Disentangling the veil line for Brazilian biodiversity: An overview from two long-term research programs reveals huge gaps in ecological data reporting.

The lack of synthesized information regarding biodiversity is a major problem among researchers, leading to a pervasive cycle where ecologists make field campaigns to collect information that already exists and yet has not been made available for a broader audience. This problem leads to long-lasting effects in public policies such as spending money multiple times to conduct similar studies in the same area. We aim to identify this knowledge gap by synthesizing information available regarding two Brazilian long-term biodiversity programs and the metadata generated by them. Using a unique dataset containing 1904 metadata, we identified patterns of metadata distribution and intensity of research conducted in Brazil, as well as where we should concentrate research efforts in the next decades. We found that the majority of metadata were about vertebrates, followed by plants, invertebrates, and fungi. Caatinga was the biome with least metadata, and that there's still a lack of information regarding all biomes in Brazil, with none of them being sufficiently sampled. We hope that these results will have implications for broader conservation and management guiding, as well as to funding allocation programs.

Impacts of an Amazonian hydroelectric dam on frog assemblages.

About 90% of the Amazon's energy potential remains unexploited, with many large hydroelectric dams yet to be built, so it is important to understand how terrestrial vertebrates are affected by reservoir formation and habitat loss. We investigated the influence of the construction of the Santo Antônio Hydroelectric dam on the Madeira River in southwestern Amazonia on the structure of frog assemblages based on samples collected in two years before the dam flooded (pre-stage) and one (post1-stage) and four years (post2-stage) after its construction. We surveyed five 500-ha plot systems three times during each stage; in the pre-stage we sampled 19 plots in low-lying areas that would be flooded by the dam, (from now called flooded pre-stage plots) and 45 plots in terra-firme forest (from now called unflooded pre-stage plots). At the post1-stage we sampled the 45 unflooded plots and in the post2-stage we sampled the remaining 39 unflooded plots. We detected frogs by active visual and acoustic searches standardized by both time and sampling area. Few species recorded in the pre-stage flooded plots were not found in the pre-stage unflooded plots or in stages after flooding. However, the composition of frog assemblages based on relative densities in flooded pre-stage plots did not re-establish in plots on the new river margins. In unflooded areas, frog assemblages were distinct among the flooding stages with no tendency to return to the original assemblage compositions even four years after the dam was filled. For the areas that were not flooded, there was an increase in species richness in 82% of the plots between the surveys before dam construction and the first surveys after dam completion, and 65% between the pre-stage and surveys four years after dam completion. Lack of understanding by the controlling authorities of the long-term effects of landscape changes, such as water-table rises, means that studies covering appropriate periods post construction are not required in legislation, but the data from Santo Antônio indicate that changes due to dam construction are either long-term or difficult to distinguish from natural fluctuations. Future environmental-impact studies should follow strict BACI designs.

Key roles of paternal care and climate on offspring survival of an Amazonian poison frog.

In poison frogs (Dendrobatoidea), usually the males are territorial, care for terrestrial nests and later transport their offspring to waterbodies where they complete larval development. In some species, mothers care for their offspring or may exhibit flexible care to compensate for father absence. We conducted a multi-season field experiment with the Amazonian species Allobates paleovarzensis, in which it was possible to study the joint impact of paternal care and the El Niño climatic anomaly on offspring survival. The experiment consisted of two treatments: non-removal, and removal of the father from their territories. We observed that parental care was performed exclusively by the father, and none of the mothers of the 21 monitored nests transported the tadpoles. We also observed that the severe drought in a year under the influence of the El Niño event caused such a high mortality in all pre-metamorphic stages, that the role of parental care became irrelevant for offspring survival during that season. We found that pre-metamorphic Allobates paleovarzensis are highly vulnerable to the loss of paternal care. In addition, we showed that paternal care, when present, does not prevent offspring death under these increasingly frequent climatic anomalies.

Evaluation of benzocaine-based anesthetic gel in anuran skins extracts: A case study using the frog Lithodytes lineatus (Anura: Leptodactylidae).

