Protein identification from the parotoid macrogland secretion of Duttaphrynus melanostictus

Bufonid parotoid macrogland secretion contains several low molecular mass molecules, such as alkaloids and steroids. Nevertheless, its protein content is poorly understood. Herein, we applied a sample preparation methodology that allows the analysis of viscous matrices in order to examine its proteins. Methods: Duttaphrynus melanostictus parotoid macrogland secretion was submitted to ion-exchange batch sample preparation, yielding two fractions: salt-displaced fraction and acid-displaced fraction. Each sample was then fractionated by anionic-exchange chromatography, followed by in-solution proteomic analysis. Results: Forty-two proteins could be identified, such as acyl-CoA-binding protein, alcohol dehydrogenase, calmodulin, galectin and histone. Moreover, de novo analyses yielded 153 peptides, whereas BLAST analyses corroborated some of the proteomic-identified proteins. Furthermore, the de novo peptide analyses indicate the presence of proteins related to apoptosis, cellular structure, catalysis and transport processes. Conclusions: Proper sample preparation allowed the proteomic and de novo identification of different proteins in the D. melanostictus parotoid macrogland secretion. These results may increase the knowledge about the universe of molecules that compose amphibian skin secretion, as well as to understand their biological/physiological role in the granular gland.(AU)

Diet and resource partitioning among anurans in irrigated rice fields in Pantanal, Brazil / Dieta e partição de recursos entre anuros em campos de rizicultura no Pantanal, Brasil

Artificial ponds or irrigated systems scattered throughout farmlands can offer important habitats for anurans and can be interesting sites for research on species resources use in a changing landscape. This study describes the diet and resource partitioning among anurans inhabiting irrigated rice fields in the Pantanal region. Twenty categories of prey were found in the stomachs of Leptodactylus chaquensis, L. elenae, L. podicipinus and Rhinella bergi, the most frequent being Coleoptera, Hymenoptera, larvae of Hexapoda, Hemiptera, Diptera and Orthoptera. The great differences found in the diet of these species in rice fields compared to other locations, according to available records in the literature, was the increased importance of Hemipitera and Orthoptera and the decrease in importance of Hymenoptera in the diet of leptodactylids. These differences might be attributed to changes in the availability of resources in response to habitat modification. Although diet composition was very similar among species, niche overlap was larger than expected by chance, suggesting that the competition for food resources is not, or has not been, a significant force in determining the structure of this frog community. Two non-exclusive hypotheses could be considered as a justification for this result: 1) the high niche overlap could result from resource availability, which is sufficient to satisfy all species without any strong competition; 2) or the high values of niche overlap could be a selective force driving species to compete, but there has not been enough time to express a significant divergence in the species diet because the study area is characterised as a dynamic habitat influenced by frequent and cyclical changes.

Lagoas artificiais e sistemas irrigados em áreas de produção agrícola podem oferecer habitats importantes a anuros e ser significativos para investigações sobre o uso de recursos pelas espécies em ambientes modificados. Este estudo descreve a dieta e partição de recursos tróficos entre anuros de campos de rizicultura na região do Pantanal. Nos estômagos de Leptodactylus chaquensis, L. elenae, L. podicipinus e Rhinella bergi foram encontradas 20 categorias de presas, sendo Coleoptera, Hymenoptera, larvas de Hexápoda, Hemiptera, Diptera e Orthoptera as mais frequentes. Maiores diferenças encontradas na dieta dessas espécies em campos de arroz comparadas a de outros locais, de acordo com dados da literatura, foram o aumento da importância de Hemipitera e Orthoptera e a diminuição na importância de Hymenopteras na dieta dos leptodactilídeos. Essas diferenças talvez sejam reflexos de mudanças na disponibilidade de recursos de presas, em resposta às modificações ambientais. Apesar de as espécies apresentarem dietas bastante similares, a sobreposição de nicho foi maior que a esperada ao acaso, sugerindo que a competição por recursos alimentares não é, ou não tem sido, uma força significante na determinação na estrutura dessa comunidade de anuros. Duas respostas não exclusivas podem ser consideradas para justificar esse resultado: 1) a alta sobreposição de nicho poderia resultar da grande disponibilidade de recursos alimentares, a qual seria suficiente para satisfazer as necessidades de todas as espécies sem que haja forte competição; 2) ou os altos valores de sobreposição poderiam ser uma força seletiva que levasse as espécies à competição, mas que não têm tempo suficiente para se expressar em divergência na dieta das espécies pelo fato de a área de estudo ser um ambiente altamente dinâmico, sofrendo bruscas e cíclicas mudanças.

