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Sample records for virulence gene regulation

  1. Brucella abortus: pathogenicity and gene regulation of virulence

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    Olga Rivas-Solano

    2015-06-01

    Full Text Available Brucella abortus is a zoonotic intracellular facultative pathogen belonging to the subdivision α2 of class Proteobacteria. It causes a worldwide distributed zoonotic disease called brucellosis. The main symptoms are abortion and sterility in cattle, as well as an undulant febrile condition in humans. In endemic regions like Central America, brucellosis has a high socioeconomic impact. A basic research project was recently conducted at the ITCR with the purpose of studying gene regulation of virulence, structure and immunogenicity in B. abortus. The present review was written as part of this project. B. abortus virulence seems to be determined by its ability to invade, survive and replicate inside professional and non-professional phagocytes. It reaches its intracellular replicative niche without the activation of host antimicrobial mechanisms of innate immunity. It also has gene regulation mechanisms for a rapid adaptation to an intracellular environment such as the two-component signal transduction system BvrR/BvrS and the quorum sensing regulator called Vjbr, as well as other transcription factors. All of them integrate a complex gene regulation network.

  2. Lvr, a Signaling System That Controls Global Gene Regulation and Virulence in Pathogenic Leptospira

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    Adhikarla, Haritha; Wunder, Elsio A.; Mechaly, Ariel E.; Mehta, Sameet; Wang, Zheng; Santos, Luciane; Bisht, Vimla; Diggle, Peter; Murray, Gerald; Adler, Ben; Lopez, Francesc; Townsend, Jeffrey P.; Groisman, Eduardo; Picardeau, Mathieu; Buschiazzo, Alejandro; Ko, Albert I.

    2018-01-01

    Leptospirosis is an emerging zoonotic disease with more than 1 million cases annually. Currently there is lack of evidence for signaling pathways involved during the infection process of Leptospira. In our comprehensive genomic analysis of 20 Leptospira spp. we identified seven pathogen-specific Two-Component System (TCS) proteins. Disruption of two these TCS genes in pathogenic Leptospira strain resulted in loss-of-virulence in a hamster model of leptospirosis. Corresponding genes lvrA and lvrB (leptospira virulence regulator) are juxtaposed in an operon and are predicted to encode a hybrid histidine kinase and a hybrid response regulator, respectively. Transcriptome analysis of lvr mutant strains with disruption of one (lvrB) or both genes (lvrA/B) revealed global transcriptional regulation of 850 differentially expressed genes. Phosphotransfer assays demonstrated that LvrA phosphorylates LvrB and predicted further signaling downstream to one or more DNA-binding response regulators, suggesting that it is a branched pathway. Phylogenetic analyses indicated that lvrA and lvrB evolved independently within different ecological lineages in Leptospira via gene duplication. This study uncovers a novel-signaling pathway that regulates virulence in pathogenic Leptospira (Lvr), providing a framework to understand the molecular bases of regulation in this life-threatening bacterium. PMID:29600195

  3. A combination of independent transcriptional regulators shapes bacterial virulence gene expression during infection.

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    Samuel A Shelburne

    2010-03-01

    Full Text Available Transcriptional regulatory networks are fundamental to how microbes alter gene expression in response to environmental stimuli, thereby playing a critical role in bacterial pathogenesis. However, understanding how bacterial transcriptional regulatory networks function during host-pathogen interaction is limited. Recent studies in group A Streptococcus (GAS suggested that the transcriptional regulator catabolite control protein A (CcpA influences many of the same genes as the control of virulence (CovRS two-component gene regulatory system. To provide new information about the CcpA and CovRS networks, we compared the CcpA and CovR transcriptomes in a serotype M1 GAS strain. The transcript levels of several of the same genes encoding virulence factors and proteins involved in basic metabolic processes were affected in both DeltaccpA and DeltacovR isogenic mutant strains. Recombinant CcpA and CovR bound with high-affinity to the promoter regions of several co-regulated genes, including those encoding proteins involved in carbohydrate and amino acid metabolism. Compared to the wild-type parental strain, DeltaccpA and DeltacovRDeltaccpA isogenic mutant strains were significantly less virulent in a mouse myositis model. Inactivation of CcpA and CovR alone and in combination led to significant alterations in the transcript levels of several key GAS virulence factor encoding genes during infection. Importantly, the transcript level alterations in the DeltaccpA and DeltacovRDeltaccpA isogenic mutant strains observed during infection were distinct from those occurring during growth in laboratory medium. These data provide new knowledge regarding the molecular mechanisms by which pathogenic bacteria respond to environmental signals to regulate virulence factor production and basic metabolic processes during infection.

  4. The FTF gene family regulates virulence and expression of SIX effectors in Fusarium oxysporum.

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    Niño-Sánchez, Jonathan; Casado-Del Castillo, Virginia; Tello, Vega; De Vega-Bartol, José J; Ramos, Brisa; Sukno, Serenella A; Díaz Mínguez, José María

    2016-09-01

    The FTF (Fusarium transcription factor) gene family comprises a single copy gene, FTF2, which is present in all the filamentous ascomycetes analysed, and several copies of a close relative, FTF1, which is exclusive to Fusarium oxysporum. An RNA-mediated gene silencing system was developed to target mRNA produced by all the FTF genes, and tested in two formae speciales: F. oxysporum f. sp. phaseoli (whose host is common bean) and F. oxysporum f. sp. lycopersici (whose host is tomato). Quantification of the mRNA levels showed knockdown of FTF1 and FTF2 in randomly isolated transformants of both formae speciales. The attenuation of FTF expression resulted in a marked reduction in virulence, a reduced expression of several SIX (Secreted In Xylem) genes, the best studied family of effectors in F. oxysporum, and lower levels of SGE1 (Six Gene Expression 1) mRNA, the presumptive regulator of SIX expression. Moreover, the knockdown mutants showed a pattern of colonization of the host plant similar to that displayed by strains devoid of FTF1 copies (weakly virulent strains). Gene knockout of FTF2 also resulted in a reduction in virulence, but to a lesser extent. These results demonstrate the role of the FTF gene expansion, mostly the FTF1 paralogues, as a regulator of virulence in F. oxysporum and suggest that the control of effector expression is the mechanism involved. © 2016 The Authors Molecular Plant Pathology Published by British Society for Plant Pathology and John Wiley & Sons Ltd.

  5. Virulence meets metabolism: Cra and KdpE gene regulation in enterohemorrhagic Escherichia coli.

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    Njoroge, Jacqueline W; Nguyen, Y; Curtis, Meredith M; Moreira, Cristiano G; Sperandio, Vanessa

    2012-10-16

    Gastrointestinal (GI) bacteria sense diverse environmental signals as cues for differential gene regulation and niche adaptation. Pathogens such as enterohemorrhagic Escherichia coli (EHEC), which causes bloody diarrhea, use these signals for the temporal and energy-efficient regulation of their virulence factors. One of the main virulence strategies employed by EHEC is the formation of attaching and effacing (AE) lesions on enterocytes. Most of the genes necessary for the formation of these lesions are grouped within a pathogenicity island, the locus of enterocyte effacement (LEE), whose expression requires the LEE-encoded regulator Ler. Here we show that growth of EHEC in glycolytic environments inhibits the expression of ler and consequently all other LEE genes. Conversely, growth within a gluconeogenic environment activates expression of these genes. This sugar-dependent regulation is achieved through two transcription factors: KdpE and Cra. Both Cra and KdpE directly bind to the ler promoter, and Cra's affinity to this promoter is catabolite dependent. Moreover, we show that the Cra and KdpE proteins interact in vitro and that KdpE's ability to bind DNA is enhanced by the presence of Cra. Cra is important for AE lesion formation, and KdpE contributes to this Cra-dependent regulation. The deletion of cra and kdpE resulted in the ablation of AE lesions. One of the many challenges that bacteria face within the GI tract is to successfully compete for carbon sources. Linking carbon metabolism to the precise coordination of virulence expression is a key step in the adaptation of pathogens to the GI environment. IMPORTANCE An appropriate and prompt response to environmental cues is crucial for bacterial survival. Cra and KdpE are two proteins found in both nonpathogenic and pathogenic bacteria that regulate genes in response to differences in metabolite concentration. In this work, we show that, in the deadly pathogen enterohemorrhagic Escherichia coli (EHEC) O157:H7

  6. Haemophilus ducreyi Hfq contributes to virulence gene regulation as cells enter stationary phase.

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    Gangaiah, Dharanesh; Labandeira-Rey, Maria; Zhang, Xinjun; Fortney, Kate R; Ellinger, Sheila; Zwickl, Beth; Baker, Beth; Liu, Yunlong; Janowicz, Diane M; Katz, Barry P; Brautigam, Chad A; Munson, Robert S; Hansen, Eric J; Spinola, Stanley M

    2014-02-11

    To adapt to stresses encountered in stationary phase, Gram-negative bacteria utilize the alternative sigma factor RpoS. However, some species lack RpoS; thus, it is unclear how stationary-phase adaptation is regulated in these organisms. Here we defined the growth-phase-dependent transcriptomes of Haemophilus ducreyi, which lacks an RpoS homolog. Compared to mid-log-phase organisms, cells harvested from the stationary phase upregulated genes encoding several virulence determinants and a homolog of hfq. Insertional inactivation of hfq altered the expression of ~16% of the H. ducreyi genes. Importantly, there were a significant overlap and an inverse correlation in the transcript levels of genes differentially expressed in the hfq inactivation mutant relative to its parent and the genes differentially expressed in stationary phase relative to mid-log phase in the parent. Inactivation of hfq downregulated genes in the flp-tad and lspB-lspA2 operons, which encode several virulence determinants. To comply with FDA guidelines for human inoculation experiments, an unmarked hfq deletion mutant was constructed and was fully attenuated for virulence in humans. Inactivation or deletion of hfq downregulated Flp1 and impaired the ability of H. ducreyi to form microcolonies, downregulated DsrA and rendered H. ducreyi serum susceptible, and downregulated LspB and LspA2, which allow H. ducreyi to resist phagocytosis. We propose that, in the absence of an RpoS homolog, Hfq serves as a major contributor of H. ducreyi stationary-phase and virulence gene regulation. The contribution of Hfq to stationary-phase gene regulation may have broad implications for other organisms that lack an RpoS homolog. Pathogenic bacteria encounter a wide range of stresses in their hosts, including nutrient limitation; the ability to sense and respond to such stresses is crucial for bacterial pathogens to successfully establish an infection. Gram-negative bacteria frequently utilize the alternative sigma

  7. The metabolic regulator CodY links L. monocytogenes metabolism to virulence by directly activating the virulence regulatory gene, prfA

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    Lobel, Lior; Sigal, Nadejda; Borovok, Ilya; Belitsky, Boris R.; Sonenshein, Abraham L.; Herskovits, Anat A.

    2015-01-01

    Summary Metabolic adaptations are critical to the ability of bacterial pathogens to grow within host cells and are normally preceded by sensing of host-specific metabolic signals, which in turn can influence the pathogen's virulence state. Previously, we reported that the intracellular bacterial pathogen Listeria monocytogenes responds to low availability of branched-chain amino acids (BCAA) within mammalian cells by up-regulating both BCAA biosynthesis and virulence genes. The induction of virulence genes required the BCAA-responsive transcription regulator, CodY, but the molecular mechanism governing this mode of regulation was unclear. In this report, we demonstrate that CodY directly binds the coding sequence of the L. monocytogenes master virulence activator gene, prfA, 15 nt downstream of its start codon, and that this binding results in up-regulation of prfA transcription specifically under low concentrations of BCAA. Mutating this site abolished CodY binding and reduced prfA transcription in macrophages, and attenuated bacterial virulence in mice. Notably, the mutated binding site did not alter prfA transcription or PrfA activity under other conditions that are known to activate PrfA, such as during growth in the presence of glucose-1-phosphate. This study highlights the tight crosstalk between L. monocytogenes metabolism and virulence' while revealing novel features of CodY-mediated regulation. PMID:25430920

  8. Effect of Negative Pressure on Proliferation, Virulence Factor Secretion, Biofilm Formation, and Virulence-Regulated Gene Expression of Pseudomonas aeruginosa In Vitro

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    Guo-Qi Wang

    2016-01-01

    Full Text Available Objective. To investigate the effect of negative pressure conditions induced by NPWT on P. aeruginosa. Methods. P. aeruginosa was cultured in a Luria–Bertani medium at negative pressure of −125 mmHg for 24 h in the experimental group and at atmospheric pressure in the control group. The diameters of the colonies of P. aeruginosa were measured after 24 h. ELISA kit, orcinol method, and elastin-Congo red assay were used to quantify the virulence factors. Biofilm formation was observed by staining with Alexa Fluor® 647 conjugate of concanavalin A (Con A. Virulence-regulated genes were determined by quantitative RT-PCR. Results. As compared with the control group, growth of P. aeruginosa was inhibited by negative pressure. The colony size under negative pressure was significantly smaller in the experimental group than that in the controls (p<0.01. Besides, reductions in the total amount of virulence factors were observed in the negative pressure group, including exotoxin A, rhamnolipid, and elastase. RT-PCR results revealed a significant inhibition in the expression level of virulence-regulated genes. Conclusion. Negative pressure could significantly inhibit the growth of P. aeruginosa. It led to a decrease in the virulence factor secretion, biofilm formation, and a reduction in the expression level of virulence-regulated genes.

  9. Structure of Rot, a global regulator of virulence genes in Staphylococcus aureus.

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    Zhu, Yuwei; Fan, Xiaojiao; Zhang, Xu; Jiang, Xuguang; Niu, Liwen; Teng, Maikun; Li, Xu

    2014-09-01

    Staphylococcus aureus is a highly versatile pathogen that can infect human tissue by producing a large arsenal of virulence factors that are tightly regulated by a complex regulatory network. Rot, which shares sequence similarity with SarA homologues, is a global regulator that regulates numerous virulence genes. However, the recognition model of Rot for the promoter region of target genes and the putative regulation mechanism remain elusive. In this study, the 1.77 Å resolution X-ray crystal structure of Rot is reported. The structure reveals that two Rot molecules form a compact homodimer, each of which contains a typical helix-turn-helix module and a β-hairpin motif connected by a flexible loop. Fluorescence polarization results indicate that Rot preferentially recognizes AT-rich dsDNA with ~30-base-pair nucleotides and that the conserved positively charged residues on the winged-helix motif are vital for binding to the AT-rich dsDNA. It is proposed that the DNA-recognition model of Rot may be similar to that of SarA, SarR and SarS, in which the helix-turn-helix motifs of each monomer interact with the major grooves of target dsDNA and the winged motifs contact the minor grooves. Interestingly, the structure shows that Rot adopts a novel dimerization model that differs from that of other SarA homologues. As expected, perturbation of the dimer interface abolishes the dsDNA-binding ability of Rot, suggesting that Rot functions as a dimer. In addition, the results have been further confirmed in vivo by measuring the transcriptional regulation of α-toxin, a major virulence factor produced by most S. aureus strains.

  10. Pleiotropic Regulation of Virulence Genes in Streptococcus mutans by the Conserved Small Protein SprV.

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    Shankar, Manoharan; Hossain, Mohammad S; Biswas, Indranil

    2017-04-15

    Streptococcus mutans , an oral pathogen associated with dental caries, colonizes tooth surfaces as polymicrobial biofilms known as dental plaque. S. mutans expresses several virulence factors that allow the organism to tolerate environmental fluctuations and compete with other microorganisms. We recently identified a small hypothetical protein (90 amino acids) essential for the normal growth of the bacterium. Inactivation of the gene, SMU.2137, encoding this protein caused a significant growth defect and loss of various virulence-associated functions. An S. mutans strain lacking this gene was more sensitive to acid, temperature, osmotic, oxidative, and DNA damage-inducing stresses. In addition, we observed an altered protein profile and defects in biofilm formation, bacteriocin production, and natural competence development, possibly due to the fitness defect associated with SMU.2137 deletion. Transcriptome sequencing revealed that nearly 20% of the S. mutans genes were differentially expressed upon SMU.2137 deletion, thereby suggesting a pleiotropic effect. Therefore, we have renamed this hitherto uncharacterized gene as sprV ( s treptococcal p leiotropic r egulator of v irulence). The transcript levels of several relevant genes in the sprV mutant corroborated the phenotypes observed upon sprV deletion. Owing to its highly conserved nature, inactivation of the sprV ortholog in Streptococcus gordonii also resulted in poor growth and defective UV tolerance and competence development as in the case of S. mutans Our experiments suggest that SprV is functionally distinct from its homologs identified by structure and sequence homology. Nonetheless, our current work is aimed at understanding the importance of SprV in the S. mutans biology. IMPORTANCE Streptococcus mutans employs several virulence factors and stress resistance mechanisms to colonize tooth surfaces and cause dental caries. Bacterial pathogenesis is generally controlled by regulators of fitness that are

  11. Quorum-sensing regulators control virulence gene expression in Vibrio cholerae

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    Zhu, Jun; Miller, Melissa B.; Vance, Russell E.; Dziejman, Michelle; Bassler, Bonnie L.; Mekalanos, John J.

    2002-01-01

    The production of virulence factors including cholera toxin and the toxin-coregulated pilus in the human pathogen Vibrio cholerae is strongly influenced by environmental conditions. The well-characterized ToxR signal transduction cascade is responsible for sensing and integrating the environmental information and controlling the virulence regulon. We show here that, in addition to the known components of the ToxR signaling circuit, quorum-sensing regulators are involved in regulation of V. ch...

  12. Metabolic Genetic Screens Reveal Multidimensional Regulation of Virulence Gene Expression in Listeria monocytogenes and an Aminopeptidase That Is Critical for PrfA Protein Activation.

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    Friedman, Sivan; Linsky, Marika; Lobel, Lior; Rabinovich, Lev; Sigal, Nadejda; Herskovits, Anat A

    2017-06-01

    Listeria monocytogenes is an environmental saprophyte and intracellular bacterial pathogen. Upon invading mammalian cells, the bacterium senses abrupt changes in its metabolic environment, which are rapidly transduced to regulation of virulence gene expression. To explore the relationship between L. monocytogenes metabolism and virulence, we monitored virulence gene expression dynamics across a library of genetic mutants grown under two metabolic conditions known to activate the virulent state: charcoal-treated rich medium containing glucose-1-phosphate and minimal defined medium containing limiting concentrations of branched-chain amino acids (BCAAs). We identified over 100 distinct mutants that exhibit aberrant virulence gene expression profiles, the majority of which mapped to nonessential metabolic genes. Mutants displayed enhanced, decreased, and early and late virulence gene expression profiles, as well as persistent levels, demonstrating a high plasticity in virulence gene regulation. Among the mutants, one was noteworthy for its particularly low virulence gene expression level and mapped to an X-prolyl aminopeptidase (PepP). We show that this peptidase plays a role in posttranslational activation of the major virulence regulator, PrfA. Specifically, PepP mediates recruitment of PrfA to the cytoplasmic membrane, a step identified as critical for PrfA protein activation. This study establishes a novel step in the complex mechanism of PrfA activation and further highlights the cross regulation of metabolism and virulence. Copyright © 2017 American Society for Microbiology.

  13. Unfolded Protein Response (UPR Regulator Cib1 Controls Expression of Genes Encoding Secreted Virulence Factors in Ustilago maydis.

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    Martin Hampel

    Full Text Available The unfolded protein response (UPR, a conserved eukaryotic signaling pathway to ensure protein homeostasis in the endoplasmic reticulum (ER, coordinates biotrophic development in the corn smut fungus Ustilago maydis. Exact timing of UPR activation is required for virulence and presumably connected to the elevated expression of secreted effector proteins during infection of the host plant Zea mays. In the baker's yeast Saccharomyces cerevisiae, expression of UPR target genes is induced upon binding of the central regulator Hac1 to unfolded protein response elements (UPREs in their promoters. While a role of the UPR in effector secretion has been described previously, we investigated a potential UPR-dependent regulation of genes encoding secreted effector proteins. In silico prediction of UPREs in promoter regions identified the previously characterized effector genes pit2 and tin1-1, as bona fide UPR target genes. Furthermore, direct binding of the Hac1-homolog Cib1 to the UPRE containing promoter fragments of both genes was confirmed by quantitative chromatin immunoprecipitation (qChIP analysis. Targeted deletion of the UPRE abolished Cib1-dependent expression of pit2 and significantly affected virulence. Furthermore, ER stress strongly increased Pit2 expression and secretion. This study expands the role of the UPR as a signal hub in fungal virulence and illustrates, how biotrophic fungi can coordinate cellular physiology, development and regulation of secreted virulence factors.

  14. Accessory Gene Regulator-1 Locus Is Essential for Virulence and Pathogenesis of Clostridium difficile

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    Charles Darkoh

    2016-08-01

    Full Text Available Clostridium difficile infection (CDI is responsible for most of the definable cases of antibiotic- and hospital-associated diarrhea worldwide and is a frequent cause of morbidity and mortality in older patients. C. difficile, a multidrug-resistant anaerobic pathogen, causes disease by producing toxins A and B, which are controlled by an accessory gene regulator (Agr quorum signaling system. Some C. difficile strains encode two Agr loci in their genomes, designated agr1 and agr2. The agr1 locus is present in all of the C. difficile strains sequenced to date, whereas the agr2 locus is present in a few strains. The functional roles of agr1 and agr2 in C. difficile toxin regulation and pathogenesis were unknown until now. Using allelic exchange, we deleted components of both agr loci and examined the mutants for toxin production and virulence. The results showed that the agr1 mutant cannot produce toxins A and B; toxin production can be restored by complementation with wild-type agr1. Furthermore, the agr1 mutant is able to colonize but unable to cause disease in a murine CDI model. These findings have profound implications for CDI treatment because we have uncovered a promising therapeutic target for the development of nonantibiotic drugs to treat this life-threatening emerging pathogen by targeting the toxins directly responsible for disease.

  15. The MogR Transcriptional Repressor Regulates Nonhierarchal Expression of Flagellar Motility Genes and Virulence in Listeria monocytogenes.

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    2006-04-01

    Full Text Available Flagella are surface structures critical for motility and virulence of many bacterial species. In Listeria monocytogenes, MogR tightly represses expression of flagellin (FlaA during extracellular growth at 37 degrees C and during intracellular infection. MogR is also required for full virulence in a murine model of infection. Using in vitro and in vivo infection models, we determined that the severe virulence defect of MogR-negative bacteria is due to overexpression of FlaA. Specifically, overproduction of FlaA in MogR-negative bacteria caused pleiotropic defects in bacterial division (chaining phenotype, intracellular spread, and virulence in mice. DNA binding and microarray analyses revealed that MogR represses transcription of all known flagellar motility genes by binding directly to a minimum of two TTTT-N(5-AAAA recognition sites positioned within promoter regions such that RNA polymerase binding is occluded. Analysis of MogR protein levels demonstrated that modulation of MogR repression activity confers the temperature-specificity to flagellar motility gene expression. Epistasis analysis revealed that MogR repression of transcription is antagonized in a temperature-dependent manner by the DegU response regulator and that DegU further regulates FlaA levels through a posttranscriptional mechanism. These studies provide the first known example to our knowledge of a transcriptional repressor functioning as a master regulator controlling nonhierarchal expression of flagellar motility genes.

  16. Triethylene Glycol Up-Regulates Virulence-Associated Genes and Proteins in Streptococcus mutans.

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    Sadeghinejad, Lida; Cvitkovitch, Dennis G; Siqueira, Walter L; Santerre, J Paul; Finer, Yoav

    2016-01-01

    Triethylene glycol dimethacrylate (TEGDMA) is a diluent monomer used pervasively in dental composite resins. Through hydrolytic degradation of the composites in the oral cavity it yields a hydrophilic biodegradation product, triethylene glycol (TEG), which has been shown to promote the growth of Streptococcus mutans, a dominant cariogenic bacterium. Previously it was shown that TEG up-regulated gtfB, an important gene contributing to polysaccharide synthesis function in biofilms. However, molecular mechanisms related to TEG's effect on bacterial function remained poorly understood. In the present study, S. mutans UA159 was incubated with clinically relevant concentrations of TEG at pH 5.5 and 7.0. Quantitative real-time PCR, proteomics analysis, and glucosyltransferase enzyme (GTF) activity measurements were employed to identify the bacterial phenotypic response to TEG. A S. mutans vicK isogenic mutant (SMΔvicK1) and its associated complemented strain (SMΔvicK1C), an important regulatory gene for biofilm-associated genes, were used to determine if this signaling pathway was involved in modulation of the S. mutans virulence-associated genes. Extracted proteins from S. mutans biofilms grown in the presence and absence of TEG were subjected to mass spectrometry for protein identification, characterization and quantification. TEG up-regulated gtfB/C, gbpB, comC, comD and comE more significantly in biofilms at cariogenic pH (5.5) and defined concentrations. Differential response of the vicK knock-out (SMΔvicK1) and complemented strains (SMΔvicK1C) implicated this signalling pathway in TEG-modulated cellular responses. TEG resulted in increased GTF enzyme activity, responsible for synthesizing insoluble glucans involved in the formation of cariogenic biofilms. As well, TEG increased protein abundance related to biofilm formation, carbohydrate transport, acid tolerance, and stress-response. Proteomics data was consistent with gene expression findings for the selected

  17. The Hos2 Histone Deacetylase Controls Ustilago maydis Virulence through Direct Regulation of Mating-Type Genes.

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    Alberto Elías-Villalobos

    2015-08-01

    Full Text Available Morphological changes are critical for host colonisation in plant pathogenic fungi. These changes occur at specific stages of their pathogenic cycle in response to environmental signals and are mediated by transcription factors, which act as master regulators. Histone deacetylases (HDACs play crucial roles in regulating gene expression, for example by locally modulating the accessibility of chromatin to transcriptional regulators. It has been reported that HDACs play important roles in the virulence of plant fungi. However, the specific environment-sensing pathways that control fungal virulence via HDACs remain poorly characterised. Here we address this question using the maize pathogen Ustilago maydis. We find that the HDAC Hos2 is required for the dimorphic switch and pathogenic development in U. maydis. The deletion of hos2 abolishes the cAMP-dependent expression of mating type genes. Moreover, ChIP experiments detect Hos2 binding to the gene bodies of mating-type genes, which increases in proportion to their expression level following cAMP addition. These observations suggest that Hos2 acts as a downstream component of the cAMP-PKA pathway to control the expression of mating-type genes. Interestingly, we found that Clr3, another HDAC present in U. maydis, also contributes to the cAMP-dependent regulation of mating-type gene expression, demonstrating that Hos2 is not the only HDAC involved in this control system. Overall, our results provide new insights into the role of HDACs in fungal phytopathogenesis.

  18. Epigenetic control of virulence gene expression in Pseudomonas aeruginosa by a LysR-type transcription regulator.

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    Keith H Turner

    2009-12-01

    Full Text Available Phenotypic variation within an isogenic bacterial population is thought to ensure the survival of a subset of cells in adverse conditions. The opportunistic pathogen Pseudomonas aeruginosa variably expresses several phenotypes, including antibiotic resistance, biofilm formation, and the production of CupA fimbriae. Here we describe a previously unidentified bistable switch in P. aeruginosa. This switch controls the expression of a diverse set of genes, including aprA, which encodes the secreted virulence factor alkaline protease. We present evidence that bistable expression of PA2432, herein named bexR (bistable expression regulator, which encodes a LysR-type transcription regulator, controls this switch. In particular, using DNA microarrays, quantitative RT-PCR analysis, chromatin immunoprecipitation, and reporter gene fusions, we identify genes directly under the control of BexR and show that these genes are bistably expressed. Furthermore, we show that bexR is itself bistably expressed and positively autoregulated. Finally, using single-cell analyses of a GFP reporter fusion, we present evidence that positive autoregulation of bexR is necessary for bistable expression of the BexR regulon. Our findings suggest that a positive feedback loop involving a LysR-type transcription regulator serves as the basis for an epigenetic switch that controls virulence gene expression in P. aeruginosa.

  19. Structure of Vibrio cholerae ToxT reveals a mechanism for fatty acid regulation of virulence genes

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    Lowden, Michael J.; Skorupski, Karen; Pellegrini, Maria; Chiorazzo, Michael G.; Taylor, Ronald K.; Kull, F. Jon (Dartmouth)

    2010-03-04

    Cholera is an acute intestinal infection caused by the bacterium Vibrio cholerae. In order for V. cholerae to cause disease, it must produce two virulence factors, the toxin-coregulated pilus (TCP) and cholera toxin (CT), whose expression is controlled by a transcriptional cascade culminating with the expression of the AraC-family regulator, ToxT. We have solved the 1.9 {angstrom} resolution crystal structure of ToxT, which reveals folds in the N- and C-terminal domains that share a number of features in common with AraC, MarA, and Rob as well as the unexpected presence of a buried 16-carbon fatty acid, cis-palmitoleate. The finding that cis-palmitoleic acid reduces TCP and CT expression in V. cholerae and prevents ToxT from binding to DNA in vitro provides a direct link between the host environment of V. cholerae and regulation of virulence gene expression.

  20. Staphylococcus aureus Quorum Regulator SarA Targeted Compound, 2-[(Methylaminomethyl]phenol Inhibits Biofilm and Down-Regulates Virulence Genes

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    P. Balamurugan

    2017-07-01

    Full Text Available Staphylococcus aureus is a widely acknowledged Gram-positive pathogen for forming biofilm and virulence gene expressions by quorum sensing (QS, a cell to cell communication process. The quorum regulator SarA of S. aureus up-regulates the expression of many virulence factors including biofilm formation to mediate pathogenesis and evasion of the host immune system in the late phases of growth. Thus, inhibiting the production or blocking SarA protein might influence the down-regulation of biofilm and virulence factors. In this context, here we have synthesized 2-[(Methylaminomethyl]phenol, which was specifically targeted toward the quorum regulator SarA through in silico approach in our previous study. The molecule has been evaluated in vitro to validate its antibiofilm activity against clinical S. aureus strains. In addition, antivirulence properties of the inhibitor were confirmed with the observation of a significant reduction in the expression of representative virulence genes like fnbA, hla and hld that are governed under S. aureus QS. Interestingly, the SarA targeted inhibitor showed negligible antimicrobial activity and markedly reduced the minimum inhibitory concentration of conventional antibiotics when used in combination making it a more attractive lead for further clinical tests.

  1. Spaceflight Alters Bacterial Gene Expression and Virulence and Reveals Role for Global Regulator Hfq

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    Wilson, J. W.; Ott, C. M.; zuBentrup, K. Honer; Ramamurthy R.; Quick, L.; Porwollik, S.; Cheng, P.; McClellan, M.; Tsaprailis, G.; Radabaugh, T.; hide

    2007-01-01

    A comprehensive analysis of both the molecular genetic and phenotypic responses of any organism to the spaceflight environment has never been accomplished due to significant technological and logistical hurdles. Moreover, the effects of spaceflight on microbial pathogenicity and associated infectious disease risks have not been studied. The bacterial pathogen Salmonella typhimurium was grown aboard Space Shuttle mission STS-115 and compared to identical ground control cultures. Global microarray and proteomic analyses revealed 167 transcripts and 73 proteins changed expression with the conserved RNA-binding protein Hfq identified as a likely global regulator involved in the response to this environment. Hfq involvement was confirmed with a ground based microgravity culture model. Spaceflight samples exhibited enhanced virulence in a murine infection model and extracellular matrix accumulation consistent with a biofilm. Strategies to target Hfq and related regulators could potentially decrease infectious disease risks during spaceflight missions and provide novel therapeutic options on Earth.

  2. SuhB Is a Regulator of Multiple Virulence Genes and Essential for Pathogenesis of Pseudomonas aeruginosa

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    Li, Kewei; Xu, Chang; Jin, Yongxin; Sun, Ziyu; Liu, Chang; Shi, Jing; Chen, Gukui; Chen, Ronghao; Jin, Shouguang; Wu, Weihui

    2013-01-01

    ABSTRACT During initial colonization and chronic infection, pathogenic bacteria encounter distinct host environments. Adjusting gene expression accordingly is essential for the pathogenesis. Pseudomonas aeruginosa has evolved complicated regulatory networks to regulate different sets of virulence factors to facilitate colonization and persistence. The type III secretion system (T3SS) and motility are associated with acute infections, while biofilm formation and the type VI secretion system (T6SS) are associated with chronic persistence. To identify novel regulatory genes required for pathogenesis, we screened a P. aeruginosa transposon (Tn) insertion library and found suhB to be an essential gene for the T3SS gene expression. The expression of suhB was upregulated in a mouse acute lung infection model, and loss of suhB resulted in avirulence. Suppression of T3SS gene expression in the suhB mutant is linked to a defective translation of the T3SS master regulator, ExsA. Further studies demonstrated that suhB mutation led to the upregulation of GacA and its downstream small RNAs, RsmY and RsmZ, triggering T6SS expression and biofilm formation while inhibiting the T3SS. Our results demonstrate that an in vivo-inducible gene, suhB, reciprocally regulates genes associated with acute and chronic infections and plays an essential role in the pathogenesis of P. aeruginosa. PMID:24169572

  3. Involvement of Trichoderma harzianum Epl-1 Protein in the Regulation of Botrytis Virulence- and Tomato Defense-Related Genes

    Directory of Open Access Journals (Sweden)

    Eriston V. Gomes

    2017-05-01

    Full Text Available Several Trichoderma spp. are well known for their ability to: (i act as important biocontrol agents against phytopathogenic fungi; (ii function as biofertilizers; (iii increase the tolerance of plants to biotic and abiotic stresses; and (iv induce plant defense responses via the production and secretion of elicitor molecules. In this study, we analyzed the gene-regulation effects of Trichoderma harzianum Epl-1 protein during the interactions of mutant Δepl-1 or wild-type T. harzianum strains with: (a the phytopathogen Botrytis cinerea and (b with tomato plants, on short (24 h hydroponic cultures and long periods (4-weeks old plants after Trichoderma inoculation. Our results indicate that T. harzianum Epl-1 protein affects the in vitro expression of B. cinerea virulence genes, especially those involved in the botrydial biosynthesis (BcBOT genes, during the mycoparasitism interaction. The tomato defense-related genes were also affected, indicating that Epl-1 is involved in the elicitation of the salicylic acid pathway. Moreover, Epl-1 also regulates the priming effect in host tomato plants and contributes to enhance the interaction with the host tomato plant during the early stage of root colonization.

  4. Involvement of Trichoderma harzianum Epl-1 Protein in the Regulation of Botrytis Virulence- and Tomato Defense-Related Genes.

    Science.gov (United States)

    Gomes, Eriston V; Ulhoa, Cirano J; Cardoza, Rosa E; Silva, Roberto N; Gutiérrez, Santiago

    2017-01-01

    Several Trichoderma spp. are well known for their ability to: (i) act as important biocontrol agents against phytopathogenic fungi; (ii) function as biofertilizers; (iii) increase the tolerance of plants to biotic and abiotic stresses; and (iv) induce plant defense responses via the production and secretion of elicitor molecules. In this study, we analyzed the gene-regulation effects of Trichoderma harzianum Epl-1 protein during the interactions of mutant Δ epl-1 or wild-type T. harzianum strains with: (a) the phytopathogen Botrytis cinerea and (b) with tomato plants, on short (24 h hydroponic cultures) and long periods (4-weeks old plants) after Trichoderma inoculation. Our results indicate that T. harzianum Epl-1 protein affects the in vitro expression of B. cinerea virulence genes, especially those involved in the botrydial biosynthesis ( BcBOT genes), during the mycoparasitism interaction. The tomato defense-related genes were also affected, indicating that Epl-1 is involved in the elicitation of the salicylic acid pathway. Moreover, Epl-1 also regulates the priming effect in host tomato plants and contributes to enhance the interaction with the host tomato plant during the early stage of root colonization.

  5. Exploring potential virulence regulators in Paracoccidioides brasiliensis isolates of varying virulence through quantitative proteomics.

    Science.gov (United States)

    Castilho, Daniele G; Chaves, Alison F A; Xander, Patricia; Zelanis, André; Kitano, Eduardo S; Serrano, Solange M T; Tashima, Alexandre K; Batista, Wagner L

    2014-10-03

    Few virulence factors have been identified for Paracoccidioides brasiliensis, the agent of paracoccidioidomycosis. In this study, we quantitatively evaluated the protein composition of P. brasiliensis in the yeast phase using minimal and rich media to obtain a better understanding of its virulence and to gain new insights into pathogen adaptation strategies. This analysis was performed on two isolates of the Pb18 strain showing distinct infection profiles in B10.A mice. Using liquid chromatography/tandem mass spectrometry (LC-MS/MS) analysis, we identified and quantified 316 proteins in minimal medium, 29 of which were overexpressed in virulent Pb18. In rich medium, 29 out of 295 proteins were overexpressed in the virulent fungus. Three proteins were found to be up-regulated in both media, suggesting the potential roles of these proteins in virulence regulation in P. brasiliensis. Moreover, genes up-regulated in virulent Pb18 showed an increase in its expression after the recovery of virulence of attenuated Pb18. Proteins up-regulated in both isolates were grouped according to their functional categories. Virulent Pb18 undergoes metabolic reorganization and increased expression of proteins involved in fermentative respiration. This approach allowed us to identify potential virulence regulators and provided a foundation for achieving a molecular understanding of how Paracoccidioides modulates the host-pathogen interaction to its advantage.

  6. Control of Virulence Gene Expression by the Master Regulator, CfaD, in the Prototypical Enterotoxigenic Escherichia coli Strain, H10407

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    Carla Hodson

    2017-08-01

    Full Text Available Enterotoxigenic Escherichia coli (ETEC is the most common bacterial cause of diarrhea in children in developing countries, as well as in travelers to these countries. To cause disease, ETEC needs to produce a series of virulence proteins including enterotoxins, colonization factors and secretion pathways, which enable this pathogen to colonize the human small intestine and deliver enterotoxins to epithelial cells. Previously, a number of studies have demonstrated that CfaD, an AraC-like transcriptional regulator, plays a key role in virulence gene expression by ETEC. In this study, we carried out a transcriptomic analysis of ETEC strain, H10407, grown under different conditions, and determined the complete set of genes that are regulated by CfaD. In this way, we identified a number of new target genes, including rnr-1, rnr-2, etpBAC, agn43, flu, traM and ETEC_3214, whose expression is strongly activated by CfaD. Using promoter-lacZ reporters, primer extension and electrophoretic mobility shift assays, we characterized the CfaD-mediated activation of several selected target promoters. We also showed that the gut-associated environmental signal, sodium bicarbonate, stimulates CfaD-mediated upregulation of its virulence target operons. Finally, we screened a commercial small molecule library and identified a compound (CH-1 that specifically inhibited the regulatory function of CfaD, and by 2-D analoging, we identified a second inhibitor (CH-2 with greater potency.

  7. The Acinetobacter baumannii Two-Component System AdeRS Regulates Genes Required for Multidrug Efflux, Biofilm Formation, and Virulence in a Strain-Specific Manner

    Directory of Open Access Journals (Sweden)

    Grace E. Richmond

    2016-04-01

    Full Text Available The opportunistic pathogen Acinetobacter baumannii is able to persist in the environment and is often multidrug resistant (MDR, causing difficulties in the treatment of infections. Here, we show that the two-component system AdeRS, which regulates the production of the AdeABC multidrug resistance efflux pump, is required for the formation of a protective biofilm in an ex vivo porcine mucosal model, which mimics a natural infection of the human epithelium. Interestingly, deletion of adeB impacted only on the ability of strain AYE to form a biofilm on plastic and only on the virulence of strain Singapore 1 for Galleria mellonella. RNA-Seq revealed that loss of AdeRS or AdeB significantly altered the transcriptional landscape, resulting in the changed expression of many genes, notably those associated with antimicrobial resistance and virulence interactions. For example, A. baumannii lacking AdeRS displayed decreased expression of adeABC, pil genes, com genes, and a pgaC-like gene, whereas loss of AdeB resulted in increased expression of pil and com genes and decreased expression of ferric acinetobactin transport system genes. These data define the scope of AdeRS-mediated regulation, show that changes in the production of AdeABC mediate important phenotypes controlled by AdeRS, and suggest that AdeABC is a viable target for antimicrobial drug and antibiofilm discovery.

  8. RegA, an AraC-Like Protein, Is a Global Transcriptional Regulator That Controls Virulence Gene Expression in Citrobacter rodentium▿

    Science.gov (United States)

    Hart, Emily; Yang, Ji; Tauschek, Marija; Kelly, Michelle; Wakefield, Matthew J.; Frankel, Gad; Hartland, Elizabeth L.; Robins-Browne, Roy M.

    2008-01-01

    Citrobacter rodentium is an attaching and effacing pathogen which causes transmissible colonic hyperplasia in mice. Infection with C. rodentium serves as a model for infection of humans with enteropathogenic and enterohemorrhagic Escherichia coli. To identify novel colonization factors of C. rodentium, we screened a signature-tagged mutant library of C. rodentium in mice. One noncolonizing mutant had a single transposon insertion in an open reading frame (ORF) which we designated regA because of its homology to genes encoding members of the AraC family of transcriptional regulators. Deletion of regA in C. rodentium resulted in markedly reduced colonization of the mouse intestine. Examination of lacZ transcriptional fusions using promoter regions of known and putative virulence-associated genes of C. rodentium revealed that RegA strongly stimulated transcription of two newly identified genes located close to regA, which we designated adcA and kfcC. The cloned adcA gene conferred autoaggregation and adherence to mammalian cells to E. coli strain DH5α, and a kfc mutation led to a reduction in the duration of intestinal colonization, but the kfc mutant was far less attenuated than the regA mutant. These results indicated that other genes of C. rodentium whose expression required activation by RegA were required for colonization. Microarray analysis revealed a number of RegA-regulated ORFs encoding proteins homologous to known colonization factors. Transcription of these putative virulence determinants was activated by RegA only in the presence of sodium bicarbonate. Taken together, these results show that RegA is a global regulator of virulence in C. rodentium which activates factors that are required for intestinal colonization. PMID:18765720

  9. RegA, an AraC-like protein, is a global transcriptional regulator that controls virulence gene expression in Citrobacter rodentium.

    Science.gov (United States)

    Hart, Emily; Yang, Ji; Tauschek, Marija; Kelly, Michelle; Wakefield, Matthew J; Frankel, Gad; Hartland, Elizabeth L; Robins-Browne, Roy M

    2008-11-01

    Citrobacter rodentium is an attaching and effacing pathogen which causes transmissible colonic hyperplasia in mice. Infection with C. rodentium serves as a model for infection of humans with enteropathogenic and enterohemorrhagic Escherichia coli. To identify novel colonization factors of C. rodentium, we screened a signature-tagged mutant library of C. rodentium in mice. One noncolonizing mutant had a single transposon insertion in an open reading frame (ORF) which we designated regA because of its homology to genes encoding members of the AraC family of transcriptional regulators. Deletion of regA in C. rodentium resulted in markedly reduced colonization of the mouse intestine. Examination of lacZ transcriptional fusions using promoter regions of known and putative virulence-associated genes of C. rodentium revealed that RegA strongly stimulated transcription of two newly identified genes located close to regA, which we designated adcA and kfcC. The cloned adcA gene conferred autoaggregation and adherence to mammalian cells to E. coli strain DH5alpha, and a kfc mutation led to a reduction in the duration of intestinal colonization, but the kfc mutant was far less attenuated than the regA mutant. These results indicated that other genes of C. rodentium whose expression required activation by RegA were required for colonization. Microarray analysis revealed a number of RegA-regulated ORFs encoding proteins homologous to known colonization factors. Transcription of these putative virulence determinants was activated by RegA only in the presence of sodium bicarbonate. Taken together, these results show that RegA is a global regulator of virulence in C. rodentium which activates factors that are required for intestinal colonization.

  10. The effect of mutation on Rhodococcus equi virulence plasmid gene expression and mouse virulence.

    Science.gov (United States)

    Ren, Jun; Prescott, John F

    2004-11-15

    An 81 kb virulence plasmid containing a pathogenicity island (PI) plays a crucial role in the pathogenesis of Rhodococcus equi pneumonia in foals but its specific function in virulence and regulation of plasmid-encoded virulence genes is unclear. Using a LacZ selection marker developed for R. equi in this study, in combination with an apramycin resistance gene, an efficient two-stage homologous recombination targeted gene mutation procedure was used to mutate three virulence plasmid genes, a LysR regulatory gene homologue (ORF4), a ResD-like two-component response regulator homologue (ORF8), and a gene (ORF10) of unknown function that is highly expressed by R. equi inside macrophages, as well as the chromosomal gene operon, phoPR. Virulence testing by liver clearance after intravenous injection in mice showed that the ORF4 and ORF8 mutants were fully attenuated, that the phoPR mutant was hypervirulent, and that virulence of the ORF10 mutant remained unchanged. A virulence plasmid DNA microarray was used to compare the plasmid gene expression profile of each of the four gene-targeted mutants against the parental R. equi strain. Changes were limited to PI genes and gene induction was observed for all mutants, suggesting that expression of virulence plasmid genes is dominated by a negative regulatory network. The finding of attenuation of ORF4 and ORF8 mutants despite enhanced transcription of vapA suggests that factors other than VapA are important for full expression of virulence. ORF1, a putative Lsr antigen gene, was strongly and similarly induced in all mutants, implying a common regulatory pathway affecting this gene for all four mutated genes. ORF8 is apparently the centre of this common pathway. Two distinct highly correlated gene induction patterns were observed, that of the ORF4 and ORF8 mutants, and that of the ORF10 and phoPR mutants. The gene induction pattern distinguishing these two groups paralleled their virulence in mice.

  11. Temperature Regulation of Shigella Virulence: Identification of Temperature-Regulated Shigella Invasion Genes by the Isolation of inv::lacZ Operon Fusions and the Characterization of the Virulence Gene Regulator virR

    Science.gov (United States)

    1991-04-10

    conserved in virulent strains of Shigella and EIEC. Evaluation of the serum Immune response to Shigella proteins in Rhesus monkeys and humans revealed...sera from both humans and monkeys following a Shigella infection (Hale et al. , 1985; Oaks et al., 1986). Moreover, Tn5 insertions In various...Microbiology, Washington, D.C. 108. Oaks, E, V,, T, L. Hale, and S. B. Formal. 1986. Serum Immune response to Shigella protein antigens In Rhesus monkeys

  12. Single Nucleotide Polymorphisms in Regulator-Encoding Genes Have an Additive Effect on Virulence Gene Expression in a Vibrio cholerae Clinical Isolate.

    Science.gov (United States)

    Carignan, Bailey M; Brumfield, Kyle D; Son, Mike S

    2016-01-01

    Vibrio cholerae is the etiological agent of the infectious disease cholera, which is characterized by vomiting and severe watery diarrhea. Recently, V. cholerae clinical isolates have demonstrated increased virulence capabilities, causing more severe symptoms with a much higher rate of disease progression than previously observed. We have identified single nucleotide polymorphisms (SNPs) in four virulence-regulatory genes (hapR, hns, luxO, and vieA) of a hypervirulent V. cholerae clinical isolate, MQ1795. Herein, all SNPs and SNP combinations of interest were introduced into the prototypical El Tor reference strain N16961, and the effects on the production of numerous virulence-related factors, including cholera toxin (CT), the toxin-coregulated pilus (TCP), and ToxT, were analyzed. Our data show that triple-SNP (hapR hns luxO and hns luxO vieA) and quadruple-SNP combinations produced the greatest increases in CT, TCP, and ToxT production. The hns and hns luxO SNP combinations were sufficient for increased TCP and ToxT production. Notably, the hns luxO vieA triple-SNP combination strain produced TCP and ToxT levels similar to those of MQ1795. Certain SNP combinations (hapR and hapR vieA) had the opposite effect on CT, TCP, and ToxT expression. Interestingly, the hns vieA double-SNP combination strain increased TCP production while decreasing CT production. Our findings suggest that SNPs identified in the four regulatory genes, in various combinations, are associated with increased virulence capabilities observed in V. cholerae clinical isolates. These studies provide insight into the evolution of highly virulent strains. IMPORTANCE Cholera, an infectious disease of the small intestine caused by the aquatic bacterium Vibrio cholerae, often results in vomiting and acute watery diarrhea. If left untreated or if the response is too slow, the symptoms can quickly lead to extreme dehydration and ultimately death of the patient. Recent anecdotal evidence of cholera

  13. Virulence regulation in Staphylococcus aureus: the need for in vivo analysis of virulence factor regulation.

    Science.gov (United States)

    Pragman, Alexa A; Schlievert, Patrick M

    2004-10-01

    Staphylococcus aureus is a pathogenic microorganism that is responsible for a wide variety of clinical infections. These infections can be relatively mild, but serious, life-threatening infections may result from the expression of staphylococcal virulence factors that are coordinated by virulence regulators. Much work has been done to characterize the actions of staphylococcal virulence regulators in broth culture. Recently, several laboratories showed that transcriptional analyses of virulence regulators in in vivo animal models or in human infection did not correlate with transcriptional analyses accomplished in vitro. In describing the differences between in vitro and in vivo transcription of staphylococcal virulence regulators, we hope to encourage investigators to study virulence regulators using infection models whenever possible.

  14. Transcription and translation products of the cytolysin gene psm-mec on the mobile genetic element SCCmec regulate Staphylococcus aureus virulence.

    Directory of Open Access Journals (Sweden)

    Chikara Kaito

    Full Text Available The F region downstream of the mecI gene in the SCCmec element in hospital-associated methicillin-resistant Staphylococcus aureus (HA-MRSA contains two bidirectionally overlapping open reading frames (ORFs, the fudoh ORF and the psm-mec ORF. The psm-mec ORF encodes a cytolysin, phenol-soluble modulin (PSM-mec. Transformation of the F region into the Newman strain, which is a methicillin-sensitive S. aureus (MSSA strain, or into the MW2 (USA400 and FRP3757 (USA300 strains, which are community-acquired MRSA (CA-MRSA strains that lack the F region, attenuated their virulence in a mouse systemic infection model. Introducing the F region to these strains suppressed colony-spreading activity and PSMα production, and promoted biofilm formation. By producing mutations into the psm-mec ORF, we revealed that (i both the transcription and translation products of the psm-mec ORF suppressed colony-spreading activity and promoted biofilm formation; and (ii the transcription product of the psm-mec ORF, but not its translation product, decreased PSMα production. These findings suggest that both the psm-mec transcript, acting as a regulatory RNA, and the PSM-mec protein encoded by the gene on the mobile genetic element SCCmec regulate the virulence of Staphylococcus aureus.

  15. Involvement of Trichoderma harzianum Epl-1 Protein in the Regulation of Botrytis Virulence- and Tomato Defense-Related Genes

    OpenAIRE

    Gomes, Eriston V.; Ulhoa, Cirano J.; Cardoza, Rosa E.; Silva, Roberto N.; Guti?rrez, Santiago

    2017-01-01

    Several Trichoderma spp. are well known for their ability to: (i) act as important biocontrol agents against phytopathogenic fungi; (ii) function as biofertilizers; (iii) increase the tolerance of plants to biotic and abiotic stresses; and (iv) induce plant defense responses via the production and secretion of elicitor molecules. In this study, we analyzed the gene-regulation effects of Trichoderma harzianum Epl-1 protein during the interactions of mutant Δepl-1 or wild-type T. harzianum stra...

  16. Pathogenomic inference of virulence-associated genes in Leptospira interrogans.

    Directory of Open Access Journals (Sweden)

    Jason S Lehmann

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

  17. Pathogenomic inference of virulence-associated genes in Leptospira interrogans.

    Science.gov (United States)

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

    2013-01-01

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

  18. Genetic Regulation of Virulence and Antibiotic Resistance in Acinetobacter baumannii

    Science.gov (United States)

    Kröger, Carsten; Kary, Stefani C.; Schauer, Kristina; Cameron, Andrew D. S.

    2016-01-01

    Multidrug resistant microorganisms are forecast to become the single biggest challenge to medical care in the 21st century. Over the last decades, members of the genus Acinetobacter have emerged as bacterial opportunistic pathogens, in particular as challenging nosocomial pathogens because of the rapid evolution of antimicrobial resistances. Although we lack fundamental biological insight into virulence mechanisms, an increasing number of researchers are working to identify virulence factors and to study antibiotic resistance. Here, we review current knowledge regarding the regulation of virulence genes and antibiotic resistance in Acinetobacter baumannii. A survey of the two-component systems AdeRS, BaeSR, GacSA and PmrAB explains how each contributes to antibiotic resistance and virulence gene expression, while BfmRS regulates cell envelope structures important for pathogen persistence. A. baumannii uses the transcription factors Fur and Zur to sense iron or zinc depletion and upregulate genes for metal scavenging as a critical survival tool in an animal host. Quorum sensing, nucleoid-associated proteins, and non-classical transcription factors such as AtfA and small regulatory RNAs are discussed in the context of virulence and antibiotic resistance. PMID:28036056

  19. Genes for carbon metabolism and the ToxA virulence factor in Pseudomonas aeruginosa are regulated through molecular interactions of PtxR and PtxS.

    Directory of Open Access Journals (Sweden)

    Abdelali Daddaoua

    Full Text Available Homologs of the transcriptional regulator PtxS are omnipresent in Pseudomonas, whereas PtxR homologues are exclusively found in human pathogenic Pseudomonas species. In all Pseudomonas sp., PtxS with 2-ketogluconate is the regulator of the gluconate degradation pathway and controls expression from its own promoter and also from the P(gad and P(kgu for the catabolic operons. There is evidence that PtxS and PtxR play a central role in the regulation of exotoxin A expression, a relevant primary virulence factor of Pseudomonas aeruginosa. We show using DNaseI-footprint analysis that in P. aeruginosa PtxR binds to the -35 region of the P(toxA promoter in front of the exotoxin A gene, whereas PtxS does not bind to this promoter. Bioinformatic and DNaseI-footprint analysis identified a PtxR binding site in the P(kgu and P(gad promoters that overlaps the -35 region, while the PtxS operator site is located 50 bp downstream from the PtxR site. In vitro, PtxS recognises PtxR with nanomolar affinity, but this interaction does not occur in the presence of 2-ketogluconate, the specific effector of PtxS. DNAaseI footprint assays of P(kgu and P(gad promoters with PtxS and PtxR showed a strong region of hyper-reactivity between both regulator binding sites, indicative of DNA distortion when both proteins are bound; however in the presence of 2-ketogluconate no protection was observed. We conclude that PtxS modulates PtxR activity in response to 2-ketogluconate by complex formation in solution in the case of the P(toxA promoter, or via the formation of a DNA loop as in the regulation of gluconate catabolic genes. Data suggest two different mechanisms of control exerted by the same regulator.

  20. Regulation of the Vibrio vulnificus hupA gene by temperature alteration and cyclic AMP receptor protein and evaluation of its role in virulence.

    Science.gov (United States)

    Oh, Man Hwan; Lee, Sung Min; Lee, Dong Hwan; Choi, Sang Ho

    2009-03-01

    Availability of free iron is extremely limited in the mammalian host, and the acquisition of iron in the host is essential for successful infection by pathogenic bacteria. Expression of many genes involved in acquiring iron is regulated in response to the level of iron availability, and iron regulation is mediated by Fur. In this study, cellular levels of Vibrio vulnificus HupA, a heme receptor protein, and the hupA transcript were found to increase in cells grown at 40 degrees C compared to cells grown at 30 degrees C. The results suggested that change in growth temperature, in addition to iron availability, is an environmental cue controlling the expression of the hupA gene. The influence of global regulatory proteins on the expression of hupA was examined, and the cyclic AMP receptor protein (CRP) was found to activate the expression of hupA at the transcriptional level. CRP exerts its effects by directly binding to DNA upstream of the hupA promoter P(hupA), and a CRP binding site, centered at 174 bp upstream of the transcription start site, was identified by a DNase I protection assay. Finally, a hupA mutant showed reduced virulence in mice and in tissue cultures, in which growth of the hupA mutant was impaired, indicating that HupA of V. vulnificus is essential for survival and multiplication during infection.

  1. ClpP deletion causes attenuation of Salmonella Typhimurium virulence through mis-regulation of RpoS and indirect control of CsrA and the SPI genes

    DEFF Research Database (Denmark)

    Knudsen, Gitte Maegaard; Olsen, John E.; Aabo, Søren

    2013-01-01

    , suggesting the repression of invasion was directed through RpoS. The expression of the csrA virulence regulator was increased in the ΔclpP mutant and decreased in the rpoS : : amp and ΔclpP/rpoS : : amp mutants, indicating that ClpP affects the csrA expression level as well. Thus, this study suggests...... the proteolytic component ClpP, the stationary phase regulator RpoS and the carbon-storage regulator CsrA. However, the mechanism behind the ClpP regulation is not fully understood. To elucidate this we examined differentially expressed genes in a ΔclpP mutant compared with WT using global transcriptomic analysis...... that ClpP affects SPI1 expression and thereby virulence indirectly through its regulation of both RpoS and CsrA....

  2. Transcriptional Analysis of the MrpJ Network: Modulation of Diverse Virulence-Associated Genes and Direct Regulation of mrp Fimbrial and flhDC Flagellar Operons in Proteus mirabilis

    Science.gov (United States)

    Bode, Nadine J.; Debnath, Irina; Kuan, Lisa; Schulfer, Anjelique; Ty, Maureen

    2015-01-01

    The enteric bacterium Proteus mirabilis is associated with a significant number of catheter-associated urinary tract infections (UTIs). Strict regulation of the antagonistic processes of adhesion and motility, mediated by fimbriae and flagella, respectively, is essential for disease progression. Previously, the transcriptional regulator MrpJ, which is encoded by the mrp fimbrial operon, has been shown to repress both swimming and swarming motility. Here we show that MrpJ affects an array of cellular processes beyond adherence and motility. Microarray analysis found that expression of mrpJ mimicking levels observed during UTIs leads to differential expression of 217 genes related to, among other functions, bacterial virulence, type VI secretion, and metabolism. We probed the molecular mechanism of transcriptional regulation by MrpJ using transcriptional reporters and chromatin immunoprecipitation (ChIP). Binding of MrpJ to two virulence-associated target gene promoters, the promoters of the flagellar master regulator flhDC and mrp itself, appears to be affected by the condensation state of the native chromosome, although both targets share a direct MrpJ binding site proximal to the transcriptional start. Furthermore, an mrpJ deletion mutant colonized the bladders of mice at significantly lower levels in a transurethral model of infection. Additionally, we observed that mrpJ is widely conserved in a collection of recent clinical isolates. Altogether, these findings support a role of MrpJ as a global regulator of P. mirabilis virulence. PMID:25847961

  3. Phosphotyrosine-Mediated Regulation of Enterohemorrhagic Escherichia coli Virulence

    Science.gov (United States)

    Robertson, Colin D.; Hazen, Tracy H.; Kaper, James B.

    2018-01-01

    ABSTRACT Enteric pathogens with low infectious doses rely on the ability to orchestrate the expression of virulence and metabolism-associated genes in response to environmental cues for successful infection. Accordingly, the human pathogen enterohemorrhagic Escherichia coli (EHEC) employs a complex multifaceted regulatory network to link the expression of type III secretion system (T3SS) components to nutrient availability. While phosphorylation of histidine and aspartate residues on two-component system response regulators is recognized as an integral part of bacterial signaling, the involvement of phosphotyrosine-mediated control is minimally explored in Gram-negative pathogens. Our recent phosphotyrosine profiling study of E. coli identified 342 phosphorylated proteins, indicating that phosphotyrosine modifications in bacteria are more prevalent than previously anticipated. The present study demonstrates that tyrosine phosphorylation of a metabolite-responsive LacI/GalR family regulator, Cra, negatively affects T3SS expression under glycolytic conditions that are typical for the colonic lumen environment where production of the T3SS is unnecessary. Our data suggest that Cra phosphorylation affects T3SS expression by modulating the expression of ler, which encodes the major activator of EHEC virulence gene expression. Phosphorylation of the Cra Y47 residue diminishes DNA binding to fine-tune the expression of virulence-associated genes, including those of the locus of enterocyte effacement pathogenicity island that encode the T3SS, and thereby negatively affects the formation of attaching and effacing lesions. Our data indicate that tyrosine phosphorylation provides an additional mechanism to control the DNA binding of Cra and other LacI/GalR family regulators, including LacI and PurR. This study describes an initial effort to unravel the role of global phosphotyrosine signaling in the control of EHEC virulence potential. PMID:29487233

  4. Phosphotyrosine-Mediated Regulation of Enterohemorrhagic Escherichia coli Virulence

    Directory of Open Access Journals (Sweden)

    Colin D. Robertson

    2018-02-01

    Full Text Available Enteric pathogens with low infectious doses rely on the ability to orchestrate the expression of virulence and metabolism-associated genes in response to environmental cues for successful infection. Accordingly, the human pathogen enterohemorrhagic Escherichia coli (EHEC employs a complex multifaceted regulatory network to link the expression of type III secretion system (T3SS components to nutrient availability. While phosphorylation of histidine and aspartate residues on two-component system response regulators is recognized as an integral part of bacterial signaling, the involvement of phosphotyrosine-mediated control is minimally explored in Gram-negative pathogens. Our recent phosphotyrosine profiling study of E. coli identified 342 phosphorylated proteins, indicating that phosphotyrosine modifications in bacteria are more prevalent than previously anticipated. The present study demonstrates that tyrosine phosphorylation of a metabolite-responsive LacI/GalR family regulator, Cra, negatively affects T3SS expression under glycolytic conditions that are typical for the colonic lumen environment where production of the T3SS is unnecessary. Our data suggest that Cra phosphorylation affects T3SS expression by modulating the expression of ler, which encodes the major activator of EHEC virulence gene expression. Phosphorylation of the Cra Y47 residue diminishes DNA binding to fine-tune the expression of virulence-associated genes, including those of the locus of enterocyte effacement pathogenicity island that encode the T3SS, and thereby negatively affects the formation of attaching and effacing lesions. Our data indicate that tyrosine phosphorylation provides an additional mechanism to control the DNA binding of Cra and other LacI/GalR family regulators, including LacI and PurR. This study describes an initial effort to unravel the role of global phosphotyrosine signaling in the control of EHEC virulence potential.

  5. Catabolite and Oxygen Regulation of Enterohemorrhagic Escherichia coli Virulence

    Directory of Open Access Journals (Sweden)

    Kimberly M. Carlson-Banning

    2016-11-01

    Full Text Available The biogeography of the gut is diverse in its longitudinal axis, as well as within specific microenvironments. Differential oxygenation and nutrient composition drive the membership of microbial communities in these habitats. Moreover, enteric pathogens can orchestrate further modifications to gain a competitive advantage toward host colonization. These pathogens are versatile and adept when exploiting the human colon. They expertly navigate complex environmental cues and interkingdom signaling to colonize and infect their hosts. Here we demonstrate how enterohemorrhagic Escherichia coli (EHEC uses three sugar-sensing transcription factors, Cra, KdpE, and FusR, to exquisitely regulate the expression of virulence factors associated with its type III secretion system (T3SS when exposed to various oxygen concentrations. We also explored the effect of mucin-derived nonpreferred carbon sources on EHEC growth and expression of virulence genes. Taken together, the results show that EHEC represses the expression of its T3SS when oxygen is absent, mimicking the largely anaerobic lumen, and activates its T3SS when oxygen is available through Cra. In addition, when EHEC senses mucin-derived sugars heavily present in the O-linked and N-linked glycans of the large intestine, virulence gene expression is initiated. Sugars derived from pectin, a complex plant polysaccharide digested in the large intestine, also increased virulence gene expression. Not only does EHEC sense host- and microbiota-derived interkingdom signals, it also uses oxygen availability and mucin-derived sugars liberated by the microbiota to stimulate expression of the T3SS. This precision in gene regulation allows EHEC to be an efficient pathogen with an extremely low infectious dose.

  6. Regulation of bacterial virulence by Csr (Rsm) systems.

    Science.gov (United States)

    Vakulskas, Christopher A; Potts, Anastasia H; Babitzke, Paul; Ahmer, Brian M M; Romeo, Tony

    2015-06-01

    Most bacterial pathogens have the remarkable ability to flourish in the external environment and in specialized host niches. This ability requires their metabolism, physiology, and virulence factors to be responsive to changes in their surroundings. It is no surprise that the underlying genetic circuitry that supports this adaptability is multilayered and exceedingly complex. Studies over the past 2 decades have established that the CsrA/RsmA proteins, global regulators of posttranscriptional gene expression, play important roles in the expression of virulence factors of numerous proteobacterial pathogens. To accomplish these tasks, CsrA binds to the 5' untranslated and/or early coding regions of mRNAs and alters translation, mRNA turnover, and/or transcript elongation. CsrA activity is regulated by noncoding small RNAs (sRNAs) that contain multiple CsrA binding sites, which permit them to sequester multiple CsrA homodimers away from mRNA targets. Environmental cues sensed by two-component signal transduction systems and other regulatory factors govern the expression of the CsrA-binding sRNAs and, ultimately, the effects of CsrA on secretion systems, surface molecules and biofilm formation, quorum sensing, motility, pigmentation, siderophore production, and phagocytic avoidance. This review presents the workings of the Csr system, the paradigm shift that it generated for understanding posttranscriptional regulation, and its roles in virulence networks of animal and plant pathogens. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  7. Systems analysis of multiple regulator perturbations allows discovery of virulence factors in Salmonella

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Hyunjin; Ansong, Charles; McDermott, Jason E.; Gritsenko, Marina A.; Smith, Richard D.; Heffron, Fred; Adkins, Joshua N.

    2011-06-28

    Background: Systemic bacterial infections are highly regulated and complex processes that are orchestrated by numerous virulence factors. Genes that are coordinately controlled by the set of regulators required for systemic infection are potentially required for pathogenicity. Results: In this study we present a systems biology approach in which sample-matched multi-omic measurements of fourteen virulence-essential regulator mutants were coupled with computational network analysis to efficiently identify Salmonella virulence factors. Immunoblot experiments verified network-predicted virulence factors and a subset was determined to be secreted into the host cytoplasm, suggesting that they are virulence factors directly interacting with host cellular components. Two of these, SrfN and PagK2, were required for full mouse virulence and were shown to be translocated independent of either of the type III secretion systems in Salmonella or the type III injectisome-related flagellar mechanism. Conclusions: Integrating multi-omic datasets from Salmonella mutants lacking virulence regulators not only identified novel virulence factors but also defined a new class of translocated effectors involved in pathogenesis. The success of this strategy at discovery of known and novel virulence factors suggests that the approach may have applicability for other bacterial pathogens.

  8. Transcriptional analysis of the MrpJ network: modulation of diverse virulence-associated genes and direct regulation of mrp fimbrial and flhDC flagellar operons in Proteus mirabilis.

    Science.gov (United States)

    Bode, Nadine J; Debnath, Irina; Kuan, Lisa; Schulfer, Anjelique; Ty, Maureen; Pearson, Melanie M

    2015-06-01

    The enteric bacterium Proteus mirabilis is associated with a significant number of catheter-associated urinary tract infections (UTIs). Strict regulation of the antagonistic processes of adhesion and motility, mediated by fimbriae and flagella, respectively, is essential for disease progression. Previously, the transcriptional regulator MrpJ, which is encoded by the mrp fimbrial operon, has been shown to repress both swimming and swarming motility. Here we show that MrpJ affects an array of cellular processes beyond adherence and motility. Microarray analysis found that expression of mrpJ mimicking levels observed during UTIs leads to differential expression of 217 genes related to, among other functions, bacterial virulence, type VI secretion, and metabolism. We probed the molecular mechanism of transcriptional regulation by MrpJ using transcriptional reporters and chromatin immunoprecipitation (ChIP). Binding of MrpJ to two virulence-associated target gene promoters, the promoters of the flagellar master regulator flhDC and mrp itself, appears to be affected by the condensation state of the native chromosome, although both targets share a direct MrpJ binding site proximal to the transcriptional start. Furthermore, an mrpJ deletion mutant colonized the bladders of mice at significantly lower levels in a transurethral model of infection. Additionally, we observed that mrpJ is widely conserved in a collection of recent clinical isolates. Altogether, these findings support a role of MrpJ as a global regulator of P. mirabilis virulence. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

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

    Science.gov (United States)

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

    2010-12-01

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

  10. Molecular Detection of Virulence Genes and Antibiotic Resistance ...

    African Journals Online (AJOL)

    Pathogen, E. coli O157:H7, virulence genes, antibiotic-resistance, beef meat. Correspondence: ... box to the laboratory for further processing. Isolation and identification of ... Technologies (IDT) Inc, U.S.A. The sequences and annealing ...

  11. Regulators Involved in Dickeya solani Virulence, Genetic Conservation and Functional Variability.

    Science.gov (United States)

    Potrykus, Marta; Golanowska, Małgorzata; Hugouvieux-Cotte-Pattat, Nicole; Lojkowska, Ewa

    2015-01-01

    Bacteria from the genus Dickeya (formerly Erwinia chrysanthemi) are plant pathogens causing severe diseases in many economically important crops. A majority of the strains responsible for potato disease in Europe belong to a newly identified Dickeya solani species. Although some ecological and epidemiological studies have been carried out, little is known about the regulation of D. solani virulence. The characterization of four D. solani strains indicates significant differences in their virulence on potato although they are genetically similar based on genomic fingerprinting profiles. A phenotypic examination included an analysis of virulence on potato, growth rate in culture, motility, Fe 3+ chelation, and pectate lyase, cellulase, protease, biosurfactant and blue pigment production. Mutants of four D. solani strains were constructed by inactivating the genes coding either for one of the main negative regulators of D. dadantii virulence (kdgR, pecS and pecT) or for the synthesis and perception of signaling molecules (expI and expR). Analysis of these mutants indicated that PecS, PecT and KdgR play a similar role in both species, repressing to different degrees the synthesis of virulence factors. The thermoregulator PecT seems to be a major regulator of D. solani virulence. This work also reveals the role of quorum sensing mediated by ExpI and ExpR in D. solani virulence on potato.

  12. Integrative genomic analysis identifies isoleucine and CodY as regulators of Listeria monocytogenes virulence.

    Directory of Open Access Journals (Sweden)

    Lior Lobel

    2012-09-01

    Full Text Available Intracellular bacterial pathogens are metabolically adapted to grow within mammalian cells. While these adaptations are fundamental to the ability to cause disease, we know little about the relationship between the pathogen's metabolism and virulence. Here we used an integrative Metabolic Analysis Tool that combines transcriptome data with genome-scale metabolic models to define the metabolic requirements of Listeria monocytogenes during infection. Twelve metabolic pathways were identified as differentially active during L. monocytogenes growth in macrophage cells. Intracellular replication requires de novo synthesis of histidine, arginine, purine, and branch chain amino acids (BCAAs, as well as catabolism of L-rhamnose and glycerol. The importance of each metabolic pathway during infection was confirmed by generation of gene knockout mutants in the respective pathways. Next, we investigated the association of these metabolic requirements in the regulation of L. monocytogenes virulence. Here we show that limiting BCAA concentrations, primarily isoleucine, results in robust induction of the master virulence activator gene, prfA, and the PrfA-regulated genes. This response was specific and required the nutrient responsive regulator CodY, which is known to bind isoleucine. Further analysis demonstrated that CodY is involved in prfA regulation, playing a role in prfA activation under limiting conditions of BCAAs. This study evidences an additional regulatory mechanism underlying L. monocytogenes virulence, placing CodY at the crossroads of metabolism and virulence.

  13. A novel anti-virulence gene revealed by proteomic analysis in Shigella flexneri 2a

    Directory of Open Access Journals (Sweden)

    Ying Tianyi

    2010-06-01

    Full Text Available Abstract Background Shigella flexneri is a gram-negative, facultative pathogen that causes the majority of communicable bacterial dysenteries in developing countries. The virulence factors of S. flexneri have been shown to be produced at 37 degrees C but not at 30 degrees C. To discover potential, novel virulence-related proteins of S. flexneri, we performed differential in-gel electrophoresis (DIGE analysis to measure changes in the expression profile that are induced by a temperature increase. Results The ArgT protein was dramatically down-regulated at 37 degrees C. In contrast, the ArgT from the non-pathogenic E. coli did not show this differential expression as in S. flexneri, which suggested that argT might be a potential anti-virulence gene. Competitive invasion assays in HeLa cells and in BALB/c mice with argT mutants were performed, and the results indicated that the over-expression of ArgTY225D would attenuate the virulence of S. flexneri. A comparative proteomic analysis was subsequently performed to investigate the effects of ArgT in S. flexneri at the molecular level. We show that HtrA is differentially expressed among different derivative strains. Conclusion Gene argT is a novel anti-virulence gene that may interfere with the virulence of S. flexneri via the transport of specific amino acids or by affecting the expression of the virulence factor, HtrA.

  14. Two-component regulators involved in the global control of virulence in Erwinia carotovora subsp. carotovora.

    Science.gov (United States)

    Eriksson, A R; Andersson, R A; Pirhonen, M; Palva, E T

    1998-08-01

    Production of extracellular, plant cell wall degrading enzymes, the main virulence determinants of the plant pathogen Erwinia carotovora subsp. carotovora, is coordinately controlled by a complex regulatory network. Insertion mutants in the exp (extracellular enzyme production) loci exhibit pleiotropic defects in virulence and the growth-phase-dependent transcriptional activation of genes encoding extracellular enzymes. Two new exp mutations, designated expA and expS, were characterized. Introduction of the corresponding wild-type alleles to the mutants complemented both the lack of virulence and the impaired production of plant cell wall degrading enzymes. The expA gene was shown to encode a 24-kDa polypeptide that is structurally and functionally related to the uvrY gene product of Escherichia coli and the GacA response regulator of Pseudomonas fluorescens. Functional similarity of expA and uvrY was demonstrated by genetic complementation. The expA gene is organized in an operon together with a uvrC-like gene, identical to the organization of uvrY and uvrC in E. coli. The unlinked expS gene encodes a putative sensor kinase that shows 92% identity to the recently described rpfA gene product from another E. carotovora subsp. carotovora strain. Our data suggest that ExpS and ExpA are members of two-component sensor kinase and response regulator families, respectively. These two proteins might interact in controlling virulence gene expression in E. carotovora subsp. carotovora.

  15. The LuxS/AI-2 Quorum-Sensing System of Streptococcus pneumoniae Is Required to Cause Disease, and to Regulate Virulence- and Metabolism-Related Genes in a Rat Model of Middle Ear Infection

    Directory of Open Access Journals (Sweden)

    Mukesh K. Yadav

    2018-05-01

    DNA replication and repair, ATP synthesis, capsule biosynthesis, cell division, the cell cycle, signal transduction, transcription regulation, competence, virulence, and carbohydrate metabolism were downregulated in the absence of LuxS/AI-2.Conclusion: The S. pneumoniae LuxS/AI-2 quorum-sensing system is necessary for biofilm formation and the colonization of the ear epithelium, and caused middle ear infection in the rat model. LuxS/AI-2 regulates the expression of the genes involved in virulence and bacterial fitness during pneumococcal biofilm formation.

  16. Influence of sublethal concentrations of common disinfectants on expression of virulence genes in Listeria monocytogenes

    DEFF Research Database (Denmark)

    Kastbjerg, Vicky Gaedt; Larsen, M. H.; Gram, Lone

    2010-01-01

    Listeria monocytogenes is a food-borne human pathogen that causes listeriosis, a relatively rare infection with a high fatality rate. The regulation of virulence gene expression is influenced by several environmental factors, and the aim of the present study was to determine how disinfectants use......, such as antibiotic resistance....... by Northern blot analysis. Eleven disinfectants representing four different groups of active components were evaluated in this study. Disinfectants with the same active ingredients had a similar effect on gene expression. Peroxy and chlorine compounds reduced the expression of the virulence genes...

