Ticks parasitizing cold-blooded animals from three different Brazilian biomes; with note for males of Amblyomma rotundatum.

Ticks parasitize a wide variety of wild animals, including amphibians and reptiles. In addition to the possibility of microorganism transmission to these hosts, ticks can also cause severe bleeding, and high parasitism can lead to death. Therefore, knowing the diversity of ticks parasitizing amphibians and reptiles is important for conservation and preservation measures for these vertebrates. In the present study, we report parasitism by ticks in amphibians and reptiles from different Brazilian biomes (Amazon, Caatinga, Cerrado and Atlantic Forest). Ticks were collected from amphibians and reptiles deposited from the Herpetological Collection of the Federal University of Maranhão (UFMA), São Luís (Maranhão State), the State University Santa Cruz (UESC), Ilhéus (Bahia State), and the Federal University of São Francisco Valley (Univasf), Petrolina (Pernambuco State). Additionally, ticks were collected from amphibians and reptiles captured and road-killed in the Amazon biome, at Maranhão and Amapá States. Specimens of ticks were photographed under a Zeiss stereomicroscope (5.1 zoom). Map with the locations were made using the Qgis program. Overall, 1973 specimens of amphibians and reptiles were examined. A total of 927 ticks were collected: 98 larvae, 421 nymphs and 408 adults. Six species of ticks were identified: Amblyomma rotundatum and Amblyomma dissimile the most frequent, and Amblyomma cajennense sensu stricto, Amblyomma sculptum, Amblyomma nodosum and Amblyomma humerale, occasionally. Surprisingly, a total of twelve males of A. rotundatum were collected. Here we report new records of association between cold-blooded animals and ticks and reinforce the absence of A. dissimile in the Caatinga, Cerrado and Atlantic Forest biomes. Additionally, we report new records of A. rotundatum males on reptiles in the Amazon biome. This last record allows us to speculate about a possible association of A. rotundatum males with reptiles and the Amazon biome.

Validation of enzyme immunoassays for quantifying sex steroid hormones in tropical screech owls (Megascops choliba).

Androgens and estrogens are steroid hormones that regulate reproductive processes in both males and females. Monitoring plasma levels of these steroids or their metabolites present in feces, offers diagnostic support for assessing the reproductive status of animals. Immunoassays are commonly used methods for quantifying these hormones, but their protocols require species-specific validation to ensure reliability. The objective of this study was to perform analytically and biologically validation of enzyme immunoassay (EIA) kits for measuring testosterone (T), estradiol (E2), faecal androgen metabolites (fAM), and faecal estrogenic metabolites (fEM) in the tropical screech owl (Megascops choliba). Serum and fecal samples were collected from six adult females and six males both before and during breeding season, with males' gonadal activity assessed using electroejaculation (EE). The parallelism test confirmed the immunogenic similarity of the antigens in the estradiol and testosterone standards and the antigens in the serum samples and fecal extracts of M. choliba. Additionally, the EIA kits displayed nearly 100% recovery rates, and showed coefficients of variation ranging from 8% to 14% at the intra-assay level and from 10% to 16% at the inter-assay level, underscoring result reliability and consistency. In males, the highest serum T and fAM levels were recorded concurrently with the presence of spermatozoa in samples collected via EE. Although females did not exhibit oviposition events, significantly higher E2 and fEM levels were observed in August compared to May, suggesting potential seasonal variations in estrogenic hormone production. Fecal androgen and estrogen levels were significantly different between sexes in August, with males having higher fAM and females having higher fEM levels. Overall, the immunoassays validated in this study were found to be efficient in diagnosing reproductive activity in owls.

