David Tarkhnishvili

Geographic and genetic boundaries of brown bear (Ursus arctos) population in the Caucasus

The taxonomic status of brown bears in the Caucasus remains unclear. Several morphs or subspecies have been identified from the morphological (craniological) data, but the status of each of these subspecies has never been verified by molecular genetic methods. We analysed mitochondrial DNA sequences (control region) to reveal phylogenetic relationships and infer divergence time between brown bear subpopulations in the Caucasus. We estimated migration and gene flow from both mitochondrial DNA and microsatellite allele frequencies, and identified possible barriers to gene flow among the subpopulations. Our suggestion is that all Caucasian bears belong to the nominal subspecies of Ursus arctos. Our results revealed two genetically and geographically distinct maternal haplogroups: one from the Lesser Caucasus and the other one from the Greater Caucasus. The genetic divergence between these haplogroups dates as far back as the beginning of human colonization of the Caucasus. Our analysis of the least‐cost distances between the subpopulations suggests humans as a major barrier to gene flow. The low genetic differentiation inferred from microsatellite allele frequencies indicates that gene flow between the two populations in the Caucasus is maintained through the movements of male brown bears. The Likhi Ridge that connects the Greater and Lesser Caucasus mountains is the most likely corridor for this migration.

Gene Flow between Wolf and Shepherd Dog Populations in Georgia (Caucasus)

We studied the distribution of the mitochondrial DNA haplotypes and microsatellite genotypes at 8 loci in 102 gray wolves, 57 livestock guarding dogs, and 9 mongrel dogs from Georgia (Caucasus). Most of the studied dogs had mitochondrial haplotypes clustered with presumably East Asian dog lineages, and most of the studied wolves had the haplotypes clustered with European wolves, but 20% of wolves and 37% of dogs shared the same mitochondrial haplotypes. Bayesian inference with STRUCTURE software suggested that more than 13% of the studied wolves had detectable dog ancestry and more than 10% of the dogs had detectable wolf ancestry. About 2-3% of the sampled wolves and dogs were identified, with a high probability, as first-generation hybrids. These results were supported by the relatedness analysis, which showed that 10% of wolves and 20% of dogs had closest relatives from an opposite group. The results of the study suggest that wolf-dog hybridization is a common event in the areas where large livestock guarding dogs are held in a traditional way, and that gene flow between dogs and gray wolves was an important force influencing gene pool of dogs for millennia since early domestication events. This process may have been terminated 1) in areas outside the natural range of gray wolves and 2) since very recent time, when humans started to more tightly control contacts of purebred dogs.

Modelling the range of Syrian spadefoot toad (Pelobates syriacus) with combination of GIS-based approaches

Syrian spadefoot toads (Pelobates syriacus) are found from Balkan Peninsula throughout the Caucasus, Middle East, and Northern Iran. However, the range of this species is sufficiently mapped only in Balkans. Based on the detailed mapping of the species in Georgia, we analysed the environmental factors influencing species occurrence and produced spatial models for the entire range. Nine models based on the logistic regression, discriminant function analysis, and classification tree analysis, all showed significant influence of the vegetation index and maximum summer temperature on the species distribution. The habitat suitability models based on the presence-only data (Mahalanobis distance estimation and Maximum Entropy approach) had a better performance than the models based on the presence-absence data. The habitat suitability models included two variables only, identified as significant in all preliminary analyses. The area identified as suitable by presence-only models strongly increases when a few environmental variables are included in the analysis. When only two variables are considered, the model predicted large areas north of the actual range. Competition with the related common spadefoot toad (P. fuscus) might have a role on the position of the northern limit of the range of P. syriacus. Within the predicted range, use of breeding sites is negatively associated with presence of fish. Breeding pond preferences of P. syriacus were particularly similar to those of the lake frog (Rana ridibunda), although the latter species uses appropriate habitats more extensively.

