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Equine piroplasmosisis, a tick-borne disease caused by the intra-erythrocytic protozoans Babesia caballi and Theileria equi, has economic importance due to the international trade and the increased movement of horses all over the world. The goal of this study was to evaluate the occurrence of phylogenetic diversity of T. equi and B. caballi genotypes among infected equids from São Luís Island, state of Maranhão, northeastern Brazil. Between December of 2011 and June of 2012, EDTA-blood and serum samples were collected from 139 equids (90 donkeys, 39 horses and 10 mules). From 139 serum samples submitted to ELISA assay, IgG antibodies to T. equi and B. caballi were detected in 19.4% (27/139) and 25.2% (35/139), respectively. Among sampled animals, 21.6% (30/139) and 55.4% (77/139) were positive for cPCR assays for T. equi and B. caballi, based on ema-1 and rap-1 genes, respectively. Overall, the T. equi sequences (n=7) submitted to Maximum Likelihood analysis (based on a 18S rRNA fragment of 1700 bp after alignment) grouped into three main groups, which were subdivided in eight clusters. The present work showed that different genotypes of T. equi and B. caballi circulate among equids in Brazil. Resumo A piroplasmose equina, uma doença transmitida por carrapatos e causada pelos protozoários intra-eritrocíticos Babesia caballi e Theileria equi, tem importância econômica devido ao comércio internacional e ao aumento do movimento de cavalos em todo o mundo. O objetivo do presente estudo foi mostrar a diversidade filogenética de T. equi e B. caballi infectando cavalos, burros e jumentos na Ilha de São Luís, Estado do Maranhão, Nordeste do Brasil. Entre dezembro de 2011 e junho de 2012, amostras de sangue com EDTA e soro de foram coletadas de 139 equídeos (90 jumentos, 39 cavalos e 10 burros). Dentre as 139 amostras de soro submetidas ao ensaio de ELISA, foram detectados anticorpos IgG contra T. equi e B. caballi em 19,4% (27/139) e 25,2% (35/139), respectivamente. Entre os animais amostrados, 21,6% (30/139) e 55,4% (77/139) foram positivos por meio dos ensaios de cPCR para T. equi e B. caballi, com base nos genes ema-1 e rap-1, respectivamente. No geral, as sequências T. equi (n = 7) submetidas à análise de Máxima Verossimilhança (baseada em um fragmento do 18S rRNA de 1700 pb, após o alinhamento) foram agrupadas em três grupos principais, os quais foram subdivididos em oito grupos. O presente trabalho mostrou que diferentes genótipos de T. equi e B. caballi circulam entre equídeos no Brasil.

Made available in DSpace on 2018-12-11T17:34:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-07-01. Added 1 bitstream(s) on 2019-10-09T18:33:00Z : No. of bitstreams: 1 S1984-29612017000300352.pdf: 2139150 bytes, checksum: 0300af9bba769c4ea378914230bfcf26 (MD5) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Hepatozoon species are the most common intracellular hemoparasite found in reptiles. Hepatozoon caimani, whose vectors are Culex mosquitoes, has been detected in a high prevalence among caimans in Brazil by blood smears examinations. The present work aimed to detect and characterize the Hepatozoon spp. found in 33 caimans (24 free-ranging and 9 captive; 28 males and 5 females) (Caiman crocodilus yacare) sampled at Poconé, North Pantanal, state of Mato Grosso, Brazil, using blood smears examinations and molecular techniques. Hepatozoon spp.-gametocytes were found in 70.8% (17/24) and 88.8% (8/9) of blood smears from free-ranging and captive caimans, respectively. Hepatozoon spp. 18S rRNA DNA was found in 79.2% (19/24) and 88.8% (8/9) of free-ranging and captive caimans, respectively. Comparative analysis of parasitized and non-parasitized erythrocytes showed that all analyzed features were significantly different (P

Made available in DSpace on 2018-12-11T17:32:12Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-06-01 New genotypes of Anaplasmataceae agents have been detected in wild carnivores, birds and deer in Brazil. The present work aimed to investigate the presence of Ehrlichia and Anaplasma species in rodents sampled in Brazil. Additionally, a newly designed quantitative 5′ nuclease real-time multiplex PCR for Ehrlichia and Anaplasma spp. detection based on groEL gene amplification was designed, showing high specificity and sensitivity (10 groEL fragment copy/μL). Between 2000 and 2011, different rodent species [n = 60] were trapped in 5 Brazilian biomes. Among 458 rodent spleen samples, 0.4% (2/458) and 2.4% (11/458) were positive for Ehrlichia and Anaplasma spp., respectively. Of 458 samples, 2.0% (9/458) and 1.1% (5/458) were positive for Anaplasma sp. and Ehrlichia sp., respectively, using conventional 16S rRNA PCR assays. Maximum Likelihood phylogenetic analyse based on a small region of 16S rRNA genes positioned the Anaplasma genotypes in rodents near Anaplasma phagocytophilum or Anaplasma marginale and Anaplasma odocoilei isolates. Ehrlichia genotypes were closely related to E. canis. There was a low occurrence of Anaplasma and Ehrlichia in wild and synanthropic rodents in Brazil, suggesting the circulation of new genotypes of these agents in rodents in the studied areas. Universidade Estadual Paulista (Unesp) Faculdade de Ciências Agrárias e Veterinárias, Jaboticabal Johns Hopkins School of Medicine Uniformed Services University Laboratório de Hantaviroses e Rickettsioses Instituto Oswaldo Cruz/Fiocruz Laboratório de Biologia de Tripanosomatídeos Instituto Oswaldo Cruz/Fiocruz Laboratório de Biologia e Parasitologia de Mamíferos Silvestres Reservatórios Instituto Oswaldo Cruz/Fiocruz Universidade Estadual Paulista (Unesp) Faculdade de Ciências Agrárias e Veterinárias, Jaboticabal

Dioctophyme renale is a nematode that can be found parasitizing the kidney, peritoneal cavity and, rarely, other organs of canids and mustelids. This disease has high occurrence in the municipality of Três Barras, state of Santa Catarina, thus making this an interesting area to study the epidemiological aspects of infection by D. renale in dogs. Among 197 dogs, 14.2% showed the parasite eggs in urine and 16.4% showed IgG antibodies anti-D. renale in serum samples according to the indirect ELISA method; among seropositive dogs, 15 (37.5%) animals did not show any parasite eggs in their urine. Parasitism was more frequent in females, and there was no finding of interference from age on parasitism. Factors such as water potential and presence of paratenic hosts in the studied region were reported by the owners of dogs and may have contributed to the occurrence of parasitism. Resumo Dioctophyme renale é nematoda parasita de rins, cavidade peritoneal e, mais raramente, de outros órgãos de canídeos e mustelídeos. Esta parasitose tem ocorrência elevada no Município de Três Barras - SC, tornando essa área interessante para o estudo de aspectos epidemiológicos da infecção por D. renale em cães. De 197 cães examinados, 14,2% apresentaram ovos do parasita na urina e 16,4% apresentaram anticorpos IgG anti-D. renale no soro pelo método de ELISA indireto e, entre estes, 15 (37,5%) não apresentavam ovos do parasita sendo eliminados pela urina. O parasitismo foi mais frequente nas fêmeas, e não houve constatação de interferência da idade no parasitismo. Fatores como potencial hídrico e presença de hospedeiros paratênicos na região estudada foram relatados pelos tutores dos cães e podem contribuir para a ocorrência do parasitismo.

This study aimed to detect Mycoplasma spp. in naturally infected cats from Rio de Janeiro and to evaluate hematological abnormalities and factors associated with this infection. Out of the 197 cats sampled, 11.2% presented structures compatible with hemoplasma organisms on blood smears. In contrast, 22.8% were positive for Mycoplasma spp. by means of 16S rRNA gene real-time polymerase chain reaction, which reflects the weak concordance between techniques. The infection rates, by means of 16S rRNA gene conventional polymerase chain reaction, was 4.6%, 4.6% and 11.7% for Mycoplasma haemofelis (Mhf), ‘Candidatus Mycoplasma turicensis’ (CMt) and ‘Candidatus Mycoplasma haemominutum’ (CMhm), respectively. Mhf and CMhm infections are more frequent in the summer (p>0.05). Presence of anemia (p < 0.02), lymphocytosis (p < 0.03), thrombocytopenia (p < 0.04) and activated monocytes (p < 0.04) was associated with Mhf infection. No hematological abnormality was associated with CMt or CMhm infection. Male cats were more prone to be infected by Mhf or CMhm (p < 0.01). Adult cats had more chance to be infected by CMhm. Three hemoplasma species occur in the metropolitan region of Rio de Janeiro and Mhf seems to be the most pathogenic of them. Anemia is the most important hematological abnormality. Resumo Este estudo teve por objetivo detectar Mycoplasma spp. em gatos naturalmente infectados do Rio de Janeiro e avaliar as alterações hematológicas e fatores associados à infecção. Dos 197 gatos amostrados, 11,2% apresentaram estruturas compatíveis com hemoplasmas em esfregaços de sangue. Em contraste, 22,8% foram positivas para Mycoplasma spp. por meio da reação em cadeia da polimerase em tempo real (qPCR), baseado no gene 16S rRNA, o que reflete a fraca concordância entre as técnicas. As taxas de infecção, por meio da reação em cadeia da polimerase convencional baseada no gene 16S rRNA, foi de 4,6%, 4,6% e 11,7% para Mycoplasma haemofelis (Mhf), 'Candidatus Mycoplasma turicensis' (CMt) e 'Candidatus Mycoplasma haemominutum' (CMhm), respectivamente. Infecção por Mhf e CMhm foram mais frequentes no verão (p> 0,05). Anemia (p

Water buffalo (Bubalus bubalis) is a potential reservoir for Babesia bovis and B. bigemina in tropical regions, but the epidemiological evidence of their reservoir competence is limited, especially due to the lack of diagnostic tests capable of detecting and quantifying the low-level parasitemia present in the carrier animals. In this paper we present the standardization process of a SYBR Green based real-time PCR system (qPCR), consisting of two single qPCR assays, for the detection and quantification of B. bovis and/or B. bigemina. Both assays were optimized in similar protocols, including reagent concentrations and thermocycling parameters, so it is possible its use as a multiple qPCR in a single run. Both single assays showed a suitable analytical performance, especially by allowing detection of a greater number of carrier animals when compared with nested PCR assays (nPCR) against a reference panel of 60 DNA samples extracted from blood of both, infected- and non-infected buffaloes. Furthermore, a mathematical algorithm to convert the qPCR outcomes in percent of infected red blood cell was used, and was found that the estimated parasitemia in carrier buffaloes within the reference sample panels were close to those described in carrier cattle. This method could be a useful tool for epidemiological studies on the participation of the bubaline specie in the epidemic process of bovine babesiosis.

Made available in DSpace on 2020-12-12T01:49:16Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-10-01. Added 1 bitstream(s) on 2021-07-15T15:25:44Z : No. of bitstreams: 1 S1984-29612019000400661.pdf: 2536900 bytes, checksum: 4c798b71b1ab142f8c99e3da2ee244d4 (MD5) We report the first documented case of endocarditis associated with Bartonella clarridgeiae in a dog in Latin America. Infective vegetative valvular aortic endocarditis was diagnosed in a 10-year-old male mixed breed dog. The dog presented grade V/VI systolic and diastolic murmur, hyperthermia, and progressive weight loss. Cardiomegaly and presence of diffuse alveolar pattern in the lung fields were observed in the thorax radiography evaluation. Irregular and hyperechogenic structures adhered to the aortic leaflets, causing obstruction of the left ventricular outflow tract and severe aortic insufficiency, were observed in the echocardiography evaluation. A vegetative, whitish, hardened structure measuring 1.0 cm in diameter was observed in aortic semilunar valve at necropsy. Based on a combination of pre-enrichment insect-based medium liquid culture, quantitative real-time and conventional PCR assays based on nuoG and gltA genes, respectively, followed by sequencing and phylogenetic inferences, B. clarridgeiae DNA was detected in the patient’s aortic valve lesions. Clinical, echocardiographic, anatomopathologic and molecular features supported the diagnosis of severe aortic vegetative endocarditis possibly caused by B. clarridgeiae in a dog in Brazil. Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP Serviço de Cardiologia Veterinária Hospital Veterinário e Departamento de Clínica e Cirurgia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP Serviço de Patologia Veterinária Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP Serviço de Cardiologia Veterinária Hospital Veterinário e Departamento de Clínica e Cirurgia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP Serviço de Patologia Veterinária Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP

