Karl Ocius

Gut Microbes. 2026 Dec 31;18(1):2611603. doi: 10.1080/19490976.2025.2611603. Epub 2026 Jan 9.

ABSTRACT

The mechanisms by which gut microbiota modulate host immune responses remain incompletely understood. Here, we screened Lactobacillus and Bifidobacterium strains isolated from healthy individuals to identify symbionts capable of suppressing gut inflammation. Among them, Bifidobacterium adolescentis (Bifi-94) induced IL-10 production in mononuclear cells in vitro. Oral administration of Bifi-94 to mice treated with dextran sulfate sodium attenuated weight loss and reduced colonic inflammation scores. In wild-type C57BL/6 mice, Bifi-94 increased IL-10 levels in colonic tissue homogenates without altering the frequency of regulatory T cells. Instead, CD19⁺CD11b⁺ regulatory B (Breg) cells emerged as the primary source of IL-10, with their numbers significantly increasing in the peritoneal cavity (PEC) after treatment. IL-10 secretion by PEC cells was robustly activated by live, heat-killed, and formalin-fixed Bifi-94. Bifi-94-derived peptidoglycan (PG) selectively stimulated IL-10 production in CD19⁺CD11b⁺ Breg cells, and multi-omics analyses showed that Bifi-94 exhibits increased expression of PG biosynthetic enzymes (MurE, MurD, Alr, UppP) relative to the type strain. Mechanistically, Bifi-94-derived PG promoted TLR2-dependent activation of ERK and p38 MAPK signaling in Breg cells. Notably, PG similarly enhanced IL-10 production in CD19⁺ B cells from human colonic tissue. These findings demonstrate that Bifi-94-derived PG promotes IL-10 production in Breg cells via TLR2-mediated signaling, thereby contributing to the attenuation of gut inflammation.

PMID:41511071 | PMC:PMC12795277 | DOI:10.1080/19490976.2025.2611603

Antibiotics (Basel). 2025 Dec 1;14(12):1210. doi: 10.3390/antibiotics14121210.

ABSTRACT

LD-Transpeptidases (LDTs) are a widely conserved class of peptidoglycan (PG) crosslinking enzymes in bacteria. They are sometimes overlooked as they often act secondary to penicillin binding proteins (PBPs) under standard conditions. However, LDTs are essential in key pathogens such as Clostridioides difficile and are responsible for β-lactam resistance in Mycobacterium tuberculosis and Enterococcus faecium due their low affinity for penicillins and cephalosporins, allowing them to form LD-crosslinks when DD-crosslinking PBPs are inactivated. This role makes LDTs a promising target when developing new treatments for these pathogens. LDTs can perform different enzymatic reactions. Most commonly they reinforce the PG with 3,3-LD-crosslinks or, in a few cases, 1,3-LD-crosslinks, during stationary phase or stress responses. Some LDTs also incorporate endogenous and exogenous non-canonical D-amino acids into the PG. In many Gram-negative bacteria, specialised LDTs tether lipoproteins or outer membrane proteins (OMPs) to the PG to maintain cell envelope integrity; in some cases this regulates virulence factors. Specialised LDTs have also been implied to have roles in polar growth, toxin secretion, and symbiotic colonisation. Recent discoveries include novel subgroups of the major YkuD family and the identification of the VanW family; this has opened new research directions surrounding LDTs. We aim to understand LDTs and their roles to expand our knowledge of PG synthesis and modification and how these enzymes can be targeted for antibiotic treatment.

PMID:41463713 | PMC:PMC12729407 | DOI:10.3390/antibiotics14121210

Expert Opin Ther Targets. 2026 Jan 6:1-18. doi: 10.1080/14728222.2025.2608020. Online ahead of print.

ABSTRACT

INTRODUCTION: Despite the fact that the progress in developing new antibiotics with novel mechanism of action is limited owing to the considerable challenges involved and the modest financial returns, we are urgently needing new molecules with new mode of actions to overcome the increasing burden of multidrug resistance. Half of all antibiotics prescribed for human use target the bacterial cell wall. Targeting the cell wall has many advantages as it is essential for bacterial survival, with many steps readily accessible from the environment to small molecules and despite the success of beta-lactams, the pathway still has many underexploited targets.

