Expression of a Clostridium perfringens Type IV Pilin by Neisseria gonorrhoeae Mediates Adherence to Muscle Cells

ABSTRACTClostridium perfringensis an anaerobic, Gram-positive bacterium that causes a range of diseases in humans, including lethal gas gangrene. We have recently shown that strains ofC. perfringensmove across the surface of agar plates by a unique type IV pilus (TFP)-mediated social motility that had not been previously described. Based on sequence homology to pilins in Gram-negative bacteria,C. perfringensappears to have two pilin subunits, PilA1 and PilA2. Structural prediction analysis indicated PilA1 is similar to the pseudopilin found inKlebsiella oxytoca, while PilA2 is more similar to true pilins found in the Gram-negative pathogensPseudomonas aeruginosaandNeisseria gonorrhoeae. Strains ofN. gonorrhoeaethat were genetically deficient in the native pilin, PilE, but supplemented with inducible expression of PilA1 and PilA2 ofC. perfringenswere constructed. Genetic competence, wild-type twitching motility, and attachment to human urogenital epithelial cells were not restored by expression of either pilin. However, attachment to mouse and rat myoblast (muscle) cell lines was observed with theN. gonorrhoeaestrain expressing PilA2. Significantly, wild-typeC. perfringenscells adhered to mouse myoblasts under anaerobic conditions, and adherence was 10-fold lower in apilTmutant that lacked functional TFP. These findings implicateC. perfringensTFP in the ability ofC. perfringensto adhere to and move along muscle fibersin vivo, which may provide a therapeutic approach to limiting this rapidly spreading and highly lethal infection.

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Lipid A’s Structure Mediates Neisseria gonorrhoeae Fitness during Experimental Infection of Mice and Men

mBio ◽

10.1128/mbio.00892-13 ◽

2013 ◽

Vol 4 (6) ◽

Cited By ~ 44

Author(s):

Marcia M. Hobbs ◽

James E. Anderson ◽

Jacqueline T. Balthazar ◽

Justin L. Kandler ◽

Russell W. Carlson ◽

...

Keyword(s):

Neisseria Gonorrhoeae ◽

Genital Tract ◽

Lipid A ◽

Female Genital Tract ◽

Treatment Strategies ◽

Wild Type ◽

Genital Tract Infection ◽

Content Type

ABSTRACT Phosphoethanolamine (PEA) on Neisseria gonorrhoeae lipid A influences gonococcal inflammatory signaling and susceptibility to innate host defenses in in vitro models. Here, we evaluated the role of PEA-decorated gonococcal lipid A in competitive infections in female mice and in male volunteers. We inoculated mice and men with mixtures of wild-type N. gonorrhoeae and an isogenic mutant that lacks the PEA transferase, LptA. LptA production conferred a marked survival advantage for wild-type gonococci in the murine female genital tract and in the human male urethra. Our studies translate results from test tube to animal model and into the human host and demonstrate the utility of the mouse model for studies of virulence factors of the human-specific pathogen N. gonorrhoeae that interact with non-host-restricted elements of innate immunity. These results validate the use of gonococcal LptA as a potential target for development of novel immunoprophylactic strategies or antimicrobial treatments. IMPORTANCE Gonorrhea is one of the most common bacterial sexually transmitted infections, and increasing antibiotic resistance threatens the use of currently available antimicrobial therapies. In this work, encompassing in vitro studies and in vivo studies of animal and human models of experimental genital tract infection, we document the importance of lipid A’s structure, mediated by a single bacterial enzyme, LptA, in enhancing the fitness of Neisseria gonorrhoeae. The results of these studies suggest that novel agents targeting LptA may offer urgently needed prevention or treatment strategies for gonorrhea.

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Cyclic Di-GMP Binding by an Assembly ATPase (PilB2) and Control of Type IV Pilin Polymerization in the Gram-Positive Pathogen Clostridium perfringens

Journal of Bacteriology ◽

10.1128/jb.00034-17 ◽

2017 ◽

Vol 199 (10) ◽

Cited By ~ 17

Author(s):

William A. Hendrick ◽

Mona W. Orr ◽

Samantha R. Murray ◽

Vincent T. Lee ◽

Stephen B. Melville

Keyword(s):

