Relationship between Vancomycin MIC and Virulence Gene Expression in Clonal Complexes of Methicillin-Susceptible Staphylococcus aureus Strains Isolated from Left-Sided Endocarditis

ABSTRACT Higher vancomycin MICs have been associated with more complicated courses and higher mortality rates in patients with Staphylococcus aureus bacteremia and infective endocarditis (IE). The aim of this study was to investigate whether the strains belonging to the cohort of 93 patients from a previously published study in which patients with strains with vancomycin MICs of ≥1.5 μg/ml presented higher mortality rates and systemic emboli than patients with strains with vancomycin MICs of <1.5 μg/ml had specific patterns of virulence factors, clonal complex (CC) types, or the ability to form biofilms. Vancomycin MICs were determined by Etest, and the isolates underwent spa typing to infer the CC, biofilm studies, a thrombin-induced platelet microbicidal assay, and multiplex PCR for the presence of virulence genes. We found no differences in genes encoding adhesins, toxins, or other putative virulence genes according to the vancomycin MIC group. CC30, CC34, and CC45 represented nearly half of the isolates, and there was no association with the vancomycin MIC. agr subgroups I and III predominated, with no association with the vancomycin MIC. Isolates with higher vancomycin MICs exhibited a poorer ability to form biofilms with and without the presence of vancomycin (2.03 versus 2.48 [P < 0.001], respectively, for isolates with higher vancomycin MICs and 2.60 versus 2.87 [P = 0.022], respectively, for isolates with lower vancomycin MICs). In the multivariable analysis, efb and V8 were risk factors for major emboli (adjusted odds ratio [aOR] = 7.5 and 95% confidence interval [CI] = 1.2 to 46.6 for efb, and aOR = 3.9 and 95% CI = 1.1 to 14.1 for V8), whereas no genotypic predictors of in-hospital mortality were found. No clear associations between genes encoding virulence factors, agr type, clonal complexes, mortality, and major embolic events according to vancomycin MIC group were found.

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Inhibition of Rho Activity Increases Expression of SaeRS-Dependent Virulence Factor Genes inStaphylococcus aureus, Showing a Link between Transcription Termination, Antibiotic Action, and Virulence

mBio ◽

10.1128/mbio.01332-18 ◽

2018 ◽

Vol 9 (5) ◽

Cited By ~ 6

Author(s):

Anna Nagel ◽

Stephan Michalik ◽

Michel Debarbouille ◽

Tobias Hertlein ◽

Manuela Gesell Salazar ◽

...

Keyword(s):

Gene Expression ◽

Staphylococcus Aureus ◽

Virulence Factors ◽

Transcription Termination ◽

Virulence Gene ◽

Antisense Transcription ◽

Termination Factor ◽

Content Type ◽

Virulence Gene Expression ◽

Transcription Termination Factor

ABSTRACTStaphylococcus aureuscauses various diseases ranging from skin and soft tissue infections to life-threatening infections. Adaptation to the different host niches is controlled by a complex network of transcriptional regulators. Global profiling of condition-dependent transcription revealed adaptation ofS. aureusHG001 at the levels of transcription initiation and termination. In particular, deletion of the gene encoding the Rho transcription termination factor triggered a remarkable overall increase in antisense transcription and gene expression changes attributable to indirect regulatory effects. The goal of the present study was a detailed comparative analysis ofS. aureusHG001 and its isogenicrhodeletion mutant. Proteome analysis revealed significant differences in cellular and extracellular protein profiles, most notably increased amounts of the proteins belonging to the SaeR regulon in the Rho-deficient strain. The SaeRS two-component system acts as a major regulator of virulence gene expression in staphylococci. Higher levels of SaeRS-dependent virulence factors such as adhesins, toxins, and immune evasion proteins in therhomutant resulted in higher virulence in a murine bacteremia model, which was alleviated in arhocomplemented strain. Inhibition of Rho activity by bicyclomycin, a specific inhibitor of Rho activity, also induced the expression of SaeRS-dependent genes, at both the mRNA and protein levels, to the same extent as observed in therhomutant. Taken together, these findings indicate that activation of the Sae system in the absence of Rho is directly linked to Rho’s transcription termination activity and establish a new link between antibiotic action and virulence gene expression inS. aureus.IMPORTANCEThe major human pathogenStaphylococcus aureusis a widespread commensal bacterium but also the most common cause of nosocomial infections. It adapts to the different host niches through a complex gene regulatory network. We show here that the Rho transcription termination factor, which represses pervasive antisense transcription in various bacteria, includingS. aureus, plays a role in controlling SaeRS-dependent virulence gene expression. A Rho-deficient strain produces larger amounts of secreted virulence factorsin vitroand shows increased virulence in mice. We also show that treatment ofS. aureuswith the antibiotic bicyclomycin, which inhibits Rho activity and is effective against Gram-negative bacteria, induces the same changes in the proteome as observed in the Rho-deficient strain. Our results reveal for the first time a link between transcription termination and virulence regulation inS. aureus, which implies a novel mechanism by which an antibiotic can modulate the expression of virulence factors.

