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The order Boletales is a group of fungi with complex life styles, which include saprophytic and ectomycorrhizal mushroom-forming fungi. In the present study, the complete mitogenomes of two saprophytic Boletales species, Coniophora olivacea, and C. puteana, were assembled and compared with mitogenomes of ectomycorrhizal Boletales. Both mitogenomes comprised circular DNA molecules with sizes of 78,350 bp and 79,655 bp, respectively. Comparative mitogenomic analysis indicated that the two saprophytic Boletales species contained more plasmid-derived (7 on average) and unknown functional genes (12 on average) than the four ectomycorrhizal Boletales species previously reported. In addition, the core protein coding genes, nad2 and rps3, were found to be subjected to positive selection pressure between some Boletales species. Frequent intron gain/loss events were detected in Boletales and Basidiomycetes, and several novel intron classes were found in two Coniophora species. A total of 33 introns were detected in C. olivacea, and most were found to have undergone contraction in the C. olivacea mitogenome. Mitochondrial genes of Coniophora species were found to have undergone large-scale gene rearrangements, and the accumulation of intra-genomic repeats in the mitogenome was considered as one of the main contributing factors. Based on combined mitochondrial gene sets, we obtained a well-supported phylogenetic tree for 76 Basidiomycetes, demonstrating the utility of mitochondrial gene analysis for inferring Basidiomycetes phylogeny. The study served as the first report on the mitogenomes of the family Coniophorineae, which will help to understand the origin and evolution patterns of Boletales species with complex lifestyles.
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