Fejervarya Cancrivora Research Articles

Enhanced salt tolerance of euryhaline tadpoles depends on increased Na+, K+-ATPase expression after salinity acclimation

Understanding physiological responses and osmoregulatory mechanisms for dealing with salinity stress is essential to clarify how amphibians living in coastal areas adapt to fluctuating salinity levels. Euryhaline species are rare among reported tadpole species inhabiting saline habitats, and few studies addressed the osmoregulatory mechanisms. We quantified the effects of salinity acclimation on survival, osmolality, water content, ion concentration, and gill Na+, K+-ATPase (NKA) expression of euryhaline tadpoles of Fejervarya cancrivora, to examine time-course changes of osmoregulatory responses of tadpoles subjected to salinity stress and how osmoregulatory mechanisms were involved in the process. Acclimation to 10 ppt for 24 h increased tadpole survival of F. cancrivora in 15 ppt, and it activated osmoregulatory mechanisms such as increase in NKA expression, which enabled them to maintain a stable osmolality below that of the surrounding media, to reach lower sodium and chloride concentrations of body fluid, and to modulate dehydration at higher salinities. The minimum required acclimation period is shorter than that reported previously on this species and non-euryhaline tadpoles. This study highlights that these physiological mechanisms are ecologically relevant and critical for tadpoles living in coastal brackish waters, improving their survival in coastal microhabitats with highly variable salinity levels.

Habitat preference based on herpetofauna spatial distribution in Coal Mining Area of PT Singlurus Pratama, East Kalimantan

Herpetofauna communities are formed due to the suitability of the habitat or adaptation process that forces a species to survive. The opening of the land mines result in missing or changing the habitat of herpetofauna. This research was conducted to know the preferences of the herpetofauna of habitat based on spatial distribution of herpetofauna in the vicinity of mining area of PT Singlurus Pratama. The survey was conducted in the area of revegetasi, forest fragmentation, the district adjacent to the mining area, rivers and water spots or natural and man-made on two mining blocks i.e. block Pearl and block independence. The method of election of purposive sampling location with direct search survey methods (Visual Encounter Surveys). The main species found 4 that its spread most widely with populations high. The most frequent species of herpetofauna and many found among others: Eutropis multifasciata, Enhydris enhydris, Fejervarya cancrivora, Polypedates leucomystax and Duttaphrynus melanostictus. Based on the value of the index indicates that the diversity of herpetofauna communities distress with low. Types of herpetofauna more land cover found in the undergrowth, secondary forest and agricultural areas that there are sources of water in it.

Full compensation of growth in salt‐tolerant tadpoles after release from salinity stress

AbstractThe benefit of compensatory growth is to minimize the negative effects of stress experienced by organisms early in development once the conditions improve. Tadpoles of crab‐eating frog Fejervarya cancrivora are euryhaline, have the highest salt tolerance among reported anuran species and may exhibit a different pattern of compensatory growth after stress release. We examined how F. cancrivora tadpoles responded and possibly recovered from salinity stresses. Specifically, we tested whether tadpoles expressed compensatory growth, what degree of compensation they expressed after release from salinity stress and whether the stage at and duration during which they experienced salt stress influenced the size at and time to metamorphosis of tadpoles. We found that the effects of salinity stress on the growth and development of tadpoles are quite different between early and late exposure to high salinity (LHH vs. LLH). Exposure to high salinity late in development (LLH) reduced growth but not the rate of development, resulting in smaller metamorphs but a similar time to metamorphosis when compared to the control treatment (LLL). By contrast, exposure to high salinity early in development (LHH) retarded rates of growth and development of tadpoles, but the resulting prolonged larval period enabled tadpoles to catch up and reach similar metamorphic sizes to LLL treatment metamorphs. Most importantly, tadpoles released from salinity stress (LHL) exhibited similar time to and size at metamorphosis when compared to the LLL tadpoles, suggesting that they can exhibit full compensation of growth. Compensatory growth may be a useful mechanism for F. cancrivora tadpoles inhabiting brackish habitats, allowing them to overcome the effects of dramatically increased salt levels by expressing catch‐up growth when the salt stress ceases.

