High-Resolution Core Gene-Associated Multiple Nucleotide Polymorphism (cgMNP) Markers for Strain Identification in the Wine Cap Mushroom Stropharia rugosoannulata

Stropharia rugosoannulata, an ecologically valuable and economically important edible mushroom, faces challenges in strain-level identification and breeding due to limited genomic resources and the lack of high-resolution molecular markers. In this study, we generated high-quality genomic data for 105 S. rugosoannulata strains and identified over 2.7 million SNPs, unveiling substantial genetic diversity within the species. Using core gene-associated multiple nucleotide polymorphism (cgMNP) markers, we developed an efficient and transferable framework for strain discrimination. The analysis revealed pronounced genetic differentiation among cultivars, clustering them into two distinct phylogenetic groups. Nucleotide diversity (π) across 83 core genes varied significantly, highlighting both highly conserved loci under purifying selection and highly variable loci potentially associated with adaptive evolution. Phylogenetic analysis of the most variable gene, Phosphatidate cytidylyltransferase mitochondrial, identified 865 SNPs, enabling precise differentiation of all 85 cultivars. Our findings underscore the utility of cgMNP markers in addressing challenges posed by horizontal gene transfer and phylogenetic noise, demonstrating their robustness in cross-species applications. By providing insights into genetic diversity, evolutionary dynamics, and marker utility, this study establishes a foundation for advancing breeding programs, conservation strategies, and functional genomics in S. rugosoannulata. Furthermore, the adaptability of cgMNP markers offers a universal tool for high-resolution strain identification across diverse fungal taxa, contributing to broader fungal phylogenomics and applied mycology.