Integrative species delimitation helps to find the hidden diversity of the leaf-litter frog Ischnocnema manezinho (Garcia, 1996) (Anura, Brachycephalidae), endemic to the southern Atlantic Forest.

Background: The delimitation of cryptic species is a challenge for biodiversity conservation. Anurans show high cryptic diversity levels, and molecular species delimitation methods could help identify putative new species. Additionally, species delimitation approaches can provide important results for cryptic species conservation, with integrative methods adding robustness to results. Ischnocnema manezinho was described from Santa Catarina Island (SCI), southern Brazil. More recently, some inventories indicated continental populations supposedly similar in morphology to it. If these records are confirmed as I. manezinho, it would likely change its endangered status on National Red List, removing the species from conservation agendas. We investigated the threatened frog Ischnocnema manezinho, to evaluate if the continental populations belong to this species or if they form an undescribed species complex. Methods: We used coalescent, distance, and allele-sharing-based species delimitation methods and integrative analyses of morphometric and bioacoustics traits to test evolutionary independence between I. manezinho from SCI, Arvoredo Island, and continental populations. Results: Ischnocnema manezinho is restricted to Santa Catarina Island, while the five remaining lineages should be further investigated through a taxonomic review. Our results point to a small geographic range of Ischnocnema manezinho. Additionally, the species occurs in isolated fragments of forest in SCI surrounded by expanding urban areas, confirming its status as Endangered. Thus, the protection and monitoring of I. manezinho and the taxonomic description of the continental and Arvoredo Island candidate species should be priorities.

Tadpole of Nyctimantis galeata (Anura: Hylidae: Lophyohylini), a narrow endemic casque-headed frog from Bahia, Brazil.

Nyctimantis galeata (Pombal, Menezes, Fontes, Nunes, Rocha Van Sluys) is a casque-headed frog member of the Lophyohylini tribe (Blotto et al. 2020), narrow-endemic to the municipality of Morro do Chapéu, Bahia state, Brazil (Pombal et al. 2012). This species was initially described as sister of Corythomantis greeningi Boulenger based on shared osteological autapomorphies within Hylidae (see Faivovich et al. 2005; Pombal et al. 2012). However, in the most recent molecular phylogenetic analysis of Lophyohylini, Blotto et al. (2020) redefined the genus Nyctimantis to accommodate species from the former genus Aparasphenodon, Argenteohyla and "Corythomantis galeata" (as initially described; Pombal et al. 2012). Currently, the genus Nyctimantis comprises seven species: N. arapapa (Pimenta, Napoli Haddad), N. bokermanni (Pombal), N. brunoi (Miranda-Ribeiro), N. galeata, N. pomba (Assis, Santana, Silva, Quintela Feio), N. rugiceps Boulenger, and N. siemersi (Mertens). Among these, N. arapapa (Lourenço-de-Moraes et al. 2013), N. brunoi (Wogel et al. 2006) and N. siemersi (Céspedez 2000; Cajade et al. 2010) have their tadpoles described, while oral cavity anatomy was only reported for those of N. brunoi and N. siemersi (Wogel et al. 2006; Cajade et al. 2010). Considering that larval morphology generally provides reliable information for anuran systematic and taxonomic studies (Wassersug 1980; McDiarmid Altig 1999; Haas 2003), a formal comparison of these larvae might highlight additional non-molecular evidence that support them as congeners (Blotto et al. 2020). Herein, we describe the external morphology and oral cavity anatomy of tadpoles of N. galeata and compare it with those of related species.

Quaternary climatic fluctuations influence the demographic history of two species of sky-island endemic amphibians in the Neotropics.

We evaluated the role of Quaternary climatic fluctuations on the demographic history and population structure of amphibian species endemic to the 'campo rupestre' in the Neotropics, evaluating their distributional shifts, demographic changes, and lineage formation from the end of Pleistocene to present. We chose two anurans endemic to the high-elevation 'campo rupestre' in the Espinhaço Range (ER) in northeastern and southeastern Brazil (Bokermannohyla alvarengai and Bokermannohyla oxente), as models to test the role of Quaternary climatic fluctuations over their distribution range in this region. We collected tissue samples throughout their distribution range and used statistical phylogeography to examine processes of divergence and population demography. We generated spatial-temporal reconstructions using Bayesian inference in a coalescent framework in combination with hind-cast projections of species distribution models (SDMs). We also used the results and literature information to test alternative diversification scenarios via approximate Bayesian computation (ABC). Our results show that Quaternary climatic fluctuations influenced the geographic ranges of both species showing population expansion during the last glacial maximum (LGM) and range contraction during interglacial periods, as inferred from selected ABC models and from past projections of SDMs. We recovered Pleistocene diversification for both species occuring in distinctly unique periods for each taxon. An older and range-restricted lineage was recovered in a geographically isolated geological massif, deserving conservation and further taxonomic study. The diversification and distribution of these amphibian species endemic to the Neotropical 'campo rupestre' were influenced by Quaternary climatic fluctuations. The expansion of cold adapted species restricted to higher elevations during glacial periods and their concomitant retraction during interglacial periods may have been crucial for producing patterns of species richness and endemism along elevation gradients in tropical and subtropical domains. Such processes may influence the evolution of the biota distributed in heterogeneous landscapes with varied topography.

