Morphological and molecular characterization of Cosmocercoides amapari n. sp. (Nematoda: Cosmocercidae), parasitic in hylid frogs from the Brazilian Amazon.

Cosmocercoides Wilkie, 1930 are gastrointestinal parasites commonly found in amphibians and reptiles, with 4 species reported from the Neotropical region. In the present study, a new species of Cosmocercoides, namely Cosmocercoides amapari n. sp. is described using integrated approaches such as light and scanning microscopy, and DNA sequencing of the mitochondrial cytochrome c oxidase subunit 1 gene. The specimens were collected from the large intestine of 3 species of hylid frogs in Amapá, Brazil. The new species can be distinguished from its congeners by morphological traits, including the pattern of caudal papillae, absence of the gubernaculum, 2 poorly sclerotized spicules, presence of lateral alae and somatic papillae along the body. In addition, our molecular analyses and phylogenetic reconstructions strongly support the status of the new taxon, which clustered poorly with a large clade of Cosmocerca spp. Cosmocercoides amapari n. sp. is the 29th species of the genus, the 5th from the Neotropical region, the third reported in Brazil, the second described from the Amazon region and the first belonging to the Neotropical region with molecular data.

Molecular phylogeny of two Rhinoxenus species (Monogenea: Dactylogyridae) from the nasal cavities of serrasalmids (Characiformes: Serrasalmidae) from Brazil.

The dactylogyrid genus Rhinoxenus (Monogenea: Dactylogyridae) is composed of specialized monogeneans infecting the nasal cavities of freshwater fishes from the Neotropical region. This taxon currently comprises 11 species and is easily distinguished from other monogeneans by the absence of the dorsal bar, ventral anchor with inconspicuous roots covered by a sclerotized cap, dorsal anchor greatly modified into a needle-like shape, and hook pair 2 located into bilateral lobes of the trunk. Specimens of Rhinoxenus euryxenus and Rhinoxenus paranaensis were found infecting the nasal cavities of Serrasalmus marginatus and Serrasalmus maculatus, respectively, from the Paraná River basin in Brazil. For the first time, molecular data of Rhinoxenus spp. were obtained and used as a basis for phylogenetic analyses of the genus. Furthermore, our findings represent the first record of R. paranaensis in Brazil.

A new species of Rhabdias (Nematoda: Rhabdiasidae), a lung parasite of Pristimantis chiastonotus (Anura: Strabomantidae) from the Brazilian Amazon: description and phylogenetic analyses.

Rhabdias Stiles and Hassal, 1905 comprises approximately 90 species of parasitic lung nematodes of amphibians and reptiles that have a wide distribution, with 21 species occurring in the Neotropics. In the present study, we describe Rhabdias waiapi n. sp. found parasitizing the lungs of the anuran species Pristimantis chiastonotus from the Amazon Biome in the Amapá State, Northern Brazil. The new species is characterized by having an elongated body, expansions of the cuticular inflation in the anterior end that become more discrete along the body, an anterior end with a slight constriction at the level of the esophageal apex with four rounded subapical elevations of the body wall, six lips, four near the edge of the oral opening and two more distant lateral ones, and a gradually tapering elongated tail. In addition, molecular analyses and phylogenetic reconstructions were made, with sequences from the coding region of the mitochondrial cytochrome c oxidase subunit I gene. Those results strongly support the status of the new taxon, which formed a poorly supported clade with Rhabdias sp. 5 from Anolis brasiliensis from Northeast Brazil. Rhabdias waiapi n. sp. is the 19th species of the genus described in the Neotropics for amphibians, the 10th in Brazil, the second described from hosts of the family Strabomantidae from the Neotropical region, and the first amphibian nematode species described in the Amapá State.

First molecular assessment of two digenean parasites of the lancehead snake Bothrops moojeni Hoge, 1966 (Serpentes, Viperidae) in Brazil.