Clostridium Perfringens ◽

Biochemical Characterization ◽

Core Protein ◽

Type Iv Pili ◽

Host Cells ◽

Dependent Manner ◽

Gram Positive ◽

Content Type ◽

Type Iv

ABSTRACT The Gram-positive pathogen Clostridium perfringens possesses type IV pili (TFP), which are extracellular fibers that are polymerized from a pool of pilin monomers in the cytoplasmic membrane. Two proteins that are essential for pilus functions are an assembly ATPase (PilB) and an inner membrane core protein (PilC). Two homologues each of PilB and PilC are present in C. perfringens, called PilB1/PilB2 and PilC1/PilC2, respectively, along with four pilin proteins, PilA1 to PilA4. The gene encoding PilA2, which is considered the major pilin based on previous studies, is immediately downstream of the pilB2 and pilC2 genes. Purified PilB2 had ATPase activity, bound zinc, formed hexamers even in the absence of ATP, and bound the second messenger molecule cyclic di-GMP (c-di-GMP). Circular dichroism spectroscopy of purified PilC2 indicated that it retained its predicted degree of alpha-helical secondary structure. Even though no direct interactions between PilB2 and PilC2 could be detected in vivo or in vitro even in the presence of c-di-GMP, high levels of expression of a diguanylate cyclase from C. perfringens (CPE1788) stimulated polymerization of PilA2 in a PilB2- and PilC2-dependent manner. These results suggest that PilB2 activity is controlled by c-di-GMP levels in vivo but that PilB2-PilC2 interactions are either transitory or of low affinity, in contrast to results reported previously from in vivo studies of the PilB1/PilC1 pair in which PilC1 was needed for polar localization of PilB1. This is the first biochemical characterization of a c-di-GMP-dependent assembly ATPase from a Gram-positive bacterium. IMPORTANCE Type IV pili (TFP) are protein fibers involved in important bacterial functions, including motility, adherence to surfaces and host cells, and natural transformation. All clostridia whose genomes have been sequenced show evidence of the presence of TFP. The genetically tractable species Clostridium perfringens was used to study proteins involved in polymerizing the pilin, PilA2, into a pilus. The assembly ATPase PilB2 and its cognate membrane protein partner, PilC2, were purified. PilB2 bound the intracellular signal molecule c-di-GMP. Increased levels of intracellular c-di-GMP led to increased polymerization of PilA2, indicating that Gram-positive bacteria use this molecule to regulate pilus synthesis. These findings provide valuable information for understanding how pathogenic clostridia regulate TFP to cause human diseases.

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In VivoActivities of Ceftolozane, a New Cephalosporin, with and without Tazobactam against Pseudomonas aeruginosa and Enterobacteriaceae, Including Strains with Extended-Spectrum β-Lactamases, in the Thighs of Neutropenic Mice

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01590-12 ◽

2012 ◽

Vol 57 (4) ◽

pp. 1577-1582 ◽

Cited By ~ 88

Author(s):

W. A. Craig ◽

D. R. Andes

Keyword(s):

Pseudomonas Aeruginosa ◽

Infection Model ◽

Wild Type ◽

Gram Negative ◽

Content Type ◽

Peak Dose ◽

Extended Spectrum ◽

The Impact ◽

Primary Index

ABSTRACTCeftolozane is a new cephalosporin with potent activity againstPseudomonas aeruginosaandEnterobacteriaceae. A neutropenic murine thigh infection model was used to determine which pharmacokinetic/pharmacodynamic index and magnitude drives the efficacy of ceftolozane with Gram-negative bacilli, to compare the rates ofin vivokilling ofP. aeruginosaby ceftolozane and ceftazidime, and to determine the impact of different ratios of ceftolozane plus tazobactam onEnterobacteriaceaecontaining extended-spectrum β-lactamases (ESBLs). Neutropenic mice had 106.2-7.1CFU/thigh when treated with ceftolozane for 24 h with (i) various doses (3.12 to 1,600 mg/kg) and dosage intervals (3, 6, 12, and 24 h) against twoEnterobacteriaceaestrains, (ii) 0.39 to 800 mg/kg every 6 h for fourEnterobacteriaceaeand fourP. aeruginosastrains, and (iii) 400 or 800 mg/kg with 2:1. 4:1, and 8:1 ratios of tazobactam against fiveEnterobacteriaceaestrains with ESBLs. The pharmacokinetics of ceftolozane at 25, 100, and 400 mg/kg were linear with peak/dose values of 1.0 to 1.4 and half-lives of 12 to 14 min. T>MIC was the primary index driving efficacy. For stasis (1 log kill), T>MIC was 26.3% ± 2.1% (31.6% ± 1.6%) for wild-typeEnterobacteriaceae, 31.1% ± 4.9% (34.8% ± 4.4%) forEnterobacteriaceaewith ESBLs, and 24.0% ± 3.3% (31.5% ± 3.9%) forP. aeruginosa. At 200 mg/kg every 3 h, the rate ofin vivokilling ofP. aeruginosawas faster with ceftolozane than with ceftazidime (−0.34 to −0.41 log10CFU/thigh/h versus −0.21 to −0.24 log10CFU/thigh/h). The 2:1 ratio of ceftolozane with tazobactam was the most potent combination studied. The T>MIC required for ceftolozane is less than with other cephalosporins and may be due to more rapid killing.

