Molecular survey of potentially pathogenic microorganisms in ticks collected from coatis (Nasua nasua) in Iguaçu National Park, Atlantic Forest biome, southern Brazil.

Human contact with wild animals in synanthropic habits is often mediated by arthropod vectors such as ticks. This is an important method of spreading infectious agents that pose a risk to human health. Thus, this study aimed to molecularly detect Ehrlichia spp., Anaplasma spp., Borrelia spp., and protozoa of the order Piroplasmida in ticks collected from coatis of Iguaçu National Park (PNI), Paraná, Brazil. This study involved 553 ticks DNA, including Amblyomma spp. larvae, Haemaphysalis juxtakochi nymphs, Amblyomma brasiliense, Amblyomma coelebs, and adults of Amblyomma ovale. The DNA extracted from each sample was subjected to polymerase chain reaction (PCR) targeting the genes 23S rRNA for the Anaplasmataceae family, 16S rRNA for Anaplasma spp., dsb for Ehrlichia spp., flaB, 16S rRNA, hpt, and glpQ for Borrelia spp., and 18S rRNA for Piroplasmid protozoans. DNA from Anaplasma sp. was detected in ticks of the species A. coelebs (4/553); Borrelia sp. DNA was detected in A. coelebs (3/553), A. ovale (1/553), and Amblyomma larvae (1/553); and Theileria sp. was detected in A. coelebs (2/553). All tested samples were negative for Ehrlichia spp. Our study constitutes the newest report in South America of these microorganisms, which remain poorly studied.

Detection of Rickettsia spp. in ring-tailed coatis (Nasua nasua) and ticks of the Iguaçu National Park, Brazilian Atlantic Rainforest.

Wild animals are of considerable importance in the ecology of infectious agents, as they can function as hosts and even as possible vectors. In this study, DNA from Rickettsia spp. was detected on ticks and fragments of skin collected from wild coatis with synanthropic habits in the Iguaçu National Park (INP) in the state of Paraná in southern Brazil. Testing was carried out on a total of 566 ticks, comprising Amblyomma spp. larvae, nymphs of Haemaphysalis juxtakochi, Amblyomma brasiliense, Amblyomma coelebs, and adults of Amblyomma ovale. The samples were tested by polymerase chain reaction (PCR) by amplifying htrA, gltA, ompA, and ompB gene fragments to detect Rickettsia spp. A fragment of each positive sample was sequenced in both directions, submitted to Genbank for a homology search, and also used for phylogenetic analyses. Samples of A. coelebs (1.90%, 8/420), A. ovale (13%, 6/45), and ring-tailed coati skin (1%, 1/75) amplified Rickettsia spp. DNA. Through sequencing, Rickettsia bellii was observed in A. ovale, Rickettsia amblyommatis in A. coelebs, while Rickettsia rhipicephali was detected in the skin samples. Wild ring-tailed coatis with synanthropic habits in the INP and their ticks are infected by Rickettsia spp., and associations with new hosts have been described.

Serological prevalence of Toxoplasma gondii infection in cats (Belém, Pará, Brazil).

We evaluated the prevalence of anti-Toxoplasma gondii antibodies in the serum samples collected from domestic cats in Belém, Pará, Brazil. We also correlated the presence of T. gondii antibodies with environmental variables and cat-owner habits. Four-hundred and forty-seven serum samples from domestic cats were analyzed. The sera were tested using an indirect immunofluorescence assay. Among the animals analyzed, 21.92% (98/447) were seropositive. A statistically significant association was found in relation to age and serology among the animals over 1 year old (p<0.01): in the group up to 1 year old, 12.82% (20/156) of the animals were positive, and in the group over 1 year old, 26.80% (78/291) were positive. Our results show that the cats in Belém, Pará region have anti-T. gondii antibodies, and their owners are not aware of toxoplasmosis or how to prevent its transmission.

