Frog Species Research Articles

Comparing skin swabs, buccal swabs, and toe clips for amphibian genetic sampling, a case study with a small anuran (Acris blanchardi).

Multiple methods for collecting genetic samples from amphibians exist, each with their own implications for study design, animal welfare, and costs. Toe clipping is one common method, but there is ongoing debate regarding its potential detriment. Less invasive methods should be implemented, if efficacious, as amphibians are a particularly vulnerable vertebrate group. Skin and buccal swabbing are less invasive methods for genetic sampling, but the potential for contamination and a lower yield of DNA may exist. To compare these methods, we gathered skin swabs, buccal swabs, and toe clips from the same individuals of a relatively small anuran species, Blanchard's Cricket Frog (Acris blanchardi). We then compared DNA yield, DNA purity, amplification success rate, and genotypic data quality among sample types. We found toe clips and buccal swabs generated similar DNA yield and purity, with skin swabs yielding significantly less DNA of significantly lower purity than the other sample types. Amplification success rate was significantly higher using toe clips compared to the other sample types, though buccal swab samples amplified more readily than skin swabs. Genotypic data from toe clips and buccal swabs did not differ significantly in quality, but skin swab data quality was significantly lowest among sample types. Thus, skin swabbing could produce erroneous data in some situations, but buccal swabbing is likely an effective substitute to toe clipping, even for small species. Our results can help future researchers select which genetic sampling method might best suit their research needs.

Advertisement call variation is related to environmental and geographic distances in two anuran species inhabiting highland forests in northeastern Brazil

AbstractHighlands are of paramount importance to the study of evolution as they are frequently implicated in historical and ecological processes that generate and maintain biological diversity. In northeastern Brazil, sparse rainforest remnants located in highlands north of the São Francisco River are surrounded by the dry and open landscape of the Caatinga biome. Earlier studies suggested that these forests were historical refuges to the rainforest fauna and flora during Pleistocene's climatic cycles. However, it is still unclear whether populations in distinct highlands experienced phenotypic differentiation as a result of adaptation to environmental conditions of each forest remnant. Herein, we used two frog species widely distributed and ecologically different, Dendropsophus oliveirai, a habitat specialist, and Physalaemus cuvieri, a habitat generalist, to investigate the relationships between environmental variation, geographic, genetic, and body size distance with advertisement call variation among populations inhabiting different highlands. Our results indicated that acoustic variation among P. cuvieri populations is strongly influenced by environmental variation, but also by the geographic distance among populations. In D. oliveirai, environment is also the most influent factor on acoustic variation, followed by a lower influence of genetic and morphological variation. Associations between environmental and geographic factors suggest indirect effects of geographic distance on acoustic variation in both species through an environmental gradient. We believe that selective processes and isolation by distance possibly act together in driving interpopulational acoustic variation with habitat‐specific species being more affected by geographic isolation in suitable habitats.Abstract in Portuguese is available with online material.

Physiological thermal niches, elevational ranges and thermal stress in dendrobatid frogs: An integrated approach

AbstractAimWe investigated the relationship between thermal physiology, elevational distribution and thermal stress among nine closely related dendrobatid frogs during their aquatic stage by employing an integrated approach, combining thermal physiology, environmental temperature modelling and predictive assessments of current and future exposure to thermal variation.LocationEcuador.TaxonAmphibians; Anura, Dendrobatidae, Epipedobates, Hyloxalus.MethodsWe determined the thermal performance curves (TPCs) of larval growth for each species and modelled the thermal variation in contrasting aquatic larval environments for both present and future times. This allowed us to estimate the expected elevational distributions and forecast periods of exposure to stressful temperatures that inhibit larval growth due to elevation and global warming.ResultsWe found significant correlations between optimum temperature (Topt), 50% maximum performance temperature (maxB50), 50% minimum performance temperature (minB50) and cold resistance (survival at 9°C) with the current elevational distributions. However, thermal physiology predicted lower than observed distributions for high‐elevation dendrobatids and higher than observed maximum elevations for lowland species. Nonetheless, our models predicted that low thermal variability habitats (i.e. streams and deep permanent ponds) can buffer the future temperature increase for all taxa, even when considering the most extreme scenario. In contrast, all species within high thermal variation habitats (open forest temporary ponds) are expected to experience stressful temperatures under present conditions.Main ConclusionsThe findings indicate that thermal physiology may not be a limiting factor for dendrobatid frog species' ranges in this equatorial mountain gradient. Highland species may need to adapt to suboptimal performance, while some lowland species could occupy higher elevations. This study emphasizes the importance of habitat buffering to mitigate thermal stress in the face of climate change for amphibians in tropical mountains.

