Ongoing declines for the world's amphibians in the face of emerging threats.

Systematic assessments of species extinction risk at regular intervals are necessary for informing conservation action1,2. Ongoing developments in taxonomy, threatening processes and research further underscore the need for reassessment3,4. Here we report the findings of the second Global Amphibian Assessment, evaluating 8,011 species for the International Union for Conservation of Nature Red List of Threatened Species. We find that amphibians are the most threatened vertebrate class (40.7% of species are globally threatened). The updated Red List Index shows that the status of amphibians is deteriorating globally, particularly for salamanders and in the Neotropics. Disease and habitat loss drove 91% of status deteriorations between 1980 and 2004. Ongoing and projected climate change effects are now of increasing concern, driving 39% of status deteriorations since 2004, followed by habitat loss (37%). Although signs of species recoveries incentivize immediate conservation action, scaled-up investment is urgently needed to reverse the current trends.

Reconciling molecular phylogeny, morphological divergence and classification of Madagascan narrow-mouthed frogs (Amphibia: Microhylidae).

A recent study clarified several aspects of microhylid phylogeny by combining DNA sequences from Sanger sequencing and anchored phylogenomics, although numerous aspects of tree topology proved highly susceptible to data partition and chosen model. Although the phylogenetic results of the study were in conflict with previous publications, the authors made several changes to the taxonomy of Madagascar's cophyline microhylids. We re-analyzed part of their data together with our own molecular and morphological data. Based on a supermatrix of 11 loci, we propose a new phylogeny of the Cophylinae, and discuss it in the context of a newly generated osteological dataset. We found several sample misidentifications, partially explaining their deviant results, and propose to resurrect the genera Platypelis and Stumpffia from the synonymy of Cophyla and Rhombophryne, respectively. We provide support for the previous genus-level taxonomy of this subfamily, and erect a new genus, Anilany gen. nov., in order to eliminate paraphyly of Stumpffia and to account for the osteological differences observed among these groups. Deep nodes in our phylogeny remain poorly supported, and future works will certainly refine our classification, but we are confident that these will not produce large-scale rearrangements.

Mitochondrial introgressive hybridization following a demographic expansion in the tomato frogs of Madagascar, genus Dyscophus.

Madagascar is a biodiversity hotspot with a unique fauna and flora largely endemic at the species level and highly threatened by habitat destruction. The processes underlying population-level differentiation in Madagascar's biota are poorly understood and have been proposed to be related to Pleistocene climatic cycles, yet the levels of genetic divergence observed are often suggestive of ancient events. We combined molecular markers of different variability to assess the phylogeography of Madagascar's emblematic tomato frogs (Dyscophus guineti and D. antongilii) and interpret the observed pattern as resulting from ancient and recent processes. Our results suggest that the initial divergence between these taxa is probably old as reflected by protein-coding nuclear genes and by a strong mitochondrial differentiation of the southernmost population. Dramatic changes in their demography appear to have been triggered by the end of the last glacial period and possibly by the short return of glacial conditions known as the 8K event. This dramatic change resulted in an approximately 50-fold reduction of the effective population size in various populations of both species. We hypothesize these species' current mitochondrial DNA diversity distribution reflects a swamping of the mitochondrial genetic diversity of D. guineti by that of D. antongilii previous to the populations' bottlenecks during the Holocene, and probably as a consequence of D. antongilii demographic expansion approximately 1 million years ago. Our data support the continued recognition of D. antongilii and D. guineti as separate species and flag D. guineti as the more vulnerable species to past and probably also future environmental changes.

Malagasy Amphibian Wildlife Trade Revisited: Improving Management Knowledge of the Trade.

Madagascar is a biodiversity hotspot with a long history of trading in its wildlife, especially its hyper-diverse amphibian taxa. Due to globally raised concerns over the conservation of harvested species, CITES was introduced as a global mechanism with which to monitor and regulate the trade. Utilising data collated from the CITES Trade database, this study sought to investigate the trade and CITES' effectiveness in managing the trade with respect to Madagascar. Over a 28-year period, 20 known amphibian species were exported from Madagascar, constituting a total of nearly 271,000 individuals. Formal descriptions of Malagasy amphibian species have increased and continue to increase greatly over time. However, there was no longitudinal relationship regarding the numbers of individuals traded as new species were described. Overall, the number of individuals traded has declined over time, but where assessments were provided by the IUCN Redlist, population declines were reported in all but one species of Malagasy amphibian. Mantella (97.5%) continues to be the predominantly traded genus, with certain, high-conservation-concern, species continuing to be traded. Despite initial concerns over the effectiveness of CITES's actions, after concerted efforts, it appears that CITES' actions were having positive impacts on regulating the trade. However, going forward, concerns remain over the appropriateness of the quotas set and the robustness of their underpinning NDFs. Furthermore, with the increase in the number of recognised species, the potential for incorrect species labelling on the CITES permits increases and requires greater attention.