Ticks in wild animals at a resort and the first documentation of Amblyomma sculptum infesting Emu (Dromaius novaehollandiae) / Carrapatos em animais selvagens em um resort e o primeiro registro de Amblyomma sculptum infestando Emu (Dromaius novaehollandiae)

Abstract The study aimed to identify species of ticks present in the environment and among captive animals, in Mangaratiba, Rio de Janeiro, Brazil. Ticks were isolated from captive animals by manual examination and free-living ticks in the environment were captured using the flannel drag technique. A total of 91 ticks were obtained (51 adults, 25 nymphs and 15 larvae). The specimens were identified morphologically as Amblyomma sp., Amblyomma dubitatum, Amblyomma ovale and Amblyomma sculptum , and were distributed among five species of native mammals and an exotic bird. This study also reports the first case of infestation of the Australian emu (Dromaius novaehollandiae) by A. sculptum.

Resumo O estudo teve como objetivo identificar espécies de carrapatos presentes no ambiente e entre animais de cativeiro em Mangaratiba, Rio de Janeiro, Brasil. Os carrapatos foram removidos manualmente de animais de cativeiro e no ambiente usando a técnica de arrasto em flanela. Um total de 91 carrapatos foram capturados (51 adultos, 25 ninfas e 15 larvas). Os espécimes foram identificados morfologicamente como Amblyomma sp., Amblyomma dubitatum, Amblyomma ovale e Amblyomma sculptum, e foram distribuídos entre cinco espécies de mamíferos nativos e uma ave exótica. Este estudo também relata o primeiro caso de infestação do emu australiano (Dromaius novaehollandiae) por A. sculptum .

Ticks in wild animals at a resort and the first documentation of Amblyomma sculptum infesting Emu (Dromaius novaehollandiae).

The study aimed to identify species of ticks present in the environment and among captive animals, in Mangaratiba, Rio de Janeiro, Brazil. Ticks were isolated from captive animals by manual examination and free-living ticks in the environment were captured using the flannel drag technique. A total of 91 ticks were obtained (51 adults, 25 nymphs and 15 larvae). The specimens were identified morphologically as Amblyomma sp., Amblyomma dubitatum, Amblyomma ovale and Amblyomma sculptum , and were distributed among five species of native mammals and an exotic bird. This study also reports the first case of infestation of the Australian emu (Dromaius novaehollandiae) by A. sculptum.

Morphological, molecular and phylogenetic characterization of Borrelia theileri in Rhipicephalus microplus.

This study aimed to perform a morphological, molecular and phylogenetic characterization of Borrelia theileri obtained from infected Rhipicephalus microplus in Brazil. Fifty engorged R. microplus females from cattle in the municipality of Seropédica, Rio de Janeiro, were analyzed for spirochetes by hemolymph smear. Macerated eggs and positive ticks, as well as blood from the bovine infested by these ticks, were analyzed the glpQ, flaB and hpt genes by PCR. The PCR products were purified and sequenced for analysis and construction of a phylogenetic tree. Only 2% (1/50) of the ticks generated a positive result by both smear and PCR. The spiral forms (n = 50) had (media ± SD) a mean length of 19.17 ± 4.12 µm, diameter of 0.2935 ± 0.0469 and number of turns 8.44 ± 2.59. Sequence alignments of the three evaluated genes exhibited 98% similarity to B. theileri isolates, occurring in a clade highly related to B. theileri strain KAT. Egg maceration samples were positive for the three evaluated genes, whereas bovine blood was negative by PCR. This is the most detailed characterization of B. theileri in the Americas to-date, presenting morphological, molecular and phylogenetic data, including the transovarial transmission of the spirochete in the host tick.