Two digenean species, Infidum infidum Faria, 1910 (Dicrocoeliidae) and Travtrema stenocotyle Cohn, 1902 (Plagiorchiidae), were collected in the large pit viper Bothrops moojeni Hoge, 1966 from Reserva Particular do Patrimônio Natural Cisalpina, municipality of Brasilândia, Mato Grosso do Sul State, Brazil. In this study, we provide the first molecular characterisation using the 28S rDNA and phylogenetic position data of these two common digeneans from B. moojeni. The molecular framework revealed topologies with strongly supported clades using maximum likelihood and Bayesian inference methods, positioned I. infidum among Plagiorchiidae and not among Dicrocoeliidae as expected and T. stenocotyle (Plagiorchiidae) surprisingly grouped as a sister group to Allassogonoporidae, Microphallidae, Pleurogenidae, and Prosthogonimidae, not related to plagiorchids. Our molecular phylogenetic data showed that these species may not correspond to their assigned families and encourage future studies on the systematic of these understudied groups.

Molecular and morphological characterization of the metacercariae of two species of diplostomid trematodes (Platyhelminthes, Digenea) in freshwater fishes of the Batalha River, Brazil.

The Diplostomidae include a large group of flatworms with complex life cycles and are frequently found parasitizing the eyes and central nervous system of freshwater fishes. The morphological identification of the metacercariae at species level is not always possible. Thus, molecular tools have become essential to assist in the parasite species determination. This study was aimed at describing two diplostomid metacercariae found in freshwater fish in São Paulo, Brazil, based on morphological characters and in the genetic characterization of COI sequences. Our results showed that the two recognized taxa (Tylodelphys sp. and Diplostomidae gen. sp.) appear to be different from the species already described in South America. Tylodelphys sp. differs morphologically from Tylodelphys xenopi, T. mashonense, T. jenynsiae, and T. scheuringi. The metacercariae of T. clavata and T. conifera are smaller than Tylodelphys sp., while T. podicipina is larger than the metacercariae described here. The phylogenetic analysis of COI sequences yielded Tylodelphys sp. as the sister species of Tylodelphys sp. 4, a species reported from the brain of the eleotrid Gobiomorus maculatus in Oaxaca, Mexico. The metacercariae identified as Diplostomidae gen. sp. are morphologically different from the known diplostomid metacercariae and did not match with other diplostomid sequences available. Diplostomidae gen. sp. is recovered as the sister species of Diplostomum ardeae. Although the morphological evidence and the COI sequences differentiate the metacercariae found, the absence of adult specimens of both species precludes the specific designation. This is one of the first papers that use an integrative taxonomy approach to describe the species diversity of diplostomid trematodes in Brazil.

Prevalence, intensity, and phylogenetic analysis of Henneguya piaractus and Myxobolus cf. colossomatis from farmed Piaractus mesopotamicus in Brazil.

Henneguya piaractus and Myxobolus colossomatis (Myxosporea: Myxobolidae) are commonly found in the characid Piaractus mesopotamicus, an important fish farm species in Brazil. This paper describes the prevalence, mean intensity, molecular phylogeny, ultrastructure, and histology of H. piaractus and M. cf. colossomatis found infecting specimens of P. mesopotamicus collected from fish farms in the state of São Paulo, Brazil. A total of 278 fish were collected from 3 fish farms between February 2008 and July 2010. Parasite prevalence and mean intensity varied throughout the study period, and according to location and year. A phylogenetic tree, placing South American species in a global context, showed a clear tendency among myxosporean species to cluster according to host families. Ultrastructural analysis for M. cf. colossomatis showed the plasmodial wall with numerous projections toward host cells and phagocytic activity. Histopathological data showed hyperplasia caused by H. piaractus in highly infected fish. Histological and ultrastructural analysis of H. piaractus showed results similar to those that have previously been reported.

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.

Revisiting the taxonomy of Rhabdias fuelleborni Travassos, 1928 (Nematoda, Rhabdiasidae) with approaches to delimitation of species and notes on molecular phylogeny.