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MetR-Regulated Vibrio cholerae Metabolism Is Required for Virulence

mBio ◽

10.1128/mbio.00236-12 ◽

2012 ◽

Vol 3 (5) ◽

Cited By ~ 28

Author(s):

Ryan W. Bogard ◽

Bryan W. Davies ◽

John J. Mekalanos

Keyword(s):

Amino Acid ◽

Vibrio Cholerae ◽

Virulence Factor ◽

Current Knowledge ◽

Intestinal Colonization ◽

Wild Type ◽

Pathogenic Potential ◽

Content Type ◽

Mouse Intestine

ABSTRACTLysR-type transcriptional regulators (LTTRs) are the largest, most diverse family of prokaryotic transcription factors, with regulatory roles spanning metabolism, cell growth and division, and pathogenesis. Using a sequence-defined transposon mutant library, we screened a panel ofV. choleraeEl Tor mutants to identify LTTRs required for host intestinal colonization. Surprisingly, out of 38 LTTRs, only one severely affected intestinal colonization in the suckling mouse model of cholera: the methionine metabolism regulator, MetR. Genetic analysis of genes influenced by MetR revealed thatglyA1andmetJwere also required for intestinal colonization. Chromatin immunoprecipitation of MetR and quantitative reverse transcription-PCR (qRT-PCR) confirmed interaction with and regulation ofglyA1, indicating that misregulation ofglyA1is likely responsible for the colonization defect observed in themetRmutant. TheglyA1mutant was auxotrophic for glycine but exhibited wild-type trimethoprim sensitivity, making folate deficiency an unlikely cause of its colonization defect. MetJ regulatory mutants are not auxotrophic but are likely altered in the regulation of amino acid-biosynthetic pathways, including those for methionine, glycine, and serine, and this misregulation likely explains its colonization defect. However, mutants defective in methionine, serine, and cysteine biosynthesis exhibited wild-type virulence, suggesting that these amino acids can be scavenged in vivo. Taken together, our results suggest that glycine biosynthesis may be required to alleviate an in vivo nutritional restriction in the mouse intestine; however, additional roles for glycine may exist. Irrespective of the precise nature of this requirement, this study illustrates the importance of pathogen metabolism, and the regulation thereof, as a virulence factor.IMPORTANCEVibrio choleraecontinues to be a severe cause of morbidity and mortality in developing countries. Identification ofV. choleraefactors critical to disease progression offers the potential to develop or improve upon therapeutics and prevention strategies. To increase the efficiency of virulence factor discovery, we employed a regulator-centric approach to multiplex our in vivo screening capabilities and allow whole regulons inV. choleraeto be interrogated for pathogenic potential. We identified MetR as a new virulence regulator and serine hydroxymethyltransferase GlyA1 as a new MetR-regulated virulence factor, both required byV. choleraeto colonize the infant mouse intestine. Bacterial metabolism is a prerequisite to virulence, and current knowledge of in vivo metabolism of pathogens is limited. Here, we expand the known role of amino acid metabolism and regulation in virulence and offer new insights into the in vivo metabolic requirements ofV. choleraewithin the mouse intestine.

