Sequencing of laser captured Z and W chromosomes of the tocantins paradoxical frog (Pseudis tocantins) provides insights on repeatome and chromosomal homology.

Pseudis tocantins is the only frog species of the hylid genus Pseudis that possesses highly heteromorphic sex chromosomes. Z and W chromosomes of Ps. tocantins differ in size, morphology, position of the nucleolar organizer region (NOR) and the amount and distribution of heterochromatin. A chromosomal inversion and heterochromatin amplification on the W chromosome were previously inferred to be involved in the evolution of this sex chromosome pair. Despite these findings, knowledge related to the molecular composition of the large heterochromatic band of this W chromosome is restricted to the PcP190 satellite DNA, and no data are available regarding the gene content of either the W or the Z chromosome of Ps. tocantins. Here, we sequenced microdissected Z and W chromosomes of this species to further resolve their molecular composition. Comparative genomic analysis suggests that Ps. tocantins sex chromosomes are likely homologous to chromosomes 4 and 10 of Xenopus tropicalis. Analyses of the repetitive DNA landscape in the Z and W assemblies allowed for the identification of several transposable elements and putative satellite DNA sequences. Finally, some transposable elements from the W assembly were found to be highly diverse and divergent from elements found elsewhere in the genome, suggesting a rapid amplification of these elements on the W chromosome.

Rapid karyotypic evolution with high diploid number variation in a rare genus of bromeligenous frogs.

Bromeligenous Crossodactylodes is a leptodactylid genus closely related to Paratelmatobius and Scythrophrys. The diploid number in all karyotyped species of these two latter genera is 24, which diverges from the modal diploid number (2n = 22) in the family. Here, we analyzed three species of Crossodactylodes and found karyotypes with 2n = 30, 2n = 32, and 2n = 36, diploid numbers that have not been reported in any other diploid leptodactylid species to date. Reconstruction of the ancestral chromosome number indicated that the diploid number changed from 22 to 24 in the common ancestor of Crossodactylodes, Paratelmatobius, and Scythrophrys, and that progressive increases in diploid number have occurred in Crossodactylodes. The large number of telocentric/subtelocentric chromosomes in karyotypes with higher diploid numbers raises the possibility that centric fissions may have occurred during the evolution of Paratelmatobiinae. Three metacentric chromosomes, probably involved in fission events, were inferred to be present in the common ancestor of all species of Crossodactylodes, but in C. bokermanni. Chromosome mapping of the satellite DNA PcP190 suggests homology between one arm of metacentric chromosome 1 of Crossodactylodes sp. 3 and telocentric chromosome 2 of C. itambe, supporting one of the presumed centric fission events.

Cytogenetics of four foam-nesting frog species of the Physalaemus gracilis group (Anura, Leptodactylidae).

Intending to increase the knowledge about cytogenetics of Physalaemus and the sparsely studied P. gracilis group, we analyzed the karyotypes of P. carrizorum, P. gracilis, P. lisei, and P. sp. aff. gracilis. We studied chromosome morphology, heterochromatin patterns, Ag-NORs location and mapped the repetitive DNA sequence PcP190. All species showed diploid karyotypes composed of 22 bi-armed chromosomes and similar C- bands and Ag-NOR patterns. C-bands were mainly centromeric and pericentromeric; non-centromeric C-bands were detected on the telomeres of pair 1 in P. lisei, although polymorphic, and interstitially on pair 10 of P. gracilis. This last character is useful to distinguish P. gracilis from its sibling species P. sp. aff. gracilis. The Ag-NOR sites were detected on the long arms of chromosome pair 8 but with a variable position among species. Clusters of PcP190 showed centromeric and pericentromeric positions coincident with conspicuous C-bands, on pairs 2 and 9 in P. gracilis and P. sp. aff. gracilis, pair 3 in P. carrizorum, and pair 7 in P. lisei. These results significantly increase the knowledge about Physalaemus cytogenetics and encourage further studies on the satellite PcP190 in other genera of Leiuperinae to better understand its taxonomic distribution and the evolutionary dynamics.

Sex chromosome evolution in frogs-helpful insights from chromosome painting in the genus Engystomops.