  17. Natural Selection in Virulence Genes of Francisella tularensis.

    Science.gov (United States)

    Gunnell, Mark K; Robison, Richard A; Adams, Byron J

    2016-06-01

    A fundamental tenet of evolution is that alleles that are under negative selection are often deleterious and confer no evolutionary advantage. Negatively selected alleles are removed from the gene pool and are eventually extinguished from the population. Conversely, alleles under positive selection do confer an evolutionary advantage and lead to an increase in the overall fitness of the organism. These alleles increase in frequency until they eventually become fixed in the population. Francisella tularensis is a zoonotic pathogen and a potential biothreat agent. The most virulent type of F. tularensis, Type A, is distributed across North America with Type A.I occurring mainly in the east and Type A.II appearing mainly in the west. F. tularensis is thought to be a genome in decay (losing genes) because of the relatively large number of pseudogenes present in its genome. We hypothesized that the observed frequency of gene loss/pseudogenes may be an artifact of evolution in response to a changing environment, and that genes involved in virulence should be under strong positive selection. To test this hypothesis, we sequenced and compared whole genomes of Type A.I and A.II isolates. We analyzed a subset of virulence and housekeeping genes from several F. tularensis subspecies genomes to ascertain the presence and extent of positive selection. Eleven previously identified virulence genes were screened for positive selection along with 10 housekeeping genes. Analyses of selection yielded one housekeeping gene and 7 virulence genes which showed significant evidence of positive selection at loci implicated in cell surface structures and membrane proteins, metabolism and biosynthesis, transcription, translation and cell separation, and substrate binding and transport. Our results suggest that while the loss of functional genes through disuse could be accelerated by negative selection, the genome decay in Francisella could also be the byproduct of adaptive evolution

  18. Distribution of the type III DNA methyltransferases modA, modB and modD among Neisseria meningitidis genotypes: implications for gene regulation and virulence.

    Science.gov (United States)

    Tan, Aimee; Hill, Dorothea M C; Harrison, Odile B; Srikhanta, Yogitha N; Jennings, Michael P; Maiden, Martin C J; Seib, Kate L

    2016-02-12

    Neisseria meningitidis is a human-specific bacterium that varies in invasive potential. All meningococci are carried in the nasopharynx, and most genotypes are very infrequently associated with invasive meningococcal disease; however, those belonging to the 'hyperinvasive lineages' are more frequently associated with sepsis or meningitis. Genome content is highly conserved between carriage and disease isolates, and differential gene expression has been proposed as a major determinant of the hyperinvasive phenotype. Three phase variable DNA methyltransferases (ModA, ModB and ModD), which mediate epigenetic regulation of distinct phase variable regulons (phasevarions), have been identified in N. meningitidis. Each mod gene has distinct alleles, defined by their Mod DNA recognition domain, and these target and methylate different DNA sequences, thereby regulating distinct gene sets. Here 211 meningococcal carriage and >1,400 disease isolates were surveyed for the distribution of meningococcal mod alleles. While modA11-12 and modB1-2 were found in most isolates, rarer alleles (e.g., modA15, modB4, modD1-6) were specific to particular genotypes as defined by clonal complex. This suggests that phase variable Mod proteins may be associated with distinct phenotypes and hence invasive potential of N. meningitidis strains.

  19. SlyA regulates motA and motB, virulence and stress-related genes under conditions induced by the PhoP-PhoQ system in Dickeya dadantii 3937.

    Science.gov (United States)

    Haque, Md Manjurul; Hirata, Hisae; Tsuyumu, Shinji

    2015-01-01

    We previously showed that SlyA of Dickeya dadantii 3937 plays an important role in virulence toward plants, and that the ΔslyA mutant is hypermotile, whereas flagellum synthesis and flagellin production are indistinguishable from the wild type. Here we show that motility factors, including the distance of continuous directed movement, time for that movement and speed, were significantly higher in the ΔslyA mutant than in the wild type. Remarkably, transcription levels of motA and motB, that are involved in flagellar rotation, were elevated in the ΔslyA mutant, suggesting that the mutant's hypermotility was due to an increase in flagellar rotation. In low (10 μM) magnesium medium that activates the PhoP-PhoQ system, growth and virulence of the ΔslyA mutant were much lower than for the wild type; expression of motA, motB, mgtA, pelA, pelB, pelC, pelD, pelE, pelI, indA, tolC, sodC, acsA and hrpN were also reduced in the mutant. Interestingly, motA, motB, pelD, pelE, pelI, sodC and indA were also reduced in phoP and phoQ mutants. Because the SlyA protein directly binds to the promoter region of PhoP, SlyA regulates virulence by controlling multiple pathogenicity-related genes directly and/or at least by controlling PhoP in D. dadantii 3937 when magnesium is low. Copyright © 2015 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  20. Prevalence of clonal complexes and virulence genes among commensal and invasive Staphylococcus aureus isolates in Sweden.

    Directory of Open Access Journals (Sweden)

    Gunlög Rasmussen

    Full Text Available Staphylococcus aureus encodes a remarkable number of virulence factors which may contribute to its pathogenicity and ability to cause invasive disease. The main objective of this study was to evaluate the association between S. aureus invasiveness and bacterial genotype, in terms of the presence of virulence genes and affiliation to clonal complexes. Also, the significance of different virulence genes, mainly adhesins, for the development of infective endocarditis was investigated. DNA microarray technology was used to analyze 134 S. aureus isolates, all methicillin-susceptible, derived from three groups of clinically well-characterized patients: nasal carriers (n=46, bacteremia (n=55, and bacteremia with infective endocarditis (n=33. Invasive isolates were dominant in four of the major clonal complexes: 5, 8, 15, and 25. Of the 170 virulence genes examined, those encoding accessory gene regulator group II (agr II, capsule polysaccharide serotype 5 (cap5, and adhesins such as S. aureus surface protein G (sasG and fibronectin-binding protein B (fnbB were found to be associated with invasive disease. The same was shown for the leukocidin genes lukD/lukE, as well as the genes encoding serine protease A and B (splA/splB, staphylococcal complement inhibitor (scn and the staphylococcal exotoxin-like protein (setC or selX. In addition, there was a trend of higher prevalence of certain genes or gene clusters (sasG, agr II, cap5 among isolates causing infective endocarditis compared to other invasive isolates. In most cases, the presence of virulence genes was linked to clonal complex affiliation. In conclusion, certain S. aureus clonal lineages harboring specific sets of virulence genes seem to be more successful in causing invasive disease.

  1. Coordinated Regulation of Virulence during Systemic Infection of Salmonella enterica serovar Typhimurium

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Hyunjin; McDermott, Jason E.; Porwollik, Steffen; Mcclelland, Michael; Heffron, Fred

    2009-02-20

    Salmonella must respond to a myriad of environmental cues during infection of a mouse and express specific subsets of genes in a temporal and spatial manner to subvert the host defense mechanisms but these regulatory pathways are poorly established. To unravel how micro-environmental signals are processed and integrated into coordinated action, we constructed in-frame non-polar deletions of 84 regulators inferred to play a role in Salmonella typhimurium virulence and tested them in three virulence assays (intraperitoneal (i.p.), and intragastric (i.g.) infection in BALB/c mice, and persistence in SvJ129 mice). Overall 36 regulators were identified that were less virulent in at least one assay, and of those, 15 regulators were required for systemic mouse infection in an acute infection model. As a first step towards understanding the interplay between a pathogen and its host from a systems biology standpoint we focused on these 15 genes. Transcriptional profiles were obtained for each of these 15 regulators from strains grown under four different environmental conditions. These results as well as publicly available transcriptional profiles were analyzed using both network inference and cluster analysis algorithms. The analysis predicts a regulatory network in which all 15 regulators control a specific set of genes necessary for Salmonella to cause systemic infection. We tested the regulatory model by expressing a subset of the regulators in trans and monitoring transcription of 7 known virulence factors located within Salmonella pathogenicity island 2 (SPI-2). These experiments validated the regulatory model and showed that, for these 7 genes, the response regulator SsrB and the marR type regulator SlyA co-regulate in a regulatory cascade by integrating multiple signals.

  2. Detection of virulence-associated genes in Brucella melitensis ...

    African Journals Online (AJOL)

    The current study involved detection of three virulence genes (bvfA, virB, ure) by PCR in 52 isolates of Brucella melitensis biovar 3, recovered from different animal species (28 sheep, 10 goats, 9 cattle and 5 buffaloes). Of the 52 B. melitensis strains; 48 (92.3%) isolates carried bvfA genes, 51 (98.1%) isolates had virB genes ...

  3. Regulation of Yersina pestis Virulence by AI-2 Mediated Quorum Sensing

    Energy Technology Data Exchange (ETDEWEB)

    Segelke, B; Hok, S; Lao, V; Corzett, M; Garcia, E

    2010-03-29

    The proposed research was motivated by an interest in understanding Y. pestis virulence mechanisms and bacteria cell-cell communication. It is expected that a greater understanding of virulence mechanisms will ultimately lead to biothreat countermeasures and novel therapeutics. Y. pestis is the etiological agent of plague, the most devastating disease in human history. Y. pestis infection has a high mortality rate and a short incubation before mortality. There is no widely available and effective vaccine for Y. pestis and multi-drug resistant strains are emerging. Y. pestis is a recognized biothreat agent based on the wide distribution of the bacteria in research laboratories around the world and on the knowledge that methods exist to produce and aerosolize large amounts of bacteria. We hypothesized that cell-cell communication via signaling molecules, or quorum sensing, by Y. pestis is important for the regulation of virulence factor gene expression during host invasion, though a causative link had never been established. Quorum sensing is a mode of intercellular communication which enables orchestration of gene expression for many bacteria as a function of population density and available evidence suggests there may be a link between quorum sensing and regulation of Y. pesits virulence. Several pathogenic bacteria have been shown to regulate expression of virulence factor genes, including genes encoding type III secretion, via quorum sensing. The Y. pestis genome encodes several cell-cell signaling pathways and the interaction of at least three of these are thought to be involved in one or more modes of host invasion. Furthermore, Y. pestis gene expression array studies carried out at LLNL have established a correlation between expression of known virulence factors and genes involved in processing of the AI-2 quorum sensing signal. This was a basic research project that was intended to provide new insights into bacterial intercellular communication and how it is

  4. Gene encoding virulence markers among Escherichia coli isolates ...

    African Journals Online (AJOL)

    River water sources and diarrhoeic stools of residents in the Venda Region, Limpopo Province of South Africa were analysed for the prevalence of Escherichia coli (E. coli) and the presence of virulence genes among the isolates. A control group of 100 nondiarrhoeic stool samples was included. Escherichia coli was ...

  5. Prevalence of Escherichia coli virulence genes in patients with ...

    African Journals Online (AJOL)

    In this study, we investigated the prevalence of the virulence genes specific for five major pathogroups of diarrheagenic Escherichia coli (DEC) in primary cultures from diarrhoeagenic patients in Burkina Faso. Methodology: From September 2016 to Mars 2017, a total of 211 faecal samples from diarrhoeagenic patients from ...

  6. Molecular Detection of Virulence Genes and Antibiotic Resistance ...

    African Journals Online (AJOL)

    Escherichia coli O157:H7 is an important food-borne pathogen that can cause diarrhea, haemorrhagic colitis and haemolytic uremic syndrome. This study was conducted to investigate the prevalence, virulence genes and antibiotic resistance patterns of E. coli O157:H7 in raw beef meat sold in Abeokuta, South west Nigeria ...

  7. The virulence regulator PrfA promotes biofilm formation by Listeria monocytogenes.

    Science.gov (United States)

    Lemon, Katherine P; Freitag, Nancy E; Kolter, Roberto

    2010-08-01

    Listeria monocytogenes is a food-borne facultative intracellular pathogen. It is widespread in the environment and has several distinct life-styles. The key transcriptional activator PrfA positively regulates L. monocytogenes virulence genes to mediate the transition from extracellular, flagellum-propelled cell to intracellular pathogen. Here we report the first evidence that PrfA also has a significant positive impact on extracellular biofilm formation. Mutants lacking prfA were defective in surface-adhered biofilm formation. The DeltaprfA mutant exhibited wild-type flagellar motility, and its biofilm defect occurred after initial surface adhesion. We also observed that mutations that led to the constitutive expression of PrfA-dependent virulence genes had a minimal impact on biofilm formation. Furthermore, biofilm development was enhanced in a mutant encoding a PrfA protein variant unable to fully transition from the extracellular form to the virulent, intracellular activity conformation. These results indicate that PrfA positively regulates biofilm formation and suggest that PrfA has a global role in modulating the life-style of L. monocytogenes. The requirement of PrfA for optimal biofilm formation may provide selective pressure to maintain this critical virulence regulator when L. monocytogenes is outside host cells in the environment.

  8. Virulence Genes and Antibiotic Susceptibilities of Uropathogenic E. coli Strains.

    Science.gov (United States)

    Uzun, Cengiz; Oncül, Oral; Gümüş, Defne; Alan, Servet; Dayioğlu, Nurten; Küçüker, Mine Anğ

    2015-01-01

    The aim of this study is to detect the presence of and possible relation between virulence genes and antibiotic resistance in E. coli strains isolated from patients with acute, uncomplicated urinary tract infections (UTI). 62 E. coli strains isolated from patients with acute, uncomplicated urinary tract infections (50 strains isolated from acute uncomplicated cystitis cases (AUC); 12 strains from acute uncomplicated pyelonephritis cases (AUP)) were screened for virulence genes [pap (pyelonephritis-associated pili), sfa/foc (S and F1C fimbriae), afa (afimbrial adhesins), hly (hemolysin), cnf1 (cytotoxic necrotizing factor), aer (aerobactin), PAI (pathogenicity island marker), iroN (catecholate siderophore receptor), ompT (outer membrane protein T), usp (uropathogenic specific protein)] by PCR and for antimicrobial resistance by disk diffusion method according to CLSI criteria. It was found that 56 strains (90.3%) carried at least one virulence gene. The most common virulence genes were ompT (79%), aer (51.6%), PAI (51.6%) and usp (56.5%). 60% of the strains were resistant to at least one antibiotic. The highest resistance rates were against ampicillin (79%) and co-trimoxazole (41.9%). Fifty percent of the E. coli strains (31 strains) were found to be multiple resistant. Eight (12.9%) out of 62 strains were found to be ESBL positive. Statistically significant relationships were found between the absence of usp and AMP - SXT resistance, iroN and OFX - CIP resistance, PAI and SXT resistance, cnf1 and AMP resistance, and a significant relationship was also found between the presence of the afa and OFX resistance. No difference between E. coli strains isolated from two different clinical presentations was found in terms of virulence genes and antibiotic susceptibility.

  9. Functional analysis of the conserved transcriptional regulator CfWor1 in Cladosporium fulvum reveals diverse roles in the virulence of plant pathogenic fungi

    NARCIS (Netherlands)

    Ökmen, B.; Collemare, J.; Griffiths, S.A.; Burgt, van der A.; Cox, R.; Wit, de P.J.G.M.

    2014-01-01

    Fungal Wor1-like proteins are conserved transcriptional regulators that are reported to regulate the virulence of several plant pathogenic fungi by affecting the expression of virulence genes. Here, we report the functional analysis of CfWor1, the homologue of Wor1 in Cladosporium fulvum. ¿cfwor1

  10. A functional gene array for detection of bacterial virulence elements

    Energy Technology Data Exchange (ETDEWEB)

    Jaing, C

    2007-11-01

    We report our development of the first of a series of microarrays designed to detect pathogens with known mechanisms of virulence and antibiotic resistance. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples. To validate our approach, we developed a first generation array targeting genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for microorganism detection and discrimination, measured the required target concentration, and assessed tolerance for mismatches between probe and target sequences. Mismatch tolerance is a priority for this application, due to DNA sequence variability among members of gene families. Arrays were created using the NimbleGen Maskless Array Synthesizer at Lawrence Livermore National Laboratory. Purified genomic DNA from combinations of one or more of the four target organisms, pure cultures of four related organisms, and environmental aerosol samples with spiked-in genomic DNA were hybridized to the arrays. Based on the success of this prototype, we plan to design further arrays in this series, with the goal of detecting all known virulence and antibiotic resistance gene families in a greatly expanded set of organisms.

  11. Genes involved in virulence of the entomopathogenic fungus Beauveria bassiana.

    Science.gov (United States)

    Valero-Jiménez, Claudio A; Wiegers, Harm; Zwaan, Bas J; Koenraadt, Constantianus J M; van Kan, Jan A L

    2016-01-01

    Pest insects cause severe damage to global crop production and pose a threat to human health by transmitting diseases. Traditionally, chemical pesticides (insecticides) have been used to control such pests and have proven to be effective only for a limited amount of time because of the rapid spread of genetic insecticide resistance. The basis of this resistance is mostly caused by (co)dominant mutations in single genes, which explains why insecticide use alone is an unsustainable solution. Therefore, robust solutions for insect pest control need to be sought in alternative methods such as biological control agents for which single-gene resistance is less likely to evolve. The entomopathogenic fungus Beauveria bassiana has shown potential as a biological control agent of insects, and insight into the mechanisms of virulence is essential to show the robustness of its use. With the recent availability of the whole genome sequence of B. bassiana, progress in understanding the genetics that constitute virulence toward insects can be made more quickly. In this review we divide the infection process into distinct steps and provide an overview of what is currently known about genes and mechanisms influencing virulence in B. bassiana. We also discuss the need for novel strategies and experimental methods to better understand the infection mechanisms deployed by entomopathogenic fungi. Such knowledge can help improve biocontrol agents, not only by selecting the most virulent genotypes, but also by selecting the genotypes that use combinations of virulence mechanisms for which resistance in the insect host is least likely to develop. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Definition of a consensus DNA-binding site for PecS, a global regulator of virulence gene expression in Erwinia chrysanthemi and identification of new members of the PecS regulon.

    Science.gov (United States)

    Rouanet, Carine; Reverchon, Sylvie; Rodionov, Dmitry A; Nasser, William

    2004-07-16

    In Erwinia chrysanthemi, production of pectic enzymes is modulated by a complex network involving several regulators. One of them, PecS, which belongs to the MarR family, also controls the synthesis of various other virulence factors, such as cellulases and indigoidine. Here, the PecS consensus-binding site is defined by combining a systematic evolution of ligands by an exponential enrichment approach and mutational analyses. The consensus consists of a 23-base pair palindromic-like sequence (C(-11)G(-10)A(-9)N(-8)W(-7)T(-6)C(-5)G(-4)T(-3)A(-2))T(-1)A(0)T(1)(T(2)A(3)C(4)G(5)A(6)N(7)N(8)N(9)C(10)G(11)). Mutational experiments revealed that (i) the palindromic organization is required for the binding of PecS, (ii) the very conserved part of the consensus (-6 to 6) allows for a specific interaction with PecS, but the presence of the relatively degenerated bases located apart significantly increases PecS affinity, (iii) the four bases G, A, T, and C are required for efficient binding of PecS, and (iv) the presence of several binding sites on the same promoter increases the affinity of PecS. This consensus is detected in the regions involved in PecS binding on the previously characterized target genes. This variable consensus is in agreement with the observation that the members of the MarR family are able to bind various DNA targets as dimers by means of a winged helix DNA-binding motif. Binding of PecS on a promoter region containing the defined consensus results in a repression of gene transcription in vitro. Preliminary scanning of the E. chrysanthemi genome sequence with the consensus revealed the presence of strong PecS-binding sites in the intergenic region between fliE and fliFGHIJKLMNOPQR which encode proteins involved in the biogenesis of flagellum. Accordingly, PecS directly represses fliE expression. Thus, PecS seems to control the synthesis of virulence factors required for the key steps of plant infection.

  13. Genome-wide identification of Pseudomonas aeruginosa virulence-related genes using a Caenorhabditis elegans infection model.

    Directory of Open Access Journals (Sweden)

    Rhonda L Feinbaum

    Full Text Available Pseudomonas aeruginosa strain PA14 is an opportunistic human pathogen capable of infecting a wide range of organisms including the nematode Caenorhabditis elegans. We used a non-redundant transposon mutant library consisting of 5,850 clones corresponding to 75% of the total and approximately 80% of the non-essential PA14 ORFs to carry out a genome-wide screen for attenuation of PA14 virulence in C. elegans. We defined a functionally diverse 180 mutant set (representing 170 unique genes necessary for normal levels of virulence that included both known and novel virulence factors. Seven previously uncharacterized virulence genes (ABC transporters PchH and PchI, aminopeptidase PepP, ATPase/molecular chaperone ClpA, cold shock domain protein PA0456, putative enoyl-CoA hydratase/isomerase PA0745, and putative transcriptional regulator PA14_27700 were characterized with respect to pigment production and motility and all but one of these mutants exhibited pleiotropic defects in addition to their avirulent phenotype. We examined the collection of genes required for normal levels of PA14 virulence with respect to occurrence in P. aeruginosa strain-specific genomic regions, location on putative and known genomic islands, and phylogenetic distribution across prokaryotes. Genes predominantly contributing to virulence in C. elegans showed neither a bias for strain-specific regions of the P. aeruginosa genome nor for putatively horizontally transferred genomic islands. Instead, within the collection of virulence-related PA14 genes, there was an overrepresentation of genes with a broad phylogenetic distribution that also occur with high frequency in many prokaryotic clades, suggesting that in aggregate the genes required for PA14 virulence in C. elegans are biased towards evolutionarily conserved genes.

  14. Genome-wide identification of Pseudomonas aeruginosa virulence-related genes using a Caenorhabditis elegans infection model.

    Science.gov (United States)

    Feinbaum, Rhonda L; Urbach, Jonathan M; Liberati, Nicole T; Djonovic, Slavica; Adonizio, Allison; Carvunis, Anne-Ruxandra; Ausubel, Frederick M

    2012-01-01

    Pseudomonas aeruginosa strain PA14 is an opportunistic human pathogen capable of infecting a wide range of organisms including the nematode Caenorhabditis elegans. We used a non-redundant transposon mutant library consisting of 5,850 clones corresponding to 75% of the total and approximately 80% of the non-essential PA14 ORFs to carry out a genome-wide screen for attenuation of PA14 virulence in C. elegans. We defined a functionally diverse 180 mutant set (representing 170 unique genes) necessary for normal levels of virulence that included both known and novel virulence factors. Seven previously uncharacterized virulence genes (ABC transporters PchH and PchI, aminopeptidase PepP, ATPase/molecular chaperone ClpA, cold shock domain protein PA0456, putative enoyl-CoA hydratase/isomerase PA0745, and putative transcriptional regulator PA14_27700) were characterized with respect to pigment production and motility and all but one of these mutants exhibited pleiotropic defects in addition to their avirulent phenotype. We examined the collection of genes required for normal levels of PA14 virulence with respect to occurrence in P. aeruginosa strain-specific genomic regions, location on putative and known genomic islands, and phylogenetic distribution across prokaryotes. Genes predominantly contributing to virulence in C. elegans showed neither a bias for strain-specific regions of the P. aeruginosa genome nor for putatively horizontally transferred genomic islands. Instead, within the collection of virulence-related PA14 genes, there was an overrepresentation of genes with a broad phylogenetic distribution that also occur with high frequency in many prokaryotic clades, suggesting that in aggregate the genes required for PA14 virulence in C. elegans are biased towards evolutionarily conserved genes.

  15. Down-regulation of MHC class I by the Marek's disease virus (MDV) UL49.5 gene product mildly affects virulence in a haplotype-specific fashion.

    Science.gov (United States)

    Jarosinski, Keith W; Hunt, Henry D; Osterrieder, Nikolaus

    2010-09-30

    Marek's disease is a devastating neoplastic disease of chickens caused by Marek's disease virus (MDV). MDV down-regulates surface expression of MHC class I molecules, although the mechanism has remained elusive. MDV harbors a UL49.5 homolog that has been shown to down-regulate MHC class I expression in other Varicelloviruses. Using in vitro assays, we showed that MDV pUL49.5 down-regulates MHC class I directly and identified its cytoplasmic tail as essential for this function. In vivo, viruses lacking the cytoplasmic tail of pUL49.5 showed no differences in MD pathogenesis compared to revertant viruses in highly susceptible chickens of the B(19)B(19) MHC class I haplotype, while there was a mild reduction in pathogenic potential of the deletion viruses in chickens more resistant to MD pathogenesis (MHC:B(21)B(21)). We concluded that the pathogenic effect of MHC class I down-regulation mediated by pUL49.5 is small because virus immune evasion possibly requires more than one viral protein. Copyright 2010 Elsevier Inc. All rights reserved.

  16. Down-regulation of MHC Class I by the Marek's Disease Virus (MDV) UL49.5 Gene Product Mildly Affects Virulence in a Haplotype-specific Fashion

    Science.gov (United States)

    Marek’s disease is a devastating neoplastic disease of chickens caused by gallid herpesvirus 2 or Marek’s disease virus (MDV), which is characterized by massive visceral tumors, immune suppression, neurologic syndromes, and peracute deaths. It has been reported that MDV down-regulates surface expre...

  17. Responsiveness to acidity via metal ion regulators mediates virulence in the gastric pathogen Helicobacter pylori.

    Science.gov (United States)

    Bury-Moné, Stéphanie; Thiberge, Jean-Michel; Contreras, Monica; Maitournam, Aboubakar; Labigne, Agnès; De Reuse, Hilde

    2004-07-01

    The virulence of pathogenic bacteria is dependent on their adaptation to and survival in the stressful conditions encountered in their hosts. Helicobacter pylori exclusively colonizes the acid stomach of primates, making it an ideal study model. Little is known about how H. pylori responds to the moderately acidic conditions encountered at its colonization site, the gastric mucus layer. Thus, we compared gene expression profiles of H. pylori 26695 grown at neutral and acidic pH, and validated the data for a selection of genes by real-time polymerase chain reaction, dot-blots or enzymatic assays. During growth in acidic conditions, 56 genes were upregulated and 45 genes downregulated. We found that acidity is a signal modulating the expression of several virulence factors. Regulation of genes related to metal ion homeostasis suggests protective mechanisms involving diminished transport and enhanced storage. Genes encoding subunits of the F0F1 ATPase and of a newly identified Na+/H+ antiporter (NhaC-HP0946) were downregulated, revealing that this bacterium uses original mechanisms to control proton entry. Five of the upregulated genes encoded proteins controlling intracellular ammonia synthesis, including urease, amidase and formamidase, underlining the major role of this buffering compound in the protection against acidity in H. pylori. Regulatory networks and transcriptome analysis as well as enzymatic assays implicated two metal-responsive transcriptional regulators (NikR and Fur) and an essential two-component response regulator (HP0166, OmpR-like) as effectors of the H. pylori acid response. Finally, a nikR-fur mutant is attenuated in the mouse model, emphasizing the link between response to acidity, metal metabolism and virulence in this gastric pathogen.

  18. Antibiotic resistance and virulence genes in coliform water isolates.

    Science.gov (United States)

    Stange, C; Sidhu, J P S; Tiehm, A; Toze, S

    2016-11-01

    Widespread fecal pollution of surface water may present a major health risk and a significant pathway for dissemination of antibiotic resistance bacteria. The River Rhine is one of the longest and most important rivers in Europe and an important raw water source for drinking water production. A total of 100 coliform isolates obtained from River Rhine (Germany) were examined for their susceptibility to seven antimicrobial agents. Resistances against amoxicillin, trimethoprim/sulfamethoxazole and tetracycline were detected in 48%, 11% and 9% of isolates respectively. The antibiotic resistance could be traced back to the resistance genes bla TEM , bla SHV , ampC, sul1, sul2, dfrA1, tet(A) and tet(B). Whereby, the ampC gene represents a special case, because its presence is not inevitably linked to a phenotypic antibiotic resistance. Multiple antibiotics resistance was often accompanied by the occurrence of class 1 or 2 integrons. E. coli isolates belonging to phylogenetic groups A and B1 (commensal) were more predominant (57%) compared to B2 and D groups (43%) which are known to carry virulent genes. Additionally, six E. coli virulence genes were also detected. However, the prevalence of virulence genes in the E. coli isolates was low (not exceeding 4.3% per gene) and no diarrheagenic E. coli pathotypes were detected. This study demonstrates that surface water is an important reservoir of ARGs for a number of antibiotic classes such as sulfonamide, trimethoprim, beta-lactam-antibiotics and tetracycline. The occurrence of antibiotic resistance in coliform bacteria isolated from River Rhine provides evidence for the need to develop management strategies to limit the spread of antibiotic resistant bacteria in aquatic environment. Copyright © 2016 Elsevier GmbH. All rights reserved.

  19. The Central Metabolism Regulator EIIAGlc Switches Salmonella from Growth Arrest to Acute Virulence through Activation of Virulence Factor Secretion

    Directory of Open Access Journals (Sweden)

    Alain Mazé

    2014-06-01

    Full Text Available The ability of Salmonella to cause disease depends on metabolic activities and virulence factors. Here, we show that a key metabolic protein, EIIAGlc, is absolutely essential for acute infection, but not for Salmonella survival, in a mouse typhoid fever model. Surprisingly, phosphorylation-dependent EIIAGlc functions, including carbohydrate transport and activation of adenylate cyclase for global regulation, do not explain this virulence phenotype. Instead, biochemical studies, in vitro secretion and translocation assays, and in vivo genetic epistasis experiments suggest that EIIAGlc binds to the type three secretion system 2 (TTSS-2 involved in systemic virulence, stabilizes its cytoplasmic part including the crucial TTSS-2 ATPase, and activates virulence factor secretion. This unexpected role of EIIAGlc reveals a striking direct link between central Salmonella metabolism and a crucial virulence mechanism.

  20. Allele-dependent differences in quorum-sensing dynamics result in variant expression of virulence genes in Staphylococcus aureus.

    Science.gov (United States)

    Geisinger, Edward; Chen, John; Novick, Richard P

    2012-06-01

    Agr is an autoinducing, quorum-sensing system that functions in many Gram-positive species and is best characterized in the pathogen Staphylococcus aureus, in which it is a global regulator of virulence gene expression. Allelic variations in the agr genes have resulted in the emergence of four quorum-sensing specificity groups in S. aureus, which correlate with different strain pathotypes. The basis for these predilections is unclear but is hypothesized to involve the phenomenon of quorum-sensing interference between strains of different agr groups, which may drive S. aureus strain isolation and divergence. Whether properties intrinsic to each agr allele directly influence virulence phenotypes within S. aureus is unknown. In this study, we examined group-specific differences in agr autoinduction and virulence gene regulation by utilizing congenic strains, each harboring a unique S. aureus agr allele, enabling a dissection of agr locus-dependent versus genotype-dependent effects on quorum-sensing dynamics and virulence factor production. Employing a reporter fusion to the principal agr promoter, P3, we observed allele-dependent differences in the timing and magnitude of agr activation. These differences were mediated by polymorphisms within the agrBDCA genes and translated to significant variations in the expression of a key transcriptional regulator, Rot, and of several important exoproteins and surface factors involved in pathogenesis. This work uncovers the contribution of divergent quorum-sensing alleles to variant expression of virulence determinants within a bacterial species.

  1. Sigma E regulators control hemolytic activity and virulence in a shrimp pathogenic Vibrio harveyi.

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    Pimonsri Rattanama

    Full Text Available Members of the genus Vibrio are important marine and aquaculture pathogens. Hemolytic activity has been identified as a virulence factor in many pathogenic vibrios including V. cholerae, V. parahaemolyticus, V. alginolyticus, V. harveyi and V. vulnificus. We have used transposon mutagenesis to identify genes involved in the hemolytic activity of shrimp-pathogenic V. harveyi strain PSU3316. Out of 1,764 mutants screened, five mutants showed reduced hemolytic activity on sheep blood agar and exhibited virulence attenuation in shrimp (Litopenaeus vannamei. Mutants were identified by comparing transposon junction sequences to a draft of assembly of the PSU3316 genome. Surprisingly none of the disrupted open reading frames or gene neighborhoods contained genes annotated as hemolysins. The gene encoding RseB, a negative regulator of the sigma factor (σ(E, was interrupted in 2 out of 5 transposon mutants, in addition, the transcription factor CytR, a threonine synthetase, and an efflux-associated cytoplasmic protein were also identified. Knockout mutations introduced into the rpoE operon at the rseB gene exhibited low hemolytic activity in sheep blood agar, and were 3-to 7-fold attenuated for colonization in shrimp. Comparison of whole cell extracted proteins in the rseB mutant (PSU4030 to the wild-type by 2-D gel electrophoresis revealed 6 differentially expressed proteins, including two down-regulated porins (OmpC-like and OmpN and an upregulated protease (DegQ which have been associated with σ(E in other organisms. Our study is the first report linking hemolytic activity to the σ(E regulators in pathogenic Vibrio species and suggests expression of this virulence-linked phenotype is governed by multiple regulatory pathways within the V. harveyi.

  2. A bistable switch and anatomical site control Vibrio cholerae virulence gene expression in the intestine.

    Directory of Open Access Journals (Sweden)

    Alex T Nielsen

    2010-09-01

    Full Text Available A fundamental, but unanswered question in host-pathogen interactions is the timing, localization and population distribution of virulence gene expression during infection. Here, microarray and in situ single cell expression methods were used to study Vibrio cholerae growth and virulence gene expression during infection of the rabbit ligated ileal loop model of cholera. Genes encoding the toxin-coregulated pilus (TCP and cholera toxin (CT were powerfully expressed early in the infectious process in bacteria adjacent to epithelial surfaces. Increased growth was found to co-localize with virulence gene expression. Significant heterogeneity in the expression of tcpA, the repeating subunit of TCP, was observed late in the infectious process. The expression of tcpA, studied in single cells in a homogeneous medium, demonstrated unimodal induction of tcpA after addition of bicarbonate, a chemical inducer of virulence gene expression. Striking bifurcation of the population occurred during entry into stationary phase: one subpopulation continued to express tcpA, whereas the expression declined in the other subpopulation. ctxA, encoding the A subunit of CT, and toxT, encoding the proximal master regulator of virulence gene expression also exhibited the bifurcation phenotype. The bifurcation phenotype was found to be reversible, epigenetic and to persist after removal of bicarbonate, features consistent with bistable switches. The bistable switch requires the positive-feedback circuit controlling ToxT expression and formation of the CRP-cAMP complex during entry into stationary phase. Key features of this bistable switch also were demonstrated in vivo, where striking heterogeneity in tcpA expression was observed in luminal fluid in later stages of the infection. When this fluid was diluted into artificial seawater, bacterial aggregates continued to express tcpA for prolonged periods of time. The bistable control of virulence gene expression points to a

  3. The Mechanisms of Virulence Regulation by Small Noncoding RNAs in Low GC Gram-Positive Pathogens

    Directory of Open Access Journals (Sweden)

    Stephanie Pitman

    2015-12-01

    Full Text Available The discovery of small noncoding regulatory RNAs (sRNAs in bacteria has grown tremendously recently, giving new insights into gene regulation. The implementation of computational analysis and RNA sequencing has provided new tools to discover and analyze potential sRNAs. Small regulatory RNAs that act by base-pairing to target mRNAs have been found to be ubiquitous and are the most abundant class of post-transcriptional regulators in bacteria. The majority of sRNA studies has been limited to E. coli and other gram-negative bacteria. However, examples of sRNAs in gram-positive bacteria are still plentiful although the detailed gene regulation mechanisms behind them are not as well understood. Strict virulence control is critical for a pathogen’s survival and many sRNAs have been found to be involved in that process. This review outlines the targets and currently known mechanisms of trans-acting sRNAs involved in virulence regulation in various gram-positive pathogens. In addition, their shared characteristics such as CU interaction motifs, the role of Hfq, and involvement in two-component regulators, riboswitches, quorum sensing, or toxin/antitoxin systems are described.

  4. Method for Screening Compounds That Influence Virulence Gene Expression in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Nielsen, A.; Nielsen, Kristian Fog; Frees, D.

    2010-01-01

    We present a simple assay to examine effects of compounds on virulence gene expression in the human pathogen Staphylococcus aureus. The assay employs transcriptional reporter strains carrying lacZ fused to central virulence genes. Compounds affecting virulence gene expression and activity...... of the agr locus are scored based on color change in the presence of a chromogenic beta-galactosidase substrate. The assay can be used to screen for novel antivirulence compounds from many different sources, such as fungi, as demonstrated here....

  5. [Virulent gene prevalence of foodborne Listeria monocytogenes in China in 2005].

    Science.gov (United States)

    Yang, Yang; Fu, Ping; Guo, Yun-Chang; Pei, Xiao-Yan; Liu, Xiu-Mei

    2010-12-01

    To study the virulent gene prevalence of foodborne Listeria monocytogenes (LM) isolated from China. 78 LM isolates derived from raw meat, cooked food, aquatic products and vegetables of 13 provinces and cities.LM isolates were investigated for prevalence of virulence genes (LIPI-1 (prfA, plcA, hly, mpl, actA, plcB); LIPI-2 (inlA, inlB), and iap) by PCR method. 87.2% (68/78) of the isolates were prfA positive, 98.7% (77/78) of the isolates were plcA, actA and plcB positive, 97.4% (76/78) of the isolates were hly positive, 87.2% (68/78) of the isolates were mpl positive, 92.3% (72/78) of the isolates were inlA positive, 100% (78/78) of the isolates were inlB positive, 98.7% (77/78) of the isolates were iap positive. Among 21 virulent gene negative isolates, there was 7 isolates lack of two or more virulence genes. The rate of virulence genes deletion isolates from cooked meat was 31.3% (10/32), the rate of virulence genes deletion isolates from raw meat was 16.1% (5/31), the rate of virulence genes deletion isolates from vegetables was 36.4% (4/11) and rate of virulence genes deletion isolates from seafood was 50% (2/4). No significant difference was found (χ(2) = 3.721, P > 0.05). The virulence gene array-1 strains were dominant among these isolates. Among 78 LM isolates, prevalent of virulent genes were different except inlB, virulence genes of LIP-1 were deleted prevalently among isolates, virulence gene deletion patterns were diverse.

  6. Mechanosensing regulates virulence in Escherichia coli O157:H7.

    Science.gov (United States)

    Islam, Md Shahidul; Krachler, Anne Marie

    2016-01-01

    Enterohemorrhagic Escherichia coli O157:H7 is a food-borne pathogen transmitted via the fecal-oral route, and can cause bloody diarrhea and hemolytic uremic syndrome (HUS) in the human host. Although a range of colonization factors, Shiga toxins and a type III secretion system (T3SS) all contribute to disease development, the locus of enterocyte effacement (LEE) encoded T3SS is responsible for the formation of lesions in the intestinal tract. While a variety of chemical cues in the host environment are known to up-regulate LEE expression, we recently demonstrated that changes in physical forces at the site of attachment are required for localized, full induction of the system and thus spatial regulation of virulence in the intestinal tract. Here, we discuss our findings in the light of other recent studies describing mechanosensing of the host and force-dependent induction of virulence mechanisms. We discuss potential mechanisms of mechanosensing and mechanotransduction, and the level of conservation across bacterial species.

  7. Evaluating virulence of waterborne and clinical Aeromonas isolates using gene expression and mortality in neonatal mice followed by assessing cell culture’s ability to predict virulence based on transcriptional response

    Energy Technology Data Exchange (ETDEWEB)

    Hayes, S L; Rodgers, M R; Lye, D J; Stelma, G N; McKinstry, Craig A.; Malard, Joel M.; Vesper, Sephen J.

    2007-10-01

    Aims: To assess the virulence of Aeromonas spp. using two models, a neonatal mouse assay and a mouse intestinal cell culture. Methods and Results: After artificial infection with a variety of Aeromonas spp., mRNA extracts from the two models were processed and hydridized to murine microarrays to determine host gene response. Definition of virulence was determined based on host mRNA production in murine neonatal intestinal tissue and mortality of infected animals. Infections of mouse intestinal cell cultures were then performed to determine whether this simpler model system’s mRNA responses correlated to neonatal results and therefore be predictive of virulence of Aeromonas spp. Virulent aeromonads up-regulated transcripts in both models including multiple host defense gene products (chemokines, regulation of transcription and apoptosis and cell signalling). Avirulent species exhibited little or no host response in neonates. Mortality results correlated well with both bacterial dose and average fold change of up-regulated transcripts in the neonatal mice. Conclusions: Cell culture results were less discriminating but showed promise as potentially being able to be predictive of virulence. Jun oncogene up-regulation in murine cell culture is potentially predictive of Aeromonas virulence. Significance and Impact of the Study: Having the ability to determine virulence of waterborne pathogens quickly would potentially assist public health officials to rapidly assess exposure risks.

  8. Antimicrobial medium- and long-chain free fatty acids prevent PrfA-dependent activation of virulence genes in Listeria monocytogenes

    DEFF Research Database (Denmark)

    Sternkopf Lillebæk, Eva Maria; Lambert Nielsen, Stine; Scheel Thomasen, Rikke

    2017-01-01

    of virulence factors required for bacterial entry, intracellular replication and cell-to-cell spread. PrfA-dependent activation of virulence genes occurs primarily in the blood and during intracellular infection. In contrast, PrfA does not play a significant role in regulation of virulence gene expression...... antimicrobial free fatty acids act to downregulate transcription of PrfA-activated virulence genes. Interestingly, the inhibitory effect is also evident in cells encoding a constitutively active variant of PrfA. Collectively, our data suggest that antimicrobial medium- and long-chain free fatty acids may act...... as signals to prevent PrfA-mediated activation of virulence genes in environments where PrfA activation is not required, such as in food and the gastrointestinal tract....

  9. Disruption of tetR type regulator adeN by mobile genetic element confers elevated virulence in Acinetobacter baumannii.

    Science.gov (United States)

    Saranathan, Rajagopalan; Pagal, Sudhakar; Sawant, Ajit R; Tomar, Archana; Madhangi, M; Sah, Suresh; Satti, Annapurna; Arunkumar, K P; Prashanth, K

    2017-10-03

    Acinetobacter baumannii is an important human pathogen and considered as a major threat due to its extreme drug resistance. In this study, the genome of a hyper-virulent MDR strain PKAB07 of A. baumannii isolated from an Indian patient was sequenced and analyzed to understand its mechanisms of virulence, resistance and evolution. Comparative genome analysis of PKAB07 revealed virulence and resistance related genes scattered throughout the genome, instead of being organized as an island, indicating the highly mosaic nature of the genome. Many intermittent horizontal gene transfer events, insertion sequence (IS) element insertions identified were augmenting resistance machinery and elevating the SNP densities in A. baumannii eventually aiding in their swift evolution. ISAba1, the most widely distributed insertion sequence in A. baumannii was found in multiple sites in PKAB07. Out of many ISAba1 insertions, we identified novel insertions in 9 different genes wherein insertional inactivation of adeN (tetR type regulator) was significant. To assess the significance of this disruption in A. baumannii, adeN mutant and complement strains were constructed in A. baumannii ATCC 17978 strain and studied. Biofilm levels were abrogated in the adeN knockout when compared with the wild type and complemented strain of adeN knockout. Virulence of the adeN knockout mutant strain was observed to be high, which was validated by in vitro experiments and Galleria mellonella infection model. The overexpression of adeJ, a major component of AdeIJK efflux pump observed in adeN knockout strain could be the possible reason for the elevated virulence in adeN mutant and PKB07 strain. Knocking out of adeN in ATCC strain led to increased resistance and virulence at par with the PKAB07. Disruption of tetR type regulator adeN by ISAba1 consequently has led to elevated virulence in this pathogen.

  10. Identification of an essential virulence gene of cyprinid herpesvirus 3.

    Science.gov (United States)

    Boutier, Maxime; Gao, Yuan; Vancsok, Catherine; Suárez, Nicolás M; Davison, Andrew J; Vanderplasschen, Alain

    2017-09-01

    The genus Cyprinivirus consists of a growing list of phylogenetically related viruses, some of which cause severe economic losses to the aquaculture industry. The archetypal member, cyprinid herpesvirus 3 (CyHV-3) causes mass mortalities worldwide in koi and common carp. A CyHV-3 mutant was described previously that is attenuated in vivo by a deletion affecting two genes (ORF56 and ORF57). The relative contributions of ORF56 and ORF57 to the safety and efficacy profile of this vaccine candidate have now been assessed by analysing viruses individually deleted for ORF56 or ORF57. Inoculation of these viruses into carp demonstrated that the absence of ORF56 did not affect virulence, whereas the absence of ORF57 led to an attenuation comparable to, though slightly less than, that of the doubly deleted virus. To demonstrate further the role of ORF57 as a key virulence factor, a mutant retaining the ORF57 region but unable to express the ORF57 protein was produced by inserting multiple in-frame stop codons into the coding region. Analysis of this virus in vivo revealed a safety and efficacy profile comparable to that of the doubly deleted virus. These findings show that ORF57 encodes an essential CyHV-3 virulence factor. They also indicate that ORF57 orthologues in other cypriniviruses may offer promising targets for the rational design of attenuated recombinant vaccines. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  11. Regulation of virulence by a two-component system in group B streptococcus.

    Science.gov (United States)

    Jiang, Sheng-Mei; Cieslewicz, Michael J; Kasper, Dennis L; Wessels, Michael R

    2005-02-01

    Group B Streptococcus (GBS) is frequently carried in the gastrointestinal or genitourinary tract as a commensal organism, yet it has the potential to cause life-threatening infection in newborn infants, pregnant women, and individuals with chronic illness. Regulation of virulence factor expression may affect whether GBS behaves as an asymptomatic colonizer or an invasive pathogen, but little is known about how such factors are controlled in GBS. We now report the characterization of a GBS locus that encodes a two-component regulatory system similar to CsrRS (or CovRS) in Streptococcus pyogenes. Inactivation of csrR, encoding the putative response regulator, in two unrelated wild-type strains of GBS resulted in a marked increase in production of beta-hemolysin/cytolysin and a striking decrease in production of CAMP factor, an unrelated cytolytic toxin. Quantitative RNA hybridization experiments revealed that these two phenotypes were associated with a marked increase and decrease in expression of the corresponding genes, cylE and cfb, respectively. The CsrR mutant strains also displayed increased expression of scpB encoding C5a peptidase. Similar, but less marked, changes in gene expression were observed in CsrS (putative sensor component) mutants, evidence that CsrR and CsrS constitute a functional two-component system. Experimental infection studies in mice demonstrated reduced virulence of both CsrR and CsrS mutant strains relative to the wild type. Together, these results indicate that CsrRS regulates expression of multiple GBS virulence determinants and is likely to play an important role in GBS pathogenesis.

  12. [Research on the relevance between the virulent genes differential expression and pathogenecity of Leptospira with microarray].

    Science.gov (United States)

    Yu, De-li; Bao, Lang

    2015-01-01

    To find the change of virulent gene expression and to analyze the relevance between the virulent change and the gene expression. Grouped guinea pigs were inoculated with 1 mL Leptospira cultured in vivo, Leptospira cultured in vitro and the Leptospira culture medium through abdominal subcutaneous respectively. The survival rate, body mass and temperature change of guinea pigs in different groups were measured within 15 d after the inoculation, then the survived guinea pigs were scarified, and the organ coefficient was also measured to know the virulence of Leptospira cultured in different environment. The amplified gene segments from Leptospira were used as probes and wrote the microarray. The total RNA was extracted from Leptospira standard strain cultured in culture medium and guinea pigs. After reverse transcription to cDNA, they were labeled with Cy3 and Cy5 respectively. Labeled cDNA was mixed and hybridized with the microarray. The hybridized mircroarray was scanned and analysed. The survival rate of inoculated guinea pig was different from group to group (in vivo group: 0%; in vitro group: 88.9%; culture medium group: 100%). The guinea pigs in vivo group had a higher temperature (PLeptospira: LA1027, LA1029, LA4004, LA3050, LA3540, LA0327, LA0378, LA1650, LA3937, LA2089, LA2144, LA3576, LA0011 and gene of Loa22 were up regulation after continuously cultured in guinea pigs. The pathogenic ability of Leptospira cultured in different environment is different and the gene expression of Leptospira is different between in vivo and in vitro as well. The understanding of the meaning of this change might help to know the pathogenecity of Leptospira.

  13. PecS is an important player in the regulatory network governing the coordinated expression of virulence genes during the interaction between Dickeya dadantii 3937 and plants.

    Science.gov (United States)

    Mhedbi-Hajri, Nadia; Malfatti, Pierrette; Pédron, Jacques; Gaubert, Stéphane; Reverchon, Sylvie; Van Gijsegem, Frédérique

    2011-11-01

    Successful infection of a pathogen relies on the coordinated expression of numerous virulence factor-encoding genes. In plant-bacteria interactions, this control is very often achieved through the integration of several regulatory circuits controlling cell-cell communication or sensing environmental conditions. Dickeya dadantii (formerly Erwinia chrysanthemi), the causal agent of soft rot on many crops and ornamentals, provokes maceration of infected plants mainly by producing and secreting a battery of plant cell wall-degrading enzymes. However, several other virulence factors have also been characterized. During Arabidopsis infection, most D. dadantii virulence gene transcripts accumulated in a coordinated manner during infection. This activation requires a functional GacA-GacS two-component regulatory system but the Gac system is not involved in the growth phase dependence of virulence gene expression. Here we show that, contrary to Pectobacterium, the AHL-mediated ExpIR quorum-sensing system does not play a major role in the growth phase-dependent control of D. dadantii virulence genes. On the other hand, the global regulator PecS participates in this coordinated expression since, in a pecS mutant, an early activation of virulence genes is observed both in vitro and in planta. This correlated with the known hypervirulence phenotype of the pecS mutant. Analysis of the relationship between the regulatory circuits governed by the PecS and GacA global regulators indicates that these two regulators act independently. PecS prevents a premature expression of virulence genes in the first stages of colonization whereas GacA, presumably in conjunction with other regulators, is required for the activation of virulence genes at the onset of symptom occurrence. © 2011 Society for Applied Microbiology and Blackwell Publishing Ltd.

  14. The TOR signaling pathway regulates vegetative development and virulence in Fusarium graminearum.

    Science.gov (United States)

    Yu, Fangwei; Gu, Qin; Yun, Yingzi; Yin, Yanni; Xu, Jin-Rong; Shim, Won-Bo; Ma, Zhonghua

    2014-07-01

    The target of rapamycin (TOR) signaling pathway plays critical roles in controlling cell growth in a variety of eukaryotes. However, the contribution of this pathway in regulating virulence of plant pathogenic fungi is unknown. We identified and characterized nine genes encoding components of the TOR pathway in Fusarium graminearum. Biological, genetic and biochemical functions of each component were investigated. The FgFkbp12-rapamycin complex binds to the FgTor kinase. The type 2A phosphatases FgPp2A, FgSit4 and FgPpg1 were found to interact with FgTap42, a downstream component of FgTor. Among these, we determined that FgPp2A is likely to be essential for F. graminearum survival, and FgSit4 and FgPpg1 play important roles in cell wall integrity by positively regulating the phosphorylation of FgMgv1, a key MAP kinase in the cell wall integrity pathway. In addition, the FgPpg1 interacting protein, FgTip41, is involved in regulating mycelial growth and virulence. Notably, FgTip41 does not interact with FgTap42 but with FgPpg1, suggesting the existence of FgTap42:FgPpg1:FgTip41 heterotrimer in F. graminearum, a complex not observed in the yeast model. Collectively, we defined a genetic regulatory framework that elucidates how the TOR pathway regulates virulence and vegetative development in F. graminearum. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  15. Virulence and Stress Responses of Shigella flexneri Regulated by PhoP/PhoQ

    Directory of Open Access Journals (Sweden)

    Zhiwei Lin

    2018-01-01

    Full Text Available The two-component signal transduction system PhoP/PhoQ is an important regulator for stress responses and virulence in most Gram-negative bacteria, but characterization of PhoP/PhoQ in Shigella has not been thoroughly investigated. In the present study, we found that deletion of phoPQ (ΔphoPQ from Shigella flexneri 2a 301 (Sf301 resulted in a significant decline (reduced by more than 15-fold in invasion of HeLa cells and Caco-2 cells, and less inflammation (− or + compared to Sf301 (+++ in the guinea pig Sereny test. In low Mg2+ (10 μM medium or pH 5 medium, the ΔphoPQ strain exhibited a growth deficiency compared to Sf301. The ΔphoPQ strain was more sensitive than Sf301 to polymyxin B, an important antimicrobial agent for treating multi-resistant Gram-negative infections. By comparing the transcriptional profiles of ΔphoPQ and Sf301 using DNA microarrays, 117 differentially expressed genes (DEGs were identified, which were involved in Mg2+ transport, lipopolysaccharide modification, acid resistance, bacterial virulence, respiratory, and energy metabolism. Based on the reported PhoP box motif [(T/G GTTTA-5nt-(T/G GTTTA], we screened 38 suspected PhoP target operons in S. flexneri, and 11 of them (phoPQ, mgtA, slyB, yoaE, yrbL, icsA, yhiWX, rstA, hdeAB, pagP, and shf–rfbU-virK-msbB2 were demonstrated to be PhoP-regulated genes based on electrophoretic mobility shift assays and β-galactosidase assays. One of these PhoP-regulated genes, icsA, is a well-known virulence factor in S. flexneri. In conclusion, our data suggest that the PhoP/PhoQ system modulates S. flexneri virulence (in an icsA-dependent manner and stress responses of Mg2+, pH and antibacterial peptides.

  16. Main functions and taxonomic distribution of virulence genes in Brucella melitensis 16 M.

    Directory of Open Access Journals (Sweden)

    Aniel Jessica Leticia Brambila-Tapia

    Full Text Available Many virulence genes have been detected in attenuated mutants of Brucella melitensis 16 M; nevertheless, a complete report of these genes, including the main Cluster of Orthologous Groups (COG represented as well as the taxonomical distribution among all complete bacterial and archaeal genomes, has not been analyzed. In this work a total of 160 virulence genes that have been reported in attenuated mutants in B. melitensis were included and analyzed. Additionally, we obtained 250 B. melitensis randomly selected genes as a reference group for the taxonomical comparisons. The COGs and the taxonomical distribution profile for 789 nonredundant bacterial and archaeal genomes were obtained and compared with the whole-genome COG distribution and with the 250 randomly selected genes, respectively. The main COGs associated with virulence genes corresponded to the following: intracellular trafficking, secretion and vesicular transport (U; cell motility (N; nucleotide transport and metabolism (F; transcription (K; and cell wall/membrane/envelope biogenesis (M. In addition, we found that virulence genes presented a higher proportion of orthologs in the Euryarchaeota and Proteobacteria phyla, with a significant decrease in Chlamydiae, Bacteroidetes, Tenericutes, Firmicutes and Thermotogae. In conclusion, we found that genes related to specific functions are more relevant to B. melitensis virulence, with the COG U the most significant. Additionally, the taxonomical distribution of virulence genes highlights the importance of these genes in the related Proteobacteria, being less relevant in distant groups of organisms with the exception of Euryarchaeota.

  17. Diverse Genetic Regulon of the Virulence-Associated Transcriptional Regulator MucR in Brucella abortus 2308

    Science.gov (United States)

    Caswell, Clayton C.; Elhassanny, Ahmed E. M.; Planchin, Emilie E.; Roux, Christelle M.; Weeks-Gorospe, Jenni N.; Ficht, Thomas A.; Dunman, Paul M.

    2013-01-01

    The Ros-type regulator MucR is one of the few transcriptional regulators that have been linked to virulence in Brucella. Here, we show that a Brucella abortus in-frame mucR deletion strain exhibits a pronounced growth defect during in vitro cultivation and, more importantly, that the mucR mutant is attenuated in cultured macrophages and in mice. The genetic basis for the attenuation of Brucella mucR mutants has not been defined previously, but in the present study the genes regulated by MucR in B. abortus have been elucidated using microarray analysis and real-time reverse transcription-PCR (RT-PCR). In B. abortus 2308, MucR regulates a wide variety of genes whose products may function in establishing and maintaining cell envelope integrity, polysaccharide biosynthesis, iron homeostasis, genome plasticity, and transcriptional regulation. Particularly notable among the MucR-regulated genes identified is arsR6 (nolR), which encodes a transcriptional regulator previously linked to virulence in Brucella melitensis 16 M. Importantly, electrophoretic mobility shift assays (EMSAs) determined that a recombinant MucR protein binds directly to the promoter regions of several genes repressed by MucR (including arsR6 [nolR]), and in Brucella, as in other alphaproteobacteria, MucR binds to its own promoter to repress expression of the gene that encodes it. Overall, these studies have uncovered the diverse genetic regulon of MucR in Brucella, and in doing so this work has begun to define the MucR-controlled genetic circuitry whose misregulation contributes to the virulence defect of Brucella mucR mutants. PMID:23319565

  18. Crystal structure of Bacillus anthracis virulence regulator AtxA and effects of phosphorylated histidines on multimerization and activity

    OpenAIRE

    Hammerstrom, Troy G.; Horton, Lori B.; Swick, Michelle C.; Joachimiak, Andrzej; Osipiuk, Jerzy; Koehler, Theresa M.

    2014-01-01

    The Bacillus anthracis virulence regulator AtxA controls transcription of the anthrax toxin genes and capsule biosynthesis operon. AtxA activity is elevated during growth in media containing glucose and CO2/bicarbonate, and there is a positive correlation between the CO2/bicarbonate signal, AtxA activity, and homomultimerization. AtxA activity is also affected by phosphorylation at specific histidines. We show that AtxA crystallizes as a dimer. Distinct folds associated with predicted DNA-bin...

  19. Opposing Roles of the Staphylococcus aureus Virulence Regulators, Agr and Sar, in Triton X-100- and Penicillin-Induced Autolysis

    OpenAIRE

    Fujimoto, David F.; Bayles, Kenneth W.

    1998-01-01

    The regulation of murein hydrolases is a critical aspect of peptidoglycan growth and metabolism. In the present study, we demonstrate that mutations within the Staphylococcus aureus virulence factor regulatory genes, agr and sar, affect autolysis, resulting in decreased and increased autolysis rates, respectively. Zymographic analyses of these mutant strains suggest that agr and sar exert their effects on autolysis, in part, by modulating murein hydrolase expression and/or activity.

  20. Bacterial Human Virulence Genes across Diverse Habitats As Assessed by In silico Analysis of Environmental Metagenomes

    DEFF Research Database (Denmark)

    Søborg, Ditte A; Hendriksen, Niels B; Kilian, Mogens

    2016-01-01

    of natural environments in the evolution of bacterial virulence. Twenty four bacterial virulence genes were analyzed in 46 diverse environmental metagenomic datasets, representing various soils, seawater, freshwater, marine sediments, hot springs, the deep-sea, hypersaline mats, microbialites, gutless worms......The occurrence and distribution of clinically relevant bacterial virulence genes across natural (non-human) environments is not well understood. We aimed to investigate the occurrence of homologs to bacterial human virulence genes in a variety of ecological niches to better understand the role...... in non-human environments point to an important ecological role of the genes for the activity and survival of environmental bacteria. Furthermore, the high degree of sequence conservation between several of the environmental and clinical genes suggests common ancestral origins....

  1. ChIP-seq and in vivo transcriptome analyses of the Aspergillus fumigatus SREBP SrbA reveals a new regulator of the fungal hypoxia response and virulence.

    Directory of Open Access Journals (Sweden)

    Dawoon Chung

    2014-11-01

    Full Text Available The Aspergillus fumigatus sterol regulatory element binding protein (SREBP SrbA belongs to the basic Helix-Loop-Helix (bHLH family of transcription factors and is crucial for antifungal drug resistance and virulence. The latter phenotype is especially striking, as loss of SrbA results in complete loss of virulence in murine models of invasive pulmonary aspergillosis (IPA. How fungal SREBPs mediate fungal virulence is unknown, though it has been suggested that lack of growth in hypoxic conditions accounts for the attenuated virulence. To further understand the role of SrbA in fungal infection site pathobiology, chromatin immunoprecipitation followed by massively parallel DNA sequencing (ChIP-seq was used to identify genes under direct SrbA transcriptional regulation in hypoxia. These results confirmed the direct regulation of ergosterol biosynthesis and iron uptake by SrbA in hypoxia and revealed new roles for SrbA in nitrate assimilation and heme biosynthesis. Moreover, functional characterization of an SrbA target gene with sequence similarity to SrbA identified a new transcriptional regulator of the fungal hypoxia response and virulence, SrbB. SrbB co-regulates genes involved in heme biosynthesis and demethylation of C4-sterols with SrbA in hypoxic conditions. However, SrbB also has regulatory functions independent of SrbA including regulation of carbohydrate metabolism. Loss of SrbB markedly attenuates A. fumigatus virulence, and loss of both SREBPs further reduces in vivo fungal growth. These data suggest that both A. fumigatus SREBPs are critical for hypoxia adaptation and virulence and reveal new insights into SREBPs' complex role in infection site adaptation and fungal virulence.

  2. Detection of virulence-associated genes in Brucella melitensis ...

    African Journals Online (AJOL)

    Ibrahim Eldaghayes

    2018-03-20

    Mar 20, 2018 ... isolated from goats. This discrepancies may indicate that B. melitensis field strains prevailing in Egypt are more virulent than the strains of B. melitensis isolated from caprines in Iran. As, it was emphasized that the. T4SS of Brucella encoded by the virB operon is a major virulence factor (Delrue et al., 2005).

  3. Nigribactin, a Novel Siderophore from Vibrio nigripulchritudo, Modulates Staphylococcus aureus Virulence Gene Expression

    DEFF Research Database (Denmark)

    Nielsen, Anita; Månsson, Maria; Wietz, Matthias

    2012-01-01

    Staphylococcus aureus is a serious human pathogen that employs a number of virulence factors as part of its pathogenesis. The purpose of the present study was to explore marine bacteria as a source of compounds that modulate virulence gene expression in S. aureus. During the global marine Galathea...... 3 expedition, a strain collection was established comprising bacteria that express antimicrobial activity against Vibrio anguillarum and/or Staphylococcus aureus. Within this collection we searched colony material, culture supernatants, and cell extracts for virulence modulating activity showing......, enterobactin, failed to influence S. aureus virulence gene expression. This study shows that marine microorganisms produce compounds with potential use in therapeutic strategies targeting virulence rather than viability of human pathogens....

  4. Uropathogenic Escherichia coli virulence genes: invaluable approaches for designing DNA microarray probes.

    Science.gov (United States)

    Jahandeh, Nadia; Ranjbar, Reza; Behzadi, Payam; Behzadi, Elham

    2015-01-01

    The pathotypes of uropathogenic Escherichia coli (UPEC) cause different types of urinary tract infections (UTIs). The presence of a wide range of virulence genes in UPEC enables us to design appropriate DNA microarray probes. These probes, which are used in DNA microarray technology, provide us with an accurate and rapid diagnosis and definitive treatment in association with UTIs caused by UPEC pathotypes. The main goal of this article is to introduce the UPEC virulence genes as invaluable approaches for designing DNA microarray probes. Main search engines such as Google Scholar and databases like NCBI were searched to find and study several original pieces of literature, review articles, and DNA gene sequences. In parallel with in silico studies, the experiences of the authors were helpful for selecting appropriate sources and writing this review article. There is a significant variety of virulence genes among UPEC strains. The DNA sequences of virulence genes are fabulous patterns for designing microarray probes. The location of virulence genes and their sequence lengths influence the quality of probes. The use of selected virulence genes for designing microarray probes gives us a wide range of choices from which the best probe candidates can be chosen. DNA microarray technology provides us with an accurate, rapid, cost-effective, sensitive, and specific molecular diagnostic method which is facilitated by designing microarray probes. Via these tools, we are able to have an accurate diagnosis and a definitive treatment regarding UTIs caused by UPEC pathotypes.

  5. Wide distribution of virulence genes among Enterococcus faecium and Enterococcus faecalis clinical isolates.

    Science.gov (United States)

    Soheili, Sara; Ghafourian, Sobhan; Sekawi, Zamberi; Neela, Vasanthakumari; Sadeghifard, Nourkhoda; Ramli, Ramliza; Hamat, Rukman Awang

    2014-01-01

    Enterococcus, a Gram-positive facultative anaerobic cocci belonging to the lactic acid bacteria of the phylum Firmicutes, is known to be able to resist a wide range of hostile conditions such as different pH levels, high concentration of NaCl (6.5%), and the extended temperatures between 5(°)C and 65(°)C. Despite being the third most common nosocomial pathogen, our understanding on its virulence factors is still poorly understood. The current study was aimed to determine the prevalence of different virulence genes in Enterococcus faecalis and Enterococcus faecium. For this purpose, 79 clinical isolates of Malaysian enterococci were evaluated for the presence of virulence genes. pilB, fms8, efaAfm, and sgrA genes are prevalent in all clinical isolates. In conclusion, the pathogenicity of E. faecalis and E. faecium could be associated with different virulence factors and these genes are widely distributed among the enterococcal species.

  6. Protocols for screening antimicrobial peptides that influence virulence gene expression in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Bojer, Martin Saxtorph; Baldry, Mara; Ingmer, Hanne

    2017-01-01

    Compounds that inhibit virulence gene expression in bacterial pathogens have received increasing interest as possible alternatives to the traditional antibiotic treatment of infections. For the human pathogen Staphylococcus aureus, we have developed two simple assays based on reporter gene fusions...... to central virulence genes that are easily applicable for screening various sources of natural and synthetic peptides for anti-virulence effects. The plate assay is qualitative but simultaneously assesses the effect of gradient concentrations of the investigated compound, whereas the liquid assay...... is quantitative and can be employed to address whether a compound is acting on the central quorum sensing regulatory system, agr, that controls a large number of virulence genes in S. aureus....

  7. Proteolysis of virulence regulator ToxR is associated with entry of Vibrio cholerae into a dormant state.

    Directory of Open Access Journals (Sweden)

    Salvador Almagro-Moreno

    2015-04-01

    Full Text Available Vibrio cholerae O1 is a natural inhabitant of aquatic environments and causes the diarrheal disease, cholera. Two of its primary virulence regulators, TcpP and ToxR, are localized in the inner membrane. TcpP is encoded on the Vibrio Pathogenicity Island (VPI, a horizontally acquired mobile genetic element, and functions primarily in virulence gene regulation. TcpP has been shown to undergo regulated intramembrane proteolysis (RIP in response to environmental conditions that are unfavorable for virulence gene expression. ToxR is encoded in the ancestral genome and is present in non-pathogenic strains of V. cholerae, indicating it has roles outside of the human host. In this study, we show that ToxR undergoes RIP in V. cholerae in response to nutrient limitation at alkaline pH, a condition that occurs during the stationary phase of growth. This process involves the site-2 protease RseP (YaeL, and is dependent upon the RpoE-mediated periplasmic stress response, as deletion mutants for the genes encoding these two proteins cannot proteolyze ToxR under nutrient limitation at alkaline pH. We determined that the loss of ToxR, genetically or by proteolysis, is associated with entry of V. cholerae into a dormant state in which the bacterium is normally found in the aquatic environment called viable but nonculturable (VBNC. Strains that can proteolyze ToxR, or do not encode it, lose culturability, experience a change in morphology associated with cells in VBNC, yet remain viable under nutrient limitation at alkaline pH. On the other hand, mutant strains that cannot proteolyze ToxR remain culturable and maintain the morphology of cells in an active state of growth. Overall, our findings provide a link between the proteolysis of a virulence regulator and the entry of a pathogen into an environmentally persistent state.

  8. Identification of virulence genes carried by bacteriophages obtained from clinically isolated methicillin-resistant Staphylococcus aureus.

    Science.gov (United States)

    Karasartova, Djursun; Cavusoglu, Zeynep Burcin; Turegun, Buse; Ozsan, Murat T; Şahin, Fikret

    2016-12-01

    Bacteriophages play an important role in the pathogenicity of Staphylococcus aureus (S. aureus) either by carrying accessory virulence factors or several superantigens. Despite their importance, there are not many studies showing the actual distribution of the virulence genes carried by the prophages obtained from the clinically isolated Staphylococcus. In this study, we investigated prophages obtained from methicillin-resistant S. aureus (MRSA) strains isolated from hospital- and community-associated (HA-CA) infections for the virulence factors. In the study, 43 phages isolated from 48 MRSA were investigated for carrying toxin genes including the sak, eta, lukF-PV, sea, selp, sek, seg, seq chp, and scn virulence genes using polymerase chain reaction (PCR) and Southern blot. Restriction fragment length polymorphism was used to analyze phage genomes to investigate the relationship between the phage profiles and the toxin genes' presence. MRSA strains isolated from HA infections tended to have higher prophage presence than the MRSA strains obtained from the CA infections (97% and 67%, respectively). The study showed that all the phages with the exception of one phage contained one or more virulence genes in their genomes with different combinations. The most common toxin genes found were sea (83%) followed by sek (77%) and seq (64%). The study indicates that prophages encode a significant proportion of MRSA virulence factors.

  9. Post-transcriptional regulation of gene expression in Yersinia species

    Directory of Open Access Journals (Sweden)

    Chelsea A Schiano

    2012-11-01

    Full Text Available Proper regulation of gene expression is required by bacterial pathogens to respond to continually changing environmental conditions and the host response during the infectious process. While transcriptional regulation is perhaps the most well understood form of controlling gene expression, recent studies have demonstrated the importance of post-transcriptional mechanisms of gene regulation that allow for more refined management of the bacterial response to host conditions. Yersinia species of bacteria are known to use various forms of post-transcriptional regulation for control of many virulence-associated genes. These include regulation by cis- and trans-acting small non-coding RNAs, RNA-binding proteins, RNases, and thermoswitches. The effects of these and other regulatory mechanisms on Yersinia physiology can be profound and have been shown to influence type III secretion, motility, biofilm formation, host cell invasion, intracellular survival and replication, and more. In this review, we will discuss these and other post-transcriptional mechanisms and their influence on virulence gene regulation, with a particular emphasis on how these processes influence the virulence of Yersinia in the host.

  10. Cytokine responses in primary chicken embryo intestinal cells infected with Campylobacter jejuni strains of human and chicken origin and the expression of bacterial virulence-associated genes

    DEFF Research Database (Denmark)

    Li, Yiping; Ingmer, Hanne; Madsen, Mogens

    2008-01-01

    of the bacterial genes. We have investigated the invasiveness of primary chicken embryo intestinal cells (CEICs) by C. jejuni strains of human and chicken origins and the production of pro-inflammatory cytokines as well as the expression of the bacterial virulence-associated genes during co-cultivation. Results C......-free media from another co-cultivation experiment also increased the expression of the virulence-associated genes in the C. jejuni chicken isolate, indicating that the expression of bacterial genes is regulated by component(s) secreted upon co-cultivation of bacteria and CEICs. Conclusion We show that under...... in vitro culture condition C. jejuni strains of both human and chicken origins can invade avian host cells with a pro-inflammatory response and that the virulence-associated genes of C. jejuni may play a role in this process....

  11. Three regulators of G protein signaling differentially affect mating, morphology and virulence in the smut fungus Ustilago maydis.

    Science.gov (United States)

    Moretti, Marino; Wang, Lei; Grognet, Pierre; Lanver, Daniel; Link, Hannes; Kahmann, Regine

    2017-09-01

    Regulators of G protein signaling (RGS) proteins modulate heterotrimeric G protein signaling negatively. To broaden an understanding of the roles of RGS proteins in fungal pathogens, we functionally characterized the three RGS protein-encoding genes (rgs1, rgs2 and rgs3) in the phytopathogenic fungus Ustilago maydis. It was found that RGS proteins played distinct roles in the regulation of development and virulence. rgs1 had a minor role in virulence when deleted in a solopathogenic strain. In crosses, rgs1 was dispensable for mating and filamentation, but was required for teliospore production. Haploid rgs2 mutants were affected in cell morphology, growth, mating and were unable to cause disease symptoms in crosses. However, virulence was unaffected when rgs2 was deleted in a solopathogenic strain, suggesting an exclusive involvement in pre-fusion events. These rgs2 phenotypes are likely connected to elevated intracellular cAMP levels. rgs3 mutants were severely attenuated in mating, in their response to pheromone, virulence and formation of mature teliospores. The mating defect could be traced back to reduced expression of the transcription factor rop1. It was speculated that the distinct roles of the three U. maydis RGS proteins were achieved by direct modulation of the Gα subunit-activated signaling pathways as well as through Gα-independent functions. © 2017 John Wiley & Sons Ltd.

  12. Iron regulation of the major virulence factors in the AIDS-associated pathogen Cryptococcus neoformans.

    Directory of Open Access Journals (Sweden)

    Won Hee Jung

    2006-11-01

    Full Text Available Iron overload is known to exacerbate many infectious diseases, and conversely, iron withholding is an important defense strategy for mammalian hosts. Iron is a critical cue for Cryptococcus neoformans because the fungus senses iron to regulate elaboration of the polysaccharide capsule that is the major virulence factor during infection. Excess iron exacerbates experimental cryptococcosis and the prevalence of this disease in Sub-Saharan Africa has been associated with nutritional and genetic aspects of iron loading in the background of the HIV/AIDS epidemic. We demonstrate that the iron-responsive transcription factor Cir1 in Cr. neoformans controls the regulon of genes for iron acquisition such that cir1 mutants are "blind" to changes in external iron levels. Cir1 also controls the known major virulence factors of the pathogen including the capsule, the formation of the anti-oxidant melanin in the cell wall, and the ability to grow at host body temperature. Thus, the fungus is remarkably tuned to perceive iron as part of the disease process, as confirmed by the avirulence of the cir1 mutant; this characteristic of the pathogen may provide opportunities for antifungal treatment.

  13. Induction of virulence gene expression in Staphylococcus aureus by pulmonary surfactant.

    Science.gov (United States)

    Ishii, Kenichi; Adachi, Tatsuo; Yasukawa, Jyunichiro; Suzuki, Yutaka; Hamamoto, Hiroshi; Sekimizu, Kazuhisa

    2014-04-01

    We performed a genomewide analysis using a next-generation sequencer to investigate the effect of pulmonary surfactant on gene expression in Staphylococcus aureus, a clinically important opportunistic pathogen. RNA sequence (RNA-seq) analysis of bacterial transcripts at late log phase revealed 142 genes that were upregulated >2-fold following the addition of pulmonary surfactant to the culture medium. Among these genes, we confirmed by quantitative reverse transcription-PCR analysis that mRNA amounts for genes encoding ESAT-6 secretion system C (EssC), an unknown hypothetical protein (NWMN_0246; also called pulmonary surfactant-inducible factor A [PsiA] in this study), and hemolysin gamma subunit B (HlgB) were increased 3- to 10-fold by the surfactant treatment. Among the major constituents of pulmonary surfactant, i.e., phospholipids and palmitate, only palmitate, which is the most abundant fatty acid in the pulmonary surfactant and a known antibacterial substance, stimulated the expression of these three genes. Moreover, these genes were also induced by supplementing the culture with detergents. The induction of gene expression by surfactant or palmitate was not observed in a disruption mutant of the sigB gene, which encodes an alternative sigma factor involved in bacterial stress responses. Furthermore, each disruption mutant of the essC, psiA, and hlgB genes showed attenuation of both survival in the lung and host-killing ability in a murine pneumonia model. These findings suggest that S. aureus resists membrane stress caused by free fatty acids present in the pulmonary surfactant through the regulation of virulence gene expression, which contributes to its pathogenesis within the lungs of the host animal.

