Brazilian Anurans Research Articles

Artificial intelligence enables unified analysis of historical and landscape influences on genetic diversity

While genetic variation in any species is potentially shaped by a range of processes, phylogeography and landscape genetics are largely concerned with inferring how environmental conditions and landscape features impact neutral intraspecific diversity. However, even as both disciplines have come to utilize SNP data over the last decades, analytical approaches have remained for the most part focused on either broad-scale inferences of historical processes (phylogeography) or on more localized inferences about environmental and/or landscape features (landscape genetics). Here we demonstrate that an artificial intelligence model-based analytical framework can consider both deeper historical factors and landscape-level processes in an integrated analysis. We implement this framework using data collected from two Brazilian anurans, the Brazilian sibilator frog (Leptodactylus troglodytes) and granular toad (Rhinella granulosa). Our results indicate that historical demographic processes shape most the genetic variation in the sibulator frog, while landscape processes primarily influence variation in the granular toad. The machine learning framework used here allows both historical and landscape processes to be considered equally, rather than requiring researchers to make an a priori decision about which factors are important.

A new species of Creptotrema (Allocreadiidae) in Cambeva davisi (Siluriformes) from river streams in fragments of the Atlantic Rainforest, southern Brazil.

We described Creptotrema cambeve n. sp. from the intestine of the Brazilian catfish Cambeva davisi (Haseman, 1911) (= Trichomycterus davisi) based on integrative analyses using morphological and molecular approaches. Creptotrema cambeve n. sp. closely resembles morphologically Creptotrema paraense Vicente, Santos & Souza, 1978, which was described from a siluriform fish in Northern Brazil; however, the two species differ mainly in the oral and ventral sucker sizes and the distribution of the vitellaria. The phylogenetic analyses of the 28S rDNA placed the sequences of the new species in a monophyletic clade together with all other Creptotrema spp. and as sister taxon of Creptotrema cruste Alcantara, Ebert, Ferreira-Silva, Forti, Morais, Pérez-Ponce de León & Silva, 2024, a parasite from a Brazilian anuran. Genetic divergences between the new species and other Creptotrema spp. varied from 0.2% to 4.3%, further corroborating the distinction of the new taxon. Creptotrema cambeve n. sp. is the 18th nominal Creptotrema species known from South America and the 22nd erected in the genus. This is the first study reporting a trematode in Cambeva spp. hosts and the second parasitological survey carried out for Ca. davisi, a poorly known small endemic fish.

Integrative taxonomy in the genus Rhabdias stiles et Hassall, 1905 from anuran in Brazil, description of two new species and phylogenetic analyses.

About 20 valid species of the genus Rhabdias are known in the Neotropical region. The present study aimed to describe two new species of Rhabdias parasitizing the lungs of Leptodactylus macrosternum and Leptodactylus podicipinus from Brazil. Distinctive characteristics between these species are numerous and based on body size, size of the buccal capsule, shape and size of the oesophagus, and position of the vulva. Molecular data based on ribosomal genes 28S and ITS region and mitochondrial COI of the two species are presented. Molecular analysis and comparison of the partial mitochondrial COI sequence of Rhabdias matogrossensis n. sp. and Rhabdias guaianensis n. sp. revealed a genetic divergence between these new species and the sequences of Rhabdias spp. previously deposited in GenBank. In the phylogenetic analysis, R. matogrossensis n. sp. was grouped with R. breviensis species complex, and R. guaianensis n. sp. was grouped as a sister group of R. cf. stenochepala. This study contributes to improving the diversity of known species of Rhabdias described in Brazilian anurans.

Vocal repertoire and acoustic variation in a treefrog (Boana ericae) (Amphibia, Anura, Hylidae) endemic to the Chapada dos Veadeiros, Central Brazil

ABSTRACT Despite the great diversity of Brazilian anuran species, there are still many knowledge gaps about the acoustic communication of these animals. Among the species, those classified as ‘Data Deficient’ have the greatest gaps in knowledge on their behavior. This is the case of Boana ericae, a species belonging to the Boana pulchella group, endemic to the Chapada dos Veadeiros National Park region, in the northeast of the state of Goiás, Central Brazil. Three distinct call types (A, B, and C) are currently known for B. ericae, with the type A call associated with attracting reproductive partners and the type C call presenting a territorial function. We recorded the vocalizations of 10 individuals of this species and found the presence of vocalizations composing three new note types (D, E, and F). Our results indicate that air temperature and body condition influenced the acoustic parameters of the advertisement and aggressive calls of this species. We also observed that some acoustic parameters of the advertisement calls can be used for individual recognition for showing greater variation among individuals than intra-individually. Our results add new information regarding the vocal repertoire in this species, expanding their known intra-individual classification of the acoustic parameters.

