathogen Agrobacterium tumefaciens supports the replication of tomato leaf curl geminivirus (Australian isolate) (TLCV) DNA. However, the reproducibility of this observation with other geminiviruses has been questioned. Here, we show that replicative DNA forms of three other geminiviruses also accumulate at varying levels in Agrobacterium. Geminiviral DNA constructs that lacked the ability to replicate in Agrobacterium were rendered replication-competent by changing their configuration so that two copies of the viral ori were present. Furthermore, we report that low-level replication of TLCV DNA can occur in Escherichia coli containing a dimeric TLCV construct in a high copy number plasmid. These findings were reinforced by expression studies using beta-glucuronidase which revealed that all six TLCV promoters are active in Agrobacterium, and two are functional in E. coli.

Structural and Biochemical Characterization of Gentisate 1,2-Dioxygenase from Escherichia coli O157:H7.

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

Adams,M.; Singh, V.; Keller, B.

2006-01-01

Gentisic acid (2,5-dihydroxybenzoic acid) is a key intermediate in aerobic bacterial pathways that are responsible for the metabolism of a large number of aromatic compounds. The critical step of these pathways is the oxygen-dependent reaction catalysed by gentisate 1,2-dioxygenase which opens the aromatic ring of gentisate to form maleylpyruvate. From gentisic acid, the cell derives carbon and energy through the conversion of maleylpyruvate to central metabolites. We have confirmed the annotation of a gentisate 1,2-dioygenase from the pathogenic O157:H7 Escherichia coli strain and present the first structural characterization of this family of enzymes. The identity of the reaction product wasmore » revealed using tandem mass spectroscopy. The operon responsible for the degradation of gentisate in this organism exhibits a high degree of conservation with the gentisate-degrading operons of other pathogenic bacteria, including the Shiga toxin-producing E. coli O103:H2, but does not appear to be present in non-pathogenic strains. The acquisition of the gentisate operon may represent a special adaptation to meet carbon source requirements under conditions of environmental stress and may provide a selective advantage for enterohaemorrhagic E. coli relative to their non-pathogenic counterparts.« less

The effect of interactions between a bacterial strain isolated from drinking water and a pathogen surrogate on biofilms formation diverged under static vs flow conditions.

PubMed

Dai, D; Raskin, L; Xi, C

2017-12-01

Interactions with water bacteria affect the incorporation of pathogens into biofilms and thus pathogen control in drinking water systems. This study was to examine the impact of static vs flow conditions on interactions between a pathogen and a water bacterium on pathogen biofilm formation under laboratory settings. A pathogen surrogate Escherichia coli and a drinking water isolate Stenotrophomonas maltophilia was selected for this study. Biofilm growth was examined under two distinct conditions, in flow cells with continuous medium supply vs in static microtitre plates with batch culture. E. coli biofilm was greatly stimulated (c. 2-1000 times faster) with the presence of S. maltophilia in flow cells, but surprisingly inhibited (c. 65-95% less biomass) in microtitre plates. These divergent effects were explained through various aspects including surface attachment, cellular growth, extracellular signals and autoaggregation. Interactions with the same water bacterium resulted in different effects on E. coli biofilm formation when culture conditions changed from static to flow. This study highlights the complexity of species interactions on biofilm formation and suggests that environmental conditions such as the flow regime can be taken into consideration for the management of microbial contamination in drinking water systems. © 2017 The Society for Applied Microbiology.

Detection of Waterborne Protozoa, Viruses, and Bacteria in Groundwater and Other Water Samples in the Kathmandu Valley, Nepal

NASA Astrophysics Data System (ADS)

Haramoto, E.

2018-03-01

In this study, the prevalence of various waterborne pathogens in water samples collected in the Kathmandu Valley, Nepal, and the applicability of Escherichia coli as an indicator of pathogen contamination in groundwater were assessed. Fifty-three water samples, including shallow groundwater and river water, were analyzed to examine the presence of protozoan (oo)cysts via fluorescence microscopy and that of viral and bacterial genomes via quantitative PCR. At least one of the seven types of pathogens tested (i.e., Cryptosporidium, Giardia, human adenoviruses, noroviruses of genogroups I and II, group A rotaviruses, and Vibrio cholerae) was detected in 68% (15/22) of the shallow dug well water samples; groundwater in the shallow dug wells was more contaminated compared with that in shallow tube wells (8/15, 53%). River water and sewage samples were contaminated with extremely high concentrations of multiple pathogens, whereas a tap water sample supplied by a water tanker tested positive for human adenoviruses and V. cholerae. The detection of host-specific Bacteroidales genetic markers revealed the effects of human and animal feces on groundwater contamination. The tested pathogens were sometimes detected even in E. coli-negative groundwater samples, indicative of the limitations of using E. coli as an indicator for waterborne pathogens in groundwater.

Geographic setting influences Great Lakes beach microbiological water quality

USGS Publications Warehouse

Haack, Sheridan K.; Fogarty, Lisa R.; Stelzer, Erin A.; Fuller, Lori M.; Brennan, Angela K.; Isaacs, Natasha M.; Johnson, Heather E.