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Dynamics ofMycobacterium tuberculosisAg85B Revealed by a Sensitive Enzyme-Linked Immunosorbent Assay

mBio ◽

10.1128/mbio.00611-19 ◽

2019 ◽

Vol 10 (2) ◽

Cited By ~ 2

Author(s):

Joel D. Ernst ◽

Amber Cornelius ◽

Miriam Bolz

Keyword(s):

Protein Secretion ◽

Immunosorbent Assay ◽

Bacterial Population ◽

Enzyme Linked Immunosorbent Assay ◽

Bacterial Protein ◽

Gram Positive ◽

Gram Negative ◽

Content Type ◽

Bacterial Proteins

ABSTRACTSecretion of specific proteins contributes to pathogenesis and immune responses in tuberculosis and other bacterial infections, yet the kinetics of protein secretion and fate of secreted proteinsin vivoare poorly understood. We generated new monoclonal antibodies that recognize theMycobacteriumtuberculosissecreted protein Ag85B and used them to establish and characterize a sensitive enzyme-linked immunosorbent assay (ELISA) to quantitate Ag85B in samples generatedin vitroandin vivo. We found that nutritional or culture conditions had little impact on the secretion of Ag85B and that there is considerable variation in Ag85B secretion by distinct strains in theM. tuberculosiscomplex: compared with the commonly used H37Rv strain (lineage 4),Mycobacteriumafricanum(lineage 6) secretes less Ag85B, and two strains from lineage 2 secrete more Ag85B. We also used the ELISA to determine that the rate of secretion of Ag85B is 10- to 100-fold lower than that of proteins secreted by Gram-negative and Gram-positive bacteria, respectively. ELISA quantitation of Ag85B in lung homogenates ofM. tuberculosisH37Rv-infected mice revealed that although Ag85B accumulates in the lungs as the bacterial population expands, the amount of Ag85B per bacterium decreases nearly 10,000-fold at later stages of infection, coincident with the development of T cell responses and arrest of bacterial population growth. These results indicate that bacterial protein secretionin vivois dynamic and regulated, and quantitation of secreted bacterial proteins can contribute to the understanding of pathogenesis and immunity in tuberculosis and other infections.IMPORTANCEBacterial protein secretion contributes to host-pathogen interactions, yet the process and consequences of bacterial protein secretion during infection are poorly understood. We developed a sensitive ELISA to quantitate a protein (termed Ag85B) secreted byM. tuberculosisand used it to find that Ag85B secretion occurs with slower kinetics than for proteins secreted by Gram-positive and Gram-negative bacteria and that accumulation of Ag85B in the lungs is markedly regulated as a function of the bacterial population density. Our results demonstrate that quantitation of bacterial proteins during infection can reveal novel insights into host-pathogen interactions.

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Protein Nanowires: the Electrification of the Microbial World and Maybe Our Own

Journal of Bacteriology ◽

10.1128/jb.00331-20 ◽

2020 ◽

Vol 202 (20) ◽

Cited By ~ 2

Author(s):

Derek R. Lovley ◽

Dawn E. Holmes

Keyword(s):

Electron Transport ◽

Long Range ◽

Electronic Devices ◽

Electricity Production ◽

Microbial World ◽

Electrically Conductive ◽

Anaerobic Digesters ◽

Content Type ◽

Type Iv ◽

Type Iv Pilin

ABSTRACT Electrically conductive protein nanowires appear to be widespread in the microbial world and are a revolutionary “green” material for the fabrication of electronic devices. Electrically conductive pili (e-pili) assembled from type IV pilin monomers have independently evolved multiple times in microbial history as have electrically conductive archaella (e-archaella) assembled from homologous archaellin monomers. A role for e-pili in long-range (micrometer) extracellular electron transport has been demonstrated in some microbes. The surprising finding of e-pili in syntrophic bacteria and the role of e-pili as conduits for direct interspecies electron transfer have necessitated a reassessment of routes for electron flux in important methanogenic environments, such as anaerobic digesters and terrestrial wetlands. Pilin monomers similar to those found in e-pili may also be a major building block of the conductive “cables” that transport electrons over centimeter distances through continuous filaments of cable bacteria consisting of a thousand cells or more. Protein nanowires harvested from microbes have many functional and sustainability advantages over traditional nanowire materials and have already yielded novel electronic devices for sustainable electricity production, neuromorphic memory, and sensing. e-pili can be mass produced with an Escherichia coli chassis, providing a ready source of material for electronics as well as for studies on the basic mechanisms for long-range electron transport along protein nanowires. Continued exploration is required to better understand the electrification of microbial communities with microbial nanowires and to expand the “green toolbox” of sustainable materials for wiring and powering the emerging “Internet of things.”

