Cytogenetic studies in six species of Scinax (Anura,Hylidae) clade Scinax ruber from northern and northeastern Brazil

Scinax species are still underrepresented in cytogenetic studies, mainly with respect to populations from northeastern and northern Brazil. In this study, we provide new chromosomal information on Scinax boesemani, S. camposseabrai, S. garbei, S. pachycrus, S. trilineatus and S. x-signatus, all belonging to clade S. ruber. They were collected at two locations in the Caatinga biome (northeastern Brazil) and at one in the Amazon (northern Brazil) biomes. Chromosomes were analyzed by conventional staining, C-banding, Ag-NOR staining, and fluorochrome staining. All species shared a modal diploid value of 2n = 24 and fundamental arm number (FN) of 48. Moreover, both chromosomal size and morphology were similar to other species in this Scinaxclade. C-banding revealed centromeric heterochromatin in all species, along with terminal species-specific C-bands in some species. Active nucleolar organizer regions (Ag-NORs) were identified at 11q in most species, except for S. boesemani and S. garbei (Ag-NORs at interstitial region of 8q). Differing from most anurans, GC-rich regions were not restricted to NORs, but also coincident with some centromeric and terminal C-bands. These data contribute to the cytotaxonomy of Scinax by providing chromosomal markers and demonstrating the occurrence of microstructural rearrangements and inversions on chromosomal evolution of Scinax.     

DNA barcode reveals candidate species of Scinax and Ololygon (Anura: Hylidae) in Atlantic Forest

Molecular species delimitation methods are efficient tools to identify species, including the discovery of new taxa and cryptic organisms, thus being useful to biodiversity studies. In the present work, 16S mitochondrial sequences and cytochrome oxidase I (COI) were used to evaluate the richness of species in the genus Scinax and Ololygon from a biodiversity hotspot in Atlantic Forest. A total of 109 specimens formally belonging to eight species of Scinax and three species of Ololygon were collected in 13 localities along the state of Bahia (northeastern Brazil) and one site in Espírito Santo (southeastern Brazil). Of the Scinax species collected in this study, three were morphologically differentiated from other described species and identified as putative new species (Scinax sp.1, Scinax sp.2 and Scinax sp.3). The species delimitations were inferred using three different methods: ABGD, PTP and mPTP which allowed recognizing 11 Scinax species and five Ololygon species. Scinax sp. 1, Scinax sp. 2 and Scinax sp. 3, have been confirmed as new putative species and Ololygon argyreornata possibly contains cryptic species. We suggest additional studies, including morphological and bioacoustic data to validate these new putative species.     

The hyolaryngeal apparatus of three species of Scinax (Scinaxinae: Hylidae: Anura) and comparison with other Hylidae species

The hyolaryngeal apparatus includes the arytenoid, cricoid and hyoid cartilages, and varied sizes and shapes can be distinguished between species. Although the larynx is an important morphological structure for sound production in frogs, few studies describe the structures and processes of its parts in hylids. We studied the hyolaryngeal anatomy of both males and females of Scinax ruber (Laurenti, 1768), using clearing and double staining methods, comparing it with that of Scinax wandae (Pyburn and Fouquette, 1971), Scinax kennedyi (Pyburn, 1973) and other hylid species. We found that S. ruber has the largest arytenoid cartilage of any species of the subfamily Scinaxinae studied to date. We described both the laryngeal differences among three Scinax species and the interspecific variability in the shapes of the projections of the arytenoid in the males of these species. The taxonomic characters (presence/absence of processes of the cartilage of Santorini, oesophageal process or bronchial processes) described here can play a key role in differentiating these species from Scinax species and from other species. In addition, we want to contribute to the natural history and comparison of the larynges in Scinax and within the genus and the Hylidae.     

The tadpole of Scinax aromothyella (Anura: Hylidae) from Uruguay

We describe the tadpole of Scinax aromothyella from the surroundings of Quebrada de los Cuervos, Departamento de Treinta y Tres, Uruguay. It shares with other larvae of the Scinax catharinae clade the slightly depressed body, a dextral vent tube that reaches the free margin of the ventral fin, and the oral disc with a dorsal gap in the marginal row of papillae and with a concave posterior edge. It is most similar to the tadpole of Scinax berthae, from which it differs in the larger total length of S. aromothyella.