Morphological, molecular and phylogenetic characterization of Borrelia theileri in Rhipicephalus microplus / Caracterização morfológica, molecular e filogenética de Borrelia theileri em Rhipicephalus microplus

Abstract This study aimed to perform a morphological, molecular and phylogenetic characterization of Borrelia theileri obtained from infected Rhipicephalus microplus in Brazil. Fifty engorged R. microplus females from cattle in the municipality of Seropédica, Rio de Janeiro, were analyzed for spirochetes by hemolymph smear. Macerated eggs and positive ticks, as well as blood from the bovine infested by these ticks, were analyzed the glpQ, flaB and hpt genes by PCR. The PCR products were purified and sequenced for analysis and construction of a phylogenetic tree. Only 2% (1/50) of the ticks generated a positive result by both smear and PCR. The spiral forms (n = 50) had (media ± SD) a mean length of 19.17 ± 4.12 µm, diameter of 0.2935 ± 0.0469 and number of turns 8.44 ± 2.59. Sequence alignments of the three evaluated genes exhibited 98% similarity to B. theileri isolates, occurring in a clade highly related to B. theileri strain KAT. Egg maceration samples were positive for the three evaluated genes, whereas bovine blood was negative by PCR. This is the most detailed characterization of B. theileri in the Americas to-date, presenting morphological, molecular and phylogenetic data, including the transovarial transmission of the spirochete in the host tick.

Resumo O estudo teve como objetivo realizar a caracterização morfológica, molecular e filogenética de Borrelia theileri obtida de Rhipicephalus microplus naturalmente infectado em bovino no estado do Rio de Janeiro, Brasil. Um total de 50 fêmeas de R. microplus ingurgitadas foram analisadas para espiroquetas por meio de esfregaço de hemolinfa. Ovos macerados e carrapatos, assim como sangue de bovinos infectados por esses carrapatos, foram analisados os genes glpQ, flaB e hpt por PCR. Os produtos de PCR foram purificados e sequenciados para análise e construção de uma árvore filogenética. Apenas 2% (1/50) dos carrapatos geraram um resultado positivo tanto pelo esfregaço como pela PCR. As formas espirais (n = 50) apresentaram (média ± DP) comprimento médio de 19,17 ± 4,12, diâmetro de 0,2935 ± 0,0469 e número de voltas de 8,44 ± 2,59. Os alinhamentos das sequências dos três genes avaliados exibiram 98% de similaridade aos isolados de B. theileri, ocorrendo em um clado altamente relacionado à linhagem de B. theileri KAT. As amostras de maceração de ovos foram positivas para os três genes avaliados, enquanto o sangue bovino foi negativo pela PCR. Esta é a mais completa caracterização de B. theileri nas Américas, apresentando dados morfológicos, moleculares e filogenéticos, incluindo a transmissão transovarial da espiroqueta no carrapato hospedeiro.

Trypanosoma cruzi infection in captive Neotropical primates in the Brazilian Amazon.

The aim of this study was to detect the infection by Trypanosoma cruzi in captive Neotropical primates in the Brazilian Amazon. From February 2013 to July 2014, 112 blood samples were collected from Neotropical primates from the Amazonas, Amapá, and Pará States, north of Brazil. The subjects belonged to the families Cebidae (N = 59), Atelidae (N = 41), Callitrichidae (N = 5), Pitheciidae (N = 4), and Aotidae (N = 3). Blood smears also were examined for the presence of trypomastigotes by optical microscopy. For the detection of T. cruzi DNA, a Nested-PCR with primers TCZ1/TCZ2 and TCZ3/TCZ4 was performed. T. cruzi DNA was detected in 12.5% (14/112) of Neotropical primates examined. Positive samples were detected in 16%, 12.5%, and 11.11% of the different species of primates sampled from the Amapá, Pará, and Amazonas states, respectively. The analysis of the blood smears did not reveal trypomastigote forms of T. cruzi. In conclusion, Neotropical primates kept in captivity were infected by T. cruzi in the studied areas. We recommend that a health management protocol be put into place to prevent the transmission of infectious agents among captive populations, captive and wild populations, and between NHPs and the technicians who handle these animals.