Made available in DSpace on 2020-12-12T01:49:10Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-10-01. Added 1 bitstream(s) on 2021-07-15T15:25:44Z : No. of bitstreams: 1 S1984-29612019000400816.pdf: 636244 bytes, checksum: 20f143aba89c4bc602ed57410f222396 (MD5) Toxoplasma gondii and Neospora caninum are Apicomplexan intracellular protozoan parasites that affect numerous animal species, thus leading to severe diseases and economic losses, depending on the vertebrate species involved. The role of the avian species in maintaining and transmission of these coccidia has been studied for several years as they tend to serve as a potential source of infection for mammals and humans. The present study aimed to assess the serological exposure of Orinoco goose (Neochen jubata) to T. gondii and N. caninum. Between 2010 and 2013, 41 free-ranging Orinoco geese were captured in the Araguaia River, Brazil. The presence and titration of IgY antibodies to both coccidia were assayed via indirect immunofluorescent antibody test (IFAT). While IgY antibodies for N. caninum were present in 5 animals, with titers of 20, the antibodies for T. gondii were found in 35 animals, with titers ranging from 20 to 640. Considering that the Orinoco goose’s meat is consumed by the local population in the studied area, it may represent an important source of T. gondii infection for humans. Due to its migratory behavior, this goose may play a pivotal role in the natural dispersion of both parasites. Furthermore, molecular studies are required for genotyping the isolates of T. gondii that occurs in this avian species. Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias-FCAV Universidade Estadual Paulista-UNESP Laboratório de Patologia de Animais Selvagens Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias-FCAV Universidade Estadual Paulista-UNESP Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias-FCAV Universidade Estadual Paulista-UNESP Laboratório de Patologia de Animais Selvagens Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias-FCAV Universidade Estadual Paulista-UNESP

Made available in DSpace on 2019-10-06T16:41:56Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-10-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Hemotropic mycoplasmas (hemoplasmas) are Gram-negative bacteria that parasitize the erythrocyte surface of a wide variety of mammals. The present study aimed at investigating the occurrence of hemoplasmas in beef cattle in the Brazilian Pantanal, an area endemic for bovine trypanosomiasis in South America. Additionally, the objective of this study was to characterize molecularly the genotypes of the found hemoplasmas. For this purpose, blood and serum samples of 400 beef cattle were collected from five properties in Corumbá, Nhecolândia sub-region, Mato Grosso do Sul, in Midwest Brazil. Blood samples underwent DNA extraction and standard 16S rRNA gene-based PCR assays for hemoplasmas. The sequences obtained were submitted to phylogenetic inferences, distance analysis, and genotype diversity. The Indirect Enzyme-Linked Immunoabsorbent Assay (iELISA) indicated the presence of anti-Trypanosoma vivax IgG antibodies in 89.75% of the animals sampled, confirming the endemicity of said agent in the studied region. Among the 400 bovine blood samples tested, 2.25% (9/400) were positive for hemoplasmas in cPCR. The phylogenetic analysis of the obtained sequences confirmed the presence of 'Candidatus Mycoplasma haemobos' and Mycoplasma wenyonii DNA in 0.5% (2/400) and 1.75% (7/400) animals, respectively. Five genotypes of M. wenyonii and one of 'Candidatus M. haemobos' were detected among the sequenced amplicons. The present study showed low molecular occurrence of haemoplasmas in beef cattle sampled in the Brazilian Pantanal, an area endemic for bovine trypanosomiasis. Despite of the conservation of the 16S rRNA gene, there was considerable diversity of hemoplasma genotypes infecting the sampled beef cattle. Graduate Program in Agricultural Microbiology Universidade Estadual Paulista (FCAV/UNESP) Laboratory of Immunoparasitology Department of Veterinary Pathology Faculdade de Ciências Agrárias e Veterinárias/ Universidade Estadual Paulista (FCAV/UNESP) Universidade Católica Dom Bosco (UCDB) Graduate Program in Agricultural Microbiology Universidade Estadual Paulista (FCAV/UNESP) Laboratory of Immunoparasitology Department of Veterinary Pathology Faculdade de Ciências Agrárias e Veterinárias/ Universidade Estadual Paulista (FCAV/UNESP)

Made available in DSpace on 2019-10-06T17:10:52Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-09-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coxiella burnetii (order Legionellales, family Coxiellaceae), the etiological agent of Q fever, is a pleomorphic, obligate Gram-negative intracellular bacillococcus that can infect humans and animals. Among the mammals hosting this agent, both domestic and wild ruminants are of particular economic and public health importance. Ticks and other hematophagous arthropods or aerosols are incriminated in the transmission between reservoirs and susceptible hosts. This study used serological and molecular methods to investigate the C. burnetii occurrence in blood samples from free-living deer (143 Blastocerus dichotomus, 27 Mazama gouazoubira, 4 M. bororo, 3 M. americana, and 11 Ozotocerus bezoarticus)sampled in Mato-Grosso do Sul, São Paulo, Goiás and Paraná states in Brazil. The DNA extracted from the blood samples of 188 cervids was submitted to nested (n)PCR for C. burnetii based on the IS1111 repetitive insertion element of the heat shock protein (htpAB)gene. Additionally, 169 serum samples were submitted to Indirect Immunofluorescence Assay (IFAT)to detect Ig antibodies to C. burnetii. The nPCR results indicated that all blood samples were negative, evidencing the absence of circulating C. burnetii DNA in cervids from the studied regions or, alternatively, the C. burnetii DNA concentration in the deer blood samples was below the threshold of the used PCR technique. On the other hand, 5.32% (9/169)of deer were seropositive for C. burnetii by IFAT, with titers ranging from 256 and 16,384. In conclusion, the present work showed the first evidence of exposure of free-living deer to C. burnetii in Brazil. Programa de Pós-graduação em Microbiologia Agropecuária Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Patologia Veterinária Universidade Estadual Paulista “Júlio de Mesquita Filho” (UNESP) Departamento de Zootecnia Universidade Estadual Paulista - Júlio de Mesquita Filho Departamento de Medicina Veterinária Preventiva e Saúde Animal Universidade de São Paulo Programa de Pós-graduação em Microbiologia Agropecuária Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Patologia Veterinária Universidade Estadual Paulista “Júlio de Mesquita Filho” (UNESP) Departamento de Zootecnia Universidade Estadual Paulista - Júlio de Mesquita Filho

Made available in DSpace on 2019-10-06T16:30:41Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-07-01 Babesia bovis is the etiological agent of bovine babesiosis, a disease transmitted by Rhipicephalus microplus, which affects cattle herds in tropical and subtropical regions of the world, causing significant economic losses due to decreasing meat and milk yield. This study used molecular techniques to determine the occurrence and genetic diversity of B. bovis, based on the genes encoding the spherical body protein (sbp-2) and the merozoite surface antigens (MSAs) genes, in a herd of 400 Nellore (Bos indicus) sampled from beef cattle farms in the Pantanal region, state of Mato Grosso do Sul, Midwestern Brazil. The results of the nested PCR assays based on the sbp-2 gene indicated that 18 (4.5%) calves were positive for B. bovis; out of them, while 77.7% (14/18) were positive for the B. bovis msa-2b fragment, 66.6% (12/18) were positive for the msa-2c fragment. The phylogenetic analysis based on the maximum likelihood method using 14 sequences from msa-2b clones and 13 sequences from msa-2c clones indicated that the sequences detected in this study are clearly distributed in different cladograms. These findings corroborated the diversity analysis of the same sequences, which revealed the presence of 14 and 11 haplotypes of the msa-2b and msa-2c genes, respectively. Furthermore, the entropy analyses of amino acid sequences revealed 78 and 44 high entropy peaks with values ranging from 0.25 to 1.53 and from 0.27 to 1.09 for MSA-2B and MSA-2C, respectively. Therefore, the results indicate a low molecular occurrence of B. bovis in beef cattle sampled in the Brazilian Pantanal. Despite this, a high degree of genetic diversity was found in the analyzed B. bovis population, with possibly different haplotypes coexisting in the same animal and/or in the same studied herd. Agricultural and Livestock Microbiology Graduation Program School of Agricultural and Veterinarian Sciences São Paulo State University (UNESP) Laboratory of Immunoparasitology Department of Veterinary Pathology Faculdade de Ciências Agrárias e Veterinárias/Universidade Estadual Paulista (FCAV/UNESP), Via de Acesso Prof. Paulo Donato Castellane, s/n, Zona Rural Universidade Católica Dom Bosco (UCDB) Agricultural and Livestock Microbiology Graduation Program School of Agricultural and Veterinarian Sciences São Paulo State University (UNESP) Laboratory of Immunoparasitology Department of Veterinary Pathology Faculdade de Ciências Agrárias e Veterinárias/Universidade Estadual Paulista (FCAV/UNESP), Via de Acesso Prof. Paulo Donato Castellane, s/n, Zona Rural

Livestock infections by Trypanosoma vivax have been occurring with increasing frequency, mainly due to the presence of animals with subclinical infections and without apparent parasitaemia, making diagnosis challenging. The aim of the present study was to evaluate several techniques used for T. vivax diagnosis in order to assess the best way of using them during the course of the disease. Molecular methods demonstrated higher rates of detection than parasitological methods, detecting 33 of the 54 (61.1%) known positive samples, while the hematocrit centrifugation technique (best parasitological test) detected only 44.4%. The serological methods, IFAT and ELISA, detected seropositivity in 51 of the 54 (94.4%) and 49 of the 54 (90.7%) known positive samples, respectively. Despite being highly sensitive, the latter only demonstrates exposure to the infectious agent and does not indicate whether the infection is active. The present study was the first to use the qPCR for a South American isolate, improving disease detection and quantification. Furthermore, the analyses revealed that the patent phase of the disease may extend up to 42 days, longer than previously reported. The combination of several diagnostic techniques can lower the frequency of false negative results and contributes toward better disease control. Resumo Infecções por Trypanosoma vivax têm ocorrido com frequência crescente em animais de produção, principalmente pela aquisição de animais com infecções subclínicas e sem aparente parasitemia, o que dificulta o diagnóstico. O objetivo do presente estudo foi avaliar várias técnicas empregadas para o diagnóstico de T. vivax, a fim de verificar a melhor maneira de utilizá-las durante o curso da doença. Os métodos moleculares demonstraram maiores taxas de detecção que os métodos parasitológicos, detectando 33 das 54 (61,1%) amostras sabidamente positivas, enquanto a técnica de hemoconcentração (melhor teste parasitológico) detectou apenas 44,4%. Os métodos sorológicos, RIFI e ELISA, detectaram soropositividade em 51 das 54 (94,4%) e 49 das 54 (90,7%) amostras sabidamente positivas, respectivamente. Apesar de serem altamente sensíveis, estes testes apenas demonstram a exposição ao agente infeccioso, e não indicam se a infecção permanece ativa. O presente estudo foi o primeiro a utilizar a qPCR para um isolado sul-americano, melhorando sua detecção e quantificação. Além disso, as análises revelaram que a fase patente da doença pode se estender por até 42 dias após a infecção, sendo maior que anteriormente relatado. A combinação de várias técnicas de diagnóstico pode evitar a frequência de resultados falso-negativos e contribuir para um melhor controle da doença.

Made available in DSpace on 2019-10-03T17:31:48Z (GMT). No. of bitstreams: 0 Previous issue date: 2019. Added 1 bitstream(s) on 2019-10-04T16:20:52Z : No. of bitstreams: 1 S1984-29612019005015105.pdf: 964122 bytes, checksum: f7759749575a21f127f915f0287fcde3 (MD5) Resumo Um lobo guará (Chrysocyon brachyurus) adulto, macho, de vida livre foi encaminhado para atendimento no hospital veterinário Governador “Laudo Natel” – FCAV/Unesp após ser encontrado com lesões de pele e fratura em membro pélvico direito, sendo amputado devido a comprometimento da integridade do membro. Aproximadamente 20 dias após a chegada ao hospital, foi notado acentuado aumento de volume bilateral em região de articulação úmero-rádio-ulnar. O líquido sinovial foi drenado e enviado para análise citológica com coloração de Rosenfeld, revelando a presença de neutrófilos degenerados com cromatina cariolítica associados a inclusões intracelulares sugestivas de gametócitos de Hepatozoon sp. Amostras de sangue e líquido sinovial foram submetidas a análises moleculares visando amplificar um fragmento do gene 18S rRNA de Hepatozoon spp. Apesar da sequência de Hepatozoon detectada se posicionar filogeneticamente no mesmo clado que H. canis previamente detectado em carnívoros domésticos, o resultado da análise do BLAST mostrou somente 98% de identidade com H. canis. De acordo com o conhecimento dos autores, esta é a primeira vez que Hepatozoon foi detectado no líquido sinovial por meio de patologia clínica e análises moleculares. Abstract A free-living, adult male maned wolf (Chrysocyon brachyurus) was referred to the Governador “Laudo Natel” – FCAV/Unesp veterinary hospital after being found with skin lesions and a fracture on the right pelvic limb, which had to be amputated due to compromised integrity. Around 20 days later, bilateral accentuated swollen on humerus-radius-ulna articulation was observed. The synovial liquid was drained and sent to the laboratory for synovial cytology with Rosenfeld staining that revealed predominantly degenerated neutrophils with karyolytic chromatin associated with intracellular inclusions suggestive of Hepatozoon sp. gametocytes. Blood and synovial liquid samples were submitted to molecular analysis, aiming to amplify the Hepatozoon spp. 18S rRNA gene fragment. Despite the positioning of the found Hepatozoon sequence together with Hepatozoon canis previously detected in domestic carnivores, the BLAST analysis showed only 98% identity with H. canis. To the best of the authors’ knowledge, this is the first time a Hepatozoon was detected in the synovial liquid by clinical pathology and molecular analyses. Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Patologia Veterinária Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Clínica e Cirurgia Veterinária Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Patologia Veterinária Universidade Estadual Paulista Júlio de Mesquita Filho Departamento de Clínica e Cirurgia Veterinária

