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Sample records for citrus pathogen alternaria

  1. Worldwide Phylogeography of the Citrus Brown Spot Pathogen, Alternaria alternata.

    PubMed

    Peever, T L; Ibañez, A; Akimitsu, K; Timmer, L W

    2002-07-01

    ABSTRACT Sixty-five isolates of Alternaria alternata were sampled from brown spot lesions on tangerines and mandarins (Citrus reticulata) and tangerine x grapefruit (C. reticulata x C. paradisi) hybrids in the United States, Colombia, Australia, Turkey, South Africa, and Israel to investigate the worldwide phylogeography of the fungus. Genetic variation was scored at 15 putative random amplified polymorphic DNA (RAPD) loci and 465 bp of an endo-polygalacturonase (endo-PG) gene was sequenced for each isolate. Cluster analysis of RAPD genotypes revealed significant differentiation between United State and Colombia isolates and Turkey, South Africa, Israel, and Australia isolates. Sequencing of endo-PG revealed 21 variable sites when the outgroup A. gaisen (AK-toxin-producing pathogen of Japanese pear) was included and 13 variable sites among the sampled isolates. Nucleotide substitutions at 10 of 13 variable sites represented silent mutations when endo-PG was translated in frame. Eight distinct endo-PG haplotypes were found among the sampled isolates and estimation of a phylogeny with endo-PG sequence data revealed three clades, each with strong bootstrap support. The most basal clade (clade 1) was inferred based on its similarity to the outgroup A. gaisen and consisted exclusively of pathogenic isolates from the United States and Colombia. Clade 2 consisted of pathogenic and nonpathogenic isolates from the United States, Australia, South Africa, and Israel and clade 3 contained pathogenic and nonpathogenic isolates from Australia, South Africa, Israel, and Turkey. Quantitative estimates of virulence (disease incidence) were obtained for isolates from the United States, Colombia, South Africa, Israel, and Turkey by spray inoculating detached citrus leaves and counting the number of lesions 24 h after inoculation. Large differences in virulence were detected among isolates within each location and isolates from the United States were significantly more virulent than

  2. Overexpression of citrus polygalacturonase-inhibiting protein in citrus black rot pathogen Alternaria citri.

    PubMed

    Katoh, Hiroshi; Nalumpang, Sarunya; Yamamoto, Hiroyuki; Akimitsu, Kazuya

    2007-05-01

    The rough lemon (Citrus jambhiri) gene encoding polygalacturonase-inhibiting protein (RlemPGIPA) was overexpressed in the pathogenic fungus Alternaria citri. The overexpression mutant AcOPI6 retained the ability to utilize pectin as a sole carbon source, and the overexpression of polygalacturonase-inhibiting protein did not have any effect on the growth of AcOPI6 in potato dextrose and pectin medium. The pathogenicity of AcOPI6 to cause a black rot symptom in citrus fruits was also unchanged. Polygalacturonase-inhibiting protein was secreted together with endopolygalacturonase into culture filtrates of AcOPI6, and oligogalacturonides were digested from polygalacturonic acid by both proteins in the culture filtrates. The reaction mixture containing oligogalacturonides possessed activity for induction of defense-related gene, RlemLOX, in rough lemon leaves.

  3. A Virulence-Reducing Mutation in the Postharvest Citrus Pathogen Alternaria citri.

    PubMed

    Katoh, H; Isshiki, A; Masunaka, A; Yamamoto, H; Akimitsu, K

    2006-09-01

    ABSTRACT Alternaria citri causes Alternaria black rot, a postharvest fruit disease, on a broad range of citrus cultivars. We previously described that an endopolygalacturonase minus mutant of A. citri caused significantly less black rot in citrus fruit. To search for other essential factors causing symptoms in addition to endopolygalacturonase, a random mutation analysis of pathogenicity was performed using restriction enzyme-mediated integration. Three isolates among 1,694 transformants of A. citri had a loss in pathogenicity in a citrus peel assay, and one of these three mutants was a histidine auxotroph. Gene AcIGPD that encodes imidazole glycerol phosphate dehydratase, the sixth enzyme in the histidine biosynthetic pathway, was cloned, and the mutant containing the disrupted target gene, AcIGPD, caused less black rot.

  4. A nonribosomal peptide synthetase mediates siderophore production and virulence in the citrus fungal pathogen Alternaria alternata.

    PubMed

    Chen, Li-Hung; Lin, Ching-Hsuan; Chung, Kuang-Ren

    2013-06-01

    Alternaria species produce and excrete dimethyl coprogen siderophores to acquire iron. The Alternaria alternata gene AaNPS6, encoding a polypeptide analogous to fungal nonribosomal peptide synthetases, was found to be required for the production of siderophores and virulence on citrus. Siderophores purified from culture filtrates of the wild-type strain did not induce any phytotoxicity on the leaves of citrus. Fungal strains lacking AaNPS6 produced little or no detectable extracellular siderophores and displayed an increased sensitivity to H₂O₂, superoxide-generating compounds (KO₂ and menadione) and iron depletion. Δnps6 mutants were also defective for the production of melanin and conidia. The introduction of a wild-type AaNPS6 under the control of its endogenous promoter to a Δnps6 null mutant at least partially restored siderophore production and virulence to citrus, demonstrating a functional link between iron uptake and fungal pathogenesis. Elevated sensitivity to H₂O₂, seen for the Δnps6 null strain could be relieved by exogenous application of ferric iron. The expression of the AaNPS6 gene was highly up-regulated under low-iron conditions and apparently controlled by the redox-responsive yeast transcriptional regulator YAP1. Hence, the maintenance of iron homeostasis via siderophore-mediated iron uptake also plays an important role in resistance to toxic reactive oxygen species (ROS). Our results demonstrate further the critical role of ROS detoxification for the pathogenicity of A. alternata in citrus.

  5. Signatures of recombination in clonal lineages of the citrus brown spot pathogen, Alternaria alternata sensu lato.

    PubMed

    Stewart, Jane E; Thomas, Kalyn A; Lawrence, Christopher B; Dang, Ha; Pryor, Barry M; Timmer, L M Pete; Peever, Tobin L

    2013-07-01

    Most Alternaria spp. are considered asexual but recent molecular evolution analyses of Alternaria mating-type genes show that the mating locus is under strong purifying selection, indicating a possible role in sexual reproduction. The objective of this study was to determine the mode of reproduction of an Alternaria alternata sensu lato population causing citrus brown spot in central Florida. Mating type of each isolate was determined, and isolates were sequenced at six putatively unlinked loci. Three genetically distinct subpopulations (SH1, SH4A, and SH4B) were identified using network and Bayesian population structure analyses. Results demonstrate that most subpopulations of A. alternata associated with citrus are clonal but some have the ability to extensively recombine through a cryptic sexual cycle or parasexual cycle. Although isolates were sampled in close physical proximity (≈2,500-m² area), we were able to reject a random mating model using multilocus gametic disequilibrium tests for two subpopulations, SH1 and SH4B, suggesting that these subpopulations were predominantly asexual. However, three recombination events were identified in SH1 and SH4B and localized to individuals of opposite mating type, possibly indicating meiotic recombination. In contrast, in the third subpopulation (SH4A), where only one mating type was present, extensive reticulation was evident in network analyses, and multilocus gametic disequilibrium tests were consistent with recombination. Recombination among isolates of the same mating type suggests that a nonmeiotic mechanism of recombination such as the parasexual cycle may be operating in this subpopulation. The level of gene flow detected among subpopulations does not appear to be sufficient to prevent differentiation, and perhaps future speciation, of these A. alternata subpopulations.

  6. Roles for SKN7 response regulator in stress resistance, conidiation and virulence in the citrus pathogen Alternaria alternata.

    PubMed

    Chen, Li-Hung; Lin, Ching-Hsuan; Chung, Kuang-Ren

    2012-10-01

    "Two-component" histidine kinase (HSK1) is the primary regulator of resistance to sugar osmotic stress and sensitivity to dicarboximide or phenylpyrrole fungicides in the citrus fungal pathogen Alternaria alternata. On the other hand, the mitogen-activated protein kinase HOG1 confers resistance solely to salts and oxidative stress. We report here independent and shared functions of the SKN7-mediated signaling pathway with HSK1 and HOG1. SKN7, a putative transcription downstream regulator of HSK1, is primarily required for cellular resistance to oxidative and sugar-induced osmotic stress. SKN7, perhaps acting in parallel with HOG1, is required for resistance to H(2)O(2), tert-butyl hydroperoxide, and cumyl peroxide, but not to the superoxide-generating compounds - menadione, potassium superoxide, and diamide. Because of phenotypic commonalities, SKN7 is likely involved in resistance to sugar-induced osmotic stress via the HSK1 signaling pathway. However, mutants lacking SKN7 displayed wild-type sensitivity to NaCl and KCl salts. SKN7 is constitutively localized in the nucleus regardless of H(2)O(2) treatment. When compared to the wild type, skn7 mutants exhibited lower catalase, peroxidase, and superoxide dismutase activities and induced significantly fewer necrotic lesions on the susceptible citrus cultivar. The skn7 mutant exhibited fungicide resistance at levels between the hsk1 and the hog1 mutant strains. Skn7/hog1 double mutants exhibited fungicide resistance, similar to the strain with a single AaHSK1 gene mutation. Moreover, the A. alternata SKN7 plays a role in conidia formation. Conidia produced by the skn7 mutant are smaller and have fewer transverse septae than those produced by wild type. All altered phenotypes in the mutant were restored by introducing and expressing a wild-type copy of SKN7 under control of the endogenous promoter. Copyright © 2012 Elsevier Inc. All rights reserved.

  7. Molecular systematics of citrus-associated Alternaria species.

    PubMed

    Peever, T L; Su, G; Carpenter-Boggs, L; Timmer, L W

    2004-01-01

    The causal agents of Alternaria brown spot of tangerines and tangerine hybrids, Alternaria leaf spot of rough lemon and Alternaria black rot of citrus historically have been referred to as Alternaria citri or A. alternata. Ten species of Alternaria recently were described among a set of isolates from leaf lesions on rough lemon (Citrus jambhiri) and tangelo (C. paradisi × C. reticulata), and none of these isolates was considered representative of A. alternata or A. citri. To test the hypothesis that these newly described morphological species are congruent with phylogenetic species, selected Alternaria brown spot and leaf spot isolates, citrus black rot isolates (post-harvest pathogens), isolates associated with healthy citrus tissue and reference species of Alternaria from noncitrus hosts were scored for sequence variation at five genomic regions and used to estimate phylogenies. These data included 432 bp from the 5' end of the mitochondrial ribosomal large subunit (mtLSU), 365 bp from the 5' end of the beta-tubulin gene, 464 bp of an endopolygalacturonase gene (endoPG) and 559 and 571 bp, respectively, of two anonymous genomic regions (OPA1-3 and OPA2-1). The mtLSU and beta-tubulin phylogenies clearly differentiated A. limicola, a large-spored species causing leaf spot of Mexican lime, from the small-spored isolates associated with citrus but were insufficiently variable to resolve evolutionary relationships among the small-spored isolates from citrus and other hosts. Sequence analysis of translation elongation factor alpha, calmodulin, actin, chitin synthase and 1, 3, 8-trihydroxynaphthalene reductase genes similarly failed to uncover significant variation among the small-spored isolates. Phylogenies estimated independently from endoPG, OPA1-3 and OPA2-1 data were congruent, and analysis of the combined data from these regions revealed nine clades, eight of which contained small-spored, citrus-associated isolates. Lineages inferred from analysis of the

  8. Molecular, ecological and evolutionary approaches to understanding Alternaria diseases of citrus.

    PubMed

    Akimitsu, Kazuya; Peever, Tobin L; Timmer, L W

    2003-11-01

    SUMMARY Alternaria fungi cause four different diseases of citrus: Alternaria brown spot of tangerines, Alternaria leaf spot of rough lemon, Alternaria black rot of several citrus fruits and Mancha foliar of Mexican lime. The first three diseases are caused by the small-spored species, Alternaria alternata and the causal agents can only be differentiated using pathogenicity tests, toxin assays or genetic markers. Mancha foliar is caused by the morphologically distinct, large-spored species A. limicola. Substantial progress has been made in understanding the biology, ecology, population biology, systematics, molecular biology and biochemistry of the interactions between these pathogens and citrus. Epidemiological studies have focused on brown spot of tangerines and their hybrids and have contributed to the development of a model of disease development which has improved control and reduced fungicide use. Studies of the population genetics, host specificity and ecology of A. alternata from different ecological niches on citrus have revealed host specific forms of the pathogen which cause disease on different citrus species, the existence of three phylogenetic lineages of the fungus which cause brown spot world-wide, and closely related non-pathogenic isolates which colonize healthy citrus tissue. The role of host-specific toxins in Alternaria diseases of citrus has been extensively studied for over 20 years, and these pathosystems have become model systems for host-pathogen interactions. Recent molecular research has started to unravel the genetic basis of toxin production and the host susceptibility to toxin, and the role of extracellular, degradative enzymes in disease.

  9. Characterization of Citrus-Associated Alternaria Species in Mediterranean Areas

    PubMed Central

    Garganese, Francesca; Schena, Leonardo; Siciliano, Ilenia; Prigigallo, Maria Isabella; Spadaro, Davide; De Grassi, Anna; Ippolito, Antonio

    2016-01-01

    Alternaria brown spot is one of the most important diseases of tangerines and their hybrids worldwide. Recently, outbreaks in Mediterranean areas related to susceptible cultivars, refocused attention on the disease. Twenty representatives were selected from a collection of 180 isolates of Alternaria spp. from citrus leaves and fruit. They were characterized along with reference strains of Alternaria spp. Micro- and macroscopic characteristics separated most Alternaria isolates into six morphotypes referable to A. alternata (5) and A. arborescens (1). Phylogenetic analyses, based on endopolygalacturonase (endopg) and internal transcribed spacer (ITS), confirmed this finding. Moreover, a five-gene phylogeny including two anonymous genomics regions (OPA 1–3 and OPA 2–1), and the beta-tubulin gene (ß-tub), produced a further clustering of A. alternata into three clades. This analysis suggested the existence of intra-species molecular variability. Investigated isolates showed different levels of virulence on leaves and fruit. In particular, the pathogenicity on fruit seemed to be correlated with the tissue of isolation and the clade. The toxigenic behavior of Alternaria isolates was also investigated, with tenuazonic acid (TeA) being the most abundant mycotoxin (0.2–20 mg/L). Isolates also synthesized the mycotoxins alternariol (AOH), its derivate alternariol monomethyl ether (AME), and altenuene (ALT), although to a lesser extent. AME production significantly varied among the six morphotypes. The expression of pksJ/pksH, biosynthetic genes of AOH/AME, was not correlated with actual toxin production, but it was significantly different between the two genotypes and among the four clades. Finally, ten isolates proved to express the biosynthetic genes of ACTT1 phytotoxin, and thus to be included in the Alternaria pathotype tangerine. A significant correlation between pathogenicity on leaves and ACTT1 gene expression was recorded. The latter was significantly

  10. Characterization of Citrus-Associated Alternaria Species in Mediterranean Areas.

    PubMed

    Garganese, Francesca; Schena, Leonardo; Siciliano, Ilenia; Prigigallo, Maria Isabella; Spadaro, Davide; De Grassi, Anna; Ippolito, Antonio; Sanzani, Simona Marianna

    2016-01-01

    Alternaria brown spot is one of the most important diseases of tangerines and their hybrids worldwide. Recently, outbreaks in Mediterranean areas related to susceptible cultivars, refocused attention on the disease. Twenty representatives were selected from a collection of 180 isolates of Alternaria spp. from citrus leaves and fruit. They were characterized along with reference strains of Alternaria spp. Micro- and macroscopic characteristics separated most Alternaria isolates into six morphotypes referable to A. alternata (5) and A. arborescens (1). Phylogenetic analyses, based on endopolygalacturonase (endopg) and internal transcribed spacer (ITS), confirmed this finding. Moreover, a five-gene phylogeny including two anonymous genomics regions (OPA 1-3 and OPA 2-1), and the beta-tubulin gene (ß-tub), produced a further clustering of A. alternata into three clades. This analysis suggested the existence of intra-species molecular variability. Investigated isolates showed different levels of virulence on leaves and fruit. In particular, the pathogenicity on fruit seemed to be correlated with the tissue of isolation and the clade. The toxigenic behavior of Alternaria isolates was also investigated, with tenuazonic acid (TeA) being the most abundant mycotoxin (0.2-20 mg/L). Isolates also synthesized the mycotoxins alternariol (AOH), its derivate alternariol monomethyl ether (AME), and altenuene (ALT), although to a lesser extent. AME production significantly varied among the six morphotypes. The expression of pksJ/pksH, biosynthetic genes of AOH/AME, was not correlated with actual toxin production, but it was significantly different between the two genotypes and among the four clades. Finally, ten isolates proved to express the biosynthetic genes of ACTT1 phytotoxin, and thus to be included in the Alternaria pathotype tangerine. A significant correlation between pathogenicity on leaves and ACTT1 gene expression was recorded. The latter was significantly dependent on

  11. Resistance to oxidative stress via regulating siderophore-mediated iron acquisition by the citrus fungal pathogen Alternaria alternata.

    PubMed

    Chen, Li-Hung; Yang, Siwy Ling; Chung, Kuang-Ren

    2014-05-01

    The ability of the necrotrophic fungus Alternaria alternata to detoxify reactive oxygen species (ROS) is crucial for pathogenesis to citrus. We report regulation of siderophore-mediated iron acquisition and ROS resistance by the NADPH oxidase (NOX), the redox activating yes-associated protein 1 (YAP1) regulator, and the high-osmolarity glycerol 1 (HOG1) mitogen-activated protein kinase (MAPK). The A. alternata nonribosomal peptide synthetase (NPS6) is essential for the biosynthesis of siderophores, contributing to iron uptake under low-iron conditions. Fungal strains impaired for NOX, YAP1, HOG1 or NPS6 all display increased sensitivity to ROS. Exogenous addition of iron at least partially rescues ROS sensitivity seen for NPS6, YAP1, HOG1, and NOX mutants. Importantly, expression of the NPS6 gene and biosynthesis of siderophores are regulated by NOX, YAP1 and HOG1, supporting a functional link among these regulatory pathways. Although iron fully rescues H2O2 sensitivity seen in mutants impaired for the response regulator SKN7, neither expression of NPS6 nor biosynthesis of siderophores is controlled by SKN7. Our results indicate that the acquisition of environmental iron has profound effects on ROS detoxification.

  12. Calcineurin phosphatase and phospholipase C are required for developmental and pathological functions in the citrus fungal pathogen Alternaria alternata.

    PubMed

    Tsai, Hsieh-Chin; Chung, Kuang-Ren

    2014-07-01

    Excessive Ca(2+) or compounds interfering with phosphoinositide cycling have been found to inhibit the growth of the tangerine pathotype of Alternaria alternata, suggesting a crucial role of Ca(2+) homeostasis in this pathotype. The roles of PLC1, a phospholipase C-coding gene and CAL1, a calcineurin phosphatase-coding gene were investigated. Targeted gene disruption showed that both PLC1 and CAL1 were required for vegetative growth, conidial formation and pathogenesis in citrus. Fungal strains lacking PLC1 or CAL1 exhibited extremely slow growth and induced small lesions on calamondin leaves. Δplc1 mutants produced fewer conidia, which germinated at slower rates than wild-type. Δcal1 mutants produced abnormal hyphae and failed to produce any mature conidia, but instead produced highly melanized bulbous hyphae with distinct septae. Fluorescence microscopy using Fluo-3 dye as a Ca(2+) indicator revealed that the Δplc1 mutant hyphae emitted stronger cytosolic fluorescence, and the Δcal1 mutant hyphae emitted less cytosolic fluorescence, than those of wild-type. Infection assessed on detached calamondin leaves revealed that application of CaCl2 or neomycin 24 h prior to inoculation provided protection against Alt. alternata. These data indicate that a dynamic equilibrium of cellular Ca(2+) is critical for developmental and pathological processes of Alt. alternata. © 2014 The Authors.

  13. Epoxide hydrolase: a mRNA induced by the fungal pathogen Alternaria alternata on rough lemon (Citrus jambhiri Lush).

    PubMed

    Gomi, Kenji; Yamamato, Hiroyuki; Akimitsu, Kazuya

    2003-09-01

    An expression profile of genes induced by non-pathogenic Alternaria alternata on rough lemon leaves was obtained by sequencing 500 subtractive PCR clones generated from mRNA of leaves inoculated with the fungus after subtraction with that of non-inoculated leaves. About 6% of the cDNA sequences had homology to known putative defense-related genes including epoxide hydrolase. A full-length cDNA (951 bp) from rough lemon that encoded epoxy hydrolase was isolated by random amplification of cDNA ends (RACEs), based on sequence information from subtractive PCR, and designated as RlemEH. The product of this gene expressed with an in vitro translation system with Escherichia coli also had activity of a soluble type of epoxide hydrolase. The transcript of rough lemon RlemEH was not detected in flowers, fruits, stems or leaves, but was induced after inoculation of leaves with conidia of Alternaria alternata, wounding, or treatment with C6 volatiles, including trans-2-hexenol and cis-3-hexenol, and methyl jasmonate. The response of the epoxide hydrolase gene correlated well with the activation of defense mechanisms induced in plant-fungus interactions.

  14. Endopolygalacturonase is essential for citrus black rot caused by Alternaria citri but not brown spot caused by Alternaria alternata.

    PubMed

    Isshiki, A; Akimitsu, K; Yamamoto, M; Yamamoto, H

    2001-06-01

    Alternaria citri, the cause of Alternaria black rot, and Alternaria alternata rough lemon pathotype, the cause of Alternaria brown spot, are morphologically indistinguishable pathogens of citrus: one causes rot by macerating tissues and the other causes necrotic spots by producing a host-selective toxin. To evaluate the role of endopolygalacturonase (endoPG) in pathogenicity of these two Alternaria spp. pathogens, their genes for endoPG were mutated by gene targeting. The endoPGs produced by these fungi have similar biochemical properties, and the genes are highly similar (99.6% nucleotide identity). The phenotypes of the mutants, however, are completely different. An endoPG mutant of A. citri was significantly reduced in its ability to cause black rot symptoms on citrus as well as in the maceration of potato tissue and could not colonize citrus peel segments. In contrast, an endoPG mutant of A. alternata was unchanged in pathogenicity. The results indicate that a cell wall-degrading enzyme can play different roles in the pathogenicity of fungal pathogens. The role of a cell wall-degrading enzyme depends upon the type of disease but not the taxonomy of the fungus.

  15. The glutathione peroxidase-mediated reactive oxygen species resistance, fungicide sensitivity and cell wall construction in the citrus fungal pathogen Alternaria alternata.

    PubMed

    Yang, Siwy Ling; Yu, Pei-Ling; Chung, Kuang-Ren

    2016-03-01

    The ability to detoxify reactive oxygen species (ROS) is critical for pathogenicity in the necrotrophic fungus Alternaria alternata. We report a glutathione peroxidase 3 (AaGPx3) involved in the complex signalling network that is essential for the detoxification of cellular stresses induced by ROS and for A. alternata pathogenesis in citrus. AaGPx3 deletion mutants displayed increased sensitivity to H2 O2 and many ROS-generating compounds. AaGPx3 is required for correct fungal development as the AaGPx3 mutant strains showed a severe reduction in conidiation. AaGPx3 mutants accumulated higher chitin content than the wild-type and were less sensitive to the cell wall-targeting compounds calcofluor white and Congo red, as well as the fungicides fludioxonil and vinclozolin, suggesting a role of the glutathione systems in fungal cell wall construction. Virulence assays revealed that AaGPx3 is required for full virulence. The expression of AaGPx3 was downregulated in fungal strains carrying defective NADPH oxidase (Nox) or the oxidative stress responsive regulators YAP1 and HOG1, all implicated in ROS resistance. These results further support the important role of ROS detoxification during A. alternata pathogenesis in citrus. Overall, our study provides genetic evidence to define the central role of AaGPx3 in the biological and pathological functions of A. alternata. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  16. The YAP1 homolog-mediated oxidative stress tolerance is crucial for pathogenicity of the necrotrophic fungus Alternaria alternata in citrus.

    PubMed

    Lin, Ching-Hsuan; Yang, Siwy Ling; Chung, Kuang-Ren

    2009-08-01

    Citrus brown spot disease is caused by the necrotrophic fungus Alternaria alternata. Its pathogenic capability has been thought to depend exclusively on the production of host-selective ACT toxin. However, circumvention of plant defenses is also likely to be important for the disease process. To investigate the fungal response to host-generated reactive oxygen species (ROS), we cloned and characterized the AaAP1 gene of A. alternata, which encodes a polypeptide resembling yeast YAP1-like transcriptional activators implicated in cellular responses to stress. Expression of the AaAP1 gene in a wild-type strain was primarily induced by H(2)O(2) or ROS-generating oxidants. Using a loss-of-function mutation in the AaAP1 gene, we demonstrated an essential requirement for oxidative tolerance during the host invasion step. Mutants lacking AaAP1 showed increased sensitivity to H(2)O(2) and loss of fungal pathogenicity. The DeltaAaAP1 null mutant did not cause any visible necrotic lesions on wounded or unwounded leaves of citrus cv. Minneola. Compared with the wild type, the null mutant displayed lower catalase, peroxidase, and superoxide dismutase activities. All mutant phenotypes were restored to the wild type in fungal strains expressing a functional copy of AaAP1. Upon exposure to H(2)O(2), the AaAP1::sGFP (synthetic green fluorescent protein) fusion protein became localized in the nucleus. Inoculation of the mutant with NADPH oxidase inhibitors partially restored fungal pathogenicity. Our results highlight the global regulatory role of a YAP1 homolog in response to oxidative stress in A. alternata and provide insights into the critical role of ROS detoxification in the pathogenicity of A. alternata.

  17. Citrus Black Rot is Caused by Phylogenetically Distinct Lineages of Alternaria alternata.

    PubMed

    Peever, T L; Carpenter-Boggs, L; Timmer, L W; Carris, L M; Bhatia, A

    2005-05-01

    ABSTRACT Phylogenetic analysis revealed that isolates of Alternaria alternata causing black rot of citrus were associated with six well-supported evolutionary lineages. Isolates recovered from brown spot lesions on Minneola tangelo, leaf spot lesions on rough lemon, and healthy citrus tissue and noncitrus hosts were related closely to isolates from black-rotted fruit. Phylogenies estimated independently from DNA sequence data from an endopolygalacturonase gene (endoPG) and two anonymous regions of the genome (OPA1-3 and OPA2-1) had similar topologies, and phylogenetic analysis was performed on the combined data set. In the combined phylogeny, isolates from diverse ecological niches on citrus and noncitrus hosts were distributed in eight clades. Isolates from all lineages, regardless of ecological or host association, caused black rot in fruit inoculation assays, demonstrating that small-spored Alternaria isolates associated with different ecological niches on citrus and other plant hosts are potential black rot pathogens. These data also indicated that the fungi associated with black-rotted fruit do not form a natural evolutionary group distinct from other Alternaria pathogens and saprophytes associated with citrus. The use of the name A. citri to describe fungi associated with citrus black rot is not justified and it is proposed that citrus black rot fungi be referred to as A. alternata.

  18. Specialized and shared functions of the histidine kinase- and HOG1 MAP kinase-mediated signaling pathways in Alternaria alternata, a filamentous fungal pathogen of citrus.

    PubMed

    Lin, Ching-Hsuan; Chung, Kuang-Ren

    2010-10-01

    Signal transduction pathways are critical for the coordination of complex cellular processes in cells. In Alternaria alternata, a necrotrophic fungal pathogen of citrus, cloning and characterization of a gene coding a Group III histidine kinase (AaHSK1) and the yeast HOG1 ortholog (AaHOG1) showed the two genes to operate, both uniquely and synergistically, in a number of physiological and pathological functions. Systemic loss-of-function genetics in A. alternata revealed that AaHSK1 is a primary regulator for cellular resistance to sugar osmotic stress and for sensitivity to dicarboximide or phenylpyrrole fungicides. These functions were likely modulated by unknown mechanisms rather than solely by the AaHOG1-mediated pathway. AaHOG1, which conferred cellular resistance to salts and oxidative stress, also bypassed AaHSK1, even though deletion of AaHSK1 affected AaHOG1 phosphorylation. Phosphorylation of AaHOG1 was increased when the fungus was treated with osmotic stress, fungicides or H(2)O(2). Fungal mutants impaired in AaHSK1, AaHOG1, AaAP1 (encoding a redox-responsive transcription factor) or AaFUS3 (encoding a MAP kinase) were all hypersensitive to 2-chloro-5-hydroxypyridine (CHP) or 2,3,5-triiodobenzoic acid (TIBA). An AaHOG1::sGFP (synthetic green fluorescent protein) fusion protein became localized in the nucleus in response to H(2)O(2), CHP, TIBA, fungicides, but not glucose. Glucose, however, enhanced AaHOG1 phosphorylation and nuclear localization in the AaHSK1 deficient background. Accumulation of the AaHSK1 gene transcript was negatively regulated by AaHOG1, AaAP1 or AaFUS3. AaHOG1 was necessary for fungal pathogenicity, yet AaHSK1 was completely dispensable for pathogenicity. Our results highlight a dramatic flexibility and uniqueness in the signaling pathways that are involved in responding to diverse environmental stimuli in A. alternata. Copyright © 2010 Elsevier Inc. All rights reserved.

  19. A G alpha subunit gene is essential for conidiation and potassium efflux but dispensable for pathogenicity of Alternaria alternata on citrus.

    PubMed

    Wang, Nan-Yi; Lin, Ching-Hsuan; Chung, Kuang-Ren

    2010-02-01

    Heterotrimeric G proteins play a profound role in the recognition and transduction of extracellular signals in eukaryotic cells. We characterized the AaG alpha1 gene, encoding a fungal Class I G alpha subunit of the GTP-binding protein, in Alternaria alternata of citrus. Interruption of AaG alpha1 with a marker gene resulted in fungal transformants producing fewer conidia, becoming hypersensitive to KCl, and displaying elevated lipolytic and pectolytic activities. Expression of a functional copy of AaG alpha1 in a null mutant restored all altered phenotypes to the wild type. The AaG alpha1 mutants, whose conidia germinate normally, caused necrotic lesions on citrus indistinguishable from wild type. Application of cAMP, its inhibitors (atropine and theophylline), or 3-isobutyl-1-methylxanthine (IBMX) decreased conidiation in the fungal strains carrying a functional AaG alpha1. In contrast, conidial formation in the null mutants was restored by dibutyryl-cAMP or by a low concentration of cAMP or theophylline (1 mM). Unlike the oxidative stress-responsive AaAP1 transcription activator or the AaFUS3 mitogen-activated protein kinase (MAPK), AaG alpha1 is not required for cellular resistance to oxidative, osmotic, or chemical stress. AaFUS3 has also been demonstrated to be essential for conidial formation, suggesting a possible interaction between AaG alpha1 and AaFUS3 during conidiation even though expression of AaG alpha1 was not affected by AaFUS3 and vice versa. Inactivation of AaG alpha1 suppressed accumulation of transcripts of the AaAP1 gene and the AaHSK1 gene encoding a histidine kinase. These are novel features that have not been previously characterized to be associated with the GTP-binding protein.

  20. A Major Facilitator Superfamily Transporter-Mediated Resistance to Oxidative Stress and Fungicides Requires Yap1, Skn7, and MAP Kinases in the Citrus Fungal Pathogen Alternaria alternata.

    PubMed

    Chen, Li-Hung; Tsai, Hsieh-Chin; Yu, Pei-Ling; Chung, Kuang-Ren

    2017-01-01

    Major Facilitator Superfamily (MFS) transporters play an important role in multidrug resistance in fungi. We report an AaMFS19 gene encoding a MFS transporter required for cellular resistance to oxidative stress and fungicides in the phytopathogenic fungus Alternaria alternata. AaMFS19, containing 12 transmembrane domains, displays activity toward a broad range of substrates. Fungal mutants lacking AaMFS19 display profound hypersensitivities to cumyl hydroperoxide, potassium superoxide, many singlet oxygen-generating compounds (eosin Y, rose Bengal, hematoporphyrin, methylene blue, and cercosporin), and the cell wall biosynthesis inhibitor, Congo red. AaMFS19 mutants also increase sensitivity to copper ions, clotrimazole, fludioxonil, and kocide fungicides, 2-chloro-5-hydroxypyridine (CHP), and 2,3,5-triiodobenzoic acid (TIBA). AaMFS19 mutants induce smaller necrotic lesions on leaves of a susceptible citrus cultivar. All observed phenotypes in the mutant are restored by introducing and expressing a wild-type copy of AaMFS19. The wild-type strain of A. alternata treated with either CHP or TIBA reduces radial growth and formation and germination of conidia, increases hyphal branching, and results in decreased expression of the AaMFS19 gene. The expression of AaMFS19 is regulated by the Yap1 transcription activator, the Hog1 and Fus3 mitogen-activated protein (MAP) kinases, the 'two component' histidine kinase, and the Skn7 response regulator. Our results demonstrate that A. alternata confers resistance to different chemicals via a membrane-bound MFS transporter.

  1. Green Fluorescent Detection of Fungal Colonization and Endopolygalacturonase Gene Expression in the Interaction of Alternaria citri with Citrus.

    PubMed

    Isshiki, A; Ohtani, K; Kyo, M; Yamamoto, H; Akimitsu, K

    2003-07-01

    ABSTRACT Alternaria citri, a postharvest pathogen, produces endopolygalacturonase (endoPG) and causes black rot on citrus fruit. We previously described that an endoPG-disrupted mutant of Alternaria citri was significantly reduced in its ability to macerate plant tissue and cause black rot symptoms on citrus. In order to investigate colonization of citrus fruit tissues by Alternaria citri, pTEFEGFP carrying a green fluorescent protein (GFP) gene was introduced into wild-type Alternaria citri and its endoPG-disrupted mutant (M60). Green fluorescence was observed in spores, germ tubes, appressoria, and infection hyphae of transformants G1 (derived from wild type) and GM4 (derived from M60). Hyphae of G1 but not GM4 vertically penetrated the peel, but the hyphae of both G1 and GM4 spread equally in the juice sac area of citrus fruit. Green fluorescence of Alternaria citri transformant EPG7 carrying a GFP gene under control of the endoPG gene promoter of Alternaria citri was induced by pectin in the peel during the infection stage, but repressed completely in the juice sac area, likely by carbon catabolite repression by sugars in the juice.

  2. A Major Facilitator Superfamily Transporter-Mediated Resistance to Oxidative Stress and Fungicides Requires Yap1, Skn7, and MAP Kinases in the Citrus Fungal Pathogen Alternaria alternata

    PubMed Central

    Chen, Li-Hung; Tsai, Hsieh-Chin; Yu, Pei-Ling

    2017-01-01

    Major Facilitator Superfamily (MFS) transporters play an important role in multidrug resistance in fungi. We report an AaMFS19 gene encoding a MFS transporter required for cellular resistance to oxidative stress and fungicides in the phytopathogenic fungus Alternaria alternata. AaMFS19, containing 12 transmembrane domains, displays activity toward a broad range of substrates. Fungal mutants lacking AaMFS19 display profound hypersensitivities to cumyl hydroperoxide, potassium superoxide, many singlet oxygen-generating compounds (eosin Y, rose Bengal, hematoporphyrin, methylene blue, and cercosporin), and the cell wall biosynthesis inhibitor, Congo red. AaMFS19 mutants also increase sensitivity to copper ions, clotrimazole, fludioxonil, and kocide fungicides, 2-chloro-5-hydroxypyridine (CHP), and 2,3,5-triiodobenzoic acid (TIBA). AaMFS19 mutants induce smaller necrotic lesions on leaves of a susceptible citrus cultivar. All observed phenotypes in the mutant are restored by introducing and expressing a wild-type copy of AaMFS19. The wild-type strain of A. alternata treated with either CHP or TIBA reduces radial growth and formation and germination of conidia, increases hyphal branching, and results in decreased expression of the AaMFS19 gene. The expression of AaMFS19 is regulated by the Yap1 transcription activator, the Hog1 and Fus3 mitogen-activated protein (MAP) kinases, the ‘two component’ histidine kinase, and the Skn7 response regulator. Our results demonstrate that A. alternata confers resistance to different chemicals via a membrane-bound MFS transporter. PMID:28060864

  3. The FUS3 MAPK signaling pathway of the citrus pathogen Alternaria alternata functions independently or cooperatively with the fungal redox-responsive AP1 regulator for diverse developmental, physiological and pathogenic processes.

    PubMed

    Lin, Ching-Hsuan; Yang, Siwy Ling; Wang, Nan-Yi; Chung, Kuang-Ren

    2010-04-01

    Alternaria alternata, the fungus that causes citrus brown spot, invades its hosts primarily through the production and action of a host-selective ACT toxin that kills citrus cells prior to invasion. In this study, we show that, in the tangerine pathotype of A. alternata, a mitogen-activated protein kinase (MAPK)-mediated signaling pathway governs a number of biological functions, either separately or in a cooperative manner, with the AaAP1 gene encoding a transcription regulator. The reported MAPK is encoded by the AaFUS3 gene, which we show to be necessary for conidial development, resistance to copper fungicides, melanin biosynthesis, and particularly, for elaboration of the penetration process. In contrast, AaFUS3 negatively controls salt tolerance and production of several hydrolytic enzymes. AaFUS3 has no apparent role in the biosynthesis of host-selective toxin or in resistance to oxidative stress. Both AaAP1 and AaFUS3 are required for fungal resistance to 2,3,5-triiodobenzoic acid (TIBA), 2-chloro-5-hydroxypyridine (CHP), diethyl maleate (DEM), and many pyridine-containing compounds. A strain with mutations in both AaAP1 and AaFUS3 displayed an increased sensitivity to these compounds. Expression of the AaAP1 and AaFUS3 genes and phosphorylation of AaFUS3 were also induced by CHP, DEM, or TIBA. Expression of two genes coding for a putative MFS transporter was coordinately regulated by AaAP1 and AaFUS3. The AaAP1::sGFP (synthetic green fluorescent protein) fusion protein became localized in the nucleus in response to CHP or TIBA. Inactivation of the AaAP1 gene, however, promoted phosphorylation of AaFUS3. Taken together, our results indicate that A. alternata utilizes specialized or synergistic regulatory interactions between the AP1 and MAPK signaling pathways for diverse physiological functions.

  4. Involvement of an extracellular fungus laccase in the flavonoid metabolism in Citrus fruits inoculated with Alternaria alternata.

    PubMed

    Díaz, Licinio; Del Río, José Antonio; Pérez-Gilabert, Manuela; Ortuño, Ana

    2015-04-01

    Fungi of the genus Alternaria are responsible for substantial pre-harvest losses in Citrus. In this study a degradative metabolism of flavonoids (flavanones, flavones and polymethoxyflavones) was observed when 'Fortune' mandarin, Citrus limon and Citrus paradisi, fruits were inoculated with Alternaria alternata, a pre-harvest pathogenic fungus. Associated to this flavonic metabolism the de novo synthesis of the phytoalexin scoparone was detected. This metabolism of flavonoids is caused by an extracellular fungus laccase. The kinetic characterisation of this enzyme revealed that the activity was induced by Citrus flavonoids and was dependent on flavonoid concentrations. The enzyme exhibited a Km of 1.9 mM using ABTS as substrate with an optimum pH of 3.5 in citrate buffer 100 mM. The enzyme is active between 15 and 45 °C, the optimum temperature being around 35 °C, although 50% of the initial activity is lost after 45 min at 35 °C. The A. alternata laccase was inhibited by 0.5 mM l-cysteine and by caffeic acid. Study of the substrate specificity of this enzyme revealed that Citrus flavonoids are substrates of A. alternata laccase. These results suggest that the laccase enzyme could be involved in the pathogenesis of A. alternata in Citrus.

  5. The NADPH oxidase-mediated production of hydrogen peroxide (H(2)O(2)) and resistance to oxidative stress in the necrotrophic pathogen Alternaria alternata of citrus.

    PubMed

    Yang, Siwy Ling; Chung, Kuang-Ren

    2012-10-01

    It has become increasingly apparent that the production of reactive oxygen species (ROS) by the NADPH oxidase (Nox) complex is vital for cellular differentiation and signalling in fungi. We cloned and characterized an AaNoxA gene of the necrotrophic fungus Alternaria alternata, which encodes a polypeptide analogous to mammalian gp91(phox) and fungal Noxs implicated in the generation of ROS. Genetic analysis confirmed that AaNoxA is responsible for the production of ROS. Moreover, deletion of AaNoxA in A. alternata resulted in an elevated hypersensitivity to hydrogen peroxide (H(2)O(2)), menadione, potassium superoxide (KO(2)), diamide and many ROS-generating compounds. The results implicate the involvement of AaNoxA in cellular resistance to ROS stress. The impaired phenotypes strongly resemble those previously seen for the ap1 null mutant defective in a YAP1-like transcriptional regulator and for the hog1 mutant defective in a HOG1-like mitogen-activated protein (MAP) kinase. The noxA null mutant was also hypersensitive to Nox inhibitors, nitric oxide (NO(·)) donors and NO(·) synthase inhibitors, implying a role of AaNoxA in the NO(·) signalling pathway. Expression of AaNoxA was activated by H(2)O(2), menadione, KO(2), NO(·) donors and L-arginine (a substrate for NO(·) synthase). AaNoxA may be able to sense and respond to both ROS and nitric oxide. Moreover, AaNoxA is required for normal conidiation and full fungal virulence. AaNoxA promoted the expression of the AaAP1 and AaHOG1 genes in A. alternata. Inactivation of AaNoxA greatly reduced the transcriptional activation of AaAP1 in response to ROS stress. Thus, we conclude that the regulatory functions of AaNoxA conferring ROS resistance are modulated partially through the activation of the YAP1- and HOG1 MAP kinase-mediated signalling pathways. © 2012 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

  6. Sensitivity to Alternaria alternata toxin in citrus because of altered mitochondrial RNA processing

    PubMed Central

    Ohtani, Kouhei; Yamamoto, Hiroyuki; Akimitsu, Kazuya

    2002-01-01

    Specificity in the interaction between rough lemon (Citrus jambhiri Lush.) and the fungal pathogen Alternaria alternata rough lemon pathotype is determined by a host-selective toxin, ACR-toxin. Mitochondria from rough lemon are sensitive to ACR-toxin whereas mitochondria from resistant plants, including other citrus species, are resistant. We have identified a C. jambhiri mitochondrial DNA sequence, designated ACRS (ACR-toxin sensitivity gene), that confers toxin sensitivity to Escherichia coli. ACRS is located in the group II intron of the mitochondrial tRNA-Ala and is translated into a SDS-resistant oligomeric protein in C. jambhiri mitochondria but is not translated in the toxin-insensitive mitochondria. ACRS is present in the mitochondrial genome of both toxin-sensitive and -insensitive citrus. However, in mitochondria of toxin-insensitive plants, the transcripts from ACRS are shorter than those in mitochondria of sensitive plants. These results demonstrate that sensitivity to ACR-toxin and hence specificity of the interaction between A. alternata rough lemon pathotype and C. jambhiri is due to differential posttranscriptional processing of a mitochondrial gene. PMID:11842194

  7. Identification of a novel phylogenetic lineage of Alternaria alternata causing citrus brown spot in China.

    PubMed

    Huang, Feng; Fu, Yushi; Nie, Danni; Stewart, Jane E; Peever, Tobin L; Li, Hongye

    2015-05-01

    Alternaria alternata sensu lato, casual agent of citrus brown spot, first identified in Yunnan province in 2010 and subsequently found in Zhejiang, Hunan, Guangdong provinces, Chongqing municipality andGuangxi autonomous region in China. During 2010-2012, 86 isolates were collected from diseased citrus, of which 85 % isolates were pathogenic to Ponkan tangerine. Phylogenetic analyses of Chinese and worldwide isolates using partial sequences of an endopolygalacturonase gene (endoPG) and combined dataset ofendoPG and two anonymous loci (OPA1-3, OPA2-1) found that Chinese isolates fell into two of three previously described clades. One clade ('clade 3') contained isolates from Turkey and Israel, and the other clade ('clade 1') contained isolates from Florida, USA. None of the isolates from China fell into the last previously described clade ('clade 2'). However, 24 isolates from Hunan, Guangdong and Guangxi fell into a fourth clade ('clade 4') not previously reported to be associated with citrus brown spot. This clade included multilocus haplotypes known to infect Japanese pear and strawberry. The observation that Chinese brown spot isolates fell into only two of three known worldwide lineages suggests that this fungus may not have co-evolved with its host in China but elsewhere in Southeast Asia and introduced to China. Copyright © 2014 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  8. Sensitivity to Alternaria alternata toxin in citrus because of altered mitochondrial RNA processing.

    PubMed

    Ohtani, Kouhei; Yamamoto, Hiroyuki; Akimitsu, Kazuya

    2002-02-19

    Specificity in the interaction between rough lemon (Citrus jambhiri Lush.) and the fungal pathogen Alternaria alternata rough lemon pathotype is determined by a host-selective toxin, ACR-toxin. Mitochondria from rough lemon are sensitive to ACR-toxin whereas mitochondria from resistant plants, including other citrus species, are resistant. We have identified a C. jambhiri mitochondrial DNA sequence, designated ACRS (ACR-toxin sensitivity gene), that confers toxin sensitivity to Escherichia coli. ACRS is located in the group II intron of the mitochondrial tRNA-Ala and is translated into a SDS-resistant oligomeric protein in C. jambhiri mitochondria but is not translated in the toxin-insensitive mitochondria. ACRS is present in the mitochondrial genome of both toxin-sensitive and -insensitive citrus. However, in mitochondria of toxin-insensitive plants, the transcripts from ACRS are shorter than those in mitochondria of sensitive plants. These results demonstrate that sensitivity to ACR-toxin and hence specificity of the interaction between A. alternata rough lemon pathotype and C. jambhiri is due to differential posttranscriptional processing of a mitochondrial gene.

  9. Large-spored Alternaria pathogens in section Porri disentangled

    PubMed Central

    Woudenberg, J.H.C.; Truter, M.; Groenewald, J.Z.; Crous, P.W.

    2014-01-01

    The omnipresent fungal genus Alternaria was recently divided into 24 sections based on molecular and morphological data. Alternaria sect. Porri is the largest section, containing almost all Alternaria species with medium to large conidia and long beaks, some of which are important plant pathogens (e.g. Alternaria porri, A. solani and A. tomatophila). We constructed a multi-gene phylogeny on parts of the ITS, GAPDH, RPB2, TEF1 and Alt a 1 gene regions, which, supplemented with morphological and cultural studies, forms the basis for species recognition in sect. Porri. Our data reveal 63 species, of which 10 are newly described in sect. Porri, and 27 species names are synonymised. The three known Alternaria pathogens causing early blight on tomato all cluster in one clade, and are synonymised under the older name, A. linariae. Alternaria protenta, a species formerly only known as pathogen on Helianthus annuus, is also reported to cause early blight of potato, together with A. solani and A. grandis. Two clades with isolates causing purple blotch of onion are confirmed as A. allii and A. porri, but the two species cannot adequately be distinguished based on the number of beaks and branches as suggested previously. This is also found among the pathogens of Passifloraceae, which are reduced from four to three species. In addition to the known pathogen of sweet potato, A. bataticola, three more species are delineated of which two are newly described. A new Alternaria section is also described, comprising two large-spored Alternaria species with concatenate conidia. PMID:25492985

  10. Comparative Genomics of Pathogens Causing Brown Spot Disease of Tobacco: Alternaria longipes and Alternaria alternata

    PubMed Central

    Wan, Wenting; Long, Ni; Zhang, Jing; Tan, Yuntao; Duan, Shengchang; Zeng, Yan; Dong, Yang

    2016-01-01

    The genus Alternaria is a group of infectious/contagious pathogenic fungi that not only invade a wide range of crops but also induce severe allergic reactions in a part of the human population. In this study, two strains Alternaria longipes cx1 and Alternaria alternata cx2 were isolated from different brown spot lesions on infected tobacco leaves. Their complete genomes were sequenced, de novo assembled, and comparatively analyzed. Phylogenetic analysis revealed that A. longipes cx1 and A. alternata cx2 diverged 3.3 million years ago, indicating a recent event of speciation. Seventeen non-ribosomal peptide synthetase (NRPS) genes and 13 polyketide synthase (PKS) genes in A. longipes cx1 and 13 NRPS genes and 12 PKS genes in A. alternata cx2 were identified in these two strains. Some of these genes were predicted to participate in the synthesis of non-host specific toxins (non-HSTs), such as tenuazonic acid (TeA), alternariol (AOH) and alternariol monomethyl ether (AME). By comparative genome analysis, we uncovered that A. longipes cx1 had more genes putatively involved in pathogen-plant interaction, more carbohydrate-degrading enzymes and more secreted proteins than A. alternata cx2. In summary, our results demonstrate the genomic distinction between A. longipes cx1 and A. altenata cx2. They will not only improve the understanding of the phylogenetic relationship among genus Alternaria, but more importantly provide valuable genomic resources for the investigation of plant-pathogen interaction. PMID:27159564

  11. Stress Response and Pathogenicity of the Necrotrophic Fungal Pathogen Alternaria alternata

    PubMed Central

    Chung, Kuang-Ren

    2012-01-01

    The production of host-selective toxins by the necrotrophic fungus Alternaria alternata is essential for the pathogenesis. A. alternata infection in citrus leaves induces rapid lipid peroxidation, accumulation of hydrogen peroxide (H2O2), and cell death. The mechanisms by which A. alternata avoids killing by reactive oxygen species (ROS) after invasion have begun to be elucidated. The ability to coordinate of signaling pathways is essential for the detoxification of cellular stresses induced by ROS and for pathogenicity in A. alternata. A low level of H2O2, produced by the NADPH oxidase (NOX) complex, modulates ROS resistance and triggers conidiation partially via regulating the redox-responsive regulators (YAP1 and SKN7) and the mitogen-activated protein (MAP) kinase (HOG1) mediated pathways, which subsequently regulate the genes required for the biosynthesis of siderophore, an iron-chelating compound. Siderophore-mediated iron acquisition plays a key role in ROS detoxification because of the requirement of iron for the activities of antioxidants (e.g., catalase and SOD). Fungal strains impaired for the ROS-detoxifying system severely reduce the virulence on susceptible citrus cultivars. This paper summarizes the current state of knowledge of signaling pathways associated with cellular responses to multidrugs, oxidative and osmotic stress, and fungicides, as well as the pathogenicity/virulence in the tangerine pathotype of A. alternata. PMID:24278721

  12. Stress Response and Pathogenicity of the Necrotrophic Fungal Pathogen Alternaria alternata.

    PubMed

    Chung, Kuang-Ren

    2012-01-01

    The production of host-selective toxins by the necrotrophic fungus Alternaria alternata is essential for the pathogenesis. A. alternata infection in citrus leaves induces rapid lipid peroxidation, accumulation of hydrogen peroxide (H2O2), and cell death. The mechanisms by which A. alternata avoids killing by reactive oxygen species (ROS) after invasion have begun to be elucidated. The ability to coordinate of signaling pathways is essential for the detoxification of cellular stresses induced by ROS and for pathogenicity in A. alternata. A low level of H2O2, produced by the NADPH oxidase (NOX) complex, modulates ROS resistance and triggers conidiation partially via regulating the redox-responsive regulators (YAP1 and SKN7) and the mitogen-activated protein (MAP) kinase (HOG1) mediated pathways, which subsequently regulate the genes required for the biosynthesis of siderophore, an iron-chelating compound. Siderophore-mediated iron acquisition plays a key role in ROS detoxification because of the requirement of iron for the activities of antioxidants (e.g., catalase and SOD). Fungal strains impaired for the ROS-detoxifying system severely reduce the virulence on susceptible citrus cultivars. This paper summarizes the current state of knowledge of signaling pathways associated with cellular responses to multidrugs, oxidative and osmotic stress, and fungicides, as well as the pathogenicity/virulence in the tangerine pathotype of A. alternata.

  13. The major Alternaria alternata allergen, Alt a 1: A reliable and specific marker of fungal contamination in citrus fruits.

    PubMed

    Gabriel, M F; Uriel, N; Teifoori, F; Postigo, I; Suñén, E; Martínez, J

    2017-09-18

    The ubiquitously present spores of Alternaria alternata can spoil a wide variety of foodstuffs, including a variety of fruits belonging to the Citrus genus. The major allergenic protein of A. alternata, Alt a 1, is a species-specific molecular marker that has been strongly associated with allergenicity and phytopathogenicity of this fungal species. This study aimed to evaluate the potential of the detection of Alt a 1 as a reliable indicator of A. alternata contamination in citrus fruits. To accomplish this aim, sixty oranges were artificially infected with a spore suspension of A. alternata. Internal fruit material was collected at different incubation times (one, two and three weeks after the fungal inoculation) and used for both total RNA extraction and protein extraction. Alt a 1 detection was then performed by polymerase chain reaction (PCR) amplification using Alt a 1 specific primers and by enzyme-linked immunosorbent assay (ELISA). The experimental model presented in this work was effective to simulate the typical Alternaria black rot phenotype and its progression. Although both PCR and ELISA techniques have been successfully carried out for detecting Alt a 1 allergen in A. alternata infected oranges, the PCR method was found to be more sensitive than ELISA. Nevertheless, ELISA results were highly valuable to demonstrate that considerable amounts of Alt a 1 are produced during A. alternata fruit infection process, corroborating the recently proposed hypothesis that this protein plays a role in the pathogenicity and virulence of Alternaria species. Such evidence suggests that the detection of Alt a 1 by PCR-based assay may be used as a specific indicator of the presence of pathogenic and allergenic fungal species, A. alternata, in fruits. This knowledge can be employed to control the fungal infection and mitigate agricultural losses as well as human exposure to A. alternata allergens and toxins. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Comparison of methodologies for conidia production by Alternaria alternata from citrus

    PubMed Central

    Carvalho, Daniel D. C.; Alves, Eduardo; Batista, Tereza R. S.; Camargos, Renato B.; Lopes, Eloísa A. G. L.

    2008-01-01

    Conidia production is a problem in the study of Alternaria alternata from citrus. Thus, this study aimed to compare existing methodologies for conidial production of A. alternata isolated from Ponkan tangerine (2 isolates), Cravo lemon (1 isolate), Pêra orange (2 isolates) and Murcott tangor (1 isolate). The methodologies used were conidia production with 12 and 24 hours under white fluorescent light, evaluation with 24 and 48 hours after applying fungal mycelium stress technique, cold stress followed by injury of mycelium and evaluation with 24 hours, using healthy vegetable tissue and the use of black fluorescent near ultraviolet (NUV) lamp. Satisfactory result was obtained with A. alternata isolate from Murcott tangor, with the production of 2.8 × 105 conidia mL-1, when fungal mycelium was stressed (Petri dish with 66.66% of fungi growth) and subsequently 24 h of growth. The use of white light (24 h) and black fluorescent NUV lamp also induced expressive conidia production by one isolate of Ponkan tangerine, which produced 17.2 × 105 and 10.1 × 105conidia mL-1 and another of Murcott tangor, which produced 13.9 × 105 and 10.1 × 105 conidia mL-1, respectively. The remaining methodologies analyzed in this study were not able to induce conidia production in satisfactory quantity. The use of both mycelium stress technique and white light (24 h) and black fluorescent NUV lamp allowed the production of enough quantities of conidia to be used in vitro (detection of fungitoxic substances) and in vivo (pathogenicity test) assays, respectively. PMID:24031309

  15. Comparison of methodologies for conidia production by Alternaria alternata from citrus.

    PubMed

    Carvalho, Daniel D C; Alves, Eduardo; Batista, Tereza R S; Camargos, Renato B; Lopes, Eloísa A G L

    2008-10-01

    Conidia production is a problem in the study of Alternaria alternata from citrus. Thus, this study aimed to compare existing methodologies for conidial production of A. alternata isolated from Ponkan tangerine (2 isolates), Cravo lemon (1 isolate), Pêra orange (2 isolates) and Murcott tangor (1 isolate). The methodologies used were conidia production with 12 and 24 hours under white fluorescent light, evaluation with 24 and 48 hours after applying fungal mycelium stress technique, cold stress followed by injury of mycelium and evaluation with 24 hours, using healthy vegetable tissue and the use of black fluorescent near ultraviolet (NUV) lamp. Satisfactory result was obtained with A. alternata isolate from Murcott tangor, with the production of 2.8 × 10(5) conidia mL(-1), when fungal mycelium was stressed (Petri dish with 66.66% of fungi growth) and subsequently 24 h of growth. The use of white light (24 h) and black fluorescent NUV lamp also induced expressive conidia production by one isolate of Ponkan tangerine, which produced 17.2 × 10(5) and 10.1 × 10(5)conidia mL(-1) and another of Murcott tangor, which produced 13.9 × 10(5) and 10.1 × 10(5) conidia mL(-1), respectively. The remaining methodologies analyzed in this study were not able to induce conidia production in satisfactory quantity. The use of both mycelium stress technique and white light (24 h) and black fluorescent NUV lamp allowed the production of enough quantities of conidia to be used in vitro (detection of fungitoxic substances) and in vivo (pathogenicity test) assays, respectively.

  16. Assessment of protein silver nanoparticles toxicity against pathogenic Alternaria solani.

    PubMed

    Abdel-Hafez, Sobhy I I; Nafady, Nivien A; Abdel-Rahim, Ismail R; Shaltout, Abeer M; Daròs, José-Antonio; Mohamed, Mohamed A

    2016-12-01

    Mycogenic synthesis of silver nanoparticles (AgNPs) was carried out in the present investigation using an aqueous extract of endophytic non-pathogenic Alternaria solani F10 (KT721914). The mycosynthesized AgNPs were characterized by means of spectroscopic and microscopic techniques. The surface plasmon resonance found at 430 nm confirmed the formation of stable AgNPs for several weeks at room temperature. Also, the results revealed the formation of spherical and monodispersed AgNPs with an average size of 14.8 ± 1.2 nm. The FT-IR spectrum suggested that the fungal extracellular proteins and secondary metabolites had the role in Ag reduction and AgNPs capping of which protein Ag nanoconjugates were formed. Furthermore, the mycosynthesized AgNPs exhibited potent antifungal activity against different pathogenic isolates of the same Alternaria solani fungus, the causal pathogen of tomato early blight disease. The antifungal efficiency of the AgNPs at 1, 5 and 10 ppm were evaluated for 8 days after incubation by measuring the inhibition rate of fungal radial growth. The results were further supported by investigating fungal hyphae morphology alteration by scanning and transmission electron microscopy. Treated fungal hyphae showed formation of pits and pores. Also, the mycosynthesized AgNPs were able to pass and distribute throughout the fungal cell area and interact with the cell components.

  17. Participation of the phosphoinositide metabolism in the hypersensitive response of Citrus limon against Alternaria alternata.

    PubMed

    Ortega, X; Pérez, L M

    2001-01-01

    Lemon seedlings inoculated with Alternaria alternata develop a hypersensitive response (HR) that includes the induction of Phenylalanine ammonia-lyase (PAL, E. C. 4.3.1.5) and the synthesis of scoparone. The signal transduction pathway involved in the development of this response is unknown. We used several inhibitors of the Phosphoinositide (PI) animal system to study a possible role of Inositol-1,4,5-triphosphate (IP3) in the transduction of the fungal conidia signal in Citrus limon. The HR was only partially inhibited by EGTA, suggesting that not only external but internal calcium as well are necessary for a complete development of the HR. In this plant system, Alternaria alternata induced an early accumulation of the second messenger IP3. When lemon seedlings were watered long term with LiCl, an inhibitor of the phosphoinositide cycle, the IP3 production was reduced, and the LiCl-watered plants could neither induce PAL nor synthesize scoparone in response to fungal conidia. Furthermore, neomycin, a Phospholipase C (PLC, E. C. 3.1.4.3) inhibitor, also inhibited PAL induction and scoparone synthesis in response to A. alternata. These results suggest that IP3 could be involved in the signal transduction pathway for the development of the HR of Citrus limon against A. alternata.

  18. Natural occurrence of four Alternaria mycotoxins in tomato- and citrus-based foods in China.

    PubMed

    Zhao, Kai; Shao, Bing; Yang, Dajin; Li, Fengqin

    2015-01-14

    A total of 70 tomato-based and 86 citrus-based products collected in China were analyzed for alternariol, alternariol monomethyl ether, tentoxin, and tenuazonic acid by ultraperformance liquid chromatography–electrospray ionization–tandem mass spectrometry. No toxins were found in any fresh tomato or citrus fruit samples. Tenuazonic acid was the predominant toxin detected in all tomato ketchup (10.2–1787 μg/kg) and tomato juice samples (7.4–278 μg/kg). Alternariol was quantitated at higher level than alternariol monomethyl ether with the ratio of alternariol/alternariol monomethyl ether ranging from 0.37 to 104 in 14 alternariol-positive tomato ketchup samples. Tentoxin was detected at much lower levels in all samples analyzed. Some citrus juice samples were positive for tenuazonic acid and alternariol monomethyl ether. It is necessary to conduct a systemic surveillance of Alternaria toxins in raw and processed foods to provide the scientific basis for risk assessment of dietary exposure to these toxins in Chinese populations.

  19. Digital PCR for detection of citrus pathogens

    USDA-ARS?s Scientific Manuscript database

    Citrus trees are often infected with multiple pathogens of economic importance, especially those with insect or mite vectors. Real-time/quantitative PCR (qPCR) has been used for high-throughput detection and relative quantification of pathogens; however, target reference or standards are required. I...

  20. Alternaria redefined

    PubMed Central

    Woudenberg, J.H.C.; Groenewald, J.Z.; Binder, M.; Crous, P.W.

    2013-01-01

    Alternaria is a ubiquitous fungal genus that includes saprobic, endophytic and pathogenic species associated with a wide variety of substrates. In recent years, DNA-based studies revealed multiple non-monophyletic genera within the Alternaria complex, and Alternaria species clades that do not always correlate to species-groups based on morphological characteristics. The Alternaria complex currently comprises nine genera and eight Alternaria sections. The aim of this study was to delineate phylogenetic lineages within Alternaria and allied genera based on nucleotide sequence data of parts of the 18S nrDNA, 28S nrDNA, ITS, GAPDH, RPB2 and TEF1-alpha gene regions. Our data reveal a Pleospora/Stemphylium clade sister to Embellisia annulata, and a well-supported Alternaria clade. The Alternaria clade contains 24 internal clades and six monotypic lineages, the assemblage of which we recognise as Alternaria. This puts the genera Allewia, Brachycladium, Chalastospora, Chmelia, Crivellia, Embellisia, Lewia, Nimbya, Sinomyces, Teretispora, Ulocladium, Undifilum and Ybotromyces in synonymy with Alternaria. In this study, we treat the 24 internal clades in the Alternaria complex as sections, which is a continuation of a recent proposal for the taxonomic treatment of lineages in Alternaria. Embellisia annulata is synonymised with Dendryphiella salina, and together with Dendryphiella arenariae, are placed in the new genus Paradendryphiella. The sexual genera Clathrospora and Comoclathris, which were previously associated with Alternaria, cluster within the Pleosporaceae, outside Alternaria s. str., whereas Alternariaster, a genus formerly seen as part of Alternaria, clusters within the Leptosphaeriaceae. Paradendryphiella is newly described, the generic circumscription of Alternaria is emended, and 32 new combinations and 10 new names are proposed. A further 10 names are resurrected, while descriptions are provided for 16 new Alternaria sections. Taxonomic novelties: New

  1. Alternaria redefined.

    PubMed

    Woudenberg, J H C; Groenewald, J Z; Binder, M; Crous, P W

    2013-06-30

    Alternaria is a ubiquitous fungal genus that includes saprobic, endophytic and pathogenic species associated with a wide variety of substrates. In recent years, DNA-based studies revealed multiple non-monophyletic genera within the Alternaria complex, and Alternaria species clades that do not always correlate to species-groups based on morphological characteristics. The Alternaria complex currently comprises nine genera and eight Alternaria sections. The aim of this study was to delineate phylogenetic lineages within Alternaria and allied genera based on nucleotide sequence data of parts of the 18S nrDNA, 28S nrDNA, ITS, GAPDH, RPB2 and TEF1-alpha gene regions. Our data reveal a Pleospora/Stemphylium clade sister to Embellisia annulata, and a well-supported Alternaria clade. The Alternaria clade contains 24 internal clades and six monotypic lineages, the assemblage of which we recognise as Alternaria. This puts the genera Allewia, Brachycladium, Chalastospora, Chmelia, Crivellia, Embellisia, Lewia, Nimbya, Sinomyces, Teretispora, Ulocladium, Undifilum and Ybotromyces in synonymy with Alternaria. In this study, we treat the 24 internal clades in the Alternaria complex as sections, which is a continuation of a recent proposal for the taxonomic treatment of lineages in Alternaria. Embellisia annulata is synonymised with Dendryphiella salina, and together with Dendryphiella arenariae, are placed in the new genus Paradendryphiella. The sexual genera Clathrospora and Comoclathris, which were previously associated with Alternaria, cluster within the Pleosporaceae, outside Alternaria s. str., whereas Alternariaster, a genus formerly seen as part of Alternaria, clusters within the Leptosphaeriaceae. Paradendryphiella is newly described, the generic circumscription of Alternaria is emended, and 32 new combinations and 10 new names are proposed. A further 10 names are resurrected, while descriptions are provided for 16 new Alternaria sections. New combinations - Alternaria

  2. Cellular responses required for oxidative stress tolerance, colonization, and lesion formation by the necrotrophic fungus Alternaria alternata in citrus.

    PubMed

    Lin, Ching-Hsuan; Yang, Siwy Ling; Chung, Kuang-Ren

    2011-03-01

    The pathogenic capability of the tangerine pathotype of Alternaria alternata relies on the production of host-selective ACT toxin. Inoculation of A. alternata in leaves of the citrus quickly induced rapid lipid peroxidation, accumulation of hydrogen peroxide (H(2)O(2)), and cell death, indicative of host defensive response. We previously demonstrated an essential role of the A. alternata AaAP1 gene, encoding a redox-responsive YAP1-like transcription factor, to contribute to fungal pathogenicity. The AaAP1 null mutant fails to incite necrotic lesions. In this study, we show further that the fungal mutant defective at the AaAP1 locus displayed reduced activities for glutathione-S-transferase, glutathione peroxidase, glutathione reductase, and ligninolytic peroxidase, yet retained normal production of ACT toxin. In contrast to the wild-type progenitor and the genetically reverted strain, the mutant strain was unable to detoxify H(2)O(2) effectively and was killed upon exposure to H(2)O(2). The mutant strain induced lower levels of H(2)O(2) accumulation in citrus leaves, compared to those induced by the wild-type or by the genetically reverted strain. Upon exposure to H(2)O(2), A. alternata apparently changed expression of a wide array of the genes regulated by AaAP1. Thus, the impairment of the AaAP1 null mutants to incite necrotic lesions is apparently a consequence of their inability to alleviate the toxicity of ROS, and circumvention of plant defenses is important for the disease process.

  3. Fine Mapping for Identification of Citrus Alternaria Brown Spot Candidate Resistance Genes and Development of New SNP Markers for Marker-Assisted Selection.

    PubMed

    Cuenca, Jose; Aleza, Pablo; Garcia-Lor, Andres; Ollitrault, Patrick; Navarro, Luis

    2016-01-01

    Alternaria brown spot (ABS) is a serious disease affecting susceptible citrus genotypes, which is a strong concern regarding citrus breeding programs. Resistance is conferred by a recessive locus (ABSr) previously located by our group within a 3.3 Mb genome region near the centromere in chromosome III. This work addresses fine-linkage mapping of this region for identifying candidate resistance genes and develops new molecular markers for ABS-resistance effective marker-assisted selection (MAS). Markers closely linked to ABSr locus were used for fine mapping using a 268-segregating diploid progeny derived from a heterozygous susceptible × resistant cross. Fine mapping limited the genomic region containing the ABSr resistance gene to 366 kb, flanked by markers at 0.4 and 0.7 cM. This region contains nine genes related to pathogen resistance. Among them, eight are resistance (R) gene homologs, with two of them harboring a serine/threonine protein kinase domain. These two genes along with a gene encoding a S-adenosyl-L-methionine-dependent-methyltransferase protein, should be considered as strong candidates for ABS-resistance. Moreover, the closest SNP was genotyped in 40 citrus varieties, revealing very high association with the resistant/susceptible phenotype. This new marker is currently used in our citrus breeding program for ABS-resistant parent and cultivar selection, at diploid, triploid and tetraploid level.

  4. Fine Mapping for Identification of Citrus Alternaria Brown Spot Candidate Resistance Genes and Development of New SNP Markers for Marker-Assisted Selection

    PubMed Central

    Cuenca, Jose; Aleza, Pablo; Garcia-Lor, Andres; Ollitrault, Patrick; Navarro, Luis

    2016-01-01

    Alternaria brown spot (ABS) is a serious disease affecting susceptible citrus genotypes, which is a strong concern regarding citrus breeding programs. Resistance is conferred by a recessive locus (ABSr) previously located by our group within a 3.3 Mb genome region near the centromere in chromosome III. This work addresses fine-linkage mapping of this region for identifying candidate resistance genes and develops new molecular markers for ABS-resistance effective marker-assisted selection (MAS). Markers closely linked to ABSr locus were used for fine mapping using a 268-segregating diploid progeny derived from a heterozygous susceptible × resistant cross. Fine mapping limited the genomic region containing the ABSr resistance gene to 366 kb, flanked by markers at 0.4 and 0.7 cM. This region contains nine genes related to pathogen resistance. Among them, eight are resistance (R) gene homologs, with two of them harboring a serine/threonine protein kinase domain. These two genes along with a gene encoding a S-adenosyl-L-methionine-dependent-methyltransferase protein, should be considered as strong candidates for ABS-resistance. Moreover, the closest SNP was genotyped in 40 citrus varieties, revealing very high association with the resistant/susceptible phenotype. This new marker is currently used in our citrus breeding program for ABS-resistant parent and cultivar selection, at diploid, triploid and tetraploid level. PMID:28066498

  5. Phylogenetic, Morphological, and Pathogenic Characterization of Alternaria Species Associated with Fruit Rot of Blueberry in California.

    PubMed

    Zhu, X Q; Xiao, C L

    2015-12-01

    Fruit rot caused by Alternaria spp. is one of the most important factors affecting the postharvest quality and shelf life of blueberry fruit. The aims of this study were to characterize Alternaria isolates using morphological and molecular approaches and test their pathogenicity to blueberry fruit. Alternaria spp. isolates were collected from decayed blueberry fruit in the Central Valley of California during 2012 and 2013. In total, 283 isolates were obtained and five species of Alternaria, including Alternaria alternata, A. tenuissima, A. arborescens, A. infectoria, and A. rosae, were identified based on DNA sequences of the plasma membrane ATPase, Alt a1 and Calmodulin gene regions in combination with morphological characters of the culture and sporulation. Of the 283 isolates, 61.5% were identified as A. alternata, 32.9% were A. arborescens, 5.0% were A. tenuissima, and only one isolate of A. infectoria and one isolate of A. rosae were found. These fungi were able to grow at temperatures from 0 to 35°C, and mycelial growth was arrested at 40°C. Optimal radial growth occurred between 20 to 30°C. Pathogenicity tests showed that all five Alternaria spp. were pathogenic on blueberry fruit at 0, 4, and 20°C, with A. alternata, A. arborescens, and A. tenuissima being the most virulent species, followed by A. infectoria and A. rosae. Previously A. tenuissima has been reported to be the primary cause of Alternaria fruit rot of blueberry worldwide. Our results indicated that the species composition of Alternaria responsible for Alternaria fruit rot in blueberry can be dependent on geographical region. A. alternata, A. arborescens, A. infectoria, and A. rosae are reported for the first time on blueberry in California. This is also the first report of A. infectoria and A. rosae infecting blueberry fruit.

  6. New Pseudomonas spp. Are Pathogenic to Citrus

    PubMed Central

    Beiki, Farid; Busquets, Antonio; Gomila, Margarita; Rahimian, Heshmat; Lalucat, Jorge; García-Valdés, Elena

    2016-01-01

    Five putative novel Pseudomonas species shown to be pathogenic to citrus have been characterized in a screening of 126 Pseudomonas strains isolated from diseased citrus leaves and stems in northern Iran. The 126 strains were studied using a polyphasic approach that included phenotypic characterizations and phylogenetic multilocus sequence analysis. The pathogenicity of these strains against 3 cultivars of citrus is demonstrated in greenhouse and field studies. The strains were initially grouped phenotypically and by their partial rpoD gene sequences into 11 coherent groups in the Pseudomonas fluorescens phylogenetic lineage. Fifty-three strains that are representatives of the 11 groups were selected and analyzed by partial sequencing of their 16S rRNA and gyrB genes. The individual and concatenated partial sequences of the three genes were used to construct the corresponding phylogenetic trees. The majority of the strains were identified at the species level: P. lurida (5 strains), P. monteilii (2 strains), P. moraviensis (1 strain), P. orientalis (16 strains), P. simiae (7 strains), P. syringae (46 strains, distributed phylogenetically in at least 5 pathovars), and P. viridiflava (2 strains). This is the first report of pathogenicity on citrus of P. orientalis, P. simiae, P. lurida, P. moraviensis and P. monteilii strains. The remaining 47 strains that could not be identified at the species level are considered representatives of at least 5 putative novel Pseudomonas species that are not yet described. PMID:26919540

  7. New Pseudomonas spp. Are Pathogenic to Citrus.

    PubMed

    Beiki, Farid; Busquets, Antonio; Gomila, Margarita; Rahimian, Heshmat; Lalucat, Jorge; García-Valdés, Elena

    2016-01-01

    Five putative novel Pseudomonas species shown to be pathogenic to citrus have been characterized in a screening of 126 Pseudomonas strains isolated from diseased citrus leaves and stems in northern Iran. The 126 strains were studied using a polyphasic approach that included phenotypic characterizations and phylogenetic multilocus sequence analysis. The pathogenicity of these strains against 3 cultivars of citrus is demonstrated in greenhouse and field studies. The strains were initially grouped phenotypically and by their partial rpoD gene sequences into 11 coherent groups in the Pseudomonas fluorescens phylogenetic lineage. Fifty-three strains that are representatives of the 11 groups were selected and analyzed by partial sequencing of their 16S rRNA and gyrB genes. The individual and concatenated partial sequences of the three genes were used to construct the corresponding phylogenetic trees. The majority of the strains were identified at the species level: P. lurida (5 strains), P. monteilii (2 strains), P. moraviensis (1 strain), P. orientalis (16 strains), P. simiae (7 strains), P. syringae (46 strains, distributed phylogenetically in at least 5 pathovars), and P. viridiflava (2 strains). This is the first report of pathogenicity on citrus of P. orientalis, P. simiae, P. lurida, P. moraviensis and P. monteilii strains. The remaining 47 strains that could not be identified at the species level are considered representatives of at least 5 putative novel Pseudomonas species that are not yet described.