  14. The prevalence of virulence genes of E. coli strains isolated from children with urinary tract infection

    Directory of Open Access Journals (Sweden)

    Farshad Shohreh

    2009-01-01

    Full Text Available To evaluate the prevalence of virulence genes in E. coli strains isolated from urine samples of children with urinary tract infection(UTI and their correlation with clinical data, we iso-lated E. coli strains from urine samples of children with UTI during the period of August 2005 - August 2006 and studied them for the presence of the virulence genes by PCR. A total of 96 E. coli strains were isolated. The prevalence of genes, pyelonephritis associated pili (pap genes, S-family adhesions (sfa gene, hemolysin (hly gene, and cytotoxic nercotizing factor type 1 (cnf-1-1 gene among the isolated strains was 27.1%, 14.6%, 13.5% and 22.9 %, respectively. Pyelonephritis was more prevalent in the cases with positive virulence genes. The results showed significant correlation bet-ween age of the patient and the presence of the genes (P< 0.05. Cnf-1 gene was significantly more common in samples of patients with abnormal finding on the ultrasound of kidneys (P= 0.049. Our study demonstrated higher prevalence of pyelonephritis in the presence of E. coli virulence genes. Detection of the genes in urine samples may help in the management of UTI.

  15. The prevalence of virulence genes of E. coli strains isolated from children with urinary tract infection

    International Nuclear Information System (INIS)

    Farshad, Shohreh; Emamghorashi, Fatemeh

    2009-01-01

    To evaluate the prevalence of virulence genes in E. coli strains isolated from urine samples of children with urinary tract infection(UTI) and their correlation with clinical data, we isolated E. coli strains from urine samples of children with UTI during the period of August 2005 - August 2006 and studied them for the presence of the virulence genes by PCR. A total of 96 E. coli strains were isolated. The prevalence of genes, pyelonephritis associated pili (pap genes), S-family adhesions (sfa gene), hemolysin (hly gene), and cytotoxic nercotizing factor type 1 (cnf-1-1 gene) among the isolated strains was 27.1%, 14.6%, 13.5% and 22.9 %, respectively. Pyelonephritis was more prevalent in the cases with positive virulence genes. The results showed significant correlation between age of the patient and the presence of the genes (P< 0.05). Cnf-1 gene was significantly more common in samples of patients with abnormal finding on the ultrasound of kidneys (P0.049). Our study demonstrated higher prevalence of pyelonephritis in the presence of E. coli virulence genes. Detection of the genes in urine samples may help in the management of UTI. (author)

  16. Deletion analysis of Streptococcus pneumoniae late competence genes distinguishes virulence determinants that are dependent or independent of competence induction.

    Science.gov (United States)

    Zhu, Luchang; Lin, Jingjun; Kuang, Zhizhou; Vidal, Jorge E; Lau, Gee W

    2015-07-01

    The competence regulon of Streptococcus pneumoniae (pneumococcus) is crucial for genetic transformation. During competence development, the alternative sigma factor ComX is activated, which in turn, initiates transcription of 80 'late' competence genes. Interestingly, only 16 late genes are essential for genetic transformation. We hypothesized that these late genes that are dispensable for competence are beneficial to pneumococcal fitness during infection. These late genes were systematically deleted, and the resulting mutants were examined for their fitness during mouse models of bacteremia and acute pneumonia. Among these, 14 late genes were important for fitness in mice. Significantly, deletion of some late genes attenuated pneumococcal fitness to the same level in both wild-type and ComX-null genetic backgrounds, suggesting that the constitutive baseline expression of these genes was important for bacterial fitness. In contrast, some mutants were attenuated only in the wild-type genetic background but not in the ComX-null background, suggesting that specific expression of these genes during competence state contributed to pneumococcal fitness. Increased virulence during competence state was partially caused by the induction of allolytic enzymes that enhanced pneumolysin release. These results distinguish the role of basal expression versus competence induction in virulence functions encoded by ComX-regulated late competence genes. © 2015 John Wiley & Sons Ltd.

  17. Identifying Virulence-Associated Genes Using Transcriptomic and Proteomic Association Analyses of the Plant Parasitic Nematode Bursaphelenchus mucronatus

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    Lifeng Zhou

    2016-09-01

    Full Text Available Bursaphelenchus mucronatus (B. mucronatus isolates that originate from different regions may vary in their virulence, but their virulence-associated genes and proteins are poorly understood. Thus, we conducted an integrated study coupling RNA-Seq and isobaric tags for relative and absolute quantitation (iTRAQ to analyse transcriptomic and proteomic data of highly and weakly virulent B. mucronatus isolates during the pathogenic processes. Approximately 40,000 annotated unigenes and 5000 proteins were gained from the isolates. When we matched all of the proteins with their detected transcripts, a low correlation coefficient of r = 0.138 was found, indicating probable post-transcriptional gene regulation involved in the pathogenic processes. A functional analysis showed that five differentially expressed proteins which were all highly expressed in the highly virulent isolate were involved in the pathogenic processes of nematodes. Peroxiredoxin, fatty acid- and retinol-binding protein, and glutathione peroxidase relate to resistance against plant defence responses, while β-1,4-endoglucanase and expansin are associated with the breakdown of plant cell walls. Thus, the pathogenesis of B. mucronatus depends on its successful survival in host plants. Our work adds to the understanding of B. mucronatus’ pathogenesis, and will aid in controlling B. mucronatus and other pinewood nematode species complexes in the future.

  18. Virulence gene profiles of avian pathogenic Escherichia coli isolated from chickens with colibacillosis in Bulawayo, Zimbabwe

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    Joshua Mbanga

    2015-04-01

    Full Text Available Colibacillosis, a disease caused by avian pathogenic Escherichia coli (APEC, is one of the main causes of economic losses in the poultry industry worldwide. This study was carried out in order to determine the APEC-associated virulence genes contained by E. coli isolates causing colibacillosis in chickens. A total of 45 E. coli isolates were obtained from the diagnostics and research branch of the Central Veterinary Laboratories, Bulawayo, Zimbabwe. These isolates were obtained from chickens with confirmed cases of colibacillosis after postmortem examination. The presence of the iutA, hlyF, ompT, frz, sitD, fimH, kpsM, sitA, sopB, uvrY, pstB and vat genes were investigated by multiplex polymerase chain reaction (PCR assay. Of the 45 isolates, 93% were positive for the presence of at least one virulence gene. The three most prevalent virulence genes were iutA (80%, fimH (33.3% and hlyF (24.4%. The kpsM, pstB and ompT genes had the lowest prevalence, having been detected in only 2.2% of the isolates. All 12 virulence genes studied were detected in the 45 APEC isolates. Virulence gene profiles were constructed for each APEC isolate from the multiplex data. The APEC isolates were profiled as 62.2% fitting profile A, 31.1% profile B and 6.7% profile C. None of the isolates had more than seven virulence genes. Virulence profiles of Zimbabwean APEC isolates are different from those previously reported. Zimbabwean APEC isolates appear to be less pathogenic and may rely on environmental factors and stress in hosts to establish infection.

  19. Staphylococcus aureus hyaluronidase is a CodY-regulated virulence factor.

    Science.gov (United States)

    Ibberson, Carolyn B; Jones, Crystal L; Singh, Shweta; Wise, Matthew C; Hart, Mark E; Zurawski, Daniel V; Horswill, Alexander R

    2014-10-01

    Staphylococcus aureus is a Gram-positive pathogen that causes a diverse range of bacterial infections. Invasive S. aureus strains secrete an extensive arsenal of hemolysins, immunomodulators, and exoenzymes to cause disease. Our studies have focused on the secreted enzyme hyaluronidase (HysA), which cleaves the hyaluronic acid polymer at the β-1,4 glycosidic bond. In the study described in this report, we have investigated the regulation and contribution of this enzyme to S. aureus pathogenesis. Using the Nebraska Transposon Mutant Library (NTML), we identified eight insertions that modulate extracellular levels of HysA activity. Insertions in the sigB operon, as well as in genes encoding the global regulators SarA and CodY, significantly increased HysA protein levels and activity. By altering the availability of branched-chain amino acids, we further demonstrated CodY-dependent repression of HysA activity. Additionally, through mutation of the CodY binding box upstream of hysA, the repression of HysA production was lost, suggesting that CodY is a direct repressor of hysA expression. To determine whether HysA is a virulence factor, a ΔhysA mutant of a community-associated methicillin-resistant S. aureus (CA-MRSA) USA300 strain was constructed and found to be attenuated in a neutropenic, murine model of pulmonary infection. Mice infected with this mutant strain exhibited a 4-log-unit reduction in bacterial burden in their lungs, as well as reduced lung pathology and increased levels of pulmonary hyaluronic acid, compared to mice infected with the wild-type, parent strain. Taken together, these results indicate that S. aureus hyaluronidase is a CodY-regulated virulence factor. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  20. Screening of virulence genes in Staphylococcus aureus isolates from rabbits

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    David Viana Martín

    2015-09-01

    Full Text Available Staphylococcus aureus is a versatile pathogen able to cause disease in both humans and animals. In rabbits, this bacterium infects animals of different ages, producing several purulent lesions. The ability of S. aureus to cause disease depends on a combination of virulence factors. The aim of this study was therefore to investigate the distribution of bacterial virulence determinants in 69 S. aureus isolates from rabbits. Some virulence factors (7 adhesins, 1 toxin and 1 protease were positive in all rabbit S. aureus isolates analysed, while others (1 adhesin and 10 toxins were always negative. The remaining virulence factors were more variable among isolates. An association between genotype and the different profiles of virulence factors was observed, but not with the type of lesion (P<0.05. One strain of each genotype was further analysed by multilocus sequence typing, generating ST121, ST96 and ST2951, determining a greater number of enterotoxins in ST121 isolates compared to ST96 and ST2951 isolates, which could justify the different pathogenicity between strains. 

  1. Norlichexanthone Reduces Virulence Gene Expression and Biofilm Formation in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Baldry, Mara; Nielsen, Anita; Bojer, Martin S.

    2016-01-01

    characterise the mode of action of norlichexanthone and its effect on biofilm formation. We find that norlichexanthone reduces expression of both hla and RNAIII also in strain USA300. Structurally, norlichexanthone resembles ω-hydroxyemodin that recently was shown to bind the agr two component response......-hydroxyemodin however, norlichexanthone reduces staphylococcal biofilm formation. Transcriptomic analysis revealed that genes regulated by the SaeRS two-component system are repressed by norlichexanthone when compared to untreated cells, an effect that was mitigated in strain Newman carrying a partially constitutive...... SaeRS system. Our data show that norlichexanthone treatment reduces expression of key virulence factors in CA-MRSA strain USA300 via AgrA binding and represses biofilm formation....

  2. Virulence Gene Pool Detected in Bovine Group C Streptococcus dysgalactiae subsp. dysgalactiae Isolates by Use of a Group A S. pyogenes Virulence Microarray ▿

    Science.gov (United States)

    Rato, Márcia G.; Nerlich, Andreas; Bergmann, René; Bexiga, Ricardo; Nunes, Sandro F.; Vilela, Cristina L.; Santos-Sanches, Ilda; Chhatwal, Gursharan S.

    2011-01-01

    A custom-designed microarray containing 220 virulence genes of Streptococcus pyogenes (group A Streptococcus [GAS]) was used to test group C Streptococcus dysgalactiae subsp. dysgalactiae (GCS) field strains causing bovine mastitis and group C or group G Streptococcus dysgalactiae subsp. equisimilis (GCS/GGS) isolates from human infections, with the latter being used for comparative purposes, for the presence of virulence genes. All bovine and all human isolates carried a fraction of the 220 genes (23% and 39%, respectively). The virulence genes encoding streptolysin S, glyceraldehyde-3-phosphate dehydrogenase, the plasminogen-binding M-like protein PAM, and the collagen-like protein SclB were detected in the majority of both bovine and human isolates (94 to 100%). Virulence factors, usually carried by human beta-hemolytic streptococcal pathogens, such as streptokinase, laminin-binding protein, and the C5a peptidase precursor, were detected in all human isolates but not in bovine isolates. Additionally, GAS bacteriophage-associated virulence genes encoding superantigens, DNase, and/or streptodornase were detected in bovine isolates (72%) but not in the human isolates. Determinants located in non-bacteriophage-related mobile elements, such as the gene encoding R28, were detected in all bovine and human isolates. Several virulence genes, including genes of bacteriophage origin, were shown to be expressed by reverse transcriptase PCR (RT-PCR). Phylogenetic analysis of superantigen gene sequences revealed a high level (>98%) of identity among genes of bovine GCS, of the horse pathogen Streptococcus equi subsp. equi, and of the human pathogen GAS. Our findings indicate that alpha-hemolytic bovine GCS, an important mastitis pathogen and considered to be a nonhuman pathogen, carries important virulence factors responsible for virulence and pathogenesis in humans. PMID:21525223

  3. Role and regulation of the Flp/Tad pilus in the virulence of Pectobacterium atrosepticum SCRI1043 and Pectobacterium wasabiae SCC3193.

    Science.gov (United States)

    Nykyri, Johanna; Mattinen, Laura; Niemi, Outi; Adhikari, Satish; Kõiv, Viia; Somervuo, Panu; Fang, Xin; Auvinen, Petri; Mäe, Andres; Palva, E Tapio; Pirhonen, Minna

    2013-01-01

    In this study, we characterized a putative Flp/Tad pilus-encoding gene cluster, and we examined its regulation at the transcriptional level and its role in the virulence of potato pathogenic enterobacteria of the genus Pectobacterium. The Flp/Tad pilus-encoding gene clusters in Pectobacterium atrosepticum, Pectobacterium wasabiae and Pectobacterium aroidearum were compared to previously characterized flp/tad gene clusters, including that of the well-studied Flp/Tad pilus model organism Aggregatibacter actinomycetemcomitans, in which this pilus is a major virulence determinant. Comparative analyses revealed substantial protein sequence similarity and open reading frame synteny between the previously characterized flp/tad gene clusters and the cluster in Pectobacterium, suggesting that the predicted flp/tad gene cluster in Pectobacterium encodes a Flp/Tad pilus-like structure. We detected genes for a novel two-component system adjacent to the flp/tad gene cluster in Pectobacterium, and mutant analysis demonstrated that this system has a positive effect on the transcription of selected Flp/Tad pilus biogenesis genes, suggesting that this response regulator regulate the flp/tad gene cluster. Mutagenesis of either the predicted regulator gene or selected Flp/Tad pilus biogenesis genes had a significant impact on the maceration ability of the bacterial strains in potato tubers, indicating that the Flp/Tad pilus-encoding gene cluster represents a novel virulence determinant in Pectobacterium. Soft-rot enterobacteria in the genera Pectobacterium and Dickeya are of great agricultural importance, and an investigation of the virulence of these pathogens could facilitate improvements in agricultural practices, thus benefiting farmers, the potato industry and consumers.

  4. Evaluation of Antimicrobial Resistance and Virulence Genes in Uropathogenic Escherichia coli in Pediatric and Adult Patients

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    Kerem YILMAZ

    2017-06-01

    Full Text Available We aimed to evaluate the antimicrobial resistance patterns and the prevalence of certain virulence genes in uropathogenic E. coli isolated from pediatric and adult patients with uncomplicated urinary tract infection.We examined nonduplicate 83 uropathogenic E. coli isolated from mid-stream clean-catch urine samples of the pediatric and adult outpatients with the diagnosis of acute uncomplicated urinary tract infection. VITEK® 2 automated system (bioMerieux, Marcy l’Etoile, France was used for identification and determination of antimicrobial resistance. We examined the isolates in respect to their antimicrobial resistance patterns and the presence of virulence genes (pap, aer, sfa, hly and cnf-1. Antimicrobial susceptibility testing results of the E. coli isolates revealed that commonly used empiric antimicrobials (ciprofloxacin, trimethoprim–sulfamethoxazole, gentamicin, ampicillin and cephalothin for urinary tract infections were less effective than others. Most frequently detected virulence genes were pap and aer in both age groups. Sfa and hly genes were the least frequently detected genes in the pediatric age group; hly gene was the also the least common in the adult age group. There was no association with virulence factors and antimicrobial resistance patterns of the uropathogenic E. coli isolates in contrary to literature. More comprehensive studies with larger sample groups are needed to demonstrate the relation between virulence factors with antimicrobial drugs in different age groups.

  5. Reciprocal interaction between dental alloy biocorrosion and Streptococcus mutans virulent gene expression.

    Science.gov (United States)

    Zhang, Songmei; Qiu, Jing; Ren, Yanfang; Yu, Weiqiang; Zhang, Fuqiang; Liu, Xiuxin

    2016-04-01

    Corrosion of dental alloys is a major concern in dental restorations. Streptococcus mutans reduces the pH in oral cavity and induces demineralization of the enamel as well as corrosion of restorative dental materials. The rough surfaces of dental alloys induced by corrosion enhance the subsequent accumulation of plaque. In this study, the corrosion process of nickel-chromium (Ni-Cr) and cobalt-chromium (Co-Cr) alloys in a nutrient-rich medium containing S. mutans was studied using inductively coupled plasma atomic emission spectrometry (ICP-AES), X-ray photoelectron spectroscopy (XPS) and electrochemical corrosion test. Our results showed that the release of Ni and Co ions increased, particularly after incubation for 3 days. The electrochemical corrosion results showed a significant decrease in the corrosion resistance (Rp) value after the alloys were immersed in the media containing S. mutans for 3 days. Correspondingly, XPS revealed a reduction in the relative dominance of Ni, Co, and Cr in the surface oxides after the alloys were immersed in the S. mutans culture. After removal of the biofilm, the pre-corroded alloys were re-incubated in S. mutans medium, and the expressions of genes associated with the adhesion and acidogenesis of S. mutans, including gtfBCD, gbpB, fif and ldh, were evaluated by detecting the mRNA levels using real-time reverse transcription polymerase chain reaction (RT-PCR). We found that the gtfBCD, gbpB, ftf and Idh expression of S. mutans were noticeably increased after incubation with pre-corroded alloys for 24 h. This study demonstrated that S. mutans enhanced the corrosion behavior of the dental alloys, on the other hand, the presence of corroded alloy surfaces up-regulated the virulent gene expression in S. mutans. Compared with smooth surfaces, the rough corroded surfaces of dental alloys accelerated the bacteria-adhesion and corrosion process by changing the virulence gene expression of S. mutans.

  6. A Bistable Switch and Anatomical Site Control Vibrio cholerae Virulence Gene Expression in the Intestine

    DEFF Research Database (Denmark)

    Nielsen, Alex Toftgaard; Dolganov, N. A.; Rasmussen, Thomas

    2010-01-01

    A fundamental, but unanswered question in host-pathogen interactions is the timing, localization and population distribution of virulence gene expression during infection. Here, microarray and in situ single cell expression methods were used to study Vibrio cholerae growth and virulence gene...... expression during infection of the rabbit ligated ileal loop model of cholera. Genes encoding the toxin-coregulated pilus (TCP) and cholera toxin (CT) were powerfully expressed early in the infectious process in bacteria adjacent to epithelial surfaces. Increased growth was found to co......, a chemical inducer of virulence gene expression. Striking bifurcation of the population occurred during entry into stationary phase: one subpopulation continued to express tcpA, whereas the expression declined in the other subpopulation. ctxA, encoding the A subunit of CT, and toxT, encoding the proximal...

  7. Coronavirus gene 7 counteracts host defenses and modulates virus virulence.

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    Jazmina L G Cruz

    2011-06-01

    Full Text Available Transmissible gastroenteritis virus (TGEV genome contains three accessory genes: 3a, 3b and 7. Gene 7 is only present in members of coronavirus genus a1, and encodes a hydrophobic protein of 78 aa. To study gene 7 function, a recombinant TGEV virus lacking gene 7 was engineered (rTGEV-Δ7. Both the mutant and the parental (rTGEV-wt viruses showed the same growth and viral RNA accumulation kinetics in tissue cultures. Nevertheless, cells infected with rTGEV-Δ7 virus showed an increased cytopathic effect caused by an enhanced apoptosis mediated by caspase activation. Macromolecular synthesis analysis showed that rTGEV-Δ7 virus infection led to host translational shut-off and increased cellular RNA degradation compared with rTGEV-wt infection. An increase of eukaryotic translation initiation factor 2 (eIF2α phosphorylation and an enhanced nuclease, most likely RNase L, activity were observed in rTGEV-Δ7 virus infected cells. These results suggested that the removal of gene 7 promoted an intensified dsRNA-activated host antiviral response. In protein 7 a conserved sequence motif that potentially mediates binding to protein phosphatase 1 catalytic subunit (PP1c, a key regulator of the cell antiviral defenses, was identified. We postulated that TGEV protein 7 may counteract host antiviral response by its association with PP1c. In fact, pull-down assays demonstrated the interaction between TGEV protein 7, but not a protein 7 mutant lacking PP1c binding motif, with PP1. Moreover, the interaction between protein 7 and PP1 was required, during the infection, for eIF2α dephosphorylation and inhibition of cell RNA degradation. Inoculation of newborn piglets with rTGEV-Δ7 and rTGEV-wt viruses showed that rTGEV-Δ7 virus presented accelerated growth kinetics and pathology compared with the parental virus. Overall, the results indicated that gene 7 counteracted host cell defenses, and modified TGEV persistence increasing TGEV survival. Therefore, the

  8. Antimicrobial medium- and long-chain free fatty acids prevent PrfA-dependent activation of virulence genes in Listeria monocytogenes.

    Science.gov (United States)

    Sternkopf Lillebæk, Eva Maria; Lambert Nielsen, Stine; Scheel Thomasen, Rikke; Færgeman, Nils J; Kallipolitis, Birgitte H

    The foodborne pathogen Listeria monocytogenes is the causative agent of the invasive disease listeriosis. Infection by L. monocytogenes involves bacterial crossing of the intestinal barrier and intracellular replication in a variety of host cells. The PrfA protein is the master regulator of virulence factors required for bacterial entry, intracellular replication and cell-to-cell spread. PrfA-dependent activation of virulence genes occurs primarily in the blood and during intracellular infection. In contrast, PrfA does not play a significant role in regulation of virulence gene expression in the intestinal environment. In the gastrointestinal phase of infection, the bacterium encounters a variety of antimicrobial agents, including medium- and long-chain free fatty acids that are commonly found in our diet and as active components of bile. Here we show that subinhibitory concentrations of specific antimicrobial free fatty acids act to downregulate transcription of PrfA-activated virulence genes. Interestingly, the inhibitory effect is also evident in cells encoding a constitutively active variant of PrfA. Collectively, our data suggest that antimicrobial medium- and long-chain free fatty acids may act as signals to prevent PrfA-mediated activation of virulence genes in environments where PrfA activation is not required, such as in food and the gastrointestinal tract. Copyright © 2017 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  9. Virulence factor genes possessing Enterococcus faecalis strains from rabbits and their sensitivity to enterocins

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    M. Pogány Simonová

    2017-03-01

    Full Text Available Information concerning the virulence factor genes and antibiotic resistance of rabbit enterococci is limited, so in this study we tested the virulence factor genes in Enterococcus faecalis strains from rabbits. Moreover, their resistance/sensitivity to antibiotics and sensitivity to enterocins was also tested, with the aim of contributing to our enterocin spectra study and to indicate the possibility of enterocin application in prevention or contaminant elimination in rabbit husbandry. A total of 144 rabbit samples were treated using a standard microbiological method. Thirty-one pure colonies of the species Enterococcus faecalis were identified, using the MALDI-TOF identification system and confirmed using phenotyping, among which 15 strains were virulence factor gene absent. The gelE gene was the most detected (42%; however, the expression of gelatinase phenotype did not always correlate with the detection of gelE. Strains did not show ß-haemolysis and were mostly resistant to tested antibiotics, but sensitive to enterocins (Ent, mainly to Ents EK13=A (P, 2019 and Ent M. Rabbit E. faecalis strains displayed antibiotic resistant traits and the presence of expressed and silent virulence genes, but they showed high levels of sensitivity to natural antimicrobials-enterocins, which indicates the possible prevention of multidrug and virulent enterococcal contaminants by enterocins.

  10. The Composition and Spatial Patterns of Bacterial Virulence Factors and Antibiotic Resistance Genes in 19 Wastewater Treatment Plants.

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    Bing Zhang

    Full Text Available Bacterial pathogenicity and antibiotic resistance are of concern for environmental safety and public health. Accumulating evidence suggests that wastewater treatment plants (WWTPs are as an important sink and source of pathogens and antibiotic resistance genes (ARGs. Virulence genes (encoding virulence factors are good indicators for bacterial pathogenic potentials. To achieve a comprehensive understanding of bacterial pathogenic potentials and antibiotic resistance in WWTPs, bacterial virulence genes and ARGs in 19 WWTPs covering a majority of latitudinal zones of China were surveyed by using GeoChip 4.2. A total of 1610 genes covering 13 virulence factors and 1903 genes belonging to 11 ARG families were detected respectively. The bacterial virulence genes exhibited significant spatial distribution patterns of a latitudinal biodiversity gradient and a distance-decay relationship across China. Moreover, virulence genes tended to coexist with ARGs as shown by their strongly positive associations. In addition, key environmental factors shaping the overall virulence gene structure were identified. This study profiles the occurrence, composition and distribution of virulence genes and ARGs in current WWTPs in China, and uncovers spatial patterns and important environmental variables shaping their structure, which may provide the basis for further studies of bacterial virulence factors and antibiotic resistance in WWTPs.

  11. Association between antimicrobial resistance and virulence genes in Escherichia coli obtained from blood and faeces

    DEFF Research Database (Denmark)

    Bagger-Skjøt, Line; Sandvang, Dorthe; Frimodt-Møller, Niels

    2007-01-01

    Escherichia coli isolates obtained from faeces (n = 85) and blood (n = 123) were susceptibility tested against 17 antimicrobial agents and the presence of 9 virulence genes was determined by PCR. Positive associations between several antimicrobial resistances and 2 VF genes (iutA and traT) were...

  12. Subinhibitory concentrations of antibiotics affect stress and virulence gene expression in Listeria monocytogenes and cause enhanced stress sensitivity but do not affect Caco‐2 cell invasion

    DEFF Research Database (Denmark)

    Knudsen, Gitte Maegaard; Holch, Anne; Gram, Lone

    2012-01-01

    with promoter fusions, 14 of 16 antibiotics induced or repressed expression of one or more stress and/or virulence genes. Despite ampicillin‐induced up‐regulation of PinlA‐lacZ expression, Caco‐2 cell invasion was not affected. Subinhibitory concentrations of ampicillin and tetracycline caused up‐ and down...

  13. Regulation of hemolysin expression and virulence of Staphylococcus aureus by a serine/threonine kinase and phosphatase.

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    Kellie Burnside

    2010-06-01

    Full Text Available Exotoxins, including the hemolysins known as the alpha (alpha and beta (beta toxins, play an important role in the pathogenesis of Staphylococcus aureus infections. A random transposon library was screened for S. aureus mutants exhibiting altered hemolysin expression compared to wild type. Transposon insertions in 72 genes resulting in increased or decreased hemolysin expression were identified. Mutations inactivating a putative cyclic di-GMP synthetase and a serine/threonine phosphatase (Stp1 were found to reduce hemolysin expression, and mutations in genes encoding a two component regulator PhoR, LysR family transcriptional regulator, purine biosynthetic enzymes and a serine/threonine kinase (Stk1 increased expression. Transcription of the hla gene encoding alpha toxin was decreased in a Deltastp1 mutant strain and increased in a Deltastk1 strain. Microarray analysis of a Deltastk1 mutant revealed increased transcription of additional exotoxins. A Deltastp1 strain is severely attenuated for virulence in mice and elicits less inflammation and IL-6 production than the Deltastk1 strain. In vivo phosphopeptide enrichment and mass spectrometric analysis revealed that threonine phosphorylated peptides corresponding to Stk1, DNA binding histone like protein (HU, serine-aspartate rich fibrinogen/bone sialoprotein binding protein (SdrE and a hypothetical protein (NWMN_1123 were present in the wild type and not in the Deltastk1 mutant. Collectively, these studies suggest that Stk1 mediated phosphorylation of HU, SrdE and NWMN_1123 affects S. aureus gene expression and virulence.

  14. Virulence of Klebsiella pneumoniae isolates harboring bla KPC-2 carbapenemase gene in a Caenorhabditis elegans model.

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    Jean-Philippe Lavigne

    Full Text Available Klebsiella pneumoniae carbapenemase (KPC is a carbapenemase increasingly reported worldwide in Enterobacteriaceae. The aim of this study was to analyze the virulence of several KPC-2-producing K. pneumoniae isolates. The studied strains were (i five KPC-2 clinical strains from different geographical origins, belonging to different ST-types and possessing plasmids of different incompatibility groups; (ii seven transformants obtained after electroporation of either these natural KPC plasmids or a recombinant plasmid harboring only the bla KPC-2 gene into reference strains K. pneumoniae ATCC10031/CIP53153; and (iii five clinical strains cured of plasmids. The virulence of K. pneumoniae isolates was evaluated in the Caenorhabditis elegans model. The clinical KPC producers and transformants were significantly less virulent (LT50: 5.5 days than K. pneumoniae reference strain (LT50: 4.3 days (p<0.01. However, the worldwide spread KPC-2 positive K. pneumoniae ST258 strains and reference strains containing plasmids extracted from K. pneumoniae ST258 strains had a higher virulence than KPC-2 strains belonging to other ST types (LT50: 5 days vs. 6 days, p<0.01. The increased virulence observed in cured strains confirmed this trend. The bla KPC-2 gene itself was not associated to increased virulence.

  15. Use of Metarhizium anisopliae Chitinase Genes for Genotyping and Virulence Characterization

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    Saliou Niassy

    2013-01-01

    Full Text Available Virulence is the primary factor used for selection of entomopathogenic fungi (EPF for development as biopesticides. To understand the genetic mechanisms underlying differences in virulence of fungal isolates on various arthropod pests, we compared the chitinase genes, chi2 and chi4, of 8 isolates of Metarhizium anisopliae. The clustering of the isolates showed various groups depending on their virulence. However, the analysis of their chitinase DNA sequences chi2 and chi4 did not reveal major divergences. Although their protein translates have been implicated in fungal virulence, the predicted protein structure of chi2 was identical for all isolates. Despite the critical role of chitin digestion in fungal infection, we conclude that chi2 and chi4 genes cannot serve as molecular markers to characterize observed variations in virulence among M. anisopliae isolates as previously suggested. Nevertheless, processes controlling the efficient upregulation of chitinase expression might be responsible for different virulence characteristics. Further studies using comparative “in vitro” chitin digestion techniques would be more appropriate to compare the quality and the quantity of chitinase production between fungal isolates.

  16. A Dual Role for the Bacillus anthracis Master Virulence Regulator AtxA: Control of Sporulation and Anthrax Toxin Production.

    Science.gov (United States)

    Dale, Jennifer L; Raynor, Malik J; Ty, Maureen C; Hadjifrangiskou, Maria; Koehler, Theresa M

    2018-01-01

    Bacillus anthracis is an endemic soil bacterium that exhibits two different lifestyles. In the soil environment, B. anthracis undergoes a cycle of saprophytic growth, sporulation, and germination. In mammalian hosts, the pathogenic lifestyle of B. anthracis is spore germination followed by vegetative cell replication, but cells do not sporulate. During infection, and in specific culture conditions, transcription of the structural genes for the anthrax toxin proteins and the biosynthetic operon for capsule synthesis is positively controlled by the regulatory protein AtxA. A critical role for the atxA gene in B. anthracis virulence has been established. Here we report an inverse relationship between toxin production and sporulation that is linked to AtxA levels. During culture in conditions favoring sporulation, B. anthracis produces little to no AtxA. When B. anthracis is cultured in conditions favoring toxin gene expression, AtxA is expressed at relatively high levels and sporulation rate and efficiency are reduced. We found that a mutation within the atxA promoter region resulting in AtxA over-expression leads to a marked sporulation defect. The sporulation phenotype of the mutant is dependent upon pXO2-0075 , an atxA -regulated open reading frame located on virulence plasmid pXO2. The predicted amino acid sequence of the pXO2-0075 protein has similarity to the sensor domain of sporulation sensor histidine kinases. It was shown previously that pXO2-0075 overexpression suppresses sporulation. We have designated pXO2-0075 " skiA " for "sporulation kinase inhibitor." Our results indicate that in addition to serving as a positive regulator of virulence gene expression, AtxA modulates B. anthracis development.

  17. Virulence Factors and Antibiotic Susceptibility of Staphylococcus aureus Isolates in Ready-to-Eat Foods: Detection of S. aureus Contamination and a High Prevalence of Virulence Genes

    Directory of Open Access Journals (Sweden)

    Suat Moi Puah

    2016-02-01

    Full Text Available Staphylococcus aureus is one of the leading causes of food poisoning. Its pathogenicity results from the possession of virulence genes that produce different toxins which result in self-limiting to severe illness often requiring hospitalization. In this study of 200 sushi and sashimi samples, S. aureus contamination was confirmed in 26% of the food samples. The S. aureus isolates were further characterized for virulence genes and antibiotic susceptibility. A high incidence of virulence genes was identified in 96.2% of the isolates and 20 different virulence gene profiles were confirmed. DNA amplification showed that 30.8% (16/52 of the S. aureus carried at least one SE gene which causes staphylococcal food poisoning. The most common enterotoxin gene was seg (11.5% and the egc cluster was detected in 5.8% of the isolates. A combination of hla and hld was the most prevalent coexistence virulence genes and accounted for 59.6% of all isolates. Antibiotic resistance studies showed tetracycline resistance to be the most common at 28.8% while multi-drug resistance was found to be low at 3.8%. In conclusion, the high rate of S. aureus in the sampled sushi and sashimi indicates the need for food safety guidelines.

  18. Detection of virulence genes and the phylogenetic groups of Escherichia coli isolated from dogs in Brazil

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    Fernanda Morcatti Coura

    2018-02-01

    Full Text Available ABSTRACT: This study identified the virulence genes, pathovars, and phylogenetic groups of Escherichia coli strains obtained from the feces of dogs with and without diarrhea. Virulence genes and phylogenetic group identification were studied using polymerase chain reaction. Thirty-seven E. coli isolates were positive for at least one virulence factor gene. Twenty-one (57.8% of the positive isolates were isolated from diarrheal feces and sixteen (43.2% were from the feces of non-diarrheic dogs. Enteropathogenic E. coli (EPEC were the most frequently (62.2% detected pathovar in dog feces and were mainly from phylogroup B1 and E. Necrotoxigenic E. coli were detected in 16.2% of the virulence-positive isolates and these contained the cytotoxic necrotizing factor 1 (cnf1 gene and were classified into phylogroups B2 and D. All E. coli strains were negative for the presence of enterotoxigenic E. coli (ETEC enterotoxin genes, but four strains were positive for ETEC-related fimbriae 987P and F18. Two isolates were Shiga toxin-producing E. coli strains and contained the toxin genesStx2 or Stx2e, both from phylogroup B1. Our data showed that EPEC was the most frequent pathovar and B1 and E were the most common phylogroups detected in E. coli isolated from the feces of diarrheic and non-diarrheic dogs.

  19. Detection of virulence genes in Uropathogenic E. coli (UPEC strains by Multiplex-PCR method

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    Javad Mohammadi

    2017-06-01

    Full Text Available Background & Objectives: Urinary tract infection caused by E. coli is one of the most common illnesses in all age groups worldwide. Presence of virulence genes is a key factor in bacterial pathogens in uroepithelial cells. The present study was performed to detect iha, iroN, ompT genes in the Uropathogenic E.coli isolates from clinical samples using multiplex-PCR method in Kerman. Materials & Methods: In this descriptive cross-sectional study, 200 samples of patients with urinary tract infections in Kerman hospitals were collected. After biochemical and microbiological tests, all strains were tested with regard to the presence of iha, iroN, and ompT genes using multiplex-PCR method. Results: The results of Multiplex-PCR showed that all specimens had one, two, or three virulence genes simultaneously. The highest and lowest frequency distribution of genes was related to iha (56.7% and iroN (20% respectively. Conclusion: According to the prevalence of urinary tract infection in the community and distribution of resistance and virulence factors, the fast and accurate detection of the strains and virulence genes is necessary

  20. Selected lactic acid-producing bacterial isolates with the capacity to reduce Salmonella translocation and virulence gene expression in chickens.

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    Xiaojian Yang

    Full Text Available BACKGROUND: Probiotics have been used to control Salmonella colonization/infection in chickens. Yet the mechanisms of probiotic effects are not fully understood. This study has characterized our previously-selected lactic acid-producing bacterial (LAB isolates for controlling Salmonella infection in chickens, particularly the mechanism underlying the control. METHODOLOGY/PRINCIPAL FINDINGS: In vitro studies were conducted to characterize 14 LAB isolates for their tolerance to low pH (2.0 and high bile salt (0.3-1.5% and susceptibility to antibiotics. Three chicken infection trials were subsequently carried out to evaluate four of the isolates for reducing the burden of Salmonella enterica serovar Typhimurium in the broiler cecum. Chicks were gavaged with LAB cultures (10(6-7 CFU/chick or phosphate-buffered saline (PBS at 1 day of age followed by Salmonella challenge (10(4 CFU/chick next day. Samples of cecal digesta, spleen, and liver were examined for Salmonella counts on days 1, 3, or 4 post-challenge. Salmonella in the cecum from Trial 3 was also assessed for the expression of ten virulence genes located in its pathogenicity island-1 (SPI-1. These genes play a role in Salmonella intestinal invasion. Tested LAB isolates (individuals or mixed cultures were unable to lower Salmonella burden in the chicken cecum, but able to attenuate Salmonella infection in the spleen and liver. The LAB treatments also reduced almost all SPI-1 virulence gene expression (9 out of 10 in the chicken cecum, particularly at the low dose. In vitro treatment with the extracellular culture fluid from a LAB culture also down-regulated most SPI-1 virulence gene expression. CONCLUSIONS/SIGNIFICANCE: The possible correlation between attenuation of Salmonella infection in the chicken spleen and liver and reduction of Salmonella SPI-1 virulence gene expression in the chicken cecum by LAB isolates is a new observation. Suppression of Salmonella virulence gene expression in

  1. Antibiotic resistance profile and virulence genes of uropathogenic Escherichia coli isolates in relation to phylogeny.

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    Adib, N; Ghanbarpour, R; Solatzadeh, H; Alizade, H

    2014-03-01

    Escherichia coli (E. coli) strains are the major cause of urinary tract infections (UTI) and belong to the large group of extra-intestinal pathogenic E. coli. The purposes of this study were to determine the antibiotic resistance profile, virulence genes and phylogenetic background of E. coli isolates from UTI cases. A total of 137 E. coli isolates were obtained from UTI samples. The antimicrobial susceptibility of confirmed isolates was determined by disk diffusion method against eight antibiotics. The isolates were examined to determine the presence and prevalence of selected virulence genes including iucD, sfa/focDE, papEF and hly. ECOR phylo-groups of isolates were determined by detection of yjaA and chuA genes and fragment TspE4.C2. The antibiogram results showed that 71% of the isolates were resistant to cefazolin, 60.42% to co-trimoxazole, 54.16% to nalidixic acid, 36.45% to gentamicin, 29.18% to ciprofloxacin, 14.58% to cefepime, 6.25% to nitrofurantoin and 0.00% to imipenem. Twenty-two antibiotic resistance patterns were observed among the isolates. Virulence genotyping of isolates revealed that 58.39% isolates had at least one of the four virulence genes. The iucD gene was the most prevalent gene (43.06%). The other genes including sfa/focDE, papEF and hly genes were detected in 35.76%, 18.97% and 2.18% isolates, respectively. Nine combination patterns of the virulence genes were detected in isolates. Phylotyping of 137 isolates revealed that the isolates fell into A (45.99%), B1 (13.14%), B2 (19.71%) and D (21.16%) groups. Phylotyping of multidrug resistant isolates indicated that these isolates are mostly in A (60.34%) and D (20.38%) groups. In conclusion, the isolates that possessed the iucD, sfa/focDE, papEF and hly virulence genes mostly belonged to A and B2 groups, whereas antibiotic resistant isolates were in groups A and D. Escherichia coli strains carrying virulence factors and antibiotic resistance are distributed in specific phylogenetic

  2. Regulation of eucaryotic gene expression

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    Brent, R.; Ptashne, M.S

    1989-05-23

    This patent describes a method of regulating the expression of a gene in a eucaryotic cell. The method consists of: providing in the eucaryotic cell, a peptide, derived from or substantially similar to a peptide of a procaryotic cell able to bind to DNA upstream from or within the gene, the amount of the peptide being sufficient to bind to the gene and thereby control expression of the gene.

  3. Steps toward broad-spectrum therapeutics: discovering virulence-associated genes present in diverse human pathogens

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    de Rochefort Anna

    2009-10-01

    Full Text Available Abstract Background New and improved antimicrobial countermeasures are urgently needed to counteract increased resistance to existing antimicrobial treatments and to combat currently untreatable or new emerging infectious diseases. We demonstrate that computational comparative genomics, together with experimental screening, can identify potential generic (i.e., conserved across multiple pathogen species and novel virulence-associated genes that may serve as targets for broad-spectrum countermeasures. Results Using phylogenetic profiles of protein clusters from completed microbial genome sequences, we identified seventeen protein candidates that are common to diverse human pathogens and absent or uncommon in non-pathogens. Mutants of 13 of these candidates were successfully generated in Yersinia pseudotuberculosis and the potential role of the proteins in virulence was assayed in an animal model. Six candidate proteins are suggested to be involved in the virulence of Y. pseudotuberculosis, none of which have previously been implicated in the virulence of Y. pseudotuberculosis and three have no record of involvement in the virulence of any bacteria. Conclusion This work demonstrates a strategy for the identification of potential virulence factors that are conserved across a number of human pathogenic bacterial species, confirming the usefulness of this tool.

  4. Co-regulation of Iron Metabolism and Virulence Associated Functions by Iron and XibR, a Novel Iron Binding Transcription Factor, in the Plant Pathogen Xanthomonas

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    Pandey, Sheo Shankar; Patnana, Pradeep Kumar; Lomada, Santosh Kumar; Tomar, Archana; Chatterjee, Subhadeep

    2016-01-01

    Abilities of bacterial pathogens to adapt to the iron limitation present in hosts is critical to their virulence. Bacterial pathogens have evolved diverse strategies to coordinately regulate iron metabolism and virulence associated functions to maintain iron homeostasis in response to changing iron availability in the environment. In many bacteria the ferric uptake regulator (Fur) functions as transcription factor that utilize ferrous form of iron as cofactor to regulate transcription of iron metabolism and many cellular functions. However, mechanisms of fine-tuning and coordinated regulation of virulence associated function beyond iron and Fur-Fe2+ remain undefined. In this study, we show that a novel transcriptional regulator XibR (named X anthomonas iron binding regulator) of the NtrC family, is required for fine-tuning and co-coordinately regulating the expression of several iron regulated genes and virulence associated functions in phytopathogen Xanthomonas campestris pv. campestris (Xcc). Genome wide expression analysis of iron-starvation stimulon and XibR regulon, GUS assays, genetic and functional studies of xibR mutant revealed that XibR positively regulates functions involved in iron storage and uptake, chemotaxis, motility and negatively regulates siderophore production, in response to iron. Furthermore, chromatin immunoprecipitation followed by quantitative real-time PCR indicated that iron promoted binding of the XibR to the upstream regulatory sequence of operon’s involved in chemotaxis and motility. Circular dichroism spectroscopy showed that purified XibR bound ferric form of iron. Electrophoretic mobility shift assay revealed that iron positively affected the binding of XibR to the upstream regulatory sequences of the target virulence genes, an effect that was reversed by ferric iron chelator deferoxamine. Taken together, these data revealed that how XibR coordinately regulates virulence associated and iron metabolism functions in Xanthomonads in

  5. Co-regulation of Iron Metabolism and Virulence Associated Functions by Iron and XibR, a Novel Iron Binding Transcription Factor, in the Plant Pathogen Xanthomonas.

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    Sheo Shankar Pandey

    2016-11-01

    Full Text Available Abilities of bacterial pathogens to adapt to the iron limitation present in hosts is critical to their virulence. Bacterial pathogens have evolved diverse strategies to coordinately regulate iron metabolism and virulence associated functions to maintain iron homeostasis in response to changing iron availability in the environment. In many bacteria the ferric uptake regulator (Fur functions as transcription factor that utilize ferrous form of iron as cofactor to regulate transcription of iron metabolism and many cellular functions. However, mechanisms of fine-tuning and coordinated regulation of virulence associated function beyond iron and Fur-Fe2+ remain undefined. In this study, we show that a novel transcriptional regulator XibR (named Xanthomonas iron binding regulator of the NtrC family, is required for fine-tuning and co-coordinately regulating the expression of several iron regulated genes and virulence associated functions in phytopathogen Xanthomonas campestris pv. campestris (Xcc. Genome wide expression analysis of iron-starvation stimulon and XibR regulon, GUS assays, genetic and functional studies of xibR mutant revealed that XibR positively regulates functions involved in iron storage and uptake, chemotaxis, motility and negatively regulates siderophore production, in response to iron. Furthermore, chromatin immunoprecipitation followed by quantitative real-time PCR indicated that iron promoted binding of the XibR to the upstream regulatory sequence of operon's involved in chemotaxis and motility. Circular dichroism spectroscopy showed that purified XibR bound ferric form of iron. Electrophoretic mobility shift assay revealed that iron positively affected the binding of XibR to the upstream regulatory sequences of the target virulence genes, an effect that was reversed by ferric iron chelator deferoxamine. Taken together, these data revealed that how XibR coordinately regulates virulence associated and iron metabolism functions in

  6. Accessory genes confer a high replication rate to virulent feline immunodeficiency virus.

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    Troyer, Ryan M; Thompson, Jesse; Elder, John H; VandeWoude, Sue

    2013-07-01

    Feline immunodeficiency virus (FIV) is a lentivirus that causes AIDS in domestic cats, similar to human immunodeficiency virus (HIV)/AIDS in humans. The FIV accessory protein Vif abrogates the inhibition of infection by cat APOBEC3 restriction factors. FIV also encodes a multifunctional OrfA accessory protein that has characteristics similar to HIV Tat, Vpu, Vpr, and Nef. To examine the role of vif and orfA accessory genes in FIV replication and pathogenicity, we generated chimeras between two FIV molecular clones with divergent disease potentials: a highly pathogenic isolate that replicates rapidly in vitro and is associated with significant immunopathology in vivo, FIV-C36 (referred to here as high-virulence FIV [HV-FIV]), and a less-pathogenic strain, FIV-PPR (referred to here as low-virulence FIV [LV-FIV]). Using PCR-driven overlap extension, we produced viruses in which vif, orfA, or both genes from virulent HV-FIV replaced equivalent genes in LV-FIV. The generation of these chimeras is more straightforward in FIV than in primate lentiviruses, since FIV accessory gene open reading frames have very little overlap with other genes. All three chimeric viruses exhibited increased replication kinetics in vitro compared to the replication kinetics of LV-FIV. Chimeras containing HV-Vif or Vif/OrfA had replication rates equivalent to those of the virulent HV-FIV parental virus. Furthermore, small interfering RNA knockdown of feline APOBEC3 genes resulted in equalization of replication rates between LV-FIV and LV-FIV encoding HV-FIV Vif. These findings demonstrate that Vif-APOBEC interactions play a key role in controlling the replication and pathogenicity of this immunodeficiency-inducing virus in its native host species and that accessory genes act as mediators of lentiviral strain-specific virulence.

  7. The interacting Cra and KdpE regulators are involved in the expression of multiple virulence factors in enterohemorrhagic Escherichia coli.

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    Njoroge, Jacqueline W; Gruber, Charley; Sperandio, Vanessa

    2013-06-01

    The human pathogen enterohemorrhagic Escherichia coli (EHEC) O157:H7 codes for two interacting DNA binding proteins, Cra and KdpE, that coregulate expression of the locus of enterocyte effacement (LEE) genes in a metabolite-dependent manner. Cra is a transcription factor that uses fluctuations in the concentration of carbon metabolism intermediates to positively regulate virulence of EHEC. KdpE is a response regulator that activates the transcription of homeostasis genes in response to salt-induced osmolarity and virulence genes in response to changes in metabolite concentrations. Here, we probed the transcriptional profiles of the Δcra, ΔkdpE, and Δcra ΔkdpE mutant strains and show that Cra and KdpE share several targets besides the LEE, but both Cra and KdpE also have independent targets. Several genes within O-islands (genomic islands present in EHEC but absent from E. coli K-12), such as Z0639, Z0640, Z3388, Z4267, and espFu (encoding an effector necessary for formation of attaching and effacing lesions on epithelial cells), were directly regulated by both Cra and KdpE, while Z2077 was only regulated by Cra. These studies identified and confirmed new direct targets for Cra and KdpE that included putative virulence factors as well as characterized virulence factors, such as EspFu and EspG. These results map out the role of the two interacting regulators, Cra and KdpE, in EHEC pathogenesis and global gene regulation.

  8. Virulence properties of methicillin-susceptible Staphylococcus aureus food isolates encoding Panton-Valentine Leukocidin gene.

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    Sudagidan, Mert; Aydin, Ali

    2010-04-15

    In this study, three Panton-Valentine Leukocidin gene carrying methicillin-susceptible Staphylococcus aureus (MSSA) strains (M1-AAG42B, PY30C-b and YF1B-b) were isolated from different food samples in Kesan-Edirne, Turkey. These strains were characterized on the basis of MLST type, spa type, virulence factor gene contents, antibiotic susceptibilities against 21 antibiotics and biofilm formation. The genetic relatedness of the strains was determined by PFGE. In addition, the complete gene sequences of lukS-PV and lukF-PV were also investigated. All strains were found to be susceptible to tested antibiotics and they were mecA negative. Three strains showed the same PFGE band pattern, ST152 clonal type and t355 spa type. In the detection of virulence factor genes, sea, seb, sec, sed, see, seg, seh, sei, sej, sek, sel, sem, sen, seo, sep, seq, seu, eta, etb, set1, geh and tst genes were not detected. All strains showed the positive results for alpha- and beta-haemolysin genes (hla and hlb), protease encoding genes (sspA, sspB and aur), lukE and lukD leukocidin genes (lukED). The strains were found to be non-biofilm formers. By this study, the virulence properties of the strains were described and this is one of the first reports regarding PVL-positive MSSA strains from food. (c) 2010 Elsevier B.V. All rights reserved.

  9. Identification of a novel gene in ROD9 island of Salmonella Enteritidis involved in the alteration of virulence-associated genes expression.

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    Das, Susmita; Ray, Shilpa; Ryan, Daniel; Sahu, Bikash; Suar, Mrutyunjay

    2018-01-01

    Salmonella enterica subsp. I serovar Enteritidis (S. Enteritidis), one of the causative agents for non-typhoidal gastrointestinal diseases in humans is an intracellular bacterium and mechanism for its invasion into host cells is critical to cause infection. The virulence of the pathogen is explained by the expression of genes located on its pathogenicity islands, mostly encoded under SPI-1 and SPI-2. However, S. Typhimurium SL1344, despite sharing ∼98% of its genome with S. Enteritidis P125109, lacks few regions of differences (ROD) that are hypothesized to impart virulence potential to S. Enteritidis. In this study, we created different mutants in the ROD9 island of S. Enteritidis, also referred as SPI-19 and identified a novel locus, SEN1005, encoding a hypothetical protein that is involved in its pathogenesis. ΔSEN1005 displayed significantly reduced entry into cultured epithelial cells as well as uptake by macrophages and failed to cause acute colitis in C57BL/6 mice at day 3 post-infection (p.i.). Additionally, the global transcriptome analysis revealed a highly repressed SPI-1 and other down-regulated genes responsible for flagellar assembly, chemotaxis and motility in the mutant which correlated with decreased invasion and abated inflammation as compared to the wild-type. Therefore, our findings revealed that ΔSEN1005 was attenuated in vitro as well as in vivo and we propose this hypothetical protein to play a role in altering the expression of genes involved in Salmonella virulence.

  10. Multiplex reverse transcription polymerase chain reaction to study the expression of virulence and stress response genes in Staphylococcus aureus.

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    Shrihari, Rohinishree Yadahalli; Singh, Negi Pradeep

    2012-02-01

    Staphylococcus aureus survives well in different stress conditions. The ability of this organism to adapt to various stresses is the result of a complex regulatory response, which is attributed to regulation of multiple genes. The aims of the present study were (1) to develop a multiplex PCR for the detection of genes which are involved in stress adaptation (asp23, dnaK, and groEL); alternative sigma factor (sigB) and virulence determination (entB and spa) and (2) to study the expression of these genes during stress conditions for S. aureus culture collection strains (FRI 722 and ATCC 6538) and S. aureus food isolates at mRNA level using multiplex reverse transcription polymerase chain reaction (RT-PCR). During heat shock treatment groEL, dnaK, asp23, sodA, entB, spa, and sigB genes were up regulated up to 2.58, 2.07, 2.76, 2.55, 3.55, 2.71, and 2.62- folds, respectively, whereas in acid shock treatment, sodA and groEL were up regulated; dnaK was downregulated; and entB and sigB genes were not expressed in food isolates. Multiplex PCR assay standardized in this study offers an inexpensive alternative to uniplex PCR for detection of various virulence and stress response genes. This study is relevant to rapid and accurate detection of potential pathogenic S. aureus in foods. © 2012 Institute of Food Technologists®

  11. Virulence Genes Profile of Multidrug Resistant Pseudomonas aeruginosa Isolated from Iranian Children with UTIs

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    Zohreh Heidary

    2016-04-01

    Full Text Available Virulent and resistant strains Pseudomonas aeruginosa (P. aeruginosa is one of the most important cause of UTIs in pediatrics. The present study was carried to investigate the frequency of virulence factors in the multi-drug resistant strains of P. aeruginosa isolated from pediatrics hospitalized due to the UTIs. One - hundred and forty three urine samples were collected from pediatric patients suffered from UTIs. Samples were cultured and those that were P. aeruginosa positive were analyzed for the presence of putative virulence genes. Seventy one out of 143 samples (49.65% were positive for P. aeruginosa. Monthly, sex and age-dependent prevalence were seen for P. aeruginosa. Bacterial strains had the highest levels of resistance against ampicillin (95.77%, gentamicin (92.95% and ciprofloxacin (81.69%. Of 71 P. aeruginosa isolates, 12 strains were resistant to more than 9 antibiotics (16.90%. The most commonly detected virulence factors in the cases of urethral infections were exoU and plcH while those of pyelonephritis and cystitis were were exoS and lasB. Our findings should raise awareness about antibiotic resistance in hospitalized pediatrics with UTIs in Iran. Clinicians should exercise caution in prescribing antibiotics, especially in cases of UTIs. Such information can help in identifying these virulence genes as useful diagnostic markers for clinical P. aeruginosa strains isolated from UTIs.

  12. Natural mutations in a Staphylococcus aureus virulence regulator attenuate cytotoxicity but permit bacteremia and abscess formation

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    Das, Sudip; Lindemann, Claudia; Young, Bernadette C.; Muller, Julius; Österreich, Babett; Ternette, Nicola; Winkler, Ann-Cathrin; Paprotka, Kerstin; Reinhardt, Richard; Allen, Elizabeth; Flaxman, Amy; Yamaguchi, Yuko; Rollier, Christine S.; van Diemen, Pauline; Blättner, Sebastian; Remmele, Christian W.; Selle, Martina; Dittrich, Marcus; Müller, Tobias; Vogel, Jörg; Ohlsen, Knut; Crook, Derrick W.; Massey, Ruth; Wilson, Daniel J.; Rudel, Thomas; Wyllie, David H.; Fraunholz, Martin J.

    2016-01-01

    Staphylococcus aureus is a major bacterial pathogen, which causes severe blood and tissue infections that frequently emerge by autoinfection with asymptomatically carried nose and skin populations. However, recent studies report that bloodstream isolates differ systematically from those found in the nose and skin, exhibiting reduced toxicity toward leukocytes. In two patients, an attenuated toxicity bloodstream infection evolved from an asymptomatically carried high-toxicity nasal strain by loss-of-function mutations in the gene encoding the transcription factor repressor of surface proteins (rsp). Here, we report that rsp knockout mutants lead to global transcriptional and proteomic reprofiling, and they exhibit the greatest signal in a genome-wide screen for genes influencing S. aureus survival in human cells. This effect is likely to be mediated in part via SSR42, a long-noncoding RNA. We show that rsp controls SSR42 expression, is induced by hydrogen peroxide, and is required for normal cytotoxicity and hemolytic activity. Rsp inactivation in laboratory- and bacteremia-derived mutants attenuates toxin production, but up-regulates other immune subversion proteins and reduces lethality during experimental infection. Crucially, inactivation of rsp preserves bacterial dissemination, because it affects neither formation of deep abscesses in mice nor survival in human blood. Thus, we have identified a spontaneously evolving, attenuated-cytotoxicity, nonhemolytic S. aureus phenotype, controlled by a pleiotropic transcriptional regulator/noncoding RNA virulence regulatory system, capable of causing S. aureus bloodstream infections. Such a phenotype could promote deep infection with limited early clinical manifestations, raising concerns that bacterial evolution within the human body may contribute to severe infection. PMID:27185949

  13. Wide Distribution of Virulence Genes among Enterococcus faecium and Enterococcus faecalis Clinical Isolates

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    Sara Soheili

    2014-01-01

    Full Text Available Enterococcus, a Gram-positive facultative anaerobic cocci belonging to the lactic acid bacteria of the phylum Firmicutes, is known to be able to resist a wide range of hostile conditions such as different pH levels, high concentration of NaCl (6.5%, and the extended temperatures between 5°C and 65°C. Despite being the third most common nosocomial pathogen, our understanding on its virulence factors is still poorly understood. The current study was aimed to determine the prevalence of different virulence genes in Enterococcus faecalis and Enterococcus faecium. For this purpose, 79 clinical isolates of Malaysian enterococci were evaluated for the presence of virulence genes. pilB, fms8, efaAfm, and sgrA genes are prevalent in all clinical isolates. In conclusion, the pathogenicity of E. faecalis and E. faecium could be associated with different virulence factors and these genes are widely distributed among the enterococcal species.

  14. Hemolytic porcine intestinal Escherichia coli without virulence-associated genes typical of intestinal pathogenic E. coli.

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    Schierack, Peter; Weinreich, Joerg; Ewers, Christa; Tachu, Babila; Nicholson, Bryon; Barth, Stefanie

    2011-12-01

    Testing 1,666 fecal or intestinal samples from healthy and diarrheic pigs, we obtained hemolytic Escherichia coli isolates from 593 samples. Focusing on hemolytic E. coli isolates without virulence-associated genes (VAGs) typical for enteropathogens, we found that such isolates carried a broad variety of VAGs typical for extraintestinal pathogenic E. coli.

  15. Carbapenem-resistant Pseudomonas aeruginosa: association with virulence genes and biofilm formation

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    Iara Rossi Gonçalves

    Full Text Available Abstract Pseudomonas aeruginosa is an opportunistic pathogen that causes frequently nosocomial infections, currently becoming more difficult to treat due to the various resistance mechanisms and different virulence factors. The purpose of this study was to determine the risk factors independently associated with the development of bacteremia by carbapenem-resistant P. aeruginosa, the frequency of virulence genes in metallo-β-lactamases producers and to evaluate their ability to produce biofilm. We conducted a case–control study in the Uberlândia Federal University – Hospital Clinic, Brazil. Polymerase Chain Reaction was performed for metallo-β-lactamases and virulence genes. Adhesion and biofilm assays were done by quantitative tests. Among the 157 strains analyzed, 73.9% were multidrug-resistant, 43.9% were resistant to carbapenems, 16.1% were phenotypically positive for metallo-β-lactamases, and of these, 10.7% were positive for blaSPM gene and 5.3% positive for blaVIM. The multivariable analysis showed that mechanical ventilation, enteral/nasogastric tubes, primary bacteremia with unknown focus, and inappropriate therapy were independent risk factors associated with bacteremia. All tested strains were characterized as strongly biofilm producers. A higher mortality was found among patients with bacteremia by carbapenem-resistant P. aeruginosa strains, associated independently with extrinsic risk factors, however it was not evident the association with the presence of virulence and metallo-β-lactamases genes.

  16. Comparative genome analysis of 24 bovine-associated Staphylococcus isolates with special focus on the putative virulence genes

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    Åvall-Jääskeläinen, Silja; Paulin, Lars; Blom, Jochen

    2018-01-01

    Non-aureus staphylococci (NAS) are most commonly isolated from subclinical mastitis. Different NAS species may, however, have diverse effects on the inflammatory response in the udder. We determined the genome sequences of 20 staphylococcal isolates from clinical or subclinical bovine mastitis, belonging to the NAS species Staphylococcus agnetis, S. chromogenes, and S. simulans, and focused on the putative virulence factor genes present in the genomes. For comparison we used our previously published genome sequences of four S. aureus isolates from bovine mastitis. The pan-genome and core genomes of the non-aureus isolates were characterized. After that, putative virulence factor orthologues were searched in silico. We compared the presence of putative virulence factors in the NAS species and S. aureus and evaluated the potential association between bacterial genotype and type of mastitis (clinical vs. subclinical). The NAS isolates had much less virulence gene orthologues than the S. aureus isolates. One third of the virulence genes were detected only in S. aureus. About 100 virulence genes were present in all S. aureus isolates, compared to about 40 to 50 in each NAS isolate. S. simulans differed the most. Several of the virulence genes detected among NAS were harbored only by S. simulans, but it also lacked a number of genes present both in S. agnetis and S. chromogenes. The type of mastitis was not associated with any specific virulence gene profile. It seems that the virulence gene profiles or cumulative number of different virulence genes are not directly associated with the type of mastitis (clinical or subclinical), indicating that host derived factors such as the immune status play a pivotal role in the manifestation of mastitis. PMID:29610707

  17. Comparative genome analysis of 24 bovine-associated Staphylococcus isolates with special focus on the putative virulence genes

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    Silja Åvall-Jääskeläinen

    2018-03-01

    Full Text Available Non-aureus staphylococci (NAS are most commonly isolated from subclinical mastitis. Different NAS species may, however, have diverse effects on the inflammatory response in the udder. We determined the genome sequences of 20 staphylococcal isolates from clinical or subclinical bovine mastitis, belonging to the NAS species Staphylococcus agnetis, S. chromogenes, and S. simulans, and focused on the putative virulence factor genes present in the genomes. For comparison we used our previously published genome sequences of four S. aureus isolates from bovine mastitis. The pan-genome and core genomes of the non-aureus isolates were characterized. After that, putative virulence factor orthologues were searched in silico. We compared the presence of putative virulence factors in the NAS species and S. aureus and evaluated the potential association between bacterial genotype and type of mastitis (clinical vs. subclinical. The NAS isolates had much less virulence gene orthologues than the S. aureus isolates. One third of the virulence genes were detected only in S. aureus. About 100 virulence genes were present in all S. aureus isolates, compared to about 40 to 50 in each NAS isolate. S. simulans differed the most. Several of the virulence genes detected among NAS were harbored only by S. simulans, but it also lacked a number of genes present both in S. agnetis and S. chromogenes. The type of mastitis was not associated with any specific virulence gene profile. It seems that the virulence gene profiles or cumulative number of different virulence genes are not directly associated with the type of mastitis (clinical or subclinical, indicating that host derived factors such as the immune status play a pivotal role in the manifestation of mastitis.

  18. Mycobacterium tuberculosis nuoG is a virulence gene that inhibits apoptosis of infected host cells.

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    Kamalakannan Velmurugan

    2007-07-01

    Full Text Available The survival and persistence of Mycobacterium tuberculosis depends on its capacity to manipulate multiple host defense pathways, including the ability to actively inhibit the death by apoptosis of infected host cells. The genetic basis for this anti-apoptotic activity and its implication for mycobacterial virulence have not been demonstrated or elucidated. Using a novel gain-of-function genetic screen, we demonstrated that inhibition of infection-induced apoptosis of macrophages is controlled by multiple genetic loci in M. tuberculosis. Characterization of one of these loci in detail revealed that the anti-apoptosis activity was attributable to the type I NADH-dehydrogenase of M. tuberculosis, and was mainly due to the subunit of this multicomponent complex encoded by the nuoG gene. Expression of M. tuberculosis nuoG in nonpathogenic mycobacteria endowed them with the ability to inhibit apoptosis of infected human or mouse macrophages, and increased their virulence in a SCID mouse model. Conversely, deletion of nuoG in M. tuberculosis ablated its ability to inhibit macrophage apoptosis and significantly reduced its virulence in mice. These results identify a key component of the genetic basis for an important virulence trait of M. tuberculosis and support a direct causal relationship between virulence of pathogenic mycobacteria and their ability to inhibit macrophage apoptosis.

  19. Genetic diversity and virulence genes in Streptococcus uberis strains isolated from bovine mastitis

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    Rafael Ambrósio Loures

    2017-08-01

    Full Text Available Mastitis is one of the most common and costly infectious diseases in dairy cattle worldwide. This is a multifactorial illness caused by different microorganisms, including virus, yeasts, algae, parasites, and several species of bacteria. Among these bacteria, Streptococcus uberis is an important environmental pathogen that is responsible for a large range of clinical and subclinical mammary infections, especially in intensively managed herds. Despite the increasing importance of this pathogen in the etiology of bovine mastitis, data on its virulence and diversity in Brazilian dairy herds are scarce. The aims of the present study were to investigate the virulence characteristics of S. uberis isolated from bovine mastitis and to assess the molecular epidemiology of the Brazilian isolates using pulsed-field gel electrophoresis (PFGE. In this work, 46 strains of S. uberis isolated from bovine mastitis from 26 Brazilian dairy herds were evaluated regarding their genetic diversity by PFGE using with the SmaI enzyme. Additionally, the presence of the virulence genes skc and pauA, which encode plasminogen activators, and the gene sua, which encodes an adhesion molecule in mammary epithelial cells, were assessed by PCR. Our results showed a high genetic diversity in the population, displaying many different patterns in the PFGE analysis. A high proportion of strains was positive for virulence genes in the sampled population (sua [100%], pauA [91%], and skc [91%]. The high frequency of skc, pauA, and sua genes among the studied strains suggests the importance of these virulence factors, possibly helping S. uberis in the colonization of the bovine mammary gland. Surveys of the genetic and molecular characteristics of this pathogen can improve our knowledge of bacterial activity and identify molecules that have roles in the establishment of the infection. This might help in the development of more effective measures to control and prevent bovine mastitis.

  20. Characterization of Aeromonas hydrophila wound pathotypes by comparative genomic and functional analyses of virulence genes.

    Science.gov (United States)

    Grim, Christopher J; Kozlova, Elena V; Sha, Jian; Fitts, Eric C; van Lier, Christina J; Kirtley, Michelle L; Joseph, Sandeep J; Read, Timothy D; Burd, Eileen M; Tall, Ben D; Joseph, Sam W; Horneman, Amy J; Chopra, Ashok K; Shak, Joshua R

    2013-04-23

    Aeromonas hydrophila has increasingly been implicated as a virulent and antibiotic-resistant etiologic agent in various human diseases. In a previously published case report, we described a subject with a polymicrobial wound infection that included a persistent and aggressive strain of A. hydrophila (E1), as well as a more antibiotic-resistant strain of A. hydrophila (E2). To better understand the differences between pathogenic and environmental strains of A. hydrophila, we conducted comparative genomic and functional analyses of virulence-associated genes of these two wound isolates (E1 and E2), the environmental type strain A. hydrophila ATCC 7966(T), and four other isolates belonging to A. aquariorum, A. veronii, A. salmonicida, and A. caviae. Full-genome sequencing of strains E1 and E2 revealed extensive differences between the two and strain ATCC 7966(T). The more persistent wound infection strain, E1, harbored coding sequences for a cytotoxic enterotoxin (Act), a type 3 secretion system (T3SS), flagella, hemolysins, and a homolog of exotoxin A found in Pseudomonas aeruginosa. Corresponding phenotypic analyses with A. hydrophila ATCC 7966(T) and SSU as reference strains demonstrated the functionality of these virulence genes, with strain E1 displaying enhanced swimming and swarming motility, lateral flagella on electron microscopy, the presence of T3SS effector AexU, and enhanced lethality in a mouse model of Aeromonas infection. By combining sequence-based analysis and functional assays, we characterized an A. hydrophila pathotype, exemplified by strain E1, that exhibited increased virulence in a mouse model of infection, likely because of encapsulation, enhanced motility, toxin secretion, and cellular toxicity. Aeromonas hydrophila is a common aquatic bacterium that has increasingly been implicated in serious human infections. While many determinants of virulence have been identified in Aeromonas, rapid identification of pathogenic versus nonpathogenic

  1. Systems level analysis of two-component signal transduction systems in Erwinia amylovora: Role in virulence, regulation of amylovoran biosynthesis and swarming motility

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    Sundin George W

    2009-05-01

    Full Text Available Abstract Background Two-component signal transduction systems (TCSTs, consisting of a histidine kinase (HK and a response regulator (RR, represent a major paradigm for signal transduction in prokaryotes. TCSTs play critical roles in sensing and responding to environmental conditions, and in bacterial pathogenesis. Most TCSTs in Erwinia amylovora have either not been identified or have not yet been studied. Results We used a systems approach to identify TCST and related signal transduction genes in the genome of E. amylovora. Comparative genomic analysis of TCSTs indicated that E. amylovora TCSTs were closely related to those of Erwinia tasmaniensis, a saprophytic enterobacterium isolated from apple flowers, and to other enterobacteria. Forty-six TCST genes in E. amylovora including 17 sensor kinases, three hybrid kinases, 20 DNA- or ligand-binding RRs, four RRs with enzymatic output domain (EAL-GGDEF proteins, and two kinases were characterized in this study. A systematic TCST gene-knockout experiment was conducted, generating a total of 59 single-, double-, and triple-mutants. Virulence assays revealed that five of these mutants were non-pathogenic on immature pear fruits. Results from phenotypic characterization and gene expression experiments indicated that several groups of TCST systems in E. amylovora control amylovoran biosynthesis, one of two major virulence factors in E. amylovora. Both negative and positive regulators of amylovoran biosynthesis were identified, indicating a complex network may control this important feature of pathogenesis. Positive (non-motile, EnvZ/OmpR, negative (hypermotile, GrrS/GrrA, and intermediate regulators for swarming motility in E. amylovora were also identified. Conclusion Our results demonstrated that TCSTs in E. amylovora played major roles in virulence on immature pear fruit and in regulating amylovoran biosynthesis and swarming motility. This suggested presence of regulatory networks governing

  2. Differences in virulence genes and genome patterns of mastitis-associated Staphylococcus aureus among goat, cow, and human isolates in Taiwan.

    Science.gov (United States)

    Chu, Chishih; Wei, Yajiun; Chuang, Shih-Te; Yu, Changyou; Changchien, Chih-Hsuan; Su, Yaochi

    2013-03-01

    A total of 117 mastitis-associated Staphylococcus aureus isolates from cow, goat, and human patients were analyzed for differences in antibiotic susceptibility, virulence genes, and genotypes using accessory gene regulator (agr) typing, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Multidrug-resistant (MDR) S. aureus were commonly found in all sources, though they were predominantly found in human and goat isolates. Almost 70% of the isolates were resistant to ampicillin and penicillin. Host-associated virulence genes were identified as follows: tst, a gene encoding toxic shock syndrome toxin, was found in goat isolates; lukED and lukM, genes encoding leukocidin, found in cow isolates; lukPV, a gene encoding leukocidin, found in human isolates; and eta, a gene encoding for exfoliative toxin, found in both human and cow isolates. All four types of hemolysin, α, β, γ, and δ, were identified in human isolates, three types (α, γ, and δ), were identified in cow isolates, and two types (α and δ) were identified in goat isolates. Agr-typing determined agr1 to be the main subtype in human and cow isolates. PFGE and MLST analysis revealed the presence of diverse genotypes among the three sources. In conclusion, mastitis-associated, genetically diverse strains of MDR S. aureus differed in virulence genes among human, cow, and goat isolates.

  3. Virulence Genes of S. aureus from Dairy Cow Mastitis and Contagiousness Risk.

    Science.gov (United States)

    Magro, Giada; Biffani, Stefano; Minozzi, Giulietta; Ehricht, Ralf; Monecke, Stefan; Luini, Mario; Piccinini, Renata

    2017-06-21

    Staphylococcus aureus ( S. aureus ) is a major agent of dairy cow intramammary infections: the different prevalences of mastitis reported might be related to a combination of S. aureus virulence factors beyond host factors. The present study considered 169 isolates from different Italian dairy herds that were classified into four groups based on the prevalence of S. aureus infection at the first testing: low prevalence (LP), medium-low (MLP), medium-high (MHP) and high (HP). We aimed to correlate the presence of virulence genes with the prevalence of intramammary infections in order to develop new strategies for the control of S. aureus mastitis. Microarray data were statistically evaluated using binary logistic regression and correspondence analysis to screen the risk factors and the relationship between prevalence group and gene. The analysis showed: (1) 24 genes at significant risk of being detected in all the herds with infection prevalence >5%, including genes belonging to microbial surface components recognizing adhesive matrix molecules (MSCRAMMs), immune evasion and serine proteases; and (2) a significant correlation coefficient between the genes interacting with the host immune response and HP isolates against LP ones. These results support the hypothesis that virulence factors, in addition to cow management, could be related to strain contagiousness, offering new insights into vaccine development.

  4. Virulence Genes of S. aureus from Dairy Cow Mastitis and Contagiousness Risk

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    Giada Magro

    2017-06-01

    Full Text Available Staphylococcus aureus (S. aureus is a major agent of dairy cow intramammary infections: the different prevalences of mastitis reported might be related to a combination of S. aureus virulence factors beyond host factors. The present study considered 169 isolates from different Italian dairy herds that were classified into four groups based on the prevalence of S. aureus infection at the first testing: low prevalence (LP, medium–low (MLP, medium–high (MHP and high (HP. We aimed to correlate the presence of virulence genes with the prevalence of intramammary infections in order to develop new strategies for the control of S. aureus mastitis. Microarray data were statistically evaluated using binary logistic regression and correspondence analysis to screen the risk factors and the relationship between prevalence group and gene. The analysis showed: (1 24 genes at significant risk of being detected in all the herds with infection prevalence >5%, including genes belonging to microbial surface components recognizing adhesive matrix molecules (MSCRAMMs, immune evasion and serine proteases; and (2 a significant correlation coefficient between the genes interacting with the host immune response and HP isolates against LP ones. These results support the hypothesis that virulence factors, in addition to cow management, could be related to strain contagiousness, offering new insights into vaccine development.

  5. Microevolution of Virulence-Related Genes in Helicobacter pylori Familial Infection.

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    Yoshikazu Furuta

    Full Text Available Helicobacter pylori, a bacterial pathogen that can infect human stomach causing gastritis, ulcers and cancer, is known to have a high degree of genome/epigenome diversity as the result of mutation and recombination. The bacteria often infect in childhood and persist for the life of the host. One of the reasons of the rapid evolution of H. pylori is that it changes its genome drastically for adaptation to a new host. To investigate microevolution and adaptation of the H. pylori genome, we undertook whole genome sequencing of the same or very similar sequence type in multi-locus sequence typing (MLST with seven genes in members of the same family consisting of parents and children in Japan. Detection of nucleotide substitutions revealed likely transmission pathways involving children. Nonsynonymous (amino acid changing mutations were found in virulence-related genes (cag genes, vacA, hcpDX, tnfα, ggt, htrA and the collagenase gene, outer membrane protein (OMP genes and other cell surface-related protein genes, signal transduction genes and restriction-modification genes. We reconstructed various pathways by which H. pylori can adapt to a new human host, and our results raised the possibility that the mutational changes in virulence-related genes have a role in adaptation to a child host. Changes in restriction-modification genes might remodel the methylome and transcriptome to help adaptation. This study has provided insights into H. pylori transmission and virulence and has implications for basic research as well as clinical practice.

