Comparative and phylogenomic analyses of 19 Petrocodon (Gesneriaceae) species from the Guizhou karst plateau, China, featuring 13 newly assembled chloroplast genomes

The “terrestrial island” effect of the karst landscape in southwest China drives plant diversification, with Petrocodon (Gesneriaceae) being a highly specialized representative. Frequent taxonomic revisions and severe morphological convergent evolution obscure its intrageneric phylogeny. Given the low resolution of traditional markers for radiated taxa and the scarcity of molecular data from the Guizhou karst plateau, we analyzed 19 Petrocodon complete chloroplast genomes (plastomes) to characterize genomic structures, resolve interspecific relationships, and identify highly variable candidate markers. We newly assembled 13 plastomes and comparatively analyzed them with six re-annotated public sequences. Ranging from 152,378 to 153,621 bp in length, all plastomes exhibited a typical quadripartite structure containing 131–132 functional genes. They displayed a significant A/T bias, with preferred codons (relative synonymous codon usage, RSCU > 1) strongly terminating in A/U. Sliding window analysis identified 12 highly variable regions as mutational hotspots based on nucleotide diversity ( Pi ), with ycf1 and ndhF - rpl32 exhibiting the highest nucleotide diversity as the most informative candidate markers. Selection pressure analysis revealed that, despite an overall background of strong purifying selection, the ndhK and atpF genes exhibited elevated evolutionary rates, indicating relaxed purifying selection. Phylogenomic analysis strongly supported the monophyly of Petrocodon , delineating three geographically associated clades (Clades I–III) within our current sampling. Notably, Clade I exhibited distinct geographical endemism. Within Clade III, topological conflicts, extremely short branch lengths, and the non-monophyly of varieties among closely related species suggested a complex history of extremely short internal branches consistent with rapid diversification. This study reveals highly conserved plastome structures among the sampled Petrocodon species. While primarily reflecting ancestral neutral mutational pressures, the pronounced A/T bias may incidentally confer metabolic advantages in nitrogen-poor habitats. Concurrently, the accelerated evolution of ndhK and atpF suggests a potential trend of photoprotective optimization against environmental extremes. Our backbone phylogeny aligns with clade distributions, which is consistent with the hypothesis that karst habitat isolation contributes to speciation among the sampled taxa. Extensive morphological homoplasy exposes traditional taxonomic limitations. Consequently, our whole-plastome data robustly support the monophyly and phylogeny of sampled Petrocodon , including former genera ( Calcareoboea , Tengia ). These resources and candidate markers aid in elucidating radiation mechanisms and conserving karst biodiversity.