Rhabdias paraensis sp. nov.: a parasite of the lungs of Rhinella marina (Amphibia: Bufonidae) from Brazilian Amazonia

The nematode parasites of Rhinella marina include species of the genus Rhabdias (Rhabdiasidae: Rhabditoidea). The present study describes Rhabdias paraensis sp. nov., which parasitizes the lungs of R. marina in Brazilian Amazonia. Of the more than 70 known species of this genus, 18 are parasites of bufonids, of which, eight are Neotropical. The new species described here is similar to Rhabdias alabialis in the absence of lips is different by the presence of conspicuous cephalic papillae. We describe details of the four rows of pores, which are distributed equally along the whole of the length of the body and connected with hypodermal cells, using histology and scanning electron microscopy. Other histological aspects of the internal structure of this nematode are also described.

Hematología y citoquímica de las células sanguíneas de Rhinella fernandezae (Anura: Bufonidae) en Espinal y Delta-Islas del río Paraná, Argentina

Hematology and blood cell cytochemistry of Rhinella fernandezae (Amphibia: Anura) from Espinal and Delta-Islands of Paraná River, Argentina. The description of amphibian hematology is scarce and most of these studies have been done in species from North America, Asia and Europe. With the purpose to obtain basic hematological information of Rhinella fernandezae, 23 blood samples from Santa Fe and Entre Ríos natural reserves were studied. Blood of each individual was extracted by cardiac puncture and hemograms were carried out. Morphological and cytochemical description of blood cells were analyzed in slides and were inspected for extra and intra cellular parasites. Five leucocytes types were observed, being lymphocytes the predominant ones followed by basophiles. Heterophils and eosinophils were positive to PAS, Sudan B and peroxidase. The erythrocytes and its precursors were negative for cytochemical reactions. Micronuclei and nuclear alterations frequencies were scarce. No significant differences (p>0.05) were observed between sexes neither in hemograms nor in blood cells morphology. Microfilarias were the only hemoparasites found with a relative low prevalence and infection intensity. The hematological characteristics studied were similar to those reported for other amphibians, suggesting that R. fernandezae individuals present optimal nutritional and immunological status. Rev. Biol. Trop. 59 (1): 17-28. Epub 2011 March 01.

La descripción de la hematología de los anfibios anuros es escasa, habiéndose realizado la mayoría de los trabajos en especies de Norteamérica, Asia y Europa. Con el propósito de obtener datos hematológicos para Rhinella fernandezae, fueron estudiados 23 especímenes provenientes de zonas protegidas de las provincias de Santa Fe y Entre Ríos. Se les extrajo sangre por punción cardíaca y se realizaron hemogramas. En los extendidos sanguíneos, se efectuaron la descripción morfológica y citoquímica de las células sanguíneas y búsqueda de parásitos. Se observaron cinco tipos de leucocitos, donde predominaron los linfocitos pequeños. Heterófilos y eosinófilos resultaron positivos para PAS, Sudan B y peroxidasa; contrariamente, los eritrocitos y sus precursores fueron negativos. Los puntajes de las reacciones citoquímicas fueron variables para basófilos, linfocitos, monocitos y trombocitos. Las frecuencias de micronúcleos y de alteraciones nucleares fueron escasas. No se observaron diferencias significativas (p>0.05) entre sexos en el hemograma ni en la morfología sanguínea. Los únicos hemoparásitos encontrados fueron microfilarias (Nematoda: Filaroidea), cuya prevalencia e intensidad de infección fueron bajas. Las características hematológicas estudiadas fueron semejantes a los valores reportados para otros anfibios, pudiendo inferir que los individuos de R. fernandezae estudiados se encuentran en un buen estado nutricional e inmunológico.

