16S rRNA sequencing of bacterial microbiota of Ugandan participants in a project investigating the mediatory role of the gut microbiome in the association between Schistosoma mansoni and cardiovascular risk.
Microbial genomic DNA was extracted from approximately 150 mg of human stool using the QIAamp DNA Stool Mini Kit (Qiagen) following manufacturer protocols. DNA quantity, purity, and integrity were assessed using NanoDrop spectrophotometry and agarose gel electrophoresis, and only high-quality DNA was used. The V3V4 regions of the bacterial 16S rRNA gene were amplified using the Institute of Food Research protocol (GHFS-LH-039). Amplicons were quantified, pooled in equimolar concentrations, end-repaired, A-tailed, and ligated to Illumina adapters. Library quality was evaluated using Qubit, real-time PCR, and Bioanalyzer. Libraries were sequenced on an Illumina platform (250 bp paired-end). Reads were demultiplexed and merged using FLASH v1.2.11.