Describimos la larva de Scinax aromothyella de los alrededores de la Quebrada de los Cuervos, Departamento de Treinta y Tres, Uruguay. Comparte con las larvas de otras especies del clado de Scinax catharinae el tener el cuerpo levemente deprimido, el tubo proctodeal dextro que alcanza el borde libre de la aleta ventral, y el disco oral con un claro rostral en la hilera de papilas marginales y con su borde posterior cóncavo. Es muy parecida a la larva de Scinax berthae, de la que difiere en la mayor longitud total de S. aromothyella.     

High Species Richness of Scinax Treefrogs (Hylidae) in a Threatened Amazonian Landscape Revealed by an Integrative Approach

Rising habitat loss is one of the main drivers of the global amphibian decline. Nevertheless, knowledge of amphibian diversity needed for effective habitat protection is still highly inadequate in remote tropical regions, the greater part of the Amazonia. In this study we integrated molecular, morphological and bioacoustic evidence to evaluate the species richness of the treefrogs genus Scinax over a 1000 km transect across rainforest of the Purus-Madeira interfluve, and along the east bank of the upper Madeira river, Brazilian Amazonia. Analysis revealed that 82% of the regional species richness of Scinax is still undescribed; two nominal species, seven confirmed candidate species, two unconfirmed candidate species, and one deep conspecific lineage were detected in the study area. DNA barcoding based analysis of the 16s rRNA gene indicates possible existence of three discrete species groups within the genus Scinax, in addition to the already-known S. rostratus species Group. Quantifying and characterizing the number of undescribed Scinax taxa on a regional scale, we provide a framework for future taxonomic study in Amazonia. These findings indicate that the level to which Amazonian anura species richness has been underestimated is far greater than expected. Consequently, special attention should be paid both to taxonomic studies and protection of the still-neglected Amazonian Scinax treefrogs.     

Molecular and Morphological Evidence Reveals a New Species in the Phyllomedusa hypochondrialis Group (Hylidae,Phyllomedusinae) from the Atlantic Forest of the Highlands of Southern Brazil

The taxonomic status of a disjunctive population of Phyllomedusa from southern Brazil was diagnosed using molecular, chromosomal, and morphological approaches, which resulted in the recognition of a new species of the P. hypochondrialis group. Here, we describe P. rustica sp. n. from the Atlantic Forest biome, found in natural highland grassland formations on a plateau in the south of Brazil. Phylogenetic inferences placed P. rustica sp. n. in a subclade that includes P. rhodei + all the highland species of the clade. Chromosomal morphology is conservative, supporting the inference of homologies among the karyotypes of the species of this genus. Phyllomedusa rustica is apparently restricted to its type-locality, and we discuss the potential impact on the strategies applied to the conservation of the natural grassland formations found within the Brazilian Atlantic Forest biome in southern Brazil. We suggest that conservation strategies should be modified to guarantee the preservation of this species.     

The Subfamily Cytheropterinae Hanai, 1957 (Subphylum Crustacea,Class Ostracoda) from the Southern Brazilian Continental Shelf

The study of the subfamily Cytheropterinae from the southern Brazilian continental shelf reveals the presence of four genera and nine species, including two newly described (Cytheropteron sudatlanticum sp. nov., Loxoreticulatum pulchrum sp. nov.) and two left in open nomenclature (Cytheropteron sp. and Kangarina sp.). All of these species are restricted to the southern part of the Southwest Atlantic Ocean and do not extend beyond the coast of Rio de Janeiro, Brazil. Three of these species (Oculocytheropteron micropunctatum, O. reticulatum, O. macropunctatum) extend beyond the southern limit of the Brazilian Province (36°S), while the others occur inside the limits of the Brazilian Province (36°S–22/21°S). The genus Loxoreticulatum, a well-known ostracod taxon from the Antarctic Peninsula, southern Argentina and Falkland (= Malvinas) Islands, is herein reported for the first time from offshore Brazil.     

A taxonomic contribution to the genus Dolichomitus Smith (Hymenoptera,Ichneumonidae, Pimplinae) from?Brazil

In the present study, two new species of Pimplinae, Dolichomitus jatai sp. n. and Dolichomitus moacyri sp. n. are described, and the distribution range of Dolichomitus annulicornis (Cameron, 1886) is extended. The specimens were collected using Malaise traps in areas of Atlantic forest and Brazilian savannah (cerrado) in southeastern Brazil and are deposited in a Brazilian collection (DCBU).     

