[Microsatellite loci variation and investigation of gene flow between two karyoforms of Cricetulus barabensis sensu lato (Rodentia, Cricetidae)].

We examine the diversity of six microsatellite loci and partial RAG1 exon of "barabensis" and "pseudogriseus" karyoforms in Cricetulus barabensis sensu lato species complex. A total of 435 specimens from 68 localities ranging from Altai to the Far East are investigated. The results of the population structure analysis (factor analysis and NJ tree based on Nei genetic distances) support subdivision into two well-differentiated clusters corresponding to the two karyoforms. These karyoforms are also well differentiated by the level of microsatellite variability. In several "barabensis" specimens, we found microsatellite alleles that are common in "pseudogriseus" populations but are otherwise absent in "barabensis." Most of these specimens originate from a single population in one of the zones of potential contact between karyoforms, Kharkhorin in Central Mongolia. These molecular results are consistent with previously published karyological data in suggesting that rare hybridization events between the two chromosomal races occur in nature.

[Microsatellite analysis of clonality and individual heterozygosity in natural populations of aspen Populus tremula L.: Identification of highly heterozygous clone].

Aspen Populus tremula L. (Salicaceae) is the fast-growing tree species of environmental and economic value. Aspen is capable of reproduction by both seeds and vegetative means, forming root sprouts. In an adult stand, identification of ramets of one clone among the trees of seed origin based on their morphology is difficult. A panel of 14 microsatellite loci developed for individual identification of aspen was applied for the clonal structure analysis in four natural aspen stands of the European part of Russia: Moscow and Voronezh oblasts, the Mari-El Republic, and the Republic of Tatarstan. In 52 trees from the Moscow sample, 41 multilocus genotypes were identified; in the Voronezh sample, among 30 individuals, 25 different genotypes were detected; and in the sample from Mari-El, 32 trees were represented by 13 genotypes. In the stand from Sabinsky Forestry, Tatarstan, all of the examined 29 trees were represented by a single genotype. The ancestral tree carrier of this genotype which was the most heterozygous (0.929) among all studied aspen individuals (sample mean, 0.598) obviously has spread over a large territory during several cutting and reproduction cycles, currently occupying the area of 2.2 ha. For aspen, usually suffering from Aspen trunk rot, such high viability is evidence of resistance to the main pathogens. The revealed superclone deserves further study with karyological methods and flow cytometry to determine ploidy level and analysis of the growth rate and the quality of wood for possible use in plantation forest production.

[Distribution of the genetic diversity of the Siberian stone pine, Pinus sibirica Du Tour, along the latitudinal and longitudinal profiles].

The Siberian stone pine (Pinus sibirica Du Tour) is one of the main forest-forming coniferous species of the boreal ecosystems of Western Siberia. We used the isozyme method to analyze 11 ecotypes representing the latitudinal and longitudinal profiles within the species range, including samples from the geographic boundaries of the distribution. The genetic structure of the ecotypes is described on the basis of the variability for 26 isozyme loci encoding for 16 enzyme systems. The greatest genetic diversity was observed in the taiga ecotypes in the central part of the studied area, while the ecotypes along the species range boundaries were shown to be genetically depauperized. Approximately 8.1% of the observed genetic diversity is attributed to differences between the studied ecotypes. We detected high levels of genetic diversity for the Fest_2, Pgm_1, Sod_4, and a few otherloci, as well as a correlation between allele frequencies and geographical locations of the populations. The results of multivariate analysis of allelic frequencies as well as cluster analysis allowed us to discriminate three major groups of ecotypes: north-eastern, central and south-western. In view of our results, we compare two hypotheses: one which attributes the spatial distribution of genetic variations to the selectivity for some of the polymorphic allozyme loci, and the other based in the history of the formation of the range of the Siberian stone pine.

[Genetic diversity, population structure, and differentiation of Siberian larch, Gmelin larch and Cajander larch on SSR-markers data].

The genetic diversity of SSR markers was studied in six populations of Siberian larch, two populations of Gmelin larch, and four populations of Cajander larch. Seven pairs of nuclear microsatellite loci were used for this analysis. 103 allelic variants were detected in 365 individuals of three species of larch. According to the AMOVA results, the variability proportion that characterizes the differences between three Larix species was twice as higher (15%) than the proportion that accounts for among-population differences within the species (6%). The differentiation of the populations of three species of larch based on SSR markers exceeded 12% (FsT = 0.121). A significant correlation of the genetic distances with the geographic distances between populations was found (r = 0.835, P = 0.01).

[Dynamics of allozyme heterozygosity in Siberian dwarf pine Pinus pumila (Pall.) Regel populations of the Russian Far East: comparison of embryos and maternal plants].

