No evidence for nuclear introgression despite complete mtDNA replacement in the Carpathian newt (Lissotriton montandoni).

Patterns of interspecific introgression may vary geographically, and the distribution of introgressed variants can yield insight into the historical dynamics of genetic interactions between hybridizing species. Urodele amphibians, often characterized by limited mobility, deep intraspecific genetic structuring and vulnerability to climatic changes, constitute suitable models for such historical inferences. Here, we combine an extensive survey of the mitochondrial (mtDNA) and nuclear (15 microsatellites) genomes in the Carpathian newt, Lissotriton montandoni (Lm) with species distribution modelling (SDM). Populations of the smooth newt, L. vulgaris (Lv) from the areas surrounding the Lm range were also sampled to test whether gene flow between these hybridizing species extends beyond the area of strict syntopy. The extent of introgression differs dramatically between the mitochondrial genome and the nuclear genome. While multiple, spatially and temporally distinct introgression events from Lv resulted in complete mtDNA replacement in Lm, there was little evidence of recent interspecific nuclear gene flow in the assayed markers. Microsatellite differentiation within Lm defines three units, probably derived from separate glacial refugia, located in the northern, eastern and southern part of the Carpathians. In situ survival and range fragmentation of Lm are supported by SDM, corroborating the role of the Carpathians as a major refugial area. Our results, in combination with previous reports of extensive introgression of the major histocompatibility complex (MHC) genes, emphasize the complexity of historical gene exchange between Lm and Lv.

Multilocus species tree analyses resolve the radiation of the widespread Bufo bufo species group (Anura, Bufonidae).

New analytical methods are improving our ability to reconstruct robust species trees from multilocus datasets, despite difficulties in phylogenetic reconstruction associated with recent, rapid divergence, incomplete lineage sorting and/or introgression. In this study, we applied these methods to resolve the radiation of toads in the Bufo bufo (Anura, Bufonidae) species group, ranging from the Iberian Peninsula and North Africa to Siberia, based on sequences from two mitochondrial and four nuclear DNA regions (3490 base pairs). We obtained a fully-resolved topology, with the recently described Bufo eichwaldi from the Talysh Mountains in south Azerbaijan and Iran as the sister taxon to a clade including: (1) north African, Iberian, and most French populations, referred herein to Bufo spinosus based on the implied inclusion of populations from its type locality and (2) a second clade, sister to B. spinosus, including two sister subclades: one with all samples of Bufo verrucosissimus from the Caucasus and another one with samples of B. bufo from northern France to Russia, including the Apennine and Balkan peninsulas and most of Anatolia. Coalescent-based estimations of time to most recent common ancestors for each species and selected subclades allowed historical reconstruction of the diversification of the species group in the context of Mediterranean paleogeography and indicated a long evolutionary history in this region. Finally, we used our data to delimit the ranges of the four species, particularly the more widespread and historically confused B. spinosus and B. bufo, and identify potential contact zones, some of which show striking parallels with other co-distributed species.

Long-term survival of a urodele amphibian despite depleted major histocompatibility complex variation.

Depletion of polymorphism at major histocompatibility complex (MHC) genes has been hypothesized to limit the ability of populations to respond to emerging pathogens, thus putting their survival at risk. As pathogens contribute substantially to the global amphibian decline, assessing patterns of MHC variation is important in devising conservation strategies. Here, we directly compare levels of MHC class II and neutral variation between multiple populations of the great crested newt (Triturus cristatus) from refugial (REF: Romania) and postglacial expansion (PGE: Germany, Poland and UK) areas. REF populations harboured high levels of adaptive variation (24 expressed alleles), exhibiting clear signatures of historical positive selection, which points to the overall importance of MHC class II variation in this species. On the other hand, PGE populations were extremely depauperate (two alleles) but nevertheless have survived for c. 10,000 years, since the postglacial expansion. Variation in putative MHC class II pseudogenes, microsatellites and allozymes also showed a significant southern richness-northern purity pattern. The populations in the postglacial expansion area thus provide the clearest example to date of the long-term survival of populations in which MHC variation, historically under positive selection, has been depleted.

Phylogeography of two European newt species--discordance between mtDNA and morphology.

The newts Triturus vulgaris and Triturus montandoni are sister species that exhibit contrasting levels of intraspecific morphological variation. Triturus vulgaris has a broad Eurasiatic distribution encompassing both formerly glaciated and unglaciated areas and shows substantial morphological differentiation in the southern part of its range, while T. montandoni, confined to the Carpathians, is morphologically uniform. We analysed sequence variation of two mtDNA fragments of the total length of c. 1850 bp in 285 individuals of both species collected from 103 localities. Phylogenetic analysis of 200 unique haplotypes defined 12 major clades, their age estimated at c. 4.5-1.0 million years (Myr). Most of the older clades were found in the southern part of the range, and also in central Europe, mainly in Romania. The distribution of mtDNA clades points to the existence of several glacial refugia, located in the Caucasus region, Anatolia, the Balkan Peninsula, Italy, and more to the north in central Europe. The concordance between mtDNA based phylogeny and the distribution of T. vulgaris subspecies was weak. Triturus montandoni haplotypes did not form a monophyletic group. Instead they were found in six clades, in five of them mixed with T. vulgaris haplotypes, most likely as a result of past or ongoing hybridization and multiple introgression of mtDNA from T. vulgaris to T. montandoni. Patterns of sequence variation within clades suggested long-term demographic stability in the southern groups, moderate and relatively old demographic growth in the populations inhabiting central Europe, and high growth in some of the groups that colonized northern parts of Europe after the last glacial maximum.

Genetic differentiation of the two subspecies of the smooth newt inhabiting Romania, Triturus vulgaris vulgaris and T.v. ampelensis (Urodela, Salamandridae) as revealed by enzyme electrophoresis.

Starch gel electrophoresis was applied to quantify genetic variation and divergence in samples from Romanian populations of the nominal form of the smooth newt Triturus vulgaris and those of the endemic Romanian subspecies T.v. ampelensis, a population from a parapatric area was additionally included. All the samples had similar levels of genetic variation measured by the mean heterozygosity, proportion of polymorphic loci, and mean number of alleles per locus. T.v. ampelensis samples were genetically clearly different from the nominal form samples, the mean genetic distance between the two subspecies was being estimated as DN = 0.114. No fixed differences in allele composition between the two subspecies were found, although some of the alleles were found either exclusively in the nominal form (Aat-1) or in T.v. ampelensis (Mpi a). Other alleles at these loci together with Mdh-1 differed markedly in frequency. The population from the parapatric area was intermediate in allelic composition, but grouped together with the T.v. ampelensis samples in a maximum likelihood tree (99.7% bootstrap support for this grouping). The data indicate that the two subspecies interbreed in a parapatric zone. The molecular clock applied to electrophoretic data indicates that these two forms split during the Pleistocene.