Diversity of Haemogregarine Parasites Infecting Brazilian Anurans, with a Description of New Species of Dactylosoma (Apicomplexa: Adeleorina: Dactylosomatidae).

Brazilian anurans are considered the most diverse and species rich around the world. Although in recent years there has been a strong focus on research related to this group of animals, their parasites have not received the same attention. Thus, this study aimed to provide morphological and molecular data on haemogregarines biodiversity infecting Brazilian anurans. During 2020, 116 anurans were collected from four Brazilian States and their blood and fragment of organs were screened for haemogregarine parasites. From the total, seven (6.03%) animals were found infected with species of Hepatozoon and Dactylosoma. Based on the morphological and molecular analysis, four anurans were found infected with Hepatozoon latrensis. The phylogenetic analysis has shown the isolates from this study grouping with the Brazilian anuran Hepatozoon clade, also with gene similarity ranging from 99.70 to 100% to H. latrensis isolates available on GenBank. Furthermore, three specimens (Trachycephalus typhonius, Leptodactylus latrans, and Rhinella diptycha) were infected with the same species of Dactylosoma (100% genetic similarity), with a genetic similarity of 98.56% to Dactylosoma piperis the only other species described in Brazil. In support of the molecular data, different morphological characters were observed in the blood smears as compared to D. piperis, suggesting that the species of Dactylosoma from the present study infecting three different species of Brazilian anurans is an undescribed species. Thus, this study increases the knowledge of Brazilian anuran blood parasites and demonstrates the importance of using integrative approaches for the diagnosis of haemoparasites.

New insights on the diversity of Brazilian anuran blood parasites: With the description of three new species of Hepatozoon (Apicomplexa: Hepatozoidae) from Leptodactylidae anurans.

The Amphibia are considered the most threatened vertebrate class globally, yet in Brazil they are also one of the more diverse and species rich groups. Although, in recent years there has been strong focus on amphibian related research, their parasites have not received the same attention. In Brazil, only a single species of Hepatozoon, namely H. leptodactyli (Lesage, 1908) Pessoa, 1970, has been described from anuran hosts. The present study aimed to describe three new species of Hepatozoon parasitising Leptodactylus labyrinthicus and Leptodactylus latrans from Mato Grosso State, Brazil. From 66 anurans screened for haemogregarines, four belonging to the Leptodactylidae were found positive for species of Hepatozoon. Based on the morphological analysis of peripheral blood gamonts and spleen and liver tissue meronts, three different morphotypes of Hepatozoon spp. were identified. Morphotype 1 (M1) and morphotype 2 (M2) in L. labyrinthicus and morphotype 3 (M3) in L. latrans. Molecular data based on partial 18S rDNA sequences revealed an interspecific divergence, between the species ranging from 0.43% to 1.16%. Phylogenetic analysis recovered isolates from the present study monophyletic with other isolates from Brazilian reptile and anuran hosts, sister to a clade comprising species isolated from African, North American and European reptile and anuran host species. Thus, using morphological and molecular analysis three new species infecting Brazilian Leptodactylidae anurans were identified and described. This study increases the knowledge of Brazilian anurans blood parasites and demonstrates the importance of using integrative approaches for diagnosis of hemoparasites.