  8. Overexpression of a Gene Encoding a Catabolite Repression Element in Alternaria citri Causes Severe Symptoms of Black Rot in Citrus Fruit.

    PubMed

    Katoh, H; Ohtani, K; Yamamoto, H; Akimitsu, K

    2007-05-01

    ABSTRACT A gene (AcCreA) encoding a catabolite repression element (CreA) with (two zinc fingers of the Cys(2)His(2) type was isolated from the postharvest fungal pathogen Alternaria citri. The AcCreA overexpression mutant AcOEC2 of A. citri showed normal growth on pectin medium and on segments of peel or the juice sac area from citrus fruit. Production of endopolygalacturonase, an essential virulence factor of this pathogen, was similar in AcOEC2 and the wild type in pectin-containing media. However, addition of glucose to the medium showed that carbon catabolite repression of endopolygalacturonase gene (Acpg1) expression, as well as endopolygalacturonase production, was lost in AcOEC2. The wild-type strain of A. citri causes rot mainly in the central axis of citrus fruit without development of rotting in the juice sac area; however, AcOEC2 caused severe black rot symptoms in both the central axis and juice sac areas. These results indicate that AcCreA-mediated catabolite repression controls the virulence or infection of this pathogen, and that the wild-type A. citri does not cause symptoms in the juice sac area due to carbon catabolite repression by sugars in the juice of the juice sac area.

  9. A conditionally dispensable chromosome controls host-specific pathogenicity in the fungal plant pathogen Alternaria alternata.

    PubMed Central

    Hatta, Rieko; Ito, Kaoru; Hosaki, Yoshitsugu; Tanaka, Takayoshi; Tanaka, Aiko; Yamamoto, Mikihiro; Akimitsu, Kazuya; Tsuge, Takashi

    2002-01-01

    The filamentous fungus Alternaria alternata contains seven pathogenic variants (pathotypes), which produce host-specific toxins and cause diseases on different plants. Previously, the gene cluster involved in host-specific AK-toxin biosynthesis of the Japanese pear pathotype was isolated, and four genes, named AKT genes, were identified. The AKT homologs were also found in the strawberry and tangerine pathotypes, which produce AF-toxin and ACT-toxin, respectively. This result is consistent with the fact that the toxins of these pathotypes share a common 9,10-epoxy-8-hydroxy-9-methyl-decatrienoic acid structural moiety. In this study, three of the AKT homologs (AFT1-1, AFTR-1, and AFT3-1) were isolated on a single cosmid clone from strain NAF8 of the strawberry pathotype. In NAF8, all of the AKT homologs were present in multiple copies on a 1.05-Mb chromosome. Transformation-mediated targeting of AFT1-1 and AFT3-1 in NAF8 produced AF-toxin-minus, nonpathogenic mutants. All of the mutants lacked the 1.05-Mb chromosome encoding the AFT genes. This chromosome was not essential for saprophytic growth of this pathogen. Thus, we propose that a conditionally dispensable chromosome controls host-specific pathogenicity of this pathogen. PMID:12019223

  10. A conditionally dispensable chromosome controls host-specific pathogenicity in the fungal plant pathogen Alternaria alternata.

    PubMed

    Hatta, Rieko; Ito, Kaoru; Hosaki, Yoshitsugu; Tanaka, Takayoshi; Tanaka, Aiko; Yamamoto, Mikihiro; Akimitsu, Kazuya; Tsuge, Takashi

    2002-05-01

    The filamentous fungus Alternaria alternata contains seven pathogenic variants (pathotypes), which produce host-specific toxins and cause diseases on different plants. Previously, the gene cluster involved in host-specific AK-toxin biosynthesis of the Japanese pear pathotype was isolated, and four genes, named AKT genes, were identified. The AKT homologs were also found in the strawberry and tangerine pathotypes, which produce AF-toxin and ACT-toxin, respectively. This result is consistent with the fact that the toxins of these pathotypes share a common 9,10-epoxy-8-hydroxy-9-methyl-decatrienoic acid structural moiety. In this study, three of the AKT homologs (AFT1-1, AFTR-1, and AFT3-1) were isolated on a single cosmid clone from strain NAF8 of the strawberry pathotype. In NAF8, all of the AKT homologs were present in multiple copies on a 1.05-Mb chromosome. Transformation-mediated targeting of AFT1-1 and AFT3-1 in NAF8 produced AF-toxin-minus, nonpathogenic mutants. All of the mutants lacked the 1.05-Mb chromosome encoding the AFT genes. This chromosome was not essential for saprophytic growth of this pathogen. Thus, we propose that a conditionally dispensable chromosome controls host-specific pathogenicity of this pathogen.

  11. An Isolate of Alternaria alternata That Is Pathogenic to Both Tangerines and Rough Lemon and Produces Two Host-Selective Toxins, ACT- and ACR-Toxins.

    PubMed

    Masunaka, A; Ohtani, K; Peever, T L; Timmer, L W; Tsuge, T; Yamamoto, M; Yamamoto, H; Akimitsu, K

    2005-03-01

    ABSTRACT Two different pathotypes of Alternaria alternata cause Alternaria brown spot of tangerines and Alternaria leaf spot of rough lemon. The former produces the host-selective ACT-toxin and the latter produces ACR-toxin. Both pathogens induce similar symptoms on leaves or young fruits of their respective hosts, but the host ranges of these pathogens are distinct and one pathogen can be easily distinguished from another by comparing host ranges. We isolated strain BC3-5-1-OS2A from a leaf spot on rough lemon in Florida, and this isolate is pathogenic on both cv. Iyokan tangor and rough lemon and also produces both ACT-toxin and ACR-toxin. Isolate BC3-5-1-OS2A carries both genomic regions, one of which was known only to be present in ACT-toxin producers and the other was known to exist only in ACR-toxin producers. Each of the genomic regions is present on distinct small chromosomes, one of 1.05 Mb and the other of 2.0 Mb. Alternaria species have no known sexual or parasexual cycle in nature and populations of A. alternata on citrus are clonal. Therefore, the ability to produce both toxins was not likely acquired through meiotic or mitotic recombination. We hypothesize that a dispensable chromosome carrying the gene cluster controlling biosynthesis of one of the host-selective toxins was transferred horizontally and rearranged by duplication or translocation in another isolate of the fungus carrying genes for biosynthesis of the other host-selective toxin.

  12. Proteome Analysis of Pathogen-Responsive Proteins from Apple Leaves Induced by the Alternaria Blotch Alternaria alternata.

    PubMed

    Zhang, Cai-xia; Tian, Yi; Cong, Pei-hua

    2015-01-01

    Understanding the defence mechanisms used by apple leaves against Alternaria alternate pathogen infection is important for breeding purposes. To investigate the ultrastructural differences between leaf tissues of susceptible and resistant seedlings, in vitro inoculation assays and transmission electron microscopy (TEM) analysis were conducted with two different inoculation assays. The results indicated that the resistant leaves may have certain antifungal activity against A. alternate that is lacking in susceptible leaves. To elucidate the two different host responses to A. alternate infection in apples, the proteomes of susceptible and resistant apple leaves that had or had not been infected with pathogen were characterised using two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionisation time-of-flight tandem mass spectrometry (MALDI-TOF-TOF MS). MS identified 43 differentially expressed proteins in two different inoculation assays. The known proteins were categorised into 5 classes, among these proteins, some pathogenesis-related (PR) proteins, such as beta-1,3-glucanase, ascorbate peroxidase (APX), glutathione peroxidase (GPX) and mal d1, were identified in susceptible and resistant hosts and were associated with disease resistance of the apple host. In addition, the different levels of mal d1 in susceptible and resistant hosts may contribute to the outstanding anti-disease properties of resistant leaves against A. alternate. Taken together, the resistance mechanisms of the apple host against A. alternate may be a result of the PR proteins and other defence-related proteins. Given the complexity of the biology involved in the interaction between apple leaves and the A. alternate pathogen, further investigation will yield more valuable insights into the molecular mechanisms of suppression of the A. alternate pathogen. Overall, we outline several novel insights into the response of apple leaves to pathogen attacks. These findings

  13. Proteome Analysis of Pathogen-Responsive Proteins from Apple Leaves Induced by the Alternaria Blotch Alternaria alternata

    PubMed Central

    Zhang, Cai-xia; Tian, Yi; Cong, Pei-hua

    2015-01-01

    Understanding the defence mechanisms used by apple leaves against Alternaria alternate pathogen infection is important for breeding purposes. To investigate the ultrastructural differences between leaf tissues of susceptible and resistant seedlings, in vitro inoculation assays and transmission electron microscopy (TEM) analysis were conducted with two different inoculation assays. The results indicated that the resistant leaves may have certain antifungal activity against A. alternate that is lacking in susceptible leaves. To elucidate the two different host responses to A. alternate infection in apples, the proteomes of susceptible and resistant apple leaves that had or had not been infected with pathogen were characterised using two-dimensional electrophoresis (2-DE) and matrix-assisted laser desorption/ionisation time-of-flight tandem mass spectrometry (MALDI-TOF-TOF MS). MS identified 43 differentially expressed proteins in two different inoculation assays. The known proteins were categorised into 5 classes, among these proteins, some pathogenesis-related (PR) proteins, such as beta-1,3-glucanase, ascorbate peroxidase (APX), glutathione peroxidase (GPX) and mal d1, were identified in susceptible and resistant hosts and were associated with disease resistance of the apple host. In addition, the different levels of mal d1 in susceptible and resistant hosts may contribute to the outstanding anti-disease properties of resistant leaves against A. alternate. Taken together, the resistance mechanisms of the apple host against A. alternate may be a result of the PR proteins and other defence-related proteins. Given the complexity of the biology involved in the interaction between apple leaves and the A. alternate pathogen, further investigation will yield more valuable insights into the molecular mechanisms of suppression of the A. alternate pathogen. Overall, we outline several novel insights into the response of apple leaves to pathogen attacks. These findings

  14. How the Pathogenic Fungus Alternaria alternata Copes with Stress via the Response Regulators SSK1 and SHO1

    PubMed Central

    Yu, Pei-Ling; Chen, Li-Hung; Chung, Kuang-Ren

    2016-01-01

    The tangerine pathotype of Alternaria alternata is a necrotrophic fungal pathogen causing brown spot disease on a number of citrus cultivars. To better understand the dynamics of signal regulation leading to oxidative and osmotic stress response and fungal infection on citrus, phenotypic characterization of the yeast SSK1 response regulator homolog was performed. It was determined that SSK1 responds to diverse environmental stimuli and plays a critical role in fungal pathogenesis. Experiments to determine the phenotypes resulting from the loss of SSK1 reveal that the SSK1 gene product may be fulfilling similar regulatory roles in signaling pathways involving a HOG1 MAP kinase during ROS resistance, osmotic resistance, fungicide sensitivity and fungal virulence. The SSK1 mutants display elevated sensitivity to oxidants, fail to detoxify H2O2 effectively, induce minor necrosis on susceptible citrus leaves, and displays resistance to dicarboximide and phenylpyrrole fungicides. Unlike the SKN7 response regulator, SSK1 and HOG1 confer resistance to salt-induced osmotic stress via an unknown kinase sensor rather than the “two component” histidine kinase HSK1. SSK1 and HOG1 play a moderate role in sugar-induced osmotic stress. We also show that SSK1 mutants are impaired in their ability to produce germ tubes from conidia, indicating a role for the gene product in cell differentiation. SSK1 also is involved in multi-drug resistance. However, deletion of the yeast SHO1 (synthetic high osmolarity) homolog resulted in no noticeable phenotypes. Nonetheless, our results show that A. alternata can sense and react to different types of stress via SSK1, HOG1 and SKN7 in a cooperative manner leading to proper physiological and pathological functions. PMID:26863027

  15. How the Pathogenic Fungus Alternaria alternata Copes with Stress via the Response Regulators SSK1 and SHO1.

    PubMed

    Yu, Pei-Ling; Chen, Li-Hung; Chung, Kuang-Ren

    2016-01-01

    The tangerine pathotype of Alternaria alternata is a necrotrophic fungal pathogen causing brown spot disease on a number of citrus cultivars. To better understand the dynamics of signal regulation leading to oxidative and osmotic stress response and fungal infection on citrus, phenotypic characterization of the yeast SSK1 response regulator homolog was performed. It was determined that SSK1 responds to diverse environmental stimuli and plays a critical role in fungal pathogenesis. Experiments to determine the phenotypes resulting from the loss of SSK1 reveal that the SSK1 gene product may be fulfilling similar regulatory roles in signaling pathways involving a HOG1 MAP kinase during ROS resistance, osmotic resistance, fungicide sensitivity and fungal virulence. The SSK1 mutants display elevated sensitivity to oxidants, fail to detoxify H2O2 effectively, induce minor necrosis on susceptible citrus leaves, and displays resistance to dicarboximide and phenylpyrrole fungicides. Unlike the SKN7 response regulator, SSK1 and HOG1 confer resistance to salt-induced osmotic stress via an unknown kinase sensor rather than the "two component" histidine kinase HSK1. SSK1 and HOG1 play a moderate role in sugar-induced osmotic stress. We also show that SSK1 mutants are impaired in their ability to produce germ tubes from conidia, indicating a role for the gene product in cell differentiation. SSK1 also is involved in multi-drug resistance. However, deletion of the yeast SHO1 (synthetic high osmolarity) homolog resulted in no noticeable phenotypes. Nonetheless, our results show that A. alternata can sense and react to different types of stress via SSK1, HOG1 and SKN7 in a cooperative manner leading to proper physiological and pathological functions.

  16. Developing cryotherapy to eliminate graft-transmissible pathogens in citrus

    USDA-ARS?s Scientific Manuscript database

    This article summarizes research being conducted as part of a project funded by the California Citrus Research Board to develop cryotherapy (freezing buds in liquid nitrogen, and then recovering them) as a viable method for elimination of graft transmissible pathogens from Citrus. There are current...

  17. The development of Alternaria alternata is prevented by chitinases and beta-1,3-glucanases from Citrus limon seedlings.

    PubMed

    Fanta, Nora; Ortega, Ximena; Pérez, Luz M

    2003-01-01

    In addition to phytoalexin synthesis, the defense response of intact Citrus limon seedlings against Alternaria alternata involves both constitutive and induced enzyme activities such as chitinases (Ch) and beta-1,3-glucanases (Glu). A alternata conidial germination was prevented by protein extracts from inoculated lemon seedlings, but also by extracts from mock-inoculated specimens. On the other hand, degradation of mycelia was accomplished only by protein extracts from inoculated seedlings. The presence of six Ch isoenzymes and of four Glu isoenzymes was detected in protein extracts from mock-inoculated seedlings. As a result of fungal inoculation, the isoenzyme pattern of Ch and Glu changed, making possible the detection of a new Ch isoenzyme and of three new Glu. Also, some constitutive Ch and Glu increased their enzyme activity, and those Ch that increased their activity also showed a broadening of their substrate specificity. These changes were prevented by alpha-amanitin and cycloheximide, suggesting that the presence of new Ch and Glu isoenzymes was due to de novo synthesis of proteins. Results suggest that constitutive Ch and Glu could act as pre-formed defense molecules in Citrus limon preventing A. alternata germination, while those induced after fungal inoculation of lemon seedlings could act along with the former, to produce lysis of fungal mycelia, resulting in a more efficient control of A. alternata development.

  18. Genetically Based Location from Triploid Populations and Gene Ontology of a 3.3-Mb Genome Region Linked to Alternaria Brown Spot Resistance in Citrus Reveal Clusters of Resistance Genes

    PubMed Central

    Cuenca, José; Aleza, Pablo; Vicent, Antonio; Brunel, Dominique; Ollitrault, Patrick; Navarro, Luis

    2013-01-01

    Genetic analysis of phenotypical traits and marker-trait association in polyploid species is generally considered as a challenge. In the present work, different approaches were combined taking advantage of the particular genetic structures of 2n gametes resulting from second division restitution (SDR) to map a genome region linked to Alternaria brown spot (ABS) resistance in triploid citrus progeny. ABS in citrus is a serious disease caused by the tangerine pathotype of the fungus Alternaria alternata. This pathogen produces ACT-toxin, which induces necrotic lesions on fruit and young leaves, defoliation and fruit drop in susceptible genotypes. It is a strong concern for triploid breeding programs aiming to produce seedless mandarin cultivars. The monolocus dominant inheritance of susceptibility, proposed on the basis of diploid population studies, was corroborated in triploid progeny. Bulk segregant analysis coupled with genome scan using a large set of genetically mapped SNP markers and targeted genetic mapping by half tetrad analysis, using SSR and SNP markers, allowed locating a 3.3 Mb genomic region linked to ABS resistance near the centromere of chromosome III. Clusters of resistance genes were identified by gene ontology analysis of this genomic region. Some of these genes are good candidates to control the dominant susceptibility to the ACT-toxin. SSR and SNP markers were developed for efficient early marker-assisted selection of ABS resistant hybrids. PMID:24116149

  19. Genetically based location from triploid populations and gene ontology of a 3.3-mb genome region linked to Alternaria brown spot resistance in citrus reveal clusters of resistance genes.

    PubMed

    Cuenca, José; Aleza, Pablo; Vicent, Antonio; Brunel, Dominique; Ollitrault, Patrick; Navarro, Luis

    2013-01-01

    Genetic analysis of phenotypical traits and marker-trait association in polyploid species is generally considered as a challenge. In the present work, different approaches were combined taking advantage of the particular genetic structures of 2n gametes resulting from second division restitution (SDR) to map a genome region linked to Alternaria brown spot (ABS) resistance in triploid citrus progeny. ABS in citrus is a serious disease caused by the tangerine pathotype of the fungus Alternaria alternata. This pathogen produces ACT-toxin, which induces necrotic lesions on fruit and young leaves, defoliation and fruit drop in susceptible genotypes. It is a strong concern for triploid breeding programs aiming to produce seedless mandarin cultivars. The monolocus dominant inheritance of susceptibility, proposed on the basis of diploid population studies, was corroborated in triploid progeny. Bulk segregant analysis coupled with genome scan using a large set of genetically mapped SNP markers and targeted genetic mapping by half tetrad analysis, using SSR and SNP markers, allowed locating a 3.3 Mb genomic region linked to ABS resistance near the centromere of chromosome III. Clusters of resistance genes were identified by gene ontology analysis of this genomic region. Some of these genes are good candidates to control the dominant susceptibility to the ACT-toxin. SSR and SNP markers were developed for efficient early marker-assisted selection of ABS resistant hybrids.

  20. Pathogen infection drives patterns of nutrient resorption in citrus plants

    PubMed Central

    Cao, Jirong; Cheng, Chunzhen; Yang, Junjie; Wang, Qibing

    2015-01-01

    Nutrient resorption processes in the plants infected by pathogen remain poorly understood. Huanglongbing (HLB) is a destructive disease of citrus. HLB-pathogen ‘Candidatus Liberibacter asiaticus’ grows specifically in the phloem of hosts and may cause problems in the plant vascular system after infection. Therefore, it brings a great concern about the phloem nutrient transport and nutrient intra-cycling in HLB-affected plants. We investigated the effects of ‘Ca. L. asiaticus’ infection on nitrogen (N) and phosphorus (P) concentrations and resorption in different citrus species (i.e. Citrus reticulata, Citrus limon and Citrus maxima). HLB-pathogen infection had distinctive impacts on nutrient resorption in different species. P resorption efficiency substantially decreased in infected C. reticulata plants relative to the healthy plants in summer, which may account for the marked decrease in the average fruit yield. P resorption was more efficient in infected C. limon plants than in the healthy plants. However, for C. maxima plants, HLB had no significant effects on N:P ratio in live leaves and resorption efficiency as well as on fruit yield. Keeping efficient internal nutrient cycling can be a strategy of citrus species being tolerant to HLB. PMID:26419510

  1. Pathogen infection drives patterns of nutrient resorption in citrus plants.

    PubMed

    Cao, Jirong; Cheng, Chunzhen; Yang, Junjie; Wang, Qibing

    2015-09-30

    Nutrient resorption processes in the plants infected by pathogen remain poorly understood. Huanglongbing (HLB) is a destructive disease of citrus. HLB-pathogen 'Candidatus Liberibacter asiaticus' grows specifically in the phloem of hosts and may cause problems in the plant vascular system after infection. Therefore, it brings a great concern about the phloem nutrient transport and nutrient intra-cycling in HLB-affected plants. We investigated the effects of 'Ca. L. asiaticus' infection on nitrogen (N) and phosphorus (P) concentrations and resorption in different citrus species (i.e. Citrus reticulata, Citrus limon and Citrus maxima). HLB-pathogen infection had distinctive impacts on nutrient resorption in different species. P resorption efficiency substantially decreased in infected C. reticulata plants relative to the healthy plants in summer, which may account for the marked decrease in the average fruit yield. P resorption was more efficient in infected C. limon plants than in the healthy plants. However, for C. maxima plants, HLB had no significant effects on N:P ratio in live leaves and resorption efficiency as well as on fruit yield. Keeping efficient internal nutrient cycling can be a strategy of citrus species being tolerant to HLB.

  2. Enhancement of the citrus immune system provides effective resistance against Alternaria brown spot disease.

    PubMed

    Llorens, Eugenio; Fernández-Crespo, Emma; Vicedo, Begonya; Lapeña, Leonor; García-Agustín, Pilar

    2013-01-15

    In addition to basal defense mechanisms, plants are able to develop enhanced defense mechanisms such as induced resistance (IR) upon appropriate stimulation. We recently described the means by which several carboxylic acids protect Arabidopsis and tomato plants against fungi. In this work, we demonstrate the effectiveness of hexanoic acid (Hx) in the control of Alternaria brown spot (ABS) disease via enhancement of the immune system of Fortune mandarin. The application of 1mM Hx in irrigation water to 2-year-old Fortune plants clearly reduced the incidence of the disease and led to smaller lesions. We observed that several of the most important mechanisms involved in induced resistance were affected by Hx application. Our results demonstrate enhanced callose deposition in infected plants treated with Hx, which suggests an Hx priming mechanism. Plants treated with the callose inhibitor 2-DDG were more susceptible to the fungus. Moreover, polygalacturonase-inhibiting protein (PGIP) gene expression was rapidly and significantly upregulated in treated plants. However, treatment with Hx decreased the levels of reactive oxygen species (ROS) in infected plants. Hormonal and gene analyses revealed that the jasmonic acid (JA) pathway was activated due to a greater accumulation of 12-oxo-phytodienoic acid (OPDA) and JA along with a rapid accumulation of JA-isoleucine (JA-Ile). Furthermore, we observed a more rapid accumulation of abscisic acid (ABA), which could act as a positive regulator of callose deposition. Thus, our results support the hypothesis that both enhanced physical barriers and the JA signaling pathway are involved in hexanoic acid-induced resistance (Hx-IR) to Alternaria alternata. Copyright © 2012 Elsevier GmbH. All rights reserved.

  3. Characterization and pathogenicity of Alternaria spp. strains associated with grape bunch rot during post-harvest withering.

    PubMed

    Lorenzini, Marilinda; Zapparoli, Giacomo

    2014-09-01

    Alternaria is a fungal agent of grape bunch rot which occurs during withering, a process which produces passito style wines. Seven isolates of Alternaria spp. were characterized using morphological examination, genotypic analysis and pathogenicity. Six of these isolates produced conidiophores and conidia displaying sporulation patterns typical of the Alternaria alternata species-group. Variability in colony morphology and growth on different media was observed. Phylogenetic analysis of internal transcribed spacer (ITS) sequences clustered all isolates within a monophyletic clade, while intergenic spacer region (IGS)-RFLP profiles were congruent with those of A. alternata and Alternaria arborescens. RAPD-PCR proved helpful in discriminating between strains. To assay strain pathogenicity, grape berries were infected while undergoing withering conditions at different temperatures. Disease capacity was found to be strain dependent and varied consistently between the most and least aggressive strains. This study has provided interesting information on polymorphism within Alternaria spp. populations in withered grapes and on understanding the saprophytic role of this fungus during the post-harvest dehydrating process. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Role of mannitol metabolism in the pathogenicity of the necrotrophic fungus Alternaria brassicicola

    PubMed Central

    Calmes, Benoit; Guillemette, Thomas; Teyssier, Lény; Siegler, Benjamin; Pigné, Sandrine; Landreau, Anne; Iacomi, Béatrice; Lemoine, Rémi; Richomme, Pascal; Simoneau, Philippe

    2013-01-01

    In this study, the physiological functions of fungal mannitol metabolism in the pathogenicity and protection against environmental stresses were investigated in the necrotrophic fungus Alternaria brassicicola. Mannitol metabolism was examined during infection of Brassica oleracea leaves by sequential HPLC quantification of the major soluble carbohydrates and expression analysis of genes encoding two proteins of mannitol metabolism, i.e., a mannitol dehydrogenase (AbMdh), and a mannitol-1-phosphate dehydrogenase (AbMpd). Knockout mutants deficient for AbMdh or AbMpd and a double mutant lacking both enzyme activities were constructed. Their capacity to cope with various oxidative and drought stresses and their pathogenic behavior were evaluated. Metabolic and gene expression profiling indicated an increase in mannitol production during plant infection. Depending on the mutants, distinct pathogenic processes, such as leaf and silique colonization, sporulation, survival on seeds, were impaired by comparison to the wild-type. This pathogenic alteration could be partly explained by the differential susceptibilities of mutants to oxidative and drought stresses. These results highlight the importance of mannitol metabolism with respect to the ability of A. brassicicola to efficiently accomplish key steps of its pathogenic life cycle. PMID:23717316

  5. Jasmonic acid signalling mediates resistance of the wild tobacco Nicotiana attenuata to its native Fusarium, but not Alternaria, fungal pathogens.

    PubMed

    Luu, Van Thi; Schuck, Stefan; Kim, Sang-Gyu; Weinhold, Arne; Baldwin, Ian T

    2015-03-01

    We recently characterized a highly dynamic fungal disease outbreak in native populations of Nicotiana attenuata in the southwestern United States. Here, we explore how phytohormone signalling contributes to the observed disease dynamics. Single inoculation with three native Fusarium and Alternaria fungal pathogens, isolated from diseased plants growing in native populations, resulted in disease symptoms characteristic for each pathogen species. While Alternaria sp.-infected plants displayed fewer symptoms and recovered, Fusarium spp.-infected plants became chlorotic and frequently spontaneously wilted. Jasmonic acid (JA) and salicylic acid (SA) levels were differentially induced after Fusarium or Alternaria infection. Transgenic N. attenuata lines silenced in JA production or JA conjugation to isoleucine (JA-Ile), but not in JA perception, were highly susceptible to infection by F. brachygibbosum Utah 4, indicating that products derived from the JA-Ile biosynthetic pathway, but not their perception, is associated with increased Fusarium resistance. Infection assays using ov-nahG plants which were silenced in pathogen-induced SA accumulations revealed that SA may increase N. attenuata's resistance to Fusarium infection but not to Alternaria. Taken together, we propose that the dynamics of fungal disease symptoms among plants in native populations may be explained by a complex interplay of phytohormone responses to attack by multiple pathogens. © 2014 John Wiley & Sons Ltd.

  6. Phytotoxin solanapyrone A produced by Ascochyta rabiei and Alternaria solani is nonessential for pathogenicity, but likely plays ecological roles

    USDA-ARS?s Scientific Manuscript database

    Ascochyta rabiei and Alternaria solani, causal agents of chickpea and potato blights respectively, produce the same phytotoxin solanapyrone A (SolA).The toxicity of SolA to plants has been documented, but its role in pathogenicity has not been investigated. In this study, we generated solanapyrone-d...

  7. Host-selective toxins produced by the plant pathogenic fungus Alternaria alternata.

    PubMed

    Tsuge, Takashi; Harimoto, Yoshiaki; Akimitsu, Kazuya; Ohtani, Kouhei; Kodama, Motoichiro; Akagi, Yasunori; Egusa, Mayumi; Yamamoto, Mikihiro; Otani, Hiroshi

    2013-01-01

    Host-selective toxins (HSTs) produced by fungal plant pathogens are generally low-molecular-weight secondary metabolites with a diverse range of structures that function as effectors controlling pathogenicity or virulence in certain plant-pathogen interactions. There are now seven known diseases caused by Alternaria alternata in which HSTs are responsible for fungal pathogenesis. The pathogens have been defined as pathotypes of A. alternata because of morphological similarity but pathological differences. Chemical structures of HSTs from six pathotypes have been determined. The role of A. alternata HSTs in pathogenesis has been studied extensively, and discovery of the release of HSTs from germinating conidia prior to penetration aids in understanding the early participation of HSTs to induce susceptibility of host cells by suppressing their defence reactions. Many attempts have been made to find the target sites of A. alternata HSTs, and four cellular components, plasma membrane, mitochondrion, chloroplast and a metabolically important enzyme, have been identified as the primary sites of each HST action, leading to elucidation of the molecular mechanisms of HST sensitivity in host plants. Studies of the molecular genetics of HST production have identified supernumerary chromosomes encoding HST gene clusters and have provided new insights into the evolution of A. alternata pathotypes.

  8. Nuclear Ribosomal DNA Variation and Pathogenic Specialization in Alternaria Fungi Known To Produce Host-Specific Toxins †

    PubMed Central

    Kusaba, Motoaki; Tsuge, Takashi

    1994-01-01

    A total of 99 strains of 11 Alternaria species, including 68 strains of seven fungi known to produce host-specific toxins, were subjected to analysis of restriction fragment length polymorphism (RFLP) in nuclear ribosomal DNA (rDNA). Total DNA was digested with XbaI, and the Southern blots were probed with a nuclear rDNA clone of Alternaria kikuchiana. The hybridization gave 17 different RFLPs from the 99 strains. On the basis of these RFLPs, populations of host-specific toxin-producing fungi could not be differentiated from one another nor from nonpathogenic A. alternata. Each population of the toxin-producing fungi carried rDNA variants. Nine different types, named A1 to A6 and B1 to B3, were detected among the toxin-producing fungi and nonpathogenic A. alternata. All of the populations contained the type A4 variant, and the other rDNA types were also shared by different toxin-producing fungi and A. alternata. In contrast, Alternaria species that are morphologically distinguishable from A. alternata could be differentiated from A. alternata on the basis of the rDNA RFLPs. Polymorphisms in rDNA digested with HaeIII and MspI were also evaluated in 61 Alternaria strains. These restriction enzymes produced 31 variations among all of the samples. The seven toxin-producing fungi and nonpathogenic A. alternata could not be resolved by phylogenetic analysis based on the RFLPs, although they could be differentiated from the other Alternaria species studied. These results provide support for the hypothesis that Alternaria fungi known to produce host-specific toxins are intraspecific variants of A. alternata specialized in pathogenicity. Images PMID:16349367

  9. Complex regulation of secondary metabolism controlling pathogenicity in the phytopathogenic fungus Alternaria alternata.

    PubMed

    Takaoka, Shinya; Kurata, Mariko; Harimoto, Yoshiaki; Hatta, Rieko; Yamamoto, Mikihiro; Akimitsu, Kazuya; Tsuge, Takashi

    2014-06-01

    The filamentous fungus Alternaria alternata includes seven pathogenic variants (pathotypes), which produce different host-selective toxins and cause disease on different plants. The Japanese pear, strawberry and tangerine pathotypes produce AK-toxin, AF-toxin and ACT-toxin, respectively, which have a common structural moiety, 9,10-epoxy-8-hydroxy-9-methyl-decatrienoic acid (EDA). Here, we identified a new gene, AKT7 (AK-toxin biosynthetic gene 7), from the Japanese pear pathotype, which encodes a cytochrome P450 monooxygenase and functions to limit AK-toxin production. AKT7 homologs were found in the strawberry pathotype, but not the tangerine pathotype. However, the strawberry pathotype homolog appeared to include a premature stop codon. Although the Japanese pear pathotype strain has multiple copies of AKT7, a single-copy disruption resulted in mutants with increased production of AK-toxin and EDA. AKT7 overexpression in the three pathotypes caused marked reductions of toxin and EDA production, suggesting that Akt7 catalyzes a side reaction of EDA or its precursor. AKT7 overexpression caused reduced virulence in these pathotypes. We also found that AKT7 transcripts predominantly include misspliced mRNAs, which have premature stop codons. Our observations suggest that the AK-toxin production required for full virulence is regulated in a complex way by the copy number and intron information content of AKT7. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  10. Characterization of glutathione transferases involved in the pathogenicity of Alternaria brassicicola.

    PubMed

    Calmes, Benoit; Morel-Rouhier, Mélanie; Bataillé-Simoneau, Nelly; Gelhaye, Eric; Guillemette, Thomas; Simoneau, Philippe

    2015-06-18

    Glutathione transferases (GSTs) represent an extended family of multifunctional proteins involved in detoxification processes and tolerance to oxidative stress. We thus anticipated that some GSTs could play an essential role in the protection of fungal necrotrophs against plant-derived toxic metabolites and reactive oxygen species that accumulate at the host-pathogen interface during infection. Mining the genome of the necrotrophic Brassica pathogen Alternaria brassicicola for glutathione transferase revealed 23 sequences, 17 of which could be clustered into the main classes previously defined for fungal GSTs and six were 'orphans'. Five isothiocyanate-inducible GSTs from five different classes were more thoroughly investigated. Analysis of their catalytic properties revealed that two GSTs, belonging to the GSTFuA and GTT1 classes, exhibited GSH transferase activity with isothiocyanates (ITC) and peroxidase activity with cumene hydroperoxide, respectively. Mutant deficient for these two GSTs were however neither more susceptible to ITC nor less aggressive than the wild-type parental strain. By contrast mutants deficient for two other GSTs, belonging to the Ure2pB and GSTO classes, were distinguished by their hyper-susceptibility to ITC and low aggressiveness against Brassica oleracea. In particular AbGSTO1 could participate in cell tolerance to ITC due to its glutathione-dependent thioltransferase activity. The fifth ITC-inducible GST belonged to the MAPEG class and although it was not possible to produce the soluble active form of this protein in a bacterial expression system, the corresponding deficient mutant failed to develop normal symptoms on host plant tissues. Among the five ITC-inducible GSTs analyzed in this study, three were found essential for full aggressiveness of A. brassicicola on host plant. This, to our knowledge is the first evidence that GSTs might be essential virulence factors for fungal necrotrophs.

  11. Sexual Reproduction in the Citrus Black Spot Pathogen, Phyllosticta citricarpa.

    PubMed

    Tran, Nga T; Miles, Andrew K; Dietzgen, Ralf G; Dewdney, Megan M; Zhang, Ke; Rollins, Jeffrey A; Drenth, André

    2017-06-01

    Citrus black spot (Phyllosticta citricarpa) causes fruit blemishes and premature fruit drop, resulting in significant economic losses in citrus growing areas with summer rainfall across the globe. The mating type locus of P. citricarpa has recently been characterized, revealing the heterothallic nature of this pathogen. However, insight into the occurrence of mating and the impact of completing the sexual cycle of P. citricarpa was lacking. To investigate the occurrence and impact of sexual reproduction, we developed a method to reliably, and for the first time, produce ascospores of P. citricarpa on culture media. To demonstrate meiosis during the mating process, we identified recombinant genotypes through multilocus genotyping of single ascospores. Because the process of fertilization was not well understood, we experimentally determined that fertilization of P. citricarpa occurs via spermatization. Our results demonstrate that P. citricarpa is heterothallic and requires isolates of different MAT idiomorphs to be in direct physical contact, or for spermatia to fulfill their role as male elements to fertilize the receptive organs, in order to initiate the mating process. The impact of mating on the epidemiology of citrus black spot in the field is discussed.

  12. Citrus tristeza virus: a pathogen that changed the course of the citrus industry.

    PubMed

    Moreno, Pedro; Ambrós, Silvia; Albiach-Martí, Maria R; Guerri, José; Peña, Leandro

    2008-03-01

    shaping CTV populations. Measures to control CTV damage include quarantine and budwood certification programmes, elimination of infected trees, use of tristeza-tolerant rootstocks, or cross protection with mild isolates, depending on CTV incidence and on the virus strains and host varieties predominant in each region. Incorporating resistance genes into commercial varieties by conventional breeding is presently unfeasible, whereas incorporation of pathogen-derived resistance by plant transformation has yielded variable results, indicating that the CTV-citrus interaction may be more specific and complex than initially thought. A deep understanding of the interactions between viral proteins and host and vector factors will be necessary to develop reliable and sound control measures.

  13. Contribution of Peroxisomes to Secondary Metabolism and Pathogenicity in the Fungal Plant Pathogen Alternaria alternata ▿ †

    PubMed Central

    Imazaki, Ai; Tanaka, Aiko; Harimoto, Yoshiaki; Yamamoto, Mikihiro; Akimitsu, Kazuya; Park, Pyoyun; Tsuge, Takashi

    2010-01-01

    The filamentous fungus Alternaria alternata includes seven pathogenic variants (pathotypes) which produce different host-selective toxins and cause diseases on different plants. The Japanese pear pathotype produces the host-selective AK-toxin, an epoxy-decatrienoic acid ester, and causes black spot of Japanese pear. Previously, we identified four genes, AKT1, AKT2, AKT3, and AKTR, involved in AK toxin biosynthesis. AKT1, AKT2, and AKT3 encode enzyme proteins with peroxisomal targeting signal type 1 (PTS1)-like tripeptides, SKI, SKL, and PKL, respectively, at the C-terminal ends. In this study, we verified the peroxisome localization of Akt1, Akt2, and Akt3 by using strains expressing N-terminal green fluorescent protein (GFP)-tagged versions of the proteins. To assess the role of peroxisome function in AK-toxin production, we isolated AaPEX6, which encodes a peroxin protein essential for peroxisome biogenesis, from the Japanese pear pathotype and made AaPEX6 disruption-containing transformants from a GFP-Akt1-expressing strain. The ΔAaPEX6 mutant strains did not grow on fatty acid media because of a defect in fatty acid β oxidation. The import of GFP-Akt1 into peroxisomes was impaired in the ΔAaPEX6 mutant strains. These strains completely lost AK toxin production and pathogenicity on susceptible pear leaves. These data show that peroxisomes are essential for AK-toxin biosynthesis. The ΔAaPEX6 mutant strains showed a marked reduction in the ability to cause lesions on leaves of a resistant pear cultivar with defense responses compromised by heat shock. This result suggests that peroxisome function is also required for plant invasion and tissue colonization in A. alternata. We also observed that mutation of AaPEX6 caused a marked reduction of conidiation. PMID:20348386

  14. PAMPs, PRRs, effectors and R-genes associated with citrus-pathogen interactions.

    PubMed

    Dalio, Ronaldo J D; Magalhães, Diogo M; Rodrigues, Carolina M; Arena, Gabriella D; Oliveira, Tiago S; Souza-Neto, Reinaldo R; Picchi, Simone C; Martins, Paula M M; Santos, Paulo J C; Maximo, Heros J; Pacheco, Inaiara S; De Souza, Alessandra A; Machado, Marcos A

    2017-03-01

    Recent application of molecular-based technologies has considerably advanced our understanding of complex processes in plant-pathogen interactions and their key components such as PAMPs, PRRs, effectors and R-genes. To develop novel control strategies for disease prevention in citrus, it is essential to expand and consolidate our knowledge of the molecular interaction of citrus plants with their pathogens. This review provides an overview of our understanding of citrus plant immunity, focusing on the molecular mechanisms involved in the interactions with viruses, bacteria, fungi, oomycetes and vectors related to the following diseases: tristeza, psorosis, citrus variegated chlorosis, citrus canker, huanglongbing, brown spot, post-bloom, anthracnose, gummosis and citrus root rot.

  15. Effect of Alternaria solani exudates on resistant and susceptible potato cultivars from two different pathogen isolates

    USDA-ARS?s Scientific Manuscript database

    The resistance phenotypes of two potato cultivars to two isolates of Alternaria solani, causal agent of early blight, were studied under greenhouse conditions. The two isolates contain varying degrees of aggressiveness on both susceptible and resistant phenotypes of potatoes. A bioassay was used to ...

  16. The biochemical basis of pathogenicity and host-specificity of Penicillium digitatum on citrus

    USDA-ARS?s Scientific Manuscript database

    In this work, we report that volatiles emitted from wounded citrus peel play a major role in host recognition by Penicillium digitatum. Volatiles of various citrus cultivars had a pronounced stimulatory effect on germination and germ tube elongation of green mold pathogen. When exposed to volatile...

  17. Multiplexed lateral flow microarray assay for detection of citrus pathogens Xylella fastidiosa and Xanthomonas axonopodis pv citri

    SciTech Connect

    Cary,; Bruce, R; Stubben, Christopher J

    2011-03-22

    The invention provides highly sensitive and specific assays for the major citrus pathogens Xylella fastidiosa and Xanthomonas axonopodis, including a field deployable multiplexed assay capable of rapidly assaying for both pathogens simultaneously. The assays are directed at particular gene targets derived from pathogenic strains that specifically cause the major citrus diseases of citrus variegated chlorosis (Xylella fastidiosa 9a5c) and citrus canker (Xanthomonas axonopodis pv citri). The citrus pathogen assays of the invention offer femtomole sensitivity, excellent linear dynamic range, and rapid and specific detection.

  18. Characterization and Pathogenicity of Alternaria burnsii from Seeds of Cucurbita maxima (Cucurbitaceae) in Bangladesh.

    PubMed

    Paul, Narayan Chandra; Deng, Jian Xin; Lee, Hyang Burm; Yu, Seung-Hun

    2015-12-01

    In the course of survey of endophytic fungi from Bangladesh pumpkin seeds in 2011~2012, two strains (CNU111042 and CNU111043) with similar colony characteristics were isolated and characterized by their morphology and by molecular phylogenetic analysis of the internal transcribed spacer, glyceraldehydes-3-phosphate dehydrogenase (gpd), and Alternaria allergen a1 (Alt a1) sequences. Phylogenetic analysis of all three sequences and their combined dataset revealed that the fungus formed a subclade within the A. alternata clade, matching A. burnsi and showing differences with its other closely related Alternaria species, such as A. longipes, A. tomato, and A. tomaticola. Long ellipsoid, obclavate or ovoid beakless conidia, shorter and thinner conidial size (16~60 [90] × 6.5~14 [~16] µm) distinguish this fungus from other related species. These isolates showed more transverse septation (2~11) and less longitudinal septation (0~3) than did other related species. Moreover, the isolate did not produce any diffusible pigment on media. Therefore, our results reveal that the newly recorded fungus from a new host, Cucurbita maxima, is Alternaria burnsii Uppal, Patel & Kamat.

  19. Characterization and Pathogenicity of Alternaria burnsii from Seeds of Cucurbita maxima (Cucurbitaceae) in Bangladesh

    PubMed Central

    Paul, Narayan Chandra; Deng, Jian Xin

    2015-01-01

    In the course of survey of endophytic fungi from Bangladesh pumpkin seeds in 2011~2012, two strains (CNU111042 and CNU111043) with similar colony characteristics were isolated and characterized by their morphology and by molecular phylogenetic analysis of the internal transcribed spacer, glyceraldehydes-3-phosphate dehydrogenase (gpd), and Alternaria allergen a1 (Alt a1) sequences. Phylogenetic analysis of all three sequences and their combined dataset revealed that the fungus formed a subclade within the A. alternata clade, matching A. burnsi and showing differences with its other closely related Alternaria species, such as A. longipes, A. tomato, and A. tomaticola. Long ellipsoid, obclavate or ovoid beakless conidia, shorter and thinner conidial size (16~60 [90] × 6.5~14 [~16] µm) distinguish this fungus from other related species. These isolates showed more transverse septation (2~11) and less longitudinal septation (0~3) than did other related species. Moreover, the isolate did not produce any diffusible pigment on media. Therefore, our results reveal that the newly recorded fungus from a new host, Cucurbita maxima, is Alternaria burnsii Uppal, Patel & Kamat. PMID:26839497

  20. Environmental Factors Affecting Production, Release, and Field Populations of Conidia of Alternaria alternata, the Cause of Brown Spot of Citrus.

    PubMed

    Timmer, L W; Solel, Z; Gottwald, T R; Ibañez, A M; Zitko, S E

    1998-11-01

    ABSTRACT Alternaria brown spot, caused by Alternaria alternata pv. citri, affects many tangerines and their hybrids, causing loss of immature leaves and fruit and reducing the marketability of the remaining fruit. Conidial production of A. alternata was greatest on mature leaves moistened and maintained at near 100% relative humidity (RH) for 24 h, whereas leaves that had been soaked or maintained at moderate RH produced few conidia. Conidial release from filter paper cultures and infected leaves was studied in a computer-controlled environmental chamber. Release of large numbers of conidia was triggered from both substrates by sudden drops in RH or by simulated rainfall events. Vibration induced release of low numbers of conidia, but red/infrared irradiation had no effect. In field studies from 1994 to 1996, air sampling with a 7-day recording volumetric spore trap indicated that conidia were present throughout the year with periodic large peaks. The number of conidia captured was not closely related to rainfall amounts or average wind speed, but was weakly related to the duration of leaf wetness. Likewise, disease severity on trap plants placed in the field weekly during 1995 to 1996 was not closely related to conidial numbers or rainfall amounts, but was weakly related to leaf wetness duration. Sufficient inoculum appears to be available to allow infection to occur throughout the year whenever susceptible host tissue and moisture are available.

  1. Biology and management of Asian citrus psyllid, vector of the huanglongbing pathogens.

    PubMed

    Grafton-Cardwell, Elizabeth E; Stelinski, Lukasz L; Stansly, Philip A

    2013-01-01

    The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), is the most important pest of citrus worldwide because it serves as a vector of "Candidatus Liberibacter" species (Alphaproteobacteria) that cause huanglongbing (citrus greening disease). All commercially cultivated citrus is susceptible and varieties tolerant to disease expression are not yet available. Onset of disease occurs following a long latent period after inoculation, and thus the pathogen can spread widely prior to detection. Detection of the pathogen in Brazil in 2004 and Florida in 2005 catalyzed a significant increase in research on D. citri biology. Chemical control is the primary management strategy currently employed, but recently documented decreases in susceptibility of D. citri to several insecticides illustrate the need for more sustainable tools. Herein, we discuss recent advances in the understanding of D. citri biology and behavior, pathogen transmission biology, biological control, and chemical control with respect to "Candidatus Liberibacter asiaticus." Our goal is to point toward integrated and biologically relevant management of this pathosystem.

  2. Discovery of a viral pathogen in the Asian citrus psyllid

    USDA-ARS?s Scientific Manuscript database

    We used a Metagenomics approach and discovered an insect-infecting virus in adult Asian citrus psyllids in Florida. Though wide spread in nature, this is the first report of a Fijivirus in North America. The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae) is a small insect tha...

  3. Fungal-specific transcription factor AbPf2 activates pathogenicity in Alternaria brassicicola

    SciTech Connect

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

    2013-05-24

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

  4. Alternaria section Alternaria: Species, formae speciales or pathotypes?

    PubMed Central

    Woudenberg, J.H.C.; Seidl, M.F.; Groenewald, J.Z.; de Vries, M.; Stielow, J.B.; Thomma, B.P.H.J.; Crous, P.W.

    2015-01-01

    The cosmopolitan fungal genus Alternaria consists of multiple saprophytic and pathogenic species. Based on phylogenetic and morphological studies, the genus is currently divided into 26 sections. Alternaria sect. Alternaria contains most of the small-spored Alternaria species with concatenated conidia, including important plant, human and postharvest pathogens. Species within sect. Alternaria have been mostly described based on morphology and / or host-specificity, yet molecular variation between them is minimal. To investigate whether the described morphospecies within sect. Alternaria are supported by molecular data, whole-genome sequencing of nine Alternaria morphospecies supplemented with transcriptome sequencing of 12 Alternaria morphospecies as well as multi-gene sequencing of 168 Alternaria isolates was performed. The assembled genomes ranged in size from 33.3–35.2 Mb within sect. Alternaria and from 32.0–39.1 Mb for all Alternaria genomes. The number of repetitive sequences differed significantly between the different Alternaria genomes; ranging from 1.4–16.5 %. The repeat content within sect. Alternaria was relatively low with only 1.4–2.7 % of repeats. Whole-genome alignments revealed 96.7–98.2 % genome identity between sect. Alternaria isolates, compared to 85.1–89.3 % genome identity for isolates from other sections to the A. alternata reference genome. Similarly, 1.4–2.8 % and 0.8–1.8 % single nucleotide polymorphisms (SNPs) were observed in genomic and transcriptomic sequences, respectively, between isolates from sect. Alternaria, while the percentage of SNPs found in isolates from different sections compared to the A. alternata reference genome was considerably higher; 8.0–10.3 % and 6.1–8.5 %. The topology of a phylogenetic tree based on the whole-genome and transcriptome reads was congruent with multi-gene phylogenies based on commonly used gene regions. Based on the genome and transcriptome data, a set of core

  5. Protein interaction networks at the host-microbe interface in Diaphorina citri, the insect vector of the citrus greening pathogen

    USDA-ARS?s Scientific Manuscript database

    The Asian citrus psyllid (Diaphorina citri) is the insect vector responsible for the worldwide spread of Candidatus Liberibacter asiaticus, the bacterial pathogen associated with citrus greening disease. Developmental changes in the insect vector impact pathogen transmission, such that D. citri tra...

  6. Rapid screening for citrus canker resistance employing pathogen-associated molecular pattern-triggered immunity responses

    PubMed Central

    Pitino, Marco; Armstrong, Cheryl M; Duan, Yongping

    2015-01-01

    Citrus canker, caused by the bacterial pathogen Xanthomonas citri ssp. citri (Xcc), has been attributed to millions of dollars in loss or damage to commercial citrus crops in subtropical production areas of the world. Since identification of resistant plants is one of the most effective methods of disease management, the ability to screen for resistant seedlings plays a key role in the production of a long-term solution to canker. Here, an inverse correlation between reactive oxygen species (ROS) production by the plant and the ability of Xcc to grow and form lesions on infected plants is reported. Based on this information, a novel screening method that can rapidly identify citrus seedlings that are less susceptible to early infection by Xcc was devised by measuring ROS accumulation triggered by a 22-amino acid sequence of the conserved N-terminal part of flagellin (flg22) from X. citri ssp. citri (Xcc-flg22). In addition to limiting disease symptoms, ROS production was also correlated with the expression of basal defense-related genes such as the pattern recognition receptors LRR8 and FLS2, the leucine-rich repeat receptor-like protein RLP12, and the defense-related gene PR1, indicating an important role for pathogen-associated molecular pattern-triggered immunity (PTI) in determining resistance to citrus canker. Moreover, the differential expression patterns observed amongst the citrus seedlings demonstrated the existence of genetic variations in the PTI response among citrus species/varieties. PMID:26504581

  7. Genetic variability of Brazilian isolates of Alternaria alternata detected by AFLP and RAPD techniques

    PubMed Central

    Dini-Andreote, Francisco; Pietrobon, Vivian Cristina; Andreote, Fernando Dini; Romão, Aline Silva; Spósito, Marcel Bellato; Araújo, Welington Luiz

    2009-01-01

    The Alternaria brown spot (ABS) is a disease caused in tangerine plants and its hybrids by the fungus Alternaria alternata f. sp. citri which has been found in Brazil since 2001. Due to the recent occurrence in Brazilian orchards, the epidemiology and genetic variability of this pathogen is still an issue to be addressed. Here it is presented a survey about the genetic variability of this fungus by the characterization of twenty four pathogenic isolates of A. alternata f. sp. citri from citrus plants and four endophytic isolates from mango (one Alternaria tenuissima and three Alternaria arborescens). The application of two molecular markers Random Amplified Polymorphic DNA (RAPD) and Amplified Fragment Length Polymorphism (AFLP) had revealed the isolates clustering in distinct groups when fingerprintings were analyzed by Principal Components Analysis (PCA). Despite the better assessment of the genetic variability through the AFLP, significant modifications in clusters components were not observed, and only slight shifts in the positioning of isolates LRS 39/3 and 25M were observed in PCA plots. Furthermore, in both analyses, only the isolates from lemon plants revealed to be clustered, differently from the absence of clustering for other hosts or plant tissues. Summarizing, both RAPD and AFLP analyses were both efficient to detect the genetic variability within the population of the pathogenic fungus Alternaria spp., supplying information on the genetic variability of this species as a basis for further studies aiming the disease control. PMID:24031413

  8. Genetic variability of Brazilian isolates of Alternaria alternata detected by AFLP and RAPD techniques.

    PubMed

    Dini-Andreote, Francisco; Pietrobon, Vivian Cristina; Andreote, Fernando Dini; Romão, Aline Silva; Spósito, Marcel Bellato; Araújo, Welington Luiz

    2009-07-01

    The Alternaria brown spot (ABS) is a disease caused in tangerine plants and its hybrids by the fungus Alternaria alternata f. sp. citri which has been found in Brazil since 2001. Due to the recent occurrence in Brazilian orchards, the epidemiology and genetic variability of this pathogen is still an issue to be addressed. Here it is presented a survey about the genetic variability of this fungus by the characterization of twenty four pathogenic isolates of A. alternata f. sp. citri from citrus plants and four endophytic isolates from mango (one Alternaria tenuissima and three Alternaria arborescens). The application of two molecular markers Random Amplified Polymorphic DNA (RAPD) and Amplified Fragment Length Polymorphism (AFLP) had revealed the isolates clustering in distinct groups when fingerprintings were analyzed by Principal Components Analysis (PCA). Despite the better assessment of the genetic variability through the AFLP, significant modifications in clusters components were not observed, and only slight shifts in the positioning of isolates LRS 39/3 and 25M were observed in PCA plots. Furthermore, in both analyses, only the isolates from lemon plants revealed to be clustered, differently from the absence of clustering for other hosts or plant tissues. Summarizing, both RAPD and AFLP analyses were both efficient to detect the genetic variability within the population of the pathogenic fungus Alternaria spp., supplying information on the genetic variability of this species as a basis for further studies aiming the disease control.

  9. Draft Genome Sequence of 11399, a Transformable Citrus-Pathogenic Strain of Xylella fastidiosa

    PubMed Central

    Niza, Bárbara; Merfa, Marcus V.; Alencar, Valquíria C.; Menegidio, Fabiano B.; Nunes, Luiz R.; Machado, Marcos A.; Takita, Marco A.

    2016-01-01

    The draft genome of Xylella fastidiosa subsp. pauca strain 11399, a transformable citrus-pathogenic strain, is reported here. The 11399 genome size is 2,690,704 bp and has a G+C content of 52.7%. The draft genome of 11399 reveals the absence of four type I restriction-modification system genes. PMID:27738038

  10. Effect of citrus pulp on the viability of Saccharomyces boulardii in the presence of enteric pathogens

    USDA-ARS?s Scientific Manuscript database

    Saccharomyces cerevisiae subtype boulardii is frequently used as a dietary supplement to promote intestinal health and reduce the impact of growth of enteric pathogens in livestock, including cattle and swine. Citrus by-products are also fed as dietary supplements that have the additional benefit o...

  11. Substrate specificity and inhibition of brassinin hydrolases, detoxifying enzymes from the plant pathogens Leptosphaeria maculans and Alternaria brassicicola.

    PubMed

    Pedras, M Soledade C; Minic, Zoran; Sarma-Mamillapalle, Vijay K

    2009-12-01

    Blackleg (Leptosphaeria maculans and Leptosphaeria biglobosa) and black spot (Alternaria brassicicola) fungi are devastating plant pathogens known to detoxify the plant defence metabolite, brassinin. The significant roles of brassinin as a crucifer phytoalexin and as a biosynthetic precursor of several other plant defences make it important in plant fitness. Brassinin detoxifying enzymes produced by L. maculans and A. brassicicola catalyse the detoxification of brassinin by hydrolysis of its dithiocarbamate group to indolyl-3-methanamine. The purification and characterization of brassinin hydrolases produced by L. maculans (BHLmL2) and A. brassicicola (BHAb) were accomplished: native BHLmL2 was found to be a tetrameric protein with a molecular mass of 220 kDa, whereas native BHAb was found to be a dimeric protein of 120 kDa. Protein characterization using LC-MS/MS and sequence alignment analyses suggested that both enzymes belong to the family of amidases with the catalytic Ser/Ser/Lys triad. Furthermore, chemical modification of BHLmL2 and BHAb with selective reagents suggested that the amino acid serine was involved in the catalytic activity of both enzymes. The overall results indicated that BHs have new substrate specificities with a new catalytic activity that can be designated as dithiocarbamate hydrolase. Investigation of the effect of various phytoalexins on the activities of BHLmL2 and BHAb indicated that cyclobrassinin was a competitive inhibitor of both enzymes. On the basis of pH dependence, sequence analyses, chemical modifications of amino acid residues and identification of headspace volatiles, a chemical mechanism for hydrolysis of the dithiocarbamate group of brassinin catalysed by BHLmL2 and BHAb is proposed. The current information should facilitate the design of specific synthetic inhibitors of these enzymes for plant treatments against blackleg and black spot fungal infections.

  12. Inhibitory activity of Indian spice plant Cinnamomum zeylanicum extracts against Alternaria solani and Curvularia lunata, the pathogenic dematiaceous moulds.

    PubMed

    Mishra, Ajay K; Mishra, Amita; Kehri, H K; Sharma, Bechan; Pandey, Abhay K

    2009-03-07

    Dematiaceous moulds are pathogenic microorganisms and act as etiological agents of mycoses with different degrees of severity in humans and animals. These moulds also cause loss of food crops and storage food products. The information regarding antimicrobial efficacy of the plant preparations on these moulds is scanty. The present study reveals phytochemical characterization and the effect of bark and leaf extracts of Indian spice plant, Cinnamomum zeylanicum (Cz), against the growth of two species of dematiaceous moulds, Alternaria solani and Curvularia lunata. Cz bark and leaf samples were sequentially extracted in different solvents using Soxhlet apparatus. Phytochemical analyses of extracts were done as per standard protocols. The antifungal bioassay of extracts was done by hanging drop technique. The inhibition of fungal spore germination was monitored under influence of three different concentrations of extracts. The lowest test concentration (50 microg/ml) of extracts of Cz bark prepared into acetone and that of Cz leaf into petroleum ether and ethanol exhibited complete inhibition (100%) of spore germination in both the moulds. At 100 microg/ml concentration all the extracts showed about 50 to 100% inhibition. However, the treatment of the spores of the two fungal species with highest concentration (500 microg/ml) of bark and leaf extracts in all the solvents showed 100% fungicidal activity as it completely arrested the germination of spores. Relatively lower activity of aqueous extracts at 50 and 100 microg/ml concentrations suggests that the antifungal ingredients present in Cz bark and leaf are more soluble in organic solvents than water. The results demonstrated that the Cz bark and leaves contain certain fungicidal constituents exhibiting potential antimould activity against A. solani and C. lunata.

  13. Inhibitory activity of Indian spice plant Cinnamomum zeylanicum extracts against Alternaria solani and Curvularia lunata, the pathogenic dematiaceous moulds

    PubMed Central

    Mishra, Ajay K; Mishra, Amita; Kehri, HK; Sharma, Bechan; Pandey, Abhay K

    2009-01-01

    Background Dematiaceous moulds are pathogenic microorganisms and act as etiological agents of mycoses with different degrees of severity in humans and animals. These moulds also cause loss of food crops and storage food products. The information regarding antimicrobial efficacy of the plant preparations on these moulds is scanty. The present study reveals phytochemical characterization and the effect of bark and leaf extracts of Indian spice plant, Cinnamomum zeylanicum (Cz), against the growth of two species of dematiaceous moulds, Alternaria solani and Curvularia lunata. Methods Cz bark and leaf samples were sequentially extracted in different solvents using Soxhlet apparatus. Phytochemical analyses of extracts were done as per standard protocols. The antifungal bioassay of extracts was done by hanging drop technique. The inhibition of fungal spore germination was monitored under influence of three different concentrations of extracts. Results The lowest test concentration (50 μg/ml) of extracts of Cz bark prepared into acetone and that of Cz leaf into petroleum ether and ethanol exhibited complete inhibition (100%) of spore germination in both the moulds. At 100 μg/ml concentration all the extracts showed about 50 to 100% inhibition. However, the treatment of the spores of the two fungal species with highest concentration (500 μg/ml) of bark and leaf extracts in all the solvents showed 100% fungicidal activity as it completely arrested the germination of spores. Relatively lower activity of aqueous extracts at 50 and 100 μg/ml concentrations suggests that the antifungal ingredients present in Cz bark and leaf are more soluble in organic solvents than water. Conclusion The results demonstrated that the Cz bark and leaves contain certain fungicidal constituents exhibiting potential antimould activity against A. solani and C. lunata. PMID:19267932

  14. Validation of high-throughput real time polymerase chain reaction assays for simultaneous detection of invasive citrus pathogens

    USDA-ARS?s Scientific Manuscript database

    A number of economically important citrus pathogens are spread by nursery propagation, arthropod vector transmission and in advertent importation and dissemination of infected plants. For these reasons, citrus disease management and clean stock programs need to employ an economical and sensitive p...

  15. Structural and biochemical characteristics of citrus flowers associated with defence against a fungal pathogen

    PubMed Central

    Marques, João Paulo Rodrigues; Amorim, Lilian; Silva-Junior, Geraldo José; Spósito, Marcel Bellato; Appezzato-da Gloria, Beatriz

    2015-01-01

    The constitutive characters of plants can be structural or biochemical and play an important role in their defence against pathogens. Citrus postbloom fruit drop (PFD) caused by Colletotrichum spp. is one of the most important fungal diseases of citrus. The pathogen infects the flowers, leading to premature fruit drop and reducing citrus production. However, flower buds smaller than 8 mm long are usually not infected by Colletotrichum spp. Thus, this study investigated whether there are constitutive mechanisms in flower buds related to Colletotrichum spp. infection. We studied flower buds that were 2, 3, 4, 8, 12 and 15 mm long and petals, after anthesis, of sweet orange ‘Valência’ using light and scanning electron microscopy and histochemistry. We evaluated the effect of volatile organic compounds (VOCs) in flowers (R-limonene and linalool) on the in vitro growth of Colletotrichum acutatum. We found that the arrangement of the epidermal papillae in the petal primordia, the occurrence of prismatic crystals and the distribution of oil glands are the main differences between buds smaller than 8 mm and buds 8–15 mm long. Osmophores at the tips of petals produced and accumulated phenols, terpenes and lipophilic compounds. Flower buds smaller than 8 mm long have constitutive structural and biochemical barriers to Colletotrichum spp. infection. In addition, this is the first time that osmophores have been reported in citrus. Our study shows that natural terpenes of Citrus flowers inhibit the fungal growth in vitro, highlighting the potential use of terpenes for the chemical control of PFD in citrus. PMID:25535209

  16. Role of the host-selective ACT-toxin synthesis gene ACTTS2 encoding an enoyl-reductase in pathogenicity of the tangerine pathotype of Alternaria alternata.

    PubMed

    Ajiro, Naoya; Miyamoto, Yoko; Masunaka, Akira; Tsuge, Takashi; Yamamoto, Mikihiro; Ohtani, Kouhei; Fukumoto, Takeshi; Gomi, Kenji; Peever, Tobin L; Izumi, Yuriko; Tada, Yasuomi; Akimitsu, Kazuya

    2010-02-01

    ABSTRACT The tangerine pathotype of Alternaria alternata produces host-selective ACT-toxin and causes Alternaria brown spot disease of tangerines and tangerine hybrids. Sequence analysis of a genomic BAC clone identified a previously uncharacterized portion of the ACT-toxin biosynthesis gene cluster (ACTT). A 1,034-bp gene encoding a putative enoyl-reductase was identified by using rapid amplification of cDNA ends and polymerase chain reaction and designated ACTTS2. Genomic Southern blots demonstrated that ACTTS2 is present only in ACT-toxin producers and is carried on a 1.9 Mb conditionally dispensable chromosome by the tangerine pathotype. Targeted gene disruption of ACTTS2 led to a reduction in ACT-toxin production and pathogenicity, and transcriptional knockdown of ACTTS2 using RNA silencing resulted in complete loss of ACT-toxin production and pathogenicity. These results indicate that ACTTS2 is an essential gene for ACT-toxin biosynthesis in the tangerine pathotype of A. alternata and is required for pathogenicity of this fungus.

  17. ACTTS3 encoding a polyketide synthase is essential for the biosynthesis of ACT-toxin and pathogenicity in the tangerine pathotype of Alternaria alternata.

    PubMed

    Miyamoto, Y; Masunaka, A; Tsuge, T; Yamamoto, M; Ohtani, K; Fukumoto, T; Gomi, K; Peever, T L; Tada, Y; Ichimura, K; Akimitsu, K

    2010-04-01

    The tangerine pathotype of Alternaria alternata produces host-selective ACT-toxin and causes Alternaria brown spot disease of tangerine and tangerine hybrids. Sequence analysis of a genomic BAC clone identified part of the ACT-toxin TOX (ACTT) gene cluster, and knockout experiments have implicated several open reading frames (ORF) contained within the cluster in the biosynthesis of ACT-toxin. One of the ORF, designated ACTTS3, encoding a putative polyketide synthase, was isolated by rapid amplification of cDNA ends and genomic/reverse transcription-polymerase chain reactions using the specific primers designed from the BAC sequences. The 7,374-bp ORF encodes a polyketide synthase with putative beta-ketoacyl synthase, acyltransferase, methyltransferase, beta-ketoacyl reductase, and phosphopantetheine attachment site domains. Genomic Southern blots demonstrated that ACTTS3 is present on the smallest chromosome in the tangerine pathotype of A. alternata, and the presence of ACTTS3 is highly correlated with ACT-toxin production and pathogenicity. Targeted gene disruption of two copies of ACTTS3 led to a complete loss of ACT-toxin production and pathogenicity. These results indicate that ACTTS3 is an essential gene for ACT-toxin biosynthesis in the tangerine pathotype of A. alternata and is required for pathogenicity of this fungus.

  18. Citrus phenylpropanoids and defence against pathogens. Part I: metabolic profiling in elicited fruits.

    PubMed

    Ballester, Ana-Rosa; Lafuente, M Teresa; de Vos, Ric C H; Bovy, Arnaud G; González-Candelas, Luis

    2013-01-01

    Penicillium spp. are among the major postharvest pathogens of citrus fruit. Induction of natural resistance in fruits constitutes one of the alternatives to chemical fungicides. Here, we investigated the involvement of the phenylpropanoid pathway in the induction of resistance in Navelate oranges by examining changes in the metabolic profile of upon eliciting citrus fruits. By using both HPLC-PDA-FD and HPLC-PDA-QTOF-MS allowed the identification of several compounds that seem to be relevant for induced resistance. In elicited fruits, a greater diversity of phenolic compounds was observed in the flavedo (outer coloured part of the peel) when compared to the albedo (inner white part). Moreover, only small changes were detected in the most abundant citrus flavonoids. The coumarin scoparone was among the compounds with the highest induction upon elicitation. Two other highly induced compounds were identified as citrusnin A and drupanin aldehyde. All three compounds are known to exert antimicrobial activity. Our results suggest that phenylpropanoids and their derivatives play an important role in the induction of resistance in citrus fruit. Copyright © 2012 Elsevier Ltd. All rights reserved.

  19. Metabolomic analysis of citrus infection by 'Candidatus Liberibacter' reveals insight into pathogenicity.

    PubMed

    Slisz, Anne M; Breksa, Andrew P; Mishchuk, Darya O; McCollum, Greg; Slupsky, Carolyn M

    2012-08-03

    Huanglongbing (HLB), considered the most serious citrus disease in the world, is associated with the nonculturable bacterium 'Candidatus Liberibacter asiaticus' (Las). Infection of citrus by this pathogen leads to reduced plant vigor and productivity, ultimately resulting in death of the infected tree. It can take up to two years following initial infection before outward symptoms become apparent, making detection difficult. The existing knowledge gap in our understanding of Las and its pathogenesis leading to HLB has stymied development of treatments and methods to mitigate the pathogen's influence. To evaluate the influence of Las on fruit quality in both symptomatic and asymptomatic fruit, and gain further insight into the pathogenesis of the disease, a 1H NMR metabolomics investigation, complemented with physicochemical and analyte-specific analyses, was undertaken. Comparison of the juice obtained from oranges gathered from Las+ (symptomatic and asymptomatic) and Las- (healthy) trees revealed significant differences in the concentrations of sugars, amino and organic acids, limonin glucoside, and limonin. This study demonstrates differing metabolic profiles in the juice of oranges from Las+ and Las- and proposes how Las may be able to evade citrus defense responses.

  20. Responsiveness of different citrus genotypes to the Xanthomonas citri ssp. citri-derived pathogen-associated molecular pattern (PAMP) flg22 correlates with resistance to citrus canker.

    PubMed

    Shi, Qingchun; Febres, Vicente J; Jones, Jeffrey B; Moore, Gloria A

    2015-06-01

    The bacterial agent of citrus canker disease (Xanthomonas citri ssp. citri, Xcc) has caused tremendous economic losses to the citrus industry around the world. Pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) is important to plant immunity. In this study, we compared the defence responses of citrus canker-resistant and citrus canker-susceptible genotypes to the Xcc-derived PAMP flg22 (Xflg22) by analysing the expression of 20 citrus defence-associated genes. We showed that, in the most resistant genotype, 'Nagami' kumquat, there was significant induction of several defence genes (EDS1, NDR1, PBS1, RAR1, SGT1, PAL1, NPR2 and NPR3) as early as 6 h and up to 72 h after Xflg22 treatment. At the other end of the spectrum, highly susceptible 'Duncan' grapefruit showed no induction of the same defence genes, even 120 h after treatment. Citrus genotypes with partial levels of resistance showed intermediate levels of transcriptional reprogramming that correlated with their resistance level. Xflg22 also triggered a rapid oxidative burst in all genotypes which was higher and accompanied by the induction of PTI marker genes (WRKY22 and GST1) only in the more resistant genotypes. Pretreatment with Xflg22 prior to Xcc inoculation inhibited bacterial growth in kumquat, but not in grapefruit. A flagellin-deficient Xcc strain (XccΔfliC) showed greater growth increase relative to wild-type Xcc in kumquat than in grapefruit. Taken together, our results indicate that Xflg22 initiates strong PTI in canker-resistant genotypes, but not in susceptible ones, and that a robust induction of PTI is an important component of citrus resistance to canker.

  1. Validation of high-throughput real time polymerase chain reaction assays for simultaneous detection of invasive citrus pathogens.

    PubMed

    Saponari, Maria; Loconsole, Giuliana; Liao, Hui-Hong; Jiang, Bo; Savino, Vito; Yokomi, Raymond K

    2013-11-01

    A number of important citrus pathogens are spread by graft propagation, arthropod vector transmission and inadvertent import and dissemination of infected plants. For these reasons, citrus disease management and clean stock programs require pathogen detection systems which are economical and sensitive to maintain a healthy industry. To this end, multiplex quantitative real-time PCR (qPCR) assays were developed allowing high-throughput and simultaneous detection of some major invasive citrus pathogens. Automated high-throughput extraction comparing several bead-based commercial extraction kits were tested and compared with tissue print and manual extraction to obtain nucleic acids from healthy and pathogen-infected citrus trees from greenhouse in planta collections and field. Total nucleic acids were used as templates for pathogen detection. Multiplex reverse transcription-qPCR (RT-qPCR) assays were developed for simultaneous detection of six targets including a virus, two viroids, a bacterium associated with huanglongbing and a citrus RNA internal control. Specifically, two one-step TaqMan-based multiplex RT-qPCR assays were developed and tested with target templates to determine sensitivity and detection efficiency. The first assay included primers and probes for 'Candidatus Liberibacter asiaticus' (CLas) and Citrus tristeza virus (CTV) broad spectrum detection and genotype differentiation (VT- and T3-like genotypes). The second assay contained primers and probes for Hop stunt viroid (HSVd), Citrus exocortis viroid (CEVd) and the mitochondrial NADH dehydrogenase (nad5) mRNA as an internal citrus host control. Primers and TaqMan probes for the viroids were designed in this work; whereas those for the other pathogens were from reports of others. Based on quantitation cycle values, automated high-throughput extraction of samples proved to be as suitable as manual extraction. The multiplex RT-qPCR assays detected both RNA and DNA pathogens in the same dilution series

  2. Identification and characterization of chitin synthase genes in the postharvest citrus fruit pathogen Penicillium digitatum.

    PubMed

    Gandía, Mónica; Harries, Eleonora; Marcos, Jose F

    2012-06-01

    In this study, we carried out the isolation and characterization of chitin synthase genes (CHS) of the main citrus fruit postharvest pathogen Penicillium digitatum. Using distinct sets of degenerate primers designed from conserved regions of CHS genes of yeast and filamentous fungi, PCR methods, and a DNA genomic library, five putative CHS genes (PdigCHSI, PdigCHSII, PdigCHSIII, PdigCHSV, and PdigCHSVII) were identified, isolated, sequenced, and characterized. Phylogenetic analyses, sequence identity, and domain conservation support the annotation as CHS. A very high sequence identity and strong synteny were found with corresponding regions from the genome of Penicillium chrysogenum. Gene expression of P. digitatum CHS genes during mycelium axenic growth, under oxidative and osmotic stress conditions, and during infection of citrus fruits was confirmed and quantified using quantitative RT-PCR (qRT-PCR). PdigCHSIII had the highest expression among the five genes by one order of magnitude, while PdigCHSII had the lowest. However, PdigCHSII was strongly induced coincident with conidial production, suggesting a role in conidiogenesis. The expression of PdigCHSI, PdigCHSIII, PdigCHSV, and PdigCHSVII was upregulated during infection of citrus fruit. PdigCHSV and PdigCHSVII coexpressed in most of the experiments carried out, and they are separated by a 1.77 kb intergenic region and arranged in opposite directions.

  3. Diaphorina citri (Hemiptera: Liviidae) Vector Competence for the Citrus Greening Pathogen 'Candidatus Liberibacter Asiaticus'.

    PubMed

    Tabachnick, Walter J

    2015-06-01

    Characterizing the vector competence of Diaphorina citri Kuwayama for 'Candidatus Liberibacter asiaticus,' the pathogen causing citrus greening, is essential for understanding the epidemiology of this disease that is threatening the U.S. citrus industry. Vector competence studies have been difficult because of the biology of D. citri, the inability to culture the pathogen, and the available diagnostic methods used to detect the bacteria in plant and insect tissues. The methods employed in many studies of D. citri vector competence may have overestimated amounts of live 'Ca. L. asiaticus' in both plant and insect tissues, and it is possible that the amounts of phloem ingested by psyllids may not contain sufficient detectable pathogen using current diagnostic methods. As a result of the difficulty in characterizing D. citri vector competence, the several daunting challenges for providing D. citri that are unable to inoculate 'Ca. L. asiaticus', as a novel method to control greening are discussed. Suggestions to overcome some of these challenges are provided. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  4. Synthesis of Oxylipin Mimics and Their Antifungal Activity against the Citrus Postharvest Pathogens.