  6. Molecular serotyping, virulence gene profiling and pathogenicity of Streptococcus agalactiae isolated from tilapia farms in Thailand by multiplex PCR.

    Science.gov (United States)

    Kannika, K; Pisuttharachai, D; Srisapoome, P; Wongtavatchai, J; Kondo, H; Hirono, I; Unajak, S; Areechon, N

    2017-06-01

    This study aimed to biotype Streptococcus agalactiae isolated from tilapia farms in Thailand based on molecular biotyping methods and to determine the correlation between the serotype and virulence of bacteria. In addition to a biotyping (serotyping) technique based on multiplex PCR of cps genes, in this study, we developed multiplex PCR typing of Group B streptococcus (GBS) virulence genes to examine three clusters of virulence genes and their correlation with the pathogenicity of S. agalactiae. The epidemiology of S. agalactiae in Thailand was analysed to provide bacterial genetic information towards a future rational vaccine strategy for tilapia culture systems. Streptococcus agalactiae were isolated from diseased tilapia from different areas of Thailand. A total of 124 S. agalactiae isolates were identified by phenotypic analysis and confirmed by 16S rRNA PCR. Bacterial genotyping was conducted based on (i) molecular serotyping of the capsular polysaccharide (cps) gene cluster and (ii) virulence gene profiling using multiplex PCR analysis of 14 virulence genes (lmb, scpB, pavA, cspA, spb1, cyl, bca, rib, fbsA, fbsB, cfb, hylB, bac and pbp1A/ponA). Only serotypes Ia and III were found in this study; serotype Ia lacks the lmb, scpB and spb1 genes, whereas serotype III lacks only the bac gene. Virulence tests in juvenile Nile tilapia demonstrated a correlation between the pathogenicity of the bacteria and their virulence gene profile, with serotype III showing higher virulence than serotype Ia. Epidemiological analysis showed an almost equal distribution in all regions of Thailand, except serotype III was found predominantly in the southern areas. Only two serotypes of S. agalactiae were isolated from diseased tilapia in Thailand. Serotype Ia showed fewer virulence genes and lower virulence than serotype III. Both serotypes showed a similar distribution throughout Thailand. We identified two major serotypes of S. agalactiae isolates associated with the outbreak in

  7. Phosphorylation of KasB Regulates Virulence and Acid-Fastness in Mycobacterium tuberculosis

    Science.gov (United States)

    Vilchèze, Catherine; Molle, Virginie; Carrère-Kremer, Séverine; Leiba, Jade; Mourey, Lionel; Shenai, Shubhada; Baronian, Grégory; Tufariello, Joann; Hartman, Travis; Veyron-Churlet, Romain; Trivelli, Xavier; Tiwari, Sangeeta; Weinrick, Brian; Alland, David; Guérardel, Yann; Jacobs, William R.; Kremer, Laurent

    2014-01-01

    Mycobacterium tuberculosis bacilli display two signature features: acid-fast staining and the capacity to induce long-term latent infections in humans. However, the mechanisms governing these two important processes remain largely unknown. Ser/Thr phosphorylation has recently emerged as an important regulatory mechanism allowing mycobacteria to adapt their cell wall structure/composition in response to their environment. Herein, we evaluated whether phosphorylation of KasB, a crucial mycolic acid biosynthetic enzyme, could modulate acid-fast staining and virulence. Tandem mass spectrometry and site-directed mutagenesis revealed that phosphorylation of KasB occurred at Thr334 and Thr336 both in vitro and in mycobacteria. Isogenic strains of M. tuberculosis with either a deletion of the kasB gene or a kasB_T334D/T336D allele, mimicking constitutive phosphorylation of KasB, were constructed by specialized linkage transduction. Biochemical and structural analyses comparing these mutants to the parental strain revealed that both mutant strains had mycolic acids that were shortened by 4–6 carbon atoms and lacked trans-cyclopropanation. Together, these results suggested that in M. tuberculosis, phosphorylation profoundly decreases the condensing activity of KasB. Structural/modeling analyses reveal that Thr334 and Thr336 are located in the vicinity of the catalytic triad, which indicates that phosphorylation of these amino acids would result in loss of enzyme activity. Importantly, the kasB_T334D/T336D phosphomimetic and deletion alleles, in contrast to the kasB_T334A/T336A phosphoablative allele, completely lost acid-fast staining. Moreover, assessing the virulence of these strains indicated that the KasB phosphomimetic mutant was attenuated in both immunodeficient and immunocompetent mice following aerosol infection. This attenuation was characterized by the absence of lung pathology. Overall, these results highlight for the first time the role of Ser/Thr kinase

  8. Phosphorylation of KasB regulates virulence and acid-fastness in Mycobacterium tuberculosis.

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    Catherine Vilchèze

    2014-05-01

    Full Text Available Mycobacterium tuberculosis bacilli display two signature features: acid-fast staining and the capacity to induce long-term latent infections in humans. However, the mechanisms governing these two important processes remain largely unknown. Ser/Thr phosphorylation has recently emerged as an important regulatory mechanism allowing mycobacteria to adapt their cell wall structure/composition in response to their environment. Herein, we evaluated whether phosphorylation of KasB, a crucial mycolic acid biosynthetic enzyme, could modulate acid-fast staining and virulence. Tandem mass spectrometry and site-directed mutagenesis revealed that phosphorylation of KasB occurred at Thr334 and Thr336 both in vitro and in mycobacteria. Isogenic strains of M. tuberculosis with either a deletion of the kasB gene or a kasB_T334D/T336D allele, mimicking constitutive phosphorylation of KasB, were constructed by specialized linkage transduction. Biochemical and structural analyses comparing these mutants to the parental strain revealed that both mutant strains had mycolic acids that were shortened by 4-6 carbon atoms and lacked trans-cyclopropanation. Together, these results suggested that in M. tuberculosis, phosphorylation profoundly decreases the condensing activity of KasB. Structural/modeling analyses reveal that Thr334 and Thr336 are located in the vicinity of the catalytic triad, which indicates that phosphorylation of these amino acids would result in loss of enzyme activity. Importantly, the kasB_T334D/T336D phosphomimetic and deletion alleles, in contrast to the kasB_T334A/T336A phosphoablative allele, completely lost acid-fast staining. Moreover, assessing the virulence of these strains indicated that the KasB phosphomimetic mutant was attenuated in both immunodeficient and immunocompetent mice following aerosol infection. This attenuation was characterized by the absence of lung pathology. Overall, these results highlight for the first time the role of

  9. Virulence genes and subclone status as markers of experimental virulence in a murine sepsis model among Escherichia coli sequence type 131 clinical isolates from Spain.

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    Irene Merino

    Full Text Available To assess experimental virulence among sequence type 131 (ST131 Escherichia coli bloodstream isolates in relation to virulence genotype and subclone.We analysed 48 Spanish ST131 bloodstream isolates (2010 by PCR for ST131 subclone status (H30Rx, H30 non-Rx, or non-H30, virulence genes (VGs, and O-type. Then we compared these traits with virulence in a murine sepsis model, as measured by illness severity score (ISS and rapid lethality (mean ISS ≥ 4.Of the 48 study isolates, 65% were H30Rx, 21% H30 non-Rx, and 15% non-H30; 44% produced ESBLs, 98% were O25b, and 83% qualified as extraintestinal pathogenic E. coli (ExPEC. Of 49 VGs, ibeA and iss were associated significantly with non-H30 isolates, and sat, iha and malX with H30 isolates. Median VG scores differed by subclone, i.e., 12 (H30Rx, 10 (H30 non-Rx, and 11 (non-H30 (p < 0.01. Nearly 80% of isolates represented a described virotype. In mice, H30Rx and non-H30 isolates were more virulent than H30 non-Rx isolates (according to ISS [p = 0.03] and rapid lethality [p = 0.03], as were ExPEC isolates compared with non-ExPEC isolates (median ISS, 4.3 vs. 2.7: p = 0.03. In contrast, most individual VGs, VG scores, VG profiles, and virotypes were not associated with mouse virulence.ST131 subclone and ExPEC status, but not individual VGs, VG scores or profiles, or virotypes, predicted mouse virulence. Given the lower virulence of non-Rx H30 isolates, hypervirulence probably cannot explain the ST131-H30 clade's epidemic emergence.

  10. Regulation of Three Virulence Strategies of Mycobacterium tuberculosis: A Success Story

    Science.gov (United States)

    van Dam, Jesse C. J.; Martins dos Santos, Vitor A. P.

    2018-01-01

    Tuberculosis remains one of the deadliest diseases. Emergence of drug-resistant and multidrug-resistant M. tuberculosis strains makes treating tuberculosis increasingly challenging. In order to develop novel intervention strategies, detailed understanding of the molecular mechanisms behind the success of this pathogen is required. Here, we review recent literature to provide a systems level overview of the molecular and cellular components involved in divalent metal homeostasis and their role in regulating the three main virulence strategies of M. tuberculosis: immune modulation, dormancy and phagosomal rupture. We provide a visual and modular overview of these components and their regulation. Our analysis identified a single regulatory cascade for these three virulence strategies that respond to limited availability of divalent metals in the phagosome. PMID:29364195

  11. Polygalacturonase gene pgxB in Aspergillus niger is a virulence factor in apple fruit.

    Science.gov (United States)

    Liu, Cheng-Qian; Hu, Kang-Di; Li, Ting-Ting; Yang, Ying; Yang, Feng; Li, Yan-Hong; Liu, He-Ping; Chen, Xiao-Yan; Zhang, Hua

    2017-01-01

    Aspergillus niger, a saprophytic fungus, is widely distributed in soil, air and cereals, and can cause postharvest diseases in fruit. Polygalacturonase (PG) is one of the main enzymes in fungal pathogens to degrade plant cell wall. To evaluate whether the deletion of an exo-polygalacturonase gene pgxB would influence fungal pathogenicity to fruit, pgxB gene was deleted in Aspergillus niger MA 70.15 (wild type) via homologous recombination. The ΔpgxB mutant showed similar growth behavior compared with the wild type. Pectin medium induced significant higher expression of all pectinase genes in both wild type and ΔpgxB in comparison to potato dextrose agar medium. However, the ΔpgxB mutant was less virulent on apple fruits as the necrosis diameter caused by ΔpgxB mutant was significantly smaller than that of wild type. Results of quantitive-PCR showed that, in the process of infection in apple fruit, gene expressions of polygalacturonase genes pgaI, pgaII, pgaA, pgaC, pgaD and pgaE were enhanced in ΔpgxB mutant in comparison to wild type. These results prove that, despite the increased gene expression of other polygalacturonase genes in ΔpgxB mutant, the lack of pgxB gene significantly reduced the virulence of A. niger on apple fruit, suggesting that pgxB plays an important role in the infection process on the apple fruit.

  12. Antimicrobial resistance and virulence genes in enterococci from wild game meat in Spain.

    Science.gov (United States)

    Guerrero-Ramos, Emilia; Cordero, Jorge; Molina-González, Diana; Poeta, Patrícia; Igrejas, Gilberto; Alonso-Calleja, Carlos; Capita, Rosa

    2016-02-01

    A total of 55 enterococci (45 Enterococcus faecium, 7 Enterococcus faecalis, and three Enterococcus durans) isolated from the meat of wild game animals (roe deer, boar, rabbit, pheasant, and pigeon) in North-Western Spain were tested for susceptibility to 14 antimicrobials by the disc diffusion method. All strains showed a multi-resistant phenotype (resistance to between three and 10 antimicrobials). The strains exhibited high percentages of resistance to erythromycin (89.1%), tetracycline (67.3%), ciprofloxacin (92.7%), nitrofurantoin (67.3%), and quinupristin-dalfopristin (81.8%). The lowest values (9.1%) were observed for high-level resistance to gentamicin, kanamycin, and streptomycin. The average number of resistances per strain was 5.8 for E. faecium isolates, 7.9 for E. faecalis, and 5.7 for E. durans. Genes encoding antimicrobial resistance and virulence were studied by polymerase chain reaction. A total of 15 (57.7%) of the 26 vancomycin-resistant isolates harboured the vanA gene. Other resistance genes detected included vanB, erm(B) and/or erm(C), tet(L) and/or tet(M), acc(6')-aph(2″), and aph(3')-IIIa in strains resistant to vancomycin, erythromycin, tetracycline, gentamicin, and kanamycin, respectively. Specific genes of the Tn5397 transposon were detected in 54.8% of the tet(M)-positive enterococci. Nine virulence factors (gelE, agg, ace, cpd, frs, esp, hyl, efaAfs and efaAfm) were studied. All virulence genes, with the exception of the frs gene, were found to be present in the enterococcal isolates. At least one virulence gene was detected in 20.0% of E. faecium, 71.4% of E. faecalis and 33.3% of E. durans isolates, with ace and cpd being the most frequently detected genes (6 isolates each). This suggests that wild game meat might play a role in the spreading through the food chain of enterococci with antimicrobial resistance and virulence determinants to humans. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Erwinia amylovora expresses fast and simultaneously hrp/dsp virulence genes during flower infection on apple trees.

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    Doris Pester

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

  14. Virulence-associated gene pattern of porcine and human Yersinia enterocolitica biotype 4 isolates.

    Science.gov (United States)

    Schneeberger, M; Brodard, I; Overesch, G

    2015-04-02

    Yersinia enterocolitica 4/O:3 is the most important human pathogenic bioserotype in Europe and the predominant pathogenic bioserotype in slaughter pigs. Although many studies on the virulence of Y. enterocolitica strains have showed a broad spectrum of detectable factors in pigs and humans, an analysis based on a strict comparative approach and serving to verify the virulence capability of porcine Y. enterocolitica as a source for human yersiniosis is lacking. Therefore, in the present study, strains of biotype (BT) 4 isolated from Swiss slaughter pig tonsils and feces and isolates from human clinical cases were compared in terms of their spectrum of virulence-associated genes (yadA, virF, ail, inv, rovA, ymoA, ystA, ystB and myfA). An analysis of the associated antimicrobial susceptibility pattern completed the characterization. All analyzed BT 4 strains showed a nearly similar pattern, comprising the known fundamental virulence-associated genes yadA, virF, ail, inv, rovA, ymoA, ystA and myfA. Only ystB was not detectable among all analyzed isolates. Importantly, neither the source of the isolates (porcine tonsils and feces, humans) nor the serotype (ST) had any influence on the gene pattern. From these findings, it can be concluded that the presence of the full complement of virulence genes necessary for human infection is common among porcine BT 4 strains. Swiss porcine BT 4 strains not only showed antimicrobial susceptibility to chloramphenicol, cefotaxime, ceftazidime, ciprofloxacin, colistin, florfenicol, gentamicin, kanamycin, nalidixic acid, sulfamethoxazole, streptomycin, tetracycline and trimethoprim but also showed 100% antibiotic resistance to ampicillin. The human BT 4 strains revealed comparable results. However, in addition to 100% antibiotic resistance to ampicillin, 2 strains were resistant to chloramphenicol and nalidixic acid. Additionally, 1 of these strains was resistant to sulfamethoxazole. The results demonstrated that Y. enterocolitica BT 4

  15. Type VI Secretion is a Major Virulence Determinant in Burkholderia Mallei

    National Research Council Canada - National Science Library

    Schell, Mark A; Ulrich, Ricky L; Ribot, Wilson J; Brueggemann, Ernst E; Hines, Harry B; Chen, Dan; Lipscomb, Lyla; Kim, H. S; Mrazek, Jan; Nierman, William C; DeShazer, David

    2007-01-01

    Burkholderia mallei is a host-adapted pathogen and a category B biothreat agent. Although the B. mallei VirAG two-component regulatory system is required for virulence in hamsters, the virulence genes it regulates are unknown...

  16. Leaf Extracts of Mangifera indica L. Inhibit Quorum Sensing – Regulated Production of Virulence Factors and Biofilm in Test Bacteria

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    Iqbal Ahmad

    2017-04-01

    Full Text Available Quorum sensing (QS is a global gene regulatory mechanism in bacteria for various traits including virulence factors. Disabling QS system with anti-infective agent is considered as a potential strategy to prevent bacterial infection. Mangifera indica L. (mango has been shown to possess various biological activities including anti-QS. This study investigates the efficacy of leaf extracts on QS-regulated virulence factors and biofilm formation in Gram negative pathogens. Mango leaf (ML extract was tested for QS inhibition and QS-regulated virulence factors using various indicator strains. It was further correlated with the biofilm inhibition and confirmed by electron microscopy. Phytochemical analysis was carried out using ultra performance liquid chromatography (UPLC and gas chromatography–mass spectrometry (GC-MS analysis. In vitro evaluation of anti-QS activity of ML extracts against Chromobacterium violaceum revealed promising dose-dependent interference in violacein production, by methanol extract. QS inhibitory activity is also demonstrated by reduction in elastase (76%, total protease (56%, pyocyanin (89%, chitinase (55%, exopolysaccharide production (58% and swarming motility (74% in Pseudomonas aeruginosa PAO1 at 800 μg/ml concentration. Biofilm formation by P. aeruginosa PAO1 and Aeromonas hydrophila WAF38 was reduced considerably (36–82% over control. The inhibition of biofilm was also observed by scanning electron microscopy. Moreover, ML extracts significantly reduced mortality of Caenorhabditis elegans pre-infected with PAO1 at the tested concentration. Phytochemical analysis of active extracts revealed very high content of phenolics in methanol extract and a total of 14 compounds were detected by GC-MS and UPLC. These findings suggest that phytochemicals from the ML could provide bioactive anti-infective and needs further investigation to isolate and uncover their therapeutic efficacy.

  17. Leaf Extracts of Mangifera indica L. Inhibit Quorum Sensing – Regulated Production of Virulence Factors and Biofilm in Test Bacteria

    Science.gov (United States)

    Husain, Fohad M.; Ahmad, Iqbal; Al-thubiani, Abdullah S.; Abulreesh, Hussein H.; AlHazza, Ibrahim M.; Aqil, Farrukh

    2017-01-01

    Quorum sensing (QS) is a global gene regulatory mechanism in bacteria for various traits including virulence factors. Disabling QS system with anti-infective agent is considered as a potential strategy to prevent bacterial infection. Mangifera indica L. (mango) has been shown to possess various biological activities including anti-QS. This study investigates the efficacy of leaf extracts on QS-regulated virulence factors and biofilm formation in Gram negative pathogens. Mango leaf (ML) extract was tested for QS inhibition and QS-regulated virulence factors using various indicator strains. It was further correlated with the biofilm inhibition and confirmed by electron microscopy. Phytochemical analysis was carried out using ultra performance liquid chromatography (UPLC) and gas chromatography–mass spectrometry (GC-MS) analysis. In vitro evaluation of anti-QS activity of ML extracts against Chromobacterium violaceum revealed promising dose-dependent interference in violacein production, by methanol extract. QS inhibitory activity is also demonstrated by reduction in elastase (76%), total protease (56%), pyocyanin (89%), chitinase (55%), exopolysaccharide production (58%) and swarming motility (74%) in Pseudomonas aeruginosa PAO1 at 800 μg/ml concentration. Biofilm formation by P. aeruginosa PAO1 and Aeromonas hydrophila WAF38 was reduced considerably (36–82%) over control. The inhibition of biofilm was also observed by scanning electron microscopy. Moreover, ML extracts significantly reduced mortality of Caenorhabditis elegans pre-infected with PAO1 at the tested concentration. Phytochemical analysis of active extracts revealed very high content of phenolics in methanol extract and a total of 14 compounds were detected by GC-MS and UPLC. These findings suggest that phytochemicals from the ML could provide bioactive anti-infective and needs further investigation to isolate and uncover their therapeutic efficacy. PMID:28484444

  18. Bistable expression of virulence genes in salmonella leads to the formation of an antibiotic-tolerant subpopulation.

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    Markus Arnoldini

    2014-08-01

    Full Text Available Phenotypic heterogeneity can confer clonal groups of organisms with new functionality. A paradigmatic example is the bistable expression of virulence genes in Salmonella typhimurium, which leads to phenotypically virulent and phenotypically avirulent subpopulations. The two subpopulations have been shown to divide labor during S. typhimurium infections. Here, we show that heterogeneous virulence gene expression in this organism also promotes survival against exposure to antibiotics through a bet-hedging mechanism. Using microfluidic devices in combination with fluorescence time-lapse microscopy and quantitative image analysis, we analyzed the expression of virulence genes at the single cell level and related it to survival when exposed to antibiotics. We found that, across different types of antibiotics and under concentrations that are clinically relevant, the subpopulation of bacterial cells that express virulence genes shows increased survival after exposure to antibiotics. Intriguingly, there is an interplay between the two consequences of phenotypic heterogeneity. The bet-hedging effect that arises through heterogeneity in virulence gene expression can protect clonal populations against avirulent mutants that exploit and subvert the division of labor within these populations. We conclude that bet-hedging and the division of labor can arise through variation in a single trait and interact with each other. This reveals a new degree of functional complexity of phenotypic heterogeneity. In addition, our results suggest a general principle of how pathogens can evade antibiotics: Expression of virulence factors often entails metabolic costs and the resulting growth retardation could generally increase tolerance against antibiotics and thus compromise treatment.

  19. Temperature and Oxidative Stress as Triggers for Virulence Gene Expression in Pathogenic Leptospira spp.

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    Tricia Fraser

    2017-05-01

    Full Text Available Leptospirosis is a zooanthroponosis aetiologically caused by pathogenic bacteria belonging to the genus, Leptospira. Environmental signals such as increases in temperatures or oxidative stress can trigger response regulatory modes of virulence genes during infection. This study sought to determine the effect of temperature and oxidative stress on virulence associated genes in highly passaged Leptospira borgpeterseneii Jules and L. interrogans Portlandvere. Bacteria were grown in EMJH at 30°C, 37°C, or at 30°C before being transferred to 37°C. A total of 14 virulence-associated genes (fliY, invA, lenA, ligB, lipL32, lipL36, lipL41, lipL45, loa22, lsa21, mce, ompL1, sph2, and tlyC were assessed using endpoint PCR. Transcriptional analyses of lenA, lipL32, lipL41, loa22, sph2 were assessed by quantitative real-time RT-PCR at the temperature conditions. To assess oxidative stress, bacteria were exposed to H2O2 for 30 and 60 min with or without the temperature stress. All genes except ligB (for Portlandvere and ligB and mce (for Jules were detectable in the strains. Quantitatively, temperature stress resulted in significant changes in gene expression within species or between species. Temperature changes were more influential in gene expression for Jules, particularly at 30°C and upshift conditions; at 37°C, expression levels were higher for Portlandvere. However, compared to Jules, where temperature was influential in two of five genes, temperature was an essential element in four of five genes in Portlandvere exposed to oxidative stress. At both low and high oxidative stress levels, the interplay between genetic predisposition (larger genome size and temperature was biased towards Portlandvere particularly at 30°C and upshift conditions. While it is clear that expression of many virulence genes in highly passaged strains of Leptospira are attenuated or lost, genetic predisposition, changes in growth temperature and/or oxidative intensity and

  20. The Role of TonB Gene in Edwardsiella ictaluri Virulence

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    Hossam Abdelhamed

    2017-12-01

    Full Text Available Edwardsiella ictaluri is a Gram-negative facultative intracellular pathogen that causes enteric septicemia in catfish (ESC. Stress factors including poor water quality, poor diet, rough handling, overcrowding, and water temperature fluctuations increase fish susceptibility to ESC. The TonB energy transducing system (TonB-ExbB-ExbD and TonB-dependent transporters of Gram-negative bacteria support active transport of scarce resources including iron, an essential micronutrient for bacterial virulence. Deletion of the tonB gene attenuates virulence in several pathogenic bacteria. In the current study, the role of TonB (NT01EI_RS07425 in iron acquisition and E. ictaluri virulence were investigated. To accomplish this, the E. ictaluri tonB gene was in-frame deleted. Growth kinetics, iron utilization, and virulence of the EiΔtonB mutant were determined. Loss of TonB caused a significant reduction in bacterial growth in iron-depleted medium (p > 0.05. The EiΔtonB mutant grew similarly to wild-type E. ictaluri when ferric iron was added to the iron-depleted medium. The EiΔtonB mutant was significantly attenuated in catfish compared with the parent strain (21.69 vs. 46.91% mortality. Catfish surviving infection with EiΔtonB had significant protection against ESC compared with naïve fish (100 vs. 40.47% survival. These findings indicate that TonB participates in pathogenesis of ESC and is an important E. ictaluri virulence factor.

  1. The Staphylococcus aureus Global Regulator MgrA Modulates Clumping and Virulence by Controlling Surface Protein Expression.

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    Heidi A Crosby

    2016-05-01

    Full Text Available Staphylococcus aureus is a human commensal and opportunistic pathogen that causes devastating infections in a wide range of locations within the body. One of the defining characteristics of S. aureus is its ability to form clumps in the presence of soluble fibrinogen, which likely has a protective benefit and facilitates adhesion to host tissue. We have previously shown that the ArlRS two-component regulatory system controls clumping, in part by repressing production of the large surface protein Ebh. In this work we show that ArlRS does not directly regulate Ebh, but instead ArlRS activates expression of the global regulator MgrA. Strains lacking mgrA fail to clump in the presence of fibrinogen, and clumping can be restored to an arlRS mutant by overexpressing either arlRS or mgrA, indicating that ArlRS and MgrA constitute a regulatory pathway. We used RNA-seq to show that MgrA represses ebh, as well as seven cell wall-associated proteins (SraP, Spa, FnbB, SasG, SasC, FmtB, and SdrD. EMSA analysis showed that MgrA directly represses expression of ebh and sraP. Clumping can be restored to an mgrA mutant by deleting the genes for Ebh, SraP and SasG, suggesting that increased expression of these proteins blocks clumping by steric hindrance. We show that mgrA mutants are less virulent in a rabbit model of endocarditis, and virulence can be partially restored by deleting the genes for the surface proteins ebh, sraP, and sasG. While mgrA mutants are unable to clump, they are known to have enhanced biofilm capacity. We demonstrate that this increase in biofilm formation is partially due to up-regulation of SasG, a surface protein known to promote intercellular interactions. These results confirm that ArlRS and MgrA constitute a regulatory cascade, and that they control expression of a number of genes important for virulence, including those for eight large surface proteins.

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

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    Mukesh Meena

    2017-08-01

    Full Text Available 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.

  3. Relationship of biofilm formation and different virulence genes in uropathogenic Escherichia coli isolates from Northwest Iran

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    Fattahi, Sargol

    2015-07-01

    Full Text Available Background and objectives: The ( bacterium is one of the main causative agents of urinary tract infections (UTI worldwide. The ability of this bacterium to form biofilms on medical devices such as catheters plays an important role in the development of UTI. The aim of the present study was to investigate the possible relationship between virulence factors and biofilm formation of isolates responsible for urinary tract infection.Materials and methods: A total of 100 isolates isolated from patients with UTI were collected and characterized by routine bacteriological methods. In vitro biofilm formation by these isolates was determined using the 96-well microtiter-plate test, and the presence of , , and virulence genes was examined by PCR assay. Data analysis was performed using SPSS 16.0 software.Results: From 100 isolates isolated from UTIs, 92% were shown to be biofilm positive. The genes , , and were detected in 43%, 94% and 26% of isolates, respectively. Biofilm formation in isolates that expressed , , and genes was 100%, 93%, and 100%, respectively. A significant relationship was found between presence of the gene and biofilm formation in isolates isolated from UTI (<0.01, but there was no statistically significant correlation between presence of and genes with biofilm formation (<0.072, <0.104. Conclusion: Results showed that and genes do not seem to be necessary or sufficient for the production of biofilm in , but the presence of correlates with increased biofilm formation of urinary tract isolates. Overall, the presence of , , and virulence genes coincides with in vitro biofilm formation in uropathogenic

  4. Virulence genes in a probiotic E. coli product with a recorded long history of safe use

    Science.gov (United States)

    Zschüttig, Anke; Beimfohr, Claudia; Geske, Thomas; Auerbach, Christian; Cook, Helen; Zimmermann, Kurt; Gunzer, Florian

    2015-01-01

    The probiotic product Symbioflor2 (DSM 17252) is a bacterial concentrate of six different Escherichia coli genotypes, whose complete genome sequences are compared here, between each other as well as to other E. coli genomes. The genome sequences of Symbioflor2 E. coli components contained a number of virulence-associated genes. Their presence seems to be in conflict with a recorded history of safe use, and with the observed low frequency of adverse effects over a period of more than 6 years. The genome sequences were used to identify unique sequences for each component, for which strain-specific hybridization probes were designed. A colonization study was conducted whereby five volunteers were exposed to an exceptionally high single dose. The results showed that the probiotic E. coli could be detected for 3 months or longer in their stools, and this was in particular the case for those components containing higher numbers of virulence-associated genes. Adverse effects from this long-term colonization were absent. Thus, the presence of the identified virulence genes does not result in a pathogenic phenotype in the genetic background of these probiotic E. coli. PMID:25883796

  5. Characterization of Antimicrobial Resistance Patterns and Detection of Virulence Genes in Campylobacter Isolates in Italy

    Science.gov (United States)

    Di Giannatale, Elisabetta; Di Serafino, Gabriella; Zilli, Katiuscia; Alessiani, Alessandra; Sacchini, Lorena; Garofolo, Giuliano; Aprea, Giuseppe; Marotta, Francesca

    2014-01-01

    Campylobacter has developed resistance to several antimicrobial agents over the years, including macrolides, quinolones and fluoroquinolones, becoming a significant public health hazard. A total of 145 strains derived from raw milk, chicken faeces, chicken carcasses, cattle faeces and human faeces collected from various Italian regions, were screened for antimicrobial susceptibility, molecular characterization (SmaI pulsed-field gel electrophoresis) and detection of virulence genes (sequencing and DNA microarray analysis). The prevalence of C. jejuni and C. coli was 62.75% and 37.24% respectively. Antimicrobial susceptibility revealed a high level of resistance for ciprofloxacin (62.76%), tetracycline (55.86%) and nalidixic acid (55.17%). Genotyping of Campylobacter isolates using PFGE revealed a total of 86 unique SmaI patterns. Virulence gene profiles were determined using a new microbial diagnostic microarray composed of 70-mer oligonucleotide probes targeting genes implicated in Campylobacter pathogenicity. Correspondence between PFGE and microarray clusters was observed. Comparisons of PFGE and virulence profiles reflected the high genetic diversity of the strains examined, leading us to speculate different degrees of pathogenicity inside Campylobacter populations. PMID:24556669

  6. The regulation of virulence in phytopathogenic Erwinia species: quorum sensing, antibiotics and ecological considerations.

    Science.gov (United States)

    Whitehead, Neil A; Byers, Joseph T; Commander, Paul; Corbett, Mark J; Coulthurst, Sarah J; Everson, Lee; Harris, Abigail K P; Pemberton, Clare L; Simpson, Natalie J L; Slater, Holly; Smith, Debra S; Welch, Martin; Williamson, Neil; Salmond, George P C

    2002-08-01

    Erwinia carotovora is a Gram-negative bacterial phytopathogen that causes soft-rot disease and potato blackleg. The organism is environmentally widespread and exhibits an opportunistic plant pathogenesis. The ability to secrete multiple plant cell wall-degrading enzymes is a key virulence trait and exoenzyme production is responsive to multiple environmental and physiological cues. One important cue is the cell population density of the pathogen. Cell density is monitored via an acylated homoserine lactone (acyl HSL) signalling molecule, which is thought to diffuse between Erwinia cells in a process now commonly known as 'quorum sensing'. This molecule also acts as the chemical communication signal controlling production of a broad-spectrum beta-lactam antibiotic (1-carbapen-2-em-3-carboxylic acid; carbapenem) synthesised in concert with exoenzyme elaboration, possibly for niche defence. In antibiotic production control, quorum sensing acts at the level of transcriptional activation of the antibiotic biosynthetic cluster. This is achieved via a dedicated LuxR-type protein, CarR that is bound to the signalling molecule. The molecular relay connecting acyl HSL production and exoenzyme induction is not clear, despite the identification of a multitude of global regulatory genes, including those of the RsmA/rsmB system, impinging on enzyme synthesis. Quorum sensing control mediated by acyl HSLs is widespread in Gram-negative bacteria and is responsible for the regulation of diverse phenotypes. Although there is still a paucity of meaningful information on acyl HSL availability and in-situ biological function, there is growing evidence that such molecules play significant roles in microbial ecology.

  7. Disruption of each of the secreted aspartyl proteinase genes SAP1, SAP2, and SAP3 of Candida albicans attenuates virulence.

    OpenAIRE

    Hube, B; Sanglard, D; Odds, F C; Hess, D; Monod, M; Schäfer, W; Brown, A J; Gow, N A

    1997-01-01

    Secreted aspartyl proteinases (Saps), encoded by a gene family with at least nine members (SAP1 to SAP9), are one of the most discussed virulence factors produced by the human pathogen Candida albicans. In order to study the role of each Sap isoenzyme in pathogenicity, we have constructed strains which harbor mutations at selected SAP genes. SAP1, SAP2, and SAP3, which are regulated differentially in vitro, were mutated by targeted gene disruption. The growth rates of all homozygous null muta...

  8. A Pectate Lyase-Coding Gene Abundantly Expressed during Early Stages of Infection Is Required for Full Virulence in Alternaria brassicicola.

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    Yangrae Cho

    Full Text Available Alternaria brassicicola causes black spot disease of Brassica species. The functional importance of pectin digestion enzymes and unidentified phytotoxins in fungal pathogenesis has been suspected but not verified in A. brassicicola. The fungal transcription factor AbPf2 is essential for pathogenicity and induces 106 genes during early pathogenesis, including the pectate lyase-coding gene, PL1332. The aim of this study was to test the importance and roles of PL1332 in pathogenesis. We generated deletion strains of the PL1332 gene, produced heterologous PL1332 proteins, and evaluated their association with virulence. Deletion strains of the PL1332 gene were approximately 30% less virulent than wild-type A. brassicicola, without showing differences in colony expansion on solid media and mycelial growth in nutrient-rich liquid media or minimal media with pectins as a major carbon source. Heterologous PL1332 expressed as fusion proteins digested polygalacturons in vitro. When the fusion proteins were injected into the apoplast between leaf veins of host plants the tissues turned dark brown and soft, resembling necrotic leaf tissue. The PL1332 gene was the first example identified as a general toxin-coding gene and virulence factor among the 106 genes regulated by the transcription factor, AbPf2. It was also the first gene to have its functions investigated among the 19 pectate lyase genes and several hundred putative cell-wall degrading enzymes in A. brassicicola. These results further support the importance of the AbPf2 gene as a key pathogenesis regulator and possible target for agrochemical development.

  9. Network analysis of S. aureus response to ramoplanin reveals modules for virulence factors and resistance mechanisms and characteristic novel genes.

    Science.gov (United States)

    Subramanian, Devika; Natarajan, Jeyakumar

    2015-12-10

    Staphylococcus aureus is a major human pathogen and ramoplanin is an antimicrobial attributed for effective treatment. The goal of this study was to examine the transcriptomic profiles of ramoplanin sensitive and resistant S. aureus to identify putative modules responsible for virulence and resistance-mechanisms and its characteristic novel genes. The dysregulated genes were used to reconstruct protein functional association networks for virulence-factors and resistance-mechanisms individually. Strong link between metabolic-pathways and development of virulence/resistance is suggested. We identified 15 putative modules of virulence factors. Six hypothetical genes were annotated with novel virulence activity among which SACOL0281 was discovered to be an essential virulence factor EsaD. The roles of MazEF toxin-antitoxin system, SACOL0202/SACOL0201 two-component system and that of amino-sugar and nucleotide-sugar metabolism in virulence are also suggested. In addition, 14 putative modules of resistance mechanisms including modules of ribosomal protein-coding genes and metabolic pathways such as biotin-synthesis, TCA-cycle, riboflavin-biosynthesis, peptidoglycan-biosynthesis etc. are also indicated. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. Identification of genes preferentially expressed by highly virulent piscine Streptococcus agalactiae upon interaction with macrophages.

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    Chang-Ming Guo

    Full Text Available Streptococcus agalactiae, long recognized as a mammalian pathogen, is an emerging concern with regard to fish. In this study, we used a mouse model and in vitro cell infection to evaluate the pathogenetic characteristics of S. agalactiae GD201008-001, isolated from tilapia in China. This bacterium was found to be highly virulent and capable of inducing brain damage by migrating into the brain by crossing the blood-brain barrier (BBB. The phagocytosis assays indicated that this bacterium could be internalized by murine macrophages and survive intracellularly for more than 24 h, inducing injury to macrophages. Further, selective capture of transcribed sequences (SCOTS was used to investigate microbial gene expression associated with intracellular survival. This positive cDNA selection technique identified 60 distinct genes that could be characterized into 6 functional categories. More than 50% of the differentially expressed genes were involved in metabolic adaptation. Some genes have previously been described as associated with virulence in other bacteria, and four showed no significant similarities to any other previously described genes. This study constitutes the first step in further gene expression analyses that will lead to a better understanding of the molecular mechanisms used by S. agalactiae to survive in macrophages and to cross the BBB.

  11. Identification of Genes Preferentially Expressed by Highly Virulent Piscine Streptococcus agalactiae upon Interaction with Macrophages

    Science.gov (United States)

    Guo, Chang-Ming; Chen, Rong-Rong; Kalhoro, Dildar Hussain; Wang, Zhao-Fei; Liu, Guang-Jin; Lu, Cheng-Ping; Liu, Yong-Jie

    2014-01-01

    Streptococcus agalactiae, long recognized as a mammalian pathogen, is an emerging concern with regard to fish. In this study, we used a mouse model and in vitro cell infection to evaluate the pathogenetic characteristics of S. agalactiae GD201008-001, isolated from tilapia in China. This bacterium was found to be highly virulent and capable of inducing brain damage by migrating into the brain by crossing the blood–brain barrier (BBB). The phagocytosis assays indicated that this bacterium could be internalized by murine macrophages and survive intracellularly for more than 24 h, inducing injury to macrophages. Further, selective capture of transcribed sequences (SCOTS) was used to investigate microbial gene expression associated with intracellular survival. This positive cDNA selection technique identified 60 distinct genes that could be characterized into 6 functional categories. More than 50% of the differentially expressed genes were involved in metabolic adaptation. Some genes have previously been described as associated with virulence in other bacteria, and four showed no significant similarities to any other previously described genes. This study constitutes the first step in further gene expression analyses that will lead to a better understanding of the molecular mechanisms used by S. agalactiae to survive in macrophages and to cross the BBB. PMID:24498419

  12. Reduced Set of Virulence Genes Allows High Accuracy Prediction of Bacterial Pathogenicity in Humans

    Science.gov (United States)

    Iraola, Gregorio; Vazquez, Gustavo; Spangenberg, Lucía; Naya, Hugo

    2012-01-01

    Although there have been great advances in understanding bacterial pathogenesis, there is still a lack of integrative information about what makes a bacterium a human pathogen. The advent of high-throughput sequencing technologies has dramatically increased the amount of completed bacterial genomes, for both known human pathogenic and non-pathogenic strains; this information is now available to investigate genetic features that determine pathogenic phenotypes in bacteria. In this work we determined presence/absence patterns of different virulence-related genes among more than finished bacterial genomes from both human pathogenic and non-pathogenic strains, belonging to different taxonomic groups (i.e: Actinobacteria, Gammaproteobacteria, Firmicutes, etc.). An accuracy of 95% using a cross-fold validation scheme with in-fold feature selection is obtained when classifying human pathogens and non-pathogens. A reduced subset of highly informative genes () is presented and applied to an external validation set. The statistical model was implemented in the BacFier v1.0 software (freely available at ), that displays not only the prediction (pathogen/non-pathogen) and an associated probability for pathogenicity, but also the presence/absence vector for the analyzed genes, so it is possible to decipher the subset of virulence genes responsible for the classification on the analyzed genome. Furthermore, we discuss the biological relevance for bacterial pathogenesis of the core set of genes, corresponding to eight functional categories, all with evident and documented association with the phenotypes of interest. Also, we analyze which functional categories of virulence genes were more distinctive for pathogenicity in each taxonomic group, which seems to be a completely new kind of information and could lead to important evolutionary conclusions. PMID:22916122

  13. Prevalence of diarrheagenic Escherichia coli virulence genes in the feces of slaughtered cattle, chickens, and pigs in Burkina Faso

    Science.gov (United States)

    Kagambèga, Assèta; Martikainen, Outi; Siitonen, Anja; Traoré, Alfred S; Barro, Nicolas; Haukka, Kaisa

    2012-01-01

    We investigated the prevalence of the virulence genes specific for five major pathogroups of diarrheagenic Escherichia coli (DEC) in primary cultures from feces of animals slaughtered for human consumption in Burkina Faso. For the study, 704 feces samples were collected from cattle (n = 304), chickens (n = 350), and pigs (n = 50) during carcass processing. The presence of the virulence-associated genes in the mixed bacterial cultures was assessed using 16-plex polymerase chain reaction (PCR). Virulence genes indicating presence of DEC were detected in 48% of the cattle, 48% of the chicken, and 68% of the pig feces samples. Virulence genes specific for different DECs were detected in the following percentages of the cattle, chicken, and pig feces samples: Shiga toxin-producing E. coli (STEC) in 37%, 6%, and 30%; enteropathogenic E. coli (EPEC) in 8%, 37%, and 32%; enterotoxigenic E. coli (ETEC) in 4%, 5%, and 18%; and enteroaggregative E. coli (EAEC) in 7%, 6%, and 32%. Enteroinvasive E. coli (EIEC) virulence genes were detected in 1% of chicken feces samples only. The study was the first of its kind in Burkina Faso and revealed the common occurrence of the diarrheal virulence genes in feces of food animals. This indicates that food animals are reservoirs of DEC that may contaminate meat because of the defective slaughter and storage conditions and pose a health risk to the consumers in Burkina Faso. PMID:23170227

  14. Sugar Allocation to Metabolic Pathways is Tightly Regulated and Affects the Virulence of Streptococcus mutans

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    Miki Kawada-Matsuo

    2016-12-01

    Full Text Available Bacteria take up and metabolize sugar as a carbohydrate source for survival. Most bacteria can utilize many sugars, including glucose, sucrose, and galactose, as well as amino sugars, such as glucosamine and N-acetylglucosamine. After entering the cytoplasm, the sugars are mainly allocated to the glycolysis pathway (energy production and to various bacterial component biosynthesis pathways, including the cell wall, nucleic acids and amino acids. Sugars are also utilized to produce several virulence factors, such as capsule and lipoteichoic acid. Glutamine-fructose-6-phosphate aminotransferase (GlmS and glucosamine-6-phosphate deaminase (NagB have crucial roles in sugar distribution to the glycolysis pathway and to cell wall biosynthesis. In Streptococcus mutans, a cariogenic pathogen, the expression levels of glmS and nagB are coordinately regulated in response to the presence or absence of amino sugars. In addition, the disruption of this regulation affects the virulence of S. mutans. The expression of nagB and glmS is regulated by NagR in S. mutans, but the precise mechanism underlying glmS regulation is not clear. In Staphylococcus aureus and Bacillus subtilis, the mRNA of glmS has ribozyme activity and undergoes self-degradation at the mRNA level. However, there is no ribozyme activity region on glmS mRNA in S. mutans. In this review article, we summarize the sugar distribution, particularly the coordinated regulation of GlmS and NagB expression, and its relationship with the virulence of S. mutans.

  15. Transcriptional start site turnover in the evolution of bacterial paralogous genes - the pelE-pelD virulence genes in Dickeya.

    Science.gov (United States)

    Duprey, Alexandre; Nasser, William; Léonard, Simon; Brochier-Armanet, Céline; Reverchon, Sylvie

    2016-11-01

    After a gene duplication event, the resulting paralogous genes frequently acquire distinct expression profiles, roles, and/or functions but the underlying mechanisms are poorly understood. While transcription start site (TSS) turnover, i.e., the repositioning of the TSS during evolution, is widespread in eukaryotes, it is less documented in bacteria. Using pelD and pelE, two closely related paralogous genes encoding key virulence factors in Dickeya, a gamma proteobacterial genus of phytopathogens, we show that pelE has been selected as an initiator of bacterial aggression, while pelD acts at a later stage, thanks to modifications in the transcriptional regulation of these two genes. This expression change is linked to a few mutations that caused a shift in the position of the pelETSS and the rapid divergence in the regulation of these genes after their duplication. Genomic surveys detected additional examples of putative turnovers in other bacteria. This first report of TSS shifting in bacteria suggests that this mechanism could play a major role in paralogous genes fixation in prokaryotes. © 2016 Federation of European Biochemical Societies.

  16. PfSETvs methylation of histone H3K36 represses virulence genes in Plasmodium falciparum

    DEFF Research Database (Denmark)

    Jiang, Lubin; Mu, Jianbing; Zhang, Qingfeng

    2013-01-01

    The variant antigen Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1), which is expressed on the surface of P. falciparum-infected red blood cells, is a critical virulence factor for malaria. Each parasite has 60 antigenically distinct var genes that each code for a different PfEMP1...... parasite nuclei and their expression as proteins on the surface of individual infected red blood cells. PfSETvs-dependent H3K36me3 is present along the entire gene body, including the transcription start site, to silence var genes. With low occupancy of PfSETvs at both the transcription start site of var...... protein. During infection the clonal parasite population expresses only one gene at a time before switching to the expression of a new variant antigen as an immune-evasion mechanism to avoid the host antibody response. The mechanism by which 59 of the 60 var genes are silenced remains largely unknown...

  17. The host-encoded Heme Regulated Inhibitor (HRI facilitates virulence-associated activities of bacterial pathogens.

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    Niraj Shrestha

    Full Text Available Here we show that cells lacking the heme-regulated inhibitor (HRI are highly resistant to infection by bacterial pathogens. By examining the infection process in wild-type and HRI null cells, we found that HRI is required for pathogens to execute their virulence-associated cellular activities. Specifically, unlike wild-type cells, HRI null cells infected with the gram-negative bacterial pathogen Yersinia are essentially impervious to the cytoskeleton-damaging effects of the Yop virulence factors. This effect is due to reduced functioning of the Yersinia type 3 secretion (T3S system which injects virulence factors directly into the host cell cytosol. Reduced T3S activity is also observed in HRI null cells infected with the bacterial pathogen Chlamydia which results in a dramatic reduction in its intracellular proliferation. We go on to show that a HRI-mediated process plays a central role in the cellular infection cycle of the Gram-positive pathogen Listeria. For this pathogen, HRI is required for the post-invasion trafficking of the bacterium to the infected host cytosol. Thus by depriving Listeria of its intracellular niche, there is a highly reduced proliferation of Listeria in HRI null cells. We provide evidence that these infection-associated functions of HRI (an eIF2α kinase are independent of its activity as a regulator of protein synthesis. This is the first report of a host factor whose absence interferes with the function of T3S secretion and cytosolic access by pathogens and makes HRI an excellent target for inhibitors due to its broad virulence-associated activities.

  18. The response regulator expM is essential for the virulence of Erwinia carotovora subsp. carotovora and acts negatively on the sigma factor RpoS (sigma s).

    Science.gov (United States)

    Andersson, R A; Palva, E T; Pirhonen, M

    1999-07-01

    The main virulence factors of Erwinia carotovora subsp. carotovora, the secreted, extracellular cell-wall-degrading enzymes, are controlled by several regulatory mechanisms. We have isolated transposon mutants with reduced virulence on tobacco. One of these mutants, with a mutation in a gene designated expM, was characterized in this study. This mutant produces slightly reduced amounts of extracellular enzymes in vitro and the secretion of the enzymes is also affected. The expM wild-type allele was cloned together with an upstream gene, designated expL, that has an unknown function. The expM gene was sequenced and found to encode a protein with similarity to the RssB/SprE protein of Escherichia coli and the MviA protein of Salmonella typhimurium. These proteins belong to a new type of two-component response regulators that negatively regulate the stability of the Sigma factor RpoS (sigma s) at the protein level. The results of this study suggest that ExpM has a similar function in E. carotovora subsp. carotovora. We also provide evidence that the overproduction of RpoS in the expM mutant is an important factor for the reduced virulence phenotype and that it partly causes the observed phenotype seen in vitro. However, an expM/rpoS double mutant is still affected in secretion of extracellular enzymes, suggesting that ExpM in addition to RpoS also acts on other targets.

  19. Porcine E. coli: virulence-associated genes, resistance genes and adhesion and probiotic activity tested by a new screening method.

    Science.gov (United States)

    Schierack, Peter; Rödiger, Stefan; Kuhl, Christoph; Hiemann, Rico; Roggenbuck, Dirk; Li, Ganwu; Weinreich, Jörg; Berger, Enrico; Nolan, Lisa K; Nicholson, Bryon; Römer, Antje; Frömmel, Ulrike; Wieler, Lothar H; Schröder, Christian

    2013-01-01

    We established an automated screening method to characterize adhesion of Escherichia coli to intestinal porcine epithelial cells (IPEC-J2) and their probiotic activity against infection by enteropathogenic E. coli (EPEC). 104 intestinal E. coli isolates from domestic pigs were tested by PCR for the occurrence of virulence-associated genes, genes coding for resistances to antimicrobial agents and metals, and for phylogenetic origin by PCR. Adhesion rates and probiotic activity were examined for correlation with the presence of these genes. Finally, data were compared with those from 93 E. coli isolates from wild boars. Isolates from domestic pigs carried a broad variety of all tested genes and showed great diversity in gene patterns. Adhesions varied with a maximum of 18.3 or 24.2 mean bacteria adherence per epithelial cell after 2 or 6 hours respectively. Most isolates from domestic pigs and wild boars showed low adherence, with no correlation between adhesion/probiotic activity and E. coli genes or gene clusters. The gene sfa/foc, encoding for a subunit of F1C fimbriae did show a positive correlative association with adherence and probiotic activity; however E. coli isolates from wild boars with the sfa/foc gene showed less adhesion and probiotic activity than E. coli with the sfa/foc gene isolated from domestic pigs after 6 hour incubation. In conclusion, screening porcine E. coli for virulence associated genes genes, adhesion to intestinal epithelial cells, and probiotic activity revealed a single important adhesion factor, several probiotic candidates, and showed important differences between E. coli of domestic pigs and wild boars.

  20. Porcine E. coli: virulence-associated genes, resistance genes and adhesion and probiotic activity tested by a new screening method.

    Directory of Open Access Journals (Sweden)

    Peter Schierack

    Full Text Available We established an automated screening method to characterize adhesion of Escherichia coli to intestinal porcine epithelial cells (IPEC-J2 and their probiotic activity against infection by enteropathogenic E. coli (EPEC. 104 intestinal E. coli isolates from domestic pigs were tested by PCR for the occurrence of virulence-associated genes, genes coding for resistances to antimicrobial agents and metals, and for phylogenetic origin by PCR. Adhesion rates and probiotic activity were examined for correlation with the presence of these genes. Finally, data were compared with those from 93 E. coli isolates from wild boars. Isolates from domestic pigs carried a broad variety of all tested genes and showed great diversity in gene patterns. Adhesions varied with a maximum of 18.3 or 24.2 mean bacteria adherence per epithelial cell after 2 or 6 hours respectively. Most isolates from domestic pigs and wild boars showed low adherence, with no correlation between adhesion/probiotic activity and E. coli genes or gene clusters. The gene sfa/foc, encoding for a subunit of F1C fimbriae did show a positive correlative association with adherence and probiotic activity; however E. coli isolates from wild boars with the sfa/foc gene showed less adhesion and probiotic activity than E. coli with the sfa/foc gene isolated from domestic pigs after 6 hour incubation. In conclusion, screening porcine E. coli for virulence associated genes genes, adhesion to intestinal epithelial cells, and probiotic activity revealed a single important adhesion factor, several probiotic candidates, and showed important differences between E. coli of domestic pigs and wild boars.

  1. A sensor kinase recognizing the cell-cell signal BDSF (cis-2-dodecenoic acid) regulates virulence in Burkholderia cenocepacia

    DEFF Research Database (Denmark)

    McCarthy, Y.; Yang, Liang; Twomey, K.B.

    2010-01-01

    Xanthomonas campestris. The mechanism of perception of this signal and the range of functions regulated in B. cenocepacia are, however, unknown. A screen for transposon mutants unable to respond to exogenous signal identified BCAM0227 as a potential BDSF sensor. BCAM0227 is a histidine sensor kinase...... with an input domain unrelated to that of RpfC, the DSF sensor found in xanthomonads. Transcriptome profiling established the scope of the BDSF regulon and demonstrated that the sensor controls expression of a subset of these genes. A chimeric sensor kinase in which the input domain of BCAM0227 replaced...... the input domain of RpfC was active in BDSF signal perception when expressed in X. campestris. Mutation of BCAM0227 gave rise to reduced cytotoxicity to Chinese hamster ovary cells and reduced virulence to Wax moth larvae and in the agar-bead mouse model of pulmonary infection. The findings identify BCAM...

  2. Effect of the Streptococcus agalactiae Virulence Regulator CovR on the Pathogenesis of Urinary Tract Infection.

    Science.gov (United States)

    Sullivan, Matthew J; Leclercq, Sophie Y; Ipe, Deepak S; Carey, Alison J; Smith, Joshua P; Voller, Nathan; Cripps, Allan W; Ulett, Glen C

    2017-02-01

    Streptococcus agalactiae can cause urinary tract infection (UTI). The role of the S. agalactiae global virulence regulator, CovR, in UTI pathogenesis is unknown. We used murine and human bladder uroepithelial cell models of UTI and S. agalactiae mutants in covR and related factors, including β-hemolysin/cytolysin (β-h/c), surface-anchored adhesin HvgA, and capsule to study the role of CovR in UTI. We found that covR-deficient serotype III S. agalactiae 874391 was significantly attenuated for colonization in mice and adhesion to uroepithelial cells. Mice infected with covR-deficient S. agalactiae produced less proinflammatory cytokines than those infected with wild-type 874391. Acute cytotoxicity in uroepithelial cells triggered by covR-deficient but not wild-type 874391 was associated with significant caspase 3 activation. Mechanistically, covR mutation significantly altered the expression of several genes in S. agalactiae 874391 that encode key virulence factors, including β-h/c and HvgA, but not capsule. Subsequent mutational analyses revealed that HvgA and capsule, but not the β-h/c, exerted significant effects on colonization of the murine urinary tract in vivo. S. agalactiae CovR promotes bladder infection and inflammation, as well as adhesion to and viability of uroepithelial cells. The pathogenesis of S. agalactiae UTI is complex, multifactorial, and influenced by virulence effects of CovR, HvgA, and capsule. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

  3. Prophage Rs551 and Its Repressor Gene orf14 Reduce Virulence and Increase Competitive Fitness of Its Ralstonia solanacearum Carrier Strain UW551.

    Science.gov (United States)

    Ahmad, Abdelmonim Ali; Stulberg, Michael J; Huang, Qi

    2017-01-01

    We previously characterized a filamentous lysogenic bacteriophage, ϕRs551, isolated directly from the race 3 biovar 2 phylotype IIB sequevar 1 strain UW551 of Ralstonia solanacearum grown under normal culture conditions. The genome of ϕRs551 was identified with 100% identity in the deposited genomes of 11 race 3 biovar 2 phylotype IIB sequevar 1 strains of R. solanacearum , indicating evolutionary and biological importance, and ORF14 of ϕRs551 was annotated as a putative type-2 repressor. In this study, we determined the effect of the prophage and its ORF14 on the virulence and competitive fitness of its carrier strain UW551 by deleting the orf14 gene only (the UW551 orf14 mutant), and nine of the prophage's 14 genes including orf14 and six out of seven structural genes (the UW551 prophage mutant), respectively, from the genome of UW551. The two mutants were increased in extracellular polysaccharide production, twitching motility, expression of targeted virulence and virulence regulatory genes ( pilT, egl, pehC, hrPB, and phcA ), and virulence, suggesting that the virulence of UW551 was negatively regulated by ϕRs551, at least partially through ORF14. Interestingly, we found that the wt ϕRs551-carrying strain UW551 of R. solanacearum significantly outcompeted the wt strain RUN302 which lacks the prophage in tomato plants co-inoculated with the two strains. When each of the two mutant strains was co-inoculated with RUN302, however, the mutants were significantly out-competed by RUN302 for the same colonization site. Our results suggest that ecologically, ϕRs551 may play an important role by regulating the virulence of and offering a competitive fitness advantage to its carrier bacterial strain for persistence of the bacterium in the environment, which in turn prolongs the symbiotic relationship between the phage ϕRs551 and the R. solanacearum strain UW551. Our study is the first toward a better understanding of the co-existence between a lysogenic phage and

  4. Prophage Rs551 and Its Repressor Gene orf14 Reduce Virulence and Increase Competitive Fitness of Its Ralstonia solanacearum Carrier Strain UW551

    Directory of Open Access Journals (Sweden)

    Abdelmonim Ali Ahmad

    2017-12-01

    Full Text Available We previously characterized a filamentous lysogenic bacteriophage, ϕRs551, isolated directly from the race 3 biovar 2 phylotype IIB sequevar 1 strain UW551 of Ralstonia solanacearum grown under normal culture conditions. The genome of ϕRs551 was identified with 100% identity in the deposited genomes of 11 race 3 biovar 2 phylotype IIB sequevar 1 strains of R. solanacearum, indicating evolutionary and biological importance, and ORF14 of ϕRs551 was annotated as a putative type-2 repressor. In this study, we determined the effect of the prophage and its ORF14 on the virulence and competitive fitness of its carrier strain UW551 by deleting the orf14 gene only (the UW551 orf14 mutant, and nine of the prophage’s 14 genes including orf14 and six out of seven structural genes (the UW551 prophage mutant, respectively, from the genome of UW551. The two mutants were increased in extracellular polysaccharide production, twitching motility, expression of targeted virulence and virulence regulatory genes (pilT, egl, pehC, hrPB, and phcA, and virulence, suggesting that the virulence of UW551 was negatively regulated by ϕRs551, at least partially through ORF14. Interestingly, we found that the wt ϕRs551-carrying strain UW551 of R. solanacearum significantly outcompeted the wt strain RUN302 which lacks the prophage in tomato plants co-inoculated with the two strains. When each of the two mutant strains was co-inoculated with RUN302, however, the mutants were significantly out-competed by RUN302 for the same colonization site. Our results suggest that ecologically, ϕRs551 may play an important role by regulating the virulence of and offering a competitive fitness advantage to its carrier bacterial strain for persistence of the bacterium in the environment, which in turn prolongs the symbiotic relationship between the phage ϕRs551 and the R. solanacearum strain UW551. Our study is the first toward a better understanding of the co-existence between a

  5. Prophage Rs551 and Its Repressor Gene orf14 Reduce Virulence and Increase Competitive Fitness of Its Ralstonia solanacearum Carrier Strain UW551

    Science.gov (United States)

    Ahmad, Abdelmonim Ali; Stulberg, Michael J.; Huang, Qi

    2017-01-01

    We previously characterized a filamentous lysogenic bacteriophage, ϕRs551, isolated directly from the race 3 biovar 2 phylotype IIB sequevar 1 strain UW551 of Ralstonia solanacearum grown under normal culture conditions. The genome of ϕRs551 was identified with 100% identity in the deposited genomes of 11 race 3 biovar 2 phylotype IIB sequevar 1 strains of R. solanacearum, indicating evolutionary and biological importance, and ORF14 of ϕRs551 was annotated as a putative type-2 repressor. In this study, we determined the effect of the prophage and its ORF14 on the virulence and competitive fitness of its carrier strain UW551 by deleting the orf14 gene only (the UW551 orf14 mutant), and nine of the prophage’s 14 genes including orf14 and six out of seven structural genes (the UW551 prophage mutant), respectively, from the genome of UW551. The two mutants were increased in extracellular polysaccharide production, twitching motility, expression of targeted virulence and virulence regulatory genes (pilT, egl, pehC, hrPB, and phcA), and virulence, suggesting that the virulence of UW551 was negatively regulated by ϕRs551, at least partially through ORF14. Interestingly, we found that the wt ϕRs551-carrying strain UW551 of R. solanacearum significantly outcompeted the wt strain RUN302 which lacks the prophage in tomato plants co-inoculated with the two strains. When each of the two mutant strains was co-inoculated with RUN302, however, the mutants were significantly out-competed by RUN302 for the same colonization site. Our results suggest that ecologically, ϕRs551 may play an important role by regulating the virulence of and offering a competitive fitness advantage to its carrier bacterial strain for persistence of the bacterium in the environment, which in turn prolongs the symbiotic relationship between the phage ϕRs551 and the R. solanacearum strain UW551. Our study is the first toward a better understanding of the co-existence between a lysogenic phage and

  6. Enhanced Virulence Gene Activity of Agrobacterium in Muskmelon (Cucumis melo L. cv. ‘Birdie’

    Directory of Open Access Journals (Sweden)

    Abul K.M. MOHIUDDIN

    2011-05-01

    Full Text Available Muskmelon (Cucumis melo L. cultivar ‘Birdie’, was evaluated for its response to the tumorigenic Agrobacterium tumefaciens and the oncogenic A. rhizogenes strains. Stem and petiole of three week-old in vitro-grown muskmelon plants were inoculated with five strains of A. tumefaciens and A. rhizogenes each and observed phenotypic expressions i.e. induction of crown galls and hairy roots. This phenotypic expression was efficaciously increased when virulence gene activity of different strains of two Agrobacterium species was enhanced. Intensive studies on enhancement of virulence gene activity of Agrobacterium found to be correlated to the appropriate light intensity (39.3 μmol m-2 s-1 with a specific concentration of monocyclic phenolic compound, acetosyringone (20 μM. The gene activity was also influenced by several other physical factors e.g. plant tissue type, Agrobacterium species and their strains, and plant tissue-Agrobacterium interaction. Among the different A. tumefaciens strains, LBA4404 showed the best virulence gene activity in both stem and petiole through the formation of higher rate of crown galls. On the other hand, strain 15834 of A. rhizogenes showed better gene activity in stem and 8196 in petiole through the formation of higher rate of hairy roots as well as higher average number of hairy roots. Among the two different types of explants, petiole was more susceptible to both Agrobacterium species. Thus it was concluded that future muskmelon transformation study can efficiently be carried out with LBA4404, 15834 and 8196 strains using petiole explants by adding 20 μM of acetosyringone in the medium.

  7. Plasmid fingerprinting and virulence gene detection among indigenous strains of salmonella enterica serovar enteritidis

    International Nuclear Information System (INIS)

    Sajid, S.U.; Schwarz, S.

    2009-01-01

    Salmonella enterica serovar Enteritidis is an important frequently reported zoonotic pathogen and a common cause of human gastroenteritis worldwide. The highly conserved Serospecific plasmids (SSPs) and Salmonella plasmid virulence (Spv) genes have been shown to mediate extra-intestinal colonization and systemic infection. The objective of current study was to document the presence of SSPs and SpvB/SpvC genes prevailing in the indigenous population of serovar Enteritidis. A total of 48 epidemiologically unrelated strains of Salmonella enteritidis were included in the study. Preparation of plasmids DNA suitable for endonuclease digestion and separation of respective fragments by agarose gel electrophoresis followed previously described protocols. The plasmids of Escherichia coli V517, 1-kbp ladder, and lambda DNA HindIII fragments served as DNA size standards. Transfer of DNA fragments from agarose gels to nitrocellulose membranes was achieved by capillary blot procedure. An ECL labeled 3.6 kbp HindIII fragment of plasmid PRQ 51 was used as probe for SpvB/SpvC gene detection. Plasmid DNA fingerprinting revealed the presence of two different profiles of approximately 55 kbp and 90 kbp and were identified as virulence plasmids by DNA hybridization. The SpvB/SpvC genes were located on HindIII fragments of 3.6 kbp in each of the two types of virulence plasmids. The study confirms the presence of SSPs and SpvB/SpvC genes in indigenous strains of S. enteritidis isolated from Northern Punjab area of Pakistan and substantiate the previous data on such findings from other parts of the world. (author)

  8. Comparison of phenotypic and virulence genes characteristics in human and chicken isolates of Proteus mirabilis.

    Science.gov (United States)

    Barbour, Elie K; Hajj, Zahi G; Hamadeh, Shadi; Shaib, Houssam A; Farran, Mohamad T; Araj, George; Faroon, Obaid; Barbour, Kamil E; Jirjis, Faris; Azhar, Esam; Kumosani, Taha; Harakeh, Steve

    2012-10-01

    The objective of this work is to compare the phenotypic and virulence genes characteristics in human and chicken isolates of Proteus mirabilis. The bacterial examination of 50 livers of individual broilers, marketed by four major outlets, revealed a high recovery of P. mirabilis (66%), and a low recovery frequency of Salmonella spp. (4%), Serratia odorifera (2%), Citrobacter brakii (2%), and Providencia stuartii (2%). The phenotypic biochemical characterization of the recovered 33 chicken isolates of P. mirabilis were compared to 30 human isolates (23 urinary and six respiratory isolates). The comparison revealed significant differences in the presence of gelatinase enzyme (100% presence in chicken isolates versus 91.3 and 83.3% presence in human urinary and respiratory isolates, respectively, P,0.05). The H(2)S production occurred in 100% of chicken isolates versus 95.6 and 66.7% presence in human urinary and respiratory isolates, respectively, P,0.05). The other 17 biochemical characteristics did not differ significantly among the three groups of isolates (P.0.05). Two virulence genes, the mrpA and FliL, were having a typical 100% presence in randomly selected isolates of P. mirabilis recovered from chicken livers (N510) versus isolates recovered from urinary (N55) and respiratory specimens of humans (N55) (P.0.05). The average percentage similarity of mrpA gene nucleotide sequence of poultry isolates to human urinary and respiratory isolates was 93.2 and 97.5-%, respectively. The high similarity in phenotypic characteristics, associated with typical frequency of presence of two virulence genes, and high similarity in sequences of mrpA gene among poultry versus human P. mirabilis isolates justifies future investigations targeting the evaluation of adaptable pathogenicity of avian Proteus mirabilis isolates to mammalian hosts.

  9. Crystal Structures of SlyA Protein, a Master Virulence Regulator of Salmonella, in Free and DNA-bound States

    Energy Technology Data Exchange (ETDEWEB)

    Dolan, Kyle T.; Duguid, Erica M.; He, Chuan (UC)

    2011-11-17

    SlyA is a master virulence regulator that controls the transcription of numerous genes in Salmonella enterica. We present here crystal structures of SlyA by itself and bound to a high-affinity DNA operator sequence in the slyA gene. SlyA interacts with DNA through direct recognition of a guanine base by Arg-65, as well as interactions between conserved Arg-86 and the minor groove and a large network of non-base-specific contacts with the sugar phosphate backbone. Our structures, together with an unpublished structure of SlyA bound to the small molecule effector salicylate (Protein Data Bank code 3DEU), reveal that, unlike many other MarR family proteins, SlyA dissociates from DNA without large conformational changes when bound to this effector. We propose that SlyA and other MarR global regulators rely more on indirect readout of DNA sequence to exert control over many genes, in contrast to proteins (such as OhrR) that recognize a single operator.

  10. Different distribution patterns of ten virulence genes in Legionella reference strains and strains isolated from environmental water and patients.

    Science.gov (United States)

    Zhan, Xiao-Yong; Hu, Chao-Hui; Zhu, Qing-Yi

    2016-04-01

    Virulence genes are distinct regions of DNA which are present in the genome of pathogenic bacteria and absent in nonpathogenic strains of the same or related species. Virulence genes are frequently associated with bacterial pathogenicity in genus Legionella. In the present study, an assay was performed to detect ten virulence genes, including iraA, iraB, lvrA, lvrB, lvhD, cpxR, cpxA, dotA, icmC and icmD in different pathogenicity islands of 47 Legionella reference strains, 235 environmental strains isolated from water, and 4 clinical strains isolated from the lung tissue of pneumonia patients. The distribution frequencies of these genes in reference or/and environmental L. pneumophila strains were much higher than those in reference non-L. pneumophila or/and environmental non-L. pneumophila strains, respectively. L. pneumophila clinical strains also maintained higher frequencies of these genes compared to four other types of Legionella strains. Distribution frequencies of these genes in reference L. pneumophila strains were similar to those in environmental L. pneumophila strains. In contrast, environmental non-L. pneumophila maintained higher frequencies of these genes compared to those found in reference non-L. pneumophila strains. This study illustrates the association of virulence genes with Legionella pathogenicity and reveals the possible virulence evolution of non-L. pneumophia strains isolated from environmental water.

  11. Gene regulation by growth factors

    International Nuclear Information System (INIS)

    Metz, R.; Gorham, J.; Siegfried, Z.; Leonard, D.; Gizang-Ginsberg, E.; Thompson, M.A.; Lawe, D.; Kouzarides, T.; Vosatka, R.; MacGregor, D.; Jamal, S.; Greenberg, M.E.; Ziff, E.B.

    1988-01-01

    To coordinate the proliferation and differentiation of diverse cell types, cells of higher eukaryotes communicate through the release of growth factors. These peptides interact with specific transmembrane receptors of other cells and thereby generate intracellular messengers. The many changes in cellular physiology and activity that can be induced by growth factors imply that growth factor-induced signals can reach the nucleus and control gene activity. Moreover, current evidence also suggests that unregulated signaling along such pathways can induce aberrant proliferation and the formation of tumors. This paper reviews investigations of growth factor regulation of gene expression conducted by the authors' laboratory

  12. Multiple plasmid-borne virulence genes of Clavibacter michiganensis ssp. capsici critical for disease development in pepper.

    Science.gov (United States)

    Hwang, In Sun; Oh, Eom-Ji; Kim, Donghyuk; Oh, Chang-Sik

    2018-02-01

    Clavibacter michiganensis ssp. capsici is a Gram-positive plant-pathogenic bacterium causing bacterial canker disease in pepper. Virulence genes and mechanisms of C. michiganensis ssp. capsici in pepper have not yet been studied. To identify virulence genes of C. michiganensis ssp. capsici, comparative genome analyses with C. michiganensis ssp. capsici and its related C. michiganensis subspecies, and functional analysis of its putative virulence genes during infection were performed. The C. michiganensis ssp. capsici type strain PF008 carries one chromosome (3.056 Mb) and two plasmids (39 kb pCM1 Cmc and 145 kb pCM2 Cmc ). The genome analyses showed that this bacterium lacks a chromosomal pathogenicity island and celA gene that are important for disease development by C. michiganensis ssp. michiganensis in tomato, but carries most putative virulence genes in both plasmids. Virulence of pCM1 Cmc -cured C. michiganensis ssp. capsici was greatly reduced compared with the wild-type strain in pepper. The complementation analysis with pCM1 Cmc -located putative virulence genes showed that at least five genes, chpE, chpG, ppaA1, ppaB1 and pelA1, encoding serine proteases or pectate lyase contribute to disease development in pepper. In conclusion, C. michiganensis ssp. capsici has a unique genome structure, and its multiple plasmid-borne genes play critical roles in virulence in pepper, either separately or together. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  13. The Influences of Bacillus subtilis on the Virulence of Aeromonas hydrophila and Expression of luxS Gene of Both Bacteria Under Co-cultivation.

    Science.gov (United States)

    Ren, Yuwei; Li, Sisi; Wu, Zhixin; Zhou, Chengchong; Zhang, Ding; Chen, Xiaoxuan

    2017-06-01

    The aim of this study was to explore the influence of Bacillus subtilis CH9 on Aeromonas hydrophila SC2005. The transcription level of virulence genes of A. hydrophila SC2005 and its hemolysin activity as well as its cytotoxicity were analyzed when B. subtilis CH9 and A. hydrophila SC2005 were co-cultured. The results indicated that the transcription levels of four virulence genes of A. hydrophila, including aer, ahyB, hcp, and emp, decreased when A. hydrophila was cultured with B. subtilis CH9. Furthermore, the extracellular products of A. hydrophila showed attenuated hemolysin activity as well as cytotoxicity when A. hydrophila was cultured with B. subtilis CH9. Finally, the transcriptional levels of luxS genes of B. subtilis CH9 and A. hydrophila SC2005 were determined when these two species were co-cultured. RT-qPCR results suggested that the transcription level of A. hydrophila was down-regulated significantly. On the contrary, the transcription level of B. subtilis CH9 was up-regulated significantly. These results suggested that the probiotic role of B. subtilis CH9 is related to the inhibition of growth and virulence of A. hydrophila SC2005, and quorum sensing may be involved.

  14. Staphylococcus aureus nasal carriage in Ukraine: antibacterial resistance and virulence factor encoding genes.

    Science.gov (United States)

    Netsvyetayeva, Irina; Fraczek, Mariusz; Piskorska, Katarzyna; Golas, Marlena; Sikora, Magdalena; Mlynarczyk, Andrzej; Swoboda-Kopec, Ewa; Marusza, Wojciech; Palmieri, Beniamino; Iannitti, Tommaso

    2014-03-05

    The number of studies regarding the incidence of multidrug resistant strains and distribution of genes encoding virulence factors, which have colonized the post-Soviet states, is considerably limited. The aim of the study was (1) to assess the Staphylococcus (S.) aureus nasal carriage rate, including Methicillin Resistant S. aureus (MRSA) strains in adult Ukrainian population, (2) to determine antibiotic resistant pattern and (3) the occurrence of Panton Valentine Leukocidine (PVL)-, Fibronectin-Binding Protein A (FnBPA)- and Exfoliative Toxin (ET)-encoding genes. Nasal samples for S. aureus culture were obtained from 245 adults. The susceptibility pattern for several classes of antibiotics was determined by disk diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The virulence factor encoding genes, mecA, lukS-lukF, eta, etb, etd, fnbA, were detected by Polymerase Chain Reaction (PCR). The S. aureus nasal carriage rate was 40%. The prevalence of nasal MRSA carriage in adults was 3.7%. LukS-lukF genes were detected in over 58% of the strains. ET-encoding genes were detected in over 39% of the strains and the most prevalent was etd. The fnbA gene was detected in over 59% of the strains. All MRSA isolates tested were positive for the mecA gene. LukS-lukF genes and the etd gene were commonly co-present in MRSA, while lukS-lukF genes and the fnbA gene were commonly co-present in Methicillin Sensitive S. aureus (MSSA) isolates. No significant difference was detected between the occurrence of lukS-lukF genes (P > 0.05) and the etd gene (P > 0.05) when comparing MRSA and MSSA. The occurrence of the fnbA gene was significantly more frequent in MSSA strains (P aureus is a common cause of infection. The prevalence of S. aureus nasal carriage in our cohort of patients from Ukraine was 40.4%. We found that 9.1% of the strains were classified as MRSA and all MRSA isolates tested positive for the mecA gene

  15. Detection and sequence analysis of accessory gene regulator genes of Staphylococcus pseudintermedius isolates

    Directory of Open Access Journals (Sweden)

    M. Ananda Chitra

    2015-07-01

    Full Text Available Background: Staphylococcus pseudintermedius (SP is the major pathogenic species of dogs involved in a wide variety of skin and soft tissue infections. The accessory gene regulator (agr locus of Staphylococcus aureus has been extensively studied, and it influences the expression of many virulence genes. It encodes a two-component signal transduction system that leads to down-regulation of surface proteins and up-regulation of secreted proteins during in vitro growth of S. aureus. The objective of this study was to detect and sequence analyzing the AgrA, B, and D of SP isolated from canine skin infections. Materials and Methods: In this study, we have isolated and identified SP from canine pyoderma and otitis cases by polymerase chain reaction (PCR and confirmed by PCR-restriction fragment length polymorphism. Primers for SP agrA and agrBD genes were designed using online primer designing software and BLAST searched for its specificity. Amplification of the agr genes was carried out for 53 isolates of SP by PCR and sequencing of agrA, B, and D were carried out for five isolates and analyzed using DNAstar and Mega5.2 software. Results: A total of 53 (59% SP isolates were obtained from 90 samples. 15 isolates (28% were confirmed to be methicillinresistant SP (MRSP with the detection of the mecA gene. Accessory gene regulator A, B, and D genes were detected in all the SP isolates. Complete nucleotide sequences of the above three genes for five isolates were submitted to GenBank, and their accession numbers are from KJ133557 to KJ133571. AgrA amino acid sequence analysis showed that it is mainly made of alpha-helices and is hydrophilic in nature. AgrB is a transmembrane protein, and AgrD encodes the precursor of the autoinducing peptide (AIP. Sequencing of the agrD gene revealed that the 5 canine SP strains tested could be divided into three Agr specificity groups (RIPTSTGFF, KIPTSTGFF, and RIPISTGFF based on the putative AIP produced by each strain

  16. A Csr-type regulatory system, including small non-coding RNAs, regulates the global virulence regulator RovA of Yersinia pseudotuberculosis through RovM.