Made available in DSpace on 2019-10-06T16:40:06Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-04-01. Added 1 bitstream(s) on 2019-10-09T18:35:32Z : No. of bitstreams: 1 S1984-29612019000200306.pdf: 593217 bytes, checksum: 48766ce7e53219a76e4b29b5eededae8 (MD5) Fundação de Amparo à Pesquisa e ao Desenvolvimento Científico e Tecnológico do Maranhão Universidade Estadual Paulista Mycoplasma suis is a bacterium that causes hemoplasmosis in pigs. This agent is capable of adhering to the surface of porcine erythrocytes, inducing structural changes on these cells. In Brazil, there are few reports about the disease, its causal agent, and the economic impact of this pathogen on pig production systems and farm sanitation. The present study aimed to investigate the occurrence of M. suis in extensive swine farms located in the counties of Itapecuru Mirim, Santa Rita and Rosario, State of Maranhão, northeast Brazil. For such purpose, 64 blood samples of pigs from these facilities were tested for M. suis using a 16S rRNA gene-based quantitative real-time PCR (qPCR); 82.3%, 65.2% and 25% of blood samples of swine from farms in the cities of Itapecuru Mirim, Santa Rita and Rosario were positive for M. suis by qPCR, respectively. This study shows, for the first time, that M. suis circulates in pig populations from the state of Maranhão, Northeast Brazil. Universidade Estadual do Maranhão – UEMA Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista – UNESP Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista – UNESP

The msp4 gene of A. marginale is unicodon, stable and mostly homogeneous, being considered as a useful marker for phylogeographic characterization of this bacterium. The objective of this work was to analyze the phylogeography of A. marginale based on the msp4 gene in beef cattle from the Brazilian Pantanal, compared to those found in other regions worldwide. The blood samples investigated were collected from 400 animals (200 cows and 200 calves) reared in five extensive breeding farms in this region. The results indicated that of the evaluated samples, 56.75% (227/400) were positive for A. marginale based on the msp1β gene by quantitatitve PCR (qPCR), while 8.37% (19/227) were positive for the msp4 gene in the conventional PCR. In the Network distance analysis, 14 sequences from the Brazilian Pantanal were grouped into a single group with those from Thailand, India, Spain, Colombia, Parana (Brazil), Mexico, Portugal, Argentina, China, Venezuela, Australia, Italy and Minas Gerais (Brazil). Among 68 sequences from Brazil and the world, 15 genotypes were present while genotype number one (#1) was the most distributed worldwide. Both Splitstree and network analyses showed that the A. marginale msp4 sequences detected in beef cattle from the Brazilian Pantanal showed low polymorphism, with the formation of one genogroup phylogenetically related to those found in ruminants from South and Central America, Europe, and Asia. Resumo O gene msp4 de A. marginale é unicodon, estável e pouco heterogêneo, sendo considerado como um marcador útil para caracterização filogeográfica desta bactéria. Este trabalho teve como objetivo analisar a filogeografia de A. marginale com base no gene msp4 em bovinos de corte do Pantanal Brasileiro, comparativamente a outra regiões do mundo. Alíquotas de sangue foram colhidas de 400 bovinos (200 vacas e 200 bezerros) em cinco propriedades de cria e recria extensiva. Como resultado, 56,75% (227/400) mostraram-se positivas para A. marginale pela qPCR para o gene msp1β e destas, 8,37% (19/227) amostras foram positivas na PCR convencional para o gene msp4. Na análise de distância Network, 14 sequências do Pantanal brasileiro foram agrupadas em um único grupo com as da Thailândia, Índia, Espanha, Colômbia, Paraná (Brasil), México, Portugal, Argentina, China, Venezuela, Austrália, Italia e Minas Gerais (Brasil). Dentre 68 sequências do Brasil e do mundo, constatou-se a presença de 15 genótipos, sendo o genótipo número um (#1) o mais distribuído. As sequências msp4 de A. marginale detectadas em bovinos de corte no Pantanal brasileiro apresentaram baixo polimorfismo com formação de dois genogrupos filogeneticamente relacionados àqueles encontrados em ruminantes de países das América do Sul e Central, Europa e Ásia.

Recently, an increasing number of Bartonella species have been emerged to cause human diseases. Among animal reservoirs for Bartonella spp., bats stand out due to their high mobility, wide distribution, social behaviour and long-life span. Although studies on the role of vampire bats in the epidemiology of rabies have been extensively investigated in Latin America, information on the circulation and genetic diversity of Bartonella species in these bat species is scarce. In the present work, 208 vampire bats, namely Desmodus rotundus (the common vampire bat; n = 167), Diphylla ecaudata (the hairy-legged vampire bat; n = 32) and Diaemus youngii (the white-winged vampire bat; n = 9) from 15 different states in Brazil were sampled. DNA was extracted from liver tissue samples and submitted to real-time PCR (qPCR) and conventional PCR (cPCR) assays for Bartonella spp. targeting five genetic loci, followed by phylogenetic and genotype network analyses. Fifty-one out of 208 liver samples (24.51%) were positive for Bartonella DNA in the ITS real-time PCR assay [40 (78.43%) of them were from D. rotundus from 11 states, and 11 (21.57%) samples from D. ecaudata from three states. Eleven genotypes were found for each gltA and rpoB genes. Several ITS sequences detected in the present study clustered within the lineage that includes B. bacilliformis and B. ancachensis. The Bayesian phylogenetic inference based on the gltA gene positioned the obtained sequences in six different clades, closely related to Bartonella genotypes previously detected in D. rotundus and associated ectoparasites sampled in Latin America. On the other hand, the Bartonella rpoB genotypes clustered together with the ruminant species, B. schoenbuchensis and B. chomelii. The present study describes for the first time the molecular detection of Bartonella spp. in D. ecaudata bats. It also indicates that Bartonella spp. of vampire bats are genetically diverse and geographically widespread in Brazil.

Trypanosoma vivax is the most pathogenic ruminant’s hemoparasite, causing huge economic losses to the producer when prevalent in the herd. This study aims to characterize the trypanosomiasis in naturally infected cattle in order to assess the clinical findings, epidemiological risk factors and diagnosis of this disease in outbreaks occurred in the states of Pernambuco and Alagoas, northeastern Brazil. For this purpose, historical and clinical examinations, blood collection and clinical monitoring were performed in 109 animals presenting illness for a period of one year. The main clinical findings were fever, apathy, anorexia, diarrhea, progressive weight loss, lymphadenopathy, pale mucous, incoordination, aggressiveness, abortion, decrease in milk production and high mortality. Out of 109 animals, 94% (103/109) were seropositive to Trypanosoma vivax by ELISA, 92% (100/109) were seropositive for IFAT, 15% (17/109) were positive in conventional PCR assay based on cathepsine L gene, and 9% (10/109) were showed the presence of Trypanosoma vivax trypomastigotes in stained-blood smears. In 13 samples, blood counts revealed that 69.23% (9/13) showed normocytic normochromic anemia and 53.84% (7/13) had leukocytosis, in 46.15% (6/13) neutrophilia and shift left regenerative. In 72 samples also examined whether it hematocrit, plasma protein and fibrinogen. Hematocrit presented average 22% (10% to 37%) to plasma protein had a mean value of 7.55 g/dl (5.4 g/dl to 10.0 g/dl) and plasma fibrinogen had a mean value 700mg/dl (200mg/dl to 1600mg/dl). The sequencing of DNA samples revealed 100% identicalness to T. vivax by BLAST analysis. The main factors involved in the spread of the disease were: the ingress of animals without prior tests and quarantine in the properties, application of oxytocin with sharing needles contaminated by blood at the time of lactation and the restriction of the disease in the lactation herd. The transmission of T. vivax occurred with the sharing of needles among animal’s lactation during application of oxytocin after the entry of carrier animals in the herd. Serological tests demonstrated a high rate of seropositive animals, however, conventional PCR and blood smear revealed a low rate positive animal’s because the animals were already being treated with trypanocide drugs. It is concluded that trypanosomiasis is a disease to be considered in the cattle regions studied as the cause of outbreaks. Hygienic and security actions must be taken during the administration ofoxytocin in lactating cows. In fact, this practice, when carried incorrectly and without hygienic criterions and sharing needles, may play a role as the main risk factor in the transmission of trypanosomiasis in dairy herds among regions where the disease is prevalent, increasing the challenge of reinfection of animals., O Tripanosoma vivax é o hemoparasito mais patogênico para ruminantes, causando grandes perdas econômicas para o produtor quando prevalente no rebanho. Este trabalho se propõe a caracterizar a tripanossomose em bovinos através dos achados clínicos, fatores epidemiológicos de risco e diagnóstico dessa doença em surtos ocorridos no agreste pernambucano e alagoano, Brasil. Nas propriedades foram realizados levantamentos dos históricos e exames clínicos, coletas de sangue e acompanhamento clínico de 109 animais doentes por um período de um ano. Os achados clínicos foram: febre, apatia, anorexia, diarreia, emagrecimento progressivo, linfoadenomegalia, mucosas pálidas, incoordenação motora, agressividade, aborto, queda na produção de leite e alta mortalidade. Das 109 amostras utilizadas, 94% (103/109) foram soropositivas no ELISA, 92% (100/109) foram soropositivas para RIFI, 15% (17/109) positivas para PCR convencional e 9% (10/109) foram evidenciadas presença do Tripanosoma vivax no esfregaço sanguíneo. Em 13 amostras, foram realizados hemogramas o que revelou que 69,23% (9/13) apresentaram anemia normocítica normocromica e 53,84% (7/13) apresentaram leucocitose, em 46,15% (6/13) neutrofilia e desvio a esquerda regenerativo. Em 72 amostras analisou-se também o hematócrito, proteína plasmática e o fibrinogênio. O hematócrito, teve valor médio de 22% (10% a 37%), a proteína plasmática teve valor médio de 7,55 g/dl (5,4g/dl a 10,0g/dl) e o fibrinogênio plasmático teve valor médio de 700mg/dl (200mg/dl a 1600mg/dl). O sequenciamento das amostras de DNA revelou 100% de similaridade com T. vivax. Os principais fatores envolvidos na propagação da doença foram: a entrada de animais portadores do T. vivax nas propriedades sem exames prévios e quarentena, a aplicação de ocitocina com compartilhamento de agulhas contaminadas por sangue no momento da ordenha e a restrição da doença ao rebanho em lactação. Os testes sorológicos apresentaram uma melhor taxa de detecção de animais soropositivos, entretanto, o PCR e o esfregaço sanguíneo revelaram uma baixa taxa de detecção em virtude de os animais já estarem sendo medicados continuamente com drogas tripanocidas. Conclui-se que a tripanossomose é uma doença a ser considerada na bovinocultura das regiões estudadas como causadora dos surtos. Medidas higiênicas e de seguridade devem ser adotadas durante a administração de ocitocina nas vacas em lactação, sendo esta prática, quando realizada de forma errada e sem critérios higiênicos e com compartilhamento de agulhas, o principal fator de risco na transmissão da tripanossomose em rebanhos leiteiros em regiões onde a doença é prevalente, aumentando o desafio de reinfecções entre os animais.