    PubMed

    Ma, Jimei; Li, Yupeng; Chen, Hangwei; Zeng, Zhen; Li, Zi-Long; Jiang, Hong

    2016-02-22

    Nine oxylipin mimics were designed and synthesized starting from d-mannose. Their antifungal activity against three citrus postharvest pathogens was evaluated by spore germination assay. The results indicated that all the compounds significantly inhibited the growth of Penicillium digitatum, Penicillium italicum and Aspergillus niger. The compound (3Z,6Z,8S,9R,10R)-octadeca-3,6-diene-8,9,10-triol (3) exhibited excellent inhibitory effect on both Penicillium digitatum (IC50 = 34 ppm) and Penicillium italicum (IC50 = 94 ppm). Their in vivo antifungal activities against citrus postharvest blue mold were tested with fruit inoculated with the pathogen Penicillium italicum. The compound (3R,4S)-methyl 3,4-dihydroxy-5-octyltetrahydrofuran-2-carboxylate (9) demonstrated significant efficacy by reducing the disease severity to 60%. The antifungal mechanism of these oxylipin mimics was postulated in which both inhibition of pathogenic mycelium and stimuli of the host oxylipin-mediated defense response played important roles.

  5. Prophage-mediated dynamics of 'Candidatus Liberibacter asiaticus' populations, the destructive bacterial pathogens of citrus huanglongbing.

    PubMed

    Zhou, Lijuan; Powell, Charles A; Li, Wenbin; Irey, Mike; Duan, Yongping

    2013-01-01

    Prophages are highly dynamic components in the bacterial genome and play an important role in intraspecies variations. There are at least two prophages in the chromosomes of Candidatus Liberibacter asiaticus' (Las) Floridian isolates. Las is both unculturable and the most prevalent species of Liberibacter pathogens that cause huanglongbing (HLB), a worldwide destructive disease of citrus. In this study, seven new prophage variants resulting from two hyper-variable regions were identified by screening clone libraries of infected citrus, periwinkle and psyllids. Among them, Types A and B share highly conserved sequences and localize within the two prophages, FP1 and FP2, respectively. Although Types B and C were abundant in all three libraries, Type A was much more abundant in the libraries from the Las-infected psyllids than from the Las-infected plants, and Type D was only identified in libraries from the infected host plants but not from the infected psyllids. Sequence analysis of these variants revealed that the variations may result from recombination and rearrangement events. Conventional PCR results using type-specific molecular markers indicated that A, B, C and D are the four most abundant types in Las-infected citrus and periwinkle. However, only three types, A, B and C are abundant in Las-infected psyllids. Typing results for Las-infected citrus field samples indicated that mixed populations of Las bacteria present in Floridian isolates, but only the Type D population was correlated with the blotchy mottle symptom. Extended cloning and sequencing of the Type D region revealed a third prophage/phage in the Las genome, which may derive from the recombination of FP1 and FP2. Dramatic variations in these prophage regions were also found among the global Las isolates. These results are the first to demonstrate the prophage/phage-mediated dynamics of Las populations in plant and insect hosts, and their correlation with insect transmission and disease development.

  6. Endophytic and pathogenic Phyllosticta species, with reference to those associated with Citrus Black Spot.

    PubMed

    Glienke, C; Pereira, O L; Stringari, D; Fabris, J; Kava-Cordeiro, V; Galli-Terasawa, L; Cunnington, J; Shivas, R G; Groenewald, J Z; Crous, P W

    2011-06-01

    We investigated the identity and genetic diversity of more than 100 isolates belonging to Phyllosticta (teleomorph Guignardia), with particular emphasis on Phyllosticta citricarpa and Guignardia mangiferae s.l. occurring on Citrus. Phyllosticta citricarpa is the causal agent of Citrus Black Spot and is subject to phytosanitary legislation in the EU. This species is frequently confused with a taxon generally referred to as G. mangiferae, the presumed teleomorph of P. capitalensis, which is a non-pathogenic endophyte, commonly isolated from citrus leaves and fruits and a wide range of other hosts. DNA sequence analysis of the nrDNA internal transcribed spacer region (ITS1, 5.8S nrDNA, ITS2) and partial translation elongation factor 1-alpha (TEF1), actin and glyceraldehyde-3-phosphate dehydrogenase (GPDH) genes resolved nine clades correlating to seven known, and two apparently undescribed species. Phyllosticta citribraziliensis is newly described as an endophytic species occurring on Citrus in Brazil. An epitype is designated for P. citricarpa from material newly collected in Australia, which is distinct from P. citriasiana, presently only known on C. maxima from Asia. Phyllosticta bifrenariae is newly described for a species causing leaf and bulb spots on Bifrenaria harrisoniae (Orchidaceae) in Brazil. It is morphologically distinct from P. capitalensis, which was originally described from Stanhopea (Orchidaceae) in Brazil; an epitype is designated here. Guignardia mangiferae, which was originally described from Mangifera indica (Anacardiaceae) in India, is distinguished from the non-pathogenic endophyte, P. brazilianiae sp. nov., which is common on M. indica in Brazil. Furthermore, a combined phylogenetic tree revealed the P. capitalensis s.l. clade to be genetically distinct from the reference isolate of G. mangiferae. Several names are available for this clade, the oldest being P. capitalensis. These results suggest that endophytic, non-pathogenic isolates

  7. Insecticidal suppression of Asian citrus psyllid Diaphorina citri (Hemiptera: Liviidae) vector of huanglongbing pathogens.

    PubMed

    Qureshi, Jawwad A; Kostyk, Barry C; Stansly, Philip A

    2014-01-01

    Diaphorina citri vectors pathogens that cause 'huanglongbing' or citrus greening disease which poses a serious threat to citrus production worldwide. Vector suppression is critical to reduce disease spread. Efficacy is a main concern when choosing an insecticide. Insecticidal treatments of 49 products or 44 active ingredients (a.i) labeled or experimental were field tested between 2005-2013 as foliar sprays (250 treatments, 39 a.i) or soil applications (47 treatments, 9 a.i) to control D. citri in citrus. A combined effect of nymphal and adult suppression in response to sprays of 23 insecticides representing 9 modes of action (MoA) groups and 3 unknown MoA provided more than 90% reduction of adult D. citri over 24-68 days. Observable effects on nymphs were generally of shorter duration due to rapid maturation of flush. However, reduction of 76-100% nymphs or adults over 99-296 days was seen on young trees receiving drenches of the neonicotinoids imidacloprid, thiamethoxam or clothianidin (MoA 4A) and a novel anthranilic diamide, cyantraniliprole (MoA 28). Effective products identified for foliar sprays to control D. citri provide sufficient MoA groups for rotation to delay evolution of insecticide resistance by D. citri and other pests. However, cyantraniliprole is now the only available alternative for rotation with neonicotinoids in soil application to young trees. Sprays of up to eight of the most effective insecticides could be rotated over a year without repetition of any MoA and little or no recourse to neonicotinoids or cyantraniliprole, so important for protection of young trees. Other considerations effecting decisions of what and when to spray include prevalence of huanglongbing, pest pressure, pre-harvest intervals, overall budget, equipment availability, and conservation of beneficial arthropods. Examples of spray programs utilizing broad-spectrum and relatively selective insecticides are provided to improve vector management and may vary depending on

  8. Identification of Novel Source of Resistance and Differential Response of Allium Genotypes to Purple Blotch Pathogen, Alternaria porri (Ellis) Ciferri

    PubMed Central

    Nanda, Satyabrata; Chand, Subodh Kumar; Mandal, Purander; Tripathy, Pradyumna; Joshi, Raj Kumar

    2016-01-01

    Purple blotch, caused by Alternaria porri (Ellis) Cifferi, is a serious disease incurring heavy yield losses in the bulb and seed crop of onion and garlic worldwide. There is an immediate need for identification of effective resistance sources for use in host resistance breeding. A total of 43 Allium genotypes were screened for purple blotch resistance under field conditions. Allium cepa accession ‘CBT-Ac77’ and cultivar ‘Arka Kalyan’ were observed to be highly resistant. In vitro inoculation of a selected set of genotypes with A. porri, revealed that 7 days after inoculation was suitable to observe the disease severity. In vitro screening of 43 genotypes for resistance to A. porri revealed two resistant lines. An additional 14 genotypes showed consistent moderate resistance in the field as well as in vitro evaluations. Among the related Allium species, A. schoenoprasum and A. roylei showed the least disease index and can be used for interspecific hybridization with cultivated onion. Differential reaction analysis of three A. porri isolates (Apo-Chiplima, Apn-Nasik, Apg-Guntur) in 43 genotypes revealed significant variation among the evaluated Allium species (P = 0.001). All together, the present study suggest that, the newly identified resistance sources can be used as potential donors for ongoing purple blotch resistance breeding program in India. PMID:27904458

  9. Identification of Novel Source of Resistance and Differential Response of Allium Genotypes to Purple Blotch Pathogen, Alternaria porri (Ellis) Ciferri.

    PubMed

    Nanda, Satyabrata; Chand, Subodh Kumar; Mandal, Purander; Tripathy, Pradyumna; Joshi, Raj Kumar

    2016-12-01

    Purple blotch, caused by Alternaria porri (Ellis) Cifferi, is a serious disease incurring heavy yield losses in the bulb and seed crop of onion and garlic worldwide. There is an immediate need for identification of effective resistance sources for use in host resistance breeding. A total of 43 Allium genotypes were screened for purple blotch resistance under field conditions. Allium cepa accession 'CBT-Ac77' and cultivar 'Arka Kalyan' were observed to be highly resistant. In vitro inoculation of a selected set of genotypes with A. porri, revealed that 7 days after inoculation was suitable to observe the disease severity. In vitro screening of 43 genotypes for resistance to A. porri revealed two resistant lines. An additional 14 genotypes showed consistent moderate resistance in the field as well as in vitro evaluations. Among the related Allium species, A. schoenoprasum and A. roylei showed the least disease index and can be used for interspecific hybridization with cultivated onion. Differential reaction analysis of three A. porri isolates (Apo-Chiplima, Apn-Nasik, Apg-Guntur) in 43 genotypes revealed significant variation among the evaluated Allium species (P = 0.001). All together, the present study suggest that, the newly identified resistance sources can be used as potential donors for ongoing purple blotch resistance breeding program in India.

  10. Induction of protection against the necrotrophic pathogens Phytophthora citrophthora and Alternaria solani in Lycopersicon esculentum Mill. by a novel synthetic glycoside combined with amines.

    PubMed

    Flors, Víctor; Miralles, M Carmen; González-Bosch, Carmen; Carda, Miguel; García-Agustín, Pilar

    2003-04-01

    1,2,3,4-tetra-O-acetyl-6-ethyladipate-beta- D-glucopyranose (TOGE), a glycoside derivative of adipic acid monoethyl ester and 1,2,3,4-tetra-O-acetyl-beta- D-glucopyranose, was synthesized and the resistance-inducing activity shown by TOGE-1 (TOGE plus furfurylamine) and TOGE-2 (TOGE plus 1,3-diaminepropane) was assayed. TOGE-1 and TOGE-2 protected tomato plants against two different fungal pathogens Phytophthora citrophthora and Alternaria solani. Foliar treatments at very low concentrations (2.5 mg l(-1) TOGE, 0.5 mg l(-1) amine) clearly reduced the disease incidence for both pathogens. TOGE-2 application was the most effective on intact plants as well as on detached leaves, reducing fungal growth by more than 46% with respect to control plants. On the other hand TOGE-1 treatment reduced fungal advance by 21%. These results demonstrate a high protective effect against fungal infections for both chemicals. A possible direct antimicrobial effect was discounted due to the weak activity observed in vitro against these pathogens at the low concentrations used in plants. TOGE-2 treatment clearly activates resistance against both pathogens and improves the protective effect previously shown by FGA mixture (adipic acid monoethyl ester, 1,2,3,4-tetra-O-acetyl-beta- D-glucopyranose and furfurylamine) [V. Flors et al. (2001) J Agric Food Chem 49:2569-2575]. The obtained results indicate that TOGE-1 and TOGE-2 act as resistance inducers. Although their mode of action is still unknown, pre-challenge studies have demonstrated the induction of the phenylpropanoid pathway and antioxidant activities. Both chemicals have demonstrated a beneficial effect on plant development, increasing chlorophyll and protein contents, photosynthetic rate and water-use efficiency. The improvement of plant growth and development produced by these treatments suggests crop tolerance to these chemicals, although effective formulations that are safe to humans must be developed before this technology can

  11. Alternaria Species and Their Associated Mycotoxins.

    PubMed

    Pinto, Virginia Elena Fernández; Patriarca, Andrea

    2017-01-01

    The genus Alternaria includes more than 250 species. The traditional methods for identification of Alternaria species are based on morphological characteristics of the reproductive structures and sporulation patterns under controlled culture conditions. Cladistics analyses of "housekeeping genes" commonly used for other genera, failed to discriminate among the small-spored Alternaria species. The development of molecular methods achieving a better agreement with morphological differences is still needed. The production of secondary metabolites has also been used as a means of classification and identification. Alternaria spp. can produce a wide variety of toxic metabolites. These metabolites belong principally to three different structural groups: (1) the dibenzopyrone derivatives, alternariol (AOH), alternariol monomethyl ether (AME), and altenuene (ALT); (2) the perylene derivative altertoxins (ATX-I, ATX-II, and ATX II); and (3) the tetramic acid derivative, tenuazonic acid (TeA). TeA, AOH, AME, ALT, and ATX-I are the main. Certain species in the genus Alternaria produce host-specific toxins (HSTs) that contribute to their pathogenicity and virulence. Alternaria species are plant pathogens that cause spoilage of agricultural commodities with consequent mycotoxin accumulation and economic losses. Vegetable foods infected by Alternaria rot could introduce high amounts of these toxins to the human diet. More investigations on the toxic potential of these toxins and their hazard for human consumption are needed to make a reliable risk assessment of dietary exposure.

  12. Genetic Diversity and Pathogenicity of Xanthomonas axonopodis Strains Inducing Citrus Canker Disease in Iran and South Korea.

    PubMed

    Khodakaramian, G; Swings, J

    2011-06-01

    For the first time in 1989 citrus bacterial canker disease has seen on Citrusaurantiifolia in southern Iran. A total of 43 strains from affected citrus trees, ten strains from South Korea and representative from all known five pathotypes of Xanthomonas axonopodis pathogenic on citrus trees were used in this study. Isolated strains from Iran were indistinguishable by phenotypic, FAMEs, and SDS-PAGE analyses but showed different host range. First group were pathogenic on all tested citrus seedlings including C. aurantiifolia, C. limettioides, C. limon, C. jambhiri, Poncirus trifoliata X C. paradisi, C. aurantium, C. paradise, C. medica, P. trifoliate, C. grandis, C. sinensis, C. reticulate and C. sinensis X P. trifoliate. Pathogenicity of the second group were limited to C. aurantiifolia, C. limettioides, C. limon, C. jambhiri, P. trifoliata X C. paradis, and C. aurantium. Among the strains studied by AFLP fingerprinting six clusters were found. These clusters were: (1) strains of pathotype C; (2) strains of pathotypes B and D; (3) strains of pathotype A together with the main group of the Iranian strains; (4) strains isolated from Korea; (5) strains of pathotype E; and (6) seven strains from Iran which made a completely separate cluster. Strains from pathotypes B and D could not be differentiated by AFLP. The tested Iranian strains belongs to the two different groups and strains from Korea grouped as a subcluster from main cluster of Iranian strains belong to the pathotype A.

  13. Analysis of alternariol and alternariol monomethyl ether on flavedo and albedo tissues of tangerines (Citrus reticulata) with symptoms of alternaria brown spot.

    PubMed

    Magnani, Rodrigo F; De Souza, Gezimar D; Rodrigues-Filho, Edson

    2007-06-27

    A method was developed for the quantification of alternariol and alternariol monomethyl ether on tangerines with and without symptoms of Alternaria brown spot disease. The method employs solid-phase extraction for cleanup, followed by high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS) for detection. This method was validated on flavedo (exocarp or epicarp, exterior yellow peel) and on albedo tissue (mesocarp, interior white peel). An excellent linearity over a range of 0.50-20.0 mg/kg was achieved, with r2 >or= 0.997. The limits of detection (LOD) and quantification (LOQ) were fewer than 0.13 and 0.50 microg/kg, respectively. The relative standard deviations (RSDs) were

  14. Genetic Differentiation and Host Specificity Among Populations of Alternaria spp. Causing Brown Spot of Grapefruit and Tangerine x Grapefruit Hybrids in Florida.

    PubMed

    Peever, T L; Olsen, L; Ibañez, A; Timmer, L W

    2000-04-01

    ABSTRACT Alternaria spp. were sampled from brown spot lesions in several geographically separated citrus groves and different grapefruit and tangerine x grapefruit hybrid cultivars in Florida and screened for variation at 16 putative random amplified polymorphic DNA loci. Populations of the pathogen on two hybrids, Minneola and Orlando, in five locations throughout Florida were moderately differentiated (Nei's coefficient of gene differentiation [G(ST)] = 0.12) among locations. The hypothesis that host-specialized forms of Alternaria spp. cause brown spot on different Citrus spp. and cultivars was tested by estimating genetic differentiation among isolates sampled from different hosts and by pathogenicity assays. Isolates sampled from grapefruit and the hybrid cv. Nova were genetically distinct from isolates sampled from other hybrid cultivars including Robinson, Sunburst, Minneola, Orlando, and Murcott. No differentiation could be detected among isolates sampled from this latter group of hybrids. Quantitative pathogenicity assays on leaves using spray inoculation revealed that 'Nova' isolates were not significantly more pathogenic on 'Nova' compared with isolates from 'Minneola' and 'Orlando'. Similarly, grapefruit isolates were not significantly more pathogenic on grapefruit compared with isolates from 'Minneola'. Isolates from all hosts had similar disease rankings on each inoculated cultivar, with 'Minneola' the most susceptible, followed in decreasing order of susceptibility by 'Orlando', 'Sunburst', 'Nova', and 'Duncan' grapefruit. Rough lemon was generally immune to all isolates tested; however, occasional brown spot lesions were observed on leaves of this host with isolates from grapefruit. No evidence was found to support the hypothesis that unique genotypes of the pathogen, which are more virulent on 'Sunburst' or grapefruit, have been introduced to Florida. Populations of Alternaria spp. causing brown spot of citrus on grapefruit and 'Nova' in Florida

  15. The citrus postharvest pathogen Penicillium digitatum depends on the PdMpkB kinase for developmental and virulence functions.

    PubMed

    Ma, Haijie; Sun, Xuepeng; Wang, Mingshuang; Gai, Yunpeng; Chung, Kuang-Ren; Li, Hongye

    2016-11-07

    The postharvest pathogen Penicillium digitatum causes green mold decay on citrus fruit, resulting in severe economic losses. To explore possible factors involved in fungal pathogenesis, phenotypic characterization of the budding yeast Fus3/Kiss1 mitogen-activated protein (MAP) kinase homolog was carried out. The P. digitatum MAP kinase B coding gene, designated PdMpkB, was functionally inactivated via homologous recombination. The fungal strain (∆PdMpkB) carrying a PdMpkBdeletion demonstrated altered gene expression profiles, reduced growth and conidiogenesis, elevated resistance to osmotic stress, and failed to induce green mold decay on citrus fruit. ∆PdMpkB was more resistant to CaCl2, NaCl and sorbitol than its progenitor strain, indicating a negative regulatory function of PdMpkB in osmotic stress adaptation. Fungal infection assays on citrus fruit revealed that ∆PdMpkB proliferated poorly within host tissues, induced water-soaking lesions, failed to break through host cuticle layers and thus, failed to produce aerial hyphae and conidia. Introduction of a functional copy of PdMpkB into a null mutant restored all defective phenotypes. Transcriptome analysis revealed that inactivation of PdMpkB impacted expression of the genes associated with cell wall-degrading enzyme activities, carbohydrate and amino acid metabolisms, conidial formation, and numerous metabolic processes. Our results define pivotal roles of the PdMpkB-mediated signaling pathway in developmental and pathological functions in the citrus postharvest pathogen P. digitatum.

  16. Insecticidal Suppression of Asian Citrus Psyllid Diaphorina citri (Hemiptera: Liviidae) Vector of Huanglongbing Pathogens

    PubMed Central

    Qureshi, Jawwad A.; Kostyk, Barry C.; Stansly, Philip A.

    2014-01-01

    Diaphorina citri vectors pathogens that cause ‘huanglongbing’ or citrus greening disease which poses a serious threat to citrus production worldwide. Vector suppression is critical to reduce disease spread. Efficacy is a main concern when choosing an insecticide. Insecticidal treatments of 49 products or 44 active ingredients (a.i) labeled or experimental were field tested between 2005–2013 as foliar sprays (250 treatments, 39 a.i) or soil applications (47 treatments, 9 a.i) to control D. citri in citrus. A combined effect of nymphal and adult suppression in response to sprays of 23 insecticides representing 9 modes of action (MoA) groups and 3 unknown MoA provided more than 90% reduction of adult D. citri over 24–68 days. Observable effects on nymphs were generally of shorter duration due to rapid maturation of flush. However, reduction of 76–100% nymphs or adults over 99–296 days was seen on young trees receiving drenches of the neonicotinoids imidacloprid, thiamethoxam or clothianidin (MoA 4A) and a novel anthranilic diamide, cyantraniliprole (MoA 28). Effective products identified for foliar sprays to control D. citri provide sufficient MoA groups for rotation to delay evolution of insecticide resistance by D. citri and other pests. However, cyantraniliprole is now the only available alternative for rotation with neonicotinoids in soil application to young trees. Sprays of up to eight of the most effective insecticides could be rotated over a year without repetition of any MoA and little or no recourse to neonicotinoids or cyantraniliprole, so important for protection of young trees. Other considerations effecting decisions of what and when to spray include prevalence of huanglongbing, pest pressure, pre-harvest intervals, overall budget, equipment availability, and conservation of beneficial arthropods. Examples of spray programs utilizing broad-spectrum and relatively selective insecticides are provided to improve vector management and may vary

  17. Toxigenic Alternaria species from Argentinean blueberries.

    PubMed

    Greco, M; Patriarca, A; Terminiello, L; Fernández Pinto, V; Pose, G

    2012-03-15

    Blueberries are traditionally consumed in North America, some European countries and Japan. In Argentina, the blueberry crop is profitable because production starts in November, when the northern hemisphere lacks fresh fruit. Fungal contaminants can grow and produce mycotoxins in fresh fruit. The aims of this work were to identify the main genera of the mycobiota of blueberries grown in Argentina and to determine the toxicogenic potential, pathogenicity and host specificity of the species isolated. The genus Alternaria was the main component of the blueberry mycobiota (95%); minor proportions of Phoma spp. (4%) and Penicillium spp. (1%) were also isolated. According to their sporulation patterns, 127 Alternaria isolates belonged to the Alternaria tenuissima species-group, 5 to the Alternaria alternata species-group and 2 to the Alternaria arborescens species-group. The last mentioned species-group was not isolated at 5°C. Of the 134 isolates, 61% were toxicogenic in autoclaved rice; 97% of these produced alternariol (AOH) in a range from 0.14 to 119.18 mg/kg, 95% produced alternariol methylether (AME) in a range from 1.23 to 901.74 mg/kg and 65% produced tenuazonic acid (TA) in a range from 0.13 to 2778 mg/kg. Fifty two isolates co-produced the three mycotoxins. According to the size of the lesion that they caused on blueberries, the isolates were classified as slightly pathogenic, moderately pathogenic and very pathogenic. No significant differences in pathogenicity were found on different blueberry varieties. In this work, high incidence and toxicogenic potential of the Alternaria isolates from blueberries were demonstrated. Thus, more studies should be done to evaluate the health risk posed by the presence of the Alternaria toxins in blueberries and in the manufactured by-products.

  18. Sequence diversity on four ORFs of citrus tristeza virus correlates with pathogenicity

    PubMed Central

    Herrera-Isidrón, Lisset; Ochoa-Sánchez, Juan Carlos; Rivera-Bustamante, Rafael; Martínez-Soriano, Juan Pablo

    2009-01-01

    The molecular characterization of isolates of citrus tristeza virus (CTV) from eight locations in Mexico was undertaken by analyzing five regions located at the opposite ends of the virus genome. Two regions have been previously used to study CTV variability (coat protein and p23), while the other three correspond to other genomic segments (p349-B, p349-C and p13). Our comparative nucleotide analyses included CTV sequences from different geographical origins already deposited in the GenBank databases. The largest nucleotide differences were located in two fragments located at the 5' end of the genome (p349-B and p349-C). Phylogenetic analyses on those five regions showed that the degree of nucleotide divergence among strains tended to correlate with their pathogenicity. Two main groups were defined: mild, with almost no noticeable effects on the indicator plants and severe, with drastic symptoms. Mild isolates clustered together in every analyzed ORF sharing a genetic distance below 0.022, in contrast with the severe isolates, which showed a more disperse distribution and a genetic distance of 0.276. Analyses of the p349-B and p349-C regions evidenced two lineages within the severe group: severe common subgroup (most of severe isolates) and severe divergent subgroup (T36-like isolates). This study represents the first attempt to analyze the genetic variability of CTV in Mexico by constructing phylogenetic trees based on new genomic regions that use group-specific nucleotide and amino acid sequences. These results may be useful to implement specific assays for strain discrimination. Moreover, it would be an excellent reference for the CTV situation in México to face the recent arrival of brown citrus aphid. PMID:19642988

  19. Sequence diversity on four ORFs of citrus tristeza virus correlates with pathogenicity.

    PubMed

    Herrera-Isidrón, Lisset; Ochoa-Sánchez, Juan Carlos; Rivera-Bustamante, Rafael; Martínez-Soriano, Juan Pablo

    2009-07-30

    The molecular characterization of isolates of citrus tristeza virus (CTV) from eight locations in Mexico was undertaken by analyzing five regions located at the opposite ends of the virus genome. Two regions have been previously used to study CTV variability (coat protein and p23), while the other three correspond to other genomic segments (p349-B, p349-C and p13). Our comparative nucleotide analyses included CTV sequences from different geographical origins already deposited in the GenBank databases. The largest nucleotide differences were located in two fragments located at the 5' end of the genome (p349-B and p349-C). Phylogenetic analyses on those five regions showed that the degree of nucleotide divergence among strains tended to correlate with their pathogenicity. Two main groups were defined: mild, with almost no noticeable effects on the indicator plants and severe, with drastic symptoms. Mild isolates clustered together in every analyzed ORF sharing a genetic distance below 0.022, in contrast with the severe isolates, which showed a more disperse distribution and a genetic distance of 0.276. Analyses of the p349-B and p349-C regions evidenced two lineages within the severe group: severe common subgroup (most of severe isolates) and severe divergent subgroup (T36-like isolates). This study represents the first attempt to analyze the genetic variability of CTV in Mexico by constructing phylogenetic trees based on new genomic regions that use group-specific nucleotide and amino acid sequences. These results may be useful to implement specific assays for strain discrimination. Moreover, it would be an excellent reference for the CTV situation in México to face the recent arrival of brown citrus aphid.

  20. Inhibitory effects of Fortunella japonica var. margarita and Citrus sunki essential oils on nitric oxide production and skin pathogens.

    PubMed

    Yang, Eun-Jin; Kim, Sang Suk; Moon, Ji-Young; Oh, Tae-Heon; Baik, Jong Seok; Lee, Nam Ho; Hyun, Chang-Gu

    2010-03-01

    A number of essential oils from citrus peels are claimed to have biological activities. Citrus peel, called 'Jin-Pi', is used in traditional medicine for digestion, severe cold, and fever. However, the antibacterial activities against skin pathogens and anti-inflammatory effects of the essential oils of Citrus sunki (JinGyul) and Fortunella japonica var. margarita (GumGyul) have not yet been described. Therefore, in this study, the essential oils of the citrus species C. sunki (CSE) and F. japonica var. margarita (FJE), both native to the island of Jeju, Korea, were examined for their anti-inflammatory and antimicrobial activities against skin pathogens. Four human skin pathogenic microorganisms, Staphylococcus epidermidis CCARM 3709, Propionibacterium acnes CCARM 0081, Malassezia furfur KCCM 12679, and Candida albicans KCCM 11282, were studied. CSE and FJE exhibited strong antimicrobial activity against most of the pathogenic bacteria and yeast strains that were tested. Interestingly, CSE and FJE even showed antimicrobial activity against antibiotic-resistant S. epidermidis CCARM 3710, S. epidermidis CCARM 3711, P. acnes CCARM9009, and P. acnes CCARM9010 strains. In addition, CSE and FJE reduced the lipopolysaccharide (LPS)-induced secretion of nitric oxide (NO) in RAW 264.7 cells, indicating that they have anti-inflammatory effects. We also analysed the chemical composition of the oils by gas chromatography-mass spectrometry (GC-MS) and identified several major components, including dl-limonene (68.18%) and beta-myrcene (4.36%) for CSE, and dl-limonene (61.58%) and carvone (6.36%) for FJE. Taken together, these findings indicate that CSE and FJE have great potential to be used in human skin health applications.

  1. Seedling Albinism Induced by an Extract of Alternaria tenuis.

    PubMed

    Ryan, G F; Greenblatt, G; Al-Delaimy, K A

    1961-09-22

    The chlorophyll deficiency in citrus seedlings known as albinism was induced by inoculating seed with certain clones of Alternaria tenuis, or by germination of the seed in contact with extracts from the fungus. The active material in the extracts apparently has a rather specific inhibiting effect on chlorophyll formation in seedlings of various species.

  2. Infection Density Dynamics of the Citrus Greening Bacterium “Candidatus Liberibacter asiaticus” in Field Populations of the Psyllid Diaphorina citri and Its Relevance to the Efficiency of Pathogen Transmission to Citrus Plants

    PubMed Central

    Ukuda-Hosokawa, Rie; Sadoyama, Yasutsune; Kishaba, Misaki; Kuriwada, Takashi; Anbutsu, Hisashi

    2015-01-01

    Huanglongbing, or citrus greening, is a devastating disease of citrus plants recently spreading worldwide, which is caused by an uncultivable bacterial pathogen, “Candidatus Liberibacter asiaticus,” and vectored by a phloem-sucking insect, Diaphorina citri. We investigated the infection density dynamics of “Ca. Liberibacter asiaticus” in field populations of D. citri with experiments using field-collected insects to address how “Ca. Liberibacter asiaticus” infection density in the vector insect is relevant to pathogen transmission to citrus plants. Of 500 insects continuously collected from “Ca. Liberibacter asiaticus”-infected citrus trees with pathological symptoms in the spring and autumn of 2009, 497 (99.4%) were “Ca. Liberibacter asiaticus” positive. The infections were systemic across head-thorax and abdomen, ranging from 103 to 107 bacteria per insect. In spring, the infection densities were low in March, at ∼103 bacteria per insect, increasing up to 106 to 107 bacteria per insect in April and May, and decreasing to 105 to 106 bacteria per insect in late May, whereas the infection densities were constantly ∼106 to 107 bacteria per insect in autumn. Statistical analysis suggested that several factors, such as insect sex, host trees, and collection dates, may be correlated with “Ca. Liberibacter asiaticus” infection densities in field D. citri populations. Inoculation experiments with citrus seedlings using field-collected “Ca. Liberibacter asiaticus”-infected insects suggested that (i) “Ca. Liberibacter asiaticus”-transmitting insects tend to exhibit higher infection densities than do nontransmitting insects, (ii) a threshold level (∼106 bacteria per insect) of “Ca. Liberibacter asiaticus” density in D. citri is required for successful transmission to citrus plants, and (iii) D. citri attaining the threshold infection level transmits “Ca. Liberibacter asiaticus” to citrus plants in a stochastic manner. These

  3. Elucidation of the biochemical basis of specificity and pathogenicity of Penicillium digitatum on citrus fruit

    USDA-ARS?s Scientific Manuscript database

    Green mold caused by Penicillium digitatum is the most damaging postharvest diseases of citrus fruit. This Penicillium species is specific to citrus fruit and do not cause progressive decay in any other fresh fruit or vegetable crops. While the etiology of P. digitatum is well understood, the phys...

  4. Alternaria infection in a patient with acquired immunodeficiency syndrome: case report and review of invasive alternaria infections.

    PubMed

    Wiest, P M; Wiese, K; Jacobs, M R; Morrissey, A B; Abelson, T I; Witt, W; Lederman, M M

    1987-01-01

    A 31-year-old man with AIDS developed a necrotic lesion on his nasal septum due to Alternaria alternata. Excision and treatment with amphotericin B resulted in cure. This case expands the spectrum of opportunistic pathogens that infect patients with AIDS. Visceral and mucosal infections due to Alternaria have been reported in at least seven other patients.

  5. Expression of ß-1,3-glucanase and ß-1,4-glucanase in two potato cultivars following challenge by the fungal pathogen Alternaria solani

    USDA-ARS?s Scientific Manuscript database

    Early blight of potato, caused by Alternaria solani, is a ubiquitous disease in many countries around the world. We have previously found that variation in resistance phenotypes exist between two different Iranian cultivars of potato. Cultivar ‘Diamond’ is more resistant to multiple isolates of A. s...

  6. Functional analyses of the Diels-Alderase gene sol5 of Ascochyta rabiei and Alternaria solani indicate that the Solanapyrone phytotoxins are not required for pathogenicity

    USDA-ARS?s Scientific Manuscript database

    Ascochyta rabiei and Alternaria solani, the causal agents of Ascochyta blight of chickpea (Cicer arietinum) and early blight of potato (Solanum tuberosum), respectively, produce a set of phytotoxic compounds incuding solanapyrones A, B, and C. Although both the phytotoxicity of solanopyrones and the...

  7. Dose response of soilborne plant pathogens and Meloidogyne incognita to citrus-based experimental compounds.

    USDA-ARS?s Scientific Manuscript database

    Two novel citrus-based compounds have been tested in vitro against Colletotrichum gleosporioides, Fusarium oxysporum, Sclerotinia sclerotiorum, Sclerotium rolfsii, Rhizoctonia solani, Verticillium albo-atrum, Pythium aphanidermatum, P. myriotilum, Phytophthora nicotianae and P. capsici. One of the...

  8. Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response

    PubMed Central

    2014-01-01

    Background Transcriptional activator-like (TAL) effectors, formerly known as the AvrBs3/PthA protein family, are DNA-binding effectors broadly found in Xanthomonas spp. that transactivate host genes upon injection via the bacterial type three-secretion system. Biologically relevant targets of TAL effectors, i.e. host genes whose induction is vital to establish a compatible interaction, have been reported for xanthomonads that colonize rice and pepper; however, citrus genes modulated by the TAL effectors PthA“s” and PthC“s” of the citrus canker bacteria Xanthomonas citri (Xc) and Xanthomonas aurantifolii pathotype C (XaC), respectively, are poorly characterized. Of particular interest, XaC causes canker disease in its host lemon (Citrus aurantifolia), but triggers a defense response in sweet orange. Results Based on, 1) the TAL effector-DNA binding code, 2) gene expression data of Xc and XaC-infiltrated sweet orange leaves, and 3) citrus hypocotyls transformed with PthA2, PthA4 or PthC1, we have identified a collection of Citrus sinensis genes potentially targeted by Xc and XaC TAL effectors. Our results suggest that similar with other strains of Xanthomonas TAL effectors, PthA2 and PthA4, and PthC1 to some extent, functionally converge. In particular, towards induction of genes involved in the auxin and gibberellin synthesis and response, cell division, and defense response. We also present evidence indicating that the TAL effectors act as transcriptional repressors and that the best scoring predicted DNA targets of PthA“s” and PthC“s” in citrus promoters predominantly overlap with or localize near to TATA boxes of core promoters, supporting the idea that TAL effectors interact with the host basal transcriptional machinery to recruit the RNA pol II and start transcription. Conclusions The identification of PthA“s” and PthC“s” targets, such as the LOB (LATERAL ORGAN BOUNDARY) and CCNBS genes that we report here, is key for the understanding

  9. Identification of putative TAL effector targets of the citrus canker pathogens shows functional convergence underlying disease development and defense response.

    PubMed

    Pereira, Andre L A; Carazzolle, Marcelo F; Abe, Valeria Y; de Oliveira, Maria L P; Domingues, Mariane N; Silva, Jaqueline C; Cernadas, Raul A; Benedetti, Celso E

    2014-02-25

    Transcriptional activator-like (TAL) effectors, formerly known as the AvrBs3/PthA protein family, are DNA-binding effectors broadly found in Xanthomonas spp. that transactivate host genes upon injection via the bacterial type three-secretion system. Biologically relevant targets of TAL effectors, i.e. host genes whose induction is vital to establish a compatible interaction, have been reported for xanthomonads that colonize rice and pepper; however, citrus genes modulated by the TAL effectors PthA"s" and PthC"s" of the citrus canker bacteria Xanthomonas citri (Xc) and Xanthomonas aurantifolii pathotype C (XaC), respectively, are poorly characterized. Of particular interest, XaC causes canker disease in its host lemon (Citrus aurantifolia), but triggers a defense response in sweet orange. Based on, 1) the TAL effector-DNA binding code, 2) gene expression data of Xc and XaC-infiltrated sweet orange leaves, and 3) citrus hypocotyls transformed with PthA2, PthA4 or PthC1, we have identified a collection of Citrus sinensis genes potentially targeted by Xc and XaC TAL effectors. Our results suggest that similar with other strains of Xanthomonas TAL effectors, PthA2 and PthA4, and PthC1 to some extent, functionally converge. In particular, towards induction of genes involved in the auxin and gibberellin synthesis and response, cell division, and defense response. We also present evidence indicating that the TAL effectors act as transcriptional repressors and that the best scoring predicted DNA targets of PthA"s" and PthC"s" in citrus promoters predominantly overlap with or localize near to TATA boxes of core promoters, supporting the idea that TAL effectors interact with the host basal transcriptional machinery to recruit the RNA pol II and start transcription. The identification of PthA"s" and PthC"s" targets, such as the LOB (lateral organ boundary) and CCNBS genes that we report here, is key for the understanding of the canker symptoms development during host

  10. Prophage-Mediated Dynamics of ‘Candidatus Liberibacter asiaticus’ Populations, the Destructive Bacterial Pathogens of Citrus Huanglongbing

    PubMed Central

    Zhou, Lijuan; Powell, Charles A.; Li, Wenbin; Irey, Mike; Duan, Yongping

    2013-01-01

    Prophages are highly dynamic components in the bacterial genome and play an important role in intraspecies variations. There are at least two prophages in the chromosomes of Candidatus Liberibacter asiaticus’ (Las) Floridian isolates. Las is both unculturable and the most prevalent species of Liberibacter pathogens that cause huanglongbing (HLB), a worldwide destructive disease of citrus. In this study, seven new prophage variants resulting from two hyper-variable regions were identified by screening clone libraries of infected citrus, periwinkle and psyllids. Among them, Types A and B share highly conserved sequences and localize within the two prophages, FP1 and FP2, respectively. Although Types B and C were abundant in all three libraries, Type A was much more abundant in the libraries from the Las-infected psyllids than from the Las-infected plants, and Type D was only identified in libraries from the infected host plants but not from the infected psyllids. Sequence analysis of these variants revealed that the variations may result from recombination and rearrangement events. Conventional PCR results using type-specific molecular markers indicated that A, B, C and D are the four most abundant types in Las-infected citrus and periwinkle. However, only three types, A, B and C are abundant in Las-infected psyllids. Typing results for Las-infected citrus field samples indicated that mixed populations of Las bacteria present in Floridian isolates, but only the Type D population was correlated with the blotchy mottle symptom. Extended cloning and sequencing of the Type D region revealed a third prophage/phage in the Las genome, which may derive from the recombination of FP1 and FP2. Dramatic variations in these prophage regions were also found among the global Las isolates. These results are the first to demonstrate the prophage/phage-mediated dynamics of Las populations in plant and insect hosts, and their correlation with insect transmission and disease

  11. Protein interaction networks at the host–microbe interface in Diaphorina citri, the insect vector of the citrus greening pathogen

    PubMed Central

    Chavez, J. D.; Johnson, R.; Hosseinzadeh, S.; Mahoney, J. E.; Mohr, J. P.; Robison, F.; Zhong, X.; Hall, D. G.; MacCoss, M.; Bruce, J.; Cilia, M.

    2017-01-01

    The Asian citrus psyllid (Diaphorina citri) is the insect vector responsible for the worldwide spread of ‘Candidatus Liberibacter asiaticus’ (CLas), the bacterial pathogen associated with citrus greening disease. Developmental changes in the insect vector impact pathogen transmission, such that D. citri transmission of CLas is more efficient when bacteria are acquired by nymphs when compared with adults. We hypothesize that expression changes in the D. citri immune system and commensal microbiota occur during development and regulate vector competency. In support of this hypothesis, more proteins, with greater fold changes, were differentially expressed in response to CLas in adults when compared with nymphs, including insect proteins involved in bacterial adhesion and immunity. Compared with nymphs, adult insects had a higher titre of CLas and the bacterial endosymbionts Wolbachia, Profftella and Carsonella. All Wolbachia and Profftella proteins differentially expressed between nymphs and adults are upregulated in adults, while most differentially expressed Carsonella proteins are upregulated in nymphs. Discovery of protein interaction networks has broad applicability to the study of host–microbe relationships. Using protein interaction reporter technology, a D. citri haemocyanin protein highly upregulated in response to CLas was found to physically interact with the CLas coenzyme A (CoA) biosynthesis enzyme phosphopantothenoylcysteine synthetase/decarboxylase. CLas pantothenate kinase, which catalyses the rate-limiting step of CoA biosynthesis, was found to interact with a D. citri myosin protein. Two Carsonella enzymes involved in histidine and tryptophan biosynthesis were found to physically interact with D. citri proteins. These co-evolved protein interaction networks at the host–microbe interface are highly specific targets for controlling the insect vector responsible for the spread of citrus greening. PMID:28386418

  12. Protein interaction networks at the host-microbe interface in Diaphorina citri, the insect vector of the citrus greening pathogen.

    PubMed

    Ramsey, J S; Chavez, J D; Johnson, R; Hosseinzadeh, S; Mahoney, J E; Mohr, J P; Robison, F; Zhong, X; Hall, D G; MacCoss, M; Bruce, J; Cilia, M

    2017-02-01

    The Asian citrus psyllid (Diaphorina citri) is the insect vector responsible for the worldwide spread of 'Candidatus Liberibacter asiaticus' (CLas), the bacterial pathogen associated with citrus greening disease. Developmental changes in the insect vector impact pathogen transmission, such that D. citri transmission of CLas is more efficient when bacteria are acquired by nymphs when compared with adults. We hypothesize that expression changes in the D. citri immune system and commensal microbiota occur during development and regulate vector competency. In support of this hypothesis, more proteins, with greater fold changes, were differentially expressed in response to CLas in adults when compared with nymphs, including insect proteins involved in bacterial adhesion and immunity. Compared with nymphs, adult insects had a higher titre of CLas and the bacterial endosymbionts Wolbachia, Profftella and Carsonella. All Wolbachia and Profftella proteins differentially expressed between nymphs and adults are upregulated in adults, while most differentially expressed Carsonella proteins are upregulated in nymphs. Discovery of protein interaction networks has broad applicability to the study of host-microbe relationships. Using protein interaction reporter technology, a D. citri haemocyanin protein highly upregulated in response to CLas was found to physically interact with the CLas coenzyme A (CoA) biosynthesis enzyme phosphopantothenoylcysteine synthetase/decarboxylase. CLas pantothenate kinase, which catalyses the rate-limiting step of CoA biosynthesis, was found to interact with a D. citri myosin protein. Two Carsonella enzymes involved in histidine and tryptophan biosynthesis were found to physically interact with D. citri proteins. These co-evolved protein interaction networks at the host-microbe interface are highly specific targets for controlling the insect vector responsible for the spread of citrus greening.

  13. Fungicidal activity of silver nanoparticles against Alternaria brassicicola

    NASA Astrophysics Data System (ADS)

    Gupta, Deepika; Chauhan, Pratima

    2016-04-01

    This work highlighted the fungicidal properties of silver nanoparticles against Alternaria brassicicola. Alternaria brassicicola causes Black spot of Cauliflower, radish, cabbage, kale which results in sever agricultural loss. We treat the synthesised silver nanoparticles (AgNPs) of 10, 25, 50, 100 and 110 ppm concentrations against Alternaria brassicicola on PDA containing Petri dish. We calculated inhibitory rate (%) in order to evaluate the antifungal efficacy of silver nanoparticles against pathogens. Treatment with 100ppm AgNPs resulted in maximum inhibition of Alternaria brassicicola i.e.92.2%. 110ppm of AgNPS also shows the same result, therefore 100ppm AgNPs was treated as optimize concentration. AgNPs effectively inhibited the growth of a Alternaria brassicicola, which suggests that AgNPs could be used as fungicide in plant disease management. Further research and development are necessary to translate this technology into plant disease management strategies.

  14. Alternaria in Food: Ecophysiology, Mycotoxin Production and Toxicology

    PubMed Central

    Patriarca, Andrea; Magan, Naresh

    2015-01-01

    Alternaria species are common saprophytes or pathogens of a wide range of plants pre- and post-harvest. This review considers the relative importance of Alternaria species, their ecology, competitiveness, production of mycotoxins and the prevalence of the predominant mycotoxins in different food products. The available toxicity data on these toxins and the potential future impacts of Alternaria species and their toxicity in food products pre- and post-harvest are discussed. The growth of Alternaria species is influenced by interacting abiotic factors, especially water activity (aw), temperature and pH. The boundary conditions which allow growth and toxin production have been identified in relation to different matrices including cereal grain, sorghum, cottonseed, tomato, and soya beans. The competitiveness of Alternaria species is related to their water stress tolerance, hydrolytic enzyme production and ability to produce mycotoxins. The relationship between A. tenuissima and other phyllosphere fungi has been examined and the relative competitiveness determined using both an Index of Dominance (ID) and the Niche Overlap Index (NOI) based on carbon-utilisation patterns. The toxicology of some of the Alternaria mycotoxins have been studied; however, some data are still lacking. The isolation of Alternaria toxins in different food products including processed products is reviewed. The future implications of Alternaria colonization/infection and the role of their mycotoxins in food production chains pre- and post-harvest are discussed. PMID:26190916

  15. Infection Density Dynamics of the Citrus Greening Bacterium "Candidatus Liberibacter asiaticus" in Field Populations of the Psyllid Diaphorina citri and Its Relevance to the Efficiency of Pathogen Transmission to Citrus Plants.

    PubMed

    Ukuda-Hosokawa, Rie; Sadoyama, Yasutsune; Kishaba, Misaki; Kuriwada, Takashi; Anbutsu, Hisashi; Fukatsu, Takema

    2015-06-01

    Huanglongbing, or citrus greening, is a devastating disease of citrus plants recently spreading worldwide, which is caused by an uncultivable bacterial pathogen, "Candidatus Liberibacter asiaticus," and vectored by a phloem-sucking insect, Diaphorina citri. We investigated the infection density dynamics of "Ca. Liberibacter asiaticus" in field populations of D. citri with experiments using field-collected insects to address how "Ca. Liberibacter asiaticus" infection density in the vector insect is relevant to pathogen transmission to citrus plants. Of 500 insects continuously collected from "Ca. Liberibacter asiaticus"-infected citrus trees with pathological symptoms in the spring and autumn of 2009, 497 (99.4%) were "Ca. Liberibacter asiaticus" positive. The infections were systemic across head-thorax and abdomen, ranging from 10(3) to 10(7) bacteria per insect. In spring, the infection densities were low in March, at ∼ 10(3) bacteria per insect, increasing up to 10(6) to 10(7) bacteria per insect in April and May, and decreasing to 10(5) to 10(6) bacteria per insect in late May, whereas the infection densities were constantly ∼ 10(6) to 10(7) bacteria per insect in autumn. Statistical analysis suggested that several factors, such as insect sex, host trees, and collection dates, may be correlated with "Ca. Liberibacter asiaticus" infection densities in field D. citri populations. Inoculation experiments with citrus seedlings using field-collected "Ca. Liberibacter asiaticus"-infected insects suggested that (i) "Ca. Liberibacter asiaticus"-transmitting insects tend to exhibit higher infection densities than do nontransmitting insects, (ii) a threshold level (∼ 10(6) bacteria per insect) of "Ca. Liberibacter asiaticus" density in D. citri is required for successful transmission to citrus plants, and (iii) D. citri attaining the threshold infection level transmits "Ca. Liberibacter asiaticus" to citrus plants in a stochastic manner. These findings provide

  16. Discord between morphological and phylogenetic species boundaries: incomplete lineage sorting and recombination results in fuzzy species boundaries in an asexual fungal pathogen

    PubMed Central

    2014-01-01

    Background Traditional morphological and biological species concepts are difficult to apply to closely related, asexual taxa because of the lack of an active sexual phase and paucity of morphological characters. Phylogenetic species concepts such as genealogical concordance phylogenetic species recognition (GCPSR) have been extensively used; however, methods that incorporate gene tree uncertainty into species recognition may more accurately and objectively delineate species. Using a worldwide sample of Alternaria alternata sensu lato, causal agent of citrus brown spot, the evolutionary histories of four nuclear loci including an endo-polygalacturonase gene, two anonymous loci, and one microsatellite flanking region were estimated using the coalescent. Species boundaries were estimated using several approaches including those that incorporate uncertainty in gene genealogies when lineage sorting and non-reciprocal monophyly of gene trees is common. Results Coalescent analyses revealed three phylogenetic lineages strongly influenced by incomplete lineage sorting and recombination. Divergence of the citrus 2 lineage from the citrus 1 and citrus 3 lineages was supported at most loci. A consensus of species tree estimation methods supported two species of Alternaria causing citrus brown spot worldwide. Based on substitution rates at the endo-polygalacturonase locus, divergence of the citrus 2 and the 1 and 3 lineages was estimated to have occurred at least 5, 400 years before present, predating the human-mediated movement of citrus and associated pathogens out of SE Asia. Conclusions The number of Alternaria species identified as causing brown spot of citrus worldwide using morphological criteria has been overestimated. Little support was found for most of these morphospecies using quantitative species recognition approaches. Correct species delimitation of plant-pathogenic fungi is critical for understanding the evolution of pathogenicity, introductions of pathogens to

  17. Discord between morphological and phylogenetic species boundaries: incomplete lineage sorting and recombination results in fuzzy species boundaries in an asexual fungal pathogen.

    PubMed

    Stewart, Jane E; Timmer, Lavern W; Lawrence, Christopher B; Pryor, Barry M; Peever, Tobin L

    2014-03-03

    Traditional morphological and biological species concepts are difficult to apply to closely related, asexual taxa because of the lack of an active sexual phase and paucity of morphological characters. Phylogenetic species concepts such as genealogical concordance phylogenetic species recognition (GCPSR) have been extensively used; however, methods that incorporate gene tree uncertainty into species recognition may more accurately and objectively delineate species. Using a worldwide sample of Alternaria alternata sensu lato, causal agent of citrus brown spot, the evolutionary histories of four nuclear loci including an endo-polygalacturonase gene, two anonymous loci, and one microsatellite flanking region were estimated using the coalescent. Species boundaries were estimated using several approaches including those that incorporate uncertainty in gene genealogies when lineage sorting and non-reciprocal monophyly of gene trees is common. Coalescent analyses revealed three phylogenetic lineages strongly influenced by incomplete lineage sorting and recombination. Divergence of the citrus 2 lineage from the citrus 1 and citrus 3 lineages was supported at most loci. A consensus of species tree estimation methods supported two species of Alternaria causing citrus brown spot worldwide. Based on substitution rates at the endo-polygalacturonase locus, divergence of the citrus 2 and the 1 and 3 lineages was estimated to have occurred at least 5, 400 years before present, predating the human-mediated movement of citrus and associated pathogens out of SE Asia. The number of Alternaria species identified as causing brown spot of citrus worldwide using morphological criteria has been overestimated. Little support was found for most of these morphospecies using quantitative species recognition approaches. Correct species delimitation of plant-pathogenic fungi is critical for understanding the evolution of pathogenicity, introductions of pathogens to new areas, and for regulating

  18. Cyclic AMP-dependent protein kinase A negatively regulates conidia formation by the tangerine pathotype of Alternaria alternata.

    PubMed

    Tsai, Hsieh-Chin; Yang, Siwy Ling; Chung, Kuang-Ren

    2013-02-01

    The necrotrophic fungal pathogen Alternaria alternata causes brown spot diseases in many citrus cultivars. The FUS3 and SLT2 mitogen-activated protein kinases (MAPK)-mediated signaling pathways have been shown to be required for conidiation. Exogenous application of cAMP to this fungal pathogen decreased conidia formation considerably. This study determined whether a cAMP-activated protein kinase A (PKA) is required for conidiation. Using loss-of-function mutations in PKA catalytic and regulatory subunit-coding genes, we demonstrated that PKA negatively regulates conidiation. Fungal mutants lacking PKA catalytic subunit gene (PKA ( cat )) reduced growth, lacked detectable PKA activity, and produced higher amounts of conidia compared to wild-type. Introduction of a functional copy of PKA ( cat ) into a null mutant partially restored PKA activity and produced wild-type level of conidia. In contrast, fungi lacking PKA regulatory subunit gene (PKA ( reg )) produced detectable PKA activity, exhibited severe growth reduction, formed swelling hyphal segments, and produced no mature conidia. Introduction of the PKA ( reg ) gene to a regulatory subunit mutant restored all phenotypes to wild type. PKA ( reg )-null mutants induced fewer necrotic lesions on citrus compared to wild-type, whereas PKA ( cat ) mutant displayed wild-type virulence. Overall, our studies indicate that PKA and FUS3-mediated signaling pathways apparently have very different roles in the regulation of conidia production and A. alternata pathogenesis in citrus.

  19. Biotransformations of 6',7'-dihydroxybergamottin and 6',7'-epoxybergamottin by the citrus-pathogenic fungi diminish cytochrome P450 3A4 inhibitory activity

    USDA-ARS?s Scientific Manuscript database

    Penicillium digitatum, as well as five other citrus pathogenic species, (P. ulaiense Link, Geotrichum citri Link, Botrytis cinerea P. Micheli ex Pers., Lasiodiplodia theobromae (Pat.)Griffon & Maubl. and Phomopsis citri (teleomorph Diaporthe citri)) was observed to convert 6',7'-epoxybergamottin (1)...

  20. Additive roles of PthAs in bacterial growth and pathogenicity associated with nucleotide polymorphisms in effector-binding elements of citrus canker susceptibility genes.

    PubMed

    Abe, Valeria Yukari; Benedetti, Celso Eduardo

    2016-10-01

    Citrus canker, caused by Xanthomonas citri, affects most commercial citrus varieties. All X. citri strains possess at least one transcription activator-like effector of the PthA family that activates host disease susceptibility (S) genes. The X. citri strain 306 encodes four PthA effectors; nevertheless, only PthA4 is known to elicit cankers on citrus. As none of the PthAs act as avirulence factors on citrus, we hypothesized that PthAs 1-3 might also contribute to pathogenicity on certain hosts. Here, we show that, although PthA4 is indispensable for canker formation in six Brazilian citrus varieties, PthAs 1 and 3 contribute to canker development in 'Pera' sweet orange, but not in 'Tahiti' lemon. Deletions in two or more pthA genes reduce bacterial growth in planta more pronouncedly than single deletions, suggesting an additive role of PthAs in pathogenicity and bacterial fitness. The contribution of PthAs 1 and 3 in canker formation in 'Pera' plants does not correlate with the activation of the canker S gene, LOB1 (LATERAL ORGAN BOUNDARIES 1), but with the induction of other PthA targets, including LOB2 and citrus dioxygenase (DIOX). LOB1, LOB2 and DIOX show differential PthA-dependent expression between 'Pera' and 'Tahiti' plants that appears to be associated with nucleotide polymorphisms found at or near PthA-binding sites. We also present evidence that LOB1 activation alone is not sufficient to elicit cankers on citrus, and that DIOX acts as a canker S gene in 'Pera', but not 'Tahiti', plants. Our results suggest that the activation of multiple S genes, such as LOB1 and DIOX, is necessary for full canker development. © 2015 BSPP and John Wiley & Sons Ltd.

  1. Bacterial Diversity Analysis of Huanglongbing Pathogen-Infected Citrus, Using PhyloChip Arrays and 16S rRNA Gene Clone Library Sequencing▿ †

    PubMed Central

    Sagaram, Uma Shankar; DeAngelis, Kristen M.; Trivedi, Pankaj; Andersen, Gary L.; Lu, Shi-En; Wang, Nian

    2009-01-01

    The bacterial diversity associated with citrus leaf midribs was characterized for citrus groves that contained the Huanglongbing (HLB) pathogen, which has yet to be cultivated in vitro. We employed a combination of high-density phylogenetic 16S rRNA gene microarrays and 16S rRNA gene clone library sequencing to determine the microbial community composition for symptomatic and asymptomatic citrus midribs. Our results revealed that citrus leaf midribs can support a diversity of microbes. PhyloChip analysis indicated that 47 orders of bacteria in 15 phyla were present in the citrus leaf midribs, while 20 orders in 8 phyla were observed with the cloning and sequencing method. PhyloChip arrays indicated that nine taxa were significantly more abundant in symptomatic midribs than in asymptomatic midribs. “Candidatus Liberibacter asiaticus” was detected at a very low level in asymptomatic plants but was over 200 times more abundant in symptomatic plants. The PhyloChip analysis results were further verified by sequencing 16S rRNA gene clone libraries, which indicated the dominance of “Candidatus Liberibacter asiaticus” in symptomatic leaves. These data implicate “Candidatus Liberibacter asiaticus” as the pathogen responsible for HLB disease. PMID:19151177

  2. Cellular toxicity of elsinochrome phytotoxins produced by the pathogenic fungus, Elsinoë fawcettii causing citrus scab.

    PubMed

    Liao, Hui-Ling; Chung, Kuang-Ren

    2008-01-01

    Elsinochromes are the red/orange pigments produced by many Elsinoë fungal species and are structurally similar to the phytotoxin, cercosporin. Here, pigments were extracted from cultures of a citrus pathogen, Elsinoë fawcettii and tested for cellular toxicity. On irradiation with light, elsinochromes rapidly killed suspension cultured citrus and tobacco cells. The toxicity was decreased by adding the singlet oxygen ((1)O(2)) quenchers (bixin (carotenoid carboxylic acid), DABCO (1, 4-diazabicyco octane), ascorbate or reduced glutathione). Application of elsinochromes onto rough lemon leaves resulted in necrotic lesions, whereas lesion development was inhibited by the addition of bixin, DABCO or ascorbate, but not a-tocopherol. Incubation of rough lemon leaf discs with elsinochromes in the light induced a steady increase of electrolyte leakage. Compared with two photosensitizing compounds, hematoporphyrin and cercosporin, the accumulation of (1)O(2) induced by elsinochromes after irradiation was indicated by successful detection of the cholesterol oxidation product, 5a-hydroperoxide. Addition of a potent quencher, beta-carotene prevented 5alpha-hydroperoxide production. Elsinochromes generated superoxide ions (O(2)(*-)), whereas accumulation of O(2)(*-)was blocked by addition of the superoxide dismutase, a scavenger of O(2)(*-), but not the (1)O(2)-quencher, DABCO. Our study indicated that elsinochromes are functioning as photosensitizing compounds that produce (1)O(2)and O(2)(*-), and exert toxicity to plant cells.

  3. Bridgehead invasion of a monomorphic plant pathogenic bacterium: Xanthomonas citri pv. citri, an emerging citrus pathogen in Mali and Burkina Faso.

    PubMed

    Leduc, A; Traoré, Y N; Boyer, K; Magne, M; Grygiel, P; Juhasz, C C; Boyer, C; Guerin, F; Wonni, I; Ouedraogo, L; Vernière, C; Ravigné, V; Pruvost, O

    2015-11-01

    Molecular epidemiology studies further our understanding of migrations of phytopathogenic bacteria, the major determining factor in their emergence. Asiatic citrus canker, caused by Xanthomonas citri pv. citri, was recently reported in Mali and Burkina Faso, a region remote from other contaminated areas. To identify the origin and pathways of these emergences, we used two sets of markers, minisatellites and microsatellites, for investigating different evolutionary scales. Minisatellite typing suggested the introduction of two groups of strains in Mali (DAPC 1 and DAPC 2), consistent with microsatellite typing. DAPC 2 was restricted to Bamako district, whereas DAPC 1 strains were found much more invasive. The latter strains formed a major clonal complex based on microsatellite data with the primary and secondary founders detected in commercial citrus nurseries and orchards. This suggests that human activities played a major role in the spread of DAPC 1 strains via the movement of contaminated propagative material, further supported by the frequent lack of differentiation between populations from geographically distant nurseries and orchards. Approximate Bayesian Computation analyses supported the hypothesis that strains from Burkina Faso resulted from a bridgehead invasion from Mali. Multi-locus variable number of tandem repeat analysis and Approximate Bayesian Computation are useful for understanding invasion routes and pathways of monomorphic bacterial pathogens.

  4. Past and future of a century old Citrus Tristeza virus collection: A California citrus germplasm tale

    USDA-ARS?s Scientific Manuscript database

    The California Citrus Clonal Protection Program (CCPP) provides a mechanism for introduction and distribution of pathogen-free citrus varieties to California for use in research, variety improvement, or commercial production. Citrus tristeza virus (CTV) is a serious citrus pathogen worldwide. The pr...

  5. Occurrence and function of fungal antifungal proteins: a case study of the citrus postharvest pathogen Penicillium digitatum.

    PubMed

    Garrigues, Sandra; Gandía, Mónica; Marcos, Jose F

    2016-03-01

    Antifungal proteins (AFPs) of fungal origin have been described in filamentous fungi. AFPs are small, highly stable, cationic cysteine-rich proteins (CRPs) that are usually secreted in high amounts and show potent antifungal activity against non-self fungi. The role of AFPs in the biology of the producer fungus remains unclear. AFPs have been proposed as promising lead compounds for the development of new antifungals. The analyses of available antifungal CRP sequences from fungal origin and their phylogenetic reconstruction led us to propose a new classification of AFPs in three distinct classes: A, B and C. We initiate for the first time the characterization of an AFP in a fungal pathogen, by analysing the functional role of the unique afpB gene in the citrus fruit pathogen Penicillium digitatum. Null ΔafpB mutants revealed that this gene is dispensable for vegetative growth and fruit infection. However, strains that artificially express afpB in a constitutive way (afpB (C)) showed a phenotype of restricted growth, distortion of hyphal morphology and strong reduction in virulence to citrus fruits. These characteristics support an antifungal role for AfpB. Surprisingly, we did not detect the AfpB protein in any of the P. digitatum strains and growth conditions that were analysed in this study, regardless of high gene expression. The afpB (C) phenotype is not stable and occasionally reverts to a wild type-like phenotype but molecular changes were not detected with this reversion. The reduced virulence of afpB (C) strains correlated with localized fruit necrosis and altered timing of expression of fruit defence genes.

  6. Distribution and Characterization of AKT Homologs in the Tangerine Pathotype of Alternaria alternata.

    PubMed

    Masunaka, A; Tanaka, A; Tsuge, T; Peever, T L; Timmer, L W; Yamamoto, M; Yamamoto, H; Akimitsu, K

    2000-07-01

    ABSTRACT The tangerine pathotype of Alternaria alternata produces a host-selective toxin (HST), known as ACT-toxin, and causes Alternaria brown spot disease of citrus. The structure of ACT-toxin is closely related to AK- and AF-toxins, which are HSTs produced by the Japanese pear and strawberry pathotypes of A. alternata, respectively. AC-, AK-, and AF-toxins are chemically similar and share a 9,10-epoxy-8-hydroxy-9-methyl-decatrienoic acid moiety. Two genes controlling AK-toxin biosynthesis (AKT1 and AKT2) were recently cloned from the Japanese pear pathotype of A. alternata. Portions of these genes were used as heterologous probes in Southern blots, that detected homologs in 13 isolates of A. alternata tangerine pathotype from Minneola tangelo in Florida. Partial sequencing of the homologs in one of these isolates demonstrated high sequence similarity to AKT1 (89.8%) and to AKT2 (90.7%). AKT homologs were not detected in nine isolates of A. alternata from rough lemon, six isolates of nonpathogenic A. alternata, and one isolate of A. citri that causes citrus black rot. The presence of homologs in the Minneola isolates and not in the rough lemon isolates, nonpathogens or black rot isolates, correlates perfectly to pathogenicity on Iyo tangerine and ACT-toxin production. Functionality of the homologs was demonstrated by detection of transcripts using reverse transcription-polymerase chain reaction (RT-PCR) in total RNA of the tangerine pathotype of A. alternata. The high sequence similarity of AKT and AKT homologs in the tangerine patho-type, combined with the structural similarity of AK-toxin and ACT-toxin, may indicate that these homologs are involved in the biosynthesis of the decatrienoic acid moiety of ACT-toxin.

  7. Effect of X-irradiation on Citrus Canker Pathogen Xanthomonas citri subsp. citri of Satsuma Mandarin Fruits

    PubMed Central

    Song, Min-A; Park, Jae Sin; Kim, Ki Deok; Jeun, Yong Chull

    2015-01-01

    Citrus canker caused by Xanthomonas citri subsp. citri (Xcc) is one of the most important bacterial diseases of citrus. Because citrus canker is not found in many countries including European Union and Australia, Xcc is strictly regulated in order to prevent its spread. In this study, the effects of X-irradiation on Xcc growth either in the suspension or on the surface of citrus fruits were investigated. The suspension containing 1×107 cfu/ml of Xcc was irradiated with different absorbed doses of X-irradiation ranging from 50 to 400 Gy. The results showed that Xcc was fully dead at 400 Gy of X-irradiation. To determine the effect of X-irradiation on quarantine, the Xcc-inoculated citrus fruits were irradiated with different X-ray doses at which Xcc was completely inhibited by an irradiation dose of 250 Gy. The D10 value for Xcc on citrus fruits was found to be 97 Gy, indicating the possibility of direct application on citrus quarantine without any side sterilizer. Beside, presence of Xcc on the surface of asymptomatic citrus fruits obtained from citrus canker-infected orchards was noted. It indicated that the exporting citrus fruits need any treatment so that Xcc on the citrus fruits should be completely eliminated. Based on these results, ionizing radiation can be considered as an alternative method of eradicating Xcc for export of citrus fruits. PMID:26672670

  8. Annotation of the Asian citrus psyllid genome reveals a reduced innate immune system

    USDA-ARS?s Scientific Manuscript database

    Citrus production worldwide is currently facing significant losses due to citrus greening disease, also known as huanglongbing. The citrus greening bacteria, Candidatus Liberibacter asiaticus (CLas), is a persistent propagative pathogen transmitted by the Asian citrus psyllid, Diaphorina citri Kuway...

  9. Improved annotation of the insect vector of citrus greening disease: Biocuration by a diverse genomics community

    USDA-ARS?s Scientific Manuscript database

    The Asian citrus psyllid (Diaphorina citri Kuwayama) is the insect vector of the bacterium Candidatus Liberibacter asiaticus (CLas), the pathogen associated with citrus Huanglongbing (HLB, citrus greening). HLB threatens citrus production worldwide. Suppression or reduction of the insect vector usin...

  10. Pathogenic Interactions Between Xanthomonas axonopodis pv. citri and Cultivars of Pummelo (Citrus grandis).

    PubMed

    Shiotani, H; Ozaki, K; Tsuyumu, S

    2000-12-01

    ABSTRACT The aggressiveness of strains of Xanthomonas axonopodis pv. citri on seven Citrus species, including Citrus sinensis (navel orange), C. paradisi (grapefruit), C. unshiu (Satsuma mandarin), C. junos (Yuzu), C. aurantifolia ('Mexican' lime), C. tachibana (Tachibana), and C. grandis (pummelo: 'Otachibana', 'Banpeiyu', and 'Anseikan'), were assessed by comparing lesion expansion and growth in planta, using a prick inoculation method. The existence of two groups distinct in aggressiveness was demonstrated on the pummelo cultivars, whereas the remaining species tested were uniformly susceptible. The two groups of strains were distinct in lesion expansion and growth in planta; however, both caused canker lesions on the 'Otachibana' pummelo. The sensitivity of the bacterial strains to phages Cp1 and Cp2 was associated with differences in aggressiveness. Namely, all the strains sensitive to Cp2 but resistant to Cp1 were aggressive to 'Otachibana', whereas all the strains sensitive to Cp1 but resistant to Cp2 were weakly aggressive. When a repetitive sequence-based polymerase chain reaction amplification was carried out by enterobacterial repetitive intergeneric consensus (ERIC) sequences (ERIC1R and ERIC2) as the primers, these two groups were also distinguishable by the presence or absence of a 1.8-kb DNA fragment among otherwise identical fragments. The 1.8-kb fragment was amplified only from the strains aggressive to C. grandis.

  11. The value of citrus genebanking

    USDA-ARS?s Scientific Manuscript database

    Access to diverse citrus genetic resources is critical for breeding new citrus cultivars that have higher quality fruit and improved resistance to pathogens and changing environmental conditions. The USDA-ARS National Clonal Repository for Citrus and Dates (NCGRCD) maintains a very diverse collectio...

  12. Genomic and transcriptomic analyses of the tangerine pathotype of Alternaria alternata in response to oxidative stress

    PubMed Central

    Wang, Mingshuang; Sun, Xuepeng; Yu, Dongliang; Xu, Jianping; Chung, Kuangren; Li, Hongye

    2016-01-01

    The tangerine pathotype of Alternaria alternata produces the A. citri toxin (ACT) and is the causal agent of citrus brown spot that results in significant yield losses worldwide. Both the production of ACT and the ability to detoxify reactive oxygen species (ROS) are required for A. alternata pathogenicity in citrus. In this study, we report the 34.41 Mb genome sequence of strain Z7 of the tangerine pathotype of A. alternata. The host selective ACT gene cluster in strain Z7 was identified, which included 25 genes with 19 of them not reported previously. Of these, 10 genes were present only in the tangerine pathotype, representing the most likely candidate genes for this pathotype specialization. A transcriptome analysis of the global effects of H2O2 on gene expression revealed 1108 up-regulated and 498 down-regulated genes. Expressions of those genes encoding catalase, peroxiredoxin, thioredoxin and glutathione were highly induced. Genes encoding several protein families including kinases, transcription factors, transporters, cytochrome P450, ubiquitin and heat shock proteins were found associated with adaptation to oxidative stress. Our data not only revealed the molecular basis of ACT biosynthesis but also provided new insights into the potential pathways that the phytopathogen A. alternata copes with oxidative stress. PMID:27582273

  13. Genomic and transcriptomic analyses of the tangerine pathotype of Alternaria alternata in response to oxidative stress.

    PubMed

    Wang, Mingshuang; Sun, Xuepeng; Yu, Dongliang; Xu, Jianping; Chung, Kuangren; Li, Hongye

    2016-09-01

    The tangerine pathotype of Alternaria alternata produces the A. citri toxin (ACT) and is the causal agent of citrus brown spot that results in significant yield losses worldwide. Both the production of ACT and the ability to detoxify reactive oxygen species (ROS) are required for A. alternata pathogenicity in citrus. In this study, we report the 34.41 Mb genome sequence of strain Z7 of the tangerine pathotype of A. alternata. The host selective ACT gene cluster in strain Z7 was identified, which included 25 genes with 19 of them not reported previously. Of these, 10 genes were present only in the tangerine pathotype, representing the most likely candidate genes for this pathotype specialization. A transcriptome analysis of the global effects of H2O2 on gene expression revealed 1108 up-regulated and 498 down-regulated genes. Expressions of those genes encoding catalase, peroxiredoxin, thioredoxin and glutathione were highly induced. Genes encoding several protein families including kinases, transcription factors, transporters, cytochrome P450, ubiquitin and heat shock proteins were found associated with adaptation to oxidative stress. Our data not only revealed the molecular basis of ACT biosynthesis but also provided new insights into the potential pathways that the phytopathogen A. alternata copes with oxidative stress.

  14. Development of a Loop-Mediated Isothermal Amplification Assay for Rapid and Specific Identification of ACT Producing Alternaria alternata, the Agent of Brown Spot Disease in Tangerine.

    PubMed

    Moghimi, Hamid; Moradi, Amir; Hamedi, Javad; Basiri, Mina

    2016-03-01

    Rapid, accurate, and easy identification of pathogenic agents has always been important in medicine, veterinary, and agriculture. The brown spot infection is among the most common diseases in tangerine caused by Alternaria alternata. Due to the existence of seven various pathotypes of A. alternata species, it is challenging and time consuming to detect a pathotype responsible for citrus brown spot. In this study, we were seeking a rapid and specific approach to identify the tangerine pathotype within the A. alternata-pathogenic species, using the loop-mediated isothermal amplification (LAMP) method and actts2 gene as a marker molecule. Nine pathogenic samples were obtained from the region of Ramsar, Iran, and certified as A. alternata-pathogenic isolates. Specific primers were designed for regions coding for Alternaria citri toxin (ACT), and the PCR and LAMP reactions were performed. Our data showed that the primers designed for the tangerine pathotype of A. alternata were specific, and in both reactions, positive results were only observed in desired pathotypes. In the other pathotypes of this species as well as other standard fungal samples as negative controls, no positive result was observed. Therefore, our results suggest the possibility to detect the tangerine-specific A. alternata pathotype from other related species with a high accuracy and in early stages of the disease.

  15. 75 FR 17289 - Citrus Seed Imports; Citrus Greening and Citrus Variegated Chlorosis

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-04-06

    ... Animal and Plant Health Inspection Service 7 CFR Part 319 RIN 0579-AD07 Citrus Seed Imports; Citrus Greening and Citrus Variegated Chlorosis AGENCY: Animal and Plant Health Inspection Service, USDA. ACTION...'' that attack the vascular system of host plants. The pathogens are phloem-limited, inhabiting the...

  16. The Response of Citrus limon Seedlings to a Symbiont, Glomus etunicatus, and a Pathogen, Radopholus similis.

    PubMed

    O'Bannon, J H; Nemec, S

    1979-07-01

    The influences of a vesicular-arbuscular mycorrhiza (Glomus etunicatus) and burrowing nematode (Radophohts similis), alone and in combination, on the growth of rough lemon (Citrus limon) seedlings were studied in the greenhouse. Growth of mycorrhizal seedlings was significantly greater than that of nonmycorrhizal seedlings or seedlings inoculated with R. sindlis. Mycorrhizal stimulation of seedling growth was inhibited by nematode infection. When seedlings were inoculated with G. etunicatus arid R. similis, suppression of seedling growth by R. similis was less on VAM seedlings than on nonmycorrhizal seedlings, Nonmycorrhizal seedlings infected with R. similis were significantly smaller than nonmycorrhizal seedlings free of R. similis. Vesicle formation and mycelia growth were less in nematode-infected roots.

  17. The Response of Citrus limon Seedlings to a Symbiont, Glomus etunicatus, and a Pathogen, Radopholus similis

    PubMed Central

    O'Bannon, J. H.; Nemec, S.

