Genitourinary Microbiome and Volatilome: A Pilot Study in Patients with Prostatic Adenocarcinoma Submitted to Radical Prostatectomy

<b>Background/Objectives</b>: The genitourinary microbiome and metabolome may contribute to prostate cancer (PC) biology, but evidence remains limited. This pilot study characterizes the urinary microbiota and volatilome in men with PC and investigates microbial and viral DNA in prostate tissue, comparing findings with benign prostatic hyperplasia (BPH). <b>Methods</b>: We prospectively enrolled 21 non-metastatic PC patients undergoing radical prostatectomy and 17 BPH controls. Lesional and non-lesional prostate tissues and urine were collected from PC patients, as well as urine samples from BPH participants. DNA samples were tested for sexually transmitted pathogens by multiplex real-time PCR. Urine and prostate tissue were analyzed for human polyomaviruses (JCPyV, BKPyV, MCPyV) by qPCR, bacterial profiles via 16S rRNA gene sequencing, and urinary volatile organic metabolites (VOMs) using HS-SPME/GC-MS. Microbial and metabolic profiles were compared, and taxa-metabolites were assessed. <b>Results</b>: JCPyV and BKPyV were detected in urine and tissue from PC patients; MCPyV was detected only in tissue, at low frequency. In BPH, viral prevalence was lower and MCPyV was absent. JCPyV/BKPyV co-infection was common in cancer. No sexually transmitted pathogen emerged. PC patients showed greater urinary microbial diversity and five enriched genera, along with specific metabolic pathways. 36 urinary VOMs were identified, with 14 differing significantly, with positive correlations between PC-associated genera and metabolites. In contrast, prostate tissue was low-biomass, dominated by <i>Pseudomonas</i>, and showed no significant differences between lesional and non-lesional areas. <b>Conclusions</b>: This preliminary, hypothesis-generating study indicates that urinary, rather than tissue, microbial and volatilome signatures show clearer differences between PC and BPH. These findings suggest possible microbiota-metabolite interactions in PC but require validation in larger cohorts.