Epizootic amebiasis outbreak in wild black howler monkeys (Alouatta caraya) in a wildlife facility during captivity prior to translocation - Recommendations to wildlife management programs / Surto epizoótico de amebíase em Bugio preto (Alouatta caraya) provenientes de vida livre durante o cativeiro prévio a translocação - Recomendações a programas de manejo de fauna

The capture and quarantine of non-human primates could be necessary in some circumstances including those required under environmental permit. Mortality is undesirable for wildlife management programs and could be related to opportunistic pathogens, for example, deaths due to intestinal protozoa infection outbreaks as described here. Parasitological, necroscopic, microbiological, and molecular tests were used in the diagnosis of severe necrotic enteritis leading to death of three female and two male black howler monkeys (Alouatta caraya) captured and held in quarantine prior to translocation. Parasitological tests showed the presence of cysts of Entamoeba histolytica/dispar (5/5), Entamoeba coli (5/5) and Giardia duodenalis (1/5). Necroscopic assessment revealed areas of severe multifocal necrosis in the intestinal mucosa and submucosa. Histopathological examination revealed the presence of structures morphologically compatible to Entamoeba spp. in all individuals examined. Furthermore, G. duodenalis (1/5) was demonstrated by the nested PCR technique. During temporary captivity of non-human primates in management programs, proper handling protocols, including fast or immediate destination, are suggested in order to mitigate the negative effects of stress and decrease the risk of infections.(AU)

Óbitos por infecções causadas por protozoários intestinais são indesejáveis para programas de manejo de fauna, exigidos no âmbito do licenciamento ambiental e que demandam a captura e quarentena de primatas não humanos. Exames coproparasitológicos, necroscópicos, microbiológicos e moleculares foram utilizados para a confirmação do diagnóstico de uma severa enterite necrótica que levou a óbito três fêmeas e dois machos de Alouatta (A.) caraya capturados e mantidos em quarentena prévia à translocação. Exames coproparasitológicos revelaram a presença de cistos de Entamoeba (E.) histolytica/dispar (5/5), Entamoeba (E.) coli (5/5) e Giardia (G.) duodenalis (1/5). A avaliação necroscópica revelou áreas de necrose multifocal severa na mucosa e submucosa intestinal. A análise microscópica revelou a presença de estruturas morfologicamente compatíveis com trofozoítos do gênero Entamoeba spp. em todos os indivíduos examinados. Além disso, G. duodenalis (1/5) foi demonstrada pela técnica de Nested PCR. Sugere-se que quando for necessário o cativeiro temporário de primatas, deverão ser adotados protocolos de manejo adequados buscando a destinação imediata dos animais, de moto a mitigar os efeitos negativos do estresse e reduzir o risco da ocorrência de infecções.(AU)

Glial cells of the central nervous system of Bothrops jararaca (Reptilia, Ofidae): an ultrastructural study / Células gliais do sistema nervoso central de Bothrops jararaca (Reptilia, Ofidae): um estudo ultraestrutural

Although ultrastructural characteristics of mature neuroglia in the central nervous system (CNS) are very well described in mammals, much less is known in reptiles, especially serpents. In this context, two specimens of Bothrops jararaca were euthanized for morphological analysis of CNS glial cells. Samples from telencephalon, mesencephalon and spinal cord were collected and processed for light and transmission electron microscopy investigation. Astrocytes, oligodendrocytes, microglial cells and ependymal cells, as well as myelin sheaths, presented similar ultrastructural features to those already observed in mammals and tended to maintain their general aspect all over the distinct CNS regions observed. Morphological similarities between reptilian and mammalian glia are probably linked to their evolutionary conservation throughout vertebrate phylogeny...

Muito embora as características ultraestruturais da neuróglia madura do sistema nervoso central (SNC) sejam bem descritas em mamíferos, muito pouco é conhecido em répteis, especialmente em serpentes. Neste contexto, dois espécimes de Bothrops jararaca foram eutanasiados para análise morfológica das células gliais presentes no SNC. Amostras de telencéfalo, mesencéfalo e medula espinhal foram coletadas e processadas para investigação por microscopia de luz e eletrônica de transmissão. Astrócitos, oligodendócitos, células microgliais e ependimárias, bem como bainhas de mielina, apresentaram características ultraestruturais similares àquelas já observadas em mamíferos e tenderam a manter seu aspecto geral pelas diferentes regiões observadas no SNC. Similaridades morfológicas entre as células gliais de mamíferos e de répteis estão provavelmente ligadas a sua conservação evolutiva ao longo da filogenia dos vertebrados...

