Background: The gut microbiome plays a pivotal role in the progression of sepsis. However, the specific mechanism of gut microbiota and its metabolites involved in the process of sepsis remains elusive, which limits its translational application. Method: In this study, we used a combination of the microbiome and untargeted metabolomics to analyze stool samples from patients with sepsis enrolled at admission, then microbiota, metabolites, and potential signaling pathways that might play important roles in disease outcome were screened out. Finally, the above results were validated by the microbiome and transcriptomics analysis in an animal model of sepsis. Results: Patients with sepsis showed destruction of symbiotic flora and elevated abundance of Enterococcus, which were validated in animal experiments. Additionally, patients with a high burden of Bacteroides, especially B. vulgatus, had higher Acute Physiology and Chronic Health Evaluation II scores and longer stays in the intensive care unit. The intestinal transcriptome in CLP rats illustrated that Enterococcus and Bacteroides had divergent profiles of correlation with differentially expressed genes, indicating distinctly different roles for these bacteria in sepsis. Furthermore, patients with sepsis exhibited disturbances in gut amino acid metabolism compared with healthy controls; namely, tryptophan metabolism was tightly related to an altered microbiota and the severity of sepsis. Conclusion: Alterations in microbial and metabolic features in the gut corresponded with the progression of sepsis. Our findings may help to predict the clinical outcome of patients in the early stage of sepsis and provide a translational basis for exploring new therapies.

Clostridioides difficile is a pathogen contributing to increased morbidity and mortality of patients with inflammatory bowel disease (IBD). To determine how C. difficile affects the severity of colitis, we constructed a dextran sulfate solution-induced colitis model challenged with C. difficile. Without antibiotic administration, C. difficile led to transient colonization in mice with colitis, but still significantly enhanced disease severity as assessed by weight loss, histopathological damages, and inflammatory cytokine concentrations. Because this effect is independent of toxin production as shown by infection with a non-toxigenic strain, we focused on changes in the gut microbiota. The microbiota altered by C.difficile, featured with reduced proportions of g_Prevotellaceae_UCG-001 and g_Muribaculaceae, were confirmed to contribute to disease severity in colitis mice via fecal microbiota transplantations. The inflamed colon showed neutrophil accumulation by flow cytometric analysis and myeloperoxidase immunochemical staining. There was enrichment of upregulated genes in leukocyte chemotaxis or migration as shown by RNA sequencing analysis. The isolated neutrophils from C. difficile-infected mice with colitis showed a robust migratory ability and had enhanced expression of cytokines and chemokines. We observed a detrimental role of neutrophils in the progress of disease by hindering neutrophil recruitment with the CXCR2 inhibitor SB225002. Furthermore, neutrophil recruitment appeared to be regulated by interleukin (IL)-1β, as inhibition of IL-1β production by MCC950 markedly ameliorated inflammation with decreased neutrophil accumulation and neutrophil-derived chemokine expression. In conclusion, our study provides information on the complicated interaction between microbiota and immune responses in C. difficile-induced inflammation in mice with colitis. Our findings could help determine potential therapeutic targets for patients with IBD concurrent with C. difficile infection.

Inflammasome activity is important for the immune response and is instrumental in numerous clinical conditions. Here we identify a mechanism that modulates the central Caspase-1 and NLR (Nod-like receptor) adaptor protein ASC (apoptosis-associated speck-like protein containing a CARD). We show that the function of ASC in assembling the inflammasome is controlled by its modification with SUMO (small ubiquitin-like modifier) and identify that the nuclear ZBTB16 (zinc-finger and BTB domain-containing protein 16) promotes this SUMOylation. The physiological significance of this activity is demonstrated through the reduction of acute inflammatory pathogenesis caused by a constitutive hyperactive inflammasome by ablating ZBTB16 in a mouse model of Muckle-Wells syndrome. Together our findings identify an further mechanism by which ZBTB16-dependent control of ASC SUMOylation assembles the inflammasome to promote this pro-inflammatory response.

Intestinal colonization by pathogenic bacteria is a risk factor for infection, and contributes to environmental contamination and disease dissemination. Alteration of gut microbiota also plays a pivotal role in the development of disease. Although Clostridium difficile and Staphylococcus aureus are well-recognized pathogens causing nosocomial and community infections, the intestinal colonization was not fully investigated. Herein, we explored their overall carriage rates in healthy adults from the community, and characterized the gut microbiomes of C. difficile and S. aureus carriers. Fecal samples were collected from 1709 healthy volunteers from communities in Shanghai, China, and tested for the presence of C. difficile, methicillin-sensitive S. aureus (MSSA), and methicillin-resistant S. aureus (MRSA) using culture-based techniques. To explore differences in the gut microbiome, 16S rRNA gene sequencing was conducted using samples from non-carriers (CH), C. difficile carriers (CCD), MRSA carriers (CM), and MSSA carriers (CS). Overall, we detected 12 C. difficile and 60 S. aureus isolates, accounting for 0.70% and 3.51% of total isolates, respectively. Eight isolates were determined to be MRSA, accounting for 13.3% of the S. aureus population. Sequencing data revealed that the microbial diversity and richness were similar among the four groups. However, at the phylum level, carriage of C. difficile or MRSA was associated with a paucity of Bacteroidetes and an overabundance of Proteobacteria compared with non-carriers. At the genus level, the prevalence of the genera Bacteroides, Prevotella, Faecalibacterium, and Roseburia was decreased in C. difficile-positive samples compared with the controls, while the proportion of Clostridium cluster XIVa species was increased. MRSA carriers exhibited a higher proportion of the genera Parasutterella and Klebsiella, but a decreased prevalence of Bacteroides. Compared with MSSA carriers, Klebsiella was the only genus found to be significantly enriched in MRSA carriers. In healthy adults, colonization by C. difficile or S. aureus did not significantly affect gut microbiota diversity. However, the alteration of the gut microbiota composition in C. difficile carriers could indicate a predisposition to further infection. Our study provides essential data on the prevalence and effects of C. difficile and S. aureus colonization on gut microbiota composition in healthy adults.