I. V. Kartavtseva

Comparative analysis of micro and macro B chromosomes in the Korean field mouse Apodemus peninsulae (Rodentia, Murinae) performed by chromosome microdissection and FISH

Comparative analysis of micro B and macro B chromosomes of the Korean field mouse Apodemus peninsulae, collected in populations from Siberia and the Russian Far East, was performed with Giemsa, DAPI, Ag-NOR staining and chromosome painting with whole and partial chromosome probes generated by microdissection and DOP-PCR. DNA composition of micro B chromosomes was different from that of macro B chromosomes. All analyzed micro B chromosomes contained clusters of DNA repeats associated with regions characterized by an uncondensed state in mitosis. Giemsa and DAPI staining did not reveal these regions. Their presence in micro B chromosomes led to their special morphology and underestimation in size. DNA repeat clusters homologous to DNA of micro B chromosome arms were also revealed in telomeric regions of some macro B chromosomes of specimens captured in Siberian regions. Neither active NORs nor clusters of ribosomal DNA were found in the uncondensed regions of micro B chromosomes. Possible evolutionary pathways for the origin of macro and micro B chromosomes are discussed.

A spatial aspect on mitochondrial DNA genealogy in Apodemus peninsulae from East Asia.

Apodemus peninsulae is a field mouse that inhabits the broad-leafed forests of temperate Eurasia. We examined the mitochondrial cytochrome b gene in 57 individuals of A. peninsulae from northeastern Asia, including Siberia, Primorye, Magadan region, Sakhalin, Hokkaido, and the Korean Peninsula. The genealogy of the mitochondrial DNA (mtDNA) in A. peninsulae was shown to have substantial geographic affinity, suggesting geographic architecture of northeastern Asia, including the islands of Sakhalin and Hokkaido, played important roles on the cladogenesis. Taking into account the presence of region-specific anciently divergent mtDNA types, three parts of the regions of Primorye, Siberia, and the Korean Peninsula can be denoted as refugia for A. peninsulae during the substantial period of the Quaternary glacial ages. Among the geographic regions examined, Primorye is likely to be the most influential one, from which the mtDNA is thought to have migrated to the neighboring regions of Sakhalin, Hokkaido, the Magadan region, and Siberia during the evolution of this species.

Geographic Patterns of Cytochrome b and Sry Gene Lineages in the Gray Red-Backed Vole Clethrionomys rufocanus from Far East Asia Including Sakhalin and Hokkaido

Abstract The gray red-backed vole, Clethrionomys rufocanus, from Far East Asia including Sakhalin and Hokkaido is known to harbor intraspecific morphological and cytogenetic variations. Here we analyzed geographic variation in the mitochondrial cytochrome b (Cytb) gene and Y chromosome specific Sry gene by a PCR direct sequencing technique. Determination of sequences in the Cytb gene (1140 bp) in 29 individuals provided 28 haplotypes and construction of a neighbor joining tree clearly indicated that they were grouped into four major lineages, which predominated in Primorskyi territory, Kamchatska-Magadan, Sakhalin, and Hokkaido-Kuril, with 0.02–0.04 sequence divergence (Kimuras distance, all substitutions, all codon positions). The sequences for the Sry region (336–366 bp) included a variable repetitious region of microsatellites such as TG(TC)2. In total five sequences were recognized which can be grouped into two forms, continental and insular (Sakhalin and Hokkaido) based on the presence or absence of a segment of TG(TC)3TG(TC)4. It was revealed that, therefore, the population of Sakhalin possessed their own type of mitochondrial DNA but the same Sry gene as Hokkaido. A similar trend can be seen in the continental populations. Our results suggested that the population of C. rufocanus in each of the geographic domains accumulated own genetic elements in part but genetic exchanges between neighboring populations occurred during the course of evolution. It is noteworthy that the insular domains, Sakhalin and Hokkaido, have played an important role in raising the amount of genetic diversity in small rodent species.