Geographic variation in the acoustic signals of lesser treefrogs Dendropsophus minutus (Anura, Hylidae)

Biological sound archives are a major source to investigate geographic variation in animal acoustic signals and their evolutionary drivers. The acoustic signals of anuran species with wide distribution ranges often vary geographically as a result of isolation by distance and climate amongst other factors. We examined whether the acoustic structure of call notes would vary geographically in lesser treefrogs Dendropsophus minutus using recordings from bioacoustics repositories. We also tested whether climate (mean annual temperature and annual precipitation) drive geographic variation in those signals. The acoustic distance was unrelated to geographic distance, suggesting that isolation by distance solely cannot explain geographic variation in call structure. Overall, lesser treefrogs uttered call notes with lower frequencies and bandwidths in the west of their range. In addition, frogs produced shorter call notes in hotter, wetter sites and narrow-bandwidth notes in hotter environments. We suggest that frogs produce more calls (not measured here) of shorter durations to maximize transmission and minimize the metabolic costs of calling at high air temperatures. We also suggest that hotter environments favor the propagation of lower-bandwidth calls. This study reinforces the feasibility and power of citizen science and acoustic data repositories for understanding the evolution of anuran acoustic signals.

Sexual Size Dimorphism in Multiple Traits: An Integrative Perspective in Several Anuran Species

Sexual Size Dimorphism in Multiple Traits: An Integrative Perspective in Several Anuran Species

Zoogeographical regions in the Atlantic Forest: patterns and potential drivers

AbstractAimTo delineate present‐day zoogeographical regions of terrestrial vertebrates (frogs, lizards, snakes, birds and non‐volant mammals) in the Atlantic Forest. Within each taxonomic group, we examine the relative importance of abrupt climatic transitions, orographic barriers, past climate change and rivers in shaping zoogeographical boundaries.LocationSouth America's Atlantic Forest.MethodsWe applied a network‐based method to delineate zoogeographical regions, using distribution data (range maps) for 455 species of frogs, 103 lizards, 220 snakes, 917 birds and 202 non‐volant mammals, in 50 × 50 km grid cells. We used hierarchical generalized linear mixed‐effects models to test environmental predictors associated with zoogeographical boundaries. Finally, we intersected the bioregion maps delineated for each group to identify general patterns across all vertebrates.ResultsWe identified four zoogeographical regions for birds and snakes, and five for frogs, lizards and non‐volant mammals. Depending on the group, contemporary and past climate conditions, elevation variation and/or rivers were associated with zoogeographical boundaries. The combined maps indicate that the Atlantic Forest retains four spatially cohesive zoogeographical regions based on present‐day distribution of vertebrates.Main conclusionsCross‐taxon congruence indicates that the geographical and environmental characteristics of the Atlantic Forest have a strong influence on the location of zoogeographical regions for vertebrates. In contrast, transition zones appear to be associated with the spatial distribution of life history traits of each group, potentially explaining the observed differences in the number of bioregions across groups and the position of zoogeographical boundaries. This work paves the way for further research into the evolutionary assembly of the Atlantic Forest's zoogeographical regions and may help inform conservation priorities for maintaining their distinctive faunas.

It's not easy being green: Comparing typical skin colouration among amphibians with colour abnormalities associated with chromatophore deficits.

Amphibians can obtain their colour from a combination of several different pigment and light reflecting cell types called chromatophores, with defects in one or several of the cells leading to colour abnormalities. There is a need for better recording of colour abnormalities within wild amphibian populations, as this may provide baseline data that can be used to determine changes in environmental conditions and population dynamics, such as inbreeding. In this study, we provide records of several types of chromatophore deficiencies, including those involving iridophores, xanthophores and melanophores, among two Australian tree frog species; the green and golden bell frog, Litoria aurea, and the eastern dwarf tree frog, L. fallax. We explore these colour abnormalities in terms of the chromatophores that have likely been affected and associated with their expression, in combination with typical colour phenotypes, colour variations and colour changes for these species. We intend for our photographs to be used as a visual guide that addresses the need for more accessible information regarding the physical manifestation of different chromatophore defects among amphibians.