Made available in DSpace on 2020-12-12T01:53:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-01-01 Cervids represent a mammal group which plays an important role in the maintenance of ecological balance. Recent studies have highlighted the role of these species as reservoirs for several arthropods-borne pathogens. Globally, hemotropic mycoplasmas (haemoplasmas) are emerging or remerging bacteria that attach to red blood cells of several mammals species causing hemolytic anaemia. Therefore, the aim of this study was to investigate the occurrence and assess the phylogenetic positioning of Mycoplasma ovis in free-ranging deer from Brazil. Using a polymerase chain reaction targeting the 16S rRNA region, 18 (40%) out of 45 sampled deer were positive to M. ovis. Among the nine sequences analysed, four distinct genotypes were identified. The sequences detected in the present study were closely related to sequences previously identified in deer from Brazil and the USA. On the other hand, the Neighbour-Net network analysis showed that the human-associated M. ovis genotypes were related to genotypes detected in sheep and goats. The present study shows, for the first time, the occurrence of M. ovis in Mazama gouazoubira and Mazama bororo deer species, expanding the knowledge on the hosts harbouring this haemoplasma species. Once several deer species have your population in decline, additional studies are needed to evaluate the pathogenicity of M. ovis among deer populations around the world and assess its potential as reservoir hosts to human infections. Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Universidade Estadual Paulista (Unesp) Faculdade de Ciencias Agrarias e Veterinarias Núcleo de Pesquisa e Conservacąo de Cervídeos Departamento de Zootecnia Universidade Estadual Paulista (Unesp) Faculdade de Ciências Agrárias e Veterinárias Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Universidade Estadual Paulista (Unesp) Faculdade de Ciencias Agrarias e Veterinarias Núcleo de Pesquisa e Conservacąo de Cervídeos Departamento de Zootecnia Universidade Estadual Paulista (Unesp) Faculdade de Ciências Agrárias e Veterinárias

This is a cross-sectional study to assess the presence of antibodies in ruminants against selected pathogens associated with reproductive disorders in cattle in four Brazilian states, including the zoonotic agent Coxiella burnetii. The used tests were Virus Neutralization Assay for IBR and BVD, Microscopic Agglutination Test for Leptospira spp., Indirect Fluorescent Antibody Test (IFAT) for C. burnetii and Toxoplasma gondii, and Enzyme-Linked Immunosorbent Assay for Neospora caninum and Trypanosoma vivax. Seropositivity for C. burnetii was 13.7% with titers from 128 to 131,072; 57.8% for BoHV-1, with titers between 2 and 1,024; 47.1% for BVDV-1a, with titers from 10 to 5,120; 89.2% for N. caninum; 50% for T. vivax; and 52.0% for Leptospira spp., with titers between 100 to 800 (the following serovars were found: Tarassovi, Grippotyphosa, Canicola, Copenhageni, Wolffi, Hardjo, Pomona and Icterohaemorrhagiae); 19.6% for T. gondii with titer of 40. This is the first study that has identified C. burnetii in cattle associated with BoHV and BVDV, N. caninum, Leptospira spp., T. gondii and T. vivax. Thus, future studies should be conducted to investigate how widespread this pathogen is in Brazilian cattle herds. Resumo Este é um estudo transversal para avaliar a presença de anticorpos em ruminantes contra patógenos selecionados e associados a distúrbios reprodutivos em bovinos de quatro estados brasileiros, incluindo o agente zoonótico Coxiella burnetii. Os testes utilizados foram Teste de Vírus-Neutralização para BoHV e BVDV, teste de Aglutinação Microscópica para Leptospira spp., Reação de Imunofluorescência Indireta for C. burnetii e Toxoplasma gondii, e Ensaio de Imunoabsorção Enzimática para Neospora caninum e Trypanosoma vivax. A soropositividade para C. burnetii foi de 13,7% com títulos de 128 a 131.072; 57,8% para BoHV-1, com títulos entre 2 a 1.024; 47,1% para BVDV-1a, com títulos de 10 a 5.120; 89,2% para N. caninum; 50% para T. vivax; e 52,0% para Leptospira spp., com títulos entre 100 a 800 (sorovares encontrados: Tarassovi, Grippotyphosa, Canicola, Copenhageni, Wolffi, Hardjo, Pomona e Icterohaemorrhagiae) 19,6% para T. gondii com título de 40. Este é o primeiro estudo que evidencia a participação de C. burnetii em bovinos associada ao Vírus da Rinotraqueíte bovina infecciosa e da diarreia viral bovina, N. caninum, Leptospira spp., T. gondii e T. vivax em bovinos. Desta forma, futuros estudos devem ser conduzidos a fim de investigar o quão disseminado se encontra este patógeno em rebanhos bovinos brasileiros.

Objectives The objective of this study was to develop a quantitative 5' nuclease real-time polymerase chain reaction (PCR) assay to diagnose infections caused by Bartonella species. Methods Between January and April 2013 whole blood samples were collected by convenience from 151 cats (86 domiciled and 65 stray cats). The feline blood samples were subjected to a novel quantitative 5' nuclease real-time PCR (qPCR) for Bartonella species targeting the nictonamide adenine dinucleotide dehydrogenase gamma subunit ( nuoG) and conventional PCR assays targeting intergenic transcribed spacer, ribC, gltA, pap31 and rpoB, followed by sequencing and basic local alignment search tool analysis. Results The qPCR assay detected as few as 10 copies of plasmid per reaction. Forty-six (54.4% domiciled and 45.6% stray cats) of 151 sampled cats showed positive results in nuoG qPCR for Bartonella species. The absolute quantification of nuoG Bartonella DNA in sampled cats ranged from 1.1 × 104 to 1.3 × 104. Eighteen (39.1%) of 46 positive samples in the qPCR were also positive in conventional PCR assays. The sequencing confirmed that Bartonella henselae and Bartonella clarridgeiae circulate in cats in midwestern Brazil. Conclusions and relevance The present work provides details of a novel qPCR assay to diagnose infections caused by Bartonella species.

Major surface protein 1a has been used as a marker for genetic stability in identifying geographical isolates of Anaplasma marginale. We conducted a longitudinal study to ascertain the genetic diversity of A. marginale in a dairy cattle herd with a history of clinical anaplasmosis. A total of 20 calves were evaluated every 3 months, from birth to 1 year of age. They were evaluated using blood smears, IFAT, ELISA, and qPCR. Additionally, samples positive for the msp1a gene using nPCR were sequenced. The detection of A. marginale ranged from 20 to 90% using blood smears, 20-80% using ELISA/IFAT, and 15-100% using qPCR. We found evidence that suggested transplacental transmission of A. marginale in 15% (3/20) of the calves based on qPCR and 20% (4/20) based on blood smears. Additionally, these four animals were A. marginale ELISA/IFAT-positive. The A. marginale strains found were α-β(3)-Γ (n=7), α-β(2) (n=1), and α-β(2)-190-Γ (n=1). The new MSP1a tandem repeat 190 was described. The results showed that the genetic diversity of A. marginale in a group of calves up to 1 year of age from Taiaçu (SP) was low, with only three different strains identified, showing the microsatellite genotype E. Eighty percent of the animals evaluated had clinical signs of bovine anaplasmosis and were treated using oxytetracycline and imidocarb dipropionate. Additionally, it was found that 30% (6/20) and 10% (2/20) of the animals required a second and third treatment, respectively, based on clinical signs. Four animals infected with the α-β(3)-Γ strain died, showing that this strain is also involved in the clinical anaplasmosis cases in Brazil.

Babesiosis is a tick-transmitted disease that causes severe economic losses to the cattle industry in Brazil. Water buffaloes (Bubalus bubalis) are often carriers of Babesia spp., but there are no studies that provide an accurate estimation of this infection in animals raised in regions of endemic stability. This study was conducted to investigate Babesia bovis and B. bigemina infections in 108 water buffaloes (50 calves and 58 adult females) located in areas of S&atildeo Paulo state, where the animals were continuously exposed to Rhipicephalus microplus ticks. B. bovis and B. bigemina infections were screened by microscopic examination of blood smears, nested PCR (nPCR) and quantitative real-time PCR (qPCR), which were also used to estimate the number of copies (NC) of the cytochrome b (mt-cytB) gene in the blood samples. B. bigemina was found in blood smears of three calves from Alambari herd (all with less than 0.1% parasitemia). Molecular techniques were more sensitive than blood smears to diagnose piroplasms in water buffaloes: 20.37% and 100.00% for B. bovis-infected animals and 59.26% and 100.00% for B. bigemina-infected animals, respectively for nPCR and qPCR. The NC of mt-cytB gene of B. bovis and B. bigemina in blood samples revealed significant effects (p < 0.05) of herd-age, species and their interaction. The NC values were higher (p ≤ 0.05) for B. bigemina (2.80 ± 0.06) than for B. bovis (2.61 ± 0.05). Within each herd-age, differences between the species’ NC values were found only in Alambari calves, which showed significantly higher (p ≤ 0.05) NC of B. bigemina (3.48 ± 0.13). The calves and cows from Ibate showed the lowest NC of B. bigemina (2.29 ± 0.13 and 2.63 ± 0.14) and B. bovis (2.54 ± 0.11 and 2.37 ± 0.12), respectively. These data suggest a high prevalence of B. bovis and B. bigemina infection in the buffalo population in endemic areas of S&atildeo Paulo state.

Trypanosoma vivax affects cattle herds in Africa and Americas and has been spreading rapidly in Brazil, through introduction of animals with subclinical infections and without apparent parasitemia, which makes its diagnosis challenging. PCR and LAMP are effective in detecting the presence of T. vivax DNA in situations of low parasitemia. LAMP is simpler and faster technique than PCR, and can be performed in the field, with limited resources. In this study, the capacities of conventional PCR and LAMP for detecting T. vivax in bovine blood samples classified as aparasitemic were evaluated. The capacity of conventional PCR (56.25%) for detecting positive samples was lower than that of LAMP (93.73%). This may influence the choice of screening tests for cattle herds infected with T. vivax.

Made available in DSpace on 2020-12-12T02:11:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2020-07-01 Universidade Estadual Paulista Worldwide, Bartonella species are known to infect a wide range of mammalian and arthropod hosts, including humans. The current study aimed to investigate the prevalence of Bartonella spp. in synanthropic mammals captured in peri-urban areas from Central-Western and Southern Brazil and their ectoparasites. For this aim, 160 mammals belonging to four species, and 218 associated arthropods were sampled. DNA was extracted and subjected to different Bartonella screening assays. Additionally, blood samples from 48 small rodents were submitted to liquid BAPGM culture followed by qPCR assay and solid culture. Two out of 55 Rattus captured in Santa Catarina state were PCR-positive for Bartonella when targeting the nuoG, 16S, and ITS loci. Sequences showed high homology with Bartonella coopersplainsensis. Conversely, all 48 small rodents, 14 capybaras and 43 opossum DNA samples from animals trapped in Mato Grosso do Sul were Bartonella negative in the HRM real time PCR assays targeting the ITS locus and gltA gene. Additionally, all mammal-associated ectoparasites showed negativity results based on HRM real time PCR assays. The present study showed, for the first time, the occurrence of B. coopersplainsensis in Brazil, shedding some light on the distribution of rats-related Bartonella in South America. In addition, the majority of rodents and marsupials were negative for Bartonella spp. Since B. coopersplainsensis reservoirs - Rattus spp. - are widely dispersed around the globe, their zoonotic potential should be further investigated. Graduate Program of Agricultural and Livestock Microbiology Faculty of Agrarian and Veterinary Sciences (FCAV/UNESP) Department of Veterinary Pathology Faculty of Agrarian and Veterinary Sciences (FCAV/UNESP) Koret School of Veterinary Medicine The Robert H. Smith Faculty of Agriculture Food and Environment. The Hebrew University of Jerusalem Dom Bosco Catholic University Department of Veterinary Medicine University of Contestado Federal University of Mato Grosso do Sul Graduate Program of Agricultural and Livestock Microbiology Faculty of Agrarian and Veterinary Sciences (FCAV/UNESP) Department of Veterinary Pathology Faculty of Agrarian and Veterinary Sciences (FCAV/UNESP)

Made available in DSpace on 2019-10-06T16:22:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-06-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) There are few studies on the genetic diversity of Anaplasma marginale in Brazilian cattle herds, especially about beef cattle. The objective of this study was to evaluate the genetic diversity of A. marginale, based on the msp1α gene in Bos taurus indicus sampled from the Brazilian Pantanal. Aliquots of blood with and without EDTA were taken from 400 cattle (200 cows and 200 calves) across five extensive farms. The samples were submitted to the indirect immunoenzymatic assay (iELISA), quantitative real-time PCR (qPCR) for the msp1β gene and to the semi-nested (sn) PCR for the msp1α gene. Positive samples were sequenced by the Sanger method and subjected to diversity analysis using the RepeatAnalyser software. The percentage of positive animals by iELISA, qPCR and (sn) PCR was 72.2% (289/400), 56.7% (227/400) and 23% (52/227), respectively. Cows (154/200) showed to be significantly more seropositive than calves (135/200). In qPCR, the number of calves and average quantification value (138/200; 1.3 × 106) A. marginale msp1α copies per μL proved to be higher when compared to that found for the cows (89/200; 3.9 × 104). The microsatellite analysis of the 26 sequences obtained from the msp1α gene revealed the presence of E (77%), C (15.4%) and B (7.7%) genotypes. Fourteen A. marginale strains were identified in the studied region, with eight that have never before been described in the literature (τ-10-13−13-18; τ-27-18; EV8-EV8-17; α-β-β-β-100; EV7-11−10-15; τ-11-11−27-18; τ-11-10-15; τ-27-13-18). Beef cattle are highly exposed to A. marginale in the Brazilian Pantanal. Moreover, a high genetic diversity of A. marginale, with eight new strains, was found in the studied region. While cows may act as chronic carriers, perpetuating the pathogen within the herd, male beef calves sold to other regions may disperse these strains. Universidade Estadual Paulista (Unesp) Faculty of Agrarian and Veterinary Sciences Don Bosco Catholic University (UCDB), Campo Grande Graduate Program in Veterinary Medicine Animal Pathology Host-Parasite Relationship State University of São Paulo (UNESP) Faculty of Agrarian and Veterinary Sciences UNESP Jaboticabal Universidade Estadual Paulista (Unesp) Faculty of Agrarian and Veterinary Sciences Graduate Program in Veterinary Medicine Animal Pathology Host-Parasite Relationship State University of São Paulo (UNESP) Faculty of Agrarian and Veterinary Sciences UNESP Jaboticabal