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Nutritional Regulation of the Sae Two-Component System by CodY inStaphylococcus aureus

Journal of Bacteriology ◽

10.1128/jb.00012-18 ◽

2018 ◽

Vol 200 (8) ◽

Cited By ~ 14

Author(s):

Kevin D. Mlynek ◽

William E. Sause ◽

Derek E. Moormeier ◽

Marat R. Sadykov ◽

Kurt R. Hill ◽

...

Keyword(s):

Gene Expression ◽

Virulence Factors ◽

Virulence Gene ◽

Nutrient Depletion ◽

Global Regulator ◽

Two Component System ◽

Component System ◽

Content Type ◽

Virulence Gene Expression ◽

Two Component

ABSTRACTStaphylococcus aureussubverts innate defenses during infection in part by killing host immune cells to exacerbate disease. This human pathogen intercepts host cues and activates a transcriptional response via theS. aureusexoprotein expression (SaeR/SaeS [SaeR/S]) two-component system to secrete virulence factors critical for pathogenesis. We recently showed that the transcriptional repressor CodY adjusts nuclease (nuc) gene expression via SaeR/S, but the mechanism remained unknown. Here, we identified two CodY binding motifs upstream of thesaeP1 promoter, which suggested direct regulation by this global regulator. We show that CodY shares a binding site with the positive activator SaeR and that alleviating direct CodY repression at this site is sufficient to abrogate stochastic expression, suggesting that CodY repressessaeexpression by blocking SaeR binding. Epistasis experiments support a model that CodY also controlssaeindirectly through Agr and Rot-mediated repression of thesaeP1 promoter. We also demonstrate that CodY repression ofsaerestrains production of secreted cytotoxins that kill human neutrophils. We conclude that CodY plays a previously unrecognized role in controlling virulence gene expression via SaeR/S and suggest a mechanism by which CodY acts as a master regulator of pathogenesis by tying nutrient availability to virulence gene expression.IMPORTANCEBacterial mechanisms that mediate the switch from a commensal to pathogenic lifestyle are among the biggest unanswered questions in infectious disease research. Since the expression of most virulence genes is often correlated with nutrient depletion, this implies that virulence is a response to the lack of nourishment in host tissues and that pathogens likeS. aureusproduce virulence factors in order to gain access to nutrients in the host. Here, we show that specific nutrient depletion signals appear to be funneled to the SaeR/S system through the global regulator CodY. Our findings reveal a strategy by whichS. aureusdelays the production of immune evasion and immune-cell-killing proteins until key nutrients are depleted.

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Role of BrnQ1 and BrnQ2 in Branched-Chain Amino Acid Transport and Virulence in Staphylococcus aureus

Infection and Immunity ◽

10.1128/iai.02542-14 ◽

2014 ◽

Vol 83 (3) ◽

pp. 1019-1029 ◽

Cited By ~ 21

Author(s):

Julienne C. Kaiser ◽

Sameha Omer ◽

Jessica R. Sheldon ◽

Ian Welch ◽

David E. Heinrichs

Keyword(s):

