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.

A reappraisal of Bufo levicristatus Boettger, 1885 (Anura: Bufonidae), a long forgotten toad from Paraguay.

In this work, we show that Bufo levicristatus Boettger, 1885 is a senior synonym of Bufo scitulus Caramaschi Niemeyer, 2003, and not a junior synonym of Bufo ornatus Spix, 1824, as previously considered. In addition, we present evidence that Bufo scitulus Caramaschi Niemeyer, 2003 complies with the International Code of Zoological Nomenclature requirements for a reversal of precedence over the name Bufo levicristatus Boettger, 1885, and so the name Bufo scitulus is to be maintained for this species.

A new frog of the Leptodactylus fuscus species group (Anura: Leptodactylidae), endemic from the South American Gran Chaco.

A new species of Leptodactylus frog (Anura: Leptodactylidae) from the South American Gran Chaco, morphologically similar and previously confused with the widespread Leptodactylus mystacinus, is described through the use of multiple sources of evidence (molecular, external morphology, coloration, osteology, bioacoustics, and behavior). The phylogenetic analysis with partial sequences of mitochondrial rDNA genes (12S and 16S) recovered the new species within the L. fuscus group, being highly divergent (>3% genetic distance in 16S). The new species was recovered as sister taxa of L. mystacinus, from which it is distinguished by tympanum coloration, cephalic index, dorsum and legs coloration, and some osteological differences in nasals and prevomers. This new frog is characterized by a moderate body size (SVL 46.80-66.21 mm), distinctive color pattern (reddish dorsal surfaces of body with noticeable black stripes in the dorsolateral folds), a circular and dark tympanum with dark tympanic annuli, and behavior of males that call on top of fallen logs and tree branches close to the ground.

On the identity of Bufo diptychus Cope, 1862 (Anura: Bufonidae).

The enigmatic toad Bufo diptychus was described by Cope (1862) based on a single individual (USNM 5841, now lost) of about 25 mm of SVL, collected during the expedition to La Plata River and tributaries, conducted by Captain Page between 1853 and 1856. As no dwarf species of toad was ever recorded in the surveyed area, and based on some tips that arise from Page's narrative, we postulate that the description was based on a toadlet. With this hypothesis in mind, we compared Cope's characterization of B. diptychus with juveniles of all species of Rhinella present in the region, finding an exact match in almost all characters shown by the juveniles of the common "cururú" or "rococo" toad, Rhinella schneideri (Werner 1894). Henceforth, we postulate that R. schneideri is a junior synonym of B. diptychus, under the combination Rhinella diptycha (Cope 1862).

What happened in the South American Gran Chaco? Diversification of the endemic frog genus Lepidobatrachus Budgett, 1899 (Anura: Ceratophryidae).

The Chaco is one the most neglected and least studied regions of the world. This highly-seasonal semiarid biome is an extensive continuous plain without any geographic barrier, and in spite of its high species diversity, the events and processes responsible have never been assessed. Miocene marine introgressions and Pleistocene glaciations have been mentioned as putative drivers of diversification for some groups of vertebrates in adjacent biomes of southern South America. Here we used multilocus data (one mitochondrial and six nuclear loci) from the three species of the endemic frog genus Lepidobatrachus (Lepidobatrachus asper, Lepidobatrachus laevis, and Lepidobatrachus llanensis) to determine if any of the historical events suggested as drivers of vertebrate diversification in southern South America are related to the diversification of the genus and if the Chaco is indeed a biome without barriers. Using fossil calibration in a coalescent framework we estimated that the genus diversified in the second half of the Miocene, coinciding with marine introgressions. Genetic patterns and historical demography suggest an important role of old archs and cratons as refuges during floods. In one species of the genus, L. llanensis, genetic structure reveals some breaks along the landscape, the main one of which corresponds to an area of the central Chaco that may act as a climatic barrier. Additionally, we found differential effects of the main Chacoan rivers on species of Lepidobatrachus that could be related to the time of persistence of populations in the areas influenced by these rivers.