AREAS COVERED: Here, we review the limitations and the gaps that remain to be filled and the latest developments in new targets, strategies and drugs aimed at inhibiting the assembly of a viable cell wall, as this pathway remains a formidable and accessible source of new targets.

EXPERT OPINION: The future of the field should benefit immensely from computational and AI-driven methodologies, combinatorial therapies reducing the risk of development of resistance and acknowledging that we have been screening poorly adapted chemical spaces for antibiotic discovery.

PMID:41481307 | DOI:10.1080/14728222.2025.2608020

Infect Immun. 2026 Feb 10;94(2):e0047225. doi: 10.1128/iai.00472-25. Epub 2025 Dec 19.

ABSTRACT

Pathogenic chlamydial species restrict their peptidoglycan (PG) to the division septum of their replicative forms. PG is a microbe-associated molecular pattern, and two of its major pattern recognition receptors in human cells are nucleotide-binding oligomerization domain-containing proteins 1 and 2 (NOD1 and NOD2, respectively). It has been proposed that this unique morphological feature is evidence of pathoadaptation by the microbe, permitting PG-dependent cell division while also reducing the bacterium's recognition by innate immune receptors. Chlamydia trachomatis-infected cells activate NOD1 signaling within 8-12 hours of exposure to the bacterium, roughly coinciding with the microbe's transition from its infectious to replicative forms. Here, we report that, unlike NOD1 signaling, Chlamydia-induced NOD2 signaling does not occur until later in the pathogen's developmental cycle. Both C. trachomatis and the related murine pathogen Chlamydia muridarum signal late in infection in HEK293 reporter cell lines expressing either human or murine-derived NOD2 receptors. NOD2 signaling can be modulated by disruption of the chlamydial amidase enzyme, AmiA_CT, interrupting the microbe's developmental cycle, and treatment with inhibitors of lipooligosaccharide or PG biosynthesis/assembly. These results mirror prior observations with Chlamydia-induced TLR9 signaling, leading us to hypothesize that Chlamydia-induced NOD2 signaling results from lytic events that occur sporadically during the transition between the pathogen's developmental forms. Given our finding that pre-treating cells with NOD2-stimulatory ligands reduces chlamydial inclusion size and delays the developmental cycle, we hypothesize that the microbe preferentially degrades its PG during development to reduce the generation of NOD2 ligands.

PMID:41416804 | PMC:PMC12890027 | DOI:10.1128/iai.00472-25

Nat Microbiol. 2025 Dec 19. doi: 10.1038/s41564-025-02210-5. Online ahead of print.

ABSTRACT

Class A penicillin-binding proteins (aPBPs) are involved in the biosynthesis and remodelling of peptidoglycan (PG). The human bacterial pathogen Streptococcus pneumoniae produces three aPBPs, which are regulated to maintain the bacterium's ovoid shape. Evidence suggests that PBP1a and PBP2a activities are closely coordinated; however, their precise functions remain unclear. Here we characterized the pneumococcal S protein, which contains a LysM-PG-binding domain and a GpsB-interacting domain. Using S protein fusion constructs or mutant bacterial strains, we show that S protein localizes to the division ring and is required to prevent premature cell lysis and minicell formation due to aberrant division site placement. S protein interacts with PBP1a and activates its PG synthesis activity. Co-immunoprecipitation experiments combined with biochemical, genetic, structural prediction and microscopy analyses suggest that S protein is part of a larger multiprotein complex containing aPBPs and PG-modifying enzymes, and coordinated by the scaffolding protein GpsB. Together, these findings suggest that a GpsB-associated complex orchestrates PG biosynthesis and remodelling in S. pneumoniae.

PMID:41420061 | DOI:10.1038/s41564-025-02210-5

Infect Immun. 2025 Dec 19:e0047225. doi: 10.1128/iai.00472-25. Online ahead of print.