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ZN148 Is a Modular Synthetic Metallo-β-Lactamase Inhibitor That Reverses Carbapenem Resistance in Gram-Negative Pathogens In Vivo

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02415-19 ◽

2020 ◽

Vol 64 (6) ◽

Cited By ~ 3

Author(s):

Ørjan Samuelsen ◽

Ove Alexander Høgmoen Åstrand ◽

Christopher Fröhlich ◽

Adam Heikal ◽

Susann Skagseth ◽

...

Keyword(s):

Active Site ◽

Therapeutic Potential ◽

Treatment Options ◽

Carbapenem Resistance ◽

Functional Space ◽

Bactericidal Effect ◽

Gram Negative ◽

Content Type

ABSTRACT Carbapenem-resistant Gram-negative pathogens are a critical public health threat and there is an urgent need for new treatments. Carbapenemases (β-lactamases able to inactivate carbapenems) have been identified in both serine β-lactamase (SBL) and metallo-β-lactamase (MBL) families. The recent introduction of SBL carbapenemase inhibitors has provided alternative therapeutic options. Unfortunately, there are no approved inhibitors of MBL-mediated carbapenem-resistance and treatment options for infections caused by MBL-producing Gram-negatives are limited. Here, we present ZN148, a zinc-chelating MBL-inhibitor capable of restoring the bactericidal effect of meropenem and in vitro clinical susceptibility to carbapenems in >98% of a large international collection of MBL-producing clinical Enterobacterales strains (n = 234). Moreover, ZN148 was able to potentiate the effect of meropenem against NDM-1-producing Klebsiella pneumoniae in a murine neutropenic peritonitis model. ZN148 showed no inhibition of the human zinc-containing enzyme glyoxylase II at 500 μM, and no acute toxicity was observed in an in vivo mouse model with cumulative dosages up to 128 mg/kg. Biochemical analysis showed a time-dependent inhibition of MBLs by ZN148 and removal of zinc ions from the active site. Addition of exogenous zinc after ZN148 exposure only restored MBL activity by ∼30%, suggesting an irreversible mechanism of inhibition. Mass-spectrometry and molecular modeling indicated potential oxidation of the active site Cys221 residue. Overall, these results demonstrate the therapeutic potential of a ZN148-carbapenem combination against MBL-producing Gram-negative pathogens and that ZN148 is a highly promising MBL inhibitor that is capable of operating in a functional space not presently filled by any clinically approved compound.

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Release of collagen type IV degrading activity from C6 astrocytoma cells and cell density

Journal of Neurosurgery ◽

10.3171/jns.1996.84.6.1013 ◽

1996 ◽

Vol 84 (6) ◽

pp. 1013-1019 ◽

Cited By ~ 8

Author(s):

Masashi Tamaki ◽

Warren McDonald ◽

Rolando F. Del Maestro

Keyword(s):

Cell Density ◽

Collagen Type ◽

Cell Communication ◽

Collagen Type Iv ◽

Major Protein ◽

Content Type ◽

Type Iv ◽

Astrocytoma Cells ◽

C6 Astrocytoma

✓ Type IV collagen is a major protein component of the vascular basement membrane and its degradation is crucial to the initiation of tumor-associated angiogenesis. The authors have investigated the influence of cell density on the release of collagen type IV degrading activity by C6 astrocytoma cells in monolayer culture. The release of collagen type IV degrading activity was assessed biochemically, immunocytochemically, and by Western blot analysis. The results demonstrate that increasing plating density and increasing cell density are associated with decreased collagen type IV degrading activity released per tumor cell. These findings indicate the existence of regulatory mechanisms dependent on cell—cell communication, which modulate release of collagen type IV degrading activity. The extrapolation of these results to the in vivo tumor microenvironment would suggest that individual and/or small groups of invading tumor cells, distant from the main tumor mass, would release substantial collagen type IV degrading activity, which may be crucial to their continued invasion and to angiogenesis.

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Abrogation of Neuraminidase Reduces Biofilm Formation, Capsule Biosynthesis, and Virulence of Porphyromonas gingivalis

Infection and Immunity ◽

10.1128/iai.05773-11 ◽

2011 ◽

Vol 80 (1) ◽

pp. 3-13 ◽

Cited By ~ 27

Author(s):

Chen Li ◽

Kurniyati ◽

Bo Hu ◽

Jiang Bian ◽

Jianlan Sun ◽

...