Filarial nematodes with zoonotic potential in ring-tailed coatis (Nasua nasua Linnaeus, 1766, Carnivora: Procyonidae) and domestic dogs from Iguaçu National Park, Brazil.

Iguaçu National Park, which contains the largest remnant of Atlantic Forest in southern Brazil and Iguaçu Falls as one of its main visiting points, is a year-round attraction for thousands of domestic and international tourists. Ring-tailed coatis are generalist, opportunistic carnivores that benefit from human association. These animals are the most abundant carnivores in this park, where they come into close contact with tourists and the resident population around the park. Moreover, as the park is surrounded by human dwellings, free roaming domestic dogs are frequently present, favoring the exchange of pathogens with wild animals. Wild carnivores are known to be infected with several pathogens, from viruses to arthropods, some of them passed on by domestic carnivores. Among the nematodes that infect wild carnivores, those of the Onchocercidae family are of concern due to their zoonotic potential. The objective of this research was to assess the prevalence of filarial nematodes in coatis and dogs in Iguaçu National Park, as well as the potential impact of infection on host health and body condition. To this end, 75 coatis and 50 adult dogs were captured in the aforementioned area. Seven species of filarioids, Dirofilaria immitis, Dirofilaria repens, Acanthocheilonema reconditum, Brugia sp., Mansonella sp. and two undetermined species, confirmed by histochemical tests, were diagnosed in the area. Knott's concentration test in coatis and dogs showed a prevalence of 81.6% and 16.0%, respectively. Seven microfilarial morphotypes were diagnosed in the coatis and two morphotypes were found in the dogs. A specific immunoassay test for Dirofilaria immitis revealed the presence of D. immitis infection in 1.33% of the coatis and 22% of the domestic dogs. The parasitic infection had negligible effects on the body condition of both dog and coati hosts, but an increase was found in eosinophil counts in coatis with filarial infection, as well as decreases in hematocrit and hemoglobin in the infected domestic dogs. These findings represent new locality and host records for all the filarioids diagnosed in the area of this study, with some of the diagnosed parasites, given their zoonotic potential, representing a health risk for park visitors and the local population.

Occurrence of eggs and oocysts of gastrointestinal parasites in passerine birds kept in captivity in Para State, Brazil / Ocorrência de ovos e oocistos de parasitos gastrointestinais em aves Passeriformes mantidas em cativeiro no estado do Pará, Brasil

ABSTRACT: The objective of this study was to detect helminth eggs and protozoan oocysts in samples of feces from birds of the order Passeriformes in Para State, Brazil. Fecal samples were collected individually from 403 passerine birds seized and kept in captivity in Para State. Samples were processed by the double centrifugation technique in saturated sucrose solution and the coccidial oocyst-positive samples were submitted to sporulation in potassium dichromate 2.0%. Helminth eggs and/or protozoan oocysts were observed in 43.18% (174/403) of the fecal samples examined. Coccidial oocysts were detected in 93.68% (163/174) of the positive samples, whereas helminth eggs were observed in 10.34% (18/174) of the positive samples. Oocyst sporulation occurred in 43.56% (71/163) of the samples, and only Isospora spp. oocysts were detected. Nematode eggs of the superfamilies Trichostrongyloidea (4.60%; 8/174), Ascaridoidea (0.57%; 1/174), and Trichuroidea (0.57%; 1/174) were diagnosed in the positive samples. Cestoda eggs were diagnosed in 2.87% (5/174), whereas Trematoda eggs were detected in 2.30% (4/174) of positive samples. Passerine birds seized and kept in captivity in the visited local presented parasitism by intestinal helminths and protozoan, with a predominance of infection with coccidia of the gender Isospora.

