Journal of Bacteriology最新文献

{"title":"Contribution of PonA2 to biofilm and colony formation independent of its catalytic domains in <i>Mycobacterium smegmatis</i>.","authors":"Bryan A Montero-Gutierrez, Jathin K Bande, Mercedes D Stites, Takehiro Kado","doi":"10.1128/jb.00301-26","DOIUrl":"https://doi.org/10.1128/jb.00301-26","url":null,"abstract":"<p><p>Biofilm and colony growth create microenvironments that require coordinated localization and function of cell envelope components. However, the mechanisms by which membrane organization contributes to cell envelope organization during multicellular growth remain poorly understood. <i>Mycobacterium smegmatis</i> contains an inner membrane domain (IMD) at the subpolar regions of the cell that contributes to cell envelope synthesis. Prior research identified PonA2 as important for <i>de novo</i> formation of distinct plasma membrane domains during planktonic growth. PonA2 is a penicillin-binding protein with transglycosylase (TG) and transpeptidase (TP) activities involved in peptidoglycan synthesis. To investigate the role of PonA2 during colony and pellicle biofilm growth, wild-type, Δ<i>ponA2</i>, the complement strain (c<i>ponA2</i>), and catalytically inactive PonA2 variants (TG-, TP-, and TG-/TP-) were analyzed. Subpolar IMD localization was maintained in wild-type cells during both colony and pellicle biofilm growth, demonstrating that IMD localization is not exclusive to planktonic growth. Loss of <i>ponA2</i> resulted in smooth colony and early pellicle biofilm architecture and was associated with reduced abundance of trehalose monomycolate, trehalose dimycolate, and glycopeptidolipids. Δ<i>ponA2</i> cells in colony growth exhibited shortened cell morphology and lacked detectable IMD localization, whereas Δ<i>ponA2</i> cells in pellicle biofilm growth retained bacillary morphology and subpolar IMD localization. In contrast, the catalytically inactive PonA2 variants maintained rugose multicellular architecture, bacillary morphology, and subpolar IMD localization similar to that of wild-type. Together, these findings identify a previously unrecognized noncatalytic role for PonA2 in maintaining normal colony and pellicle biofilm development in <i>M. smegmatis</i>.IMPORTANCEPlanktonic, pellicle biofilm, and colony growth expose mycobacteria to distinct environmental conditions that can affect cell-envelope organization and survival. Prior research has shown that mycobacteria form subpolar plasma membrane domains that support polar cell elongation in planktonic growth, but it remains unclear whether this organization is conserved in multicellular biofilm and colony growth, where cells experience nutrient and oxygen gradients and altered cell-to-cell interactions. Our current study analyzed the plasma membrane domain formation across the two growth states and showed that membrane domain localization is conserved, while the mechanisms required to maintain these domains differ depending on growth conditions. These findings suggest that mycobacteria use growth state-specific mechanisms to coordinate membrane organization.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0030126"},"PeriodicalIF":3.8,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182113","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Thank you to our recently retired editors Joe Bondy-Denomy and Tina Henkin.","authors":"George A O'Toole","doi":"10.1128/jb.00507-26","DOIUrl":"https://doi.org/10.1128/jb.00507-26","url":null,"abstract":"","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0050726"},"PeriodicalIF":3.8,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182051","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Correction for Robin et al., \"The CpxAR signaling system confers a fitness advantage for flea gut colonization by the plague bacillus\".","authors":"Brandon Robin, Amélie Dewitte, Véronique Alaimo, Cecile Lecoeur, François Pierre, Gabriel Billon, Florent Sebbane, Sébastien Bontemps-Gallo","doi":"10.1128/jb.00426-26","DOIUrl":"https://doi.org/10.1128/jb.00426-26","url":null,"abstract":"","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0042626"},"PeriodicalIF":3.8,"publicationDate":"2026-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182093","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The virulence regulator ToxT influences <i>Vibrio cholerae</i> motility through the expression of the methyl-accepting chemoreceptors AcfB and TcpI.","authors":"Carlos Candanosa, Michael Cole Carter, Venus Stanton, Mark Eppinger, Karl E