Extracts made from the skin of dead Lithodytes lineatus frog individuals with the application of the benzocaine-based anesthetic gel, introduced into the oral cavity, were analyzed by 1H Nuclear Magnetic Resonance to investigate whether the application of this product (oral) can make studies that use extracts from the skins of these animals unfeasible. For comparison, we used skins of another species of anuran following the same death protocol. No trace of the benzocaine substance was found in the 1H-NMR spectra of the skin extracts from any of the tested anuran species. Still, using the hierarchical clustering model, it was possible to observe the formation of well-defined groups between the skin extracts of anurans and the anesthetic used to kill these animals. Our results suggest that the lethal dose of benzocaine in gel used inside the mouth of frogs may have no influence on potential results regarding the chemical composition or even bioassays using extracts made from the skin of these animals killed under this protocol since there was no detection of this substance for the analyzed samples.

Hierarchical effects of historical and environmental factors on lizard assemblages in the upper Madeira River, Brazilian Amazonia.

Investigating the role of historical and ecological factors structuring assemblages is relevant to understand mechanisms and processes affecting biodiversity across heterogeneous habitats. Considering that community assembly often involves scale-dependent processes, different spatial scales may reveal distinct factors structuring assemblages. In this study we use arboreal and leaf-litter lizard abundance data from 83 plots to investigate assemblage spatial structure at two distinct scales in southwestern Brazilian Amazonia. At a regional scale, we test the general hypothesis that the Madeira River acts as a barrier to dispersal of some lizard species, which results in distinct assemblages between river banks. At a local scale, we test the hypothesis that assemblages are not evenly distributed across heterogeneous habitats but respond to a continuum of inadequate-to-optimal portions of environmental predictors. Our results show that regional lizard assemblages are structured by the upper Madeira River acting as a regional barrier to 29.62% of the species sampled. This finding suggests species have been historically isolated at one of the river banks, or that distinct geomorphological features influence species occurrence at each river bank. At a local scale, different sets of environmental predictors affected assemblage composition between river banks or even along a river bank. These findings indicate that environmental filtering is a major cause of lizard assemblage spatial structure in the upper Madeira River, but predictor variables cannot be generalized over the extensive (nearly 500 km) study area. Based on a single study system we demonstrate that lizard assemblages along the forests near the banks of the upper Madeira River are not randomly structured but respond to multiple factors acting at different and hierarchical spatial scales.

A new nurse frog of Allobates masniger-nidicola complex (Anura, Aromobatidae) from the east bank of Tapajós River, eastern Amazonia.

The small nurse frogs of the genus Allobates (Anura, Aromobatidae) represent one of the most challenging taxonomic issues of the Neotropics. During several amphibian surveys in the Middle Tapajós River region, state of Pará, Brazil, we collected phenotypic, ecological, and molecular data on species of this genus, leading to the identification of a new species included in the Allobates masniger-nidicola complex. The new species is characterized by a large body size (snout-vent length 19.2-21.7 mm in males and 19.3-22.0 mm in females), finger III not swollen in adult males; cryptic external coloration, with dorsum uniformly ochre; a dark brown lateral stripe and a pale cream ventrolateral stripe; limbs ranging from ochre to orange; throat and chest violaceous in males and yellowish in females. The advertisement call is usually arranged in bouts of four closely spaced notes, which we term 4-pulsed units of repetition (UR), 0.317 s long on average, followed by silent intervals, and an average dominant frequency of 4.163 kHz. The new species also has exotrophic tadpoles with a unique fin morphology, which begins after the body-tail insertion and is deeper posteriorly to half of the caudal length. Sequencing of the 16S and COI regions of the mitochondrial DNA show a genetic p-distance of approximately 6-10% compared to closely related congeners. We discuss the biogeography of the new species based on phylogenetic relationships of the species within the Allobates masniger-nidicola complex and the allopatric geographic distribution in relation to sister taxa. Functional characteristics and geographic restrictions make this species particularly sensitive to the increasing human impact in eastern Amazonia.

A new species of Amazophrynella (Anura: Bufonidae) with two distinct advertisement calls.