Spatial Ecology of an Arboreal Iguana (Oplurus cyclurus) in a Treeless Landscape.

Understanding the spatial ecology of species has important implications for conservation, as it helps identify suitable habitats and minimum requirements for biodiversity monitoring and management. The spiny-tailed lizard Oplurus cyclurus is a widespread endemic iguanid occurring in dry areas of southern and western Madagascar. While the species is known to be mostly arboreal, populations of the Isalo sandstone massif suggest local adaptation to a less forested savannah and a more exposed habitat. We radio-tracked 19 spiny-tailed lizards to investigate the species' rock-dwelling behaviour and spatial ecology at Isalo National Park. Tracked individuals showed high site and burrow fidelity, and a basking behaviour mostly tied to the accessibility of their burrow, the time of day, and their life stage. Activity peaked during the sunniest hours, while juveniles were more active than adults with unfavourable weather conditions. Despite high burrow fidelity, lizards used shelters non-exclusively, regularly changing (approx. once a week) with neighbouring burrows (average distance between burrows = 13.6 m). However, there was no obvious relation between lizards' body and/or tail size and the width and depth of selected burrows. Dynamic Brownian Bridge Movement Models estimated frequented areas over 247.8 m2 (95% isopleth), where territorial overlap is common. Our results challenge the notion that burrow-site fidelity is the sole driving factor behind space utilization in the studied population. We argue that the apparently unusual saxicolous habits imposed by habitat features (the absence of trees) may lead to local behavioural adjustments influencing antipredatory and foraging strategies, as well as intraspecific interactions.

Spatial ecology of the invasive Asian common toad in Madagascar and its implications for invasion dynamics.

Invasion dynamics are determined, among other aspects, by the spatial behaviour of invasive populations. The invasive toad Duttaphrynus melanostictus is spreading inland from the eastern coast of Madagascar, causing considerable ecological impacts. Understanding the basic factors determining the spread dynamics can inform management strategies and provide insights into spatial evolutionary processes. We radio-tracked 91 adult toads in three localities along the invasion gradient to determine whether spatial sorting of dispersive phenotypes is occurring, and investigate intrinsic and extrinsic determinants of spatial behaviour. Overall, toads in our study appeared as habitat generalists, and their sheltering behaviour was tied to water proximity, with toads changing shelter more frequently closer to waterbodies. Toads showed low displacement rates (mean = 4.12 m/day) and quite a philopatric behaviour but were able to perform daily movements of over 50 m. We did not detect any spatial sorting of dispersal-relevant traits nor sex- or size-biased dispersal. Our results suggest that toads are more likely to expand their range during the wet season, and that the range expansion is probably dominated by short-distance dispersal at this stage of the invasion, although a future increase in invasion speed is expected, due to the capacity for long-distance movements of this species.

Integrative revision of the Blommersia wittei complex, with description of a new species of frog from western and north-western Madagascar.

Frogs of the Blommersia wittei complex are widespread in western and northern Madagascar, and are one of two clades of the family Mantellidae that have colonized the Comoran island of Mayotte. Based on a comprehensive set of DNA sequences of the mitochondrial 16S rRNA gene and the nuclear-encoded RAG1 and SACS genes, integrated with morphological and bioacoustic data, we here analyze the genetic differentiation of populations of this complex across Madagascar. We confirm that a candidate species named B. sp. Ca5 in previous studies represents a genetically well-defined evolutionary lineage distributed over much of western Madagascar, which we describe herein as Blommeria bara sp. nov. based on its molecular and bioacoustic differentiation. Blommersia wittei occurs across northern Madagascar but its type locality Ambanja, at the lower Sambirano river, is very close to the range of another, newly discovered microendemic lineage that was only found at two sites along the upper Sambirano river (here named as candidate species B. sp. Ca12). The B. wittei complex thus provides an example of a clade of closely related Malagasy frogs that contains species widespread over hundreds of kilometers, as well as extreme microendemics. For a full resolution of this species complex, more data need to be collected on the geographical contact among these two lineages, on the morphology and bioacoustics of B. sp. Ca12, and on the north-eastern populations of B. wittei at Sambava, which are weakly differentiated in mitochondrial genes but differ in bioacoustics and possibly in the extent of foot webbing.