Klose","doi":"10.1128/jb.00425-26","DOIUrl":"https://doi.org/10.1128/jb.00425-26","url":null,"abstract":"<p><p><i>Vibrio cholerae</i> is the causative agent of the human diarrheal disease cholera. Previous studies identified \"hyperswarming\" (HS) mutant <i>V. cholerae</i> with enhanced motility and decreased virulence factor expression, but the genetic basis of HS was unknown. Through whole-genome sequencing, we have identified point mutations within <i>toxT</i>, which encodes the master virulence regulator, as being responsible for the HS phenotype. These mutations are loss of function, and they eliminate transcription of cholera toxin and toxin-coregulated pilus genes. Using single-cell video tracking, these HS strains were also observed to exhibit enhanced motility/chemotaxis (increased speed and decreased directional change). Two methyl-accepting chemoreceptors (methyl-accepting chemotaxis proteins), AcfB and TcpI, are positively regulated by ToxT, and expression of both AcfB and TcpI within HS mutants restores wild-type motility. Moreover, a <i>V. cholerae</i> strain with a hyperactive ToxT (N106A) exhibits reduced motility (decreased speed and increased directional change), and knocking out <i>acfB</i> and <i>tcpI</i> in this strain restores wild-type motility. AcfB and TcpI localize to the bacterial cell poles, suggesting they incorporate into the chemotaxis arrays to modulate swimming behavior. Our findings demonstrate that the absence of the ToxT-dependent MCPs AcfB and TcpI is responsible for the HS phenotype, demonstrating a direct role of ToxT in the swimming behavior of <i>V. cholerae</i>.IMPORTANCE<i>Vibrio cholerae</i> causes the human diarrheal disease cholera, and the master virulence regulator ToxT is responsible for the expression of the most important virulence factors, cholera toxin and the toxin-coregulated pilus, necessary for disease. Motility is also important for the bacteria to cause disease, and in this study, we provide evidence that ToxT also modulates motility, providing a regulatory link between motility and the other virulence factors required for the pathogenic lifestyle of <i>V. cholerae</i>.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0042526"},"PeriodicalIF":3.8,"publicationDate":"2026-10-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182078","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The evolution of <i>Brucella</i> as a model intracellular pathogen.","authors":"Jeannette E Farnham, Sean Crosson","doi":"10.1128/jb.00404-26","DOIUrl":"https://doi.org/10.1128/jb.00404-26","url":null,"abstract":"<p><p><i>Brucella</i> species are intracellular bacterial pathogens that cause brucellosis, one of the most common zoonoses worldwide. This review traces how <i>Brucella</i>, first discovered in the 19th century through investigations of human and bovine disease, became a modern experimental model system. Work on <i>Brucella</i> has illuminated how intracellular bacteria evade and adapt to host defenses and how they remodel host cell biology to establish a replicative niche. It has also yielded insights that extend beyond pathogenesis into bacterial cell envelope architecture and development. Though long regarded as a small, homogeneous group of mammalian pathogens, the genus has expanded in recent years as \"non-classical\" <i>Brucella</i> have been isolated from unexpected animal hosts, opening new opportunities to understand the fascinating host preferences and evolutionary history of these closely related Alphaproteobacteria. More than a century after their discovery, <i>Brucella</i> species remain versatile models for the study of intracellular infection, bacterial cell biology, and physiology, and the evolution of host association.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0040426"},"PeriodicalIF":3.8,"publicationDate":"2026-10-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182105","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Molecular mechanisms and regulatory networks governing <i>Yersinia</i> biofilm formation and host adaptation.","authors":"Haisheng Wu, Hailian Wu, Wenxuan Peng, Yiquan Zhang, Qingwen Zhang","doi":"10.1128/jb.00418-26","DOIUrl":"https://doi.org/10.1128/jb.00418-26","url":null,"abstract":"<p><p>The genus <i>Yersinia</i> includes three human pathogens-<i>Y</i>. <i>pestis</i>, <i>Y. pseudotuberculosis</i>, and <i>Y. enterocolitica</i>-where biofilm formation plays distinct roles in transmission and environmental persistence. All three species share the core biofilm machinery-the <i>hmsHFRS</i> operon, encoding enzymes for poly-β-1,6-N-acetylglucosamine (PNAG) exopolysaccharide production, and the c-di-GMP signaling controlling this production-but their regulatory networks and biofilm outputs differ. Transcriptional control of <i>hms</i> involves an integrated network that includes the Rcs phosphorelay (reshaped by <i>rcsA</i> pseudogenization), PhoP/PhoQ, EnvZ/OmpR, BarA/UvrY, Fur/Zur, CRP-cAMP, RovM/RovA, and multiple sigma factors. Post-transcriptionally, the Hfq-sRNA and CsrA-CsrB/CsrC systems modulate gene expression; the latter is directly validated in <i>Y. pestis</i> and <i>Y. pseudotuberculosis</i>, whereas its role in <i>Y. enterocolitica</i> is inferred from genomic conservation. At the translational and enzymatic levels, polyamines sustain Hms protein production, while redox signals govern the HmsC-HmsD disulfide switch, controlling diguanylate cyclase activity. This multi-layer architecture ensures that biofilm output is tuned to species-specific ecological niches. In host adaptation, <i>Y. pestis</i> transitioned from a foodborne enteric pathogen to a flea-borne lifestyle through key genetic events-acquisition of <i>ymt</i>, pseudogenization of <i>rcsA</i>, inactivation of <i>y2909</i>/<i>y3389,</i> and <i>ureD</i>-which are not shared with <i>Y. pseudotuberculosis</i> or <i>Y. enterocolitica</i>, thereby explaining their different biofilm-dependent transmission strategies. Studies in <i>Caenorhabditis elegans</i> have identified host O-glycosylation modifications as receptors for biofilm attachment. This review summarizes progress on the molecular mechanisms, regulatory networks, and host-adaptive functions of <i>Yersinia</i> biofilm formation and proposes future research directions to address knowledge gaps. Future priorities include the identification of c-di-GMP effectors, flea-specific signals, 3D biofilm imaging, and CRISPR-based detection.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0041826"},"PeriodicalIF":3.8,"publicationDate":"2026-10-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182062","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Anti-mycobacterial activity of phage-encoded lysterases.","authors":"Jindanuch Maneekul, Krista G Freeman, Elizabeth D Amarh, Julia M Kovalski, Emma Skeel, Manisha Vannan, Matthew T Montgomery, Casey Moyer, Rebecca A Garlena, Amit K Singh, Pallavi Ghosh, Anil K Ojha, Graham F Hatfull","doi":"10.1128/jb.00410-26","DOIUrl":"https://doi.org/10.1128/jb.00410-26","url":null,"abstract":"<p><p><i>Mycobacterium abscessus</i> is a non-tuberculous <i>Mycobacterium</i> (NTM) that is often associated with respiratory infections in persons with cystic fibrosis. High levels of antibiotic resistance make <i>M. abscessus</i> infections difficult to treat, highlighting the urgent need for alternative therapeutic strategies. Mycobacteriophage therapy-treatment of infections with viruses that infect <i>Mycobacterium</i> hosts-shows considerable promise, but the narrow host specificity of the current mycobacteriophage repertoire limits this approach to personalized therapies. Here, we characterize the anti-<i>M</i>. <i>abscessus</i> activity of mycobacteriophage-encoded Lysin B enzymes (lysterases). Lysterases are serine-esterases that cleave the ester bonds linking mycolic acids to either arabinogalactan or trehalose in the mycobacterial cell envelope to complete the lysis process. In this study, we explored their ability to kill a spectrum of clinically isolated <i>M. abscessus</i> strains when administered exogenously. Six phylogenetically distinct lysterases were purified, characterized on the surrogate <i>M. smegmatis</i>, and their activity against a diverse panel of 24 <i>M. abscessus</i> clinical strains was assessed. The enzymes exhibit a range of activities against these strains, but most have broad-spectrum activity with only minor variations among the <i>M. abscessus</i> isolates. However, the activities can be quite potent, resulting in up to six orders of magnitude of reduction in viability within 4 h with one activity unit of the enzymes, and act synergistically with amikacin. Overall, mycobacteriophage-encoded lysterases show considerable potential for antimycobacterial control, especially for topical applications.IMPORTANCE<i>Mycobacterium</i> infections are clinically challenging, and there is a high failure of resolving <i>Mycobacterium abscessus</i> infections using antibiotics. Mycobacteriophages show promise in treating <i>M. abscessus</i> infections, but there is substantial variation among clinical isolates in their phage infection profiles. Mycobacteriophages are unusual in coding for lysis enzymes-lysterases-that target cell wall exposed constituents, and these can have potential antibacterial activity. Exploring the vast sequence diversity of thousands of phage-encoded lysterases identifies a subset that was expressed, purified, and tested for their ability to kill a broad range of <i>M. abscessus</i> clinical strains. Lysterases with the broadest and greatest activity offer substantial potential for treatment of <i>M. abscessus</i> infection, especially where they display synergism with current antibiotics.