Rhabdias fuelleborni was described by Travassos (1926), who identified it in the lungs of Rhinella gr. marina (=Bufo marinus) from São Paulo. Later, Kloss (1971) added new information on the morphology and diagnosis of the species and new data on host and localities. However, its taxonomic status remains uncertain, due in particular to the morphological uniformity of Rhabdias spp., while the original description lacks important morphological data. In the present study, we found and reevaluated the type series of R. fuelleborni, as well as examining fresh material obtained from the type host and type locality, using light and scanning electron microscopy and molecular tools. The type series was deposited by Travassos (1926), and according to museum records the type locality is Paraty, Rio de Janeiro, Brazil. Our morphological and molecular analyses reinforce the taxonomic validity of this species, while molecular phylogeny identified a close relationship between R. fuelleborni and Rhabdias sp.4 and Rhabdias sp. from other studies, and from bufonids. Also R. fuelleborni are sister taxa with R. cf. stenocephala from Leptodactylidae from the northeast of Brazil and R. vencesi from Boophis madagascarariensis, Madagascar, according to mitochondrial COI gene.

Morphological and molecular data of new species of Characithecium and Diaphorocleidus (Monogenea: Dactylogyridae) from Neotropical characid fishes.

The present study describes three new species of monogenean parasites of characid fishes from the Upper Paraná River basin, Brazil: Characithecium paranapanemense n. sp. on Psalidodon paranae and Psalidodon bockmanni, Diaphorocleidus magnus n. sp. on Astyanax lacustris and Psalidodon fasciatus, and Diaphorocleidus neotropicalis n. sp. on Astyanax lacustris and P. bockmanni. An amendment for Diaphorocleidus is proposed, since additional characters observed in the new species required to extend the generic diagnostic features mainly to include: articulation process connecting the base of the MCO with accessory piece present or absent, and accessory piece with variable shapes (plate-like, pincer-shaped, wrench-shaped, sheath-shaped), divided or not into subunits. Characithecium paranapanemense n. sp. can be distinguished from other congeners by the morphology of its MCO and accessory piece. Diaphorocleidus magnus n. sp. differs from most of its congeners by the morphology of its accessory piece, the presence of articulation process connecting the base of the MCO with accessory piece, and the morphology of the sclerotized structures of the haptor. Diaphorocleidus neotropicalis n. sp. can be easily distinguished from its congeners by the morphology of the accessory piece, the sclerotized structures of the haptor and the morphology of the vagina. Molecular data of the new species (partial 28S rDNA and mitochondrial cytochrome oxidase I) were obtained and the first phylogenetic analysis based on 28S rDNA gene sequences for species of Characithecium and Diaphorocleidus are provided. Although Diaphorocleidus and Characithecium share some morphological similarities, phylogenetic analysis indicates that species of these two genera are not closely related.

A new species of Cacatuocotyle (Monogenea, Dactylogyridae) parasitizing Astyanax spp. (Characiformes, Characidae) from Brazil, including molecular data and a key to species identification.

The present study describes Cacatuocotyle papilionis n. sp. (Monogenea, Dactylogyridae) from the skin of the characid fishes Astyanax lacustris (Lütken, 1875) (=Astyanax altiparanae Garutti & Britski, 2000) and Astyanax fasciatus (Cuvier, 1819) (Characiformes, Characidae) from the Southeast of Brazil, supported by morphological and molecular data. The new species differs from all congeners, mainly due to the morphology of the ventral bar (resembling a butterfly), accessory piece, and the number of rings of the male copulatory organ (MCO), comprising a coiled tube with 4.5-5.5 counterclockwise rings. The first molecular data for this monogenean genus is provided in this study, using the partial sequences of the ribosomal gene (28S), as well as providing an identification key to the species.

Molecular phylogenetic position of Haplometroides intercaecalis (Digenea, Plagiorchiidae).

Three valid species of Haplometroides Odhner, 1910 parasitise snakes and amphisbaenians from South America. This study provides additional data on morphometric and molecular phylogenetic position inferred from the nuclear ribosomal gene 28S (partial). DNA sequences were isolated from Haplometroides intercaecalis Silva, Ferreira and Strüssmann, 2007 found in one specimen of Phalotris matogrossensis Lema, D'Agostini and Cappellari, 2005. Five digenean specimens were recovered from the esophagus of this snake, and four specimens were used for morphometrical studies and one specimen for molecular analysis. Phylogenetic analysis using maximum likelihood and Bayesian methods was conducted with sequences available for the order Plagiorchiida and its phylogenetic position places H. intercaecalis among the brachycoeliids Brachycoelium (Dujardin, 1845) Stiles and Hassall, 1898 and Parabrachycoelium Pérez-Ponce de León, Mendoza-Garfias, Razo-Mendivil and Parra-Olea, 2011, and the mesocoeliid Mesocoelium Odhner, 1910, not closely related to plagiorchids as expected. Due to morphological differences among these families, it may be necessary to create a new family to accommodate Haplometroides spp. However, more genera/taxa as well as other molecular markers should be added in future studies to confirm our results and resolve this matter. This is the first phylogenetic positioning of digeneans of the genus Haplometroides, contributing to the systematic analysis of the helminthological biodiversity of Neotropical snakes.