The phylogenetic position of the Loimoidae Price, 1936 (Monogenoidea: Monocotylidea) based on analyses of partial rDNA sequences and morphological data

Phylogenetic analyses of partial sequences of 18S and 28S rDNA of some monogenoids, including monocotylids and a specimen of Loimosina sp. collected from a hammerhead shark off Brazil, indicated that the Loimoidae (as represented by the specimen of Loimosina sp.) represents an in-group taxon of the Monocotylidae. In all analyses, the Loimoidae fell within a major monocotylid clade including species of the Heterocotylinae, Decacotylinae, and Monocotylinae. The Loimoidae formed a terminal clade with two heterocotyline species, Troglocephalus rhinobatidis and Neoheterocotyle rhinobatis, for which it represented the sister taxon. The following morphological characters supported the clade comprising the Loimoidae, Heterocotylinae, Decacotylinae and Monocotylinae: single vagina present, presence of a narrow deep anchor root, and presence of a marginal haptoral membrane. The presence of cephalic pits was identified as a putative synapomorphy for the clade (Loimoidae (T. rhinobatidis, N. rhinobatis)). Although rDNA sequence data support the rejection of the Loimoidae and incorporating its species into the Monocotylidae, this action was not recommended pending a full phylogenetic analysis of morphological data.     

Zooxanthellae genotypes in the coral Siderastrea stellata from coastal reefs in northeastern Brazil

Siderastrea stellata is a common scleractinian coral that inhabits shallow reefs off the coast of Brazil. This species is considered to be very resistant to temperature and salinity variations and water turbidity, demonstrating great ecological plasticity and adaptability to environmental changes. Samples of S. stellata were taken from the Cabo Branco coastal reefs near João Pessoa, Brazil, every month for two years and analyzed using PCR and Restriction Fragment Length Polymorphisms of SSU rDNA techniques. The data indicated that during the study period S. stellata hosted only one SSU rDNA genotype of Symbiodinium with the RFLP pattern of clade C. The presence of clade C zooxanthellae in S. stellata in northeastern Brazilian reefs shows the wide geographical distribution of this clade, and it may aid bleaching recovery in S. stellata. Furthermore, the association of S. stellata with a zooxanthellae clade considered to be one of most resistant to bleaching may help to explain the high ecological plasticity of this scleractinian species, its capacity to reverse bleaching, and its high resistance and resilience to environmental disturbances.     

Taxonomic review of Scinax fuscomarginatus (Lutz, 1925) and related species (Anura; Hylidae)

Scinax fuscomarginatus, Scinax parkeri, Scinax trilineatus, Scinax lutzorum, and Scinax pusillus are morphologically similar species with controversial taxonomy associated with open formations of South America east of the Andes. We used external morphology, advertisement call, and molecular sequences to assess the taxonomy of these species. Phylogenetic analysis of molecular data showed a well‐supported monophyletic group divided into two main clades and several subclades. Specimens of Hyla madeirae (synonym of S. fuscomarginatus) and those from Serra do Cachimbo correspond to distinct subclades, whereas the currently valid species were grouped into a single clade. This clade showed a marked substructure, but concordance of the recognized species to subclades was limited, showing S. fuscomarginatus to be paraphyletic with respect to S. parkeri, S. trilineatus, S. lutzorum, and S. pusillus. The high intrapopulation and intraspecific variation found in morphological characters did not allow us to differentiate amongst the currently recognized species. Similarly, none of them was distinguishable by morphometric analysis or advertisement call. Based on multiple evidence, we propose the synonymy of S. parkeri, S. trilineatus, S. lutzorum, and S. pusillus with S. fuscomarginatus, the revalidation of H. madeirae, and describe the specimens from Serra do Cachimbo as a new species. © 2014 The Linnean Society of London     

Phylogenetic relationships of Ochoterenatrema Caballero, 1943 (Digenea: Lecithodendriidae) with descriptions of two new species