    Science.gov (United States)

    Heroven, Ann Kathrin; Böhme, Katja; Rohde, Manfred; Dersch, Petra

    2008-06-01

    The MarR-type regulator RovA controls expression of virulence genes of Yersinia pseudotuberculosis in response to environmental signals. Using a genetic strategy to discover components that influence rovA expression, we identified new regulatory factors with homology to components of the carbon storage regulator system (Csr). We showed that overexpression of a CsrB- or a CsrC-type RNA activates rovA, whereas a CsrA-like protein represses RovA synthesis. We further demonstrate that influence of the Csr system on rovA is indirect and occurs through control of the LysR regulator RovM, which inhibits rovA transcription. The CsrA protein had also a major influence on the motility of Yersinia, which was independent of RovM. The CsrB and CsrC RNAs are differentially expressed in Yersinia. CsrC is highly induced in complex but not in minimal media, indicating that medium-dependent rovM expression is mediated through CsrC. CsrB synthesis is generally very low. However, overexpression of the response regulator UvrY was found to activate CsrB production, which in turn represses CsrC synthesis independent of the growth medium. In summary, the post-transcriptional Csr-type components were shown to be key regulators in the co-ordinated environmental control of physiological processes and virulence factors, which are crucial for the initiation of Yersinia infections.

  17. Crystal structure of Bacillus anthracis virulence regulator AtxA and effects of phosphorylated histidines on multimerization and activity.

    Science.gov (United States)

    Hammerstrom, Troy G; Horton, Lori B; Swick, Michelle C; Joachimiak, Andrzej; Osipiuk, Jerzy; Koehler, Theresa M

    2015-02-01

    The Bacillus anthracis virulence regulator AtxA controls transcription of the anthrax toxin genes and capsule biosynthetic operon. AtxA activity is elevated during growth in media containing glucose and CO(2)/bicarbonate, and there is a positive correlation between the CO(2)/bicarbonate signal, AtxA activity and homomultimerization. AtxA activity is also affected by phosphorylation at specific histidines. We show that AtxA crystallizes as a dimer. Distinct folds associated with predicted DNA-binding domains (HTH1 and HTH2) and phosphoenolpyruvate: carbohydrate phosphotransferase system-regulated domains (PRD1 and PRD2) are apparent. We tested AtxA variants containing single and double phosphomimetic (His→Asp) and phosphoablative (His→Ala) amino acid changes for activity in B. anthracis cultures and for protein-protein interactions in cell lysates. Reduced activity of AtxA H199A, lack of multimerization and activity of AtxAH379D variants, and predicted structural changes associated with phosphorylation support a model for control of AtxA function. We propose that (i) in the AtxA dimer, phosphorylation of H199 in PRD1 affects HTH2 positioning, influencing DNA-binding; and (ii) phosphorylation of H379 in PRD2 disrupts dimer formation. The AtxA structure is the first reported high-resolution full-length structure of a PRD-containing regulator, and can serve as a model for proteins of this family, especially those that link virulence to bacterial metabolism. © 2014 John Wiley & Sons Ltd.

  18. Yersinia enterocolitica of porcine origin: carriage of virulence genes and genotypic diversity.

    Science.gov (United States)

    Tadesse, Daniel A; Bahnson, Peter B; Funk, Julie A; Morrow, W E Morgan; Abley, Melanie J; Ponte, Valeria A; Thakur, Siddhartha; Wittum, Thomas; DeGraves, Fred J; Rajala-Schultz, Paivi J; Gebreyes, Wondwossen A

    2013-01-01

    Yersinia enterocolitica is an important foodborne pathogen, and pigs are recognized as a major reservoir and potential source of pathogenic strains to humans. A total of 172 Y. enterocolitica recovered from conventional and antimicrobial-free pig production systems from different geographic regions (North Carolina, Ohio, Michigan, Wisconsin, and Iowa) were investigated to determine their pathogenic significance to humans. Phenotypic and genotypic diversity of the isolates was assessed using antibiogram, serogrouping, and amplified fragment length polymorphism (AFLP). Carriage of chromosomal and plasmid-borne virulence genes were investigated using polymerase chain reaction. A total of 12 antimicrobial resistance patterns were identified. More than two-thirds (67.4%) of Y. enterocolitica were pan-susceptible, and 27.9% were resistant against β-lactams. The most predominant serogroup was O:3 (43%), followed by O:5 (25.6%) and O:9 (4.1%). Twenty-two of 172 (12.8%) isolates were found to carry Yersinia adhesion A (yadA), a virulence gene encoded on the Yersinia virulence plasmid. Sixty-nine (40.1%) isolates were found to carry ail gene. The ystA and ystB genes were detected in 77% and 26.2% of the strains, respectively. AFLP genotyping of isolates showed wide genotypic diversity and were grouped into nine clades with an overall genotypic similarity of 66.8-99.3%. AFLP analysis revealed that isolates from the same production system showed clonal relatedness, while more than one genotype of Y. enterocolitica circulates within a farm.

  19. Virulence-associated gene profiling of Streptococcus suis isolates by PCR

    NARCIS (Netherlands)

    Silva, L.M.G.; Baums, C.G.; Rehm, T.; Wisselink, H.J.; Goethe, R.; Valentin-Weigand, P.

    2006-01-01

    Definition of virulent Streptococcus suis strains is controversial. One successful approach for identification of virulent European strains is differentiation of capsular serotypes (or the corresponding cps types) and subsequent detection of virulence-associated factors, namely the extracellular

  20. Cold Plasma Inactivation of Bacterial Biofilms and Reduction of Quorum Sensing Regulated Virulence Factors.

    Directory of Open Access Journals (Sweden)

    Dana Ziuzina

    Full Text Available The main objectives of this work were to investigate the effect of atmospheric cold plasma (ACP against a range of microbial biofilms commonly implicated in foodborne and healthcare associated human infections and against P. aeruginosa quorum sensing (QS-regulated virulence factors, such as pyocyanin, elastase (Las B and biofilm formation capacity post-ACP treatment. The effect of processing factors, namely treatment time and mode of plasma exposure on antimicrobial activity of ACP were also examined. Antibiofilm activity was assessed for E. coli, L. monocytogenes and S. aureus in terms of reduction of culturability and retention of metabolic activity using colony count and XTT assays, respectively. All samples were treated 'inpack' using sealed polypropylene containers with a high voltage dielectric barrier discharge ACP generated at 80 kV for 0, 60, 120 and 300 s and a post treatment storage time of 24 h. According to colony counts, ACP treatment for 60 s reduced populations of E. coli to undetectable levels, whereas 300 s was necessary to significantly reduce populations of L. monocytogenes and S. aureus biofilms. The results obtained from XTT assay indicated possible induction of viable but non culturable state of bacteria. With respect to P. aeruginosa QS-related virulence factors, the production of pyocyanin was significantly inhibited after short treatment times, but reduction of elastase was notable only after 300 s and no reduction in actual biofilm formation was achieved post-ACP treatment. Importantly, reduction of virulence factors was associated with reduction of the cytotoxic effects of the bacterial supernatant on CHO-K1 cells, regardless of mode and duration of treatment. The results of this study point to ACP technology as an effective strategy for inactivation of established biofilms and may play an important role in attenuation of virulence of pathogenic bacteria. Further investigation is warranted to propose direct evidence

  1. Detection of virulence-associated genes in pathogenic and commensal avian Escherichia coli isolates.

    Science.gov (United States)

    Paixão, A C; Ferreira, A C; Fontes, M; Themudo, P; Albuquerque, T; Soares, M C; Fevereiro, M; Martins, L; Corrêa de Sá, M I

    2016-07-01

    Poultry colibacillosis due to Avian Pathogenic Escherichia coli (APEC) is responsible for several extra-intestinal pathological conditions, leading to serious economic damage in poultry production. The most commonly associated pathologies are airsacculitis, colisepticemia, and cellulitis in broiler chickens, and salpingitis and peritonitis in broiler breeders. In this work a total of 66 strains isolated from dead broiler breeders affected with colibacillosis and 61 strains from healthy broilers were studied. Strains from broiler breeders were typified with serogroups O2, O18, and O78, which are mainly associated with disease. The serogroup O78 was the most prevalent (58%). All the strains were checked for the presence of 11 virulence genes: 1) arginine succinyltransferase A (astA); ii) E.coli hemeutilization protein A (chuA); iii) colicin V A/B (cvaA/B); iv) fimbriae mannose-binding type 1 (fimC); v) ferric yersiniabactin uptake A (fyuA); vi) iron-repressible high-molecular-weight proteins 2 (irp2); vii) increased serum survival (iss); viii) iron-uptake systems of E.coli D (iucD); ix) pielonefritis associated to pili C (papC); x) temperature sensitive haemaglutinin (tsh), and xi) vacuolating autotransporter toxin (vat), by Multiplex-PCR. The results showed that all genes are present in both commensal and pathogenic E. coli strains. The iron uptake-related genes and the serum survival gene were more prevalent among APEC. The adhesin genes, except tsh, and the toxin genes, except astA, were also more prevalent among APEC isolates. Except for astA and tsh, APEC strains harbored the majority of the virulence-associated genes studied and fimC was the most prevalent gene, detected in 96.97 and 88.52% of APEC and AFEC strains, respectively. Possession of more than one iron transport system seems to play an important role on APEC survival. © 2016 Poultry Science Association Inc.

  2. Virulence factors genes of Staphylococcus spp. isolated from caprine subclinical mastitis.

    Science.gov (United States)

    Salaberry, Sandra Renata Sampaio; Saidenberg, André Becker Simões; Zuniga, Eveline; Melville, Priscilla Anne; Santos, Franklin Gerônimo Bispo; Guimarães, Ednaldo Carvalho; Gregori, Fábio; Benites, Nilson Roberti

    2015-08-01

    The aim of this study was to investigate genes involved in adhesion expression, biofilm formation, and enterotoxin production in isolates of Staphylococcus spp. from goats with subclinical mastitis and associate these results with the staphylococcal species. One hundred and twenty-four isolates were identified and polymerase chain reaction (PCR) was performed to detect the following genes: cna, ebpS, eno, fib, fnbA, fnbB, bap, sea, seb, sec, sed and see. The most commonly Staphylococcus species included S. epidermidis, S. lugdunensis, S. chromogenes, S. capitis ss capitis and S. intermedius. With the exception of fnbB, the genes were detected in different frequencies of occurrence in 86.3% of the Staphylococcus spp. isolates. Eno (73.2%) and bap (94.8%) were more frequently detected in coagulase-negative staphylococci (CNS); ebpS (76%), fib (90.9%) and fnbA (87%) were the most frequent genes in coagulase-positive staphylococci (CPS). Regarding enterotoxins, genes sed (28.2%) and see (24.2%) had a higher frequency of occurrence; sec gene was more frequently detected in CPS (58.8%). There was no association between the presence of the genes and the Staphylococcus species. Different virulence factors genes can be detected in caprine subclinical mastitis caused by CNS and CPS. The knowledge of the occurrence of these virulence factors is important for the development of effective control and prevention measures of subclinical mastitis caused by CNS and CPS in goats. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. A mutation in the aroE gene affects pigment production, virulence, and chemotaxis in Xanthomonas oryzae pv. oryzae.

    Science.gov (United States)

    Kim, Hong-Il; Noh, Tae-Hwan; Lee, Chang-Soo; Park, Young-Jin

    2015-01-01

    Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial blight (BB) in rice. To study its function, a random insertion mutation library of Xoo was constructed using the Tn5 transposon. A mutant strain with decreased virulence against the susceptible rice cultivar IR24 was isolated from the library (aroE mutant), which also had extremely low pigment production. Thermal asymmetric interlaced-polymerase chain reaction (TAIL-PCR) and sequence analysis of the mutant revealed that the transposon was inserted into the aroE gene (encoding shikimate dehydrogenase). To investigate gene expression changes in the pigment- and virulence-deficient mutant, DNA microarray analysis was performed, which showed downregulation of 20 genes involved in the chemotaxis of Xoo. Our findings reveal that mutation of the aroE gene affects virulence and pigment production, as well as expression of genes involved in Xoo chemotaxis. Copyright © 2014 Elsevier GmbH. All rights reserved.

  4. Quorum-sensing-regulated virulence factors in Pseudomonas aeruginosa are toxic to Lucilia sericata maggots

    DEFF Research Database (Denmark)

    Andersen, A S; Joergensen, B; Bjarnsholt, T

    2010-01-01

    Maggot debridement therapy (MDT) is widely used for debridement of chronic infected wounds; however, for wounds harbouring specific bacteria limited effect or failure of the treatment has been described. Here we studied the survival of Lucilia sericata maggots encountering Pseudomonas aeruginosa...... PAO1 in a simple assay with emphasis on the quorum-sensing (QS)-regulated virulence. The maggots were challenged with GFP-tagged P. aeruginosa wild-type (WT) PAO1 and a GFP-tagged P. aeruginosa DeltalasR rhlR (DeltaRR) QS-deficient mutant in different concentrations. Maggots were killed...

  5. Helicobacter pylori virulence genes and microevolution in host and the clinical outcome: review article

    Directory of Open Access Journals (Sweden)

    Seyedeh Zahra Bakhti

    2014-12-01

    Full Text Available Helicobacter pylori (H. pylori is the causative agent in development of gastroduode-nal diseases, such as chronic atrophic gastritis, peptic ulcers, mucosa associated lym-phoid tissue (MALT lymphoma, and gastric cancer. H. pylori has been associated with inflammation in cardia, showing the fact that infection with this bacterium could also be a risk factor for gastric cardia cancer. Gastric cancer is the fourth most common cancer worldwide. This is the second leading cause of cancer-related deaths, and ap-proximately 700,000 people succumb each year to gastric adenocarcinoma. It has been estimated that 69% of the Iranian population currently harbor H. pylori infection. The prevalence of duodenal ulcer and gastric cancer is high in Iranian populations. However, this has been largely influenced by geographic and/or ethnic origin. Epidemi-ology studies have shown that host, environmental, and bacterial factors determine the outcome of H. pylori infection. The bacterium contains allelic diversity and high genet-ic variability into core- and virulence-genes and that this diversity is geographically and ethnically structured. The genetic diversity within H. pylori is greater than within most other bacteria, and its diversity is more than 50-fold higher than that of human DNA. The maintenance of high diversification makes this bacterium to cope with particular challenges in individual hosts. It has been reported that the recombination contributed to the creation of new genes and gene family. Furthermore, the microevolution in cagA and vacA genes is a common event, leading to a change in the virulence phenotype. These factors contribute to the bacterial survival in acidic conditions in stomach and protect it from host immune system, causing tissue damage and clinical disease. In this review article, we discussed the correlation between H. pylori virulence factors and clin-ical outcomes, microevolution of H. pylori virulence genes in a single host

  6. A Zinc-Finger-Family Transcription Factor, AbVf19, Is Required for the Induction of a Gene Subset Important for Virulence in Alternaria brassicicola

    Energy Technology Data Exchange (ETDEWEB)

    Srivastava, Akhil [Univ. of Hawaii, Manoa, HI (United States); Ohm, Robin A. [USDOE Joint Genome Inst., Walnut Creek, CA (United States); Oxiles, Lindsay [Univ. of Hawaii, Manoa, HI (United States); Brooks, Fred [Univ. of Hawaii, Manoa, HI (United States); Lawrence, Christopher B. [Virginia Polytechnic Inst. and State Univ. (Virginia Tech), Blacksburg, VA (United States); Grigoriev, Igor V. [USDOE Joint Genome Inst., Walnut Creek, CA (United States); Cho, Yangrae [Univ. of Hawaii, Manoa, HI (United States)

    2011-10-26

    Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen with a broad host range within the family Brassicaceae. It produces secondary metabolites that marginally affect virulence. Cell wall degrading enzymes (CDWE) have been considered important for pathogenesis but none of them individually have been identified as significant virulence factors in A. brassicicola. In this study, knockout mutants of a gene, AbVf19, were created and produced considerably smaller lesions than the wild type on inoculated host plants. The presence of tandem zinc-finger domains in the predicted amino acid sequence and nuclear localization of AbVf19- reporter protein suggested that it was a transcription factor. Gene expression comparisons using RNA-seq identified 74 genes being downregulated in the mutant during a late stage of infection. Among the 74 downregulated genes, 28 were putative CWDE genes. These were hydrolytic enzyme genes that composed a small fraction of genes within each family of cellulases, pectinases, cutinases, and proteinases. The mutants grew slower than the wild type on an axenic medium with pectin as a major carbon source. This study demonstrated the existence and the importance of a transcription factor that regulates a suite of genes that are important for decomposing and utilizing plant material during the late stage of plant infection.

  7. Salmonella Modulates Metabolism During Growth under Conditions that Induce Expression of Virulence Genes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Young-Mo; Schmidt, Brian; Kidwai, Afshan S.; Jones, Marcus B.; Deatherage, Brooke L.; Brewer, Heather M.; Mitchell, Hugh D.; Palsson, Bernhard O.; McDermott, Jason E.; Heffron, Fred; Smith, Richard D.; Peterson, Scott N.; Ansong, Charles; Hyduke, Daniel R.; Metz, Thomas O.; Adkins, Joshua N.

    2013-04-05

    Salmonella enterica serovar Typhimurium (S. Typhimurium) is a facultative pathogen that uses complex mechanisms to invade and proliferate within mammalian host cells. To investigate possible contributions of metabolic processes in S. Typhimurium grown under conditions known to induce expression of virulence genes, we used a metabolomics-driven systems biology approach coupled with genome scale modeling. First, we identified distinct metabolite profiles associated with bacteria grown in either rich or virulence-inducing media and report the most comprehensive coverage of the S. Typhimurium metabolome to date. Second, we applied an omics-informed genome scale modeling analysis of the functional consequences of adaptive alterations in S. Typhimurium metabolism during growth under our conditions. Excitingly, we observed possible sequestration of metabolites recently suggested to have immune modulating roles. Modeling efforts highlighted a decreased cellular capability to both produce and utilize intracellular amino acids during stationary phase culture in virulence conditions, despite significant abundance increases for these molecules as observed by our metabolomics measurements. Model-guided analysis suggested that alterations in metabolism prioritized other activities necessary for pathogenesis instead, such as lipopolysaccharide biosynthesis.

  8. Virulence Factor Genes in Staphylococcus aureus Isolated From Diabetic Foot Soft Tissue and Bone Infections.

    Science.gov (United States)

    Víquez-Molina, Gerardo; Aragón-Sánchez, Javier; Pérez-Corrales, Cristian; Murillo-Vargas, Christian; López-Valverde, María Eugenia; Lipsky, Benjamin A

    2018-03-01

    The aim of this study is to describe the presence of genes encoding for 4 virulence factors (pvl, eta, etb, and tsst), as well as the mecA gene conferring resistance to beta-lactam antibiotics, in patients with diabetes and a staphylococcal foot infection. We have also analyzed whether isolates of Staphylococcus aureus from bone infections have a different profile for these genes compared with those from exclusively soft tissue infections. In this cross-sectional study of a prospectively recruited series of patients admitted to the Diabetic Foot Unit, San Juan de Dios Hospital, San José, Costa Rica with a moderate or severe diabetic foot infection (DFI), we collected samples from infected soft tissue and from bone during debridement. During the study period (June 1, 2014 to May 31, 2016), we treated 379 patients for a DFI. S aureus was isolated from 101 wound samples, of which 43 were polymicrobial infections; we only included the 58 infections that were monomicrobial S aureus for this study. Infections were exclusively soft tissue in 17 patients (29.3%) while 41 (70.7%) had bone involvement (osteomyelitis). The mecA gene was detected in 35 cases (60.3%), pvl gene in 4 cases (6.9%), and tsst gene in 3 (5.2%). We did not detect etA and etB in any of the cases. There were no differences in the profile of S aureus genes encoding for virulence factors (pvl, etA, etB, and tsst) recovered from DFIs between those with just soft tissue compared to those with osteomyelitis. However, we found a significantly higher prevalence of pvl+ strains of S aureus associated with soft tissue compared with bone infections. Furthermore, we observed a significantly longer time to healing among patients infected with mecA+ (methicillin-resistant) S aureus (MRSA).

  9. Cotton plants export microRNAs to inhibit virulence gene expression in a fungal pathogen.

    Science.gov (United States)

    Zhang, Tao; Zhao, Yun-Long; Zhao, Jian-Hua; Wang, Sheng; Jin, Yun; Chen, Zhong-Qi; Fang, Yuan-Yuan; Hua, Chen-Lei; Ding, Shou-Wei; Guo, Hui-Shan

    2016-09-26

    Plant pathogenic fungi represent the largest group of disease-causing agents on crop plants, and are a constant and major threat to agriculture worldwide. Recent studies have shown that engineered production of RNA interference (RNAi)-inducing dsRNA in host plants can trigger specific fungal gene silencing and confer resistance to fungal pathogens 1-7 . Although these findings illustrate efficient uptake of host RNAi triggers by pathogenic fungi, it is unknown whether or not such an uptake mechanism has been evolved for a natural biological function in fungus-host interactions. Here, we show that in response to infection with Verticillium dahliae (a vascular fungal pathogen responsible for devastating wilt diseases in many crops) cotton plants increase production of microRNA 166 (miR166) and miR159 and export both to the fungal hyphae for specific silencing. We found that two V. dahliae genes encoding a Ca 2+ -dependent cysteine protease (Clp-1) and an isotrichodermin C-15 hydroxylase (HiC-15), and targeted by miR166 and miR159, respectively, are both essential for fungal virulence. Notably, V. dahliae strains expressing either Clp-1 or HiC-15 rendered resistant to the respective miRNA exhibited drastically enhanced virulence in cotton plants. Together, our findings identify a novel defence strategy of host plants by exporting specific miRNAs to induce cross-kingdom gene silencing in pathogenic fungi and confer disease resistance.

  10. Evidence of cross gene regulation of some virulence factors of ...

    African Journals Online (AJOL)

    Periodontal disease has been associated with poor dental care, which promotes the accumulation of bacteria and the development of diseases of the mouth. Porphyromonas gingivalis are anaerobic Gramnegative bacteria found in the subgingival plaque. They are largely responsible for chronic periodontal disease.

  11. Gene Regulation in Streptococcus pneumoniae: interplay between nutrition and virulence

    NARCIS (Netherlands)

    W.T. Hendriksen (Wouter)

    2010-01-01

    textabstractStreptococcus pneumoniae (the pneumococcus) is a Gram-positive bacterium, which belongs to the species of streptococci. Other pathogenic bacteria belonging to this class include Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus suis, Streptococcus uberis, Streptococcus

  12. Evidence of cross gene regulation of some virulence factors of ...

    African Journals Online (AJOL)

    David Mtz

    2013-07-10

    Jul 10, 2013 ... They are largely responsible for chronic periodontal disease. .... nucleotide sequence is not reported by the manufacturer (Roche). Previous to the use of .... thesis of different metabolic pathways; in the case of downregulated ...

  13. Identification of anti-virulence compounds that disrupt quorum-sensing regulated acute and persistent pathogenicity.

    Directory of Open Access Journals (Sweden)

    Melissa Starkey

    2014-08-01

    Full Text Available Etiological agents of acute, persistent, or relapsing clinical infections are often refractory to antibiotics due to multidrug resistance and/or antibiotic tolerance. Pseudomonas aeruginosa is an opportunistic Gram-negative bacterial pathogen that causes recalcitrant and severe acute chronic and persistent human infections. Here, we target the MvfR-regulated P. aeruginosa quorum sensing (QS virulence pathway to isolate robust molecules that specifically inhibit infection without affecting bacterial growth or viability to mitigate selective resistance. Using a whole-cell high-throughput screen (HTS and structure-activity relationship (SAR analysis, we identify compounds that block the synthesis of both pro-persistence and pro-acute MvfR-dependent signaling molecules. These compounds, which share a benzamide-benzimidazole backbone and are unrelated to previous MvfR-regulon inhibitors, bind the global virulence QS transcriptional regulator, MvfR (PqsR; inhibit the MvfR regulon in multi-drug resistant isolates; are active against P. aeruginosa acute and persistent murine infections; and do not perturb bacterial growth. In addition, they are the first compounds identified to reduce the formation of antibiotic-tolerant persister cells. As such, these molecules provide for the development of next-generation clinical therapeutics to more effectively treat refractory and deleterious bacterial-human infections.

  14. Small-Molecule Inhibitor of the Shigella flexneri Master Virulence Regulator VirF

    Science.gov (United States)

    Koppolu, Veerendra; Osaka, Ichie; Skredenske, Jeff M.; Kettle, Bria; Hefty, P. Scott; Li, Jiaqin

    2013-01-01

    VirF is an AraC family transcriptional activator that is required for the expression of virulence genes associated with invasion and cell-to-cell spread by Shigella flexneri, including multiple components of the type three secretion system (T3SS) machinery and effectors. We tested a small-molecule compound, SE-1 (formerly designated OSSL_051168), which we had identified as an effective inhibitor of the AraC family proteins RhaS and RhaR, for its ability to inhibit VirF. Cell-based reporter gene assays with Escherichia coli and Shigella, as well as in vitro DNA binding assays with purified VirF, demonstrated that SE-1 inhibited DNA binding and transcription activation (likely by blocking DNA binding) by VirF. Analysis of mRNA levels using real-time quantitative reverse transcription-PCR (qRT-PCR) further demonstrated that SE-1 reduced the expression of the VirF-dependent virulence genes icsA, virB, icsB, and ipaB in Shigella. We also performed eukaryotic cell invasion assays and found that SE-1 reduced invasion by Shigella. The effect of SE-1 on invasion required preincubation of Shigella with SE-1, in agreement with the hypothesis that SE-1 inhibited the expression of VirF-activated genes required for the formation of the T3SS apparatus and invasion. We found that the same concentrations of SE-1 had no detectable effects on the growth or metabolism of the bacterial cells or the eukaryotic host cells, respectively, indicating that the inhibition of invasion was not due to general toxicity. Overall, SE-1 appears to inhibit transcription activation by VirF, exhibits selectivity toward AraC family proteins, and has the potential to be developed into a novel antibacterial agent. PMID:24002059

  15. QB1 - Stochastic Gene Regulation

    Energy Technology Data Exchange (ETDEWEB)

    Munsky, Brian [Los Alamos National Laboratory

    2012-07-23

    Summaries of this presentation are: (1) Stochastic fluctuations or 'noise' is present in the cell - Random motion and competition between reactants, Low copy, quantization of reactants, Upstream processes; (2) Fluctuations may be very important - Cell-to-cell variability, Cell fate decisions (switches), Signal amplification or damping, stochastic resonances; and (3) Some tools are available to mode these - Kinetic Monte Carlo simulations (SSA and variants), Moment approximation methods, Finite State Projection. We will see how modeling these reactions can tell us more about the underlying processes of gene regulation.

  16. Biochemical basis for activation of virulence genes by bile salts in Vibrio parahaemolyticus.

    Science.gov (United States)

    Rivera-Cancel, Giomar; Orth, Kim

    2017-07-04

    Bile salts act as a stressor to bacteria that transit the intestinal tract. Enteric pathogens have hijacked bile as an intestinal signal to regulate virulence factors. We recently demonstrated that Vibrio parahemolyticus senses bile salts via a heterodimeric receptor formed by the periplasmic domains of inner-membrane proteins VtrA and VtrC. Crystal structures of the periplasmic complex reveal that VtrA and VtrC form a β-barrel that binds bile salts in its hydrophobic interior to activate the VtrA cytoplasmic DNA-binding domain. Proteins with the same domain arrangement as VtrA and VtrC are widespread in Vibrio and related bacteria, where they are involved in regulating virulence and other unknown functions. Here we discuss our findings and review current knowledge on VtrA and VtrC homologs. We propose that signaling by these membrane-bound transcription factors can be advantageous for the regulation of membrane and secretory proteins.

  17. Biochemical basis for activation of virulence genes by bile salts in Vibrio parahaemolyticus

    Science.gov (United States)

    2017-01-01

    ABSTRACT Bile salts act as a stressor to bacteria that transit the intestinal tract. Enteric pathogens have hijacked bile as an intestinal signal to regulate virulence factors. We recently demonstrated that Vibrio parahemolyticus senses bile salts via a heterodimeric receptor formed by the periplasmic domains of inner-membrane proteins VtrA and VtrC. Crystal structures of the periplasmic complex reveal that VtrA and VtrC form a β-barrel that binds bile salts in its hydrophobic interior to activate the VtrA cytoplasmic DNA-binding domain. Proteins with the same domain arrangement as VtrA and VtrC are widespread in Vibrio and related bacteria, where they are involved in regulating virulence and other unknown functions. Here we discuss our findings and review current knowledge on VtrA and VtrC homologs. We propose that signaling by these membrane-bound transcription factors can be advantageous for the regulation of membrane and secretory proteins. PMID:28129014

  18. Associations between Antimicrobial Resistance Phenotypes, Antimicrobial Resistance Genes, and Virulence Genes of Fecal Escherichia coli Isolates from Healthy Grow-Finish Pigs ▿

    OpenAIRE

    Rosengren, Leigh B.; Waldner, Cheryl L.; Reid-Smith, Richard J.

    2009-01-01

    Escherichia coli often carries linked antimicrobial resistance genes on transmissible genetic elements. Through coselection, antimicrobial use may select for unrelated but linked resistance or virulence genes. This study used unconditional statistical associations to investigate the relationships between antimicrobial resistance phenotypes and antimicrobial resistance genes in 151 E. coli isolates from healthy pigs. Phenotypic resistance to each drug was significantly associated with phenotyp...

  19. Agrobacterium tumefaciens-Mediated Transformation of Pseudocercospora fijiensis to Determine the Role of PfHog1 in Osmotic Stress Regulation and Virulence Modulation.

    Science.gov (United States)

    Onyilo, Francis; Tusiime, Geoffrey; Chen, Li-Hung; Falk, Bryce; Stergiopoulos, Ioannis; Tripathi, Jaindra N; Tushemereirwe, Wilberforce; Kubiriba, Jerome; Changa, Charles; Tripathi, Leena

    2017-01-01

    Black Sigatoka disease, caused by Pseudocercospora fijiensis is a serious constraint to banana production worldwide. The disease continues to spread in new ecological niches and there is an urgent need to develop strategies for its control. The high osmolarity glycerol (HOG) pathway in Saccharomyces cerevisiae is well known to respond to changes in external osmolarity. HOG pathway activation leads to phosphorylation, activation and nuclear transduction of the HOG1 mitogen-activated protein kinases (MAPKs). The activated HOG1 triggers several responses to osmotic stress, including up or down regulation of different genes, regulation of protein translation, adjustments to cell cycle progression and synthesis of osmolyte glycerol. This study investigated the role of the MAPK-encoding PfHog1 gene on osmotic stress adaptation and virulence of P. fijie nsis. RNA interference-mediated gene silencing of PfHog1 significantly suppressed growth of P. fijiensis on potato dextrose agar media supplemented with 1 M NaCl, indicating that PfHog1 regulates osmotic stress. In addition, virulence of the PfHog1 -silenced mutants of P. fijiensis on banana was significantly reduced, as observed from the low rates of necrosis and disease development on the infected leaves. Staining with lacto phenol cotton blue further confirmed the impaired mycelial growth of the PfHog1 in the infected leaf tissues, which was further confirmed with quantification of the fungal biomass using absolute- quantitative PCR. Collectively, these findings demonstrate that PfHog1 plays a critical role in osmotic stress regulation and virulence of P. fijiensis on its host banana. Thus, PfHog1 could be an interesting target for the control of black Sigatoka disease in banana.

  20. Agrobacterium tumefaciens-Mediated Transformation of Pseudocercospora fijiensis to Determine the Role of PfHog1 in Osmotic Stress Regulation and Virulence Modulation

    Directory of Open Access Journals (Sweden)

    Francis Onyilo

    2017-05-01

    Full Text Available Black Sigatoka disease, caused by Pseudocercospora fijiensis is a serious constraint to banana production worldwide. The disease continues to spread in new ecological niches and there is an urgent need to develop strategies for its control. The high osmolarity glycerol (HOG pathway in Saccharomyces cerevisiae is well known to respond to changes in external osmolarity. HOG pathway activation leads to phosphorylation, activation and nuclear transduction of the HOG1 mitogen-activated protein kinases (MAPKs. The activated HOG1 triggers several responses to osmotic stress, including up or down regulation of different genes, regulation of protein translation, adjustments to cell cycle progression and synthesis of osmolyte glycerol. This study investigated the role of the MAPK-encoding PfHog1 gene on osmotic stress adaptation and virulence of P. fijiensis. RNA interference-mediated gene silencing of PfHog1 significantly suppressed growth of P. fijiensis on potato dextrose agar media supplemented with 1 M NaCl, indicating that PfHog1 regulates osmotic stress. In addition, virulence of the PfHog1-silenced mutants of P. fijiensis on banana was significantly reduced, as observed from the low rates of necrosis and disease development on the infected leaves. Staining with lacto phenol cotton blue further confirmed the impaired mycelial growth of the PfHog1 in the infected leaf tissues, which was further confirmed with quantification of the fungal biomass using absolute- quantitative PCR. Collectively, these findings demonstrate that PfHog1 plays a critical role in osmotic stress regulation and virulence of P. fijiensis on its host banana. Thus, PfHog1 could be an interesting target for the control of black Sigatoka disease in banana.

  1. SigB is a dominant regulator of virulence in Staphylococcus aureus small-colony variants.

    Science.gov (United States)

    Mitchell, Gabriel; Fugère, Alexandre; Pépin Gaudreau, Karine; Brouillette, Eric; Frost, Eric H; Cantin, André M; Malouin, François

    2013-01-01

    Staphylococcus aureus small-colony variants (SCVs) are persistent pathogenic bacteria characterized by slow growth and, for many of these strains, an increased ability to form biofilms and to persist within host cells. The virulence-associated gene expression profile of SCVs clearly differs from that of prototypical strains and is often influenced by SigB rather than by the agr system. One objective of this work was to confirm the role of SigB in the control of the expression of virulence factors involved in biofilm formation and intracellular persistence of SCVs. This study shows that extracellular proteins are involved in the formation of biofilm by three SCV strains, which, additionally, have a low biofilm-dispersing activity. It was determined that SigB activity modulates biofilm formation by strain SCV CF07-S and is dominant over that of the agr system without being solely responsible for the repression of proteolytic activity. On the other hand, the expression of fnbA and the control of nuclease activity contributed to the SigB-dependent formation of biofilm of this SCV strain. SigB was also required for the replication of CF07-S within epithelial cells and may be involved in the colonization of lungs by SCVs in a mouse infection model. This study methodically investigated SigB activity and associated mechanisms in the various aspects of SCV pathogenesis. Results confirm that SigB activity importantly influences the production of virulence factors, biofilm formation and intracellular persistence for some clinical SCV strains.

  2. Staphylococcus aureus Isolates from Bovine Mastitis in Eight Countries: Genotypes, Detection of Genes Encoding Different Toxins and Other Virulence Genes

    Directory of Open Access Journals (Sweden)

    Valentina Monistero

    2018-06-01

    Full Text Available Staphylococcus aureus is recognized worldwide as one of the major agents of dairy cow intra-mammary infections. This microorganism can express a wide spectrum of pathogenic factors used to attach, colonize, invade and infect the host. The present study evaluated 120 isolates from eight different countries that were genotyped by RS-PCR and investigated for 26 different virulence factors to increase the knowledge on the circulating genetic lineages among the cow population with mastitis. New genotypes were observed for South African strains while for all the other countries new variants of existing genotypes were detected. For each country, a specific genotypic pattern was found. Among the virulence factors, fmtB, cna, clfA and leucocidins genes were the most frequent. The sea and sei genes were present in seven out of eight countries; seh showed high frequency in South American countries (Brazil, Colombia, Argentina, while sel was harboured especially in one Mediterranean country (Tunisia. The etb, seb and see genes were not detected in any of the isolates, while only two isolates were MRSA (Germany and Italy confirming the low diffusion of methicillin resistance microorganism among bovine mastitis isolates. This work demonstrated the wide variety of S. aureus genotypes found in dairy cattle worldwide. This condition suggests that considering the region of interest might help to formulate strategies for reducing the infection spreading.

  3. Coliform bacteria isolated from recreational lakes carry class 1 and class 2 integrons and virulence-associated genes.

    Science.gov (United States)

    Koczura, R; Krysiak, N; Taraszewska, A; Mokracka, J

    2015-08-01

    To characterize the integron-harbouring Gram-negative bacteria in recreational lakes, with focus on the genetic content of integrons, antimicrobial resistance profiles and virulence-associated genes. The presence and structure of integrons in coliform bacteria isolated from the water of four recreational lakes located in Poznań, Poland, was determined by PCR method. Antimicrobial resistance testing was done by disc diffusion method. Virulence-associated genes in integron-bearing Escherichia coli isolates were detected by PCR. A total of 155 integron-bearing strains of coliform bacteria were cultured. Sequence analysis showed the presence of dfrA7, aadA1, dfrA1-aadA1, dfrA17-aadA5 and dfrA12-orfF-aadA2 gene cassette arrays in class 1 integrons and dfrA1-sat2-aadA1 in class 2 integrons. Higher frequency of integron-positive bacteria and higher antimicrobial resistance ranges were noted in colder months (January and November) compared with spring and summer months. The integron-harbouring E. coli carried up to nine virulence-associated genes, with the highest frequency of kpsMT (84.6%) and traT (783%), coding for group 2 capsule and determining human serum resistance respectively. Integron-bearing multidrug resistant coliform bacteria carrying virulence genes are present in waters of recreational lakes. This study presents antimicrobial resistance and virulence-associated genes in integron-bearing coliform bacteria present in the waters of recreational lakes, which showed that multidrug resistant bacteria with virulence traits might pose a threat to public health. Moreover, the presence of genes typical for enterotoxigenic and Shiga toxin-producing E. coli is a concern. © 2015 The Society for Applied Microbiology.

  4. Diversity of virulence genes in Brucella melitensis and Brucella abortus detected from patients with rheumatoid arthritis.

    Science.gov (United States)

    Rahdar, Hossein Ali; Golmohammadi, Reza; Mirnejad, Reza; Ataee, Ramezan Ali; Alishiri, Gholam Hossein; Kazemian, Hossein

    2018-03-22

    The presence of Brucella melitensis and Brucella abortus genomes were investigated in the synovial fluid (SF) samples from 90 patients with rheumatoid arthritis (RA). DNA extraction and PCR assay were performed for simultaneous identification and discrimination of B. melitensis and B. abortus from the SF using three specific primers. After gel electrophoresis, the PCR products were confirmed by DNA sequencing. The cbg, omp31, manA, virB, and znuA virulence genes typing were performed by multiplex-PCR. Of the 90 samples, 14 were positive for B. melitensis (n = 9; 10%) and B. abortus (n = 5; 5.5%). The virulotyping of positive samples revealed the presence of all five virulence genes in B. melitensis. The virB, cbg, and om31 were detected in all five samples of B. abortus. In addition, zhuA and manA were detected in three (60%) and four (80%) samples, respectively, of the B. abortus-positive samples. Moreover, a total of 94.2% and 89.2% of the 14 positive samples were also found positive for manA and znuA, respectively. Our findings revealed that the Brucella spp. genomes can be detected in the SF of RA patients by the PCR-based method. We thus suggest that physicians should consider the Brucella spp. as indicators of potential RA for the timely diagnosis and treatment of RA. Copyright © 2018. Published by Elsevier Ltd.

  5. Population structure and virulence gene profiles of Streptococcus agalactiae collected from different hosts worldwide.

    Science.gov (United States)

    Morach, Marina; Stephan, Roger; Schmitt, Sarah; Ewers, Christa; Zschöck, Michael; Reyes-Velez, Julian; Gilli, Urs; Del Pilar Crespo-Ortiz, María; Crumlish, Margaret; Gunturu, Revathi; Daubenberger, Claudia A; Ip, Margaret; Regli, Walter; Johler, Sophia

    2018-03-01

    Streptococcus agalactiae is a leading cause of morbidity and mortality among neonates and causes severe infections in pregnant women and nonpregnant predisposed adults, in addition to various animal species worldwide. Still, information on the population structure of S. agalactiae and the geographical distribution of different clones is limited. Further data are urgently needed to identify particularly successful clones and obtain insights into possible routes of transmission within one host species and across species borders. We aimed to determine the population structure and virulence gene profiles of S. agalactiae strains from a diverse set of sources and geographical origins. To this end, 373 S. agalactiae isolates obtained from humans and animals from five different continents were typed by DNA microarray profiling. A total of 242 different S. agalactiae strains were identified and further analyzed. Particularly successful clonal lineages, hybridization patterns, and strains were identified that were spread across different continents and/or were present in more than one host species. In particular, several strains were detected in both humans and cattle, and several canine strains were also detected in samples from human, bovine, and porcine hosts. The findings of our study suggest that although S. agalactiae is well adapted to various hosts including humans, cattle, dogs, rodents, and fish, interspecies transmission is possible and occurs between humans and cows, dogs, and rabbits. The virulence and resistance gene profiles presented enable new insights into interspecies transmission and make a crucial contribution to the identification of suitable targets for therapeutic agents and vaccines.

  6. Comparisons of Salmonella conjugation and virulence gene hyperexpression mediated by rumen protozoa from domestic and exotic ruminants.

    Science.gov (United States)

    Brewer, Matt T; Xiong, Nalee; Dier, Jeffery D; Anderson, Kristi L; Rasmussen, Mark A; Franklin, Sharon K; Carlson, Steve A

    2011-08-05

    Recent studies have identified a phenomenon in which ciliated protozoa engulf Salmonella and the intra-protozoal environment hyperactivates virulence gene expression and provides a venue for conjugal transfer of antibiotic resistance plasmids. The former observation is relegated to Salmonella bearing the SGI1 multiresistance integron while the latter phenomenon appears to be a more generalized event for recipient Salmonella. Our previous studies have assessed virulence gene hyperexpression only with protozoa from the bovine rumen while conjugal transfer has been demonstrated in rumen protozoa from cattle and goats. The present study examined virulence gene hyperexpression for Salmonella exposed to rumen protozoa obtained from cattle, sheep, goats, or two African ruminants (giraffe and bongo). Conjugal transfer was also assessed in these protozoa using Salmonella as the recipient. Virulence gene hyperexpression was only observed following exposure to the rumen protozoa from cattle and sheep while elevated virulence was also observed in these animals. Conjugal transfer events were, however, observed in all protozoa evaluated. It therefore appears that the protozoa-based hypervirulence is not universal to all ruminants while conjugal transfer is more ubiquitous. Copyright © 2011 Elsevier B.V. All rights reserved.

  7. Virulence genes and genetic diversity of Streptococcus suis serotype 2 isolates from Thailand.

    Science.gov (United States)

    Maneerat, K; Yongkiettrakul, S; Kramomtong, I; Tongtawe, P; Tapchaisri, P; Luangsuk, P; Chaicumpa, W; Gottschalk, M; Srimanote, P

    2013-11-01

    Isolates of Streptococcus suis from different Western countries as well as those from China and Vietnam have been previously well characterized. So far, the genetic characteristics and relationship between S. suis strains isolated from both humans and pigs in Thailand are unknown. In this study, a total of 245 S. suis isolates were collected from both human cases (epidemic and sporadic) and pigs (diseased and asymptomatic) in Thailand. Bacterial strains were identified by biochemical tests and PCR targeting both, the 16S rRNA and gdh genes. Thirty-six isolates were identified as serotype 2 based on serotyping and the cps2-PCR. These isolates were tested for the presence of six virulence-associated genes: an arginine deiminase (arcA), a 38-kDa protein and protective antigen (bay046), an extracellular factor (epf), an hyaluronidase (hyl), a muramidase-released protein (mrp) and a suilysin (sly). In addition, the genetic diversities of these isolates were studied by RAPD PCR and multilocus sequence typing (MLST) analysis. Four virulence-associated gene patterns (VAGP 1 to 4) were obtained, and the majority of isolates (32/36) carried all genes tested (VAGP1). Each of the three OPB primers used provided 4 patterns designated RAPD-A to RAPD-D. Furthermore, MLST analysis could also distinguish the 36 isolates into four sequence types (STs): ST1 (n = 32), ST104 (n = 2), ST233 (n = 1) and a newly identified ST, ST336 (n = 1). Dendrogram constructions based on RAPD patterns indicated that S. suis serotype 2 isolates from Thailand could be divided into four groups and that the characteristics of the individual groups were in complete agreement with the virulence gene profiles and STs. The majority (32/36) of isolates recovered from diseased pigs, slaughterhouse pigs or human patients could be classified into a single group (VAGP1, RAPD-A and ST1). This genetic information strongly suggests the transmission of S. suis isolates from pigs to humans in Thailand. Our findings are

  8. Inactivation and Gene Expression of a Virulent WastewaterEscherichia coliStrain and the Nonvirulent CommensalEscherichia coliDSM1103 Strain upon Solar Irradiation

    KAUST Repository

    Aljassim, Nada I.; Mantilla-Calderon, David; Wang, Tiannyu; Hong, Pei-Ying

    2017-01-01

    This study examined the decay kinetics and molecular responses of two Escherichia coli strains upon solar irradiation. The first is E. coli PI-7, a virulent and antibiotic-resistant strain that was isolated from wastewater and carries the emerging NDM-1 antibiotic resistance gene. The other strain, E. coli DSM1103, displayed lower virulence and antibiotic resistance than E. coli PI-7. In a buffer solution, E. coli PI-7 displayed a longer lag phase prior to decay and a longer half-life compared with E. coli DSM1103 (6.64 ± 0.63 h and 2.85 ± 0.46 min vs 1.33 ± 0.52 h and 2.04 ± 0.36 min). In wastewater, both E. coli strains decayed slower than they did in buffer. Although solar irradiation remained effective in reducing the numbers of both strains by more than 5-log10 in <24 h, comparative genomics and transcriptomics revealed differences in the genomes and overall regulation of genes between the two E. coli strains. A wider arsenal of genes related to oxidative stress, cellular repair and protective mechanisms were upregulated in E. coli PI-7. Subpopulations of E. coli PI-7 expressed genes related to dormancy and persister cell formation during the late decay phase, which may have accounted for its prolonged persistence. Upon prolonged solar irradiation, both E. coli strains displayed upregulation of genes related to horizontal gene transfer and antibiotic resistance. Virulence functions unique to E. coli PI-7 were also upregulated. Our findings collectively indicated that, whereas solar irradiation is able to reduce total cell numbers, viable E. coli remained and expressed genes that enable survival despite solar treatment. There remains a need for heightened levels of concern regarding risks arising from the dissemination of E. coli that may remain viable in wastewater after solar irradiation.

  9. Inactivation and Gene Expression of a Virulent WastewaterEscherichia coliStrain and the Nonvirulent CommensalEscherichia coliDSM1103 Strain upon Solar Irradiation

    KAUST Repository

    Aljassim, Nada I.

    2017-03-06

    This study examined the decay kinetics and molecular responses of two Escherichia coli strains upon solar irradiation. The first is E. coli PI-7, a virulent and antibiotic-resistant strain that was isolated from wastewater and carries the emerging NDM-1 antibiotic resistance gene. The other strain, E. coli DSM1103, displayed lower virulence and antibiotic resistance than E. coli PI-7. In a buffer solution, E. coli PI-7 displayed a longer lag phase prior to decay and a longer half-life compared with E. coli DSM1103 (6.64 ± 0.63 h and 2.85 ± 0.46 min vs 1.33 ± 0.52 h and 2.04 ± 0.36 min). In wastewater, both E. coli strains decayed slower than they did in buffer. Although solar irradiation remained effective in reducing the numbers of both strains by more than 5-log10 in <24 h, comparative genomics and transcriptomics revealed differences in the genomes and overall regulation of genes between the two E. coli strains. A wider arsenal of genes related to oxidative stress, cellular repair and protective mechanisms were upregulated in E. coli PI-7. Subpopulations of E. coli PI-7 expressed genes related to dormancy and persister cell formation during the late decay phase, which may have accounted for its prolonged persistence. Upon prolonged solar irradiation, both E. coli strains displayed upregulation of genes related to horizontal gene transfer and antibiotic resistance. Virulence functions unique to E. coli PI-7 were also upregulated. Our findings collectively indicated that, whereas solar irradiation is able to reduce total cell numbers, viable E. coli remained and expressed genes that enable survival despite solar treatment. There remains a need for heightened levels of concern regarding risks arising from the dissemination of E. coli that may remain viable in wastewater after solar irradiation.

  10. Distribution of genes encoding virulence factors and molecular analysis of Shigella spp. isolated from patients with diarrhea in Kerman, Iran.

    Science.gov (United States)

    Hosseini Nave, Hossein; Mansouri, Shahla; Emaneini, Mohammad; Moradi, Mohammad

    2016-03-01

    Shigella is one of the important causes of diarrhea worldwide. Shigella has several virulence factors contributing in colonization and invasion of epithelial cells and eventually death of host cells. The present study was performed in order to investigate the distribution of virulence factors genes in Shigella spp. isolated from patients with acute diarrhea in Kerman, Iran as well as the genetic relationship of these isolates. A total of 56 isolates including 31 S. flexneri, 18 S. sonnei and 7 S. boydii were evaluated by polymerase chain reaction (PCR) for the presence of 11 virulence genes (ipaH, ial, set1A, set1B, sen, virF, invE, sat, sigA, pic and sepA). Then, the clonal relationship of these strains was analyzed by multilocus variable-number tandem repeat analysis (MLVA) method. All isolates were positive for ipaH gene. The other genes include ial, invE and virF were found in 80.4%, 60.7% and 67.9% of the isolates, respectively. Both set1A and set1B were detected in 32.3% of S. flexneri isolates, whereas 66.1% of the isolates belonging to different serogroup carried sen gene. The sat gene was present in all S. flexneri isolates, but not in the S. sonnei and S. boydii isolates. The result showed, 30.4% of isolates were simultaneously positive and the rest of the isolates were negative for sepA and pic genes. The Shigella isolates were divided into 29 MLVA types. This study, for the first time, investigated distribution of 11 virulence genes in Shigella spp. Our results revealed heterogeneity of virulence genes in different Shigella serogroups. Furthermore, the strains belonging to the same species had little diversity. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Carbon storage regulator A contributes to the virulence of Haemophilus ducreyi in humans by multiple mechanisms.

    Science.gov (United States)

    Gangaiah, Dharanesh; Li, Wei; Fortney, Kate R; Janowicz, Diane M; Ellinger, Sheila; Zwickl, Beth; Katz, Barry P; Spinola, Stanley M

    2013-02-01

    The carbon storage regulator A (CsrA) controls a wide variety of bacterial processes, including metabolism, adherence, stress responses, and virulence. Haemophilus ducreyi, the causative agent of chancroid, harbors a homolog of csrA. Here, we generated an unmarked, in-frame deletion mutant of csrA to assess its contribution to H. ducreyi pathogenesis. In human inoculation experiments, the csrA mutant was partially attenuated for pustule formation compared to its parent. Deletion of csrA resulted in decreased adherence of H. ducreyi to human foreskin fibroblasts (HFF); Flp1 and Flp2, the determinants of H. ducreyi adherence to HFF cells, were downregulated in the csrA mutant. Compared to its parent, the csrA mutant had a significantly reduced ability to tolerate oxidative stress and heat shock. The enhanced sensitivity of the mutant to oxidative stress was more pronounced in bacteria grown to stationary phase compared to that in bacteria grown to mid-log phase. The csrA mutant also had a significant survival defect within human macrophages when the bacteria were grown to stationary phase but not to mid-log phase. Complementation in trans partially or fully restored the mutant phenotypes. These data suggest that CsrA contributes to virulence by multiple mechanisms and that these contributions may be more profound in bacterial cell populations that are not rapidly dividing in the human host.

  12. Exploring new roles for the rpoS gene in the survival and virulence of the fire blight pathogen Erwinia amylovora.

    Science.gov (United States)

    Santander, Ricardo D; Monte-Serrano, Mercedes; Rodríguez-Herva, José J; López-Solanilla, Emilia; Rodríguez-Palenzuela, Pablo; Biosca, Elena G

    2014-12-01

    Erwinia amylovora causes fire blight in economically important plants of the family Rosaceae. This bacterial pathogen spends part of its life cycle coping with starvation and other fluctuating environmental conditions. In many Gram-negative bacteria, starvation and other stress responses are regulated by the sigma factor RpoS. We obtained an E. amylovora rpoS mutant to explore the role of this gene in starvation responses and its potential implication in other processes not yet studied in this pathogen. Results showed that E. amylovora needs rpoS to develop normal starvation survival and viable but nonculturable (VBNC) responses. Furthermore, this gene contributed to stationary phase cross-protection against oxidative, osmotic, and acid stresses and was essential for cross-protection against heat shock, but nonessential against acid shock. RpoS also mediated regulation of motility, exopolysaccharide synthesis, and virulence in immature loquats, but not in pear plantlets, and contributed to E. amylovora survival in nonhost tissues during incompatible interactions. Our results reveal some unique roles for the rpoS gene in E. amylovora and provide new knowledge on the regulation of different processes related to its ecology, including survival in different environments and virulence in immature fruits. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  13. The MAP kinase-activated protein kinase Rck2p regulates cellular responses to cell wall stresses, filamentation and virulence in the human fungal pathogen Candida albicans.

    Science.gov (United States)

    Li, Xichuan; Du, Wei; Zhao, Jingwen; Zhang, Lilin; Zhu, Zhiyan; Jiang, Linghuo

    2010-06-01

    Rck2p is the Hog1p-MAP kinase-activated protein kinase required for the attenuation of protein synthesis in response to an osmotic challenge in Saccharomyces cerevisiae. Rck2p also regulates rapamycin sensitivity in both S. cerevisiae and Candida albicans. In this study, we demonstrate that the deletion of CaRCK2 renders C. albicans cells sensitive to, and CaRck2p translocates from the cytosol to the nucleus in response to, cell wall stresses caused by Congo red, Calcoflor White, elevated heat and zymolyase. However, the kinase activity of CaRck2p is not required for the cellular response to these cell wall stresses. Furthermore, transcripts of cell wall protein-encoding genes CaBGL2, CaHWP1 and CaXOG1 are reduced in C. albicans cells lacking CaRCK2. The deletion of CaRCK2 also reduces the in vitro filamentation of C. albicans and its virulence in a mouse model of systemic candidasis. The kinase activity of CaRck2p is required for the virulence, but not for the in vitro filamentation, in C. albicans. Therefore, Rck2p regulates cellular responses to cell wall stresses, filamentation and virulence in the human fungal pathogen C. albicans.

  14. Catecholamines promote the expression of virulence and oxidative stress genes in Porphyromonas gingivalis.

    Science.gov (United States)

    Graziano, T S; Closs, P; Poppi, T; Franco, G C; Cortelli, J R; Groppo, F C; Cogo, K

    2014-10-01

    Stress has been identified as an important risk factor in the development of many infectious diseases, including periodontitis. Porphyromonas gingivalis, a gram-negative oral anaerobic bacterium, is considered an important pathogen in chronic periodontitis. Microorganisms, including P. gingivalis, that participate in infectious diseases have been shown to respond to catecholamines released during stress processes by modifying their growth and virulence. Therefore, the purpose of this study was to evaluate the effects of adrenaline and noradrenaline on the growth, antimicrobial susceptibility and gene expression in P. gingivalis. P. gingivalis was incubated in the presence of adrenaline and noradrenaline (100 μm) for different time-periods in rich (Tryptic soy broth supplemented with 0.2% yeast extract, 5 μg/mL of hemin and 1 μg/mL of menadione) and poor (serum-SAPI minimal medium and serum-SAPI minimal medium supplemented with 5 μg/mL of hemin and 1 μg/mL of menadione) media, and growth was evaluated based on absorbance at 660 nm. Bacterial susceptibility to metronidazole was examined after exposure to adrenaline and noradrenaline. The expression of genes involved in iron acquisition, stress oxidative protection and virulence were also evaluated using RT-quantitative PCR. Catecholamines did not interfere with the growth of P. gingivalis, regardless of nutritional or hemin conditions. In addition, bacterial susceptibility to metronidazole was not modified by exposure to adrenaline or noradrenaline. However, the expression of genes related to iron acquisition (hmuR), oxidative stress (tpx, oxyR, dps, sodB and aphC) and pathogenesis (hem, hagA and ragA) were stimulated upon exposure to adrenaline and/or noradrenaline. Adrenaline and noradrenaline can induce changes in gene expression related to oxidative stress and virulence factors in P. gingivalis. The present study is, in part, a step toward understanding the stress-pathogen interactions that may

  15. MiR-155 induction by F. novicida but not the virulent F. tularensis results in SHIP down-regulation and enhanced pro-inflammatory cytokine response.

    Directory of Open Access Journals (Sweden)

    Thomas J Cremer

    2009-12-01

    Full Text Available The intracellular gram-negative bacterium Francisella tularensis causes the disease tularemia and is known for its ability to subvert host immune responses. Previous work from our laboratory identified the PI3K/Akt pathway and SHIP as critical modulators of host resistance to Francisella. Here, we show that SHIP expression is strongly down-regulated in monocytes and macrophages following infection with F. tularensis novicida (F.n.. To account for this negative regulation we explored the possibility that microRNAs (miRs that target SHIP may be induced during infection. There is one miR that is predicted to target SHIP, miR-155. We tested for induction and found that F.n. induced miR-155 both in primary monocytes/macrophages and in vivo. Using luciferase reporter assays we confirmed that miR-155 led to down-regulation of SHIP, showing that it specifically targets the SHIP 3'UTR. Further experiments showed that miR-155 and BIC, the gene that encodes miR-155, were induced as early as four hours post-infection in primary human monocytes. This expression was dependent on TLR2/MyD88 and did not require inflammasome activation. Importantly, miR-155 positively regulated pro-inflammatory cytokine release in human monocytes infected with Francisella. In sharp contrast, we found that the highly virulent type A SCHU S4 strain of Francisella tularensis (F.t. led to a significantly lower miR-155 response than the less virulent F.n. Hence, F.n. induces miR-155 expression and leads to down-regulation of SHIP, resulting in enhanced pro-inflammatory responses. However, impaired miR-155 induction by SCHU S4 may help explain the lack of both SHIP down-regulation and pro-inflammatory response and may account for the virulence of Type A Francisella.

  16. Prevalence, Virulence Genes and Antimicrobial Resistance Profiles of Salmonella Serovars from Retail Beef in Selangor, Malaysia

    Directory of Open Access Journals (Sweden)

    Tze Y. Thung

    2018-01-01

    Full Text Available The aim of the present study was to investigate the prevalence of Salmonella spp., Salmonella Enteritidis and Salmonella Typhimurium in retail beef from different retail markets of Selangor area, as well as, to assess their pathogenic potential and antimicrobial resistance. A total of 240 retail beef meat samples (chuck = 60; rib = 60; round = 60; sirloin = 60 were randomly collected. The multiplex polymerase chain reaction (mPCR in combination with the most probable number (MPN method was employed to detect Salmonella spp., S. Enteritidis and S. Typhimurium in the meat samples. The prevalence of Salmonella spp., S. Enteritidis and S. Typhimurium in 240 beef meat samples were 7.50, 1.25, and 0.83%, respectively. The microbial loads of total Salmonella was found in the range of <3 to 15 MPN/g. Eight different serovars of Salmonella were identified among the 23 isolates, and S. Agona was the predominant serovar (26.09%. Interestingly, all the Salmonella isolates were resistant to penicillin, erythromycin and vancomycin, but the sensitivity was observed for tetracycline, gentamicin and amoxicillin/clavulanic acid. All 23 isolates were resistant to at least three antibiotics. Two S. Typhimurium isolates (8.70% exhibited the highest multiple antibiotic resistance (MAR index value of 0.56 which shown resistance to nine antibiotics. PCR analysis of virulence genes showed that all Salmonella isolates (100% were positive for the invA gene. Meanwhile, pefA was only identified in S. Enteritidis and S. Typhimurium. The findings in this study indicate that retail beef products tested were widely contaminated with multi-drug resistant (MDR Salmonella and various virulence genes are present among the isolated Salmonella serovars.

  17. Relationship between oviposition, virulence gene expression and parasitism success in Cotesia typhae nov. sp. parasitoid strains.

    Science.gov (United States)

    Benoist, R; Chantre, C; Capdevielle-Dulac, C; Bodet, M; Mougel, F; Calatayud, P A; Dupas, S; Huguet, E; Jeannette, R; Obonyo, J; Odorico, C; Silvain, J F; Le Ru, B; Kaiser, L

    2017-12-01

    Studying mechanisms that drive host adaptation in parasitoids is crucial for the efficient use of parasitoids in biocontrol programs. Cotesia typhae nov. sp. (Fernández-Triana) (Hymenoptera: Braconidae) is a newly described parasitoid of the Mediterranean corn borer Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae). Braconidae are known for their domesticated bracovirus, which is injected with eggs in the host larva to overcome its resistance. In this context, we compared reproductive success traits of four Kenyan strains of C. typhae on a French and a Kenyan populations of its host. Differences were found between the four strains and the two most contrasted ones were studied more thoroughly on the French host population. Parasitoid offspring size was correlated with parasitism success and the expression of bracovirus virulence genes (CrV1 and Cystatin) in the host larva after parasitism. Hybrids between these two parasitoid strains showed phenotype and gene expression profiles similar to the most successful parental strain, suggesting the involvement of dominant alleles in the reproductive traits. Ovary dissections revealed that the most successful strain injected more eggs in a single host larva than the less successful one, despite an equal initial ovocyte number in ovaries. It can be expected that the amount of viral particles increase with the number of eggs injected. The ability to bypass the resistance of the allopatric host may in consequence be related to the oviposition behaviour (eggs allocation). The influence of the number of injected eggs on parasitism success and on virulence gene expression was evaluated by oviposition interruption experiments.

  18. Occurrence of Putative Virulence Genes in Arcobacter Species Isolated from Humans and Animals

    Science.gov (United States)

    Douidah, Laid; de Zutter, Lieven; Baré, Julie; De Vos, Paul; Vandamme, Peter; Vandenberg, Olivier; Van den Abeele, Anne-Marie

    2012-01-01

    Interest in arcobacters in veterinary and human public health has increased since the first report of the isolation of arcobacters from food of animal origin. Since then, studies worldwide have reported the occurrence of arcobacters on food and in food production animals and have highlighted possible transmission, especially of Arcobacter butzleri, to the human population. In humans, arcobacters are associated with enteritis and septicemia. To assess their clinical relevance for humans and animals, evaluation of potential virulence factors is required. However, up to now, little has been known about the mechanisms of pathogenicity. Because of their close phylogenetic affiliation to the food-borne pathogen Campylobacter and their similar clinical manifestations, the presence of nine putative Campylobacter virulence genes (cadF, ciaB, cj1349, hecA, hecB, irgA, mviN, pldA, and tlyA) previously identified in the recent Arcobacter butzleri ATCC 49616 genome sequence was determined in a large set of human and animal Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii strains after the development of rapid and accurate PCR assays and confirmed by sequencing and dot blot hybridization. PMID:22170914

  19. Phenotypic and genotypic antimicrobial resistance and virulence genes of Salmonella enterica isolated from pet dogs and cats

    Science.gov (United States)

    Srisanga, Songsak; Angkititrakul, Sunpetch; Sringam, Patcharee; Le Ho, Phuong T.; Vo, An T. T.

    2017-01-01

    Salmonella enterica isolates (n = 122), including 32 serotypes from 113 dogs and 9 cats, were obtained from household dogs (n = 250) and cats (n = 50) during 2012–2015. The isolates were characterized by serotyping, antimicrobial resistance phenotyping and genotyping, and virulence gene screening. Serovars Weltevreden (15.6%) and Typhimurium (13.9%) were the most common. The majority (43%) of the isolates were multidrug resistant. The dog isolates (12.3%) harbored class 1 integrons, of which the dfrA12-aadA2 cassette was most frequent (66.7%). The only class integron in serovar Albany was located on a conjugative plasmid. Two ESBL-producing isolates (i.e., a serovar Krefeld and a serovar Enteritridis) carried blaTEM and blaCTX-M, and the blaTEM gene in both was horizontally transferred. Of the plasmid-mediated quinolone resistance genes tested, only qnrS (4.9%) was detected. Most Salmonella isolates harbored invA (100%), prgH (91.8%), and sipB (91%). Positive associations between resistance and virulence genes were observed for blaPSE-1/orgA, cmlA/spaN, tolC, and sul1/tolC (p resistance and virulence genes and that antimicrobial use in companion animals may select for the examined Salmonella virulence factors. PMID:27586467

  20. New Insight into Biofilm Formation Ability, the Presence of Virulence Genes and Probiotic Potential of Enterococcus sp. Dairy Isolates

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    Nikola Popović

    2018-01-01

    Full Text Available Enterococci have controversial status due to their emerging role in nosocomial infections and transmission of antibiotic resistance genes, while some enterococci strains are used as probiotics for humans and animals and starter cultures in dairy industry. In order to improve our understanding of factors involved in the safe use of enterococci as potential probiotics, the antibiotic susceptibility, virulence and probiotic traits of 75 dairy enterococci isolates belonging to Enterococcus durans (50, En. faecium (15, En. faecalis (6, En. italicus (3, and En. hirae (1 were evaluated. The results revealed that ciprofloxacin resistance and biofilm formation are correlated with isolates originated from Golija mountain (Serbia, while gelatinase activity was more common in isolates from Prigorje region (Croatia, pointing to uncontrolled use of antibiotics and anthropogenic impact on dairy products' microbiota in these regions. The virulence genes were sporadically present in 13 selected dairy enterococci isolates. Interestingly, biofilm formation was correlated with higher ability of strains to reduce the adhesion of E. coli and Salmonella Enteritidis to HT29-MTX cells. To our knowledge this is the first study reporting the presence of the esp gene (previously correlated with pathogenesis in dairy enterococci isolates, mostly associated with the genes involved in adhesion property. Hence, the results of this study revealed that the virulence genes are sporadically present in dairy isolates and more correlated to adhesion properties and biofilm formation, implicating their role in gut colonization rather than to the virulence traits.

  1. Phylogenetic diversity, antimicrobial susceptibility and virulence gene profiles of Brachyspira hyodysenteriae isolates from pigs in Germany.

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    Jessica Joerling

    Full Text Available Swine dysentery (SD is an economically important diarrheal disease in pigs caused by different strongly hemolytic Brachyspira (B. species, such as B. hyodysenteriae, B. suanatina and B. hampsonii. Possible associations of epidemiologic data, such as multilocus sequence types (STs to virulence gene profiles and antimicrobial susceptibility are rather scarce, particularly for B. hyodysenteriae isolates from Germany. In this study, B. hyodysenteriae (n = 116 isolated from diarrheic pigs between 1990 and 2016 in Germany were investigated for their STs, susceptibility to the major drugs used for treatment of SD (tiamulin and valnemulin and genes that were previously linked with virulence and encode for hemolysins (tlyA, tlyB, tlyC, hlyA, BHWA1_RS02885, BHWA1_RS09085, BHWA1_RS04705, and BHWA1_RS02195, outer membrane proteins (OMPs (bhlp16, bhlp17.6, bhlp29.7, bhmp39f, and bhmp39h as well as iron acquisition factors (ftnA and bitC. Multilocus sequence typing (MLST revealed that 79.4% of the isolates belonged to only three STs, namely ST52 (41.4%, ST8 (12.1%, and ST112 (25.9% which have been observed in other European countries before. Another 24 isolates belonged to twelve new STs (ST113-118, ST120-123, ST131, and ST193. The temporal distribution of STs revealed the presence of new STs as well as the regular presence of ST52 over three decades (1990s-2000s. The proportion of strains that showed resistance to both tiamulin und valnemulin (39.1% varied considerably among the most frequent STs ranging from 0% (0/14 isolates resistant in ST8 isolates to 46.7% (14/30, 52.1% (25/48, and 85.7% (6/7 in isolates belonging to ST112, ST52, and ST114, respectively. All hemolysin genes as well as the iron-related gene ftnA and the OMP gene bhlp29.7 were regularly present in the isolates, while the OMP genes bhlp17.6 and bhmp39h could not be detected. Sequence analysis of hemolysin genes of selected isolates revealed co-evolution of tlyB, BHWA1_RS02885, BHWA1_RS

  2. Phylogenetic diversity, antimicrobial susceptibility and virulence gene profiles of Brachyspira hyodysenteriae isolates from pigs in Germany

    Science.gov (United States)

    Joerling, Jessica; Barth, Stefanie A.; Schlez, Karen; Willems, Hermann

    2018-01-01

    Swine dysentery (SD) is an economically important diarrheal disease in pigs caused by different strongly hemolytic Brachyspira (B.) species, such as B. hyodysenteriae, B. suanatina and B. hampsonii. Possible associations of epidemiologic data, such as multilocus sequence types (STs) to virulence gene profiles and antimicrobial susceptibility are rather scarce, particularly for B. hyodysenteriae isolates from Germany. In this study, B. hyodysenteriae (n = 116) isolated from diarrheic pigs between 1990 and 2016 in Germany were investigated for their STs, susceptibility to the major drugs used for treatment of SD (tiamulin and valnemulin) and genes that were previously linked with virulence and encode for hemolysins (tlyA, tlyB, tlyC, hlyA, BHWA1_RS02885, BHWA1_RS09085, BHWA1_RS04705, and BHWA1_RS02195), outer membrane proteins (OMPs) (bhlp16, bhlp17.6, bhlp29.7, bhmp39f, and bhmp39h) as well as iron acquisition factors (ftnA and bitC). Multilocus sequence typing (MLST) revealed that 79.4% of the isolates belonged to only three STs, namely ST52 (41.4%), ST8 (12.1%), and ST112 (25.9%) which have been observed in other European countries before. Another 24 isolates belonged to twelve new STs (ST113-118, ST120-123, ST131, and ST193). The temporal distribution of STs revealed the presence of new STs as well as the regular presence of ST52 over three decades (1990s–2000s). The proportion of strains that showed resistance to both tiamulin und valnemulin (39.1%) varied considerably among the most frequent STs ranging from 0% (0/14 isolates resistant) in ST8 isolates to 46.7% (14/30), 52.1% (25/48), and 85.7% (6/7) in isolates belonging to ST112, ST52, and ST114, respectively. All hemolysin genes as well as the iron-related gene ftnA and the OMP gene bhlp29.7 were regularly present in the isolates, while the OMP genes bhlp17.6 and bhmp39h could not be detected. Sequence analysis of hemolysin genes of selected isolates revealed co-evolution of tlyB, BHWA1_RS02885, BHWA1_RS

  3. Antimicrobial susceptibility, virulence genes, and randomly amplified polymorphic DNA analysis of Staphylococcus aureus recovered from bovine mastitis in Ningxia, China.

    Science.gov (United States)

    Wang, Dong; Zhang, Limei; Zhou, Xuezhang; He, Yulong; Yong, Changfu; Shen, Mingliang; Szenci, Otto; Han, Bo

    2016-12-01

    Staphylococcus aureusis the leading pathogen involved inbovine mastitis, but knowledgeabout antimicrobial resistance, virulence factors, and genotypes of Staphylococcus aureus resulting in bovine mastitis in Ningxia, China, is limited. Therefore, antimicrobial susceptibility, virulence gene, and randomly amplified polymorphic DNA (RAPD) analyses of Staph. aureus were carried out. A total of 327 milk samples from cows with clinical and subclinical mastitis in 4 regions of Ningxia were used for the isolation and identification of pathogens according to phenotypic and molecular characteristics. Antimicrobial susceptibility against 22 antimicrobial agents was determined by disk diffusion. The presence of 8 virulence genes in Staph. aureus isolates was tested by PCR. Genotypes of isolates were investigated based on RAPD. Results showed that 35 isolates obtained from mastitis milk samples were identified as Staph. aureus. The isolates were resistant to sulfamethoxazole (100%), penicillin G (94.3%), ampicillin (94.3%), erythromycin (68.6%), azithromycin (68.6%), clindamycin (25.7%), amoxicillin (11.4%), and tetracycline (5.7%). All of the isolates contained one or more virulence genes with average (standard deviation) of 6.6±1.6. The most prevalent virulence genes were hlb (97.1%), followed by fnbpA, hla, coa (94.3% each), nuc (85.7%), fnbpB (80%), clfA (77.1%), and tsst-1 (40%). Nine different gene patterns were found and 3 of them were the dominant gene combinations (77.1%). Staphylococcus aureus isolates (n=35) were divided into 6 genotypes by RAPD tying, the genotypes III and VI were the most prevalent genotypes. There was greatvariation in genotypes of Staph. aureus isolates, not only among different farms, but also within the same herd in Ningxia province. The study showed a high incidence of Staph. aureus with genomic variation of resistance genes, which is matter of great concern in public and animal health in Ningxia province of China. Copyright © 2016 American

  4. Intermediate filaments and gene regulation.

    Science.gov (United States)

    Traub, P

    1995-01-01

    The biological role of intermediate filaments (IFs) of eukaryotic cells is still a matter of conjecture. On the basis of immunofluorescence and electron microscopic observations, they appear to play a cytoskeletal role in that they stabilize cellular structure and organize the distribution and interactions of intracellular organelles and components. The expression of a large number of cell type-specific and developmentally regulated subunit proteins is believed to provide multicellular organisms with different IF systems capable of differential interactions with the various substructures and components of their multiple, differentiated cells. However, the destruction of distinct IF systems by manipulation of cultured cells or by knock-out mutation of IF subunit proteins in transgenic mice exerts relatively little influence on cellular morphology and physiology and on development of mutant animals. In order to rationalize this dilemma, the cytoskeletal concept of IF function has been extended to purport that cytoplasmic (c) IFs and their subunit proteins also play fundamental roles in gene regulation. It is based on the in vitro capacity of cIF(protein)s to interact with guanine-rich, single-stranded DNA, supercoiled DNA and histones, as well as on their close structural relatedness to gene-regulatory DNA-binding and nuclear matrix proteins. Since cIF proteins do not possess classical nuclear localization signals, it is proposed that cIFs directly penetrate the double nuclear membrane, exploiting the amphiphilic, membrane-active character of their subunit proteins. Since they can establish metastable multisite contacts with nuclear matrix structures and/or chromatin areas containing highly repetitive DNA sequence elements at the nuclear periphery, they are supposed to participate in chromosome distribution and chromatin organization in interphase nuclei of differentiated cells. Owing to their different DNA-binding specificities, the various cIF systems may in this

  5. ChLae1 and ChVel1 regulate T-toxin production, virulence, oxidative stress response, and development of the maize pathogen Cochliobolus heterostrophus.