Staphylococcus Aureus ◽

Virulence Gene ◽

Defined Medium ◽

Infection Model ◽

Content Type ◽

Virulence Gene Expression ◽

Branched Chain Amino Acid ◽

Branched Chain

The branched-chain amino acids (BCAAs; Ile, Leu, and Val) not only are important nutrients for the growth ofStaphylococcus aureusbut also are corepressors for CodY, which regulates virulence gene expression, implicating BCAAs as an important link between the metabolic state of the cell and virulence. BCAAs are either synthesized intracellularly or acquired from the environment.S. aureusencodes three putative BCAA transporters, designated BrnQ1, BrnQ2, and BrnQ3; their functions have not yet been formally tested. In this study, we mutated all threebrnQparalogs so as to characterize their substrate specificities and their roles in growthin vitroandin vivo. We demonstrated that in the community-associated, methicillin-resistantS. aureus(CA-MRSA) strain USA300, BrnQ1 is involved in uptake of all three BCAAs, BrnQ2 transports Ile, and BrnQ3 does not have a significant role in BCAA transport under the conditions tested. Of the three, only BrnQ1 is essential for USA300 to grow in a chemically defined medium that is limited for Leu or Val. Interestingly, we observed that abrnQ2mutant grew better than USA300 in media limited for Leu and Val, owing to the fact that this mutation leads to overexpression ofbrnQ1. In a murine infection model, thebrnQ1mutant was attenuated, but in contrast,brnQ2mutants had significantly increased virulence compared to that of USA300, a phenotype we suggest is at least partially linked to enhancedin vivoscavenging of Leu and Val through BrnQ1. These data uncover a hitherto-undiscovered connection between nutrient acquisition and virulence in CA-MRSA.

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Indole Signaling at the Host-Microbiota-Pathogen Interface

mBio ◽

10.1128/mbio.01031-19 ◽

2019 ◽

Vol 10 (3) ◽

Cited By ~ 29

Author(s):

Aman Kumar ◽

Vanessa Sperandio

Keyword(s):

Gene Expression ◽

Virulence Genes ◽

Virulence Gene ◽

Enteric Pathogens ◽

Bacterial Membrane ◽

Gi Tract ◽

Intestinal Tissue ◽

Content Type ◽

Virulence Gene Expression ◽

Membrane Bound

ABSTRACTMicrobial establishment within the gastrointestinal (GI) tract requires surveillance of the gut biogeography. The gut microbiota coordinates behaviors by sensing host- or microbiota-derived signals. Here we show for the first time that microbiota-derived indole is highly prevalent in the lumen compared to the intestinal tissue. This difference in indole concentration plays a key role in modulating virulence gene expression of the enteric pathogens enterohemorrhagicEscherichia coli(EHEC) andCitrobacter rodentium. Indole decreases expression of genes within the locus of enterocyte effacement (LEE) pathogenicity island, which is essential for these pathogens to form attaching and effacing (AE) lesions on enterocytes. We synthetically altered the concentration of indole in the GI tracts of mice by employing mice treated with antibiotics to deplete the microbiota and reconstituted with indole-producing commensalBacteroides thetaiotaomicron(B. theta) or aB. thetaΔtnaAmutant (does not produce indole) or by engineering an indole-producingC. rodentiumstrain. This allowed us to assess the role of self-produced versus microbiota-produced indole, and the results show that decreased indole concentrations promote bacterial pathogenesis, while increased levels of indole decrease bacterial virulence gene expression. Moreover, we identified the bacterial membrane-bound histidine sensor kinase (HK) CpxA as an indole sensor. Enteric pathogens sense a gradient of indole concentrations in the gut to probe different niches and successfully establish an infection.IMPORTANCEPathogens sense and respond to several small molecules within the GI tract to modulate expression of their virulence repertoire. Indole is a signaling molecule produced by the gut microbiota. Here we show that indole concentrations are higher in the lumen, where the microbiota is present, than in the intestinal tissue. The enteric pathogens EHEC andC. rodentiumsense indole to downregulate expression of their virulence genes, as a read-out of the luminal compartment. We also identified the bacterial membrane-bound HK CpxA as an indole sensor. This regulation ensures that EHEC andC. rodentiumexpress their virulence genes only at the epithelial lining, which is the niche they colonize.