Ochoterenatrema is a small genus of lecithodendriid digeneans that includes six species parasitic in Neotropical bats in North and South America. Members of this genus are characterized by the presence of a pseudogonotyl formed by thickened tegument on the left side of the ventral sucker. We examined morphology of specimens belonging to five species of Ochoterenatrema from Brazil, Ecuador and the USA and describe two new Ochoterenatrema spp. (Ochoterenatrema sphaerula n. sp. and Ochoterenatrema gracilis n. sp.). These new species are readily differentiated from their congeners by a combination of morphological characters, including the distribution of vitelline follicles and the body shape. We have provided the first DNA sequences for Ochoterenatrema. Newly generated partial nuclear 28S rDNA and mitochondrial cox1 gene sequences were used to differentiate among species and study the phylogenetic affinities of Ochoterenatrema spp., including one of the new species, O. gracilis n. sp., as well as Ochoterenatrema diminutum, Ochoterenatrema fraternum and Ochoterenatrema cf. labda. The phylogeny of the Microphalloidea based on 28S had well-supported topology, particularly at the family level. The Ochoterenatrema clade was strongly supported; however, the internal topology of the clade was weakly supported. Comparison of sequences revealed 0.4–1.3% interspecific divergence in 28S and 9.1–19.7% interspecific divergence in cox1 among Ochoterenatrema spp. We hypothesize that extremely diverse fauna of bats in South and Central America harbors multiple undescribed species of Ochoterenatrema. Several lecithodendriid genera from bats and other hosts are yet to be included in future molecular phylogenetic analyses to test the monophyly of this extremely diverse digenean family and analyze evolutionary affinities of its constituent taxa.     

An ultrastructural comparative study of the sperm of<Emphasis Type="Italic"> Hyla pseudopseudis</Emphasis>,<Emphasis Type="Italic"> Scinax rostratus</Emphasis>, and<Emphasis Type="Italic"> S. squalirostris</Emphasis> (Amphibia: Anura: Hylidae)

We make detailed comparisons of the ultrastructure of the spermatozoon among three species of the family Hylidae, Hyla pseudopseudis, Scinax rostratus, and S. squalirostris. The acrosome complex consists of two conical structures covering the nuclear rostrum, the acrosome vesicle, and the subacrosomal cone. The nucleus has a moderately condensed chromatin with a conical shape in longitudinal sections and a circular shape in cross-sections. In H. pseudopseudis, mitochondria are numerous and circular, and in S. rostratus and S. squalirostris there are fewer mitochondria that are more elongate in longitudinal and transverse sections. In H. pseudopseudis, the mitochondrial collar starts adjacent to the distal centriole, occupying the whole midpiece, whereas in both Scinax species the mitochondrial collar starts only at the posterior one-third of the midpiece. In both Scinax species, the presence of juxta-axonemal fiber, axial sheath, and axial fiber in the tail are seemingly plesiomorphic characters, widespread among bufonoid frogs. In H. pseudopseudis, however, the absence of axial fiber and axial sheath seems to be derived from the typical bufonoid condition. The differences between Hyla and Scinax sperm endorse the separation of the two genera and suggest that sperm ultrastructure can be a useful tool to investigate relationships at the intrafamily level.     

The true identity of Obama (Platyhelminthes: Geoplanidae) flatworm spreading across Europe

Since 2008 there have been many records in Europe (British Isles, Spain, France, Italy) of a large terrestrial planarian morphologically very similar to the Brazilian species Obama marmorata. Sequences of mitochondrial (Cox1) and nuclear (18S, 28S, ITS‐1 and EF) genes from European specimens and some from Brazil indicate that they belong to a species different from that of other specimens also collected in Brazil. Moreover, the phylogenetic results show that they are not sister‐species. Histological sections of both Brazilian and European specimens reveal subtle morphological differences between the two species. Obama marmorata is confined to Brazil, and the second, herein described new species, O bama nungara sp. nov. , is found in Brazil and Europe. These cryptic species may be syntopic in areas in Brazil. The new species occurs in human‐modified environments both in Brazil and in Europe. We also conclude that the specimens from Spain and Argentina identified previously as Obama marmorata belong to the new species.     

