[Heat resistance of the skeletal muscle in western palearctic green frogs (Rana esculenta complex)].

Heat resistance of the gastrocnemius muscle was studied in five species of the Rana esculenta complex. It was similar in R. bedriagae, R. lessonae, and in the European form of R. ridibunda; while North African R. saharica demonstrated a lower heat resistance. No heterosis was expressed in R. esculenta, a clonal hybrid of R. lessonae and R. ridibunda, for the heat resistance of the muscle. Moreover, this species demonstrated low heat resistance at the highest test temperatures (42 degrees C). Comparison of diploid and triploid R. esculenta syntopically occurring in the same water bodies demonstrated no heat resistances between them, thus, suggesting that polyploidy has no effect on this parameter at least in this case.

[Variation of microsatellites BM224 and Bcal7 in populations of green toads (Bufo viridis complex) with various nuclear DNA content and ploidy].

We studied variation of microsatellites BM224 and Bcal7 in three species of the Bufo viridis diploid-polyploid complex. We found that locus Bcal7 in all examined samples was monomorphic. Three alleles of microsatellite BM224 were found. Among tetraploid toads, the western species B. oblongus had only one allele variant, whereas the eastern species B. pewzowi had two other alleles. Similar distribution of alleles was observed in triploid specimens, collected in the area borders of tetraploid and diploid species. Among samples of diploid toad B. viridis, we found all three allele variants of microsatellite BM224. Their distribution was geographically determined. A comparison of allele distribution with genome size variation in diploid toads showed very similar patterns.

Mitochondrial phylogeography of the moor frog, Rana arvalis.

The moor frog Rana arvalis is a lowland species with a broad Eurasiatic distribution, from arctic tundra through forest to the steppe zone. Its present-day range suggests that glacial refugia of this frog were located outside southern European peninsulas. We studied the species-wide phylogeographical pattern using sequence variation in a 682 base pairs fragment of mtDNA cytochrome b gene; 223 individuals from 73 localities were analysed. Two main clades, A and B, differing by c. 3.6% sequence divergence were detected. The A clade is further subdivided into two subclades, AI and AII differing by 1.0%. All three lineages are present in the Carpathian Basin (CB), whereas the rest of the species range, including huge expanses of Eurasian lowlands, are inhabited solely by the AI lineage. We infer that AII and B lineages survived several glacial cycles in the CB but did not expand, at least in the present interglacial, to the north. The geographical distribution and genealogical relationships between haplotypes from the AI lineage indicate that this group had two glacial refugia, one located in the eastern part of the CB and the other probably in southern Russia. Populations from both refugia contributed to the colonization of the western part of the range, whereas the eastern part was colonized from the eastern refugium only. The effective population size as evidenced by theta(ML) is an order of magnitude higher in the AI lineage than in the AII and B lineages. Demographic expansion was detected in all three lineages.

[Genetic variation in two forms of the common spadefoot toad Pelobates fuscus (Pelobatidae, Anura, Amphibia) distinguished by genome size]. / Geneticheskaia izmenchivost' u dvukh form obyknovennoi chesnochnitsy Pelobates fuscus (pelobatidae, anura, amphibia), razlichaiushchikhsia po razmeru genoma.

Genetic differences (presumed 23 loci) between two cryptic forms of Pelobates fuscus, differing in genome size, were examined by means of polyacrylamide-gel electrophoresis. This method allowed to discriminate between these forms. Average genetic distance (DN) between both the forms was equal to 0.311, ranging from 0.055 to 0.563. As a rule, differences within these groups were smaller (0.021-0.388). The data show obvious genetic differentiation between these two cryptic forms of P. fuscus. Differences between these forms and P. syriacus were significantly higher (in average 0.943). Genetic distances in relation to speciation and species concepts are discussed.

Karyological and flow cytometric evidence of triploid specimens in Bufo viridis (Amphibia Anura).

Karyological and flow cytometric (FCM) analyses were performed on a group of 14 green toads of the Bufo viridis species from seven Eurasian populations. Both approaches gave concordant results concerning the DNA ploidy level. All the populations examined were represented exclusively by diploid or tetraploid specimens, except one, where triploids were found. Results evidenced an interpopulation variability in DNA content against the same ploidy level, as well as an unusually high number of triploids in a particular reproductive place. The origin of polyploidy and the presence and persistence of a high number of triploids in a particular population are discussed.