ABSTRACT

Pathogenic chlamydial species restrict their peptidoglycan (PG) to the division septum of their replicative forms. PG is a microbe-associated molecular pattern, and two of its major pattern recognition receptors in human cells are nucleotide-binding oligomerization domain-containing proteins 1 and 2 (NOD1 and NOD2, respectively). It has been proposed that this unique morphological feature is evidence of pathoadaptation by the microbe, permitting PG-dependent cell division while also reducing the bacterium's recognition by innate immune receptors. Chlamydia trachomatis-infected cells activate NOD1 signaling within 8-12 hours of exposure to the bacterium, roughly coinciding with the microbe's transition from its infectious to replicative forms. Here, we report that, unlike NOD1 signaling, Chlamydia-induced NOD2 signaling does not occur until later in the pathogen's developmental cycle. Both C. trachomatis and the related murine pathogen Chlamydia muridarum signal late in infection in HEK293 reporter cell lines expressing either human or murine-derived NOD2 receptors. NOD2 signaling can be modulated by disruption of the chlamydial amidase enzyme, AmiA_CT, interrupting the microbe's developmental cycle, and treatment with inhibitors of lipooligosaccharide or PG biosynthesis/assembly. These results mirror prior observations with Chlamydia-induced TLR9 signaling, leading us to hypothesize that Chlamydia-induced NOD2 signaling results from lytic events that occur sporadically during the transition between the pathogen's developmental forms. Given our finding that pre-treating cells with NOD2-stimulatory ligands reduces chlamydial inclusion size and delays the developmental cycle, we hypothesize that the microbe preferentially degrades its PG during development to reduce the generation of NOD2 ligands.

PMID:41416804 | DOI:10.1128/iai.00472-25

Int J Biol Macromol. 2025 Dec 18:149784. doi: 10.1016/j.ijbiomac.2025.149784. Online ahead of print.

ABSTRACT

Ruminal microbiota disturbance is an important endogenous trigger of mastitis, but the underlying causal mechanisms and key regulatory pathways remain incompletely elucidated. Pathogen-associated molecular patterns derived from the gut microbiota can circulate into distal organs outside the gastrointestinal tract and regulate disease processes. Muramyl dipeptide (MDP), a product of bacterial peptidoglycan (PGN) degradation, is derived from lytic bacteria in the gut and can be recognized by nucleotide oligomerization domain 2 (NOD2), promoting the inflammatory response. In this study, using a model of ruminal microbiota dysbiosis induced by a high-concentrate diet (HCD), we investigated the role of MDP-mediated NOD2 in the pathogenesis of ruminal microbiota dysbiosis-derived mastitis. The results showed that HCD-induced dysregulation of the ruminal ecology caused mastitis and systemic inflammation in goats. MDP expression was up-regulated in the rumen and mammary tissues, which was positively correlated with the incidence of mastitis. Transplantation of the ruminal microbiota from goats with mastitis induced mastitis symptoms and increased MDP levels in recipient mice. In addition, we found that rumen ecological dysbiosis-derived MDP mediates mammary NF-κB signaling activation and mastitis development through the activation of NOD2-RIP2 signaling. The present study elucidates the molecular mechanism of the ruminal microbiota-induced mastitis and reveals the key active molecules and potential targets of the ruminal microbiota in regulating mastitis.

PMID:41421696 | DOI:10.1016/j.ijbiomac.2025.149784

ACS Bio Med Chem Au. 2025 Sep 19;5(6):966-981. doi: 10.1021/acsbiomedchemau.5c00158. eCollection 2025 Dec 17.