Amplification and Analysis of Cytocrome Oxidase I of <i>Polypedates leucomystax</i> from Bogor Agricultural University Area

DNA barcoding has become a useful tool for identifying and confirming of species within a known taxonomic framework. A large-scale effort is underway to barcode all amphibian species using the universally sequenced DNA region, a partial fragment of mitochondrial cytochrome oxidase subunit I (COI). This study was aimed to use DNA barcoding technique to identify and confirm species of Polypedates leucomystax and to analyze their phylogenetic relationship. Samples of Polypedates leucomystax were collected from Campus Area of Bogor Agricultural University. The cytochrome oxidase I gene of 600-700 nucleotides were amplified and observed in agarose gel electrophoresis. Forward sequence (604 base pairs) of COI gene was used for phylogenetic analyses. BLAST analysis against BOLD System database showed 95.75% identity with sequences of Polypedates leucomystax. The pairwise genetic distances of Polypedates leucomystax with Rhacophorus schlegelii, Limnonectes fujianensis, Fejervarya cancrivora, and Bufo melanostictus were 0.274, 0.352, 0.339, 0.339, 0.393, respectively. These results illustrated that the genetic identification is congruence with the morphological identification. Phylogenetic tree analysis showed that the samples were in one clade with other tree frogs. The DNA barcoding technique based on the sequence of COI gene can therefore be used to identify and confirm species of Polypedates leucomystax.

The dominant species of herpetofauna in the coal mining area at East Kalimantan, Indonesia

Muslim T, Sari UK, Yassir I. 2017. The dominant species of herpetofauna in the coal mining area at East Kalimantan, Indonesia. Biodiversitas 18: 773-779. Changing of habitat condition influences the variety of species including Herpetofauna. Herpetofauna is one of the components in the ecosystem that plays an important role, both from the ecological and economical aspect. From the ecological aspect, it has a specific role in the food chain, and it can also be used as a bio-indicator of the environment’s condition. Furthermore, herpetofauna found in reclamation area of coal mining not only stabilizes the ecosystem in the food chain, but also it can be used as a bioindicator of environment changes and the success rate of reclamation. The aim of this research was to find out the community of herpetofauna comprising and dominating the coal mining reclamation area, especially the wet and dry areas. The research location was located at PT. Singlurus Pratama Kalimantan Timur. The methods used in this research were visual encounter survey and patch sampling. This research found 10 species from 5 families of reptiles and 11 species from 6 families of amphibians. The highest dominant index score for reptile was 0.71, found in the fragmented forests. Meanwhile, the dominant reptile species were Eutropis multifasciata (20 individuals), Enhydris enhydris (4 individuals), and Dendrelaphis pictus (4 individuals). The highest dominant index score for amphibian was 1, found in the settling pond. Moreover, the dominant amphibian species were Polypedates leucomystax (42 individuals) and Fejervarya cancrivora (41 individuals). These two species were the ones able to survive in the degraded areas and they were easily found in the areas with disrupted habitats.

Herpetofauna community establishment on the micro habitat as a result of land mines fragmentation in East Kalimantan, Indonesia

Muslim T. 2017. Herpetofauna community establishment on the microhabitat as a result of land mines fragmentation in East Kalimantan, Indonesia. Biodiversitas 18: 709-714. Herpetofauna community is established because of habitat suitability or adaptation process that forces a species to survive. In the mining region, herpetofauna habitat change has occurred. The loss of natural habitat and the creation of an artificial habitat result in severance of corridor as a connecting line between herpetofauna habitats so that only microhabitat remains. Microhabitats of herpetofauna survive on limited food resources. The process of surviving of herpetofauna in microhabitats is reflected on the phenomenon of the food chain i.e. reptiles act as predators (top) and amphibians are as preys and as insect predators (middle). In the mining area, an area with a source of water is the destination for all animals, especially herpetofauna, and it even becomes the microhabitat for certain species. The purpose of this study is to determine the species of herpetofauna, its communities and its domination on the water area its microhabitats within the mine area. The exploration is carried out by direct observation (visual encounter surveys). The survey location determination is done intentionally (purposive sampling) on the potential habitat in some locations of water sources, such as pit dam, settling pond, man-made lake (embung), rivers and swamps. Based on the similarity index, there are five species of herpetofauna dominating the life in microhabitats and playing role in the formation of food chain community. Enhydris enhydris and Dendrelaphis pictus are species of snake and act as predator of frog Fejervarya cancrivora and Polypedates leucomystax.