The differentiation of sex chromosomes is thought to be interrupted by relatively frequent sex chromosome turnover and/or occasional recombination between sex chromosomes (fountain-of-youth model) in some vertebrate groups as fishes, amphibians, and lizards. As a result, we observe the prevalence of homomorphic sex chromosomes in these groups. Here, we provide evidence for the loss of sex chromosome heteromorphism in the Amazonian frogs of the genus Engystomops, which harbors an intriguing history of sex chromosome evolution. In this species complex composed of two named species, two confirmed unnamed species, and up to three unconfirmed species, highly divergent karyotypes are present, and heteromorphic X and Y chromosomes were previously found in two species. We describe the karyotype of a lineage estimated to be the sister of all remaining Amazonian Engystomops (named Engystomops sp.) and perform chromosome painting techniques using one probe for the Y chromosome and one probe for the non-centromeric heterochromatic bands of the X chromosome of E. freibergi to compare three Engystomops karyotypes. The Y probe detected the Y chromosomes of E. freibergi and E. petersi and one homolog of chromosome pair 11 of Engystomops sp., suggesting their common evolutionary origin. The X probe showed no interspecific hybridization, revealing that X chromosome heterochromatin is strongly divergent among the studied species. In the light of the phylogenetic relationships, our data suggest that sex chromosome heteromorphism may have occurred early in the evolution of the Amazonian Engystomops and have been lost in two unnamed but confirmed candidate species.

Comparative mapping of a new repetitive DNA sequence and chromosome region-specific probes unveiling rearrangements in an Amazonian frog complex.

The frog species Physalaemus ephippifer exists in the Amazonian region and harbors heteromorphic Z and W chromosomes. A genetic lineage closely related to this species was recognized based on its mitochondrial DNA and RADseq-style markers, but its taxonomic status is still unclear and has been referred to as Lineage 1 of "P. cuvieri". The heteromorphic sex chromosomes found in P. ephippifer are not present in this lineage and which of its chromosome pairs is homologous to the sex chromosomes of P. ephippifer remain to be elucidated as well as the role of such a karyotypic divergence in the evolution of these frogs. Here, we described a new family of repetitive DNA and used its chromosomal sites along with the markers detected by a probe constructed from the microdissected segment of the Z chromosome of P. ephippifer to infer chromosomal homology. We also analyzed an unnamed species that is considered to be the sister group of the clade composed of Lineage 1 of "P. cuvieri" and P. ephippifer. Our results suggest that complex rearrangements involving the chromosomes that were inferred to be homeologous to the sex chromosomes of P. ephippifer have occurred during the divergence of this group of frogs.

Cytogenetic and genetic data support Crossodactylus aeneus Müller, 1924 as a new junior synonym of C. gaudichaudii Duméril and Bibron, 1841 (Amphibia, Anura).

The nominal anuran species Crossodactylus gaudichaudii Duméril and Bibron, 1841 and Crossodactylus aeneus Müller, 1924 are indistinguishable based on adult and larval morphology, being subject of taxonomic doubts. Here, we describe the karyotypes of C. gaudichaudii and C. aeneus, using classical and molecular cytogenetic markers. In addition, we used sequences of the H1 mitochondrial DNA to infer their phylogenetic relationships by Maximum Likelihood (ML) and Maximum Parsimony (MP) approaches and species delimitation test (by bPTP approach). The karyotypic data do not differentiate C. gaudichaudii and C. aeneus in any of the chromosome markers assessed. In both phylogenetic analyses, C. gaudichaudii and C. aeneus were recovered into a strongly supported clade. The species delimitation analysis recovered the specimens assigned to C. gaudichaudii and C. aeneus as a single taxonomic unit. Taken the cytogenetic and genetic results together with previous studies of internal and external morphology of tadpoles and biacoustic pattern, C. gaudichaudii and C. aeneus could not be differentiated, which supports the hypothesis that they correspond to the same taxonomic unit, with C. aeneus being a junior synonym of C. gaudichaudii.

Managing the Aesthetic Patient

wide variety of therapeutic options, some challenges have arisen: designing an adequate treatment plan, electing the most suitable procedures for the patient, taking into consideration the rheological characteristics of the products, considering the amount needed to achieve a natural look, but also defining the safest way to deliver the treatment. Aging of the facial structural tissue layers occurs at a different pace and the requirements for the aesthetic approach of a 30-year-old patient are different from a 60-year-old patient. A group of twelve experts in minimally invasive injectable procedures sought to identify common changes observed in different age groups in order to help in establishing a plan of treatment for patients of different ages. The individualized aesthetic plan should consider the main pillars of the aesthetic treatment -- aging process, facial assessment, and facial anatomy, to identify and systematize common changes observed in different age groups, as well as the safest and most reproducible techniques, especially for new injectors. J Drugs Dermatol. 2019;18(1):92-102.

