Search

Espinilho savanna ("seasonal steppe savanna") is a unique vegetation formation of the Pampas biome that is found near the tri-border of Brazil, Uruguay, and Argentina. The Yellow Cardinal (Gubernatrix cristata) is a flagship species of this ecosystem, but it is classified as "critically endangered" in Brazil due to habitat loss and poaching for the illegal trade. Population supplementation through the release of individuals that were captive-bred or apprehended by authorities from the illegal trade has been considered as a conservation strategy for this species; however, the risk of pathogen introduction is a critical concern. We used microscopy and molecular methods to investigate the occurrence of blood parasites in wild passerines (n = 64, including three Yellow Cardinals) at Espinilho State Park, Rio Grande do Sul, Brazil, and in captive Yellow Cardinals (n = 30) at three facilities in Brazil. Haemosporidian parasites were detected in the blood smears of 10.9% of the wild passerines, comprising the morphospecies Haemoproteus erythrogravidus in Rufous-collared Sparrow (Zonotrichia capensis), H. quiscalus in Grayish Baywing (Agelaioides badius), and H. tyranni in Great Kiskadee (Pitangus sulphuratus); these are the southernmost records for these morphospecies and their first record for the Pampas biome. No haemosporidian parasites were detected in the blood smears of the Yellow Cardinals, wild or captive. Microfilariae were detected in the blood smears of 14.1% of the wild passerines, including all wild Yellow Cardinals, and in 43.3% of captive Yellow Cardinals. Trypanosoma sp. was detected in the blood smear of one captive Yellow Cardinal. Nested PCR and gene sequencing of the cyt-b gene of Haemoproteus/Plasmodium was used to test a subset of wild passerines and captive Yellow Cardinals, allowing for the molecular barcoding of H. quiscalus lineage AGEBAD04 and H. tyranni lineage PITSUL01; additionally, DNA identical to that of lineage PITSUL01 was detected in the blood of one captive Yellow Cardinal. This study provides valuable data to support the conservation management of the Yellow Cardinal and other threatened passerines from the Pampas and highlights the need for further studies on the epidemiology and pathology of filarioid worms and trypanosomes in passerines from this biome.

Made available in DSpace on 2022-05-01T09:47:17Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-12-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Universidade Federal do Paraná Fundação Araucária Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Accidents involving Brown spiders are reported throughout the world. In the venom, the major toxins involved in the deleterious effects are phospholipases D (PLDs). In this work, recombinant mutated phospholipases D from three endemic species medically relevant in South America (Loxosceles intermedia, L. laeta and L. gaucho) were tested as antigens in a vaccination protocol. In such isoforms, key amino acid residues involved in catalysis, magnesium-ion coordination, and binding to substrates were replaced by Alanine (H12A-H47A, E32A-D34A and W230A). These mutations eliminated the phospholipase activity and reduced the generation of skin necrosis and edema to residual levels. Molecular modeling of mutated isoforms indicated that the three-dimensional structures, topologies, and surface charges did not undergo significant changes. Mutated isoforms were recognized by sera against the crude venoms. Vaccination protocols in rabbits using mutated isoforms generated a serum that recognized the native PLDs of crude venoms and neutralized dermonecrosis and edema induced by L. intermedia venom. Vaccination of mice prevented the lethal effects of L. intermedia crude venom. Furthermore, vaccination of rabbits prevented the cutaneous lesion triggered by the three venoms. These results indicate a great potential for mutated recombinant PLDs to be employed as antigens in developing protective vaccines for Loxoscelism. Department of Cell Biology Federal University of Paraná (UFPR) Production and Research Center of Immunobiological Products (CPPI) State Department of Health Veterinary Hospital Federal University of Paraná (UFPR) Department of Structural Molecular Biology and Genetics State University of Ponta Grossa (UEPG) Multiuser Center for Biomolecular Innovation Departament of Physics Universidade Estadual Paulista (UNESP) Multiuser Center for Biomolecular Innovation Departament of Physics Universidade Estadual Paulista (UNESP) Universidade Federal do Paraná: 02/2020 Universidade Federal do Paraná: 04/2019 Fundação Araucária: 057/2017 CNPq: 303868/2016-3 CNPq: 408633/2018-2

Loxoscelism is the clinical condition triggered after the bite of spiders of the genus Loxosceles. The main species involved in accidents in South America are L. intermedia, L. laeta, and L. gaucho. The only specific treatment is the anti-Loxosceles serum produced with crude venoms. As phospholipases D (PLDs) trigger most of the effects observed in accidents, we developed and evaluated second-generation sera using mutated PLDs as antigens. Three isoforms of PLDs with site-directed mutations without biological activities were used for rabbit immunizations: D32A-E34A (L. gaucho), W230A (L. intermedia), and H12A-H47A (L. laeta). Sera were produced using crude venoms of three species of Loxosceles enriched with mutated recombinant PLDs (MIX) or using only mutated PLDs (REC). Immunizations stimulated the immune system from the second immunization with higher antibody production in the REC group. In vivo neutralization assays demonstrated that both sera reduced edema and dermonecrosis caused by Loxosceles intermedia crude venom. Follow-up of animals during the immunization protocols and in the neutralization assays demonstrated that the mutated proteins and the sera are safe. Results demonstrate the potential of using mutated recombinant PLDs in total or partial replacement of Loxosceles venoms in animal immunizations to produce anti-Loxosceles sera for treatments of Loxoscelism.