Cytotoxicity potential of chemical constituents isolated and derivatised from Rhinella marina venom

Chemical compounds from skin secretions from toads of Bufonidae family have been long-studied. In the search for new molecules with pharmacological action, the 3β-OH groups of bufadienolides are commonly derivatised using acetyl groups. This work described the isolation and/or structural elucidation of isolated and derivatised compounds from the venom of the Brazilian anuran Rhinella marina, and their evaluation in in vitro assays. In the methanolic extract of the R. marina venom, compound cholesterol (1) was isolated from the CRV-52 fraction by classic column chromatography, dehydrobufotenine (2) by Sephadex LH-20 from the CRV-28 fraction, and a mix of suberoyl arginine (3) and compound 2 was obtained from the CRV-6-33 fraction. The compounds marinobufagin (4), telocionbufagin (5) and bufalin (6) were isolated by classic column chromatography, followed by separation via HPLC in the CRV-70 fraction, and the compound marinobufotoxin (9) was isolated by classic column chromatography in the CRV-6 fraction, here being isolated for the first time in R. marina specimens. Compounds 4 and 5 were submitted for acetylation with acetic anhydride, in the presence of pyridine and 4-dimethyilaminopiridine (DMAP), in order to obtain the compounds 3-acetyl-marinobufagin (7) and 3-acetyl-telocinobufogin (8). The isolated and derivatised compounds were identified by 1H and 13C NMR, and their molecular mass confirmed by mass spectrometry. All compounds (except 1 and 3) were tested in cytotoxic assays by the MTT method and presented cytotoxic potential against human cancer cell lines, as well as against non-tumoral human embryonic kidney HEK-293 cells. With the exception of compound 2, all molecules presented IC50 values < 4 μM, and none caused hemolysis of human erythrocytes, demonstrating a promising cytotoxic potential of natural and chemically-modified bufadienolides. This study presents a detailed contribution of bioactive chemicals from Brazilian Amazon Rhinella species, and indicates promising areas for further studies and pharmaceutical investments.

A new species, Dactylosoma piperis n. sp. (Apicomplexa, Dactylosomatidae), from the pepper frog Leptodactylus labyrinthicus (Anura, Leptodactylidae) from Mato Grosso State, Brazil.

The Dactylosomatidae Jakowska and Negrelli, 1955 are one of four families belonging to adeleorinid coccidia and comprise the genera Babesiosoma Jakowska and Nigrelli, 1956 and Dactylosoma Labbé, 1894. These blood protozoa occur in peripheral blood of lower vertebrates, and are commonly reported parasitising amphibians. The present study describes Dactylosoma piperis n. sp. from the pepper frog Leptodactylus labyrinthicus (Spix, 1824) (Anura: Leptodactylidae), collected in 2018 at the municipality of Araguaiana, Mato Grosso State, Brazil, based on morphology of intra-erythrocytic trophozoite, primary and secondary merogonic stages and a molecular analysis (partial 18S rDNA). Dactylosoma piperis n. sp. forms a well-supported clade with other Dactylosomatidae. This is the first molecular characterization of a species of Dactylosoma from a Brazilian anuran.

Polystoma knoffi n. sp. and Polystoma travassosi n. sp. (Monogenea: Polystomatidae): naming museum-archived specimens from Brazil.

In 1978, Kohn and co-workers deposited several polystome (Monogenea) specimens infecting several Brazilian anurans [Trachycephalus mesophaeus (Hensel), T. nigromaculatus Tschudi and Leptodactylus pentadactylus (Laurenti)] within the Helminthological Collection of the Instituto Oswaldo Cruz, Brazil. No specimen was formally described but we herein identified three morphotypes and formally describe two of them (Polystoma knoffi n. sp. and P. tavassosi n. sp.). These are respectively the 12th and 13th species of Polystoma described from South America. For both species, the intestine forms a reticulated network, a characteristic unique to most Neotropical species of Polystoma.

The advertisement calls of Brazilian anurans: Historical review, current knowledge and future directions.