Stable isotope analyses-A method to distinguish intensively farmed from wild frogs.

Consumption of frog legs is increasing worldwide, with potentially dramatic effects for ecosystems. More and more functioning frog farms are reported to exist. However, due to the lack of reliable methods to distinguish farmed from wild‐caught individuals, the origin of frogs in the international trade is often uncertain. Here, we present a new methodological approach to this problem. We investigated the isotopic composition of legally traded frog legs from suppliers in Vietnam and Indonesia. Muscle and bone tissue samples were examined for δ15N, δ13C, and δ18O stable isotope compositions, to elucidate the conditions under which the frogs grew up. We used DNA barcoding (16S rRNA) to verify species identities. We identified three traded species (Hoplobatrachus rugulosus, Fejervarya cancrivora and Limnonectes macrodon); species identities were partly deviating from package labeling. Isotopic values of δ15N and δ18O showed significant differences between species and country of origin. Based on low δ15N composition and generally little variation in stable isotope values, our results imply that frogs from Vietnam were indeed farmed. In contrast, the frogs from the Indonesian supplier likely grew up under natural conditions, indicated by higher δ15N values and stronger variability in the stable isotope composition. Our results indicate that stable isotope analyses seem to be a useful tool to distinguish between naturally growing and intensively farmed frogs. We believe that this method can be used to improve the control in the international trade of frog legs, as well as for other biological products, thus supporting farming activities and decreasing pressure on wild populations. However, we examined different species from different countries and had no access to samples of individuals with confirmed origin and living conditions. Therefore, we suggest improving this method further with individuals of known origin and history, preferably including samples of the respective nutritive bases.

Which frog's legs do froggies eat? The use of DNA barcoding for identification of deep frozen frog legs (Dicroglossidae, Amphibia) commercialized in France

Several millions frogs captured in the wild in Indonesia are sold for food yearly in French supermarkets, as deep frozen frog legs. They are commercialized as Rana macrodon, but up to 15 look-alike species might also be concerned by this trade. From December 2012 to May 2013, we bought 209 specimens of deep frozen frog legs, and identified them through a barcoding approach based on the 16S gene. Our results show that 206 out of the 209 specimens belong to Fejervarya cancrivora, two to Limnonectes macrodon and one to F. moodiei. Thus only 0.96 % of the frogs were correctly identified. Unless misclassification was intentional, it seems that Indonesian frog leg exporters are not able to discriminate between the species. The quasi absence of L. macrodon in our samples might be an indication of its rarity, confirming that its natural populations are declining rapidly, in agreement with its “vulnerable” status according to the IUCN Red List. Our results show that the genetic and morphological diversity of the frogs in trade is much higher than the genetic and morphological diversity measured so far by scientific studies. These results underline the need for large scale studies to assess the status of wild populations.

Kepadatan Populasi Kodok Fejervarya cancrivora di Persawahan Kabupaten Karawang, Jawa Barat

ABSTRACT Karawang District is the largest producer of frog meat in West Java. To find out how the frog harvesting impacts to the represent condition of population in the wild, on 16-27 May 2016 population survey was conducted on paddy field in East Karawang by implementing transect method with total length was 3000 meters. The results of this transect method showed that the population of juvenile was the highest in most areas of transects, followed by pre-adult population. The population of adult individuals was at the lowest and most adult male individuals were detected by the call that they make. Conditions of water supply in rice fields also determine the conditions of juvenile and pre-adult populations; newly planted rice fields with high water availability obtained the highest population per unit area compared to the condition of paddy fields with a little water availability; but exceptions to the plowed rice fields, despite the availability of a lot of water but disturbance from human activities was also high. Average density per unit area m2 for a little watery rice fields to dry with a total length of 1500 meters transect were as follows: (1) juvenil density was 0.33 individuals/m2, (2) pre-adult density was 0.04 individuals/m2, (3) adult density was 0.005 individuals/m2. Average density for much watery rice fields with a total length of 1200 meters transect were as follows: (1) juvenile density was 0.89 individuals/m2, (2) pre-adult density was 0.08 individuals/m2, (3) adult density was 0.01 individuals/m2. Key words: population density, Fejervarya cancrivora, paddy field, Karawang District

Amphibians of the Mangrove Areas of Odisha with a New Record to India

The amphibian fauna of the mangrove areas of Odisha was found to be quite rich with 14 species belonging to 9 genera and 5 families of Anurans. An important aspect of the study is that 5 species were recorded for the first time from the mangrove areas of Odisha. These included Duttaphrynus stomaticus (Bufonidae), Fejervarya moodiei, Hoplobatrachus crassus and Sphaerotheca rolandae (Dicroglossidae) and Hylarana tytleri (Ranidae). The Taylor's crab-eating frog (Fejervarya moodiei) was the first record of the species from India.

