LAUSR

Microbiota-derived metabolites that elicit DNA damage can contribute to colorectal cancer (CRC). However, the full spectrum of genotoxic chemicals produced by indigenous gut microbes remains to be defined. We established a pipeline to systematically evaluate the genotoxicity of an extensive collection of gut commensals from inflammatory bowel disease patients. We identified isolates from divergent phylogenies whose metabolites caused DNA damage and discovered a distinctive family of genotoxins—termed the indolimines—produced by the CRC-associated species Morganella morganii. A non–indolimine-producing M. morganii mutant lacked genotoxicity and failed to exacerbate colon tumorigenesis in mice. These studies reveal the existence of a previously unexplored universe of genotoxic small molecules from the microbiome that may affect host biology in homeostasis and disease. Description A new class of bacterial genotoxins Individuals with inflammatory bowel disease are at increased risk of developing colorectal cancer compared with the general population. The gut microbiome is among the many factors that can influence tumorigenesis, in part by modulating the immune system and producing microbial metabolites. Cao et al. developed a functional screen to test whether gut bacteria from patients with inflammatory bowel disease have genotoxic effects (see the Perspective by Puschhof and Sears). The authors discovered a family of DNA damage–inducing microbial metabolites called indolimines, which were produced by the Gram-negative bacteria Morganella morganii. In a mouse model of colon cancer, M. morganii exacerbated tumor burden, but a mutant form of the bacteria unable to produce indolimine did not. This diverse series of genotoxic small molecules from the human microbiome may play a role in intestinal tumorigenesis. —PNK Functional screening of commensal gut microbiota reveals small-molecule genotoxins in patients with inflammatory bowel disease. INTRODUCTION Gut microbiota can potentially contribute to the development and progression of colorectal cancer (CRC) by producing small-molecule genotoxins. For example, select commensal Escherichia coli strains produce the canonical microbiota-derived genotoxin colibactin, which engenders the formation of DNA double-strand breaks (DSBs) in intestinal epithelial cells and exacerbates CRC in mouse models. Furthermore, human CRCs harbor colibactin-associated mutational signatures, implying a direct role for microbiota-induced DNA damage in CRC. However, the impacts of microbiota-derived genotoxins beyond colibactin remain largely unexplored. RATIONALE Given the extensive diversity of small-molecule metabolites produced by bacteria, we hypothesized that additional taxa from the human gut microbiome might produce previously undiscovered small molecules that cause DNA damage in host cells. Identifying and characterizing such genotoxins and their respective biosynthetic pathways may reveal causal roles of gut microbes in shaping host biology and disease susceptibility. Thus, we designed a large-scale electrophoresis-based DNA damage screening pipeline to evaluate the genotoxicity of a collection of more than 100 gut commensals isolated from patients with inflammatory bowel disease (IBD). RESULTS We identified diverse bacteria from the human microbiota whose small-molecule metabolites caused genotoxicity in both cell-free and cell-based DNA damage assays. For example, small-molecule metabolites from gram-positive bacteria (including Clostridium perfringens and Clostridium ramosum strains) and gram-negative bacteria (including multiple Morganella morganii strains) directly damaged DNA in cell-free assays and induced the expression of the DSB marker γ-H2AX and cell-cycle arrest in epithelial cells. However, the DNA damage patterns caused by these metabolites were distinct from colibactin-induced cross-links, and these isolates lacked the biosynthetic machinery to produce colibactin or other known genotoxins. These data thus implied the existence of previously unrecognized microbiota-derived genotoxins. M. morganii is enriched in the gut microbiota of both IBD and CRC patients. Using a combination of comparative metabolomics and bioactivity-guided natural product-discovery techniques, we discovered a family of M. morganii-derived small-molecule genotoxins—termed the indolimines—that elicited DNA damage in cell-based and cell-free assays. Furthermore, we identified a previously uncharacterized bacterial decarboxylase (annotated as an aspartate aminotransferase encoded by the aat gene) that was essential for indolimine synthesis and constructed an isogenic aat mutant M. morganii that lacked genotoxicity in both cell-free and cell-based DNA damage assays. Compared with the non–indolimine-producing mutant, wild-type M. morganii caused increased intestinal permeability and induced transcriptional signatures associated with abnormal DNA replication and intestinal epithelial cell proliferation in gnotobiotic mice. Furthermore, indolimine-producing M. morganii induced increased colonic tumor burden in the context of a mock microbial community in a mouse model of CRC. CONCLUSION By leveraging function-based assessments of the microbiome, we uncovered the existence of a broader universe of microbiota-derived small-molecule genotoxins. We found that diverse bacterial strains isolated from IBD patients exhibited DNA-damaging activities and discovered a previously undescribed family of genotoxins, termed the indolimines, produced by the IBD- and CRC-associated species M. morganii. Indolimine-producing M. morganii caused increased intestinal permeability and exacerbated colon tumorigenesis in gnotobiotic mice. Overall, these studies imply an expanded role for microbiota-derived genotoxins in shaping host biology and disease susceptibility. Human gut microbes isolated from IBD patients produce small-molecule genotoxins. Diverse gut microbes isolated from patients with IBD exhibit direct genotoxicity. M. morganii produces a family of genotoxic small molecule metabolites, termed the indolimines. Indolimine-producing M. morganii induces DNA damage in intestinal epithelial cells (IECs) and increased colon tumor burdens in gnotobiotic mouse models.