Siberian dwarf pine, or Japanese stone pine, Pinus pumila (Pall.) Regel is widespread in eastern Siberia and the Russian Far East; the species is bird-dispersed and has a unique spreading or shrub living form. A mixed mating system (predominant outcrossing with self-pollination and matings of close relatives) leads to the formation of partly inbred progenies in P. pumila, as in the majority of other conifers. The question arises as to whether inbred individuals persist in the reproductive part of a population, which can have negative genetic consequences. The ADH, FDH, FEST, GDH, GOT, IDH, LAP, MNR, MDH, PEPCA, 6-PGD, PGI, PGM, SKDH, and SOD isozyme systems were analyzed to study the dynamics of heterozygosity in four P. pumila natural populations from the Pacific region, optimal for the species. Samples were collected in northern Koryakia, southern Kamchatka (two samples), and the Kunashir Island (Kurils). Wright's fixation index was used to estimate the level of inbreeding in embryos from dormant seeds resulting from free pollination and in maternal plants. A substantial level of inbreeding (F(IS) = 0.124-0.342) was observed in the embryo samples but not in three out of the four adult samples. The inbreeding level at the reproductive age was high only in the sample from Koryakia, which was explained by a relatively young age of plants in the population frequently affected by fires. A general increase in heterozygosity, characteristic of other conifers as well, was attributed to elimination of inbred progenies and by balancing selection for heterozygotes, which is a key factor maintaining allozyme polymorphism in populations.

[Evaluation of the state of the gene pool of natural populations of vertebrate animals inhabiting the fragmented landscape of Moscow and Moscow district (with special reference to brown frogs)].

A system of urban ecological genetic monitoring has been developed for the first time in Russia and in the world. As a model, two species of brown frogs, Rana arvalis Nilss. and R. temporaria L. (16 populations, seven isozyme loci) were used. An evaluation of the gene pool state in urban frog populations has shown that the diversity in the Moscow frog populations was lower than that in natural populations (up to 80 and 50% in respectively R. anvalis and R. temporaria). Mean heterozygosities per locus were higher in large natural populations than in small urban isolates: in R. arvalis, these values were 0.16 and 0.06; in R. temporaria, 0.34 and 0.18, respectively. The number of polymorphic loci was also higher in natural populations than in the urban ones: 4 versus 2 in R. arvalis and 5 versus 4 in R. temporaria. Using superoxiddismutase as an example, fixation of different alleles of the same locus in different small isolates was shown. The gene pool condition of all but one urban populations of brown frog was evaluated as unsatisfactory, and that of the R. arvalis populations, as critical. These changes of the gene pool are explained mainly by gene drift accompanied by inbreeding, which was caused by human-induced fragmentation of the range and a decrease in population size of the species. The results of this study was employed in the development of the Moscow governmental program on restoration of the gene pools of vanishing animal species on specially protected natural urban territories. The series of works on long-term monitoring and assessment of the state of natural populations of model species in anthropogenic landscapes of Moscow and Moscow district has laid a foundation for a new branch of science, gene urbanology.

[Estimating the gene pool condition in natural populations of invertebrates in the fragmented landscape of Moscow and Moscow district with special reference to shrub snail Bradybaena fruticum Müll].

Using shrub snail Bradybaena fruticum Müll (20 populations) as a model, we were the first in Russia and in the world to develop a system of urban ecological genetic monitoring of the gene pool of an invertebrate species. The results of isozyme polymorphism studies in shrub snail populations showed a dramatic (up to 70%) reduction in genetic diversity in small isolates from the urbanized environment as compared to large natural populations. In urban populations, genetic diversity parameters were demonstrated to be lower than in natural ones: the mean heterozygosity per locus was reduced up to 0.08 (0.15-0.20 in natural populations); the mean allele number, to 1.9 (2.7 in natural populations); and the number of polymorphic loci, to four, i.e., 2.2-fold (nine in natural populations). In Moscow district, the number of polymorphic loci also decreased to five in the population subjected to anthropogenic pressure. The changes in the population gene pool (as shown by the number of polymorphic loci) were different in Moscow and Moscow district. The percentage of populations with the number of polymorphic loci as low as four to six was 76.9 in Moscow and 23.1% in Moscow district. The gene pool quality of 80% of the urban snail populations was estimated as unsatisfactory, and in half of them, as critical. The main reason for these changes seems to be genetic drift accompanied by inbreeding, caused by fragmentation of the range and reduction in the abundance of populations of the species, due to the anthropogenic pressure. The results of the study were employed in the program of the Moscow government for restoring the gene pools of endangered animals species on the preserved territories of the city.

[Allozyme polymorphism of Swiss stone pine Pinus cembra L. in mountainous populations of the Alps and the Eastern Carpathians].