Here we describe a new species of the toad genus Amazophrynella (Bufonidae) from the south margin of the Amazon River in Santarém, state of Pará, Brazilian Amazonia. The new species can be diagnosed by (1) medium body size for the genus: adult females 19.5-20.4 mm SVL, adult males 13.0-14.5 mm SVL; (2) snout acuminate in lateral view; (3) Finger I larger and robust; (4) Finger I nearly equal in size than Finger II; (5) palmar tubercle rounded, covering » of hand; (6) pigmented callus in Finger I in males; in life: (7) brown stain on throat and chest; (8) black dots on venter. Males emit two distinct advertisement calls: the call type 1 consists of a relatively long note (mean 0.248 s) with a mean dominant frequency of 3526 Hz. The call type 2 is arranged in bouts of 6 to 22 notes with mean dominant frequency of 3450 Hz and much shorter note durations (mean 0.034 s). In this study we also redescribe the advertisement call of A. minuta, which also emits a second call type.

Comparative cytogenetics of nine populations of the Rhinella genus (Anura, Bufonidae) with a highlight on their conservative karyotype.

The genus Rhinella is one of the most diverse groups of bufonid toads, currently composed by 93 valid species and naturally distributed throughout different Neotropical ecoregions. Here, we analyze nine Brazilian populations of toads representing species of the Rhinella margaritifera and Rhinella marina groups. These new data include the first description of the R. hoogmoedi and R. proboscidae karyotypes, as well as other taxonomically unresolved forms. Chromosomal analysis of the populations revealed pronounced chromosomal uniformity (2n=22), including the diploid number and chromosomal morphology. Three different NOR-bearing chromosomes were identified: in the subterminal region of pair 10q in R. hoogmoedi, Rhinella sp. 1 and Rhinella sp. 2, in subterminal region of 7p in R. proboscidae and Rhinella cf. margaritifera while in R. henseli and R. icterica was detected in interstitial region of 7p. Karyotypic uniformity of the genus permits the inference of interspecific chromosome homologies and evolutionary changes in the NOR-bearing chromosome may represent an informative character in species group level. The review of the cytogenetic data of the Rhinella species together with the new karyotypes reported here contributes to the understanding of the chromosomal evolution of these toads, which karyotypes are highly conserved despite the ample distribution of many forms.

A new species of Scinax from the Purus-Madeira interfluve, Brazilian Amazonia (Anura, Hylidae).

A new tree frog species of the genus Scinax from the interfluve between the Purus and Madeira rivers, Brazilian Amazonia, is described and illustrated. The new species is diagnosed by medium body size, snout truncate in dorsal view, ulnar and tarsal tubercles absent, nuptial pads poorly developed, skin on dorsum shagreen, dorsum light brown with dark brown spots and markings, white groin with black spots, anterior and posterior surfaces of thighs black, and iris bright orange. The advertisement call consists of a single short note, with 16-18 pulses and dominant frequency at 1572-1594 Hz. Tadpoles are characterized by body ovoid in dorsal view and triangular in lateral view, tail higher than body, oral disc located anteroventrally and laterally emarginated, dorsum of body uniformly grey-brown with dark brown eye-snout stripe in preservative, fins translucent with small to large irregular diffuse dark brown spots.

Conspicuousness, color resemblance, and toxicity in geographically diverging mimicry: The pan-Amazonian frog Allobates femoralis.

Predation risk is allegedly reduced in Batesian and Müllerian mimics, because their coloration resembles the conspicuous coloration of unpalatable prey. The efficacy of mimicry is thought to be affected by variation in the unpalatability of prey, the conspicuousness of the signals, and the visual system of predators that see them. Many frog species exhibit small colorful patches contrasting against an otherwise dark body. By measuring toxicity and color reflectance in a geographically variable frog species and the syntopic toxic species, we tested whether unpalatability was correlated with between-species color resemblance and whether resemblance was highest for the most conspicuous components of coloration pattern. Heterospecific resemblance in colorful patches was highest between species at the same locality, but unrelated to concomitant variation in toxicity. Surprisingly, resemblance was lower for the conspicuous femoral patches compared to the inconspicuous dorsum. By building visual models, we further tested whether resemblance was affected by the visual system of model predators. As predicted, mimic-model resemblance was higher under the visual system of simulated predators compared to no visual system at all. Our results indicate that femoral patches are aposematic signals and support a role of mimicry in driving phenotypic divergence or mimetic radiation between localities.

Evaluation of the taxonomic status of populations assigned to Phyllomedusa hypochondrialis (Anura, Hylidae, Phyllomedusinae) based on molecular, chromosomal, and morphological approach.

