Donkeys’ gut microbe is critical for their health and adaptation to the environment. Little research has been conducted on the donkey gut microbiome compared with other domestic animals. The Tibetan Plateau is an extreme environment. In this study, 6 Qinghai donkeys (QH) from the Tibetan Plateau and 6 Dezhou donkeys (DZ) were investigated, and the contents of 4 parts—stomach, small intestine, cecum, and rectum—were collected. 16S rRNA sequencing and metagenomic sequencing were used to analyze the composition and diversity of gut microbial communities in donkeys. The results showed that the flora diversity and richness of the hindgut were significantly higher than those of the foregut (p < 0.01), with no sex differences, and the community structure and composition of the same or adjacent regions (stomach, small intestine, cecum, and rectum) were similar. Besides, the flora diversity and richness of QH on the Tibetan Plateau were significantly higher than those of DZ (p < 0.05). The major pathways associated with QH were signal transduction mechanisms and carbohydrate transport and metabolism, and Bacteroidales were the major contributors to these functions. Our study provides novel insights into the contribution of microbiomes to the adaptive evolution of donkeys.

Background: Gut microbiota plays a significant role in host survival, health, and diseases; however, compared to other livestock, research on the gut microbiome of donkeys is limited. Results: In this study, a total of 30 donkey samples of rectal contents from six regions, including Shigatse, Changdu, Yunnan, Xinjiang, Qinghai, and Dezhou, were collected for metagenomic sequencing. The results of the species annotation revealed that the dominant phyla were Firmicutes and Bacteroidetes, and the dominant genera were Bacteroides, unclassified_o_Clostridiales (short for Clostridiales) and unclassified_f_Lachnospiraceae (short for Lachnospiraceae). The dominant phyla, genera and key discriminators were Bacteroidetes, Clostridiales and Bacteroidetes in Tibet donkeys (Shigatse); Firmicutes, Clostridiales and Clostridiales in Tibet donkeys (Changdu); Firmicutes, Fibrobacter and Tenericutes in Qinghai donkeys; Firmicutes, Clostridiales and Negativicutes in Yunnan donkeys; Firmicutes, Fibrobacter and Fibrobacteres in Xinjiang donkeys; Firmicutes, Clostridiales and Firmicutes in Dezhou donkeys. In the functional annotation, it was mainly enriched in the glycolysis and gluconeogenesis of carbohydrate metabolism, and the abundance was the highest in Dezhou donkeys. These results combined with altitude correlation analysis demonstrated that donkeys in the Dezhou region exhibited strong glucose-conversion ability, those in the Shigatse region exhibited strong glucose metabolism and utilization ability, those in the Changdu region exhibited a strong microbial metabolic function, and those in the Xinjiang region exhibited the strongest ability to decompose cellulose and hemicellulose. Conclusion: According to published literature, this is the first study to construct a dataset with multi-regional donkey breeds. Our study revealed the differences in the composition and function of gut microbes in donkeys from different geographic regions and environmental settings and is valuable for donkey gut microbiome research.

Donkeys, with high economic value for meat, skin and milk production, are important livestock. However, the current insights into reproduction of donkeys are far from enough. To obtain a deeper understanding, the differential expression analysis and weighted gene co-expression network analysis (WGCNA) of transcriptomic data of testicular and epididymis tissues in donkeys were performed. In the result, there were 4313 differentially expressed genes (DEGs) in the two tissues, including 2047 enriched in testicular tissue and 2266 in epididymis tissue. WGCNA identified 1081 hub genes associated with testis development and 6110 genes with epididymal development. Next, the tissue-specific genes were identified with the above two methods, and the gene ontology (GO) analysis revealed that the epididymal-specific genes were associated with gonad development. On the other hand, the testis-specific genes were involved in the formation of sperm flagella, meiosis period, ciliary assembly, ciliary movement, etc. In addition, we found that eca-Mir-711 and eca-Mir-143 likely participated in regulating the development of epididymal tissue. Meanwhile, eca-Mir-429, eca-Mir-761, eca-Mir-200a, eca-Mir-191 and eca-Mir-200b potentially played an important role in regulating the development of testicular tissue. In short, these results will contribute to functional studies of the male reproductive trait in donkeys.

This paper represents the fundamental report of the survey of genome-wide changes of four Chinese indigenous donkey breeds, Dezhou (DZ), Guangling (GL), North China (NC), and Shandong Little donkey (SDL), and the findings will prove usefully for identification of biomarkers that perhaps predict or characterize the growth and coat color patterns. Three genomic regions in CYP3A12, TUBGCP5, and GSTA1 genes, were identified as putative selective sweeps in all researched donkey populations. The loci of candidate genes that may have contributed to the phenotypes in body size (ACSL4, MSI2, ADRA1B, and CDKL5) and coat color patterns (KITLG and TBX3) in donkey populations would be found in underlying strong selection signatures when compared between large and small donkey types, and between different coat colors. The results of the phylogenetic analysis, F_ST, and principal component analysis (PCA) supported that each population cannot clearly deviate from each other, showing no obvious population structure. We can conclude from the population history that the formation processes between DZS and NC, GL, and SDL are completely different. The genetic variants discovered here provide a rich resource to help identify potential genomic markers and their associated molecular mechanisms that impact economically important traits for Chinese donkey breeding programs.