Microbial community characterization
MP
Stool microbiome was assessed using non-targeted shotgun metagenome sequencing and taxonomic and functional gene profiling. Total DNA was extracted from stool samples (250 mg) utilizing the FastDNA bead-beating Spin Kit for Soil (MP Biomedicals, Solon, OH, USA), and verified with fluorometric quantitation (Qubit 3.0, Life Technologies, Grand Island, NY). Library preparation was performed using a Swift 2 Turbo DNA Library kit (Swift Biosciences, Ann Arbor, MI) with 50 ng of input DNA and five cycles of PCR for indexing with unique dual indices. Libraries were sequenced on an Illumina NovaSeq6000 instrument employing an SP flow cell (paired-end 2 × 150 base reads). Libraries were created in the Genomics and Microbiome Core Facility at Rush University Medical Center, and sequencing was performed at the DNA Services Lab at the University of Illinois at Urbana-Champaign. Raw FASTQ files were checked for quality using FastQC (v 0.11.9). Sequence reads were quality filtered and trimmed using the algorithm bbduk (Department of Energy Joint Genome Institute)