Prevalência de Cryptosporidium serpentis em serpentes de cativeiro / Prevalence of Cryptosporidium serpentis in captive snakes

Cryptosporidium é um protozoário encontrado em uma grande variedade de espécies animais como responsável por casos de gastrite e enterite, porém com epidemiologia pouco conhecida em animais silvestres. A presente investigação teve como objetivo avaliar a prevalência de Cryptosporidium serpentis em lavado gástrico de serpentes mantidas em cativeiro no serpentário do Instituto Butantan (São Paulo, Brasil). A coleta foi realizada uma semana após alimentação, evitando, assim, a regurgitação devido à manipulação. Foram realizados esfregaços do sedimento do lavado gástrico, obtido por centrifugação, corados pela técnica de coloração de Kinyoun. Parte do sedimento foi submetido à técnica de RFLP-PCR para identificação da espécie de Cryptosporidium. O serpentário é dividido em três seções, por espécie - a primeira com oito jibóias (Boa constrictor amarali), a segunda com dez jararacas (Bothropoides jararaca) e a última com sete cascavéis (Caudisona durissa). A prevalência de C. serpentis encontrada neste estudo para as serpentes C. durissa, B. jararaca e Boa c. amarali, foi de 57,14 por cento (04/07), 40 por cento (04/10) e 37,5 por cento (03/08), respectivamente, revelando importante ocorrência desse protozoário em serpentes de cativeiro. Apesar da alta prevalência encontrada, apenas as jiboias apresentaram sintomas como perda de peso e regurgitação, refletindo uma sensibilidade diferente da espécie para C. serpentis.

Cryptosporidium is a protozoan found in a wide variety of animal species which is responsible for gastritis and enteritis, but its epidemiology is poorly known in wild animals. The present investigation aimed to evaluate the prevalence of Cryptosporidium serpentis in gastric aspirate of captive snakes from the public serpentarium of the Butantan Institute (São Paulo, Brazil). Sampling was performed preferably one week after feeding, thereby preventing regurgitation due to manipulation. Smears were done from the gastric aspirate sediment obtained by centrifugation and stained by Kinyoun technique. Part of the pellet was submitted to RFLP-PCR technique for amplification of Cryptosporidium segment (833bp, CSP01) of SSU rRNA gene. The serpentarium was divided in three sections by species - the first housing eight Amaral´s Boa (Boa constrictor amarali), the second with ten jararacas (Bothropoides jararaca) and the last one with seven south american rattlesnakes (Caudisona durissa). The prevalence of C. serpentis found in this study for the snakes C. durissa, B. jararaca and B. constrictor was 57.14 percent (04/07), 40 percent (04/10) and 37.5 percent (03/08), respectively, thus revealing the high occurrence of this protozoan among captive snakes. Despite the high prevalence found, only B. constrictor amarali presented symptoms as regurgitation and weight loss, probably due to a different sensibility of this species to C. serpentis.

Semen collection and evaluation in free-ranging Brazilian rattlesnakes (Crotalus durissus terrificus).

Two hundred-ninety species of reptiles are estimated to need urgent action for conservation, with at least 113 threatened species worldwide. The International Union for Conservation of Nature and Natural Resources (IUCN) Red List of Threatened Species includes 80 species of snakes, with six native Brazilian species, a number likely to be an underestimation. Some authors believe that assisted reproduction would be an important tool to improve reproduction in captivity of some reptiles. An efficient technique for semen collection and evaluation is an important step in development of protocols for cryopreservation of semen or artificial insemination in snakes, contributing to the conservation of endangered species. Although these techniques are important, some basic semen parameters are described for four of the approximately 2,900 snake species in the world. The Brazilian rattlesnake (Crotalus durissus terrificus) was chosen as a model for semen collection in snakes because it is found quite often in Sao Paulo State. Semen was collected once from each animal by the same investigator during the mating season of this species in Brazil. After antiseptic cleansing of the skin around the cloaca, the snakes were injected subcutaneously with a dose of 15 mg/kg of 1% solution of lidocaine around the cloaca. Semen was collected with ventral massages after cloacal relaxation and directly from genital papilla inside the cloaca. A total of 28 ejaculates from 39 animals were obtained, representing collection efficiency of 71.80%. Semen volume and concentration in Brazilian rattlesnakes ranged from 3-70 microl and from 0.94-2.23 x 10(9) spermatozoa/ml, respectively. Zoo Biol 0:1-6, 2007. (c) 2007 Wiley-Liss, Inc.