[Differences in muscle thermostability between diploid and tetraploid species of green toads (Bufo viridis complex) of Middle Asia]. / Razlichiia v termorezistentnosti skeletnykh myshts mezhdu diploidnym i tetraploidnym vidami zelenykh zhab (Bufo viridis complex) Srednei Asii.

Significant differences were found between diploid (Bufo viridis) and tetraploid (B. danatensis) toad species in their skeletal muscle thermostability. The lower muscle thermostability in tetraploids may be associated with the adaptation of B. danatensis to colder conditions of arid mountain area in the Middle Asia.

Lithium secretion in kidneys of amphibians and reptiles under hydrated conditions.

Renal lithium transport was studied at different hydration levels in five species of anuran amphibians (Bufo bufo, B. danatensis, B. viridis, Rana ridibunda, and R. temporaria), two species of urodeles (Triturus vulgaris and T. cristatus) and four species of reptiles (lizards Eremias multiocellata, Lacerta vivipara, Trapelus sanguinolentus, and Teratoscincus scincus). Under dehydration conditions, Li+ was reabsorbed in the kidneys of amphibians ans reptiles, but to a lesser degree than in mammalian kidneys: the ratio of lithium clearance (CLi) to glomerular filtration rate (GFR)--fractional lithium excretion--in dehydrated animals was in the range 0.5-0.8. The transition to the hydrated state resulted in a cessation of net renal lithium reabsorption. Under condition of high hydration, all the animals studied, except for urodeles, showed net renal secretion of Li+, i.e., CLi exceeded GFR. The ratio CLi/GFR was 1.2-1.3 in hydrated anurans and 1.7-2.3 in hydrated lizards. In urodeles, this ratio was approximately unity. It is suggested that renal lithium secretion in hydrated amphibians and reptiles reflects fluid secretion in the proximal tubule, which is additional to the glomerular filtration mechanism of fluid delivery to nephron under water loading.

Allometry of base pair specific-DNA contents in Tetrapoda.

Cells of 82 species of Tetrapoda were stained with DNA base pair specific fluorochromes (Hoechst 33258 and olivomycin) and studied by means of flow cytometry. The genome size range was about 50-fold. The class Amphibia, which had the widest range of genome size variation (about 20-fold), exhibited linear allometry in their base pair specific DNA contents (bps-C-values), i.e., the more DNA they had, the lower the quotient of AT-pairs (C(AT) = 0.13 + 0.87 x C(GC), r = +0.998). Data for Mammalia, pooled with amphibians, fell on the same allometric line at the lower extreme end of genome size range, supporting the correlation. Reptilia-Aves (or Reptilia alone) pooled with Amphibia did not conform with this relationship. Reptilia-Aves form their own line (zone); pooled with Mammalia, this group showed no regularities in the relationship of their bps-DNA contents. Besides revealing the allometry of bps-C-values, these data indicate an integral genomic feature, localization within the same regression line, which Mammalia share with Amphibia but not with recent Reptilia (and Aves). These data also suggest that the relationship between DNA base frequencies and genome size is nonlinear (reciprocal); to obtain a linear relationship, the bps-C-values should be used. It is also concluded that caution is needed when DNA-content is measured for comparative purposes using a fluorescent dye which is known to be base-pair specific. DNA content values obtained with fluorochromes with different specificity may differ as much as by a factor of 1.8, the average discrepancy level is about 14%.

Two germ cell lineages with genomes of different species in one and the same animal.

Cytological evidence for two germ cell lineages, each with the DNA-content of one of the parental species only, was obtained for certain males of the hybridogenic water frog Rana esculenta (R. ridibunda x R. lessonae) by means of DNA flow cytometry. The approach was based on the 16 +/- 2% difference in the DNA-content of genomes of the parental species. Two types of germ cells were observed in six out of 39 males studied from populations where such males did exist. The term 'hybrid amphispermy' is proposed for the phenomenon. Occurrence of the 'hybrid amphispermy' suggests that the direction of elimination (i.e., which genome being eliminated) is determined in R. esculenta by the relative activity of putative elimination-inducing factor(s) in each of the parental genomes, which can alternate from cell to cell.

Genome elimination in diploid and triploid Rana esculenta males: cytological evidence from DNA flow cytometry.