BACKGROUND: The taxonomic and phylogenetic relationships of the genus Phyllomedusa have been amply discussed. The marked morphological similarities among some species hamper the reliable identification of specimens and may often lead to their incorrect taxonomic classification on the sole basis of morphological traits. Phenotypic variation was observed among populations assigned to either P. azurea or P. hypochondrialis. In order to evaluate whether the variation observed in populations assigned to P. hypochondrialis is related to that in genotypes, a cytogenetic analysis was combined with phylogenetic inferences based on mitochondrial and nuclear sequences. RESULTS: The inter- and intra-population variation in the external morphology observed among the specimens analyzed in the present study do not reflect the phylogenetic relationships among populations. A monophyletic clade was recovered, grouping all the specimens identified as P. hypochondrialis and specimens assigned P. azurea from Minas Gerais state. This clade is characterized by conserved chromosomal morphology and a common C-banding pattern. Extensive variation in the nucleolar organizing region (NOR) was observed among populations, with four distinct NOR positions being recognized in the karyotypes. Intra-population polymorphism of the additional rDNA clusters observed in specimens from Barreiras, Bahia state, also highlights the marked genomic instability of the rDNA in the genome of this group. Based on the topology obtained in the phylogenetic analyses, the re-evaluation of the taxonomic status of the specimens from the southernmost population known in Brazil is recommended. CONCLUSIONS: The results of this study support the need for a thorough revision of the phenotypic features used to discriminate P. azurea and P. hypochondrialis. The phylogenetic data presented here also contribute to an extension of the geographic range of P. hypochondrialis, which is known to occur in the Amazon basin and neighboring areas of the Cerrado savanna, where it may be sympatric with P. azurea, within contact zones. The misidentification of specimens may have led to inconsistencies in the original definition of the geographic range of P. azurea. The variability observed in the NOR of P. hypochondrialis reinforces the conclusion that these sites represent hotspots of rearrangement. Intraspecific variation in the location of these sites is the result of constant rearrangements that are not detected by classical cytogenetic methods or are traits of an ancestral, polymorphic karyotype, which would not be phylogenetically informative for this group.

Comparative cytogenetic analysis of some species of the Dendropsophus microcephalus group (Anura, Hylidae) in the light of phylogenetic inferences.

BACKGROUND: Dendropsophus is a monophyletic anuran genus with a diploid number of 30 chromosomes as an important synapomorphy. However, the internal phylogenetic relationships of this genus are poorly understood. Interestingly, an intriguing interspecific variation in the telocentric chromosome number has been useful in species identification. To address certain uncertainties related to one of the species groups of Dendropsophus, the D. microcephalus group, we carried out a cytogenetic analysis combined with phylogenetic inferences based on mitochondrial sequences, which aimed to aid in the analysis of chromosomal characters. Populations of Dendropsophus nanus, Dendropsophus walfordi, Dendropsophus sanborni, Dendropsophus jimi and Dendropsophus elianeae, ranging from the extreme south to the north of Brazil, were cytogenetically compared. A mitochondrial region of the ribosomal 12S gene from these populations, as well as from 30 other species of Dendropsophus, was used for the phylogenetic inferences. Phylogenetic relationships were inferred using maximum parsimony and Bayesian analyses. RESULTS: The species D. nanus and D. walfordi exhibited identical karyotypes (2n = 30; FN = 52), with four pairs of telocentric chromosomes and a NOR located on metacentric chromosome pair 13. In all of the phylogenetic hypotheses, the paraphyly of D. nanus and D. walfordi was inferred. D. sanborni from Botucatu-SP and Torres-RS showed the same karyotype as D. jimi, with 5 pairs of telocentric chromosomes (2n = 30; FN = 50) and a terminal NOR in the long arm of the telocentric chromosome pair 12. Despite their karyotypic similarity, these species were not found to compose a monophyletic group. Finally, the phylogenetic and cytogenetic analyses did not cluster the specimens of D. elianeae according to their geographical occurrence or recognized morphotypes. CONCLUSIONS: We suggest that a taxonomic revision of the taxa D. nanus and D. walfordi is quite necessary. We also observe that the number of telocentric chromosomes is useful to distinguish among valid species in some cases, although it is unchanged in species that are not necessarily closely related phylogenetically. Therefore, inferences based on this chromosomal character must be made with caution; a proper evolutionary analysis of the karyotypic variation in Dendropsophus depends on further characterization of the telocentric chromosomes found in this group.

