Histological Comparison of the Edible Water Frog (Pelophylax ridibundus Pallas, 1771) Gonads Before and After Reproduction

In this study, testicular and ovarian structures of economically important edible Pelophylax ridibundus (Pallas, 1771) were histologically examined before and after reproduction in male and female individuals. Fourty eight (24 ♀, 24 ♂) adult frogs were collected from Gölbaşı Lake in Hatay. The average weight and length values of female frogs collected from nature were found to be 56.61±19.59 g and 79.54±7.07 mm; while, the average weight and length values of male frogs were 36.63±12.84 g and 69.29±9.15 mm, respectively. Frogs were brought to the frog farm established in Aydıncık and placed in breeding ponds with a width of 1m2. Frogs in the ponds were brought to the laboratory of Iskenderun Technical University in different periods, before breeding (March) and after breeding (June). Then, histological samples were taken from ovary and testis. The female frogs were determined ready for reproduction. Moreover, a large number of mature oocytes in the before breeding ovaries in vitellogenic stage, while after reproduction oocytes in primary structure and oocytes which have atresia status observed. Also, increase in the thickness of the theca layer was determined. In the male frog seminiferous tubules containing a large number of spermatogonia, spermatocyte, spermatid and a small number of spermatozoons including sperm bundles and leydig cells were found before reproduction. After the reproduciton, the density of spermatogonia, spermatocyte and spermatids were decreased; while, the density of spermatozoon and sperm bundle were increased in the seminiferous tubules. This study will contribute to the determination of mating and spawning in frog breeding by revealing the histological status of the gonad structure of P. ridibundus in the breeding process.

Discovery of a Pelophylax saharicus (Anura, Ranidae) population in Southern France: a new potentially invasive species of water frogs in Europe

We report the discovery of a population of the exotic North African Water Frog Pelophylax saharicus around the Etang de Berre, on the Mediterranean coast of France, about 25 km north-west of Marseille. The animals had been originally identified as P. perezi or P. kl. grafi by a combination of acoustic and morphological characters and their true identity was not revealed until three samples from one locality were included in a large-scale genomic work dedicated to the genus Pelophylax. Mitochondrial barcoding of the samples from other areas around Etang de Berre did not detect any native P. perezi or P. kl. grafi and confirmed that P. saharicus has spread to several localities, does reproduce and has been present since 2011 at least. We suggest that dedicated field work is needed as soon as possible to (i) map the extant of P. saharicus’s distribution around the Etang de Berre, (ii) establish if populations of the native P. perezi – kl. grafi system still persist around the Etang de Berre or not and (iii) check if P. saharicus has spread to neighboring areas or not. Depending on the answers to these three questions, local conservationists will need to evaluate the feasibility and relevance of any action to control the spread of this new invasive species and attempt to eradicate it.

Genetic structure of the water frog (Pelophylax esculentus complex) populations in the south of the Central Russian Upland

BACKGROUND: The water frog (Pelophylax esculentus complex) is hybrid in composition. In view of the fact that a large number of data on the species composition of the water frog and very scarce material on the genetic structure of populations are available in the literature, we aimed to analyze the genetic structure of populations of the water frog in the southern part of the Middle Russian upland, which was one of the refugia for many species during the glacial epoch and the center of dispersion in the postglacial time, based on DNA microsatellite markers. MATERIALS AND METHODS: The study involved 36 local populations. DNA variability was analyzed by multiplex SSR-PCR. Seven loci (Res 14, Res 15, Res 17, Res 22, Rrid059A, Rrid082A, and Rrid171A) were used for amplification. Fragment analysis of PCR products was performed on an ABI PRISM 3500 automated capillary DNA sequencer (Applied Biosystems, USA). RESULTS: The total number of alleles detected ranged from 13 to 41. The effective number of alleles (Ae) averaged 4.569 0.219, the Chenon index (I) 1.567 0.04, level of expected heterozygosity (Не) 0.68 0.01. According to Wrights model, the greatest contribution to genetic variability is made by the heterogeneity of individuals within populations, some of which are of a hybrid nature (Fis = 0.281 0.069, Fit = 0.413 0.053, Fst = 0.180 0.017). The average indicator of the intensity of gene exchange between populations (Nm) was 1.212 0.142 individuals per generation. The calculation of the effective abundance using the LD method indicates a high level of viability of the studied groups of the frogs. CONCLUSION: The results demonstrated a high level of genetic diversity and viability of most of the studied groups, which, due to the intense gene exchange between them, can represent a single panmictic population. The data of the genetic analysis support the active adaptation of P. esculentus complex to living in an urbanized environment.

