design: 16S rRNA regions V4 and V5 were amplified using specific primers 515F-Y (5-GTGYCAGCMGCCGCGGTAA-3) and 926R (5-CCGYCAATTYMTTTRAGTTT-3). PCR reactions were performed using Phusion High Fidelity PCR Master Mix with temperature conditions of 98 for 1 min, 30 cycles of 10 s denaturation at 98, 30 s annealing at 50, 30 s elongation at 72, and final elongation at 72 for 5 min. The amplicons were purified using magnetic beads and mixed equimolarly. Sequencing library was generated and quality was checked using Qubit, RT-PCR and bioanalyzer and sequenced on Illumina platform NovaSeq 6000 using paired-end 250bp kit at Novogene Europe.