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    Dongliang Wu

    2012-02-01

    Full Text Available LaeA and VeA coordinate secondary metabolism and differentiation in response to light signals in Aspergillus spp. Their orthologs, ChLae1 and ChVel1, were identified in the maize pathogen Cochliobolus heterostrophus, known to produce a wealth of secondary metabolites, including the host selective toxin, T-toxin. Produced by race T, T-toxin promotes high virulence to maize carrying Texas male sterile cytoplasm (T-cms. T-toxin production is significantly increased in the dark in wild type (WT, whereas Chvel1 and Chlae1 mutant toxin levels are much reduced in the dark compared to WT. Correspondingly, expression of T-toxin biosynthetic genes (Tox1 is up-regulated in the dark in WT, while dark-induced expression is much reduced/minimal in Chvel1 and Chlae1 mutants. Toxin production and Tox1 gene expression are increased in ChVEL1 overexpression (OE strains grown in the dark and in ChLAE1 strains grown in either light or dark, compared to WT. These observations establish ChLae1 and ChVel1 as the first factors known to regulate host selective toxin production. Virulence of Chlae1 and Chvel1 mutants and OE strains is altered on both T-cms and normal cytoplasm maize, indicating that both T-toxin mediated super virulence and basic pathogenic ability are affected. Deletion of ChLAE1 or ChVEL1 reduces tolerance to H(2O(2. Expression of CAT3, one of the three catalase genes, is reduced in the Chvel1 mutant. Chlae1 and Chvel1 mutants also show decreased aerial hyphal growth, increased asexual sporulation and female sterility. ChLAE1 OE strains are female sterile, while ChVEL1 OE strains are more fertile than WT. ChLae1 and ChVel1 repress expression of 1,8-dihydroxynaphthalene (DHN melanin biosynthesis genes, and, accordingly, melanization is enhanced in Chlae1 and Chvel1 mutants, and reduced in OE strains. Thus, ChLae1 and ChVel1 positively regulate T-toxin biosynthesis, pathogenicity and super virulence, oxidative stress responses, sexual development, and

  6. The flagellar master operon flhDC is a pleiotropic regulator involved in motility and virulence of the fish pathogen Yersinia ruckeri.

    Science.gov (United States)

    Jozwick, A K S; Graf, J; Welch, T J

    2017-03-01

    To investigate the function of the master flagellar operon flhDC in the fish pathogen Yersinia ruckeri and compare the effect of a constructed flhD mutation to a naturally occurring fliR mutation causing loss-of-motility in emergent biotype 2 (BT2) strains. Yersinia ruckeri flhD and fliR mutants were constructed in a motile strain. Both mutations caused loss-of-motility, ablation of flagellin synthesis and phospholipase secretion, similar to naturally occurring BT2 strains. Transcriptome analysis confirmed flhDC regulation of flagellar, chemotaxis and phospholipase loci as well as other genes of diverse function. The flhD mutation confers a competitive advantage within the fish host when compared with its parent strain, while this advantage was not seen with the naturally occurring fliR mutation. An intact flhD is necessary for expression of the flagellar secretion system as well as other diverse loci, consistent with a role for flhD as a pleiotropic regulator. The maintenance of the flhD locus in Y. ruckeri strains suggests its importance for aspects of Y. ruckeri biology other than virulence, since the flhD mutation conferred a competitive advantage during experimental challenge of rainbow trout. Yersinia ruckeri is the causative agent of enteric red mouth disease, an invasive septicaemia that affects farmed salmonid fish species. Disease outbreaks can cause severe economic losses in aquaculture. BT2 variants, which have independently emerged worldwide, are an increasing threat to farmed fish production. Knowledge of mechanisms involved in virulence, conserved functions and gene regulation among strains may be exploited for the development of novel disease control strategies to prevent pathogen growth or virulence phenotypes within aquaculture. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  7. Prevalence, Virulence Genes and Antimicrobial Resistance Profiles of Salmonella Serovars from Retail Beef in Selangor, Malaysia.

    Science.gov (United States)

    Thung, Tze Y; Radu, Son; Mahyudin, Nor A; Rukayadi, Yaya; Zakaria, Zunita; Mazlan, Nurzafirah; Tan, Boon H; Lee, Epeng; Yeoh, Soo L; Chin, Yih Z; Tan, Chia W; Kuan, Chee H; Basri, Dayang F; Wan Mohamed Radzi, Che W J

    2017-01-01

    The aim of the present study was to investigate the prevalence of Salmonella spp., Salmonella Enteritidis and Salmonella Typhimurium in retail beef from different retail markets of Selangor area, as well as, to assess their pathogenic potential and antimicrobial resistance. A total of 240 retail beef meat samples (chuck = 60; rib = 60; round = 60; sirloin = 60) were randomly collected. The multiplex polymerase chain reaction (mPCR) in combination with the most probable number (MPN) method was employed to detect Salmonella spp., S . Enteritidis and S . Typhimurium in the meat samples. The prevalence of Salmonella spp., S . Enteritidis and S . Typhimurium in 240 beef meat samples were 7.50, 1.25, and 0.83%, respectively. The microbial loads of total Salmonella was found in the range of retail beef products tested were widely contaminated with multi-drug resistant (MDR) Salmonella and various virulence genes are present among the isolated Salmonella serovars.

  8. Frequency, virulence genes and antimicrobial resistance of Listeria spp. isolated from bovine clinical mastitis.

    Science.gov (United States)

    Jamali, Hossein; Radmehr, Behrad

    2013-11-01

    The aims of this study were to determine the prevalence, characteristics and antimicrobial resistance of Listeria spp. isolated from bovine clinical mastitis in Iran. Listeria spp. were detected in 21/207 bovine mastitic milk samples from dairy farms in Iran, comprising L. monocytogenes (n=17), L. innocua (n=3) and L. ivanovii (n=1). L. monocytogenes isolates were grouped into serogroups '4b, 4d, 4e', '1/2a, 3a', '1/2b, 3b, 7' and '1/2c, 3c'; all harboured inlA, inlC and inlJ virulence genes. Listeria spp. were most frequently resistant to penicillin G (14/21 isolates, 66.7%) and tetracyclines (11/21 isolates, 52.4%). Copyright © 2013 Elsevier Ltd. All rights reserved.

  9. Biofilm formation, antimicrobial susceptibility, serogroups and virulence genes of uropathogenic E. coli isolated from clinical samples in Iran

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    Elahe Tajbakhsh

    2016-04-01

    Full Text Available Abstract Background Uropathogenic Escherichia coli O- Serogroups with their virulence factors are the most prevalent causes of UTIs. The present research performed to track common uropathogenic E.coli serogroups, antibiotic resistance pattern of strains and prevalence of virulence genes in isolations having the ability to constitute biofilm. Methods In this research 130 E.coli isolation from patients having UTI symptoms were collected and antimicrobial resistance pattern was performed by Kirby-Bauer method. Polymerase chain reaction was done using primer pairs to identify common serogroups of uropathogenic E.coli and studying virulence genes in isolations creating biofilm. Results Among 130 E.coli isolates, 80 (61.53 % were able to make biofilm that 15 isolates (18.75 % indicated strong reaction, 20 (25 % of medium and 45 (56.25 % of weak biofilm reaction. Among isolations creating biofilm, the highest resistance reported to Ampicillin (87.5 % and the lowest to Nitrofurantoin (3.75 %. The frequency of fimH, pap, sfa and afa genes in isolations having the ability to create strong biofilm reported 93.33 %, 86.66 %, 86.66 % and 66.66 %, respectively. Conclusions The findings indicated the importance of virulence genes in serogroups producing uropathogenic E.coli biofilm. It is recommended that strains producing biofilm before antibiotic use should be studied.

  10. Hemolytic Porcine Intestinal Escherichia coli without Virulence-Associated Genes Typical of Intestinal Pathogenic E. coli ▿ †

    Science.gov (United States)

    Schierack, Peter; Weinreich, Joerg; Ewers, Christa; Tachu, Babila; Nicholson, Bryon; Barth, Stefanie

    2011-01-01

    Testing 1,666 fecal or intestinal samples from healthy and diarrheic pigs, we obtained hemolytic Escherichia coli isolates from 593 samples. Focusing on hemolytic E. coli isolates without virulence-associated genes (VAGs) typical for enteropathogens, we found that such isolates carried a broad variety of VAGs typical for extraintestinal pathogenic E. coli. PMID:21965399

  11. Detection of virulence genes determining the ability to adhere and invade in Campylobacter spp. from cattle and swine in Poland.

    Science.gov (United States)

    Wysok, Beata; Wojtacka, Joanna

    2018-02-01

    The aim of the study was to determine the prevalence of virulence genes responsible for the adhesion (flaA, cadF and racR) and invasion (virB11, iam and pldA) in Campylobacter isolates from cattle and swine and determine their adherence and invasion abilities. The studies conducted revealed high prevalence rate of adherence and invasion associated genes irrespective of the isolates origin. All Campylobacter strains of swine and cattle origin adhered to HeLa cells at mean level 0.1099% ± SD 0.1341% and 0.0845% ± SD 0.1304% of starting viable inoculum, respectively. However swine isolates exhibited higher invasion abilities (0.0012% ± SD 0.0011%) compared to bovine isolates (0.00038% ± SD 0.00055%). The results obtained revealed significantly positive correlation between invasion and adherence abilities of swine origin isolates (R = 0.4867 in regard to C. jejuni and R = 0.4507 in regard to C. coli) and bovine origin isolates (R = 0.726 in regard to C. jejuni). Bacterial virulence is multifactorial and it is affected by the expression of virulence genes. Moreover the presence of virulence genes determines the ability of Campylobacter isolates to adhere and invade the cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Iron-Regulated Phospholipase C Activity Contributes to the Cytolytic Activity and Virulence of Acinetobacter baumannii.

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    Steven E Fiester

    Full Text Available Acinetobacter baumannii is an opportunistic Gram-negative pathogen that causes a wide range of infections including pneumonia, septicemia, necrotizing fasciitis and severe wound and urinary tract infections. Analysis of A. baumannii representative strains grown in Chelex 100-treated medium for hemolytic activity demonstrated that this pathogen is increasingly hemolytic to sheep, human and horse erythrocytes, which interestingly contain increasing amounts of phosphatidylcholine in their membranes. Bioinformatic, genetic and functional analyses of 19 A. baumannii isolates showed that the genomes of each strain contained two phosphatidylcholine-specific phospholipase C (PC-PLC genes, which were named plc1 and plc2. Accordingly, all of these strains were significantly hemolytic to horse erythrocytes and their culture supernatants tested positive for PC-PLC activity. Further analyses showed that the transcriptional expression of plc1 and plc2 and the production of phospholipase and thus hemolytic activity increased when bacteria were cultured under iron-chelation as compared to iron-rich conditions. Testing of the A. baumannii ATCC 19606T plc1::aph-FRT and plc2::aph isogenic insertion derivatives showed that these mutants had a significantly reduced PC-PLC activity as compared to the parental strain, while testing of plc1::ermAM/plc2::aph demonstrated that this double PC-PLC isogenic mutant expressed significantly reduced cytolytic and hemolytic activity. Interestingly, only plc1 was shown to contribute significantly to A. baumannii virulence using the Galleria mellonella infection model. Taken together, our data demonstrate that both PLC1 and PLC2, which have diverged from a common ancestor, play a concerted role in hemolytic and cytolytic activities; although PLC1 seems to play a more critical role in the virulence of A. baumannii when tested in an invertebrate model. These activities would provide access to intracellular iron stores this pathogen

  13. The effect of γ radiation on the expression of the virulence genes of Salmonella typhimurium and Vibrio spp

    International Nuclear Information System (INIS)

    Lim, Sangyong; Jung, Jinwoo; Kim, Dongho

    2007-01-01

    The principle benefit of food irradiation is the reduction of food-borne bacteria in food products. However, the microbiological safety with respect to increased virulence of surviving pathogens after irradiation remains an important issue with regard to the effectiveness of food irradiation. In this study, the transcriptional changes of virulence genes of Salmonella and Vibrio spp. after γ radiation were investigated by real-time PCR (RT-PCR). Samonella typhimurium is dependent upon the products of a large number of genes located within Salmonella pathogenicity islands (SPI) on the chromosome. The expressions of seven genes including four SPI genes, hilD, ssrB, pipB, and sopD, were measured at 1 h after 1 kGy irradiation. Compared with non-irradiated controls, the expression of hilD encoded within SPI1 and sopD encoding SPI1-related effector proteins was reduced about 4- and 16-fold, respectively. The expressions of Vibrio toxin genes, vvhA, ctxA, and tdh, were also monitored during the course of a growth cycle after re-inoculation of irradiated Vibrio spp. (0.5 and 1.0 kGy). The expressions of Vibrio toxin genes tested did not increase compared with non-irradiated counterparts. Results from this study indicate that γ radiation is much more likely to reduce the virulence gene expression of surviving pathogens

  14. The effect of {gamma} radiation on the expression of the virulence genes of Salmonella typhimurium and Vibrio spp

    Energy Technology Data Exchange (ETDEWEB)

    Lim, Sangyong; Jung, Jinwoo [Radiation Food Science and Biotechnology Team, Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongup 580-185 (Korea, Republic of); Kim, Dongho [Radiation Food Science and Biotechnology Team, Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongup 580-185 (Korea, Republic of)], E-mail: fungikim@kaeri.re.kr

    2007-11-15

    The principle benefit of food irradiation is the reduction of food-borne bacteria in food products. However, the microbiological safety with respect to increased virulence of surviving pathogens after irradiation remains an important issue with regard to the effectiveness of food irradiation. In this study, the transcriptional changes of virulence genes of Salmonella and Vibrio spp. after {gamma} radiation were investigated by real-time PCR (RT-PCR). Samonella typhimurium is dependent upon the products of a large number of genes located within Salmonella pathogenicity islands (SPI) on the chromosome. The expressions of seven genes including four SPI genes, hilD, ssrB, pipB, and sopD, were measured at 1 h after 1 kGy irradiation. Compared with non-irradiated controls, the expression of hilD encoded within SPI1 and sopD encoding SPI1-related effector proteins was reduced about 4- and 16-fold, respectively. The expressions of Vibrio toxin genes, vvhA, ctxA, and tdh, were also monitored during the course of a growth cycle after re-inoculation of irradiated Vibrio spp. (0.5 and 1.0 kGy). The expressions of Vibrio toxin genes tested did not increase compared with non-irradiated counterparts. Results from this study indicate that {gamma} radiation is much more likely to reduce the virulence gene expression of surviving pathogens.

  15. Virulence Genes and Antimicrobial Resistance Profiles of Pasteurella multocida Strains Isolated from Rabbits in Brazil

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    Thais Sebastiana Porfida Ferreira

    2012-01-01

    Full Text Available Pasteurella multocida is responsible for a wide range of diseases in domestic animals. In rabbits, the agent is related to nasal discharge, pneumonia, otitis media, pyometra, orchitis, abscess, and septicemia. One hundred and forty rabbits with respiratory diseases from four rabbitries in São Paulo State, Brazil were evaluated for the detection of P. multocida in their nasal cavities. A total of twenty-nine animals were positive to P. multocida isolation, and 46 strains were selected and characterized by means of biochemical tests and PCR. P. multocida strains were tested for capsular type, virulence genes, and resistance profile. A total of 45.6% (21/46 of isolates belonged to capsular type A, and 54.34% (25/46 of the isolates were untypeable. None of the strains harboured toxA or pfhA genes. The frequency of the other twenty genes tested was variable, and the data generated was used to build a dendrogram, showing the relatedness of strains, which were clustered according to origin. Resistance revealed to be more common against sulfonamides and cotrimoxazole, followed by erythromycin, penicillin, and amoxicillin.

  16. Effect of co-culture with enterocinogenic E. faecium on L. monocytogenes key virulence gene expression

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    Eleftherios H. Drosinos

    2016-08-01

    Full Text Available The aim of the present study was to assess the expression of key virulence genes during co-culture of L. monocytogenes with a bacteriocinogenic E. faecium strain in liquid growth medium. For that purpose, BHI broth was inoculated with 7 log CFU·mL–1 L. monocytogenes and 4, 5 or 6 log CFU·mL–1 E. faecium. Sampling took place after 8 and 24 h of incubation, corresponding to the maximum and minimum of enterocin production, respectively. The RNA was extracted, stabilized and expression of prfA, sigB, hly, plcA, plcB, inlA, inlB, inlC and inlJ, was assessed by RT-qPCR. Most of the genes were downregulated during co-culture at 5 °C. Moreover, a statistically significant effect of the inoculum level was evident in most of the cases. On the contrary, no effect on the transcription level of most of the genes was observed during co-culture at 37 °C.

  17. Comparative genomic analysis of Brucella abortus vaccine strain 104M reveals a set of candidate genes associated with its virulence attenuation.

    Science.gov (United States)

    Yu, Dong; Hui, Yiming; Zai, Xiaodong; Xu, Junjie; Liang, Long; Wang, Bingxiang; Yue, Junjie; Li, Shanhu

    2015-01-01

    The Brucella abortus strain 104M, a spontaneously attenuated strain, has been used as a vaccine strain in humans against brucellosis for 6 decades in China. Despite many studies, the molecular mechanisms that cause the attenuation are still unclear. Here, we determined the whole-genome sequence of 104M and conducted a comprehensive comparative analysis against the whole genome sequences of the virulent strain, A13334, and other reference strains. This analysis revealed a highly similar genome structure between 104M and A13334. The further comparative genomic analysis between 104M and A13334 revealed a set of genes missing in 104M. Some of these genes were identified to be directly or indirectly associated with virulence. Similarly, a set of mutations in the virulence-related genes was also identified, which may be related to virulence alteration. This study provides a set of candidate genes associated with virulence attenuation in B.abortus vaccine strain 104M.

  18. Genotypes, Virulence Factors and Antimicrobial Resistance Genes of Staphylococcus aureus Isolated in Bovine Subclinical Mastitis from Eastern China

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    Javed Memon§, Yongchun Yang§, Jam Kashifa, Muhammad Yaqoob, Rehana Buriroa, Jamila Soomroa, Wang Liping and Fan Hongjie*

    2013-11-01

    Full Text Available This study was carried out to determine the genotypes, virulence factors and antimicrobial resistance traits of 34 Staphylococcus aureus isolated from subclinical mastitis in Eastern China. Minimal inhibitory concentration (MIC results showed resistance to erythromycin in all isolates. A high frequency of Methicillin resistant S. aureus (MRSA; 29% was observed and these isolates were also highly resistant to penicillin, oxacillin, oxytetracycline and chloramphenicol than methicillin sensitive S. aureus (MSSA isolates. Thirteen pathogenic factors and seven resistance genes including mecA and blaZ gene were checked through PCR. The spaX gene was found in all isolates, whereas cna, spaIg, nuc, clfA, fnbpB, hlA, hlB and seA were present in 35, 79, 85, 59, 35, 85, 71 and 38% isolates, respectively. Nine isolates carried a group of 8 different virulence genes. Moreover, macrolide resistance genes ermB and ermC were present in all isolates. High resistance rate against methicillin was found but no isolate was positive for mecA gene, whereas blaZ and tetK were detected in 82 and 56% isolates, respectively. Genes; fnbpA, seB, seC, seD, dfrK and tetM were not found in any isolate. The statistical association between phenotypic resistance and virulence genes showed, clfA, fnbpB, hlB and seA, were potentially associated with penicillin G, ciprofloxacin, methicillin, chloramphenicol, trimethoprim and oxytetracycline resistance (P≤0.05. REP-PCR based genotyping showed seven distinct genotypes (A-G prevalent in this region. This study reports the presence of multidrug resistant S. aureus in sub-clinical mastitis which were also highly virulent that could be a major obstacle in the treatment of mastitis in this region of China.

  19. Characterization of virulence factor regulation by SrrAB, a two-component system in Staphylococcus aureus.

    Science.gov (United States)

    Pragman, Alexa A; Yarwood, Jeremy M; Tripp, Timothy J; Schlievert, Patrick M

    2004-04-01

    Workers in our laboratory have previously identified the staphylococcal respiratory response AB (SrrAB), a Staphylococcus aureus two-component system that acts in the global regulation of virulence factors. This system down-regulates production of agr RNAIII, protein A, and toxic shock syndrome toxin 1 (TSST-1), particularly under low-oxygen conditions. In this study we investigated the localization and membrane orientation of SrrA and SrrB, transcription of the srrAB operon, the DNA-binding properties of SrrA, and the effect of SrrAB expression on S. aureus virulence. We found that SrrA is localized to the S. aureus cytoplasm, while SrrB is localized to the membrane and is properly oriented to function as a histidine kinase. srrAB has one transcriptional start site which results in either an srrA transcript or a full-length srrAB transcript; srrB must be cotranscribed with srrA. Gel shift assays of the agr P2, agr P3, protein A (spa), TSST-1 (tst), and srr promoters revealed SrrA binding at each of these promoters. Analysis of SrrAB-overexpressing strains by using the rabbit model of bacterial endocarditis demonstrated that overexpression of SrrAB decreased the virulence of the organisms compared to the virulence of isogenic strains that do not overexpress SrrAB. We concluded that SrrAB is properly localized and oriented to function as a two-component system. Overexpression of SrrAB, which represses agr RNAIII, TSST-1, and protein A in vitro, decreases virulence in the rabbit endocarditis model. Repression of these virulence factors is likely due to a direct interaction between SrrA and the agr, tst, and spa promoters.

  20. GigA and GigB are Master Regulators of Antibiotic Resistance, Stress Responses, and Virulence in Acinetobacter baumannii.

    Science.gov (United States)

    Gebhardt, Michael J; Shuman, Howard A

    2017-05-15

    A critical component of bacterial pathogenesis is the ability of an invading organism to sense and adapt to the harsh environment imposed by the host's immune system. This is especially important for opportunistic pathogens, such as Acinetobacter baumannii , a nutritionally versatile environmental organism that has recently gained attention as a life-threatening human pathogen. The emergence of A. baumannii is closely linked to antibiotic resistance, and many contemporary isolates are multidrug resistant (MDR). Unlike many other MDR pathogens, the molecular mechanisms underlying A. baumannii pathogenesis remain largely unknown. We report here the characterization of two recently identified virulence determinants, GigA and GigB, which comprise a signal transduction pathway required for surviving environmental stresses, causing infection and antibiotic resistance. Through transcriptome analysis, we show that GigA and GigB coordinately regulate the expression of many genes and are required for generating an appropriate transcriptional response during antibiotic exposure. Genetic and biochemical data demonstrate a direct link between GigA and GigB and the nitrogen phosphotransferase system (PTS Ntr ), establishing a novel connection between a novel stress response module and a well-conserved metabolic-sensing pathway. Based on the results presented here, we propose that GigA and GigB are master regulators of a global stress response in A. baumannii , and coupling this pathway with the PTS Ntr allows A. baumannii to integrate cellular metabolic status with external environmental cues. IMPORTANCE Opportunistic pathogens, including Acinetobacter baumannii , encounter many harsh environments during the infection cycle, including antibiotic exposure and the hostile environment within a host. While the development of antibiotic resistance in A. baumannii has been well studied, how this organism senses and responds to environmental cues remain largely unknown. Herein, we

  1. Cyclic di-GMP regulation of the bvg-repressed genes and the orphan response regulator RisA in Bordetella pertussis

    Science.gov (United States)

    Expression of Bordetella pertussis virulence factors is activated by the BvgAS two-component system. Under modulating growth conditions BvgAS indirectly represses another set of genes through the action of BvgR, a bvg-activated protein. BvgR blocks activation of the response regulator RisA which is ...

  2. Regulation of gene expression in Streptococcus pneumoniae by response regulator 09 is strain dependent

    NARCIS (Netherlands)

    W.T. Hendriksen (Wouter); N. Silva (Nuno); H.J. Bootsma (Hester); C.E. Blue (Clare); G.K. Paterson (Gavin); A.R. Kerr (Alison); A.S. de Jong (Arjan); O.P. Kuipers (Oscar); P.W.M. Hermans (Peter); T.J. Mitchell

    2007-01-01

    textabstractRecent murine studies have demonstrated that the role of response regulator 09 (RR09) of Streptococcus pneumoniae in virulence is different in different strains. In the present study, we used a murine pneumonia model of infection to assess the virulence of a TIGR4 rr09 mutant, and we

  3. Regulation of gene expression in Streptococcus pneumoniae by response regulator 09 is strain dependent

    NARCIS (Netherlands)

    Hendriksen, Wouter T.; Silva, Nuno; Bootsma, Hester J.; Blue, Clare E.; Paterson, Gavin K.; Kerr, Alison R.; de Jong, Anne; Kuipers, Oscar P.; Hermans, Peter W. M.; Mitchell, Tim J.

    Recent murine studies have demonstrated that the role of response regulator 09 (RR09) of Streptococcus pneumoniae in virulence is different in different strains. In the present study, we used a murine pneumonia model of infection to assess the virulence of a TIGR4 rr09 mutant, and we found that

  4. The majority of genes in the pathogenic Neisseria species are present in non-pathogenic Neisseria lactamica, including those designated as 'virulence genes'

    Directory of Open Access Journals (Sweden)

    Saunders Nigel J

    2006-05-01

    Full Text Available Abstract Background Neisseria meningitidis causes the life-threatening diseases meningococcal meningitis and meningococcal septicemia. Neisseria gonorrhoeae is closely related to the meningococcus, but is the cause of the very different infection, gonorrhea. A number of genes have been implicated in the virulence of these related yet distinct pathogens, but the genes that define and differentiate the species and their behaviours have not been established. Further, a related species, Neisseria lactamica is not associated with either type of infection in normally healthy people, and lives as a harmless commensal. We have determined which of the genes so far identified in the genome sequences of the pathogens are also present in this non-pathogenic related species. Results Thirteen unrelated strains of N. lactamica were investigated using comparative genome hybridization to the pan-Neisseria microarray-v2, which contains 2845 unique gene probes. The presence of 127 'virulence genes' was specifically addressed; of these 85 are present in N. lactamica. Of the remaining 42 'virulence genes' only 11 are present in all four of the sequenced pathogenic Neisseria. Conclusion Assessment of the complete dataset revealed that the vast majority of genes present in the pathogens are also present in N. lactamica. Of the 1,473 probes to genes shared by all four pathogenic genome sequences, 1,373 hybridize to N. lactamica. These shared genes cannot include genes that are necessary and sufficient for the virulence of the pathogens, since N. lactamica does not share this behaviour. This provides an essential context for the interpretation of gene complement studies of the pathogens.

  5. Correlation between Group B Streptococcal Genotypes, Their Antimicrobial Resistance Profiles, and Virulence Genes among Pregnant Women in Lebanon

    Directory of Open Access Journals (Sweden)

    Antoine Hannoun

    2009-01-01

    Full Text Available The antimicrobial susceptibility profiles of 76 Streptococcus agalactiae (Group B Streptococci [GBS] isolates from vaginal specimens of pregnant women near term were correlated to their genotypes generated by Random Amplified Polymorphic DNA analysis and their virulence factors encoding genes cylE, lmb, scpB, rib, and bca by PCR. Based on the distribution of the susceptibility patterns, six profiles were generated. RAPD analysis detected 7 clusters of genotypes. The cylE gene was present in 99% of the isolates, the lmb in 96%, scpB in 94.7%, rib in 33%, and bca in 56.5% of isolates. The isolates demonstrated a significant correlation between antimicrobial resistance and genotype clusters denoting the distribution of particular clones with different antimicrobial resistance profiles, entailing the practice of caution in therapeutic options. All virulence factors encoding genes were detected in all seven genotypic clusters with rib and bca not coexisting in the same genome.

  6. Identification of let-7-regulated oncofetal genes

    DEFF Research Database (Denmark)

    Boyerinas, Benjamin; Park, Sun-Mi; Shomron, Noam

    2008-01-01

    -regulated at the end of embryonic development. Let-7 is often down-regulated early during cancer development, suggesting that let-7-regulated oncofetal genes (LOG) may become reexpressed in cancer cells. Using comparative bioinformatics, we have identified 12 conserved LOGs that include HMGA2 and IMP-1/CRD-BP. IMP-1...

  7. Natural variation in the VELVET gene bcvel1 affects virulence and light-dependent differentiation in Botrytis cinerea.

    Science.gov (United States)

    Schumacher, Julia; Pradier, Jean-Marc; Simon, Adeline; Traeger, Stefanie; Moraga, Javier; Collado, Isidro González; Viaud, Muriel; Tudzynski, Bettina

    2012-01-01

    Botrytis cinerea is an aggressive plant pathogen causing gray mold disease on various plant species. In this study, we identified the genetic origin for significantly differing phenotypes of the two sequenced B. cinerea isolates, B05.10 and T4, with regard to light-dependent differentiation, oxalic acid (OA) formation and virulence. By conducting a map-based cloning approach we identified a single nucleotide polymorphism (SNP) in an open reading frame encoding a VELVET gene (bcvel1). The SNP in isolate T4 results in a truncated protein that is predominantly found in the cytosol in contrast to the full-length protein of isolate B05.10 that accumulates in the nuclei. Deletion of the full-length gene in B05.10 resulted in the T4 phenotype, namely light-independent conidiation, loss of sclerotial development and oxalic acid production, and reduced virulence on several host plants. These findings indicate that the identified SNP represents a loss-of-function mutation of bcvel1. In accordance, the expression of the B05.10 copy in T4 rescued the wild-type/B05.10 phenotype. BcVEL1 is crucial for full virulence as deletion mutants are significantly hampered in killing and decomposing plant tissues. However, the production of the two best known secondary metabolites, the phytotoxins botcinic acid and botrydial, are not affected by the deletion of bcvel1 indicating that other factors are responsible for reduced virulence. Genome-wide expression analyses of B05.10- and Δbcvel1-infected plant material revealed a number of genes differentially expressed in the mutant: while several protease- encoding genes are under-expressed in Δbcvel1 compared to the wild type, the group of over-expressed genes is enriched for genes encoding sugar, amino acid and ammonium transporters and glycoside hydrolases reflecting the response of Δbcvel1 mutants to nutrient starvation conditions.

  8. Absence of YbeY RNase compromises the growth and enhances the virulence plasmid gene expression of Yersinia enterocolitica O:3.

    Science.gov (United States)

    Leskinen, Katarzyna; Varjosalo, Markku; Skurnik, Mikael

    2015-02-01

    YbeY was recently recognized as an endoribonuclease playing a role in ribosome biosynthesis. In Escherichia coli it functions as a single-strand-specific RNase that processes the 3' end of the 16S rRNA and is crucial for the late-stage 70S ribosome quality control system. Here we report that YbeY is not essential in Yersinia enterocolitica serotype O:3, yet its absence strongly compromised the bacterium. The lack of YbeY resulted in misprocessing of 16S rRNA and a severe decrease of growth rate with complete growth arrest observed at elevated temperatures. Moreover, a ybeY mutation severely disturbed regulation of the Yersinia virulence plasmid (pYV) genes and affected the expression of regulatory small RNA species. Transcription of the pYV genes was upregulated in the ybeY mutant at 22 °C; the same genes were repressed in the wild-type bacterium. Furthermore, ybeY inactivation impaired many virulence-related features, such as resistance to elevated temperature and acid, and hindered utilization of different carbohydrates. In addition, the ybeY mutant strain showed decreased infectivity in a tissue culture infection model, especially at the stage of cell adhesion. Taken together, this study demonstrates the crucial role of YbeY in Y. enterocolitica O:3 physiology and pathogenicity. © 2015 The Authors.

  9. MITEs in the promoters of effector genes allow prediction of novel virulence genes in Fusarium oxysporum

    NARCIS (Netherlands)

    Schmidt, S.M.; Houterman, P.M.; Schreiver, I.; Ma, L.; Amyotte, S.; Chellappan, B.; Boeren, S.; Takken, F.L.W.; Rep, M.

    2013-01-01

    Background The plant-pathogenic fungus Fusarium oxysporum f.sp.lycopersici (Fol) has accessory, lineage-specific (LS) chromosomes that can be transferred horizontally between strains. A single LS chromosome in the Fol4287 reference strain harbors all known Fol effector genes. Transfer of this

  10. Cloning of Beauveria bassiana chitinase gene Bbchit1 and its application to improve fungal strain virulence.

    Science.gov (United States)

    Fang, Weiguo; Leng, Bo; Xiao, Yuehua; Jin, Kai; Ma, Jincheng; Fan, Yanhua; Feng, Jing; Yang, Xingyong; Zhang, Yongjun; Pei, Yan

    2005-01-01

    Entomopathogenic fungi can produce a series of chitinases, some of which act synergistically with proteases to degrade insect cuticle. However, chitinase involvement in insect fungus pathogenesis has not been fully characterized. In this paper, an endochitinase, Bbchit1, was purified to homogeneity from liquid cultures of Beauveria bassiana grown in a medium containing colloidal chitin. Bbchit1 had a molecular mass of about 33 kDa and pI of 5.4. Based on the N-terminal amino acid sequence, the chitinase gene, Bbchit1, and its upstream regulatory sequence were cloned. Bbchit1 was intronless, and there was a single copy in B. bassiana. Its regulatory sequence contained putative CreA/Crel carbon catabolic repressor binding domains, which was consistent with glucose suppression of Bbchit1. At the amino acid level, Bbchit1 showed significant similarity to a Streptomyces avermitilis putative endochitinase, a Streptomyces coelicolor putative chitinase, and Trichoderma harzianum endochitinase Chit36Y. However, Bbchit1 had very low levels of identity to other chitinase genes previously isolated from entomopathogenic fungi, indicating that Bbchit1 was a novel chitinase gene from an insect-pathogenic fungus. A gpd-Bbchit1 construct, in which Bbchit1 was driven by the Aspergiullus nidulans constitutive promoter, was transformed into the genome of B. bassiana, and three transformants that overproduced Bbchit1 were obtained. Insect bioassays revealed that overproduction of Bbchit1 enhanced the virulence of B. bassiana for aphids, as indicated by significantly lower 50% lethal concentrations and 50% lethal times of the transformants compared to the values for the wild-type strain.

  11. A study of Staphylococcus aureusnasal carriage, antibacterial resistance and virulence factor encoding genes in a tertiary care hospital, Kayseri, Turkey.

    Science.gov (United States)

    Oguzkaya-Artan, M; Artan, C; Baykan, Z; Sakalar, C; Turan, A; Aksu, H

    2015-01-01

    This study was to determine the virulence encoding genes, and the antibiotic resistance patterns of the Staphylococcus aureus isolates, which were isolated from the nasal samples of chest clinic patients. The nasal samples of the in-patients (431) and out-patients (1857) in Kayseri Training and Research Hospital's Chest Clinic, Kayseri, Turkey, were cultured on CHROMagar (Biolife, Italiana) S. aureus, and subcultured on sheep blood agar for the isolation of S. aureus. Disc diffusion method was used for antimicrobial susceptibility testing. The occurrence of the staphylococcal virulence encoding genes (enterotoksins [sea, seb, sec, see, seg, seh, sei, sej], fibronectin-binding proteins A, B [fnbA, fnbB], toxic shock syndrome toxin-1 [tst]) were detected by polymerase chain reaction. Forty-five of the 55 (81.8%) S. aureus isolates from inpatients, and 319 (90.6%) isolates from tested 352 out-patient's isolates were suspected to all the antibiotics tested. methicillin-resistant S. aureus (MRSA) was detected in 1.2% of S. aureus isolates. Rifampin, trimethoprim-sulfamethoxazole, clindamycin, erythromycin, gentamicin resistance rates were 1.2%, 1.7%, 2.0%, 8.8%, and 1.2%, respectively. The isolates were susceptible to teicoplanin and vancomycin. The genes most frequently found were tst (92.7%), seg (85.8%), sea (83.6%), fnbA (70.9%). There was no statistical significance detected between MRSA and mecA-negative S. aureus isolates in encoding genes distribution (P > 0.05). Our results show that virulence factor encoding genes were prevalent in patients with S. aureus carriage, whereas antibiotic resistance was low. These virulence determinants may increase the risk for subsequent invasive infections in carriers.

  12. HD-GYP domain proteins regulate biofilm formation and virulence in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Ryan, Robert P.; Lucey, Jean; O'Donovan, Karen

    2009-01-01

    residues (YN-GYP). Here we have investigated the role of these proteins in biofilm formation, virulence factor synthesis and virulence of P. aeruginosa. Mutation of PA4108 and PA4781 led to an increase in the level of cyclic-di-GMP in P. aeruginosa, consistent with the predicted activity of the encoded......2572 had a negative influence on swarming that was cryptic and was revealed only after removal of an uncharacterized C-terminal domain. Mutation of PA4108, PA4781 and PA2572 had distinct effects on biofilm formation and architecture of P. aeruginosa. All three proteins contributed to virulence of P...

  13. The importance of virulence prediction and gene networks in microbial risk assessment

    DEFF Research Database (Denmark)

    Wassenaar, Gertrude Maria; Gamieldien, Junaid; Shatkin, JoAnne

    2007-01-01

    For microbial risk assessment, it is necessary to recognize and predict Virulence of bacterial pathogens, including their ability to contaminate foods. Hazard characterization requires data on strain variability regarding virulence and survival during food processing. Moreover, information...... and characterization of microbial hazards, including emerging pathogens, in the context of microbial risk assessment....

  14. Proteomic analysis of growth phase-dependent expression of Legionella pneumophila proteins which involves regulation of bacterial virulence traits.

    Directory of Open Access Journals (Sweden)

    Tsuyoshi Hayashi

    Full Text Available Legionella pneumophila, which is a causative pathogen of Legionnaires' disease, expresses its virulent traits in response to growth conditions. In particular, it is known to become virulent at a post-exponential phase in vitro culture. In this study, we performed a proteomic analysis of differences in expression between the exponential phase and post-exponential phase to identify candidates associated with L. pneumophila virulence using 2-Dimentional Fluorescence Difference Gel Electrophoresis (2D-DIGE combined with Matrix-Assisted Laser Desorption/Ionization-Mass Spectrometry (MALDI-TOF-MS. Of 68 identified proteins that significantly differed in expression between the two growth phases, 64 were up-regulated at a post-exponential phase. The up-regulated proteins included enzymes related to glycolysis, ketone body biogenesis and poly-3-hydroxybutyrate (PHB biogenesis, suggesting that L. pneumophila may utilize sugars and lipids as energy sources, when amino acids become scarce. Proteins related to motility (flagella components and twitching motility-associated proteins were also up-regulated, predicting that they enhance infectivity of the bacteria in host cells under certain conditions. Furthermore, 9 up-regulated proteins of unknown function were found. Two of them were identified as novel bacterial factors associated with hemolysis of sheep red blood cells (SRBCs. Another 2 were found to be translocated into macrophages via the Icm/Dot type IV secretion apparatus as effector candidates in a reporter assay with Bordetella pertussis adenylate cyclase. The study will be helpful for virulent analysis of L. pneumophila from the viewpoint of physiological or metabolic modulation dependent on growth phase.

  15. RNA- and protein-mediated control of Listeria monocytogenes virulence gene expression

    Science.gov (United States)

    Lebreton, Alice; Cossart, Pascale

    2017-01-01

    ABSTRACT The model opportunistic pathogen Listeria monocytogenes has been the object of extensive research, aiming at understanding its ability to colonize diverse environmental niches and animal hosts. Bacterial transcriptomes in various conditions reflect this efficient adaptability. We review here our current knowledge of the mechanisms allowing L. monocytogenes to respond to environmental changes and trigger pathogenicity, with a special focus on RNA-mediated control of gene expression. We highlight how these studies have brought novel concepts in prokaryotic gene regulation, such as the ‘excludon’ where the 5′-UTR of a messenger also acts as an antisense regulator of an operon transcribed in opposite orientation, or the notion that riboswitches can regulate non-coding RNAs to integrate complex metabolic stimuli into regulatory networks. Overall, the Listeria model exemplifies that fine RNA tuners act together with master regulatory proteins to orchestrate appropriate transcriptional programmes. PMID:27217337

  16. GlmS and NagB regulate amino sugar metabolism in opposing directions and affect Streptococcus mutans virulence.

    Directory of Open Access Journals (Sweden)

    Miki Kawada-Matsuo

    Full Text Available Streptococcus mutans is a cariogenic pathogen that produces an extracellular polysaccharide (glucan from dietary sugars, which allows it to establish a reproductive niche and secrete acids that degrade tooth enamel. While two enzymes (GlmS and NagB are known to be key factors affecting the entrance of amino sugars into glycolysis and cell wall synthesis in several other bacteria, their roles in S. mutans remain unclear. Therefore, we investigated the roles of GlmS and NagB in S. mutans sugar metabolism and determined whether they have an effect on virulence. NagB expression increased in the presence of GlcNAc while GlmS expression decreased, suggesting that the regulation of these enzymes, which functionally oppose one another, is dependent on the concentration of environmental GlcNAc. A glmS-inactivated mutant could not grow in the absence of GlcNAc, while nagB-inactivated mutant growth was decreased in the presence of GlcNAc. Also, nagB inactivation was found to decrease the expression of virulence factors, including cell-surface protein antigen and glucosyltransferase, and to decrease biofilm formation and saliva-induced S. mutans aggregation, while glmS inactivation had the opposite effects on virulence factor expression and bacterial aggregation. Our results suggest that GlmS and NagB function in sugar metabolism in opposing directions, increasing and decreasing S. mutans virulence, respectively.

  17. Virulence-associated and antibiotic resistance genes of microbial populations in cattle feces analyzed using a metagenomic approach.

    Science.gov (United States)

    Durso, Lisa M; Harhay, Gregory P; Bono, James L; Smith, Timothy P L

    2011-02-01

    The bovine fecal microbiota impacts human food safety as well as animal health. Although the bacteria of cattle feces have been well characterized using culture-based and culture-independent methods, techniques have been lacking to correlate total community composition with community function. We used high throughput sequencing of total DNA extracted from fecal material to characterize general community composition and examine the repertoire of microbial genes present in beef cattle feces, including genes associated with antibiotic resistance and bacterial virulence. Results suggest that traditional 16S sequencing using "universal" primers to generate full-length sequence may under represent Acitinobacteria and Proteobacteria. Over eight percent (8.4%) of the sequences from our beef cattle fecal pool sample could be categorized as virulence genes, including a suite of genes associated with resistance to antibiotic and toxic compounds (RATC). This is a higher proportion of virulence genes found in Sargasso sea, chicken cecum, and cow rumen samples, but comparable to the proportion found in Antarctic marine derived lake, human fecal, and farm soil samples. The quantitative nature of metagenomic data, combined with the large number of RATC classes represented in samples from widely different habitats indicates that metagenomic data can be used to track relative amounts of antibiotic resistance genes in individual animals over time. Consequently, these data can be used to generate sample-specific and temporal antibiotic resistance gene profiles to facilitate an understanding of the ecology of the microbial communities in each habitat as well as the epidemiology of antibiotic resistant gene transport between and among habitats. Published by Elsevier B.V.

  18. Discovery and biophysical characterization of 2-amino-oxadiazoles as novel antagonists of PqsR, an important regulator of Pseudomonas aeruginosa virulence.

    Science.gov (United States)

    Zender, Michael; Klein, Tobias; Henn, Claudia; Kirsch, Benjamin; Maurer, Christine K; Kail, Dagmar; Ritter, Christiane; Dolezal, Olan; Steinbach, Anke; Hartmann, Rolf W

    2013-09-12

    The human pathogen Pseudomonas aeruginosa employs alkyl quinolones for cell-to-cell communication. The Pseudomonas quinolone signal (PQS) regulates various virulence factors via interaction with the transcriptional regulator PqsR. Therefore, we consider the development of PqsR antagonists a novel strategy to limit the pathogenicity of P. aeruginosa. A fragment identification approach using surface plasmon resonance screening led to the discovery of chemically diverse PqsR ligands. The optimization of the most promising hit (5) resulted in the oxadiazole-2-amine 37 showing pure antagonistic activity in Escherichia coli (EC50 = 7.5 μM) and P. aeruginosa (EC50 = 38.5 μM) reporter gene assays. 37 was able to diminish the production of the PQS precursor HHQ in a PqsH-deficient P. aeruginosa mutant. The level of the major virulence factor pyocyanin was significantly reduced in wild-type P. aeruginosa. In addition, site-directed mutagenesis in combination with isothermal titration calorimetry and NMR INPHARMA experiments revealed that the identified ligands bind to the same site of PqsR by adopting different binding modes. These findings will be utilized in a future fragment-growing approach aiming at novel therapeutic options for the treatment of P. aeruginosa infections.

  19. Combinatorial gene regulation in Plasmodium falciparum.

    NARCIS (Netherlands)

    Noort, V. van; Huynen, M.A.

    2006-01-01

    The malaria parasite Plasmodium falciparum has a complicated life cycle with large variations in its gene expression pattern, but it contains relatively few specific transcriptional regulators. To elucidate this paradox, we identified regulatory sequences, using an approach that integrates the

  20. Rice bacterial blight pathogen Xanthomonas oryzae pv. oryzae produces multiple DSF-family signals in regulation of virulence factor production

    Directory of Open Access Journals (Sweden)

    Cha Jae-Soon

    2010-07-01

    Full Text Available Abstract Background Xanthomonas oryzae pv. oryzae (Xoo is the causal agent of rice bacterial blight disease. Xoo produces a range of virulence factors, including EPS, extracellular enzyme, iron-chelating siderophores, and type III-secretion dependent effectors, which are collectively essential for virulence. Genetic and genomics evidence suggest that Xoo might use the diffusible signal factor (DSF type quorum sensing (QS system to regulate the virulence factor production. However, little is known about the chemical structure of the DSF-like signal(s produced by Xoo and the factors influencing the signal production. Results Xoo genome harbours an rpf cluster comprising rpfB, rpfF, rpfC and rpfG. The proteins encoded by these genes are highly homologous to their counterparts in X. campestris pv. campestris (Xcc, suggesting that Xcc and Xoo might use similar mechanisms for DSF biosynthesis and autoregulation. Consistent with in silico analysis, the rpfF mutant was DSF-deficient and the rpfC mutant produced about 25 times higher DSF-like activity than the wild type Xoo strain KACC10331. From the supernatants of rpfC mutant, we purified three compounds showing strong DSF-like activity. Mass spectrometry and NMR analysis revealed that two of them were the previously characterized DSF and BDSF; the third one was a novel unsaturated fatty acid with 2 double bonds and was designated as CDSF in this study. Further analysis showed that all the three DSF-family signals were synthesized via the enzyme RpfF encoded by Xoo2868. DSF and BDSF at a final concentration of 3 μM to the rpfF mutant could fully restore its extracellular xylanase activity and EPS production to the wild type level, but CDSF was less active than DSF and BDSF in induction of EPS and xylanase. DSF and CDSF shared a similar cell density-dependent production time course with the maximum production being detected at 42 h after inoculation, whereas the maximum production of BDSF was observed

  1. Divergent regulation of Arabidopsis SAUR genes

    NARCIS (Netherlands)

    Mourik, van Hilda; Dijk, van Aalt D.J.; Stortenbeker, Niek; Angenent, Gerco C.; Bemer, Marian

    2017-01-01

    Background: Small Auxin-Upregulated RNA (SAUR) genes encode growth regulators that induce cell elongation. Arabidopsis contains more than 70 SAUR genes, of which the growth-promoting function has been unveiled in seedlings, while their role in other tissues remained largely unknown. Here, we

  2. Influenza A virus NS1 gene mutations F103L and M106I increase replication and virulence

    Directory of Open Access Journals (Sweden)

    Ping Jihui

    2011-01-01

    Full Text Available Abstract Background To understand the evolutionary steps required for a virus to become virulent in a new host, a human influenza A virus (IAV, A/Hong Kong/1/68(H3N2 (HK-wt, was adapted to increased virulence in the mouse. Among eleven mutations selected in the NS1 gene, two mutations F103L and M106I had been previously detected in the highly virulent human H5N1 isolate, A/HK/156/97, suggesting a role for these mutations in virulence in mice and humans. Results To determine the selective advantage of these mutations, reverse genetics was used to rescue viruses containing each of the NS1 mouse adapted mutations into viruses possessing the HK-wt NS1 gene on the A/PR/8/34 genetic backbone. Both F103L and M106I NS1 mutations significantly enhanced growth in vitro (mouse and canine cells and in vivo (BALB/c mouse lungs as well as enhanced virulence in the mouse. Only the M106I NS1 mutation enhanced growth in human cells. Furthermore, these NS1 mutations enhanced early viral protein synthesis in MDCK cells and showed an increased ability to replicate in mouse interferon β (IFN-β pre-treated mouse cells relative to rPR8-HK-NS-wt NS1. The double mutant, rPR8-HK-NS-F103L + M106I, demonstrated growth attenuation late in infection due to increased IFN-β induction in mouse cells. We then generated a rPR8 virus possessing the A/HK/156/97 NS gene that possesses 103L + 106I, and then rescued the L103F + I106M mutant. The 103L + 106I mutations increased virulence by >10 fold in BALB/c mice. We also inserted the avian A/Ck/Beijing/1/95 NS1 gene (the source lineage of the A/HK/156/97 NS1 gene that possesses 103L + 106I, onto the A/WSN/33 backbone and then generated the L103F + I106M mutant. None of the H5N1 and H9N2 NS containing viruses resulted in increased IFN-β induction. The rWSN-A/Ck/Beijing/1/95-NS1 gene possessing 103L and 106I demonstrated 100 fold enhanced growth and >10 fold enhanced virulence that was associated with increased tropism for lung

  3. Differentiation of the virulence potential of Campylobacter jejuni strains by use of gene transcription analysis and a caco-2 assay

    DEFF Research Database (Denmark)

    Poli, Vanessa Fadanelli Schoenardie; Thorsen, Line; Olesen, Inger

    2012-01-01

    properties were evaluated by analyzing transcriptions of the virulence genes cdtB, ciaB, cadF and the stress associated genes clpP, htrB using reverse transcription quantitative PCR (RT-qPCR) and by the ability of the C. jejuni strains to adhere to and invade Caco-2 cells. Similar cell survival and no growth...... gene, cipA between DFVF1099 and NCTC11168 resulting in a 14 amino acid deletion and 28 amino acid addition at the N and C terminal ends respectively of the CipA protein of DFVF1099. In contrast to DFVF1099, strains NCTC1168 and TB1048 were able to invade Caco-2 cells. Invasion ability was not affected...... expression of C. jejuni. The clinical strains appeared to be more virulent than the chicken isolate as measured by the Caco-2 invasion assay which could be due to differences in CipA functionality. The RT-qPCR analysis and Caco-2 assay showed to be useful tools for differentiating virulence potentials...

  4. Posttranscriptional Regulation of the Neurofibromatosis 2 Gene

    Science.gov (United States)

    2006-07-01

    signaling and division were downregulated, including an apoptosis - related, putative tumor suppressor gene, LUCA-15, which was downregulated in seven of... embryologically from the outgrowth of the developing brain (Martinez-Morales et al., 2004). It is comprised of two major layers, the inner layer (prospective...eight genes involved with cell signaling and division were down- regulated. These include an apoptosis -related, putative tumor suppressor gene LUCA-15

  5. Phasevarion mediated epigenetic gene regulation in Helicobacter pylori.

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    Yogitha N Srikhanta

    Full Text Available Many host-adapted bacterial pathogens contain DNA methyltransferases (mod genes that are subject to phase-variable expression (high-frequency reversible ON/OFF switching of gene expression. In Haemophilus influenzae and pathogenic Neisseria, the random switching of the modA gene, associated with a phase-variable type III restriction modification (R-M system, controls expression of a phase-variable regulon of genes (a "phasevarion", via differential methylation of the genome in the modA ON and OFF states. Phase-variable type III R-M systems are also found in Helicobacter pylori, suggesting that phasevarions may also exist in this key human pathogen. Phylogenetic studies on the phase-variable type III modH gene revealed that there are 17 distinct alleles in H. pylori, which differ only in their DNA recognition domain. One of the most commonly found alleles was modH5 (16% of isolates. Microarray analysis comparing the wild-type P12modH5 ON strain to a P12ΔmodH5 mutant revealed that six genes were either up- or down-regulated, and some were virulence-associated. These included flaA, which encodes a flagella protein important in motility and hopG, an outer membrane protein essential for colonization and associated with gastric cancer. This study provides the first evidence of this epigenetic mechanism of gene expression in H. pylori. Characterisation of H. pylori modH phasevarions to define stable immunological targets will be essential for vaccine development and may also contribute to understanding H. pylori pathogenesis.

  6. A Surfactant-Induced Functional Modulation of a Global Virulence Regulator from Staphylococcus aureus.

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    Sukhendu Mandal

    Full Text Available Triton X-100 (TX-100, a useful non-ionic surfactant, reduced the methicillin resistance in Staphylococcus aureus significantly. Many S. aureus proteins were expressed in the presence of TX-100. SarA, one of the TX-100-induced proteins, acts as a global virulence regulator in S. aureus. To understand the effects of TX-100 on the structure, and function of SarA, a recombinant S. aureus SarA (rSarA and its derivative (C9W have been investigated in the presence of varying concentrations of this surfactant using various probes. Our data have revealed that both rSarA and C9W bind to the cognate DNA with nearly similar affinity in the absence of TX-100. Interestingly, their DNA binding activities have been significantly increased in the presence of pre-micellar concentration of TX-100. The increase of TX-100 concentrations to micellar or post-micellar concentration did not greatly enhance their activities further. TX-100 molecules have altered the secondary and tertiary structures of both proteins to some extents. Size of the rSarA-TX-100 complex appears to be intermediate to those of rSarA and TX-100. Additional analyses show a relatively moderate interaction between C9W and TX-100. Binding of TX-100 to C9W has, however, occurred by a cooperative pathway particularly at micellar and higher concentrations of this surfactant. Taken together, TX-100-induced structural alteration of rSarA and C9W might be responsible for their increased DNA binding activity. As TX-100 has stabilized the somewhat weaker SarA-DNA complex effectively, it could be used to study its structure in the future.

  7. Bursal transcriptome profiling of different inbred chicken lines reveals key differentially expressed genes at 3 days post-infection with very virulent infectious bursal disease virus.

    Science.gov (United States)

    Farhanah, Mohd Isa; Yasmin, Abd Rahaman; Mat Isa, Nurulfiza; Hair-Bejo, Mohd; Ideris, Aini; Powers, Claire; Oladapo, Omobolanle; Nair, Venugopal; Khoo, Jia-Shiun; Ghazali, Ahmad-Kamal; Yee, Wai-Yan; Omar, Abdul Rahman

    2018-01-01

    Infectious bursal disease is a highly contagious disease in the poultry industry and causes immunosuppression in chickens. Genome-wide regulations of immune response genes of inbred chickens with different genetic backgrounds, following very virulent infectious bursal disease virus (vvIBDV) infection are poorly characterized. Therefore, this study aims to analyse the bursal tissue transcriptome of six inbred chicken lines 6, 7, 15, N, O and P following infection with vvIBDV strain UK661 using strand-specific next-generation sequencing, by highlighting important genes and pathways involved in the infected chicken during peak infection at 3 days post-infection. All infected chickens succumbed to the infection without major variations among the different lines. However, based on the viral loads and bursal lesion scoring, lines P and 6 can be considered as the most susceptible lines, while lines 15 and N were regarded as the least affected lines. Transcriptome profiling of the bursa identified 4588 genes to be differentially expressed, with 2985 upregulated and 1642 downregulated genes, in which these genes were commonly or uniquely detected in all or several infected lines. Genes that were upregulated are primarily pro-inflammatory cytokines, chemokines and IFN-related. Various genes that are associated with B-cell functions and genes related to apoptosis were downregulated, together with the genes involved in p53 signalling. In conclusion, bursal transcriptome profiles of different inbred lines showed differential expressions of pro-inflammatory cytokines and chemokines, Th1 cytokines, JAK-STAT signalling genes, MAPK signalling genes, and their related pathways following vvIBDV infection.

  8. Identification of Xylella fastidiosa antivirulence genes: hemagglutinin adhesins contribute a biofilm maturation to X. fastidios and colonization and attenuate virulence.

    Science.gov (United States)

    Guilhabert, Magalie R; Kirkpatrick, Bruce C

    2005-08-01

    Xylella fastidosa, a gram-negative, xylem-limited bacterium, is the causal agent of several economically important plant diseases, including Pierce's disease (PD) and citrus variegated chlorosis (CVC). Until recently, the inability to transform or produce transposon mutants of X. fastidosa had been a major impediment to identifying X. fastidosa genes that mediate pathogen and plant interactions. A random transposon (Tn5) library of X. fastidosa was constructed and screened for mutants showing more severe symptoms and earlier grapevine death (hypervirulence) than did vines infected with the wild type. Seven hypervirulent mutants identified in this screen moved faster and reached higher populations than the wild type in grapevines. These results suggest that X. fastidosa attenuates its virulence in planta and that movement is important in X. fastidosa virulence. The mutated genes were sequenced and none had been described previously as antivirulence genes, although six of them showed similarity with genes of known functions in other organisms. One transposon insertion inactivated a hemagglutinin adhesin gene (PD2118), which we named HxfA. Another mutant in a second putative X. fastidosa hemagglutinin gene, PD1792 (HxfB), was constructed, and further characterization of these hxf mutants suggests that X. fastidosa hemagglutinins mediate contact between X. fastidosa cells, which results in colony formation and biofilm maturation within the xylem vessels.

  9. Role of the Genes of Type VI Secretion System in Virulence of Rice Bacterial Brown Stripe Pathogen Acidovorax avenae subsp. avenae Strain RS-2

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    Md. Mahidul Islam Masum

    2017-09-01

    Full Text Available The Type VI secretion system (T6SS is a class of macromolecular machine that is required for the virulence of gram-negative bacteria. However, it is still not clear what the role of T6SS in the virulence of rice bacterial brown stripe pathogen Acidovorax avenae subsp. avenae (Aaa is. The aim of the current study was to investigate the contribution of T6SS in Aaa strain RS2 virulence using insertional deletion mutation and complementation approaches. This strain produced weak virulence but contains a complete T6SS gene cluster based on a genome-wide analysis. Here we compared the virulence-related phenotypes between the wild-type (RS-2 and 25 T6SS mutants, which were constructed using homologous recombination methods. The mutation of 15 T6SS genes significantly reduced bacterial virulence and the secretion of Hcp protein. Additionally, the complemented 7 mutations ΔpppA, ΔclpB, Δhcp, ΔdotU, ΔicmF, ΔimpJ, and ΔimpM caused similar virulence characteristics as RS-2. Moreover, the mutant ΔpppA, ΔclpB, ΔicmF, ΔimpJ and ΔimpM genes caused by a 38.3~56.4% reduction in biofilm formation while the mutants ΔpppA, ΔclpB, ΔicmF and Δhcp resulted in a 37.5~44.6% reduction in motility. All together, these results demonstrate that T6SS play vital roles in the virulence of strain RS-2, which may be partially attributed to the reductions in Hcp secretion, biofilm formation and motility. However, differences in virulence between strain RS-1 and RS-2 suggest that other factors may also be involved in the virulence of Aaa.

  10. Role of the Genes of Type VI Secretion System in Virulence of Rice Bacterial Brown Stripe Pathogen Acidovorax avenae subsp. avenae Strain RS-2.

    Science.gov (United States)

    Masum, Md Mahidul Islam; Yang, Yingzi; Li, Bin; Olaitan, Ogunyemi Solabomi; Chen, Jie; Zhang, Yang; Fang, Yushi; Qiu, Wen; Wang, Yanli; Sun, Guochang

    2017-09-21

    The Type VI secretion system (T6SS) is a class of macromolecular machine that is required for the virulence of gram-negative bacteria. However, it is still not clear what the role of T6SS in the virulence of rice bacterial brown stripe pathogen Acidovorax avenae subsp. avenae (Aaa) is. The aim of the current study was to investigate the contribution of T6SS in Aaa strain RS2 virulence using insertional deletion mutation and complementation approaches. This strain produced weak virulence but contains a complete T6SS gene cluster based on a genome-wide analysis. Here we compared the virulence-related phenotypes between the wild-type (RS-2) and 25 T6SS mutants, which were constructed using homologous recombination methods. The mutation of 15 T6SS genes significantly reduced bacterial virulence and the secretion of Hcp protein. Additionally, the complemented 7 mutations Δ pppA , Δ clpB , Δ hcp , Δ dotU , Δ icmF , Δ impJ , and Δ impM caused similar virulence characteristics as RS-2. Moreover, the mutant Δ pppA , Δ clpB , Δ icmF , Δ impJ and Δ impM genes caused by a 38.3~56.4% reduction in biofilm formation while the mutants Δ pppA , Δ clpB , Δ icmF and Δ hcp resulted in a 37.5~44.6% reduction in motility. All together, these results demonstrate that T6SS play vital roles in the virulence of strain RS-2, which may be partially attributed to the reductions in Hcp secretion, biofilm formation and motility. However, differences in virulence between strain RS-1 and RS-2 suggest that other factors may also be involved in the virulence of Aaa.

  11. Involvement of protein tyrosine phosphatases BcPtpA and BcPtpB in regulation of vegetative development, virulence and multi-stress tolerance in Botrytis cinerea.

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    Qianqian Yang

    Full Text Available Tyrosine phosphorylation and dephosphorylation have emerged as fundamentally important mechanisms of signal transduction and regulation in eukaryotic cells, governing many processes, but little has been known about their functions in filamentous fungi. In this study, we deleted two putative protein tyrosine phosphatase (PTP genes (BcPTPA and BcPTPB in Botrytis cinerea, encoding the orthologs of Saccharomyces cerevisiae Ptp2 and Ptp3, respectively. Although BcPtpA and BcPtpB have opposite functions in conidiation, they are essential for sclerotial formation in B. cinerea. BcPTPA and BcPTPB deletion mutants ΔBcPtpA-10 and ΔBcPtpB-4 showed significantly increased sensitivity to osmotic and oxidative stresses, and to cell wall damaging agents. Inoculation tests showed that both mutants exhibited dramatically decreased virulence on tomato leaves, apples and grapes. In S. cerevisiae, it has been shown that Ptp2 and Ptp3 negatively regulate the high-osmolarity glycerol (HOG pathway and the cell wall integrity (CWI pathway. Although both BcPtpA and BcPtpB were able to inactive Hog1 and Mpk1 in S. cerevisiae, in contrast to S. cerevisiae, they positively regulate phosphorylation of BcSak1 (the homologue of Hog1 and BcBmp3 (the homologue of Mpk1 in B. cinerea under stress conditions. These results demonstrated that functions of PTPs in B. cinerea are different from those in S. cerevisiae, and BcPtpA and BcPtpB play important roles in regulation of vegetative development, virulence and in adaptation to oxidative, osmotic and cell-wall damage stresses in B. cinerea.

  12. Beyond the chromosome: the prevalence of unique extra-chromosomal bacteriophages with integrated virulence genes in pathogenic Staphylococcus aureus.

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    Bryan Utter

    Full Text Available In Staphylococcus aureus, the disease impact of chromosomally integrated prophages on virulence is well described. However, the existence of extra-chromosomal prophages, both plasmidial and episomal, remains obscure. Despite the recent explosion in bacterial and bacteriophage genomic sequencing, studies have failed to specifically focus on extra-chromosomal elements. We selectively enriched and sequenced extra-chromosomal DNA from S. aureus isolates using Roche-454 technology and uncovered evidence for the widespread distribution of multiple extra-chromosomal prophages (ExPΦs throughout both antibiotic-sensitive and -resistant strains. We completely sequenced one such element comprised of a 43.8 kbp, circular ExPΦ (designated ФBU01 from a vancomycin-intermediate S. aureus (VISA strain. Assembly and annotation of ФBU01 revealed a number of putative virulence determinants encoded within a bacteriophage immune evasion cluster (IEC. Our identification of several potential ExPΦs and mobile genetic elements (MGEs also revealed numerous putative virulence factors and antibiotic resistance genes. We describe here a previously unidentified level of genetic diversity of stealth extra-chromosomal elements in S. aureus, including phages with a larger presence outside the chromosome that likely play a prominent role in pathogenesis and strain diversity driven by horizontal gene transfer (HGT.

  13. Phage-mediated dispersal of biofilm and distribution of bacterial virulence genes is induced by quorum sensing.

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    Friederike S Rossmann

    2015-02-01

    Full Text Available The microbiome and the phage meta-genome within the human gut are influenced by antibiotic treatments. Identifying a novel mechanism, here we demonstrate that bacteria use the universal communication molecule AI-2 to induce virulence genes and transfer them via phage release. High concentrations (i.e. 100 μM of AI-2 promote dispersal of bacteria from already established biofilms, and is associated with release of phages capable of infecting other bacteria. Enterococcus faecalis V583ΔABC harbours 7 prophages in its genome, and a mutant deficient in one of these prophages (i.e. prophage 5 showed a greatly reduced dispersal of biofilm. Infection of a probiotic E. faecalis strain without lytic prophages with prophage 5 resulted in increased biofilm formation and also in biofilm dispersal upon induction with AI-2. Infection of the probiotic E. faecalis strain with phage-containing supernatants released through AI-2 from E. faecalis V583ΔABC resulted in a strong increase in pathogenicity of this strain. The polylysogenic probiotic strain was also more virulent in a mouse sepsis model and a rat endocarditis model. Both AI-2 and ciprofloxacin lead to phage release, indicating that conditions in the gastrointestinal tract of hospitalized patients treated with antibiotics might lead to distribution of virulence genes to apathogenic enterococci and possibly also to other commensals or even to beneficial probiotic strains.

  14. An eight-year study of Shigella species in Beijing, China: serodiversity, virulence genes, and antimicrobial resistance.

    Science.gov (United States)

    Qu, Mei; Zhang, Xin; Liu, Guirong; Huang, Ying; Jia, Lei; Liang, Weili; Li, Xitai; Wu, Xiaona; Li, Jie; Yan, Hanqiu; Kan, Biao; Wang, Quanyi

    2014-07-14

    This study was conducted to determine the prevalence of serotypes, virulence factors, and antimicrobial resistance patterns of Shigella spp. in Beijing, China, from 2004 to 2011. Real-time PCR assays were used to detect virulent genes, and the Kirby-Bauer disk diffusion method was used to evaluate antimicrobial resistance. Among the total of 1,652 Shigella isolates, S. sonnei (57.1%) was the predominant species, followed by S. flexneri (42.3%), S. dysenteriae (0.4%), and S. boydii (0.2%). Nineteen serotypes were discovered among S. flexneri strains. The virulence gene ipaH was the most frequent, followed by sen and set. The presence of set showed significant difference in two dominant serogroups, S. flexneri and S. sonnei. Over 90% of Shigella isolates showed resistance to at least three drugs with widened spectrum. High-level antimicrobial resistance to single and multiple antibiotics was more common among S. sonnei than S. flexneri. There was an obvious serotype change and a dramatic increase of antibiotic resistance in Shigella prevalence in Beijing.

  15. Reduction of Aspergillus niger Virulence in Apple Fruits by Deletion of the Catalase Gene cpeB.

    Science.gov (United States)

    Zhang, Meng-Ke; Tang, Jun; Huang, Zhong-Qin; Hu, Kang-Di; Li, Yan-Hong; Han, Zhuo; Chen, Xiao-Yan; Hu, Lan-Ying; Yao, Gai-Fang; Zhang, Hua

    2018-05-30

    Aspergillus niger, a common saprophytic fungus, causes rot in many fruits. We studied the role of a putative catalase-peroxidase-encoding gene, cpeB, in oxidative stress and virulence in fruit. The cpeB gene was deleted in A. niger by homologous recombination, and the Δ cpeB mutant showed decreased CAT activity compared with that of the wild type. The cpeB gene deletion caused increased sensitivity to H 2 O 2 stress, and spore germination was significantly reduced; in addition, the reactive-oxygen-species (ROS) metabolites superoxide anions (·O 2 - ), hydrogen peroxide (H 2 O 2 ), and malondialdehyde (MDA) accumulated in the Δ cpeB mutant during H 2 O 2 stress. Furthermore, ROS metabolism in A. niger infected apples was determined, and our results showed that the Δ cpeB mutant induced an attenuated response in apple fruit during the fruit-pathogen interaction; the cpeB gene deletion significantly reduced the development of lesions, suggesting that the cpeB gene in A. niger is essential for full virulence in apples.

  16. Genome Plasticity and Polymorphisms in Critical Genes Correlate with Increased Virulence of Dutch Outbreak-Related Coxiella burnetii Strains

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    Runa Kuley

    2017-08-01

    Full Text Available Coxiella burnetii is an obligate intracellular bacterium and the etiological agent of Q fever. During 2007–2010 the largest Q fever outbreak ever reported occurred in The Netherlands. It is anticipated that strains from this outbreak demonstrated an increased zoonotic potential as more than 40,000 individuals were assumed to be infected. The acquisition of novel genetic factors by these C. burnetii outbreak strains, such as virulence-related genes, has frequently been proposed and discussed, but is not proved yet. In the present study, the whole genome sequence of several Dutch strains (CbNL01 and CbNL12 genotypes, a few additionally selected strains from different geographical locations and publicly available genome sequences were used for a comparative bioinformatics approach. The study focuses on the identification of specific genetic differences in the outbreak related CbNL01 strains compared to other C. burnetii strains. In this approach we investigated the phylogenetic relationship and genomic aspects of virulence and host-specificity. Phylogenetic clustering of whole genome sequences showed a genotype-specific clustering that correlated with the clustering observed using Multiple Locus Variable-number Tandem Repeat Analysis (MLVA. Ortholog analysis on predicted genes and single nucleotide polymorphism (SNP analysis of complete genome sequences demonstrated the presence of genotype-specific gene contents and SNP variations in C. burnetii strains. It also demonstrated that the currently used MLVA genotyping methods are highly discriminatory for the investigated outbreak strains. In the fully reconstructed genome sequence of the Dutch outbreak NL3262 strain of the CbNL01 genotype, a relatively large number of transposon-linked genes were identified as compared to the other published complete genome sequences of C. burnetii. Additionally, large numbers of SNPs in its membrane proteins and predicted virulence-associated genes were identified

  17. Characterisation of virulence genes in methicillin susceptible and resistant Staphylococcus aureus isolates from a paediatric population in a university hospital of Medellín, Colombia

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    Judy Natalia Jiménez

    2011-12-01

    Full Text Available Virulence and antibiotic resistance are significant determinants of the types of infections caused by Staphylococcus aureus and paediatric groups remain among the most commonly affected populations. The goal of this study was to characterise virulence genes of methicillin-susceptible S. aureus (MSSA and methicillin-resistant S. aureus (MRSA strains isolated from a paediatric population of a Colombian University Hospital during 2009. Sixty MSSA and MRSA isolates were obtained from paediatric patients between zero-14 years. We identified the genes encoding virulence factors, which included Panton-Valentine leucocidine (PVL, staphylococcal enterotoxins A-E, exfoliative toxins A and B and toxic shock syndrome toxin 1. Typing of the staphylococcal chromosome cassette mec (SCCmec was performed in MRSA strains. The virulence genes were more diverse and frequent in MSSA than in MRSA isolates (83% vs. 73%. MRSA strains harboured SCCmec types IVc (60%, I (30%, IVa (7% and V (3%. SCCmec type IVc isolates frequently carried the PVL encoding genes and harboured virulence determinants resembling susceptible strains while SCCmec type I isolates were often negative. PVL was not exclusive to skin and soft tissue infections. As previously suggested, these differences in the distribution of virulence factor genes may be due to the fitness cost associated with methicillin resistance.

  18. Characterisation of virulence genes in methicillin susceptible and resistant Staphylococcus aureus isolates from a paediatric population in a university hospital of Medellín, Colombia.

    Science.gov (United States)

    Jiménez, Judy Natalia; Ocampo, Ana María; Vanegas, Johanna Marcela; Rodríguez, Erika Andrea; Garcés, Carlos Guillermo; Patiño, Luz Adriana; Ospina, Sigifredo; Correa, Margarita María

    2011-12-01

    Virulence and antibiotic resistance are significant determinants of the types of infections caused by Staphylococcus aureus and paediatric groups remain among the most commonly affected populations. The goal of this study was to characterise virulence genes of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains isolated from a paediatric population of a Colombian University Hospital during 2009. Sixty MSSA and MRSA isolates were obtained from paediatric patients between zero-14 years. We identified the genes encoding virulence factors, which included Panton-Valentine leucocidine (PVL), staphylococcal enterotoxins A-E, exfoliative toxins A and B and toxic shock syndrome toxin 1. Typing of the staphylococcal chromosome cassette mec (SCCmec) was performed in MRSA strains. The virulence genes were more diverse and frequent in MSSA than in MRSA isolates (83% vs. 73%). MRSA strains harboured SCCmec types IVc (60%), I (30%), IVa (7%) and V (3%). SCCmec type IVc isolates frequently carried the PVL encoding genes and harboured virulence determinants resembling susceptible strains while SCCmec type I isolates were often negative. PVL was not exclusive to skin and soft tissue infections. As previously suggested, these differences in the distribution of virulence factor genes may be due to the fitness cost associated with methicillin resistance.

  19. Virulence Genes among Enterococcus faecalis and Enterococcus faecium Isolated from Coastal Beaches and Human and Nonhuman Sources in Southern California and Puerto Rico

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    Donna M. Ferguson

    2016-01-01

    Full Text Available Most Enterococcus faecalis and E. faecium are harmless to humans; however, strains harboring virulence genes, including esp, gelE, cylA, asa1, and hyl, have been associated with human infections. E. faecalis and E. faecium are present in beach waters worldwide, yet little is known about their virulence potential. Here, multiplex PCR was used to compare the distribution of virulence genes among E. faecalis and E. faecium isolated from beaches in Southern California and Puerto Rico to isolates from potential sources including humans, animals, birds, and plants. All five virulence genes were found in E. faecalis and E. faecium from beach water, mostly among E. faecalis. gelE was the most common among isolates from all source types. There was a lower incidence of asa1, esp, cylA, and hyl genes among isolates from beach water, sewage, septage, urban runoff, sea wrack, and eelgrass as compared to human isolates, indicating that virulent strains of E. faecalis and E. faecium may not be widely disseminated at beaches. A higher frequency of asa1 and esp among E. faecalis from dogs and of asa1 among birds (mostly seagull suggests that further studies on the distribution and virulence potential of strains carrying these genes may be warranted.

