Vinasse fermentation/sulfidogenesis
The universal primers 515F and 806R targeting the V4 region of the 16S rRNA gene for bacteria and archaea (Caporaso et al., 2011) were used in the polymerase chain reaction (PCR). Libraries were prepared using 2 L of stock DNA, 0.2 nM of each primer, 1x of 2x PCRBio Ultra Mix (PCR Biosystems Inc., Wayne, PA, USA) and ultrapure water to complete a final volume of 25 L. Reactions were performed in a thermocycler with an initial 3 min-denaturation step at 94C, followed by 30 cycles of 94C for 30 s, 50C for 30 s, 72C for 30 s, and a final elongation at 72C for 10 min. The holding temperature was maintained at 4C. PCR products were purified with AMPure XP Beads (Beckman Coulter Inc., Brea, CA, USA). The 16S rRNA gene amplicon libraries were further sequenced using a MiSeqTM System (Illumina, Inc., San Diego, CA) owned by NGS Solues Genmicas (Piracicaba, SP, Brazil).