Rapid molecular identification of Rana dybowskii by species-specific primers

Oviductus Ranae is the dried oviduct from Rana dybowskii, a forest frog species with medicinal, tonic, and cosmetic properties. Due to the high price and resource shortage, counterfeit varieties of Oviductus Ranae often appear in the market. However, traditional identification methods cannot accurately differentiate between Oviductus Ranae and its adulterants. In this study, a rapid molecular identification method has been established. The method involves extracting genomic DNA in just 30 s using filter paper purification, species-specific rapid polymerase chain reaction (PCR) amplification, and finally, fluorescence detection of the products. It can accurately identify Oviductus Ranae and its three common adulterants in about 30 min, making the process simple, fast, and highly specific.

Overcoming the lack of distribution data for range‐restricted habitat specialist frogs

AbstractWith over 40% of species threatened with extinction, the distributional range of most amphibians is still unknown, generating uncertainty whether species are naturally rare, under‐sampled, or difficult to detect. We implemented a modelling approach that uses bromeliads as surrogates to predict the distribution of habitat specialist frogs that lack distributional data. We aimed to predict and survey potential new sites for the occurrence of a rare and microendemic bromeliad‐dwelling frog from the Atlantic Rainforest of Brazil. We used Maxent to predict suitable areas based on climate and topographic profiles, combined with 21 occurrence records of bromeliads within which our target frog species (Crossodactylodes itambe) is strictly restricted. The bromeliad‐based models identified four areas potentially suitable, and subsequent surveys revealed an entirely new species of a rare bromeliad‐dwelling frog in one of the areas. We demonstrate that using easy‐to‐survey surrogate species that have a strong relationship with species that are hard to detect has enormous potential to reveal crucial information on the potential ranges and distribution of cryptic taxa. Such a surrogate modelling approach could be extended to other habitat specialist species that lack distribution data, such as amphibians restricted to specific refugia and/or reproduction sites. This could improve the targeting of surveys and increase understanding of the patterns of rarity and the drivers of species distribution, especially for areas with high endemicity and range‐restricted frogs.

Modelling human genetic disorders in Xenopus tropicalis.

Recent progress in human disease genetics is leading to rapid advances in understanding pathobiological mechanisms. However, the sheer number of risk-conveying genetic variants being identified demands in vivo model systems that are amenable to functional analyses at scale. Here we provide a practical guide for using the diploid frog species Xenopus tropicalis to study many genes and variants to uncover conserved mechanisms of pathobiology relevant to human disease. We discuss key considerations in modelling human genetic disorders: genetic architecture, conservation, phenotyping strategy and rigour, as well as more complex topics, such as penetrance, expressivity, sex differences and current challenges in the field. As the patient-driven gene discovery field expands significantly, the cost-effective, rapid and higher throughput nature of Xenopus make it an essential member of the model organism armamentarium for understanding gene function in development and in relation to disease.

Molecular systematics of the subgenus Gephyromantis (Phylacomantis) with description of a new subspecies

The mantellid subgenus Phylacomantis (genus Gephyromantis) currently contains four species of frogs distributed in the South (G. corvus, G. kintana), West (G. atsingy), North East, North, and Sambirano regions (G. pseudoasper) in Madagascar. We assess the molecular systematics of these amphibians based on extended sampling and analysis of multiple nuclear-encoded and mitochondrial genes. We confirm the delimitation of the four known species, which form monophyletic groups in a mitochondrial tree and separate phylogroups without haplotype sharing in haplotype networks of four nuclear markers. Newly included samples of G. pseudoasper from the North and North East regions confirm a comparatively low genetic variation of this species across its range, with pairwise genetic distance in the 16S gene not exceeding 1.4%. On the contrary, newly collected specimens from three sites in the North East and North West of the island strongly differed from all other Phylacomantis, with genetic distances exceeding 4.8% for the16S gene. Mitochondrial and nuclear markers strongly suggest them to be sister to G. corvus from the South. Due to haplotype sharing in two nuclear genes, absence of detectable morphological differences, and lack of data on bioacoustic differentiation, we describe these populations as a new subspecies, G. corvus bakilana ssp. nov., emphasizing that this taxon may be elevated to species status in the future pending new data. This biogeographic pattern, with apparently disjunct sister lineages in the South vs. North West of Madagascar, is rare among Malagasy anurans and calls for increased exploration of the remaining humid forest fragments in the North West and West of the island.

