Shotgun metagenomic sequencing of fecal specimens from mouse model

To achieve species-level resolution, shotgun metagenomic sequencing was also performed on fecal DNA from the same participants. Briefly, DNA was fragmented to an average size of ~400 bp for paired-end library construction using the NEXTFLE Rapid DNA-Seq kit. Sequencing was carried out on the Illumina NovaSeq 6000 platform (Illumina Inc., San Diego, CA, USA). Metagenomic functional annotation was performed using HUMAnN3 with the ChocoPhlAn database for human samples, and DIAMOND (v2.0.13) against the NCBI NR protein database (version 20230830) for mouse samples.