ASSEMBLAGES OF FROGS SPECIES AT BALAMBANGAN ISLAND, SABAH, MALAYSIA

Borneo is well known as a hotspot for biodiversity, yet species assemblages at smaller islands in the East Malaysian states of Sabah and Sarawak, are not well documented. The survey of frogs in Balambangan Island, which is situated at the west coast of Sabah, was the first attempt to look at the occurrence of frogs at smaller islands in the state. Sampling of frogs was conducted at the coastal areas of Kampung Selamat and limestone forest for four nights sampling period. Visual encounter survey and transect line were used to sample the frogs which were captured by hand. A total of seven species of 32 individuals were captured. This was only about 8% of the total species that occurs on the mainland Sabah. The island was dominated by the ranids (65%), rhacophoridae (22%) and microhylids (13%). Bufonids and megophryids were absent. The Mangrove frog, Fejervarya cancrivora (9 individuals) and the Lesser swamp frog, Limnonectes paramacrodon (8 individuals) were dominant on this island. No new species and endemism were noted on this island. The results from this preliminary study did not support the initial belief that the island contains high levels of endemism. On the contrary, the results appear to imply recent isolation from the Sabah mainland. Thus, a more detailed study including genetic diversity should be conducted to cover more areas on the main island and other smaller islands surrounding the Borneo Island and to obtain overall picture of biogeography of the frog species.

Isolation and characterization of 20 microsatellite markers for the crab-eating frog Fejervarya cancrivora

We isolated 20 polymorphic microsatellite markers in the crab-eating frog Fejervarya cancrivora which has experienced drastic population declines in China. The number of alleles per locus ranged from 3 to 17, and the observed heterozygosity ranged from 0.181 to 0.939. Fourteen loci were highly polymorphic (PIC > 0.5), four are moderately polymorphic (0.2 < PIC < 0.5), and two loci showed low polymorphism (PIC < 0.2). The combined probability of identity for the 20 loci was 6.2 × 10−19. Two loci deviated from Hardy–Weinberg equilibrium. These markers will provide useful tools for population genetic studies of F. cancrivora.

Origin and Identity of Fejervarya (Anura: Dicroglossidae) on Guam

Abstract: We used morphological and molecular data to infer the identity and origin of frogs in the genus Fejervarya that have been introduced to the island of Guam. Mensural and meristic data were collected from 96 specimens from throughout their range on the island and a principal component analysis was used to investigate the distribution of these data in morphological space. We also amplified a fragment of the 16S ribosomal ribonucleic acid mitochondrial gene from 27 of these specimens and compared it to 63 published sequences of Fejervarya and the morphologically similar Zakerana. All examined Fejervarya from Guam are morphologically indistinguishable and share an identical haplotype. The molecular data identify them as Fejervarya cancrivora with a haplotype identical to F. cancrivora from Taiwan.

Transcriptomes reveal the genetic mechanisms underlying ionic regulatory adaptations to salt in the crab-eating frog.

The crab-eating frog, Fejervarya cancrivora, is the only frog that lives near seas. It tolerates increased environmental concentrations of sodium, chloride and potassium partly by raising ion and urea levels in its blood plasma. The molecular mechanism of the adaptation remains rarely documented. Herein, we analyze transcriptomes of the crab-eating frog and its closely related saline-intolerant species, F. limnocharis, to explore the molecular basis of adaptations to such extreme environmental conditions. Analyses reveal the potential genetic mechanism underlying the adaptation to salinity for the crab-eating frog. Genes in categories associated with ion transport appear to have evolved rapidly in F. cancrivora. Both positively selected and differentially expressed genes exhibit enrichment in the GO category regulation of renal sodium excretion. In this category, the positively selected sites of ANPEP and AVPR2 encode CD13 and V2 receptors, respectively; they fall precisely on conserved domains. More differentially expressed rapidly evolved genes occur in the kidney of F. cancrivora than in F. limnocharis. Four genes involved in the regulation of body fluid levels show signs of positive selection and increased expression. Significant up-regulation occurs in several genes of F. cancrivora associated with renin-angiotensin system and aldosterone-regulated sodium reabsorption pathways, which relate to osmotic regulation.