Genetic diversity of European tree frogs (Hyla arborea group): A systematic review

Amphibian populations are increasingly threatened by global change and the study of their genetic diversity is a major conservation priority. Western palearctic tree frog species of the Hyla arborea group are commonly distributed across Europe and the Middle East and many have declining populations. We performed a PRISMA systematic review to gain insight into the genetic diversity of H. arborea group. Sixteen published studies were included in the final qualitative analysis. While the genetic diversity of H. arborea group species was widely variable, it could often be explained by phylogeographic history. Populations in Western and Northern Europe had lower genetic diversity, with some populations also affected by habitat fragmentation. However, important regions of high genetic diversity were found in the Balkan peninsula for H. arborea sensu stricto and around the Black Sea for H. orientalis. Genetic diversity of H. molleri, H. savignyi, H. meridionalis, H. felixarabica, H. intermedia, H. sarda has been investigated only across extensive phylogeographical studies, while data regarding their genetic diversity at the local level are missing. Through our review, we identify knowledge gaps about the genetic diversity of the H. arborea group that require further investigation, of and illustrate how filling these gaps might translate into future conservation efforts.

The chondrocranium of the tadpole of Hymenochirus boettgeri (Anura: Pipidae) and the sequence and timing of its development

The evolutionary development of the vertebrate head has played a critical role in shaping the diversity of body plans and ecological roles within vertebrate species. This evolutionary journey is characterised by the emergence of the head compartment during the Cambrian radiation, the evolution of jaws during the Silurian, and the subsequent diversification of vertebrate groups. The cranium, consisting of the neurocranium, dermatocranium, and viscerocranium, plays a central role in craniofacial support. Research on the development of the cranium has been carried out in several vertebrate species, providing valuable insights into their unique patterns of cartilage development. While most vertebrates show typical sequences of chondrocranial development, anurans divert from these patterns. Our study focused on the chondrocranial development of Congo dwarf frog Hymenochirus boettgeri (Anura: Pipidae) tadpoles, providing a detailed sequential analysis from mesenchymal elements to the premetamorphic chondrocranium. The chondrocranium of these tadpoles showed several reductions as a consequence of their miniaturization and their unique suction feeding lifestyle. We found a mosaic-like development of anuran mandibular and hyoid arches, and we also discussed the intricate patterns of suprarostral cartilage development and deviations in branchial basket development compared to other anuran species. Our study also contrasts anuran neurocranial development with that of other osteichthyan species, highlighting unique features such as the similar direction of development of anterior and posterior neurocranial components. Overall, our research contributes to our understanding of chondrocranial development in anurans, highlighting both conserved and unique features in H. boettgeri tadpoles. Our findings enrich our knowledge of vertebrate cranial development and provide insights into the evolutionary history of anuran chondrocranial features.

Range extension and expanded description of Micryletta hekouensis Liu, Hou, Mo & Rao, 2021 (Amphibia, Anura, Microhylidae), with comments on Micryletta of Northern Vietnam

The Paddy frog species Micryletta hekouensis was described based on only two specimens from Nanxi Village, Nanxi Town, Hekou County, Honghe Prefecture, Yunnan Province, China. Herein, we report on new findings and a range extension of this species based on a re-examination of preserved specimens deposited in Duy Tan University (DTU) and Zoological Museum of Lomonosov Moscow State University (ZMMU) collected from Vietnam. All new specimens were previously identified as Micryletta cf. inornata or M. cf. steinegeri. Molecular analyses based on mitochondrial DNA supported the morphological findings. The newly identified specimens have a pairwise divergence of only 0.7% from those of the type series of M. hekouensis, based on the 16s rRNA mtDNA gene. Based on the new information, we confirm the presence of M. hekouensis in Vietnam and update the diagnostic characters of this species and its distribution. We suggest the species should be considered as Near Threatened (NT) following the IUCN’s Red List categories. Further studies reassessing the populations of the Micryletta inornata complex in the Indochinese Region (including Vietnam, Laos, Cambodia, and Thailand) are required.

