gut metagenome

The TIANamp stool DNA kit was used to extract total genomic DNA from collected caecal content. The extracted DNA was then amplified using primers which target the V3-V4 hypervariable region in bacterial 16S rRNA genes. The resulting PCR products were sequenced using Illumina Miseq platform, generating 250bp paired-end reads (SinoGenoMax Co., Beijing, China). After removing chimeric sequences, the remaining sequences were analyzed using QIIME 2.0 software to obtain effective tags. Sequences with more than 97% similarity were assigned to the sample operational taxonomic unit (OTU), and a typical sequence was selected for each OUT for further analysis. Taxonomic information was obtained by querying the SILVA Database, and the complexity of species diversity was calculated using the MOTHUR 3.0 software based on alpha diversity measures such as community richness (Chao) and diversity (Shannon and Simpson). Taxonomic community structure and phylogeny were visualized to analyze the microbial diversity and abundances across different samples.