Significance We demonstrate, to our knowledge for the first time, that bacterial biofilms are associated with colorectal cancers, one of the leading malignancies in the United States and abroad. Colon biofilms, dense communities of bacteria encased in a likely complex matrix that contact the colon epithelial cells, are nearly universal on right colon tumors. Most remarkably, biofilm presence correlates with bacterial tissue invasion and changes in tissue biology with enhanced cellular proliferation, a basic feature of oncogenic transformation occurring even in colons without evidence of cancer. Microbiome profiling revealed that biofilm communities on paired normal mucosa cluster with tumor microbiomes but lack distinct taxa differences. This work introduces a previously unidentified concept whereby microbial community structural organization exhibits the potential to contribute to disease progression.

Colorectal cancer (CRC) remains the third most common cancer worldwide, with a growing incidence among young adults. Multiple studies have presented associations between the gut microbiome and CRC, suggesting a link with cancer risk. Although CRC microbiome studies continue to profile larger patient cohorts with increasingly economical and rapid DNA sequencing platforms, few common associations with CRC have been identified, in part due to limitations in taxonomic resolution and differences in analysis methodologies. Complementing these taxonomic studies is the newly recognized phenomenon that bacterial organization into biofilm structures in the mucus layer of the gut is a consistent feature of right-sided (proximal), but not left-sided (distal) colorectal cancer. In the present study, we performed 16S rRNA gene amplicon sequencing and biofilm quantification in a new cohort of patients from Malaysia, followed by a meta-analysis of eleven additional publicly available data sets on stool and tissue-based CRC microbiota using Resphera Insight, a high-resolution analytical tool for species-level characterization. Results from the Malaysian cohort and the expanded meta-analysis confirm that CRC tissues are enriched for invasive biofilms (particularly on right-sided tumors), a symbiont with capacity for tumorigenesis (Bacteroides fragilis), and oral pathogens including Fusobacterium nucleatum, Parvimonas micra, and Peptostreptococcus stomatis. Considered in aggregate, species from the Human Oral Microbiome Database are highly enriched in CRC. Although no detected microbial feature was universally present, their substantial overlap and combined prevalence supports a role for the gut microbiota in a significant percentage (>80%) of CRC cases.

Summary IL-17-producing Th17 cells, generated through a STAT3-dependent mechanism, have been shown to promote carcinogenesis in many systems, including microbe-driven colon cancer. Additional sources of IL-17, such as γδ T cells, become available under inflammatory conditions, but their contributions to cancer development are unclear. In this study, we modeled Th17-driven colon tumorigenesis by colonizing MinApc+/- mice with the human gut bacterium, enterotoxigenic Bacteroides fragilis (ETBF), to investigate the link between inflammation and colorectal cancer. We found that ablating Th17 cells by knocking out Stat3 in CD4+ T cells delayed tumorigenesis, but failed to suppress the eventual formation of colonic tumors. However, IL-17 blockade significantly attenuated tumor formation, indicating a critical requirement for IL-17 in tumorigenesis, but from a source other than Th17 cells. Notably, genetic ablation of γδ T cells in ETBF-colonized Th17-deficient Min mice prevented the late emergence of colonic tumors. Taken together, these findings support a redundant role for adaptive Th17 cell- and innate γδT17 cell-derived IL-17 in bacteria-induced colon carcinogenesis, stressing the importance of therapeutically targeting the cytokine itself rather than its cellular sources.

Escherichia coli (E. coli) from the B2 phylogenetic group is implicated in colorectal cancer (CRC) as it possesses a genomic island, termed polyketide synthetase (pks), which codes for the synthesis of colibactin, a genotoxin that induces DNA damage, cell cycle arrest, mutations and chromosomal instability in eukaryotic cells. The aim of this study was to detect and compare the prevalence of E. coli expressing pks (pks + E. coli) in CRC patients and healthy controls followed by investigating the virulence triggered by pks + E. coli using an in-vitro model. Mucosal colon tissues were collected and processed to determine the presence of pks + E. coli. Thereafter, primary colon epithelial (PCE) and colorectal carcinoma (HCT116) cell lines were used to detect cytopathic response to the isolated pks + E. coli strains. Our results showed 16.7% and 4.3% of CRC and healthy controls, respectively were pks + E. coli. Further, PCE displayed syncytia and cell swelling and HCT116 cells, megalocytosis, in response to treatment with the isolated pks + E. coli strains. In conclusion, pks + E. coli was more often isolated from tissue of CRC patients compared to healthy individuals, and our in-vitro assays suggest these isolated strains may be involved in the initiation and development of CRC.