An integrative appraisal of the diagnosis and distribution of Allobates sumtuosus (Morales, 2002) (Anura, Aromobatidae).

We describe the advertisement calls and color in life of Allobates sumtuosus (Morales 2002) based on specimens recorded and collected at its type locality in Reserva Biológica do Rio Trombetas, Brazilian Amazonia. We also improve the species diagnosis by adding information on states of characters frequently used in current Allobates taxonomy. Finally, we analyze genetic distances and the evolutionary relationships between typical A. sumtuosus and other Allobates species distributed in Brazil and along the Guiana Shield region using a fragment of the 16S rDNA mitochondrial gene. Based on this integrative analysis, we propose the synonym of Allobates spumaponens Kok & Ernst 2007 with A. sumtuosus and provide an updated geographic distribution of the species.

Cytogenetics and sperm ultrastructure of Atelopus spumarius (Anura, Bufonidae) from the Brazilian Amazon.

The current taxonomy of most Atelopus species is based on morphological and color data only. Recent studies suggest that A. spumarius may represent a species complex assigned under the same name. Karyotypic data and description of sperm ultrastructure for 13 specimens of A. spumarius are presented here for the first time. A chromosomal analysis revealed 2n = 22 chromosomes, with centromeric heterochromatin in all pairs and a nucleolar organizer region (NOR) on the telomere of pair 7. The sperm was of the bufonoid type, presenting a filiform nucleus covered by an acrosomal complex and a mitochondrial collar in the neck region. The tail was composed of an axoneme, an undulating membrane and an axial rod. A karyotype analysis of A. spumarius showed the same chromosome number and similar chromosomal morphology as described for congeneric species, with slight differences probably resulting from pericentric inversions. The NOR location (on pair 7) was the same as that observed for species belonging to the genus Rhinella. The spermatological findings indicate a close relationship between Atelopus and the bufonoid lineage. The present data are useful for reference in future studies to determine whether more than one species are assigned to A. spumarius.

Acoustic interference and recognition space within a complex assemblage of dendrobatid frogs.

In species-rich assemblages of acoustically communicating animals, heterospecific sounds may constrain not only the evolution of signal traits but also the much less-studied signal-processing mechanisms that define the recognition space of a signal. To test the hypothesis that the recognition space is optimally designed, i.e., that it is narrower toward the species that represent the higher potential for acoustic interference, we studied an acoustic assemblage of 10 diurnally active frog species. We characterized their calls, estimated pairwise correlations in calling activity, and, to model the recognition spaces of five species, conducted playback experiments with 577 synthetic signals on 531 males. Acoustic co-occurrence was not related to multivariate distance in call parameters, suggesting a minor role for spectral or temporal segregation among species uttering similar calls. In most cases, the recognition space overlapped but was greater than the signal space, indicating that signal-processing traits do not act as strictly matched filters against sounds other than homospecific calls. Indeed, the range of the recognition space was strongly predicted by the acoustic distance to neighboring species in the signal space. Thus, our data provide compelling evidence of a role of heterospecific calls in evolutionarily shaping the frogs' recognition space within a complex acoustic assemblage without obvious concomitant effects on the signal.

Chromosome analysis of five Brazilian species of poison frogs (Anura: Dendrobatidae).

Dendrobatid frogs have undergone an extensive systematic reorganization based on recent molecular findings. The present work describes karyotypes of the Brazilian species Adelphobates castaneoticus, A. quinquevittatus, Ameerega picta, A. galactonotus and Dendrobates tinctorius which were compared to each other and with previously described related species. All karyotypes consisted of 2n = 18 chromosomes, except for A. picta which had 2n = 24. The karyotypes of the Adelphobates and D. tinctorius species were highly similar to each other and to the other 2n = 18 previously studied species, revealing conserved karyotypic characteristics in both genera. In recent phylogenetic studies, all Adelphobates species were grouped in a clade separated from the Dendrobates species. Thus, we hypothesized that their common karyotypic traits may have a distinct origin by chromosome rearrangements and mutations. In A. picta, with 2n = 24, chromosome features of pairs from 1 to 8 are shared with other previously karyotyped species within this genus. Hence, the A. picta data reinforced that the C-banding pattern and the NOR location are species-specific traits in the genus Ameerega. Moreover, the Ameerega monophyletism proposed by previous phylogenetic studies indicates that the karyotypic differences among species in this genus result from a long divergence time.