INDUKSI KEJUTAN SUHU 360 C TERHADAP PERKEMBANGAN EMBRIO DAN KEBERHASILAN POLIPLOIDISASI KATAK (Rana cancrivora)

Polyploidy in the frog Rana cancrivora can be done by providing a temperature shock. The thing need to consider in treatment temperature shock at the egg is the beginning of a shock, temperature of shock and duration of shock. The research aims to identify and test the effect of temperature shock 36⁰C to the development of the embryo, and the success of poliploidisation of frog R. Cancrivora. The research was conducted in Biology Laboratory STKIP PGRI West Sumatra. The results shows that administration of 36⁰C temperature shock on egg fertilization influences on the development of frog embryos R. Cancrivora i.e. on the treatment D (45 minutes after fertilization). Yet, the temperature shock is 36⁰C does not show any influence on the success of poliploidisation of frog R. Cancrivora.

Nematodes of amphibians from Java, Indonesia, with a description of new species, Meteterakis wonosoboensis n. sp. (Nematoda : Heterakoidea).

During a survey on the parasites of amphibians of Indonesia, toads (30 Bufo melanostictus) and 246 frogs (213 Fejervarya cancrivora, 11 F. limnocharis, 22 Rana macrodon from West Java and 68 F. cancrivora from Central Java) were examined for parasitic nematodes. Three species of nematodes were found and described, i.e. Meteterakis wonosoboensis n. sp. from Fejervaria cancrivora; Meteterakis japonica from Bufo melanostictus, F. cancrivora and F. limnocharis; and Chabaudus sp. from F. cancrivora, F. limnocharis and Rana macrodon. Meteterakis wonosoboensis n. sp. is distinguished from other species of the genus by the length and shape of spicules, the number of caudal papillae, the presence of gubernaculum in male and the presence of vulval flap in female. Bufo melanostictus and Java are recorded as new host and locality for M. japonica, respectively.

Influence of Some Environmental Parameters on Some Frog Populations and their Parasitc Fauna

Wild animals, including frogs are usually infected with several species of parasites. Frogs are important for a variety of reasons. Firstly, they control populations of insects and secondly they can act as indicator species to the environment. Studies have shown that frogs can harbor many species of parasites. Worldwide, there were numerous researches carried out on frog parasites but few had been reported from Malaysia. From this study, 300 frogs from 14 species were collected from eight study sites: Duttaphrynus melanostictus, Phrynoidis aspera, Hylarana erythraea, H. labialis, H. nigrovittata, Polypedates leucomystax, Fejervarya cancrivora, F. limnocharis, Limnonectes blythii, L. paramacrodon, L. ibanorum, L. ingeri, Microhylabutleri, and Kaloulapulchra. Eleven (11) helminthic parasites (Heterakis spp., H. vesicularis, Trichostrongylus spp., Pharyngodon spp., Ascaris spp., Oswaldocruzia spp., Rhabdias spp.,Glypthelmins staffordi, Diplodiscusm sacculosus, Manodistomum spp., and Macracanthorynchus spp.), eight blood parasites (a rickettsia, Hepatozoon spp.,Haemogregarina spp., Lankesterella spp., Trypanosoma loricatum, T. rotatorium, T. chattoni, and microfilaria) and one protozoa (Nyctotherus spp.) were recorded from frogs. There were correlations between the presence of helminthic parasites and distribution of rainfall, as well as between blood parasites and Biochemical Oxygen Demand (BOD) of water.