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MroQ Is a Novel Abi-Domain Protein That Influences Virulence Gene Expression in Staphylococcus aureus via Modulation of Agr Activity

Infection and Immunity ◽

10.1128/iai.00002-19 ◽

2019 ◽

Vol 87 (5) ◽

Cited By ~ 3

Author(s):

Stephanie Marroquin ◽

Brittney Gimza ◽

Brooke Tomlinson ◽

Michelle Stein ◽

Andrew Frey ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Virulence Gene ◽

Transcriptional Analysis ◽

Data Sets ◽

Virulence Determinants ◽

Sequencing Data ◽

Content Type ◽

Virulence Gene Expression ◽

Function Mechanism ◽

Family Protein

ABSTRACT Numerous factors have, to date, been identified as playing a role in the regulation of Agr activity in Staphylococcus aureus, including transcription factors, antisense RNAs, and host elements. Herein we investigated the product of SAUSA300_1984 (termed MroQ), a transmembrane Abi-domain/M79 protease-family protein, as a novel effector of this system. Using a USA300 mroQ mutant, we observed a drastic reduction in proteolysis, hemolysis, and pigmentation that was fully complementable. This appears to result from diminished agr activity, as transcriptional analysis revealed significant decreases in expression of both RNAII and RNAIII in the mroQ mutant. Such effects appear to be direct, rather than indirect, as known agr effectors demonstrated limited alterations in their activity upon mroQ disruption. A comparison of RNA sequencing data sets for both mroQ and agr mutants revealed a profound overlap in their regulomes, with the majority of factors affected being known virulence determinants. Importantly, the preponderance of alterations in expression were more striking in the agr mutant, indicating that MroQ is necessary, but not sufficient, for Agr function. Mechanism profiling revealed that putative residues for metalloprotease activity within MroQ are required for its Agr-controlling effect; however, this was not wielded at the level of AgrD processing. Virulence assessment demonstrated that both mroQ and agr mutants exhibited increased formation of renal abscesses but decreased skin abscess formation alongside diminished dermonecrosis. Collectively, we present the characterization of a novel agr effector in S. aureus which would appear to be a direct regulator, potentially functioning via interaction with the AgrC histidine kinase.

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18β-Glycyrrhetinic Acid Inhibits Methicillin-Resistant Staphylococcus aureus Survival and Attenuates Virulence Gene Expression

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01023-12 ◽

2012 ◽

Vol 57 (1) ◽

pp. 241-247 ◽

Cited By ~ 32

Author(s):

Danyelle R. Long ◽

Julia Mead ◽

Jay M. Hendricks ◽

Michele E. Hardy ◽

Jovanka M. Voyich

Keyword(s):

Gene Expression ◽

Staphylococcus Aureus ◽

Virulence Gene ◽

Glycyrrhetinic Acid ◽

Licorice Root ◽

Methicillin Resistant ◽

Content Type ◽

Virulence Gene Expression

ABSTRACTMethicillin-resistantStaphylococcus aureus(MRSA) has become a major source of infection in hospitals and in the community. Increasing antibiotic resistance inS. aureusstrains has created a need for alternative therapies to treat disease. A component of the licorice rootGlycyrrhizaspp., 18β-glycyrrhetinic acid (GRA), has been shown to have antiviral, antitumor, and antibacterial activity. This investigation explores thein vitroandin vivoeffects of GRA on MRSA pulsed-field gel electrophoresis (PFGE) type USA300. GRA exhibited bactericidal activity at concentrations exceeding 0.223 μM. Upon exposure ofS. aureusto sublytic concentrations of GRA, we observed a reduction in expression of key virulence genes, includingsaeRandhla. In murine models of skin and soft tissue infection, topical GRA treatment significantly reduced skin lesion size and decreased the expression ofsaeRandhlagenes. Our investigation demonstrates that at high concentrations GRA is bactericidal to MRSA and at sublethal doses it reduces virulence gene expression inS. aureusbothin vitroandin vivo.

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Detection of virulence factors in high-level gentamicin-resistant Enterococcus faecalis and Enterococcus faecium isolates from a Tunisian hospital

Canadian Journal of Microbiology ◽

10.1139/w06-136 ◽

2007 ◽

Vol 53 (3) ◽

pp. 372-379 ◽

Cited By ~ 17

Author(s):

N. Klibi ◽

K. Ben Slama ◽

Y. Sáenz ◽

A. Masmoudi ◽

S. Zanetti ◽

...