Local differentiation of Clethrionomys rutilus in northeastern Asia inferred from mitochondrial gene sequences

Summary Mitochondrial DNA cytochrome b (Cytb) gene sequences were used to assess the evolutionary history of the northern red-backed vole, Clethrionomys rutilus , in northeastern Asia. Neighbor-joining and maximum likelihood trees constructed with the Cytb gene sequences (1140 bp) of 27 samples revealed four major local lineages; those represented by haplotypes from central Siberia, far eastern Siberia, Alaska-Kamchatskasol;Sakhalin, and Hokkaido. These lineages differed from one another with sequence divergences ranging from 0.0160 to 0.0298 (Kimuras distance, all substitutions at all codon positions). These findings imply that C. rutilus has inhabited the local areas during a longterm period of the evolutionary time, such as the last one or two million years, as observed in another common species of red-backed vole C. rufocanus with similar geographic distribution. The intraspecies geographic partition, however, differs between the species, implying that these two species have experienced different evolutionary histories in geographic expansion and genetic exchanges among local populations.

MOLECULAR PHYLOGENY OF RED-BACKED VOLES IN FAR EAST ASIA BASED ON VARIATION IN RIBOSOMAL AND MITOCHONDRIAL DNA

Phylogenetic relationships among various species of red-backed voles (Clethrionomys and Eothenomys) from areas surrounding the Sea of Japan were assessed by examining variation in the nuclear ribosomal DNA (rDNA) and mitochondrial DNA (mtDNA). The rDNA data indicated at least six phylogenetically distinct lineages represented by C. rufocanus, C. rex, C. rutilus, E. andersoni, E. smithii, and E. melanogaster. Lineages of rufocanus-rex and andersoni-smithii can be regarded as sister species, respectively. The mtDNA data generally were congruent but also suggested the splitting of two additional species lineages, C. regulus from C. rufocanus and E. imaizumii from E. andersoni-smithii. Our data revealed that these species have a complex evolutionary history, which includes interspecific gene flow.

Multiple independent evolutionary losses of XY pairing at meiosis in the grey voles

In many eutherian mammals, X–Y chromosome pairing and recombination is required for meiotic progression and correct sex chromosome disjunction. Arvicoline rodents present a notable exception to this meiotic rule, with multiple species possessing asynaptic sex chromosomes. Most asynaptic vole species belong to the genus Microtus sensu lato. However, many of the species both inside and outside the genus Microtus display normal X–Y synapsis at meiosis. These observations suggest that the synaptic condition was present in the common ancestor of all voles, but gaps in current taxonomic sampling across the arvicoline phylogeny prevent identification of the lineage(s) along which the asynaptic state arose. In this study, we use electron and immunofluorescent microscopy to assess heterogametic sex chromosome pairing in 12 additional arvicoline species. Our sample includes ten species of the tribe Microtini and two species of the tribe Lagurini. This increased breadth of sampling allowed us to identify asynaptic species in each major Microtine lineage. Evidently, the ability of the sex chromosomes to pair and recombine in male meiosis has been independently lost at least three times during the evolution of Microtine rodents. These results suggest a lack of evolutionary constraint on X–Y synapsis in Microtini, hinting at the presence of alternative molecular mechanisms for sex chromosome segregation in this large mammalian tribe.

Genetic diversity of the sable (Martes zibellina, Mustelidae) in Russian Far East and Hokkaido inferred from mitochondrial NADH dehydrogenase subunit 2 gene sequences

Abstract. Intra-species genetic variations of the sable Martes zibellina (Carnivora, Mustelidae), originating from Russian Far East and Hokkadio, were assessed by using nucleotide sequences of the mitochondrial NADH dehydrogenase subunit 2 gene (976 base pairs). Evaluation of the genetic diversity of the sables demonstrated that populations in the southern Primorsky territory in Russian Far East harbors high genetic diversity. We assumed that the high genetic variations might have been due to effects of refugia, secondary admixture of allopatrically differentiated lineages, or massive anthropogenic introductions. Molecular phylogenetic (maximum likelihood and Bayesian inference approaches) and network (median joining method) analyses clarified that sables in Hokkaido was monophyletic. Bayesian-relaxed molecular dating approach estimated the date for migration of sables into Hokkaido to lie between 0.10–0.27 Myr BP. Considering the geological evidence, the Late Pleistocene was presumed to be the plausible epoch for the establishment of the sables in Hokkaido. Lower genetic diversity of the sables in Hokkaido observed in this study was probably caused by the foundation effects or anthropogenic hunting pressures. Mammalian faunal construction in Hokkaido was also discussed.