O efeito das enzimas hialuronidase e tripsina na liquefação do sêmen de macacos pregos (Cebus apella) / The effect of hyaluronidase and trypsin ensymes on liquefaction of semen in capuchin monkey (Cebus apella)

Os efeitos das enzimas hialuronidase e tripsina foram avaliados quanto à liquefação, motilidade, vigor e integridade do acrossoma, no sêmen de seis macacos pregos (Cebus apeiia), mantidos na Fundação Parque Zoológico de São Paulo. O sêmen foi colhido por eletroejaculação, após anestesia geral, e a fração líquida foi imediatamente avaliada. A fração coagulada do sêmen foi tratada com as enzimas hialuronidase e tripsina, na dose de 1mg/ml, diluídas em meio 199 (Nutricel, Campinas/SP, Brasil), na proporção 1:4 e examinada após 5 e 15 minutos. Test t de Student foi utilizado para comparar os tratamentos. Não houve diferença significativa quanto a motilidade, vigor e integridade de acrossoma (p > 0.05), entre a fração de sêmen coagulado diluído em hialuronidase e tripsina, após cinco ou quinze minutos. No entanto, houve diferença significativa quanto a motilidade e vigor entre a fração líquida e a coagulada do sêmen (p < 0.05), após quinze minutos. Não houve diferença significativa com relação à integridade de acrossoma (p > 0.05) entre a fração líquida e coagulada do sêmen, após 15 minutos. De acordo com os resultados, podemos concluir que não houve efeito aparente na fração coagulada do sêmen tratado com as enzimas hialuronidase e tripsina com relação a motilidade, vigor e integridade do acrossoma. No entanto, houve diferença significativa entre a fração líquida e coagulada do sêmen com relação a motilidade e vigor, porém não quanto à integridade do acrossoma. De maneira geral, em ambos os tratamentos, não houve a completa dissolução do coágulo.

The effect of the enzymes hyaluronidase and trypsin were recorded on the motility, vigor and acrosome integrity in the semen of capuchin monkey (Cebus apella). The animals (n=6) were maintained at Fundação Parque Zoológico de São Paulo. Under anesthesia semen samples were collected by electroejaculation. Immediately after the ejaculation, the semen liquid fraction was analyzed for volume (ml), pH, motility (%), vigor (0-5), concentration (cells/ml), defects (%) and percentage of intact acrosome (%). The coagulated fraction was treated with a solution of hyaluronidase or trypsin, 1mg/ml in commercial medium (199-Nutricel, Campinas/SP, Brazil) in a proportion of 1:4 and the samples were examined after 5 and 15 minutes. The Student T Test (95%) was used to compare the treatments. There was no significant difference in the motility, vigor or acrossome integrity (p > 0.05) between coagulated fraction diluted either in trypsin or in hyaluronidase, after 5 or 15 minutes. However, there was significant difference in motility and vigor between liquid and coagulated fraction, after 15 minutes, for both treatments (p<0.05), but there were no difference in acrosome integrity (p > 0.05). In conclusion, there were no apparent effects in the coagulum for both treatments regarding motility, vigor and acrosome integrity. There were significant differences between liquid and coagulated fractions regarding motility and vigor, but not for acrosome integrity. In both enzyme treatments there were no complete dissolution of the coagulum.