ABSTRACT

MraY is an essential bacterial enzyme for peptidoglycan synthesis in cell walls and serves as a promising but unrealized target for developing effective antibacterial drugs. Nature has provided a remarkable array of nucleoside inhibitors of MraY, and researchers have skillfully refined these structures to develop inhibitors that effectively mimic natural products. Yet, both natural products and their synthetic variants often face challenges regarding inadequate in vivo efficacy, and the intricate nature of these structures complicates their synthesis and exploration of structure-activity relationships (SAR). Here, we present our findings on the discovery of first-in-class small molecule MraY inhibitors that are non-nucleoside-derived, based on 1,2,4-triazoles, using a structure-based drug design strategy. By leveraging the structural roadmap of the MraY binding site, we discovered the initial hit compound 1 with an IC₅₀ value of 171 μM in vitro against MraY from Staphylococcus aureus (MraY _SA ) that was refined to compound 12a, exhibiting an IC₅₀ value of 25 μM. Molecular docking studies against MraY _SA provided critical insights into how the binding interactions of compounds directly influence their activity. Furthermore, we report that these compounds show broad-spectrum antibacterial activity against critical pathogens such as Enterococcus spp., methicillin-resistant S. aureus (MRSA), vancomycin-resistant Enterococci (VRE) strains, Acinetobacter baumannii, and Mycobacterium tuberculosis. This study showcases novel non-nucleoside inhibitors as a compelling proof-of-concept for crafting the next generation of antibacterial agents targeting MraY.

PMID:41425802 | PMC:PMC12715639 | DOI:10.1021/acsbiomedchemau.5c00158

Nat Commun. 2025 Dec 18;16(1):11204. doi: 10.1038/s41467-025-66221-w.

ABSTRACT

Antibiotic resistance stands as a formidable global challenge to public health. Herein, we present a bacterial nanoinducer (bacNID) designed for targeted protein degradation in treating bacterial infections. Specifically, bacNID is engineered by grafting targeting peptides of MurD and SspB onto gold nanoparticles (GNPs). MurD plays a pivotal role in peptidoglycan production for cell wall synthesis, while SspB recruits SsrA-tagged proteins for degradation by ClpXP protease. The effectiveness of bacNIDs in targeted MurD degradation via ClpXP is demonstrated across both Gram-positive and Gram-negative bacterial strains. Importantly, prolonged exposure to bacNIDs does not result in the acquisition of resistance in either Staphylococcus aureus (S. aureus) or Salmonella typhimurium (S. typhimurium), even after 25 successive treatment passages. This stands in stark contrast to the rapid emergence of robust resistance observed with norfloxacin, evidenced by a 243-fold reduction in antibacterial activity against S. aureus after just 15 passages, and a 7410-fold decrease in activity against S. typhimurium over 22 passages. Moreover, the antimicrobial potential of bacNIDs is evaluated in vivo using S. aureus-infected nonhealing skin and corneal wounds. In summary, this study unveils a potent nanotechnology-driven strategy for targeted bacterial protein degradation with promising implications for in vivo antimicrobial applications.

PMID:41413196 | PMC:PMC12715223 | DOI:10.1038/s41467-025-66221-w

Nat Commun. 2025 Dec 17;16(1):11244. doi: 10.1038/s41467-025-66095-y.

ABSTRACT

The cell wall is essential for bacterial survival. Its core component is peptidoglycan (PG), a polymer comprised of disaccharide-peptides (stem peptides) that are cross-linked to one another via transpeptidation by penicillin-binding proteins (PBPs). While much is known about how PBPs are inactivated by β-lactam antibiotics, little is known about how PBPs bind and catalyze the transpeptidation of PG. Here we show how native PG and stem peptides are recruited to PBP5 of E. faecium, a critical ESKAPE pathogen. We discovered that PG binds PBP5 at the periphery of the PBP active site cleft, not the active site, and that the D-Ala leaving group contributes minimally to PBP binding. We show that β-lactam antibiotics and stem peptides can bind PBP5 simultaneously. We also show that only the single central residue of the stem peptide (L-Lys substituted by D-iAsn in E. faecium) is both necessary and sufficient for peptide recruitment. Finally, we translate our molecular findings by demonstrating that recruitment binding variants are unable to create a PG cell wall in E. faecium. Our studies define the key molecular interactions that govern bacterial cell wall formation and provide opportunities for the development of antibiotics that do not rely on PBP inactivation.

PMID:41407687 | PMC:PMC12717099 | DOI:10.1038/s41467-025-66095-y