Phylogeny of frogs from the genus Physalaemus (Anura, Leptodactylidae) inferred from mitochondrial and nuclear gene sequences.

Although some species groups have been recognized in the leiuperine genus Physalaemus, no phylogenetic analysis has previously been performed. Here, we provide a phylogenetic study based on mitochondrial and nuclear DNA sequences from 41 of the 46 species of Physalaemus. We employed the parsimony criterion using the software TNT and POY and the Bayesian criterion using the software MrBayes. Two major clades were recovered inside the monophyletic Physalaemus: (i) the highly supported Physalaemus signifer Clade, which included P. nattereri and the species previously placed in the P. deimaticus and P. signifer Groups; and (ii) the Physalaemus cuvieri Clade, which included the remaining species of Physalaemus. Five species groups were recognized in the P. cuvieri Clade: the P. biligonigerus Group, the P. cuvieri Group, the P. henselii Group, the P. gracilis Group and the P. olfersii Group. The P. gracilis Species Group was the same as that previously proposed by Nascimento et al. (2005). The P. henselii Group includes P. fernandezae and P. henselii, and was the sister group of a clade that comprised the remaining species of the P. cuvieri Clade. The P. olfersii Group included P. olfersii, P. soaresi, P. maximus, P. feioi and P. lateristriga. The P. biligonigerus Species Group was composed of P. biligonigerus, P. marmoratus, P. santafecinus and P. riograndensis. The P. cuvieri Group inferred here differed from that recognized by Nascimento et al. (2005) only by the inclusion of P. albifrons and the exclusion of P. cicada. The paraphyly of P. cuvieri with respect to P. ephippifer was inferred in all the analyses. Distinct genetic lineages were recognized among individuals currently identified as P. cuvieri and they were congruent with cytogenetic differences reported previously, supporting the hypothesis of occurrence of formally unnamed species.

Long-time evolution and highly dynamic satellite DNA in leptodactylid and hylodid frogs.

BACKGROUND: Satellite DNA sequences are the most abundant components of heterochromatin and are repeated in tandem hundreds to thousands of times in the genome. However, the number of repeats of a specific satellite family can vary even between the genomes of related species or populations. The PcP190 satellite DNA family was identified in the genome of the leptodactylid frog Physalaemus cuvieri, which showed to be derived most likely from the 5S rDNA in an ancestral species. In this study, we investigate the presence of the PcP190 satellite DNA in several P. cuvieri populations and in four closely related species at the chromosomal and molecular level. Furthermore, we investigate the occurrence of this satellite DNA in the genomes of P. marmoratus as well as in representative species of the leptodactylid genus Leptodactylus (L. latrans) and the hylodid family (Crossodactylus gaudichaudii), all with the aim of investigating if the PcP190 satellite DNA presents or not a restricted distribution. RESULTS: The PcP190 satellite DNA was detected in all the analyzed species. Some of them exhibited particular sequence differences, allowing the identification of species-specific groups of sequences, but in other species, the sequences were more conserved. However, in a general analysis, conserved and variable domains have been recognized within the PcP190 monomer. The chromosomal analysis performed on P. cuvieri populations and closely related species revealed high variability of the satellite DNA amount and its chromosomal location, which has always been coincident with regions of centromeric/pericentromeric heterochromatin. CONCLUSION: The PcP190 satellite DNA was found in representatives of two families, Leptodactylidae and Hylodidae, indicating that these sequences are widely distributed and conserved in these frogs. There is a pattern of non-random variation within the repeating units, indicating interplay between stochastic events and selective pressure along the PcP190 sequences. Karyotypic differences involving the PcP190 satellite DNA prove to be highly dynamic on the chromosomes of the Physalaemus and its differential accumulation has contributed to the differentiation process of the Z and W sex chromosomes in P. ephippifer.

Square technique-A treatment for cellulite with large size particle hyaluronic acid.