Advertisement calls are often used as essential basic information in studies of animal behaviour, ecology, evolution, conservation, taxonomy or biodiversity inventories. Yet the description of this type of acoustic signals is far to be completed, especially in tropical regions, and is frequently non-standardized or limited in information, restricting the application of bioacoustics in science. Here we conducted a scientometric review of the described adverstisement calls of anuran species of Brazil, the world richest territory in anurans, to evaluate the amount, standard and trends of the knowledge on this key life-history trait and to identify gaps and directions for future research strategies. Based on our review, 607 studies have been published between 1960 to 2016 describing the calls of 719 Brazilian anuran species (68.8% of all species), a publication rate of 10.6 descriptions per year. From each of these studies, thirty-one variables were recorded and examined with descriptive and inferential statistics. In spite of an exponential rise over the last six decades in the number of studies, described calls, and quantity of published metadata, as revealed by regression models, clear shortfalls were identified with regard to anuran families, biomes, and categories of threat. More than 55% of these species belong to the two richest families, Hylidae or Leptodactylidae. The lowest percentage of species with described calls corresponds to the most diverse biomes, namely Atlantic Forest (65.1%) and Amazon (71.5%), and to the IUCN categories of threat (56.8%), relative to the less-than-threatened categories (74.3%). Moreover, only 52.3% of the species have some of its calls deposited in the main scientific sound collections. Our findings evidence remarkable knowledge gaps on advertisement calls of Brazilian anuran species, emphasizing the need of further efforts in standardizing and increasing the description of anuran calls for their application in studies of the behaviour, ecology, biogeography or taxonomy of the species.

Helminths of some tree frogs of the families Hylidae and Phyllomedusidae in an Atlantic rainforest fragment, Brazil

ABSTRACTDespite the fact that Brazil is the most-studied country regarding helminths of amphibians, only around 8% of Brazilian anurans have had at least one study made of their helminth fauna. Therefore, the aim of this study was to describe the helminth community of nine species of Atlantic Rainforest anurans of two different families, eight of them with no previous study, and six of which are endemic to Brazil. The analysed hosts for their parasites were: Dendropsophus minutus (n = 48), Scinax auratus (n = 36), D. branneri (n = 33), D. elegans (n = 26), Hypsiboas albomarginatus (n = 22), Pithecopus nordestinus (n = 19), D. decipiens (n = 12), D. haddadi (n = 11) and S. x-signatus (n = 11). A total of 781 helminths were collected from 106 (48.6%) of 218 analysed hosts, 656 Centrorhyncus sp. cystacanths and 125 nematodes: 76 adults (25 Cosmocerca sp.; three Cosmocerceidae gen. sp.; 20 Cosmocerca parva, 20 Cosmocercella phyllomedusae, five Aplectana sp., two Oswaldocruzia sp. and one Rhabdias sp.), 43 encysted larvae (nine Porrocaecum sp. and 34 Brevimulticaecum sp.) and six Physaloptera sp. larvae. Anurans may display many roles within the helminth life cycles, as they act as both predator and prey to a wide variety of animals. Consequently, we have found helminths that use anurans as definitive, intermediate and paratenic hosts. Opposing other surveys in which nematodes are the most prevalent and abundant parasites, acanthocephalans were the most prevalent taxa. These findings highlight the lack of knowledge regarding the helminth fauna of anurans and reveal many gaps with respect to their infection patterns in amphibians.

The last scream: the distress call of a probably extinct Brazilian anuran (Holoaden bradei Lutz, 1958).

The genus Holoaden Miranda-Ribeiro (Anura, Craugastoridae, Holoadeninae) includes four species endemic to the southeastern Brazilian Atlantic Forest, which occur in cloud forests at high elevations (Lutz 1958, Pombal et al. 2008, Martins & Zaher 2013). Out of these, two species are considered threatened by Ministério do Meio Ambiente (2014): H. bradei is classified as critically endangered (CR) and H. luederwaldti as endangered (EN). Holoaden bradei might be already extinct in the wild, as it has not been recorded in the last 40 years in spite of intense scientific activity within its original distribution range (Rocha & van Sluys 2004). The advertisement call has been described only for Holoaden luederwaldti (Martins 2010). Call descriptions, especially of advertisement calls, are important sources of evidence in taxonomic and phylogenetic studies (Roy 1996, Toledo et al. 2007, Andrade et al. 2016). However, there are other call types (see classification in Toledo et al. 2015) that can be used in absence of advertisement calls (e.g., Grenat & Martino 2013). We recently had access to a recording made in 1960's of the distress call of H. bradei. We hereby describe this call.

