Contrast and Genomic Characterisation of Ancient and Recent Interspecific Introgression Between Deeply Diverged Moustache Toads (Leptobrachium).

Recent genomic analyses have provided new insights into the process of interspecific introgression and its consequences on species evolution. Most recent studies, however, focused on hybridization between recently radiated species, with few examining the genomic outcomes of ancient hybridization across deeply diverged species. Using whole genome data of moustache toads (Leptobrachium), we identified signals of three hybridization events among nine species that diverged at the Eocene. An ancient introgression from L. leishanense to the ancestral branch (C1) of L. liui introduced adaptive variants. The highly introgressed regions include genes with important functions in odorant detection and immune responses. These genes are preserved in all three descendent populations of L. liui_C1, and these regions likely have been positively selected over a long filtering process. A recent introgression occurred from L. huashen to L. tengchongense, with the introgressed regions being mostly neutral. Furthermore, one F1 hybrid individual was detected between sympatric L. ailaonicum and L. promustache. The signals of introgression largely disappeared after removing the hybrid individual, indicating an occasional hybridization but minimal introgression. Further examination of highly divergent but low introgressed genomic regions revealed both pre-mating isolation and genetic incompatibility as potential mechanisms of resisting introgression and maintaining species boundaries. Additionally, no large X-effect was found in these introgression events. Hybridization between deeply diverged amphibian species may be common, but detectable introgressions are likely less so, with recent introgression being mostly neutral and the rare ancient one potentially adaptive. Our findings complement recent genomic work, and together they provide a better understanding of the genomic characteristics of interspecific introgression and its significance in species adaptation and evolution.

Whole-genome resequencing reveals complex effects of geographical-palaeoclimatic interactions on diversification of moustache toads in East Asia.

Geographical features and palaeoclimatic fluctuations are two classical evolutionary forces that shape genetic diversification within species. Fine-grained analysis of the mechanisms involved through population demographic processes, however, remains limited. Taking advantage of two recently published reference genomes, we resequenced the genomes and examined the evolutionary history of the moustache toads, a group endemic to East Asia where complex topography and fluctuating palaeoclimate are known to have had profound impacts on organisms. Moustache toads probably originated in southeast Yunnan, China, and diversified towards the northwestern of Yunnan, as well as central and eastern China. Further exploration based on three widespread species (Leptobrachium ailaonicum, L. boringii and L. liui) using demographic modelling and species distribution models revealed that mountains and river valleys in East Asia not only functioned as geographical barriers, but also provided dispersal corridors and facilitated continuous migration or post-glacial secondary contact among moustache toad populations. Furthermore, periodic oscillation of effective population sizes accompanying fluctuations of historical temperature and population contraction at the Last Glacial Maximum support the widespread impact of climatic changes of the Pleistocene on species diversification in East Asia. This impact was moderate for populations of L. ailaonicum and L. boringii in the southwestern mountains but severe for populations of L. liui in the eastern lowland regions of continental East Asia, which is supported by different degrees of change of their effective population sizes. Our findings reveal mechanisms underlying genetic diversification among moustache toads, and highlight the power of genomic data and demographic modelling for examining complex historical population-level processes and for understanding how geographical and palaeoclimatic factors interactively shape current intraspecific diversity.

Genetic Diversity and Population Dynamics of Leptobrachium leishanense (Anura: Megophryidae) as Determined by Tetranucleotide Microsatellite Markers Developed from Its Genome.

Persisting declination of amphibians around the world has resulted in the public attaching importance to the conservation of their biodiversity. Genetic data can be greatly helpful in conservation planning and management, especially in species that are small in size and hard to observe. It is essential to perform genetic assessments for the conservation of Leptobrachium leishanense, an endangered toad and receiving secondary protection on the list of state-protected wildlife in China. However, current molecular markers with low reliability and efficiency hinder studies. Here, we sampled 120 adult toes from the population in the Leishan Mountain, 23 of which were used to develop tetranucleotide microsatellite markers based on one reference L. leishanense genome. After primer optimization, stability detection, and polymorphism detection, we obtained 12 satisfactory microsatellite loci. Then, we used these loci to evaluate the genetic diversity and population dynamics of the 120 individuals. Our results show that there is a low degree of inbreeding in the population, and it has a high genetic diversity. Recently, the population has not experienced population bottlenecks, and the estimated effective population size was 424.3. Accordingly, stabilizing genetic diversity will be key to population sustainability. Recovering its habitat and avoiding intentional human use will be useful for conservation of this species.