Cytological aspects of hemiclonal (meroclonal) inheritance in diploid and triploid males of the hybridogenetic frog Rana esculenta (Rana ridibunda x Rana lessonae) have been studied by DNA flow cytometry. The fact that the R. ridibunda genome contains 16% more DNA than the R. lessonae genome provides the ability to discern cells containing genomes of any species from the water-frog complex under study. Data are presented showing that elimination of the R. ridibunda genome occurs in hybridogenetic males from certain populations. In triploid males, the cytogenetic mechanism of hemiclonal inheritance is simpler than in diploids: after the elimination of a genome (always the genome in the minority in the triploid set; "homogenizing elimination"), no compensatory duplication of the remaining genetic material is necessary, as it is in diploids. The process of elimination can be visualized in triploid males by using DNA flow cytometry to identify cells in the special phase of the spermatogonial cell cycle that we termed the E phase.

[RNA and protein content in cells of diploid and tetraploid toads of the Bufo viridis group]. / Soderzhanie RNK i belka v kletkakh diploidnykh i tetraploidnykkh zhab gruppy Bufo viridis.

Nucleic acids and total protein contents in hepatocytes and erythrocytes of diploid (Bufo viridis, 2n = 22) and tetraploid (B. danatensis, 2n = 44) green toads were measured cytophotometrically. The RNA (table 1) and protein (table 2) contents in the tetraploid cells are twice as much as that in diploid ones. According to these results tetraploidy in green toads may have a recent origin. The relation between structural and functional polyploidization in tetraploid amphibian species is discussed.

[Elimination of the genome of one of the parents prior to premeiotic DNA synthesis in a hybridogenic species of Rana esculenta]. / Eliminatsiia genoma odnogo iz roditelei do premeoiticheskogo sinteza DNK u gibridogennogo vida Rana esculenta.

A DNA-cytometric study was made of spermatogenesis of the hybridogenic European green frog R. esculenta, whose somatic cells have the ridibunda + lessonae genome. The DNA amount in the ridibunda genome is by 16% more than the lessonae one. The DNA content of esculenta somatic cells is exactly intermediate between those of both the parental species. On the contrary, the sperms (1c) and the primary spermatocytes (4c) of R. esculenta have the DNA content which corresponds to the size of the ridibunda genome. These findings are in a good agreement with the hypothesis of semiclonal inheritance. Furthermore, some hybridogenic males have also spermatogonia (2c) with only the ridibunda genome size, whereas the others have altogether diploid cells with the esculenta (i.e. ridibunda + lessonae) genome size. So, it can be suggested that the selective elimination of the lessonae genome and compensatory doubling of the ridibunda one may occur in spermatogonia of R. esculenta males before the premeiotic DNA synthesis. Meiosis, as it can be inferred from the DNA-cytometry data, proceeds in a usual way on the basis of the ridibunda genome.

Dysplastic lesions and abnormalities in amphibians associated with environmental conditions.

High incidence of developmental anomaly and tumor-like dysplasia of osteochondrous tissue of hind limbs has been detected in frogs and toads the larval development of which has taken place in water, contaminated with sewage. Correlation between frequency of dysplasia and the level of water oxidation has been noted. Study on histogenesis of the marked tumor-like dysplasia were performed. It is assumed that the major mechanism responsible for anomaly consists in cytotoxic effect of teratogenic agent and that the differentiation lesion of cartilaginous tissue is caused by blocking of maturation and death of chondrocytes.

[Structural differences in the collagens of Rana frog species with dissimilar temperature ecology. The contribution of carbohydrate components to collagen molecular thermostability]. / Strukturnye razlichiia kollagenov u nekotorykh vidov liagushek roda Rana s neodinakovoi temperaturnoii ékologiei. Vklad uglevodnykh komponentov v termostabil'nost' molekul kollagena.

Skin procollagens in two species groups of frogs of the genus Rana - brown frogs (4 species) and green frogs (3 species) distributed in European and Asiatic parts of the USSR have been compared. A geographic intraspecific variation in collagen characteristics (two geographic populations of the pond frog Rana nigromaculata) is revealed. Intergroup differences in the thermal denaturation of collagens established by means of methods of circular dichroism and microcalorimetry are significant. Such differences within both the groups are not so visible. With the species studied, carbohydrate components contents in procollagen vary. Interspecific differences in the procollagen thermostability correlate with the thermal ecology of frogs and seem to be associated with the availability of carbohydrates.