Neospora caninum is one of the most prevalent Apicomplexa parasites that causes abortion in cattle, as it infects dogs as its definitive host, causing subclinical disease or active neosporosis, marked by meningoencephalitis, and myopathies with muscle and neuromuscular signs of disease. This study aimed to evaluate the acute phase protein response in dogs seropositive and seronegative for N. caninum. Serum samples of 72 dogs were tested by an immunofluorescence antibody test using N. caninum NC-1 strain, and the study population was divided into four groups: symptomatic - muscular and/or neuromuscular signs - and seropositive (n = 16); symptomatic and seronegative (n = 9); asymptomatic and seropositive (n = 34); and asymptomatic and seronegative (n = 13). C-reactive protein (CRP) was measured via immunoturbidimetric assay and serum haptoglobin (Hp) via hemoglobin-binding capacity assay. In the symptomatic groups, seropositive dogs had higher levels of Hp, but not CRP, while seronegative dogs had higher CRP levels. There was no difference in CRP concentration in asymptomatic dogs. Dogs with neuromuscular signs had higher concentrations for Hp in the group seropositive. Hp concentration did not differ between dogs seropositive and seronegative dogs for each group. Serum Hp and CRP could not sufficiently alone flag subclinical infections. Measurement of CRP and Hp concentrations could be clinically valuable to the diagnosis of neurological diseases, and their relative change may indicate the stage of the infection, although their sole use is not able to support the diagnosis of canine neosporosis. Further studies are encouraged to evaluate the specific dynamics of acute phase proteins in canine neosporosis. Serum samples of 72 dogs were tested by an immunofluorescence antibody test using N. caninum NC-1 strain, and the study population was divided into four groups: (1) dogs with muscular and/or neuromuscular signs and seropositive for N. caninum; (2) dogs with muscular and/or neuromuscular signs and seronegative for N. caninum; (3) dogs seropositive for N. caninum with no neuromuscular signs; and (4) healthy dogs and seronegative for N. caninum. The study evaluated if N. caninum infection could have pathophysiological changes activating the acute phase response and an increase in the concentration of acute phase proteins in serum.

The present study characterized oligosaccharide compounds (Oligo) in Cabernet Franc red wine and investigated its antineoplastic effects against mammary tumor cells in vivo and in vitro, isolated or in combination with chemotherapy. The Oligo fraction was characterized by nuclear magnetic resonance spectroscopy and mass spectrometry. The complex mixture of Oligo showed high amounts of oligoxyloglucuronans, oligorhamnogalacturonans, oligoarabinogalactans, and oligoglucans, such as trehalose and isomaltotriose. To investigate the antineoplastic effects of Oligo, Female Swiss mice were subcutaneously inoculated with Ehrlich tumor cells and then received vehicle (distilled water, p.o.), Oligo solution (9, 35, or 70 mg kg

Brazil has the highest number of birds of prey in the world, which are important environmental quality indicators. Nevertheless, few studies of the clinical pathology of raptors have been developed in this country. The objectives of this study were to create a database of owl hematology in Brazil and to compare the values between sex in Asio clamator, Megascops spp. and Tyto furcata. Blood samples were collected from 81 captive owls of 10 species located in Rio Grande do Sul, Santa Catarina, Paraná and São Paulo states, Brazil. Hemogram and Total Plasma Protein (TPP) values were determined. Reference intervals (RIs) and descriptive statistic values were established using an Excel program with Reference Value Advisor. The reference intervals were the following: A. clamator: RBC (×106/?L) 1.0-2.5; PCV (%) 30.2-50.1; Hb (g dL-1) 6.7-15.3; MCV (fL) 123.8-355.2; MCHC (%) 17.1-38.6; WBC (× 103/?L) 1.2-23.6; Heterophils (× 103/?L) 0.6-16.6; Lymphocytes (× 103/?L) 0.6-10.2; Eosinophils (× 103/?L) 0.0-1.9; Basophils (× 103/?L) 0.0-1.0; Thrombocytes (× 103/?L) 10.8-56.6; H/L 0.0-10.8. Megascops spp.: RBC (× 106/?L) 0.8-2.3; PCV (%) 29.7-44.6; Hb (g dL-1) 6.4-12.4; MCV (fL) 131.6-374.4; MCHC (%) 12.1-34; WBC (× 103/?L) 0.7-23.1; Heterophils (× 103/?L) 1.1-10.3; Lymphocytes (× 103/?L) 0.0-11.5; Eosinophils (× 103/?L) 0.0-2.2; Basophils (× 103/?L) 0.0-0.7; Thrombocytes (× 103/?L) 10.3-43.6; H/L 0.5-7.3; TPP (g dL-1) 2.9-5.1. The parameters for Tyto furcata were presented with descriptive statistics values. Individual data were provided for the others Strigiformes species sampled. This study provides a wide database of hematological and TPP references for Megascops spp., A. clamator and T. furcata and hematological values for Athene cunicularia, Bubo virginianus, Pulsatrix perspicillata, Asio stygius, Pulsatrix koeniswaldiana, Strix virgata and Asio flammeus in Brazil.