Unraveling the conservation status of Data Deficient species

Many species are classified as Data Deficient, because there is a knowledge gap about distribution range and population size and trends.This situation may be a problem in conservation, because the extinction risk of these species is unknown. In the present study, we analyzed all Brazilian anuran species classified as Data Deficient in the IUCN Red List to propose a method to know the conservation status of Data Deficient species. We used the time since species description coupled with the known current species distribution size to indicate the potential conservation status of Data Deficient species. We considered 231 Data Deficient anuran species in Brazil, in which most species (n=166 spp.) are newly described and restricted geographically (Group D). Group A (n=9 spp.) and C (n=18 spp.) were composed by species widely distributed and Group B (n=37 spp.) was composed by species described more than 50years ago and geographically restricted. Data Deficient is not a threatened category, however it indicates a need to obtain more information about the species listed, but unfortunately financial resource is limited. We suggested that the species allocated in the group B in our analyses must be priorities in future studies, because it is possible that these are threatened. Our analysis used the amphibian anuran from Brazil as model to propose some action that may be useful to known the conservation status of Data Deficient species.

Ecological correlates in Brazilian Amazonian anurans: implications for conservation

The objective of the present study is to identify the ecological characteristics of Amazonian frogs that would make them candidates for endangered species status, and to quantify correlations between body size and geographic distribution. The study included 180 species for which information was available on life history characteristics, including reproductive mode, aquatic larval development, primary larval habitat and adult microhabitat, and activity period. Species with aquatic development were classified in eight ecological groups based on a cluster analysis. Twenty-eight of the aquatic-developing species share a stream-based larval development stage with groups of endangered or vulnerable species from Australia and Central America. The Amazonian species have geographic ranges around five times larger than their Australian and Central American counterparts. The median clutch size and geographic range in aquatic-developing anuran species were 20 percent and 3.3 percent greater, respectively, than those of terrestrial-developing species. For both aquatic- and terrestrial-developing species, body size was positively associated with clutch size. Body size was also correlated with geographic range in aquatic-developing, but not in terrestrial species. Restricted geographic ranges and a lack of populations in protected areas were recorded in both groups. Most of these species occur in the southwestern Brazilian Amazon, which not only has poor coverage of protected areas, but is becoming increasingly threatened by ongoing economic development. The systematic monitoring of the region’s anurans and the creation of new conservation units should thus be of the highest priority.

Isolation and amino acid sequencing by MALDI-TOF-MS/MS of a novel antimicrobial anionic peptide from the skin secretion of Osteocephalus taurinus (Anura, Hylidae)

By means of chromatographic techniques, a novel anionic peptide, named otacidin, was isolated from the skin secretions of tree Brazilian anurans Osteocephalus taurinus. The de novo sequencing of the amino acids was performed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS/MS) analyses. The antimicrobial potential of this novel metabolite was evaluated against Escherichia coli and Pseudomonas aeruginosa, being observed minimum inhibitory concentration (MICs) of 37.4 and 78.4 mM, respectively. This work is just the second report of an antimicrobial anionic peptide from an amphibian.

Novel, panzootic and hybrid genotypes of amphibian chytridiomycosis associated with the bullfrog trade

Global amphibian declines are linked with the presence of specific, highly virulent genotypes of the emerging fungal disease chytridiomycosis caused by Batrachochytrium dendrobatidis (Bd) known as the global panzootic lineage (Bd-GPL). The global trade in amphibians for human consumption is suspected to have facilitated emergence of the disease, but evidence to support this is largely lacking. Here, we investigated the role the Lithobates catesbeianus (North American bullfrog) trade in spreading Bd genotypes by comparing strains associated with L.catesbeianus to a global panel using 36 sequenced loci from multiple chromosomal regions. Most bullfrogs were infected with Bd-GPL genotypes, but we also detected novel, highly divergent Bd genotypes (Bd-Brazil) from a live bullfrog in a US market and from native Brazilian anurans in the Atlantic Forest where bullfrogs are widely farmed. Sexual reproduction was also detected for the first time in Bd in the form of a hybrid genotype between the Bd-GPL and Bd-Brazil lineages in the Atlantic Forest. Despite the demonstration that ribosomal RNA types in Bd fail to undergo concerted evolution (over 20 sequence types may be found in a single strain), the Bd-GPL and Bd-Brazil lineages form largely separate clusters of related internal transcribed spacer (ITS) RNA sequences. Using ITS sequences, we then demonstrate the presence of Bd-Brazil in Japan, primarily on invasive L.catesbeianus. The finding that Bd is capable of sexual reproduction between panzootic and endemic genotypes emphasizes the risk of international wildlife trade as a source of additional Bd epizootics owing to hybridization.