Diversity of haemogregarine parasites infecting Brazilian snakes from the Midwest and Southeast regions with a description of two new species of Hepatozoon (Apicomplexa: Adeleorina: Hepatozoidae)

Although Brazil is a hotspot for snake species, there is a lack of information on the biodiversity of haemoparasites infecting these hosts. Thus, the present study aimed to bring new insights on the diversity of species of Hepatozoon (Apicomplexa: Adeleorina: Hepatozoidae) infecting Brazilian snakes from the Midwest and Southeast regions. The snakes were captured from 2018 to 2020 from the states of Mato Grosso, Mato Grosso do Sul, Goiás, and São Paulo. Three to five blood smears were made and the remaining blood sample was stored for further molecular analysis. Moreover, histopathological slides of the organs were stained with haematoxylin-eosin. Regarding molecular diagnosis, PCR was performed targeting different regions of the 18S rRNA gene of apicomplexan parasites. From the 13 free-living snakes screened, ten (76.92%) were found infected with Hepatozoon spp. Based on morphological and morphometric tools, five different morphotypes of species of Hepatozoon gamonts were detected. Molecular data and phylogenetic analysis support the morphological data, identifying five species of Hepatozoon from snakes, of which three species belong to previously described species, Hepatozoon cevapii, Hepatozoon cuestensis, and Hepatozoon quagliattus, with a genetic similarity of 100% (based on the 18S rRNA genetic marker). The present study identifies and describes two new species of Hepatozoon, Hepatozoon annulatum sp. nov. infecting the snake Leptodeira annulata and Hepatozoon trigeminum sp. nov. infecting the snake Oxyrhopus trigeminus. Thus, based on morphological and molecular data the present study provides new insights on haemogregarine diversity infecting Brazilian snakes from the Midwest and Southeast regions.     

Morphological and molecular identification of helminths of the greater bulldog bat Noctilio leporinus (Quiroptera: Noctilionidae) from Campeche,Mexico

Surveys on parasites of bats from the Americas have been conducted, but information on helminths is still scarce, especially in the Neotropical region. In Mexico, there are species of bats that lack of a record for helminth species, such as members of the family Noctilionidae. The present study describes for the first time the helminths of Noctilio leporinus in Campeche, Mexico. In 2017, six specimens of N. leporinus were studied for helminths. The species identification of helminths was based on morphological studies and molecular analysis of fragments of the 28S rDNA. All bat specimens were infected for at least one helminth species. Three helminth taxa were identified: the trematode Pygidiopsis macrostomum, and the nematodes Tricholeiperia cf. proencai, and Heligmonellidae gen. sp. The morphological identification of P. macrostomum was confirmed by sequence analysis of 28S rDNA gene. The phylogeny of P. macrostomum grouped our sequence with other sequences of the same species collected in Brazil. The phylogenetic tree of Heligmonellidae gen. sp. indicated that the helminth belongs to clade formed by the species Odilia bainae, Nippostrongylus magnus and Nippostrongylus brasiliensis of the family Heligmonellidae. The phylogenetic analysis of the 28S sequences of T. cf. proencai did not show any similarity or close affinity with nematodes from which that gene has been sequenced to date. The findings of the present study increase the number of helminth species parasitizing bats in Mexico.     

Complex female genitalia indicate sperm dumping in armored goblin spiders (Arachnida,Araneae, Oonopidae)

In promiscuous females, sperm ejection from the sperm storage site can be a strong mechanism to influence sperm priority patterns. Sperm dumping is reported from different animals including birds, insects, and humans. In spiders, it has been documented for four species including the oonopid Silhouettella loricatula. Oonopidae are a diverse spider family comprising many species with peculiar female genitalia. Especially in species where studies of mating behavior are difficult, morphological investigations of the genitalia help to understand their function and evolution. In the present study, the genitalia of the oonopids Myrmopopaea sp., Grymeus sp., and Lionneta sp. are investigated by means of histological serial sections and scanning electron microscopy (SEM). The results are compared with previous findings on S. loricatula. In Myrmopopaea sp. and Grymeus sp., the same morphological components are present that are involved in sperm dumping in S. loricatula. Inside the receptaculum, sperm are enclosed in a secretory sac which can be moved to the genital opening and dumped during copulation by muscle contractions. The female genitalia of Lionneta sp. are asymmetric. They show the same characteristics as S. loricatula but all the investigated females were unmated. The results strongly suggest that sperm dumping occurs in Myrmopopaea sp., Grymeus sp., and Lionneta sp. and happens by the same mechanism as in S. loricatula. Sperm dumping might even be common within a clade of oonopids. As in S. loricatula, the sperm transfer forms in the investigated species consist of several spermatozoa. Papillae with unknown function occur on the receptacula of all females.