Época reproductiva y distribución geográfica del sapo Incilius aucoinae (Bufonidae) en Golfito, Costa Rica / Reproductive period and geographic distribution of the toad Incilius aucoinae in Golfito, Costa Rica

Reproductive period and geographic distribution of the toad Incilius aucoinae in Golfito, Costa Rica. The biology of neotropical amphibians is not well known. Some toad species of the Bufonidae family are common, thus allowing the study of their populations. We studied a population in río Cañaza, Golfito, Costa Rica, in a sector 360 m upstream of Barrio Ureña, divided in 36 sectors of 10 m. The study was carried out for five years, but taking samples of adults in January 1995, 1997, 1998 (except March), and 1999. We also studied tadpoles during four days in 1997. In total, 443 males and 7 females were marked, with 315 males recaptured and no females. Females are bigger (91.1 mm) than males (61.3 mm). Most recaptured individuals occurred in the same or adjacent sectors, with a maximum movement of 28 sectors. One individual was recaptured repeatedly in all the samples. This indicates that they survived at least five reproductive cycles. The reproductive cycle takes place during the dry season, between December and April. We found more individuals during the reproductive cycle of 1997 and less during 1995. Sector 6 had the most number of males and sector number 4 had the least. Sectors with semi-open vegetation had more males compared to sectors with more forest cover. February is the month with the highest abundance of males. There are intermediate values in December and January, and fewer individuals in March and April. In every month, except February, the abundance of individuals was greater where there were beaches. Tadpoles were found both during the day and night in the 31 river sectors, but it was less likely to find tadpoles in the river section during the night. During the day all tadpoles were found scattered around the river, at night they congregated near river margins. The reproductive cycle of this toad occurs during the dry season and females are present only in the reproductive season, resulting in an almost only male sex ratio (only seven females were found). Males can reproduce at least in 5 reproductive cycles, and -unexpectedly- day and night distribution of tadpoles varies. Rev. Biol. Trop. 57 (Suppl. 1): 301-311. Epub 2009 November 30.

Se estudian algunos aspectos de la biología reproductiva del sapo Incilius aucoinae en una población en el río Cañaza, Golfito, Costa Rica; en una sección de 360 m aguas arriba dividida en 36 sectores de 10 m. La duración del estudio con adultos fue de cuatro años, con muestreos en enero de 1995, 1997, 1998 (excepto marzo) y enero de 1999; los renacuajos se estudiaron en cuatro días de 1997. Se marcaron 443 machos (con 315 reobservaciones) y 7 hembras (no fueron recapturadas). Un individuo fue recapturado en todos los periodos de estudio, lo que indica que por los menos sobreviven a 5 épocas reproductivas. La época reproductiva corresponde a los meses secos, de diciembre a abril. En la época reproductiva de 1997 se encontró más individuos y menos en 1995; el sector 6 es el que tiene más machos y el 4 menos; la mayor cantidad de machos se encontró en sectores con vegetación semiabierta, los sectores con mayor cobertura vegetal tienen menos; febrero es el mes con mayor abundancia de machos, con valores intermedios en diciembre y enero, y con muy pocos individuos en marzo y abril. En todos los meses, excepto febrero, la abundancia fue mayor donde había playones. Este sapo se reproduce en la época seca, y en el sitio reproductivo las hembras apenas entran a reproducirse. Se da una relación de sexos de casi solo machos, pues solo hubo 7 hembras observadas, y sorprende los cambios de distribución entre el día y la noche en los renacuajos.

Desarrollo embrionario y larva del sapo Incilius aucoinae (Bufonidae) en Golfito, Costa Rica / Embryonic development and larva of the toad Incilius aucoinae (Bufonidae) in Golfito, Costa Rica

Embryonic development and larva of the toad Incilius aucoinae (Bufonidae) in Golfito, Costa Rica. The larva and developmental stages of Incilius aucoinae are described using specimens from Río Cañaza, Golfito, Costa Rica. We collected the individuals in January 1997, and January and February 1998. The total sample was 696 individuals, 317 eggs in 1-12 stages, 176 eggs with embryos in stages 15-16, and 204 tadpoles in stages 21-43. The average number of eggs per nest was 8 940 (n = 4). These eggs were laid in strings, attached to the river strata. Eggs in stages 1-12 had an average diameter of 1.75 (s = 0.38) mm and eggs with embryos in stages 15 -16 measured 1.33 (0.34) mm wide and 3.06 (1.46) mm long. The description of the tadpole based on UCR 19982 is as follows: dorsal, ventral surfaces, and tail are brown with dark brown spots. Length of tail 9.43 mm, body length 6.18 mm (total length 15.62). Translucent light brown small fins, dorsal fin with scattered spots, ventral fin without dark pigments. Dorsal-ventral flattened oval body with anteroventral snout. A few individuals had varied colors, from black (3.4%) to uniform (0.5-1.5%). Rev. Biol. Trop. 57 (Suppl. 1): 291-299. Epub 2009 November 30.