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0041026"},"PeriodicalIF":3.8,"publicationDate":"2026-10-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149182018","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Structural basis for ITP-selective hydrolysis by a CRISPR-associated Nudix hydrolase.","authors":"Wanqian Wu, Shenglan Xia, Duo Li, Junchao Lin, Chen Cao, Jianping Kong, Meiling Lu, Yibei Xiao, Zhaoxing Li","doi":"10.1128/jb.00280-26","DOIUrl":"https://doi.org/10.1128/jb.00280-26","url":null,"abstract":"<p><p>The nucleoside diphosphates linked to moiety-X (Nudix) hydrolases are a superfamily of proteins defined by the characteristic Nudix box sequence, most of which are involved in clearing the cell of potentially deleterious metabolites and in preventing the accumulation of metabolic intermediates. The Nudix hydrolase associated with CRISPR-Cas-associated adenosine deaminases (CAAD) from <i>Limisphaera ngatamarikiensis</i> (<i>Lng</i>NH) mitigates CAAD-induced toxicity by hydrolyzing ITP to IMP and pyrophosphate. Here, we determined crystal structures of <i>Lng</i>NH in the apo form and in complex with ITP using catalytic mutants and validated key interactions by site-directed mutagenesis. The hypoxanthine of ITP is deeply buried in a hydrophobic pocket and engages a distinct hydrogen-bond network, whereas the triphosphate tail is coordinated by conserved residues and divalent metal ions. Together, these findings provide a structural framework for CAAD-linked nucleotide sanitization and suggest that CRISPR-associated Nudix hydrolases function as local self-protection modules that limit toxicity from noncanonical inosine nucleotides.IMPORTANCENucleotide quality-control enzymes protect cells from harmful metabolic by-products. Here, we characterize a CRISPR-associated nucleoside diphosphates linked to moiety-X (Nudix) hydrolase that removes ITP, a toxic noncanonical nucleotide generated during antiviral defense. Biochemical and structural analyses show how this enzyme specifically recognizes and hydrolyzes ITP while avoiding related cellular nucleotides. Our findings suggest that CRISPR-associated nucleotide deaminase systems require \"self-protection\" mechanisms to prevent damage from their own antiviral activities. These results expand our understanding of how bacteria maintain nucleotide homeostasis during immune responses and reveal an additional regulatory layer within CRISPR-associated defense systems. The structural framework reported here also provides a foundation for future studies of nucleotide sanitization enzymes in microbial physiology and antiviral immunity.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0028026"},"PeriodicalIF":3.8,"publicationDate":"2026-10-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149165957","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Environmental context defines the physiological contribution of the Tol-Pal system to cell envelope integrity in <i>Acinetobacter baumannii</i>.","authors":"Roberto Jhonatan Olea-Ozuna, Berenice Furlan, Suman Tiwari, Hanling Gong, Augusto C Hunt-Serracin, Michael B Whalen, Orietta Massidda, Nicholas Dillon, Joseph M Boll","doi":"10.1128/jb.00289-26","DOIUrl":"https://doi.org/10.1128/jb.00289-26","url":null,"abstract":"<p><p>The gram-negative cell envelope requires coordinated remodeling of the peptidoglycan layer and outer membrane to preserve envelope integrity during growth and division. The conserved Tol-Pal system contributes to envelope maintenance, although the physiological consequences of its loss vary across environmental contexts and among gram-negative bacteria. Here, we examined the contribution of Tol-Pal across diverse growth environments in <i>Acinetobacter baumannii</i>. Loss of Tol-Pal did not cause a substantial population growth defect, and septal peptidoglycan incorporation remained largely preserved under standard laboratory conditions. In contrast, nutrient-rich media, conditions that alter osmotic balance, and a host-like environment exposed