Amphibian Species Research Articles

Terrestrial vertebrate fauna in a multifunctional landscape in Gabaldon, Nueva Ecija, Philippines

Abstract. Gojo-Cruz PHP, Caldito JC, Catubig YDL, Cornes HM, Corpuz JS, Maniquiz SS, Matunan KGE, Pascual FJ, Miguel ESS, Siguancia GC, Turqueza DC, Madlao JP, Cruz CF, Merculio JG. 2024. Terrestrial vertebrate fauna in a multifunctional landscape in Gabaldon, Nueva Ecija, Philippines. Biodiversitas 25: 583-597. This study was done to document the vertebrate fauna in a multifunctional landscape at the foothills of Mt. Mingan, Gabaldon, Nueva Ecija. The inventory of wild vertebrate fauna associated with the different habitat types in the area will be valuable in creating suitable management programs to reduce the impact of human activities in the area without affecting local economic activities. Standard survey techniques, including day and night transect walks, opportunistic surveys, mist-netting, and cage trapping, were employed to document vertebrate fauna in the agro-forest, around built-up areas, forest edges, lowland forest, grassland, and riparian habitats. A total of 79 species of vertebrate fauna were recorded, including 7 species of mammals, 13 species of reptiles, 9 species of amphibians, and 50 species of birds. Thirty-three (33) species were endemic to the Philippines. Regarding conservation status, eight (8) species were listed in the higher threat status under DAO 2019-09. Species richness was observed to be highest in lowland dipterocarp forests. On the other hand, species composition varies between habitats, with endemic species being more common in complex habitats such as lowland dipterocarp forests than in simpler habitats like agricultural areas. The high species richness and the presence of endemic and threatened species in the multifunctional landscape of the area highlight the need for management plans considering habitat heterogeneity and connectivity.

Does the anesthetic tricaine methanesulfonate (MS-222) distort oxidative status parameters in tadpoles?

The effect of anesthesia/euthanasia with ethyl 3-aminobenzoate methanesulfonate (MS-222) on the oxidative status of Hyla arborea tadpoles was examined to determine whether the use of the anesthetic can confound the experimental results of the oxidative stress-based investigation. The experiment was conducted on two groups of tadpoles reared at different temperatures to produce differences in antioxidant capacity between the groups. After development at different temperatures (20 °C and 25 °C), the animals were exposed to different concentrations of MS-222 (0, 0.1, 1, and 5 g/L) for 15 min. The higher temperature decreased catalase activity, glutathione and protein carbonyl levels and increased glutathione reductase activity. The glutathione level and glutathione/thiol-related parameters were significantly changed after MS-222 exposure. However, individuals from the different temperature groups responded differently to the tested anesthetic, pointing to the possible influence of the initial levels of antioxidant capacity. The analysis of the interaction between the factors (temperature and MS-222) confirmed that the anesthetic can confound the results regarding the effects of temperature on the oxidative status parameters. The concentration of 0.1 g/L MS-222 had the lowest influence on the alterations in oxidative status and the results of the effect of temperature. A brief review of the current literature on the use of MS-222 in tadpoles made clear the absence of precise information on anesthetic concentration and exposure time. Similar studies should be repeated and extended to other amphibian species and other factors of interest to provide better guidance on tadpole anesthesia/euthanasia for future experiments that consider oxidative status parameters.

EDNA Metabarcoding: Gaps of Publicly Available Reference Databases of Freshwater Vertebrates in Sri Lanka