Gynoid lipodystrophy, also called cellulite, is defined as a metabolic disorder of the subcutaneous tissue that alters body contour. It is rare in men but affects 80%-90% of adult women.The authors describe a technique for the treatment of cellulite with non-traumatic subcisionTM of septa by cannula associated with large-gel-particle hyaluronic acid (HA). It uses a special marking, a safe plan and a specific product. The product being used is Subskin Aeskins, in a subcutaneous plan as no major blood vessels were found.The authors report good aesthetic results with the proposed technique. The patients reported a high degree of satisfaction.The product chosen has high G prime and large particles, which accounts for resistance to deformation and a high lifting power. Moreover, HA is a biocompatible, absorbable product and considered a safe filler. This technique allows the treatment of an area with various surface irregularities.The Square Technique proved to be a minimally invasive technique, with fast results, no major risks and little downtime, bringing satisfaction to patients and improving their self-confidence and quality of life.

Multiple contact zones and karyotypic evolution in a neotropical frog species complex.

Previous studies of DNA sequence and karyotypic data have revealed high genetic diversity in the Physalaemus cuvieri - Physalaemus ephippifer species complex-a group of small leptodactylid frogs in South America. To date, seven major genetic lineages have been recognized in this group, with species delimitation tests supporting four to seven of them as valid species. Among these, only P. ephippifer shows heteromorphic sex chromosomes, but the implications of cytogenetic divergence for the evolution of this group are unknown. We analyzed karyotypic, mitochondrial DNA, and 3RAD genomic data to characterize a putative contact zone between P. ephippifer and P. cuvieri Lineage 1, finding evidence for admixture and karyotypic evolution. We also describe preliminary evidence for admixture between two other members of this species complex-Lineage 1 and Lineage 3 of P. cuvieri. Our study sheds new light on evolutionary relationships in the P. cuvieri - P. ephippifer species complex, suggesting an important role of karyotypic divergence in its evolutionary history and underscoring the importance of hybridization as a mechanism of sex chromosome evolution in amphibians.

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.

High diversity of 5S ribosomal DNA and evidence of recombination with the satellite DNA PcP190 in frogs.

Distinct types of 5S rDNA repeats, differing by size and composition of its non-transcribed spacer (NTS), have been found in diverse taxa. Both concerted evolution and birth-and-death evolution have proven to play important roles in the evolution of the 5S rDNA family. In anurans, however, this subject has been underexplored as only a few anuran species had their 5S rDNA characterized and evolutionary analyzed to date. In the present study, we characterized the 5S rDNA sequences from species of two anuran families using classical molecular biology techniques and bioinformatic approaches. Based on the NTS, more than one type of 5S rDNA was identified in each analyzed species, which suggests that birth-and-death processes take part in the evolution of these sequences along the Anura tree of life. In addition, closely related species shared the same types of sequences, in accordance with the model of concerted evolution. We also found evidence of recombination between 5S rDNA and PcP190 satellite DNA, a repetitive sequence that is derived from the 5S rDNA. The interplay between 5S rDNA and PcP190 satellite DNA might favor the maintenance of the latter in the genome and respond to its presence in several species of frogs. The analysis of 5S rRNA transcripts confirmed the type I 5S rDNA of leiuperine as a functional 5S rRNA gene. Finally, the chromosome mapping of 5S rDNA sequences allowed some inferences of chromosome homology in Leiuperinae. In conclusion, our study provides additional information about the organization, differentiation and functionality of 5S rDNA in anuran species, revealing the potential participation of satellite DNA in the evolution of this family of rDNA.

Molecular organization and chromosomal localization of 5S rDNA in Amazonian Engystomops (Anura, Leiuperidae).

BACKGROUND: For anurans, knowledge of 5S rDNA is scarce. For Engystomops species, chromosomal homeologies are difficult to recognize due to the high level of inter- and intraspecific cytogenetic variation. In an attempt to better compare the karyotypes of the Amazonian species Engystomops freibergi and Engystomops petersi, and to extend the knowledge of 5S rDNA organization in anurans, the 5S rDNA sequences of Amazonian Engystomops species were isolated, characterized, and mapped. RESULTS: Two types of 5S rDNA, which were readily differentiated by their NTS (non-transcribed spacer) sizes and compositions, were isolated from specimens of E. freibergi from Brazil and E. petersi from two Ecuadorian localities (Puyo and Yasuní). In the E. freibergi karyotypes, the entire type I 5S rDNA repeating unit hybridized to the pericentromeric region of 3p, whereas the entire type II 5S rDNA repeating unit mapped to the distal region of 6q, suggesting a differential localization of these sequences. The type I NTS probe clearly detected the 3p pericentromeric region in the karyotypes of E. freibergi and E. petersi from Puyo and the 5p pericentromeric region in the karyotype of E. petersi from Yasuní, but no distal or interstitial signals were observed. Interestingly, this probe also detected many centromeric regions in the three karyotypes, suggesting the presence of a satellite DNA family derived from 5S rDNA. The type II NTS probe detected only distal 6q regions in the three karyotypes, corroborating the differential distribution of the two types of 5S rDNA. CONCLUSIONS: Because the 5S rDNA types found in Engystomops are related to those of Physalaemus with respect to their nucleotide sequences and chromosomal locations, their origin likely preceded the evolutionary divergence of these genera. In addition, our data indicated homeology between Chromosome 5 in E. petersi from Yasuní and Chromosomes 3 in E. freibergi and E. petersi from Puyo. In addition, the chromosomal location of the type II 5S rDNA corroborates the hypothesis that the Chromosomes 6 of E. petersi and E. freibergi are homeologous despite the great differences observed between the karyotypes of the Yasuní specimens and the others.