Made available in DSpace on 2019-10-06T17:08:27Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-05-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Canine monocytic ehrlichiosis (CME) is a disease caused by the obligate intracellular bacterium Ehrlichia canis. Tropical lineages of Rhipicephalus sanguineus ticks play an essential role in the transmission of this pathogen. The aim of the present study was to evaluate the prevalence of E. canis DNA in tissue from R. sanguineus ticks in areas endemic for CME in Brazil and quantify levels of E. canis DNA in dissected tissues from these samples. A total of 720 ticks were collected from 72 dogs (36 dogs from the city Araçatuba in São Paulo state and 36 from Campo Grande in the state of Mato Grosso do Sul). Ticks were dissected to collect the guts, ovaries and salivary gland. A quantitative polymerase chain reaction (qPCR) targeting the disulphide bond formation (dsb) protein gene was performed to quantify the level of E. canis infection. The E. canis dsb-qPCR assay was positive for 31.9, 10, and 15.2% of the gut, ovary, and salivary glands, respectively. The average gut, ovary, and salivary gland bacterial load estimated by qPCR was 1.21 × 103, 2.60 × 103, and 4.92 × 103 gene copies/µl, respectively. This is the first report of E. canis DNA in ovaries of R. sanguineus ticks parasitizing dogs in these CME-endemic areas. These observations raise the possibility of E. canis trans-ovarial transmission. Universidade Estadual Paulista (Unesp) Faculdade de Medicina Veterinária de Araçatuba, Clóvis Pestana St., 793 - Dona Universidade Estadual Paulista Júlio de Mesquita Filho (UNESP) Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal, Via de Acesso Prof. Paulo Donato Castelane University of Campinas (UNICAMP) Cidade Universitária Zeferino Vaz, Barão Geraldo United States Department of Agriculture Agricultural Research Service Knipling-Bushland U. S. Livestock Insects Research Laboratory, 2700, Fredericksburg Road Universidade Estadual Paulista (Unesp) Faculdade de Medicina Veterinária de Araçatuba, Clóvis Pestana St., 793 - Dona Universidade Estadual Paulista Júlio de Mesquita Filho (UNESP) Faculdade de Ciências Agrárias e Veterinárias de Jaboticabal, Via de Acesso Prof. Paulo Donato Castelane FAPESP: 2014/26461-7

Made available in DSpace on 2019-10-04T12:37:14Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-04-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Mycoplasma spp. and Bartonella spp. are Gram-negative bacteria transmitted by arthropod vectors that infect red blood cells of several mammal species. This study investigated the occurrence and genetic diversity of hemoplasmas and Bartonella spp. in 68 howler monkeys kept in captivity in Sao Paulo, a southeastern state in Brazil. In addition, possible hematological, biochemical and electrophoretic changes of serum proteins associated with the occurrence of hemoplasmas and Bartonella spp. in captive primates were also investigated. The cPCR results showed that all sampled howler monkeys were negative for Bartonella spp. based on the gltA gene. The cPCR results indicated that 18 (26.47%) non-human primates (NHP) were positive for hemoplasmas based on the 16S rRNA gene. Monocyte and lymphocyte counts were higher in hemoplasma-positive howlers (P < 0.05). Platelet counts decreased in nonhuman primates (NHP) positive for hemoplasmas (P < 0.05). The results from the blood serum proteinogram and biochemistry analyses were not significantly different between NHPs positive and negative for hemotrophic mycoplasmas. Phylogenetic analysis using Bayesian Inference (BI) based on the 16S rRNA gene positioned the obtained sequences close to 'Candidatus Mycoplasma kahanei'. The analysis of sequence diversity of the 16S rRNA gene showed that 5 different genotypes are circulating in NHP in Brazil and in the world; besides, a clear separation between the sequences of hemoplasmas that infect NHP of the Sapajus and Alouatta genus in Brazil was found, probably corresponding to two different species. The pathogenic potential of this hemoplasma species in NHP should be further investigated. Univ Estadual Paulista, Dept Vet Pathol, Lab Immunoparasitol, Jaboticabal, SP, Brazil Univ Estadual Paulista, Dept Genet & Anim Breeding, Jaboticabal, SP, Brazil Ctr Univ Moura Lacerda, Dept Vet Clin Pathol, Ribeirao Preto, SP, Brazil Univ Estadual Paulista, Dept Vet Pathol, Lab Immunoparasitol, Jaboticabal, SP, Brazil Univ Estadual Paulista, Dept Genet & Anim Breeding, Jaboticabal, SP, Brazil CNPq: 302420/2017-7

Made available in DSpace on 2018-11-26T17:55:26Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-09-17 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) There are two distinct lineages of ticks, Rhipicephalus sanguineus, in South America: tropical and temperate lineages. Only the tropical lineage is recognized as competent vector for Ehrlichia canis. The epidemiological data of canine monocytic ehrlichiosis is congruent with the distribution of the two lineages of R. sanguineus. Herein, we report the infection of R. sanguineus (tropical lineage) cell cultures with E. canis, after cryopreservation. R. sanguineus (tropical lineage) cell identity was confirmed by sequencing using a 16S rDNA gene fragment. Tick cell cultures were prepared in L-15B medium supplemented with 10%, 15%, and 20% Fetal Bovine Serum (FBS), and 10% of Tryptose Phosphate Broth (TPB). Cell cultures developed better at the concentration of 20% of FBS. Cultures in the fifth harvest (approximately 7 months later) were selected for the first infections. Optimal R. sanguineus cell growth and adhesion was observed (5.0x10(6) cells/mL, and the population doubling time every 57h). Once infected with E. canis, the cultures were maintained in L-15B medium supplemented with 2% and 5% of FBS fortified with iron and 10% TPB. Infected cells were also cryopreserved. DNA was extracted from infected and noninfected cells and analyzed using quantitative real-time PCR targeting the E. canis-dsb gene. Primary culture of the fifth passage was infected by E. canis and it maintained the pathogen for at least 40 days before partial cell destruction. Subcultures of infected cells (fresh and cryopreserved cultures) onto new tick cell cultures were successful. The E. canis infection was confirmed by real-time PCR and light and transmission electron microscopy. The R. sanguineus (tropical lineage) cells infected with E. canis successfully infected new tick cell cultures, showing that these cells could be an alternative substrate for maintenance of this pathogen. State Univ Julio de Mesquita Filho UNESP, Fac Agr & Vet Sci, Dept Vet Pathol, BR-14884900 Jaboticabal, Brazil Butantan Inst, Lab Parasitol, Sao Paulo, Brazil Fed Univ Latin Amer Integrat UNILA, Foz Do Iguacu, Brazil Univ Sao Paulo, Sch Vet Med, Dept Prevent Vet Med & Anim Sci, Sao Paulo, Brazil Univ Santo Amaro, Vet Med, Sao Paulo, Brazil Univ Fed Uberlandia, Uberlandia, MG, Brazil State Univ Julio de Mesquita Filho UNESP, Fac Agr & Vet Sci, Dept Vet Pathol, BR-14884900 Jaboticabal, Brazil FAPESP: 2007/57749-2 FAPESP: 2015/26209-9 CNPq: 312331/2013-4

Made available in DSpace on 2019-10-06T16:00:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-07-01. Added 1 bitstream(s) on 2019-10-09T18:35:04Z : No. of bitstreams: 1 S1984-29612018000300363.pdf: 2770381 bytes, checksum: c15ec391d851104f6d38492a03ad8099 (MD5) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Instituto Brasileiro do Meio Ambiente e dos Recursos Naturais Renováveis The aim of this study was to identify Plasmodium spp. in blood samples from nonhuman primates (NHPs) in the state of Maranhão, using classical and alternative techniques for examination of human malaria. A total of 161 blood samples from NHPs were analyzed: 141 from captive animals at a Wildlife Screening Center (CETAS) and 20 from free-living animals in a private reserve. The techniques used were microscopy, rapid diagnostic test (RDT), Indirect fluorescent antibody test (IFAT) and molecular techniques (semi-nested PCR, quantitative real-time PCR and LAMP). Two serological methods (dot-ELISA and indirect ELISA) were also standardized with rhoptry protein-soluble antigen of P. falciparum and P. berghei. Trophozoite forms of Plasmodium sp. were identified on slides from five different animals. No samples were positive through RDT and LAMP. Four samples were seropositive for P. malariae through IFAT. The samples showed low reactivity to ELISA. Plasmodium sp. was detected in 34.16% (55/161) of the samples using qPCR based on the 18S rRNA gene. After sequencing, two samples showed 100% identityl to P. malariae, one showed 97% identity to Plasmodium sp. ZOOBH and one showed 99% identity to P. falciparum. PCR was shown to be the most sensitive technique for diagnosing Plasmodium in NHP samples. Laboratório de Parasitologia Animal Curso de Medicina Veterinária Universidade Federal de Rondônia – UNIR Centro de Estudos da Malária Superintendência de Controle de Endemias – SUCEN Departamento de Saúde do Estado de São Paulo Instituto de Medicina Tropical de São Paulo – IMTSP Universidade de São Paulo – USP Laboratório de Patologia Veterinária Curso de Medicina Veterinária Universidade Federal de Rondônia – UNIR Laboratório de Imunoparasitologia Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista – UNESP Laboratório de Imunoparasitologia Faculdade de Ciências Agrárias e Veterinárias – FCAV Universidade Estadual Paulista – UNESP FAPESP: 10/12820-4 FAPESP: 12/03961-9

INCOGARR is a thematic network recently approved to be financially supported by the Ibero-American Program of Science and Technology for Development (CYTED). The objectives of this Network are the design and evaluation of an efficient and feasible anti-tick vaccine candidate from the technical and economical points of view and also sharing experiences in the immunological control of ticks as part of an Integrated Control Program. The Network consists of seven laboratories and one company from six countries. The first meeting of the Network took place with the representation of each laboratory involved. In the meeting, general and specific objectives and activities of the Network were discussed and it was a very nice example of international collaboration to address an unsolved worldwide topic on tick control in which laboratories with different competencies and expertise join their efforts in a common goal.

Made available in DSpace on 2019-10-06T17:03:52Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-04-01 Fundação de Amparo à Pesquisa do Estado de Goiás Although species of Anaplasma are highly prevalent Rickettsiales agents in domestic and wild ruminants with a wide distribution worldwide, few studies have been conducted so far to detect and/or investigate the diversity of these agentsin cattle in Mozambique. In the present study, serological and molecular assays were used to investigate the occurrence of Anaplasma spp. in 219 bovines sampled in the districts of Boane, Magude, Matutuíne, Moamba and Namaacha in Maputo, Mozambique. In the iELISA test for detection ofIgG antibodies to A. marginale, 86.3% (189/219) of the samples were positive. In qPCR assays for the gene msp1β for A. marginale and msp2 for A. phagocytophilum, 97.3% (213/219) and 2.7% (6/219) of the animals were positive, respectively. Two different cPCR protocols based on the 16S rRNA gene showed that 100% of the samples were positive for Anaplasma spp. The DNA sequences obtained were phylogenetically related to A. platys, A. phagocytophilum, Candidatus Anaplasma boleense, A. centrale, A. marginale and A. ovis. Phylogenetic inference based on the msp4 and msp5 genes positioned the obtained sequences in the clade of A. marginale, with evidence of occurrence of 8 and 5 different haplotypes for each gene, respectively. Anaplasma sp. phylogenetically associated with A. platys was evidenced in phylogenetic analyzes based on 16S rRNA and groEL genes. It is concluded that a high diversity of species of Anaplasma spp. occurs in cattle in Mozambique. Paulista State University (Unesp)Universidade Estadual Paulista (Unesp) Faculty of Agrarian and Veterinary Sciences Directorate of Animal Sciences Imunodot Diagnósticos Ltda Paulista State University (Unesp)Universidade Estadual Paulista (Unesp) Faculty of Agrarian and Veterinary Sciences Fundação de Amparo à Pesquisa do Estado de Goiás: 2015/14896-1

Made available in DSpace on 2019-10-06T16:13:25Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-03-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Heartwater caused by Ehrlichia ruminantiumis a disease of domestic and wild ruminants and one of the most economically important tick-borne diseases in Africa. The present study aimed to investigate the occurrence and genetic diversity of E. ruminantium in blood samples from 210 cattle sampled in five districts of Maputo Province, Mozambique. DNA blood samples were initially submitted to PCR assays targeting E. ruminantium pCS20 gene fragments. Additionally, in order to assess the genetic diversity of E. ruminantium, the positive samples were submitted to a PCR assay targeting the E. ruminantium map1 gene. Finally, the amplicons were sequenced and phylogenetic position was inferred using the Maximum Likelihood method. PCR results revealed that the overall prevalence in Maputo Province was 15% of the animals sampled. E. ruminantium map1 sequences showed not to be conserved. In the phylogenetic analysis, E. ruminantium map1 genotypes were positioned into multiple-clades. This study provides information on the prevalence and genetic diversity of E. ruminantium in five localities of Maputo Province. The future immune control strategies against local E. ruminantium must be designed in the light of the genetic diversity of this parasite. Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias (FCAV) Universidade Estadual Paulista (Unesp) Direcção de Ciências Animais Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias (FCAV) Universidade Estadual Paulista (Unesp) FAPESP: 2012 ̸ 213714