Environmental DNA (eDNA) has become a rapidly emerging technology during the last decade. Although eDNA applications are used worldwide, none of the eDNA metabarcoding studies have been conducted solely in Sri Lanka yet. Detection rate of species through DNA metabarcoding technique varies with the availability of gene sequences in reference databases. Therefore, the main objective of this study was to assess the data gaps of local species in two selected reference databases (the NCBI database and the BOLD database). Local freshwater vertebrates were considered for this analysis by dividing them into three major categories as freshwater fish (97 species), freshwater amphibians (40 species) and freshwater reptiles (7 species). The availability of COI gene sequence and CytB gene sequence of each species were recorded. In the NCBI database, reference gene sequences were available only for 17.53% (17 species) of freshwater fish, 7.50% (3 species) of freshwater amphibians and 14.29% (1 species) of freshwater reptiles. In the BOLD database, reference gene sequences were available for 52.58% (51 species) of freshwater fish, 17.50% (7 species) of freshwater amphibians and 42.86% (3 species) of freshwater reptiles. Endemic species were analyzed separately. This revealed the availability of reference gene sequences for 4.92% (3 species) of endemic freshwater fish in the NCBI database. However, none of the reference gene sequence was found for endemic freshwater amphibians and reptiles. In the BOLD database, reference gene sequences were available for 36.07% (22 species) of endemic freshwater fish and 33.33% (1 species) of endemic freshwater reptiles but none for endemic freshwater amphibians. According to the results, large gaps of publicly available reference CO1 gene sequences were observed in both NCBI and BOLD databases. Not only the availability of reference gene sequences, but also the submitted countries of those sequences were also analyzed as a part of this study. Asia has contributed the most according to the country-vice analysis. Among them, many are in the South Asian region. Therefore, this study highlights the need of expanding reference databases for greater local representation as well as the importance of forging collaborations with the developed countries in the same region in order to increase international cooperation and funding to carry on further studies on neglected taxa and under‐represented taxa in Sri Lanka. 
 Keywords: eDNA (environmental DNA), Metabarcoding, Reference databases, Reference gene sequences

Is thermal sensitivity affected by predation risk? A case study in tadpoles from ephemeral environments.

Changes in environmental temperature may induce variations in thermal tolerance and sensitivity in ectotherm organisms. These variations generate plastic responses that can be analyzed by examining their Thermal Performance Curves (TPCs). Additionally, some performance traits, like locomotion, could be affected by other factors such as biological interactions (e.g., predator-prey interaction). Here, we evaluate if the risk of predation modifies TPCs in Mendoza four-eyed frog (Pleurodema nebulosum, Burmeister, 1861) and Guayapa's four-eyed frog (Pleurodema guayapae, Barrio, 1964), two amphibian species that occur in ephemeral ponds in arid environments. We measured thermal tolerances and maximum swimming velocity at six different temperatures in tadpoles under three situations: control, exposure to predator chemical cues, and exposure to conspecific alarm cues. TPCs were fitted using General Additive Mixed Models. We found that curves of tadpoles at risk of predation differed from those of control mainly in thermal sensitivity parameters. Our work confirms the importance of biotic interactions have in thermal physiology.

Enhancing predictive performance for spectroscopic studies in wildlife science through a multi-model approach: A case study for species classification of live amphibians.

Near infrared spectroscopy coupled with predictive modeling is a growing field of study for addressing questions in wildlife science aimed at improving management strategies and conservation outcomes for managed and threatened fauna. To date, the majority of spectroscopic studies in wildlife and fisheries applied chemometrics and predictive modeling with a single-algorithm approach. By contrast, multi-model approaches are used routinely for analyzing spectroscopic datasets across many major industries (e.g., medicine, agriculture) to maximize predictive outcomes for real-world applications. In this study, we conducted a benchmark modeling exercise to compare the performance of several machine learning algorithms in a multi-class problem utilizing a multivariate spectroscopic dataset obtained from live animals. Spectra obtained from live individuals representing eleven amphibian species were classified according to taxonomic designation. Seven modeling techniques were applied to generate prediction models, which varied significantly (p < 0.05) with regard to mean classification accuracy (e.g., support vector machine: 95.8 ± 0.8% vs. K-nearest neighbors: 89.3 ± 1.0%). Through the use of a multi-algorithm approach, candidate algorithms can be identified and applied to more effectively model complex spectroscopic data collected for wildlife sciences. Other key considerations in the predictive modeling workflow that serve to optimize spectroscopic model performance (e.g., variable selection and cross-validation procedures) are also discussed.