  20. Data in support of quantitative proteomics to identify potential virulence regulators in Paracoccidioides brasiliensis isolates

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    Alexandre Keiji Tashima

    2015-12-01

    Full Text Available Paracoccidioides genus are the etiologic agents of paracoccidioidomycosis (PCM, a systemic mycosis endemic in Latin America. Few virulence factors have been identified in these fungi. This paper describes support data from the quantitative proteomics of Paracoccidioides brasiliensis attenuated and virulent isolates [1]. The protein compositions of two isolates of the Pb18 strain showing distinct infection profiles were quantitatively assessed by stable isotopic dimethyl labeling and proteomic analysis. The mass spectrometry and the analysis dataset have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with identifier PXD000804.

  1. OxyR-regulated catalase CatB promotes the virulence in rice via detoxifying hydrogen peroxide in Xanthomonas oryzae pv. oryzae.

    Science.gov (United States)

    Yu, Chao; Wang, Nu; Wu, Maosen; Tian, Fang; Chen, Huamin; Yang, Fenghuan; Yuan, Xiaochen; Yang, Ching-Hong; He, Chenyang

    2016-11-08

    To facilitate infection, Xanthomonas oryzae pv. oryzae (Xoo), the bacterial blight pathogen of rice, needs to degrade hydrogen peroxide (H 2 O 2 ) generated by the host defense response via a mechanism that is mediated by the transcriptional regulator OxyR. The catalase (CAT) gene catB has previously been shown to belong to the OxyR regulon in Xoo. However, its expression patterns and function in H 2 O 2 detoxification and bacterial pathogenicity on rice remain to be elucidated. The catB gene encodes a putative catalase and is highly conserved in the sequenced strains of Xanthomonas spp. β-galactosidase analysis and electrophoretic mobility shift assays (EMSA) showed that OxyR positively regulated the transcription of catB by directly binding to its promoter region. The quantitative real-time PCR (qRT-PCR) assays revealed that the expression levels of catB and oxyR were significantly induced by H 2 O 2 . Deletion of catB or oxyR drastically impaired bacterial viability in the presence of extracellular H 2 O 2 and reduced CAT activity, demonstrating that CatB and OxyR contribute to H 2 O 2 detoxification in Xoo. In addition, ΔcatB and ΔoxyR displayed shorter bacterial blight lesions and reduced bacterial growth in rice compared to the wild-type stain, indicating that CatB and OxyR play essential roles in the virulence of Xoo. Transcription of catB is enhanced by OxyR in response to exogenous H 2 O 2 . CatB functions as an active catalase that is required for the full virulence of Xoo in rice.

  2. Characterization of Aeromonas hydrophila Wound Pathotypes by Comparative Genomic and Functional Analyses of Virulence Genes

    Science.gov (United States)

    Grim, Christopher J.; Kozlova, Elena V.; Sha, Jian; Fitts, Eric C.; van Lier, Christina J.; Kirtley, Michelle L.; Joseph, Sandeep J.; Read, Timothy D.; Burd, Eileen M.; Tall, Ben D.; Joseph, Sam W.; Horneman, Amy J.; Chopra, Ashok K.; Shak, Joshua R.

    2013-01-01

    ABSTRACT Aeromonas hydrophila has increasingly been implicated as a virulent and antibiotic-resistant etiologic agent in various human diseases. In a previously published case report, we described a subject with a polymicrobial wound infection that included a persistent and aggressive strain of A. hydrophila (E1), as well as a more antibiotic-resistant strain of A. hydrophila (E2). To better understand the differences between pathogenic and environmental strains of A. hydrophila, we conducted comparative genomic and functional analyses of virulence-associated genes of these two wound isolates (E1 and E2), the environmental type strain A. hydrophila ATCC 7966T, and four other isolates belonging to A. aquariorum, A. veronii, A. salmonicida, and A. caviae. Full-genome sequencing of strains E1 and E2 revealed extensive differences between the two and strain ATCC 7966T. The more persistent wound infection strain, E1, harbored coding sequences for a cytotoxic enterotoxin (Act), a type 3 secretion system (T3SS), flagella, hemolysins, and a homolog of exotoxin A found in Pseudomonas aeruginosa. Corresponding phenotypic analyses with A. hydrophila ATCC 7966T and SSU as reference strains demonstrated the functionality of these virulence genes, with strain E1 displaying enhanced swimming and swarming motility, lateral flagella on electron microscopy, the presence of T3SS effector AexU, and enhanced lethality in a mouse model of Aeromonas infection. By combining sequence-based analysis and functional assays, we characterized an A. hydrophila pathotype, exemplified by strain E1, that exhibited increased virulence in a mouse model of infection, likely because of encapsulation, enhanced motility, toxin secretion, and cellular toxicity. PMID:23611906

  3. In Vivo fitness associated with high virulence in a vertebrate virus is a complex trait regulated by host entry, replication, and shedding

    Science.gov (United States)

    Wargo, Andrew R.; Kurath, Gael

    2011-01-01

    The relationship between pathogen fitness and virulence is typically examined by quantifying only one or two pathogen fitness traits. More specifically, it is regularly assumed that within-host replication, as a precursor to transmission, is the driving force behind virulence. In reality, many traits contribute to pathogen fitness, and each trait could drive the evolution of virulence in different ways. Here, we independently quantified four viral infection cycle traits, namely, host entry, within-host replication, within-host coinfection fitness, and shedding, in vivo, in the vertebrate virus Infectious hematopoietic necrosis virus (IHNV). We examined how each of these stages of the viral infection cycle contributes to the fitness of IHNV genotypes that differ in virulence in rainbow trout. This enabled us to determine how infection cycle fitness traits are independently associated with virulence. We found that viral fitness was independently regulated by each of the traits examined, with the largest impact on fitness being provided by within-host replication. Furthermore, the more virulent of the two genotypes of IHNV we used had advantages in all of the traits quantified. Our results are thus congruent with the assumption that virulence and within-host replication are correlated but suggest that infection cycle fitness is complex and that replication is not the only trait associated with virulence.

  4. A Unique Fungal Two-Component System Regulates Stress Responses, Drug Sensitivity, Sexual Development, and Virulence of Cryptococcus neoformans

    Science.gov (United States)

    Bahn, Yong-Sun; Kojima, Kaihei; Cox, Gary M.

    2006-01-01

    The stress-activated mitogen-activated protein kinase (MAPK) pathway is widely used by eukaryotic organisms as a central conduit via which cellular responses to the environment effect growth and differentiation. The basidiomycetous human fungal pathogen Cryptococcus neoformans uniquely uses the stress-activated Pbs2-Hog1 MAPK system to govern a plethora of cellular events, including stress responses, drug sensitivity, sexual reproduction, and virulence. Here, we characterized a fungal “two-component” system that controls these fundamental cellular functions via the Pbs2-Hog1 MAPK cascade. A typical response regulator, Ssk1, modulated all Hog1-dependent phenotypes by controlling Hog1 phosphorylation, indicating that Ssk1 is the major upstream signaling component of the Pbs2-Hog1 pathway. A second response regulator, Skn7, governs sensitivity to Na+ ions and the antifungal agent fludioxonil, negatively controls melanin production, and functions independently of Hog1 regulation. To control these response regulators, C. neoformans uses multiple sensor kinases, including two-component–like (Tco) 1 and Tco2. Tco1 and Tco2 play shared and distinct roles in stress responses and drug sensitivity through the Hog1 MAPK system. Furthermore, each sensor kinase mediates unique cellular functions for virulence and morphological differentiation. Our findings highlight unique adaptations of this global two-component MAPK signaling cascade in a ubiquitous human fungal pathogen. PMID:16672377

  5. Molecular evolution of the two-component system BvgAS involved in virulence regulation in Bordetella.

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    Julien Herrou

    Full Text Available The whooping cough agent Bordetella pertussis is closely related to Bordetella bronchiseptica, which is responsible for chronic respiratory infections in various mammals and is occasionally found in humans, and to Bordetella parapertussis, one lineage of which causes mild whooping cough in humans and the other ovine respiratory infections. All three species produce similar sets of virulence factors that are co-regulated by the two-component system BvgAS. We characterized the molecular diversity of BvgAS in Bordetella by sequencing the two genes from a large number of diverse isolates. The response regulator BvgA is virtually invariant, indicating strong functional constraints. In contrast, the multi-domain sensor kinase BvgS has evolved into two different types. The pertussis type is found in B. pertussis and in a lineage of essentially human-associated B. bronchiseptica, while the bronchiseptica type is associated with the majority of B. bronchiseptica and both ovine and human B. parapertussis. BvgS is monomorphic in B. pertussis, suggesting optimal adaptation or a recent population bottleneck. The degree of diversity of the bronchiseptica type BvgS is markedly different between domains, indicating distinct evolutionary pressures. Thus, absolute conservation of the putative solute-binding cavities of the two periplasmic Venus Fly Trap (VFT domains suggests that common signals are perceived in all three species, while the external surfaces of these domains vary more extensively. Co-evolution of the surfaces of the two VFT domains in each type and domain swapping experiments indicate that signal transduction in the periplasmic region may be type-specific. The two distinct evolutionary solutions for BvgS confirm that B. pertussis has emerged from a specific B. bronchiseptica lineage. The invariant regions of BvgS point to essential parts for its molecular mechanism, while the variable regions may indicate adaptations to different lifestyles. The

  6. Regulation of meiotic gene expression in plants

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    Adele eZhou

    2014-08-01

    Full Text Available With the recent advances in genomics and sequencing technologies, databases of transcriptomes representing many cellular processes have been built. Meiotic transcriptomes in plants have been studied in Arabidopsis thaliana, rice (Oryza sativa, wheat (Triticum aestivum, petunia (Petunia hybrida, sunflower (Helianthus annuus, and maize (Zea mays. Studies in all organisms, but particularly in plants, indicate that a very large number of genes are expressed during meiosis, though relatively few of them seem to be required for the completion of meiosis. In this review, we focus on gene expression at the RNA level and analyze the meiotic transcriptome datasets and explore expression patterns of known meiotic genes to elucidate how gene expression could be regulated during meiosis. We also discuss mechanisms, such as chromatin organization and non-coding RNAs, that might be involved in the regulation of meiotic transcription patterns.

  7. Natural variation in the VELVET gene bcvel1 affects virulence and light-dependent differentiation in Botrytis cinerea.

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    Julia Schumacher

    Full Text Available Botrytis cinerea is an aggressive plant pathogen causing gray mold disease on various plant species. In this study, we identified the genetic origin for significantly differing phenotypes of the two sequenced B. cinerea isolates, B05.10 and T4, with regard to light-dependent differentiation, oxalic acid (OA formation and virulence. By conducting a map-based cloning approach we identified a single nucleotide polymorphism (SNP in an open reading frame encoding a VELVET gene (bcvel1. The SNP in isolate T4 results in a truncated protein that is predominantly found in the cytosol in contrast to the full-length protein of isolate B05.10 that accumulates in the nuclei. Deletion of the full-length gene in B05.10 resulted in the T4 phenotype, namely light-independent conidiation, loss of sclerotial development and oxalic acid production, and reduced virulence on several host plants. These findings indicate that the identified SNP represents a loss-of-function mutation of bcvel1. In accordance, the expression of the B05.10 copy in T4 rescued the wild-type/B05.10 phenotype. BcVEL1 is crucial for full virulence as deletion mutants are significantly hampered in killing and decomposing plant tissues. However, the production of the two best known secondary metabolites, the phytotoxins botcinic acid and botrydial, are not affected by the deletion of bcvel1 indicating that other factors are responsible for reduced virulence. Genome-wide expression analyses of B05.10- and Δbcvel1-infected plant material revealed a number of genes differentially expressed in the mutant: while several protease- encoding genes are under-expressed in Δbcvel1 compared to the wild type, the group of over-expressed genes is enriched for genes encoding sugar, amino acid and ammonium transporters and glycoside hydrolases reflecting the response of Δbcvel1 mutants to nutrient starvation conditions.

  8. Virulence-related genes, adhesion and invasion of some Yersinia enterocolitica-like strains suggests its pathogenic potential.

    Science.gov (United States)

    Imori, Priscilla F M; Passaglia, Jaqueline; Souza, Roberto A; Rocha, Lenaldo B; Falcão, Juliana P

    2017-03-01

    Yersina enterocolitica-like species have not been extensively studied regarding its pathogenic potential. This work aimed to assess the pathogenic potential of some Y. enterocolitica-like strains by evaluating the presence of virulence-related genes by PCR and their ability to adhere to and invade Caco-2 and HEp-2 cells. A total of 50 Y. frederiksenii, 55 Y. intermedia and 13 Y. kristensenii strains were studied. The strains contained the following genes: Y. frederiksenii, fepA(44%), fes(44%) and ystB(18%); Y. intermedia, ail(53%), fepA (35%), fepD(2%), fes(97%), hreP(2%), ystB(2%) and tccC(35%); Y. kristensenii, ail(62%), ystB(23%), fepA(77%), fepD(54%), fes(54%) and hreP(77%). Generally, the Y. enterocolitica-like strains had a reduced ability to adhere to and invade mammalian cells compared to the highly pathogenic Y. enterocolitica 8081. However, Y. kristensenii FCF410 and Y. frederiksenii FCF461 presented high invasion potentials in Caco-2 cells after five days of pre-incubation increased by 45- and 7.2-fold compared to Y. enterocolitica 8081, respectively; but, the ail gene was not detected in these strains. The presence of virulence-related genes in some of the Y. enterocolitica-like strains indicated their possible pathogenic potential. Moreover, the results suggest the existence of alternative virulence mechanisms and that the pathogenicity of Y. kristensenii and Y. frederiksenii may be strain-dependent. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Identification of Salmonella typhimurium Genes Required for Colonization of the Chicken Alimentary Tract and for Virulence in Newly Hatched Chicks

    Science.gov (United States)

    Turner, Arthur K.; Lovell, Margaret A.; Hulme, Scott D.; Zhang-Barber, Li; Barrow, Paul A.

    1998-01-01

    From a collection of 2,800 Tn5-TC1 transposon mutants of Salmonella typhimurium F98, 18 that showed reduced intestinal colonization of 3-week-old chicks were identified. The sites of transposon insertion were determined for most of the mutants and included insertions in the lipopolysaccharide biosynthesis genes rfaK, rfaY, rfbK, and rfbB and the genes dksA, clpB, hupA, and sipC. In addition, identification was made of an insertion into a novel gene that encodes a protein showing similarity to the IIC component of the mannose class of phosphoenolpyruvate-carbohydrate phosphotransferase systems, which we putatively called ptsC. Transduction of most of the transposon mutations to a fresh S. typhimurium F98 genetic background and construction of defined mutations in the rfbK, dksA, hupA, sipC, and ptsC genes of S. typhimurium F98 supported the role in colonization of all but the pts locus. The virulence of the rfbK, dksA, hupA, sipC, and ptsC defined mutants and clpB and rfaY transductants in 1-day-old chicks was tested. All but the ptsC and rfaY mutants were attenuated for virulence. A number of other phenotypes associated with some of the mutations are described. PMID:9573095

  10. Ellagic acid derivatives from Terminalia chebula Retz. downregulate the expression of quorum sensing genes to attenuate Pseudomonas aeruginosa PAO1 virulence.

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    Sajal Sarabhai

    Full Text Available BACKGROUND: Burgeoning antibiotic resistance in Pseudomonas aeruginosa has necessitated the development of anti pathogenic agents that can quench acylhomoserine lactone (AHL mediated QS with least risk of resistance. This study explores the anti quorum sensing potential of T. chebula Retz. and identification of probable compounds(s showing anti QS activity and the mechanism of attenuation of P. aeruginosa PAO1 virulence factors. METHODS AND RESULTS: Methanol extract of T. chebula Retz. fruit showed anti QS activity using Agrobacterium tumefaciens A136. Bioactive fraction (F7, obtained by fractionation of methanol extract using Sephadex LH20, showed significant reduction (p<0.001 in QS regulated production of extracellular virulence factors in P. aeruginosa PAO1. Biofilm formation and alginate were significantly (p<0.05 reduced with enhanced (20% susceptibility to tobramycin. Real Time PCR of F7 treated P. aeruginosa showed down regulation of autoinducer synthase (lasI and rhlI and their cognate receptor (lasR and rhlR genes by 89, 90, 90 and 93%, respectively. Electrospray Ionization Mass Spectrometry also showed 90 and 64% reduction in the production of 3-oxo-C(12HSL and C(4HSL after treatment. Decrease in AHLs as one of the mechanisms of quorum quenching by F7 was supported by the reversal of inhibited swarming motility in F7-treated P. aeruginosa PAO1 on addition of C(4HSL. F7 also showed antagonistic activity against 3-oxo-C(12HSL-dependent QS in E. coli bioreporter. C. elegans fed on F7-treated P. aeruginosa showed enhanced survival with LT50 increasing from 24 to 72 h. LC-ESI-MS of F7 revealed the presence of ellagic acid derivatives responsible for anti QS activity in T. chebula extract. CONCLUSIONS: This is the first report on anti QS activity of T. chebula fruit linked to EADs which down regulate the expression of lasIR and rhlIR genes with concomitant decrease in AHLs in P. aeruginosa PAO1 causing attenuation of its virulence factors

  11. The Riemerella anatipestifer AS87_01735 Gene Encodes Nicotinamidase PncA, an Important Virulence Factor.

    Science.gov (United States)

    Wang, Xiaolan; Liu, Beibei; Dou, Yafeng; Fan, Hongjie; Wang, Shaohui; Li, Tao; Ding, Chan; Yu, Shengqing

    2016-10-01

    Riemerella anatipestifer is a major bacterial pathogen that causes septicemic and exudative diseases in domestic ducks. In our previous study, we found that deletion of the AS87_01735 gene significantly decreased the bacterial virulence of R. anatipestifer strain Yb2 (mutant RA625). The AS87_01735 gene was predicted to encode a nicotinamidase (PncA), a key enzyme that catalyzes the conversion of nicotinamide to nicotinic acid, which is an important reaction in the NAD(+) salvage pathway. In this study, the AS87_01735 gene was expressed and identified as the PncA-encoding gene, using an enzymatic assay. Western blot analysis demonstrated that R. anatipestifer PncA was localized to the cytoplasm. The mutant strain RA625 (named Yb2ΔpncA in this study) showed a similar growth rate but decreased NAD(+) quantities in both the exponential and stationary phases in tryptic soy broth culture, compared with the wild-type strain Yb2. In addition, Yb2ΔpncA-infected ducks showed much lower bacterial loads in their blood, and no visible histological changes were observed in the heart, liver, and spleen. Furthermore, Yb2ΔpncA immunization of ducks conferred effective protection against challenge with the virulent wild-type strain Yb2. Our results suggest that the R. anatipestifer AS87_01735 gene encodes PncA, which is an important virulence factor, and that the Yb2ΔpncA mutant can be used as a novel live vaccine candidate. Riemerella anatipestifer is reported worldwide as a cause of septicemic and exudative diseases of domestic ducks. The pncA gene encodes a nicotinamidase (PncA), a key enzyme that catalyzes the conversion of nicotinamide to nicotinic acid, which is an important reaction in the NAD(+) salvage pathway. In this study, we identified and characterized the pncA-homologous gene AS87_01735 in R. anatipestifer strain Yb2. R. anatipestifer PncA is a cytoplasmic protein that possesses similar PncA activity, compared with other organisms. Generation of the pncA mutant Yb

  12. MaHog1, a Hog1-type mitogen-activated protein kinase gene, contributes to stress tolerance and virulence of the entomopathogenic fungus Metarhizium acridum.

    Science.gov (United States)

    Jin, Kai; Ming, Yue; Xia, Yu Xian

    2012-12-01

    Fungal biocontrol agents have great potential in integrated pest management. However, poor efficacy and sensitivity to various adverse factors have hampered their wide application. In eukaryotic cells, Hog1 kinase plays a critical role in stress responses. In this study, MaHog1 (GenBank accession no. EFY85878), encoding a member of the Hog1/Sty1/p38 mitogen-activated protein kinase family in Metarhizium (Me.) acridum, was identified. Targeted gene disruption was used to analyse the role of MaHog1 in virulence and tolerance of adverse factors. Mutants with MaHog1 depletion showed increased sensitivity to high osmotic stress, high temperature and oxidative stress, and exhibited remarkable resistance to cell wall-disturbing agents. These results suggest that Hog1 kinase has a conserved function in regulating multistress responses among fungi, and that MaHog1 might influence cell wall biogenesis in Me. acridum. Bioassays conducted with topical inoculation and intrahaemocoel injection revealed that MaHog1 is required for both penetration and postpenetration development of Me. acridum. MaHog1 disruption resulted in a significant reduction in virulence, likely due to the combination of a decrease in conidial germination, a reduction in appressorium formation and a decline in growth rate in insect haemolymph, which might be caused by impairing fungal tolerance of various stresses during infection.

  13. Prevalence, Virulence Genes, Antimicrobial Susceptibility, and Genetic Diversity of Bacillus cereus Isolated From Pasteurized Milk in China

    Directory of Open Access Journals (Sweden)

    Tiantian Gao

    2018-03-01

    Full Text Available Bacillus cereus is a common and important food-borne pathogen that can be found in various food products. Due to low-temperature sterilization for a short period of time, pasteurization is not sufficient for complete elimination of B. cereus in milk, thereby cause severe economic loss and food safety problems. It is therefore of paramount importance to perform risk assessment of B. cereus in pasteurized milk. In this study, we isolated B. cereus from pasteurized milk samples in different regions of China, and evaluated the contamination situation, existence of virulence genes, antibiotic resistance profile and genetic polymorphism of B. cereus isolates. Intriguingly, 70 samples (27% were found to be contaminated by B. cereus and the average contamination level was 111 MPN/g. The distribution of virulence genes was assessed toward 10 enterotoxigenic genes (hblA, hblC, hblD, nheA, nheB, nheC, cytK, entFM, bceT, and hlyII and one emetic gene (cesB. Forty five percent strains harbored enterotoxigenic genes hblACD and 93% isolates contained nheABC gene cluster. The positive rate of cytK, entFM, bceT, hlyII, and cesB genes were 73, 96, 75, 54, and 5%, respectively. Antibiotic susceptibility assessment showed that most of the isolates were resistant to β-lactam antibiotics and rifampicin, but susceptible to other antibiotics such as ciprofloxacin, gentamicin and chloramphenicol. Total multidrug-resistant population was about 34%. In addition, B. cereus isolates in pasteurized milk showed a high genetic diversity. In conclusion, our findings provide the first reference on the prevalence, contamination level and characteristics of B. cereus isolated from pasteurized milk in China, suggesting a potential high risk of B. cereus to public health and dairy industry.

  14. Comparative genomics reveals cotton-specific virulence factors in flexible genomic regions in Verticillium dahliae and evidence of horizontal gene transfer from Fusarium.

    Science.gov (United States)

    Chen, Jie-Yin; Liu, Chun; Gui, Yue-Jing; Si, Kai-Wei; Zhang, Dan-Dan; Wang, Jie; Short, Dylan P G; Huang, Jin-Qun; Li, Nan-Yang; Liang, Yong; Zhang, Wen-Qi; Yang, Lin; Ma, Xue-Feng; Li, Ting-Gang; Zhou, Lei; Wang, Bao-Li; Bao, Yu-Ming; Subbarao, Krishna V; Zhang, Geng-Yun; Dai, Xiao-Feng

    2018-01-01

    Verticillium dahliae isolates are most virulent on the host from which they were originally isolated. Mechanisms underlying these dominant host adaptations are currently unknown. We sequenced the genome of V. dahliae Vd991, which is highly virulent on its original host, cotton, and performed comparisons with the reference genomes of JR2 (from tomato) and VdLs.17 (from lettuce). Pathogenicity-related factor prediction, orthology and multigene family classification, transcriptome analyses, phylogenetic analyses, and pathogenicity experiments were performed. The Vd991 genome harbored several exclusive, lineage-specific (LS) genes within LS regions (LSRs). Deletion mutants of the seven genes within one LSR (G-LSR2) in Vd991 were less virulent only on cotton. Integration of G-LSR2 genes individually into JR2 and VdLs.17 resulted in significantly enhanced virulence on cotton but did not affect virulence on tomato or lettuce. Transcription levels of the seven LS genes in Vd991 were higher during the early stages of cotton infection, as compared with other hosts. Phylogenetic analyses suggested that G-LSR2 was acquired from Fusarium oxysporum f. sp. vasinfectum through horizontal gene transfer. Our results provide evidence that horizontal gene transfer from Fusarium to Vd991 contributed significantly to its adaptation to cotton and may represent a significant mechanism in the evolution of an asexual plant pathogen. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  15. ICESag37, a Novel Integrative and Conjugative Element Carrying Antimicrobial Resistance Genes and Potential Virulence Factors in Streptococcus agalactiae.

    Science.gov (United States)

    Zhou, Kaixin; Xie, Lianyan; Han, Lizhong; Guo, Xiaokui; Wang, Yong; Sun, Jingyong

    2017-01-01

    ICE Sag37 , a novel integrative and conjugative element carrying multidrug resistance and potential virulence factors, was characterized in a clinical isolate of Streptococcus agalactiae . Two clinical strains of S. agalactiae , Sag37 and Sag158, were isolated from blood samples of new-borns with bacteremia. Sag37 was highly resistant to erythromycin and tetracycline, and susceptible to levofloxacin and penicillin, while Sag158 was resistant to tetracycline and levofloxacin, and susceptible to erythromycin. Transfer experiments were performed and selection was carried out with suitable antibiotic concentrations. Through mating experiments, the erythromycin resistance gene was found to be transferable from Sag37 to Sag158. Sma I-PFGE revealed a new Sma I fragment, confirming the transfer of the fragment containing the erythromycin resistance gene. Whole genome sequencing and sequence analysis revealed a mobile element, ICE Sag37 , which was characterized using several molecular methods and in silico analyses. ICE Sag37 was excised to generate a covalent circular intermediate, which was transferable to S. agalactiae . Inverse PCR was performed to detect the circular form. A serine family integrase mediated its chromosomal integration into rumA , which is a known hotspot for the integration of streptococcal ICEs. The integration site was confirmed using PCR. ICE Sag37 carried genes for resistance to multiple antibiotics, including erythromycin [ erm(B) ], tetracycline [ tet(O) ], and aminoglycosides [ aadE, aphA , and ant(6) ]. Potential virulence factors, including a two-component signal transduction system ( nisK/nisR ), were also observed in ICE Sag37 . S1-PFGE analysis ruled out the existence of plasmids. ICE Sag37 is the first ICE Sa2603 family-like element identified in S. agalactiae carrying both resistance and potential virulence determinants. It might act as a vehicle for the dissemination of multidrug resistance and pathogenicity among S. agalactiae .

  16. Regulation of Metalloprotease Gene Expression in Vibrio vulnificus by a Vibrio harveyi LuxR Homologue

    Science.gov (United States)

    Shao, Chung-Ping; Hor, Lien-I

    2001-01-01

    Expression of the Vibrio vulnificus metalloprotease gene, vvp, was turned up rapidly when bacterial growth reached the late log phase. A similar pattern of expression has been found in the metalloprotease gene of Vibrio cholerae, and this has been shown to be regulated by a Vibrio harveyi LuxR-like transcriptional activator. To find out whether a LuxR homologue exists in V. vulnificus, a gene library of this organism was screened by colony hybridization using a probe derived from a sequence that is conserved in various luxR-like genes of vibrios. A gene containing a 618-bp open reading frame was identified and found to be identical to the smcR gene of V. vulnificus reported previously. An isogenic SmcR-deficient (RD) mutant was further constructed by an in vivo allelic exchange technique. This mutant exhibited an extremely low level of vvp transcription compared with that of the parent strain. On the other hand, the cytolysin gene, vvhA, was expressed at a higher level in the RD mutant than in the parent strain during the log phase of growth. These data suggested that SmcR might not only be a positive regulator of the protease gene but might also be involved in negative regulation of the cytolysin gene. Virulence of the RD mutant in either normal or iron-overloaded mice challenged by intraperitoneal injection was comparable to that of the parent strain, indicating that SmcR is not required for V. vulnificus virulence in mice. PMID:11157950

  17. Characterisation of virulence genes in methicillin susceptible and resistant Staphylococcus aureus isolates from a paediatric population in a university hospital of Medellín, Colombia

    OpenAIRE

    Jiménez,Judy Natalia; Ocampo,Ana María; Vanegas,Johanna Marcela; Rodríguez,Erika Andrea; Garcés,Carlos Guillermo; Patiño,Luz Adriana; Ospina,Sigifredo; Correa,Margarita María

    2011-01-01

    Virulence and antibiotic resistance are significant determinants of the types of infections caused by Staphylococcus aureus and paediatric groups remain among the most commonly affected populations. The goal of this study was to characterise virulence genes of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) strains isolated from a paediatric population of a Colombian University Hospital during 2009. Sixty MSSA and MRSA isolates were obtained from paediatric...

  18. Antimicrobial Effects of Blueberry, Raspberry, and Strawberry Aqueous Extracts and their Effects on Virulence Gene Expression in Vibrio cholerae.

    Science.gov (United States)

    Khalifa, Hazim O; Kamimoto, Maki; Shimamoto, Toshi; Shimamoto, Tadashi

    2015-11-01

    The antimicrobial effects of aqueous extracts of blueberry, raspberry, and strawberry on 13 pathogenic bacteria were evaluated. The minimum inhibitory concentrations and minimum bactericidal concentrations of the extracts were determined before and after neutralization to pH 7.03 ± 0.15. Both Gram-positive and Gram-negative pathogenic bacteria were selectively inhibited by the non-neutralized berries. Blueberry was the best inhibitor, and Vibrio and Listeria were the most sensitive bacteria. After neutralization, blueberry affected only Vibrio and Listeria, whereas the antimicrobial activities of raspberry and strawberry were abolished. The total contents of phenolics, flavonoids, and proanthocyanidins in the extracts were measured with colorimetric methods and were highest in strawberry, followed by raspberry, and then blueberry. We also studied the effects of sub-bactericidal concentrations of the three berry extracts on virulence gene expression in Vibrio cholerae. Real-time quantitative reverse transcription-polymerase chain reaction revealed that the three berry extracts effectively repressed the transcription of the tcpA gene. Raspberry also repressed the transcription of the ctxA gene, whereas blueberry and strawberry did not. However, the three berry extracts did not affect the transcription of toxT. These results suggest that the three berry extracts exert potent antimicrobial effects and inhibit the expression of the virulence factors of V. cholerae. Copyright © 2015 John Wiley & Sons, Ltd.

  19. Inactivation of the Haemophilus ducreyi luxS gene affects the virulence of this pathogen in human subjects.

    Science.gov (United States)

    Labandeira-Rey, Maria; Janowicz, Diane M; Blick, Robert J; Fortney, Kate R; Zwickl, Beth; Katz, Barry P; Spinola, Stanley M; Hansen, Eric J

    2009-08-01

    Haemophilus ducreyi 35000HP contains a homologue of the luxS gene, which encodes an enzyme that synthesizes autoinducer 2 (AI-2) in other gram-negative bacteria. H. ducreyi 35000HP produced AI-2 that functioned in a Vibrio harveyi-based reporter system. A H. ducreyi luxS mutant was constructed by insertional inactivation of the luxS gene and lost the ability to produce AI-2. Provision of the H. ducreyi luxS gene in trans partially restored AI-2 production by the mutant. The luxS mutant was compared with its parent for virulence in the human challenge model of experimental chancroid. The pustule-formation rate in 5 volunteers was 93.3% (95% confidence interval, 81.7%-99.9%) at 15 parent sites and 60.0% (95% confidence interval, 48.3%-71.7%) at 15 mutant sites (1-tailed P < .001). Thus, the luxS mutant was partially attenuated for virulence. This is the first report of AI-2 production contributing to the pathogenesis of a genital ulcer disease.

  20. Polyamines are essential for virulence in Salmonella enterica serovar Gallinarum despite evolutionary decay of polyamine biosynthesis genes

    DEFF Research Database (Denmark)

    Schroll, Casper; Christensen, Jens P.; Christensen, Henrik

    2014-01-01

    . Typhi and S. Gallinarum and happened through independent events. The remaining polyamine biosynthesis pathway was found to be essential for oral infection with S. Gallinarum since single and double mutants in speB and speE, encoding the pathways from agmatine to putrescine and from putrescine...... to putrescine. The first pathway is not active in S. Gallinarum and S. Typhi, and this prompted us to investigate the importance of polyamines for virulence in S. Gallinarum. Bioinformatic analysis of all sequenced genomes of Salmonella revealed that pseudogene formation of the speC gene was exclusive for S...

  1. Identification of a response regulator involved in surface attachment, cell-cell aggregation, exopolysaccharide production and virulence in the plant pathogen Xylella fastidiosa.

    Science.gov (United States)

    Voegel, Tanja M; Doddapaneni, Harshavardhan; Cheng, Davis W; Lin, Hong; Stenger, Drake C; Kirkpatrick, Bruce C; Roper, M Caroline

    2013-04-01

    Xylella fastidiosa, the causal agent of Pierce's disease of grapevine, possesses several two-component signal transduction systems that allow the bacterium to sense and respond to changes in its environment. Signals are perceived by sensor kinases that autophosphorylate and transfer the phosphate to response regulators (RRs), which direct an output response, usually by acting as transcriptional regulators. In the X. fastidiosa genome, 19 RRs were found. A site-directed knockout mutant in one unusual RR, designated XhpT, composed of a receiver domain and a histidine phosphotransferase output domain, was constructed. The resulting mutant strain was analysed for changes in phenotypic traits related to biofilm formation and gene expression using microarray analysis. We found that the xhpT mutant was altered in surface attachment, cell-cell aggregation, exopolysaccharide (EPS) production and virulence in grapevine. In addition, this mutant had an altered transcriptional profile when compared with wild-type X. fastidiosa in genes for several biofilm-related traits, such as EPS production and haemagglutinin adhesins. © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

  2. Involvement of a velvet protein FgVeA in the regulation of asexual development, lipid and secondary metabolisms and virulence in Fusarium graminearum.

    Directory of Open Access Journals (Sweden)

    Jinhua Jiang

    Full Text Available The velvet protein, VeA, is involved in the regulation of diverse cellular processes. In this study, we explored functions of FgVeA in the wheat head blight pathogen, Fusarium graminearum,using a gene replacement strategy. The FgVEA deletion mutant exhibited a reduction in aerial hyphae formation, hydrophobicity, and deoxynivalenol (DON biosynthesis. Deletion of FgVEA gene led to an increase in conidial production, but a delay in conidial germination. Pathogencity assays showed that the mutant was impaired in virulence on flowering wheat head. Sensitivity tests to various stresses exhibited that the FgVEA deletion mutant showed increased resistance to osmotic stress and cell wall-damaging agents, but increased sensitivity to iprodione and fludioxonil fungicides. Ultrastructural and histochemical analyses revealed that conidia of FgVeA deletion mutant contained an unusually high number of large lipid droplets, which is in agreement with the observation that the mutant accumulated a higher basal level of glycerol than the wild-type progenitor. Serial analysis of gene expression (SAGE in the FgVEA mutant confirmed that FgVeA was involved in various cellular processes. Additionally, six proteins interacting with FgVeA were identified by yeast two hybrid assays in current study. These results indicate that FgVeA plays a critical role in a variety of cellular processes in F. graminearum.

  3. Regulation of Gene Expression in Protozoa Parasites

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    Consuelo Gomez

    2010-01-01

    Full Text Available Infections with protozoa parasites are associated with high burdens of morbidity and mortality across the developing world. Despite extensive efforts to control the transmission of these parasites, the spread of populations resistant to drugs and the lack of effective vaccines against them contribute to their persistence as major public health problems. Parasites should perform a strict control on the expression of genes involved in their pathogenicity, differentiation, immune evasion, or drug resistance, and the comprehension of the mechanisms implicated in that control could help to develop novel therapeutic strategies. However, until now these mechanisms are poorly understood in protozoa. Recent investigations into gene expression in protozoa parasites suggest that they possess many of the canonical machineries employed by higher eukaryotes for the control of gene expression at transcriptional, posttranscriptional, and epigenetic levels, but they also contain exclusive mechanisms. Here, we review the current understanding about the regulation of gene expression in Plasmodium sp., Trypanosomatids, Entamoeba histolytica and Trichomonas vaginalis.

  4. Regulation of gene expression in protozoa parasites.

    Science.gov (United States)

    Gomez, Consuelo; Esther Ramirez, M; Calixto-Galvez, Mercedes; Medel, Olivia; Rodríguez, Mario A

    2010-01-01

    Infections with protozoa parasites are associated with high burdens of morbidity and mortality across the developing world. Despite extensive efforts to control the transmission of these parasites, the spread of populations resistant to drugs and the lack of effective vaccines against them contribute to their persistence as major public health problems. Parasites should perform a strict control on the expression of genes involved in their pathogenicity, differentiation, immune evasion, or drug resistance, and the comprehension of the mechanisms implicated in that control could help to develop novel therapeutic strategies. However, until now these mechanisms are poorly understood in protozoa. Recent investigations into gene expression in protozoa parasites suggest that they possess many of the canonical machineries employed by higher eukaryotes for the control of gene expression at transcriptional, posttranscriptional, and epigenetic levels, but they also contain exclusive mechanisms. Here, we review the current understanding about the regulation of gene expression in Plasmodium sp., Trypanosomatids, Entamoeba histolytica and Trichomonas vaginalis.

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

    Science.gov (United States)

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

    2017-03-20

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

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

    Directory of Open Access Journals (Sweden)

    Akebe Luther King Abia

    2017-03-01

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

  7. Brucella, nitrogen and virulence.

    Science.gov (United States)

    Ronneau, Severin; Moussa, Simon; Barbier, Thibault; Conde-Álvarez, Raquel; Zuniga-Ripa, Amaia; Moriyon, Ignacio; Letesson, Jean-Jacques

    2016-08-01

    The brucellae are α-Proteobacteria causing brucellosis, an important zoonosis. Although multiplying in endoplasmic reticulum-derived vacuoles, they cause no cell death, suggesting subtle but efficient use of host resources. Brucellae are amino-acid prototrophs able to grow with ammonium or use glutamate as the sole carbon-nitrogen source in vitro. They contain more than twice amino acid/peptide/polyamine uptake genes than the amino-acid auxotroph Legionella pneumophila, which multiplies in a similar vacuole, suggesting a different nutritional strategy. During these two last decades, many mutants of key actors in nitrogen metabolism (transporters, enzymes, regulators, etc.) have been described to be essential for full virulence of brucellae. Here, we review the genomic and experimental data on Brucella nitrogen metabolism and its connection with virulence. An analysis of various aspects of this metabolism (transport, assimilation, biosynthesis, catabolism, respiration and regulation) has highlighted differences and similarities in nitrogen metabolism with other α-Proteobacteria. Together, these data suggest that, during their intracellular life cycle, the brucellae use various nitrogen sources for biosynthesis, catabolism and respiration following a strategy that requires prototrophy and a tight regulation of nitrogen use.

  8. VRprofile: gene-cluster-detection-based profiling of virulence and antibiotic resistance traits encoded within genome sequences of pathogenic bacteria.

    Science.gov (United States)

    Li, Jun; Tai, Cui; Deng, Zixin; Zhong, Weihong; He, Yongqun; Ou, Hong-Yu

    2017-01-10

    VRprofile is a Web server that facilitates rapid investigation of virulence and antibiotic resistance genes, as well as extends these trait transfer-related genetic contexts, in newly sequenced pathogenic bacterial genomes. The used backend database MobilomeDB was firstly built on sets of known gene cluster loci of bacterial type III/IV/VI/VII secretion systems and mobile genetic elements, including integrative and conjugative elements, prophages, class I integrons, IS elements and pathogenicity/antibiotic resistance islands. VRprofile is thus able to co-localize the homologs of these conserved gene clusters using HMMer or BLASTp searches. With the integration of the homologous gene cluster search module with a sequence composition module, VRprofile has exhibited better performance for island-like region predictions than the other widely used methods. In addition, VRprofile also provides an integrated Web interface for aligning and visualizing identified gene clusters with MobilomeDB-archived gene clusters, or a variety set of bacterial genomes. VRprofile might contribute to meet the increasing demands of re-annotations of bacterial variable regions, and aid in the real-time definitions of disease-relevant gene clusters in pathogenic bacteria of interest. VRprofile is freely available at http://bioinfo-mml.sjtu.edu.cn/VRprofile. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  9. Effect Of Spaceflight On Microbial Gene Expression And Virulence: Preliminary Results From Microbe Payload Flown On-Board STS-115

    Science.gov (United States)

    Wilson, J. W.; HonerzuBentrup, K,; Schurr, M. J.; Buchanan, K.; Morici, L.; Hammond, T.; Allen, P.; Baker, C.; Ott, C. M.; Nelman-Gonzalez M.; hide

    2007-01-01

    Human presence in space, whether permanent or temporary, is accompanied by the presence of microbes. However, the extent of microbial changes in response to spaceflight conditions and the corresponding changes to infectious disease risk is unclear. Previous studies have indicated that spaceflight weakens the immune system in humans and animals. In addition, preflight and in-flight monitoring of the International Space Station (ISS) and other spacecraft indicates the presence of opportunistic pathogens and the potential of obligate pathogens. Altered antibiotic resistance of microbes in flight has also been shown. As astronauts and cosmonauts live for longer periods in a closed environment, especially one using recycled water and air, there is an increased risk to crewmembers of infectious disease events occurring in-flight. Therefore, understanding how the space environment affects microorganisms and their disease potential is critically important for spaceflight missions and requires further study. The goal of this flight experiment, operationally called MICROBE, is to utilize three model microbial pathogens, Salmonella typhimurium, Pseudomonas aeruginosa, and Candida albicans to examine the global effects of spaceflight on microbial gene expression and virulence attributes. Specifically, the aims are (1) to perform microarray-mediated gene expression profiling of S. typhimurium, P. aeruginosa, and C. albicans, in response to spaceflight in comparison to ground controls and (2) to determine the effect of spaceflight on the virulence potential of these microorganisms immediately following their return from spaceflight using murine models. The model microorganisms were selected as they have been isolated from preflight or in-flight monitoring, represent different degrees of pathogenic behavior, are well characterized, and have sequenced genomes with available microarrays. In particular, extensive studies of S. typhimurium by the Principal Investigator, Dr. Nickerson

  10. Quantitative proteomics unravels that the post-transcriptional regulator Crc modulates the generation of vesicles and secreted virulence determinants of Pseudomonas aeruginosa.

    Science.gov (United States)

    Reales-Calderón, Jose Antonio; Corona, Fernando; Monteoliva, Lucía; Gil, Concha; Martínez, Jose Luis

    2015-09-08

    Recent research indicates that the post-transcriptional regulator Crc modulates susceptibility to antibiotics and virulence in Pseudomonas aeruginosa. Several P. aeruginosa virulence factors are secreted or engulfed in vesicles. To decipher the Crc modulation of P. aeruginosa virulence, we constructed a crc deficient mutant and measure the proteome associated extracellular vesicles and the vesicle-free secretome using iTRAQ. Fifty vesicle-associated proteins were more abundant and 14 less abundant in the crc-defective strain, whereas 37 were more abundant and 17 less abundant in the vesicle-free secretome. Among them, virulence determinants, such as ToxA, protease IV, azurin, chitin-binding protein, PlcB and Hcp1, were less abundant in the crc-defective mutant. Transcriptomic analysis revealed that some of the observed changes were post-transcriptional and, thus, could be attributed to a direct Crc regulatory role; whereas, for other differentially secreted proteins, the regulatory role was likely indirect. We also observed that the crc mutant presented an impaired vesicle-associated secretion of quorum sensing signal molecules and less cytotoxicity than its wild-type strain. Our results offer new insights into the mechanisms by which Crc regulates P. aeruginosa virulence, through the modulation of vesicle formation and secretion of both virulence determinants and quorum sensing signals. This article is part of a Special Issue entitled: HUPO 2014. Published by Elsevier B.V.

  11. Chlamydia trachomatis In Vivo to In Vitro Transition Reveals Mechanisms of Phase Variation and Down-Regulation of Virulence Factors.

    Directory of Open Access Journals (Sweden)

    Vítor Borges

    Full Text Available Research on the obligate intracellular bacterium Chlamydia trachomatis demands culture in cell-lines, but the adaptive process behind the in vivo to in vitro transition is not understood. We assessed the genomic and transcriptomic dynamics underlying C. trachomatis in vitro adaptation of strains representing the three disease groups (ocular, epithelial-genital and lymphogranuloma venereum propagated in epithelial cells over multiple passages. We found genetic features potentially underlying phase variation mechanisms mediating the regulation of a lipid A biosynthesis enzyme (CT533/LpxC, and the functionality of the cytotoxin (CT166 through an ON/OFF mechanism. We detected inactivating mutations in CT713/porB, a scenario suggesting metabolic adaptation to the available carbon source. CT135 was inactivated in a tropism-specific manner, with CT135-negative clones emerging for all epithelial-genital populations (but not for LGV and ocular populations and rapidly increasing in frequency (~23% mutants per 10 passages. RNA-sequencing analyses revealed that a deletion event involving CT135 impacted the expression of multiple virulence factors, namely effectors known to play a role in the C. trachomatis host-cell invasion or subversion (e.g., CT456/Tarp, CT694, CT875/TepP and CT868/ChlaDub1. This reflects a scenario of attenuation of C. trachomatis virulence in vitro, which may take place independently or in a cumulative fashion with the also observed down-regulation of plasmid-related virulence factors. This issue may be relevant on behalf of the recent advances in Chlamydia mutagenesis and transformation where culture propagation for selecting mutants/transformants is mandatory. Finally, there was an increase in the growth rate for all strains, reflecting gradual fitness enhancement over time. In general, these data shed light on the adaptive process underlying the C. trachomatis in vivo to in vitro transition, and indicates that it would be prudent to

  12. Virulence Characterization of Salmonella enterica by a New Microarray: Detection and Evaluation of the Cytolethal Distending Toxin Gene Activity in the Unusual Host S. Typhimurium.

    Directory of Open Access Journals (Sweden)

    Rui Figueiredo

    Full Text Available Salmonella enterica is a zoonotic foodborne pathogen that causes acute gastroenteritis in humans. We assessed the virulence potential of one-hundred and six Salmonella strains isolated from food animals and products. A high through-put virulence genes microarray demonstrated Salmonella Pathogenicity Islands (SPI and adherence genes were highly conserved, while prophages and virulence plasmid genes were variably present. Isolates were grouped by serotype, and virulence plasmids separated S. Typhimurium in two clusters. Atypical microarray results lead to whole genome sequencing (WGS of S. Infantis Sal147, which identified deletion of thirty-eight SPI-1 genes. Sal147 was unable to invade HeLa cells and showed reduced mortality in Galleria mellonella infection model, in comparison to a SPI-1 harbouring S. Infantis. Microarray and WGS of S. Typhimurium Sal199, established for the first time in S. Typhimurium presence of cdtB and other Typhi-related genes. Characterization of Sal199 showed cdtB genes were upstream of transposase IS911, and co-expressed with other Typhi-related genes. Cell cycle arrest, cytoplasmic distension, and nuclear enlargement were detected in HeLa cells infected by Sal199, but not with S. Typhimurium LT2. Increased mortality of Galleria was detected on infection with Sal199 compared to LT2. Thus, Salmonella isolates were rapidly characterized using a high through-put microarray; helping to identify unusual virulence features which were corroborated by further characterisation. This work demonstrates that the use of suitable screening methods for Salmonella virulence can help assess the potential risk associated with certain Salmonella to humans. Incorporation of such methodology into surveillance could help reduce the risk of emergence of epidemic Salmonella strains.

  13. Antisense long noncoding RNAs regulate var gene activation in the malaria parasite Plasmodium falciparum.

    Science.gov (United States)

    Amit-Avraham, Inbar; Pozner, Guy; Eshar, Shiri; Fastman, Yair; Kolevzon, Netanel; Yavin, Eylon; Dzikowski, Ron

    2015-03-03

    The virulence of Plasmodium falciparum, the causative agent of the deadliest form of human malaria, is attributed to its ability to evade human immunity through antigenic variation. These parasites alternate between expression of variable antigens, encoded by members of a multicopy gene family named var. Immune evasion through antigenic variation depends on tight regulation of var gene expression, ensuring that only a single var gene is expressed at a time while the rest of the family is maintained transcriptionally silent. Understanding how a single gene is chosen for activation is critical for understanding mutually exclusive expression but remains a mystery. Here, we show that antisense long noncoding RNAs (lncRNAs) initiating from var introns are associated with the single active var gene at the time in the cell cycle when the single var upstream promoter is active. We demonstrate that these antisense transcripts are incorporated into chromatin, and that expression of these antisense lncRNAs in trans triggers activation of a silent var gene in a sequence- and dose-dependent manner. On the other hand, interference with these lncRNAs using complement peptide nucleic acid molecules down-regulated the active var gene, erased the epigenetic memory, and induced expression switching. Altogether, our data provide evidence that these antisense lncRNAs play a key role in regulating var gene activation and mutually exclusive expression.

  14. Inhibition of Virulence Gene Expression in Staphylococcus aureus by Novel Depsipeptides from a Marine Photobacterium

    Directory of Open Access Journals (Sweden)

    Lone Gram

    2011-12-01

    Full Text Available During a global research expedition, more than five hundred marine bacterial strains capable of inhibiting the growth of pathogenic bacteria were collected. The purpose of the present study was to determine if these marine bacteria are also a source of compounds that interfere with the agr quorum sensing system that controls virulence gene expression in Staphylococcus aureus. Using a gene reporter fusion bioassay, we recorded agr interference as enhanced expression of spa, encoding Protein A, concomitantly with reduced expression of hla, encoding α-hemolysin, and rnaIII encoding RNAIII, the effector molecule of agr. A marine Photobacterium produced compounds interfering with agr in S. aureus strain 8325-4, and bioassay-guided fractionation of crude extracts led to the isolation of two novel cyclodepsipeptides, designated solonamide A and B. Northern blot analysis confirmed the agr interfering activity of pure solonamides in both S. aureus strain 8325-4 and the highly virulent, community-acquired strain USA300 (CA-MRSA. To our knowledge, this is the first report of inhibitors of the agr system by a marine bacterium.

  15. Endogenous CO2 may inhibit bacterial growth and induce virulence gene expression in enteropathogenic Escherichia coli.

    Science.gov (United States)

    Martínez, Haydee; Buhse, Thomas; Rivera, Marco; Parmananda, P; Ayala, Guadalupe; Sánchez, Joaquín

    2012-07-01

    Analysis of the growth kinetics of enteropathogenic Escherichia coli (EPEC) revealed that growth was directly proportional to the ratio between the exposed surface area and the liquid culture volume (SA/V). It was hypothesized that this bacterial behavior was caused by the accumulation of an endogenous volatile growth inhibitor metabolite whose escape from the medium directly depended on the SA/V. The results of this work support the theory that an inhibitor is produced and indicate that it is CO(2). We also report that concomitant to the accumulation of CO(2), there is secretion of the virulence-related EspB and EspC proteins from EPEC. We therefore postulate that endogenous CO(2) may have an effect on both bacterial growth and virulence. Copyright © 2012 Elsevier Ltd. All rights reserved.

  16. Norlichexanthone Reduces Virulence Gene Expression and Biofilm Formation in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Baldry, Mara; Nielsen, Anita; Bojer, Martin S.

    2016-01-01

    Staphylococcus aureus is a serious human pathogen and antibiotic resistant, community-associated strains, such as the methicillin resistant S. aureus (MRSA) strain USA300, continue to spread. To avoid resistance, anti-virulence therapy has been proposed where toxicity is targeted rather than...... viability. Previously we have shown that norlichexanthone, a small non-reduced tricyclic polyketide produced by fungi and lichens, reduces expression of hla encoding α-hemolysin as well as the regulatory RNAIII of the agr quorum sensing system in S. aureus 8325-4. The aim of the present study was to further...... SaeRS system. Our data show that norlichexanthone treatment reduces expression of key virulence factors in CA-MRSA strain USA300 via AgrA binding and represses biofilm formation....

  17. Identification of Russian wheat aphid (Homoptera : Aphididae) populations virulent to the Dn4 resistance gene

    Czech Academy of Sciences Publication Activity Database

    Smith, C. M.; Belay, T.; Stauffer, CH.; Starý, Petr; Kubečková, I.; Starkey, S.

    2004-01-01

    Roč. 97, č. 3 (2004), s. 1112-1117 ISSN 0022-0493 R&D Projects: GA AV ČR IBS5007102 Grant - others:Kansas Agricultural Experiment Station(US) 04-120-J Institutional research plan: CEZ:AV0Z5007907 Keywords : Diuraphis noxia * barley * virulence Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 1.283, year: 2004

  18. Pandemic serotypes of Vibrio cholerae isolated from ships' ballast tanks and coastal waters: assessment of antibiotic resistance and virulence genes (tcpA and ctxA).

    Science.gov (United States)

    Dobbs, Fred C; Goodrich, Amanda L; Thomson, Frank K; Hynes, Wayne

    2013-05-01

    There is concern that ships' ballasting operations may disseminate Vibrio cholerae to ports throughout the world. Given evidence that the bacterium is indeed transported by ships, we isolated pandemic serotypes O1 and O139 from ballast tanks and characterized them with respect to antibiotic resistance and virulence genes ctxA and tcpA. We carried out concurrent studies with V. cholerae isolated from coastal waters. Of 284 isolates, 30 were serotype O1 and 59 were serotype O139. These serotypes were overrepresented in ballast tanks relative to the coastal waters sampled. All locations, whether coastal waters or ballast tanks, yielded samples from which serotype O1, O139, or both were isolated. There were three groups among the 62 isolates for which antibiotic characterization was conclusive: those exhibiting β-lactamase activity and resistance to at least one of the 12 antibiotics tested; those negative for β-lactamase but having antibiotic resistance; those negative for β-lactamase and registering no antibiotic resistance. When present, antibiotic resistance in nearly all cases was to ampicillin; resistance to multiple antibiotics was uncommon. PCR assays revealed that none of the isolates contained the ctxA gene and only two isolates, one O139 and one O1, contained the tcpA gene; both isolates originated from ballast water. These results support the bacteriological regulations proposed by the International Maritime Association for discharged ballast water.

  19. Inactivation of the spxA1 or spxA2 gene of Streptococcus mutans decreases virulence in the rat caries model

    Science.gov (United States)

    Galvão, Lívia C.C.; Rosalen, Pedro L.; Rivera-Ramos, Isamar; Franco, Gilson C.N.; Kajfasz, Jessica K; Abranches, Jacqueline; Bueno-Silva, Bruno; Koo, Hyun; Lemos, José A.

    2016-01-01

    SUMMARY In oral biofilms, the major environmental challenges encountered by Streptococcus mutans are acid and oxidative stresses. Previously, we showed that the transcriptional regulators SpxA1 and SpxA2 are involved in general stress survival of S. mutans with SpxA1 playing a primary role in activation of antioxidant and detoxification strategies whereas SpxA2 serves as a back up activator of oxidative stress genes. We have also found that spxA1 mutant strains (ΔspxA1 and ΔspxA1ΔspxA2) are outcompeted by peroxigenic oral streptococci in vitro and have impaired abilities to colonize the teeth of rats fed a highly cariogenic diet. Here, we show that the Spx proteins can also exert regulatory roles in the expression of additional virulence attributes of S. mutans. Competence activation is significantly impaired in Δspx strains and the production of mutacin IV and V is virtually abolished in ΔspxA1 strains. Unexpectedly, the ΔspxA2 strain showed increased production of glucans from sucrose, without affecting the total amount of bacteria within biofilms when compared to the parent strain. By using the rat caries model, we showed that the capacity of the ΔspxA1 and ΔspxA2 strains to cause caries on smooth tooth surfaces is significantly impaired. The ΔspxA2 strain also formed fewer lesions on sulcal surfaces. This report reveals that global regulation via Spx contributes to the cariogenic potential of S. mutans and highlights the essentiality of animal models in the characterization of bacterial traits implicated in virulence. PMID:27037617

  20. Inactivation of the spxA1 or spxA2 gene of Streptococcus mutans decreases virulence in the rat caries model.

    Science.gov (United States)

    Galvão, L C C; Rosalen, P L; Rivera-Ramos, I; Franco, G C N; Kajfasz, J K; Abranches, J; Bueno-Silva, B; Koo, H; Lemos, J A

    2017-04-01

    In oral biofilms, the major environmental challenges encountered by Streptococcus mutans are acid and oxidative stresses. Previously, we showed that the transcriptional regulators SpxA1 and SpxA2 are involved in general stress survival of S. mutans with SpxA1 playing a primary role in activation of antioxidant and detoxification strategies whereas SpxA2 serves as a back up activator of oxidative stress genes. We have also found that spxA1 mutant strains (∆spxA1 and ∆spxA1∆spxA2) are outcompeted by peroxigenic oral streptococci in vitro and have impaired abilities to colonize the teeth of rats fed a highly cariogenic diet. Here, we show that the Spx proteins can also exert regulatory roles in the expression of additional virulence attributes of S. mutans. Competence activation is significantly impaired in Δspx strains and the production of mutacin IV and V is virtually abolished in ΔspxA1 strains. Unexpectedly, the ∆spxA2 strain showed increased production of glucans from sucrose, without affecting the total amount of bacteria within biofilms when compared with the parent strain. By using the rat caries model, we showed that the capacity of the ΔspxA1 and ΔspxA2 strains to cause caries on smooth tooth surfaces is significantly impaired. The ∆spxA2 strain also formed fewer lesions on sulcal surfaces. This report reveals that global regulation via Spx contributes to the cariogenic potential of S. mutans and highlights that animal models are essential in the characterization of bacterial traits implicated in virulence. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  1. Prevalence of genes encoding extracellular virulence factors among meticillin-resistant Staphylococcus aureus isolates from the University Hospital, Olomouc, Czech Republic.

    Science.gov (United States)

    Sauer, P; Síla, J; Stosová, T; Vecerová, R; Hejnar, P; Vágnerová, I; Kolár, M; Raclavsky, V; Petrzelová, J; Lovecková, Y; Koukalová, D

    2008-04-01

    A rather fast and complicated progression of an infection caused by some strains of Staphylococcus aureus could be associated with the expression and co-action of virulence factor complexes in these strains. This study screened the antibiotic susceptibility and prevalence of virulence markers in isolates of meticillin-resistant S. aureus (MRSA) obtained from patients hospitalized at the University Hospital in Olomouc, Czech Republic. A total of 100 isolates was screened for 13 genes encoding extracellular virulence determinants (tst, pvl, eta, etb, sea, seb, sec, sed, see, seg, seh, sei and sej) and for their distribution in sample types. Eighty-nine isolates were positive for at least one of the genes. Genes for etb, pvl, see and seh were not detected in any of the MRSA isolates. No statistically significant differences in the occurrence of the determinants studied among sample types were found.

  2. Oxygen Availability Influences Expression of Dickeya solani Genes Associated With Virulence in Potato (Solanum tuberosum L. and Chicory (Cichorium intybus L.

    Directory of Open Access Journals (Sweden)

    Wioletta Lisicka

    2018-03-01

    Full Text Available Dickeya solani is a Gram-negative necrotrophic, plant pathogenic bacterium able to cause symptoms in a variety of plant species worldwide. As a facultative anaerobe, D. solani is able to infect hosts under a broad range of oxygen concentrations found in plant environments. However, little is known about oxygen-dependent gene expression in Dickeya spp. that might contribute to its success as a pathogen. Using a Tn5 transposon, harboring a promoterless gusA reporter gene, 146 mutants of D. solani IPO2222 were identified that exhibited oxygen-regulated expression of the gene into which the insertion had occurred. Of these mutants 114 exhibited higher expression under normal oxygen conditions than hypoxic conditions while 32 were more highly expressed under hypoxic conditions. The plant host colonization potential and pathogenicity as well as phenotypes likely to contribute to the ecological fitness of D. solani, including growth rate, carbon and nitrogen source utilization, production of pectinolytic enzymes, proteases, cellulases and siderophores, swimming and swarming motility and the ability to form biofilm were assessed for 37 strains exhibiting the greatest oxygen-dependent change in gene expression. Eight mutants expressed decreased ability to cause disease symptoms when inoculated into potato tubers or chicory leaves and three of these also exhibited delayed colonization of potato plants and exhibited tissue specific differences in gene expression in these various host tissues. The genes interrupted in these eight mutants encoded proteins involved in fundamental bacterial metabolism, virulence, bacteriocin and proline transport, while three encoded hypothetical or unknown proteins. The implications of environmental oxygen concentration on the ability of D. solani to cause disease symptoms in potato are discussed.

  3. Global regulation of gene expression by the MafR protein of Enterococcus faecalis

    Directory of Open Access Journals (Sweden)

    Sofía eRuiz-Cruz

    2016-01-01

    Full Text Available Enterococcus faecalis is a natural inhabitant of the human gastrointestinal tract. However, as an opportunistic pathogen, it is able to colonize other host niches and cause life-threatening infections. Its adaptation to new environments involves global changes in gene expression. The EF3013 gene (here named mafR of E. faecalis strain V583 encodes a protein (MafR, 482 residues that has sequence similarity to global response regulators of the Mga/AtxA family. The enterococcal OG1RF genome also encodes the MafR protein (gene OG1RF_12293. In this work, we have identified the promoter of the mafR gene using several in vivo approaches. Moreover, we show that MafR influences positively the transcription of many genes on a genome-wide scale. The most significant target genes encode components of PTS-type membrane transporters, components of ABC-type membrane transporters, and proteins involved in the metabolism of carbon sources. Some of these genes were previously reported to be up-regulated during the growth of E. faecalis in blood and/or in human urine. Furthermore, we show that a mafR deletion mutant strain induces a significant lower degree of inflammation in the peritoneal cavity of mice, suggesting that enterococcal cells deficient in MafR are less virulent. Our work indicates that MafR is a global transcriptional regulator. It might facilitate the adaptation of E. faecalis to particular host niches and, therefore, contribute to its potential virulence.

  4. Regulation of methane genes and genome expression

    Energy Technology Data Exchange (ETDEWEB)

    John N. Reeve

    2009-09-09

    At the start of this project, it was known that methanogens were Archaeabacteria (now Archaea) and were therefore predicted to have gene expression and regulatory systems different from Bacteria, but few of the molecular biology details were established. The goals were then to establish the structures and organizations of genes in methanogens, and to develop the genetic technologies needed to investigate and dissect methanogen gene expression and regulation in vivo. By cloning and sequencing, we established the gene and operon structures of all of the “methane” genes that encode the enzymes that catalyze methane biosynthesis from carbon dioxide and hydrogen. This work identified unique sequences in the methane gene that we designated mcrA, that encodes the largest subunit of methyl-coenzyme M reductase, that could be used to identify methanogen DNA and establish methanogen phylogenetic relationships. McrA sequences are now the accepted standard and used extensively as hybridization probes to identify and quantify methanogens in environmental research. With the methane genes in hand, we used northern blot and then later whole-genome microarray hybridization analyses to establish how growth phase and substrate availability regulated methane gene expression in Methanobacterium thermautotrophicus ΔH (now Methanothermobacter thermautotrophicus). Isoenzymes or pairs of functionally equivalent enzymes catalyze several steps in the hydrogen-dependent reduction of carbon dioxide to methane. We established that hydrogen availability determine which of these pairs of methane genes is expressed and therefore which of the alternative enzymes is employed to catalyze methane biosynthesis under different environmental conditions. As were unable to establish a reliable genetic system for M. thermautotrophicus, we developed in vitro transcription as an alternative system to investigate methanogen gene expression and regulation. This led to the discovery that an archaeal protein

  5. A Novel Small RNA Regulates Tolerance and Virulence in Shigella flexneri by Responding to Acidic Environmental Changes

    Directory of Open Access Journals (Sweden)

    Ligui eWang

    2016-03-01

    Full Text Available Shigella flexneri is an important cause of bacillary dysentery in developing countries. Small regulatory RNAs (sRNAs play essential roles in diverse cellular processes. We found a novel sRNA Ssr1 based on RT-PCR, northern blot, and 5´RACE in S. flexneri. Ssr1 responds to acidic environmental changes, as shown by a strong linear correlation between the pH value and Ssr1 expression (R = 0.785, P < 0.05 using the qRT-PCR method. Deletion of Ssr1 results in growth retardation at pH values ranging from 5.0 to 7.0 (P < 0.05, and the survival rate was reduced by 22% in acidic conditions (pH 3.0. Additionally, virulence was significantly increased in an Ssr1 mutant strain, as revealed in a murine lung invasion model and survival model assays. By using the sTarPicker method and proteomic analysis, we considered that DnaK, which is a major factor that confers acidic stress tolerance, may be a direct target of Ssr1. We also found that Ssr1 may enhance virulence by directly targeting OmpA; this leads to altered expression of genes in the type three secretion system. This work provides new insight into the mechanism of adaptation to environmental stress and into the pathogenesis of Shigella.

  6. Short communication: Emergence of a new race of leaf rust with combined virulence to Lr14a and Lr72 genes on durum wheat

    Energy Technology Data Exchange (ETDEWEB)

    Soleiman, N.H; Solis, I.; Soliman, M.H.; Sillero, J.C.; Villegas, D.; Alvaro, F.; Royo, C.; Serra, J.; Ammar, K.; Martínez-Moreno, F.

    2016-11-01

    Leaf rust is a foliar disease caused by the fungus Puccinia triticina that may severely reduce durum wheat yield. Resistance to this pathogen is common in modern durum germplasm but is frequently based on Lr72 and Lr14a. After accounts of races with virulence to Lr14a gene in France in 2000, the present study reports the detection in 2013 for the first time of a new race with virulence to Lr14a and Lr72. The aim of this work was to characterize the virulence pattern of four Spanish isolates with virulence to Lr14a, and to discuss the consequences of this presence. Rusted leaves from cultivars ‘Don Jaime’ (Lr14a) and ‘Gallareta’ (Lr72) were collected in 2013 in the field at two Spanish sites, one in the south (near Cadiz) and another in the north (near Girona). Spores from single pustule for each cultivar and site were multiplied on susceptible cultivar ‘Don Rafael’. Then, the four isolates were inoculated on a set of 19 isogenic lines Thatcher to characterize their virulence spectrum. All isolates presented the same virulence pattern. They were virulent on both Lr14a and Lr72 and the race was named DBB/BS. This race was very similar to those reported in 2009-11, but with added virulence to Lr14a. The resistance based on Lr14a has therefore been overcome in Spain, by a new race that has likely emerged via stepwise mutation from the local predominating races. This information is important to guide breeders in their breeding programmes and gene deployment strategies. (Author)

  7. The LacI–Family Transcription Factor, RbsR, Is a Pleiotropic Regulator of Motility, Virulence, Siderophore and Antibiotic Production, Gas Vesicle Morphogenesis and Flotation in Serratia

    Directory of Open Access Journals (Sweden)

    Chin M. Lee

    2017-09-01

    Full Text Available Gas vesicles (GVs are proteinaceous, gas-filled organelles used by some bacteria to enable upward movement into favorable air/liquid interfaces in aquatic environments. Serratia sp. ATCC39006 (S39006 was the first enterobacterium discovered to produce GVs naturally. The regulation of GV assembly in this host is complex and part of a wider regulatory network affecting various phenotypes, including antibiotic biosynthesis. To identify new regulators of GVs, a comprehensive mutant library containing 71,000 insertion mutants was generated by random transposon mutagenesis and 311 putative GV-defective mutants identified. Three of these mutants were found to have a transposon inserted in a LacI family transcription regulator gene (rbsR of the putative ribose operon. Each of these rbsR mutants was GV-defective; no GVs were visible by phase contrast microscopy (PCM or transmission electron microscopy (TEM. GV deficiency was caused by the reduction of gvpA1 and gvrA transcription (the first genes of the two contiguous operons in the GV gene locus. Our results also showed that a mutation in rbsR was highly pleiotropic; the production of two secondary metabolites (carbapenem and prodigiosin antibiotics was abolished. Interestingly, the intrinsic resistance to the carbapenem antibiotic was not affected by the rbsR mutation. In addition, the production of a siderophore, cellulase and plant virulence was reduced in the mutant, whereas it exhibited increased swimming and swarming motility. The RbsR protein was predicted to bind to regions upstream of at least 18 genes in S39006 including rbsD (the first gene of the ribose operon and gvrA. Electrophoretic mobility shift assays (EMSA confirmed that RbsR bound to DNA sequences upstream of rbsD, but not gvrA. The results of this study indicate that RbsR is a global regulator that affects the modulation of GV biogenesis, but also with complex pleiotropic physiological impacts in S39006.

  8. Genome-wide Analysis of Gene Regulation

    DEFF Research Database (Denmark)

    Chen, Yun

    to protein: through epigenetic modifications, transcription regulators or post-transcriptional controls. The following papers concern several layers of gene regulation with questions answered by different HTS approaches. Genome-wide screening of epigenetic changes by ChIP-seq allowed us to study both spatial...... and temporal alterations of histone modifications (Papers I and II). Coupling the data with machine learning approaches, we established a prediction framework to assess the most informative histone marks as well as their most influential nucleosome positions in predicting the promoter usages. (Papers I...... they regulated or if the sites had global elevated usage rates by multiple TFs. Using RNA-seq, 5’end-seq in combination with depletion of 5’exonuclease as well as nonsensemediated decay (NMD) factors, we systematically analyzed NMD substrates as well as their degradation intermediates in human cells (Paper V...

  9. Inhibition of expression in Escherichia coli of a virulence regulator MglB of Francisella tularensis using external guide sequence technology.

    Directory of Open Access Journals (Sweden)

    Gaoping Xiao

    Full Text Available External guide sequences (EGSs have successfully been used to inhibit expression of target genes at the post-transcriptional level in both prokaryotes and eukaryotes. We previously reported that EGS accessible and cleavable sites in the target RNAs can rapidly be identified by screening random EGS (rEGS libraries. Here the method of screening rEGS libraries and a partial RNase T1 digestion assay were used to identify sites accessible to EGSs in the mRNA of a global virulence regulator MglB from Francisella tularensis, a Gram-negative pathogenic bacterium. Specific EGSs were subsequently designed and their activities in terms of the cleavage of mglB mRNA by RNase P were tested in vitro and in vivo. EGS73, EGS148, and EGS155 in both stem and M1 EGS constructs induced mglB mRNA cleavage in vitro. Expression of stem EGS73 and EGS155 in Escherichia coli resulted in significant reduction of the mglB mRNA level coded for the F. tularensis mglB gene inserted in those cells.

  10. Molecular analysis of virulent genes (coa and spa) of staphylococcus aureus involved in natural cases of bovine mastitis

    International Nuclear Information System (INIS)

    Khan, A.; Javed, M.T.; Mahmood, F.; Hussain, R.

    2013-01-01

    The present study was undertaken to determine the distribution and genotypic characteristics of Staphylococcus aureus isolates recovered from naturally occurring mastitis in cattle and buffaloes. For this purpose a total of 1445 lactating cattle (653) and buffaloes (792) present at two experimental livestock farms Okara (Bahadarnagar) and Sahiwal (Qadiarabad), in and around district Faisalabad and slaughtered at an abattoir due to low milk yield and were screened for mastitis. California Mastitis Test (CMT) was used to detect sub clinical mastitis. The positive quarter milk samples were collected for culturing of S. aureus isolates. taphylococcus aureus isolates were identified on the basis of growth features, biochemical characteristics, coagulase test and as well as amplification of coagulase (coa) and spa (spa-X) genes specific to its virulence. S. aureus isolates (n=265) were characterized by Polymerase chain reaction to determine the frequency of coagulase (coa) and spa (spa-X) genes. From these isolates the amplification of the coagulase (coa) gene yielded three different PCR products approximately 204bp to 490bp while spa (spa-X) gene produced five different products ranging in size from 190bp to 320bp. PCR revealed that from all the coagulase positive S. aureus isolates 261(98.5%) had spa (spa-X) gene. The results of the present study indicated that S. aureus isolates recovered from bovine mastitis were genetically different within and among the various herds which may provide essential and valuable strategies to control staphylococcal infections in future. (author)

  11. Molecular analysis of virulent genes (coa and spa) of staphylococcus aureus involved in natural cases of bovine mastitis

    Energy Technology Data Exchange (ETDEWEB)

    Khan, A.; Javed, M. T.; Mahmood, F. [University of Agriculture, Faisalabad (Pakistan). Dept. of Pathology; Hussain, R. [The Islamia Univ. of Bahawalpur, Pakistan (Pakistan). Dept. of Veterinary and Animal Sciences

    2013-12-15

    The present study was undertaken to determine the distribution and genotypic characteristics of Staphylococcus aureus isolates recovered from naturally occurring mastitis in cattle and buffaloes. For this purpose a total of 1445 lactating cattle (653) and buffaloes (792) present at two experimental livestock farms Okara (Bahadarnagar) and Sahiwal (Qadiarabad), in and around district Faisalabad and slaughtered at an abattoir due to low milk yield and were screened for mastitis. California Mastitis Test (CMT) was used to detect sub clinical mastitis. The positive quarter milk samples were collected for culturing of S. aureus isolates. taphylococcus aureus isolates were identified on the basis of growth features, biochemical characteristics, coagulase test and as well as amplification of coagulase (coa) and spa (spa-X) genes specific to its virulence. S. aureus isolates (n=265) were characterized by Polymerase chain reaction to determine the frequency of coagulase (coa) and spa (spa-X) genes. From these isolates the amplification of the coagulase (coa) gene yielded three different PCR products approximately 204bp to 490bp while spa (spa-X) gene produced five different products ranging in size from 190bp to 320bp. PCR revealed that from all the coagulase positive S. aureus isolates 261(98.5%) had spa (spa-X) gene. The results of the present study indicated that S. aureus isolates recovered from bovine mastitis were genetically different within and among the various herds which may provide essential and valuable strategies to control staphylococcal infections in future. (author)

  12. Gene Transcription and Virulence Potential of Listeria monocytogenes Strains After Exposure to Acidic and NaCl Stress

    DEFF Research Database (Denmark)

    Olesen, Inger; Vogensen, Finn Kvist; Jespersen, Lene

    2009-01-01

    transcription were observed both after exposure to shock (six genes) and after long-term adaptation to stress (18 genes). In the shock experiments, a transient induction of clpC and clpE was seen for both strains, while transient induction of sigB, inlA, and inlB was observed for strain 4140 only; actA was only...... induced in EGD-e after NaCl shock. The longterm stress experiments were included to imitate the stress conditions encountered by L. monocytogenes when present in food products. Long-term adaptation of EGD-e to acidic stress induced transcription of iap and repressed flaA, while genes related to stress......Gene transcription and virulence potential of two strains of Listeria monocytogenes, EGD-e and 4140, were compared by quantitative real-time polymerase chain reaction and in a Caco-2 in vitro model after exposure to acidic (pH 5.5) and NaCl (4.5% w=v) stress. Strain-dependent differences in gene...

  13. Dynamics of Vibrio with virulence genes detected in Pacific harbor seals (Phoca vitulina richardii) off California: implications for marine mammal health.

