16S Amplicon sequencing of tissue resident bacteria

Recently, there has been increasing awareness that human tumors contain significant amount of viable commensal microbiota. To unveil the mystery of intratumor bacteria in tumor progression, we performed the optimized 16S library preparation even though the challenges of the low biomass and potential host cell contamination. We found that breast tumor tissue had distinct microbial communities with normal breast tissue. Furthermore, we identified a significant amount of intratumor bacteria predominantly residing in the tumor cell cytosol, which faithfully recapitulates the pattern of human breast cancer microbiota. Elimination of primary intratumor bacteria did not influence tumor growth but reduced lung metastasis. To answer the question that intratumor bacteria play a role in lung metastasis, we performed 16S sequencing of primary breast tumor (BT), adjacent lung containing early micro-metastasis (MAL) and lung macro-metastasis (LM), along with normal lung and normal breast tissue. Interestingly, the BT microbiota showed a similar pattern with MAL, LM rather than normal breast and lung tissue resident bacteria. We also found the core bacteria through bioinformatics analysis. Combined culturomics, scRNA seq and mouse model, we conducted functional experiments and concluded that the intratumor bacteria, albeit at low biomass, play a vital role in tumor progression. All these findings would open a new area of intratumor bacteria in cancer biology and cancer clinical.