Keyword(s):

Porphyromonas Gingivalis ◽

Biofilm Formation ◽

Adult Population ◽

Transcriptional Analysis ◽

Wild Type ◽

Content Type ◽

Multiple Organs ◽

Biochemical Analyses

ABSTRACTThe oral bacteriumPorphyromonas gingivalisis a key etiological agent of human periodontitis, a prevalent chronic disease that affects up to 80% of the adult population worldwide.P. gingivalisexhibits neuraminidase activity. However, the enzyme responsible for this activity, its biochemical features, and its role in the physiology and virulence ofP. gingivalisremain elusive. In this report, we found thatP. gingivalisencodes a neuraminidase, PG0352 (SiaPg). Transcriptional analysis showed thatPG0352is monocistronic and is regulated by a sigma70-like promoter. Biochemical analyses demonstrated that SiaPgis an exo-α-neuraminidase that cleaves glycosidic-linked sialic acids. Cryoelectron microscopy and tomography analyses revealed that thePG0352deletion mutant (ΔPG352) failed to produce an intact capsule layer. Compared to the wild type,in vitrostudies showed that ΔPG352 formed less biofilm and was less resistant to killing by the host complement.In vivostudies showed that while the wild type caused a spreading type of infection that affected multiple organs and all infected mice were killed, ΔPG352 only caused localized infection and all animals survived. Taken together, these results demonstrate that SiaPgis an important virulence factor that contributes to the biofilm formation, capsule biosynthesis, and pathogenicity ofP. gingivalis, and it can potentially serve as a new target for developing therapeutic agents againstP. gingivalisinfection.

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CodY Is a Global Regulator of Virulence-Associated Properties for Clostridium perfringens Type D Strain CN3718

mBio ◽

10.1128/mbio.00770-13 ◽

2013 ◽

Vol 4 (5) ◽

Cited By ~ 27

Author(s):

Jihong Li ◽

Menglin Ma ◽

Mahfuzur R. Sarker ◽

Bruce A. McClane

Keyword(s):

Clostridium Perfringens ◽

Null Mutant ◽

Intestinal Infection ◽

Wild Type ◽

Global Regulator ◽

Content Type ◽

Gram Positive Bacteria ◽

Type D ◽

Epsilon Toxin ◽

Virulence Properties

ABSTRACT CodY is known to regulate various virulence properties in several Gram-positive bacteria but has not yet been studied in the important histotoxic and intestinal pathogen Clostridium perfringens. The present study prepared an isogenic codY-null mutant in C. perfringens type D strain CN3718 by insertional mutagenesis using the Targetron system. Western blot analysis indicated that, relative to wild-type CN3718 or a complementing strain, this isogenic codY mutant produces reduced levels of epsilon toxin (ETX). Using supernatants from cultures of the wild-type, codY-null mutant, and complementing strains, CodY regulation of ETX production was shown to have cytotoxic consequences for MDCK cells. The CodY regulatory effect on ETX production was specific, since the codY-null mutant still made wild-type levels of alpha-toxin and perfringolysin O. Sialidase activity measurements and sialidase Western blot analysis of supernatants from CN3718 and its isogenic derivatives showed that CodY represses overall exosialidase activity due to a reduced presence of NanH in culture supernatants. Inactivation of the codY gene significantly decreased the adherence of CN3718 vegetative cells or spores to host Caco-2 cells. Finally, the codY mutant showed increased spore formation under vegetative growth conditions, although germination of these spores was impaired. Overall, these results identify CodY as a global regulator of many C. perfringens virulence-associated properties. Furthermore, they establish that, via CodY, CN3718 coordinately regulates many virulence-associated properties likely needed for intestinal infection. IMPORTANCE Clostridium perfringens is a major human and livestock pathogen because it produces many potent toxins. C. perfringens type D strains cause intestinal infections by producing toxins, especially epsilon toxin (ETX). Previous studies identified CodY as a regulator of certain virulence properties in other Gram-positive bacteria. Our study now demonstrates that CodY is a global regulator of virulence-associated properties for type D strain CN3718. It promotes production of ETX, attachment of CN3718 vegetative cells or spores to host enterocyte-like Caco-2 cells, and spore germination; the last two effects may assist intestinal colonization. In contrast, CodY represses sporulation. These results provide the first evidence that CodY can function as a global regulator of C. perfringens virulence-associated properties and that this strain coordinately regulates its virulence-associated properties using CodY to increase ETX production, host cell attachment, and spore germination but to repress sporulation, as would be optimal during type D intestinal infection.

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