Se describe la larva y los estadios del desarrollo del huevo y larva de Incilius aucoinae utilizando especímenes del río Cañaza, Golfito, Costa Rica, recolectados en enero de 1997 y enero-febrero de 1998. La muestra fue de 696 individuos, 317 huevos en estadios 1-12, 176 huevos con embriones en estadios 15-16, y 204 renacuajos en estadios 21-43. El promedio del número de huevos por nidada fue de 8 940 (n = 4), los cuales fueron colocados en una hilera, sujetos al sustrato del río. Los huevos en estadios 1-12 tienen un promedio de diámetro de 1.75 (s = 0.38) mm y los huevos con embriones en estadios 15-16 miden 1.33 (0.34) de ancho y 3.06 (1.46) mm de largo. La descripción del renacuajo basada en el espécimen UCR 19982, es como sigue: el dorso, el vientre y la cola son de color café y punteado de café oscuro; la cola mide 9.43 mm y el cuerpo 6.18, para un tamaño total de 15.62; las aletas son pequeñas café claro translúcidas, la dorsal con puntos dispersos y la ventral sin ningún tipo de pigmento oscuro. También es el único renacuajo ovoide con una tendencia a ser aplastado dorso-ventralmente, y con la boca antero-ventral. Dentro de la variación observada, algunos individuos son de color negro (3.4 %) y pocos son uniformes (0.5- 1.5 %).

An enigmatic frog of the genus Atelopus (Family Bufonidae) from Parque Nacional Chirripó, Cordillera de Talamanca, Costa Rica

A distinctive new species of Atelopus is described from Parque Nacional Chirripó Grande, Cordillera de Talamanca (3 400-3 500 m). It closely resembles populations of the Atelopus ignescens complex from the Andes of northern Ecuador and southern Colombia. it differs most significantly from these frogs in the pattern of spiculae and coni development on the throat, chest, hands and feet. The Costa Rican species appears to be an outlier of the complex inexplicably separated geographically from its nearest allies by an over land distance of about 1 600 km. Rev. Biol. Trop. 57 (1-2): 381-386. Epub 2009 June 30.

Se describe una nueva especie de Atelopus del Parque Nacional Chirripó Grande, Cordillera de Talamanca (3 400-3 500 m sobre el nivel del mar). Se parece a poblaciones del complejo de Atelopus ignescens de los Andes del norte de Ecuador y del sur de Colombia. Principalmente difiere de estas ranas en el patrón de desarrollo de espículas y conos en la garganta, pecho, manos y pies. La especie de Costa Rica es atípica dentro del complejo por estar inexplicablemente separada geográficamente de sus más cercanos representantes por una distancia aproximada de 1 600 km por tierra.

Study on Isolation of Pathogenic Dematiaceous Fungi from Wild Toads / 中华皮肤科杂志

Objective To compare the carriage of dematiaceous fungi in wild toads from both en-demic area and non-epidemic area of chromoblastomycosis.Methods A total of354and45wild toads(Bufo Chinese)were caught from two counties of Shandong Province,Zhangqiu county,an endemic area of chromoblastomycosis,and from Laiyang county,an area where the infection of dematiaceous fungi in humans has not been found,respectively.The toads were killed and the suspension of the internal organs were used to isolate dematiaceous fungi.Results Eleven species including162strains of dematiaceous fungi were iso-lated from151of354(42.7%)toads from the endemic area.The isolated strains consisted of49Phialopho-ra verrucosa,33Exsophiala,19Phialophora(nonpathogenic),19E.jenselmei,12Cladosporium carrionii,3suspected of C.carrionii,7Rhinocladiella sp.,5Fonsecaea pedrosoi,4C.trichoides,3E.spinifera,1C.mi-nourae,4unidentified.Only4nonpathogenic dematiaceous fungi were isolated from4of45(8.9%)toads obtained from non-epidemic area.Conclusions Varieties of pathogenic dematiaceous fungi,especially C.carrionii,are isolated from wild toads in endemic area of chromoblastomycosis,while none from non-endemic area.The results suggest that the colonization of pathogenic dematiaceous fungi in wild toads may be associa-ted with endemicity of chromoblastomycosis.