Masking interference and the evolution of the acoustic communication system in the Amazonian dendrobatid frog Allobates femoralis.

The efficacy of communication relies on detection of species-specific signals against the background noise. Features affecting signal detection are thus expected to evolve under selective pressures represented by masking noise. Spectral partitioning between the auditory signals of co-occurring species has been interpreted as the outcome of the selective effects of masking interference. However, masking interference depends not only on signal's frequency but on receiver's range of frequency sensitivity; moreover, selection on signal frequency can be confounded by selection on body size, because these traits are often correlated. To know whether geographic variation in communication traits agrees with predictions about masking interference effects, we tested the hypothesis that variation in the male-male communication system of the Amazonian frog, Allobates femoralis, is correlated with the occurrence of a single species calling within an overlapping frequency range, Epipedobates trivittatus. We studied frogs at eight sites, four where both species co-occur and four where A. femoralis occurs but E. trivittatus does not. To study the sender component of the communication system of A. femoralis and to describe the use of the spectral range, we analyzed the signal's spectral features of all coactive species at each site. To study the receiver component, we derived frequency-response curves from playback experiments conducted on territorial males of A. femoralis under natural conditions. Most geographic variation in studied traits was correlated with either call frequency or with response frequency range. The occurrence of E. trivittatus significantly predicted narrower and asymmetric frequency-response curves in A. femoralis, without concomitant differences in the call or in body size. The number of acoustically coactive species did not significantly predict variation in any of the studied traits. Our results strongly support that the receiver but not the sender component of the communication system changed due to masking interference by a single species.

Chromosomal study of Colostethus brunneus from the type locality and two related species (Anura--Dendrobatidae).

In this paper, we report a chromosomal study of three Brazilian species of Colostethus, C. brunneus from the type locality, Colostethus sp. (aff. trilineatus), and Colostethus sp., which is morphologically similar to C. brunneus. The diploid number for C. brunneus was 2n = 24 chromosomes, in agreement with that previously described for specimens from Peru. Colostethus sp. (aff. trilineatus) and Colostethus sp. showed a very similar karyotype with 22 chromosomes. The NOR was located on pair 3 in C. brunneus, on pair 4 in Colostethus sp. (aff. trilineatus), and on pair 2 in Colostethus sp. In one specimen of Colostethus sp., an additional NOR site was located on pair 7 in only one of the homologs. This extra Ag-NOR site was confirmed by FISH using an rDNA probe. In addition to the NOR location, the C-banding pattern was also species-specific, despite the similar chromosomal morphology of the species. These results indicate that although these species may be closely related, there is a clear dichotomy in their chromosome number.

Chromosomal differentiation of populations of Lysapsus limellus limellus, L. l. bolivianus, and of Lysapsus caraya (Hylinae, Hylidae).

Cytogenetic analysis were done on specimens from two populations of Lysapsus limellus limellus, three of L. l. bolivianus and of one of Lysapsus caraya. All animals showed a diploid chromosomal number of 2n=24. The karyotypes of the two L. limellus subspecies were very similar, differing only by the larger amount of telomeric heterochromatin and a small pericentromeric C-band on the short arms of pair 2 in L. l. limellus specimens. The karyotype of L. caraya differed from those of the two L. limellus subspecies in terms of chromosomal morphology, C-banding pattern and location of the main NOR on chromosomes 7 and 6, respectively. The karyotype of the L. l. bolivianus population from Guajará-Mirim/RO differed from those of the other populations of the same subspecies in morphology and heterochromatin pattern of chromosomes 7 and 8. Additional NORs were detected by silver staining and confirmed by FISH in one of the homologues of pairs 1 and 8 in L. l. bolivianus and in pair 7 in L. caraya. These results suggest that a reassessment of the taxonomic status of L. limellus subspecies, especially of the L. l. bolivianus populations, may be necessary.