Comparative Mitogenomics Reveals Intron Dynamics and Mitochondrial Gene Expression Shifts in Domesticated and Wild Pleurotus ostreatus

Mitochondrial genomes play a central role in fungal physiology and adaptation, yet their evolutionary dynamics during domestication remain poorly understood. Here, we performed a comparative mitogenomic and gene-expression analysis of three <i>Pleurotus ostreatus</i> dikaryotic strains differing in origin and degree of adaptation to laboratory conditions: the long-term commercial strain dkN001, the laboratory-maintained wild isolate dkF515, and the recently collected wild strain dkN009. High-throughput Illumina sequencing enabled complete assembly of circular mitochondrial genomes, revealing substantial size variation among strains, where the dkN001 strain exhibited the second smallest mitogenome reported for the genus <i>Pleurotus</i>. Comparative analyses showed >99% sequence identity between wild isolates and ~95% identity relative to the commercial strain. Variations in genome size among strains were associated with intron dynamics in the <i>cox1</i> and <i>rnl</i> genes, as well as intron loss predominantly in the commercial strain dkN001, consistent with mitochondrial genome streamlining during domestication. Expression profiling of mitochondrial protein-coding genes (PCGs) under multiple culture conditions revealed conserved transcriptional responses in dkN001 and dkF515 that contrasted sharply with those of dkN009. The differences observed, which affected components of the electron transport chain, suggested shifts in energy metabolism associated with long-term laboratory maintenance. Therefore, our results demonstrate that domestication in <i>P. ostreatus</i> involves both structural remodelling of the mitogenome and changes in regulation of mitochondrial PCGs, highlighting the importance of mitonuclear interactions in fungal adaptation to controlled environments.