    1979-01-01

    The influences of a vesicular-arbuscular mycorrhiza (Glomus etunicatus) and burrowing nematode (Radophohts similis), alone and in combination, on the growth of rough lemon (Citrus limon) seedlings were studied in the greenhouse. Growth of mycorrhizal seedlings was significantly greater than that of nonmycorrhizal seedlings or seedlings inoculated with R. sindlis. Mycorrhizal stimulation of seedling growth was inhibited by nematode infection. When seedlings were inoculated with G. etunicatus arid R. similis, suppression of seedling growth by R. similis was less on VAM seedlings than on nonmycorrhizal seedlings, Nonmycorrhizal seedlings infected with R. similis were significantly smaller than nonmycorrhizal seedlings free of R. similis. Vesicle formation and mycelia growth were less in nematode-infected roots. PMID:19300646

  18. Disruption of Glycosylphosphatidylinositol-Anchored Lipid Transfer Protein Gene Altered Cuticular Lipid Composition, Increased Plastoglobules, and Enhanced Susceptibility to Infection by the Fungal Pathogen Alternaria brassicicola1[W

    PubMed Central

    Lee, Saet Buyl; Go, Young Sam; Bae, Hyun-Jong; Park, Jong Ho; Cho, Sung Ho; Cho, Hong Joo; Lee, Dong Sook; Park, Ohkmae K.; Hwang, Inhwan; Suh, Mi Chung

    2009-01-01

    All aerial parts of vascular plants are covered with cuticular waxes, which are synthesized by extensive export of intracellular lipids from epidermal cells to the surface. Although it has been suggested that plant lipid transfer proteins (LTPs) are involved in cuticular lipid transport, the in planta evidence is still not clear. In this study, a glycosylphosphatidylinositol-anchored LTP (LTPG1) showing higher expression in epidermal peels of stems than in stems was identified from an Arabidopsis (Arabidopsis thaliana) genome-wide microarray analysis. The expression of LTPG1 was observed in various tissues, including the epidermis, stem cortex, vascular bundles, mesophyll cells, root tips, pollen, and early-developing seeds. LTPG1 was found to be localized in the plasma membrane. Disruption of the LTPG1 gene caused alterations of cuticular lipid composition, but no significant changes on total wax and cutin monomer loads were seen. The largest reduction (10 mass %) in the ltpg1 mutant was observed in the C29 alkane, which is the major component of cuticular waxes in the stems and siliques. The reduced content was overcome by increases of the C29 secondary alcohols and C29 ketone wax loads. The ultrastructure analysis of ltpg1 showed a more diffuse cuticular layer structure, protrusions of the cytoplasm into the vacuole in the epidermis, and an increase of plastoglobules in the stem cortex and leaf mesophyll cells. Furthermore, the ltpg1 mutant was more susceptible to infection by the fungus Alternaria brassicicola than the wild type. Taken together, these results indicated that LTPG1 contributed either directly or indirectly to cuticular lipid accumulation. PMID:19321705

  19. Bacterial diversity analysis of Huanglongbing pathogen-infected citrus, using PhyloChip and 16S rRNA gene clone library sequencing

    SciTech Connect

    Shankar Sagaram, U.; DeAngelis, K.M.; Trivedi, P.; Andersen, G.L.; Lu, S.-E.; Wang, N.

    2009-03-01

    The bacterial diversity associated with citrus leaf midribs was characterized 1 from citrus groves that contained the Huanglongbing (HLB) pathogen, which has yet to be cultivated in vitro. We employed a combination of high-density phylogenetic 16S rDNA microarray and 16S rDNA clone library sequencing to determine the microbial community composition of symptomatic and asymptomatic citrus midribs. Our results revealed that citrus leaf midribs can support a diversity of microbes. PhyloChip analysis indicated that 47 orders of bacteria from 15 phyla were present in the citrus leaf midribs while 20 orders from phyla were observed with the cloning and sequencing method. PhyloChip arrays indicated that nine taxa were significantly more abundant in symptomatic midribs compared to asymptomatic midribs. Candidatus Liberibacter asiaticus (Las) was detected at a very low level in asymptomatic plants, but was over 200 times more abundant in symptomatic plants. The PhyloChip analysis was further verified by sequencing 16S rDNA clone libraries, which indicated the dominance of Las in symptomatic leaves. These data implicate Las as the pathogen responsible for HLB disease. Citrus is the most important commercial fruit crop in Florida. In recent years, citrus Huanglongbing (HLB), also called citrus greening, has severely affected Florida's citrus production and hence has drawn an enormous amount of attention. HLB is one of the most devastating diseases of citrus (6,13), characterized by blotchy mottling with green islands on leaves, as well as stunting, fruit decline, and small, lopsided fruits with poor coloration. The disease tends to be associated with a phloem-limited fastidious {alpha}-proteobacterium given a provisional Candidatus status (Candidatus Liberobacter spp. later changed to Candidatus Liberibacter spp.) in nomenclature (18,25,34). Previous studies indicate that HLB infection causes disorder in the phloem and severely impairs the translocation of assimilates in host

  20. Occurrence of Leaf Blight on Cosmos Caused by Alternaria cosmosa in Korea.

    PubMed

    Deng, Jian Xin; Lee, Ji Hye; Paul, Narayan Chandra; Cho, Hye Sun; Lee, Hyang Burm; Yu, Seung Hun

    2015-03-01

    In 2011, a leaf blight disease was observed on cosmos (Cosmos bipinnatus) leaves in Nonsan, Korea. The causal pathogen was isolated and identified based on morphological and molecular approaches. Morphological characteristics of the pathogen matched well with the Alternaria cosmosa and also easily distinguishable from Alternaria zinniae reported from cosmos seeds by producing branched beak. Phylogenetically, the pathogen could not be distinguished from A. passiflorae based on the sequence analysis of a combined data set of Alt a1 and gpd genes. However, A. passiflorae was distinguished from the present species by having conidiophores with 4 to 5 conidiogenous loci. The results indicate that the present Alternaria species is A. cosmosa. Pathogenicity tests revealed that the isolate was pathogenic to the leaves of Cosmos bipinnatus. This is the first report of Alternaria blight disease caused by A. cosmosa on cosmos in Korea.

  1. Ectopic expression of Xylella fastidiosa rpfF conferring production of diffusible signal factor in transgenic tobacco and citrus alters pathogen behavior and reduces disease severity.

    PubMed

    Caserta, Raquel; Souza-Neto, Reinaldo Rodrigues; Takita, Marco Aurelio; Lindow, Steven; Souza, Alessandra De

    2017-08-04

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

  2. Structural and Physiological Analyses of the Alkanesulphonate-Binding Protein (SsuA) of the Citrus Pathogen Xanthomonas citri

    PubMed Central

    Tófoli de Araújo, Fabiano; Bolanos-Garcia, Victor M.; Pereira, Cristiane T.; Sanches, Mario; Oshiro, Elisa E.; Ferreira, Rita C. C.; Chigardze, Dimitri Y.; Barbosa, João Alexandre Gonçalves; de Souza Ferreira, Luís Carlos; Benedetti, Celso E.; Blundell, Tom L.; Balan, Andrea

    2013-01-01

    Background The uptake of sulphur-containing compounds plays a pivotal role in the physiology of bacteria that live in aerobic soils where organosulfur compounds such as sulphonates and sulphate esters represent more than 95% of the available sulphur. Until now, no information has been available on the uptake of sulphonates by bacterial plant pathogens, particularly those of the Xanthomonas genus, which encompasses several pathogenic species. In the present study, we characterised the alkanesulphonate uptake system (Ssu) of Xanthomonas axonopodis pv. citri 306 strain (X. citri), the etiological agent of citrus canker. Methodology/Principal Findings A single operon-like gene cluster (ssuEDACB) that encodes both the sulphur uptake system and enzymes involved in desulphurisation was detected in the genomes of X. citri and of the closely related species. We characterised X. citri SsuA protein, a periplasmic alkanesulphonate-binding protein that, together with SsuC and SsuB, defines the alkanesulphonate uptake system. The crystal structure of SsuA bound to MOPS, MES and HEPES, which is herein described for the first time, provides evidence for the importance of a conserved dipole in sulphate group coordination, identifies specific amino acids interacting with the sulphate group and shows the presence of a rather large binding pocket that explains the rather wide range of molecules recognised by the protein. Isolation of an isogenic ssuA-knockout derivative of the X. citri 306 strain showed that disruption of alkanesulphonate uptake affects both xanthan gum production and generation of canker lesions in sweet orange leaves. Conclusions/Significance The present study unravels unique structural and functional features of the X. citri SsuA protein and provides the first experimental evidence that an ABC uptake system affects the virulence of this phytopathogen. PMID:24282519

  3. Genome sequence of the necrotrophic fungus Penicillium digitatum, the main postharvest pathogen of citrus

    PubMed Central

    2012-01-01

    Background Penicillium digitatum is a fungal necrotroph causing a common citrus postharvest disease known as green mold. In order to gain insight into the genetic bases of its virulence mechanisms and its high degree of host-specificity, the genomes of two P. digitatum strains that differ in their antifungal resistance traits have been sequenced and compared with those of 28 other Pezizomycotina. Results The two sequenced genomes are highly similar, but important differences between them include the presence of a unique gene cluster in the resistant strain, and mutations previously shown to confer fungicide resistance. The two strains, which were isolated in Spain, and another isolated in China have identical mitochondrial genome sequences suggesting a recent worldwide expansion of the species. Comparison with the closely-related but non-phytopathogenic P. chrysogenum reveals a much smaller gene content in P. digitatum, consistent with a more specialized lifestyle. We show that large regions of the P. chrysogenum genome, including entire supercontigs, are absent from P. digitatum, and that this is the result of large gene family expansions rather than acquisition through horizontal gene transfer. Our analysis of the P. digitatum genome is indicative of heterothallic sexual reproduction and reveals the molecular basis for the inability of this species to assimilate nitrate or produce the metabolites patulin and penicillin. Finally, we identify the predicted secretome, which provides a first approximation to the protein repertoire used during invasive growth. Conclusions The complete genome of P. digitatum, the first of a phytopathogenic Penicillium species, is a valuable tool for understanding the virulence mechanisms and host-specificity of this economically important pest. PMID:23171342

  4. Genome sequence of the necrotrophic fungus Penicillium digitatum, the main postharvest pathogen of citrus.

    PubMed

    Marcet-Houben, Marina; Ballester, Ana-Rosa; de la Fuente, Beatriz; Harries, Eleonora; Marcos, Jose F; González-Candelas, Luis; Gabaldón, Toni

    2012-11-21

    Penicillium digitatum is a fungal necrotroph causing a common citrus postharvest disease known as green mold. In order to gain insight into the genetic bases of its virulence mechanisms and its high degree of host-specificity, the genomes of two P. digitatum strains that differ in their antifungal resistance traits have been sequenced and compared with those of 28 other Pezizomycotina. The two sequenced genomes are highly similar, but important differences between them include the presence of a unique gene cluster in the resistant strain, and mutations previously shown to confer fungicide resistance. The two strains, which were isolated in Spain, and another isolated in China have identical mitochondrial genome sequences suggesting a recent worldwide expansion of the species. Comparison with the closely-related but non-phytopathogenic P. chrysogenum reveals a much smaller gene content in P. digitatum, consistent with a more specialized lifestyle. We show that large regions of the P. chrysogenum genome, including entire supercontigs, are absent from P. digitatum, and that this is the result of large gene family expansions rather than acquisition through horizontal gene transfer. Our analysis of the P. digitatum genome is indicative of heterothallic sexual reproduction and reveals the molecular basis for the inability of this species to assimilate nitrate or produce the metabolites patulin and penicillin. Finally, we identify the predicted secretome, which provides a first approximation to the protein repertoire used during invasive growth. The complete genome of P. digitatum, the first of a phytopathogenic Penicillium species, is a valuable tool for understanding the virulence mechanisms and host-specificity of this economically important pest.

  5. Transcriptomic profile of tobacco in response to Alternaria longipes and Alternaria alternata infections

    PubMed Central

    Duan, Shengchang; Ma, Xiao; Chen, Wei; Wan, Wenting; He, Yuqi; Ma, Xiaoqin; Ma, Yujin; Long, Ni; Tan, Yuntao; Wang, Yangzi; Hou, Yujie; Dong, Yang

    2016-01-01

    Tobacco brown spot caused by Alternaria fungal species is one of the most damaging diseases, and results in significant yield losses. However, little is known about the systematic response of tobacco to this fungal infection. To fill this knowledge gap, de novo assemblies of tobacco leaf transcriptomes were obtained in cultivars V2 and NC89 after the inoculation of either Alternaria longipes (AL) or Alternaria alternata (AA) at three different time points. We studied the gene expression profile of each cultivar-pathogen combination, and identified eight differentially expressed genes shared among all combinations. Gene ontology enrichment analysis of the differentially expressed genes revealed key components during the fungal infection, which included regulation of gene expression (GO:0010468), regulation of RNA metabolic process (GO:0051252), tetrapyrrole binding (GO:0046906), and external encapsulating structure (GO:0030312). Further analyses of the continuously upregulated/downregulated genes and the resistance genes demonstrated that the gene expression profile upon fungal infection was contingent on the specific cultivar and pathogen. In conclusion, this study provides a solid foundation for the investigation of plant-pathogen interaction, and is of great importance for disease prevention and molecular breeding. PMID:27157477

  6. The potential for citrus cryotherapy

    USDA-ARS?s Scientific Manuscript database

    Citrus collections of pathogen-free plants are needed for breeding, research, and distribution to the user community. The Citrus Research Board funded research project “Development of cryotherapy as an improved method of eliminating graft transmissible pathogens in Citrus” sought to use cryotherapy,...

  7. Serological detection of ‘Candidatus Liberibacter asiaticus’ in citrus, and identification by GeLC-MS/MS of a chaperone protein responding to cellular pathogens

    PubMed Central

    Ding, Fang; Duan, Yongping; Yuan, Qing; Shao, Jonathan; Hartung, John S.

    2016-01-01

    We describe experiments with antibodies against ‘Candidatus Liberibacter asiaticus used to detect the pathogen in infected plants. We used scFv selected to bind epitopes exposed on the surface of the bacterium in tissue prints, with secondary monoclonal antibodies directed at a FLAG epitope included at the carboxyl end of the scFv. Unexpectedly, the anti-FLAG secondary antibody produced positive results with CaLas diseased samples when the primary scFv were not used. The anti-FLAG monoclonal antibody (Mab) also identified plants infected with other vascular pathogens. We then identified a paralogous group of secreted chaperone proteins in the HSP-90 family that contained the amino acid sequence DDDDK identical to the carboxy-terminal sequence of the FLAG epitope. A rabbit polyclonal antibody against one of the same epitopes combined with a goat anti-rabbit secondary antibody produced very strong purple color in individual phloem cells, as expected for this pathogen. These results were entirely specific for CaLas-infected citrus. The simplicity, cost and ability to scale the tissue print assay makes this an attractive assay to complement PCR-based assays currently in use. The partial FLAG epitope may itself be useful as a molecular marker for the rapid screening of citrus plants for the presence of vascular pathogens. PMID:27381064

  8. Transcriptional analysis of the sweet orange interaction with the citrus canker pathogens Xanthomonas axonopodis pv. citri and Xanthomonas axonopodis pv. aurantifolii.

    PubMed

    Cernadas, Raúl Andrés; Camillo, Luciana Rodrigues; Benedetti, Celso Eduardo

    2008-09-01

    Xanthomonas axonopodis pv. citri (Xac) and Xanthomonas axonopodis pv. aurantifolii pathotype C (Xaa) are responsible for citrus canker disease; however, while Xac causes canker on all citrus varieties, Xaa is restricted to Mexican lime, and in sweet oranges it triggers a defence response. To gain insights into the differential pathogenicity exhibited by Xac and Xaa and to survey the early molecular events leading to canker development, a detailed transcriptional analysis of sweet orange plants infected with the pathogens was performed. Using differential display, suppressed subtractive hybridization and microarrays, we identified changes in transcript levels in approximately 2.0% of the approximately 32,000 citrus genes examined. Genes with altered expression in response to Xac/Xaa surveyed at 6 and 48 h post-infection (hpi) were associated with cell-wall modifications, cell division and expansion, vesicle trafficking, disease resistance, carbon and nitrogen metabolism, and responses to hormones auxin, gibberellin and ethylene. Most of the genes that were commonly modulated by Xac and Xaa were associated with basal defences triggered by pathogen-associated molecular patterns, including those involved in reactive oxygen species production and lignification. Significantly, we detected clear changes in the transcriptional profiles of defence, cell-wall, vesicle trafficking and cell growth-related genes in Xac-infected leaves between 6 and 48 hpi. This is consistent with the notion that Xac suppresses host defences early during infection and simultaneously changes the physiological status of the host cells, reprogramming them for division and growth. Notably, brefeldin A, an inhibitor of vesicle trafficking, retarded canker development. In contrast, Xaa triggered a mitogen-activated protein kinase signalling pathway involving WRKY and ethylene-responsive transcriptional factors known to activate downstream defence genes.

  9. Citrus phenylpropanoids and defence against pathogens. Part II: gene expression and metabolite accumulation in the response of fruits to Penicillium digitatum infection.

    PubMed

    Ballester, Ana-Rosa; Teresa Lafuente, M; González-Candelas, Luis

    2013-01-01

    The effect of infection of Citrus sinensis (var. Navelina) fruits with Penicillium digitatum was studied at gene expression and metabolite levels. In this study, expression of genes involved in the phenylpropanoid pathway was studied in the flavedo (outer coloured part of the peel) and albedo (inner white part) in response to pathogen infection. Results of the time-course experiment showed that maximal expression of 10 out of 17 phenylpropanoid genes analysed occurred at 48h post-inoculation, when decay symptoms started to appear, and mRNA levels either kept constant or decreased after 72h post-inoculation. To further investigate the putative involvement of the phenylpropanoid pathway in the defence of citrus fruit, changes in the metabolic profile of both tissues infected with P. digitatum was studied by means of HPLC-PDA-FD. Metabolite accumulation levels along the time course suggest that flavanones, flavones, polymethoxylated flavones and scoparone are induced in citrus fruit in response to P. digitatum infection, although with different trends depending on the tissue.

  10. Alternaria Toxins: Potential Virulence Factors and Genes Related to Pathogenesis

    PubMed Central

    Meena, Mukesh; Gupta, Sanjay K.; Swapnil, Prashant; Zehra, Andleeb; Dubey, Manish K.; Upadhyay, Ram S.

    2017-01-01

    Alternaria is an important fungus to study due to their different life style from saprophytes to endophytes and a very successful fungal pathogen that causes diseases to a number of economically important crops. Alternaria species have been well-characterized for the production of different host-specific toxins (HSTs) and non-host specific toxins (nHSTs) which depend upon their physiological and morphological stages. The pathogenicity of Alternaria species depends on host susceptibility or resistance as well as quantitative production of HSTs and nHSTs. These toxins are chemically low molecular weight secondary metabolites (SMs). The effects of toxins are mainly on different parts of cells like mitochondria, chloroplast, plasma membrane, Golgi complex, nucleus, etc. Alternaria species produce several nHSTs such as brefeldin A, tenuazonic acid, tentoxin, and zinniol. HSTs that act in very low concentrations affect only certain plant varieties or genotype and play a role in determining the host range of specificity of plant pathogens. The commonly known HSTs are AAL-, AK-, AM-, AF-, ACR-, and ACT-toxins which are named by their host specificity and these toxins are classified into different family groups. The HSTs are differentiated on the basis of bio-statistical and other molecular analyses. All these toxins have different mode of action, biochemical reactions and signaling mechanisms to cause diseases. Different species of Alternaria produced toxins which reveal its biochemical and genetic effects on itself as well as on its host cells tissues. The genes responsible for the production of HSTs are found on the conditionally dispensable chromosomes (CDCs) which have been well characterized. Different bio-statistical methods like basic local alignment search tool (BLAST) data analysis used for the annotation of gene prediction, pathogenicity-related genes may provide surprising knowledge in present and future. PMID:28848500

  11. Phenylacetaldehyde O-methyloxime: a volatile compound produced by grapefruit leaves infected with the citrus canker pathogen, Xanthomonas axonopodis pv. citri.

    PubMed

    Zhang, Aijun; Hartung, John S

    2005-06-29

    An aldehyde oxime O-methyl ether, phenylacetaldehyde O-methyloxime, was detected using solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS) in the headspace above grapefruit leaves infected with Xanthomonas axonopodis pv. citri, the causal agent of citrus bacterial canker disease (CBCD). This disease is a major phytosanitary concern, and an eradication campaign against it is currently underway in Florida. Phenylacetaldehyde O-methyloxime has been reported to be produced by other plants and fragrant flowers, but it was not observed in the headspace above uninfected grapefruit leaves, the pathogenic bacterium X. axonopodis pv. citri itself, or grapefruit leaves infected with another closely related bacterial pathogen, X. axonopodis pv. citrumelo, which causes citrus bacterial spot, a disease of no phytosanitary significance. It was also not detected from CBCD infected fruits, including orange, lemon, grapefruit, and lime. We conclude that phenylacetaldehyde O-methyloxime may potentially be used to identify CBCD infestations. However, more intensive studies will be required to fully evaluate the potential of phenylacetaldehyde O-methyloxime as a diagnostic compound for CBCD. Using SPME and GC-MS to measure phenylacetaldehyde O-methyloxime may provide an easy and feasible tool to complement current methods used to detect X. axonopodis pv. citri in environmental samples.

  12. Calculation of diagnostic parameters of advanced serological and molecular tissue-print methods for detection of Citrus tristeza virus: a model for other plant pathogens.

    PubMed

    Vidal, E; Yokomi, R K; Moreno, A; Bertolini, E; Cambra, M

    2012-01-01

    Citrus tristeza virus (CTV) is one of the most important virus diseases that affect citrus. Control of CTV is achieved by grafting selected virus-free citrus scions onto CTV-tolerant or -resistant rootstocks. Quarantine and certification programs are essential for avoiding the entry and propagation of severe strains of CTV. Citrus nurseries in Spain and central California (United States) maintain zero-tolerance policies for CTV that require sensitive, specific, and reliable pathogen-detection methods. Tissue-print (TP) real-time reverse-transcriptase polymerase chain reaction (RT-PCR) assay was compared with the validated TP enzyme-linked immunosorbent assay (ELISA), using the CTV-specific monoclonal antibodies 3DF1 and 3CA5, for CTV detection. In total, 1,395 samples from healthy and CTV-infected nursery and mature tree plants were analyzed with both methods. The total agreement between both detection methods was substantial (Cohen's kappa index of 0.77 ± 0.03). The diagnostic parameters of each technique (i.e., the sensitivity, specificity, and likelihood ratios) were evaluated in a second test involving 658 Citrus macrophylla nursery plants. Mexican lime indexing was used to evaluate samples with discrepant results in the analysis. For TP-ELISA, a sensitivity of 0.8015, a specificity of 0.9963, and a positive and negative likelihood ratio of 216.42 and 0.199, respectively, were estimated. For TP real-time RT-PCR, a sensitivity of 0.9820, a specificity of 0.8519, and a positive and negative likelihood ratio of 6.63 and 0.021, respectively, were estimated. These diagnostic parameters show that TP real-time RT-PCR was the most sensitive technique, whereas TP-ELISA showed the highest specificity, validating the use of the molecular technique for routine CTV-detection purposes. In addition, our results show that the combination of both techniques can accurately substitute for the conventional biological Mexican lime index for the detection of CTV. The calculation of

  13. Antennal and behavioral response of the Asian citrus psyllid (Diaphorina citri Kuwayama) to degradation products of citrus volatiles

    USDA-ARS?s Scientific Manuscript database

    Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae) vectors the bacterial causal pathogen of the deadly citrus disease, Huanglongbing (citrus greening) which is a major threat to citrus industry worldwide. We studied antennal and behavioral responses to principal components of head...

  14. Citrus quarantine, sanitary and certification programs in the USA. Prevention of introduction and distribution of citrus diseases

    USDA-ARS?s Scientific Manuscript database

    Citrus germplasm originated in Australasia, the Far East, and Africa, thus all citrus grown in the New World was imported. This importation of citrus also resulted in importation of graft transmissible pathogens of citrus, many of which are latent in their original host but can cause epidemics of t...

  15. Seasonal prevalence of the insect pathogenic fungus Colletotrichum nymphaeae in Brazilian citrus groves under different chemical pesticide regimes

    USDA-ARS?s Scientific Manuscript database

    In a field study, we identified an endemic fungal entomopathogen, known as the 'salmão' fungus infecting populations of citrus scale, Praelongorthezia praelonga Douglas. The identification of this fungus is close to Colletotrichum nymphaeae (Sordariomycetes: Glomerellales) based on morphological sim...

  16. A polyketide synthase gene, ACRTS2, is responsible for biosynthesis of host-selective ACR-toxin in the rough lemon pathotype of Alternaria alternata.

    PubMed

    Izumi, Y; Ohtani, K; Miyamoto, Y; Masunaka, A; Fukumoto, T; Gomi, K; Tada, Y; Ichimura, K; Peever, T L; Akimitsu, K

    2012-11-01

    The rough lemon pathotype of Alternaria alternata produces host-selective ACR-toxin and causes Alternaria leaf spot disease of rough lemon (Citrus jambhiri). The structure of ACR-toxin I (MW = 496) consists of a polyketide with an α-dihydropyrone ring in a 19-carbon polyalcohol. Genes responsible for toxin production were localized to a 1.5-Mb chromosome in the genome of the rough lemon pathotype. Sequence analysis of this chromosome revealed an 8,338-bp open reading frame, ACRTS2, that was present only in the genomes of ACR-toxin-producing isolates. ACRTS2 is predicted to encode a putative polyketide synthase of 2,513 amino acids and belongs to the fungal reducing type I polyketide synthases. Typical polyketide functional domains were identified in the predicted amino acid sequence, including β-ketoacyl synthase, acyl transferase, methyl transferase, dehydratase, β-ketoreductase, and phosphopantetheine attachment site domains. Combined use of homologous recombination-mediated gene disruption and RNA silencing allowed examination of the functional role of multiple paralogs in ACR-toxin production. ACRTS2 was found to be essential for ACR-toxin production and pathogenicity of the rough lemon pathotype of A. alternata.

  17. Horsfall-Barratt recalibration and replicated severity estimates of citrus canker

    USDA-ARS?s Scientific Manuscript database

    Citrus canker is a serious disease of citrus in tropical and subtropical citrus growing regions. Accurate and precise assessment of citrus canker and other plant pathogens is needed to obtain good quality data. Citrus canker assessment data were used to ascertain some of the mechanics of the Horsfal...

  18. Combining 'omics and microscopy to visualize interactions between the Asian citrus psyllid vector and the Huanglongbing pathogen Candidatus Liberibacter asiaticus in the insect gut.

    PubMed

    Kruse, Angela; Fattah-Hosseini, Somayeh; Saha, Surya; Johnson, Richard; Warwick, EricaRose; Sturgeon, Kasie; Mueller, Lukas; MacCoss, Michael J; Shatters, Robert G; Cilia Heck, Michelle

    2017-01-01

    Huanglongbing, or citrus greening disease, is an economically devastating bacterial disease of citrus. It is associated with infection by the gram-negative bacterium Candidatus Liberibacter asiaticus (CLas). CLas is transmitted by Diaphorina citri, the Asian citrus psyllid (ACP). For insect transmission to occur, CLas must be ingested during feeding on infected phloem sap and cross the gut barrier to gain entry into the insect vector. To investigate the effects of CLas exposure at the gut-pathogen interface, we performed RNAseq and mass spectrometry-based proteomics to analyze the transcriptome and proteome, respectively, of ACP gut tissue. CLas exposure resulted in changes in pathways involving the TCA cycle, iron metabolism, insecticide resistance and the insect's immune system. We identified 83 long non-coding RNAs that are responsive to CLas, two of which appear to be specific to the ACP. Proteomics analysis also enabled us to determine that Wolbachia, a symbiont of the ACP, undergoes proteome regulation when CLas is present. Fluorescent in situ hybridization (FISH) confirmed that Wolbachia and CLas inhabit the same ACP gut cells, but do not co-localize within those cells. Wolbachia cells are prevalent throughout the gut epithelial cell cytoplasm, and Wolbachia titer is more variable in the guts of CLas exposed insects. CLas is detected on the luminal membrane, in puncta within the gut epithelial cell cytoplasm, along actin filaments in the gut visceral muscles, and rarely, in association with gut cell nuclei. Our study provides a snapshot of how the psyllid gut copes with CLas exposure and provides information on pathways and proteins for targeted disruption of CLas-vector interactions at the gut interface.

  19. Combining 'omics and microscopy to visualize interactions between the Asian citrus psyllid vector and the Huanglongbing pathogen Candidatus Liberibacter asiaticus in the insect gut

    PubMed Central

    Kruse, Angela; Fattah-Hosseini, Somayeh; Saha, Surya; Johnson, Richard; Warwick, EricaRose; Sturgeon, Kasie; Mueller, Lukas; MacCoss, Michael J.; Shatters, Robert G.

    2017-01-01

    Huanglongbing, or citrus greening disease, is an economically devastating bacterial disease of citrus. It is associated with infection by the gram-negative bacterium Candidatus Liberibacter asiaticus (CLas). CLas is transmitted by Diaphorina citri, the Asian citrus psyllid (ACP). For insect transmission to occur, CLas must be ingested during feeding on infected phloem sap and cross the gut barrier to gain entry into the insect vector. To investigate the effects of CLas exposure at the gut-pathogen interface, we performed RNAseq and mass spectrometry-based proteomics to analyze the transcriptome and proteome, respectively, of ACP gut tissue. CLas exposure resulted in changes in pathways involving the TCA cycle, iron metabolism, insecticide resistance and the insect’s immune system. We identified 83 long non-coding RNAs that are responsive to CLas, two of which appear to be specific to the ACP. Proteomics analysis also enabled us to determine that Wolbachia, a symbiont of the ACP, undergoes proteome regulation when CLas is present. Fluorescent in situ hybridization (FISH) confirmed that Wolbachia and CLas inhabit the same ACP gut cells, but do not co-localize within those cells. Wolbachia cells are prevalent throughout the gut epithelial cell cytoplasm, and Wolbachia titer is more variable in the guts of CLas exposed insects. CLas is detected on the luminal membrane, in puncta within the gut epithelial cell cytoplasm, along actin filaments in the gut visceral muscles, and rarely, in association with gut cell nuclei. Our study provides a snapshot of how the psyllid gut copes with CLas exposure and provides information on pathways and proteins for targeted disruption of CLas-vector interactions at the gut interface. PMID:28632769

  20. A plant natriuretic peptide-like molecule of the pathogen Xanthomonas axonopodis pv. citri causes rapid changes in the proteome of its citrus host

    PubMed Central

    2010-01-01

    Background Plant natriuretic peptides (PNPs) belong to a novel class of peptidic signaling molecules that share some structural similarity to the N-terminal domain of expansins and affect physiological processes such as water and ion homeostasis at nano-molar concentrations. The citrus pathogen Xanthomonas axonopodis pv. citri possesses a PNP-like peptide (XacPNP) uniquely present in this bacteria. Previously we observed that the expression of XacPNP is induced upon infection and that lesions produced in leaves infected with a XacPNP deletion mutant were more necrotic and lead to earlier bacterial cell death, suggesting that the plant-like bacterial PNP enables the plant pathogen to modify host responses in order to create conditions favorable to its own survival. Results Here we measured chlorophyll fluorescence parameters and water potential of citrus leaves infiltrated with recombinant purified XacPNP and demonstrate that the peptide improves the physiological conditions of the tissue. Importantly, the proteomic analysis revealed that these responses are mirrored by rapid changes in the host proteome that include the up-regulation of Rubisco activase, ATP synthase CF1 α subunit, maturase K, and α- and β-tubulin. Conclusions We demonstrate that XacPNP induces changes in host photosynthesis at the level of protein expression and in photosynthetic efficiency in particular. Our findings suggest that the biotrophic pathogen can use the plant-like hormone to modulate the host cellular environment and in particular host metabolism and that such modulations weaken host defence. PMID:20302677

  1. Adenylyl cyclase is required for cAMP production, growth, conidial germination, and virulence in the citrus green mold pathogen Penicillium digitatum.

    PubMed

    Wang, Weili; Wang, Mingshuang; Wang, Jiye; Zhu, Congyi; Chung, Kuang-Ren; Li, Hongye

    2016-11-01

    Penicillium digitatum is the causative agent of green mold decay on citrus fruit. The cAMP-mediated signaling pathway plays an important role in the transduction of extracellular signals and has been shown to regulate a wide range of developmental processes and pathogenicity in fungal pathogens. We cloned and characterized a Pdac1 gene of P. digitatum, which encodes a polypeptide similar to fungal adenylyl cyclases. Using a loss-of-function mutation in the Pdac1 gene we demonstrated a critical requirement for hyphal growth and conidial germination. Deletion of Pdac1 resulted in decreased accumulation of cAMP and down-regulation of genes encoding a G protein α subunit, both catalytic and regulatory subunits of PKA, and two transcriptional regulators StuA and Som1. Fungal mutants lacking Pdac1 produced abundant conidia, which failed to germinate effectively and displayed an elevated sensitivity to heat treatment. Pdac1 mutant failed to utilize carbohydrates effectively and thus displayed severe growth retardation on rich and synthetic media. Slow growth seen in the Pdac1 mutants could be due to a defect in nutrient sensing and acquisition. Quantitative RT-PCR analysis revealed that Pdac1 was primarily expressed at the early stage of infection. Fungal pathogenicity assayed on citrus fruit revealed that P. digitatum strains impaired for Pdac1 delayed lesion formation. Our results highlight important regulatory roles of adenylyl cyclase-mediated cAMP production in P. digitatum and provide insights into the critical role of cAMP in fungal growth, development and virulence. Copyright © 2016 Elsevier GmbH. All rights reserved.

  2. Postharvest decay control of citrus fruit by preharvest pyrimethanil spray.

    PubMed

    D'Aquino, S; Angioni, A; Suming, D; Palma, A; Schirra, M

    2013-01-01

    Preharvest infections or conidia load on fruit surface by Penicillium digitatum, P. italicum, Alternaria citri and other filamentous fungi can cause important postharvest losses of citrus fruit. Reduction in pruning frequency occurred in the last decade together with un-picked yield that eventually rots on the trees have increased the risk of postharvest decay especially when environmental conditions at picking time are favourable to pathogens' development. Sanitation procedures in the packinghouses, alternate use of postharvest fungicides with different modes of action, along with fungicide application before harvest could be an effective approach to minimize postharvest decay in citrus fruit. The present study investigated the effectiveness of a preharvest treatment with pyrimethanil (PYR), a broad spectrum fungicide, recently registered in different citrus-producing countries for postharvest treatments of citrus fruit and widely used worldwide as a preharvest treatment to control various diseases in different crops. PYR (750 mg/L) was sprayed by a hand-back sprayer at run-off on 'Fremont' mandarins. The day after the treatment, half of the trees were sprayed with a 10(4) conidial suspension of P. digitatum at run-off. Fruit were harvested following 2 or 4 weeks from treatments. Sound or either wounded 2-mm-deep and 2-mm-wide or superficial wound-scratched fruit were stored at 20 degrees C and 90% RH and inspected for decay after 1, 2 or 3 weeks of storage. In fruit harvested after 2 weeks from field treatment, PYR remarkably reduced decay development during two weeks of storage in sound fruit and in wound-scratched fruit and was fairly effective even after 4 weeks from treatment, but was ineffective in fruit wounded 2 mm deep and 2 mm wide. PYR was also effective in reducing preharvest decay incited by P. digitatum, P. italicum and Botrytis cinerea, but not by other pathogens. Results show that preharvest treatment with PYR could be a feasible approach to reduce

  3. Alernaria undulata, a new species from Citrus sinensis

    USDA-ARS?s Scientific Manuscript database

    A new species belonging to the Alternaria infectoria species-group, A undulata, is described from imported fruit of Citrus sinensis (navel Orange). A. undulata was associated with visible lesions that appeared to originate from the peduncular scars of several fruits purchased from a retail display ...

  4. Hyperspectral and Thermal Imaging of Oilseed Rape (Brassica napus) Response to Fungal Species of the Genus Alternaria

    PubMed Central

    Baranowski, Piotr; Jedryczka, Malgorzata; Mazurek, Wojciech; Babula-Skowronska, Danuta; Siedliska, Anna; Kaczmarek, Joanna

    2015-01-01

    In this paper, thermal (8-13 µm) and hyperspectral imaging in visible and near infrared (VNIR) and short wavelength infrared (SWIR) ranges were used to elaborate a method of early detection of biotic stresses caused by fungal species belonging to the genus Alternaria that were host (Alternaria alternata, Alternaria brassicae, and Alternaria brassicicola) and non-host (Alternaria dauci) pathogens to oilseed rape (Brassica napus L.). The measurements of disease severity for chosen dates after inoculation were compared to temperature distributions on infected leaves and to averaged reflectance characteristics. Statistical analysis revealed that leaf temperature distributions on particular days after inoculation and respective spectral characteristics, especially in the SWIR range (1000-2500 nm), significantly differed for the leaves inoculated with A. dauci from the other species of Alternaria as well as from leaves of non-treated plants. The significant differences in leaf temperature of the studied Alternaria species were observed in various stages of infection development. The classification experiments were performed on the hyperspectral data of the leaf surfaces to distinguish days after inoculation and Alternaria species. The second-derivative transformation of the spectral data together with back-propagation neural networks (BNNs) appeared to be the best combination for classification of days after inoculation (prediction accuracy 90.5%) and Alternaria species (prediction accuracy 80.5%). PMID:25826369

  5. Huanglongbing increases Diplodia Stem End Rot in Citrus sinensis

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB), one of the most devastating diseases of citrus is caused by the a-Proteobacteria Candidatus Liberibacter. Diplodia natalensis Pole-Evans is a fungal pathogen which has been known to cause a postharvest stem-end rot of citrus, the pathogen infects citrus fruit under the calyx, an...

  6. Transgenic Indian mustard (Brassica juncea) expressing tomato glucanase leads to arrested growth of Alternaria brassicae.

    PubMed

    Mondal, Kalyan K; Bhattacharya, R C; Koundal, K R; Chatterjee, S C

    2007-02-01

    Brassica juncea is an important oilseed crop of the Indian sub-continent. Yield loss due to fungal disease alternaria leaf spot caused by Alternaria brassicae is a serious problem in cultivation of this crop. Nonavailability of resistance genes within crossable germplasms of Brassica necessitates use of genetic engineering strategies to develop genetic resistance against this pathogen. The pathogenesis related (PR) proteins are group of plant proteins that are toxic to invading fungal pathogens, but are present in plant in trace amount. Thus, overexpression of PR proteins leads to increased resistance to pathogenic fungi in several crops. The PR protein glucanase hydrolyzes a major cell-wall component, glucan, of pathogenic fungi and acts as a plant defense barrier. We report the expression of a class I basic glucanase gene, under the control of CaMV 35S promoter, in Indian mustard and its genetic resistance against alternaria leaf spot. Southern and Northern hybridization confirmed stable integration and expression of the glucanase gene in mustard transgenics. Several independent transgenics were screened in vitro and under poly house conditions for their resistance against Alternaria brassicae. In an in vitro antifungal assay, transgenics arrested hyphal growth of Alternaria brassicae by 15-54%. Under pathogen-challenged conditions in poly house, the transgenics showed restricted number, size and spread of lesions caused by Alternaria brassicae. Also, the onset of disease was delayed in transgenics compared to untransformed parent plants. The results demonstrate potentiality of a PR protein from a heterologous source in developing alternaria leaf spot resistance in Indian mustard.

  7. Role of the pathotype-specific ACRTS1 gene encoding a hydroxylase involved in the biosynthesis of host-selective ACR-toxin in the rough lemon pathotype of Alternaria alternata.

    PubMed

    Izumi, Yuriko; Kamei, Eri; Miyamoto, Yoko; Ohtani, Kouhei; Masunaka, Akira; Fukumoto, Takeshi; Gomi, Kenji; Tada, Yasuomi; Ichimura, Kazuya; Peever, Tobin L; Akimitsu, Kazuya

    2012-08-01

    The rough lemon pathotype of Alternaria alternata produces host-selective ACR-toxin and causes Alternaria leaf spot disease of the rootstock species rough lemon (Citrus jambhiri) and Rangpur lime (C. limonia). Genes controlling toxin production were localized to a 1.5-Mb chromosome carrying the ACR-toxin biosynthesis gene cluster (ACRT) in the genome of the rough lemon pathotype. A genomic BAC clone containing a portion of the ACRT cluster was sequenced which allowed identification of three open reading frames present only in the genomes of ACR-toxin producing isolates. We studied the functional role of one of these open reading frames, ACRTS1 encoding a putative hydroxylase, in ACR-toxin production by homologous recombination-mediated gene disruption. There are at least three copies of ACRTS1 gene in the genome and disruption of two copies of this gene significantly reduced ACR-toxin production as well as pathogenicity; however, transcription of ACRTS1 and production of ACR-toxin were not completely eliminated due to remaining functional copies of the gene. RNA-silencing was used to knock down the remaining ACRTS1 transcripts to levels undetectable by reverse transcription-polymerase chain reaction. The silenced transformants did not produce detectable ACR-toxin and were not pathogenic. These results indicate that ACRTS1 is an essential gene in ACR-toxin biosynthesis in the rough lemon pathotype of A. alternata and is required for full virulence of this fungus.

  8. Calculation of diagnostic parameters of advanced serological and molecular tissue-print methods for detection of Citrus tristeza virus. A model for other plant pathogens

    USDA-ARS?s Scientific Manuscript database

    Citrus tristeza virus (CTV) is one of the most important virus diseases which affect citrus. Control of CTV in Spain and central California is achieved by planting virus-free citrus on CTV-tolerant or -resistant rootstocks. Quarantine and certification programs remain essential to avoid importation ...

  9. Production of the refolded oligopeptide-binding protein (OppA) encoded by the citrus pathogen Xanthomonas axonopodis pv. Citri.

    PubMed

    Balan, A; Ferreira, R C C; Ferreira, L C S

    2008-02-01

    The oligopeptide-binding protein, OppA, binds and ushers oligopeptide substrates to the membrane-associated oligopeptide permease (Opp), a multi-component ABC-type transporter involved in the uptake of oligopeptides expressed by several bacterial species. In the present study, we report the cloning, purification, refolding and conformational analysis of a recombinant OppA protein derived from Xanthomonas axonopodis pv. citri (X. citri), the etiological agent of citrus canker. The oppA gene was expressed in Escherichia coli BL21 (DE3) strain under optimized inducing conditions and the recombinant protein remained largely insoluble. Solubilization was achieved following refolding of the denatured protein. Circular dichroism analysis indicated that the recombinant OppA protein preserved conformational features of orthologs expressed by other bacterial species. The refolded recombinant OppA represents a useful tool for structural and functional analyses of the X. citri protein.

  10. Citrus Genomics

    PubMed Central

    Talon, Manuel; Gmitter Jr., Fred G.

    2008-01-01

    Citrus is one of the most widespread fruit crops globally, with great economic and health value. It is among the most difficult plants to improve through traditional breeding approaches. Currently, there is risk of devastation by diseases threatening to limit production and future availability to the human population. As technologies rapidly advance in genomic science, they are quickly adapted to address the biological challenges of the citrus plant system and the world's industries. The historical developments of linkage mapping, markers and breeding, EST projects, physical mapping, an international citrus genome sequencing project, and critical functional analysis are described. Despite the challenges of working with citrus, there has been substantial progress. Citrus researchers engaged in international collaborations provide optimism about future productivity and contributions to the benefit of citrus industries worldwide and to the human population who can rely on future widespread availability of this health-promoting and aesthetically pleasing fruit crop. PMID:18509486

  11. Reference genes for accurate transcript normalization in citrus genotypes under different experimental conditions.

    PubMed

    Mafra, Valéria; Kubo, Karen S; Alves-Ferreira, Marcio; Ribeiro-Alves, Marcelo; Stuart, Rodrigo M; Boava, Leonardo P; Rodrigues, Carolina M; Machado, Marcos A

    2012-01-01

    Real-time reverse transcription PCR (RT-qPCR) has emerged as an accurate and widely used technique for expression profiling of selected genes. However, obtaining reliable measurements depends on the selection of appropriate reference genes for gene expression normalization. The aim of this work was to assess the expression stability of 15 candidate genes to determine which set of reference genes is best suited for transcript normalization in citrus in different tissues and organs and leaves challenged with five pathogens (Alternaria alternata, Phytophthora parasitica, Xylella fastidiosa and Candidatus Liberibacter asiaticus). We tested traditional genes used for transcript normalization in citrus and orthologs of Arabidopsis thaliana genes described as superior reference genes based on transcriptome data. geNorm and NormFinder algorithms were used to find the best reference genes to normalize all samples and conditions tested. Additionally, each biotic stress was individually analyzed by geNorm. In general, FBOX (encoding a member of the F-box family) and GAPC2 (GAPDH) was the most stable candidate gene set assessed under the different conditions and subsets tested, while CYP (cyclophilin), TUB (tubulin) and CtP (cathepsin) were the least stably expressed genes found. Validation of the best suitable reference genes for normalizing the expression level of the WRKY70 transcription factor in leaves infected with Candidatus Liberibacter asiaticus showed that arbitrary use of reference genes without previous testing could lead to misinterpretation of data. Our results revealed FBOX, SAND (a SAND family protein), GAPC2 and UPL7 (ubiquitin protein ligase 7) to be superior reference genes, and we recommend their use in studies of gene expression in citrus species and relatives. This work constitutes the first systematic analysis for the selection of superior reference genes for transcript normalization in different citrus organs and under biotic stress.

  12. Reference Genes for Accurate Transcript Normalization in Citrus Genotypes under Different Experimental Conditions

    PubMed Central

    Mafra, Valéria; Kubo, Karen S.; Alves-Ferreira, Marcio; Ribeiro-Alves, Marcelo; Stuart, Rodrigo M.; Boava, Leonardo P.; Rodrigues, Carolina M.; Machado, Marcos A.

    2012-01-01

    Real-time reverse transcription PCR (RT-qPCR) has emerged as an accurate and widely used technique for expression profiling of selected genes. However, obtaining reliable measurements depends on the selection of appropriate reference genes for gene expression normalization. The aim of this work was to assess the expression stability of 15 candidate genes to determine which set of reference genes is best suited for transcript normalization in citrus in different tissues and organs and leaves challenged with five pathogens (Alternaria alternata, Phytophthora parasitica, Xylella fastidiosa and Candidatus Liberibacter asiaticus). We tested traditional genes used for transcript normalization in citrus and orthologs of Arabidopsis thaliana genes described as superior reference genes based on transcriptome data. geNorm and NormFinder algorithms were used to find the best reference genes to normalize all samples and conditions tested. Additionally, each biotic stress was individually analyzed by geNorm. In general, FBOX (encoding a member of the F-box family) and GAPC2 (GAPDH) was the most stable candidate gene set assessed under the different conditions and subsets tested, while CYP (cyclophilin), TUB (tubulin) and CtP (cathepsin) were the least stably expressed genes found. Validation of the best suitable reference genes for normalizing the expression level of the WRKY70 transcription factor in leaves infected with Candidatus Liberibacter asiaticus showed that arbitrary use of reference genes without previous testing could lead to misinterpretation of data. Our results revealed FBOX, SAND (a SAND family protein), GAPC2 and UPL7 (ubiquitin protein ligase 7) to be superior reference genes, and we recommend their use in studies of gene expression in citrus species and relatives. This work constitutes the first systematic analysis for the selection of superior reference genes for transcript normalization in different citrus organs and under biotic stress. PMID:22347455

  13. Trailers transporting oranges to processing plants move Asian citrus psyllids

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (citrus greening) is one of the most serious of citrus diseases. Movement of the disease occurs as a result of natural vector-borne infection and by movement of plant material. We demonstrate here that Diaphorina citri Kuwayama (vector of citrus greening pathogens) can be transported i...

  14. Degradation products of citrus volatile organic compounds (VOCs) acting as phagostimulants that increase probing behavior of Asian citrus psyllid

    USDA-ARS?s Scientific Manuscript database

    Volatile phytochemicals play a role in orientation by phytophagous insects. We studied antennal and behavioral responses of the Asian citrus psyllid, Diaphorina citri Kuwayama, vector of the citrus greening disease pathogen. Little or no response to citrus leaf volatiles was detected by electroanten...

  15. Overexpression of a modified plant thionin enhances disease resistance to citrus canker and huanglongbing (HLB, citrus greening)

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB or citrus greening disease) caused by Candidatus Liberibacter asiaticus (Las) is a great threat to the United States citrus industry. Citrus canker is also an economically important disease associated with a bacterial pathogen (Xanthomonas citri). In this study, we characterized e...

  16. Comparison of antifungal activities of Vietnamese citrus essential oils.

    PubMed

    Van Hung, Pham; Chi, Pham Thi Lan; Phi, Nguyen Thi Lan

    2013-03-01

    Citrus essential oils (EOs) are volatile compounds from citrus peels and widely used in perfumes, cosmetics, soaps and aromatherapy. In this study, inhibition of citrus EOs extracted from Vietnamese orange (Citrus sinensis), mandarin (Citrus reticulata Blanco), pomelo (Citrus grandis Osbeck) and lime (Citrus aurantifolia Swingle) on the growth of plant pathogenic fungi, Mucor hiemalis, Penicillium expansum and Fusarium proliferatum was investigated. The EOs of the citrus peels were obtained by cold-pressing method and the antifungal activity of EOs was evaluated using the agar dilution method. The results show that the EOs had significant antifungal activity. Lime EO was the best inhibitor of M. hiemalis and F. proliferatum while pomelo EO was the most effective against P. expansum. These results indicate that citrus EOs can be used as antifungal natural products in the food, pharmaceutical and cosmetic industries.

  17. Movement of Diaphorina citri Kuwayama (Hemiptera: Liviidae) adults between huanglongbing-affected and healthy citrus

    USDA-ARS?s Scientific Manuscript database

    Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Liviidae), is a vector transmitting the pathogen of citrus huanglongbing (HLB, also called yellow shoot disease or citrus greening disease). A typical symptom of citrus HLB is leaf yellowing. ACP adults behaved differently on HLB-affe...

  18. The Pathogenicity Determinant of Citrus Tristeza Virus Causing the Seedling Yellows Syndrome is Located at the 3’-Terminal Region of the Viral Genome

    USDA-ARS?s Scientific Manuscript database

    Citrus tristeza virus (CTV) (genus Closterovirus, family Closteroviridae) causes some of the more important viral diseases of citrus worldwide. The ability to map disease-inducing determinants of CTV is needed to develop better diagnostic and disease control procedures. A distinctive phenotype of s...

  19. In vivo generated Citrus exocortis viroid progeny variants display a range of phenotypes with altered levels of replication, systemic accumulation and pathogenicity

    USDA-ARS?s Scientific Manuscript database

    Citrus exocortis viroid (CEVd) exists as heterogeneous variants in plant hosts. We inoculated RNA transcripts from a CEVd cDNA clone into protoplasts, seedlings and mature plants of citrus and sequenced 240 in vivo generated progeny variants. Selected progeny variants were further used to evaluate t...

  20. Innate and conditioned responses to chemosensory and visual cues in Asian citrus psyllid, Diaphorina citri (Hemiptera: Liviidae), vector of Huanglongbing pathogens

    USDA-ARS?s Scientific Manuscript database

    Asian citrus psyllid (Diaphorina citri) transmits the causal agent of Huanglongbing, a devastating disease that threatens citrus trees worldwide. This psyllid oviposits and develops only on the emerging shoots of its rutaceous host-plants; however, little is known about the mechanisms underlying its...

  1. A MLVA genotyping scheme for global surveillance of the citrus pathogen Xanthomonas citri pv. citri suggests a worldwide geographical expansion of a single genetic lineage.

    PubMed

    Pruvost, Olivier; Magne, Maxime; Boyer, Karine; Leduc, Alice; Tourterel, Christophe; Drevet, Christine; Ravigné, Virginie; Gagnevin, Lionel; Guérin, Fabien; Chiroleu, Frédéric; Koebnik, Ralf; Verdier, Valérie; Vernière, Christian

    2014-01-01

    MultiLocus Variable number of tandem repeat Analysis (MLVA) has been extensively used to examine epidemiological and evolutionary issues on monomorphic human pathogenic bacteria, but not on bacterial plant pathogens of agricultural importance albeit such tools would improve our understanding of their epidemiology, as well as of the history of epidemics on a global scale. Xanthomonas citri pv. citri is a quarantine organism in several countries and a major threat for the citrus industry worldwide. We screened the genomes of Xanthomonas citri pv. citri strain IAPAR 306 and of phylogenetically related xanthomonads for tandem repeats. From these in silico data, an optimized MLVA scheme was developed to assess the global diversity of this monomorphic bacterium. Thirty-one minisatellite loci (MLVA-31) were selected to assess the genetic structure of 129 strains representative of the worldwide pathological and genetic diversity of X. citri pv. citri. Based on Discriminant Analysis of Principal Components (DAPC), four pathotype-specific clusters were defined. DAPC cluster 1 comprised strains that were implicated in the major geographical expansion of X. citri pv. citri during the 20th century. A subset of 12 loci (MLVA-12) resolved 89% of the total diversity and matched the genetic structure revealed by MLVA-31. MLVA-12 is proposed for routine epidemiological identification of X. citri pv. citri, whereas MLVA-31 is proposed for phylogenetic and population genetics studies. MLVA-31 represents an opportunity for international X. citri pv. citri genotyping and data sharing. The MLVA-31 data generated in this study was deposited in the Xanthomonas citri genotyping database (http://www.biopred.net/MLVA/).

  2. Use of Extract of Citrus sinensis as an antimicrobial agent for foodborne zoonotic pathogens and spoilage bacteria

    USDA-ARS?s Scientific Manuscript database

    Foodborne pathogens remain global health problems despite concerted efforts to control the transmission of these microorganisms through food. The resurgence of drug resistant bacteria has renewed interest in developing and testing new sources of antimicrobial agents to control foodborne illness. Thi...

  3. Host-Specific Effects of Toxin from the Rough Lemon Pathotype of Alternaria alternata on Mitochondria.

    PubMed

    Akimitsu, K; Kohmoto, K; Otani, H; Nishimura, S

    1989-03-01

    Host-specific toxin from the rough lemon (Citrus jambhiri Lush) pathotype of Alternaria alternata (ACR toxin) was tested for effects on mitochondria isolated from several citrus species. The toxin caused uncoupling of oxidative phosphorylation and changes in membrane potential in mitochondria from leaves of the susceptible host (rough lemon); the effects differed from those of carbonylcyanide-m-chlorophenylhydrazone, a typical protonophore. ACR toxin also inhibited malate oxidation, apparently because of lack of NAD(+) in the matrix. In contrast, the toxin had no effect on mitochondria from citrus species (Dancy tangerine and Emperor mandarin [Citrus reticulata Blanco], and grapefruit [Citrus paradisi Macf.]) that are not hosts of the fungus. The effects of the toxin on mitochondria from rough lemon are similar to the effects of a host-specific toxin from Helminthosporium maydis (HMT) on mitochondria from T-cytoplasm maize. Both ACR and HMT toxins are highly selective for the respective host plants. HMT toxin and methomyl had no effect (toxic or protective) on the activity of ACR toxin against mitochondria from rough lemon.

  4. Citrus Functional Genomics and Molecular Modeling in Relation to Citrus sinensis (Sweet Orange) Infection with Xylella fastidiosa (Citrus Variegated Chlorosis).

    PubMed

    Dwivedi, Upendra N; Tiwari, Sameeksha; Prasanna, Pragya; Awasthi, Manika; Singh, Swati; Pandey, Veda P

    2016-08-01

    Citrus are among the economically most important fruit tree crops in the world. Citrus variegated chlorosis (CVC), caused by Xylella fastidiosa infection, is a serious disease limiting citrus production at a global scale. With availability of citrus genomic resources, it is now possible to compare citrus expressed sequence tag (EST) data sets and identify single-nucleotide polymorphisms (SNPs) within and among different citrus cultivars that can be exploited for citrus resistance to infections, citrus breeding, among others. We report here, for the first time, SNPs in the EST data sets of X. fastidiosa-infected Citrus sinensis (sweet orange) and their functional annotation that revealed the involvement of eight C. sinensis candidate genes in CVC pathogenesis. Among these genes were xyloglucan endotransglycosylase, myo-inositol-1-phosphate synthase, and peroxidase were found to be involved in plant cell wall metabolism. These have been further investigated by molecular modeling for their role in CVC infection and defense. Molecular docking analyses of the wild and the mutant (SNP containing) types of the selected three enzymes with their respective substrates revealed a significant decrease in the binding affinity of substrates for the mutant enzymes, thus suggesting a decrease in the catalytic efficiency of these enzymes during infection, thereby facilitating a favorable condition for infection by the pathogen. These findings offer novel agrigenomics insights in developing future molecular targets and strategies for citrus fruit cultivation in ways that are resistant to X. fastidiosa infection, and by extension, with greater harvesting efficiency and economic value.

  5. Characterization of Alternaria infectoria extracellular vesicles

    PubMed Central

    Silva, Branca M.A.; Prados-Rosales, Rafael; Espadas-Moreno, Javier; Wolf, Julie M.; Luque-Garcia, Jose L.; Gonçalves, Teresa; Casadevall, Arturo

    2015-01-01

    Many fungi use membrane vesicles to transport complex molecules across their cell walls. Like mammalian exosomes, fungal vesicles contain lipids, proteins, and polysaccharides, many of which are associated with virulence. Here we identify and characterize extracellular vesicles (EVs) in Alternaria infectoria, a ubiquitous, environmental filamentous fungus that is also an opportunistic human pathogen. Examination of the A. infectoria EVs revealed a morphology similar to that of vesicles described in other fungal species. Of note, proteomic analysis detected a reduced number of vesicle-associated proteins. There were two prevalent categories among the 20 identified proteins, including the polysaccharide metabolism group, probably related to plant host invasion or biosynthesis/degradation of cell wall components, and the nuclear proteins, especially DNA repair enzymes. We also found enzymes related to pigment synthesis, adhesion to the host cell, and trafficking of vesicles/organelles/molecules. This is the first time EV secretions have been identified in a filamentous fungus. We believe that these vesicles might have a role in virulence. PMID:24576997

  6. Citrus pulp for cattle.

    PubMed

    Arthington, John D; Kunkle, William E; Martin, Amy M

    2002-07-01

    Citrus pulp is classified as an energy concentrate by-product feed. Citrus by-products fed to beef cattle include citrus molasses, citrus meal, wet citrus pulp, dried citrus pulp, and pelleted citrus pulp; however, in current production systems, pulp (wet, dry, and pelleted) is the only by-product commonly used. Citrus pulp production in the United States is limited to specific subtropical regions, of which south central Florida remains the largest with additional production in California and Texas.

  7. Arbuscular mycorrhiza reduces susceptibility of tomato to Alternaria solani.

    PubMed

    Fritz, Maendy; Jakobsen, Iver; Lyngkjaer, Michael Foged; Thordal-Christensen, Hans; Pons-Kühnemann, Jörn

    2006-09-01

    Mycorrhiza frequently leads to the control of root pathogens, but appears to have the opposite effect on leaf pathogens. In this study, we studied mycorrhizal effects on the development of early blight in tomato (Solanum lycopersicum) caused by the necrotrophic fungus Alternaria solani. Alternaria-induced necrosis and chlorosis of all leaves were studied in mycorrhizal and non-mycorrhizal plants over time course and at different soil P levels. Mycorrhizal tomato plants had significantly less A. solani symptoms than non-mycorrhizal plants, but neither plant growth nor phosphate uptake was enhanced by mycorrhizas. An increased P supply had no effect on disease severity in non-mycorrhizal plants, but led to a higher disease severity in mycorrhizal plants. This was parallel to a P-supply-induced reduction in mycorrhiza formation. The protective effect of mycorrhizas towards development of A. solani has some parallels to induced systemic resistance, mediated by rhizobacteria: both biocontrol agents are root-associated organisms and both are effective against necrotrophic pathogens. The possible mechanisms involved are discussed.

  8. The effects of citrus extract (Citrox©) on the naturally occurring microflora and inoculated pathogens, Bacillus cereus and Salmonella enterica, in a model food system and the traditional Greek yogurt-based salad Tzatziki.

    PubMed

    Tsiraki, Maria I; Savvaidis, Ioannis N

    2016-02-01

    The antimicrobial effect of citrus extract (at 1 mL/kg [TC1] and 2 mL/kg [TC2]) on the naturally occurring microflora and inoculated pathogens (Bacillus cereus and Salmonella enterica, at ca. 6 log cfu/g) in the traditional Greek yogurt-based salad Tzatziki during storage under vacuum at 4 or 10 °C was examined. We also examined the effect of citrus extract (Citrox(©)) against the two aforementioned pathogens in tryptic soy broth (TSB). Of the two treatments, TC2 yielded the lowest yeast counts, irrespective of temperature, resulting in approximately 2 (4 °C) and 3 (10 °C) log reductions on the final day of storage (70 and 30 days, respectively). Although panelists preferred the TC1-treated salad, the TC2-treated product was sensorily acceptable. Therefore, at the concentrations used, Citrox had no negative sensorial effect on the Tzatziki. During storage, the Bacillus populations in the Citrox-treated Tzatziki samples progressively decreased, showing major declines from days 12 and 28 (at 10 and 4 °C, respectively). Citrox, especially at 2 mL/kg, had a significant effect on the survival of B. cereus. S. enterica showed major declines in all untreated Tzatziki samples from day 0-70 (4 °C) and from day 0-30 (10 °C), with averages of 2.5 and 2.8 log cfu/g, respectively. The results indicate that Citrox (at 1 and 2 mL/kg) is effective, from a safety standpoint, for reducing Bacillus and Salmonella spp. in Tzatziki. In addition, 2% citrus extract also showed a higher inhibitory effect against B. cereus and S. enterica grown in TSB than 1% citrus extract. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Alternaria induces STAT-6 dependent acute airway eosinophila and epithelial FIZZ1 expression that promotes airway fibrosis and epithelial thickness

    PubMed Central

    Doherty, Taylor A.; Khorram, Naseem; Sugimoto, Kotaro; Sheppard, Dean; Rosenthal, Peter; Youn Cho, Jae; Pham, Alexa; Miller, Marina; Croft, Michael; Broide, David H.

    2012-01-01

    The fungal allergen, Alternaria, is specifically associated with severe asthma, including life-threatening exacerbations. To better understand the acute innate airway response to Alternaria, naïve WT mice were challenged once intranasally with Alternaria. Naïve WT mice developed significant BAL eosinophila following Alternaria challenge when analyzed 24 hours later. In contrast to Alternaria, neither Aspergillus nor Candida induced BAL eosinophilia. Gene microarray analysis of airway epithelial cell brushings demonstrated that Alternaria-challenged naïve WT mice had an over 20 fold increase level of expression of “Found in Inflammatory Zone 1” (FIZZ1/Retnla), a resistin-like molecule. Lung immunostaining confirmed strong airway epithelial FIZZ1 expression present as early as 3 hours after a single Alternaria challenge that persisted for at least 5 days and was significantly reduced in STAT6-deficient, but not PAR-2-deficient mice. Bone marrow chimera studies revealed that STAT6 expressed in lung cells was required for epithelial FIZZ1 expression, while in contrast, STAT6 present in bone marrow derived cells contributed to airway eosinophilia. Studies investigating which cells in the non-challenged lung bind FIZZ1 demonstrated that CD45+CD11c+ (macrophages and dendritic cells) as well as collagen-1 producing CD45 negative cells (fibroblasts) can bind to FIZZ1. Importantly, direct administration of recombinant FIZZ1 to naïve WT mice led to airway eosinophilia, peribronchial fibrosis, and increased thickness of the airway epithelium. Thus, Alternaria induces STAT-6 dependent acute airway eosinophila and epithelial FIZZ1 expression that promotes airway fibrosis and epithelial thickness. This may provide some insight into the uniquely pathogenic aspects of Alternaria-associated asthma. PMID:22327070

  10. Alt a 1 from Alternaria interacts with PR5 thaumatin-like proteins.

    PubMed

    Gómez-Casado, Cristina; Murua-García, Amaya; Garrido-Arandia, María; González-Melendi, Pablo; Sánchez-Monge, Rosa; Barber, Domingo; Pacios, Luis F; Díaz-Perales, Araceli

    2014-05-02

    Alt a 1 is a protein found in Alternaria alternata spores related to virulence and pathogenicity and considered to be responsible for chronic asthma in children. We found that spores of Alternaria inoculated on the outer surface of kiwifruits did not develop hyphae. Nevertheless, the expression of Alt a 1 gene was upregulated, and the protein was detected in the pulp where it co-localized with kiwi PR5. Pull-down assays demonstrated experimentally that the two proteins interact in such a way that Alt a 1 inhibits the enzymatic activity of PR5. These results are relevant not only for plant defense, but also for human health as patients with chronic asthma could suffer from an allergic reaction when they eat fruit contaminated with Alternaria. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  11. Phagostimulants for the Asian citrus psyllid also elicit volatile release from citrus leaves

    USDA-ARS?s Scientific Manuscript database

    Chemical cues that elicit orientation by the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), are of great interest because it is the primary vector of the causal pathogen of citrus greening disease. We identified an optimal blend ratio of formic and acetic acids that stimulate...

  12. Citrus huanglongbing shapes the structure of bacterial community associated with citrus roots

    USDA-ARS?s Scientific Manuscript database

    To examine the effect of pathogen on the diversity and structure of plant associated bacterial community, we carried out a molecular based analysis using citrus and huanglongbing as host-disease model. 16S rDNA clone library analysis of the citrus roots revealed shifts in the microbial diversity in ...

  13. Apparent tolerance to huanglongbing in citrus and citrus-related germplasm

    USDA-ARS?s Scientific Manuscript database

    In a Ft. Pierce, Florida field planting, growth and huanglongbing (HLB) severity were assessed as indicators of HLB-tolerance on progenies of 83 seed-source accessions of citrus and citrus-relatives from the Riverside, California genebank. The HLB-pathogen (Candidatus Liberibacter asiaticus) and vec...

  14. Swainsonine biosynthesis genes in diverse symbiotic and pathogenic fungi

    USDA-ARS?s Scientific Manuscript database

    Swainsonine, a cytotoxic fungal alkaloid and a potential cancer therapy drug, is produced by the insect pathogen and plant symbiont, Metarhizium robertsii, the clover pathogen Slafractonia leguminicola, locoweed symbionts belonging to Alternaria sect. Undifilum, and a recently discovered morning glo...

  15. Citrus Inventory

    NASA Technical Reports Server (NTRS)

    1994-01-01

    An aerial color infrared (CIR) mapping system developed by Kennedy Space Center enables Florida's Charlotte County to accurately appraise its citrus groves while reducing appraisal costs. The technology was further advanced by development of a dual video system making it possible to simultaneously view images of the same area and detect changes. An image analysis system automatically surveys and photo interprets grove images as well as automatically counts trees and reports totals. The system, which saves both time and money, has potential beyond citrus grove valuation.

  16. Citrus Inventory

    NASA Technical Reports Server (NTRS)

    1986-01-01

    Florida's Charlotte County Property Appraiser is using an aerial color infrared mapping system for inventorying citrus trees for valuation purposes. The ACIR system has significantly reduced the time and manpower required for appraisal. Aerial photographs are taken and interpreted by a video system which makes it possible to detect changes from previous years. Potential problems can be identified. KSC's TU Office has awarded a contract to the Citrus Research and Education Center to adapt a prototype system which would automatically count trees and report totals.

  17. Alternaria Keratitis after Deep Anterior Lamellar Keratoplasty

    PubMed Central

    Naik, Mekhla; Mohd. Shahbaaz; Sheth, Jay; Sunderamoorthy, S. K.

    2014-01-01

    To describe a case of Alternaria keratitis in a 30-year-old male patient who presented with bilateral vascularised central corneal opacity and underwent deep anterior lamellar keratoplasty (DALK) in the left eye. Patient was treated for recurrent epithelial defect with a bandage contact lens in the follow-up visits after DALK. Subsequently, patient presented with pigmented fungal keratitis, which on culture examination of the corneal scrapping demonstrated Alternaria species. Patient had to undergo a repeat DALK as the keratitis did not resolve with medical therapy alone. Patient did not have a recurrence for 11 months following the regraft. This case report highlights the importance of considering the Alternaria species as a possibile cause of non-resolving fungal keratitis after DALK. PMID:24669155

  18. Alternaria keratitis after deep anterior lamellar keratoplasty.

    PubMed

    Naik, Mekhla; Mohd Shahbaaz; Sheth, Jay; Sunderamoorthy, S K

    2014-01-01

    To describe a case of Alternaria keratitis in a 30-year-old male patient who presented with bilateral vascularised central corneal opacity and underwent deep anterior lamellar keratoplasty (DALK) in the left eye. Patient was treated for recurrent epithelial defect with a bandage contact lens in the follow-up visits after DALK. Subsequently, patient presented with pigmented fungal keratitis, which on culture examination of the corneal scrapping demonstrated Alternaria species. Patient had to undergo a repeat DALK as the keratitis did not resolve with medical therapy alone. Patient did not have a recurrence for 11 months following the regraft. This case report highlights the importance of considering the Alternaria species as a possibile cause of non-resolving fungal keratitis after DALK.

  19. Host-Specific Effects of Toxin from the Rough Lemon Pathotype of Alternaria alternata on Mitochondria 1

    PubMed Central

    Akimitsu, Kazuya; Kohmoto, Keisuke; Otani, Hiroshi; Nishimura, Syoyo

    1989-01-01

    Host-specific toxin from the rough lemon (Citrus jambhiri Lush) pathotype of Alternaria alternata (ACR toxin) was tested for effects on mitochondria isolated from several citrus species. The toxin caused uncoupling of oxidative phosphorylation and changes in membrane potential in mitochondria from leaves of the susceptible host (rough lemon); the effects differed from those of carbonylcyanide-m-chlorophenylhydrazone, a typical protonophore. ACR toxin also inhibited malate oxidation, apparently because of lack of NAD+ in the matrix. In contrast, the toxin had no effect on mitochondria from citrus species (Dancy tangerine and Emperor mandarin [Citrus reticulata Blanco], and grapefruit [Citrus paradisi Macf.]) that are not hosts of the fungus. The effects of the toxin on mitochondria from rough lemon are similar to the effects of a host-specific toxin from Helminthosporium maydis (HMT) on mitochondria from T-cytoplasm maize. Both ACR and HMT toxins are highly selective for the respective host plants. HMT toxin and methomyl had no effect (toxic or protective) on the activity of ACR toxin against mitochondria from rough lemon. PMID:16666643

  20. All five host-range variants of Xanthomonas citri carry one pthA homolog with 17.5 repeats that determines pathogenicity on citrus, but none determine host-range variation.

    PubMed

    Al-Saadi, Abdulwahid; Reddy, Joseph D; Duan, Yong P; Brunings, Asha M; Yuan, Qiaoping; Gabriel, Dean W

    2007-08-01

    Citrus canker disease is caused by five groups of Xanthomonas citri strains that are distinguished primarily by host range: three from Asia (A, A*, and A(w)) and two that form a phylogenetically distinct clade and originated in South America (B and C). Every X. citri strain carries multiple DNA fragments that hybridize with pthA, which is essential for the pathogenicity of wide-host-range X. citri group A strain 3213. DNA fragments that hybridized with pthA were cloned from a representative strain from all five groups. Each strain carried one and only one pthA homolog that functionally complemented a knockout mutation of pthA in 3213. Every complementing homolog was of identical size to pthA and carried 17.5 nearly identical, direct tandem repeats, including three new genes from narrow-host-range groups C (pthC), A(w) (pthAW), and A* (pthA*). Every noncomplementing paralog was of a different size; one of these was sequenced from group A* (pthA*-2) and was found to have an intact promoter and full-length reading frame but with 15.5 repeats. None of the complementing homologs nor any of the noncomplementing paralogs conferred avirulence to 3213 on grapefruit or suppressed avirulence of a group A* strain on grapefruit. A knockout mutation of pthC in a group C strain resulted in loss of pathogenicity on lime, but the strain was unaffected in ability to elicit an HR on grapefruit. This pthC- mutant was fully complemented by pthA, pthB, or pthC. Analysis of the predicted amino-acid sequences of all functional pthA homologs and nonfunctional paralogs indicated that the specific sequence of the 17th repeat may be essential for pathogenicity of X. citri on citrus.

  1. Comparative influence of Radopholus similis and Pratylenchus coffeae on citrus.

    PubMed

    O'Bannon, J H; Radewald, J D; Tomerlin, A T; Inserra, R N

    1976-01-01

    Pratylenchus coffeae was as pathogenic as Radopholus sirnilis to commercial citrus rootstocks. No rootstock resistant to R. similis was resistant to P. coffeae. Both nematodes stunted citrus in three soil types. Seedling damage by P. coffeae and R. similis was greatest in fine- and coarse-textured soils, respectively. Reproduction and survival on citrus were greater for P. coffeae than for R. similis. Mixed inoculations with R. similis and P. coffeae resulted in lower populations of each species than did separate inoculations.

  2. Nonhost-specific phytotoxicity of the polyketide-derived toxin solanapyrone A produced by Ascochyta rabiei and Alternaria solani

    USDA-ARS?s Scientific Manuscript database

    Solanapyrone A is a polyketide-derived metabolite produced by Ascochyta rabiei and Alternaria solani, which are the most destructive necrotrophic pathogens of chickpea and potato/tomato, respectively. They belong to the Order Pleosporales within the Class Dothideomycetes, but are phylogenetically di...

  3. Experiments with Biocontrol of Alternaria alternata

    USDA-ARS?s Scientific Manuscript database

    A biocontrol yeast (WRL-76) developed by USDA-ARS at the Western Regional Research lab was evaluated for control of Alternaria alternata on pistachio. The experiment was conducted over a four year period in a Madera Co., CA orchard. Counts of damaged vs. intact fruit clusters were taken on 100 contr...

  4. Dramatic Change in Citrus tristeza virus populations in the Dominican Republic

    USDA-ARS?s Scientific Manuscript database

    Citrus tristeza virus (CTV) is the most destructive viral pathogen of citrus and has been an important concern for the citrus industry in the Dominican Republic. Earlier studies documented widespread distribution of mild isolates of the T30 genotype, which caused no disease in the infected trees, an...

  5. Wind speed and wind-associated leaf injury affect severity of citrus canker on Swingle citrumelo

    USDA-ARS?s Scientific Manuscript database

    Citrus canker (caused by the bacterial pathogen Xanthomonas citri subsp. citri, Xcc) can cause severe damage to citrus. It is endemic in Florida, and occurs in other citrus growing regions. The bacterium is dispersed predominantly in rain splash. To simulate dispersal in splash, and to investigate t...

  6. Optimal strategies for the eradication of Asiatic citrus canker in heterogeneous host landscapes

    USDA-ARS?s Scientific Manuscript database

    The eradication of non-native plant pathogens is a key challenge in plant disease epidemiology. Asiatic citrus canker is an economically significant disease of citrus caused by the bacterial plant pathogen Xanthomonas citri subsp. citri. The pathogen is a major exotic disease problem in many citru...