An updated checklist of the amphibians and reptiles of Nayarit, Mexico with conservation status and comparison with adjoining States

Herein, we present an updated list of the species of amphibians and reptiles that inhabit Nayarit, Mexico. In the years since the publication of a previous list in 2016, threats to amphibians and reptiles in Nayarit have continued unabated and efforts to more fully catalogue the richness of Nayarit’s herpetofauna have continued. Nayarit harbours 162 native species of amphibians and reptiles, representing 35 families and 85 genera. These include 37 species of amphibians (35 anurans and two salamanders) and 125 species of reptiles (one crocodile, 44 lizards, 69 snakes and 11 turtles). Of the amphibian and reptile species in Nayarit, 102 are endemic to Mexico and two endemic to Nayarit. The ecoregion with the highest richness of amphibians and reptiles in Nayarit is the Trans-Mexican Volcanic Belt followed closely by the Sierra Madre Occidental and then the Sierra Madre del Sur, the Pacific Lowlands and the Nayarit Islands. Just 5.8% of the species of amphibians and reptiles in Nayarit is IUCN-listed as Vulnerable, Endangered or Critically Endangered, 11.1% are placed in a protected category by SEMARNAT and nearly a third are categorised as high risk by the Environmental Vulnerability Score. The herpetofauna of Nayarit overlaps the most with Jalisco (88.9%). One of the main conclusions of our updated list of the amphibian and reptile species of Nayarit is that our understanding of the full complement of species is not complete, especially the reptiles.

Ocular Gnathostomiasis in A Farmer From South Sulawesi, Indonesia

Introduction : Ocular Gnathostomiasis is a rare parasitic infection caused by the third-stage larvae of the spiruroid nematode. It is a food-borne zoonosis caused by the consumption of raw or undercooked freshwater species of fish, amphibians, reptiles, birds, and mammals.&#x0D; Case Illustration : A 37-year-old farmer complained of seeing moving objects in his left eye accompanied by blurred vision. The patient had a history of red left eye with increased intraocular pressure. The visual acuity of the left eye was 20/30. Slit lamp examination showed slight hyperemia of the conjunctiva and a live worm moving in the anterior chamber. The worm was immediately removed using corneal clear incision and sent for parasitological examination. By finding four rows of hooks on the head and fine spines covering the body, the worm was identified as a stage 3 larva of gnathostome sp.&#x0D; Discussion : Surgical removal of the larvae is considered the most effective treatment. In this case, the patient was then treated with Albendazole 400 mg daily for 21 days and Prednisolone 50 mg for 7 days with reduced doses of 10 mg weekly for three weeks&#x0D; Conclusion : Ocular Gnathostomiasis by third-stage larvae of spiruroid nematode was found at the eye of a man from Siwa, South Sulawesi. The intracameral worm was successfully extracted from the eye. Patients with complaints of recurrent redness of the eyes accompanied by increased intraocular pressure and a history of living in an area with a habit of eating raw freshwater fish should be considered the possibility of harboring helminth in the eye.

Herpetofauna in Ranu Darungan and Blok Ireng-Ireng, Bromo Tengger Semeru National Park: Before-After the COVID-19 and Eruption

The continuous eruption of the Bromo Tengger Semeru volcanic complex may have impacted the overall biodiversity in Bromo Tengger Semeru National Park, including amphibians and reptiles. On the other hand, restrictions on traveling during the COVID-19 pandemic may positively impact nature by minimizing the tourism-disturbance. This study analyzes the diversity of reptiles and amphibians before and after COVID-19 and Semeru eruption at Ranu Darungan and Blok Ireng-Ireng (utilization zone) of Bromo Tengger Semeru National Park during three continuous surveys (2019, 2021, and 2022). Preliminary sampling was conducted in August 2021, whereas extensive sampling was carried out in March–May 2022 using a visual encounter survey on two habitat conditions (terrestrial and aquatic); we used previous survey data for 2019. The continuous surveys in Ranu Darungan and Blok Ireng-Ireng showed the herpetofauna diversity consisting of 19 amphibian species (6 families) and 11 reptile species (5 families), where sampling effort in 2022 was adequate to reflect the overall herpetofauna diversity. There was a decrease in the species diversity index in Ranu Darungan after the Semeru eruption, indicating an increase in environmental stress for amphibians. However, the expected trend was not observed before and after the COVID-19 pandemic; other factors (besides reduced human activity at the site) may have contributed. Keywords: amphibian, diversity, herpetofauna, national park, reptile