Another step through the crux: a new microendemic rock-dwelling Paroedura (Squamata, Gekkonidae) from south-central Madagascar.

Using an integrative taxonomic approach including genetic and morphological data, we formally describe a new microendemic gecko species belonging to the Paroedurabastardi clade, previously referred to as P.bastardi D. We name this taxon currently known from Anja Reserve and Tsaranoro Valley Forest (south-central Madagascar), as P.manongavatosp. nov. The new species differs from other species of the P.bastardi clade by ≥ 12.4% uncorrected p-distance at the mitochondrial 16S rRNA gene and it forms a monophyletic group in the COI mtDNA phylogenetic tree. It lacks haplotype sharing at the nuclear KIAA1239 and CMOS genes with the other species of the same complex, including the syntopic P.rennerae. Given its limited extent of occurrence and high levels of habitat fragmentation linked to forest clearances and fires, we propose the IUCN Red List Category of Critically Endangered, based on the B1ab(iii) criterion. The conservation value of Anja Reserve and Tsaranoro Valley Forest is remarkable. Preserving the remaining deciduous forest habitat is of paramount importance to protect these narrow-range reptile species.

Survived the Glaciations, Will They Survive the Fish? Allochthonous Ichthyofauna and Alpine Endemic Newts: A Road Map for a Conservation Strategy.

The Calabrian Alpine newt (Ichthyosaura alpestris inexpectata) is a glacial relict with small and extremely localised populations in the Catena Costiera (Calabria, Southern Italy) and is considered to be "Endangered" by the Italian IUCN assessment. Climate-induced habitat loss and recent fish introductions in three lakes of the Special Area of Conservation (SAC) Laghi di Fagnano threaten the subspecies' survival in the core of its restricted range. Considering these challenges, understanding the distribution and abundance of this newt is crucial. We surveyed the spatially clustered wetlands in the SAC and neighbouring areas. First, we provide the updated distribution of this subspecies, highlighting fish-invaded and fishless sites historically known to host Calabrian Alpine newt populations and two new breeding sites that have been recently colonised. Then, we provide a rough estimate of the abundance, body size and body condition of breeding adults and habitat characteristics in fish-invaded and fishless ponds. We did not detect Calabrian Alpine newts at two historically known sites now invaded by fish. Our results indicate a reduction in occupied sites and small-size populations. These observations highlight the need for future strategies, such as fish removal, the creation of alternative breeding habitats and captive breeding, to preserve this endemic taxon.

Life History Traits and Longevity of the Invasive Asian Common Toad Duttaphrynus melanostictus (Schneider, 1799) in Madagascar.

We analyzed the body length, age structure, and age at sexual maturity of the invasive Asian common toad Duttaphrynus melanostictus from different sites in Toamasina, east Madagascar. We used skeletochronology as a proxy for age estimation, while gonads were histologically analyzed to determine the age of sexual maturity. The analysis of pooled age data from three sites investigated in 2016 showed that both sexes were larger, although not older, than those of native populations. For the individuals from Madagascar, the males were significantly smaller and younger (mean ± SD, SVL: 71.4 ± 1.6 mm; age: 1.8 ± 0.7 years) than the females (SVL: 78.42 ± 1.9 mm; age: 2.7 ± 1.3 years), when the data were pooled, but when the data were analyzed separately for each of the three sites, similar results were obtained only for one site. The oldest recorded male and female were 3 and 6 years old, respectively. Gonadal histology showed that the males and females reach sexual maturity after the first and second years of age, respectively. Further studies are needed to understand if the larger size and faster growth rates observed in the invasive population of D. melanostictus in Madagascar are a consequence of more favorable environmental conditions with respect to the native range (e.g., the availability of larger trophic niches, a lack of competitors, and lower predatory pressure), and we suggest to extend the monitoring of these life history traits to understand how they might influence the invasion.

Gray versus yellow ventral coloration: Identity, distribution, color polymorphism and molecular relationships of the microhylid frog Platypelis mavomavo Andreone, Fenolio & Walvoord, 2003.