  7. Citrus leprosis research update

    USDA-ARS?s Scientific Manuscript database

    Citrus leprosis is one of the oldest citrus diseases, but is also one of the most important emerging citrus diseases in South and Central America, and it is apparently spreading northward towards the U.S. Research in our labs and by others has shown that citrus leprosis disease is caused by a compl...

  8. Citrus blight research

    USDA-ARS?s Scientific Manuscript database

    With HLB now occurring throughout Florida citrus groves, citrus blight has been getting less attention even though the problem still exists. In fact, the combination of HLB and citrus blight has compounded the problem that the citrus industry is facing with decreased yields, small fruit size and tre...

  9. Survival of adult Tamarixia radiata subjected to different short-term storage methods prior to field releases for biological control of Asian citrus psyllid

    USDA-ARS?s Scientific Manuscript database

    Tamarixia radiata is an insect parasitoid of the Asian citrus psyllid. The psyllid is an important pest of citrus because it vectors pathogens responsible for a serious disease of citrus known as huanglongbing (also known as citrus greening or yellow shoot disease). T. radiata is regarded as one of ...

  10. Molecular diversity of Citrus tristeza virus in California

    USDA-ARS?s Scientific Manuscript database

    Citrus tristeza virus (CTV) is a serious citrus pathogen worldwide. Recent genetic studies have identified five standard CTV genotypic groups: T30, VT, T36, T3, and B165/T68. Field surveys performed in California in 2008-2010 identified primarily MCA13-negative CTV isolates with T30-like genotype. C...

  11. Behavioral plasticity in Diaphorina citri feeding on different citrus tissues

    USDA-ARS?s Scientific Manuscript database

    The Asian Citrus Psyllid (Diaphorina citri) is a vector of Candidatus Liberibacter asiaticus (CLas), a phloem-limited bacterial pathogen of citrus. While psyllid nymphs are obligate feeders on young flush leaves, adults can be found on leaves of any age. To understand risk of transmission of CLas, i...

  12. An attract-and-kill strategy for Asian citrus psyllid

    USDA-ARS?s Scientific Manuscript database

    Asian citrus psyllids (ACP) transmit the pathogen responsible for citrus greening disease. Psyllids use color, smell, taste and vibrational cues to identify their host plants and conspecifics. The main goal of this project is to develop an attract-and-kill device strategy that will exploit the psyll...

  13. A phagostimulant blend for the Asian citrus psyllid

    USDA-ARS?s Scientific Manuscript database

    Chemical cues that condition orientation by the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), are of great interest because it is the primary vector of the causal pathogen of citrus greening disease. Previous work in our lab identified a blend of formic and acetic acids as s...

  14. Associated bacteria of Asian citrus psyllid (Hemiptera: Psyllidae: Diaphorina citri)

    USDA-ARS?s Scientific Manuscript database

    Using a genetics approach two new bacterial species were identified from the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae). Psyllids are economically important in the U.S. as vectors of the pathogen associated with citrus greening disease (Huanglongbing). Huanglongbing is on...

  15. Mycotoxin production in liquid culture and on plants infected with Alternaria spp. isolated from rocket and cabbage.

    PubMed

    Siciliano, Ilenia; Ortu, Giuseppe; Gilardi, Giovanna; Gullino, Maria Lodovica; Garibaldi, Angelo

    2015-03-05

    Fungi belonging to the genus Alternaria are common pathogens of fruit and vegetables with some species able to produce secondary metabolites dangerous to human health. Twenty-eight Alternaria isolates from rocket and cabbage were investigated for their mycotoxin production. Five different Alternaria toxins were extracted from synthetic liquid media and from plant material (cabbage, cultivated rocket, cauliflower). A modified Czapek-Dox medium was used for the in vitro assay. Under these conditions, more than 80% of the isolates showed the ability to produce at least one mycotoxin, generally with higher levels for tenuazonic acid. However, the same isolates analyzed in vivo seemed to lose their ability to produce tenuazonic acid. For the other mycotoxins; alternariol, alternariol monomethyl ether, altenuene and tentoxin a good correlation between in vitro and in vivo production was observed. In vitro assay is a useful tool to predict the possible mycotoxin contamination under field and greenhouse conditions.

  16. Alternaria alternata invasive fungal infection in a patient with Fanconi's anemia after an unrelated bone marrow transplant.

    PubMed

    Ferreira, Isabelina de Sousa; Teixeira, Gilda; Abecasis, Manuel

    2013-02-01

    Alternaria spp. have emerged as opportunistic pathogens particularly in immunosuppressed patients, such as bone marrow transplant recipients. The authors present a case of Alternaria alternata in a patient with Fanconi's anemia, who received antifungal prophylaxis with posaconazole after an unrelated bone marrow transplantation, followed by empirical antifungal treatment with caspofungin when persistent fever emerged until cutaneous lesions eventually appeared. At that time there were clinical reasons to assume that the patient had an infection with an emerging fungus. This consideration triggered a change of the antifungal therapy from caspofungin to liposomal amphotericin B. After collecting sufficient evidence for the presence of an invasive fungal infection by A. alternata and given the severity of neutropenia and other immunosuppression, oral posaconazole was added to liposomal amphotericin B. The course of disease in this case suggests a possibly synergistic interaction between liposomal amphotericin B and posaconazole when administered simultaneously to treat an invasive systemic infection by Alternaria spp. in immunocompromised patients.

  17. Mycotoxin Production in Liquid Culture and on Plants Infected with Alternaria spp. Isolated from Rocket and Cabbage

    PubMed Central

    Siciliano, Ilenia; Ortu, Giuseppe; Gilardi, Giovanna; Gullino, Maria Lodovica; Garibaldi, Angelo

    2015-01-01

    Fungi belonging to the genus Alternaria are common pathogens of fruit and vegetables with some species able to produce secondary metabolites dangerous to human health. Twenty-eight Alternaria isolates from rocket and cabbage were investigated for their mycotoxin production. Five different Alternaria toxins were extracted from synthetic liquid media and from plant material (cabbage, cultivated rocket, cauliflower). A modified Czapek-Dox medium was used for the in vitro assay. Under these conditions, more than 80% of the isolates showed the ability to produce at least one mycotoxin, generally with higher levels for tenuazonic acid. However, the same isolates analyzed in vivo seemed to lose their ability to produce tenuazonic acid. For the other mycotoxins; alternariol, alternariol monomethyl ether, altenuene and tentoxin a good correlation between in vitro and in vivo production was observed. In vitro assay is a useful tool to predict the possible mycotoxin contamination under field and greenhouse conditions. PMID:25751147

  18. Characterization of small-spored Alternaria from Argentinean crops through a polyphasic approach.

    PubMed

    da Cruz Cabral, Lucía; Rodriguero, Marcela; Stenglein, Sebastián; Fog Nielsen, Kristian; Patriarca, Andrea

    2017-09-18

    Small-spored Alternaria have been isolated from a wide variety of food crops, causing both economic losses and human health risk due to the metabolites produced. Their taxonomy has been discussed widely, but no scientific consensus has been established in this field to date. Argentina is a major exporter of agricultural products, so it is essential to thoroughly understand the physiological behaviour of this pathogen in a food safety context. Thus, the objective of this work was to characterize small-spored Alternaria spp. obtained from tomato fruits, pepper fruits, wheat grains and blueberries from Argentina by a polyphasic approach involving metabolomic and phylogenetic analyses based on molecular and morphological characters. Morphological analysis divided the population studied into three groups; A. arborescens sp.-grp., A. tenuissima sp.-grp., and A. alternata sp.-grp. However, when these characters were simultaneously analysed with molecular data, no clearly separated groups were obtained. Haplotype network and phylogenetic analysis (both Bayesian and maximum parsimony) of a conserved region yielded the same result, suggesting that all isolates belong to the same species. Furthermore, no correlation could be established between morphological species-groups and a metabolite or group of metabolites synthesized. Thus, the whole set of analyses carried out in the present work supports the hypothesis that these small-spored Alternaria isolates from food belong to the same species. Identification at species level through classical morphology or modern molecular techniques does not seem to be a useful tool to predict toxicological risk in food matrices. The detection of any small-spored Alternaria from Section Alternaria (D.P. Lawr., Gannibal, Peever & B.M. Pryor 2013) in food implies a potential toxicological risk. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Metabolic interplay between the Asian citrus psyllid and its Profftella symbiont: An Achilles’ heel of the citrus greening insect vector

    USDA-ARS?s Scientific Manuscript database

    ‘Candidatus Liberibacter asiaticus’ (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid. Interactions among D. citri and its microbial endosymbionts, including ‘Candidatus Profftella armatura’, are likely to impact tra...

  20. Penicillium digitatum metabolites on synthetic media and citrus fruits.

    PubMed

    Ariza, Marta R; Larsen, Thomas O; Peterson, Bent O; Duus, Jens O; Barrero, Alejandro F

    2002-10-23

    Penicillium digitatum has been cultured on citrus fruits and yeast extract sucrose agar media (YES). Cultivation of fungal cultures on solid medium allowed the isolation of two novel tryptoquivaline-like metabolites, tryptoquialanine A (1) and tryptoquialanine B (2), also biosynthesized on citrus fruits. Their structural elucidation is described on the basis of their spectroscopic data, including those from 2D NMR experiments. The analysis of the biomass sterols led to the identification of 8-12. Fungal infection on the natural substrates induced the release of citrus monoterpenes together with fungal volatiles. The host-pathogen interaction in nature and the possible biological role of citrus volatiles are also discussed.

  1. The pathogenicity determinant of Citrus tristeza virus causing the seedling yellows syndrome maps at the 3'-terminal region of the viral genome.

    PubMed

    Albiach-Marti, Maria R; Robertson, Cecile; Gowda, Siddarame; Tatineni, Satyanarayana; Belliure, Belén; Garnsey, Stephen M; Folimonova, Svetlana Y; Moreno, Pedro; Dawson, William O

    2010-01-01

    Citrus tristeza virus (CTV) (genus Closterovirus, family Closteroviridae) causes some of the more important viral diseases of citrus worldwide. The ability to map disease-inducing determinants of CTV is needed to develop better diagnostic and disease control procedures. A distinctive phenotype of some isolates of CTV is the ability to induce seedling yellows (SY) in sour orange, lemon and grapefruit seedlings. In Florida, the decline isolate of CTV, T36, induces SY, whereas a widely distributed mild isolate, T30, does not. To delimit the viral sequences associated with the SY syndrome, we created a number of T36/T30 hybrids by substituting T30 sequences into different regions of the 3' half of the genome of an infectious cDNA of T36. Eleven T36/T30 hybrids replicated in Nicotiana benthamiana protoplasts. Five of these hybrids formed viable virions that were mechanically transmitted to Citrus macrophylla, a permissive host for CTV. All induced systemic infections, similar to that of the parental T36 clone. Tissues from these C. macrophylla source plants were then used to graft inoculate sour orange and grapefruit seedlings. Inoculation with three of the T30/T36 hybrid constructs induced SY symptoms identical to those of T36; however, two hybrids with T30 substitutions in the p23-3' nontranslated region (NTR) (nucleotides 18 394-19 296) failed to induce SY. Sour orange seedlings infected with a recombinant non-SY p23-3' NTR hybrid also remained symptomless when challenged with the parental virus (T36), demonstrating the potential feasibility of using engineered constructs of CTV to mitigate disease.

  2. Establishment of CRISPR/Cas9 in Alternaria alternata.

    PubMed

    Wenderoth, Maximilian; Pinecker, Christoph; Voß, Benjamin; Fischer, Reinhard

    2017-03-10

    The filamentous fungus Alternaria alternata is a potent producer of many secondary metabolites, some of which like alternariol or alternariol-methyl ether are toxic and/or cancerogenic. Many Alternaria species do not only cause post-harvest losses of food and feed, but are aggressive plant pathogens. Despite the great economic importance and the large number of research groups working with the fungus, the molecular toolbox is rather underdeveloped. Gene deletions often result in heterokaryotic strains and therefore, gene-function analyses are rather tedious. In addition, A. alternata lacks a sexual cycle and classical genetic approaches cannot be combined with molecular biological methods. Here, we show that CRISPR/Cas9 can be efficiently used for gene inactivation. Two genes of the melanin biosynthesis pathway, pksA and brm2, were chosen as targets. Several white mutants were obtained after several rounds of strain purification through protoplast regeneration or spore inoculation. Mutation of the genes was due to deletions from 1bp to 1.5kbp. The CRISPR/Cas9 system was also used to inactivate the orotidine-5-phosphate decarboxylase gene pyrG to create a uracil-auxotrophic strain. The strain was counter-selected with fluor-orotic acid and could be re-transformed with pyrG from Aspergillus fumigatus and pyr-4 from Neurospora crassa. In order to test the functioning of GFP, the fluorescent protein was fused to a nuclear localization signal derived from the StuA transcription factor of Aspergillus nidulans. After transformation bright nuclei were visible.

  3. Innate and Conditioned Responses to Chemosensory and Visual Cues in Asian Citrus Psyllid, Diaphorina citri (Hemiptera: Liviidae), Vector of Huanglongbing Pathogens.

    PubMed

    Patt, Joseph M; Stockton, Dara; Meikle, William G; Sétamou, Mamoudou; Mafra-Neto, Agenor; Adamczyk, John J

    2014-11-19

    Asian citrus psyllid (Diaphorina citri) transmits Huanglongbing, a devastating disease that threatens citrus trees worldwide. A better understanding of the psyllid's host-plant selection process may lead to the development of more efficient means of monitoring it and predicting its movements. Since behavioral adaptations, such as associative learning, may facilitate recognition of suitable host-plants, we examined whether adult D. citri could be conditioned to visual and chemosensory stimuli from host and non-host-plant sources. Response was measured as the frequency of salivary sheaths, the residue of psyllid probing activity, in a line of emulsified wax on the surface of a test arena. The psyllids displayed both appetitive and aversive conditioning to two different chemosensory stimuli. They could also be conditioned to recognize a blue-colored probing substrate and their response to neutral visual cues was enhanced by chemosensory stimuli. Conditioned psyllids were sensitive to the proportion of chemosensory components present in binary mixtures. Naïve psyllids displayed strong to moderate innate biases to several of the test compounds. While innate responses are probably the psyllid's primary behavioral mechanism for selecting host-plants, conditioning may enhance its ability to select host-plants during seasonal transitions and dispersal.

  4. Innate and Conditioned Responses to Chemosensory and Visual Cues in Asian Citrus Psyllid, Diaphorina citri (Hemiptera: Liviidae), Vector of Huanglongbing Pathogens

    PubMed Central

    Patt, Joseph M.; Stockton, Dara; Meikle, William G.; Sétamou, Mamoudou; Mafra-Neto, Agenor; Adamczyk, John J.

    2014-01-01

    Asian citrus psyllid (Diaphorina citri) transmits Huanglongbing, a devastating disease that threatens citrus trees worldwide. A better understanding of the psyllid’s host-plant selection process may lead to the development of more efficient means of monitoring it and predicting its movements. Since behavioral adaptations, such as associative learning, may facilitate recognition of suitable host-plants, we examined whether adult D. citri could be conditioned to visual and chemosensory stimuli from host and non-host-plant sources. Response was measured as the frequency of salivary sheaths, the residue of psyllid probing activity, in a line of emulsified wax on the surface of a test arena. The psyllids displayed both appetitive and aversive conditioning to two different chemosensory stimuli. They could also be conditioned to recognize a blue-colored probing substrate and their response to neutral visual cues was enhanced by chemosensory stimuli. Conditioned psyllids were sensitive to the proportion of chemosensory components present in binary mixtures. Naïve psyllids displayed strong to moderate innate biases to several of the test compounds. While innate responses are probably the psyllid’s primary behavioral mechanism for selecting host-plants, conditioning may enhance its ability to select host-plants during seasonal transitions and dispersal. PMID:26462949

  5. Characterization of a hydroperoxide lyase gene and effect of C6-volatiles on expression of genes of the oxylipin metabolism in Citrus.

    PubMed

    Gomi, Kenji; Yamasaki, Yumiko; Yamamoto, Hiroyuki; Akimitsu, Kazuya

    2003-10-01

    A number of C6-volatile products of the lipoxygenase (LOX) pathway was examined for their antifungal activity and a potential role as a signal molecule in citrus. trans-2-Hexenal induced the rough lemon lipoxygenase gene (RlemLOX), hydroperoxide lyase gene (RlemHPL) and AOS gene, but hexanal, and hexanol suppressed them. cis-3-Hexenol and trans-2-hexenol increased expression of the AOS gene but not RlemLOX and RlemHPL. Transcripts of the RlemHPL and AOS gene were detected constitutively in leaves by northern blot, but wounding or inoculation with nonpathogenic Alternaria alternata rapidly increased the transcript accumulation. Transcripts of the RlemHPL and AOS genes were also induced with pathogenic A. alternata, which produces the host-selective ACR-toxin, but the signal declined rapidly after inoculation. An increase in enzymatic activity of HPL after wounding or inoculation with nonpathogen was suppressed in leaves infected with the pathogen. Interestingly, vapor treatment with trans-2-hexenol delayed necrotic spot formation in the leaves inoculated with the pathogenic A. alternata. Since trans-2-hexenol has no antifungal activity to A. alternata and also did not inhibit necrosis formation by ACR-toxin alone, the delay of symptoms may be caused by activation of AOS in the LOX pathway to produce oxylipin derivatives such as methyl jasmonate for activation of defense related genes with antifungal activity.

  6. Volatile Organic Compound (VOC) profiling of Citrus tristeza virus (CTV) infection in sweet orange citrus varietals using thermal desorption gas chromatography time of flight mass spectrometry (TD-GC/TOF-MS)

    USDA-ARS?s Scientific Manuscript database

    Citrus tristeza virus (CTV) is a plant pathogen which predominately infects economically important citrus crops such as sweet orange, clementine, lime and grapefruit varietals. Within the last 70 years, an estimated 100 million citrus trees on sour orange rootstock have been destroyed due to CTV inf...

  7. Visual rating and the use of image analysis for assessing different symptoms of citrus canker on grapefruit leaves

    USDA-ARS?s Scientific Manuscript database

    Citrus canker is caused by the bacterial pathogen Xanthomonas axonopodis pv citri (Xac) and infects several citrus species in wet tropical and subtropical citrus growing regions. Accurate, precise and reproducible disease assessment is needed for monitoring epidemics and disease response in breeding...

  8. Fungitoxic activity of 12 essential oils against four postharvest citrus pathogens: chemical analysis of thymus capitatus oil and its effect in subatmospheric pressure conditions.

    PubMed

    Arras, G; Usai, M

    2001-07-01

    The fungitoxic activity against Penicillium digitatum, Penicillium italicum, Botrytis cinerea, and Alternaria citri of 12 essential oils (EOs) distilled from medicinal plants is reported. The results of the in vitro trials show strong fungitoxic activity of Thymus capitatus (L.) Hofmgg EOs, which inhibited the growth of the four fungi at a concentration of 250 ppm (vol/vol). The other 11 essences reduced the development of the fungi from 95 to 9% at 250 ppm (vol/vol). The fungitoxic activity of T. capitatus EOs (75, 150, and 250 ppm) on healthy orange fruits, inoculated with P. digitatum (10(8) conidia ml(-1)) by spraying and placed in 10-liter desiccators, was weak at atmospheric pressure (3 to 10% inhibition at all three concentrations), while in vacuum conditions (0.5 bar), conidial mortality on the exocarp was high (90 to 97% at all three concentrations). These data proved not to be statistically different from treatments with thiabendazole-TBZ (2,000 ppm). Scanning electron microscope observations showed that T. capitatus EO vapors altered the morphology of P. digitatum hyphae and conidia. The gas-chromatographic analyses of thyme EO indicated that carvacrol was present at 81 to 83%, p-cymene at 4.5 to 5%, gamma-terpinene at 2.6 to 3.3%, caryophyllene at 1.5 to 1.6%, beta-myrcene at 1.6%, and linalool at 1.1 to 1.2%. Carvacrol proved to be the most important fungitoxic compound among the thyme EO constituents, but, unlike thyme EO, it caused alterations to the fruit at the concentration of 75 ppm.

  9. Presence of adhesive vesicles in the mycoherbicide Alternaria helianthi

    USDA-ARS?s Scientific Manuscript database

    Alternaria helianthi conidia have been shown to cause disease on common cocklebur. Conidia were applied to slides made hydrophobic by coating with dimethyldicholorosilane (mimics leaf surface), then rinsed and treated with FITC-Con A to stain the adhesive material. Alternaria helianthi coni...

  10. Asian citrus psyllid genome (Diaphorina citri, Hemiptera)

    USDA-ARS?s Scientific Manuscript database

    The Psyllid genome is a scientific breakthrough in that it opens the psyllid genetic blueprint to investigations of all questions ranging from taxonomic origins to the understanding of developmental biology, to the acquisition and transmission of pathogens. The Asian citrus psyllid, Diaphorina citri...

  11. Characterization of the Asian citrus psyllid transcriptome

    USDA-ARS?s Scientific Manuscript database

    The Asian citrus psyllid (Diaphorina citri Kuwayama) and other psyllids are important agricultural pests that cause extensive economic damage by feeding and as vectors of plant pathogens. No psyllid genomes have been characterized, and little is known about the composition of psyllid genomes or the ...

  12. Genetic Transformation in Citrus

    PubMed Central

    Donmez, Dicle; Simsek, Ozhan; Izgu, Tolga; Aka Kacar, Yildiz; Yalcin Mendi, Yesim

    2013-01-01

    Citrus is one of the world's important fruit crops. Recently, citrus molecular genetics and biotechnology work have been accelerated in the world. Genetic transformation, a biotechnological tool, allows the release of improved cultivars with desirable characteristics in a shorter period of time and therefore may be useful in citrus breeding programs. Citrus transformation has now been achieved in a number of laboratories by various methods. Agrobacterium tumefaciens is used mainly in citrus transformation studies. Particle bombardment, electroporation, A. rhizogenes, and a new method called RNA interference are used in citrus transformation studies in addition to A. tumefaciens. In this review, we illustrate how different gene transformation methods can be employed in different citrus species. PMID:23983635

  13. Genetic dissection defines the roles of elsinochrome Phytotoxin for fungal pathogenesis and conidiation of the citrus pathogen Elsinoë fawcettii.

    PubMed

    Liao, Hui-Ling; Chung, Kuang-Ren

    2008-04-01

    Elsinochrome pigments produced by many phytopathogenic Elsinoë spp. are nonhost-selective toxins which react with oxygen molecules after light activation to produce highly toxic reactive oxygen species. The structures and chemical properties of four derivatives are well known. However, the biological roles of elsinochromes in fungal pathogenesis are poorly understood. Many isolates of Elsinoë fawcettii causing citrus scab are able to produce elsinochromes under axenic conditions. In this article, we report the cloning, expression, and functional characterization of the polyketide synthase-encoding gene, EfPKS1, which we show is required for the production of elsinochromes and fungal pathogenesis. Targeted disruption of EfPKS1 in E.fawcettii completely abrogated elsinochrome production, drastically reduced conidiation, and significantly decreased lesion formation on rough lemon leaves. All mutant phenotypes were restored to the wild type in fungal strains expressing a functional copy of EfPKS1. Accumulation of the EfPKS1 transcript and elsinochromes by a wild-type strain appears to be coordinately regulated by light, nutrients, and pH. The results indicate that the product of EfPKS1 is involved in the biosynthesis of elsinochromes via a fungal polyketide pathway, and that elsinochromes play an important role in fungal pathogenesis.

  14. Delivery of gene biotechnologies to plants: Pathogen and pest control

    USDA-ARS?s Scientific Manuscript database

    Treatment of oligonucleotides to plants for host delivered suppression of microbes and insect pests of citrus was successful. FANA_ASO, (2'-deoxy-2'-fluoro-D- arabinonucleic acid)_( antisense oligonucleotides- AUM LifeTech) designed to: Asian citrus psyllid; Citrus plant bacterial pathogen of citru...

  15. First Record of Alternaria simsimi Causing Leaf Spot on Sesame (Sesamum indicum L.) in Korea.

    PubMed

    Choi, Young Phil; Paul, Narayan Chandra; Lee, Hyang Burm; Yu, Seung Hun

    2014-12-01

    Leaf spot disease was observed in sesame (Sesamum indicum L.) during 2009 and 2010 in Korea. The pathogen was identified as Alternaria simsimi based on morphological and cultural characteristics. The morphological identification was well supported by phylogenetic analysis of the ribosomal DNA-internal transcribed spacer region. A. simsimi isolates caused spot symptoms on leaves and stems of sesame plants 2 wk after artificial inoculation, which were similar to those observed in the field. This is the first record of leaf spot disease in Korea caused by A. simsimi.

  16. Proteomic analysis of Alternaria alternata (Fr.) Keissler responds to COS fumigation.

    PubMed

    Liu, Tao; Li, Li; Qu, Haixia; Zhan, Guoping; Liu, Bo; Wang, Yuejin

    2010-01-01

    Carbonyl sulfide (COS) is a new fumigant which has been a potential alternative to methyl bromide and phosphine in many applications. In this study, we investigated the fungitoxicity of COS towards the pathogen of pear black spot disease Alternaria alternata (Fr.) Keissler (A. alternata). Moreover, proteomic analysis and RT-PCR was performed and our results showed that during the fumigation, the regulation of 21 proteins in protein expression and mRNA accumulation levels is involved, which respond to growth inhibition caused by COS. These results provide new clues for the mechanism of the fungitoxicity of COS.

  17. Alternethanoxins A and B, polycyclic ethanones produced by Alternaria sonchi , potential mycoherbicides for Sonchus arvensis biocontrol.

    PubMed

    Evidente, Antonio; Punzo, Biancavaleria; Andolfi, Anna; Berestetskiy, Alexander; Motta, Andrea

    2009-08-12

    Alternaria sonchi is a fungal pathogen isolated from Sonchus arvensis and proposed as a biocontrol agent of this noxious perennial weed. Different phytotoxic metabolites were isolated from solid culture of the fungus. Two new polycyclic ethanones, named alternethanoxins A and B, were characterized using essentially spectroscopic and chemical methods. Tested by leaf disk-puncture assay on the fungal host plant and a number of nonhost plants, alternethanoxins A and B were shown to be phytotoxic, whereas they did not possess antimicrobial activity tested at 100 microg/disk. Hence, alternethanoxins A and B have potential as nonselective natural herbicides. Some structure-activity relationship observations were also made.

  18. Abscisic acid enhances resistance to Alternaria solani in tomato seedlings.

    PubMed

    Song, Weiwei; Ma, Xinrong; Tan, Hong; Zhou, Jinyan

    2011-07-01

    The plant hormone abscisic acid (ABA) is an important regulator in many aspects of plant growth and development, as well as stress resistance. Here, we investigated the effects of exogenous ABA application on the interaction between tomato (Solanum lycopersicon L.) and Alternaria solani (early blight). Foliar spraying of 7.58 μM ABA was effective in reducing disease severity in tomato plants. Previously, increased activities of phenylalanine ammonia-lyase (PAL), polyphenol oxidase (PPO) and peroxidase (POD) were observed in exogenous ABA-treated tomato leaves. Moreover, these enzyme activities were maintained at higher levels in ABA-pretreated and A. solani challenged tomato plants. Tomato defense genes, such as PR1, β-1, 3-glucanase (GLU), PPO, POD, and superoxide dismutase (SOD), were rapidly and significantly up-regulated by exogenous ABA treatment. Furthermore, a subsequent challenge of ABA-pretreated plants with the pathogen A. solani resulted in higher expression of defense genes, compared to water-treated or A. solani inoculated plants. Therefore, our results suggest that exogenous ABA could enhance disease resistance against A. solani infection in tomato through the activation of defense genes and via the enhancement of defense-related enzymatic activities.

  19. Evaluation of Alternaria mycotoxins in strawberries: quantification and storage condition.

    PubMed

    Juan, Cristina; Oueslati, Souheib; Mañes, Jordi

    2016-05-01

    Alternariol (AOH), alternariol methyl ether (AME) and tentoxin (TEN) are Alternaria mycotoxins produced by the most common post-harvest pathogens of fruits. The production of these metabolites depends on several environmental factors, mainly temperature, water activity, pH and the technological treatments that have been applied to the product. In this study, the occurrence of AOH, AME and TEN was evaluated in strawberries samples stored at different temperatures ranges (at 22 ± 2 or 6 ± 2°C) and different periods (up to 1 month) simulating the current practice of consumer's storage conditions. Sample extraction was performed using a liquid-liquid extraction method prior to LC-MS/MS analysis. AOH was the most prevalent mycotoxins with a 42% at strawberries stored at (22 ± 2)°C and 37% stored at (6 ± 2)°C. The highest AOH levels were found in samples conserved at (22 ± 2)°C ranging between 26 and 752 ng g(-1). AME levels ranged between 11 and 137 ng g(-)(1), which were found mainly in stored samples at (6 ± 2)°C for more than 28 days. None sample presented levels of TEN in either of the studied conditions.

  20. Excito-Repellency of Citrus hystrix DC Leaf and Peel Essential Oils Against Aedes aegypti and Anopheles minimus (Diptera: Culicidae), Vectors of Human Pathogens.

    PubMed

    Nararak, Jirod; Sathantriphop, Sunaiyana; Kongmee, Monthathip; Bangs, Michael J; Chareonviriyaphap, Theeraphap

    2016-09-30

    The essential oils of kaffir lime (Citrus hystrix DC.) at four different concentrations (0.5, 1.0, 2.5, and 5.0% v/v) were studied for their repellency, excitation, and knockdown properties against laboratory strains of Aedes aegypti (L.) and Anopheles minimus Theobald using an excito-repellency test system. Both contact and noncontact escape responses to leaf- and peel-derived kaffir lime oils were observed. Comparing unadjusted escape responses for An. minimus, leaf oil had strong combined irritant and repellent activity responses at 1-5% concentrations (90.0-96.4% escape) and the strongest spatial repellent activity at 1% and 2% (85.9% and 87.2% escape, respectively). The peel oil exhibited good excitation with repellency at concentrations of 2.5% (89.8% escape) and 5% (96.28% escape), while concentrations 1-5% showed more moderate repellent activity against An. minimus. For Ae. aegypti, 2.5% leaf oil produced the greatest response for both contact (56.1% escape) and noncontact (63.3% escape) trials, while 2.5% produced the strongest response among all concentrations of peel oil, with 46.5% escape. However, after adjusting the contact trial escape (a measure of combined excitation and repellency), the estimated escape due to contact alone was a much weaker response than spatial repellency for both species. Knockdown responses above 50% were only observed in Ae. aegypti exposed to 5% leaf oil. Kaffir lime oils were more active against An. minimus than Ae. aegypti mosquitoes. There were statistically significant differences between leaf (more active) and peel oils at each concentration against An. minimus in contact and noncontact trials, except at the highest (5%) concentration.

  1. Excito-Repellency of Citrus hystrix DC Leaf and Peel Essential Oils Against Aedes aegypti and Anopheles minimus (Diptera: Culicidae), Vectors of Human Pathogens.

    PubMed

    Nararak, Jirod; Sathantriphop, Sunaiyana; Kongmee, Monthathip; Bangs, Michael J; Chareonviriyaphap, Theeraphap

    2017-01-01

    The essential oils of kaffir lime (Citrus hystrix DC.) at four different concentrations (0.5, 1.0, 2.5, and 5.0% v/v) were studied for their repellency, excitation, and knockdown properties against laboratory strains of Aedes aegypti (L.) and Anopheles minimus Theobald using an excito-repellency test system. Both contact and noncontact escape responses to leaf- and peel-derived kaffir lime oils were observed. Comparing unadjusted escape responses for An. minimus, leaf oil had strong combined irritant and repellent activity responses at 1-5% concentrations (90.0-96.4% escape) and the strongest spatial repellent activity at 1% and 2% (85.9% and 87.2% escape, respectively). The peel oil exhibited good excitation with repellency at concentrations of 2.5% (89.8% escape) and 5% (96.28% escape), while concentrations 1-5% showed more moderate repellent activity against An. minimus. For Ae. aegypti, 2.5% leaf oil produced the greatest response for both contact (56.1% escape) and noncontact (63.3% escape) trials, while 2.5% produced the strongest response among all concentrations of peel oil, with 46.5% escape. However, after adjusting the contact trial escape (a measure of combined excitation and repellency), the estimated escape due to contact alone was a much weaker response than spatial repellency for both species. Knockdown responses above 50% were only observed in Ae. aegypti exposed to 5% leaf oil. Kaffir lime oils were more active against An. minimus than Ae. aegypti mosquitoes. There were statistically significant differences between leaf (more active) and peel oils at each concentration against An. minimus in contact and noncontact trials, except at the highest (5%) concentration. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  2. Effect of citrus pulp on the viability of the probiotic Saccharomyces cerevisiae bouldarii (Levucell) and subsequent effects in presence of pathogens

    USDA-ARS?s Scientific Manuscript database

    The probiotic Saccharomyces cerevisiae subtype boulardii (i.e., Levucell) is commonly provided to weaned and nursing sows to promote intestinal health through limiting the colonization of pathogens. Recent research from our group identified that supplementation of this probiotic to feed containing t...

  3. Citrus huanglongbing: a newly relevant disease presents unprecedented challenges.

    PubMed

    Wang, Nian; Trivedi, Pankaj

    2013-07-01

    Citrus huanglongbing (HLB) is one of the oldest citrus diseases and has been known for over a century. HLB is caused by 'Candidatus Liberibacter' spp. that are phloem-limited, fastidious α-proteobacteria and infect hosts in different Kingdoms (i.e., Animalia and Plantae). When compared with well-characterized, cultivatable plant-pathogenic Gram-negative bacteria, the interactions of uncultured insect-vectored plant-pathogenic bacteria, including 'Ca. Liberibacter' spp., with their hosts remain poorly understood. 'Ca. Liberibacter' spp. have been known to cause HLB, which has been rapidly spreading worldwide, resulting in dramatic economic losses. HLB presents an unprecedented challenge to citrus production. In this review, we focus on the most recent research on citrus, 'Candidatus Liberibacter asiaticus', and psyllid interactions, specifically considering the following topics: evolutionary relationships among 'Ca. Liberibacter' spp., genetic diversity, host range, genome analysis, transmission, virulence mechanisms, and the ecological importance of HLB. Currently, no efficient management strategy is available to control HLB, although some promising progress has been made. Further studies are needed to understand citrus, 'Ca. L. asiaticus', and psyllid interactions to design innovative management strategies. Although HLB has been problematic for over a century, we can only win the battle against HLB with a coordinated and deliberate effort by the citrus industry, citrus growers, researchers, legislatures, and governments.

  4. Development of consumer-friendly transgenic citrus plants with potential broad spectrum resistance to HLB, Citrus canker, Phytopthora and other exotic diseases

    USDA-ARS?s Scientific Manuscript database

    The second year of this CRB funded project has started, which is focused on the development of citrus cultivars that exhibit disease resistance to multiple pathogens such as HLB, Phytophthora and citrus canker diseases. We are using precise genetic engineering to introduce into disease susceptible ...

  5. Processes involved in the dispersal of Xanthomonas citri pv. citri from canker-infectd citrus canopies, and in the infection of citrus foliage

    USDA-ARS?s Scientific Manuscript database

    Citrus canker (Xanthomonas citri subsp. citri, Xcc) is now considered endemic in Florida, and epidemics result in yield loss and market penalties both in Florida, and elsewhere where the pathogen occurs, and susceptible citrus is cultivated. The bacterium is dispersed in rain splash, and storms wit...

  6. Processess involved in the dispersal of Xanthomonas citri pv. citri from canker-infected citrus canopies, and in the infection of citrus foliage

    USDA-ARS?s Scientific Manuscript database

    Citrus canker (Xanthomonas citri subsp. citri, Xcc) is now considered endemic in Florida, and epidemics result in yield loss and market penalties both in Florida and elsewhere, where the pathogen occurs and susceptible citrus is cultivated. The bacterium is dispersed in rain splash, and storms with...

  7. Processess involved in the dispersal of Xanthomonas citri pv. citri from canker-infected citrus canopies, and in the infection of citrus foliage

    USDA-ARS?s Scientific Manuscript database

    Citrus canker (Xanthomonas citri subsp. citri, Xcc) is now considered endemic in Florida, and epidemics result in yield loss and market penalties both in Florida and elsewhere, where the pathogen occurs and susceptible citrus is cultivated. The bacterium is dispersed in rain splash, and storms with...

  8. Ontogenic variation in citrus flush shoots and its relation with host plant finding and acceptance by Asian citrus psyllid (Hemiptera: Psyllidae)

    USDA-ARS?s Scientific Manuscript database

    The Asian citrus psyllid, Diaphorina citri (Hemiptera: Psyllidae) is a destructive insect mainly because it vectors the bacterial pathogens that cause the deadly and incurable citrus greening disease. Diaphorina citri adult females lay eggs and immature development occurs exclusively on new flush sh...

  9. Citrus Waste Biomass Program

    SciTech Connect

    Karel Grohman; Scott Stevenson

    2007-01-30

    Renewable Spirits is developing an innovative pilot plant bio-refinery to establish the commercial viability of ehtanol production utilizing a processing waste from citrus juice production. A novel process based on enzymatic hydrolysis of citrus processing waste and fermentation of resulting sugars to ethanol by yeasts was successfully developed in collaboration with a CRADA partner, USDA/ARS Citrus and Subtropical Products Laboratory. The process was also successfully scaled up from laboratory scale to 10,000 gal fermentor level.

  10. Resistance evaluation of Pera (Citrus sinensis) genotypes to citrus canker in greenhouse conditions

    USDA-ARS?s Scientific Manuscript database

    Citrus canker, caused by the bacterium Xanthomonas citri subsp. citri results in serious yield losses and phytoregulation penalties. The use of resistant genotypes is recognized as an important tool to facilitate control of the pathogen. Studies have show that artificial inoculation results in typic...

  11. The SLT2 mitogen-activated protein kinase-mediated signalling pathway governs conidiation, morphogenesis, fungal virulence and production of toxin and melanin in the tangerine pathotype of Alternaria alternata.

    PubMed

    Yago, Jonar Ingan; Lin, Ching-Hsuan; Chung, Kuang-Ren

    2011-09-01

    Fungi respond and adapt to different environmental stimuli via signal transduction systems. We determined the function of a yeast SLT2 mitogen-activated protein (MAP) kinase homologue (AaSLT2) in Alternaria alternata, the fungal pathogen of citrus. Analysis of the loss-of-function mutant indicated that AaSLT2 is required for the production of a host-selective toxin, and is crucial for fungal pathogenicity. Moreover, the A. alternata slt2 mutants displayed hypersensitivity to cell wall-degrading enzymes and chemicals such as Calcofluor white and Congo red. This implicates an important role of AaSLT2 in the maintenance of cell wall integrity in A. alternata. The A. alternata slt2 mutants were also hypersensitive to a heteroaromatic compound, 2-chloro-5-hydroxypyridine, and a plant growth regulator, 2,3,5-triiodobenzoic acid. Developmentally, the AaSLT2 gene product was shown to be critical for conidial formation and hyphal elongation. Compared with the wild-type, the mutants produced fewer but slightly larger conidia with less transverse septae. The mutants also accumulated lower levels of melanin and chitin. Unlike the wild-type progenitor, the A. alternata slt2 mutants produced globose, swollen hyphae that did not elongate in a straight radial direction. All defective phenotypes in the mutant were restored by transformation and expression of a wild-type copy of AaSLT2 under the control of its endogenous promoter. This study highlights an important role of the AaSLT2 MAP kinase-mediated signalling pathway, regulating diverse physiological, developmental and pathological functions, in the tangerine pathotype of A. alternata. © 2011 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2011 BSPP AND BLACKWELL PUBLISHING LTD.

  12. Exploiting fruit byproducts for eco-friendly nanosynthesis: Citrus × clementina peel extract mediated fabrication of silver nanoparticles with high efficacy against microbial pathogens and rat glial tumor C6 cells.

    PubMed

    Saratale, Rijuta Ganesh; Shin, Han-Seung; Kumar, Gopalakrishnan; Benelli, Giovanni; Ghodake, Gajanan S; Jiang, Yuan Yuan; Kim, Dong Su; Saratale, Ganesh Dattatraya

    2017-03-17

    Process byproducts from the fruit industry may represent a cheap and reliable source of green reducing agents to be used in current bio-nanosynthesis. This study reports the use of orange (Citrus × clementina) peel aqueous extract (OPE) for one-pot green synthesis of silver nanoparticles (AgNPs) with high effectiveness against various microbial pathogens as well as rat glial tumor C6 cells. The effects of various operational parameters on the synthesis of AgNPs were systematically investigated. The morphology, particle size, and properties of synthesized AgNPs were characterized using UV-visible spectroscopy, x-ray diffraction, x-ray photoelectron spectroscopy, field emission scanning electron microscopy, energy-dispersive x-ray spectroscopy, and Fourier transform infrared spectroscopy. High-resolution transmission electron microscopy shows that the nanoparticles are mostly spherical in shape and monodispersed, with an average particle size of 15-20 nm. Notably, the OPE-synthesized AgNPs were stable up to 6 months without change in their properties. Low doses of OPE-AgNPs inhibited the growth of human pathogens Escherichia coli, Bacillus cereus, and Staphylococcus aureus. The minimum inhibitory concentration and minimum bactericidal concentration of AgNPs against selected pathogenic bacteria were determined. OPE-AgNPs exhibited strong antioxidant activity in terms of ABTS (2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid)) radical scavenging (IC50 49.6 μg/mL) and DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging (IC50 63.4 μg/mL). OPE-AgNPs showed dose-dependent response against rat glial tumor C6 cells (LD50 60 μg/mL) showing a promising potential as anticancer agents. Overall, the current investigation highlighted a cheap green technology route to synthesize AgNPs using OPE byproducts and could potentially be utilized in biomedical, cosmetic, and pharmaceutical industry.

  13. Molecular diversity of Citrus tristeza virus (CTV) strains collected over the past 50 years and maintained in CTV collections in California

    USDA-ARS?s Scientific Manuscript database

    Tristeza, caused by Citrus tristeza virus (CTV), is a serious citrus disease worldwide. Because severe strains of CTV reduce fruit production and quality, CTV has been eliminated from citrus germplasm sources by a certification program. CTV is also a regulated pathogen in quarantine zones and infec...

  14. High incidence of preharvest colonization of huanglongbing-symptomatic Citrus sinensis fruit by Lasiodiplodia theobromae (Diplodia natalensis) and exacerbation of postharvest fruit decay by that fungus

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB), presumably caused by bacterium Candidatus Liberibacter asiaticus (CLas), is a devastating citrus disease associated with excessive pre-harvest fruit drop. Lasiodiplodia theobromae (Diplodia) is the causal organism of citrus stem end rot (SER). The pathogen infects citrus fruit ...

  15. Overexpression of a Modified Plant Thionin Enhances Disease Resistance to Citrus Canker and Huanglongbing (HLB).

    PubMed

    Hao, Guixia; Stover, Ed; Gupta, Goutam

    2016-01-01

    Huanglongbing (HLB or citrus greening disease) caused by Candidatus Liberibacter asiaticus (Las) is a great threat to the US citrus industry. There are no proven strategies to eliminate HLB disease and no cultivar has been identified with strong HLB resistance. Citrus canker is also an economically important disease associated with a bacterial pathogen (Xanthomonas citri). In this study, we characterized endogenous citrus thionins and investigated their expression in different citrus tissues. Since no HLB-resistant citrus cultivars have been identified, we attempted to develop citrus resistant to both HLB and citrus canker through overexpression of a modified plant thionin. To improve effectiveness for disease resistance, we modified and synthesized the sequence encoding a plant thionin and cloned into the binary vector pBinPlus/ARS. The construct was then introduced into Agrobacterium strain EHA105 for citrus transformation. Transgenic Carrizo plants expressing the modified plant thionin were generated by Agrobacterium-mediated transformation. Successful transformation and transgene gene expression was confirmed by molecular analysis. Transgenic Carrizo plants expressing the modified thionin gene were challenged with X. citri 3213 at a range of concentrations, and a significant reduction in canker symptoms and a decrease in bacterial growth were demonstrated compared to nontransgenic plants. Furthermore, the transgenic citrus plants were challenged with HLB via graft inoculation. Our results showed significant Las titer reduction in roots of transgenic Carrizo compared with control plants and reduced scion Las titer 12 months after graft inoculation. These data provide promise for engineering citrus disease resistance against HLB and canker.

  16. Fate and control of pathogenic and spoilage micro-organisms in orange blossom (Citrus aurantium) and rose flower (Rosa centifolia) hydrosols.

    PubMed

    Labadie, C; Cerutti, C; Carlin, F

    2016-12-01

    Hydrosols are hydrodistillation products used in food and cosmetic industries, perfumery, pharmacy and aromatherapy. The ability of preservatives to control previously reported bacterial proliferation and spoilage was evaluated. All tested preservatives were authorized for food and cosmetic application. Major pathogens of concern for foods and cosmetics were poorly able to grow in rose and orange blossom hydrosols when inoculated and incubated at 30°C. Commercial antimicrobials, such as isothiazolinone, chlorphenesin and paraben solutions, benzyl alcohol and sodium benzoate at pH = 5·0, controlled the growth of Pseudomonas and Burkholderia sp. strains representative of the natural microbiota of both hydrosols for >90 days at 30°C, only at concentrations close to the authorized limits. Concentrations of some of the tested preservatives that controlled growth at 5°C were lower than at 30°C. Pathogenic micro-organisms likely represent a low risk in rose flower and orange blossom hydrosol. However, the oligotrophic character of hydrosols and the antimicrobial properties of their essential oils do not prevent microbiological spoilage by the naturally present microbiota. In the absence of aseptic conditions and microbial inactivation process, only preservatives can stabilize hydrosols for a several-month storage. Several effective preservatives have been identified. © 2016 The Society for Applied Microbiology.

  17. Cercosporoid diseases of Citrus.

    PubMed

    Huang, Feng; Groenewald, J Z; Zhu, Li; Crous, P W; Li, Hongye

    2015-01-01

    Citrus leaves and fruits exhibiting disease symptoms ranging from greasy spot, yellow spot, small or large brown spot, black dot, and brown dot were sampled from Fujian, Guangdong, Guizhou, Hunan, Jiangxi, Yunnan, Zhejiang provinces and the Guangxi Zhuang Autonomous Region in China. In total 82 isolates representing various cercosporoid genera were isolated from these disease symptoms, which were supplemented with eight Citrus cercosporoid isolates collected from other countries. Based on a morphological and phylogenetic study using sequence data from the nuclear ribosomal DNA's ITS1-5.8S-ITS2 regions (ITS), and partial actin (act), β-tubulin (tub2), 28S nuclear ribosomal RNA (28S rDNA) and translation elongation factor 1-α (tef1) genes, these strains were placed in the following genera: Cercospora, Pallidocercospora, Passalora, Pseudocercospora, Verrucisporota and Zasmidium. All isolates tended to be sterile, except the Zasmidium isolates associated with citrus greasy spot-like symptoms, which subsequently were compared with phylogenetically similar isolates occurring on Citrus and other hosts elsewhere. From these results four Zasmidium species were recognized on Citrus, namely Z. indonesianum on Citrus in Indonesia, Z. fructicola and Z. fructigenum on Citrus in China and Z. citri-griseum, which appears to have a wide host range including Acacia, Citrus, Eucalyptus and Musa, as well as a global distribution.

  18. Certification Programs for Citrus

    USDA-ARS?s Scientific Manuscript database

    Citrus certification programs designed to ensure that healthy plants of the highest genetic potential are being planted in the field are the basic building block of an integrated pest management program. Certification programs began for citrus began with the discovery that the diseases were graft t...

  19. In vitro and in vivo activity of essential oils against major postharvest pathogens of Kinnow (Citrus nobilis × C. deliciosa) mandarin.

    PubMed

    Jhalegar, Md Jameel; Sharma, R R; Singh, Dinesh

    2015-04-01

    The present study envisages the influence of essential oils namely lemon grass, eucalyptus, clove and neem on Kinnow mandarin with the objective to combat major post harvest diseases and to prolong its availability for longer time in the season. For this, in vitro and in vivo studies were conducted. Poisoned food technique was used for in vitro studies, and for in vivo studies, Kinnow fruit were pre-inoculated with pathogens (Penicillium digitatum and P. italicum), treated with different essential oils and then stored at 5 °C ±1 °C temperature and 85-90 % RH). Our results indicated that all essential oils inhibited the growth (colony diameter) of both pathogens over untreated PDA plates, but the inhibition was the strongest by lemon grass oil. Similarly, under in vivo conditions, all essential oils influenced decay incidence, decay loss, lesion diameter, respiration rate, ethylene evolution, overall acceptability and physiological loss in weight but lemon grass was the most effective. And also the incidence of Penicillium italicum was more noticed in fruits than P. digitatum, however, it was reverse under in vitro conditions. The decay rot at all stages of storage was less in EOs treated fruits than untreated fruits, thereby increasing their storage life significantly. Thus, it is evident from our studies that essential oils have the potential to control green and blue mold without causing any injury or harmful effects on Kinnow mandarin, and EOs can be recommended as a safe method for extending its storage life while maintaining fruit quality.

  20. Molecular Detection of Spiroplasma Citri Associated with Stubborn Disease in Citrus Orchards in Syria

    USDA-ARS?s Scientific Manuscript database

    Spiroplasma citri, a phloem-limited pathogen, causes citrus stubborn disease (CSD) and can be transmitted from plant to plant by several species of phloem-feeding leafhoppers. CSD is an important disorder in certain warm and arid citrus-growing areas, and its agent has been recorded from several Med...

  1. An Inexpensive and Safe Experiment to Demonstrate Koch's Postulates Using Citrus Fruit

    ERIC Educational Resources Information Center

    Jakobi, Steven

    2010-01-01

    Citrus fruit (oranges, tangerines, grapefruit or lemons) purchased in a grocery store can be experimentally infected with readily-available sources of "Penicillium digitatum" to demonstrate the four basic steps of Koch's postulates, also known as proof of pathogenicity. The mould is isolated from naturally-infected citrus fruit into pure culture…

  2. Estimation of incidence and spatial temporal distribution of Citrus Stubborn disease

    USDA-ARS?s Scientific Manuscript database

    Citrus stubborn disease (CSD) is caused by Spiroplasma citri, a culturable wall-less prokaryote. The pathogen is graft-transmissible and vectored by the beet leafhopper (BLH). The objective of this study was to determine incidence and spread of S. citri in two sweet orange citrus groves in the San J...

  3. Rapid screening for citrus canker resistance employing pattern-triggered immunity (PTI) responses

    USDA-ARS?s Scientific Manuscript database

    Citrus canker, caused by the bacterial pathogen Xanthomonas citri ssp. citri (Xcc), has been attributed to millions of dollars in loss or damage to commercial citrus crops in subtropical production areas of the world. Since identification of resistant plants is one of the most effective methods of d...

  4. Entomopathogenic fungi infecting the Asian citrus psyllid, Diaphorina citri (Homoptera: Psyllidae), in Florida

    USDA-ARS?s Scientific Manuscript database

    The Asian citrus psyllid, Diaphorina citri is an invasive pest that vectors citrus greening disease. In 2005-2006 mycosed psyllids displaying two phenotypes were collected in central Florida. The major pathogen, identified by morphological and genetic analyses, was a novel isolate related to Hirsute...

  5. An Inexpensive and Safe Experiment to Demonstrate Koch's Postulates Using Citrus Fruit

    ERIC Educational Resources Information Center

    Jakobi, Steven

    2010-01-01

    Citrus fruit (oranges, tangerines, grapefruit or lemons) purchased in a grocery store can be experimentally infected with readily-available sources of "Penicillium digitatum" to demonstrate the four basic steps of Koch's postulates, also known as proof of pathogenicity. The mould is isolated from naturally-infected citrus fruit into pure culture…

  6. Evaluation of low-volume sprayers used in asian citrus Psyllid control applications

    USDA-ARS?s Scientific Manuscript database

    The Asian citrus psyllid (Diaphorina citri Kumayama, Sternorrhyncha: Psyllidae) is a detrimental pest to citrus crops when it serves as a vector of the pathogen that causes greening (huanglongbing, or HLB). Transmission of this disease causes mottling, chlorosis, dieback and reductions in both frui...

  7. Abundance of citrus leafminer larvae on citrus and citrus-related germplasm

    USDA-ARS?s Scientific Manuscript database

    The citrus leafminer, Phyllocnistis citrella Stainton (Lepidoptera: Gracillariidae), is a key pest in most citrus growing regions worldwide. Adult citrus leafminers oviposit primarily on young elongating flush of citrus as well as other Rutaceae and some ornamental plants. Larvae feed on the epiderm...

  8. First Report of Leaf Spot Caused by Alternaria tenuissima on Black Chokeberry (Aronia melanocarpa) in Korea.

    PubMed

    Wee, Jung-In; Park, Jong-Han; Back, Chang-Gi; You, Young-Hyun; Chang, Taehyun

    2016-09-01

    In July 2015, diseased leaves of black chokeberry (Aronia melanocarpa) were observed in Danyang and Gochang, Korea. The symptoms appeared as circular or irregular brown leaf spots, from which Alternaria tenuissima was isolated. The isolates were cultured on potato dextrose agar, and their morphological characteristics were observed under a light microscope. The colonies were whitish to ash colored. The pathogenicity test on healthy black chokeberry leaves produced circular brown spots, in line with the original symptoms. Molecular analyses of the ITS, GPD, RPB2, and TEF genes were conducted to confirm the identity of the pathogen. The phylogeny of the multi-gene sequences indicated that the causal agent was A. tenuissima. This study is the first report of A. tenuissima leaf spot on black chokeberry (A. melanocarpa).

  9. First Report of Leaf Spot Caused by Alternaria tenuissima on Black Chokeberry (Aronia melanocarpa) in Korea

    PubMed Central

    Wee, Jung-In; Park, Jong-Han; Back, Chang-Gi; You, Young-Hyun

    2016-01-01

    In July 2015, diseased leaves of black chokeberry (Aronia melanocarpa) were observed in Danyang and Gochang, Korea. The symptoms appeared as circular or irregular brown leaf spots, from which Alternaria tenuissima was isolated. The isolates were cultured on potato dextrose agar, and their morphological characteristics were observed under a light microscope. The colonies were whitish to ash colored. The pathogenicity test on healthy black chokeberry leaves produced circular brown spots, in line with the original symptoms. Molecular analyses of the ITS, GPD, RPB2, and TEF genes were conducted to confirm the identity of the pathogen. The phylogeny of the multi-gene sequences indicated that the causal agent was A. tenuissima. This study is the first report of A. tenuissima leaf spot on black chokeberry (A. melanocarpa). PMID:27790070

  10. Leaf proteome profiling of transgenic mint infected with Alternaria alternata.

    PubMed

    Sinha, Ragini; Bhattacharyya, Dipto; Majumdar, Aparupa Bose; Datta, Riddhi; Hazra, Saptarshi; Chattopadhyay, Sharmila

    2013-11-20

    The genus Mentha has been widely used in food, flavor, culinary, cosmetic and pharmaceutical industries. Substantial damage to this crop happened regularly due to environmental stresses like metal toxicity and pathogen attack. Here, an approach has been taken to raise transgenic mint over-expressing γ-glutamyl-cysteine synthetase (γ-ECS), the rate-limiting enzyme of GSH biosynthesis, resulted enhanced GSH content and its in planta expression confers significant tolerance towards abiotic/biotic stresses viz. metal toxicity - Cd, Zn as well as against infection of Alternaria alternata and Rhizoctonia solani. A differential proteomic analysis through 2-DE and MALDI TOF-TOF MSMS was performed to focus on the altered abundance of functionally important protein species in control and infected transgenic mint. Results showed a significant variation in the protein profile of the infected transgenic plant as compared to the wild/control transgenic counterpart. In addition to protein species related to stress and defense, redox regulation, transcription factors and energy & metabolism, protein species related to signaling and gene regulation as well as cell division also showed differential accumulation in infected transgenic. Hence, proteomics can be used as a tool to decipher the mechanism of action of GSH in providing tolerance against a necrotrophic fungus, A. alternata in transgenic mint. The reported work describes a comparative proteomics of non-model unsequenced plants like Mentha. There is a comparative protein profile between transgenic and its wild counterparts under control and infected condition. The work has an impact in crop proteomics and also tries to explain the application of proteomic approach to decipher the mechanism by which a foreign metabolite mediates stress tolerance in plant under control and infected condition. This article is part of a Special Issue entitled: Translational Plant Proteomics. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Volatilization of Selenium by Alternaria alternata

    PubMed Central

    Thompson-Eagle, E. T.; Frankenberger, W. T.; Karlson, U.

    1989-01-01

    Seleniferous water continues to be a serious problem to wildlife in the central valley of California. Water samples collected from Kesterson Reservoir, Peck Ranch, and Lost Hills evaporation pond facilities contained between 0.005 and 5 mg of Se per liter. The objective of this study was to isolate Se-methylating organisms in evaporation pond water and to assess, through enrichment and manipulation of their optimal growth parameters, the environmental factors which govern microbial Se methylation. Alternaria alternata was isolated as an active Se-methylating organism. The volatile product was identified as dimethylselenide. The effects of pH, temperature, Se substrates, and methyl donors on the ability of A. alternata to methylate Se were investigated in liquid medium containing 100 mg of Se per liter. The optimum pH and temperature for methylation were 6.5 and 30°C, respectively. Selenate and selenite were methylated more rapidly than selenium sulfide and various organic Se compounds (6-selenoguanosine, 6-selenoinosine, seleno-dl-methionine, and 6-selenopurine). l-Methionine and methyl cobalamine (0.1 μM) stimulated dimethylselenide production. This study demonstrates that Se-methylating organisms are present in evaporation pond water and are capable of liberating substantial quantities of Se in the volatile dimethylselenide form. By determining the optimum environmental conditions which stimulate volatilization, it may be possible to design a way to remove Se from seleniferous water in situ. PMID:16347933

  12. 75 FR 34322 - Citrus Greening and Asian Citrus Psyllid; Quarantine and Interstate Movement Regulations

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-17

    ...We are quarantining the States of Florida and Georgia, Puerto Rico, the U.S. Virgin Islands, two parishes in Louisiana, and two counties in South Carolina due to the presence of citrus greening and quarantining Alabama, Florida, Georgia, Guam, Hawaii, Louisiana, Mississippi, Puerto Rico, Texas, the U.S. Virgin Islands, three counties in South Carolina, portions of one county in Arizona, and all of three and portions of an additional three counties in California due to the presence of Asian citrus psyllid, a vector of the bacterial pathogen that causes citrus greening. This action follows the discovery of these pests in the respective quarantined areas. We are also establishing restrictions on the interstate movement of regulated articles from the quarantined areas. This action is necessary on an emergency basis in order to prevent the spread of the disease and its vector to noninfested areas of the United States.

  13. Quantitative distribution of 'Candidatus Liberibacter asiaticus' in citrus plants with citrus huanglongbing.

    PubMed

    Li, Wenbin; Levy, Laurene; Hartung, John S

    2009-02-01

    Citrus huanglongbing (HLB), or greening disease, is strongly associated with any of three nonculturable gram-negative bacteria belonging to 'Candidatus Liberibacter spp.' 'Ca. Liberibacter spp.' are transmitted by citrus psyllids to all commercial cultivars of citrus. The diseases can be lethal to citrus and have recently become widespread in both São Paulo, Brazil, and Florida, United States, the locations of the largest citrus industries in the world. Asiatic HLB, the form of the disease found in Florida, is associated with 'Ca. Liberibacter asiaticus' and is the subject of this report. The nonculturable nature of the pathogen has hampered research and little is known about the distribution of 'Ca. L. asiaticus' in infected trees. In this study, we have used a quantitative polymerase chain reaction assay to systematically quantify the distribution of 'Ca. L. asiaticus' genomes in tissues of six species of citrus either identified in the field during survey efforts in Florida or propagated in a greenhouse in Beltsville, MD. The populations of 'Ca. L. asiaticus' inferred from the distribution of 16S rDNA sequences specific for 'Ca. L. asiaticus' in leaf midribs, leaf blades, and bark samples varied by a factor of 1,000 among samples prepared from the six citrus species tested and by a factor of 100 between two sweet orange trees tested. In naturally infected trees, above-ground portions of the tree averaged 10(10) 'Ca. L. asiaticus' genomes per gram of tissue. Similar levels of 'Ca. L. asiaticus' genomes were observed in some but not all root samples from the same plants. In samples taken from greenhouse-inoculated trees, levels of 'Ca. L. asiaticus' genomes varied systematically from 10(4) genomes/g at the graft inoculation site to 10(10) genomes/g in some leaf petioles. Root samples from these trees also contained 'Ca. L. asiaticus' at 10(7) genomes/g. In symptomatic fruit tissues, 'Ca. L. asiaticus' genomes were also readily detected and quantified. The highest

  14. Examination of the responses of different genotypes of citrus to huanglongbing (citrus greening) under different conditions.

    PubMed

    Folimonova, Svetlana Y; Robertson, Cecile J; Garnsey, Stephen M; Gowda, Siddarame; Dawson, William O

    2009-12-01

    ABSTRACT Citrus Huanglongbing (HLB) is one of the most devastating diseases of citrus worldwide. The causal agent of HLB in Florida is thought to be 'Candidatus Liberibacter asiaticus'. In this work, we examined the responses of 30 different genotypes of citrus to Florida isolates of 'Ca. L. asiaticus' under controlled conditions in the greenhouse or growth room. Although 'Ca. L. asiaticus' was able to multiply in all of the plants, a wide range of responses was observed among different hosts. Based on the symptoms developed and the ability of plants to continue growth, the different genotypes were grouped into four categories: sensitive, which exhibited severe chlorosis on leaves, greatly reduced growth, and eventual death; moderately tolerant, which exhibited some scattered distinct symptoms but little or no growth reduction and no plant death; tolerant, which exhibited very minimal symptoms; and genotypes, which exhibited variable reactions. Interestingly, although 'Ca. L. asiaticus' was unevenly distributed within each particular plant, comparison of titers of the bacterium in different citrus genotypes revealed that most accumulated similar levels of 'Ca. L. asiaticus', demonstrating that there is no strict correlation between bacterial titer and severity of disease. Incubation of infected plants in the growth room with continuous light greatly affected symptoms production by reducing the time before distinctive symptoms developed and significantly increasing severity of chlorosis of leaves of all citrus genotypes. These results provide additional evidence of the correlation between disruption of phloem translocation of carbohydrates during HLB infection and the appearance of chlorotic symptoms in leaves of infected trees. We also examined interaction between 'Ca. L. asiaticus' and Citrus tristeza virus, which usually occurs in trees that become infected with HLB, and found no synergistic effect of the two pathogens. We trust that observations reported here

  15. 40 CFR 180.1256 - Alternaria destruens strain 059; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... RESIDUES IN FOOD Exemptions From Tolerances § 180.1256 Alternaria destruens strain 059; exemption from the... the microbial pesticide Alternaria destruens Strain 059 when used in or on all raw...

  16. 40 CFR 180.1256 - Alternaria destruens strain 059; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... RESIDUES IN FOOD Exemptions From Tolerances § 180.1256 Alternaria destruens strain 059; exemption from the... the microbial pesticide Alternaria destruens Strain 059 when used in or on all raw...

  17. Allergic bronchopulmonary mycosis due to Alternaria: Case report and review.

    PubMed

    Singh, Bhagteshwar; Denning, David W

    2012-01-01

    While allergic bronchopulmonary aspergillosis and mycosis are well recognised, no cases have been described related to Alternaria spp. Alternaria is a common sensitising fungus in asthmatics and related to thunderstorm asthma. We report a case of an asthmatic who presented with worsening asthma control, mild eosinophilia on high dose inhaled corticosteroids (800 μg/day), a total IgE of 3800 KIU/L, an Alternaria-specific IgE of 21.3 KUa/L and positive skin prick test, negative specific IgE and skin prick test to Aspergillus fumigatus, Penicillium spp., Cladosporium spp., Trichophyton spp. and a normal CT scan of the thorax. He responded well to a short course of oral prednisolone and then oral itraconazole, given over 17 months but relapsed 1 month after stopping it.

  18. Allergic bronchopulmonary mycosis due to Alternaria: Case report and review

    PubMed Central

    Singh, Bhagteshwar; Denning, David W.

    2012-01-01

    While allergic bronchopulmonary aspergillosis and mycosis are well recognised, no cases have been described related to Alternaria spp. Alternaria is a common sensitising fungus in asthmatics and related to thunderstorm asthma. We report a case of an asthmatic who presented with worsening asthma control, mild eosinophilia on high dose inhaled corticosteroids (800 μg/day), a total IgE of 3800 KIU/L, an Alternaria-specific IgE of 21.3 KUa/L and positive skin prick test, negative specific IgE and skin prick test to Aspergillus fumigatus, Penicillium spp., Cladosporium spp., Trichophyton spp. and a normal CT scan of the thorax. He responded well to a short course of oral prednisolone and then oral itraconazole, given over 17 months but relapsed 1 month after stopping it. PMID:24371728

  19. 40 CFR 180.1256 - Alternaria destruens strain 059; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Alternaria destruens strain 059... RESIDUES IN FOOD Exemptions From Tolerances § 180.1256 Alternaria destruens strain 059; exemption from the... the microbial pesticide Alternaria destruens Strain 059 when used in or on all raw...

  20. [Research of chemotaxis response of Botrytis cinerea and Alternaria panax on total ginsenosides].

    PubMed

    Chi, Kun; Xu, Yong-hua; Lei, Feng-jie; Yin, Min-jing; Wang, Zhuang; Zhang, Ai-hua; Zhang, Lian-xue

    2015-10-01

    In this paper, three kinds of chemotactic parameters (concentration, temperature and pH) were determined by plate assay and spore germination method to research the chemotactic response of Botrytis cinerea and Alternaria panax, and their spores on total ginsenosides. The results showed that Botrytis cinerea had strong chemotactic response at the mid-concentration of total ginsenosides (cultivation temperature was 20 degrees C and pH value was 6), and the data of chemotactic migration index (CMI) was 1.293 0, chemotactic growth rate (CGR) was 0.476 0, spore germination rate (SGR) was 53%, and dry weight of mycelial (DWM) was 0.452 6 g x L(-1); however, Alternaria panax had strong chemotactic response at the low-concentration of total ginsenosides (cultivation temperature was 25 degrees C and pH value was 6), and the data of chemotactic migration index (CMI) was 1.235 4, chemotactic growth rate (CGR) was 0.537 0, spore germination rate (SGR) was 67%, and dry weight of mycelial (DWM) was 0.494 8 g x L(-1). The results indicated that the low and middle concentration (2, 20 mg x L(-1)) of total ginsenosides had significant promoting effect on chemotactic response of these two pathogens, and the spore germination, mycelial growth rate, dry weight of mycelial of them were also significantly improved by this chemotactic response, whereas it decreased as the increase of total ginsenosides concentration.

  1. Toxigenic profile and AFLP variability of Alternaria alternata and Alternaria infectoria occurring on wheat

    PubMed Central

    Oviedo, María Silvina; Sturm, María Elena; Reynoso, María Marta; Chulze, Sofia Noemi; Ramirez, María Laura

    2013-01-01

    The objectives of this study were to evaluate the ability to produce alternariol (AOH), alternariol monomethyl ether (AME) and tenuazonic acid (TA) by A. alternata and A. infectoria strains recovered from wheat kernels obtained from one of the main production area in Argentina; to confirm using AFLPs molecular markers the identify of the isolates up to species level, and to evaluate the intra and inter-specific genetic diversity of these two Alternaria species. Among all the Alternaria strains tested (254), 84% of them were able to produce mycotoxins. The most frequent profile of toxin production found was the co-production of AOH and AME in both species tested. TA was only produced by strains of A. alternata. Amplified fragment polymorphism (AFLPs) analysis was applied to a set of 89 isolates of Alternaria spp (40 were A. infectoria and 49 were A. alternata) in order to confirm the morphological identification. The results showed that AFLPs are powerful diagnostic tool for differentiating between A. alternata and A. infectoria. Indeed, in the current study the outgroup strains, A. tenuissima was consistently classified. Characteristic polymorphic bands separated these two species regardless of the primer combination used. Related to intraspecific variability, A. alternata and A. infectoria isolates evaluated seemed to form and homogeneous group with a high degree of similarity among the isolates within each species. However, there was more scoreable polymorphism within A. alternata than within A. infectoria isolates. There was a concordance between morphological identification and separation up to species level using molecular markers. Clear polymorphism both within and between species showed that AFLP can be used to asses genetic variation in A. alternata and A. infectoria. The most important finding of the present study was the report on AOH and AME production by A. infectoria strains isolated from wheat kernels in Argentina on a semisynthetic media for the

  2. Mould and yeast flora in fresh berries, grapes and citrus fruits.

    PubMed

    Tournas, V H; Katsoudas, Eugenia

    2005-11-15

    Fresh fruits are prone to fungal contamination in the field, during harvest, transport, marketing, and with the consumer. It is important to identify fungal contaminants in fresh fruits because some moulds can grow and produce mycotoxins on these commodities while certain yeasts and moulds can cause infections or allergies. In this study, 251 fresh fruit samples including several varieties of grapes, strawberries, blueberries, raspberries, blackberries, and various citrus fruits were surface-disinfected, incubated at room temperature for up to 14 days without supplemental media, and subsequently examined for mould and yeast growth. The level of contamination (percent of contaminated items/sample) varied depending on the type of fruit. All raspberry and blackberry samples were contaminated at levels ranging from 33% to 100%, whereas 95% of the blueberry samples supported mould growth at levels between 10% and 100% of the tested berries, and 97% of strawberry samples showed fungal growth on 33-100% of tested berries. The most common moulds isolated from these commodities were Botrytis cinerea, Rhizopus (in strawberries), Alternaria, Penicillium, Cladosporium and Fusarium followed by yeasts, Trichoderma and Aureobasidium. Thirty-five percent of the grape samples tested were contaminated and supported fungal growth; the levels of contamination ranged from 9% to 80%. The most common fungi spoiling grapes were Alternaria, B. cinerea and Cladosporium. Eighty-three percent of the citrus fruit samples showed fungal growth at levels ranging from 25% to 100% of tested fruits. The most common fungi in citrus fruits were Alternaria, Cladosporium, Penicillium, Fusarium and yeasts. Less common were Trichoderma, Geotrichum and Rhizopus.

  3. A Novel Alternaria Species Isolated from Peucedanum japonicum in Korea

    PubMed Central

    Deng, Jian Xin; Cho, Hye Sun; Paul, Narayan Chandra; Lee, Hyang Burm

    2014-01-01

    We isolated and examined a new Alternaria sp., which causes leaf spots on Peucedanum japonicum in Korea, by using molecular and morphological methods. Phylogenetic analysis based on a combined internal transcribed spacer region analysis and two protein-coding genes (gpd and Alt a1) demonstrated that the causal fungus was most closely related to A. cinerariae and A. sonchi, and relevant to A. brassicae. However, conidial morphology indicated that it is a novel species within the genus Alternaria, and therefore we have assigned the fungus a new name in this study. PMID:24808728

  4. A Novel Alternaria Species Isolated from Peucedanum japonicum in Korea.

    PubMed

    Deng, Jian Xin; Cho, Hye Sun; Paul, Narayan Chandra; Lee, Hyang Burm; Yu, Seung Hun

    2014-03-01

    We isolated and examined a new Alternaria sp., which causes leaf spots on Peucedanum japonicum in Korea, by using molecular and morphological methods. Phylogenetic analysis based on a combined internal transcribed spacer region analysis and two protein-coding genes (gpd and Alt a1) demonstrated that the causal fungus was most closely related to A. cinerariae and A. sonchi, and relevant to A. brassicae. However, conidial morphology indicated that it is a novel species within the genus Alternaria, and therefore we have assigned the fungus a new name in this study.

  5. Effect of climate change on Alternaria leaf spot of rocket salad and black spot of basil under controlled environment.

    PubMed

    Pugliese, M; Cogliati, E; Gullino, M L; Garibaldi, A

    2012-01-01

    Plant responses to elevated CO2 and temperature have been much studied in recent years, but effects of climate change on pathological responses are still largely unknown. The pathosystems rocket (Eruca vesicaria subsp. sativa)--Alternaria leaf spot (Alternaria japonica) and basil (Ocimum basilicum)--black spot (Colletotrichum gloeosporioides) were chosen as models to assess the potential impact of increased CO2 and temperature on disease incidence and severity under controlled environment. Potted plants were grown in phytotrons under 4 different simulated climatic conditions: (1) standard temperature (ranging from 18 degrees to 22 degrees C) and standard CO2 concentration (400 ppm); (2) standard temperature and elevated CO2 concentration (800 ppm); (3) elevated temperature (ranging from 22 degrees to 26 degrees C, 4 degrees C higher than standard) and standard CO2 concentration; (4) elevated temperature and CO2 concentration. Each plant was inoculated with a spore suspension containing 1 x 10(5) cfu/ml of the pathogen. Disease incidence and severity were assessed 14 days after inoculation. Increasing CO2 to 800 ppm showed a clear increment in the percentage of Alternaria leaf spot on rocket leaves compared to standard conditions. Basil plants grown at 800 ppm of CO2 showed increased black spot symptoms compared to 400 ppm. Disease incidence and severity were always influenced by the combination of rising CO2 and increased temperature, compared to standard conditions (400 ppm of CO2 - 22 degrees C). Considering the rising concentrations of CO2 and global temperature, we can assume that this could increase the severity of Alternaria japonica on rocket and Colletotrichum gloeosporioides on basil.

  6. Overexpression of a citrus NDR1 ortholog increases disease resistance in Arabidopsis

    PubMed Central

    Lu, Hua; Zhang, Chong; Albrecht, Ute; Shimizu, Rena; Wang, Guanfeng; Bowman, Kim D.