Broad-scale pesticide screening finds anticoagulant rodenticide and legacy pesticides in Australian frogs

Pesticide contamination poses a significant threat to non-target wildlife, including amphibians, many of which are already highly threatened. This study assessed the extent of pesticide exposure in dead frogs collected during a mass mortality event across eastern New South Wales, Australia between July 2021 and March 2022. Liver tissue from 77 individual frogs of six species were analysed for >600 legacy and contemporary pesticides, including rodenticides. More than a third (36 %) of the liver samples contained at least one of the following pesticides: brodifacoum, dieldrin, DDE, heptachlor/heptachlor epoxide, fipronil sulfone, and 2-methyl-4-chlorophenoxyacetic acid (MCPA). Brodifacoum, a second-generation anticoagulant rodenticide, was found in four of the six frog species analysed: the eastern banjo frog (Limnodynastes dumerilii), cane toad (Rhinella marina), green tree frog (Litoria caerulea) and Peron's tree frog (Litoria peronii). This is the first report of anticoagulant rodenticide detected in wild amphibians, raising concerns about potential impacts on frogs and extending the list of taxa shown to accumulate rodenticides. Dieldrin, a banned legacy pesticide, was also detected in two species: striped marsh frog (Limnodynastes peronii) and green tree frog (Litoria caerulea). The toxicological effects of these pesticides on frogs are difficult to infer due to limited comparable studies; however, due to the low frequency of detection the presence of these pesticides was not considered a major contributing factor to the mass mortality event. Additional research is needed to investigate the effects of pesticide exposure on amphibians, particularly regarding the impacts of second-generation anticoagulant rodenticides. There is also need for continued monitoring and improved conservation management strategies for the mitigation of the potential threat of pesticide exposure and accumulation in amphibian populations.

Bacteria from the skin of amphibians promote growth of Arabidopsis thaliana and Solanum lycopersicum by modifying hormone-related transcriptome response

Plants and microorganisms establish beneficial associations that can improve their development and growth. Recently, it has been demonstrated that bacteria isolated from the skin of amphibians can contribute to plant growth and defense. However, the molecular mechanisms involved in the beneficial effect for the host are still unclear. In this work, we explored whether bacteria isolated from three tropical frogs species can contribute to plant growth. After a wide screening, we identified three bacterial strains with high biostimulant potential, capable of modifying the root structure of Arabidopsis thaliana plants. In addition, applying individual bacterial cultures to Solanum lycopersicum plants induced an increase in their growth. To understand the effect that these microorganisms have over the host plant, we analysed the transcriptomic profile of A. thaliana during the interaction with the C32I bacterium, demonstrating that the presence of the bacteria elicits a transcriptional response associated to plant hormone biosynthesis. Our results show that amphibian skin bacteria can function as biostimulants to improve agricultural crops growth and development by modifying the plant transcriptomic responses.

Do aposematic species have larger range sizes? A case study with neotropical poison frogs

AbstractAimAposematic animals, i.e., those that are defended and warn potential predators through signals, are suggested to have resource‐gathering advantages against non‐aposematic ones. We here explore this in a biogeographic framework expecting that aposematic species are better dispersers, which translates into larger geographic range size.LocationSouth America.TaxonPoison frogs (Amphibia; Aromobatidae and Dendrobatidae).MethodsWe use 43 toxic and 26 non‐toxic poison frog species from the lowlands only as representatives of aposematic and non‐aposematic study organisms, respectively. Realised and potential geographic ranges are calculated using minimum convex polygon and species distribution modelling methods, respectively. Accounting for species body size and phylogeny, we test if both range and aposematism are correlated using linear mixed‐effects models.ResultsAposematic and non‐aposematic species neither differ in realised nor in potential geographic range size. There was no effect on body size.Main ConclusionsThe role of aposematism is not yet as clear as suggested and determinants of poison frog range sizes are multifaceted. A more integrative approach is needed using the information on behaviour, predation risk, and reproductive biology to assess the role of aposematism on observed species distributions. Such data are not yet available for most species, neither poison frogs nor other aposematic animals.

