Comparison of plastid genomes and ITS of two sister species in Gentiana and a discussion on potential threats for the endangered species from hybridization.

backgroundGentiana rigescens Franchet is an endangered medicinal herb from the family Gentianaceae with medicinal values. Gentiana cephalantha Franchet is a sister species to G. rigescens possessing similar morphology and wider distribution. To explore the phylogeny of the two species and reveal potential hybridization, we adopted next-generation sequencing technology to acquire their complete chloroplast genomes from sympatric and allopatric distributions, as along with Sanger sequencing to produce the nrDNA ITS sequences.

resultsThe plastid genomes were highly similar between G. rigescens and G. cephalantha. The lengths of the genomes ranged from 146,795 to 147,001 bp in G. rigescens and from 146,856 to 147,016 bp in G. cephalantha. All genomes consisted of 116 genes, including 78 protein-coding genes, 30 tRNA genes, four rRNA genes and four pseudogenes. The total length of the ITS sequence was 626 bp, including six informative sites. Heterozygotes occurred intensively in individuals from sympatric distribution. Phylogenetic analysis was performed based on chloroplast genomes, coding sequences (CDS), hypervariable sequences (HVR), and nrDNA ITS. Analysis based on all the datasets showed that G. rigescens and G. cephalantha formed a monophyly. The two species were well separated in phylogenetic trees using ITS, except for potential hybrids, but were mixed based on plastid genomes. This study supports that G. rigescens and G. cephalantha are closely related, but independent species. However, hybridization was confirmed to occur frequently between G. rigescens and G. cephalantha in sympatric distribution owing to the lack of stable reproductive barriers. Asymmetric introgression, along with hybridization and backcrossing, may probably lead to genetic swamping and even extinction of G. rigescens.

conclusionG. rigescens and G. cephalantha are recently diverged species which might not have undergone stable post-zygotic isolation. Though plastid genome shows obvious advantage in exploring phylogenetic relationships of some complicated genera, the intrinsic phylogeny was not revealed because of matrilineal inheritance here; nuclear genomes or regions are hence crucial for uncovering the truth. As an endangered species, G. rigescens faces serious threats from both natural hybridization and human activities; therefore, a balance between conservation and utilization of the species is extremely critical in formulating conservation strategies.