defects in cell division organization and cell morphology. Tol-Pal-deficient cells also exhibited modest but reproducible reductions in outer membrane barrier robustness, and disruption of the Mla phospholipid transport system further exacerbated these defects. Consistent with these phenotypes, loss of Tol-Pal reduced bacterial fitness under environmental stress and host-associated models. Together, these findings indicate that Tol-Pal contributes to outer membrane barrier robustness and that additional consequences of its loss become apparent under conditions that challenge the cell envelope.IMPORTANCEThe physiological consequences of disrupting conserved envelope systems can vary with the conditions in which bacteria grow. In <i>Acinetobacter baumannii</i>, loss of Tol-Pal has little effect on population growth under routine laboratory conditions but modestly compromises outer membrane barrier robustness, whereas specific environmental and host-associated conditions reveal additional defects in cell division organization and bacterial fitness. These findings highlight how environmental context shapes the physiological consequences of Tol-Pal deficiency and provide a framework for understanding why the contribution of this conserved envelope system differs among gram-negative bacteria.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0028926"},"PeriodicalIF":3.8,"publicationDate":"2026-10-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149168963","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"EndD mediates butyrate-dependent toxin release and sporulation in <i>Clostridioides difficile</i>.","authors":"Horia A Dobrila, Henrieta Licha, Andrew J Hryckowian","doi":"10.1128/jb.00341-26","DOIUrl":"10.1128/jb.00341-26","url":null,"abstract":"<p><p><i>Clostridioides difficile</i> is an urgent threat to human health. Current treatments for <i>C. difficile</i> infections (CDIs) are antibiotics and microbiome restoration therapy (MRT) for recurrent cases. However, antibiotics contribute to antibiotic resistance and recurrent CDIs, and the long-term sustainability and accessibility of MRTs remain to be determined. Since a dysbiotic gut microbiome is the primary risk factor for CDI, a better understanding of the interactions between <i>C. difficile</i>, the microbiome, and the host will aid development of treatments with improved precision. Emerging evidence supports that butyrate, a prominent end product of gut microbiome metabolism, is a key determinant of <i>C. difficile</i> pathogenesis. Notably, <i>C. difficile</i> releases more of its toxins (TcdA and TcdB) in butyrate-rich environments. Here, we demonstrate that butyrate-dependent toxin release is not driven by two previously characterized modes of toxin release (e.g., TcdE-dependent secretion or Cwp19-dependent autolysis). Instead, butyrate enhances the expression of a broadly conserved endolysin (EndD), which is responsible for butyrate-dependent toxin release. We additionally demonstrate that <i>endD</i>-dependent toxin release does not universally occur under all growth conditions, that its expression is dependent on the late-stage sporulation sigma factor SigK, and that <i>endD</i> enhances butyrate-dependent sporulation. Overall, our findings provide deeper insight into butyrate-dependent effects on <i>C. difficile</i> pathogenesis and set the stage for future work to better understand the molecular and genetic underpinnings of <i>endD</i> regulation.IMPORTANCE<i>Clostridioides difficile</i> is a leading cause of infectious diarrhea. The gut microbiome-derived short-chain fatty acid butyrate triggers <i>C. difficile</i> to release its major virulence factors, toxins TcdA and TcdB. Here, we show that butyrate-dependent toxin release is independent of classical secretion or autolytic pathways and is instead mediated by EndD, an endolysin broadly conserved among <i>C. difficile</i> clinical isolates. We also demonstrate that <i>endD</i> expression is governed by the sporulation factor SigK and enhances sporulation, directly linking transmission and virulence. Uncovering how <i>C. difficile</i> couples metabolic sensing to EndD-mediated toxin release and sporulation advances our understanding of pathogen-microbiome interactions and highlights potential targets for non-antibiotic therapeutics.</p>","PeriodicalId":15107,"journal":{"name":"Journal of Bacteriology","volume":" ","pages":"e0034126"},"PeriodicalIF":3.8,"publicationDate":"2026-10-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149109923","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}