Keyword(s):

Enterococcus Faecalis ◽

Virulence Factors ◽

Enterococcus Faecium ◽

Biofilm Formation ◽

Virulence Genes ◽

Virulence Gene ◽

Gelatinase Activity ◽

Genes Encoding ◽

High Level

Phenotypic and genotypic determination of virulence factors were carried out in 46 high-level gentamicin-resistant (HLGR) clinical Enterococcus faecalis (n = 34) and Enterococcus faecium (n = 12) isolates recovered from different patients in La Rabta Hospital in Tunis, Tunisia, between 2000 and 2003 (all these isolates harboured the aac(6′)–aph(2″) gene). The genes encoding virulence factors (agg, gelE, ace, cylLLS, esp, cpd, and fsrB) were analysed by PCR and sequencing. The production of gelatinase and hemolysin, the adherence to caco-2 and hep-2 cells, and the capacity for biofilm formation were investigated in all 46 HLGR enterococci. The percentages of E. faecalis isolates harbouring virulence genes were as follows: gelE, cpd, and ace (100%); fsrB (62%); agg (56%); cylLLS (41.2%); and esp (26.5%). The only virulence gene detected among the 12 HLGR E. faecium isolates was esp (58%). Gelatinase activity was detected in 22 of the 34 E. faecalis isolates (65%, most of them with the gelE+–fsrB+ genotype); the remaining 12 isolates were gelatinase-negative (with the gelE+–fsrB– genotype and the deletion of a 23.9 kb fragment of the fsr locus). Overall, 64% of the cylLLS-containing E. faecalis isolates showed β-hemolysis. A high proportion of our HLGR E. faecalis isolates, in contrast to E. faecium, showed moderate or strong biofilm formation or adherence to caco-2 and hep-2 cells.

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sarZ, a sarA Family Gene, Is Transcriptionally Activated by MgrA and Is Involved in the Regulation of Genes Encoding Exoproteins in Staphylococcus aureus

Journal of Bacteriology ◽

10.1128/jb.01555-08 ◽

2008 ◽

Vol 191 (5) ◽

pp. 1656-1665 ◽

Cited By ~ 36

Author(s):

Anand Ballal ◽

Binata Ray ◽

Adhar C. Manna

Keyword(s):

Staphylococcus Aureus ◽

Target Genes ◽

Virulence Gene ◽

Promoter Regions ◽

Regulatory Relationship ◽

Virulence Gene Expression ◽

Regulatory Systems ◽

Expression Of Genes ◽

Genes Encoding ◽

Gel Shift

ABSTRACT The expression of genes involved in the pathogenesis of Staphylococcus aureus is controlled by global regulatory loci, including two-component regulatory systems and transcriptional regulators (e.g., sar family genes). Most members of the SarA family have been partially characterized and shown to regulate a large numbers of target genes. Here, we describe the characterization of sarZ, a sarA paralog from S. aureus, and its regulatory relationship with other members of its family. Expression of sarZ was growth phase dependent with maximal expression in the early exponential phase of growth. Transcription of sarZ was reduced in an mgrA mutant and returned to a normal level in a complemented mgrA mutant strain, which suggests that mgrA acts as an activator of sarZ transcription. Purified MgrA protein bound to the sarZ promoter region, as determined by gel shift assays. Among the sarA family of genes analyzed, inactivation of sarZ increased sarS transcription, while it decreased agr transcription. The expression of potential target genes involved in virulence was evaluated in single and double mutants of sarZ with mgrA, sarX, and agr. Northern and zymogram analyses indicated that the sarZ gene product played a role in regulating several virulence genes, particularly those encoding exoproteins. Gel shift assays demonstrated nonspecific binding of purified SarZ protein to the promoter regions of the sarZ-regulated target genes. These results demonstrate the important role played by SarZ in controlling regulatory and virulence gene expression in S. aureus.