The Malagasy frog Platypelis mavomavo from Ambolokopatrika in the North East of Madagascar was originally diagnosed based on its bright yellow venter, but only limited information on this species has become available after its initial description in 2003. Several Platypelis specimens with yellow ventral color have been erroneously assigned to this species due to a lack of DNA sequences from the P. mavomavo type series. On the other hand, the candidate species Platypelis sp. Ca10 from Andranomapanga in the Northern Central East of Madagascar with gray ventral color has been defined based on its genetic differentiation from other nominal Platypelis species. Here we study the genetic variation of P. mavomavo and P. sp. Ca10 based on mitochondrial (16S rRNA) and nuclear-encoded (RAG-1) genes, including a newly determined sequence from the P. mavomavo holotype, which was studied using a museomics approach. We find only limited genetic variation among the samples studied, and this variation is unlinked to ventral coloration but instead reflects geographic distribution. We, therefore, conclude that P. sp. Ca10 is a gray-colored variant of P. mavomavo, and that P. mavomavo is rather widespread in the North East and Northern Central East of Madagascar, with populations in areas bordering the North West (Ambohitantely) and Sambirano (Ampotsidy) geographic regions, and the yellow-bellied morph restricted to the North East (Makira, Ambolokopatrika). Due to the range extension of P. mavomavo, the conservation status of the species requires re-assessment.

Vast underestimation of Madagascar's biodiversity evidenced by an integrative amphibian inventory.

Amphibians are in decline worldwide. However, their patterns of diversity, especially in the tropics, are not well understood, mainly because of incomplete information on taxonomy and distribution. We assess morphological, bioacoustic, and genetic variation of Madagascar's amphibians, one of the first near-complete taxon samplings from a biodiversity hotspot. Based on DNA sequences of 2,850 specimens sampled from over 170 localities, our analyses reveal an extreme proportion of amphibian diversity, projecting an almost 2-fold increase in species numbers from the currently described 244 species to a minimum of 373 and up to 465. This diversity is widespread geographically and across most major phylogenetic lineages except in a few previously well-studied genera, and is not restricted to morphologically cryptic clades. We classify the genealogical lineages in confirmed and unconfirmed candidate species or deeply divergent conspecific lineages based on concordance of genetic divergences with other characters. This integrative approach may be widely applicable to improve estimates of organismal diversity. Our results suggest that in Madagascar the spatial pattern of amphibian richness and endemism must be revisited, and current habitat destruction may be affecting more species than previously thought, in amphibians as well as in other animal groups. This case study suggests that worldwide tropical amphibian diversity is probably underestimated at an unprecedented level and stresses the need for integrated taxonomic surveys as a basis for prioritizing conservation efforts within biodiversity hotspots.

Comparative cytogenetics on eight Malagasy Mantellinae (Anura, Mantellidae) and a synthesis of the karyological data on the subfamily.

We performed a molecular and cytogenetic analysis on different Mantellinae species and revised the available chromosomal data on this group to provide an updated assessment of its karyological diversity and evolution. Using a fragment of the mitochondrial 16S rRNA, we performed a molecular taxonomic identification of the samples that were used for cytogenetic analyses. A comparative cytogenetic analysis, with Giemsa's staining, Ag-NOR staining and sequential C-banding + Giemsa + CMA + DAPI was performed on eight species: Gephyromantis sp. Ca19, G.striatus (Vences, Glaw, Andreone, Jesu et Schimmenti, 2002), Mantidactylus (Chonomantis) sp. Ca11, M. (Brygoomantis) alutus (Peracca, 1893), M. (Hylobatrachus) cowanii (Boulenger, 1882), Spinomantispropeaglavei "North" (Methuen et Hewitt, 1913), S.phantasticus (Glaw et Vences, 1997) and S. sp. Ca3. Gephyromantisstriatus, M. (Brygoomantis) alutus and Spinomantispropeaglavei "North" have a karyotype of 2n = 24 chromosomes while the other species show 2n = 26 chromosomes. Among the analysed species we detected differences in the number and position of telocentric elements, location of NOR loci (alternatively on the 6th, 7th or 10th pair) and in the distribution of heterochromatin, which shows species-specific patterns. Merging our data with those previously available, we propose a karyotype of 2n = 26 with all biarmed elements and loci of NORs on the 6th chromosome pair as the ancestral state in the whole family Mantellidae. From this putative ancestral condition, a reduction of chromosome number through similar tandem fusions (from 2n = 26 to 2n = 24) occurred independently in Mantidactylus Boulenger, 1895 (subgenus Brygoomantis Dubois, 1992), Spinomantis Dubois, 1992 and Gephyromantis Methuen, 1920. Similarly, a relocation of NORs, from the putative primitive configuration on the 6th chromosome, occurred independently in Gephyromantis, Blommersia Dubois, 1992, Guibemantis Dubois, 1992, Mantella Boulenger, 1882 and Spinomantis. Chromosome inversions of primitive biarmed elements likely generated a variable number of telocentric elements in Mantellanigricans Guibé, 1978 and a different number of taxa of Gephyromantis (subgenera Duboimantis Glaw et Vences, 2006 and Laurentomantis Dubois, 1980) and Mantidactylus (subgenera Brygoomantis, Chonomantis Glaw et Vences, 1994, Hylobatrachus Laurent, 1943 and Ochthomantis Glaw et Vences, 1994).