RESUMO: O objetivo do presente estudo foi detectar ovos de helmintos e oocistos de protozoários em amostras de fezes de aves da ordem Passeriformes no estado do Pará, Brasil. Amostras de fezes foram coletadas individualmente de 403 aves Passeriformes oriundas de apreensão e mantidas em cativeiro no estado do Pará. As amostras foram processadas usando a técnica de dupla centrifugação em solução saturada de sacarose e as amostras positivas para oocistos de coccídios foram submetidas à esporulação em dicromato de potássio 2,0%. Ovos de helmintos e/ou oocistos de protozoários foram observados em 43,18% (174/403) das amostras fecais examinadas. Oocistos de coccídios foram detectados em 93,68% (163/174) das amostras positivas, enquanto que ovos de helmintos foram observados em 10,34% (18/174). A esporulação de oocistos ocorreu em 43,56% (71/163) das amostras, e somente oocistos de Isospora spp. foram detectados. Ovos de nematódeos das Superfamílias Trichostrongyloidea (4,60%; 8/174), Ascaridoidea (0,57%; 1/174) e Trichuroidea (0,57%; 1/174) foram diagnosticados nas amostras positivas. Ovos de Cestoda foram diagnosticados em 2,87% (5/174), enquanto que ovos de Trematoda foram detectados em 2,30% (4/174) das amostras positivas. Aves Passeriformes oriundas de apreensão e mantidas em cativeiro nas áreas visitadas estavam parasitadas por helmintos e protozoários, predominando a infecção por coccídios do gênero Isospora.

Molecular detection of Trypanosoma evansi (Kinetoplastida: Trypanosomatidae) in procyonids (Carnivora: Procyonidae) in Eastern Amazon, Brazil / Detecção molecular de Trypanosoma evansi (Kinetoplastida: Trypanosomatidae) em procionídeos (Carnivora: Procyonidae) na Amazônia Oriental, Brasil

ABSTRACT: The present study aimed to diagnose the natural infection of captive and free-living procyonids with Trypanosoma evansi in the states of Amapá and Pará, Brazil. From February 2012 to August 2013, whole blood samples and blood smears were obtained from 45 free-living procyonids and from nine procyonids kept in captivity in wild life refuges and zoobotanical parks in the states of Amapá and Pará. Whole blood samples were collected and kept at -20ºC for the detection of T. evansi DNA by PCR using the RoTat 1.2 forward and RoTat 1.2 reverse primers. In addition, the blood smears were processed and examined for the presence of trypomastigote forms of T. evansi. T. evansi DNA was detected in 18.52% (10/54) of the procyonids, namely, in captive crab-eating raccoons and captive and free-living coatis in Pará State. No trypomastigote forms were observed in the blood smears. DNA from T. evansi was detected in P. cancrivorus and N. nasua in Pará State, being this the first such report in P. cancrivorus.

RESUMO: O objetivo do presente trabalho foi realizar o diagnóstico da infecção natural por Trypanosoma evansi em procionídeos de vida livre e de cativeiro dos estados do Amapá e Pará, Brasil. Durante o período de fevereiro de 2012 a agosto de 2013, amostras de sangue total e esfregaços sanguíneos foram obtidos de 45 procionídeos de vida livre e de nove mantidos em cativeiro em mantenedores e Parques Zoobotânicos dos estados do Amapá e Pará. As amostras de sangue total foram coletadas e mantidas a -20ºC para pesquisa de DNA de T. evansi pela PCR utilizando-se os iniciadores RoTat 1.2 forward e RoTat 1.2 reverse. Os esfregaços sanguíneos também foram processados e examinados para a pesquisa de formas tripomastigotas do agente. DNA de T. evansi foi detectado em 18,52% (10/54) dos procionídeos, ocorrendo em mãos-peladas de cativeiro e quatis de vida livre e de cativeiro no estado do Pará. Não foram observadas formas tripomastigotas nos esfregaços sanguíneos. DNA de T. evansi foi detectado em P. cancrivorus e N. nasua no estado do Pará, sendo este o primeiro relato em P. cancrivorus.