    2013-01-01

    Emerging devastating diseases, such as Huanglongbing (HLB) and citrus canker, have caused tremendous losses to the citrus industry worldwide. Genetic engineering is a powerful approach that could allow us to increase citrus resistance against these diseases. The key to the success of this approach relies on a thorough understanding of defense mechanisms of citrus. Studies of Arabidopsis and other plants have provided a framework for us to better understand defense mechanisms of citrus. Salicylic acid (SA) is a key signaling molecule involved in basal defense and resistance (R) gene-mediated defense against broad-spectrum pathogens. The Arabidopsis gene NDR1 (NON-RACE-SPECIFIC DISEASE RESISTANCE 1) is a positive regulator of SA accumulation and is specifically required for signaling mediated by a subset of R genes upon recognition of their cognate pathogen effectors. Our bioinformatic analysis identified an ortholog of NDR1 from citrus, CsNDR1. Overexpression of CsNDR1 complemented susceptibility conferred by the Arabidopsis ndr1-1 mutant to Pseudomonas syringae strains and also led to enhanced resistance to an oomycete pathogen Hyaloperonospora arabidopsidis. Such heightened resistance is associated with increased SA production and expression of the defense marker gene PATHOGENESIS RELATED 1 (PR1). In addition, we found that expression of PR1 and accumulation of SA were induced to modest levels in citrus infected with Candidatus Liberibacter asiaticus, the bacterial pathogen associated with HLB disease. Thus, our data suggest that CsNDR1 is a functional ortholog of Arabidopsis NDR1. Since Ca. L. asiaticus infection only activates modest levels of defense responses in citrus, we propose that genetically increasing SA/NDR1-mediated pathways could potentially lead to enhanced resistance against HLB, citrus canker, and other destructive diseases challenging global citrus production. PMID:23761797

  7. Overexpression of a citrus NDR1 ortholog increases disease resistance in Arabidopsis.

    PubMed

    Lu, Hua; Zhang, Chong; Albrecht, Ute; Shimizu, Rena; Wang, Guanfeng; Bowman, Kim D

    2013-01-01

    Emerging devastating diseases, such as Huanglongbing (HLB) and citrus canker, have caused tremendous losses to the citrus industry worldwide. Genetic engineering is a powerful approach that could allow us to increase citrus resistance against these diseases. The key to the success of this approach relies on a thorough understanding of defense mechanisms of citrus. Studies of Arabidopsis and other plants have provided a framework for us to better understand defense mechanisms of citrus. Salicylic acid (SA) is a key signaling molecule involved in basal defense and resistance (R) gene-mediated defense against broad-spectrum pathogens. The Arabidopsis gene NDR1 (NON-RACE-SPECIFIC DISEASE RESISTANCE 1) is a positive regulator of SA accumulation and is specifically required for signaling mediated by a subset of R genes upon recognition of their cognate pathogen effectors. Our bioinformatic analysis identified an ortholog of NDR1 from citrus, CsNDR1. Overexpression of CsNDR1 complemented susceptibility conferred by the Arabidopsis ndr1-1 mutant to Pseudomonas syringae strains and also led to enhanced resistance to an oomycete pathogen Hyaloperonospora arabidopsidis. Such heightened resistance is associated with increased SA production and expression of the defense marker gene PATHOGENESIS RELATED 1 (PR1). In addition, we found that expression of PR1 and accumulation of SA were induced to modest levels in citrus infected with Candidatus Liberibacter asiaticus, the bacterial pathogen associated with HLB disease. Thus, our data suggest that CsNDR1 is a functional ortholog of Arabidopsis NDR1. Since Ca. L. asiaticus infection only activates modest levels of defense responses in citrus, we propose that genetically increasing SA/NDR1-mediated pathways could potentially lead to enhanced resistance against HLB, citrus canker, and other destructive diseases challenging global citrus production.

  8. Citrus diseases with global ramifications including citrus canker and huanglongbing

    USDA-ARS?s Scientific Manuscript database

    Although there are a number of diseases that plague citrus production worldwide, two bacterial diseases are particularly problematic. Both are of Asian origin and currently cause severe economic damage: Asiatic citrus canker (ACC) and citrus huanglongbing (HLB). Although ACC has been found in the ...

  9. Developing Transgenic Citrus for Resistance to Huanglongbing and Citrus Canker

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB) and Citrus Bacterial Canker (CBC) are serious threats to citrus production, and resistant transgenic citrus is desirable. Genes for antimicrobial peptides (AMPs) with diverse promoters have been used to generate thousands of rootstock and scion transformants. D35S::D4E1 transfor...

  10. A zinc-binding citrus protein metallothionein can act as a plant defense factor by controlling host-selective ACR-toxin production.

    PubMed

    Nishimura, Satoshi; Tatano, Satoshi; Miyamoto, Yoko; Ohtani, Kouhei; Fukumoto, Takeshi; Gomi, Kenji; Tada, Yasuomi; Ichimura, Kazuya; Akimitsu, Kazuya

    2013-01-01

    Metallothionein is a small cysteine-rich protein known to have a metal-binding function. We isolated three different lengths of rough lemon cDNAs encoding a metallothionein (RlemMT1, RlemMT2 and RlemMT3), and only RlemMT1-recombinant protein had zinc-binding activity. Appropriate concentration of zinc is an essential micronutrient for living organisms, while excess zinc is toxic. Zinc also stimulates the production of host-selective ACR-toxin for citrus leaf spot pathogen of Alternaria alternata rough lemon pathotype. Trapping of zinc by RlemMT1-recombinant protein or by a zinc-scavenging agent in the culture medium caused suppression of ACR-toxin production by the fungus. Since ACR-toxin is the disease determinant for A. alternata rough lemon pathotype, addition of RlemMT1 to the inoculum suspension led to a significant decrease in symptoms on rough lemon leaves as a result of reduced ACR-toxin production from the zinc trap around infection sites. RlemMT1-overexpression mutant of A. alternata rough lemon pathotype also produced less ACR-toxin and reduced virulence on rough lemon. This suppression was caused by an interruption of zinc absorption by cells from the trapping of the mineral by RlemMT1 and an excess supplement of ZnSO(4) restored toxin production and pathogenicity. Based on these results, we propose that zinc adsorbents including metallothionein likely can act as a plant defense factor by controlling toxin biosynthesis via inhibition of zinc absorption by the pathogen.

  11. Modeling huanglongbing transmission within a citrus tree

    PubMed Central

    Chiyaka, Christinah; Singer, Burton H.; Halbert, Susan E.; Morris, J. Glenn; van Bruggen, Ariena H. C.

    2012-01-01

    The citrus disease huanglongbing (HLB), associated with an uncultured bacterial pathogen, is threatening the citrus industry worldwide. A mathematical model of the transmission of HLB between its psyllid vector and citrus host has been developed to characterize the dynamics of the vector and disease development, focusing on the spread of the pathogen from flush to flush (a newly developing cluster of very young leaves on the expanding terminal end of a shoot) within a tree. This approach differs from that of prior models for vector-transmitted plant diseases where the entire plant is the unit of analysis. Dynamics of vector and host populations are simulated realistically as the flush population approaches complete infection. Model analysis indicates that vector activity is essential for initial infection but is not necessary for continued infection because infection can occur from flush to flush through internal movement in the tree. Flush production, within-tree spread, and latent period are the most important parameters influencing HLB development. The model shows that the effect of spraying of psyllids depends on time of initial spraying, frequency, and efficacy of the insecticides. Similarly, effects of removal of symptomatic flush depend on the frequency of removal and the time of initiation of this practice since the start of the epidemic. Within-tree resistance to spread, possibly affected by inherent or induced resistance, is a major factor affecting epidemic development, supporting the notion that alternate routes of transmission besides that by the vector can be important for epidemic development. PMID:22783015

  12. Modeling huanglongbing transmission within a citrus tree.

    PubMed

    Chiyaka, Christinah; Singer, Burton H; Halbert, Susan E; Morris, J Glenn; van Bruggen, Ariena H C

    2012-07-24

    The citrus disease huanglongbing (HLB), associated with an uncultured bacterial pathogen, is threatening the citrus industry worldwide. A mathematical model of the transmission of HLB between its psyllid vector and citrus host has been developed to characterize the dynamics of the vector and disease development, focusing on the spread of the pathogen from flush to flush (a newly developing cluster of very young leaves on the expanding terminal end of a shoot) within a tree. This approach differs from that of prior models for vector-transmitted plant diseases where the entire plant is the unit of analysis. Dynamics of vector and host populations are simulated realistically as the flush population approaches complete infection. Model analysis indicates that vector activity is essential for initial infection but is not necessary for continued infection because infection can occur from flush to flush through internal movement in the tree. Flush production, within-tree spread, and latent period are the most important parameters influencing HLB development. The model shows that the effect of spraying of psyllids depends on time of initial spraying, frequency, and efficacy of the insecticides. Similarly, effects of removal of symptomatic flush depend on the frequency of removal and the time of initiation of this practice since the start of the epidemic. Within-tree resistance to spread, possibly affected by inherent or induced resistance, is a major factor affecting epidemic development, supporting the notion that alternate routes of transmission besides that by the vector can be important for epidemic development.

  13. Deep sequencing of small RNAs from citrus affected by graft-transmissible diseases of unknown etiology leads to discovery of two novel viruses

    USDA-ARS?s Scientific Manuscript database

    Citrus is susceptible to graft-transmissible pathogens; some of which are arthropod-transmitted. Citrus cultivars vary in susceptibility or tolerance to these pathogens and may remain symptomless when infected. Established tests for known disease agents are used in certification and disease manage...

  14. Genetic variability in the pistachio late blight fungus, Alternaria alternata

    USDA-ARS?s Scientific Manuscript database

    Genetic variation in the pistachio late blight fungus, Alternaria alternata, was investigated by restriction fragment length polymorphism (RFLP) in the rDNA region. Southern hybridization of EcoRI, HindIII, and Xbal digested fungal DNA with a RNA probe derived from Alt1, an rDNA clone isolated from ...

  15. [Cellulase and xylanase activity of phytopathogenic and endophytic fungal strains of Alternaria alternata (Fr.) Keissler].

    PubMed

    Kurchenko, I M; Sokolova, O V; Zhdanova, N M; Iarynchyn, A M; Iovenko, O M

    2008-01-01

    A comparative analysis of cellulase and xylanase activity of 25 fungal strains of phytopathogenic and endophytic Alternaria alternata had been realized for the first time using the qualitative reactions. The rate of their linear growth on the media with carboxymethylcellulose or xylane had been studied. The cellulase and xylanase activities clearly depended on the distinct strain. The absence of distinct dependence of cellulase and xylanase activities on the species and organs of host plants was demonstrated. The majority of investigated strains of A. alternata did not possess a cellulase activity or the latter was low, but as a whole the phytopathogenic strains were more active than endophytic ones. Xylanase activity was considerable for the fungal strains of all trophyc groups. It was shown that the level of xylanase activity cannot become a biochemical marker of the A. alternata isolate pathogenicity.

  16. Huanglongbing: Devastating disease of citrus

    USDA-ARS?s Scientific Manuscript database

    Citrus greening or Huanglongbing (HLB) disease is devastating the citrus industry worldwide, from Asia to the Americas. The only major citrus regions free of this disease are the Mediterranean and Australia/New Zealand. To date there is no cure, no resistance and no therapy for HLB and production ra...

  17. Genetic Architecture of Resistance to Alternaria brassicae in Arabidopsis thaliana: QTL Mapping Reveals Two Major Resistance-Conferring Loci

    PubMed Central

    Rajarammohan, Sivasubramanian; Kumar, Amarendra; Gupta, Vibha; Pental, Deepak; Pradhan, Akshay K.; Kaur, Jagreet

    2017-01-01

    Alternaria brassicae, a necrotrophic fungal pathogen, causes Alternaria blight, one of the most important diseases of oleiferous Brassica crops. The current study utilized Arabidopsis as a model to decipher the genetic architecture of defense against A. brassicae. Significant phenotypic variation that was largely genetically determined was observed among Arabidopsis accessions in response to pathogen challenge. Three biparental mapping populations were developed from three resistant accessions viz. CIBC-5, Ei-2, and Cvi-0 and two susceptible accessions – Gre-0 and Zdr-1 (commonly crossed to CIBC-5 and Ei-2). A total of six quantitative trait locus (QTLs) governing resistance to A. brassicae were identified, five of which were population-specific while one QTL was common between all the three mapping populations. Interestingly, the common QTL had varying phenotypic contributions in different populations, which can be attributed to the genetic background of the parental accessions. The presence of both common and population-specific QTLs indicate that resistance to A. brassicae is quantitative, and that different genes may mediate resistance to the pathogen in different accessions. Two of the QTLs had moderate-to-large effects, one of which explained nearly 50% of the variation. The large effect QTLs may therefore contain genes that could play a significant role in conferring resistance even in heterologous hosts. PMID:28286515

  18. Airborne fungal spores of Alternaria, meteorological parameters and predicting variables

    NASA Astrophysics Data System (ADS)

    Filali Ben Sidel, Farah; Bouziane, Hassan; del Mar Trigo, Maria; El Haskouri, Fatima; Bardei, Fadoua; Redouane, Abdelbari; Kadiri, Mohamed; Riadi, Hassane; Kazzaz, Mohamed

    2015-03-01

    Alternaria is frequently found as airborne fungal spores and is recognized as an important cause of respiratory allergies. The aerobiological monitoring of fungal spores was performed using a Burkard volumetric spore traps. To establish predicting variables for daily and weakly spore counts, a stepwise multiple regression between spore concentrations and independent variables (meteorological parameters and lagged values from the series of spore concentrations: previous day or week concentration (Alt t - 1) and mean concentration of the same day or week in other years ( C mean)) was made with data obtained during 2009-2011. Alternaria conidia are present throughout the year in the atmosphere of Tetouan, although they show important seasonal fluctuations. The highest levels of Alternaria spores were recorded during the spring and summer or autumn. Alternaria showed maximum daily values in April, May or October depending on year. When the spore variables of Alternaria, namely C mean and Alt t - 1, and meteorological parameters were included in the equation, the resulting R 2 satisfactorily predict future concentrations for 55.5 to 81.6 % during the main spore season and the pre-peak 2. In the predictive model using weekly values, the adjusted R 2 varied from 0.655 to 0.676. The Wilcoxon test was used to compare the results from the expected values and the pre-peak spore data or weekly values for 2012, indicating that there were no significant differences between series compared. This test showed the C mean, Alt t - 1, frequency of the wind third quadrant, maximum wind speed and minimum relative humidity as the most efficient independent variables to forecast the overall trend of this spore in the air.

  19. Amino acids implicated in plant defense are higher in Candidatus Liberibacter asiaticus-tolerant citrus varieties

    PubMed Central

    Killiny, Nabil; Hijaz, Faraj

    2016-01-01

    ABSTRACT Citrus Huanglongbing (HLB), also known as citrus greening, has been threatening the citrus industry since the early 1900's and up to this date there are no effective cures for this disease. Field observations and greenhouse controlled studies demonstrated that some citrus genotypes are more tolerant to Candidatus Liberibacter asiaticus (CLas) pathogen than others. However, the mechanisms underpinning tolerance has not been determined yet. The phloem sap composition of CLas-tolerant and sensitive citrus varieties was studied to identify metabolites that could be responsible for their tolerance to CLas. The citrus phloem sap was collected by centrifugation and was analyzed with gas chromatography-mass spectrometry after methyl chloroformate derivatization. Thirty-three metabolites were detected in the phloem sap of the studied varieties: twenty 20 amino acids, eight 8 organic acids, and five 5 fatty acids. Interestingly, the levels of most amino acids, especially those implicated in plantdefense to pathogens such as phenylalanine, tyrosine, tryptophan, lysine, and asparagine were higher in tolerant varieties. Although the level of organic acids varied between cultivars, this variation was not correlated with citrus resistance to CLas and could be cultivar specific. The fatty acids were found in trace amounts and in most cases their levels were not significantly different among varieties. Better understanding of the mechanisms underpinning citrus tolerance to CLas will help in developing economically tolerant varieties. PMID:27057814

  20. Amino acids implicated in plant defense are higher in Candidatus Liberibacter asiaticus-tolerant citrus varieties.

    PubMed

    Killiny, Nabil; Hijaz, Faraj

    2016-01-01

    Citrus Huanglongbing (HLB), also known as citrus greening, has been threatening the citrus industry since the early 1900's and up to this date there are no effective cures for this disease. Field observations and greenhouse controlled studies demonstrated that some citrus genotypes are more tolerant to Candidatus Liberibacter asiaticus (CLas) pathogen than others. However, the mechanisms underpinning tolerance has not been determined yet. The phloem sap composition of CLas-tolerant and sensitive citrus varieties was studied to identify metabolites that could be responsible for their tolerance to CLas. The citrus phloem sap was collected by centrifugation and was analyzed with gas chromatography-mass spectrometry after methyl chloroformate derivatization. Thirty-three metabolites were detected in the phloem sap of the studied varieties: twenty 20 amino acids, eight 8 organic acids, and five 5 fatty acids. Interestingly, the levels of most amino acids, especially those implicated in plantdefense to pathogens such as phenylalanine, tyrosine, tryptophan, lysine, and asparagine were higher in tolerant varieties. Although the level of organic acids varied between cultivars, this variation was not correlated with citrus resistance to CLas and could be cultivar specific. The fatty acids were found in trace amounts and in most cases their levels were not significantly different among varieties. Better understanding of the mechanisms underpinning citrus tolerance to CLas will help in developing economically tolerant varieties.

  1. Citrus stubborn disease incidence determined by quantitative real time PCR

    USDA-ARS?s Scientific Manuscript database

    Quantitative real-time (q) PCR was developed for detection of Spiroplasma citri, the causal agent of citrus stubborn disease (CSD), using the DNA binding fluorophore SYBR Green I. The primer pair, P58-3f/4r, developed based on sequences from the P58 putative adhesin multigene of the pathogen result...

  2. Response of Citrus Seedlings to Radopholus similis in Two Soils.

    PubMed

    O'Bannon, J H; Tomerlin, A T

    1971-07-01

    Radopholus similis was less pathogenic to greenhouse-grown citrus seedlings in Leon loamy sand than in Lakeland fine sand. This was not affected by different watering regimes. Seedling growth reduction by the burrowing nematode in either soil, compared with noninfected controls, was significant at the 1% level.

  3. Asian citrus psyllid genome (Diaphorina citri, Hemiptera)-Update 2013

    USDA-ARS?s Scientific Manuscript database

    The psyllid genome is a scientific breakthrough that opens the psyllid genetic blueprint to investigations of all questions ranging from taxonomic origins to the understanding of developmental biology, to the acquisition and transmission of pathogens. The Asian citrus psyllid, Diaphorina citri (Hemi...

  4. Similar and distinct roles of NADPH oxidase components in the tangerine pathotype of Alternaria alternata.

    PubMed

    Yang, Siwy Ling; Chung, Kuang-Ren

    2013-08-01

    The fungal nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) complex, which has been implicated in the production of low-level reactive oxygen species (ROS), contains mainly NoxA, NoxB (gp91(phox) homologues) and NoxR (p67(phox) homologue). Here, we report the developmental and pathological functions of NoxB and NoxR in the tangerine pathotype of Alternaria alternata. Loss-of-function genetics revealed that all three Nox components are required for the accumulation of cellular hydrogen peroxide (H₂O₂). Alternaria alternata strains lacking NoxA, NoxB or NoxR also displayed an increased sensitivity to H₂O₂ and many ROS-generating oxidants. These phenotypes are highly similar to those previously seen for the Δyap1 mutant lacking a YAP1 transcriptional regulator and for the Δhog1 mutant lacking a HOG1 mitogen-activated protein (MAP) kinase, implicating a possible link among them. A fungal strain carrying a NoxA NoxB or NoxA NoxR double mutation was more sensitive to the test compounds than the strain mutated at a single gene, implicating a synergistic function among Nox components. The ΔnoxB mutant strain failed to produce any conidia; both ΔnoxA and ΔnoxR mutant strains showed a severe reduction in sporulation. Mutant strains carrying defective NoxB had higher chitin content than the wild-type and were insensitive to calcofluor white, Congo red and the fungicides vinclozolin and fludioxonil. Virulence assays revealed that all three Nox components are required for the elaboration of the penetration process. The inability to penetrate the citrus host, observed for Δnox mutants, could be overcome by wounding and by reacquiring a dominant Nox gene. The A. alternata NoxR did not influence the expression of NoxB, but negatively regulated NoxA. Importantly, the expression of both YAP1 and HOG1 genes, whose products are involved in resistance to ROS, was down-regulated in fungi carrying defective NoxA, NoxB or NoxR. Our results highlight the

  5. 7 CFR 301.76-3 - Quarantined areas; citrus greening and Asian citrus psyllid.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 5 2013-01-01 2013-01-01 false Quarantined areas; citrus greening and Asian citrus...) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE DOMESTIC QUARANTINE NOTICES Citrus Greening and Asian Citrus Psyllid § 301.76-3 Quarantined areas; citrus greening and Asian citrus...

  6. 7 CFR 301.76-3 - Quarantined areas; citrus greening and Asian citrus psyllid.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 5 2014-01-01 2014-01-01 false Quarantined areas; citrus greening and Asian citrus...) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE DOMESTIC QUARANTINE NOTICES Citrus Greening and Asian Citrus Psyllid § 301.76-3 Quarantined areas; citrus greening and Asian citrus...

  7. 7 CFR 301.76-3 - Quarantined areas; citrus greening and Asian citrus psyllid.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 5 2012-01-01 2012-01-01 false Quarantined areas; citrus greening and Asian citrus...) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE DOMESTIC QUARANTINE NOTICES Citrus Greening and Asian Citrus Psyllid § 301.76-3 Quarantined areas; citrus greening and Asian citrus...

  8. Deciphering the bacterial microbiome of citrus plants in response to ‘Candidatus Liberibacter asiaticus’-infection and antibiotic treatment

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB), the most devastating citrus disease worldwide, is vectored by phloem-feeding insects, and the pathogen in the USA is Candidatus Liberibacter asiaticus (Las). The bacterial microbiome of citrus after Las-infection and treatments with ampicillin (Amp) and gentamicin (Gm) was chara...

  9. Characteristics of the perception of different severity measures of citrus canker and the relations between the various symptom types

    USDA-ARS?s Scientific Manuscript database

    Citrus canker is a disease of citrus and is caused by the bacterial pathogen Xanthomonas axonopodis pv citri (Xac). Ways of managing the disease are being sought, and accurate, precise, reproducible disease assessment is needed for monitoring epidemics. The objective of this study was to investigate...

  10. Comparative phylogenomics and multi-gene cluster analyses of the CitrusHuanglongbing (HLB)-associated bacterium Candidatus Liberibacter

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB, previously known as citrus greening), is associated with Candidatus Liberibacter species and is a serious threat to citrus production world-wide. The pathogen is a Gram negative, unculturable, phloem-limited bacterium with limited known genomic information. Expanding the genetic ...

  11. Comparative Genomics and Phylogenetic Analyses of Newly Cloned Genomic Regions From the Citrus Huanglongbing (HLB)-associated Bacterium Candidatus Liberibacter

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB), or citrus greening disease, caused by Candidatus Liberibacter species, is a serious threat to citrus production worldwide. The pathogen is a gram negative, unculturable, phloem-limited bacterium, with little known genomic information. Here, we report cloning and characterizatio...

  12. Local genetic diversity of Xanthomonas citri subsp. citri in citrus orchards in northwest Paraná state, Brazil

    USDA-ARS?s Scientific Manuscript database

    Xanthomonas citri subsp. citri, causal agent of Asiatic citrus canker, is an important pathogen of citrus in Brazil and elsewhere. The genetic diversity of X. citri subsp. citri pathtype ‘A’ has not been studied in Brazil at a local scale (up to 300 km). A total of 40 isolates were collected from le...

  13. Morphological, molecular and virulence characterization of three Lencanicillium species infecting Asian citrus psyllids in Huangyan citrus groves.

    PubMed

    Lu, Lianming; Cheng, Baoping; Du, Danchao; Hu, Xiurong; Peng, Aitian; Pu, Zhanxu; Zhang, Xiaoya; Huang, Zhendong; Chen, Guoqing

    2015-02-01

    Citrus greening or Huanglongbing (HLB) is caused by the infection of Candidatus Liberibacter spp. in citrus plants. Since Asian citrus psyllid is the primary vector of this bacterial pathogen, the spread of HLB can be mitigated by suppressing Asian citrus psyllid populations in citrus groves using entomopathogens. To expand the current data on entomopathogens infecting Asian citrus psyllids, we isolated and characterized three different entomopathogens. Strains ZJLSP07, ZJLA08, and ZJLP09 infected the Asian citrus psyllid, Diaphorina citri Kuwayama, in Huangyan citrus groves. Based on molecular and morphological analyses, two were identified as Lecanicillium attenuatum and Lecanicillium psalliotae, and the third was recognized as an unidentified species of the genus, Lecanicillium. The corrected mortalities caused by strains ZJLSP07, ZJLA08 were 100% at 7days post-inoculation, while by ZJLP09 complete mortality occurred at 6days after inoculation, with 1.0×10(8)conidia/ml at 25°C and a relative humidity of 90% in the laboratory. Under the same condition, the corrected mortalities caused by strains ZJLSP07, ZJLA08 and ZJLP09 were 100%, 92.55% and 100%, respectively at 9days post-inoculation in the greenhouse. Our findings also revealed that these fungal strains infected D. citri using hyphae that penetrated deep into the insect tissues. Further, all three strains secreted the enzymes proteinases, chitinases and lipases with a potential to destroy insect tissues. Interestingly, strain ZJLP09 had an earlier invasion time and the highest levels of enzyme activities when compared to the other two strains. These findings have expanded the existing pool of entomopathogenic fungi that infect D. citri and can be potentially used for the management of D. citri populations.

  14. The Distribution of Coumarins and Furanocoumarins in Citrus Species Closely Matches Citrus Phylogeny and Reflects the Organization of Biosynthetic Pathways.

    PubMed

    Dugrand-Judek, Audray; Olry, Alexandre; Hehn, Alain; Costantino, Gilles; Ollitrault, Patrick; Froelicher, Yann; Bourgaud, Frédéric

    2015-01-01

    Citrus plants are able to produce defense compounds such as coumarins and furanocoumarins to cope with herbivorous insects and pathogens. In humans, these chemical compounds are strong photosensitizers and can interact with medications, leading to the "grapefruit juice effect". Removing coumarins and furanocoumarins from food and cosmetics imply additional costs and might alter product quality. Thus, the selection of Citrus cultivars displaying low coumarin and furanocoumarin contents constitutes a valuable alternative. In this study, we performed ultra-performance liquid chromatography coupled with mass spectrometry analyses to determine the contents of these compounds within the peel and the pulp of 61 Citrus species representative of the genetic diversity all Citrus. Generally, Citrus peel contains larger diversity and higher concentrations of coumarin/furanocoumarin than the pulp of the same fruits. According to the chemotypes found in the peel, Citrus species can be separated into 4 groups that correspond to the 4 ancestral taxa (pummelos, mandarins, citrons and papedas) and extended with their respective secondary species descendants. Three of the 4 ancestral taxa (pummelos, citrons and papedas) synthesize high amounts of these compounds, whereas mandarins appear practically devoid of them. Additionally, all ancestral taxa and their hybrids are logically organized according to the coumarin and furanocoumarin pathways described in the literature. This organization allows hypotheses to be drawn regarding the biosynthetic origin of compounds for which the biogenesis remains unresolved. Determining coumarin and furanocoumarin contents is also helpful for hypothesizing the origin of Citrus species for which the phylogeny is presently not firmly established. Finally, this work also notes favorable hybridization schemes that will lead to low coumarin and furanocoumarin contents, and we propose to select mandarins and Ichang papeda as Citrus varieties for use in

  15. The Distribution of Coumarins and Furanocoumarins in Citrus Species Closely Matches Citrus Phylogeny and Reflects the Organization of Biosynthetic Pathways

    PubMed Central

    Dugrand-Judek, Audray; Olry, Alexandre; Hehn, Alain; Costantino, Gilles; Ollitrault, Patrick; Froelicher, Yann; Bourgaud, Frédéric

    2015-01-01

    Citrus plants are able to produce defense compounds such as coumarins and furanocoumarins to cope with herbivorous insects and pathogens. In humans, these chemical compounds are strong photosensitizers and can interact with medications, leading to the “grapefruit juice effect”. Removing coumarins and furanocoumarins from food and cosmetics imply additional costs and might alter product quality. Thus, the selection of Citrus cultivars displaying low coumarin and furanocoumarin contents constitutes a valuable alternative. In this study, we performed ultra-performance liquid chromatography coupled with mass spectrometry analyses to determine the contents of these compounds within the peel and the pulp of 61 Citrus species representative of the genetic diversity all Citrus. Generally, Citrus peel contains larger diversity and higher concentrations of coumarin/furanocoumarin than the pulp of the same fruits. According to the chemotypes found in the peel, Citrus species can be separated into 4 groups that correspond to the 4 ancestral taxa (pummelos, mandarins, citrons and papedas) and extended with their respective secondary species descendants. Three of the 4 ancestral taxa (pummelos, citrons and papedas) synthesize high amounts of these compounds, whereas mandarins appear practically devoid of them. Additionally, all ancestral taxa and their hybrids are logically organized according to the coumarin and furanocoumarin pathways described in the literature. This organization allows hypotheses to be drawn regarding the biosynthetic origin of compounds for which the biogenesis remains unresolved. Determining coumarin and furanocoumarin contents is also helpful for hypothesizing the origin of Citrus species for which the phylogeny is presently not firmly established. Finally, this work also notes favorable hybridization schemes that will lead to low coumarin and furanocoumarin contents, and we propose to select mandarins and Ichang papeda as Citrus varieties for use in

  16. Antioxidant activity of Citrus fruits.

    PubMed

    Zou, Zhuo; Xi, Wanpeng; Hu, Yan; Nie, Chao; Zhou, Zhiqin

    2016-04-01

    Citrus is well-known for its nutrition and health-promotion values. This reputation is derived from the studies on the biological functions of phytochemicals in Citrus fruits and their derived products in the past decades. In recent years, the antioxidant activity of Citrus fruits and their roles in the prevention and treatment of various human chronic and degenerative diseases have attracted more and more attention. Citrus fruits are suggested to be a good source of dietary antioxidants. To have a better understanding of the mechanism underlying the antioxidant activity of Citrus fruits, we reviewed a study on the antioxidant activity of the phytochemicals in Citrus fruits, introduced methods for antioxidant activity evaluation, discussed the factors which influence the antioxidant activity of Citrus fruits, and summarized the underlying mechanism of action. Some suggestions for future study were also presented.

  17. A unique nrdB gene can be used to improve the current HLB pathogen detection system

    USDA-ARS?s Scientific Manuscript database

    Citrus Huanglongbing (HLB), also referred to as yellow shoot disease and citrus greening disease, seriously threatens citrus production worldwide. Currently, the impact of HLB is grave, as there is no cure. A bacterium, “Candidatus Liberibacter asiaticus” (CLas), has been considered as the pathogen ...

  18. Multiplex detection, distribution, and genetic diversity of Hop stunt viroid and Citrus exocortis viroid infecting citrus in Taiwan.

    PubMed

    Lin, Chun-Yi; Wu, Meng-Ling; Shen, Tang-Long; Yeh, Hsin-Hung; Hung, Ting-Hsuan

    2015-02-03

    Two citrus viroids, Citrus exocortis viroid (CEVd) and Hop stunt viroid (HSVd), have been reported and become potential threats to the citrus industry in Taiwan. The distributions and infection rates of two viroids have not been investigated since the two diseases were presented decades ago. The genetic diversities and evolutionary relationships of two viroids also remain unclear in the mix citrus planted region. Multiplex RT-PCR was used to detect the two viroids for the first time in seven main cultivars of citrus. Multiplex real-time RT-PCR quantified the distributions of two viroids in four citrus tissues. Sequence alignment and phylogenetic analysis were performed using the ClustalW and MEGA6 (neighbor-joining with p-distance model), respectively. HSVd was found more prevalent than CEVd (32.2% vs. 30.4%). Both CEVd and HSVd were commonly found simultaneously in the different citrus cultivars (up to 55%). Results of the multiplex quantitative analysis suggested that uneven distributions of both viroids with twig bark as the most appropriate material for studies involving viroid sampling such as quarantine inspection. Sequence alignment against Taiwanese isolates, along with analysis of secondary structure, revealed the existence of 10 and 5 major mutation sites in CEVd and HSVd, respectively. The mutation sites in CEVd were located at both ends of terminal and variability domains, whereas those in HSVd were situated in left terminal and pathogenicity domains. A phylogenetic analysis incorporating worldwide viroid isolates indicated three and two clusters for the Taiwanese isolates of CEVd and HSVd, respectively. Moderately high infection and co-infection rates of two viroids in certain citrus cultivars suggest that different citrus cultivars may play important roles in viroid infection and evolution. These data also demonstrate that two multiplex molecular detection methods developed in the present study provide powerful tools to understand the genetic

  19. Huanglongbing: An overview of a complex pathosystem ravaging the world's citrus.

    PubMed

    da Graça, John V; Douhan, Greg W; Halbert, Susan E; Keremane, Manjunath L; Lee, Richard F; Vidalakis, Georgios; Zhao, Hongwei

    2016-04-01

    Citrus huanglongbing (HLB) has become a major disease and limiting factor of production in citrus areas that have become infected. The destruction to the affected citrus industries has resulted in a tremendous increase to support research that in return has resulted in significant information on both applied and basic knowledge concerning this important disease to the global citrus industry. Recent research indicates the relationship between citrus and the causal agent of HLB is shaped by multiple elements, in which host defense responses may also play an important role. This review is intended to provide an overview of the importance of HLB to a wider audience of plant biologists. Recent advances on host-pathogen interactions, population genetics and vectoring of the causal agent are discussed.

  20. Multiple phytohormone signalling pathways modulate susceptibility of tomato plants to Alternaria alternata f. sp. lycopersici

    PubMed Central

    Jia, Chengguo; Zhang, Liping; Wang, Qiaomei

    2013-01-01

    Three phytohormone molecules – ethylene (ET), jasmonic acid (JA) and salicylic acid (SA) – play key roles in mediating disease response to necrotrophic fungal pathogens. This study investigated the roles of the ET, JA, and SA pathways as well as their crosstalk during the interaction between tomato (Solanum lycopersicum) plants and a necrotrophic fungal pathogen Alternaria alternata f. sp. lycopersici (AAL). Both the ET and JASMONIC ACID INSENSITIVE1 (JAI1) receptor-dependent JA signalling pathways are necessary for susceptibility, while SA response promotes resistance to AAL infection. In addition, the role of JA in susceptibility to AAL is partly dependent on ET biosynthesis and perception, while the SA pathway enhances resistance to AAL and antagonizes the ET response. Based on these results, it is proposed that ET, JA, and SA each on their own can influence the susceptibility of tomato to AAL. Furthermore, the functions of JA and SA in susceptibility to the pathogen are correlated with the enhanced or decreased action of ET, respectively. This study has revealed the functional relationship among the three key hormone pathways in tomato defence against AAL. PMID:23264518

  1. Anastomosis Is Required for Virulence of the Fungal Necrotroph Alternaria brassicicola▿

    PubMed Central

    Craven, Kelly D.; Vélëz, Heriberto; Cho, Yangrae; Lawrence, Christopher B.; Mitchell, Thomas K.

    2008-01-01

    A fungal mycelium is typically composed of radially extending hyphal filaments interconnected by bridges created through anastomoses. These bridges facilitate the dissemination of nutrients, water, and signaling molecules throughout the colony. In this study, we used targeted gene deletion and nitrate utilization mutants of the cruciferous pathogen Alternaria brassicicola and two closely related species to investigate hyphal fusion (anastomosis) and its role in the ability of fungi to cause disease. All eight of the A. brassicicola isolates tested, as well as A. mimicula and A. japonica, were capable of self-fusion, with two isolates of A. brassicicola being capable of non-self-fusion. Disruption of the anastomosis gene homolog (Aso1) in A. brassicicola resulted in both the loss of self-anastomosis and pathogenicity on cabbage. This finding, combined with our discovery that a previously described nonpathogenic A. brassicicola mutant defective for a mitogen-activated protein kinase gene (amk1) also lacked the capacity for self-anastomosis, suggests that self-anastomosis is associated with pathogenicity in A. brassicicola. PMID:18310356

  2. Genomic characterization of the conditionally dispensable chromosome in Alternaria arborescens provides evidence for horizontal gene transfer

    PubMed Central

    2012-01-01

    Background Fungal plant pathogens cause serious agricultural losses worldwide. Alternaria arborescens is a major pathogen of tomato, with its virulence determined by the presence of a conditionally dispensable chromosome (CDC) carrying host-specific toxin genes. Genes encoding these toxins are well-studied, however the genomic content and organization of the CDC is not known. Results To gain a richer understanding of the molecular determinants of virulence and the evolution of pathogenicity, we performed whole genome sequencing of A. arborescens. Here we present the de-novo assembly of the CDC and its predicted gene content. Also presented is hybridization data validating the CDC assembly. Predicted genes were functionally annotated through BLAST. Gene ontology terms were assigned, and conserved domains were identified. Differences in nucleotide usage were found between CDC genes and those on the essential chromosome (EC), including GC3-content, codon usage bias, and repeat region load. Genes carrying PKS and NRPS domains were identified in clusters on the CDC and evidence supporting the origin of the CDC through horizontal transfer from an unrelated fungus was found. Conclusions We provide evidence supporting the hypothesis that the CDC in A. arborescens was acquired through horizontal transfer, likely from an unrelated fungus. We also identified several predicted CDC genes under positive selection that may serve as candidate virulence factors. PMID:22559316

  3. Sesquiterpene emissions from Alternaria alternata and Fusarium oxysporum: Effects of age, nutrient availability, and co-cultivation

    PubMed Central

    Weikl, Fabian; Ghirardo, Andrea; Schnitzler, Jörg-Peter; Pritsch, Karin

    2016-01-01

    Alternaria alternata is one of the most studied fungi to date because of its impact on human life – from plant pathogenicity to allergenicity. However, its sesquiterpene emissions have not been systematically explored. Alternaria regularly co-occurs with Fusarium fungi, which are common plant pathogens, on withering plants. We analyzed the diversity and determined the absolute quantities of volatile organic compounds (VOCs) in the headspace above mycelial cultures of A. alternata and Fusarium oxysporum under different conditions (nutrient rich and poor, single cultures and co-cultivation) and at different mycelial ages. Using stir bar sorptive extraction and gas chromatography–mass spectrometry, we observed A. alternata to strongly emit sesquiterpenes, particularly during the early growth stages, while emissions from F. oxysporum consistently remained comparatively low. The emission profile characterizing A. alternata comprised over 20 sesquiterpenes with few effects from nutrient quality and age on the overall emission profile. Co-cultivation with F. oxysporum resulted in reduced amounts of VOCs emitted from A. alternata although its profile remained similar. Both fungi showed distinct emission profiles, rendering them suitable biomarkers for growth-detection of their phylotype in ambient air. The study highlights the importance of thorough and quantitative evaluations of fungal emissions of volatile infochemicals such as sesquiterpenes. PMID:26915756

  4. Sesquiterpene emissions from Alternaria alternata and Fusarium oxysporum: Effects of age, nutrient availability, and co-cultivation.

    PubMed

    Weikl, Fabian; Ghirardo, Andrea; Schnitzler, Jörg-Peter; Pritsch, Karin

    2016-02-26

    Alternaria alternata is one of the most studied fungi to date because of its impact on human life - from plant pathogenicity to allergenicity. However, its sesquiterpene emissions have not been systematically explored. Alternaria regularly co-occurs with Fusarium fungi, which are common plant pathogens, on withering plants. We analyzed the diversity and determined the absolute quantities of volatile organic compounds (VOCs) in the headspace above mycelial cultures of A. alternata and Fusarium oxysporum under different conditions (nutrient rich and poor, single cultures and co-cultivation) and at different mycelial ages. Using stir bar sorptive extraction and gas chromatography-mass spectrometry, we observed A. alternata to strongly emit sesquiterpenes, particularly during the early growth stages, while emissions from F. oxysporum consistently remained comparatively low. The emission profile characterizing A. alternata comprised over 20 sesquiterpenes with few effects from nutrient quality and age on the overall emission profile. Co-cultivation with F. oxysporum resulted in reduced amounts of VOCs emitted from A. alternata although its profile remained similar. Both fungi showed distinct emission profiles, rendering them suitable biomarkers for growth-detection of their phylotype in ambient air. The study highlights the importance of thorough and quantitative evaluations of fungal emissions of volatile infochemicals such as sesquiterpenes.

  5. Functional analysis of a multicopy host-selective ACT-toxin biosynthesis gene in the tangerine pathotype of Alternaria alternata using RNA silencing.

    PubMed

    Miyamoto, Y; Masunaka, A; Tsuge, T; Yamamoto, M; Ohtani, K; Fukumoto, T; Gomi, K; Peever, T L; Akimitsu, K

    2008-12-01

    Alternaria brown spot, caused by the tangerine pathotype of Alternaria alternata, is a serious disease of commercially important tangerines and their hybrids. The pathogen produces host-selective ACT toxin, and several genes (named ACTT) responsible for ACT-toxin biosynthesis have been identified. These genes have many paralogs, which are clustered on a small, conditionally dispensable chromosome, making it difficult to disrupt entire functional copies of ACTT genes using homologous recombination-mediated gene disruption. To overcome this problem, we attempted to use RNA silencing, which has never been employed in Alternaria spp., to knock down the functional copies of one ACTT gene with a single silencing event. ACTT2, which encodes a putative hydrolase and is present in multiple copies in the genome, was silenced by transforming the fungus with a plasmid construct expressing hairpin ACTT2 RNAs. The ACTT2 RNA-silenced transformant (S-7-24-2) completely lost ACTT2 transcripts and ACT-toxin production as well as pathogenicity. These results indicated that RNA silencing may be a useful technique for studying the role of ACTT genes responsible for host-selective toxin biosynthesis in A. alternata. Further, this technique may be broadly applicable to the analysis of many genes present in multiple copies in fungal genomes that are difficult to analyze using recombination-mediated knockdowns.

  6. Citrus tissue culture employing vegetative explants.

    PubMed

    Chaturvedi, H C; Singh, S K; Sharma, A K; Agnihotri, S

    2001-11-01

    culture of shoots of C grandis without loss of regeneration capacity during 31 years, observed so far, are some other significant results. Plants of C. aurantifolia and C. sinensis raised from shoot meristem and micrografting were grown in a nethouse and those from nodal stem segments in the field along with the in vitro-raised plants of rootstocks, namely, C. jambhiri, C. karna and C. limonia. All the plants showed normal healthy growth. Significantly enough, the meristem regenerated plants of C. aurantifolia attained the reproductive phase just in 1 year of transplantation to soil similar to those raised from nodal stem segments of mature trees, which also produced normal fruits in the subsequent year while growing under field conditions. Thus, a significant fundamental concept of a maturity factor, carried over through as small a shoot meristem as 200 microm in length to cloned plants has been demonstrated. The concept is of far-reaching significance in citrus industry besides production of pathogen-free orchards.

  7. Emerging Fusarium and Alternaria Mycotoxins: Occurrence, Toxicity and Toxicokinetics

    PubMed Central

    Croubels, Siska; Devreese, Mathias; Antonissen, Gunther

    2017-01-01

    Emerging Fusarium and Alternaria mycotoxins gain more and more interest due to their frequent contamination of food and feed, although in vivo toxicity and toxicokinetic data are limited. Whereas the Fusarium mycotoxins beauvericin, moniliformin and enniatins particularly contaminate grain and grain-based products, Alternaria mycotoxins are also detected in fruits, vegetables and wines. Although contamination levels are usually low (µg/kg range), higher contamination levels of enniatins and tenuazonic acid may occasionally occur. In vitro studies suggest genotoxic effects of enniatins A, A1 and B1, beauvericin, moniliformin, alternariol, alternariol monomethyl ether, altertoxins and stemphyltoxin-III. Furthermore, in vitro studies suggest immunomodulating effects of most emerging toxins and a reproductive health hazard of alternariol, beauvericin and enniatin B. More in vivo toxicity data on the individual and combined effects of these contaminants on reproductive and immune system in both humans and animals is needed to update the risk evaluation by the European Food Safety Authority. Taking into account new occurrence data for tenuazonic acid, the complete oral bioavailability, the low total body clearance in pigs and broiler chickens and the limited toxicity data, a health risk cannot be completely excluded. Besides, some less known Alternaria toxins, especially the genotoxic altertoxins and stemphyltoxin III, should be incorporated in risk evaluation as well. PMID:28718805

  8. Emerging Fusarium and Alternaria Mycotoxins: Occurrence, Toxicity and Toxicokinetics.

    PubMed

    Fraeyman, Sophie; Croubels, Siska; Devreese, Mathias; Antonissen, Gunther

    2017-07-18

    Emerging Fusarium and Alternaria mycotoxins gain more and more interest due to their frequent contamination of food and feed, although in vivo toxicity and toxicokinetic data are limited. Whereas the Fusarium mycotoxins beauvericin, moniliformin and enniatins particularly contaminate grain and grain-based products, Alternaria mycotoxins are also detected in fruits, vegetables and wines. Although contamination levels are usually low (µg/kg range), higher contamination levels of enniatins and tenuazonic acid may occasionally occur. In vitro studies suggest genotoxic effects of enniatins A, A1 and B1, beauvericin, moniliformin, alternariol, alternariol monomethyl ether, altertoxins and stemphyltoxin-III. Furthermore, in vitro studies suggest immunomodulating effects of most emerging toxins and a reproductive health hazard of alternariol, beauvericin and enniatin B. More in vivo toxicity data on the individual and combined effects of these contaminants on reproductive and immune system in both humans and animals is needed to update the risk evaluation by the European Food Safety Authority. Taking into account new occurrence data for tenuazonic acid, the complete oral bioavailability, the low total body clearance in pigs and broiler chickens and the limited toxicity data, a health risk cannot be completely excluded. Besides, some less known Alternaria toxins, especially the genotoxic altertoxins and stemphyltoxin III, should be incorporated in risk evaluation as well.

  9. Comparison of enzyme immunoassay–based assays for environmental Alternaria alternata

    PubMed Central

    Barnes, Charles; Portnoy, Jay; Sever, Michelle; Arbes, Samuel; Vaughn, Ben; Zeldin, Darryl C.

    2007-01-01

    Background Alternaria alternata–derived allergenic materials are causes of human disease. Several immunoassays exist to quantify these materials. Objective To compare methods for evaluating Alternaria content. Methods Four methods, including 1 monoclonal antibody (MAb)–based assay specific for recombinant Alt a 1, 1 MAb-based assay for chromatographically purified Alt a 1, 1 polyclonal antibody (PAb)–based assay for chromatographically purified Alt a 1, and 1 PAb-based assay for whole Alternaria extract, were evaluated. Environmental samples collected as part of the National Survey of Lead and Allergens in Housing were examined. Alternaria spore counts were determined in dust by observation. Results The MAb-based assay for recombinant Alt a 1 detected Alternaria in few samples (25%); the PAb-based assay for whole Alternaria proteins detected antigen in 97% of the samples. The PAb- and MAb-based assays for purified Alt a 1 detected antigen in 100% of the samples. There was a significant positive correlation between the 2 assays directed against purified Alt a 1. There was a positive correlation between the PAb-based assay for whole Alternaria and the PAb-based assay for Alt a 1. Nearly all the dust samples contained Alternaria spores, and there was a strong positive correlation between counts and all assays. Conclusion Because of the multifaceted nature of Alternaria, the disparities between methods for quantifying Alternaria, the cross-reactivity between fungal allergens, and the documented genetic promiscuity of this fungus, enzyme immunoassays using PAbs against a range of Alternaria proteins will probably produce the most reliable estimation of overall Alternaria exposure in house dust. PMID:17042141

  10. Overexpression of a Modified Plant Thionin Enhances Disease Resistance to Citrus Canker and Huanglongbing (HLB)

    PubMed Central

    Hao, Guixia; Stover, Ed; Gupta, Goutam

    2016-01-01

    Huanglongbing (HLB or citrus greening disease) caused by Candidatus Liberibacter asiaticus (Las) is a great threat to the US citrus industry. There are no proven strategies to eliminate HLB disease and no cultivar has been identified with strong HLB resistance. Citrus canker is also an economically important disease associated with a bacterial pathogen (Xanthomonas citri). In this study, we characterized endogenous citrus thionins and investigated their expression in different citrus tissues. Since no HLB-resistant citrus cultivars have been identified, we attempted to develop citrus resistant to both HLB and citrus canker through overexpression of a modified plant thionin. To improve effectiveness for disease resistance, we modified and synthesized the sequence encoding a plant thionin and cloned into the binary vector pBinPlus/ARS. The construct was then introduced into Agrobacterium strain EHA105 for citrus transformation. Transgenic Carrizo plants expressing the modified plant thionin were generated by Agrobacterium-mediated transformation. Successful transformation and transgene gene expression was confirmed by molecular analysis. Transgenic Carrizo plants expressing the modified thionin gene were challenged with X. citri 3213 at a range of concentrations, and a significant reduction in canker symptoms and a decrease in bacterial growth were demonstrated compared to nontransgenic plants. Furthermore, the transgenic citrus plants were challenged with HLB via graft inoculation. Our results showed significant Las titer reduction in roots of transgenic Carrizo compared with control plants and reduced scion Las titer 12 months after graft inoculation. These data provide promise for engineering citrus disease resistance against HLB and canker. PMID:27499757

  11. Daily variations of Alternaria spores in the city of Murcia (semi-arid southeastern Spain)

    NASA Astrophysics Data System (ADS)

    Munuera Giner, M.; Carrión García, J. S.

    1995-12-01

    Annual variations in the abundance of Alternaria spores were related to the length of the spore period for data from Murcia (southeastern Spain). To understand the relationship between the number of spores and climatic factors, Alternaria spore counts for March 1993 to February 1994 were examined by means of correlation and regression analyses with fourteen different weather parameters. The results indicated that there was a tendency for Alternaria spore concentrations to increase with increases in temperature, wind speed and hours of sunshine. Negative correlations were observed with air pressure, wind direction and humidity. Theoretical curves for Alternaria spore counts are given in relation to temperatures during the period studied.

  12. Innate Type-2 Response to Alternaria Extract Enhances Ryegrass-induced Lung Inflammation

    PubMed Central

    Kim, Hee-Kyoo; Lund, Sean; Baum, Rachel; Rosenthal, Peter; Khorram, Naseem; Doherty, Taylor A.

    2014-01-01

    Background Exposure to the fungal allergen Alternaria alternata as well as ryegrass pollen has been implicated in severe asthma symptoms during thunderstorms. We have previously shown that Alternaria extract induces innate type 2 lung inflammation in mice. We hypothesized that the innate eosinophilic response to Alternaria extract may enhance lung inflammation induced by ryegrass. Methods Mice were sensitized to ryegrass allergen and administered a single challenge with Alternaria alternata extract before or after final ryegrass challenges. Levels of BAL eosinophils, neutrophils, Th2 cells, innate lymphoid cells (ILC2), IL-5 and IL-13 as well as inflammation and mucus were assessed. Results Mice receiving ryegrass sensitization and challenge developed an eosinophilic lung response. A single challenge with Alternaria extract given 3 days before or 3 days after ryegrass challenges resulted in increased eosinophils, peribronchial inflammation and mucus production in the airway compared with ryegrass only challenges. Type 2 innate lymphoid cell (ILC2) and Th2 cell recruitment to the airway was increased after Alternaria extract exposure in ryegrass challenged mice. Innate challenges with Alternaria extract induced BAL eosinophilia, Th2 cell recruitment as well as ILC2 expansion and proliferation. Conclusions A single exposure of Alternaria extract in ryegrass sensitized and challenged mice enhances the type-2 lung inflammatory response including airway eosinophilia, peribronchial infiltrate, and mucus production possibly through Th2 cell recruitment and ILC2 expansion. If translated to humans, exposures to both grass pollen and Alternaria may be a potential cause of thunderstorm-related asthma. PMID:24296722

  13. Air-borne fungi in the air of Barcelona (Spain). II. The genus alternaria.

    PubMed

    Calvo, M A; Guarro, J; Suarez, G; Ramírez, C

    1979-12-28

    The genus Alternaria Nees formed one of the more important components of the fungus population of the air at Barcelone (Spain), during a two-year study, from February 1976 through January 1978. Results were based only on studied of colonies obtained by gravity-exposed plates. The occurrence of this genus was greatly affected by climatic conditions. In general. however, Alternaria appeared to prefer warmer weather. A total of six species of Alternaria was identified, of which, Alternaria alternata (Fries) Keissler was by far the commonest, representing 71.3% of all colonies.

  14. Transmission efficiency of Xylella fastidiosa by sharpshooters (Hemiptera: Cicadellidae) in coffee and citrus.

    PubMed

    Marucci, Rosangela C; Lopes, João R S; Cavichioli, Rodney R

    2008-08-01

    Xylella fastidiosa (Wells, Raju, Hung, Weisburg, Mandelco-Paul, and Brenner) is a bacterial pathogen transmitted by several sharpshooters in two tribes of Cicadellinae (Proconiini and Cicadellini). Here, we compared the transmission efficiency of X. fastidiosa in coffee (Coffea arabica L.) and citrus [Citrus sinensis (L.) Osbeck] by Cicadellini [Bucephalogonia xanthophis (Berg) and Dilobopterus costalimai Young] and Proconiini [Homalodisca ignorata Melichar and Oncometopia facialis (Signoret)] sharpshooters that occur in both crops. At different seasons, healthy adults of each species were submitted to a 48-h acquisition access period on citrus or coffee source plants infected with X. fastidiosa isolates that cause Citrus variegated chlorosis (CVC) and Coffee leaf scorch (CLS), respectively, and then confined on healthy seedlings of the corresponding host plant for a 48-h inoculation access period. No significant effect of inoculation season was observed when comparing infection rates of citrus or coffee plants inoculated by vectors at different times of the year. In citrus, the transmission rate by single insects was significantly higher for H. ignorata (30%) in relation to B. xanthophis (5%) and O. facialis (1.1%), but there was no difference among vector species in coffee, whose transmission rates ranged from 1.2 to 7.2%. Comparing host plants, H. ignorata was more effective in transmitting X. fastidiosa to citrus (30%) in relation to coffee (2.2%), whereas the other vectors transmitted the bacterium to both hosts with similar efficiencies. Despite these variations, vector efficiency in coffee and citrus is lower than that reported in other hosts.

  15. Antimicrobial activity of acid-hydrolyzed Citrus unshiu peel extract in milk.

    PubMed

    Min, Keun Young; Kim, Hyun Jung; Lee, Kyoung Ah; Kim, Kee-Tae; Paik, Hyun-Dong

    2014-01-01

    Citrus fruit (Citrus unshiu) peels were extracted with hot water and then acid-hydrolyzed using hydrochloric acid. Antimicrobial activities of acid-hydrolyzed Citrus unshiu peel extract were evaluated against pathogenic bacteria, including Bacillus cereus, Staphylococcus aureus, and Listeria monocytogenes. Antilisterial effect was also determined by adding extracts at 1, 2, and 4% to whole, low-fat, and skim milk. The cell numbers of B. cereus, Staph. aureus, and L. monocytogenes cultures treated with acid-hydrolyzed extract for 12h at 35°C were reduced from about 8log cfu/mL to <1log cfu/mL. Bacillus cereus was more sensitive to acid-hydrolyzed Citrus unshiu peel extract than were the other bacteria. The addition of 4% acid-hydrolyzed Citrus unshiu extracts to all types of milk inhibited the growth of L. monocytogenes within 1d of storage at 4°C. The results indicated that Citrus unshiu peel extracts, after acid hydrolysis, effectively inhibited the growth of pathogenic bacteria. These findings indicate that acid hydrolysis of Citrus unshiu peel facilitates its use as a natural antimicrobial agent for food products.

  16. Ex Vivo Application of Secreted Metabolites Produced by Soil-Inhabiting Bacillus spp. Efficiently Controls Foliar Diseases Caused by Alternaria spp.

    PubMed Central

    El-Sayed, Ashraf S. A.; Patel, Jaimin S.; Green, Kari B.; Ali, Mohammad; Brennan, Mary; Norman, David

    2015-01-01

    Bacterial biological control agents (BCAs) are largely used as live products to control plant pathogens. However, due to variable environmental and ecological factors, live BCAs usually fail to produce desirable results against foliar pathogens. In this study, we investigated the potential of cell-free culture filtrates of 12 different bacterial BCAs isolated from flower beds for controlling foliar diseases caused by Alternaria spp. In vitro studies showed that culture filtrates from two isolates belonging to Bacillus subtilis and Bacillus amyloliquefaciens displayed strong efficacy and potencies against Alternaria spp. The antimicrobial activity of the culture filtrate of these two biological control agents was effective over a wider range of pH (3.0 to 9.0) and was not affected by autoclaving or proteolysis. Comparative liquid chromatography-mass spectrometry (LC-MS) analyses showed that a complex mixture of cyclic lipopeptides, primarily of the fengycin A and fengycin B families, was significantly higher in these two BCAs than inactive Bacillus spp. Interaction studies with mixtures of culture filtrates of these two species revealed additive activity, suggesting that they produce similar products, which was confirmed by LC-tandem MS analyses. In in planta pre- and postinoculation trials, foliar application of culture filtrates of B. subtilis reduced lesion sizes and lesion frequencies caused by Alternaria alternata by 68 to 81%. Taken together, our studies suggest that instead of live bacteria, culture filtrates of B. subtilis and B. amyloliquefaciens can be applied either individually or in combination for controlling foliar diseases caused by Alternaria species. PMID:26519395

  17. Partial Resistance of Carrot to Alternaria dauci Correlates with In Vitro Cultured Carrot Cell Resistance to Fungal Exudates

    PubMed Central

    Voisine, Linda; Gatto, Julia; Hélesbeux, Jean-Jacques; Séraphin, Denis; Peña-Rodriguez, Luis M.; Richomme, Pascal; Boedo, Cora; Yovanopoulos, Claire; Gyomlai, Melvina; Briard, Mathilde; Simoneau, Philippe; Poupard, Pascal; Berruyer, Romain

    2014-01-01

    Although different mechanisms have been proposed in the recent years, plant pathogen partial resistance is still poorly understood. Components of the chemical warfare, including the production of plant defense compounds and plant resistance to pathogen-produced toxins, are likely to play a role. Toxins are indeed recognized as important determinants of pathogenicity in necrotrophic fungi. Partial resistance based on quantitative resistance loci and linked to a pathogen-produced toxin has never been fully described. We tested this hypothesis using the Alternaria dauci – carrot pathosystem. Alternaria dauci, causing carrot leaf blight, is a necrotrophic fungus known to produce zinniol, a compound described as a non-host selective toxin. Embryogenic cellular cultures from carrot genotypes varying in resistance against A. dauci were confronted with zinniol at different concentrations or to fungal exudates (raw, organic or aqueous extracts). The plant response was analyzed through the measurement of cytoplasmic esterase activity, as a marker of cell viability, and the differentiation of somatic embryos in cellular cultures. A differential response to toxicity was demonstrated between susceptible and partially resistant genotypes, with a good correlation noted between the resistance to the fungus at the whole plant level and resistance at the cellular level to fungal exudates from raw and organic extracts. No toxic reaction of embryogenic cultures was observed after treatment with the aqueous extract or zinniol used at physiological concentration. Moreover, we did not detect zinniol in toxic fungal extracts by UHPLC analysis. These results suggest that strong phytotoxic compounds are present in the organic extract and remain to be characterized. Our results clearly show that carrot tolerance to A. dauci toxins is one component of its partial resistance. PMID:24983469

  18. Progress on strain differentiation of Citrus tristeza virus and its application to the epidemiology of citrus tristeza disease.

    PubMed

    Niblett, C L; Genc, H; Cevik, B; Halbert, S; Brown, L; Nolasco, G; Bonacalza, B; Manjunath, K L; Febres, V J; Pappu, H R; Lee, R F

    2000-11-01

    Citrus tristeza virus (CTV) occurs in most citrus producing regions of the world, and it is the most serious viral pathogen of citrus. With the recent establishment of the brown citrus aphid, Toxoptera citricida, its most efficient vector, on Madeira Island (Portugal) and in Florida (USA) and the countries of the Caribbean Basin, the impact of CTV is likely to increase in these regions. Since there are many strains of CTV and CTV infections frequently occur as mixtures of several strains, it is necessary to be able to distinguish the strains for regulatory purposes, disease management and epidemiology. We describe the evolution of techniques developed to detect CTV and to differentiate the individual strains, and present the results of tests using these latest methods on CTV isolates from mainland Portugal, Madeira Island and Florida. Mild and decline-inducing strains of CTV were detected in mainland Portugal and mild, decline-inducing and severe stem pitting strains on Madeira Island. In Florida we demonstrated the presence of infections that reacted with probes made against stem pitting strains not previously detected there. It is concluded that CTV presents a significant threat to citrus production in mainland Portugal, on Madeira Island and in the neighbouring countries of the Mediterranean Basin, as well as in Florida, elsewhere in the USA and throughout the Caribbean Basin, especially following the widespread establishment of T. citricida throughout the region.

  19. A polyphasic approach to the taxonomy of the Alternaria infectoria species-group.

    PubMed

    Andersen, Birgitte; Sørensen, Jens Laurids; Nielsen, Kristian Fog; Gerrits van den Ende, Bert; de Hoog, Sybren

    2009-09-01

    . malorum strains do not belong in Alternaria, but in Chalastospora, however, as several distinct species. Splits Tree alignment of gpd sequences of 38 strains belonging to the A. infectoria species-group indicates that only three strains showed signs of recombination, while the remaining strains appeared to be clonal. Long term incubation at 7 degrees C in the dark showed that 12 out of 33 tested strains from the A. infectoria species-group were able to produce proascomata in axenic culture, but with no mature ascospores after 6 months. These findings suggest that Lewia/A. infectoria species-group must, at least in part, be homothallic. The results presented in this study show that ITS, tef-1alpha and gpd do not reflect ecology, secondary metabolism or morphology of the A. infectoria species-group and that molecular cladification and phylogeny cannot predict pathogenicity, host specificity or mycotoxin production.

  20. Localization of melanin in conidia of Alternaria alternata using phage display antibodies.

    PubMed

    Carzaniga, Raffaella; Fiocco, Daniela; Bowyer, Paul; O'Connell, Richard J

    2002-03-01

    Melanins derived from 1,8-dihydroxynaphthalene (DHN) are important for the pathogenicity and survival of fungi causing disease in both plants and animals. However, precise information on their location within fungal cell walls is lacking. To obtain antibodies for the immunocytochemical localization of melanin, 83 phage antibodies binding to 1,8-DHN were selected from a naive semisynthetic single-chain Fv (scFv) phage display library. Sequence analysis of the heavy chain binding domains of 17 antibodies showed a high frequency of positively charged amino acids. One antibody, designated M1, was characterized in detail. M1 bound specifically to 1,8-DHN in competitive inhibition enzyme-linked immunosorbent assays, showing no cross-reaction with nine structurally related phenolic compounds. Epitope recognition required two hydroxyl groups in a 1,8 configuration. M1 also bound to naturally occurring melanin isolated from mycelia of Alternaria alternata, suggesting that epitopes remain accessible in polymerized melanin. Transmission electron microscopy-immunogold labeling, using M1 in the form of soluble scFv fragments, showed that melanin was located in the septa and outer (primary) walls of wild-type A. alternata conidia, but not those of an albino mutant, AKT88-1. The M1 antibody provides a new tool for detecting melanized pathogens in plant and animal tissues and for precisely mapping the distribution of the polymer within spores, appressoria, and hyphae.

  1. Zinc triggers signaling mechanisms and defense responses promoting resistance to Alternaria brassicicola in Arabidopsis thaliana.

    PubMed

    Martos, Soledad; Gallego, Berta; Cabot, Catalina; Llugany, Mercè; Barceló, Juan; Poschenrieder, Charlotte

    2016-08-01

    According to the elemental defense hypothesis the accumulation of trace elements by plants may substitute for organic defenses, while the joint effects hypothesis proposes that trace elements and organic defenses can have additive or synergistic effects against pathogens or herbivores. To evaluate these hypotheses the response of the pathosystem Alternaria brassicicola-Arabidopsis thaliana to control (2μM) and surplus (12μM) Zn was evaluated using the camalexin deficient mutant pad3-1 and mtp1-1, a mutant with impaired Zn vacuolar storage, along with the corresponding wildtypes. In vitro, a 50% inhibition of fungal growth was achieved by 440μM Zn. A. thaliana leaves could accumulate equivalent concentrations without harm. In fact, surplus Zn enhanced the resistance of A. thaliana to fungal attack in Columbia (Col-0), Wassilewskija (WS), and mtp1-1. However, surplus Zn was unable to protect pad3-1 demonstrating that Zn cannot substitute for camalexin, the main organic defense in A. thaliana. High, non phytotoxic leaf Zn concentrations enhanced the resistance to A. brassicicola of A. thaliana genotypes able to produce camalexin. This was mainly due to Zn-induced enhancement of the JA/ETH signaling pathway leading to enhanced PAD3 expression. These results support the joint effects hypothesis and highlight the importance of adequate Zn supply for reinforced pathogen resistance.

  2. Population genetic analysis reveals cryptic sex in the phytopathogenic fungus Alternaria alternata

    PubMed Central

    Meng, Jing-Wen; Zhu, Wen; He, Meng-Han; Wu, E-Jiao; Duan, Guo-Hua; Xie, Ye-Kun; Jin, Yu-Jia; Yang, Li-Na; Shang, Li-Ping; Zhan, Jiasui

    2015-01-01

    Reproductive mode can impact population genetic dynamics and evolutionary landscape of plant pathogens as well as on disease epidemiology and management. In this study, we monitored the spatial dynamics and mating type idiomorphs in ~700 Alternaria alternata isolates sampled from the main potato production areas in China to infer the mating system of potato early blight. Consistent with the expectation of asexual species, identical genotypes were recovered from different locations separated by hundreds of kilometers of geographic distance and spanned across many years. However, high genotype diversity, equal MAT1-1 and MAT1-2 frequencies within and among populations, no genetic differentiation and phylogenetic association between two mating types, combined with random association amongst neutral markers in some field populations, suggested that sexual reproduction may also play an important role in the epidemics and evolution of the pathogen in at least half of the populations assayed despite the fact that no teleomorphs have been observed yet naturally or artificially. Our results indicated that A. alternata may adopt an epidemic mode of reproduction by combining many cycles of asexual propagation with fewer cycles of sexual reproduction, facilitating its adaptation to changing environments and making the disease management on potato fields even more difficult. PMID:26666175

  3. 7 CFR 301.76-2 - Regulated articles for Asian citrus psyllid and citrus greening.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 5 2014-01-01 2014-01-01 false Regulated articles for Asian citrus psyllid and citrus...) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE DOMESTIC QUARANTINE NOTICES Citrus Greening and Asian Citrus Psyllid § 301.76-2 Regulated articles for Asian citrus psyllid and...

  4. 7 CFR 301.76-2 - Regulated articles for Asian citrus psyllid and citrus greening.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 5 2013-01-01 2013-01-01 false Regulated articles for Asian citrus psyllid and citrus...) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE DOMESTIC QUARANTINE NOTICES Citrus Greening and Asian Citrus Psyllid § 301.76-2 Regulated articles for Asian citrus psyllid and...

  5. 7 CFR 301.76-2 - Regulated articles for Asian citrus psyllid and citrus greening.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 5 2012-01-01 2012-01-01 false Regulated articles for Asian citrus psyllid and citrus...) ANIMAL AND PLANT HEALTH INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE DOMESTIC QUARANTINE NOTICES Citrus Greening and Asian Citrus Psyllid § 301.76-2 Regulated articles for Asian citrus psyllid and...

  6. Agreement: Citrus College Faculty Association and Citrus Community College District.

    ERIC Educational Resources Information Center

    Citrus Community Coll. District, Glendora, CA.

    The collective bargaining agreement between the Citrus Community College District Board of Trustees and the Citrus College Faculty/California Teachers Association/National Education Association is presented. This contract, covering the period from July 1988 through June 1990, deals with the following topics: bargaining agent recognition and…

  7. Citrus Huanglongbing tolerance in Australian Citrus Relatives, Microcitrus and Eremocirus

    USDA-ARS?s Scientific Manuscript database

    Tolerance, or resistance to citrus huanglongbing will be important as a long term solution for this disease. In a field trial conducted with over 1000 plants belonging to different genera in the sub-family Aurantioideae, we observed field tolerance in many Australian citrus relatives. To confirm the...

  8. Agreement: Citrus College Faculty Association and Citrus Community College District.

    ERIC Educational Resources Information Center

    Citrus Community Coll. District, Glendora, CA.

    The collective bargaining agreement between the Citrus Community College District Board of Trustees and the Citrus College Faculty/California Teachers Association/National Education Association is presented. This contract, covering the period from July 1988 through June 1990, deals with the following topics: bargaining agent recognition and…

  9. Development of multiplex polymerase chain reaction assay for simultaneous detection of clostero-, badna- and mandari-viruses along with huanglongbing bacterium in citrus trees.

    PubMed

    Meena, Ram Prasnna; Baranwal, V K

    2016-09-01

    Citrus trees harbor a large number of viral and bacterial pathogens. Citrus yellow vein clearing virus (CYVCV), Indian citrus ringspot virus (ICRSV), Citrus yellow mosaic virus (CYMV), Citrus tristeza virus (CTV) and a bacterium, Candidatus Liberibacter asiaticus (CLa) associated with huanglongbing (HLB) disease, the most prevalent pathogens in citrus orchards of different regions in India and are responsible for debilitating citriculture. For detection of these viral and bacterial pathogens a quick, sensitive and cost effective detection method is required. With this objective a multiplex polymerase chain reaction (mPCR) assay was developed for simultaneous detection of four viruses and a bacterium in citrus. Several sets of primers were designed for each virus based on the retrieved reference sequences from the GenBank. A primer pair published previously was used for greening bacterium. Each pair of primers was evaluated for their sensitivity and differentiation by simplex and mPCR. The constant amplified products were identified on the basis of molecular size in mPCR and were compared with standard PCR. The amplicons were cloned and results were confirmed with sequencing analysis. The mPCR assay was validated using naturally infected field samples for one or more citrus viruses and the huanglongbing bacterium. The mPCR assay developed here will aid in the production of virus free planting materials and rapid indexing for certification of citrus budwood programme. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Metabolic Interplay between the Asian Citrus Psyllid and Its Profftella Symbiont: An Achilles' Heel of the Citrus Greening Insect Vector.

    PubMed

    Ramsey, John S; Johnson, Richard S; Hoki, Jason S; Kruse, Angela; Mahoney, Jaclyn; Hilf, Mark E; Hunter, Wayne B; Hall, David G; Schroeder, Frank C; MacCoss, Michael J; Cilia, Michelle

    2015-01-01

    'Candidatus Liberibacter asiaticus' (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid. Interactions among D. citri and its microbial endosymbionts, including 'Candidatus Profftella armatura', are likely to impact transmission of CLas. We used quantitative mass spectrometry to compare the proteomes of CLas(+) and CLas(-) populations of D. citri, and found that proteins involved in polyketide biosynthesis by the endosymbiont Profftella were up-regulated in CLas(+) insects. Mass spectrometry analysis of the Profftella polyketide diaphorin in D. citri metabolite extracts revealed the presence of a novel diaphorin-related polyketide and the ratio of these two polyketides was changed in CLas(+) insects. Insect proteins differentially expressed between CLas(+) and CLas(-) D. citri included defense and immunity proteins, proteins involved in energy storage and utilization, and proteins involved in endocytosis, cellular adhesion, and cytoskeletal remodeling which are associated with microbial invasion of host cells. Insight into the metabolic interdependence between the insect vector, its endosymbionts, and the citrus greening pathogen reveals novel opportunities for control of this disease, which is currently having a devastating impact on citrus production worldwide.

  11. Metabolic Interplay between the Asian Citrus Psyllid and Its Profftella Symbiont: An Achilles’ Heel of the Citrus Greening Insect Vector

    PubMed Central

    Ramsey, John S.; Johnson, Richard S.; Hoki, Jason S.; Kruse, Angela; Mahoney, Jaclyn; Hilf, Mark E.; Hunter, Wayne B.; Hall, David G.; Schroeder, Frank C.; MacCoss, Michael J.; Cilia, Michelle

    2015-01-01

    ‘Candidatus Liberibacter asiaticus’ (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid. Interactions among D. citri and its microbial endosymbionts, including ‘Candidatus Profftella armatura’, are likely to impact transmission of CLas. We used quantitative mass spectrometry to compare the proteomes of CLas(+) and CLas(-) populations of D. citri, and found that proteins involved in polyketide biosynthesis by the endosymbiont Profftella were up-regulated in CLas(+) insects. Mass spectrometry analysis of the Profftella polyketide diaphorin in D. citri metabolite extracts revealed the presence of a novel diaphorin-related polyketide and the ratio of these two polyketides was changed in CLas(+) insects. Insect proteins differentially expressed between CLas(+) and CLas(-) D. citri included defense and immunity proteins, proteins involved in energy storage and utilization, and proteins involved in endocytosis, cellular adhesion, and cytoskeletal remodeling which are associated with microbial invasion of host cells. Insight into the metabolic interdependence between the insect vector, its endosymbionts, and the citrus greening pathogen reveals novel opportunities for control of this disease, which is currently having a devastating impact on citrus production worldwide. PMID:26580079

  12. Characterization of Citrus sinensis transcription factors closely associated with the non-host response to Xanthomonas campestris pv. vesicatoria.

    PubMed

    Daurelio, Lucas D; Romero, María S; Petrocelli, Silvana; Merelo, Paz; Cortadi, Adriana A; Talón, Manuel; Tadeo, Francisco R; Orellano, Elena G

    2013-07-01

    Plants, when exposed to certain pathogens, may display a form of genotype-independent resistance, known as non-host response. In this study, the response of Citrus sinensis (sweet orange) leaves to Xanthomonas campestris pv. vesicatoria (Xcv), a pepper and tomato pathogenic bacterium, was analyzed through biochemical assays and cDNA microarray hybridization and compared with Asiatic citrus canker infection caused by Xanthomonas citri subsp. citri. Citrus leaves exposed to the non-host bacterium Xcv showed hypersensitive response (HR) symptoms (cell death), a defense mechanism common in plants but poorly understood in citrus. The HR response was accompanied by differentially expressed genes that are associated with biotic stress and cell death. Moreover, 58 transcription factors (TFs) were differentially regulated by Xcv in citrus leaves, including 26 TFs from the stress-associated families AP2-EREBP, bZip, Myb and WRKY. Remarkably, in silico analysis of the distribution of expressed sequence tags revealed that 10 of the 58 TFs, belonging to C2C2-GATA, C2H2, CCAAT, HSF, NAC and WRKY gene families, were specifically over-represented in citrus stress cDNA libraries. This study identified candidate TF genes for the regulation of key steps during the citrus non-host HR. Furthermore, these TFs might be useful in future strategies of molecular breeding for citrus disease resistance. Copyright © 2013 Elsevier GmbH. All rights reserved.

  13. Alternaria roseogrisea, a new species from achenes of Helianthus annuus (sunflower)

    USDA-ARS?s Scientific Manuscript database

    Alternaria roseogrisea was isolated from the achenes of Helianthus annuus L. during studies conducted in 1983-85 to document the diversity of fungi occurring in sunflower seeds and the possible role these fungi play in degradation of oilseed quality. A. roseogrisea was reported as "Alternaria sp. 2...

