Genetic Diversity and Population Structure of Hainan Indigenous Pig Breeds Revealed by Microsatellite and mtDNA D-Loop Analysis

This study investigated the genetic diversity and population structure of five Hainan indigenous pig breeds (147 individuals from 7 populations representing 5 breeds: 3 Duntou pig subpopulations (DT-DZ, DT-SJ, and DT-SG) and four additional breeds (Wuzhishan, Wenchang, Lingao, and Tunchang)) to address germplasm conservation needs driven by exotic crossbreeding, African swine fever, and inadequate genetic evaluation. After strict quality screening, we used 147 qualified samples for microsatellite genotyping and 104 samples for mtDNA D-loop sequencing. The analyses integrated 17 FAO-recommended microsatellite markers and mtDNA D-loop sequencing. In total, 15 out of 17 loci exhibited high polymorphism (PIC > 0.6), with Wuzhishan pigs exhibiting the highest genetic diversity (<i>He</i> = 0.666, <i>I</i> = 1.279). Pairwise Fst values indicated significant genetic differentiation among all populations (<i>p</i> < 0.05), and AMOVA attributed 87.32% of the genetic variation to within-population differences. Three complementary clustering methods (UPGMA, PCoA, and STRUCTURE with the optimal <i>K</i> value of 2 identified via the Δ<i>K</i> algorithm) divided the populations into two clades, clearly separating the Duntou subpopulations from other breeds. mtDNA D-loop sequencing of 104 individuals yielded a 1175 bp fragment, identifying 12 haplotypes and a high haplotype diversity (<i>Hd</i> = 0.688) low nucleotide diversity (<i>π</i> = 0.00193) pattern; Lingao pigs showed no genetic variation, while Duntou and Wuzhishan pigs had the highest Hd. NJ phylogenetic analysis indicated that Hainan pigs form an independent subclade within Chinese indigenous pigs, closely related to Luchuan pigs. These findings confirm the high overall genetic diversity and distinct population-level divergence in Hainan pigs, with Duntou pigs representing a unique lineage. This work provides a scientific basis for targeted conservation strategies, including prioritizing the conservation of Duntou and Wuzhishan pigs and restoring genetic variation in Lingao pigs.