Sensitive and efficient surveillance of Japanese giant salamander (Andrias japonicus) distribution in western Japan using multi-copy nuclear DNA marker

Japanese giant salamander (Andrias japonicus) is one of the largest amphibian species in the world and an iconic species in Japan. However, as its distribution has recently declined across the country, rapid and extensive monitoring of the distribution is urgently needed for its efficient conservation. Here, we used environmental DNA (eDNA) analysis to assess the Japanese giant salamander’s distribution in western Japan and, for that purpose, we collected 410 water samples from 12 rivers. We then developed a new eDNA assay for multi-copy nuclear DNA (nuDNA) of the giant salamander and compared the eDNA detectability of the nuDNA marker with that of a previous mitochondrial DNA (mtDNA) marker. Throughout the survey, we detected target eDNA from 162 water samples using either of the markers, which generally corresponded to the known natural distribution of the species. Additionally, the use of the nuDNA marker allowed for higher detection rate of target eDNA than the mtDNA marker. Moreover, the detection rate of target eDNA decreased substantially in water samples with higher conductivity and also partly in those with higher pH, suggesting their negative impacts on the salamander’s ecology. Our results demonstrated that eDNA analysis with multi-copy nuDNA marker is highly useful for efficient and sensitive surveillance of Japanese giant salamander’s distribution. Our study provided the methodology for efficiently monitoring the Japanese giant salamander’s distribution via eDNA analysis and facilitating conservation activities for them.

The cumulative niche approach: A framework to assess the performance of ecological niche model projections.

Ecological Niche Models (ENMs) are often used to project species distributions within alien ranges and in future climatic scenarios. However, ENMs depend on species-environment equilibrium, which may be absent for actively expanding species. We present a novel framework to estimate whether species have reached environmental equilibrium in their native and alien ranges. The method is based on the estimation of niche breadth with the accumulation of species occurrences. An asymptote will indicate exhaustive knowledge of the realised niches. We demonstrate the CNA framework for 26 species of mammals, amphibians, and birds. Possible outcomes of the framework include: (1) There is enough data to quantify the native and alien realised niches, allowing us to calculate niche expansion between the native and alien ranges, also indicating that ENMs can be reliably projected to new environmental conditions. (2) The data in the native range is not adequate but an asymptote is reached in the alien realised niche, indicating low confidence in our ability to evaluate niche expansion in the alien range but high confidence in model projections to new environmental conditions within the alien range. (3) There is enough data to quantify the native realised niche, but not enough knowledge about the alien realised niche, hindering the reliability of projections beyond sampled conditions. (4) Both the native and alien ranges do not reach an asymptote, and thus few robust conclusions about the species' niche or future projections can be made. Our framework can be used to detect species' environmental equilibrium in both the native and alien ranges, to quantify changes in the realised niche during the invasion processes, and to estimate the likely accuracy of model projections to new environmental conditions.

Discovery of ex situ individuals of Andrias sligoi, an extremely endangered species and one of the largest amphibians worldwide

The South China giant salamander, Andrias sligoi, is one of the largest extant amphibian species worldwide. It was recently distinguished from another Chinese species, the Chinese giant salamander, Andrias davidianus, which is considered Critically Endangered according to the International Union for Conservation of Nature (IUCN) Red List. It appears too late to save this extremely rare and large amphibian in situ. Another extant species of the same genus, Andrias japonicus, inhabits Japan. However, the introduction of Chinese giant salamanders into some areas of Japan has resulted in hybridization between the Japanese and Chinese species. During our genetic screening of giant salamanders in Japan, we unexpectedly discovered four individuals of the South China giant salamander: two were adult males in captivity, and one had recently died. The last individual was a preserved specimen. In this study, we report these extremely rare individuals of A. sligoi in Japan and discuss the taxonomic and conservational implications of these introduced individuals.

