<i>Tritrichomonas muris</i> sensitizes the intestinal epithelium to doxorubicin-induced apoptosis

Doxorubicin (DXR) is a widely used chemotherapy drug that can induce severe intestinal mucositis. Although the influence of gut bacteria on DXR-induced damage has been documented, the role of eukaryotic commensals remains unexplored. We discovered <i>Tritrichomonas muris</i> (<i>Tmu</i>) in one of our mouse colonies exhibiting abnormal tuft cell hyperplasia, prompting an investigation into its impact on DXR-induced intestinal injury. Mice from <i>Tmu</i>-colonized and <i>Tmu</i>-excluded facilities were injected with DXR. Tissue morphology and gene expression were evaluated at acute injury (6 h) and regenerative (72 h and 120 h) phases. Changes to crypt and villus morphology were more subtle than previously reported and region-specific, with significantly shorter jejunal villi in <i>Tmu</i>⁺ mice at 72 h post-DXR compared with <i>Tmu</i>^- controls. Most notably, we observed elevated rates of DXR-induced apoptosis, measured by cleaved caspase 3 (CC3) staining, in <i>Tmu</i>⁺ intestinal crypts at 6 h post-DXR. <i>Tmu</i>⁺ mice also exhibited reduced expression of active intestinal stem cell (aISC) marker <i>Lgr5</i> and facultative ISC (fISC) marker <i>Ly6</i>a at 6 h post-DXR compared with <i>Tmu</i>^- controls. <i>Tmu</i>, but not DXR, was associated with increased inflammation and expression of type 2 cytokines IL-5 and IL-13. <i>Tmu</i>⁺ mice also exhibited a decreased fecal abundance of <i>Lactobacillus</i>, which promotes gut barrier integrity, and reduced claudin expression, indicating potential barrier dysfunction that could explain the increase in DXR-induced apoptosis. These findings highlight the significant influence of commensal microbiota, particularly eukaryotic organisms like <i>Tmu</i>, on intestinal biology and response to chemotherapy, underscoring the complexity of gut microbiota interactions in drug-induced mucositis.<b>NEW & NOTEWORTHY</b> Our study found that the eukaryotic commensal <i>Tritrichomonas muris</i> (<i>Tmu</i>) significantly increases DXR-induced intestinal apoptosis in mice. <i>Tmu</i> also reduces <i>Lgr5</i> expression post-DXR injury and elevates inflammation and type 2 cytokine expression in the absence of injury. 16S sequencing identifies decreased abundance of protective <i>Lactobacillus</i> in <i>Tmu</i> colonized mice, as well as decreased expression of barrier-forming claudins, which may explain increased apoptosis. These findings emphasize the complex role of microbiota in drug-induced intestinal damage.