Integrative Study of Brazilian Anurans: Geographic Distribution, Size, Environment, Taxonomy, and Conservation

AbstractMost megadiversity regions lack basic wild animal diversity information such as species lists. This absence of basic knowledge restricts the development and effectiveness of conservation action plans, which are urgently needed for animal groups such as amphibians, the most threatened vertebrate group in the world. In this study, we estimated Brazilian anuran diversity based on species range distribution shapes and evaluated factors that likely affect species lists, such as current taxonomy knowledge, current number of specimens collected in each region, biome characteristics, and anuran size (snout–vent length). We were able to demonstrate differences among the distributional patterns of species within each biome and each Brazilian federative state and among certain factors that influence their diversity, such as latitudinal variation. In this integrative study, we provide zoogeographic information about the richest (in terms of anuran biodiversity) country in the world (at least 849 described species, about 60% of which are endemic), which may be fundamental to future conservation actions in Brazil.

Interaction between breeding habitat and elevation affects prevalence but not infection intensity of Batrachochytrium dendrobatidis in Brazilian anuran assemblages

Chytridiomycosis, an infectious disease of amphibians, is caused by the fungus Batrachochytrium dendrobatidis (Bd) and has been linked to declining amphibian populations worldwide. The susceptibility of amphibians to chytridiomycosis-induced population declines is potentially influenced by many factors, including environmental characteristics, differences among host species and the growth of the pathogen itself. We investigated the effects of elevation and breeding habitat on Bd prevalence and individual infection intensity (zoospore loads) in 3 anuran assemblages of the Atlantic Coastal Forest of Brazil. Bd infection intensity was strongly influenced by elevation and breeding habitat, but we found no evidence of an interaction between those 2 variables in explaining the number of zoospores sampled from individual frogs. In contrast, Bd infection odds were predicted by elevation and by an interaction between elevation and breeding habitat, such that frogs had a higher probability of Bd infection in lotic habitats at low elevations. Our results indicate that Bd persists across a wide variety of habitats and elevations in the Atlantic Coastal Forest. Prevalence and infection intensity of Bd are highest at high elevations where overall environmental conditions for Bd are most favorable. In addition, at low elevations amphibian host habitat choice is also an important determinant of infection. Our study highlights the need to investigate interacting variables of host ecology and the environment simultaneously.

Amphibia, Anura, Hylidae Rafinesque, 1815 and Hylodidae Günther, 1858: distribution extension and new records for Santa Catarina, southern Brazil

In the present study we report new records of the anurans Hypsiboas curupi, Scinax littoralis, Dendropsophus elegans, and Crossodactylus schmidti for the state of Santa Catarina. These records expand the geographic distribution currently known for these species and contribute for the knowledge of the southern Brazilian anuran fauna.

Is the Chytrid Fungus Really Responsible for Amphibian Decline?~!2008-08-02~!2009-03-17~!2009-04-21~!

Besides other pathogens, the chytrid fungus Batrachochytrium dendrobatidis has been considered the main etiologic agent that causes chytridiomycosis and associated with amphibian die-offs. Chytrid fungus was first described in a living specimen of a Physalaemus signifer population (Anura, Leiuperidae), a common frog in natural environment in Rio de Janeiro, Brazil. The P. signifier integument is formed by a poorly keratinized epidermis supported by a dermis, which is subdivided into a spongious and compact layers. The granular and mucous glands are located in the spongy dermis. Although P. signifer showed no macroscopic lesions, microscopic analysis revealed hyperkeratosis (epidermal disruptions associated with thickening of horny layer), and some oval to spherical sporangia in the horny layer. The occurrence of chytrid fungus indicates that this pathogen is still active in Brazilian anurans and can be spreading. Considering the few sporangia in the epidermis and the strong epidermal disruption, the results suggest that the integument lesion occurs before the colonization by the chytrid fungus. Thus, the chytrid may be an opportunist fungus and not the main cause of amphibian decline.