16S rRNA Metagenome analysis
Raw paired-end Illumina reads were trimmed for 16S rRNA gene primer sequences using Cutadapt (https://cutadapt.readthedocs.org/en/stable/). Pre-processed sequence data were filtered using the Quantitative Insights Into Microbial Ecology (QIIME), software suite. Sequences were clustered into OTUs on the basis of similarity to known bacterial sequences available in the Greengenes database (v13.8; http://greengenes.secondgenome.com/;) (at 97% sequence similarity cut-off) using UCLUST clustering software. Sequences that could not be matched to references in the Greengenes databases were clustered de novo based on pair-wise sequence identity. Total sum scaling (TSS) normalization was applied followed by CSS to correct biases introduced by TSS, and log2 transformation to account for the non-normal distribution of taxonomic counts. Statistical analyses were executed using the Calypso software V8.7241(cgenome.net/calypso/).