Comparative Genomic Analysis of Coding Sequence-Derived Microsatellites Reveals Evolutionary Conservation and Genetic Diversity in Forest Musk Deer (Moschus berezovskii) and Related Ruminants

The FMD is an endangered species under first-class national protection in China. Comparative genomic investigation of microsatellite (SSR) in CDS may provide insights into adaptive evolutionary mechanisms and may inform conservation management strategies for captive populations. Here, we analyzed the FMD genome alongside five closely related ruminants: cattle (Bos taurus), red deer (Cervus elaphus), white-tailed deer (Odocoileus virginianus), sheep (Ovis aries), and goat (Capra hircus). Through genome-wide bioinformatic identification, we systematically compared the abundance, density, structural categories, repeat motifs, chromosomal distribution, and pathway enrichment analysis of SSR-containing genes in CDS. Furthermore, we performed synteny analysis and evaluated population genetic diversity. A total of 2509 SSRs in CDS were identified in the FMD, with a relative density of 62.61 loci/Mb. Trinucleotide SSRs were overwhelmingly dominant (88.46%) in the FMD. Notably, the FMD exhibited the highest relative abundances of both tetranucleotide and pentanucleotide repeats among the six species (2.37 and 2.18 loci/Mb, respectively), with pentanucleotide abundance approximately 5.6- to 9.1-fold higher than that of the other species. Chromosomal mapping revealed the highest SSR density in CDS regions on chromosome 27, while SSR-containing genes exhibited a heterogeneous pattern characterized by localized clustering. Synteny analysis demonstrated relatively conserved syntenic relationships between the FMD and goat, sheep, and cattle, with moderate conservation also observed with red deer and white-tailed deer, suggesting that SSR-containing genes in ruminants may remain highly conserved during chromosomal rearrangements. GO and KEGG analyses indicated that SSR-containing genes across all species were predominantly enriched in transcriptional regulation, RNA processing, and signal transduction pathways. Specifically, the FMD showed enrichment patterns associated with hypoxia response, mRNA processing, and epigenetic regulation, which may reflect lineage-specific transcriptional patterns, though the functional involvement of these SSRs remains to be experimentally validated. In addition, the five primer pairs screened in this study exhibited high polymorphism, with a mean polymorphism information content (PIC) of 0.93. The observed heterozygosity (Ho) was significantly lower than the expected heterozygosity (He), and the mean inbreeding coefficient (FIS) was 0.57, indicating heterozygote deficiency and an elevated risk of inbreeding in this captive FMD population. Collectively, our findings provide preliminary insights into the conserved patterns of microsatellite evolution and lineage-specific divergence in ruminants, offering a reference framework for comparative genomics and adaptive evolution research, as well as practical molecular markers for genetic management of captive populations.