Complete mitochondrial genomics reveals phylogenetic relationships and mitogenomic features in six ectomycorrhizal Russula species.

Russula (Russulaceae, Basidiomycota) is a widely distributed genus of ectomycorrhizal fungi. In this study, we assembled and annotated the complete mitochondrial genomes (mitogenomes) of six Russula species. The mitogenomes ranged from 41,017 bp (R. hookeri) to 57,170 bp (R. aff. cessans). This size variation appears to be partly influenced by intron content, as the smallest genome (R. hookeri) lacks introns in the cox1 gene. All mitogenomes contained 15 core protein-coding genes (PCGs) (atp6, atp8, atp9, cob, cox1-cox3, nad1-nad6, nad4L, and rps3), two rRNA genes, and 25-27 tRNA genes, with the largest intergenic spacer consistently located between rrnL and nad6. Codon usage showed a strong bias toward A/U-ending codons, with UUA (Leu) and AGA (Arg) exhibiting the highest relative synonymous codon usage (RSCU) values. Notably, trnM was newly annotated in the trnE-trnL region across all six species. Selective pressure analysis revealed that 14 of the 15 core PCGs were under purifying selection (Ka/Ks < 1), while rps3 showed elevated Ka/Ks values in some species, suggesting possible relaxed or positive selection. Phylogenetic analysis based on 57 mitochondrial sequences resolved the relationships among the six Russula species. This study enriches the mitogenomic resources of Russulaceae and provides a foundation for future phylogenetic and evolutionary studies of Russula.