Neonatal meningitis-associated Escherichia coli (NMEC) is a formidable pathogen in veterinary medicine. The emergence of atypical, multidrug-resistant (MDR) variants complicates disease control. An Escherichia coli (E. coli) strain was isolated from the brain tissue of a deceased calf with acute meningitis. Comprehensive characterizations were performed, including whole-genome sequencing (WGS), multi-locus sequence typing (MLST), antimicrobial susceptibility testing (AST), murine pathogenicity assays, and RT-qPCR evaluation of neuroinflammatory cytokines. Results: The isolate (O18ab:H14) was identified as a capsule-deficient NMEC strain belonging to phylogroup A and sequence type ST1434. WGS showed that the genome size of this strain is 5.1 Mb, containing 73 strictly defined antimicrobial resistance genes and 202 virulence factors. These may be involved in the compensatory mechanism for capsule deficiency, and further functional verification is required. Phenotypically, it exhibited a robust MDR profile. In the murine model, the strain demonstrated high lethality, and induced severe multi-organ lesions characteristic of both meningitis and systemic sepsis. While intraperitoneal injection bypasses natural colonization routes, the brain-specific bacterial persistence and neuronal pathology imply neurotropic potential. Furthermore, RT-qPCR confirmed a severe neuroinflammatory response, marked by the significant upregulation of IL-1β, IL-6, and TNF-α in the infected brains. This study characterizes a novel, highly virulent, and MDR capsule-deficient NMEC/SEPEC hybrid strain. The findings emphasize the urgent need for continuous genomic surveillance of atypical E. coli pathotypes in livestock.

This study aimed to investigate the biological characteristics of Corynebacterium hindlerae, a pathogen discovered in bovine hosts, analyze its genomic features, clarify genetic relationships and differences among strains, and provide a scientific basis for comprehensive clinical prevention and control. The environmental tolerance, biofilm formation capability, and motility of the isolated strain LSKT01 were analyzed. A total of 12 Corynebacterium strains were selected for phylogenetic analysis based on core genes, evaluation of Average Nucleotide Identity (ANI), and comparative genomic analysis covering gene families, synteny, SNPs, InDels, and structural variations (SVs). The isolate exhibited optimal growth at 37 °C, pH 5.5, and 1–2% NaCl concentration, demonstrated strong biofilm-forming ability, but showed weak or no motility. Phylogenetic analysis divided the five Corynebacterium hindlerae strains into two subgroups, with strain LSKT01 being most closely related to strain 1864 (ANI 95.80–98.70%). Comparative genomic analysis revealed highly conserved gene families among strains, while LSKT01 possessed 20 specific paralogs. Genome synteny analysis showed an average similarity >90%, and strains ISL_960a and MAL_1082b exhibited more complex structural variations. The successful isolation and purification of LSKT01 revealed its biological characteristics and genomic features, elucidating the genetic relationships and evolutionary divergence among C. hindlerae strains from diverse sources. This provides a theoretical basis for the rapid diagnosis and precise prevention and control of Corynebacterium hindlerae in Xinjiang.

Background: Enterococcus faecalis is an opportunistic pathogen that is capable of causing bacterial encephalitis under specific pathological conditions. MicroRNAs (miRNAs) are a class of small, single-stranded non-coding RNAs, typically approximately 21 nucleotides in length. As master regulators of gene expression, they orchestrate critical pathways across diverse organisms and a broad spectrum of diseases; however, their role during E. faecalis neuro-invasion remains unexplored. Methods: A lamb model of E. faecalis-induced encephalitis was established. Integrated analysis of high-throughput sequencing data identified oar-miR-29b as a key differentially expressed miRNA during infection. To first verify its association with inflammation, primary SBMECs were stimulated with lipoteichoic acid (LTA), confirming that oar-miR-29b expression was significantly upregulated under inflammatory conditions. Subsequently, independent gain- and loss-of-function experiments in SBMECs were performed, with inflammatory cytokine expression assessed by qPCR and tight-junction protein levels evaluated by Western blotting. Results: Functional studies demonstrated that oar-miR-29b acts as a pro-inflammatory mediator, significantly upregulating IL-1β, IL-6, and TNF-α while degrading tight-junction proteins (ZO-1, occludin, and claudin-5), thereby compromising endothelial barrier integrity. Mechanistically, bioinformatic prediction and dual-luciferase reporter assays confirmed C1QTNF6 as a direct target of oar-miR-29b. The oar-miR-29b/C1QTNF6 axis is thus defined as a novel regulatory pathway contributing to neuro-inflammation and blood-brain barrier disruption. Conclusions: Collectively, our findings identify the oar-miR-29b/C1QTNF6 axis as a novel pathogenic mechanism that exacerbates E. faecalis-induced neuroinflammation and blood-brain barrier disruption.

Bacterial meningitis involves complex molecular networks, including microRNA-mediated regulation of inflammatory responses; however, the specific role of Ovis aries microRNA-125b (oar-miR-125b) in this process remains poorly understood. In this study, using a lamb model of Enterococcus faecalis-induced meningitis, we observed significant downregulation of oar-miR-125b, which inversely correlated with its newly identified target, Tumor Necrosis Factor Superfamily Member 4 (TNFSF4). Dual-luciferase reporter assays confirmed that oar-miR-125b directly binds to the 3′ Untranslated Region (3′UTR) of TNFSF4 but not to the 3′UTRs of Kelch Like Family Member 31 (KLHL31) or NF-κB Inhibitor Interacting Ras Like 2 (NKIRAS2). Mechanistically, decreased oar-miR-125b expression relieves its repression of TNFSF4, leading to NF-κB pathway activation and blood–brain barrier disruption. Collectively, our results demonstrate that oar-miR-125b serves as a key anti-inflammatory regulator in bacterial meningitis by targeting TNFSF4 and constraining NF-κB signaling, highlighting its potential as a therapeutic target for attenuating neuroinflammation in meningitis.