Made available in DSpace on 2021-07-14T10:54:04Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-02-19. Added 1 bitstream(s) on 2021-07-14T11:40:50Z : No. of bitstreams: 1 S1984-29612018000100098.pdf: 1147708 bytes, checksum: 5a8974bcf974304a53e81525c4e708ae (MD5) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Recentemente, a importância de roedores selvagens na manutenção de agentes Anaplasmataceae no ambiente tem sido investigada, haja visto o papel que tais mamíferos podem desempenhar como reservatórios para os patógenos e como hospedeiros para estágios imaturos dos carrapatos vetores. O presente estudo objetivou investigar a ocorrência de Ehrlichia sp. e Anaplasma sp. em 24 cotias (Dasyprocta azarae) capturadas na Reserva Ambiental de Itapiracó, em São Luís, Maranhão, nordeste do Brazil, utilizando métodos moleculares. Quatro animais (16,7%) mostraram-se positivos nos ensaios de nested PCR para Ehrlichia spp. baseados no gene 16S rRNA gene. Na análise filogenética baseda no gene 16S rRNA e utilizando o método de Máxima Verossimilhança e modelo evolutivo GTRGAMMA+I, os genótipos de Ehrlichia sp. detectados em cotias mostraram-se filogeneticamente relacionados às sequências de E. canis e outros genótipos relacionados a E. canis detectados previamente em animais selvagens no Brasil. O presente trabalho mostrou a primeira detecção molecular de Ehrlichia sp. em cotias no Brasil. Recently, the importance of wild-living rodents for maintenance of pathogens of the family Anaplasmataceae in the environment was investigated. These mammals play a role as reservoirs for these pathogens and act as hosts for the immature stages of tick vectors. The aim of the present study was to investigate the prevalence of Ehrlichia sp. and Anaplasma sp. in 24 specimens of Azara’s agouti (Dasyprocta azarae) that had been trapped in the Itapiracó Environmental Reserve, in São Luís, Maranhão, northeastern Brazil, using molecular methods. Four animals (16.7%) were positive for Ehrlichia spp. in nested PCR assays based on the 16S rRNA gene. In a phylogenetic analysis based on the 16S rRNA gene, using the maximum likelihood method and the GTRGAMMA+I evolutionary model, Ehrlichia sp. genotypes detected in Azara’s agoutis were found to be closely related to E. canis and to genotypes relating to E. canis that had previously been detected in free-living animals in Brazil. The present work showed the first molecular detection of Ehrlichia sp. in Azara’s agoutis in Brazil. Universidade Estadual do Maranhão Universidade Estadual Paulista, Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista, Faculdade de Ciências Agrárias e Veterinárias CNPq: #2015/14896-1; #473575/2014-0

Made available in DSpace on 2018-12-11T17:37:18Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-06-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) The family Streblidae comprises a monophyletic group of Hippoboscoidea, hematophagous dipterans that parasitize bats. Bartonella spp. and Rickettsia spp. have been reported in bats sampled in Europe, Africa, Asia, North, Central and South America. However, there are few reports on the Bartonella and Rickettsia bacteria infecting Hippoboscoidea flies and mites. While Spinturnicidae mites are ectoparasites found only in bats, those belonging to the family Macronyssidae comprise mites that also parasitize other mammal species. This study investigates the occurrence and assesses the phylogenetic positioning of Bartonella spp. and Rickettsia spp. found in Streblidae flies and Spinturnicidae and Macronyssidae mites collected from bats captured in Brazil. From May 2011 to April 2012 and September 2013 to December 2014, 400 Streblidae flies, 100 Macronyssidaes, and 100 Spinturnicidae mites were collected from bats captured in two sites in northeastern Nova Iguaçu, Rio de Janeiro, southeastern Brazil. Forty (19.8%) out of 202 Streblidae flies were positive for Bartonella spp. in qPCR assays based on the nuoG gene. Among the flies positive for the bacterium, six (18%) were Paratrichobius longicrus, seven (29%) Strebla guajiro, two (40%) Aspidoptera phyllostomatis, five (11%) Aspidoptera falcata, one (10%) Trichobius anducei, one (25%) Megistopoda aranea, and 18 (32%) Trichobius joblingi, and collected from bats of the following species: Artibeus lituratus, Carollia perspicillata, Artibeus planirostris, Sturnira lilium, and Artibeus obscurus. Six sequences were obtained for Bartonella (nuoG [n = 2], gltA [n = 2], rpoB [n = 1], ribC = 1]). The phylogenetic analysis based on gltA (750pb) gene showed that the Bartonella sequences clustered with Bartonella genotypes detected in bats and ectoparasites previously sampled in Latin America, including Brazil. Only one sample (0.49%) of the species Trichobius joblingi collected from a specimen of Carollia perspicillata was positive for Rickettsia sp. in cPCR based on the gltA gene (401bp). This sequence was clustered with a ‘Candidatus Rickettsia andaenae genotype detected in an Amblyomma parvum tick collected from a rodent in the southern region of Brazilian Pantanal. The sampled Macronyssidae and Spinturnicidae mites were negative for Bartonella spp. and Rickettsia spp. This study demonstrated the first occurrence of Bartonella spp. and Rickettsia spp. DNA in Streblidae flies collected from bats in Brazil. Laboratory of Immunoparasitology Department of Veterinary Pathology School of Agricultural and Veterinarian Sciences Universidade Estadual Paulista (UNESP) Agricultural and Livestock Microbiology Graduation Program School of Agricultural and Veterinarian Sciences Universidade Estadual Paulista (UNESP) Laboratory of Parasite Arthropods Department of Animal Parasitology Institute of Veterinary Universidade Federal Rural do Rio de Janeiro–UFRRJ Laboratory of Immunoparasitology Department of Veterinary Pathology School of Agricultural and Veterinarian Sciences Universidade Estadual Paulista (UNESP) Agricultural and Livestock Microbiology Graduation Program School of Agricultural and Veterinarian Sciences Universidade Estadual Paulista (UNESP) CNPq: 302420/ 2017-7

Made available in DSpace on 2018-12-11T17:34:35Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-01-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) The genus Rickettsia comprises obligatory intracellular bacteria, well known to cause zoonotic diseases around the world. The present work aimed to investigate the occurrence of Rickettsia spp. in wild animals, domestic dogs and their respective ectoparasites in southern Pantanal region, central-western Brazil, by molecular and serological techniques. Between August 2013 and March 2015, serum, whole blood and/or spleen samples were collected from 31 coatis, 78 crab-eating foxes, seven ocelots, 42 dogs, 110 wild rodents, and 30 marsupials. Serum samples from canids, felids, rodents and marsupials were individually tested by indirect fluorescent antibody test (IFAT) in order to detect IgG antibodies to Rickettsia rickettsii, Rickettsia parkeri and Rickettsia amblyommatis. DNA samples from mammals and ectoparasites were submitted to a multiplex qPCR assay in order to detect and quantify spotted fever group (SFG) and typhus group (TG) rickettsiae and Orientia tsutsugamushi. Positive samples in qPCR assays were submitted to conventional PCR assays targeting gltA, ompA, ompB and htrA genes, followed by sequencing and phylogenetic analyses. The ticks collected (1582) from animals belonged to the species Amblyomma sculptum, Amblyomma parvum, Amblyomma ovale, Amblyomma tigrinum, Rhipicephalus (Boophilus) microplus, Rhipicephalus sanguineus sensu lato and Amblyomma auricularium. Overall, 27 (64.2%) dogs, 59 (75.6%) crab-eating foxes and six (85.7%) ocelots were seroreactive (titer ≥ 64) to at least one Rickettsia species. For 17 (40.4%) dogs, 33 (42.3%) crab-eating foxes, and two (33.3%) ocelots, homologous reactions to R. amblyommatis or a closely related organism were suggested. One hundred and sixteen (23.5%) tick samples and one (1.2%) crab-eating fox blood sample showed positivity in qPCR assays for SFG Rickettsia spp. Among SFG Rickettsia-positive ticks samples, 93 (80.2%) belonged to A. parvum, 14 (12%) belonged to A. sculptum species, three (2.5%) belonged to A. auricularim, and six (5.2%) were Amblyomma larval pools. Thirty samples out of 117 qPCR positive samples for SFG Rickettsia spp. also showed positivity in cPCR assays based on gltA, htrA and/or ompB genes. The Blast analyses showed 100% identity with ‘Candidatus Rickettsia andeanae’ in all 30 sequences obtained from gltA, htrA and/or ompB genes. The concatenated phylogenetic analysis based on gltA and 17-kDa htrA genes grouped the Rickettsia sequences obtained from tick samples in the same clade of ‘Candidatus Rickettsia andeanae’. The present study revealed that wild and domestic animals in southern Pantanal region, Brazil, are exposed to SFG rickettsiae agents. Future studies regarding the pathogenicity of these agents are necessary in order to prevent human cases of rickettsiosis in Brazilian southern Pantanal. Universidade Estadual Paulista (Unesp) Faculdade de Ciências Agrárias e Veterinárias, Jaboticabal Universidade Católica Dom Bosco Universidade Federal da Paraíba Laboratório de Ecologia Animal, Rio Tinto Universidade de São Paulo Faculdade de Medicina Veterinária e Zootecnia Universidade Estadual Paulista (Unesp) Faculdade de Ciências Agrárias e Veterinárias, Jaboticabal

The African buffalo (Syncerus caffer), a mammal species whose population is declining, can play a role as a reservoir or carrier of a wide number of arthropod-borne pathogens. Translocation procedures have been used as an alternative approach for species conservation. However, the veterinary aspects of this sort of procedures are extremely important to minimize the impact on animal health. In order to detectBartonellaand haemoplasmas, two important group of bacterial that have an impact in both human and animal health, EDTA whole-blood samples were screened for the presence of these bacterial pathogens by molecular techniques. As a result, a molecular occurrence of 4.1 and 15.4% forBartonellaspp. and haemoplasmas, respectively, was reported among 97 wild buffaloes sampled during a translocation procedure from Marromeu to Gorongosa Reserve, Mozambique. Additionally, phylogenetic analyses of the obtained sequences were conducted. At least, three bovine-associated pathogens, namelyB. bovis,M. wenyoniiand ‘CandidatusM. haemobos’, as well as a probably newBartonellagenotype/species were detected inS. caffer.Further studies are needed in order to determine whether these bacterial species may cause impact in buffaloes and other sympatric ruminant species living in the release site.

Water buffalo (Bubalus bubalis) is a potential reservoir for Anaplasma marginale in livestock ecosystems of tropical countries. However, their participation in the epidemiological process of bovine anaplasmosis in endemic areas remains unclear. In the present study, the reservoir competence of water buffalo for A. marginale was explored by focusing on the analysis of rickettsemia levels in carrier animals, and the genetic characterization of A. marginale strains from cattle and buffalo. Eight groups of cattle and water buffaloes were randomly selected from cohabiting herds in four livestock ecosystems of Cuba, together with two control groups from unrelated cattle and buffalo herds. A total of 180 adult animals (88 water buffalo and 92 cattle) were sampled. Rickettsemia in carrier animals was determined by quantitative real-time PCR. The rickettsemia (parasitemia) levels in cattle were higher than in buffaloes, however the rickettsemia in buffalo may be enough to infect R. microplus ticks. The genetic diversity of A. marginale was assessed by strain characterization and phylogenetic analysis of 27 msp1α gene sequences. The results showed genetic similarity among strains from cattle and water buffalo, suggesting the occurrence of cross-species transmission., This work was supported by funding from Fundacão de Amparo à Pesquisa do Estado de São Paulo (FAPESP), Brazil (Process 2012/21371-4); and the National Priority Program for Animal and Plant Health, Ministry of Agriculture (MINAGRI), Cuba (Process P131LH003007). Dasiel Obregon would like to acknowledge the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and the Three World Academy of Science (TWAS) by his sandwich Ph.D. grant (TWAS-CNPq 2013, Process 302999/2013-2).