Comparative FISH analysis of C-positive blocks of centromeric chromosomal regions of pygmy wood mice Sylvaemus uralensis (Rodentia, Muridae)

The composition and homology of centromeric heterochromatin DNA has been compared in representatives of the Asian race and two chromosomal forms (Eastern European and Southern European) of the European race of the pygmy wood mouse Sylvaemus uralensis by means of in situ hybridization with metaphase chromosomes of microdissection DNA probes obtained from centromeric C-blocks of mice of the Southern European chromosomal form and the Asian race. Joint hybridization of both DNA probes yielded all possible variants of centromeric regions in terms of the presence of repetitive sequences homologous to those of some or another dissection region, which indicates a diversity of centromeric regions differing in DNA composition. However, most variations of the fluorescent in situ hybridization (FISH) patterns are apparently related to quantitative differences of repetitive elements of the genome. Experiments with the DNA probe obtained from the genome of the Southern European form of the pygmy wood mouse have shown that the number of intense FISH signals roughly corresponds to the number of large C-segments in representatives of the European race, which is characterized by a large amount of the centromeric C-heterochromatin in the karyotype. However, intense signals have been also detected in experiments on hybridization of this probe with chromosomes of representatives of the Asian race, which has no large C-blocks in the karyotype; thus, DNA sequences homologous to heterochromatic ones are also present in nonheterochromatic regions adjacent to C-segments. Despite the variations of the numbers of both intense and weak FISH signals, all chromosomal forms/races of S. uralensis significantly differ of the samples from one another in these characters. The number of intense FISH signals in DNA in pygmy wood mice of the samples from eastern Turkmenistan (the Kugitang ridge) and southern Omsk oblast (the vicinity of the Talapker railway station) was intermediate between those in the European and Asian races, which is apparently related to a hybrid origin of these populations (the hybridization having occurred long ago in the former case and recently in the latter case).

[Comparative FISH analysis of C-positive regions of chromosomes of wood mice (Rodentia, Muridae, Sylvaemus)].

The homology of DNA of C-positive centromeric regions of chromosomes in wood mice of the genus Sylvaemus (S. uralensis, S. fulvipectus, S. sylvaticus, S. flavicollis, and S. ponticus) was estimated for the first time. DNA probes were generated by microdissection from the centromeric regions of individual autosomes of each species, and their fluorescence in situ hybridization (FISH) with metaphase chromosomes of representatives of all studied wood mouse species was carried out. Unlike in the chromosomal forms and races of S. uralensis, changes in the DNA composition of the chromosomal centromeric regions in the wood mouse species of the genus Sylvaemus (including closely related S. flavicollis and S. ponticus) are both quantitative and qualitative. The patterns of FISH signals after in situ hybridization of the microdissection DNA probes with chromosomes of the species involved in the study demonstrate significant differences between C-positive regions of wood mouse chromosomes in the copy number and the level of homology of repetitive sequences as well as in the localization of homologous repetitive sequences. It was shown that C-positive regions of wood mouse chromosomes can contain both homologous and distinct sets of repetitive sequences. Regions enriched with homologous repeats were detected either directly in C-positive regions of individual chromosomes or only on the short arms of acrocentrics, or at the boundary of C-positive and C-negative regions.

Voles (Microtus Schrank, 1798) of the Russian Far East: Allozymic and karyological divergence

The intraspecific and interspecific differentiations of voles of the Russian Far East with respect to allozymic and karyotypic characteristics have been studied. The morphologically similar but karyotypically different species Microtus oeconomus, M. fortis, M. maximowiczii, and M. sachaliensis are characterized by considerable allozymic differentiation. In these voles, the allozymic differentiation has been found to increase in the order adjacent populations-subspecies-species. The interspecific allozymic differentiation of the chromosomally polymorphic M. maximowiczii-M. evoronensis-M. mujanensis does not exceed the intraspecific differences in M. oeconomus, M. fortis, and M. maximowiczii. The results are analyzed in terms of the allopatric and stasipatric models of speciation. The taxonomic status of M. maximowiczii gromovi is considered.