    Science.gov (United States)

    Hughes, Stephanie N; Greig, Denise J; Miller, Woutrina A; Byrne, Barbara A; Gulland, Frances M D; Harvey, James T

    2013-05-01

    Given their coastal site fidelity and opportunistic foraging behavior, harbor seals (Phoca vitulina) may serve as sentinels for coastal ecosystem health. Seals using urbanized coastal habitat can acquire enteric bacteria, including Vibrio that may affect their health. To understand Vibrio dynamics in seals, demographic and environmental factors were tested for predicting potentially virulent Vibrio in free-ranging and stranded Pacific harbor seals (Phoca vitulina richardii) off California. Vibrio prevalence did not vary with season and was greater in free-ranging seals (29 %, n = 319) compared with stranded seals (17 %, n = 189). Of the factors tested, location, turbidity, and/or salinity best predicted Vibrio prevalence in free-ranging seals. The relationship of environmental factors with Vibrio prevalence differed by location and may be related to oceanographic or terrestrial contributions to water quality. Vibrio parahaemolyticus, Vibrio alginolyticus, and Vibrio cholerae were observed in seals, with V. cholerae found almost exclusively in stranded pups and yearlings. Additionally, virulence genes (trh and tdh) were detected in V. parahaemolyticus isolates. Vibrio cholerae isolates lacked targeted virulence genes, but were hemolytic. Three out of four stranded pups with V. parahaemolyticus (trh+ and/or tdh+) died in rehabilitation, but the role of Vibrio in causing mortality is unclear, and Vibrio expression of virulence genes should be investigated. Considering that humans share the environment and food resources with seals, potentially virulent Vibrio observed in seals also may be of concern to human health.

  14. Distribution of virulence genes and genotyping of CTX-M-15-producing Klebsiella pneumoniae isolated from patients with community-acquired urinary tract infection (CA-UTI).

    Science.gov (United States)

    Ranjbar, Reza; Memariani, Hamed; Sorouri, Rahim; Memariani, Mojtaba

    2016-11-01

    Klebsiella pneumoniae is one of the most important agents of community-acquired urinary tract infection (CA-UTI). In addition to extended-spectrum β-lactamases (ESBLs), a number of virulence factors have been shown to play an important role in the pathogenesis of K. pneumoniae, including capsule, siderophores, and adhesins. Little is known about the genetic diversity and virulence content of the CTX-M-15-producing K. pneumoniae isolated from CA-UTI in Iran. A total of 152 K. pneumoniae isolates were collected from CA-UTI patients in Tehran from September 2015 through April 2016. Out of 152 isolates, 40 (26.3%) carried bla CTX-M-15 . PCR was performed for detection of virulence genes in CTX-M-15-producing isolates. Furthermore, all of these isolates were subjected to multiple-locus variable-number of tandem repeat (VNTR) analysis (MLVA). Using MLVA method, 36 types were identified. CTX-M-15-producing K. pneumoniae isolates were grouped into 5 clonal complexes (CCs). Of these isolates, mrkD was the most prevalent virulence gene (95%), followed by kpn (60%), rmpA (37.5%), irp (35%), and magA (2.5%). No correlation between MLVA types or CCs and virulence genes or antibiotic resistance patterns was observed. Overall, it is thought that CTX-M-15-producing K. pneumoniae strains isolated from CA-UTI have arisen from different clones. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Comparison of antibiotic resistance, virulence gene profiles, and pathogenicity of methicillin-resistant and methicillin-susceptible Staphylococcus aureus using a Caenorhabditis elegans infection model

    Science.gov (United States)

    Thompson, Terissa; Brown, Paul D

    2014-01-01

    Objectives: This study compared the presence of 35 virulence genes, resistance phenotypes to 11 anti-staphylococcal antibiotics, and pathogenicity in methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA). Methods: Multiplex PCR analysis was used to differentiate Staphylococcus aureus isolates (n = 102) based on characterization of the Staphylococcal Cassette Chromosome mec (SCCmec). Singleplex and multiplex PCR assays targeting 35 virulence determinants were used to analyze the virulence repertoire of S. aureus. In vitro activities of the antibiotics were determined by the disk-diffusion method. The pathogenicity of representative isolates was assessed using Caenorhabditis elegans survival assays. Significance in virulence distribution and antibiotic resistance phenotypes was assessed using the Chi-squared tests. Kaplan–Meier survival estimates were used to analyze nematode survival and significance of survival rates evaluated using the log-rank test. Results: Except for sei (staphylococcal enterotoxin I) (P  =  0.027), all other virulence genes were not significantly associated with MRSA. Resistance to clindamycin (P  =  0.03), tetracycline (P  =  0.048), trimethoprim/sulfamethoxazole (P  =  0.038), and oxacillin (P  =  0.004) was significantly associated with MRSA. Survival assay showed MSSA having a lower median lifespan of 3 days than MRSA that had a median lifespan of 6 days. The difference in the killing time of MRSA and MSSA was significant (P virulence genes. The quicker killing potential of MSSA compared to MRSA suggests that carriage of virulence determinants per se does not determine pathogenicity in S. aureus. Pathogenicity is impacted by other factors, possibly antibiotic resistance. PMID:25319852

  16. Evolution of Regions Containing Antibiotic Resistance Genes in FII-2-FIB-1 ColV-Colla Virulence Plasmids.

    Science.gov (United States)

    Moran, Robert A; Hall, Ruth M

    2018-05-01

    Three ColV virulence plasmids carrying antibiotic resistance genes were assembled from draft genome sequences of commensal ST95, ST131, and ST2705 Escherichia coli isolates from healthy Australians. Plasmids pCERC4, pCERC5, and pCERC9 include almost identical backbones containing FII-2 and FIB-1 replicons and the conserved ColV virulence region with an additional ColIa determinant. Only pCERC5 includes a complete, uninterrupted F-like transfer region and was able to conjugate. pCERC5 and pCERC9 contain Tn1721, carrying the tet(A) tetracycline resistance determinant in the same location, with Tn2 (bla TEM ; ampicillin resistance) interrupting the Tn1721 in pCERC5. pCERC4 has a Tn1721/Tn21 hybrid transposon carrying dfrA5 (trimethoprim resistance) and sul1 (sulfamethoxazole resistance) in a class 1 integron. Four FII-2:FIB-1 ColV-ColIa plasmids in the GenBank nucleotide database have a related transposon in the same position, but an IS26 has reshaped the resistance gene region, deleting 2,069 bp of the integron 3'-CS, including sul1, and serving as a target for IS26 translocatable units containing bla TEM , sul2 and strAB (streptomycin resistance), or aphA1 (kanamycin/neomycin resistance). Another ColV-ColIa plasmid containing a related resistance gene region has lost the FII replicon and acquired a unique transfer region via recombination within the resistance region and at oriT. Eighteen further complete ColV plasmid sequences in GenBank contained FIB-1, but the FII replicons were of three types, FII-24, FII-18, and a variant of FII-36.

  17. Mutation of the cyclic di-GMP phosphodiesterase gene in Burkholderia lata SK875 attenuates virulence and enhances biofilm formation.

    Science.gov (United States)

    Jung, Hae-In; Kim, Yun-Jung; Lee, Yun-Jung; Lee, Hee-Soo; Lee, Jung-Kee; Kim, Soo-Ki

    2017-10-01

    Burkholderia sp. is a gram-negative bacterium that commonly exists in the environment, and can cause diseases in plants, animals, and humans. Here, a transposon mutant library of a Burkholderia lata isolate from a pig with swine respiratory disease in Korea was screened for strains showing attenuated virulence in Caenorhabditis elegans. One such mutant was obtained, and the Tn5 insertion junction was mapped to rpfR, a gene encoding a cyclic di-GMP phosphodiesterase that functions as a receptor. Mutation of rpfR caused a reduction in growth on CPG agar and swimming motility as well as a rough colony morphology on Congo red agar. TLC analysis showed reduced AHL secretion, which was in agreement with the results from plate-based and bioluminescence assays. The mutant strain produced significantly more biofilm detected by crystal violet staining than the parent strain. SEM of the mutant strain clearly showed that the overproduced biofilm contained a filamentous structure. These results suggest that the cyclic di-GMP phosphodiesterase RpfR plays an important role in quorum sensing modulation of the bacterial virulence and biofilm formation.

  18. Expression regulation of design process gene in product design

    DEFF Research Database (Denmark)

    Li, Bo; Fang, Lusheng; Li, Bo

    2011-01-01

    To improve the design process efficiency, this paper proposes the principle and methodology that design process gene controls the characteristics of design process under the framework of design process reuse and optimization based on design process gene. First, the concept of design process gene...... is proposed and analyzed, as well as its three categories i.e., the operator gene, the structural gene and the regulator gene. Second, the trigger mechanism that design objectives and constraints trigger the operator gene is constructed. Third, the expression principle of structural gene is analyzed...... with the example of design management gene. Last, the regulation mode that the regulator gene regulates the expression of the structural gene is established and it is illustrated by taking the design process management gene as an example. © (2011) Trans Tech Publications....

  19. Lon protease modulates virulence traits in Erwinia amylovora by direct monitoring of major regulators and indirectly through the Rcs and Gac-Csr regulatory systems.

    Science.gov (United States)

    Lee, Jae Hoon; Ancona, Veronica; Zhao, Youfu

    2018-04-01

    Lon, an ATP-dependent protease in bacteria, influences diverse cellular processes by degrading damaged, misfolded and short-lived regulatory proteins. In this study, we characterized the effects of lon mutation and determined the molecular mechanisms underlying Lon-mediated virulence regulation in Erwinia amylovora, an enterobacterial pathogen of apple. Erwinia amylovora depends on the type III secretion system (T3SS) and the exopolysaccharide (EPS) amylovoran to cause disease. Our results showed that mutation of the lon gene led to the overproduction of amylovoran, increased T3SS gene expression and the non-motile phenotype. Western blot analyses showed that mutation in lon directly affected the accumulation and stability of HrpS/HrpA and RcsA. Mutation in lon also indirectly influenced the expression of flhD, hrpS and csrB through the accumulation of the RcsA/RcsB proteins, which bind to the promoter of these genes. In addition, lon expression is under the control of CsrA, possibly at both the transcriptional and post-transcriptional levels. Although mutation in csrA abolished both T3SS and amylovoran production, deletion of the lon gene in the csrA mutant only rescued amylovoran production, but not T3SS. These results suggest that CsrA might positively control both T3SS and amylovoran production partly by suppressing Lon, whereas CsrA may also play a critical role in T3SS by affecting unknown targets. © 2017 BSPP AND JOHN WILEY & SONS LTD.

  20. Random T-DNA mutagenesis identifies a Cu-Zn-superoxide dismutase gene as a virulence factor of Sclerotinia sclerotiorum

    Science.gov (United States)

    Agrobacterium-mediated transformation (AMT) was used to identify potential virulence factors in Sclerotinia sclerotiorum. Screening AMT transformants identified two mutants showing significantly reduced virulence. The mutants showed similar growth rate, colony morphology, and sclerotial and oxalate ...

  1. Listeria monocytogenes Induces a Virulence-Dependent microRNA Signature That Regulates the Immune Response in Galleria mellonella

    Directory of Open Access Journals (Sweden)

    Gopala K. Mannala

    2017-12-01

    Full Text Available microRNAs (miRNAs coordinate several physiological and pathological processes by regulating the fate of mRNAs. Studies conducted in vitro indicate a role of microRNAs in the control of host-microbe interactions. However, there is limited understanding of miRNA functions in in vivo models of bacterial infections. In this study, we systematically explored changes in miRNA expression levels of Galleria mellonella larvae (greater-wax moth, a model system that recapitulates the vertebrate innate immunity, following infection with L. monocytogenes. Using an insect-specific miRNA microarray with more than 2000 probes, we found differential expression of 90 miRNAs (39 upregulated and 51 downregulated in response to infection with L. monocytogenes. We validated the expression of a subset of miRNAs which have mammalian homologs of known or predicted function. In contrast, non-pathogenic L. innocua failed to induce these miRNAs, indicating a virulence-dependent miRNA deregulation. To predict miRNA targets using established algorithms, we generated a publically available G. mellonella transcriptome database. We identified miRNA targets involved in innate immunity, signal transduction and autophagy, including spätzle, MAP kinase, and optineurin, respectively, which exhibited a virulence-specific differential expression. Finally, in silico estimation of minimum free energy of miRNA-mRNA duplexes of validated microRNAs and target transcripts revealed a regulatory network of the host immune response to L. monocytogenes. In conclusion, this study provides evidence for a role of miRNAs in the regulation of the innate immune response following bacterial infection in a simple, rapid and scalable in vivo model that may predict host-microbe interactions in higher vertebrates.

  2. Real-time PCR expression profiling of genes encoding potential virulence factors in Candida albicans biofilms: identification of model-dependent and -independent gene expression

    Directory of Open Access Journals (Sweden)

    Řičicová Markéta

    2010-04-01

    Full Text Available Abstract Background Candida albicans infections are often associated with biofilm formation. Previous work demonstrated that the expression of HWP1 (hyphal wall protein and of genes belonging to the ALS (agglutinin-like sequence, SAP (secreted aspartyl protease, PLB (phospholipase B and LIP (lipase gene families is associated with biofilm growth on mucosal surfaces. We investigated using real-time PCR whether genes encoding potential virulence factors are also highly expressed in biofilms associated with abiotic surfaces. For this, C. albicans biofilms were grown on silicone in microtiter plates (MTP or in the Centres for Disease Control (CDC reactor, on polyurethane in an in vivo subcutaneous catheter rat (SCR model, and on mucosal surfaces in the reconstituted human epithelium (RHE model. Results HWP1 and genes belonging to the ALS, SAP, PLB and LIP gene families were constitutively expressed in C. albicans biofilms. ALS1-5 were upregulated in all model systems, while ALS9 was mostly downregulated. ALS6 and HWP1 were overexpressed in all models except in the RHE and MTP, respectively. The expression levels of SAP1 were more pronounced in both in vitro models, while those of SAP2, SAP4 and SAP6 were higher in the in vivo model. Furthermore, SAP5 was highly upregulated in the in vivo and RHE models. For SAP9 and SAP10 similar gene expression levels were observed in all model systems. PLB genes were not considerably upregulated in biofilms, while LIP1-3, LIP5-7 and LIP9-10 were highly overexpressed in both in vitro models. Furthermore, an elevated lipase activity was detected in supernatans of biofilms grown in the MTP and RHE model. Conclusions Our findings show that HWP1 and most of the genes belonging to the ALS, SAP and LIP gene families are upregulated in C. albicans biofilms. Comparison of the fold expression between the various model systems revealed similar expression levels for some genes, while for others model-dependent expression

  3. Methicillin-resistant Staphylococcus aureus in cows with mastitis, the presence of the mecA gene and the gene for virulence

    Directory of Open Access Journals (Sweden)

    Vesna Jaki Tkalec

    2015-11-01

    Full Text Available The physiological properties of 47 Staphylococcus aureus strains were investigated. The test strains were grown on bacteriological media and identified by the ID32 STAF system for biochemical identification of bacteria. Sensitivity to antimicrobial agents was performed by the disc diffusion method. The nuc gene and the virulence factors coa, hla, hlb, hld, hlg, hlg-2, tst, eta, etb, lukF-PV and lukS-PV and mecA gene were detected by the polymerase chain reaction. Furthermore, the spa type of the studied isolates was also set. According to the obtained results, all strains had the nuc, coa, hla and hld gene. Ten strains (21.3 % had also the tst gene, while 37 strains (78.7 % had the hlg gene and 35 strains (74.5 % had the hlb and hlg-2 genes. All of the investigated S. aureus isolates were penicillin resistant (100 %, with 29 strains which were also resistant to oxacillin (61.7 %. Methicillin (oxacillin resistance was detected by the mecA gene detection, which is also the first MRSA result from the secretion samples of cows’ mammary glands in Croatia. The researched MRSA strains proved to belong to different spa types, and the most common were spa types t005, t011 and t521, and a new spa type t9498 was detected.

  4. Retrotransposons as regulators of gene expression.

    Science.gov (United States)

    Elbarbary, Reyad A; Lucas, Bronwyn A; Maquat, Lynne E

    2016-02-12

    Transposable elements (TEs) are both a boon and a bane to eukaryotic organisms, depending on where they integrate into the genome and how their sequences function once integrated. We focus on two types of TEs: long interspersed elements (LINEs) and short interspersed elements (SINEs). LINEs and SINEs are retrotransposons; that is, they transpose via an RNA intermediate. We discuss how LINEs and SINEs have expanded in eukaryotic genomes and contribute to genome evolution. An emerging body of evidence indicates that LINEs and SINEs function to regulate gene expression by affecting chromatin structure, gene transcription, pre-mRNA processing, or aspects of mRNA metabolism. We also describe how adenosine-to-inosine editing influences SINE function and how ongoing retrotransposition is countered by the body's defense mechanisms. Copyright © 2016, American Association for the Advancement of Science.

  5. Dietary methanol regulates human gene activity.

    Directory of Open Access Journals (Sweden)

    Anastasia V Shindyapina

    Full Text Available Methanol (MeOH is considered to be a poison in humans because of the alcohol dehydrogenase (ADH-mediated conversion of MeOH to formaldehyde (FA, which is toxic. Our recent genome-wide analysis of the mouse brain demonstrated that an increase in endogenous MeOH after ADH inhibition led to a significant increase in the plasma MeOH concentration and a modification of mRNA synthesis. These findings suggest endogenous MeOH involvement in homeostasis regulation by controlling mRNA levels. Here, we demonstrate directly that study volunteers displayed increasing concentrations of MeOH and FA in their blood plasma when consuming citrus pectin, ethanol and red wine. A microarray analysis of white blood cells (WBC from volunteers after pectin intake showed various responses for 30 significantly differentially regulated mRNAs, most of which were somehow involved in the pathogenesis of Alzheimer's disease (AD. There was also a decreased synthesis of hemoglobin mRNA, HBA and HBB, the presence of which in WBC RNA was not a result of red blood cells contamination because erythrocyte-specific marker genes were not significantly expressed. A qRT-PCR analysis of volunteer WBCs after pectin and red wine intake confirmed the complicated relationship between the plasma MeOH content and the mRNA accumulation of both genes that were previously identified, namely, GAPDH and SNX27, and genes revealed in this study, including MME, SORL1, DDIT4, HBA and HBB. We hypothesized that human plasma MeOH has an impact on the WBC mRNA levels of genes involved in cell signaling.

  6. Limb development: a paradigm of gene regulation.

    Science.gov (United States)

    Petit, Florence; Sears, Karen E; Ahituv, Nadav

    2017-04-01

    The limb is a commonly used model system for developmental biology. Given the need for precise control of complex signalling pathways to achieve proper patterning, the limb is also becoming a model system for gene regulation studies. Recent developments in genomic technologies have enabled the genome-wide identification of regulatory elements that control limb development, yielding insights into the determination of limb morphology and forelimb versus hindlimb identity. The modulation of regulatory interactions - for example, through the modification of regulatory sequences or chromatin architecture - can lead to morphological evolution, acquired regeneration capacity or limb malformations in diverse species, including humans.

  7. Amphibian antimicrobial peptide fallaxin analogue FL9 affects virulence gene expression and DNA replication in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Gottschalk, Sanne; Gottlieb, Caroline Trebbien; Vestergaard, Martin

    2015-01-01

    antimicrobials. In the present study, the analogue FL9, based on the amphibian AMP fallaxin, was studied to elucidate its mode of action and antibacterial activity against the human pathogen Staphylococcus aureus. Our data showed that FL9 may have a dual mode of action against S. aureus. At concentrations around...... and at alkaline pH, while it was compromised by acidic pH and exposure to serum. Furthermore, at subinhibitory concentrations of FL9, S. aureus responded by increasing the expression of two major virulence factor genes, namely the regulatory rnaIII and hla, encoding α-haemolysin. In addition, the S. aureus...... the MIC, FL9 bound DNA, inhibited DNA synthesis and induced the SOS DNA damage response, whereas at concentrations above the MIC the interaction between S. aureus and FL9 led to membrane disruption. The antibacterial activity of the peptide was maintained over a wide range of NaCl and MgCl2 concentrations...

  8. Genetic diversity, phylogroup distribution and virulence gene profile of pks positive Escherichia coli colonizing human intestinal polyps.

    Science.gov (United States)

    Sarshar, Meysam; Scribano, Daniela; Marazzato, Massimiliano; Ambrosi, Cecilia; Aprea, Maria Rita; Aleandri, Marta; Pronio, Annamaria; Longhi, Catia; Nicoletti, Mauro; Zagaglia, Carlo; Palamara, Anna Teresa; Conte, Maria Pia

    2017-11-01

    Some Escherichia coli strains of phylogroup B2 harbor a (pks) pathogenicity island that encodes a polyketide-peptide genotoxin called colibactin. It causes DNA double-strand breaks and megalocytosis in eukaryotic cells and it may contribute to cancer development. Study of bacterial community that colonizes the adenomatous polyp lesion, defined as precancerous lesions, could be helpful to assess if such pathogenic bacteria possess a role in the polyp progression to cancer. In this cross-sectional study, a total of 1500 E. coli isolates were obtained from biopsies of patients presenting adenomatous colon polyps, the normal tissues adjacent to the polyp lesion and patients presenting normal mucosa. pks island frequency, phylogenetic grouping, fingerprint genotyping, and virulence gene features of pks positive (pks + ) E. coli isolates were performed. We found pks + E. coli strongly colonize two patients presenting polypoid lesions and none were identified in patients presenting normal mucosa. Predominant phylogroups among pks + E. coli isolates were B2, followed by D. Clustering based on fragment profiles of composite analysis, typed the pks + isolates into 5 major clusters (I-V) and 17 sub-clusters, demonstrating a high level of genetic diversity among them. The most prevalent virulence genes were fimH and fyuA (100%), followed by vat (92%), hra and papA (69%), ibeA (28%), and hlyA (25%). Our results revealed that pks + E. coli can colonize the precancerous lesions, with a high distribution in both the polyp lesions and in normal tissues adjacent to the lesion. The high differences in fingerprinting patterns obtained indicate that pks + E. coli strains were genetically diverse, possibly allowing them to more easily adapt to environmental variations. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Presence and characterization of Escherichia coli virulence genes isolated from diseased pigs in the central region of Argentina

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    Fernando A. Bessone

    2017-08-01

    Full Text Available Background: The main pathogen of neonatal and post weaning diarrhea and edema disease (ED is Escherichia coli and pathotypes involved are enterotoxigenic, enteropathogenic, and shiga toxigenic (ETEC, EPEC, and STEC, respectively. Those diseases cause economic loss in pig production. Aim: The aim of this work was to evaluate the presence of strains expressing virulence markers genes and the antibiotic susceptibility profiles of E. coli from clinical cases of post weaning diarrhea and ED in farms in the central area of Argentina. Materials and Methods: Intensive pig farms from the central region of Argentina were sampled. Intestinal mucosa swabs from pigs with diarrhea were taken, seeded on MacConkey agar plates, biochemically typified and tested by polymerase chain reaction (PCR. Antibiograms were made by disk-diffusion method. Results: A total of 54 strains from clinical cases studied showed PCR findings: 88.88% (48/54 expressed at least one gene coding for a virulence factor. Colonization factors found were: 39.58% of strains had F18, 33.33% were F4 and 31.25% adhesin involved in diffuse adherence-I; 29.17%, 25%, and 2.1% expressed LT, STb, and STa, respectively. 25% were STx and 16.67% were eae positive. Only 2.1% were STx2. The most active antibiotics against most strains were gentamicin and ceftiofur, but resistance profiles against many antibiotics were found. Conclusion: High circulation of pathogens strains of E. coli among pigs with diarrhea with an extended antibiotic resistance profile.

  10. Carbon nanoparticles as detection labels in antibody microarrays. Detection of genes encoding virulence factors in Shiga toxin-producing Escherichia coli.

    NARCIS (Netherlands)

    Noguera, P.S.; Posthuma-Trumpie, G.A.; Tuil, Van M.; Wal, van der F.J.; Boer, De A.; Moers, A.P.H.A.; Amerongen, Van A.

    2011-01-01

    The present study demonstrates that carbon nanoparticles (CNPs) can be used as labels in microarrays. CNPs were used in nucleic acid microarray immunoassays (NAMIAs) for the detection of different Shiga toxin-producing Escherichia coli (STEC) virulence factors: four genes specific for STEC (vt1,

  11. Some virulence genes of Escherichia coli isolated from cloacal swabs of healthy Alagoas Curassows (Pauxi mitu in Brazil Alguns genes de virulência de Escherichia coli isoladas de mutuns-do-nordeste (Pauxi mitu sadios no Brasil

    Directory of Open Access Journals (Sweden)

    André A.B. Saidenberg

    2013-04-01

    Full Text Available Birds of the Cracidae family (curassows, guans, and chachalacas are endemic of the Neotropics and 50 species are currently classified. Brazil has 22 species, seven of which are considered threatened. The Alagoas Curassow (Pauxi mitu species is considered extinct in the wild; but about 120 birds are alive in captivity. Conservation of this species depends entirely on correct management. Health reports of both wildlife and captive curassows are rare. In this study the presence of Escherichia coli was evaluated in 23 healthy Alagoas Curassows from two private breeding centres. E. coli was isolated from cloacal swabs, and the presence of genes encoding cytotoxic necrotising factor 1 (cnf1, alpha-haemolysin (hly, aerobactin (iuc, serum resistance (iss and the following adhesions: S fimbriae (sfa, pili associated with pyelonephritis (pap and temperature-sensitive haemagglutinin (tsh were investigated. E. coli was isolated from 78.3% (18/23 of the birds, and the percentage of curassows colonized by E. coli was similar between the two facilities. From the 22 E. coli isolates, 15 (68.2% were positive for at least one virulence factor by PCR, and the most frequently found gene was iss (50%. No curassows had clinical signs of disease. Nevertheless, the presence of some E. coli strains may be a concern to the wildlife in captivity. Additional health surveillance studies are essential to guarantee successful conservation programmes for threatened cracids in Brazil.Aves da família Cracidae (mutuns, jacutingas e aracuãs são endêmicas da região Neotropical com 50 espécies atualmente classificadas. O Brasil possui 22 espécies nesta família e sete delas são consideradas ameaçadas de extinção. O mutum-do-nordeste (Pauxi mitu é considerado extinto na natureza, no entanto, aproximadamente 120 indivíduos são mantidos em cativeiro. A conservação desta espécie depende inteiramente de um manejo correto. Informações sobre o status sanitário de mutuns

  12. Subinhibitory concentrations of perilla oil affect the expression of secreted virulence factor genes in Staphylococcus aureus.

    Directory of Open Access Journals (Sweden)

    Jiazhang Qiu

    Full Text Available BACKGROUND: The pathogenicity of staphylococcus aureus is dependent largely upon its ability to secrete a number of virulence factors, therefore, anti-virulence strategy to combat S. aureus-mediated infections is now gaining great interest. It is widely recognized that some plant essential oils could affect the production of staphylococcal exotoxins when used at subinhibitory concentrations. Perilla [Perilla frutescens (L. Britton], a natural medicine found in eastern Asia, is primarily used as both a medicinal and culinary herb. Its essential oil (perilla oil has been previously demonstrated to be active against S. aureus. However, there are no data on the influence of perilla oil on the production of S. aureus exotoxins. METHODOLOGY/PRINCIPAL FINDINGS: A broth microdilution method was used to determine the minimum inhibitory concentrations (MICs of perilla oil against S. aureus strains. Hemolysis, tumour necrosis factor (TNF release, Western blot, and real-time RT-PCR assays were performed to evaluate the effects of subinhibitory concentrations of perilla oil on exotoxins production in S. aureus. The data presented here show that perilla oil dose-dependently decreased the production of α-toxin, enterotoxins A and B (the major staphylococcal enterotoxins, and toxic shock syndrome toxin 1 (TSST-1 in both methicillin-sensitive S. aureus (MSSA and methicillin-resistant S. aureus (MRSA. CONCLUSIONS/SIGNIFICANCE: The production of α-toxin, SEA, SEB, and TSST-1 in S. aureus was decreased by perilla oil. These data suggest that perilla oil may be useful for the treatment of S. aureus infections when used in combination with β-lactam antibiotics, which can increase exotoxins production by S. aureus at subinhibitory concentrations. Furthermore, perilla oil could be rationally applied in food systems as a novel food preservative both to inhibit the growth of S. aureus and to repress the production of exotoxins, particularly staphylococcal enterotoxins.

  13. Comparative transcript profiling of Candida albicans and Candida dubliniensis identifies SFL2, a C. albicans gene required for virulence in a reconstituted epithelial infection model.

    LENUS (Irish Health Repository)

    Spiering, Martin J

    2010-02-01

    Candida albicans and Candida dubliniensis are closely related species displaying differences in virulence and genome content, therefore providing potential opportunities to identify novel C. albicans virulence genes. C. albicans gene arrays were used for comparative analysis of global gene expression in the two species in reconstituted human oral epithelium (RHE). C. albicans (SC5314) showed upregulation of hypha-specific and virulence genes within 30 min postinoculation, coinciding with rapid induction of filamentation and increased RHE damage. C. dubliniensis (CD36) showed no detectable upregulation of hypha-specific genes, grew as yeast, and caused limited RHE damage. Several genes absent or highly divergent in C. dubliniensis were upregulated in C. albicans. One such gene, SFL2 (orf19.3969), encoding a putative heat shock factor, was deleted in C. albicans. DeltaDeltasfl2 cells failed to filament under a range of hypha-inducing conditions and exhibited greatly reduced RHE damage, reversed by reintroduction of SFL2 into the DeltaDeltasfl2 strain. Moreover, SFL2 overexpression in C. albicans triggered hyphal morphogenesis. Although SFL2 deletion had no apparent effect on host survival in the murine model of systemic infection, DeltaDeltasfl2 strain-infected kidney tissues contained only yeast cells. These results suggest a role for SFL2 in morphogenesis and an indirect role in C. albicans pathogenesis in epithelial tissues.

  14. Studying gene regulation in methanogenic archaea.

    Science.gov (United States)

    Rother, Michael; Sattler, Christian; Stock, Tilmann

    2011-01-01

    Methanogenic archaea are a unique group of strictly anaerobic microorganisms characterized by their ability, and dependence, to convert simple C1 and C2 compounds to methane for growth. The major models for studying the biology of methanogens are members of the Methanococcus and Methanosarcina species. Recent development of sophisticated tools for molecular analysis and for genetic manipulation allows investigating not only their metabolism but also their cell cycle, and their interaction with the environment in great detail. One aspect of such analyses is assessment and dissection of methanoarchaeal gene regulation, for which, at present, only a handful of cases have been investigated thoroughly, partly due to the great methodological effort required. However, it becomes more and more evident that many new regulatory paradigms can be unraveled in this unique archaeal group. Here, we report both molecular and physiological/genetic methods to assess gene regulation in Methanococcus maripaludis and Methanosarcina acetivorans, which should, however, be applicable for other methanogens as well. Copyright © 2011 Elsevier Inc. All rights reserved.

  15. Genomes of the most dangerous epidemic bacteria have a virulence repertoire characterized by fewer genes but more toxin-antitoxin modules.

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    Kalliopi Georgiades

    2011-03-01

    Full Text Available We conducted a comparative genomic study based on a neutral approach to identify genome specificities associated with the virulence capacity of pathogenic bacteria. We also determined whether virulence is dictated by rules, or if it is the result of individual evolutionary histories. We systematically compared the genomes of the 12 most dangerous pandemic bacteria for humans ("bad bugs" to their closest non-epidemic related species ("controls".We found several significantly different features in the "bad bugs", one of which was a smaller genome that likely resulted from a degraded recombination and repair system. The 10 Cluster of Orthologous Group (COG functional categories revealed a significantly smaller number of genes in the "bad bugs", which lacked mostly transcription, signal transduction mechanisms, cell motility, energy production and conversion, and metabolic and regulatory functions. A few genes were identified as virulence factors, including secretion system proteins. Five "bad bugs" showed a greater number of poly (A tails compared to the controls, whereas an elevated number of poly (A tails was found to be strongly correlated to a low GC% content. The "bad bugs" had fewer tandem repeat sequences compared to controls. Moreover, the results obtained from a principal component analysis (PCA showed that the "bad bugs" had surprisingly more toxin-antitoxin modules than did the controls.We conclude that pathogenic capacity is not the result of "virulence factors" but is the outcome of a virulent gene repertoire resulting from reduced genome repertoires. Toxin-antitoxin systems could participate in the virulence repertoire, but they may have developed independently of selfish evolution.

  16. Characterization of Vibrio parahaemolyticus isolated from oysters in Korea: Resistance to various antibiotics and prevalence of virulence genes.

    Science.gov (United States)

    Kang, Chang-Ho; Shin, YuJin; Jang, SeokCheol; Yu, HongSik; Kim, SuKyung; An, Sera; Park, Kunbawui; So, Jae-Seong

    2017-05-15

    Vibrio parahaemolyticus, found frequently in oysters, is the most prevalent gastroenteritis-causing pathogen in Korea and in several other Asian countries. This study monitored changes in the environmental parameters and occurrence of V. parahaemolyticus in oyster aquaculture sites. Of the 44 presumed V. parahaemolyticus isolates obtained, when tested against 16 antibiotics, 90.9, 86.4, and 75.0% of the 44 isolates exhibited resistance to vancomycin, ampicillin, and streptomycin, respectively. PCR analysis for the presence of the toxR gene confirmed 31 of the 44 isolates as being positive V. parahaemolyticus strains. The toxR positive isolates were tested for the presence of thermostable direct hemolysin (tdh) and tdh-related hemolysin (trh) virulence genes. Only 9.1% toxR positive isolate exhibit the trh gene and none of the isolates were tested positive for tdh. The occurrence of multi drug resistance strains in the environment could be an indication of excessive usage of antibiotics in agriculture and aquaculture fields. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. The frequency of genes encoding three putative group B streptococcal virulence factors among invasive and colonizing isolates

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    Borchardt Stephanie M

    2006-07-01

    Full Text Available Abstract Background Group B Streptococcus (GBS causes severe infections in very young infants and invasive disease in pregnant women and adults with underlying medical conditions. GBS pathogenicity varies between and within serotypes, with considerable variation in genetic content between strains. Three proteins, Rib encoded by rib, and alpha and beta C proteins encoded by bca and bac, respectively, have been suggested as potential vaccine candidates for GBS. It is not known, however, whether these genes occur more frequently in invasive versus colonizing GBS strains. Methods We screened 162 invasive and 338 colonizing GBS strains from different collections using dot blot hybridization to assess the frequency of bca, bac and rib. All strains were defined by serotyping for capsular type, and frequency differences were tested using the Chi square test. Results Genes encoding the beta C protein (bac and Rib (rib occurred at similar frequencies among invasive and colonizing isolates, bac (20% vs. 23%, and rib (28% vs. 20%, while the alpha (bca C protein was more frequently found in colonizing strains (46% vs, invasive (29%. Invasive strains were associated with specific serotype/gene combinations. Conclusion Novel virulence factors must be identified to better understand GBS disease.

  18. Endogenous Methanol Regulates Mammalian Gene Activity

    Science.gov (United States)

    Komarova, Tatiana V.; Petrunia, Igor V.; Shindyapina, Anastasia V.; Silachev, Denis N.; Sheshukova, Ekaterina V.; Kiryanov, Gleb I.; Dorokhov, Yuri L.

    2014-01-01

    We recently showed that methanol emitted by wounded plants might function as a signaling molecule for plant-to-plant and plant-to-animal communications. In mammals, methanol is considered a poison because the enzyme alcohol dehydrogenase (ADH) converts methanol into toxic formaldehyde. However, the detection of methanol in the blood and exhaled air of healthy volunteers suggests that methanol may be a chemical with specific functions rather than a metabolic waste product. Using a genome-wide analysis of the mouse brain, we demonstrated that an increase in blood methanol concentration led to a change in the accumulation of mRNAs from genes primarily involved in detoxification processes and regulation of the alcohol/aldehyde dehydrogenases gene cluster. To test the role of ADH in the maintenance of low methanol concentration in the plasma, we used the specific ADH inhibitor 4-methylpyrazole (4-MP) and showed that intraperitoneal administration of 4-MP resulted in a significant increase in the plasma methanol, ethanol and formaldehyde concentrations. Removal of the intestine significantly decreased the rate of methanol addition to the plasma and suggested that the gut flora may be involved in the endogenous production of methanol. ADH in the liver was identified as the main enzyme for metabolizing methanol because an increase in the methanol and ethanol contents in the liver homogenate was observed after 4-MP administration into the portal vein. Liver mRNA quantification showed changes in the accumulation of mRNAs from genes involved in cell signalling and detoxification processes. We hypothesized that endogenous methanol acts as a regulator of homeostasis by controlling the mRNA synthesis. PMID:24587296

  19. Endogenous methanol regulates mammalian gene activity.

    Directory of Open Access Journals (Sweden)

    Tatiana V Komarova

    Full Text Available We recently showed that methanol emitted by wounded plants might function as a signaling molecule for plant-to-plant and plant-to-animal communications. In mammals, methanol is considered a poison because the enzyme alcohol dehydrogenase (ADH converts methanol into toxic formaldehyde. However, the detection of methanol in the blood and exhaled air of healthy volunteers suggests that methanol may be a chemical with specific functions rather than a metabolic waste product. Using a genome-wide analysis of the mouse brain, we demonstrated that an increase in blood methanol concentration led to a change in the accumulation of mRNAs from genes primarily involved in detoxification processes and regulation of the alcohol/aldehyde dehydrogenases gene cluster. To test the role of ADH in the maintenance of low methanol concentration in the plasma, we used the specific ADH inhibitor 4-methylpyrazole (4-MP and showed that intraperitoneal administration of 4-MP resulted in a significant increase in the plasma methanol, ethanol and formaldehyde concentrations. Removal of the intestine significantly decreased the rate of methanol addition to the plasma and suggested that the gut flora may be involved in the endogenous production of methanol. ADH in the liver was identified as the main enzyme for metabolizing methanol because an increase in the methanol and ethanol contents in the liver homogenate was observed after 4-MP administration into the portal vein. Liver mRNA quantification showed changes in the accumulation of mRNAs from genes involved in cell signalling and detoxification processes. We hypothesized that endogenous methanol acts as a regulator of homeostasis by controlling the mRNA synthesis.

  20. Effective genes for resistance to stripe rust and virulence of Puccinia ...

    African Journals Online (AJOL)

    The results revealed that stripe rust resistance genes Yr3, Yr5, Yr10, Yr15, Yr26, YrSP and YrCV were resistant, while Yr18 showed moderate susceptibility at all locations. Genes YrA-, Yr2, Yr6, Yr7, Yr8, Yr9, Yr17, Yr27 and gene combinations Opata (Yr27+Yr18) and Super Kauz (Yr9, Yr27, Yr18) were found susceptible.

  1. Redox regulation of photosynthetic gene expression.

    Science.gov (United States)

    Queval, Guillaume; Foyer, Christine H

    2012-12-19

    Redox chemistry and redox regulation are central to the operation of photosynthesis and respiration. However, the roles of different oxidants and antioxidants in the regulation of photosynthetic or respiratory gene expression remain poorly understood. Leaf transcriptome profiles of a range of Arabidopsis thaliana genotypes that are deficient in either hydrogen peroxide processing enzymes or in low molecular weight antioxidant were therefore compared to determine how different antioxidant systems that process hydrogen peroxide influence transcripts encoding proteins targeted to the chloroplasts or mitochondria. Less than 10 per cent overlap was observed in the transcriptome patterns of leaves that are deficient in either photorespiratory (catalase (cat)2) or chloroplastic (thylakoid ascorbate peroxidase (tapx)) hydrogen peroxide processing. Transcripts encoding photosystem II (PSII) repair cycle components were lower in glutathione-deficient leaves, as were the thylakoid NAD(P)H (nicotinamide adenine dinucleotide (phosphate)) dehydrogenases (NDH) mRNAs. Some thylakoid NDH mRNAs were also less abundant in tAPX-deficient and ascorbate-deficient leaves. Transcripts encoding the external and internal respiratory NDHs were increased by low glutathione and low ascorbate. Regulation of transcripts encoding specific components of the photosynthetic and respiratory electron transport chains by hydrogen peroxide, ascorbate and glutathione may serve to balance non-cyclic and cyclic electron flow pathways in relation to oxidant production and reductant availability.

  2. A nitrogen response pathway regulates virulence functions in Fusarium oxysporum via the protein kinase TOR and the bZIP protein MeaB.

    Science.gov (United States)

    López-Berges, Manuel S; Rispail, Nicolas; Prados-Rosales, Rafael C; Di Pietro, Antonio

    2010-07-01

    During infection, fungal pathogens activate virulence mechanisms, such as host adhesion, penetration and invasive growth. In the vascular wilt fungus Fusarium oxysporum, the mitogen-activated protein kinase Fmk1 is required for plant infection and controls processes such as cellophane penetration, vegetative hyphal fusion, or root adhesion. Here, we show that these virulence-related functions are repressed by the preferred nitrogen source ammonium and restored by treatment with l-methionine sulfoximine or rapamycin, two specific inhibitors of Gln synthetase and the protein kinase TOR, respectively. Deletion of the bZIP protein MeaB also resulted in nitrogen source-independent activation of virulence mechanisms. Activation of these functions did not require the global nitrogen regulator AreA, suggesting that MeaB-mediated repression of virulence functions does not act through inhibition of AreA. Tomato plants (Solanum lycopersicum) supplied with ammonium rather than nitrate showed a significant reduction in vascular wilt symptoms when infected with the wild type but not with the DeltameaB strain. Nitrogen source also affected invasive growth in the rice blast fungus Magnaporthe oryzae and the wheat head blight pathogen Fusarium graminearum. We propose that a conserved nitrogen-responsive pathway might operate via TOR and MeaB to control virulence in plant pathogenic fungi.

  3. A Nitrogen Response Pathway Regulates Virulence Functions in Fusarium oxysporum via the Protein Kinase TOR and the bZIP Protein MeaB[C][W

    Science.gov (United States)

    López-Berges, Manuel S.; Rispail, Nicolas; Prados-Rosales, Rafael C.; Di Pietro, Antonio

    2010-01-01

    During infection, fungal pathogens activate virulence mechanisms, such as host adhesion, penetration and invasive growth. In the vascular wilt fungus Fusarium oxysporum, the mitogen-activated protein kinase Fmk1 is required for plant infection and controls processes such as cellophane penetration, vegetative hyphal fusion, or root adhesion. Here, we show that these virulence-related functions are repressed by the preferred nitrogen source ammonium and restored by treatment with l-methionine sulfoximine or rapamycin, two specific inhibitors of Gln synthetase and the protein kinase TOR, respectively. Deletion of the bZIP protein MeaB also resulted in nitrogen source–independent activation of virulence mechanisms. Activation of these functions did not require the global nitrogen regulator AreA, suggesting that MeaB-mediated repression of virulence functions does not act through inhibition of AreA. Tomato plants (Solanum lycopersicum) supplied with ammonium rather than nitrate showed a significant reduction in vascular wilt symptoms when infected with the wild type but not with the ΔmeaB strain. Nitrogen source also affected invasive growth in the rice blast fungus Magnaporthe oryzae and the wheat head blight pathogen Fusarium graminearum. We propose that a conserved nitrogen-responsive pathway might operate via TOR and MeaB to control virulence in plant pathogenic fungi. PMID:20639450

  4. Riboregulators: Fine-Tuning Virulence in Shigella.

    Science.gov (United States)

    Fris, Megan E; Murphy, Erin R

    2016-01-01

    Within the past several years, RNA-mediated regulation (ribo-regulation) has become increasingly recognized for its importance in controlling critical bacterial processes. Regulatory RNA molecules, or riboregulators, are perpetually responsive to changes within the micro-environment of a bacterium. Notably, several characterized riboregulators control virulence in pathogenic bacteria, as is the case for each riboregulator characterized to date in Shigella. The timing of virulence gene expression and the ability of the pathogen to adapt to rapidly changing environmental conditions is critical to the establishment and progression of infection by Shigella species; ribo-regulators mediate each of these important processes. This mini review will present the current state of knowledge regarding RNA-mediated regulation in Shigella by detailing the characterization and function of each identified riboregulator in these pathogens.

  5. Directed evolution induces tributyrin hydrolysis in a virulence factor of Xylella fastidiosa using a duplicated gene as a template.

    Science.gov (United States)

    Gouran, Hossein; Chakraborty, Sandeep; Rao, Basuthkar J; Asgeirsson, Bjarni; Dandekar, Abhaya

    2014-01-01

    Duplication of genes is one of the preferred ways for natural selection to add advantageous functionality to the genome without having to reinvent the wheel with respect to catalytic efficiency and protein stability. The duplicated secretory virulence factors of Xylella fastidiosa (LesA, LesB and LesC), implicated in Pierce's disease of grape and citrus variegated chlorosis of citrus species, epitomizes the positive selection pressures exerted on advantageous genes in such pathogens. A deeper insight into the evolution of these lipases/esterases is essential to develop resistance mechanisms in transgenic plants. Directed evolution, an attempt to accelerate the evolutionary steps in the laboratory, is inherently simple when targeted for loss of function. A bigger challenge is to specify mutations that endow a new function, such as a lost functionality in a duplicated gene. Previously, we have proposed a method for enumerating candidates for mutations intended to transfer the functionality of one protein into another related protein based on the spatial and electrostatic properties of the active site residues (DECAAF). In the current work, we present in vivo validation of DECAAF by inducing tributyrin hydrolysis in LesB based on the active site similarity to LesA. The structures of these proteins have been modeled using RaptorX based on the closely related LipA protein from Xanthomonas oryzae. These mutations replicate the spatial and electrostatic conformation of LesA in the modeled structure of the mutant LesB as well, providing in silico validation before proceeding to the laborious in vivo work. Such focused mutations allows one to dissect the relevance of the duplicated genes in finer detail as compared to gene knockouts, since they do not interfere with other moonlighting functions, protein expression levels or protein-protein interaction.

  6. The stationary phase sigma factor, RpoS, regulates the production of a carbapenem antibiotic, a bioactive prodigiosin and virulence in the enterobacterial pathogen Serratia sp. ATCC 39006.

    Science.gov (United States)

    Wilf, Nabil M; Salmond, George P C

    2012-03-01

    Serratia sp. ATCC 39006 (S39006) is a Gram-negative bacterium that is virulent in plant (potato) and invertebrate animal (Caenorhabditis elegans) models. It produces two secondary metabolite antibiotics, a prodigiosin and a carbapenem, and the exoenzymes pectate lyase and cellulase. We showed previously that deletion of the RNA chaperone Hfq abolished antibiotic production and attenuated virulence in both animal and plant hosts. Hfq and dependent small RNAs (sRNAs) are known to regulate the post-transcriptional expression of rpoS, which encodes σ(S), the stationary phase sigma factor subunit of RNA polymerase. An S39006 hfq deletion mutant showed decreased transcript levels of rpoS. Therefore, in this study we investigated whether the phenotypes regulated by Hfq were mediated through its control of rpoS. Whereas loss of Hfq abolished prodigiosin and carbapenem production and attenuated virulence in both C. elegans and potato, characterization of an S39006 rpoS mutant showed unexpectedly elevated prodigiosin and carbapenem production. Furthermore, the rpoS mutant exhibited attenuated animal pathogenesis, but not plant pathogenesis. Additionally, a homologue of the Hfq-dependent sRNA, RprA, was identified and shown to regulate prodigiosin production in a manner consistent with its role in positively regulating translation of rpoS mRNA. Combined, these results demonstrate that Hfq regulation of secondary metabolism and plant pathogenesis is independent of RpoS and establishes RpoS and RprA as regulators of antibiotic production.

  7. The diversities of staphylococcal species, virulence and antibiotic resistance genes in the subclinical mastitis milk from a single Chinese cow herd.

    Science.gov (United States)

    Xu, Jia; Tan, Xiao; Zhang, Xinyu; Xia, Xiaoli; Sun, Huaichang

    2015-11-01

    Staphylococci are the leading pathogens of bovine mastitis which is difficult to control. However, the published data on the prevalence of staphylococcal species, virulence and antibiotic resistance genes in bovine mastitis from China are limited. In this study, 104 out of 209 subclinical mastitis milk samples from a single Chinese dairy herd were cultured-positive for staphylococci (49.8%), which were further identified as coagulase-positive staphylococci (CPS) or coagulase-negative staphylococci (CNS). According to the partial tuf and/or 16S rRNA gene sequence, the 28 CPS isolates were confirmed to be Staphylococcus aureus (26.9%), and 76 CNS isolates were assigned to 13 different species (73.1%) with Staphylococcus arlettae, Staphylococcus sciuri, Staphylococcus xylosus and Staphylococcus chromogenes as the dominant species. In the 28 S. aureus isolates, the most prevalent general virulence genes were coa, Ig and eno (100%), followed by hla (96.4%), hlb (92.9%), fib (92.9%), clfA (89.3%), clfB (85.7%) and nuc (85.7%). Both exotoxin and biofilm-associated genes were significantly less prevalent than the previously reported. Although 19 different virulence gene patterns were found, only one was dominant (32.1%). The prevalence of blaZ (82.1%) or mecA gene (35.7%) was much higher than the previously reported. In the 76 CNS isolates, the virulence genes were significantly less prevalent than that in the S. aureus isolates. Among the 4 main CNS species, S. chromogenes (n = 12) was the only species with high percentage (75%) of blaZ gene, while S. sciuri (n = 12) was the only species with the high percentage (66.7%) of mecA gene. The most of antibiotic resistance genes were present as multi-resistance genes, and the antibiotic resistances were attributed by different resistance genes between resistant S. aureus and CNS isolates. These data suggest that the prevalence of staphylococcal species, virulence and antibiotic resistance in the mastitis milk from the Chinese

  8. Necrotic enteritis locus 1 diguanylate cyclase and phosphodiesterase (cyclic-di-GMP) gene mutation attenuates virulence in an avian necrotic enteritis isolate of Clostridium perfringens.

    Science.gov (United States)

    Parreira, Valeria R; Ojha, Shivani; Lepp, Dion; Mehdizadeh Gohari, Iman; Zhou, Hongzhuan; Susta, Leonardo; Gong, Jianhua; Prescott, John F

    2017-09-01

    Necrotic enteritis (NE) caused by netB-positive strains of Clostridium perfringens is an important disease of intensively-reared broiler chickens. It is widely controlled by antibiotic use, but this practice that has come under increasing scrutiny and alternative approaches are required. As part of the search for alternative approaches over the last decade, advances have been made in understanding its pathogenesis but much remains to be understood and applied to the control of NE. The objective of this work was to assess the effect on virulence of mutation of the cyclic-di-GMP signaling genes present on the large pathogenicity locus (NELoc-1) in the tcp-encoding conjugative virulence plasmid, pNetB. For this purpose, the diguanylate cyclase (dgc) and phosphodiesterase (pde) genes were individually insertionally inactivated and the two mutants were subsequently complemented with their respective genes. Southern blotting showed that a single gene insertion was present. Mutation of either gene resulted in almost total attenuation of the mutants to cause NE in experimentally-infected broiler chickens, which was fully restored in each case by complementation of the respective mutated gene. Production of NetB-associated cytotoxicity for Leghorn male hepatoma (LMH) cells was unaffected in mutants. We conclude that the cyclic-di-GMP signaling system is important in controlling virulence in a NE C. perfringens strain and might be a target for control of the disease. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Helicobacter pylori virulence genes in the five largest islands of Indonesia.

    Science.gov (United States)

    Miftahussurur, Muhammad; Syam, Ari Fahrial; Makmun, Dadang; Nusi, Iswan Abbas; Zein, Lukman Hakim; Zulkhairi; Akil, Fardah; Uswan, Willi Brodus; Simanjuntak, David; Uchida, Tomohisa; Adi, Pangestu; Utari, Amanda Pitarini; Rezkitha, Yudith Annisa Ayu; Subsomwong, Phawinee; Nasronudin; Yamaoka, Yoshio

    2015-01-01

    It remains unclear whether the low incidence of gastric cancer in Indonesia is due to low infection rates only or is also related to low Helicobacter pylori pathogenicity. We collected H. pylori strains from the five largest islands in Indonesia and evaluated genetic virulence factors. The genotypes of H. pylori virulence factors were determined by polymerase chain reaction (PCR)-based sequencing. Histological severity of the gastric mucosa was classified into 4 grades, according to the updated Sydney system. A total of 44 strains were analyzed. Forty-three (97.7 %) were cagA-positive: 26 (60.5 %) were East-Asian-type-cagA, 9 (20.9 %) were Western-type-cagA, and 8 (18.6 %) were novel ABB-type, most of which were obtained from Papuan. EPIYT sequences were more prevalent than EPIYA sequences (P = 0.01) in the EPIYA-B motif of all types of cagA. The majority of cagA-positive strains (48.8 %, 21/43) had a 6-bp deletion in the first pre-EPIYA region. Subjects infected with East-Asian-type-cagA strains with a 6-bp deletion had significantly lower inflammation and atrophy scores in the corpus than those infected with Western-type-cagA strains (both P = 0.02). In total, 70.4 % of strains possessed the vacA s1m1 genotype and 29.5 % were m2. All strains from peptic ulcer patients were of the iceA1 genotype, which occurred at a significantly higher proportion in peptic ulcer patients than that in gastritis patients (55.3 %, P = 0.04). The double positive genotype of jhp0562/β-(1,3)galT was predominant (28/44, 63.6 %), and subjects infected with this type had significantly higher inflammation scores in the corpus than those with the jhp0562 negative/β-(1,3)galT positive genotype (mean [median]; 1.43 [1] vs. 0.83 [1], P = 0.04). There were significant differences in cagA and pre-EPIYA cagA type, oipA status, and jhp0562/β-(1,3)galT type among different ethnic groups (P dupA negative or short type dupA, and the jhp0562/β-(1,3)galT double positive genotype.

  10. Distribution of putative virulence genes and antimicrobial drug resistance in Vibrio harveyi

    Digital Repository Service at National Institute of Oceanography (India)

    Parvathi, A.; Mendez, D.; Anto, C.

    zonula occludens toxin (Zot) and a hemolysin-coregulated protein gene (hcp) by polymerase chain reaction (PCR). Of the four putative reversible toxin genes, vhh-1 was detected in 31% of the isolates, vhh-2 in 46%, vhh-3 in 23% and vhh-4 was detected in 27...

  11. Polyamines are essential for virulence in Salmonella enterica serovar Gallinarum despite evolutionary decay of polyamine biosynthesis genes.

    Science.gov (United States)

    Schroll, Casper; Christensen, Jens P; Christensen, Henrik; Pors, Susanne E; Thorndahl, Lotte; Jensen, Peter R; Olsen, John E; Jelsbak, Lotte

    2014-05-14

    Serovars of Salmonella enterica exhibit different host-specificities where some have broad host-ranges and others, like S. Gallinarum and S. Typhi, are host-specific for poultry and humans, respectively. With the recent availability of whole genome sequences it has been reported that host-specificity coincides with accumulation of pseudogenes, indicating adaptation of host-restricted serovars to their narrow niches. Polyamines are small cationic amines and in Salmonella they can be synthesized through two alternative pathways directly from l-ornithine to putrescine and from l-arginine via agmatine to putrescine. The first pathway is not active in S. Gallinarum and S. Typhi, and this prompted us to investigate the importance of polyamines for virulence in S. Gallinarum. Bioinformatic analysis of all sequenced genomes of Salmonella revealed that pseudogene formation of the speC gene was exclusive for S. Typhi and S. Gallinarum and happened through independent events. The remaining polyamine biosynthesis pathway was found to be essential for oral infection with S. Gallinarum since single and double mutants in speB and speE, encoding the pathways from agmatine to putrescine and from putrescine to spermidine, were attenuated. In contrast, speB was dispensable after intraperitoneal challenge, suggesting that putrescine was less important for the systemic phase of the disease. In support of this hypothesis, a ΔspeE;ΔpotCD mutant, unable to synthesize and import spermidine, but with retained ability to import and synthesize putrescine, was attenuated after intraperitoneal infection. We therefore conclude that polyamines are essential for virulence of S. Gallinarum. Furthermore, our results point to distinct roles for putrescine and spermidine during systemic infection. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Comparative genomic analysis reveals occurrence of genetic recombination in virulent Cryptosporidium hominis subtypes and telomeric gene duplications in Cryptosporidium parvum.

    Science.gov (United States)

    Guo, Yaqiong; Tang, Kevin; Rowe, Lori A; Li, Na; Roellig, Dawn M; Knipe, Kristine; Frace, Michael; Yang, Chunfu; Feng, Yaoyu; Xiao, Lihua

    2015-04-18

    Cryptosporidium hominis is a dominant species for human cryptosporidiosis. Within the species, IbA10G2 is the most virulent subtype responsible for all C. hominis-associated outbreaks in Europe and Australia, and is a dominant outbreak subtype in the United States. In recent yearsIaA28R4 is becoming a major new subtype in the United States. In this study, we sequenced the genomes of two field specimens from each of the two subtypes and conducted a comparative genomic analysis of the obtained sequences with those from the only fully sequenced Cryptosporidium parvum genome. Altogether, 8.59-9.05 Mb of Cryptosporidium sequences in 45-767 assembled contigs were obtained from the four specimens, representing 94.36-99.47% coverage of the expected genome. These genomes had complete synteny in gene organization and 96.86-97.0% and 99.72-99.83% nucleotide sequence similarities to the published genomes of C. parvum and C. hominis, respectively. Several major insertions and deletions were seen between C. hominis and C. parvum genomes, involving mostly members of multicopy gene families near telomeres. The four C. hominis genomes were highly similar to each other and divergent from the reference IaA25R3 genome in some highly polymorphic regions. Major sequence differences among the four specimens sequenced in this study were in the 5' and 3' ends of chromosome 6 and the gp60 region, largely the result of genetic recombination. The sequence similarity among specimens of the two dominant outbreak subtypes and genetic recombination in chromosome 6, especially around the putative virulence determinant gp60 region, suggest that genetic recombination plays a potential role in the emergence of hyper-transmissible C. hominis subtypes. The high sequence conservation between C. parvum and C. hominis genomes and significant differences in copy numbers of MEDLE family secreted proteins and insulinase-like proteases indicate that telomeric gene duplications could potentially contribute to

  13. Recent urbanization in China is correlated with a Westernized microbiome encoding increased virulence and antibiotic resistance genes.

    Science.gov (United States)

    Winglee, Kathryn; Howard, Annie Green; Sha, Wei; Gharaibeh, Raad Z; Liu, Jiawu; Jin, Donghui; Fodor, Anthony A; Gordon-Larsen, Penny

    2017-09-15

    Urbanization is associated with an increased risk for a number of diseases, including obesity, diabetes, and cancer, which all also show associations with the microbiome. While microbial community composition has been shown to vary across continents and in traditional versus Westernized societies, few studies have examined urban-rural differences in neighboring communities within a single country undergoing rapid urbanization. In this study, we compared the gut microbiome, plasma metabolome, dietary habits, and health biomarkers of rural and urban people from a single Chinese province. We identified significant differences in the microbiota and microbiota-related plasma metabolites in rural versus recently urban subjects from the Hunan province of China. Microbes with higher relative abundance in Chinese urban samples have been associated with disease in other studies and were substantially more prevalent in the Human Microbiome Project cohort of American subjects. Furthermore, using whole metagenome sequencing, we found that urbanization was associated with a loss of microbial diversity and changes in the relative abundances of Viruses, Archaea, and Bacteria. Gene diversity, however, increased with urbanization, along with the proportion of reads associated with antibiotic resistance and virulence, which were strongly correlated with the presence of Escherichia and Shigella. Our data suggest that urbanization has produced convergent evolution of the gut microbial composition in American and urban Chinese populations, resulting in similar compositional patterns of abundant microbes through similar lifestyles on different continents, including a loss of potentially beneficial bacteria and an increase in potentially harmful genes via increased relative abundance of Escherichia and Shigella.

  14. Strain-specific impact of PsaR of Streptococcus pneumoniae on global gene expression and virulence

    OpenAIRE

    Hendriksen, Wouter T.; Bootsma, Hester J.; van Diepen, Angela; Estevao, Silvia; Kuipers, Oscar P.; de Groot, Ronald; Hermans, Peter W. M.

    2009-01-01

    Previous studies have indicated that PsaR of Streptococcus pneumoniae is a manganese-dependent regulator, negatively affecting the expression of at least seven genes. Here, we extended these observations by transcriptome and proteome analysis of psaR mutants in strains D39 and TIGR4. The microarray analysis identified three shared PsaR targets: the psa operon, pcpA and prtA. In addition, we found 31 genes to be regulated by PsaR in D39 only, most strikingly a cellobiose-specific phosphotrains...

  15. FRUITING GENES OF SCHIZOPHYLLUM-COMMUNE ARE TRANSCRIPTIONALLY REGULATED

    NARCIS (Netherlands)

    SCHUREN, FHJ; VANDERLENDE, TR; WESSELS, JGH

    Fruiting genes in Schizophyllum commune are controlled by the mating-type genes and other regulatory genes. To examine whether differential accumulation of mRNAs for these fruiting genes is caused by transcriptional regulation, run-on transcription assaYs were performed with nuclei isolated from

  16. Application of Chemical Genomics to Plant-Bacteria Communication: A High-Throughput System to Identify Novel Molecules Modulating the Induction of Bacterial Virulence Genes by Plant Signals.

    Science.gov (United States)

    Vandelle, Elodie; Puttilli, Maria Rita; Chini, Andrea; Devescovi, Giulia; Venturi, Vittorio; Polverari, Annalisa

    2017-01-01

    The life cycle of bacterial phytopathogens consists of a benign epiphytic phase, during which the bacteria grow in the soil or on the plant surface, and a virulent endophytic phase involving the penetration of host defenses and the colonization of plant tissues. Innovative strategies are urgently required to integrate copper treatments that control the epiphytic phase with complementary tools that control the virulent endophytic phase, thus reducing the quantity of chemicals applied to economically and ecologically acceptable levels. Such strategies include targeted treatments that weaken bacterial pathogens, particularly those inhibiting early infection steps rather than tackling established infections. This chapter describes a reporter gene-based chemical genomic high-throughput screen for the induction of bacterial virulence by plant molecules. Specifically, we describe a chemical genomic screening method to identify agonist and antagonist molecules for the induction of targeted bacterial virulence genes by plant extracts, focusing on the experimental controls required to avoid false positives and thus ensuring the results are reliable and reproducible.

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

    Science.gov (United States)

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

    2018-02-03

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

  18. A multiplex real-time PCR assay targeting virulence and resistance genes in Salmonella enterica serotype Typhimurium

    Directory of Open Access Journals (Sweden)

    Brisabois Anne

    2011-06-01

    Full Text Available Abstract Background Typhimurium is the main serotype of Salmonella enterica subsp. enterica implicated in food-borne diseases worldwide. This study aimed to detect the prevalence of ten markers combined in a macro-array based on multiplex real-time PCR. We targeted characteristic determinants located on pathogenicity islands (SPI-2 to -5, virulence plasmid pSLT and Salmonella genomic island 1 (SGI1 as well as a specific 16S-23S rRNA intergenic spacer sequence of definitive type 104 (DT104. To investigate antimicrobial resistance, the study also targeted the presence of genes involved in sulfonamide (sul1 and beta-lactam (blaTEM resistance. Finally, the intI1 determinant encoding integrase from class 1 integron was also investigated. Results A total of 538 unrelated S. Typhimurium strains isolated between 1999 and 2009 from various sources, including food animals, food products, human and environmental samples were studied. Based on the combined presence or absence of these markers, we distinguished 34 different genotypes, including three major genotypes encountered in 75% of the studied strains, Although SPI determinants were almost always detected, SGI1, intI1, sul1 and blaTEM determinants were found 47%, 52%, 54% and 12% of the time respectively, varying according to isolation source. Low-marker patterns were most often detected in poultry sources whereas full-marker patterns were observed in pig, cattle and human sources. Conclusion The GeneDisc® assay developed in this study madeit easier to explore variability within serotype Typhimurium by analyzing ten relevant gene determinants in a large collection of strains. This real-time multiplex method constitutes a valuable tool for strains characterization on epidemiological purposes.

  19. Prevalence study of Vibrio species and frequency of the virulence genes of Vibrio parahaemolyticus isolated from fresh and salted shrimps in Genaveh seaport

    Directory of Open Access Journals (Sweden)

    S Hosseini

    2014-08-01

    Full Text Available Vibrio species are important seafood-borne pathogens that are responsible for 50-70% of gasteroenteritis. The present study was carried out in order to determine the prevalence of Vibrio species and the distribution of tdh, tlh and trh virulence genes in Vibrio parahaemolyticus isolated from fresh and salted shrimp samples. Totally, 60 fresh and salted shrimp samples were collected from the Genaveh seaport. Microbial culture was used to isolate Vibrio species. In addition, the presences of Vibrio parahaemolyticus, Vibrio cholera, Vibrio vulnificus and Vibrio harveyi and the virulence genes of V. parahaemolyticus were studied using the PCR method. Results showed that 20% of fresh and 23.33% of salted shrimp samples were positive for Vibrio species. In studied samples, V. vulnificus had the highest prevalence rate (8.33%, while V. cholera had the lowest prevalence rate (1.66%. From a total of 4 detected V. parahaemolyticus, all of them had tlh gene (100%. The distribution of tdh and trh genes in isolated V. parahaemolyticus strains were 50% and 25%, respectively. High prevalence of Vibrio species and especially virulent V. parahaemolyticus in samples confirmed the lack of hygienic condition in the production and distribution centers of shrimp.

  20. Transcriptional Alterations of Virulence-Associated Genes in Extended Spectrum Beta-Lactamase (ESBL-Producing Uropathogenic Escherichia coli during Morphologic Transitions Induced by Ineffective Antibiotics

    Directory of Open Access Journals (Sweden)

    Isak Demirel

    2017-06-01

    Full Text Available It is known that an ineffective antibiotic treatment can induce morphological shifts in uropathogenic Escherichia coli (UPEC but the virulence properties during these shifts remain to be studied. The present study examines changes in global gene expression patterns and in virulence factor-associated genes in an extended spectrum beta-lactamase (ESBL-producing UPEC (ESBL019 during the morphologic transitions induced by an ineffective antibiotic and in the presence of human primary bladder epithelial cells. Microarray results showed that the different morphological states of ESBL019 had significant transcriptional alterations of a large number of genes (Transition; 7%, Filamentation; 32%, and Reverted 19% of the entities on the array. All three morphological states of ESBL019 were associated with a decreased energy metabolism, altered iron acquisition systems and altered adhesion expression. In addition, genes associated with LPS synthesis and bacterial motility was also altered in all the morphological states. Furthermore, the transition state induced a significantly higher release of TNF-α from bladder epithelial cells compared to all other morphologies, while the reverted state was unable to induce TNF-α release. Our findings show that the morphological shifts induced by ineffective antibiotics are associated with significant transcriptional virulence alterations in ESBL-producing UPEC, which may affect survival and persistence in the urinary tract.

  1. Strain-specific impact of PsaR of Streptococcus pneumoniae on global gene expression and virulence.

    NARCIS (Netherlands)

    Hendriksen, W.T.; Bootsma, H.J.; Diepen, A. van; Estevao, S.; Kuipers, O.P.; Groot, R. de; Hermans, P.W.M.

    2009-01-01

    Previous studies have indicated that PsaR of Streptococcus pneumoniae is a manganese-dependent regulator, negatively affecting the expression of at least seven genes. Here, we extended these observations by transcriptome and proteome analysis of psaR mutants in strains D39 and TIGR4. The microarray

  2. CRISPR/Cas systems: new players in gene regulation and bacterial physiology

    Directory of Open Access Journals (Sweden)

    David eWeiss

    2014-04-01

    Full Text Available CRISPR-Cas systems are bacterial defenses against foreign nucleic acids derived from bacteriophages, plasmids or other sources. These systems are targeted in an RNA-dependent, sequence-specific manner, and are also adaptive, providing protection against previously encountered foreign elements. In addition to their canonical function in defense against foreign nucleic acid, their roles in various aspects of bacterial physiology are now being uncovered. We recently revealed a role for a Cas9-based Type II CRISPR-Cas system in the control of endogenous gene expression, a novel form of prokaryotic gene regulation. Cas9 functions in association with two small RNAs to target and alter the stability of an endogenous transcript encoding a bacterial lipoprotein (BLP. Since BLPs are recognized by the host innate immune protein Toll-like Receptor 2 (TLR2, CRISPR-Cas-mediated repression of BLP expression facilitates evasion of TLR2 by the intracellular bacterial pathogen Francisella novicida, and is essential for its virulence. Here we describe the Cas9 regulatory system in detail, as well as data on its role in controlling virulence traits of Neisseria meningitidis and Campylobacter jejuni. We also discuss potential roles of CRISPR-Cas systems in the response to envelope stress and other aspects of bacterial physiology. Since ~45% of bacteria and ~83% of Archaea encode these machineries, the newly appreciated regulatory functions of CRISPR-Cas systems are likely to play broad roles in controlling the pathogenesis and physiology of diverse prokaryotes.

  3. Cell Density Control of Staphylococcal Virulence Mediated by an Octapeptide Pheromone

    Science.gov (United States)

    Ji, Guangyong; Beavis, Ronald C.; Novick, Richard P.

    1995-12-01

    Some bacterial pathogens elaborate and secrete virulence factors in response to environmental signals, others in response to a specific host product, and still others in response to no discernible cue. In this study, we have demonstrated that the synthesis of Staphylococcus aureus virulence factors is controlled by a density-sensing system that utilizes an octapeptide produced by the organism itself. The octapeptide activates expression of the agr locus, a global regulator of the virulence response. This response involves the reciprocal regulation of genes encoding surface proteins and those encoding secreted virulence factors. As cells enter the postexponential phase, surface protein genes are repressed by agr and secretory protein genes are subsequently activated. The intracellular agr effector is a regulatory RNA, RNAIII, whose transcription is activated by an agr-encoded signal transduction system for which the octapeptide is the ligand.

  4. Occurrence of diarrhoeagenic Escherichia coli virulence genes in water and bed sediments of a river used by communities in Gauteng, South Africa.

    Science.gov (United States)

    Abia, Akebe Luther King; Ubomba-Jaswa, Eunice; Momba, Maggy Ndombo Benteke

    2016-08-01

    In most developing countries, especially in Southern Africa, little is known about the presence of diarrhoeagenic Escherichia coli (DEC) pathotypes in riverbed sediments. The present study sought to investigate the presence of DEC virulence genes in riverbed sediments of the Apies River, a river used by many communities in Gauteng, South Africa. Water and sediment samples were collected from the river between July and August 2013 (dry season) and also between January and February 2014 (wet season) following standard procedures. Isolation of E. coli was done using the Colilert®-18 Quanti-Tray® 2000 system. DNA was extracted from E. coli isolates using the InstaGene™ matrix from Bio-Rad and used as template DNA for real-time PCR. Water pH, temperature, dissolved oxygen, electrical conductivity and turbidity were measured in situ. Over 59 % of 180 samples analysed were positive for at least one of the seven DEC virulence genes investigated. The eaeA gene was the most isolated gene (29.44 %) while the ipaH gene the least isolated (8.33 %). The ipaH gene (p = 0.012) and the ST gene (stIa, p = 0.0001, and stIb, p = 0.019) were positively correlated with temperature. The detection of diarrhoeagenic E. coli virulence genes in the sediments of the Apies River shows that the sediments of this river might not only be a reservoir of faecal indicator bacteria like E. coli but also pathogenic strains of this bacterium. These organisms could represent a public health risk for poor communities relying on this water source for various purposes such as drinking and recreational use. There is therefore an urgent need to monitor these DEC pathotypes especially in areas without adequate water supplies.

  5. Genomic insights into a new Citrobacter koseri strain revealed gene exchanges with the virulence-associated Yersinia pestis pPCP1 plasmid

    Directory of Open Access Journals (Sweden)

    Fabrice eArmougom

    2016-03-01

    Full Text Available The history of infectious diseases raised the plague as one of the most devastating for human beings. Far too often considered an ancient disease, the frequent resurgence of the plague has led to consider it as a reemerging disease in Madagascar, Algeria, Libya and Congo. The genetic factors associated with the pathogenicity of Yersinia pestis, the causative agent of the plague, involve the acquisition of the pPCP1 plasmid that promotes host invasion through the expression of the virulence factor Pla. The surveillance of plague foci after the 2003 outbreak in Algeria resulted in a positive detection of the specific pla gene of Y. pestis in rodents. However, the phenotypic characterization of the isolate identified a Citrobacter koseri. The comparative genomics of our sequenced C. koseri URMITE genome revealed a mosaic gene structure resulting from the lifestyle of our isolate and provided evidence for gene exchanges with different enteric bacteria. The most striking was the acquisition of a continuous 2 kb genomic fragment containing the virulence factor Pla of the Y. pestis pPCP1 plasmid; however, the subcutaneous injection of the CKU strain in mice did not produce any pathogenic effect. Our findings demonstrate that fast molecular detection of plague using solely the pla gene is unsuitable and should rather require Y. pestis gene marker combinations. We also suggest that the evolutionary force that might govern the expression of pathogenicity can occur through the acquisition of virulence genes but could also require the loss or the inactivation of resident genes such as antivirulence genes.

  6. Rapid identification of genes controlling virulence and immunity in malaria parasites

    KAUST Repository

    Abkallo, Hussein M.

    2017-07-13

    Identifying the genetic determinants of phenotypes that impact disease severity is of fundamental importance for the design of new interventions against malaria. Here we present a rapid genome-wide approach capable of identifying multiple genetic drivers of medically relevant phenotypes within malaria parasites via a single experiment at single gene or allele resolution. In a proof of principle study, we found that a previously undescribed single nucleotide polymorphism in the binding domain of the erythrocyte binding like protein (EBL) conferred a dramatic change in red blood cell invasion in mutant rodent malaria parasites Plasmodium yoelii. In the same experiment, we implicated merozoite surface protein 1 (MSP1) and other polymorphic proteins, as the major targets of strain-specific immunity. Using allelic replacement, we provide functional validation of the substitution in the EBL gene controlling the growth rate in the blood stages of the parasites.

  7. Expression of the sigma35 and cry2AB genes involved in Bacillus thuringiensis virulence Expressão dos genes sigma35 e cry2AB envolvidos na virulência de Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Ana Maria Guidelli-Thuler

    2009-06-01

    Full Text Available There are several genes involved in Bacillus thuringiensis sporulation. The regulation and expression of these genes results in an upregulation in Cry protein production, and this is responsible for the death of insect larvae infected by Bacillus thuringiensis. Gene expression was monitored in Bacillus thuringiensis during three developmental phases. DNA macroarrays were constructed for selected genes whose sequences are available in the GenBank database. These genes were hybridized to cDNA sequences from B. thuringiensis var. kurstaki HD-1. cDNA probes were synthesized by reverse transcription from B. thuringiensis RNA templates extracted during the exponential (log growth, stationary and sporulation phases, and labeled with 33PadCTP. Two genes were differentially expressed levels during the different developmental phases. One of these genes is related to sigma factor (sigma35, and the other is a cry gene (cry2Ab. There were differences between the differential levels of expression of various genes and among the expression detected for different combinations of the sigma factor and cry2Ab genes. The maximum difference in expression was observed for the gene encoding sigma35 factor in the log phase, which was also expressed at a high level during the sporulation phase. The cry2Ab gene was only expressed at a high level in the log phase, but at very low levels in the other phases when compared to the sigma35.Muitos genes estão envolvidos nos mecanismos de esporulação da bactéria Bacillus thuringiensis. A regulação e expressão desses genes resultam em uma produção massiva da proteína Cry, responsável pela morte das larvas de muitos insetos. Neste trabalho monitorou-se a expressão de genes de Bacillus thuringiensis, ao long