Dynamic ultrasound evaluation of body fillers and biostimulators in the buttocks of fresh-frozen specimen.

BACKGROUND: Ultrasound imaging is a useful tool to study the injection of fillers and biostimulators in the body. OBJECTIVE: Sonographically evaluate real-time injections of fillers and biostimulators and describe their behavior in the subcutaneous tissue of the buttocks of fresh-frozen specimen. METHODS: injection of two brands of high-density hyaluronic acid (HA), one brand of calcium hydroxyapatite (CaHA), one brand of hybrid injectable (CaHa + HA), one brand of polycaprolactone (PCL) and three brands of poly-l-lactic acid (PLLA) were performed in the subcutaneous tissue of the gluteal region under ultrasonography visualization on a fresh-frozen specimen. Videos during injection and static images immediately after injection were obtained. The sonographic aspects of the substances and their dispersion and interaction with the surrounding tissues were described. RESULTS: Real-time ultrasound showed different dispersion patterns of the two brands of HA, CaHA, hybrid injectable (CaHa + HA) and the three brands of PLLA. The sonographic aspects among the brands of PLLA were also different. CONCLUSIONS: The dynamic ultrasound evaluation of the injection of HA, CaHA, hybrid injectable (CaHa + HA), PCL and PLLA in a fresh-frozen specimen shed some light on their aspects and dispersion pattern in the buttock. If these patterns are confirmed in further studies in vivo, there will be a positive impact on the selection and development of safer and more effective techniques to enhance buttock contour.

LL body contour technique-A new way of gluteal contouring and augmentation with hyaluronic acid filler.

INTRODUCTION: Enhancing and altering body contours has increasingly been the focus of new technologies and techniques in aesthetic treatments. The buttocks are seen as a symbol of beauty, sensuality, and attraction. PATIENTS AND METHODS: The authors described a technique for female and male gluteal augmentation. It uses a special marking, a safe plan, and a specific product. The patients chosen for this technique wanted to have an augmentation or projection effect on the buttocks, but were not willing to undergo surgery. The product used was Sofiderm Subskin Aeskins. The plan of injection was the subcutaneous as no major blood vessels were found. RESULTS: The authors report good aesthetic results with the proposed technique. The patients reported a high degree of satisfaction. DISCUSSION: The product chosen has a high G prime and a larger molecule size, which accounts for resistance to deformation and a high lifting power. Thus, result can be visible in a large body area. As the anatomy between male and female buttocks are different, different techniques were used for each gender. CONCLUSION: The LL body contour technique for gluteal augmentation proved to be a minimally invasive technique, with quick results, without major risks and downtime.

Heteromorphic Z and W sex chromosomes in Physalaemus ephippifer (Steindachner, 1864) (Anura, Leiuperidae).

Heteromorphisms between sex chromosomes are rarely found in anurans and sex chromosome differentiation is considered to be a set of recent recurrent events in the evolutionary history of this group. This paper describes for the first time heteromorphic sex chromosomes Z and W in the leiuperid genus Physalaemus. They were found in P. ephippifer, a species of the P. cuvieri group, and corresponded to the eighth pair of its karyotype. The W chromosome differed from the Z chromosome by the presence of an additional segment in the short arm, composed of a distal NOR and an adjacent terminal DAPI-positive C-band. The identification of this sex chromosome pair may help in future investigations into the sex determining genes in the genus Physalaemus.

Amphibians from Serra do Cipó, Minas Gerais, Brasil. VI: A New Species of the Physalemus deimaticus Group (Anura, Leptodactylidae).

