Introduction: Tumor-residing microbiota poses a new challenge in cancer progression and therapy; however, the functional behavior of patient tumor-derived microbes remains poorly understood. We previously reported the presence of tumor microbiota in intraductal papillary mucinous neoplasms (IPMNs), which are precursors of pancreatic cancer.Methods: We examined the metabolic and pathogenic potential of clinical microbiota strains obtained from IPMN tumors using various pancreatic cell lines and 3D spheroid models.Results: Our findings revealed that several strains from IPMNs with invasive cancer or high-grade dysplasia, such as E. cloacae, E. faecalis, and K. pneumoniae, induced a cancer metabolite signature in human pancreatic cells when infected ex vivo. Bacterial invasiveness was significantly correlated with DNA damage in spheroids derived from normal and tumor-derived pancreatic cells, particularly in strains derived from advanced neoplasia IPMN and under hypoxic conditions. Additionally, microbial metabolites activate human mucosal-associated invariant T (MAIT) cells and restrict the infection, both extra- and intracellularly, in hypoxic tumor conditions and in synergy with antibiotics.Discussion: Immune sensing of tumor microbiota metabolites may have clinical implications in cancer management.