Metagenomic analysis reveals the impact of sampling methods and digestive tract segments on the composition and function of intestinal microbiota in donkeys and cows

Here, employing approximately 300 GB of metagenomic sequencing data from 91 samples representing five (donkeys) and eight (cows) digestive tract segment with swab and fecal samples of monogastric (donkeys) and polygastric (cows) animals, we assembled 4,004,115 (cow) and 2,938,653 (donkey) microbial metagenome assembled contigs, and got 9,060,744 genes with an average size of 553bp (range 102bp ~ 10,912bp). The results demonstrated significant differences in microbial composition between the two sampling methods used. The direct swab sampling method revealed a higher abundance of Bacteroidetes in both donkeys and cows, while the fecal sampling method showed a relatively higher proportion of Firmicutes. Additionally, significant variations were observed in the microbial composition between different digestive tract segments in both animals. The microbiota in the stomach differed greatly from that in other segments especially for the donkey. Besides, both dominant and endemic microorganisms showed a clear difference in the microbial community structure between donkey and cow. In the donkey, 1,942 genera were found to have high levels of Lactobacillus, Clostridium and Prevotella, while 1,744 genera were identified in the cow. Unlike the donkey, the cow had distinct dominate genera in each of its segments, including Prevotella for the stomach, Kandleria for the small intestine and Clostridium for the cecum, colon and rectum. Functional analysis of the metagenomic data revealed disparities in the functional potential of the intestinal microbiota. The direct swab sampling method was associated with higher functional diversity, particularly in carbohydrate and lipid metabolism pathways. Conversely, fecal sampling exhibited an enrichment of genes linked to amino acid biosynthesis and transport.