Urinary Microbiome Differences in Bladder Cancer, Benign Urinary Diseases and Healthy Counterparts in Adult Male Population

"Urine samples and tissue from radical cystectomy specimens will be analyzed for the assessment of the urinary microbiome.~The 16s rRNA gene will be amplified and the resulting amplicons analyzed by Illumina sequencing. To distinguish contaminants, DNA from the sample will be quantified by qPCR and compared to a known urine sample of E. coli. DNA will be extracted using the Quiagen kit and stored at -20 °C until PCR amplification. Bacterial DNA will be amplified with specific primers covering the V1-V3 hypervariable region of the 16S rRNA gene. Libraries will be prepared using the Illumina 16S microbiome sequencing protocol, and their quality will be verified by Bioanalyzer and quantity by qPCR. Sequencing will be performed in paired-end mode on Illumina sequencers. Alpha, beta diversity and species richness will be measured."