Amphibians and Reptiles Diversity in Bukit Baka Bukit Raya National Park, West Kalimantan

Studies related to herpetofauna diversity in the Bukit Baka Bukit Raya National Park (Tanakaya), West Kalimantan are very limited. Data related to the richness of amphibians and reptiles in this area are important for the management of the area in the future. We conducted amphibian and reptile surveys in Tanakaya in July-August 2019 using Visual Encounter Survey methods at five sites (Belaban Resort: Km 37 &amp; 39; Resort Rantau Malam: Batu Lintang, Sungai Mangan, and Hulu Rabang). We recorded 50 amphibian species from nine families and 25 reptile species from seven different families. Twenty of these species are new records for Tanakaya. The highest diversity (H' = 2.94) was found in Batu Lintang, while the highest evenness (E = 0.55) was in Sungai Mangan, and the highest community similarity index (IS = 0.6) was in Batu Lintang with Km 37. Combined with previous studies, there have been 142 species (78 species of amphibians and 64 species of reptiles) recorded from Tanakaya. Further study is needed to fully understand the herpetofauna in Tanakaya, but our data highlights the importance of the national park for preserving global biodiversity.

The effects of preformed vitamin A and provitamin A carotenoid supplementation on tadpoles of the poison frog Phyllobates vittatus.

Understanding the nutritional requirements of captive animals is necessary for proper animal husbandry, however, the specific dietary requirements for many amphibian species commonly kept in captivity are unknown. Like most vertebrates, frogs cannot synthesize carotenoids and must therefore obtain these essential nutrients through diet. It is unclear if amphibians can cleave provitamin A carotenoids to form vitamin A metabolically within the body, so common practice is to supplement their captive diets with both preformed vitamin A and provitamin A carotenoids. We carried out a feeding experiment in tadpoles of Phyllobates vittatus, a commonly kept poison frog species, to test the effects of supplementing a fish flake diet with a provitamin A carotenoid (2.5 mg/g β-carotene) and vitamin A (0.033-0.066 µg/mL retinyl acetate), both individually and in combination. Contrary to our expectations, supplementation had either no effect or adverse effects on tadpole growth and survivorship. Tadpoles reared under supplemented diets with vitamin A showed higher mortality rates, coupled with symptoms of hypervitaminosis A. Survivors had a smaller body size and mass at metamorphosis. β-carotene supplementation alone had no detectable effect. The vitamin A and β-carotene levels in our supplemented diet have been shown to be harmless or benefit tadpoles of other species, yet our results indicate that adding these amounts to what is found in a generalist fish flake mix can have detrimental effects on P. vittatus tadpoles. More broadly, this study highlights the importance of creating husbandry guidelines based on the specific physiological needs of the species (or species groups) being kept in captivity, rather than general ones for all amphibians, as is often done.

Anti-predator defences are linked with high levels of genetic differentiation in frogs.

Predator-prey interactions have been suggested as drivers of diversity in different lineages, and the presence of anti-predator defences in some clades is linked to higher rates of diversification. Warning signals are some of the most widespread defences in the animal world, and there is evidence of higher diversification rates in aposematic lineages. The mechanisms behind such species richness, however, are still unclear. Here, we test whether lineages that use aposematism as anti-predator defence exhibit higher levels of genetic differentiation between populations, leading to increased opportunities for divergence. We collated from the literature more than 3000 pairwise genetic differentiation values across more than 700 populations from over 60 amphibian species. We find evidence that over short geographical distances, populations of species of aposematic lineages exhibit greater genetic divergence relative to species that are not aposematic. Our results support a scenario where the use of warning signals could restrict gene flow, and suggest that anti-predator defences could impact divergence between populations and potentially have effects at a macro-evolutionary scale.