2013-01-01

Understanding of factors that influence Escherichia coli (EC) and enterococci (ENT) concentrations, pathogen occurrence, and microbial sources at Great Lakes beaches comes largely from individual beach studies. Using 12 representative beaches, we tested enrichment cultures from 273 beach water and 22 tributary samples for EC, ENT, and genes indicating the bacterial pathogens Shiga-toxin producing E. coli (STEC), Shigella spp., Salmonella spp, Campylobacter jejuni/coli, and methicillin-resistant Staphylococcus aureus, and 108–145 samples for Bacteroides human, ruminant, and gull source-marker genes. EC/ENT temporal patterns, general Bacteroides concentration, and pathogen types and occurrence were regionally consistent (up to 40 km), but beach catchment variables (drains/creeks, impervious surface, urban land cover) influenced exceedances of EC/ENT standards and detections of Salmonella and STEC. Pathogen detections were more numerous when the EC/ENT Beach Action Value (but not when the Geometric Mean and Statistical Threshold Value) was exceeded. EC, ENT, and pathogens were not necessarily influenced by the same variables. Multiple Bacteroides sources, varying by date, occurred at every beach. Study of multiple beaches in different geographic settings provided new insights on the contrasting influences of regional and local variables, and a broader-scale perspective, on significance of EC/ENT exceedances, bacterial sources, and pathogen occurrence.

THE WIDESPREAD OCCURRENCE OF THE ENTEROHEMOLYSIN GENE EHLYA AMONG ENVIRONMENTAL STRAINS OF ESCHERICHIA COLI

EPA Science Inventory

The putative virulence factor enterohemolysin, encoded for by the ehlyA gene, has been closely associated with the pathogenic enterohemorrhagic Escherichia coli (EHEC) group. E. coli isolates from effluents from seven geographically dispersed municipal ...

Pathogen profile of clinical mastitis in Irish milk-recording herds reveals a complex aetiology.

PubMed

Keane, O M; Budd, K E; Flynn, J; McCoy, F

2013-07-06

Effective mastitis control requires knowledge of the predominant pathogen challenges on the farm. In order to quantify this challenge, the aetiological agents associated with clinical mastitis in 30 milk-recording dairy herds in Ireland over a complete lactation were investigated. Standard bacteriology was performed on 630 pretreatment quarter milk samples, of which 56 per cent were culture-positive, 42 per cent culture-negative and 2 per cent contaminated. Two micro-organisms were isolated from almost 5 per cent of the culture-positive samples. The bacteria isolated were Staphylococcus aureus (23 per cent), Streptococcus uberis (17 per cent), Escherichia coli (9 per cent), Streptococcus species (6 per cent), coagulase-negative Staphylococci (4 per cent) and other species (1 per cent). A wide variety of bacterial species were associated with clinical mastitis, with S aureus the most prevalent pathogen overall, followed by S uberis. However, the bacterial challenges varied widely from farm to farm. In comparison with previous reports, in the present study, the contagious pathogens S aureus and Streptococcus agalactiae were less commonly associated with clinical mastitis, whereas, the environmental pathogens S uberis and E coli were found more commonly associated with clinical mastitis. While S aureus remains the pathogen most commonly associated with intramammary infection in these herds, environmental pathogens, such as S uberis and E coli also present a considerable challenge.

Genomic Microbial Epidemiology Is Needed to Comprehend the Global Problem of Antibiotic Resistance and to Improve Pathogen Diagnosis.

PubMed

Wyrsch, Ethan R; Roy Chowdhury, Piklu; Chapman, Toni A; Charles, Ian G; Hammond, Jeffrey M; Djordjevic, Steven P

2016-01-01

Contamination of waste effluent from hospitals and intensive food animal production with antimicrobial residues is an immense global problem. Antimicrobial residues exert selection pressures that influence the acquisition of antimicrobial resistance and virulence genes in diverse microbial populations. Despite these concerns there is only a limited understanding of how antimicrobial residues contribute to the global problem of antimicrobial resistance. Furthermore, rapid detection of emerging bacterial pathogens and strains with resistance to more than one antibiotic class remains a challenge. A comprehensive, sequence-based genomic epidemiological surveillance model that captures essential microbial metadata is needed, both to improve surveillance for antimicrobial resistance and to monitor pathogen evolution. Escherichia coli is an important pathogen causing both intestinal [intestinal pathogenic E. coli (IPEC)] and extraintestinal [extraintestinal pathogenic E. coli (ExPEC)] disease in humans and food animals. ExPEC are the most frequently isolated Gram negative pathogen affecting human health, linked to food production practices and are often resistant to multiple antibiotics. Cattle are a known reservoir of IPEC but they are not recognized as a source of ExPEC that impact human or animal health. In contrast, poultry are a recognized source of multiple antibiotic resistant ExPEC, while swine have received comparatively less attention in this regard. Here, we review what is known about ExPEC in swine and how pig production contributes to the problem of antibiotic resistance.

Different cellular origins and functions of extracellular proteins from Escherichia coli O157:H7 and O104:H4 as determined by comparative proteomic analysis

USDA-ARS?s Scientific Manuscript database

Escherichia coli is a diverse species of bacteria, including several pathotypes that cause a variety of diseases in humans. Enterohemorrhagic E. coli (EHEC) and recently emerged shigatoxingenic enteroaggregative E. coli (EAEC) produce Shigatoxins and are major foodborne pathogens that can cause hem...