Genetic and Historical Colonization Analyses of an Endemic Savanna Tree, Qualea grandiflora, Reveal Ancient Connections Between Amazonian Savannas and Cerrado Core.

The evolutionary processes underlying the high diversity and endemism in the Cerrado, the most extensive Neotropical savanna, remain unclear, including the factors promoting the presence and evolution of savanna enclaves in the Amazon forest. In this study, we investigated the effects of past climate changes on genetic diversity, dynamics of species range and the historical connections between the savanna enclaves and Cerrado core for Qualea grandiflora, a tree species widely distributed in the biome. Totally, 40 populations distributed in the Cerrado core and Amazon savannas were analyzed using chloroplast and nuclear DNA sequences. We used phylogeographic, coalescent and ecological niche modeling approaches. Genetic data revealed a phylogeographic structure shaped by Pleistocene climatic oscillations. An eastern-western split in the Cerrado core was observed. The central portion of the Cerrado core harbored most of the sampled diversity for cpDNA. Ecological niche models predicted the presence of a large historical refuge in this region and multiple small refuges in peripheral areas. Relaxed Random Walk (RRW) models indicated the ancestral population in the north-western border of the central portion of the Cerrado core and cyclical dynamics of colonization related to Pleistocene climatic oscillations. Central and western ancient connections between Cerrado core and Amazonian savannas were observed. No evidence of connections among the Amazonian savannas was detected. Our study highlights the importance of Pleistocene climatic oscillations for structuring the genetic diversity of Q. grandiflora and complex evolutionary history of ecotonal areas in the Cerrado. Our results do not support the recent replacement of a large area in the Amazon forest by savanna vegetation. The Amazonian savannas appear to be fragmented and isolated from each other, evolving independently a long ago.

The larvae of two species of Bokermannohyla (Anura, Hylidae, Cophomantini) endemic to the highlands of central Brazil.

Bokermannohyla (tribe Cophomantini) is an endemic genus of Brazilian treefrogs containing 32 species arranged into four phenetic groups. The B. pseudopseudis group includes nine species, which are typically found in rupestrian ecosystems of disjunct Brazilian mountain ranges. Tadpoles have been shown to be important sources of information for the taxonomy of this genus, although careful analyses of character variation and descriptions of internal anatomy remain scarce. Since the first description of B. pseudopseudis tadpoles was based on a mixed series containing specimens of B. pseudopseudis and B. sapiranga, we describe the larvae of these two species from their type localities, including their external morphology, oral cavity, and skeletal system. Tadpoles of B. pseudopseudis are easily distinguished from those of B. sapiranga by aspects of their coloration and characters of their oral disc (more labial tooth rows, more numerous submarginal papillae and flaps with labial teeth). These morphological differences may be diagnostic for these two similar species, whereas their chondrocranium, hyobranchial apparatus, and oral cavity are alike, suggesting less variability in these character systems between closely related species. Finally, we provide the first comparison of data on the skeletal system of Bokermannohyla and other Cophomantini, highlighting characters of potential relevance to the systematics of the tribe.

Evolutionarily significant units of the critically endangered leaf frog Pithecopus ayeaye (Anura, Phyllomedusidae) are not effectively preserved by the Brazilian protected areas network.

Protected areas (PAs) are essential for biodiversity conservation, but their coverage is considered inefficient for the preservation of all species. Many species are subdivided into evolutionarily significant units (ESUs) and the effectiveness of PAs in protecting them needs to be investigated. We evaluated the usefulness of the Brazilian PAs network in protecting ESUs of the critically endangered Pithecopus ayeaye through ongoing climate change. This species occurs in a threatened mountaintop ecosystem known as campos rupestres. We used multilocus DNA sequences to delimit geographic clusters, which were further validated as ESUs with a coalescent approach. Ecological niche modeling was used to estimate spatial changes in ESUs' potential distributions, and a gap analysis was carried out to evaluate the effectiveness of the Brazilian PAs network to protect P. ayeaye in the face of climate changes. We tested the niche overlap between ESUs to gain insights for potential management alternatives for the species. Pithecopus ayeaye contains at least three ESUs isolated in distinct mountain regions, and one of them is not protected by any PA. There are no climatic niche differences between the units, and only 4% of the suitable potential area of the species is protected in present and future projections. The current PAs are not effective in preserving the intraspecific diversity of P. ayeaye in its present and future range distributions. The genetic structure of P. ayeaye could represent a typical pattern in campos rupestres endemics, which should be considered for evaluating its conservation status.