Recent shifts in taxonomic compositions of water frog populations (Anura: Pelophylax) inhabiting fish ponds in southern Poland

In Central Europe, water frog species coexist in assemblages consisting of Pelophylax lessonae, P. ridibundus and their hybridogenetic hybrid, P. esculentus. Population compositions are poorly understood, partly because of difficulties in distinguishing hybrids from parentals by means of morphology alone. Environmental change and human-mediated, cryptic introductions of non-native water frog species have modified local assemblages. In this contribution we examined the structure of nine water frog populations inhabiting mostly large fish ponds in the Upper Vistula river valley of southern Poland using morphology and PCR-based genotyping of single loci in the nuclear and mitochondrial genomes. We found four different water frog population compositions: two with only P. ridibundus, one composed of P. lessonae and P. esculentus, three with P. ridibundus and P. esculentus, and three containing all three water frog taxa. Historical data show that the area was previously inhabited by lessonae-esculentus populations. We propose that both ecological and genetic replacement by expanding P. ridibundus may have contributed to the decline of the former. Overall, 18% of P. ridibundus frogs contained introgressed P. lessonae mtDNA, however, the frequency of introgressants was most pronounced in populations with a high proportion of P. esculentus. Exotic water frogs were not detected in the study area. Our results bear significance for water frog population dynamics and conservation in Central Europe and highlight the importance of long-term monitoring for the detection of changes in population composition in these amphibians.

Rediscovery of the enigmatic Andean frog Telmatobius halli Noble (Anura: Telmatobiidae), re-description of the tadpole and comments on new adult’s characters, type locality and conservation status

We report the rediscovery of Telmatobius halli (Hall’s water frog), which had not been found since its description (over 80 years) since its type locality was not clearly established. “Aguas Calientes” near Ollagüe is hypothesized as the original type locality where Frank Gregory Hall collected the type material in 1935. The tadpole is re-described, and new data on the external and internal morphology of adults is provided. These new morphological data are compared with Telmatobius spp. inhabiting geographically close to T. halli in Chile and Bolivia. In addition, comments on its ecology, conservation, and taxonomic status in relation with other Telmatobius spp. inhabiting nearby areas in Ascotán and Carcote salt pans are provided. No evidence of Batrachochytrium dendrobatidis and Ranavirus infection was found in T. halli and a sympatric amphibian species. Our work supports the validity of T. halli and suggests this species should be considered as Data Deficient in the IUCN Red List assessment until taxonomic issues are resolved.

Micronuclei in germ cells of hybrid frogs from Pelophylax esculentus complex contain gradually eliminated chromosomes

In most organisms, cells typically maintain genome integrity, as radical genome reorganization leads to dramatic consequences. However, certain organisms, ranging from unicellular ciliates to vertebrates, are able to selectively eliminate specific parts of their genome during certain stages of development. Moreover, partial or complete elimination of one of the parental genomes occurs in interspecies hybrids reproducing asexually. Although several examples of this phenomenon are known, the molecular and cellular processes involved in selective elimination of genetic material remain largely undescribed for the majority of such organisms. Here, we elucidate the process of selective genome elimination in water frog hybrids from the Pelophylax esculentus complex reproducing through hybridogenesis. Specifically, in the gonads of diploid and triploid hybrids, but not those of the parental species, we revealed micronuclei in the cytoplasm of germ cells. In each micronucleus, only one centromere was detected with antibodies against kinetochore proteins, suggesting that each micronucleus comprises a single chromosome. Using 3D-FISH with species-specific centromeric probe, we determined the role of micronuclei in selective genome elimination. We found that in triploid LLR hybrids, micronuclei preferentially contain P. ridibundus chromosomes, while in diploid hybrids, micronuclei preferentially contain P. lessonae chromosomes. The number of centromere signals in the nuclei suggested that germ cells were aneuploid until they eliminate the whole chromosomal set of one of the parental species. Furthermore, in diploid hybrids, misaligned P. lessonae chromosomes were observed during the metaphase stage of germ cells division, suggesting their possible elimination due to the inability to attach to the spindle and segregate properly. Additionally, we described gonocytes with an increased number of P. ridibundus centromeres, indicating duplication of the genetic material. We conclude that selective genome elimination from germ cells of diploid and triploid hybrids occurs via the gradual elimination of individual chromosomes of one of the parental genomes, which are enclosed within micronuclei.

New records of the anomaly P syndrome in two water frog species (Pelophylax ridibundus and P. lessonae) in Russia

The “anomaly P” was described in Palearctic water frogs of the genus Pelophylax by Jean Rostand as complex morphological anomalies of water frogs, including polydactyly, brachymely, hind limb oedema, bone outgrowths, spikes, flexions and additional limbs in the inguinal region. In 2016, the anomaly P syndrome was rediscovered in central Russia, confirming the hypothesis concerning its wider distribution. Here, three new records of this syndrome in two species of western Palearctic water frog from Russia are described.