Swiss stone pine Pinus cembra L. is a species with fragmented range, occurring in the Alpine-East Carpathian mountain system. Seeds of P. cembra are dispersed by nutcrackers, which offers potential possibilities for gene exchange among populations. Using isozyme analysis, we have examined five samples from two parts of the Swiss stone pine range: the Alps (Switzerland and Austria) and the Carpathians (two samples from the northern macroslope of the Gorgany Ridge, Eastern Carpathians, Ivano-Frankovsk oblast and one sample from Trans-Carpathian oblast of Ukraine). The allele frequencies of 30 isozyme loci, coding for enzymes ADH, FDH, FEST, GDH, GOT, IDH, LAP, MNR, MDH, PEPCA, 6PGD, PGI, PGM, SDH, SKDH, SOD, were analyzed using cluster analysis and methods of principal components. Two clusters, corresponding to the isolated Alpine and Carpathian parts of the range, were found. The main contribution to these differences were made by loci Adh-1, Adh-2, Fest-2, Lap-3, Mdh-4, and Sod-4. The interpopulation differentiation proved to be somewhat higher than that typical for pines (F(ST) = 7.4%), but within the limits characteristic for taxonomically close species. Thus, isolation of the populations did not lead to their marked differentiation, which may be explained by gene flow and balancing selection, which equalizes gene frequencies across the fragmented species area. Interlocus F(ST) heterogeneity (from 0.003 to 0.173) suggests adaptive significance of some of the allozyme polymorphisms or linkage of some loci with adaptive genes. The Carpathian populations were shown to have higher gene diversity than the Alpine ones (expected heterozygosities 0.095-0.114 and 0.060-0.080, respectively). A deficit of heterozygotes (as compared to the Hardy-Weinberg proportions), observed in the embryo sample, was probably explained by inbreeding. The reduction in the area of Carpathian pine forests in Holocene, caused by the global climatic changes and the anthropogenic impact, is hazardous for the gene pool of the species. The maintenance of genetic uniqueness of both Carpathian populations of P. cembra in general, and individual stands in particular, requires special measures for protection of Swiss stone pine in the Eastern Carpathians.

[Comparative analysis of Pleurotus ostreatus natural isolates].

A comparative analysis is performed of the polymorphism of the Pleurotus ostreatus (Fr.) Kumm naturally occurring strains isolated from the natural substrates found in two geographically remote Russian natural preserves, the Central Arboreal Biosphere Tver State Preserve (CABTSP) and the Moscow State University Zvenigorod Biological Station (ZBS, Moscow oblast), and within the city of Moscow. The results of the frequency analysis for the isozyme loci alleles and for the sexual and vegetative incompatibility groups are presented; the genetic structure and the interpopulation relations among 58 P. ostreatus dikaryotic strains are estimated. The natural samples from the Moscow and Tver oblasts are shown to have a high degree of polymorphism with a genetic differentiation of 0.743; in spite of their territorial remoteness, they are, however, actively exchanging genetic material. The natural fungal isolates form two reproductively isolated groups.

[Interspecific genetic variability of the oyster mushroom Pleurotus ostreatus as revealed by allozyme gene analysis]. / Vnutrividovoe gennoe raznoobrazie veshenki ustrichnoi, Pleurotus ostreatus, izuchennoe po sovokupnosti allozimnykh genov.

Allozyme variation in natural populations of basidiomycetes fungus Pleurotus ostreatus (88 individuals) from three regions of central Russia was studied. The species was shown to have 92.86% of polymorphic allozyme loci and expected heterozygosity He = 0.49. The mean number of alleles per locus was 3.5. The genetic differences among populations were supported by F-statistics (FST = 0.750). The low level of inbreeding (FIS = 0.018) suggests that the P. ostreatus populations are panmictic, and the main reproduction mode involves basidiospores dispersing at long distances. Using cluster analysis, geographically isolated populations and intersterile groups were differentiated within the complex P. ostreatus species.

[Use of molecular markers for differentiation of cultivated strains of oyster and button mushrooms]. / Ispol'zovanie molekuliarnykh markerov dlia differentsiatsii kul'tiviruemykh shtammov veshenki i shampin'ona.

Analysis of commercial strains of two edible mushrooms, Pleurotus ostreatus and Agaricus bisporus, using PCR and isozyme electrophoresis techniques allowed us to differentiate groups of genetically similar and distant strains. Among the commercial strains of P. ostreatus, the level of genetic variation was higher suggesting a broader genetic basis employed in breeding of this mushroom. The cultivars and hybrids of, A. bisporus, showed a higher level of homology. The isozyme markers (nonspecific esterase, leucinaminopeptidase, and phosphoglucoisomerase) are recommended for identification of the commercial strains of edible mushrooms.