Italian natural history museums need specimen digitization and much more: a reply to Benvenuti et al.

We reply to the comments made by Benvenuti et al. (2022) about our paper on the Italian natural history museums and scientific collections and the need of a centralized hub and repository. While agreeing that digitization is a useful tool to valorize each museum and collection, we still believe that the suggestion of a centralized hub is valid and necessary. This would largely help in boosting coordination among museums, sharing personnel and resources, and in providing a place to deposit scientific collections that do not fit the scope of smaller museums.

Reconnecting research and natural history museums in Italy and the need of a national collection biorepository.

In Italy, differently from other countries, a national museum of natural history is not present. This absence is due, among other reasons, to its historical political fragmentation up to 1870, which led to the establishment of medium-sized museums, mostly managed by local administrations or universities. Moreover, a change of paradigm in biological research, at the beginning of the 20th century, contributed to privilege experimental studies in universities and facilitated the dismissal of descriptive and exploratory biology, which formed the basis of the taxonomic research carried out by natural history museums. Consequently, only a few museums have a provision of curatorial staff, space and material resources adequate to maintain their original mission of discovering the natural world, by conducting a regular research activity accompanied by field campaigns. The creation of a national research centre for the study of biodiversity, facilitating interconnections among the existing natural history museums could be a solution and is here supported, together with a centralised biorepository to host collections and vouchers, to the benefit of current and future taxonomic research and environmental conservation. Such an institution should find place and realisation within the recently proposed National Biodiversity Future Center (NBFC) planned within the National Plan of Recovery and Resilience (PNRR). Pending upon the creation of this new national centre, a network among the existing museums should coordinate their activities.

Aliens Coming by Ships: Distribution and Origins of the Ocellated Skink Populations in Peninsular Italy.

The ocellated skink (Chalcides ocellatus) is a widespread lizard, naturally distributed between the Maghreb and coastal Pakistan, with few insular populations in the Mediterranean coastal area. Some populations of this species have also been recorded in peninsular Italy, Campania and Southern Tuscany due to accidental introductions via touristic and commercial routes. In this work, we conducted genetic analyses on mitochondrial DNA COXI, cytb and 16S mtDNA genes on a sample of Italian insular and peninsular populations. Differently from what previously suggested, the nucleus in Portici (Southern Italy) may have originated from Sardinia. The intense trade and touristic traffic between Sardinia and Southern Tuscany may have been responsible for the introduction of this lizard also to Central Italy.

The Terrific Skink bite force suggests insularity as a likely driver to exceptional resource use.

Natural history museum collections hold extremely rare, extinct species often described from a single known specimen. On occasions, rediscoveries open new opportunities to understand selective forces acting on phenotypic traits. Recent rediscovery of few individuals of Bocourt´s Terrific Skink Phoboscincus bocourti, from a small and remote islet in New Caledonia allowed to genetically identify a species of land crab in its diet. To explore this further, we CT- and MRI-scanned the head of the holotype, the only preserved specimen dated to about 1870, segmented the adductor muscles of the jaw and bones, and estimated bite force through biomechanical models. These data were compared with those gathered for 332 specimens belonging to 44 other skink species. Thereafter we recorded the maximum force needed to generate mechanical failure of the exoskeleton of a crab specimen. The bite force is greater than the prey hardness, suggesting that predation on hard-shelled crabs may be an important driver of performance. The high bite force seems crucial to overcome low or seasonal variations in resource availability in these extreme insular environments. Phoboscincus bocourti appears to be an apex predator in a remote and harsh environment and the only skink known to predate on hard-shelled land crabs.