As the most abundant group of mammals, rodents possess a very rich ecotype, which makes them ideal for studying the relationship between diet and host gut microecology. Zokors are specialized herbivorous rodents adapted to living underground. Unlike more generalized herbivorous rodents, they feed on the underground parts of grassland plants. There are two species of the genus Myospalax in the Eurasian steppes in China: one is Myospalax psilurus, which inhabits meadow grasslands and forest edge areas, and the other is M. aspalax, which inhabits typical grassland areas. How are the dietary choices of the two species adapted to long-term subterranean life, and what is the relationship of this diet with gut microbes? Are there unique indicator genera for their gut microbial communities? Relevant factors, such as the ability of both species to degrade cellulose, are not yet clear. In this study, we analyzed the gut bacterial communities and diet compositions of two species of zokors using 16S amplicon technology combined with macro-barcoding technology. We found that the diversity of gut microbial bacterial communities in M. psilurus was significantly higher than that in M. aspalax, and that the two species of zokors possessed different gut bacterial indicator genera. Differences in the feeding habits of the two species of zokors stem from food composition rather than diversity. Based on the results of Mantel analyses, the gut bacterial community of M. aspalax showed a significant positive correlation with the creeping-rooted type food, and there was a complementary relationship between the axis root-type-food- and the rhizome-type-food-dominated (containing bulb types and tuberous root types) food groups. Functional prediction based on KEGG found that M. psilurus possessed a stronger degradation ability in the same cellulose degradation pathway. Neutral modeling results show that the gut flora of the M. psilurus has a wider ecological niche compared to that of the M. aspalax. This provides a new perspective for understanding how rodents living underground in grassland areas respond to changes in food conditions.

Background/Objectives: Seasonal reproduction in mammals is primarily regulated by the hypothalamic–pituitary–ovarian (HPO) axis, yet its molecular mechanisms in subterranean rodents living in light-restricted environments remain poorly understood. This study aimed to characterize the transcriptional regulation of the HPO axis during seasonal estrus in the Manchurian zokor (Myospalax psilurus, M. psilurus), a fossorial rodent exhibiting distinct breeding cycles despite perpetual darkness. Methods: Hypothalamic, pituitary, and ovarian tissues were collected from female zokors during estrus and anestrus (n = 5 per group). RNA sequencing was performed, followed by de novo transcriptome assembly and bioinformatic analyses. Differentially expressed genes (DEGs) were identified using edgeR, and functional enrichment was assessed via GO and KEGG analyses. Key DEGs were validated by RT-qPCR. Results: A total of 513, 292, and 138 DEGs were identified in the hypothalamus, pituitary, and ovary, respectively. GO analysis highlighted enrichment in G-protein-coupled receptor signaling, oxidation–reduction processes, and calcium ion binding. KEGG pathway analysis revealed significant enrichment of the neuroactive ligand–receptor interaction pathway across all three tissues. Key candidate genes included Trh and Mc3r in the hypothalamus, Pitx2 and NR4A2 in the pituitary, and PTGER2 and Sphk1 in the ovary. Conclusions: This study provides the first comprehensive transcriptomic profile of the HPO axis in Manchurian zokors during seasonal estrus. The neuroactive ligand–receptor interaction pathway appears central to reproductive regulation, and several tissue-specific genes were identified as potential regulators of seasonal breeding. These findings enhance our understanding of reproductive adaptation in subterranean mammals and offer a foundation for further functional studies.