Supplementary studies on Anacanthorus penilabiatus and Mymarothecium viatorum (Monogenea: Dactylogyridae) from Piaractus mesopotamicus (Characiformes: Serrasalmidae) in Brazil.

The dactylogyrid monogeneans Anacanthorus penilabiatus (Boeger, Husak and Martins, 1995) and Mymarothecium viatorum(Boeger, Piasecki and Sobecka, 2002), commonly found in large quantities in farmed Piaractus mesopotamicus (Holmberg, 1887) in Brazil, were collected from three fish farms located in the State of São Paulo, Brazil. Morphological and molecular identification, scanning electron microscopy, histopathological and descriptive parameters data were performed. A total of 278 specimens of P. mesopotamicus were gathered from February 2008 and July 2010. Prevalence (P) and mean intensity of infection (MII) data was taken at each location. The data showed prevalence and mean intensity of infection annually, with numbers such as P = 91.2% and MII = 50 worms/fish (Pirassununga), P = 60% and MII = 39.2 worms/fish (Mogi Mirim), and P = 100% and MII = 204.8 worms/fish (Itapira). Histopathological results showed hyperplasia of the secondary lamellae, and hypersecretion of mucus was also observed. A phylogenetic topology was inferred using complete SSU (18S) ribosomal rDNA, positioning Anacanthorus penilabiatus and Mymarothecium viatorum among other monopisthocotyleans available in GenBank.

Copépodos Notodiaptomus sp. Kiefer (Crustacea, Calanoida) naturalmente infectados com metacestódeos no reservatório do Juqueri, São Paulo, Brasil / Copepods Notodiaptomus sp. Kiefer (Crustacea, Calanoida) naturally infected by metacestodes in the Juqueri reservoir, São Paulo, Brazil

The aim of this work was to identify the components of zooplankton that act as intermediate hosts of cestodes. One hundred and ninety four copepods of the suborder Calanoida, 317 copepods of the suborder Cyclopoida and 4240 cladocerans were collected in the Juqueri reservoir, in the state of São Paulo, from January to August, 2003. Only Copepods Calanoida of the genus Notodiaptomus sp. Kiefer were found to be infected and contained two distinct forms of metacestodes. The metacestodes, denominated Met 1 (order Proteocephalidea) and Met 2 (order Cyclophyllidea), had the following rates of prevalence and mean intensities of infection: Met 1 - 2.06 percent and 64 larvae/copepod and Met 2 - 0.52 percent and one larvae/copepod. The positive copepods were collected at the margins of the reservoir during the day. This finding suggest that parasitism may lead to a change in the behavior of the copepods and make them more susceptible to predation in shallow water.

Este trabalho teve o objetivo de identificar os componentes do zooplâncton que atuam como hospedeiros intermediários de cestódeos. Foram examinados 194 copépodos da subordem Calanoida, 317 copépodos da subordem Cyclopoida e 4240 cladóceros coletados no reservatório do Juqueri, Estado de São Paulo, entre janeiro e agosto de 2003. Apenas copépodos Calanoida do gênero Notodiaptomus sp. Kiefer encontravam-se positivos, sendo relatada duas formas morfológicas distintas de metacestódeos. Os metacestódeos, denominados Met 1 (Proteocephalidea) e Met 2 (Cyclophyllidea) apresentaram as seguintes prevalências e intensidades médias de infecção: Met 1 - 2,06 por cento e 64 larvas/copépodo e Met 2 - 0,52 por cento e uma larva/copépodo. Os copépodos positivos foram coletados na região litorânea do reservatório durante o dia, o que leva a crer que o parasitismo pode modificar o comportamento dos copépodos tornando-os mais acessíveis à predação.