Morphometric, biochemical and molecular traits in Caucasian wood mice (Apodemus/Sylvaemus), with remarks on species divergence

We analysed Caucasian wood mice from Georgia (n = 60) and supplementary reference material of theApodemus/Sylvaemus species group to evaluate the reliability of taxon identification. Traditional “expert knowledge” plus three different methodological approaches were employed and combined to perceive their discriminatory power for a reliable taxon assignment. Graphs of principal component scores derived from the analysis of 14 skull metrics displayed taxon membership of individuals. Individual multi--locus (L = 18) electrophoretic profiles were used to re-assess specimens to a specific genepool by an assignment test based on allele frequencies indicative of populational taxon samples of the respective sampling locations. Genotyped individuals were re-allocated to those taxa, for which they yielded the highest probability score. Genetic distances among the taxa were computed and clustered in a neighbour-joining tree. PCR-fragments of 1074bp amplified from the mitochondrial cytochromeb gene were cut with 2 six- and 4 four-cutter restriction enzymes, and resulting RFLP patterns were analysed phenetically to classify the specimens according to their molecular similarity. Partial cytochromeb sequences were used to construct a phylogenetic tree by computing neighbour-joining clusters from a matrix of percent nucleotide differences. The power of the combined classification approaches and their congruence is discussed. It is concluded that the joint application of traditional, morphometric and biochemical or genetic techniques for taxon allocation of specimens of wood mice encountered problems in species delimitation. The mtDNA topology obtained was not congruent with protein polymorphism that indicated differential historical and/or recent introgression and incomplete lineage sorting in substructured populations. Cytochromeb sequence DNA data analysed were not as adequate as expected to resolve phylogenetic relationships among Caucasian and European members of theApodemus-Sylvaemus complex. Altogether, morphometric, biochemical and sequence data sets did not support the hypothesis of the evolutionary independence of European and Caucasian lineages of wood mice. Nonetheless, extended combined morphological and genetic analyses are considered necessary prerequisites to an in-depth study of the evolutionary lineages of theApodemus/Sylvaemus group. More sequence data of a variety of genes (and plenty of nuclear markers) are needed to resolve the various levels of differentiation of the extant lineages.

Systematics and evolutionary history of large endemic snails from the Caucasus (Helix buchii, and H. goderdziana) (Helicidae)

Abstract: Two species of genus Helix Linnaeus, 1758 (Mollusca: Gastropoda: Helicidae) endemic to the Caucasus region are known from Georgia and northeastern Turkey: Helix buchii Dubois de Montpereux, 1839 and the recently-described but disputed Helix goderdziana Mumladze, Tarkhnishvili and Pokryszko, 2008. The latter species is the largest land snail throughout non-tropical Eurasia. We compared shell shapes and genital morphology of the two species. We analyzed mitochondrial COI and nuclear 18S ribosomal RNA and ITS1gene fragments in 39 specimens of H. buchii and H. goderdziana from ten locations from the entire distribution range of these species, together with 13 specimens of the widespread H. lucorum Linnaeus, 1758 and H. pomatia Linnaeus, 1758. Based on shell morphology alone, most of the individuals of the two species can be discriminated using multivariate approaches. The species have different flagellum/diverticulum ratios, and the foot coloration is a fully diagnostic morphological character. Molecular genetic analysis revealed little variation in 18S+ITS1 fragment, and eleven COI haplotypes. Phylogenetic analyses support reciprocal monophyly of H. buchii and H. goderdziana. The genetic distances significantly correlate with the geographic and morphological distances; correlation of morphological distances with geography is insignificant. The basal lineages of both species are found within two distinct glacial refugia, a result which matches the separation of eastern and western evolutionary lineages of other relicts of the Western Caucasus. The present distribution of H. goderdziana coincides with the expected refugiai borders, whereas H. buchii is likely to have extended its geographical range since the last glaciation.

Human Paternal Lineages, Languages, and Environment in the Caucasus

ABSTRACT Publications that describe the composition of the human Y-DNA haplogroup in different ethnic or linguistic groups and geographic regions provide no explicit explanation of the distribution of human paternal lineages in relation to specific ecological conditions. Our research attempts to address this topic for the Caucasus, a geographic region that encompasses a relatively small area but harbors high linguistic, ethnic, and Y-DNA haplogroup diversity. We genotyped 224 men that identified themselves as ethnic Georgian for 23 Y-chromosome short tandem-repeat markers and assigned them to their geographic places of origin. The genotyped data were supplemented with published data on haplogroup composition and location of other ethnic groups of the Caucasus. We used multivariate statistical methods to see if linguistics, climate, and landscape accounted for geographical differences in frequencies of the Y-DNA haplogroups G2, R1a, R1b, J1, and J2. The analysis showed significant associations of (1) G2 with well-forested mountains, (2) J2 with warm areas or poorly forested mountains, and (3) J1 with poorly forested mountains. R1b showed no association with environment. Haplogroups J1 and R1a were significantly associated with Daghestanian and Kipchak speakers, respectively, but the other haplogroups showed no such simple associations with languages. Climate and landscape in the context of competition over productive areas among different paternal lineages, arriving in the Caucasus in different times, have played an important role in shaping the present-day spatial distribution of patrilineages in the Caucasus. This spatial pattern had formed before linguistic subdivisions were finally shaped, probably in the Neolithic to Bronze Age. Later historical turmoil had little influence on the patrilineage composition and spatial distribution. Based on our results, the scenario of postglacial expansions of humans and their languages to the Caucasus from the Middle East, western Eurasia, and the East European Plain is plausible.