Made available in DSpace on 2019-10-06T16:03:16Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-01-01 The aim of this study was to perform a molecular survey and determine the genetic diversity of Bartonella spp. (Rhizobiales: Bartonellaceae) and Rickettsia spp. (Rickettsiales: Rickettsiaceae) in cat feas (Siphonaptera: Pulicidae) from Southern Chile. Fleas (n = 251) were collected from 150 cats in Valdivia city and identified using morphological keys. Fleas belonging to the same cat were pooled (two to seven feas per pool). DNA was purified from individual (n = 92) and pooled (n = 58) feas and submitted to conventional polymerase chain reaction assays targeting Bartonella spp. (gltA) and Rickettsia spp. (ompA). Selected positive samples were sequenced for Bartonella gltA (n = 19), Rickettsia ompA (n = 14), and Rickettsia gltA (n = 11) for speciation, phylogenetic, and diversity analyses. All feas (n = 251) were identified as Ctenocephalides felis felis (Bouché) (Siphonaptera: Pulicidae). Bartonella and Rickettsia occurrences in C. felis felis were 39.3% (59/150) and 76.6% (115/150), respectively. From sequenced Bartonella spp., 47.3% (9/19) were identified as Bartonella clarridgeiae, 42.1% (8/19) as Bartonella henselae, 5.3% (1/19) as Bartonella koehlerae, and 5.3% (1/19) as Bartonella sp. Rickettsia felis was the only Rickettsiaceae species identified in both ompA (14/14) and gltA (11/11) products. B. henselae and B. clarridgeiae presented five genotypes. R. felis ompA sequences presented seven genotypes. On the other hand, R. felis gltA sequences showed only one genotype. Bartonella spp. and R. felis are described for the first time in C. felis felis feas from Southern Chile, highlighting the importance of these vectors as a source of zoonotic agents. Instituto de Ciencias Clínicas Veterinarias Facultad de Ciencias Veterinarias Universidad Austral de Chile Escuela de Medicina Veterinaria Facultad de Ciencias Universidad Mayor Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (FCAV/UNESP) Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (FCAV/UNESP)

An available enzyme-linked immunosorbent assay (ELISA) was studied for the detection of anti-Dioctophyma renale antibodies in the sera of dogs using, detection of parasite eggs in urine sediment as a reference test. ELISA uses a soluble antigenic preparation of esophagus of D. renale and the optimal dilutions of the antigen, serum and conjugate were determined by means of checker board titration, using positive (n=13) and negative (n=27) reference serum. The specificity and sensitivity of the ELISA were 93.8% and 92.3% respectively and the kappa index was good (0.76). These results suggest that ELISA described may prove to be an effective serological test for detecting dogs infected and exposed to this parasite mainly dogs that are not eliminating parasite eggs through their urine.

Comparative studies between brown dog tick Rhipicephalus sanguineus populations from Brazil (Jaboticabal, São Paulo) and Argentina (Rafaela, Santa Fé) showed significant biological, morphological and genetic differences between them. This work aimed to study, in a comparative way, the acquisition of resistance in domestic dogs to R. sanguineus from Jaboticabal and Rafaela, after successive and controlled infestations. Ticks were kept in a BOD incubator under controlled conditions (27 °C, 80 % relative humidity, 12-h photoperiod). Ten dogs, Dachshund breed, males and females, 6 months old, short- or long-haired, without prior contact with ticks, were used as hosts. They were distributed into two experimental groups composed of five animals each: G1 infested with ten adult couples of R. sanguineus (Jaboticabal) per animal, and G2 infested with ten adult couples of R. sanguineus (Rafaela) per animal. Ticks' biological parameters and titration of antibodies from the dogs' sera by ELISA test were used for comparison between the strains. Results of the biological parameters showed that the dogs did not acquire immunity to either of the R. sanguineus strains after repeated infestations. The ELISA test showed low antibody titers in sera of dogs from G2, in successive infestations, and higher antibody responses post second and third infestations in G1. It also demonstrated cross-reactivity between sera of dogs infested with R. sanguineus (Jaboticabal) and antigens from R. sanguineus (Rafaela) and vice versa. We conclude that Dachshund dogs did not develop resistance against neither Jaboticabal nor Rafaela strains of R. sanguineus.

Made available in DSpace on 2015-11-03T15:29:03Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-10-01. Added 1 bitstream(s) on 2015-11-04T10:14:47Z : No. of bitstreams: 1 S1984-29612014000400443.pdf: 604594 bytes, checksum: f2dd2f1f3e36bfd53ab7d2708b790958 (MD5) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Pombos silvestres (Zenaida auriculata), comuns em áreas urbanas, rurais e selvagens em muitas regiões do Brasil, são frequentemente predados por gatos domésticos. Sendo assim, os isolados de T. gondii obtidos de pombos podem refletir uma maior diversidade ambiental do que os outros hospedeiros. O objetivo do presente estudo foi avaliar a soroprevalência, isolar e genotipar T. gondii de Z. auriculata. Amostras de soro e tecido foram coletadas de 206 pombos para o teste de aglutinação modificado (MAT) e o bioensaio em camundongos. A prevalência de anticorpos contra T. gondii em pombos foi 22,3% (46/206), com títulos variando de 16 a 4096, e T. gondii foi isolado de 12 pombos. Cinco genótipos foram detectados por PCR-RFLP, incluindo os genótipos ToxoDB #1, #6, #17, #65 e um genótipo não descrito anteriormente (ToxoDB#182). Esse é o primeiro relato de isolamento de T. gondii de Z. auriculata. Este estudo também confirmou a diversidade dos isolados de T. gondii e a presença de tipo clonal II (ToxoDB #1) no Brasil. Eared doves (Zenaida auriculata), which are common in urban, rural and wild areas in many regions of Brazil, are frequently prey for domestic cats. Therefore Toxoplasma gondii isolates obtained from doves may reflect greater environmental diversity than those from other hosts. The aim of the present study was to evaluate T. gondii seroprevalence, isolate and genotype strains from Z. auriculata. Serum and tissue samples were collected from 206 doves for use in the modified agglutination test (MAT) and mouse bioassay. The prevalence of T. gondii antibodies in the doves was 22.3% (46/206), with titers ranging from 16 to 4096, and T. gondii strains were isolated from 12 of these doves. Five genotypes were detected by means of PCR-RFLP, including ToxoDB genotypes #1, #6, #17 and #65, and one genotype that had not previously been described (ToxoDB#182). This was the first report on isolation of T. gondii from Z. auriculata. This study confirmed the genetic diversity of T. gondii isolates and the existence of clonal type II (ToxoDB genotype #1) in Brazil. Univ Estadual Londrina, Dept Vet Prevent Med, Lab Protozool, BR-86051990 Londrina, PR, Brazil Univ Tennessee, Dept Microbiol, Knoxville, TN 37996 USA Univ Estadual Paulista, Sch Agr &Vet Sci, Dept Vet Pathol, UNESP, Jaboticabal, SP, Brazil Univ Estadual Paulista, Sch Agr &Vet Sci, Dept Vet Pathol, UNESP, Jaboticabal, SP, Brazil CAPES: 1345/2011 CAPES: 10259/12-0 FAPESP: 2010/01597-2

Made available in DSpace on 2018-12-11T17:20:39Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-08-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) The present work aimed to investigate the genetic diversity of Bartonella in mammals and ectoparasites in Pantanal wetland, Brazil. For this purpose, 31 Nasua nasua, 78 Cerdocyon thous, 7 Leopardus pardalis, 110 wild rodents, 30 marsupials, and 42 dogs were sampled. DNA samples were submitted to a quantitative real-time PCR assay (qPCR). Positive samples in qPCR were submitted to conventional PCR assays targeting other five protein-coding genes. Thirty-five wild rodents and three Polygenis (P.) bohlsi bohlsi flea pools showed positive results in qPCR for Bartonella spp. Thirty-seven out of 38 positive samples in qPCR were also positive in cPCR assays based on ftsZ gene, nine in nuoG-cPCR, and six in gltA-cPCR. Concatenated phylogenetic analyses showed that two main genotypes circulate in rodents and ectoparasites in the studied region. While one of them was closely related to Bartonella spp. previously detected in Cricetidae rodents from North America and Brazil, the other one was related to Bartonella alsatica, Bartonella pachyuromydis, Bartonella birtlesii, Bartonella acomydis, Bartonella silvatica, and Bartonella callosciuri. These results showed that at least two Bartonella genotypes circulate among wild rodents. Additionally, the present study suggests that Polygenis (P.) bohlsi bohlsi fleas could act as possible Bartonella vectors among rodents in Pantanal wetland, Brazil. Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (Unesp) Universidade Católica Dom Bosco Laboratório de Ecologia Animal Universidade Federal da Paraíba Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias Júlio de Mesquita Filho (UNESP) Campus de Jaboticabal, Via de Acesso Prof. Paulo Donato Castellane, s/n, Zona Rural, CEP: 14884-900, Jaboticabal Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (Unesp) Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Faculdade de Ciências Agrárias e Veterinárias Júlio de Mesquita Filho (UNESP) Campus de Jaboticabal, Via de Acesso Prof. Paulo Donato Castellane, s/n, Zona Rural, CEP: 14884-900, Jaboticabal FAPESP: 2015/14896-1 CNPq: 473575/2014-0

Made available in DSpace on 2018-12-11T17:35:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-03-01 Equine piroplasmosis is a disease caused by the hemoparasites Babesia caballi and Theileria equi and is considered to be the most important parasitic infection affecting Equidae. The objective of the present study was to carry out an epidemiological molecular and serological survey for the presence of these two protozoal organisms in equids from the northwestern region of the State of Rio Grande do Sul (RS), south Brazil. For this purpose, blood samples were collected from 90 equids in the city of Passo Fundo, RS, Brazil. Those were animals used for sport activities, outdoor recreational riding, and work including cattle herding and mounted patrol. Anti-T. equi and anti-B. caballi IgG antibodies were detected in the sera of those animals by commercial ELISA kits. The molecular diagnosis of equine piroplasmosis due to T. equi or B. caballi (or both) consisted in the amplification of the 18S rRNA gene by nested PCR followed by sequencing of the amplified PCR product and sequence comparison and phylogenetic analysis of the isolates; 17 (18.9%) and 5 (5.55%) out of the 90 serum samples tested in this study were positive for T. equi and B. caballi, respectively. Piroplasmid 18S rRNA gene fragments were detected by PCR in 24.4% (22/90) of the samples analysed and shared 99–100% identity with sequences of T. equi by BLASTn. Samples for the phylogenetic analysis were divided into 2 groups. In group A, there was close phylogenetic relationship between 4 sequences and sequences previously reported along the US-Mexico border, in South Africa, and in Brazil. There was a phylogenetic proximity between 5 samples from group B and samples tested by other authors in the US and Spain. Variation of the 18S rRNA gene allowed the identification of 9 new T. equi genotypes in the geographical region studied. Professor do Programa de Pós-Graduação em Bioexperimentação UPF Professor do curso de Medicina Veterinária UPF Estudante de Mestrado do Programa de Pós-Graduação em Bioexperimentação da Universidade de Passo Fundo (UPF) Estudante de Doutorado do Programa de Pós-Graduação em Microbiologia Agropecuária UNESP Professor no Programa de Pós-Graduação em Medicina Veterinária e Microbiologia Agropecuária UNESP Estudante de Doutorado do Programa de Pós-Graduação em Microbiologia Agropecuária UNESP Professor no Programa de Pós-Graduação em Medicina Veterinária e Microbiologia Agropecuária UNESP

Made available in DSpace on 2018-12-11T17:36:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-01-01 Trypanosoma vivax is a parasite widespread across Africa and South America. Immunological methods using recombinant antigens have been developed aiming at specific and sensitive detection of infections caused by T. vivax. Here, we sequenced for the first time the transcriptome of a virulent T. vivax strain (Lins), isolated from an outbreak of severe disease in South America (Brazil) and performed a computational integrated analysis of genome, transcriptome and in silico predictions to identify and characterize putative linear B-cell epitopes from African and South American T. vivax. A total of 2278, 3936 and 4062 linear B-cell epitopes were respectively characterized for the transcriptomes of T. vivax LIEM-176 (Venezuela), T. vivax IL1392 (Nigeria) and T. vivax Lins (Brazil) and 4684 for the genome of T. vivax Y486 (Nigeria). The results presented are a valuable theoretical source that may pave the way for highly sensitive and specific diagnostic tools. Laboratório Nacional de Computação Científica (LNCC), Av. Getúlio Vargas, 333 Grupo Hermes Pardini Setor de Pesquisa e Desenvolvimento Departamento de Parasitologia Instituto de Ciências Biomédicas Universidade de São Paulo Trypanosomatids Infectious Processes Laboratory Department of Infection and Epidemiology Institut Pasteur, 25 rue du Dr Roux Departamento Clínica Cirurgia e Reprodução Animal Faculdade de Odontologia e Curso de Medicina Veterinária Universidade Estadual Paulista – UNESP Laboratório de Immnoparasitologia Faculdade de Ciências Agrárias e Veterinárias (FCAV) Universidade Estadual Paulista (UNESP), Campus Jaboticabal Departamento Clínica Cirurgia e Reprodução Animal Faculdade de Odontologia e Curso de Medicina Veterinária Universidade Estadual Paulista – UNESP Laboratório de Immnoparasitologia Faculdade de Ciências Agrárias e Veterinárias (FCAV) Universidade Estadual Paulista (UNESP), Campus Jaboticabal