A new species of the genus Amolops (Amphibia: Ranidae) and the first national record of Amolops vitreus from China

Abstract The torrent frogs of the genus Amolops represent a great anuran diversification in southern China and Southeast Asia. Previous studies have shown that, the diversity of this genus still remains underestimated. During herpetological surveys from 2021 to 2022, several Amolops specimens were collected from the international border regions of southwestern Yunnan Province, China. Herein, we utilized molecular phylogenetic and morphological data to identify these specimens. Our findings indicate the presence of a separate and previously unknown lineage in the A. viridimaculatus group, which we formally describe as a new species. Furthermore, the specimen from Xishuangbanna National Nature Reserve clustered with A. vitreus from the paratype, supporting the morphological diagnosis. Therefore, we describe a new species and a new species record for China. Our study contributes to the species richness of the genus Amolops as well as the diversity of amphibians in China. Notably, our discovery brings the total number of Amolops species to 85 and the total number of torrent frog species known to occur in China to 53. In addition, our study further confirmed that Yunnan and Indochina Peninsula have similar faunal composition, implying that more studies are needed to achieve a complete understanding of the species diversity and distribution pattern.

Unexpected Formation of 11(9→7)-abeo-Steroid Skeleton in Synthetic Studies toward Batrachotoxin.

Batrachotoxin (1) is a potent cardio- and neurotoxic steroid isolated from certain species of frogs, birds, and beetles. We previously disclosed two synthetic routes to 1. During our synthetic studies toward 1, we explored an alternative strategy for efficiently assembling its 6/6/6/5-membered steroidal skeleton (ABCD-ring). Here we report the application of intermolecular Weix and intramolecular pinacol coupling reactions. While Pd/Ni-promoted Weix coupling linked the AB-ring and D-ring fragments, SmI2-mediated pinacol coupling did not cyclize the C-ring. Instead, we discovered that SmI2 promoted a 1,4-addition of the α-alkoxy radical intermediate to produce the unusual 11(9→7)-abeo-steroid skeleton. Thus, this study demonstrates the convergent assembly of the skeleton of the natural product matsutakone in 11 steps from 2-allyl-3-hydroxycyclopent-2-en-1-one.

Batrachochytrium dendrobatidis and Hannemania mite’s relationships with Mexican amphibians in disturbed environments

The rapid transformation and pollution of ecosystems have severely impacted biodiversity. Specifically, anthropogenic activities have imposed adverse effects on amphibians, with evidence suggesting that these activities alter parasite and pathogen interactions within their hosts. To investigate these interactions in areas affected by different anthropogenic activities, our study focused on analyzing a pathogen and a parasite known to interact within the amphibian skin (spongy epidermis layer) and both compromising amphibian health: Batrachochytrium dendrobatidis (Bd), a fungus responsible for chytridiomycosis, a disease associated with massive population declines in amphibians and the Hannemania sp. mite in Mexico. Four sampling areas along the Sonora River were selected, representing different human activities: mining, livestock, wastewater discharge, agriculture, and one in an urban zone. We analyzed 135 amphibians across 10 anuran species. Among these, the most abundant species (Lithobates yavapaiensis) exhibited the highest prevalence of both pathogen and parasite (90.1% and 27.3%, respectively) and was significantly associated with the intensity of Bd-infection. The prevalence of Hannemania mites varied significantly across sampling sites as did Bd prevalence and infection load, with the highest Bd load found at the wastewater discharge site. A significant association between the intensity of Bd-infection and both mite abundance and amphibian species was observed when the sampling site was considered. Additionally, sites with Bd-positive individuals and Hannemania parasitism coincide with refractory elements characterized by mechanical or corrosion resistance. The persistence of these elements in the environment, along with the small particle size (<850 nm) found in sediments, poses a potential risk of internalization, bioaccumulation (e.g., Fe, Co, and Ti), and their transfer through the food chain. It is thus essential to consider monitoring environmental and biotic factors that modulate the relationships between parasites, pathogens, and amphibians if we are to propose conservation strategies adapted to disturbed environments.