Gene introgression between Gazella subgutturosa and G. marica: limitations of maternal inheritance analysis for species identification with conservation purposes

It has recently been suggested that goitered gazelles (Gazella subgutturosa and Gazella marica) have paraphyletic maternal origin, and that the mitochondrial cytochrome b gene fragment can be used for species identification prior to reintroduction of the gazelles. Although there is a large geographic area where the gazelles have intermediate morphology, previous researchers have not inferred any signs of mitochondrial haplotype introgression, and it is thought that the introgression, if it exists, is male-biased. We studied mitochondrial haplotypes of morphologically typical G. subgutturosa from two geographic locations. Goitered gazelles from eastern Turkey, morphologically identical to G. subgutturosa, had haplotypes identical to G. marica. This finding confirms ongoing maternal gene introgression from G. marica to G. subgutturosa. Our suggestion is that there is a natural gene flow between these two nominal species, and morphological characters together with recombinant genetic markers rather than mitochondrial DNA should be used to differentiate among individuals from areas close to the contact zone.

Phylogeography and morphological variation in a narrowly distributed Caucasian rock lizard, Darevskia mixta

The Caucasian rock lizard Darevskia mixta was sampled and studied from throughout its range, using mitochondrial cytochrome b sequences and scalation. The populations of the Greater and the Lesser Caucasus are reciprocally monophyletic matrilineally, and the respective lineages have been separated since the mid-Pleistocene. The lizards from the Greater Caucasus commonly have an unpaired preanal scale, whereas the lizards from the Lesser Caucasus have an additional scale behind the central temporal and subdivided interparietal scale more commonly than those from the Greater Caucasus. The Lesser Caucasus populations are further subdivided into two geographically distinct matrilineages, and are more diverse genetically and morphologically than the Greater Caucasus populations. The central part of the Lesser Caucasus is suggested to be the ancestral area for the entire D. mixta lineage. Successive Pleistocene periods of glaciation appear to be responsible both for the isolation of D. mixta from its common relatives, and for subdivision within D. mixta. Presence of D. mixta in NE Turkey is challenged.

Landscape Complexity in the Caucasus Impedes Genetic Assimilation of Human Populations More Effectively than Language or Ethnicity

ABSTRACT The analyses of 15 autosomal and 23 Y-chromosome DNA single-tandem-repeat loci in five rural populations from the Caucasus (four ethnically Georgian and one ethnically Armenian) indicated that two Georgian populations, one from the west and the other from the east of the Greater Caucasus Mountains, were both patrilineally and autosomally most differentiated from each other, and the other populations of Georgians and Armenians held an intermediate position between those two. This pattern may be due to human dispersal from two distinct glacial refugia in the last glacial period and the early Holocene, followed by less gene flow among the populations from the Greater Caucasus than among those from the rest of the Caucasus, where the populations have undergone substantial admixture in historical time. This hypothesis is supported by a strong correlation between genetic differentiation among the populations and landscape permeability to human migrations as determined by terrain ruggedness, forest cover, and snow cover. Although geographic patterns of autosomal and Y-chromosome DNA are not fully concordant, both are influenced by landscape permeability and show a similar east-west gradient. Our results suggest that this permeability was a stronger factor limiting gene flow among human populations in the Caucasus than were ethnic or linguistic boundaries.

Some water chemistry parameters of breeding habitats of the Caucasian salamander, Mertensiella caucasica in the Western Lesser Caucasus

Selection of breeding habitat plays a fundamental role in the reproductive success of urodeles and anurans. We studied the influence of water chemistry variables on the selection of a specific water resource as breeding habitat in Mertensiella caucasica. To determine the influence of water chemistry parameters on their habitat selection, we surveyed a total of 45 small river, streams and brooks in the Western Lesser Caucasus (northeastern Turkey and southwestern Georgia). The water samples taken from these localities were analyzed for 14 chemical variables and the results submitted to multiple logistic regression analysis in order to evaluate the influence of these parameters on the presence or absence of the species in the localities. Of these parameters, chloride concentration influenced the breeding habitat selection of Mertensiella caucasica significantly.