GENETIC DIVERGENCE AND GEOGRAPHIC DISTRIBUTION OF FROGS IN GENUS FEJERVARYA FROM INDONESIA INFERRED FROM MITOCHONDRIAL 16S rRNA GENE ANALYSIS

The Indonesian archipelago is an ideal setting for the study of speciation and biogeography. This archipelago is divided into three island groups based on zoogeography: Sundaland, Wallaceaand the Australian region. In this paper we used frogs in genus Fejervarya (Bolkay) to study biogeography and examine patterns of gene flow across proposed zoogeographic boundaries. Severalmolecular studies on Fejervarya species from Indonesia have been carried out, but comparative studies among members of the genus Fejervarya have yet to be performed. In order to elucidate genetic divergence and geographic distribution of these frogs, we conducted a molecular analysis of the mitochondrial 16S rRNA gene using 179 frogs from five Fejervarya species. In total we collected from 32 localities in Sumatra, Kalimantan (Indonesian part of Borneo), Java, Bali, Sulawesi and Lesser Sunda Islands in Indonesia. Molecular phylogenetic analysis recovered 35 haplotypes and showed that frogs in the genus Fejervarya were divided into two well-supported clades. The first group were of three species, F. limnocharis, F. iskandari and F. cf. verruculosa and the other group clade consisted of Fejervarya cancrivora and Fejervarya sp. (Sulawesi-type). The average sequence divergence among these four species ranged from 1.09 to 16.03% (mean = 11.29±2.83%). The present results clearly show that there are five Fejervarya species in the Indonesian archipelago. Fejervarya limnocharis and F. cancrivora are widely distributed and sympatric in Sumatra, Borneo and Java. Fejervarya iskandari is not endemic to Java and also occurs in the Lesser Sundas. Fejervarya cf. verruculosa and Fejervarya sp. (Sulawesi-type) are endemic to Lesser Sunda and Sulawesi Island, respectively. Key words: Fejervarya, genetic divergence, geographic distribution, 16S rRNA gene

INDUKSI KEJUTAN SUHU 360 C TERHADAP PERKEMBANGAN EMBRIO DAN KEBERHASILAN POLIPLOIDISASI KATAK (RANA CANCRIVORA)

Polyploidy in the frog Rana cancrivora can be done by providing a temperature shock. The thing need to consider in treatment temperature shock at the egg is the beginning of a shock, temperature of shock and duration of shock. The research aims to identify and test the effect of temperature shock 36⁰C to the development of the embryo, and the success of poliploidisation of frog R. Cancrivora. The research was conducted in Biology Laboratory STKIP PGRI West Sumatra. The results shows that administration of 36⁰C temperature shock on egg fertilization influences on the development of frog embryos R. Cancrivora i.e. on the treatment D (45 minutes after fertilization). Yet, the temperature shock is 36⁰C does not show any influence on the success of poliploidisation of frog R. Cancrivora.

Changes in plasma angiotensin II, aldosterone, arginine vasotocin, corticosterone, and electrolyte concentrations during acclimation to dry condition and seawater in the crab-eating frog

The crab-eating frog Fejervarya cancrivora inhabits mangrove swamps and marshes in Southeast Asia. In the present study, circulating angiotensin II (Ang II), aldosterone (Aldo), arginine vasotocin (AVT), and corticosterone (Cort) concentrations as well as various blood parameters were studied under osmotically stressful conditions. Following acclimation to hyperosmotic seawater and dry condition for 5days, body weight was significantly decreased. Under both conditions, plasma Na+, Cl−, and urea concentrations, hematocrit values (Ht; blood volume indicator), and osmolality were significantly increased. Dehydration associated with hypovolemic and hyperosmotic states of body fluids was induced during acclimation to hyperosmotic seawater and dry condition in the crab-eating frogs. Ang II, Aldo, AVT, and Cort were maintained within relatively narrow concentration ranges in the control frogs; however, in frogs under dry and hyperosmotic seawater conditions, large variations were observed among individuals in each group. Mean plasma Ang II and Aldo concentrations significantly increased in hyperosmotic seawater-acclimated and desiccated frogs. Although mean plasma AVT concentrations in dehydrated frogs of both the groups were approximately 2.0–3.5 times higher than those in the control frogs, the differences were not significant because of the variation. There was a significant correlation between plasma osmolality and AVT as well as Ang II but not Aldo. A significant correlation was also observed between Ht and AVT as well as Ang II. Plasma Ang II was significantly correlated with plasma Aldo. These results indicate that the crab-eating frogs may exhibit similar physiological responses to both seawater-acclimated and dry conditions. It appears that under dehydrated conditions, osmoregulatory mechanisms participate in stabilization of the situation. The renin–angiotensin system may have pivotal roles in body fluid regulation under volemic and osmotic stress in the Fejervarya species with unique osmoregulation.