Effect of yeast extract and fumarate on bioremoval of nonyphenol ethoxylate under anaerobic condition
Soil DNA Extraction (MP Biomedicals). DNA quantification (ng/L) and purity was performed using Nanodrop 2000 Spectrophotometers (ThermoFisher Scientific) and the integrity was analyzed using 0.8% agarose gel. The massive sequencing was performed by the GenOne Biotechnologies company in Rio de Janeiro, RJ (www. genone.com.br). The bacterial 16S rRNA genes V3V4 regions were amplified from the microbial genomic DNA using specific primer (341F/806R). Polymerase Chain Reaction (PCR) was carried out with Phusion High-Fidelity PCR Master Mix (New England Biolabs). Sequencing was performed on PE250 Illumina platform and 250 bp paired-end reads were generated. Bioinformatics steps in order to evaluate and filter the sequences quality, as well as paired-end merging Operational Taxonomic Units (OTUs) clustering (?97%) were conducted using the DADA2 pipeline version 1.16 (Callahan et al., 2016) R package, as well as the ?-diversity indexes (Chao1, Observed taxa, Shannon-Wiener and Simpson). The SILVA SSU 138.1 rRNA Database Project (version March 10th, 2021) (Francioli et al., 2021) was used for taxonomic assignment. The potential genes encoding enzymes, especially regarding CH4 production, were inferred using the Tax4Fun2 version 1.1.6 (Wemheuer et al., 2020) package in R programming language, based on the Kyoto Encyclopedia of Genes and Genomes (KEGG), with 97% identity cutoff.