We describe a new species of Physalaemus assigned to the Physalaemus signifer Clade, and it is morphologically similar to P. rupestris, from the highlands of the Serra do Cipó in the southern Espinhaço Range, State of Minas Gerais, Southeastern Brazil. The new species is diagnosed by using the following combination of character states: presence of an arrow-shaped blotch on the dorsum of body; presence of a median longitudinal light stripe over urostyle region; belly and ventral surface of thigh marbled with dark gray irregularly shaped blotches on a bluish background; absence of reddish coloration over axillary and inguinal regions in live individuals; bright orange dots scattered over head, upper lip, and dorsum of body in live individuals; ventral surface of hand and foot red in live individuals; small size (adult male SVL=16.2-18.2 mm); presence of brown, not divided, nuptial pad in males; END/ED in males ranging from 0.85-0.93; supernumerary tubercles on foot absent; tarsal fold absent; tarsal tubercle absent; texture of posterior region of belly and ventral surface of thigh smooth; advertisement call composed of two note types (note A + B); advertisement call duration of 0.80-1.28 s; note A with ascending amplitude until mid-note then descending towards the end of the note; note B with pulses arranged in 5-7 groups; and dominant frequency of note A from 1734.4-2765.6 Hz and of note B from 1507.3-2859.4 Hz. A phylogenetic analysis based on mitochondrial DNA sequences recovered the new species nested within the Physalaemus deimaticus species group. Additionally, we redescribe the call of Physalaemus rupestris and provide a review of the geographic distribution and conservation status of the species belonging to the P. deimaticus species group.

Polytypic and polymorphic cytogenetic variations in the widespread anuran Physalaemus cuvieri (Anura, Leiuperidae) with emphasis on nucleolar organizing regions.

We investigated the NOR distribution in ten populations of Physalaemus cuvieri from different regions of Brazil and Argentina. A high variability in NOR pattern was observed and provided a useful tool in grouping several populations. The specimens from the state of Tocantins, northern Brazil, could easily be distinguished from all the other analyzed populations, since its karyotype presented NORs in the chromosome pairs 1,3,4 and 10 (and sometimes also in chromosome 5), and several pericentromeric C-bands. A NOR-site in chromosome 9 characterized three populations from the northeastern region of Brazil. Interestingly, the P. cuvieri populations located in opposite extremes of the geographic distribution had, as a fixed condition, the presence of NORs in 8q int and llp. Besides interpopulational divergences, intrapopulational variability was observed in the number of NORs, except for populations from the states of Bahia and Minas Gerais, which are boundary states respectively in the northeastern and southeastern regions of Brazil. In relation to NOR size, interindividual variations occurred in all Brazilian and Argentinean populations. Additionally, intraindividual variability in NOR size was detected in specimens from Minas Gerais. The data presented herein revealed substantial geographic polytypic variation in P. cuvieri and indicated that a taxonomic reexamination of this species is necessary.

Satellite DNA Mapping in Pseudis fusca (Hylidae, Pseudinae) Provides New Insights into Sex Chromosome Evolution in Paradoxical Frogs.

In the frog genus Pseudis, previous works found a sex-linked heteromorphism of the PcP190 satellite DNA in the nucleolus organizer region (NOR)-bearing chromosome pairs of Pseudis bolbodactyla and Pseudis tocantins, which possess a ZZ/ZW sex determination system. A pericentromeric inversion was inferred to have occurred during W chromosome evolution, moving a chromosomal cluster enriched by the PcP190 from the short arm (as observed in P. bolbodactyla) to the NOR-bearing long arm (as observed in P. tocantins). However, whether such an inversion happened in P. tocantins or in the common ancestor of Pseudis fusca and P. tocantins remained unclear. To assess this question, we mapped PcP190 in the karyotype of P. fusca from three distinct localities. Southern blotting was used to compare males and females. The mitochondrial H1 fragment (which contains the 12S ribosomal RNA (rRNA), tRNAval, and 16S rRNA genes) and cytochrome b gene were partially sequenced, and a species tree was inferred to guide our analysis. Pseudis fusca specimens were placed together as the sister group of P. tocantins, but based on genetic distance, one of the analyzed populations is probably an undescribed species. A cluster of PcP190, located in the long arm of chromosome 7, is sex linked in this putative new species but not in the remaining P. fusca. We could infer that the pericentromeric inversion that moved the PcP190 site to the NOR-bearing chromosome arm (long arm) occurred in the common ancestor of P. fusca, the putative undescribed species, and P. tocantins.