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Ethanolamine Controls Expression of Genes Encoding Components Involved in Interkingdom Signaling and Virulence in Enterohemorrhagic Escherichia coli O157:H7

mBio ◽

10.1128/mbio.00050-12 ◽

2012 ◽

Vol 3 (3) ◽

Cited By ~ 92

Author(s):

Melissa M. Kendall ◽

Charley C. Gruber ◽

Christopher T. Parker ◽

Vanessa Sperandio

Keyword(s):

Gene Expression ◽

Escherichia Coli ◽

Cell Signaling ◽

Nitrogen Source ◽

Virulence Genes ◽

Virulence Gene ◽

Human Pathogen ◽

Signaling Molecule ◽

Content Type ◽

Virulence Gene Expression

ABSTRACTBacterial pathogens must be able to both recognize suitable niches within the host for colonization and successfully compete with commensal flora for nutrients in order to establish infection. Ethanolamine (EA) is a major component of mammalian and bacterial membranes and is used by pathogens as a carbon and/or nitrogen source in the gastrointestinal tract. The deadly human pathogen enterohemorrhagicEscherichia coliO157:H7 (EHEC) uses EA in the intestine as a nitrogen source as a competitive advantage for colonization over the microbial flora. Here we show that EA is not only important for nitrogen metabolism but that it is also used as a signaling molecule in cell-to-cell signaling to activate virulence gene expression in EHEC. EA in concentrations that cannot promote growth as a nitrogen source can activate expression of EHEC’s repertoire of virulence genes. The EutR transcription factor, known to be the receptor of EA, is only partially responsible for this regulation, suggesting that yet another EA receptor exists. This important link of EA with metabolism, cell-to-cell signaling, and pathogenesis, highlights the fact that a fundamental means of communication within microbial communities relies on energy production and processing of metabolites. Here we show for the first time that bacterial pathogens not only exploit EA as a metabolite but also coopt EA as a signaling molecule to recognize the gastrointestinal environment and promote virulence expression.IMPORTANCEIn order to successfully cause disease, a pathogen must be able to sense a host environment and modulate expression of its virulence genes as well as compete with the indigenous microbiota for nutrients. Ethanolamine (EA) is present in the large intestine due to the turnover of intestinal cells. Here, we show that the human pathogenEscherichia coliO157:H7, which causes bloody diarrhea and hemolytic-uremic syndrome, regulates virulence gene expression through EA metabolism and by responding to EA as a signal. These findings provide the first information directly linking EA with bacterial pathogenesis.

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Diverse Virulence Gene Content of Shiga Toxin-Producing Escherichia coli from Finishing Swine

Applied and Environmental Microbiology ◽

10.1128/aem.01761-14 ◽

2014 ◽

Vol 80 (20) ◽

pp. 6395-6402 ◽

Cited By ~ 25

Author(s):

Marion Tseng ◽

Pina M. Fratamico ◽

Lori Bagi ◽

Sabine Delannoy ◽

Patrick Fach ◽

...

Keyword(s):

Escherichia Coli ◽

Shiga Toxin ◽

Virulence Genes ◽

Genetic Relatedness ◽

Virulence Gene ◽

Health Concern ◽

Limited Information ◽

Content Type ◽

Genes Encoding ◽

Critical Public Health

ABSTRACTShiga toxin-producingEscherichia coli(STEC) infections are a critical public health concern because they can cause severe clinical outcomes, such as hemolytic uremic syndrome, in humans. Determining the presence or absence of virulence genes is essential in assessing the potential pathogenicity of STEC strains. Currently, there is limited information about the virulence genes carried by swine STEC strains; therefore, this study was conducted to examine the presence and absence of 69 virulence genes in STEC strains recovered previously from finishing swine in a longitudinal study. A subset of STEC strains was analyzed by pulsed-field gel electrophoresis (PFGE) to examine their genetic relatedness. Swine STEC strains (n= 150) were analyzed by the use of a high-throughput real-time PCR array system, which included 69 virulence gene targets. Three major pathotypes consisted of 16 different combinations of virulence gene profiles, and serotypes were determined in the swine STEC strains. The majority of the swine STEC strains (n= 120) belonged to serotype O59:H21 and carried the same virulence gene profile, which consisted of 9 virulence genes:stx2e,iha,ecs1763,lpfAO113,estIa(STa),ehaA,paa,terE, andureD. Theeae,nleF, andnleH1-2genes were detected in one swine STEC strain (O49:H21). Other genes encoding adhesins, includingiha, were identified (n= 149). The PFGE results demonstrated that swine STEC strains from pigs raised in the same finishing barn were closely related. Our results revealed diverse virulence gene contents among the members of the swine STEC population and enhance understanding of the dynamics of transmission of STEC strains among pigs housed in the same barn.

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