Amphibian community structure in Isau-Isau Wildlife Reserve, South Sumatra, Indonesia

Abstract. Tanjung RD, Kusrini MD, Mardiastuti A, Yustian I, Setiawan A, Iqbal M. 2023. Amphibian community structure in Isau-Isau Wildlife Reserve, South Sumatra, Indonesia. Biodiversitas 24: 6836-6843. Amphibian community structures in Isau-Isau Wildlife Reserve, South Sumatra, were investigated from Mei until July 2023. The data were obtained by Visual Encounter Survey combination with line transect. The data analyzed were Species Richness (Margalef Index), Diversity Index (H', Shannon-Wiener), Dominance (D, Simpson Index), and Evenness Index (J', Pielou) calculated using PAST 4.0 software. In addition, we also analyzed abiotic factors such as air temperature, humidity, water temperature, pH, total dissolved solids (TDS), and dissolved oxygen (DO). The abiotic data were included as explanatory variables in Canonical Correspondence Analysis (CCA). A total of 12 amphibian species consisting of 11 genera and 4 families were recorded. Of them, 11 species are classified as least concern, while one species, Theloderma licin, is classified as data deficient by the IUCN Red List of Threatened Species.

The Binding, Infection, and Promoted Growth of Batrachochytrium dendrobatidis by the Ranavirus FV3.

Increasing reports suggest the occurrence of co-infection between Ranaviruses such as Frog Virus 3 (FV3) and the chytrid fungus Batrachochytrium dendrobatidis (Bd) in various amphibian species. However, the potential direct interaction of these two pathogens has not been examined to date. In this study, we investigated whether FV3 can interact with Bd in vitro using qPCR, conventional microscopy, and immunofluorescent microscopy. Our results reveal the unexpected ability of FV3 to bind, promote aggregation, productively infect, and significantly increase Bd growth in vitro. To extend these results in vivo, we assessed the impact of FV3 on Xenopus tropicalis frogs previously infected with Bd. Consistent with in vitro results, FV3 exposure to previously Bd-infected X. tropicalis significantly increased Bd loads and decreased the host's survival.

Identification of signalling pathways involved in gill regeneration in zebrafish.

The occurrence of regeneration of the organs involved in respiratory gas exchange amongst vertebrates is heterogeneous. In some species of amphibians and fishes, the gills regenerate completely following resection or amputation, whereas in mammals, only partial, facultative regeneration of lung tissue occurs following injury. Given the homology between gills and lungs, the capacity of gill regeneration in aquatic species is of major interest in determining the underlying molecular or signalling pathways involved in respiratory organ regeneration. In the present study, we used adult zebrafish (Danio rerio) to characterize signalling pathways involved in the early stages of gill regeneration. Regeneration of the gills was induced by resection of gill filaments and observed over a period of up to 10 days. We screened for the effects on regeneration of the drugs SU5402, dorsomorphin and LY411575, which inhibit FGF, BMP or Notch signalling pathways, respectively. Exposure to each drug for 5 days significantly reduced regrowth of filament tips in regenerating tissue, compared with unresected controls. In separate experiments under normal conditions of regeneration, we used reverse transcription quantitative PCR and observed an increased expression of genes encoding for the bone morphogenetic factor, Bmp2b, fibroblast growth factor, Fgf8a, a transcriptional regulator (Her6) involved in Notch signalling, and Sonic Hedgehog (Shha), in regenerating gills at 10 day post-resection, compared with unresected controls. In situ hybridization confirmed that all four genes were expressed in regenerating gill tissue. This study implicates BMP, FGF, Notch and Shh signalling in gill regeneration in zebrafish.

Longitudinal characterization of the captive adult and tadpole Wyoming toad (Anaxyrus baxteri) microbiome

IntroductionAt one time thought to be extinct in the wild, the Wyoming toad (Anaxyrus baxteri) is one of the most critically endangered North American amphibian species. Despite approximately 20 years of ex situ breeding and reintroduction programs, this species remains functionally extinct in the wild. There is concern among those working in these programs that individuals bred in captivity fail to develop a microbiome that withstands the stressors of their native habitat following release. In related species, the skin microbiome has been shown to have a defensive function against common pathogens affecting these animals. However, the early-life microbiome of developing tadpoles in this species remains unknown and, therefore, this defensive function is unexplored in the Wyoming toad.MethodsThis study employed 16S rRNA amplicon sequencing to document the baseline microbiome of tadpole and adult Wyoming toads bred for release at Omaha’s Henry Doorly Zoo and Aquarium (Omaha, Nebraska, USA). We characterized the development of the mucosal microbiome in tadpoles and the mucosal and cloacal microbiome of adult toads.ResultsOur results revealed significant differences between tadpole and adult microbiomes, as well as significant sex-dependent differences within the adult Wyoming toads, in terms of richness and composition.DiscussionThese findings have identified the baseline microbiome of this endangered species and variables significantly influencing its composition in captivity. Ongoing studies of the only extant wild population are expected to identify taxa not found in captive toads, and potentially help 100 design husbandry modifications to maximize survivability following reintroduction to the wild.