  14. Innate type 2 response to Alternaria extract enhances ryegrass-induced lung inflammation.

    PubMed

    Kim, Hee-Kyoo; Lund, Sean; Baum, Rachel; Rosenthal, Peter; Khorram, Naseem; Doherty, Taylor A

    2014-01-01

    Exposure to the fungal allergen Alternaria alternata as well as ryegrass pollen has been implicated in severe asthma symptoms during thunderstorms. We have previously shown that Alternaria extract induces innate type 2 lung inflammation in mice. We hypothesized that the innate eosinophilic response to Alternaria extract may enhance lung inflammation induced by ryegrass. Mice were sensitized to ryegrass allergen and administered a single challenge with A. alternata extract before or after final ryegrass challenges. Levels of eosinophils, neutrophils, Th2 cells, innate lymphoid cells (ILC2), interleukin (IL)-5 and IL-13 in bronchoalveolar lavage (BAL) as well as inflammation and mucus were assessed. Mice receiving ryegrass sensitization and challenge developed an eosinophilic lung response. A single challenge with Alternaria extract given 3 days before or 3 days after ryegrass challenges resulted in increased eosinophils, peribronchial inflammation and mucus production in the airways compared with ryegrass-only challenges. Type 2 ILC2 and Th2 cell recruitment to the airways was increased after Alternaria extract exposure in ryegrass-challenged mice. Innate immune challenges with Alternaria extract induced BAL eosinophilia, Th2 cell recruitment as well as ILC2 expansion and proliferation. A single exposure to Alternaria extract in ryegrass-sensitized and -challenged mice enhances the type 2 lung inflammatory response, including airway eosinophilia, peribronchial infiltrate, and mucus production, possibly through Th2 cell recruitment and ILC2 expansion. If translated to humans, exposure to both grass pollen and Alternaria may be a potential cause of thunderstorm-related asthma.

  15. Alternaria spp. from sugarbeet tolerant to four fungicide classes in Michigan

    USDA-ARS?s Scientific Manuscript database

    Alternaria species have caused minor foliar disease issues on sugarbeet historically. Recently, increased incidence and severity of Alternaria leaf spot (ALS) has been observed in several growing regions. Once defoliation is evident there is a potential to cause yield loss. One possible factor in th...

  16. Zinc treatment increases the titre of 'Candidatus Liberibacter asiaticus' in huanglongbing-affected citrus plants while affecting the bacterial microbiomes.

    PubMed

    Zhang, M Q; Guo, Y; Powell, C A; Doud, M S; Yang, C Y; Zhou, H; Duan, Y P

    2016-06-01

    Huanglongbing (HLB)-affected citrus often display zinc deficiency symptoms. In this study, supplemental zinc was applied to citrus to determine its effect on Candidatus Liberibacter asiaticus (Las) titre, HLB symptoms, and leaf microbiome. HLB-affected citrus were treated with various amounts of zinc. The treatments promoted Las growth and affected microbiomes in citrus leaves. Phylochip(™) -based results indicated that 5475 of over 50 000 known Operational Taxonomic Units (OTUs) in 52 phyla were detected in the midribs of HLB-affected citrus, of which Proteobacteria was the most abundant, followed by Firmicutes and Actinobacteria. In comparison, the microbiomes of zinc-treated diseased plants had overall more OTUs with higher amounts of Proteobacteria, but decreased percentages of Firmicutes and Actinobacteria. In addition, more OTUs of siderophore-producing bacteria were present. Only zinc-sensitive Staphylococcaceae had higher OTU's in the diseased plants without zinc treatments. Although HLB-affected citrus appear zinc deficient, zinc amendments increased the pathogen levels and shifted the microbiome. HLB is currently the most devastating disease of citrus worldwide. Zinc is often applied to HLB-affected citrus due to zinc deficiency symptoms. This study provided new insights into the potential effects of zinc on HLB and the microbial ecology of citrus. © 2016 The Society for Applied Microbiology.

  17. Widespread applications of citrus cryopreservation

    USDA-ARS?s Scientific Manuscript database

    Citrus genetic resources can now be successfully cryopreserved, which means that they can be placed into long-term storage at liquid nitrogen temperatures. This cryopreservation technology was specifically developed to address the immediate need to have secure long-term back-up storage for citrus co...

  18. The flavor of citrus fruit

    USDA-ARS?s Scientific Manuscript database

    Citrus is the largest cultivated fruit tree crop in the world, with total production of more than 100 million tons per year. The genus Citrus consists of different species, including several producing economically important crops, such as oranges, mandarins, grapefruit, pummelo, lemons and limes, c...

  19. Integrated Management of Citrus Canker

    USDA-ARS?s Scientific Manuscript database

    Fruit losses due to citrus canker, caused by Xanthomonas citri subsp. citri (Xcc), vary each crop season depending on citrus variety, tree age, flushing condition, leafminer control, and coincidence of weather events with occurrence of susceptible fruit and foliage. In 2012, crop losses in Hamlin f...

  20. Influence of environmental parameters on mycotoxin production by Alternaria arborescens.

    PubMed

    Vaquera, Sandra; Patriarca, Andrea; Fernández Pinto, V

    2016-02-16

    Alternaria arborescens has been reported as a common fungal species invading tomatoes and is capable of producing several mycotoxins in infected plants, fruits and in agricultural commodities. Alternariol (AOH), alternariol monomethyl ether (AME), and tenuazonic acid (TeA) are some of the main Alternaria mycotoxins that can be found as contaminants of food. This species can produce these toxic metabolites together with AAL toxins (Alternaria alternata f. sp. lycopersicum toxins), which can act as inhibitors of sphingolipid biosynthesis. The objective of this study was to determine the effect of water activity (aw, 0.995, 0.975, 0.950) and temperature (6, 15, 20, 25 and 30 °C) on mycotoxin production by A. arborescens on a synthetic tomato medium. The optimum production of AOH and AME occurred at 0.975 aw after 40 days of incubation at 30 °C. The maximum TeA accumulation was observed at 0.975 aw and 25 °C and at 0.950 aw and 30 °C. AAL TA was produced in higher quantities at 0.995 aw and 30 °C. At 6 °C no quantifiable levels of AOH or AME were detected, but significant amounts of TeA were produced at 0.975 aw. In general, high aw levels and high temperatures were favorable for mycotoxin production. The greatest accumulation of all four toxins occurred at 0.975 aw and 30 °C. The results obtained here could be extrapolated to evaluate the risk of tomato fruits and tomato products contamination caused by these toxins.

  1. Comparison of gene expression changes in susceptible, tolerant, and resistant hosts in response to infection with citrus tristeza virus and huanglongbing

    USDA-ARS?s Scientific Manuscript database

    The pathogens Candidatus Liberibacter asiaticus (Las) and Citrus tristeza virus (CTV) are both phloem limited and have significant economic impact on citrus production wherever they are found. Studies of host resistance have indicated that Poncirus trifoliata has tolerance or resistance to both path...

  2. Agrobacterium-mediated transformation of grapefruit with the wild-type and mutant RNA-dependent RNA polymerase genes of Citrus tristeza virus

    USDA-ARS?s Scientific Manuscript database

    Citrus paradisi Macf. cv. Duncan was transformed with constructs coding for the wild-type and mutant RNA-dependent RNA polymerase (RdRp) of Citrus tristeza virus (CTV) for exploring replicase-mediated pathogen-derived resistance (RM-PDR). The RdRp gene was amplified from CTV genome and used to gener...

  3. Improved real-time PCR detection of Candidatus Liberibacter asiaticus from citrus and psyllid hosts by targeting the intragenic tandem repeats of its prophage genes

    USDA-ARS?s Scientific Manuscript database

    Candidatus Liberibacter asiaticus (Las) is the most prevalent species of Liberibacter associated with citrus huanglongbing (HLB) worldwide. Residing in phloem sieve cells of host plants and vectored by the Asian citrus psyllid (Diaphorina citri), this fastidious bacterium lives as a pathogen or sym...

  4. Use of micro-CT to elucidate details of the anatomy and feeding of the Asian citrus psyllid Diaphorina citri Kuwayama, 1908 (Insecta: Hemiptera, Liviidae)

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB), also known as citrus greening disease, is caused by plant-infecting bacteria. The most prominent pathogen within the Americas: United States of America, Mexico, and Brazil, is Candidatus Liberibacter asiaticus, which affects plants of the Family: Rutaceae, in particularly citrus...

  5. Sclerenchymatous ring as a barrier to phloem feeding by Asian citrus psyllid: Evidence from electrical penetration graph and visualization of stylet pathways

    USDA-ARS?s Scientific Manuscript database

    Asian citrus psyllid (ACP) feeding behaviors play a significant role in the transmission of the phloem-limited Candidatus Liberibacter asiaticus (CLas) bacteria that cause citrus greening disease. Sustained phloem ingestion by ACP on CLas infected plants is very important in pathogen acquisition and...

  6. Diversity and movement of indoor Alternaria alternata across the mainland USA.

    PubMed

    Woudenberg, J H C; van der Merwe, N A; Jurjević, Ž; Groenewald, J Z; Crous, P W

    2015-08-01

    Alternaria spp. from sect. Alternaria are frequently associated with hypersensitivity pneumonitis, asthma and allergic fungal rhinitis and sinusitis. Since Alternaria is omnipresent in the outdoor environment, it is thought that the indoor spore concentration is mainly influenced by the outdoor spore concentration. However, few studies have investigated indoor Alternaria isolates, or attempted a phylogeographic or population genetic approach to investigate their movement. Therefore, the aim of the current study was to investigate the molecular diversity of indoor Alternaria isolates in the USA, and to test for recombination, using these approaches. Alternaria isolates collected throughout the USA were identified using ITS, gapdh and endoPG gene sequencing. This was followed by genotyping and population genetic inference of isolates belonging to Alternaria sect. Alternaria together with 37 reference isolates, using five microsatellite markers. Phylogenetic analyses revealed that species of Alternaria sect. Alternaria represented 98% (153 isolates) of the indoor isolates collected throughout the USA, of which 137 isolates could be assigned to A. alternata, 15 to the A. arborescens species complex and a single isolate to A. burnsii. The remaining 2% (3 isolates) represented sect. Infectoriae (single isolate) and sect. Pseudoulocladium (2 isolates). Population assignment analyses of the 137 A. alternata isolates suggested that subpopulations did not exist within the sample. The A. alternata isolates were thus divided into four artificial subpopulations to represent four quadrants of the USA. Forty-four isolates representing the south-western quadrant displayed the highest level of uniqueness based on private alleles, while the highest level of gene flow was detected between the south-eastern (32 isolates) and south-western quadrants. Genotypic diversity was high for all quadrants, and a test for linkage disequilibrium suggested that A. alternata has a cryptic sexual

  7. Antimicrobial effects of Citrus sinensis peel extracts against dental caries bacteria: An in vitro study

    PubMed Central

    Shetty, Sapna B.; Mahin-Syed-Ismail, Prabu; Varghese, Shaji; Thomas-George, Bibin; Kandathil- Thajuraj, Pathinettam; Baby, Deepak; Haleem, Shaista; Sreedhar, Sreeja

    2016-01-01

    Background Ethnomedicine is gaining admiration since years but still there is abundant medicinal flora which is unrevealed through research. The study was conducted to assess the in vitro antimicrobial potential and also determine the minimum inhibitory concentration (MIC) of Citrus sinensis peel extracts with a view of searching a novel extract as a remedy for dental caries pathogens. Material and Methods Aqueous and ethanol (cold and hot) extracts prepared from peel of Citrus sinensis were screened for in vitro antimicrobial activity against Streptococcus mutans and Lactobacillus acidophilus, using agar well diffusion method. The lowest concentration of every extract considered as the minimal inhibitory concentration (MIC) values were determined for both test organisms. One way ANOVA with Post Hoc Bonferroni test was applied for statistical analysis. Confidence level and level of significance were set at 95% and 5% respectively. Results Dental caries pathogens were inhibited most by hot ethanolic extract of Citrus sinensispeel followed by cold ethanolic extract. Aqueous extracts were effective at very high concentrations. Minimum inhibitory concentration of hot and cold ethanolic extracts of Citrus sinensis peel ranged between 12-15 mg/ml against both the dental caries pathogens. Conclusions Citrus sinensispeels extract was found to be effective against dental caries pathogens and contain compounds with therapeutic potential. Nevertheless, clinical trials on the effect of these plants are essential before advocating large-scale therapy. Key words:Agar well diffusion, antimicrobial activity, dental caries, Streptococcus mutans, Lactobacillus acidophilus. PMID:26855710

  8. Antimicrobial effects of Citrus sinensis peel extracts against dental caries bacteria: An in vitro study.

    PubMed

    Shetty, Sapna B; Mahin-Syed-Ismail, Prabu; Varghese, Shaji; Thomas-George, Bibin; Kandathil-Thajuraj, Pathinettam; Baby, Deepak; Haleem, Shaista; Sreedhar, Sreeja; Devang-Divakar, Darshan

    2016-02-01

    Ethnomedicine is gaining admiration since years but still there is abundant medicinal flora which is unrevealed through research. The study was conducted to assess the in vitro antimicrobial potential and also determine the minimum inhibitory concentration (MIC) of Citrus sinensis peel extracts with a view of searching a novel extract as a remedy for dental caries pathogens. Aqueous and ethanol (cold and hot) extracts prepared from peel of Citrus sinensis were screened for in vitro antimicrobial activity against Streptococcus mutans and Lactobacillus acidophilus, using agar well diffusion method. The lowest concentration of every extract considered as the minimal inhibitory concentration (MIC) values were determined for both test organisms. One way ANOVA with Post Hoc Bonferroni test was applied for statistical analysis. Confidence level and level of significance were set at 95% and 5% respectively. Dental caries pathogens were inhibited most by hot ethanolic extract of Citrus sinensispeel followed by cold ethanolic extract. Aqueous extracts were effective at very high concentrations. Minimum inhibitory concentration of hot and cold ethanolic extracts of Citrus sinensis peel ranged between 12-15 mg/ml against both the dental caries pathogens. Citrus sinensispeels extract was found to be effective against dental caries pathogens and contain compounds with therapeutic potential. Nevertheless, clinical trials on the effect of these plants are essential before advocating large-scale therapy. Agar well diffusion, antimicrobial activity, dental caries, Streptococcus mutans, Lactobacillus acidophilus.

  9. Seasonal Movement Patterns and Long-Range Dispersal of Asian Citrus Psyllid in Florida Citrus.

    PubMed

    Lewis-Rosenblum, Hannah; Martini, Xavier; Tiwari, Siddharth; Stelinski, Lukasz L

    2015-02-01

    The Asian citrus psyllid, Diaphorina citri Kuwayama, is the vector of the bacterial pathogen, Candidatus Liberibacter asiaticus, which is the causal agent of huanglongbing (HLB) in the United States. Both short-range and long-range dispersal of D. citri adults affect the spread of HLB; however, little is known about the long-range dispersal capabilities of D. citri in the field or the seasonality of flight behavior. In the present study, an in situ protein marking technique was used to determine the dispersal of D. citri by trapping marked adults under natural field conditions. D. citri movement from abandoned citrus groves to adjacent managed citrus groves was greatest during the spring and summer months and decreased significantly during the colder months (September-March). D. citri were able to traverse potential geographic barriers such as roads and fallow fields. In an experiment conducted to determine long-range dispersal capacity in the absence of severe weather events, D. citri were able to disperse at least 2 km within 12 d. Wind direction was not correlated with the number of marked psyllids captured, indicating substantial flight capability by D. citri. Finally, the number of marked psyllids captured increased with the density of emerging young leaves on surrounding trees. The results confirm that abandoned citrus groves in Florida serve as reservoirs for D. citri, which can disperse across long distances despite geographical barriers. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  10. Activation of Melanin Synthesis in Alternaria infectoria by Antifungal Drugs.

    PubMed

    Fernandes, Chantal; Prados-Rosales, Rafael; Silva, Branca M A; Nakouzi-Naranjo, Antonio; Zuzarte, Mónica; Chatterjee, Subhasish; Stark, Ruth E; Casadevall, Arturo; Gonçalves, Teresa

    2015-12-28

    The importance of Alternaria species fungi to human health ranges from their role as etiological agents of serious infections with poor prognoses in immunosuppressed individuals to their association with respiratory allergic diseases. The present work focuses on Alternaria infectoria, which was used as a model organism of the genus, and was designed to unravel melanin production in response to antifungals. After we characterized the pigment produced by A. infectoria, we studied the dynamics of 1,8-dihydroxynaphthalene (DHN)-melanin production during growth, the degree of melanization in response to antifungals, and how melanization affected susceptibility to several classes of therapeutic drugs. We demonstrate that A. infectoria increased melanin deposition in cell walls in response to nikkomycin Z, caspofungin, and itraconazole but not in response to fluconazole or amphotericin B. These results indicate that A. infectoria activates DHN-melanin synthesis in response to certain antifungal drugs, possibly as a protective mechanism against these drugs. Inhibition of DHN-melanin synthesis by pyroquilon resulted in a lower minimum effective concentration (MEC) of caspofungin and enhanced morphological changes (increased hyphal balloon size), characterized by thinner and less organized A. infectoria cell walls. In summary, A. infectoria synthesizes melanin in response to certain antifungal drugs, and its susceptibility is influenced by melanization, suggesting the therapeutic potential of drug combinations that affect melanin synthesis.

  11. Activation of Melanin Synthesis in Alternaria infectoria by Antifungal Drugs

    PubMed Central

    Fernandes, Chantal; Prados-Rosales, Rafael; Silva, Branca M. A.; Nakouzi-Naranjo, Antonio; Zuzarte, Mónica; Chatterjee, Subhasish; Stark, Ruth E.; Casadevall, Arturo

    2015-01-01

    The importance of Alternaria species fungi to human health ranges from their role as etiological agents of serious infections with poor prognoses in immunosuppressed individuals to their association with respiratory allergic diseases. The present work focuses on Alternaria infectoria, which was used as a model organism of the genus, and was designed to unravel melanin production in response to antifungals. After we characterized the pigment produced by A. infectoria, we studied the dynamics of 1,8-dihydroxynaphthalene (DHN)-melanin production during growth, the degree of melanization in response to antifungals, and how melanization affected susceptibility to several classes of therapeutic drugs. We demonstrate that A. infectoria increased melanin deposition in cell walls in response to nikkomycin Z, caspofungin, and itraconazole but not in response to fluconazole or amphotericin B. These results indicate that A. infectoria activates DHN-melanin synthesis in response to certain antifungal drugs, possibly as a protective mechanism against these drugs. Inhibition of DHN-melanin synthesis by pyroquilon resulted in a lower minimum effective concentration (MEC) of caspofungin and enhanced morphological changes (increased hyphal balloon size), characterized by thinner and less organized A. infectoria cell walls. In summary, A. infectoria synthesizes melanin in response to certain antifungal drugs, and its susceptibility is influenced by melanization, suggesting the therapeutic potential of drug combinations that affect melanin synthesis. PMID:26711773

  12. Mapping Resistance to Alternaria cucumerina in Cucumis melo.

    PubMed

    Daley, James; Branham, Sandra; Levi, Amnon; Hassell, Richard; Wechter, Patrick

    2017-04-01

    Infection with Alternaria cucumerina causes Alternaria leaf blight (ALB), a disease characterized by lesion formation on leaves, leading to substantial yield and quality losses in Cucumis melo (melon). Although fungicides are effective against ALB, reduction in the frequency of application would be economically and environmentally beneficial. Resistant melon lines have been identified but the genetic basis of this resistance has not been determined. A saturated melon genetic map was constructed with markers developed through genotyping by sequencing of a recombinant inbred line population (F6 to F10; n = 82) derived from single-seed descent of a F2 population from a cross between the ALB-resistant parent MR-1 and the ALB-susceptible parent Ananas Yokneum. The population was evaluated for A. cucumerina resistance with an augmented block greenhouse study using inoculation with the wounded-leaf method. Multiple quantitative trait loci (QTL) mapping identified two QTL that explained 33.9% of variation in lesion area. Several candidate genes within range of these QTL were identified using the C. melo v3.5 genome. Markers linked to these QTL will be used to accelerate efforts to breed melon cultivars resistant to ALB.

  13. Plant extracts to control Alternaria alternata in Murcott tangor fruits.

    PubMed

    Carvalho, Daniel Diego Costa; Alves, Eduardo; Barbosa Camargos, Renato; Ferreira Oliveira, Denilson; Soares Scolforo, José Roberto; de Carvalho, Douglas Antônio; Sâmia Batista, Tereza Raquel

    2011-01-01

    Alternaria alternata causes the Alternaria brown spot disease (ABS) in many tangerines and their hybrids worldwide. Plant extracts offer an alternative method for controlling this disease, which control is based on chemical fungicides. To identify plant species with antifungal properties against A. alternata, the causal agent of the ABS. Plant extracts prepared from leaves, barks, flowers, and stalks collected from 105 plant species in the State of Minas Gerais, Brazil, were tested for activity against the fungus A. alternata in vitro and in vivo. The most promising extract was obtained from Anadenanthera colubrina, which reduced the disease on Murcott tangor fruits to levels obtained with commercial fungicides. Artemisia annua, Cariniana estrelensis, Ficus carica, and Ruta graveolens presented moderate in vitro antifungal activity, but no effects were observed on the disease when the extracts were applied to fruits inoculated with the fungus. Besides, A. colubrina was the most active extract against A. alternata in the in vitro assay. The results obtained in the in vitro and in vivo assays suggested that the fungal growth test, which uses 96-well polypropylene plates, seems to be appropriate for selecting potential plant species for testing new methods to control ABS. Copyright © 2011 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  14. The Evolutionary History and Spatiotemporal Dynamics of the NC Lineage of Citrus Tristeza Virus.

    PubMed

    Benítez-Galeano, María José; Castells, Matías; Colina, Rodney

    2017-10-12

    Citrus tristeza virus (CTV) is a major pathogen affecting citrus trees worldwide. However, few studies have focused on CTV's evolutionary history and geographic behavior. CTV is locally dispersed by an aphid vector and long distance dispersion due to transportation of contaminated material. With the aim to delve deeper into the CTV-NC (New Clade) genotype evolution, we estimated an evolution rate of 1.19 × 10(-3) subs/site/year and the most common recent ancestor in 1977. Furthermore, the place of origin of the genotype was in the United States, and a great expansion of the population was observed in Uruguay. This expansion phase could be a consequence of the increment in the number of naïve citrus trees in Uruguayan orchards encompassing citrus industry growth in the past years.

  15. Tale of the Huanglongbing Disease Pyramid in the Context of the Citrus Microbiome.

    PubMed

    Wang, Nian; Stelinski, Lukasz L; Pelz-Stelinski, Kirsten S; Graham, James H; Zhang, Yunzeng

    2017-02-06

    The Huanglongbing (HLB) disease pyramid is composed of Liberibacters, psyllid vectors, citrus hosts, and the environment. The epidemiological outcomes for Liberibacter-associated plant diseases are collectively determined by the inherent relationships among plant-Liberibacters-psyllids, and how various environmental factors affect plant-Liberibacter-psyllid interactions. Citrus-Liberibacter-psyllid interactions occur in a complex microbiome system. In this review, we focus on the progress in understanding the HLB disease pyramid, and how the microbiome affects the HLB disease pyramid including the interaction between HLB and the citrus microbiome; the interaction between Liberibacters and psyllids; the interaction between Liberibacters and gut microbiota in psyllids; and the effect of HLB on selected above- and belowground citrus pathogens. Their implications for HLB management are also discussed.

  16. [Determination of the profiles of secondary metabolites characteristic of Alternaria strains isolated from tomato].

    PubMed

    Benavidez Rozo, Martha Elizabeth; Patriarca, Andrea; Cabrera, Gabriela; Fernández Pinto, Virginia E

    2014-01-01

    Many Alternaria species have been studied for their ability to produce bioactive secondary metabolites, such as tentoxin (TEN), some of which have toxic properties. The main food contaminant toxins are tenuazonic acid, alternariol (AOH), alternariol monomethyl ether (AME), altenuene, and altertoxins i, ii and iii. To determine the profiles of secondary metabolites characteristic of Alternaria strains isolated from tomato for their chemotaxonomic classification. The profiles of secondary metabolites were determined by HPLC MS. The Alternaria isolates obtained from spoiled tomatoes belong, according to their morphological characteristics, to the species groups Alternaria alternata, Alternaria tenuissima and Alternaria arborescens, with A. tenuissima being the most frequent. The most frequent profiles of secondary metabolites belonging to the species groups A. alternata (AOH, AME, TEN), A. tenuissima (AOH, AME, TEN, tenuazonic acid) and A. arborescens (AOH, AME, TEN, tenuazonic acid) were determined, with some isolates of the latter being able to synthesize AAL toxins. Secondary metabolite profiles are a useful tool for the differentiation of small spored Alternaria isolates not easily identifiable by their morphological characteristics. Copyright © 2013 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  17. Development of loop-mediated isothermal amplification and SYBR green real-time PCR methods for the detection of Citrus yellow mosaic badnavirus in citrus species.

    PubMed

    Anthony Johnson, A M; Dasgupta, I; Sai Gopal, D V R

    2014-07-01

    Citrus yellow mosaic badnavirus (CMBV) is an important pathogen in southern India spread by infected citrus propagules. One of the measures to arrest the spread of CMBV is to develop methods to screen and certify citrus propagules as CMBV-free. The methods loop-mediated isothermal amplification (LAMP) and SYBR green real-time PCR (SGRTPCR) have been developed for the efficient detection of CMBV in citrus propagules. This paper compares the sensitivities of LAMP and SGRTPCR with polymerase chain reaction (PCR) for the detection of CMBV. Whereas PCR and LAMP were able to detect CMBV from a minimum of 10 ng of total DNA of infected leaf samples, SGRTPCR could detect the same from 1 ng of total DNA. Using SGRTPCR, the viral titres were estimated to be the highest in rough lemon and lowest in Nagpur Mandarin of the five naturally infected citrus species tested. The results will help in designing suitable strategies for the sensitive detection of CMBV from citrus propagules. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Chemical constituents of marine mangrove-derived endophytic fungus Alternaria tenuissima EN-192

    NASA Astrophysics Data System (ADS)

    Sun, Hong; Gao, Shushan; Li, Xiaoming; Li, Chunshun; Wang, Bingui

    2013-03-01

    A chemical investigation of the ethyl acetate extract of the fermentation broth of Alternaria tenuissima EN-192, an endophytic fungus obtained from the stems of the marine mangrove plant Rhizophora stylosa, resulted in the isolation of nine known secondary metabolites, including four indole-diterpenoids: penijanthine A ( 1), paspaline ( 2), paspalinine ( 3), and penitrem A ( 4); three tricycloalternarene derivatives: tricycloalternarene 3a ( 5), tricycloalternarene 1b ( 6), and tricycloalternarene 2b ( 7); and two alternariol congeners: djalonensone ( 8) and alternariol ( 9). The chemical structures of these metabolites were characterized through a combination of detailed spectroscopic analyses and their comparison with reports from the literature. The inhibitory activities of each isolated compound against four bacteria were evaluated and compounds 5 and 8 displayed moderate activity against the aquaculture pathogenic bacterium Vibrio anguillarum, with inhibition zone diameters of 8 and 9 mm, respectively, at 100 μg/disk. To the best of our knowledge, this is the first report on the secondary metabolites of mangrove-derived A lternaria tenuissima and also the first report of the isolation of indole-diterpenoids from fungal genus A lternaria.

  19. Resistance to Alternaria solani in hybrids between a Solanum tuberosum haploid and S. raphanifolium.

    PubMed

    Weber, B N; Jansky, S H

    2012-02-01

    Early blight of potato (Solanum tuberosum), caused by the foliar fungal pathogen Alternaria solani, is a major cause of economic loss in many potato-growing regions. Genetic resistance offers an opportunity to decrease fungicide usage while maintaining yield and quality. In this study, an early blight resistant clone of the diploid wild species S. raphanifolium was crossed as a male to a haploid (2n=2x) of cultivated potato. Hybrids were backcrossed to both parents. Eight families were created and evaluated for early blight resistance in the field. Families created by backcrossing to the wild species parent exhibited significantly lower relative area under the disease progress curve means than those from backcrossing to the cultivated parent, leading to the conclusion that S. raphanifolium contributes genes for early blight resistance. The mechanism of resistance in S. raphanifolium is unique because A. solani could not be recovered from lesions. Clones were identified with high levels of resistance and adaptation to the photoperiod of a temperate production region.

  20. Cytological and physiological basis for tomato varietal resistance against Alternaria alternata.

    PubMed

    Ahmad, Aqeel; Shafique, Shazia; Shafique, Sobiya

    2013-07-01

    Since tomato is an important food component, it is imperative to enhance its yield against the activities of many devastating fungal pathogens such as Alternaria alternata. The exploitation of plant innate resistance by cultivation of resistant varieties is an effective measure in this regard. In the present study, 28 tomato varieties were tested against 32 A. alternata isolates, and representative varieties were further evaluated to determine the extent and basis of their antifungal resistance. A significant increase (104.7%) in polyphenols was recorded in the resistant variety Dinaar compared with the susceptible variety Red Tara. Dinaar also exhibited 100% enhancement of alkaloids and terpenoids along with a 30.7% increase in cell wall hemicellulose content. Significant differences were found in physical barriers (cellulose, lignin and pectin) of the representative varieties when stained tissue sections were subjected to colorimetric analysis. Similarly, polyphenol oxidase, peroxidase and phenylalanine ammonia lyase showed increases of 78.37, 114.67 and 125.11% respectively in the resistant variety. Higher expression of glucanase genes was evident from native gel analysis, in which not only the number of isozymes but also the quantity of individual isozymes was significantly increased. The resistant variety Dinaar had strong antifungal resistance and can therefore be recommended as suitable for cultivation in the agricultural system of Pakistan. © 2013 Society of Chemical Industry.

  1. Arabidopsis Responds to Alternaria alternata Volatiles by Triggering Plastid Phosphoglucose Isomerase-Independent Mechanisms1[OPEN

    PubMed Central

    Sánchez-López, Ángela María; Bahaji, Abdellatif; De Diego, Nuria; Baslam, Marouane; Li, Jun; Almagro, Goizeder; García-Gómez, Pablo; Ricarte-Bermejo, Adriana; Novák, Ondřej; Spíchal, Lukáš; Ciordia, Sergio; Mena, María Carmen

    2016-01-01

    Volatile compounds (VCs) emitted by phylogenetically diverse microorganisms (including plant pathogens and microbes that do not normally interact mutualistically with plants) promote photosynthesis, growth, and the accumulation of high levels of starch in leaves through cytokinin (CK)-regulated processes. In Arabidopsis (Arabidopsis thaliana) plants not exposed to VCs, plastidic phosphoglucose isomerase (pPGI) acts as an important determinant of photosynthesis and growth, likely as a consequence of its involvement in the synthesis of plastidic CKs in roots. Moreover, this enzyme plays an important role in connecting the Calvin-Benson cycle with the starch biosynthetic pathway in leaves. To elucidate the mechanisms involved in the responses of plants to microbial VCs and to investigate the extent of pPGI involvement, we characterized pPGI-null pgi1-2 Arabidopsis plants cultured in the presence or absence of VCs emitted by Alternaria alternata. We found that volatile emissions from this fungal phytopathogen promote growth, photosynthesis, and the accumulation of plastidic CKs in pgi1-2 leaves. Notably, the mesophyll cells of pgi1-2 leaves accumulated exceptionally high levels of starch following VC exposure. Proteomic analyses revealed that VCs promote global changes in the expression of proteins involved in photosynthesis, starch metabolism, and growth that can account for the observed responses in pgi1-2 plants. The overall data show that Arabidopsis plants can respond to VCs emitted by phytopathogenic microorganisms by triggering pPGI-independent mechanisms. PMID:27663407

  2. Rhamnolipids induce oxidative stress responses in cherry tomato fruit to Alternaria alternata.

    PubMed

    Yan, Fujie; Hu, Hao; Lu, Laifeng; Zheng, Xiaodong

    2016-08-01

    Rhamnolipids showed an antimicrobial activity applicable to a variety of pathogenic microorganisms, but mechanisms were mostly focused on their direct inhibitory effect. This study showed that disease incidence obviously decreased when cherry tomatoes were treated with rhamnolipids, whether before or after Alternaria alternata inoculation. The activities of antioxidant enzymes, including superoxide dismutase and catalase, were increased in rhamnolipid-pretreated cherry tomato inoculated with A. alternata within 12 h, while contents of reactive oxygen species decreased. Moreover, resistant response of cherry tomato treated with rhamnolipids and A. alternata was also attributed to activity stimulation of ascorbate peroxidase and glutathione reductase, accompanied with an increase in reduced glutathione, which is beneficial for scavenging excessive H2 O2 . Results indicated that rhamnolipids could effectively reduce fungal disease of harvested cherry tomato by inducing fruit resistance and mechanisms involved in elicitation of antioxidative reactions such as the ability to scavenge excess reactive oxygen species. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

  3. [Effects of Alternaria tenuis nees on Trifolium repens L. under Cu stress].

    PubMed

    Chu, Ling; Shao, Deng-hui; Jin, Song; Wu, Xue-feng

    2007-11-01

    With pot culture, this paper studied the effects of Alternaria tenuis Nees inoculation on the eco-physiological indices of Trifolium repens L. leaf under Cu stress. The results showed that in the control (not inoculated with the pathogen), the contents of photosynthetic pigments (chlorophyll a, b, a + b and carotenoid) and soluble protein in T. repens leaves decreased markedly with increasing Cu concentration (0-3000 mg x kg(-1)). The enhancement of cellular membrane lipids peroxidation with the increase of Cu concentration led to a rapid accumulation of malondialdehyde (MDA), the damage on cellular membrane structure, and an increase of electric conductivity. The balance of active oxygen metabolism systems was broken, SOD and CAT activities decreased, while POD activity increased. After inoculation with A. tenuis, the damages of Cu on plant membrane systems and active oxygen metabolism systems aggravated, the contents of photosynthetic pigments and soluble protein and the activities of SOD and CAT decreased to different degree, while the electric conductivity, MDA content and POD activity increased markedly, compared with the control.

  4. Mold and Alternaria skin test reactivity and asthma in children in Connecticut.

    PubMed

    Lyons, Todd W; Wakefield, Dorothy B; Cloutier, Michelle M

    2011-04-01

    Sensitivity to mold has been associated with asthma incidence, persistence, and severity. To examine the relationship between skin test reactivity (STR) to molds and specifically to Alternaria and asthma severity in a group of ethnically diverse children in Connecticut. Demographics and STR to 14 local allergens, including Alternaria, Penicillium, and mold mix, were obtained for 914 Puerto Rican, African American, and non-Hispanic white children. A total of 126 children (14%) had a positive skin test result to mold, and 58 (6%) demonstrated STR to Alternaria. Compared with non-Hispanic white children, there was no difference in the likelihood of being sensitized to Alternaria for Puerto Rican and African American children (odds ratio [OR], 0.7; 95% confidence interval [CI], 0.3-1.5; and OR, 0.9; 95% CI, 0.4-2.2; respectively). In an adjusted analysis, Alternaria STR was associated with severe, persistent asthma (OR, 3.4; 95% CI, 1.2-8.6) but did not predict increasing asthma severity. STR to cat (OR, 2.5; 95% CI, 1.3-4.9) and dog (OR, 2.9; 95% CI, 1.3-6.0) was also associated with severe persistent asthma. Alternaria STR was associated with severe persistent asthma independent of the total number of positive skin test results. Mold and Alternaria STR were uncommon among children in Connecticut. Alternaria STR was not associated with increasing asthma severity but was associated with severe, persistent asthma independent of the total number of positive skin test results. There was no association between ethnicity and Alternaria STR. Copyright © 2011 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  5. Mold and Alternaria skin test reactivity and asthma in children in Connecticut

    PubMed Central

    Lyons, Todd W.; Wakefield, Dorothy B.; Cloutier, Michelle M.

    2013-01-01

    Background Sensitivity to mold has been associated with asthma incidence, persistence, and severity. Objective To examine the relationship between skin test reactivity (STR) to molds and specifically to Alternaria and asthma severity in a group of ethnically diverse children in Connecticut. Methods Demographics and STR to 14 local allergens, including Alternaria, Penicillium, and mold mix, were obtained for 914 Puerto Rican, African American, and non-Hispanic white children. Results A total of 126 children (14%) had a positive skin test result to mold, and 58 (6%) demonstrated STR to Alternaria. Compared with non-Hispanic white children, there was no difference in the likelihood of being sensitized to Alternaria for Puerto Rican and African American children (odds ratio [OR], 0.7; 95% confidence interval [CI], 0.3–1.5; and OR, 0.9; 95% CI, 0.4–2.2; respectively). In an adjusted analysis, Alternaria STR was associated with severe, persistent asthma (OR, 3.4; 95% CI, 1.2–8.6) but did not predict increasing asthma severity. STR to cat (OR, 2.5; 95% CI, 1.3–4.9) and dog (OR, 2.9; 95% CI, 1.3–6.0) was also associated with severe persistent asthma. Alternaria STR was associated with severe persistent asthma independent of the total number of positive skin test results. Conclusions Mold and Alternaria STR were uncommon among children in Connecticut. Alternaria STR was not associated with increasing asthma severity but was associated with severe, persistent asthma independent of the total number of positive skin test results. There was no association between ethnicity and Alternaria STR. PMID:21457878

  6. Impaired induction of allergic lung inflammation by Alternaria alternata mutant MAPK homologue Fus3

    PubMed Central

    Kim, Hee-Kyoo; Baum, Rachel; Lund, Sean; Khorram, Naseem; Yang, Siwy Ling; Chung, Kuang-Ren; Doherty, Taylor A.

    2014-01-01

    The fungal allergen Alternaria alternata is associated with development of asthma, though the mechanisms underlying the allergenicity of Alternaria are largely unknown. The aim of this study was to identify whether the MAP kinase homologue Fus3 of Alternaria contributed to allergic airway responses. Wild-type (WT) and Fus3 deficient Alternaria extracts were given intranasal to mice. Extracts from Fus3 deficient Alternaria that had a functional copy of Fus3 introduced were also administered (CpFus3). Mice were challenged once and levels of BAL eosinophils and innate cytokines IL-33, thymic stromal lymphopoeitin (TSLP), and IL-25 (IL-17E) were assessed. Alternaria extracts or protease-inhibited extract were administered with (OVA) during sensitization prior to ovalbumin only challenges to determine extract adjuvant activity. Levels of BAL inflammatory cells, Th2 cytokines, and OX40-expressing Th2 cells as well as airway infiltration and mucus production were measured. WT Alternaria induced innate airway eosinophilia within 3 days. Mice given Fus3 deficient Alternaria were signifcantly impaired in developing airway eosinophilia that was largely restored by CpFus3. Further, BAL IL-33, TSLP, and Eotaxin-1 levels were reduced after challenge with Fus3 mutant extract compared with WT and CpFus3 extracts. WT and CpFus3 extracts demonstrated strong adjuvant activity in vivo as levels of BAL eosinophils, Th2 cytokines, and OX40-expressing Th2 cells as well as peribronchial inflammation and mucus production were induced. In contrast, the adjuvant activity of Fus3 extract or protease-inhibited WT extract was largely impaired. Finally, protease activity and Alt a1 levels were reduced in Fus3 mutant extract. Thus, Fus3 contributes to the Th2-sensitizing properties of Alternaria. PMID:24102366

  7. Epidemic asthma and the role of the fungal mold Alternaria alternata.

    PubMed

    Pulimood, Thomas B; Corden, Julie M; Bryden, Clare; Sharples, Linda; Nasser, Shuaib M

    2007-09-01

    After July 29, 2002, an epidemic of asthma admissions was associated with a thunderstorm in the United Kingdom. We sought to study the cause of epidemics of asthma associated with thunderstorms. We performed a case-control study of 26 patients presenting to Cambridge University Hospital with asthma after the thunderstorm. Control subjects were 31 patients with summer seasonal asthma. Subjects underwent skin tests and specific IgE serology to inhaled aeroallergens. Meteorologic and aerobiologic data correlated with asthma admissions were analyzed. Twenty-three of 26 cases had IgE sensitization to Alternaria species. Eleven of 31 control subjects gave a history of asthma exacerbation during thunderstorms. Ten of these 11 control subjects were sensitive to Alternaria species on skin testing, but Alternaria species sensitivity was only identified in 4 of the 20 remaining control subjects who did not report thunderstorm-related asthma symptoms. The odds ratio of having epidemic thunderstorm-related asthma if sensitive to Alternaria species was 9.31 (95% CI, 2.305-37.601; P = .0008) and 63.966 (95% CI, 3.577-1143.9; P < .0001) if sensitive to Alternaria species, Cladosporium species, or both. Poisson regression analysis showed that counts of broken Alternaria species and Didymella and Cladosporium species were significantly correlated with each other and with asthma admissions. The thunderstorm was associated with increased levels of Alternaria, Cladosporium, and Didymella species. Alternaria alternata sensitivity is a compelling predictor of epidemic asthma in patients with seasonal asthma and grass pollen allergy and is likely to be the important factor in thunderstorm-related asthma. Alternaria species sensitization in asthmatic subjects with grass pollen sensitivity predicts susceptibility to thunderstorm-associated asthma.

  8. Effects of soil-applied imidacloprid on Asian citrus psyllid (Hemiptera: Psyllidae) feeding behavior.

    PubMed

    Serikawa, R H; Backus, E A; Rogers, M E

    2012-10-01

    The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), is one of the most important pests of citrus (Citrus spp.) because of its status as a vector of Candidatus Liberibacter asiaticus (Las), the bacterium associated with citrus greening disease. The use of insecticides for vector control is the primary method of managing the spread of this pathogen. Imidacloprid is an insecticide commonly applied to the root zone of young citrus trees to provide systemic protection from pests. The effects of imidacloprid on feeding behavior of D. citri have not been studied in much detail. The purpose of this study was to examine the effects of imidacloprid application on feeding behavior of D. citri and to determine whether use of this systemic insecticide could have any effect on pathogen transmission by D. citri. A direct current electrical penetration graph monitor was used to record D. citri feeding behaviors for 12-h periods on mature and young leaves of imidacloprid-treated and -untreated citrus seedlings. Overall, compared with untreated plants, the feeding behavior of D. citri was disrupted on imidacloprid-treated plants via reduction in the number of probes, as well as durations of average probes, initial stylet contact with phloem, phloem salivation, and phloem ingestion. The results of this study demonstrate that soil applications of imidacloprid can reduce the probability of citrus plants becoming inoculated with Las through a reduction in the number and duration of phloem salivation events by D. citri. Furthermore, Las acquisition from infected citrus is greatly reduced as a result of decreased phloem ingestion by D. citri on imidacloprid-treated plants.

  9. Two previously unknown Phytophthora species associated with brown rot of Pomelo (Citrus grandis) fruits in Vietnam

    PubMed Central

    Schena, Leonardo; Jung, Thomas; Evoli, Maria; Pane, Antonella; Van Hoa, Nguyen; Van Tri, Mai; Wright, Sandra; Ramstedt, Mauritz; Olsson, Christer; Faedda, Roberto; Magnano di San Lio, Gaetano

    2017-01-01

    Two distinct Phytophthora taxa were found to be associated with brown rot of pomelo (Citrus grandis), a new disease of this ancestral Citrus species, in the Vinh Long province, Mekong River Delta area, southern Vietnam. On the basis of morphological characters and using the ITS1-5.8S-ITS2 region of the rDNA and the cytochrome oxidase subunit 1 (COI) as barcode genes, one of the two taxa was provisionally named as Phytophthora sp. prodigiosa, being closely related to but distinct from P. insolita, a species in Phytophthora Clade 9, while the other one, was closely related to but distinct from the Clade 2 species P. meadii and was informally designated as Phytophthora sp. mekongensis. Isolates of P. sp. prodigiosa and P. sp. mekongensis were also obtained from necrotic fibrous roots of Volkamer lemon (C. volkameriana) rootstocks grafted with ‘King’ mandarin (Citrus nobilis) and from trees of pomelo, respectively, in other provinces of the Mekong River Delta, indicating a widespread occurrence of both Phytophthora species in this citrus-growing area. Koch’s postulates were fulfilled via pathogenicity tests on fruits of various Citrus species, including pomelo, grapefruit (Citrus x paradisi), sweet orange (Citrus x sinensis) and bergamot (Citrus x bergamia) as well as on the rootstock of 2-year-old trees of pomelo and sweet orange on ‘Carrizo’ citrange (C. sinensis ‘Washington Navel’ x Poncirus trifoliata). This is the first report of a Phytophthora species from Clade 2 other than P. citricola and P. citrophthora as causal agent of fruit brown rot of Citrus worldwide and the first report of P. insolita complex in Vietnam. Results indicate that likely Vietnam is still an unexplored reservoir of Phytophthora diversity. PMID:28208159

  10. Two previously unknown Phytophthora species associated with brown rot of Pomelo (Citrus grandis) fruits in Vietnam.

    PubMed

    Puglisi, Ivana; De Patrizio, Alessandro; Schena, Leonardo; Jung, Thomas; Evoli, Maria; Pane, Antonella; Van Hoa, Nguyen; Van Tri, Mai; Wright, Sandra; Ramstedt, Mauritz; Olsson, Christer; Faedda, Roberto; Magnano di San Lio, Gaetano; Cacciola, Santa Olga

    2017-01-01

    Two distinct Phytophthora taxa were found to be associated with brown rot of pomelo (Citrus grandis), a new disease of this ancestral Citrus species, in the Vinh Long province, Mekong River Delta area, southern Vietnam. On the basis of morphological characters and using the ITS1-5.8S-ITS2 region of the rDNA and the cytochrome oxidase subunit 1 (COI) as barcode genes, one of the two taxa was provisionally named as Phytophthora sp. prodigiosa, being closely related to but distinct from P. insolita, a species in Phytophthora Clade 9, while the other one, was closely related to but distinct from the Clade 2 species P. meadii and was informally designated as Phytophthora sp. mekongensis. Isolates of P. sp. prodigiosa and P. sp. mekongensis were also obtained from necrotic fibrous roots of Volkamer lemon (C. volkameriana) rootstocks grafted with 'King' mandarin (Citrus nobilis) and from trees of pomelo, respectively, in other provinces of the Mekong River Delta, indicating a widespread occurrence of both Phytophthora species in this citrus-growing area. Koch's postulates were fulfilled via pathogenicity tests on fruits of various Citrus species, including pomelo, grapefruit (Citrus x paradisi), sweet orange (Citrus x sinensis) and bergamot (Citrus x bergamia) as well as on the rootstock of 2-year-old trees of pomelo and sweet orange on 'Carrizo' citrange (C. sinensis 'Washington Navel' x Poncirus trifoliata). This is the first report of a Phytophthora species from Clade 2 other than P. citricola and P. citrophthora as causal agent of fruit brown rot of Citrus worldwide and the first report of P. insolita complex in Vietnam. Results indicate that likely Vietnam is still an unexplored reservoir of Phytophthora diversity.

  11. A family of small cyclic amphipathic peptides (SCAmpPs) genes in citrus.

    PubMed

    Belknap, William R; McCue, Kent F; Harden, Leslie A; Vensel, William H; Bausher, Michael G; Stover, Ed

    2015-04-16

    Citrus represents a crop of global importance both in economic impact and significance to nutrition. Citrus production worldwide is threatened by the disease Huanglongbing (HLB), caused by the phloem-limited pathogen Candidatus Liberibacter spp.. As a source of stable HLB-resistance has yet to be identified, there is considerable interest in characterization of novel disease-associated citrus genes. A gene family of Small Cyclic Amphipathic Peptides (SCAmpPs) in citrus is described. The citrus genomes contain 100-150 SCAmpPs genes, approximately 50 of which are represented in the citrus EST database. These genes encode small ~50 residue precursor proteins that are post-translationally processed, releasing 5-10 residue cyclic peptides. The structures of the SCAmpPs genes are highly conserved, with the small coding domains interrupted by a single intron and relatively extended untranslated regions. Some family members are very highly transcribed in specific citrus tissues, as determined by representation in tissue-specific cDNA libraries. Comparison of the ESTs of related SCAmpPs revealed an unexpected evolutionary profile, consistent with targeted mutagenesis of the predicted cyclic peptide domain. The SCAmpPs genes are displayed in clusters on the citrus chromosomes, with apparent association with receptor leucine-rich repeat protein arrays. This study focused on three SCAmpPs family members with high constitutive expression in citrus phloem. Unexpectedly high sequence conservation was observed in the promoter region of two phloem-expressed SCAmpPs that encode very distinct predicted cyclic products. The processed cyclic product of one of these phloem SCAmpPs was characterized by LC-MS-MS analysis of phloem tissue, revealing properties consistent with a K(+) ionophore. The SCAmpPs amino acid composition, protein structure, expression patterns, evolutionary profile and chromosomal distribution are consistent with designation as ribosomally synthesized defense

  12. Ultrastructure of the salivary glands, alimentary canal and bacteria-like organisms in the Asian citrus psyllid, vector of citrus huanglongbing-disease bacteria

    USDA-ARS?s Scientific Manuscript database

    Several psyllids (Hemiptera: Psylloidea) are known as vectors of some economically important viral and bacterial plant pathogens. The Asian citrus psyllid (ACP, Diaphorina citri, Hemiptera, Liviidae) is the principal vector of ‘Candidatus Liberibacter asiaticus’ (Las), the putative bacterial causal ...

  13. Transgenic expression in citrus of single-chain antibody fragments specific to Citrus tristeza virus confers virus resistance.

    PubMed

    Cervera, Magdalena; Esteban, Olga; Gil, Maite; Gorris, M Teresa; Martínez, M Carmen; Peña, Leandro; Cambra, Mariano

    2010-12-01

    Citrus tristeza virus (CTV) causes one of the most destructive viral diseases of citrus worldwide. Generation of resistant citrus genotypes through genetic engineering could be a good alternative to control CTV. To study whether production of single-chain variable fragment (scFv) antibodies in citrus could interfere and immunomodulate CTV infection, transgenic Mexican lime plants expressing two different scFv constructs, separately and simultaneously, were generated. These constructs derived from the well-referenced monoclonal antibodies 3DF1 and 3CA5, specific against CTV p25 major coat protein, whose mixture is able to detect all CTV isolates characterized so far. ScFv accumulation levels were low and could be readily detected just in four transgenic lines. Twelve homogeneous and vigorous lines were propagated and CTV-challenged by graft inoculation with an aggressive CTV strain. A clear protective effect was observed in most transgenic lines, which showed resistance in up to 40-60% of propagations. Besides, both a delay in symptom appearance and attenuation of symptom intensity were observed in infected transgenic plants compared with control plants. This effect was more evident in lines carrying the 3DF1scFv transgene, being probably related to the biological functions of the epitope recognized by this antibody. This is the first report describing successful protection against a pathogen in woody transgenic plants by ectopic expression of scFv recombinant antibodies.

  14. Characterization of a Proposed Dichorhavirus Associated with the Citrus Leprosis Disease and Analysis of the Host Response

    PubMed Central

    Cruz-Jaramillo, José Luis; Ruiz-Medrano, Roberto; Rojas-Morales, Lourdes; López-Buenfil, José Abel; Morales-Galván, Oscar; Chavarín-Palacio, Claudio; Ramírez-Pool, José Abrahán; Xoconostle-Cázares, Beatriz

    2014-01-01

    The causal agents of Citrus leprosis are viruses; however, extant diagnostic methods to identify them have failed to detect known viruses in orange, mandarin, lime and bitter orange trees with severe leprosis symptoms in Mexico, an important citrus producer. Using high throughput sequencing, a virus associated with citrus leprosis was identified, belonging to the proposed Dichorhavirus genus. The virus was termed Citrus Necrotic Spot Virus (CNSV) and contains two negative-strand RNA components; virions accumulate in the cytoplasm and are associated with plasmodesmata—channels interconnecting neighboring cells—suggesting a mode of spread within the plant. The present study provides insights into the nature of this pathogen and the corresponding plant response, which is likely similar to other pathogens that do not spread systemically in plants. PMID:25004279

  15. Characterization of a proposed dichorhavirus associated with the citrus leprosis disease and analysis of the host response.

    PubMed

    Cruz-Jaramillo, José Luis; Ruiz-Medrano, Roberto; Rojas-Morales, Lourdes; López-Buenfil, José Abel; Morales-Galván, Oscar; Chavarín-Palacio, Claudio; Ramírez-Pool, José Abrahán; Xoconostle-Cázares, Beatriz

    2014-07-07

    The causal agents of Citrus leprosis are viruses; however, extant diagnostic methods to identify them have failed to detect known viruses in orange, mandarin, lime and bitter orange trees with severe leprosis symptoms in Mexico, an important citrus producer. Using high throughput sequencing, a virus associated with citrus leprosis was identified, belonging to the proposed Dichorhavirus genus. The virus was termed Citrus Necrotic Spot Virus (CNSV) and contains two negative-strand RNA components; virions accumulate in the cytoplasm and are associated with plasmodesmata-channels interconnecting neighboring cells-suggesting a mode of spread within the plant. The present study provides insights into the nature of this pathogen and the corresponding plant response, which is likely similar to other pathogens that do not spread systemically in plants.

  16. Past and future of a century old Citrus Tristeza Virus collection: A California citrus germplasm tale

    USDA-ARS?s Scientific Manuscript database

    The California Citrus Clonal Protection Program (CCPP), Riverside, CA provides a mechanism for introduction and distribution of citrus germplasm from any citrus-growing area of the world to California for use in research, variety improvement, or by industry. Citrus tristeza virus (CTV) is a serious ...

  17. Exacerbation of citrus canker by citrus leafminer, Phyllocnistis citrella in Florida

    USDA-ARS?s Scientific Manuscript database

    Citrus canker (caused by Xanthomonas citri subsp. citri, Xcc) is an important bacterial disease of citrus that is spread naturally by rain and wind. Damage to citrus leaves by the citrus leafminer (CLM) , Phyllocnistis citrella Stainton (Lepidoptera: Gracillariidae), has been shown to promote infect...

  18. Morphogenetic Alterations of Alternaria alternata Exposed to Dicarboximide Fungicide, Iprodione.

    PubMed

    Kim, Eunji; Lee, Hye Min; Kim, Young Ho

    2017-02-01

    Fungicide-resistant Alternaria alternata impede the practical control of the Alternaria diseases in crop fields. This study aimed to investigate cytological fungicide resistance mechanisms of A. alternata against dicarboximide fungicide iprodione. A. alternata isolated from cactus brown spot was cultured on potato-dextrose agar (PDA) with or without iprodione, and the fungal cultures with different growth characteristics from no, initial and full growth were observed by light and electron microscopy. Mycelia began to grow from one day after incubation (DAI) and continued to be in full growth (control-growth, Con-G) on PDA without fungicide, while on PDA with iprodione, no fungal growth (iprodione-no growth, Ipr-N) occurred for the first 3 DAI, but once the initial growth (iprodione-initial growth, Ipr-I) began at 4-5 DAI, the colonies grew and expanded continuously to be in full growth (iprodione-growth, Ipr-G), suggesting Ipr-I may be a turning moment of the morphogenetic changes resisting fungicidal toxicity. Con-G formed multicellular conidia with cell walls and septa and intact dense cytoplasm. In Ipr-N, fungal sporulation was inhibited by forming mostly undeveloped unicellular conidia with degraded and necrotic cytoplasm. However, in Ipr-I, conspicuous cellular changes occurred during sporulation by forming multicellular conidia with double layered (thickened) cell walls and accumulation of proliferated lipid bodies in the conidial cytoplasm, which may inhibit the penetration of the fungicide into conidial cells, reducing fungicide-associated toxicity, and may be utilized as energy and nutritional sources, respectively, for the further fungal growth to form mature colonies as in Ipr-G that formed multicellular conidia with cell walls and intact cytoplasm with lipid bodies as in Con-G.

  19. Morphogenetic Alterations of Alternaria alternata Exposed to Dicarboximide Fungicide, Iprodione

    PubMed Central

    Kim, Eunji; Lee, Hye Min; Kim, Young Ho

    2017-01-01

    Fungicide-resistant Alternaria alternata impede the practical control of the Alternaria diseases in crop fields. This study aimed to investigate cytological fungicide resistance mechanisms of A. alternata against dicarboximide fungicide iprodione. A. alternata isolated from cactus brown spot was cultured on potato-dextrose agar (PDA) with or without iprodione, and the fungal cultures with different growth characteristics from no, initial and full growth were observed by light and electron microscopy. Mycelia began to grow from one day after incubation (DAI) and continued to be in full growth (control-growth, Con-G) on PDA without fungicide, while on PDA with iprodione, no fungal growth (iprodione-no growth, Ipr-N) occurred for the first 3 DAI, but once the initial growth (iprodione-initial growth, Ipr-I) began at 4–5 DAI, the colonies grew and expanded continuously to be in full growth (iprodione-growth, Ipr-G), suggesting Ipr-I may be a turning moment of the morphogenetic changes resisting fungicidal toxicity. Con-G formed multicellular conidia with cell walls and septa and intact dense cytoplasm. In Ipr-N, fungal sporulation was inhibited by forming mostly undeveloped unicellular conidia with degraded and necrotic cytoplasm. However, in Ipr-I, conspicuous cellular changes occurred during sporulation by forming multicellular conidia with double layered (thickened) cell walls and accumulation of proliferated lipid bodies in the conidial cytoplasm, which may inhibit the penetration of the fungicide into conidial cells, reducing fungicide-associated toxicity, and may be utilized as energy and nutritional sources, respectively, for the further fungal growth to form mature colonies as in Ipr-G that formed multicellular conidia with cell walls and intact cytoplasm with lipid bodies as in Con-G. PMID:28167893

  20. Characterization of a sabinene synthase gene from rough lemon (Citrus jambhiri).

    PubMed

    Kohzaki, Keisuke; Gomi, Kenji; Yamasaki-Kokudo, Yumiko; Ozawa, Rika; Takabayashi, Junji; Akimitsu, Kazuya

    2009-10-15

    We previously isolated two putative monoterpene synthase genes, RlemTPS1 and RlemTPS2, from rough lemon (Citrus jambhiri) and showed that gene expression of RlemTPS2 was induced by microbial attack. The protein product of RlemTPS2 was obtained using a prokaryotic expression system, and GC and GC-MS of monoterpene synthesis by RlemTPS2 determined that RlemTPS2 encodes a sabinene synthase. Sabinene has antifungal activity toward Alternaria alternata. Furthermore, site-directed mutagenesis identified one amino acid, Ile, located at the front of the metal ion binding motif as an important residue for the product specificity of sabinene synthase.

  1. Microbes Associated with Freshly Prepared Juices of Citrus and Carrots

    PubMed Central

    Aneja, Kamal Rai; Dhiman, Romika; Aggarwal, Neeraj Kumar; Kumar, Vikas; Kaur, Manpreeet

    2014-01-01

    Fruit juices are popular drinks as they contain antioxidants, vitamins, and minerals that are essential for human being and play important role in the prevention of heart diseases, cancer, and diabetes. They contain essential nutrients which support the growth of acid tolerant bacteria, yeasts, and moulds. In the present study, we have conducted a microbiological examination of freshly prepared juices (sweet lime, orange, and carrot) by serial dilution agar plate technique. A total of 30 juice samples were examined for their microbiological quality. Twenty-five microbial species including 9 bacterial isolates, 5 yeast isolates, and 11 mould isolates were isolated from juices. Yeasts and moulds were the main cause of spoilage of juices. Aspergillus flavus and Rhodotorula mucilaginosa were observed in the maximum number of juice samples. Among bacteria Bacillus cereus and Serratia were dominant. Escherichia coli and Staphylococcus aureus were detected in few samples. Candida sp., Curvularia, Colletotrichum, and Acetobacter were observed only in citrus juice samples. Alternaria, Aspergillus terreus, A. niger, Cladosporium, and Fusarium were also observed in tested juice samples. Some of the microorganisms detected in these juice samples can cause disease in human beings, so there is need for some guidelines that can improve the quality of fruit juices. PMID:26904628

  2. Microbes Associated with Freshly Prepared Juices of Citrus and Carrots.

    PubMed

    Aneja, Kamal Rai; Dhiman, Romika; Aggarwal, Neeraj Kumar; Kumar, Vikas; Kaur, Manpreeet

    2014-01-01

    Fruit juices are popular drinks as they contain antioxidants, vitamins, and minerals that are essential for human being and play important role in the prevention of heart diseases, cancer, and diabetes. They contain essential nutrients which support the growth of acid tolerant bacteria, yeasts, and moulds. In the present study, we have conducted a microbiological examination of freshly prepared juices (sweet lime, orange, and carrot) by serial dilution agar plate technique. A total of 30 juice samples were examined for their microbiological quality. Twenty-five microbial species including 9 bacterial isolates, 5 yeast isolates, and 11 mould isolates were isolated from juices. Yeasts and moulds were the main cause of spoilage of juices. Aspergillus flavus and Rhodotorula mucilaginosa were observed in the maximum number of juice samples. Among bacteria Bacillus cereus and Serratia were dominant. Escherichia coli and Staphylococcus aureus were detected in few samples. Candida sp., Curvularia, Colletotrichum, and Acetobacter were observed only in citrus juice samples. Alternaria, Aspergillus terreus, A. niger, Cladosporium, and Fusarium were also observed in tested juice samples. Some of the microorganisms detected in these juice samples can cause disease in human beings, so there is need for some guidelines that can improve the quality of fruit juices.

  3. Antimicrobial activity of commercial citrus-based natural extracts against Escherichia coli O157:H7 isolates and mutant strains

    USDA-ARS?s Scientific Manuscript database

    Due to increasing concerns about the development of antimicrobial resistance amongst pathogenic bacteria, alternative strategies have been sought that do not use antibiotics to reduce pathogenic bacteria from foods and patients. A natural compound that has potent antimicrobial properties is citrus ...

  4. Citrus Grove Mapping

    NASA Technical Reports Server (NTRS)

    1980-01-01

    Citrus growers have long used aerial photography to inventory the number of groves in production. A new development at Kennedy Space Center, aerial mapping of groves with color infrared (CIR) film, affords an important advance in grove management by detecting and locating unhealthy trees long before they could be detected by ground survey methods. Aerial CIR photography picks up light reflected from foliage-- light not visible to the human eye--and enables differentiation between healthy and "stressed" (diseased) trees of a Florida orange/grapefruit grove. Computer aided photo interpretation techniques permit grading diseased trees lightly, moderately or severely stressed or dead. Method of grove mapping has offered advantage to growers in early disease warning, possible savings through water regulation and provision of a permanent record of grove growth patterns.

  5. Internal extracellular bacteria of Diaphorina citri Kuwayama (Hemiptera: Psyllidae), the Asian citrus psyllid.

    PubMed

    Kolora, Lakshmi D; Powell, Christopher M; Hunter, W; Bextine, B; Lauzon, C R

    2015-05-01

    The Asian Citrus Psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), is an invasive insect pest that transmits Candidatus Liberibacter spp. This insect/pathogen system was first identified in North America in the early 2000's and has become the top threat to the citrus industry. Limited options for management of this problem exist; therefore, innovative pest management strategies are being developed. In this study, we describe the first step toward a paratransgenic approach (also referred to symbiotic control) for control of the insect vector or the pathogen. Culturable bacteria from the gut of Asian Citrus Psyllids were identified using standard culture techniques followed by sequencing of the cultured microorganisms. Further, 454 pyrosequencing of the gut was performed to audit bacterial presence in order to begin to identify any relationship between psyllid symbionts and C. Liberibacter spp.

  6. Selection of Small Synthetic Antimicrobial Peptides Inhibiting Xanthomonas citri subsp. citri Causing Citrus Canker

    PubMed Central

    Choi, Jeahyuk; Park, Euiho; Lee, Se-Weon; Hyun, Jae-Wook; Baek, Kwang-Hyun

    2017-01-01

    Citrus canker disease decreases the fruit quality and yield significantly, furthermore, emerging of streptomycin-resistant pathogens threatens the citrus industry seriously because of a lack of proper control agents. Small synthetic antimicrobial peptides (AMPs) could be a promising alternative. Fourteen hexapeptides were selected by using positional scanning of synthetic peptide combinatorial libraries. Each hexapeptide showed different antimicrobial spectrum against Bacillus, Pseudomonas, Xanthomonas, and Candida species. Intriguingly, BHC10 showed bactericidal activity exclusively on Xanthomonas citri subsp. citri (Xcc), while BHC7 was none-active exclusively against two Pseudomonas spp. at concentration of 100 μg/ml suggesting potential selectivity constrained in hexapeptide frame. Three hexapeptides, BHC02, 06 and 11, showed bactericidal activities against various Xcc strains at concentration of 10 μg/ml. When they were co-infiltrated with pathogens into citrus leaves the disease progress was suppressed significantly. Further study would be needed to confirm the actual disease control capacity of the selected hexapeptides. PMID:28167892

  7. Control of virus diseases of citrus.

    PubMed

    Lee, Richard F

    2015-01-01

    Citrus is thought to have originated in Southeast Asia and horticulturally desirable clonal selections have been clonally cultivated for hundreds of years. While some citrus species have nucellar embryony, most cultivation of citrus has been by clonal propagation to ensure that propagated plants have the same traits as the parent selection. Clonal propagation also avoids juvenility, and the propagated plants produce fruit sooner. Because of the clonal propagation of citrus, citrus has accumulated a large number of viruses; many of these viruses are asymptomatic until a susceptible rootstock and/or scion is encountered. The viruses reported to occur in citrus will be summarized in this review. Methods of therapy to clean selected clones from viruses will be reviewed; the use of quarantine, clean stock, and certification programs for control of citrus viruses and other strategies to control insect spread citrus viruses, such as mild strain cross-protection and the use of pest management areas will be discussed.

  8. Application of ultraviolet-C light on oranges for the inactivation of postharvest wound pathogens

    USDA-ARS?s Scientific Manuscript database

    Germicidal effects of ultraviolet-C (UV-C) light on the postharvest wound pathogens of citrus fruits namely Penicillium digitatum and Penicillium italicum were investigated. P. digitatum and P. italicum spores were inoculated (4.00 – 4.50 log cfu/ orange) onto Washington navel oranges (Citrus sinens...

  9. Increased detection sensitivity of HLB pathogen by targeting a multi-copy gene

    USDA-ARS?s Scientific Manuscript database

    Citrus huanglongbing (HLB, yellow shoot disease) is a highly destructive disease in citrus production worldwide. “Candidatus Liberibacter asiaticus” is the pathogen of HLB based on strong evidence linking this bacterium with the disease. A single incidence of HLB has been reported in California. Alt...

  10. Phytochemical, antimicrobial, and antioxidant activities of different citrus juice concentrates.

    PubMed

    Oikeh, Ehigbai I; Omoregie, Ehimwenma S; Oviasogie, Faith E; Oriakhi, Kelly

    2016-01-01

    The search for new antimicrobial compounds is ongoing. Its importance cannot be overemphasized in an era of emerging resistant pathogenic organisms. This study therefore investigated the phytochemical composition and antioxidant and antimicrobial activities of different citrus juice concentrates. Fruit juices of Citrus tangerine (tangerine), Citrus paradisi (grape), Citrus limon (lemon), and Citrus aurantifolia (lime) were evaluated. Antimicrobial activities against five bacterial and three fungal strains were evaluated. The results revealed the presence of alkaloids, flavonoids, steroids, terpenoids, saponins, cardiac glycosides, and reducing sugars in all the juice concentrates. DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging capacities varied with tangerine and grape juices having better scavenging capacities than lemon and lime juices. Grape juice was observed to have a significantly higher (P < 0.05) ferric-reducing antioxidant potential (FRAP) value (364.2 ± 10.25 μmol/L Fe(II)/g of the extract) than the reference antioxidant, ascorbic acid (312.88 ± 5.61 μmol/L). Antimicrobial studies revealed differential antimicrobial activities against different microbial strains. Zones of inhibition ranging from 4 to 26 mm were observed for the antibacterial tests with 0-24 mm for antifungal test. Minimum inhibitory concentrations (MIC) and minimum bacteriostatic concentrations (MBC) for concentrates against bacterial strains ranged from 12.5 to 200 μg/mL. Lemon and lime juice concentrates had lower MIC and MBC values with orange and tangerine having the highest values. Minimum fungicidal concentrations ranged from 50 to 200 μg/mL. The results of this study suggest that these juice concentrates may have beneficial antimicrobial roles that can be exploited in controlling unwanted microbial growth.

  11. STAT6 regulates natural helper cell proliferation during lung inflammation initiated by Alternaria

    PubMed Central

    Khorram, Naseem; Chang, Jinny E.; Kim, Hee-Kyoo; Rosenthal, Peter; Croft, Michael; Broide, David H.

    2012-01-01

    Asthma exacerbations can be caused by a number of factors, including the fungal allergen Alternaria, which is specifically associated with severe and near-fatal attacks. The mechanisms that trigger lung responses are unclear and might vary between allergens. A comparison between Alternaria, Aspergillus, Candida, and house dust mite, all allergens in humans, showed that only Alternaria promoted immediate innate airway eosinophilia within 12 h of inhalation in nonsensitized mice. Alternaria, but not the other allergens, induced a rapid increase in airway levels of IL-33, accompanied by IL-33 receptor (IL-33R)-positive natural helper cell (NHC) production of IL-5 and IL-13. NHCs in the lung and bone marrow constitutively expressed transcription factors [GATA-3 and E26 transformation-specific sequence-1 (ETS-1)] that could allow for rapid induction of T helper type 2 (Th2) cytokines. Lung NHC numbers and proliferation (%Ki-67), but not IL-5 or GATA-3 expression, were significantly reduced in STAT6-deficient mice 3 days after one challenge with Alternaria. Alternaria induced NHC expression of the EGF receptor ligand amphiregulin (partially dependent on STAT6), as well as EGF receptor signaling in the airway epithelium. Finally, human peripheral blood NHCs (CRTH2+CD127+ lineage-negative lymphocytes) from allergic individuals highly expressed GATA-3 and ETS-1, similar to lung NHCs in mice. In summary, Alternaria-induced lung NHC proliferation and expression of amphiregulin are regulated by STAT6. In addition, NHCs in mouse and humans are primed to express Th2 cytokines through constitutive expression of GATA-3 and ETS-1. Thus several transcription factor pathways (STAT6, GATA-3, and ETS-1) may contribute to NHC proliferation and Th2-type responses in Alternaria-induced asthma. PMID:22865552

  12. Development and validation of a multiplex reverse transcription quantitative PCR (RT-qPCR) assay for the rapid detection of Citrus tristeza virus, Citrus psorosis virus, and Citrus leaf blotch virus.

    PubMed

    Osman, Fatima; Hodzic, Emir; Kwon, Sun-Jung; Wang, Jinbo; Vidalakis, Georgios

    2015-08-01

    A single real-time multiplex reverse transcription quantitative polymerase chain reaction (RT-qPCR) assay for the simultaneous detection of Citrus tristeza virus (CTV), Citrus psorosis virus (CPsV), and Citrus leaf blotch virus (CLBV) was developed and validated using three different fluorescently labeled minor groove binding qPCR probes. To increase the detection reliability, coat protein (CP) genes from large number of different isolates of CTV, CPsV and CLBV were sequenced and a multiple sequence alignment was generated with corresponding CP sequences from the GenBank and a robust multiplex RT-qPCR assay was designed. The capacity of the multiplex RT-qPCR assay in detecting the viruses was compared to singleplex RT-qPCR designed specifically for each virus and was assessed using multiple virus isolates from diverse geographical regions and citrus species as well as graft-inoculated citrus plants infected with various combination of the three viruses. No significant difference in detection limits was found and specificity was not affected by the inclusion of the three assays in a multiplex RT-qPCR reaction. Comparison of the viral load for each virus using singleplex and multiplex RT-qPCR assays, revealed no significant differences between the two assays in virus detection. No significant difference in Cq values was detected when using one-step and two-step multiplex RT-qPCR detection formats. Optimizing the RNA extraction technique for citrus tissues and testing the quality of the extracted RNA using RT-qPCR targeting the cytochrome oxidase citrus gene as an RNA specific internal control proved to generate better diagnostic assays. Results showed that the developed multiplex RT-qPCR can streamline viruses testing of citrus nursery stock by replacing three separate singleplex assays, thus reducing time and labor while retaining the same sensitivity and specificity. The three targeted RNA viruses are regulated pathogens for California's mandatory "Section 3701

  13. Citrus tristeza virus-host interactions

    USDA-ARS?s Scientific Manuscript database

    Citrus tristeza virus (CTV) is a phloem-limited virus whose natural host range is restricted to citrus and related species. Although the virus has killed millions of trees, almost destroying whole industries, and continually limits production in many citrus growing areas, most isolates are mild or s...

  14. Citrus tristeza virus-aphid interactions

    USDA-ARS?s Scientific Manuscript database

    A review chapter on aphid transmission of Citrus tristeza virus is provided for a book on “Vector-Mediated Transmission of Plant Pathogens”. Earliest uses of citrus goes back over two millennia as items of trade, gifts and medicinal compounds. Citrus propagation during this period was by seed and si...

  15. Weeping dragon, a unique ornamenal citrus

    USDA-ARS?s Scientific Manuscript database

    ‘Weeping Dragon’ is a new ornamental citrus cultivar developed by intercrossing of two unusual and unique citrus types, Poncirus trifoliata cultivated variety (cv.) Flying Dragon, and Citrus sinensis cv. ‘Cipo’. This new hybrid cultivar combines strongly contorted and weeping growth traits in a smal...

  16. Citrus Limonoids: Analysis, Bioactivity, and Biomedical Prospects

    USDA-ARS?s Scientific Manuscript database

    This publication is a review of the chemistry, biochemistry and bioactivity of limonoids occurring in citrus. The review chronologically relates the evolution of research in citrus limonoids beginning with their association with bitterness development in citrus juices. The chemical and biochemical...

  17. Botanicals, selective insecticides, and predators to control Diaphorina citri (Hemiptera: Liviidae) in citrus orchards.

    PubMed

    Khan, Azhar A; Afzal, Muhammad; Qureshi, Jawwad A; Khan, Arif M; Raza, Abubakar M

    2014-12-01

    The Asian citrus psyllid (ACP) Diaphorina citri Kuwayama vectors pathogens that cause huanglongbing (HLB) or citrus greening devastating and economically important disease present in most citrus growing regions. Young citrus shoots are required for psyllid reproduction and development. During winter citrus trees produce little or no new growth. Overwintering adults reproduce in spring on newly emerging shoots also attractive to other pests and beneficial insects. Botanicals and relatively selective insecticides could help to conserve beneficial insects and reduce pest resistance to insecticides. Sprays of Azadirachtin (Neem), Tropane (Datura), Spirotetramat, Spinetoram, and broad-spectrum Imidacloprid were evaluated to control ACP in spring and summer on 10-year-old "Kinow" Citrus reticulata Blanco trees producing new growth. Psyllid populations were high averaging 5-9 nymphs or adults per sample before treatment application. Nymphs or adults were significantly reduced to 0.5-1.5 per sample in all treatments for 3 weeks, average 61%-83% reduction. No significant reduction in ladybeetles Adalia bipunctata, Aneglei scardoni, Cheilomenes sexmaculata, and Coccinella septempunctata was observed. Syrphids, spiders and green lacewings were reduced in treated trees except with Tropane. Studies are warranted to assess impact of these predators on ACP and interaction with insecticides. Observed reduction in ACP populations may not be enough considering its reproductive potential and role in the spread of HLB. Follow-up sprays may be required to achieve additional suppression using rotations of different insecticides.