Made available in DSpace on 2018-12-11T17:34:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-07-01. Added 1 bitstream(s) on 2021-07-15T15:22:25Z : No. of bitstreams: 1 S1984-29612017000300366.pdf: 1029343 bytes, checksum: a7ed0de574ac9c7d372f6d45dca287a1 (MD5) Trypanosomiasis caused by Trypanosoma vivax has increased the reports in Brazil in the last decade. An outbreak is herein first reported in the state of Goiás, from May 2016 to January 2017. The outbreak start occurred in the city of Ipameri (Goiás) after the introduction of 18 auctioned cows from the state of Minas Gerais. Direct parasitological test (blood smears) and polymerase chain reactions targeting the catL genes diagnosed T. vivax infection. Fifty six cows from a herd of 161 were infected; 12 died during the outbreak and 44 animals persistently positive (by blood smears) even after chemical treatment were discarded. After this first case, five other cases were detected in state of Goiás. The spread of this disease can be linked to the commercialization of animals carrying T. vivax, allied to the iatrogenic transmission practice, using a single needle and syringe for all cows, during oxytocin administration before each milking. Departamento de Parasitologia Veterinária Escola de Veterinária e Zootecnia – EVZ Universidade Federal de Goiás – UFG Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP Instituto de Patologia Tropical e Saúde Pública – IPTSP Universidade Federal de Goiás – UFG Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista “Júlio de Mesquita Filho” – UNESP

Wild animals play an important role in carrying vectors that may potentially transmit pathogens. Several reports highlighted the participation of wild animals on the Anaplasma phagocytophilum cycle, including as hosts of the agent. The aim of this study was to report the molecular detection of an agent phylogenetically related to A. phagocytophilum isolated from a wild bird in the Midwest of the state of Paraná, Brazil. Fifteen blood samples were collected from eleven different bird species in the Guarapuava region. One sample collected from a Penelope obscura bird was positive in nested PCR targeting the 16S rRNA gene of Anaplasma spp. The phylogenetic tree based on the Maximum Likelihood analysis showed that the sequence obtained was placed in the same clade with A. phagocytophilum isolated from domestic cats in Brazil. The present study reports the first molecular detection of a phylogenetically related A. phagocytophilum bacterium in a bird from Paraná State. Resumo Animais selvagens possuem participação importante como carreadores dos vetores responsáveis por transmitir doenças e vários relatos destacam a participação de animais silvestres no ciclo do Anaplasma phagocytophilum, inclusive como hospedeiros do agente. O presente trabalho tem por objetivo relatar pela primeira vez a detecção molecular da infecção por um agente filogeneticamente associado a A. phagocytophilum em uma ave silvestre no interior do Paraná, Brasil. Foram colhidas 15 amostras de sangue originadas de onze espécies diferentes de aves, todas provenientes da região de Guarapuava. Apenas uma amostra pertencente a uma ave da espécie Penelope obscura foi positiva para o ensaio de nested PCR baseado no gene 16S rRNA. A árvore filogenética baseada na análise por máxima verossimilhança demonstrou que a sequência obtida no presente estudo se posicionou no mesmo clado com cepas de A. phagocytophilum isoladas de gatos domésticos no Brasil. O presente trabalho relata pela primeira vez a detecção molecular de Anaplasma sp. filogeneticamente relacionado à A. phagocytophilum, em um animal da espécie P. obscura, assim como a presença do parasita em uma ave silvestre do Estado do Paraná, Brasil.

Made available in DSpace on 2018-12-11T16:47:19Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-07-01 The order Chiroptera is considered the second largest group of mammals in the world, hosting important zoonotic virus and bacteria. Bartonella and hemotropic mycoplasmas are bacteria that parasite different mammals' species, including humans, causing different clinical manifestations. The present work aimed investigating the occurrence and assessing the phylogenetic positioning of Bartonella spp. and Mycoplasma spp. in neotropical bats sampled from Brazil. Between December 2015 and April 2016, 325 blood and/or tissues samples were collected from 162 bats comprising 19 different species sampled in five states of Brazil. Out of 322 bat samples collected, while 17 (5·28%) were positive to quantitative PCR for Bartonella spp. based on nuoG gene, 45 samples (13·97%) were positive to cPCR assays for hemoplasmas based on 16S rRNA gene. While seven sequences were obtained for Bartonella (nuoG) (n = 3), gltA (n = 2), rpoB (n = 1), ftsZ (n = 1), five 16S rRNA sequences were obtained for hemoplasmas. In the phylogenetic analysis, the Bartonella sequences clustered with Bartonella genotypes detected in bats sampled in Latin America countries. All five hemoplasmas sequences clustered together as a monophyletic group by Maximum Likelihood and Bayesian Inference analyses. The present work showed the first evidence of circulation of Bartonella spp. and hemoplasmas among bats in Brazil. Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (Unesp), Via de Acesso Prof. Paulo Donato Castellane, s/n Universidade Estadual Do Centro-Oeste Campus CEDETEG Faculdade de Medicina Veterinária e Zootecnia Universidade Estadual Paulista (Unesp) Instituto de Ciências Biomédicas Universidade de São Paulo (USP) Faculdade de Ciências Agrárias e Veterinárias Universidade Estadual Paulista (Unesp), Via de Acesso Prof. Paulo Donato Castellane, s/n Faculdade de Medicina Veterinária e Zootecnia Universidade Estadual Paulista (Unesp)

Made available in DSpace on 2018-12-11T17:11:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-04-26 Background: Considering the diversity of feeding habits that females of some species of anophelines present, it is important to understand which vertebrates are part of blood food sources and how important is the role of each in the ecoepidemiology of malaria. There are many vector species for Plasmodium spp. in the State of Maranhão, Brazil. In São Luís Island, Anopheles aquasalis is the main vector for human malaria; this species is abundant in areas with primates that are positive for Plasmodium. Anopheles aquasalis has natural exophilic and zoophilic feeding behavior, but in cases of high density and absence of animals, presents quite varied behavior, and feeds on human blood. In this context, the objective of the present study was to identify Plasmodium spp. and the blood meal sources of anophelines in two environmental reserves on São Luís Island, state of Maranhão, using molecular methods. Methods: Between June and July 2013, female anophelines were collected in the Sítio Aguahy Private Reserve, in the municipality of São José de Ribamar, and in the Sítio Mangalho Reserve, located within the Maracanã Environmental Protection Area, in the municipality of São Luís. CDC-type light traps, Shannon traps and protected human bait were used during three consecutive hours in peridomestic and wooded areas. Pools of anophelines were formed using mosquitoes of the same species that had been caught at the same site on the same date. A genus-specific amplification protocol based on the 18S rRNA gene was used for qPCR and cPCR. Results: A total of 416 anophelines were collected, of the following species: An. aquasalis (399), An. mediopunctatus (3), An. shannoni (1), An. nuneztovari (sensu lato) (1), An. goeldii (1), An. evansae (2) and An. (Nyssorhynchus) sp. (9), comprising 54 pools. Two pools were positive for Plasmodium (2/54) based on the 18S rRNA gene. In the phylogenetic analysis using the maximum likelihood method, based on a 240 bp fragment of the 18S rRNA gene, it was found that the sequences of Plasmodium sp. amplified from pools of An. aquasalis (pool 2) and An. nuneztovari (s.l.) (pool 10) were phylogenetically related to a clade of P. falciparum isolates from India, and to a clade of Plasmodium sp. isolates from psittacines in Brazil, respectively. Cat, dog and human DNA were identified in the blood meals of the anophelines sampled. Conclusion: The species An. aquasalis was the most abundant anopheline species in São Luís Island. Plasmodium spp. DNA was detected, thus confirming the importance of this species as the main vector on São Luís Island, Brazil. In addition, the presence of An. nuneztovari (s.l.) with DNA positive for Plasmodium spp. confirms its importance as a secondary vector. Immunoparasitology Laboratory School of Agrarian and Veterinary Sciences (FCAV) Universidade Estadual Paulista (UNESP) Jaboticabal Campus Center for Malaria Studies Superintendence of Control of Endemic Diseases State Secretariat of Health of São Paulo Institute of Tropical Medicine of São Paulo (IMT-SP) University of São Paulo (USP) Veterinary Pathology Laboratory Brazil University Descalvado Campus Immunoparasitology Laboratory School of Agrarian and Veterinary Sciences (FCAV) Universidade Estadual Paulista (UNESP) Jaboticabal Campus

Made available in DSpace on 2018-12-11T17:31:43Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-04-15 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Hepatozoon parasites comprise intracellular apicomplexan parasites transmitted to vertebrate animals by ingestion of arthropods definitive hosts. The present work aimed to investigate the occurrence of Hepatozoon spp. in wild animals, domestic dogs and their respective ectoparasites, in southern Pantanal region, central-western Brazil, by molecular techniques. Between August 2013 and March 2015, 31 coatis (Nasua nasua), 78 crab-eating foxes (Cerdocyon thous), seven ocelots (Leopardus pardalis), 42 dogs (Canis lupus familiaris), 110 wild rodents (77 Thichomys fosteri, 25 Oecomys mamorae, and 8 Clyomys laticeps), 30 marsupials (14 Thylamys macrurus, 11 Gracilinanus agilis, 4 Monodelphis domestica and 1 Didelphis albiventris), and 1582 ticks and 80 fleas collected from the sampled animals were investigated. DNA samples were submitted to PCR assays for Hepatozoon spp. targeting 18S rRNA gene. Purified amplicons were directly sequenced and submitted to phylogenetic analysis. A high prevalence of Hepatozoon among carnivores (C. thous [91.02%], dogs [45.23%], N. nasua [41.9%] and L. pardalis [71.4%]) was found. However, ticks and fleas were negative to Hepatozoon PCR assays. By phylogenetic analysis based on 18S rRNA sequences, Hepatozoon sequences amplified from crab-eating foxes, dogs, coatis and ocelots clustered with sequences of H. canis, H. americanum and H. felis. The closely related positioning of Hepatozoon sequences amplified from wild rodents and T. macrurus marsupial to Hepatozoon from reptiles and amphibians suggest a possible transmission of those Hepatozoon species between hosts by ectoparasites or by predation. Hepatozoon haplotypes found circulating in wild rodents seem to present a higher degree of polymorphism when compared to those found in other groups of animals. Although rodents seem not to participate as source of Hepatozoon infection to wild carnivores and domestic dogs, they may play an important role in the transmission of Hepatozoon to reptiles and amphibians in Pantanal biome. Faculdade de Ciências Agrárias e Veterinárias/Universidade Estadual Paulista UNESP Universidade Católica Dom Bosco Universidade Federal da Paraíba Laboratório de Ecologia Animal Universidade de São Paulo, Faculdade de Medicina Veterinária e Zootecnia Laboratório de Parasitologia Instituto Butantan São Paulo Universidade Federal de Mato Grosso do Sul Embrapa gado de corte Faculdade de Ciências Agrárias e Veterinárias/Universidade Estadual Paulista UNESP

Made available in DSpace on 2018-12-11T17:31:37Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-03-01 Bacteria in the genus Anaplasma are responsible for diseases in animals and humans. Studies carried out in Brazil have demonstrated that Brazilian deer are able to act as hosts of agents in the family Anaplasmataceae and are possibly potential reservoirs of these pathogens. Molecular and phylogenetic studies have been carried out on samples of two gray brocket specimens (Mazama gouazoubira) from the city of Guarapuava, state of Paraná, Brazil, for the detection of Anaplasma sp. in these animals. Partial nucleotide sequences of the genes 16S rRNA and groESL were used for phylogenetic analyses and compared with other 13 and 17 partial sequences of the respective genes obtained in GenBank. These assessments showed topological incongruence among the trees generated in the phylogenetic analyses. Phylogenetic analysis based on the gene 16S rRNA of the genotypes amplified in the samples of this study was similar to those of A. bovis detected in dogs and wild deer in Japan, whereas studies carried out on gene groESL indicated proximity with sequences of Anaplasma sp. that were also isolated in deer in Japan and allocated in the same clade of partial sequences of A. phagocytophilum. As the 16S rRNA gene is highly conserved, with few polymorphic positions, it may show low reliability for studies on phylogenetic positioning. The present study detected an Anaplasma sp. genotype in two specimens of M. gouazoubira in southern Brazil, which may mean that this agent possibly circulates in deer populations, and demonstrated the need for studies related to the possible role of deer in enzootic cycles of Anaplasmataceae in Brazil. Laboratório de Doenças Infecciosas e Parasitárias Departamento de Medicina Veterinária Universidade Estadual Do Centro-Oeste (UNICENTRO), Rua Simeao Varela de Sá, 03, Vila Carli Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Universidade Estadual Paulista Júlio de Mesquita Filho (UNESP) Laboratório de Imunoparasitologia Departamento de Patologia Veterinária Universidade Estadual Paulista Júlio de Mesquita Filho (UNESP)