Machine learning and phylogenetic models identify predictors of genetic variation in Neotropical amphibians

AbstractAimIntraspecific genetic variation is key for adaptation and survival in changing environments and is known to be influenced by many factors, including population size, dispersal and life‐history traits. We investigated genetic variation within Neotropical amphibian species to provide insights into how natural history traits, phylogenetic relatedness, climatic and geographic characteristics can explain intraspecific genetic diversity.LocationNeotropics.TaxonAmphibians.MethodsWe assembled data sets using open‐access databases for natural history traits, genetic sequences, phylogenetic trees, climatic and geographic data. For each species, we calculated overall nucleotide diversity (π) and tested for isolation by distance (IBD) and isolation by environment (IBE). We then identified predictors of π, IBD and IBE using random forest (RF) regression or RF classification. We also fitted phylogenetic generalized linear mixed models (PGLMMs) to predict π, IBD and IBE.ResultsWe compiled 4052 mitochondrial DNA sequences from 256 amphibian species (230 frogs and 26 salamanders), georeferencing 2477 sequences from 176 species that were not linked to occurrence data. RF regressions and PGLMMs were congruent in identifying range size and precipitation (σ) as the most important predictors of π, influencing it positively. RF classification and PGLMMs identified minimum elevation as an important predictor of IBD; most species without IBD tended to occur at higher elevations. Maximum latitude and precipitation (σ) were the best predictors of IBE, and most species without IBE occur at lower latitudes and in areas with more variable precipitation.Main ConclusionsThis study identified predictors of genetic variation in Neotropical amphibians using both machine learning and phylogenetic methods. This approach was valuable to determine which predictors were congruent between methods. We found that species with small ranges or living in zones with less variable precipitation tended to have low genetic diversity. We also showed that Western Mesoamerica, Andes and Atlantic Forest biogeographic units harbour high diversity across many species that should be prioritized for protection. These results could play a key role in the development of conservation strategies for Neotropical amphibians.

Risk from future climate change to Pakistan's protected area network: A composite analysis for hotspot identification

As climate change becomes a primary driver of global ecosystem deterioration and biodiversity loss, protected areas (PAs) are posed to play a crucial conservation role. At a global scale, 17 % of land is currently covered by PAs; a figure expected to reach 30 % by 2030 under the UN post-2020 global biodiversity framework. However, focusing only on the percent coverage of PAs without assessing their efficacy may not accomplish the intended conservation goals. Here, we present the first assessment of the risk from climate change to existing PAs and non-protected lands across Pakistan by combining data on the local exposure and vulnerability of 409 species of birds, mammals, reptiles and amphibians to multidimensional changes in climate by mid (2040–2060) and late (2061–2080) century under two climate emission scenarios (RCP4.5 and RCP8.5). We find that between 7 % (2050 RCP4.5) and 19 % (2080 RCP8.5) of the current network of PAs, mostly located in the eastern and southeastern parts of the country, are projected to be under future extreme risk (i.e., highly exposed areas containing highly vulnerable communities). Importantly, hotspots of risk within these PAs are projected to significantly expand over time and with increasing severity of the scenario. In contrast, PAs in the northern part of the country are projected to remain under moderate to low risk. Results are subject to variability across the country reflecting interesting differences in climate change exposure and species vulnerability between protected and non-protected lands. Importantly, significantly lower level of risks from future climate change are projected for PAs than non-protected lands across emission scenarios and periods suggesting potential candidate areas for the future expansion of the country's PA network. Our analysis provides novel insights that can help inform conservation decisions and management at a time when the country is investing in ambitious efforts to expand its network of protected areas.