  18. The Arabidopsis thaliana-Alternaria brassicicola pathosystem: A model interaction for investigating seed transmission of necrotrophic fungi

    PubMed Central

    2012-01-01

    Background Seed transmission constitutes a major component of the parasitic cycle for several fungal pathogens. However, very little is known concerning fungal or plant genetic factors that impact seed transmission and mechanisms underlying this key biological trait have yet to be clarified. Such lack of available data could be probably explained by the absence of suitable model pathosystem to study plant-fungus interactions during the plant reproductive phase. Results Here we report on setting up a new pathosystem that could facilitate the study of fungal seed transmission. Reproductive organs of Arabidopsis thaliana were inoculated with Alternaria brassicicola conidia. Parameters (floral vs fruit route, seed collection date, plant and silique developmental stages) that could influence the seed transmission efficiency were tested to define optimal seed infection conditions. Microscopic observations revealed that the fungus penetrates siliques through cellular junctions, replum and stomata, and into seed coats either directly or through cracks. The ability of the osmosensitive fungal mutant nik1Δ3 to transmit to A. thaliana seeds was analyzed. A significant decrease in seed transmission rate was observed compared to the wild-type parental strain, confirming that a functional osmoregulation pathway is required for efficient seed transmission of the fungus. Similarly, to test the role of flavonoids in seed coat protection against pathogens, a transparent testa Arabidopsis mutant (tt4-1) not producing any flavonoid was used as host plant. Unexpectedly, tt4-1 seeds were infected to a significantly lower extent than wild-type seeds, possibly due to over-accumulation of other antimicrobial metabolites. Conclusions The Arabidopsis thaliana-Alternaria brassicicola pathosystem, that have been widely used to study plant-pathogen interactions during the vegetative phase, also proved to constitute a suitable model pathosystem for detailed analysis of plant-pathogen

  19. The Arabidopsis thaliana-Alternaria brassicicola pathosystem: A model interaction for investigating seed transmission of necrotrophic fungi.

    PubMed

    Pochon, Stephanie; Terrasson, Emmanuel; Guillemette, Thomas; Iacomi-Vasilescu, Beatrice; Georgeault, Sonia; Juchaux, Marjorie; Berruyer, Romain; Debeaujon, Isabelle; Simoneau, Philippe; Campion, Claire

    2012-05-09

    Seed transmission constitutes a major component of the parasitic cycle for several fungal pathogens. However, very little is known concerning fungal or plant genetic factors that impact seed transmission and mechanisms underlying this key biological trait have yet to be clarified. Such lack of available data could be probably explained by the absence of suitable model pathosystem to study plant-fungus interactions during the plant reproductive phase. Here we report on setting up a new pathosystem that could facilitate the study of fungal seed transmission. Reproductive organs of Arabidopsis thaliana were inoculated with Alternaria brassicicola conidia. Parameters (floral vs fruit route, seed collection date, plant and silique developmental stages) that could influence the seed transmission efficiency were tested to define optimal seed infection conditions. Microscopic observations revealed that the fungus penetrates siliques through cellular junctions, replum and stomata, and into seed coats either directly or through cracks. The ability of the osmosensitive fungal mutant nik1Δ3 to transmit to A. thaliana seeds was analyzed. A significant decrease in seed transmission rate was observed compared to the wild-type parental strain, confirming that a functional osmoregulation pathway is required for efficient seed transmission of the fungus. Similarly, to test the role of flavonoids in seed coat protection against pathogens, a transparent testa Arabidopsis mutant (tt4-1) not producing any flavonoid was used as host plant. Unexpectedly, tt4-1 seeds were infected to a significantly lower extent than wild-type seeds, possibly due to over-accumulation of other antimicrobial metabolites. The Arabidopsis thaliana-Alternaria brassicicola pathosystem, that have been widely used to study plant-pathogen interactions during the vegetative phase, also proved to constitute a suitable model pathosystem for detailed analysis of plant-pathogen interactions during the reproductive

  20. Gender differences and effect of photophase on Asian citrus psyllid (Diaphorina citri Kuwayama) feeding behavior

    USDA-ARS?s Scientific Manuscript database

    Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), thought to be primarily a phloem-feeding insect, transmits the presumptive pathogen for Huanglongbing, ‘Candidatus Liberibacter asiaticus’. Because this bacterium is restricted to the phloem and bacterial transmission is the res...

  1. Spatial Genetic Structure of Coffee-Associated Xylella fastidiosa Populations Indicates that Cross Infection Does Not Occur with Sympatric Citrus Orchards.

    PubMed

    Francisco, Carolina S; Ceresini, Paulo C; Almeida, Rodrigo P P; Coletta-Filho, Helvécio D

    2017-04-01

    Xylella fastidiosa, an economically important plant-pathogenic bacterium, infects both coffee and citrus trees in Brazil. Although X. fastidiosa in citrus is well studied, knowledge about the population structure of this bacterium infecting coffee remains unknown. Here, we studied the population structure of X. fastidiosa infecting coffee trees in São Paulo State, Brazil, in four regions where citrus is also widely cultivated. Genotyping of over 500 isolates from coffee plants using 14 genomic microsatellite markers indicated that populations were largely geographically isolated, as previously found with populations of X. fastidiosa infecting citrus. These results were supported by a clustering analysis, which indicated three major genetic groups among the four sampled regions. Overall, approximately 38% of isolates showed significant membership coefficients not related to their original geographical populations (i.e., migrants), characterizing a significant degree of genotype flow among populations. To determine whether admixture occurred between isolates infecting citrus and coffee plants, one site with citrus and coffee orchards adjacent to each other was selected; over 100 isolates were typed from each host plant. No signal of natural admixture between citrus- and coffee-infecting isolates was found; artificial cross-infection assays with representative isolates also yielded no successful cross infection. A comparison determined that X. fastidiosa populations from coffee have higher genetic diversity and allelic richness compared with citrus. The results showed that coffee and citrus X. fastidiosa populations are effectively isolated from each other and, although coffee populations are spatially structured, migration has an important role in shaping diversity.

  2. Characterization of Pseudomonas syringae pv. syringae, Causal Agent of Citrus Blast of Mandarin in Montenegro

    PubMed Central

    Ivanović, Žarko; Perović, Tatjana; Popović, Tatjana; Blagojević, Jovana; Trkulja, Nenad; Hrnčić, Snježana

    2017-01-01

    Citrus blast caused by bacterium Pseudomonas syringae is a very important disease of citrus occuring in many areas of the world, but with few data about genetic structure of the pathogen involved. Considering the above fact, this study reports genetic characterization of 43 P. syringae isolates obtained from plant tissue displaying citrus blast symptoms on mandarin (Citrus reticulata) in Montenegro, using multilocus sequence analysis of gyrB, rpoD, and gap1 gene sequences. Gene sequences from a collection of 54 reference pathotype strains of P. syringae from the Plant Associated and Environmental Microbes Database (PAMDB) was used to establish a genetic relationship with our isolates obtained from mandarin. Phylogenetic analyses of gyrB, rpoD, and gap1 gene sequences showed that P. syringae pv. syringae causes citrus blast in mandarin in Montenegro, and belongs to genomospecies 1. Genetic homogeneity of isolates suggested that the Montenegrian population might be clonal which indicates a possible common source of infection. These findings may assist in further epidemiological studies of this pathogen and for determining mandarin breeding strategies for P. syringae control. PMID:28167885

  3. Characterization of Pseudomonas syringae pv. syringae, Causal Agent of Citrus Blast of Mandarin in Montenegro.

    PubMed

    Ivanović, Žarko; Perović, Tatjana; Popović, Tatjana; Blagojević, Jovana; Trkulja, Nenad; Hrnčić, Snježana

    2017-02-01

    Citrus blast caused by bacterium Pseudomonas syringae is a very important disease of citrus occuring in many areas of the world, but with few data about genetic structure of the pathogen involved. Considering the above fact, this study reports genetic characterization of 43 P. syringae isolates obtained from plant tissue displaying citrus blast symptoms on mandarin (Citrus reticulata) in Montenegro, using multilocus sequence analysis of gyrB, rpoD, and gap1 gene sequences. Gene sequences from a collection of 54 reference pathotype strains of P. syringae from the Plant Associated and Environmental Microbes Database (PAMDB) was used to establish a genetic relationship with our isolates obtained from mandarin. Phylogenetic analyses of gyrB, rpoD, and gap1 gene sequences showed that P. syringae pv. syringae causes citrus blast in mandarin in Montenegro, and belongs to genomospecies 1. Genetic homogeneity of isolates suggested that the Montenegrian population might be clonal which indicates a possible common source of infection. These findings may assist in further epidemiological studies of this pathogen and for determining mandarin breeding strategies for P. syringae control.

  4. Dustborne Alternaria alternata antigens in U.S. homes: Results from the National Survey of Lead and Allergens in Housing

    PubMed Central

    Salo, Päivi M.; Yin, Ming; Arbes, Samuel J.; Cohn, Richard D.; Sever, Michelle; Muilenberg, Michael; Burge, Harriet A.; London, Stephanie J.; Zeldin, Darryl C.

    2005-01-01

    Background: Alternaria alternata is one of the most common fungi associated with allergic disease. However, Alternaria exposure in indoor environments is not well characterized. Objective: The primary goals of this study were to examine the prevalence of Alternaria exposure and identify independent predictors of Alternaria antigen concentrations in U.S. homes. Methods: Data for this cross-sectional study were obtained from the National Survey of Lead and Allergens in Housing. A nationally representative sample of 831 housing units in 75 different locations throughout the U.S. completed the survey. Information on housing and household characteristics was obtained by questionnaire and environmental assessments. Concentrations of Alternaria antigens in dust collected from various indoor sites were assessed with a polyclonal anti-Alternaria antibody assay. Results: Alternaria antigens were detected in most (95-99%) of the dust samples. The geometric mean concentration, reflecting the average Alternaria concentration in homes, was 4.88 μg/g (SE=0.13 μg/g). In the multivariable linear regression analysis, the age of the housing unit, geographic region, urbanization, poverty, family race, observed mold and moisture problems, use of dehumidifier, and presence of cats and dogs were independent predictors of Alternaria antigen concentrations. Less frequent cleaning and smoking indoors also contributed to higher Alternaria antigen levels in homes. Conclusion: Exposure to Alternaria alternata antigens in U.S. homes is common. Antigen levels in homes are not only influenced by regional factors but also by residential characteristics. Preventing mold and moisture problems, avoiding smoking indoors, and regular household cleaning may help reduce exposure to Alternaria antigens indoors. PMID:16159634

  5. Characterization of Alternaria strains from Argentinean blueberry, tomato, walnut and wheat.

    PubMed

    Andersen, Birgitte; Nielsen, Kristian F; Fernández Pinto, Virginia; Patriarca, Andrea

    2015-03-02

    Alternaria species have the ability to produce a variety of secondary metabolite, which plays important roles in food safety. Argentina is the second largest exporter of fresh and processed food products to Europe, however, few studies on Alternaria mycotoxins and other bioactive secondary metabolites have been carried out on Argentinean cereals, fruit and vegetables. Knowing the full chemical potential and the distribution of Alternaria spp. on crops, it is necessary to establish a toxicological risk assessment for food products for human consumption. In the present study, 87 Alternaria strains from different substrates (tomato, wheat, blueberries and walnuts) were characterized according to morphology and metabolite production. Aggressive dereplication (accurate mass, isotopic patterns and lists of all described compounds from Alternaria) was used for high-throughput evaluation of the chemical potential. Four strains belonged to the Alternaria infectoria sp.-grp., 6 to the Alternaria arborescens sp.-grp., 6 showed a sporulation pattern similar to that of "M" according to Simmons, 1 to that of Alternaria vaccinii, and the remaining 70 constituted a diverse group belonging to morphological groups "G" and "H". The cluster analysis yielded 16 almost identical dendrograms and grouped the Alternaria strains into four clusters and 11 singletons and outlier groups. The chemical analysis showed that AOH and AME were the most common metabolites produced, followed by TEN, ALXs and TeA. The A. infectoria sp.-grp. had no metabolites in common with the rest of the strains. Several secondary metabolites isolated from large-spored Alternaria species or other fungal genera were detected, such as dehydrocurvularin, pyrenochaetic acid and alternarienonic acid. The strains isolated from tomato produced lower amounts of metabolites than strains from blueberries, walnut and wheat, although individual strains from tomato produced the highest amount of some metabolites. The A

  6. [The importance of genus Alternaria in mycotoxins production and human diseases].

    PubMed

    Pavón Moreno, M Á; González Alonso, I; Martín de Santos, R; García Lacarra, T

    2012-01-01

    Alternaría is a cosmopolitan fungal genus that includes saprophytic, endophytic and pathogenic species, widely distributed in soil and organic matter in decomposition. Plant pathogenic species affect cereals, vegetables and fruit crops in the field and during storage. Alternaría spp. contamination is responsible for some of the world's most devastating plant diseases, causing serious reduction of crop yields and considerable economic losses. Alternaría species produce more than 70 secondary metabolites which are toxic to plants, and some of these phytotoxins have been chemically characterised and reported to act as mycotoxins to humans and animals. Exposure to Alternaría spp. toxins has been linked to a variety of adverse effects on human and animal health, including genotoxic, mutagenic, carcinogenic and cytotoxic effects. Alternaría spp. mycotoxins have been isolated from fruits (apple, pear, melon, apricot, grapes, raisins, strawberry, olive, citrus fruits and dried figs), vegetables (tomato, pepper and carrot) and tubers (potato), as well as from several processed foodstuffs manufactured with damaged raw materials (juices, preserves, sauces, etc.). Moreover, Alternaría spp. are frequently associated with allergic reactions in sensitized individuals.

  7. LRR-RLK family from two Citrus species: genome-wide identification and evolutionary aspects.

    PubMed

    Magalhães, Diogo M; Scholte, Larissa L S; Silva, Nicholas V; Oliveira, Guilherme C; Zipfel, Cyril; Takita, Marco A; De Souza, Alessandra A

    2016-08-12

    Leucine-rich repeat receptor-like kinases (LRR-RLKs) represent the largest subfamily of plant RLKs. The functions of most LRR-RLKs have remained undiscovered, and a few that have been experimentally characterized have been shown to have important roles in growth and development as well as in defense responses. Although RLK subfamilies have been previously studied in many plants, no comprehensive study has been performed on this gene family in Citrus species, which have high economic importance and are frequent targets for emerging pathogens. In this study, we performed in silico analysis to identify and classify LRR-RLK homologues in the predicted proteomes of Citrus clementina (clementine) and Citrus sinensis (sweet orange). In addition, we used large-scale phylogenetic approaches to elucidate the evolutionary relationships of the LRR-RLKs and further narrowed the analysis to the LRR-XII group, which contains several previously described cell surface immune receptors. We built integrative protein signature databases for Citrus clementina and Citrus sinensis using all predicted protein sequences obtained from whole genomes. A total of 300 and 297 proteins were identified as LRR-RLKs in C. clementina and C. sinensis, respectively. Maximum-likelihood phylogenetic trees were estimated using Arabidopsis LRR-RLK as a template and they allowed us to classify Citrus LRR-RLKs into 16 groups. The LRR-XII group showed a remarkable expansion, containing approximately 150 paralogs encoded in each Citrus genome. Phylogenetic analysis also demonstrated the existence of two distinct LRR-XII clades, each one constituted mainly by RD and non-RD kinases. We identified 68 orthologous pairs from the C. clementina and C. sinensis LRR-XII genes. In addition, among the paralogs, we identified a subset of 78 and 62 clustered genes probably derived from tandem duplication events in the genomes of C. clementina and C. sinensis, respectively. This work provided the first comprehensive

  8. Systemic jasmonic acid modulation in mycorrhizal tomato plants and its role in induced resistance against Alternaria alternata.

    PubMed

    Nair, A; Kolet, S P; Thulasiram, H V; Bhargava, S

    2015-05-01

    Tomato plants colonised with the arbuscular mycorrhizal (AM) fungus Glomus fasciculatum show systemic induced resistance to the foliar pathogen Alternaria alternata, as observed in interactions of other AM-colonised plants with a range of pathogens. The role of jasmonic (JA) and salicylic (SA) acid in expression of this mycorrhiza-induced resistance (MIR) against A. alternata was studied by measuring: (i) activity of enzymes reported to be involved in their biosynthesis, namely lipoxygenase (LOX) and phenylammonia lyase (PAL); and (ii) levels of methyl jasmonate (MeJA) and SA. Transcript abundance of some defence genes associated with JA and SA response pathways were also studied. Both LOX and PAL activity increased twofold in response to pathogen application to control plants. AM-colonised plants had three-fold higher LOX activity compared to control plants, but unlike controls, this did not increase further in response to pathogen application. Higher LOX activity in AM-colonised plants correlated with four-fold higher MeJA in leaves of AM-colonised plants compared to controls. Treatment of plants with the JA biosynthesis inhibitor salicylhydroxamic acid (SHAM) led to 50% lower MeJA in both control and AM-colonised plants and correlated with increased susceptibility to A. alternata, suggesting a causal role for JA in expression of MIR against the pathogen. Genes involved in JA biosynthesis (OPR3) and response (COI1) showed six- and 42-fold higher expression, respectively, in leaves of AM-colonised plants compared to controls. AM-colonised plants also showed increased expression of the SA response gene PR1 and that of the wound-inducible polypeptide prosystemin. Our results suggest that the systemic increase in JA in response to AM colonisation plays a key role in expression of MIR against A. alternata.

  9. Spectral sensitivity of the Asian citrus psyllid, Diaphorina citri

    USDA-ARS?s Scientific Manuscript database

    The Asian Citrus psyllid, Diaphorina citri, as a vector of the bacteria causing citrus greening, is considered one of the most important citrus pests globally. Movement of infected psyllids onto uninfected young citrus remains a key concern for the maintenance of citrus production. Attraction of d...

  10. Induced resistance against the Asian citrus psyllid, Diaphorina citri, by β-aminobutyric acid in citrus.

    PubMed

    Tiwari, Siddharth; Meyer, Wendy L; Stelinski, Lukasz L

    2013-10-01

    β-Aminobutyric acid (BABA) is known to induce resistance to microbial pathogens, nematodes and insects in several host plant/pest systems. The present study was undertaken to determine whether a similar effect of BABA occurred against the Asian citrus psyllid, Diaphorina citri Kuwayama, in citrus. A 25 mM drench application of BABA significantly reduced the number of eggs/plant as compared with a water control, whereas 200 and 100 mM applications of BABA reduced the numbers of nymphs/plant and adults/plants, respectively. A 5 mM foliar application of BABA significantly reduced the number of adults but not eggs or nymphs when compared with a water control treatment. In addition, leaf-dip bioassays using various concentrations (25–500 mM) of BABA indicated no direct toxic effect on 2nd and 5th instar nymphs or adult D. citri. BABA-treated plants were characterized by significantly lower levels of iron, magnesium, phosphorus, sodium, sulfur and zinc as compared with control plants. The expression level of the PR-2 gene (β-1,3-glucanase) in BABA-treated plants that were also damaged by D. citri adult feeding was significantly higher than in plants exposed to BABA, D. citri feeding alone or control plants. Our results indicate the potential for using BABA as a systemic acquired resistance management tool for D. citri.

  11. Transcription Factor Amr1 Induces Melanin Biosynthesis and Suppresses Virulence in Alternaria brassicicola

    SciTech Connect

    Cho, Yangrae; Srivastava, Akhil; Ohm, Robin A.; Lawrence, Christopher B.; Wang, Koon-Hui; Grigoriev, Igor V.; Marahatta, Sharadchandra P.

    2012-05-01

    Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Several A. brassicicola genes have been characterized as affecting pathogenesis of Brassica species. To study regulatory mechanisms of pathogenesis, we mined 421 genes in silico encoding putative transcription factors in a machine-annotated, draft genome sequence of A. brassicicola. In this study, targeted gene disruption mutants for 117 of the transcription factor genes were produced and screened. Three of these genes were associated with pathogenesis. Disruption mutants of one gene (AbPacC) were nonpathogenic and another gene (AbVf8) caused lesions less than half the diameter of wild-type lesions. Unexpectedly, mutants of the third gene, Amr1, caused lesions with a two-fold larger diameter than the wild type and complementation mutants. Amr1 is a homolog of Cmr1, a transcription factor that regulates melanin biosynthesis in several fungi. We created gene deletion mutants of ?amr1 and characterized their phenotypes. The ?amr1 mutants used pectin as a carbon source more efficiently than the wild type, were melanin-deficient, and more sensitive to UV light and glucanase digestion. The AMR1 protein was localized in the nuclei of hyphae and in highly melanized conidia during the late stage of plant pathogenesis. RNA-seq analysis revealed that three genes in the melanin biosynthesis pathway, along with the deleted Amr1 gene, were expressed at low levels in the mutants. In contrast, many hydrolytic enzyme-coding genes were expressed at higher levels in the mutants than in the wild type during pathogenesis. The results of this study suggested that a gene important for survival in nature negatively affected virulence, probably by a less efficient use of plant cell-wall materials. We speculate that the functions of the Amr1 gene are important to the success of A. brassicicola as a competitive saprophyte and plant parasite.

  12. Transcription Factor Amr1 Induces Melanin Biosynthesis and Suppresses Virulence in Alternaria brassicicola

    PubMed Central

    Cho, Yangrae; Srivastava, Akhil; Ohm, Robin A.; Lawrence, Christopher B.; Wang, Koon-Hui; Grigoriev, Igor V.; Marahatta, Sharadchandra P.

    2012-01-01

    Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Several A. brassicicola genes have been characterized as affecting pathogenesis of Brassica species. To study regulatory mechanisms of pathogenesis, we mined 421 genes in silico encoding putative transcription factors in a machine-annotated, draft genome sequence of A. brassicicola. In this study, targeted gene disruption mutants for 117 of the transcription factor genes were produced and screened. Three of these genes were associated with pathogenesis. Disruption mutants of one gene (AbPacC) were nonpathogenic and another gene (AbVf8) caused lesions less than half the diameter of wild-type lesions. Unexpectedly, mutants of the third gene, Amr1, caused lesions with a two-fold larger diameter than the wild type and complementation mutants. Amr1 is a homolog of Cmr1, a transcription factor that regulates melanin biosynthesis in several fungi. We created gene deletion mutants of Δamr1 and characterized their phenotypes. The Δamr1 mutants used pectin as a carbon source more efficiently than the wild type, were melanin-deficient, and more sensitive to UV light and glucanase digestion. The AMR1 protein was localized in the nuclei of hyphae and in highly melanized conidia during the late stage of plant pathogenesis. RNA-seq analysis revealed that three genes in the melanin biosynthesis pathway, along with the deleted Amr1 gene, were expressed at low levels in the mutants. In contrast, many hydrolytic enzyme-coding genes were expressed at higher levels in the mutants than in the wild type during pathogenesis. The results of this study suggested that a gene important for survival in nature negatively affected virulence, probably by a less efficient use of plant cell-wall materials. We speculate that the functions of the Amr1 gene are important to the success of A. brassicicola as a competitive saprophyte and plant parasite. PMID:23133370

  13. Dehydrin-like proteins in the necrotrophic fungus Alternaria brassicicola have a role in plant pathogenesis and stress response.

    PubMed

    Pochon, Stéphanie; Simoneau, Philippe; Pigné, Sandrine; Balidas, Samuel; Bataillé-Simoneau, Nelly; Campion, Claire; Jaspard, Emmanuel; Calmes, Benoît; Hamon, Bruno; Berruyer, Romain; Juchaux, Marjorie; Guillemette, Thomas

    2013-01-01

    In this study, the roles of fungal dehydrin-like proteins in pathogenicity and protection against environmental stresses were investigated in the necrotrophic seed-borne fungus Alternaria brassicicola. Three proteins (called AbDhn1, AbDhn2 and AbDhn3), harbouring the asparagine-proline-arginine (DPR) signature pattern and sharing the characteristic features of fungal dehydrin-like proteins, were identified in the A. brassicicola genome. The expression of these genes was induced in response to various stresses and found to be regulated by the AbHog1 mitogen-activated protein kinase (MAPK) pathway. A knock-out approach showed that dehydrin-like proteins have an impact mainly on oxidative stress tolerance and on conidial survival upon exposure to high and freezing temperatures. The subcellular localization revealed that AbDhn1 and AbDhn2 were associated with peroxisomes, which is consistent with a possible perturbation of protective mechanisms to counteract oxidative stress and maintain the redox balance in AbDhn mutants. Finally, we show that the double deletion mutant ΔΔabdhn1-abdhn2 was highly compromised in its pathogenicity. By comparison to the wild-type, this mutant exhibited lower aggressiveness on B. oleracea leaves and a reduced capacity to be transmitted to Arabidopsis seeds via siliques. The double mutant was also affected with respect to conidiation, another crucial step in the epidemiology of the disease.

  14. Natural Occurrence of Alternaria Toxins in the 2015 Wheat from Anhui Province, China

    PubMed Central

    Xu, Wenjing; Han, Xiaomin; Li, Fengqin; Zhang, Lishi

    2016-01-01

    The exposure to Alternaria toxins from grain and grain-based products has been reported to be related to human esophageal cancer in China. In this study, a total of 370 freshly harvested wheat kernel samples collected from Anhui province of China in 2015 were analyzed for the four Alternaria toxins tenuazonic acid (TeA), tentoxin (TEN), alternariol (AOH) and alternariol monomethyl ether (AME) by high performance liquid chromatography-tandem mass spectrometry method (HPLC-MS/MS). TeA was the predominant toxin detected followed by TEN, AOH and AME. The concentrations of the four Alternaria toxins varied geographically. The samples from Fuyang district showed higher TEN concentration levels than the other regions studied (p < 0.05). Furthermore, 95% (352/370) of the wheat samples were positive for more than one type of Alternaria toxins. Positive correlation was observed between concentration levels of TeA and TEN, AOH and AME, TeA and AOH, and the total dibenzopyrone derivatives (AOH + AME) and TeA. Results indicate that there is a need to set the tolerance limit for Alternaria toxins in China, and more data on the contamination of these toxins in agro-products is required. PMID:27792182

  15. Alternaria toxins in wheat during the 2004 to 2005 Argentinean harvest.

    PubMed

    Azcarate, M P; Patriarca, A; Terminiello, L; Fernández Pinto, V

    2008-06-01

    The natural occurrence of Alternaria mycotoxins in Argentinean wheat from the zone 5 South during the 2004 to 2005 harvest was investigated in 64 wheat samples. All samples were highly contaminated with a wide range of fungal species. Alternaria was found as the main component of the mycota, with an infection percentage of 100%. Three mycotoxins produced by species of Alternaria were determined in wheat: alternariol, alternariol monomethyl ether, and tenuazonic acid. Alternariol was detected in 4 (6%) of 64 samples, with a range of 645 to 1,388 microg/kg (mean of 1,054 microg/kg); alternariol monomethyl ether, with a range of 566 to 7,451 microg/kg (mean of 2,118 microg/kg) in 15 (23%) of 64 samples; and tenuazonic acid in 12 (19%) of 64 samples, with a range of 1,001 to 8,814 microg/kg (mean, 2,313 microg/kg). Alternariol monomethyl ether was the predominant toxin, but tenuazonic acid was detected in higher concentrations. Alternariol was present in fewer samples and in lower levels than were the other toxins. Tenuazonic acid and alternariol monomethyl ether occurred together in four samples, while tenuazonic acid and alternariol co-occurred in one sample. This the first report of the natural occurrence of Alternaria mycotoxins in Argentinean wheat. Toxin levels were high, probably due to the heavy infection with Alternaria species found in the samples.

  16. Isolation and characterization of an Isaria fumosorosea isolate infecting the Asian citrus psyllid in Florida.

    PubMed

    Meyer, Jason M; Hoy, Marjorie A; Boucias, Drion G

    2008-09-01

    A fungal pathogen that killed adult Diaphorina citri Kuwayama (Asian citrus psyllid) (Hemiptera: Psyllidae) in Florida citrus groves during the fall of 2005 was identified and characterized. Investigation of this pathogen is important because D. citri vectors citrus greening disease (Huanglongbing), which was reported in Florida in 2005. The morphological and genetic data generated herein support identification of the fungus as Isaria fumosorosea Wize (Ifr) (=Paecilomyces fumosoroseus) (Hypocreales: Cordycipitaceae) from the Asian citrus psyllid (Ifr AsCP). Koch's postulates were fulfilled after the fungus was isolated in vitro and transmitted to healthy psyllids, which then exhibited a diseased-phenotype similar to that observed in the field. Both in vitro growth characteristics and two Ifr AsCP-specific molecular markers discriminated the psyllid pathogen from another local Ifr isolate, Ifr 97 Apopka. These molecular markers will be useful to track the dynamics of this disease in D. citri populations. The potential for utilizing Ifr to complement existing psyllid pest management strategies is discussed.

  17. Isolation and characterization of the grain mold fungi, Cochliobolus and Alternaria spp., from sorghum using semi-selective media and DNA sequence analyses

    USDA-ARS?s Scientific Manuscript database

    Mold diseases, caused by fungal complexes including Alternaria, Cochliobolus and Fusarium species, limit sorghum grain production. Media were tested by plating Fusarium thapsinum, Alternaria sp. and Curvularia lunata, individually and competitively. Dichloran chloramphenicol rose bengal (DRBC) and m...

  18. Site Plan & Site Section of Citrus Landscape (Showing Relationship ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Site Plan & Site Section of Citrus Landscape (Showing Relationship of Victoria Avenue to Citrus Groves) - Arlington Heights Citrus Landscape, Southwestern portion of city of Riverside, Riverside, Riverside County, CA

  19. In vitro activity of thimerosal against ocular pathogenic fungi.

    PubMed

    Xu, Yan; Pang, Guangren; Zhao, Dongqing; Gao, Chuanwen; Zhou, Lutan; Sun, Shengtao; Wang, Bingliang

    2010-01-01

    The in vitro activity of thimerosal versus those of amphotericin B and natamycin was assessed against 244 ocular fungal isolates. The activity of thimerosal against Fusarium spp., Aspergillus spp., and Alternaria alternata was 256 times, 512 times, and 128 times, respectively, greater than that of natamycin and 64 times, 32 times, and 32 times, respectively, greater than that of amphotericin B. Thimerosal's antifungal activity was significantly superior to those of amphotericin B and natamycin against ocular pathogenic fungi in vitro.

  20. In Vitro Activity of Thimerosal against Ocular Pathogenic Fungi▿

    PubMed Central

    Xu, Yan; Pang, Guangren; Zhao, Dongqing; Gao, Chuanwen; Zhou, Lutan; Sun, Shengtao; Wang, Bingliang

    2010-01-01

    The in vitro activity of thimerosal versus those of amphotericin B and natamycin was assessed against 244 ocular fungal isolates. The activity of thimerosal against Fusarium spp., Aspergillus spp., and Alternaria alternata was 256 times, 512 times, and 128 times, respectively, greater than that of natamycin and 64 times, 32 times, and 32 times, respectively, greater than that of amphotericin B. Thimerosal's antifungal activity was significantly superior to those of amphotericin B and natamycin against ocular pathogenic fungi in vitro. PMID:19841144

  1. Chipping citrus wood for gasification

    SciTech Connect

    Churchill, D.B.; Hedden, S.L.; Whitney, J.D.; Shaw, L.N.

    1985-01-01

    Non-productive citrus trees were chipped with a portable fly-wheel-type chipper powered by a 45 kW engine. Chips were air dried under an open shed to 14% (w.b.) moisture content. By weight, approximately 50% of the total tree could be made into usable chips. The root system averaged 36% of the total tree weight.

  2. 7 CFR 301.76-3 - Quarantined areas; citrus greening and Asian citrus psyllid.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ....usda.gov/services/report_pest_disease/report_pest_disease.shtml). After a change is made to the... interstate spread of citrus greening or Asian citrus psyllid. (c) Criteria for designation of a State, or a...

  3. First report of QoI resistance in Alternaria spp. infecting sugar beet (Beta vulgaris) in Michigan, USA

    USDA-ARS?s Scientific Manuscript database

    Alternaria leaf spot (ALS) of sugar beet is caused by Alternaria spp. in the A. alternata species complex. ALS is common wherever sugar beet is grown, but historically has been a minor issue for sugar beet production in the USA with damage usually not affecting crop yield significantly. Occurrence o...

  4. Synergistic estrogenic effects of Fusarium and Alternaria mycotoxins in vitro.

    PubMed

    Vejdovszky, Katharina; Hahn, Kathrin; Braun, Dominik; Warth, Benedikt; Marko, Doris

    2017-03-01

    Mycotoxins are toxic secondary metabolites formed by various fungal species that are found as natural contaminants in food. This very heterogeneous group of compounds triggers multiple toxic mechanisms, including endocrine disruptive potential. Current risk assessment of mycotoxins, as for most chemical substances, is based on the effects of single compounds. However, concern on a potential enhancement of risks by interactions of single substances in naturally occurring mixtures has greatly increased recently. In this study, the combinatory effects of three mycoestrogens were investigated in detail. This includes the endocrine disruptors zearalenone (ZEN) and α-zearalenol (α-ZEL) produced by Fusarium fungi and alternariol (AOH), a cytotoxic and estrogenic mycotoxin formed by Alternaria species. For evaluation of effects, estrogen-dependent activation of alkaline phosphatase (AlP) and cell proliferation were tested in the adenocarcinoma cell line Ishikawa. The estrogenic potential varied among the single substances. Half maximum effect concentrations (EC50) for AlP activation were evaluated for α-ZEL, ZEN and AOH as 37 pM, 562 pM and 995 nM, respectively. All three mycotoxins were found to act as partial agonists. The majority of binary combinations, even at very low concentrations in the case of α-ZEL, showed strong synergism in the AlP assay. These potentiating phenomena of mycotoxin mixtures highlight the urgent need to incorporate combinatory effects into future risk assessment, especially when endocrine disruptors are involved. To the best of our knowledge, this study presents the first investigation on synergistic effects of mycoestrogens.

  5. Liquid chromatographic determination of Alternaria toxins in carrots.

    PubMed

    Solfrizzo, Michele; De Girolamo, Annalisa; Vitti, Carolina; Visconti, Angelo; van den Bulk, Ruud

    2004-01-01

    A liquid chromatographic (LC) method was developed for the determination of Alternaria radicina and A. alternata toxins in carrots. Toxins were extracted from carrot with an acidified mixture of water-methanol-acetonitrile. The filtered extract was divided in 2 parts that were purified by solid-phase extraction on a C18 column for the analysis of radicinin (RAD), altertoxin-I (ATX-I), alternariol (AOH), and alternariol methyl ether (AME), and on a polymeric Oasis HLB column for tenuazonic acid (TeA). Toxins were quantified by reversed-phase LC with UV diode array detection by using 2 consecutive isocratic mixtures of acetonitrile-sodium dihydrogen phosphate solution. Mean recoveries of TeA, ATX-I, AME, RAD, and AOH from carrots spiked at levels between 0.5 and 3.0 microg/g were 69, 71, 90, 36, and 78%, with mean within-laboratory repeatability of 14, 5, 4, 6, and 18%, respectively. The mean between-laboratory reproducibilities for the determination of TeA, ATX-I, AME, and RAD in spiked samples were 25, 22, 6, and 12%, respectively. Limits of detection (signal-to-noise ratio of 3) for RAD, TeA, ATX-I, AME, and AOH were 0.006, 0.02, 0.02, 0.01, and 0.005 microg/g, respectively. RAD was detected (0.16-13.9 microg/g) in 3 out of 266 carrot samples produced under organic conditions in 3 European locations, whereas A. alternata mycotoxins were not found in any tested samples.

  6. Signal transduction in lemon seedlings in the hypersensitive response against Alternaria alternata: participation of calmodulin, G-protein and protein kinases.

    PubMed

    Ortega, Ximena; Polanco, Rubén; Castañeda, Patricia; Perez, Luz M

    2002-01-01

    The development of an effective hypersensitive response (HR) in any plant system relies, not only in their gene composition and expression, but also on an effective and rapid signal transduction system. Lemon seedlings induce the phenylpropanoid pathway, which results in the de novo biosynthesis of the phytoalexin scoparone, as part of the hypersensitive response against Alternaria alternata. In order to elucidate some of the signaling elements that participate in the development of HR in lemon seedlings, we used several compounds that are known as activators or inhibitors of signal transduction elements in plants or in animal cells. Lemon seedlings treated either with cholera toxin or with phorbol 12-myristate 13-acetate (PMA), in the absence of A. alternata induced phenylalanine ammonia-lyase (PAL, E. C. 4.3.1.5) and the synthesis of scoparone, suggesting the participation of a G-protein and of a serine/threonine kinase, respectively, in signal transduction. The use of trifluoperazine (TFP), W-7, staurosporine, lavendustin A or 2,5-dihydroximethyl cinnamate (DHMC) prevented PAL induction as well as scoparone biosynthesis in response to the fungal inoculation, thus allowing us to infer the participation of Calmodulin (CaM), of serine/threonine and of tyrosine protein kinases (TPK) for signal transduction in Citrus limon in response to A. alternata.

  7. A Phagostimulant Blend for the Asian Citrus Psyllid.

    PubMed

    Lapointe, Stephen L; Hall, David G; George, Justin

    2016-09-01

    Chemical cues that elicit orientation by the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Liviidae), are of interest because it is the primary vector of the causal pathogen of citrus greening disease. Non-pesticidal control methods for D. citri remain a high priority for the citrus industry. While searching for semiochemicals that may be involved in orientation to host plants, we previously identified a blend of formic and acetic acids that stimulated substrate probing by D. citri. Here, we applied geometric mixture designs and response surface modeling to identify and optimize a 3-component blend that further increased the number of salivary sheaths produced by D. citri on a wax substrate containing a 3.5:1.6:1 blend of formic acid, acetic acid, and p-cymene, respectively. No evidence was found for remote orientation by D. citri adults through olfaction to the phagostimulant blends. Increased probing in response to the presence of phagostimulants in the wax matrix occurred after contact with the substrate. Yellow wax beads always attracted more D. citri adults and received more probes compared with white wax beads. Yellow beads containing the 3-component blend of phagostimulants were probed by D. citri 2 to 3 times more often compared with yellow beads alone. The phagostimulant effect also was tested by covering wax beads containing the 3-component blend with a plastic film to minimize olfaction or contact chemoreception by antennation. The plastic film did not affect the probing response, thus suggesting that chemosensation was associated with mouthparts and not olfactory receptors. Salivary sheaths produced in wax beads containing the phagostimulant blend were 4.5 times longer than sheaths produced in beads without tastants. This phenomenon might be used to improve a trap, design an attract-and-kill product, or enhance other means of managing D. citri and citrus greening disease.

  8. Cutaneous infection with Alternaria triticina in a Bilateral lung transplant recipient.

    PubMed

    González-Vela, M C; Armesto, S; Unda-Villafuerte, F; Val-Bernal, J F

    2014-10-01

    We report the case of a 60-year-old man who was receiving immunosuppressive therapy for a bilateral lung transplant and presented with a crusted, violaceous plaque on the left hand. Based on histopathology and microbiological culture the patient was diagnosed with infection by Alternaria species. Treatment with itraconazole led to complete resolution of the skin lesion. Forty months later he developed four reddish, nodular, skin lesions on the left leg. Analysis of a biopsy from one of these lesions using histopathologic and molecular techniques identified a mold that shared 98% homology with a strain of Alternaria triticina. Alternaria species belong to a group of dematiaceous fungi that cause opportunistic infections in humans. The incidence of these infections is increasing, mainly in transplant centers. To the best of our knowledge, this is the first reported case of a human infection caused by A. triticina. Copyright © 2013 Elsevier España, S.L.U. and AEDV. All rights reserved.

  9. Impact of the UPR on the virulence of the plant fungal pathogen A. brassicicola

    PubMed Central

    Guillemette, Thomas; Calmes, Benoit; Simoneau, Philippe

    2014-01-01

    The fungal genus Alternaria contains many destructive plant pathogens, including Alternaria brassicicola, which causes black spot disease on a wide range of Brassicaceae plants and which is routinely used as a model necrotrophic pathogen in studies with Arabidopsis thaliana. During host infection, many fungal proteins that are critical for disease progression are processed in the endoplasmic reticulum (ER)/Golgi system and secreted in planta. The unfolded protein response (UPR) is an essential part of ER protein quality control that ensures efficient maturation of secreted and membrane-bound proteins in eukaryotes. This review highlights the importance of the UPR signaling pathway with respect to the ability of A. brassicicola to efficiently accomplish key steps of its pathogenic life cycle. Understanding the pathogenicity mechanisms that fungi uses during infection is crucial for the development of new antifungal therapies. Therefore the UPR pathway has emerged as a promising drug target for plant disease control. PMID:24189567

  10. Development of rapid, sensitive and non-radioactive tissue-blot diagnostic method for the detection of citrus greening.

    PubMed

    Nageswara-Rao, Madhugiri; Miyata, Shin-Ichi; Ghosh, Dilip; Irey, Mike; Garnsey, Stephen M; Gowda, Siddarame

    2013-01-01

    Citrus huanglongbing (HLB or citrus greening) is one of the most devastating diseases of citrus worldwide. The disease is caused by Gram-negative, phloem-limited α-proteobacterium, 'Candidatus Liberibacter asiaticus', vectored by the psyllid, Diaphorina citri Kuwayama. Citrus plants infected by the HLB bacterium may not show visible symptoms sometimes for years following infection and non-uniform distribution within the tree makes the detection of the pathogen very difficult. Efficient management of HLB disease requires rapid and sensitive detection early in the infection followed by eradication of the source of pathogen and the vector. The polymerase chain reaction (PCR) based method is most commonly employed for screening the infected/suspected HLB plants and psyllids. This is time consuming, cumbersome and not practical for screening large number of samples in the field. To overcome this, we developed a simple, sensitive, non-radioactive, tissue-blot diagnostic method for early detection and screening of HLB disease. Digoxigenin labeled molecular probes specific to 'Ca. L. asiaticus' nucleotide sequences have been developed and used for the detection of the pathogen of the HLB disease. The copy number of the target genes was also assessed using real-time PCR experiments and the optimized real-time PCR protocol allowed positive 'Ca. L. asiaticus' detection in citrus samples infected with 'Ca. L. asiaticus' bacterium. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Thymus vulgaris essential oil and thymol against Alternaria alternata (Fr.) Keissler: effects on growth, viability, early infection and cellular mode of action.

    PubMed

    Perina, Fabiano J; Amaral, Douglas C; Fernandes, Rafael S; Labory, Claudia Rg; Teixeira, Glauco A; Alves, Eduardo

    2015-10-01

    In initial assays, Thymus vulgaris essential oil (TEO) has demonstrated activity against several plant-pathogenic fungi and has reduced the fungal diseases to levels comparable with commercial fungicides. Thus, the goal of this work was to identify the mode of action in fungi of TEO and its major compound thymol (TOH) at the cellular level using an ultrastructure approach. TEO from leaves and TOH had minimum inhibitory concentrations (MICs) of 500 and 250 µg mL(-1) respectively against A. alternata; under the same conditions, MIC for a commercial fungicide was 1250 µg mL(-1) . Ultrastructure analysis showed that TOH phenolic substance prevented fungal growth, reduced fungal viability and prevented the penetration in fruits by a cell wall/plasma membrane interference mode of action with organelles targeted for destruction in the cytoplasm. Such mode of action differs from protective and preventive-curative commercial fungicides used as pattern control. These findings suggest that TOH was responsible for the antifungal activity of TEO. Therefore, both the essential oil and its major substance have potential for use in the development of new phenolic structures and analogues to control Alternaria brown spot disease caused by Alternaria alternata. © 2014 Society of Chemical Industry.

  12. Development of a Semi-nested PCR-Based Method for Specific and Rapid Detection of Alternaria solani Causing Potato Early Blight in Soil.

    PubMed

    Gu, Qing; Yang, Zhi-Hui; Zhao, Dong-Mei; Zhang, Dai; Wang, Qian; Ma, Li-Song; Zhu, Jie-Hua

    2017-09-01

    Early blight, caused by Alternaria solani, is one of the most devastating diseases of potato that causes severe yield loss worldwide. The infected potato debris existed in the soil serve as the initial infection sources for the next growing potato. Current identification of A. solani in soil relies primarily on cultural and morphological characteristics, which are time-consuming and inaccurate. In this study, a semi-nested PCR method was developed using primers based on internal transcribed spacer region that is specific to A. solani. 20 isolates including 6 Alternaria species and 10 other species of common potato pathogens were used to examine the specificity of the primers. The primer set ptAsQ-F/ptAs-R was highly specific to A. solani, as a product of 251 bp was amplified only from A. solani isolates and no amplification signal was observed from other tested species. The sensitivity of this method determined using A. solani genomic DNA was 10 fg. This PCR assay was also successfully employed to detect A. solani in soil with the detection sensitivity of one conidia spore in 0.5 g of soil. To the best of our knowledge, this is the first report of molecular detection of A. solani in soil, which provides a useful tool for early and rapid detection of early blight in soil before next growing season.

  13. Application of natural colorants on citrus fruit as alternatives to Citrus Red II

    USDA-ARS?s Scientific Manuscript database

    Warm field temperatures can often result in poor peel color of some citrus varieties, especially early in the harvest season. Under these conditions, Florida oranges, temples, tangelos, and K-Early citrus fruit are allowed to be treated with Citrus Red No.2 (CR2) to help produce a more acceptable pe...

  14. Molecular characterization and phylogenetic analysis of Citrus viroid VI variants from citrus in China

    USDA-ARS?s Scientific Manuscript database

    Citrus viroid VI (CVd-VI) was originally found from citrus and persimmon in Japan. We report here the identification and molecular characterization of CVd-VI from four production regions of China. A total of 90 cDNA clones from nine infected citrus cultivars were sequenced. The sequence homologies o...

  15. Citrus Research Board-sponsored review of the University of California Riverside citrus breeding

    USDA-ARS?s Scientific Manuscript database

    In October 2015 the Citrus Research Board (CRB) assembled a panel of experts to review the Citrus Research Board-sponsored Citrus Research and Genetics Programs at University of California Riverside (UCR). The panel consisted of: Gennaro Fazio, USDA/ARS, Geneva, NY; Maria Angeles Forner-Giner, Insti...

  16. Evaluation of resistance to asiatic citrus canker among selections of pera sweet orange (Citrus sinensis)

    USDA-ARS?s Scientific Manuscript database

    Asiatic citrus canker (ACC, caused by the bacterium Xanthomonas citri subsp. citri) is a destructive disease of citrus in Brazil and in several other citrus-producing countries. ACC management is problematic, and bactericides such as copper can be reasonably efficacious but do not completely control...

  17. Field ID guide to citrus relative hosts of Asian citrus psyllid & Huanglongbing

    USDA-ARS?s Scientific Manuscript database

    The Rutaceae family of plants includes not only species within the genus Citrus, but also several other genera and species that may not be easily recognized as having any relationship to citrus at all. However, many of these citrus relatives are used for ornamental, culinary, or religious purposes. ...

  18. Production of transgenic citrus resistant to citrus canker and Huanglongbing diseases

    USDA-ARS?s Scientific Manuscript database

    Huanglongbing (HLB or citrus greening disease) caused by Candidatus Liberibacter asiaticus (Las) is a great threat to the U.S. citrus industry. There are no proven strategies to eliminate HLB disease and no cultivars identified with strong HLB resistance. Citrus canker is also an economically import...

  19. 76 FR 23449 - Citrus Canker, Citrus Greening, and Asian Citrus Psyllid; Interstate Movement of Regulated...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-04-27

    ... conditions, the articles are treated with methyl bromide and shipped in a container that has been sealed with... of any regulated article, including citrus nursery stock, provided that: The article is treated with methyl bromide in accordance with 7 CFR part 305. That part contains our phytosanitary treatment...

  20. 78 FR 63369 - Citrus Canker, Citrus Greening, and Asian Citrus Psyllid; Interstate Movement of Regulated...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-10-24

    ..., and Asian Citrus Psyllid; Interstate Movement of Regulated Nursery Stock AGENCY: Animal and Plant... nonsubstantive changes, an interim rule that amended the regulations governing the interstate movement of... (ACP) to allow the movement of regulated nursery stock under a certificate to any area within the...

  1. Possible role of plant volatiles in tolerance against huanglongbing in citrus

    PubMed Central

    Hijaz, Faraj; Nehela, Yasser; Killiny, Nabil

    2016-01-01

    abstract Volatile organic compounds (VOC) play an important role in protecting plants from insect and pathogen attack. In this study, we investigated the leaf volatile profiles of 14 citrus varieties. The VOC in citrus leaves were extracted with n-hexane and analyzed using gas chromatography-mass spectrometry (GC-MS). Overall, 4six volatile compounds were identified in the n-hexane extract from citrus leaves. Most of the detected compounds belonged to 3 main groups (monoterpenes, sesquiterpenes, and aliphatic aldehydes). Principle component analysis was used to examine the relative distribution of the studied varieties to each other. Interestingly, volatile profiles of varieties that are tolerant to Candidatus Liberibacter asiaticus (CLas) were different from those of the susceptible ones. Tolerant and moderately-tolerant cultivars contained relatively higher amounts of volatiles than susceptible varieties. In addition, tolerant varieties were also higher in specific compounds which are known for their antimicrobial activities. These compounds include Aldehydes (undecanal, neral, geranial, and citronellal) and some monoterpenes such as linalool, d-limonene, myrcene, α- and β- phellandrene. In addition, some sesquiterpene compounds including t-caryophellene, γ-elemene, β-elemene, germacrene D, and geranyl acetate were higher in tolerant and moderately tolerant cultivars. Severinia buxifolia which is known for its tolerance to CLas and many other pathogens contained higher levels of santalenes and coumarins. Our results indicated that citrus leaf volatiles might play a role in citrus tolerance to CLas. The results of this study may help in understanding of the mechanism of citrus tolerance against CLas. PMID:26829496

  2. Possible role of plant volatiles in tolerance against huanglongbing in citrus.

    PubMed

    Hijaz, Faraj; Nehela, Yasser; Killiny, Nabil

    2016-01-01

    Volatile organic compounds (VOC) play an important role in protecting plants from insect and pathogen attack. In this study, we investigated the leaf volatile profiles of 14 citrus varieties. The VOC in citrus leaves were extracted with n-hexane and analyzed using gas chromatography-mass spectrometry (GC-MS). Overall, 4six volatile compounds were identified in the n-hexane extract from citrus leaves. Most of the detected compounds belonged to 3 main groups (monoterpenes, sesquiterpenes, and aliphatic aldehydes). Principle component analysis was used to examine the relative distribution of the studied varieties to each other. Interestingly, volatile profiles of varieties that are tolerant to Candidatus Liberibacter asiaticus (CLas) were different from those of the susceptible ones. Tolerant and moderately-tolerant cultivars contained relatively higher amounts of volatiles than susceptible varieties. In addition, tolerant varieties were also higher in specific compounds which are known for their antimicrobial activities. These compounds include Aldehydes (undecanal, neral, geranial, and citronellal) and some monoterpenes such as linalool, d-limonene, myrcene, α- and β- phellandrene. In addition, some sesquiterpene compounds including t-caryophellene, γ-elemene, β-elemene, germacrene D, and geranyl acetate were higher in tolerant and moderately tolerant cultivars. Severinia buxifolia which is known for its tolerance to CLas and many other pathogens contained higher levels of santalenes and coumarins. Our results indicated that citrus leaf volatiles might play a role in citrus tolerance to CLas. The results of this study may help in understanding of the mechanism of citrus tolerance against CLas.

  3. Incidence of Candidatus Liberibacter asiaticus infection in abandoned citrus occurring in proximity to commercially managed groves.

    PubMed

    Tiwari, Siddharth; Lewis-Rosenblum, Hannah; Pelz-Stelinski, Kirsten; Stelinski, Lukasz L

    2010-12-01

    Huanglongbing is one of the most devastating diseases of citrus (Citrus spp.). One management tactic against huanglongbing is aggressive management of the vector, the Asian citrus psyllid (Diaphorina citri Kuwayama), with insecticide applications. However, D. citri in abandoned groves are not controlled and therefore pose a risk of reinfestation for nearby commercial citrus. These abandoned groves could serve as a reservoir for the vector, as well as a source of the presumed causal agent for huanglongbing in Florida, Candidatus Liberibacter asiaticus (Las). The current study was conducted to determine the degree to which Las is present in abandoned Florida citrus groves and to compare relative inoculum levels in nearby managed and abandoned groves during times of the year when D. citri are abundant (June, July, and August). In addition, the movement of Las by dispersing D. citri adults from inner and edge rows of abandoned grove plots to the corresponding rows of managed plots was quantified during the same 3 mo. The results of the current study confirmed the presence of Las in both D. citri and plant tissue in abandoned groves at statistically equivalent levels to those in nearby managed groves. The mean number of D. citri adults dispersing from abandoned to managed grove plots ranged from 7.25 +/- 1.70 to 70.25 +/- 21.25 per 4-d intervals. Of those, the mean number of dispersing D. citri adults that were carrying the Las pathogen ranged from 1.00 +/- 0.58 to 1.50 +/- 0.50. Our results indicate that abandoned citrus groves are a significant source of Ca. Las and that dispersing D. citri move this pathogen into nearby managed groves.

  4. Development of a full-genome cDNA clone of Citrus leaf blotch virus and infection of citrus plants.

    PubMed

    Vives, María Carmen; Martín, Susana; Ambrós, Silvia; Renovell, Agueda; Navarro, Luis; Pina, Jose Antonio; Moreno, Pedro; Guerri, José

    2008-11-01

    Citrus leaf blotch virus (CLBV), a member of the family Flexiviridae, has a ~9-kb single-stranded, positive-sense genomic RNA encapsidated by a 41-kDa coat protein. CLBV isolates are associated with symptom production in citrus including leaf blotching of Dweet tangor and stem pitting in Etrog citron (Dweet mottle disease), and some isolates are associated with bud union crease on trifoliate rootstocks, but Koch's postulates for this virus were not fulfilled. A full-genome cDNA of CLBV isolate SRA-153, which induces bud union crease, was placed under the T7 promoter (clone T7-CLBV), or between the 35S promoter and the Nos-t terminator, with or without a ribozyme sequence downstream of the CLBV sequence (clones 35SRbz-CLBV and 35S-CLBV). RNA transcripts from T7-CLBV failed to infect Etrog citron and Nicotiana occidentalis and N. benthamiana plants, whereas agro-inoculation with binary vectors carrying 35SRbz-CLBV or 35S-CLBV, and the p19 silencing suppressor, caused systemic infection and production of normal CLBV virions. Virus accumulation was similar in citron plants directly agro-infiltrated, or mechanically inoculated with wild-type or 35SRbz-CLBV-derived virions from Nicotiana, and the three sources incited the symptoms characteristic of Dweet mottle disease, but not bud union crease. Our results show that (1) virions derived from an infectious clone show the same replication, movement and pathogenicity characteristics as the wild-type CLBV; (2) CLBV is the causal agent of Dweet mottle disease but not of the bud union crease syndrome; and (3) for the first time an RNA virus could be successfully agro-inoculated on citrus plants. This infectious clone may become a useful viral vector for citrus genomic studies.

  5. Genome editing of the disease susceptibility gene CsLOB1 in citrus confers resistance to citrus canker.

    PubMed

    Jia, Hongge; Zhang, Yunzeng; Orbović, Vladimir; Xu, Jin; White, Frank F; Jones, Jeffrey B; Wang, Nian

    2016-12-09

    Citrus is a highly valued tree crop worldwide, while, at the same time, citrus production faces many biotic challenges, including bacterial canker and Huanglongbing (HLB). Breeding for disease-resistant varieties is the most efficient and sustainable approach to control plant diseases. Traditional breeding of citrus varieties is challenging due to multiple limitations, including polyploidy, polyembryony, extended juvenility and long crossing cycles. Targeted genome editing technology has the potential to shorten varietal development for some traits, including disease resistance. Here, we used CRISPR/Cas9/sgRNA technology to modify the canker susceptibility gene CsLOB1 in Duncan grapefruit. Six independent lines, DLOB 2, DLOB 3, DLOB 9, DLOB 10, DLOB 11 and DLOB 12, were generated. Targeted next-generation sequencing of the six lines showed the mutation rate was 31.58%, 23.80%, 89.36%, 88.79%, 46.91% and 51.12% for DLOB 2, DLOB 3, DLOB 9, DLOB 10, DLOB 11 and DLOB 12, respectively, of the cells in each line. DLOB 2 and DLOB 3 showed canker symptoms similar to wild-type grapefruit, when inoculated with the pathogen Xanthomonas citri subsp. citri (Xcc). No canker symptoms were observed on DLOB 9, DLOB 10, DLOB 11 and DLOB 12 at 4 days postinoculation (DPI) with Xcc. Pustules caused by Xcc were observed on DLOB 9, DLOB 10, DLOB 11 and DLOB 12 in later stages, which were much reduced compared to that on wild-type grapefruit. The pustules on DLOB 9 and DLOB 10 did not develop into typical canker symptoms. No side effects and off-target mutations were detected in the mutated plants. This study indicates that genome editing using CRISPR technology will provide a promising pathway to generate disease-resistant citrus varieties.

  6. Alternaria toxins in wheat from the Autonomous Province of Vojvodina, Serbia: a preliminary survey.

    PubMed

    Janić Hajnal, Elizabet; Orčić, Dejan; Torbica, Aleksandra; Kos, Jovana; Mastilović, Jasna; Škrinjar, Marija

    2015-01-01

    Although Fusarium species remain a main source of mycotoxin contamination of wheat, in recent years, due to the evident climatic changes, other mycotoxigenic fungi have been recognised as important wheat contaminants. Alternaria species, especially A. alternata, have been found as contaminants of wheat as well as wheat-based products. Under favourable conditions A. alternata very often produce alternariol (AOH), alternariol monomethyl ether (AME), tenuazonic acid (TeA) and others Alternaria toxins. The aim of the present study was to examine the presence of three Alternaria toxins (AOH, AME and TeA) in wheat samples harvested during three years (2011-13). To this end, 92 samples were collected during wheat harvesting from different growing regions of the Autonomous Province of Vojvodina, which represents the most important wheat-growing area in Serbia. The presence of Alternaria toxins was analysed by HPLC with electrospray ionisation triple quadrupole mass spectrometry (LC-ESI-MS/MS). Among all the analysed wheat samples, 63 (68.5%) were contaminated with TeA, 11 (12.0%) with AOH and 6 (6.5%) with AME. Furthermore, the maximum and mean toxin concentrations were 2676 and 92.4 µg kg(-1), 48.9 and 18.6 µg kg(-1), and 70.2 and 39.0 µg kg(-1) for TeA, AOH and AME, respectively. Co-occurrence of three Alternaria toxins in wheat samples was detected in six samples; a combination of two toxins was found in two samples; and 64 samples contained one toxin. The results showed that among 92 analysed wheat samples, only 20 (21.7%) samples were without Alternaria toxins. The presence of Alternaria toxins was also investigated in terms of weather conditions recorded during the period of investigation, as well as with the sampling region. This study represents the first preliminary report of the natural occurrence of Alternaria toxins in wheat (Triticum aestivum) from Serbia.

  7. Primary subcutaneous Alternaria alternata infection of the hand in an immunocompromised host.

    PubMed

    Kpodzo, Dzifa S; Calderwood, Michael S; Ruchelsman, David E; Abramson, Jeremy S; Piris, Adriano; Winograd, Jonathan M; Kotton, Camille N

    2011-07-01

    We describe a case of a progressive subcutaneous Alternaria alternata infection in the hand of a patient with chronic lymphocytic leukemia (CLL). The diagnosis was based upon the examination of tissue biopsy and isolation of the etiologic agent in culture. The identity of the isolate was determined by phenotypic characteristics and by sequencing the ITS and D1/D2 regions of the rDNA. Despite combination therapy with voriconazole and micafungin, the lesion continued to progress. Posaconazole therapy, along with surgical excision of the infected tissue, resulted in the eradication of infection. The limitations of the clinical management of invasive Alternaria infections are discussed.

  8. Photographic Remote Sensing of Sick Citrus Trees

    NASA Technical Reports Server (NTRS)

    Gausman, H. W.

    1971-01-01

    Remote sensing with infrared color aerial photography (Kodak Ektachrome Infrared Aero 8443 film) for detecting citrus tree anomalies is described. Illustrations and discussions are given for detecting nutrient toxicity symptoms, for detecting foot rot and sooty mold fungal diseases, and for distinguishing among citrus species. Also, the influence of internal leaf structure on light reflectance, transmittance, and absorptance are considered; and physiological and environmental factors that affect citrus leaf light reflectance are reviewed briefly and illustrated.

  9. Antifeedant and sublethal effects of imidacloprid on Asian citrus psyllid, Diaphorina citri.

    PubMed

    Boina, Dhana Raj; Onagbola, Ebenezer O; Salyani, Masoud; Stelinski, Lukasz L

    2009-08-01

    Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, transmits the causal bacteria of the devastating citrus disease huanglongbing (HLB). Because of the variation in spatial and temporal uptake and systemic distribution of imidacloprid applied to citrus trees and its degradation over time in citrus trees, ACP adults and nymphs are exposed to concentrations that may not cause immediate mortality but rather sublethal effects. The objective of this laboratory study was to determine the effects of sublethal concentrations of imidacloprid on ACP life stages. Feeding by ACP adults and nymphs on plants treated daily with a sublethal concentration (0.1 microg mL(-1)) of imidacloprid significantly decreased adult longevity (8 days), fecundity (33%) and fertility (6%), as well as nymph survival (12%) and developmental rate compared with untreated controls. The magnitude of these negative effects was directly related to exposure duration and concentration. Furthermore, ACP adults that fed on citrus leaves treated systemically with lethal and sublethal concentrations of imidacloprid excreted significantly less honeydew (7-94%) compared with controls in a concentration-dependent manner suggesting antifeedant activity of imidacloprid. Sublethal concentrations of imidacloprid negatively affect development, reproduction, survival and longevity of ACP, which likely contributes to population reductions over time. Also, reduced feeding by ACP adults on plants treated with sublethal concentrations of imidacloprid may potentially decrease the capacity of ACP to successfully acquire and transmit the HLB causal pathogen.

  10. Exogenous nitric oxide-induced postharvest disease resistance in citrus fruit to Colletotrichum gloeosporioides.

    PubMed

    Zhou, Yahan; Li, Shunmin; Zeng, Kaifang

    2016-01-30

    Nitric oxide (NO) is an important signaling molecule involved in numerous plant responses to biotic and abiotic stresses. To investigate the effects of NO on the control of postharvest anthracnose caused by Colletotrichum gloeosporioides in citrus fruit and its possible mechanisms, citrus fruit were treated with an NO donor. The results showed that exogenous NO released from 50 µmol L(-1) sodium nitroprusside aqueous solution could effectively reduce the disease incidence and lesion diameter of citrus fruit inoculated with C. gloeosporioides during storage at 20 °C. Exogenous NO could regulate hydrogen peroxide levels, stimulate the synthesis of phenolic compounds, and induce phenylalanine ammonia-lyase, peroxidase, polyphenol oxidase, catalase activities, and the ascorbate-glutathione cycle. Furthermore, exogenous NO could inhibit weight loss, improve the ascorbic acid and titratable acidity content, and delay the increase in total soluble solids content in citrus fruit during storage at 20 °C. The results suggest that the use of exogenous NO is a potential method for inducing the disease resistance of fruit to fungal pathogens and for extending the postharvest life of citrus fruit. © 2015 Society of Chemical Industry.

  11. Citrus Allergy from Pollen to Clinical Symptoms

    PubMed Central

    Iorio, Rosa Anna; Del Duca, Stefano; Calamelli, Elisabetta; Pula, Chiara; Lodolini, Magda; Scamardella, Fortuna; Pession, Andrea; Ricci, Giampaolo

    2013-01-01

    Allergy to citrus fruits is often associated with pollinosis and sensitization to other plants due to a phenomenon of cross-reactivity. The aims of the present study were to highlight the cross-reactivity among citrus and the major allergenic pollens/fruits, throughout clinical and molecular investigations, and to evaluate the sensitization frequency to citrus fruits in a population of children and adults with pollinosis. We found a relevant percentage of sensitisation (39%) to citrus fruits in the patients recruited and in all of them the IgE-mediated mechanism has been confirmed by the positive response to the prick-to-prick test. RT-PCR experiments showed the expression of Cit s 1, Cit s 3 and a profilin isoform, already described in apple, also in Citrus clementine pollen. Data of multiple sequence alignments demonstrated that Citrus allergens shared high percentage identity values with other clinically relevant species (i.e. Triticum aestivum, Malus domestica), confirming the possible cross-allergenicity citrus/grasses and citrus/apple. Finally, a novelty of the present work has been the expression of two phospholipaseA2 isoforms (PLA2 α and β) in Citrus as well as in Triticum pollens; being PLA2 able to generate pro-inflammatory factors, this enzyme could participate in the activation of the allergenic inflammatory cascade. PMID:23308273

  12. Occurrence of citrus viroids in Costa Rica.

    PubMed

    Villalobos, W; Rivera, C; Hammond, R W

    1997-09-01

    A survey for citrus viroids was conducted in the major citrus commercial growing areas in Costa Rica. Screening of 36 sweet orange and 12 lemon trees resulted in the detection of members of four of the five citrus viroid groups as determined by nucleic acid hybridization using specific RNA probes and polymerase chain reaction (PCR) using specific oligonucleotide primers. CEVd, CVd-IIa, CVD-IIb and CVd-III viroids were found to be widespread in the three main regions of commercial citrus production. CVd-Ib was only found in lemon in Nicoya.

  13. The relationship between PthA expression and the pathogenicity of Xanthomonas axonopodis pv. citri.

    PubMed

    Li, Na; Huang, Long; Liu, Liping; Li, Dazhi; Dai, Suming; Deng, Ziniu

    2014-02-01

    Citrus canker disease, caused by Xanthomonas axonopodis pv. citri, affects almost all citrus species and cultivars and hascaused severe damage to the citrus industry worldwide. PthA is considered the main pathogenesis effector of the pathogen. This research aimed to temporally and spatially analyze the expression of the PthA protein of the bactrium during its culture, and then try to understand the relationship between the PthA expression levels and the pathogenicity. The relationship between the expression of PthA and the pathogenicity of X. axonopodis pv. citri was fully investigated by using SDS-PAGE, Western blot, ELISA and field inoculation, It was found that bacteria cultured for 36 h had the highest expression of PthA and showed the most virulent pathogenicity. The conservation duration of the pathogen isolates influenced their PthA expression and the pathogenicity, and negative relationship between the duration and the expression of PthA and pathogenicity. When the stored pathogen bacteria were cultured in liquid LB medium, they were able to regain activated, showing higher PthA expression level and enhanced pathogenicity, even though the activity was inferior, in terms of both PthA expression and pathogenicity, than the freshly isolated ones. Seven isolates from different citrus orchards displayed almost identical protein expression profiles. It could conclude that the expressions of PthA was positively related to pathogenicity.

  14. Rapid differentiation of citrus Hop stunt viroid variants by use of real-time RT-PCR and high resolution melting analysis

    USDA-ARS?s Scientific Manuscript database

    The RNA genome of Hop stunt viroid (HSVd) contains five to six nucleotides in a variable (V) domain, called the cachexia expression motif, which is associated with pathogenic and non-pathogenic variants in citrus. Current methods to differentiate HSVd variants rely on lengthy greenhouse biological i...

  15. Molecular mechanisms behind the accumulation of adenosine triphosphate (ATP) and H2O2 in citrus plants in response to ‘Candidatus Liberibacter asiaticus’ infection

    USDA-ARS?s Scientific Manuscript database

    Candidatus Liberibacter asiaticus (Las) is a fastidious, phloem-restricted pathogen with a significantly reduced genome, and attacks all citrus species with no immune cultivars documented to date. Like other plant bacterial pathogens, Las deploys effector proteins into the organelles of plant cells,...

  16. Biopriming of Infected Carrot Seed with an Antagonist, Clonostachys rosea, Selected for Control of Seedborne Alternaria spp.

    PubMed

    Jensen, Birgit; Knudsen, Inge M B; Madsen, Mette; Jensen, Dan Funck

    2004-06-01

    ABSTRACT An ecological approach was used to select fungal antagonists effective against the seedborne pathogens Alternaria dauci and A. radicina on carrot. Twenty-five and 105 isolates originating from cereal and carrot habitats were screened against the pathogens in planta, respectively. Irrespective of isolate origin, fungal isolates belonging to Clonostachys rosea controlled pre- and postemergence death caused by A. dauci and A. radicina as effectively as the fungicide iprodione. Isolate IK726 of C. rosea was used in biopriming a seed lot with 29% A. radicina and 11% A. dauci (highly infected), and a seed lot with 4% A. radicina and 7% A. dauci (low infection). Seeds were primed with water alone (hydropriming) or with addition of C. rosea IK726 (biopriming). The occurrence of A. radicina and A. dauci increased twofold and fivefold, respectively, during 14 days hydropriming, irrespective of the initial infection level. On highly infected seed, biopriming reduced the incidence of A. radicina to <2.3% and that of A. dauci to <4.8% while the level of both pathogens was <0.5% on bioprimed seed with a low initial infection rate. In sand stand establishment tests, hydroprimed seeds had a lower healthy seedling stand than nonprimed seeds, mainly due to a high degree of postemergence seedling death. In contrast, biopriming resulted in a seedling stand that was better than that of both nonprimed and hydroprimed seeds. C. rosea IK726 multiplied fivefold to eightfold, and microscopic observations using C. rosea IK726 transformed with a green fluorescent protein (GFP) reporter gene showed that seeds were covered with a fine web of sporulating mycelium of C. rosea. The positive effect of biopriming on healthy seedling stand remained after 5 months of storage at 4 degrees C and IK726 survived at high numbers on these seed. In this study, we demonstrated that bio-priming with the biocontrol strain C. rosea IK726 facilitates priming of infected seeds without risking adverse

  17. Assessing the mycotoxigenic threat of necrotrophic pathogens of wheat.

    PubMed

    Solomon, Peter S

    2011-11-01

    Pathogenic fungi are the causal agents of many significant plant diseases around the world. These diseases often result in significant yield reductions, leading to lower food production rates and economic losses. Several of these pathogenic fungi also produce mycotoxins during infection, which are harmful to human and animal health. Whilst some of these toxins and the fungi that produce them have been studied intensively, the mycotoxigenic potential of many of these pathogens remains largely unknown. Included within these fungi are the necrotrophic pathogens of wheat, Stagonospora nodorum, Pyrenophora tritici-repentis and Alternaria alternata. Recent studies have demonstrated that each of these pathogens is capable of synthesizing an array of mycotoxic compounds during disease development, questioning their status as non-mycotoxin producers. This review summarises recent mycotoxin findings in these necrotrophic wheat pathogens by briefly discussing the mycotoxins identified, their toxicity and their synthesis. Future and emerging threats are also considered.

  18. Huanglongbing, a systemic disease, restructures the bacterial community associated with citrus roots.

    PubMed

    Trivedi, Pankaj; Duan, Yongping; Wang, Nian

    2010-06-01

    To examine the effect of pathogens on the diversity and structure of plant-associated bacterial communities, we carried out a molecular analysis using citrus and huanglongbing as a host-disease model. 16S rRNA gene clone library analysis of citrus roots revealed shifts in microbial diversity in response to pathogen infection. The clone library of the uninfected root samples has a majority of phylotypes showing similarity to well-known plant growth-promoting bacteria, including Caulobacter, Burkholderia, Lysobacter, Pantoea, Pseudomonas, Stenotrophomonas, Bacillus, and Paenibacillus. Infection by "Candidatus Liberibacter asiaticus" restructured the native microbial community associated with citrus roots and led to the loss of detection of most phylotypes while promoting the growth of bacteria such as Methylobacterium and Sphingobacterium. In pairwise comparisons, the clone library from uninfected roots contained significantly higher 16S rRNA gene diversity, as reflected in the higher Chao 1 richness estimation (P pathogen. These data indicate that infection of citrus by "Ca. Liberibacter asiaticus" has a profound effect on the structure and composition of the bacterial community associated with citrus roots.

  19. Huanglongbing, a Systemic Disease, Restructures the Bacterial Community Associated with Citrus Roots▿

    PubMed Central

    Trivedi, Pankaj; Duan, Yongping; Wang, Nian

    2010-01-01

    To examine the effect of pathogens on the diversity and structure of plant-associated bacterial communities, we carried out a molecular analysis using citrus and huanglongbing as a host-disease model. 16S rRNA gene clone library analysis of citrus roots revealed shifts in microbial diversity in response to pathogen infection. The clone library of the uninfected root samples has a majority of phylotypes showing similarity to well-known plant growth-promoting bacteria, including Caulobacter, Burkholderia, Lysobacter, Pantoea, Pseudomonas, Stenotrophomonas, Bacillus, and Paenibacillus. Infection by “Candidatus Liberibacter asiaticus” restructured the native microbial community associated with citrus roots and led to the loss of detection of most phylotypes while promoting the growth of bacteria such as Methylobacterium and Sphingobacterium. In pairwise comparisons, the clone library from uninfected roots contained significantly higher 16S rRNA gene diversity, as reflected in the higher Chao 1 richness estimation (P ≤ 0.01) of 237.13 versus 42.14 for the uninfected and infected clone libraries, respectively. Similarly, the Shannon index of the uninfected clone library (4.46) was significantly higher than that of the infected clone library (2.61). Comparison of the uninfected clone library with the infected clone library using LIBSHUFF statistics showed a significant difference (P ≤ 0.05). Quantitative PCR analysis revealed that the bacterial community changes not only qualitatively but also quantitatively. The relative proportions of different groups of bacteria changed significantly after infection with the pathogen. These data indicate that infection of citrus by “Ca. Liberibacter asiaticus” has a profound effect on the structure and composition of the bacterial community associated with citrus roots. PMID:20382817

  20. Comparison of FTIR spectra between Huanglongbing (citrus greening) and other citrus maladies.

    PubMed

    Hawkins, Samantha A; Park, Bosoon; Poole, Gavin H; Gottwald, Tim R; Windham, William R; Albano, Joseph; Lawrence, Kurt C

    2010-05-26

    Fourier transform infrared (FTIR) spectroscopy has the ability to quickly identify the presence of specific carbohydrates in plant materials. The presence of the disease huanglongbing (HLB) in the leaves of infected citrus plants has a distinctive spectrum that can be used to distinguish an infected plant from a healthy plant. However, many citrus diseases display similar visible symptoms and are of concern to citrus growers. In this study several citrus diseases (citrus leaf rugose virus, citrus tristeza virus, citrus psorosis virus, and Xanthomonas axonopodis ) and nutrient deficiencies (iron, copper, zinc, manganese, and magnesium) were compared with HLB using FTIR spectroscopy to determine if the spectra alone can be used to identify plants that are infected with HLB instead of another disease. The results indicate that the spectra of some diseases and deficiencies more closely resemble those of apparently healthy plants and some share the carbohydrate transformation that has been seen in the spectra of HLB-infected plants.