The Paraguayan Rhinella toad venom: Implications in the traditional medicine and proliferation of breast cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Toads belonging to genus Rhinella are used in Paraguayan traditional medicine to treat cancer and skin infections. AIM OF THE STUDY: The objective of the study was to determine the composition of venoms obtained from three different Paraguayan Rhinella species, to establish the constituents of a preparation sold in the capital city of Paraguay to treat cancer as containing the toad as ingredient, to establish the effect of the most active Rhinella schneideri venom on the cell cycle using human breast cancer cells and to assess the antiprotozoal activity of the venoms. METHODS: The venom obtained from the toads parotid glands was analyzed by HPLC-MS-MS. The preparation sold in the capital city of Paraguay to treat cancer that is advertised as made using the toad was analyzed by HPLC-MS-MS. The effect of the R. schneideri venom and the preparation was investigated on human breast cancer cells. The antiprotozoal activity was evaluated on Leishmania braziliensis, L. infantum and murine macrophages. RESULTS: From the venoms of R. ornata, R. schneideri and R. scitula, some 40 compounds were identified by spectroscopic and spectrometric means. Several minor constituents are reported for the first time. The preparation sold as made from the toad did not contained bufadienolides or compounds that can be associated with the toad but plant compounds, mainly phenolics and flavonoids. The venom showed activity on human breast cancer cells and modified the cell cycle proliferation. The antiprotozoal effect was higher for the R. schneideri venom and can be related to the composition and relative ratio of constituents compared with R. ornata and R. scitula. CONCLUSIONS: The preparation sold in the capital city of Paraguay as containing the toad venom, used popularly to treat cancer did not contain the toad venom constituents. Consistent with this, this preparation was inactive on proliferation of human breast cancer cells. In contrast, the toad venoms of Rhinella species altered the cell cycle progression, affecting the proliferation of malignant cells. The findings suggest that care should be taken with the providers of the preparation and that the crude drug present a strong activity towards human breast cancer cell lines. The antiprotozoal effect of the R. schneideri venom was moderate while the venom of R. ornata was devoid of activity and that of R. scitula was active at very high concentration.

High levels of diversity uncovered in a widespread nominal taxon: continental phylogeography of the neotropical tree frog Dendropsophus minutus.

Species distributed across vast continental areas and across major biomes provide unique model systems for studies of biotic diversification, yet also constitute daunting financial, logistic and political challenges for data collection across such regions. The tree frog Dendropsophus minutus (Anura: Hylidae) is a nominal species, continentally distributed in South America, that may represent a complex of multiple species, each with a more limited distribution. To understand the spatial pattern of molecular diversity throughout the range of this species complex, we obtained DNA sequence data from two mitochondrial genes, cytochrome oxidase I (COI) and the 16S rhibosomal gene (16S) for 407 samples of D. minutus and closely related species distributed across eleven countries, effectively comprising the entire range of the group. We performed phylogenetic and spatially explicit phylogeographic analyses to assess the genetic structure of lineages and infer ancestral areas. We found 43 statistically supported, deep mitochondrial lineages, several of which may represent currently unrecognized distinct species. One major clade, containing 25 divergent lineages, includes samples from the type locality of D. minutus. We defined that clade as the D. minutus complex. The remaining lineages together with the D. minutus complex constitute the D. minutus species group. Historical analyses support an Amazonian origin for the D. minutus species group with a subsequent dispersal to eastern Brazil where the D. minutus complex originated. According to our dataset, a total of eight mtDNA lineages have ranges >100,000 km2. One of them occupies an area of almost one million km2 encompassing multiple biomes. Our results, at a spatial scale and resolution unprecedented for a Neotropical vertebrate, confirm that widespread amphibian species occur in lowland South America, yet at the same time a large proportion of cryptic diversity still remains to be discovered.