Sarcocystis spp. are cyst forming apicomplexan parasites that infect many vertebrates including birds. Sarcocystis spp. infection was investigated in tissue samples (pectoral muscles, heart, and brain) of 47 dead seabirds collected from the coastline of Santa Catarina State SC - Brazil, between August 2019 and March 2020. A portion of each tissue was fixed in 10% buffered formalin for histopathologic analysis while DNA was extracted from another portion and screened using nested-PCR targeting ITS1. Based on molecular analysis, Sarcocystis spp. were identified in 15/47 (31.9%) seabirds of five species, kelp gull (Larus dominicanus), manx shearwater (Puffinus puffinus), neotropic cormorant (Phalacrocorax brasilianus), brown booby (Sula leucogaster) and great skua (Stercorarius skua). Microscopically visible sarcocysts were observed only in the pectoral muscle of four seabirds 8.5% (4/47), while in one brown booby, sarcocysts were seen in both pectoral and cardiac muscles. Two types of sarcocysts, thin walled (≤1 μm) and thick-walled (≥ 2 μm) were identified. Based on ITS1 sequence comparison, S. halieti, S. falcatula and three not yet described Sarcocystis spp. were detected. Phylogenetically, S. falcatula isolates were classified as two distinct clusters. This is the first confirmation of S. halieti in seabird's species in South America and S. falcatula in birds of the order Charadriiformes. Further molecular studies are needed to understand the epidemiology of the Sarcocystis spp. infection and its impact on the health of seabirds.

The present study investigated the antineoplastic effects of pectic polysaccharides that were extracted from green sweet pepper (Capsicum annuum [CAP]) in the Ehrlich carcinoma in mice and in human mammary tumor lineages. After the subcutaneous inoculation of 2 × 106 Ehrlich tumor cells, Female Swiss mice received 50, 100, or 150 mg/kg CAP or vehicle orally once daily or methotrexate (2.5 mg/kg, i.p., every 5 days) for 21 days. CAP dose-dependently reduced Ehrlich tumor growth. It also reduced the viability of MCF-7, MDA-MB-231, and MDA-MB-436 human mammary cell lineages. Treatment with CAP reduced the gene expression of vascular endothelial growth factor in vivo and in vitro, reduced vessel areas of the tumors, and induced necrosis in Ehrlich solid tumors. CAP treatment significantly increased Interleukin-6 in tumors. The antineoplastic effect of CAP appears to depend on the regulation of inflammation and angiogenesis. Further studies are encouraged to better understand the CAP potential for the treatment of breast tumors.

O objetivo do presente estudo foi desenvolver um diluente de sêmen de touros eficiente no controle de Neospora caninum. O teste dos antimicrobianos (experimento 1) - trimetoprim isolado (TRI) e associado a sulfadiazina (SDT) e ao sulfametoxazol (SMT), claritromicina (CLA), eritromicina (ERI), azitromicina (AZI) e clindamicina (CLI) - foi realizado utilizando cultivo celular e avaliado quanto à presença de taquizoítas livres e ao efeito citopático na monocamada. Os antimicrobianos que se mostraram eficazes no controle dos taquizoítos in vitro foram utilizados na preparação do diluente para o sêmen. Cada antimicrobiano foi avaliado quanto à toxicidade dos espermatozóides (experimento 2) sendo 8 grupos (7 grupos receberam antibiótico no diluente, exceto 1 grupo- grupo controle). Em relação à análise morfológica, os grupos SMT e AZI apresentaram mais defeitos de acrossoma e de cauda.Os grupos SDT, SMT e AZI apresentaram valores mais baixos da Velocidade de Trajeto. A Velocidade Progressiva foi menor nos grupos SMT e AZI e a Frequência de Batimentos de cauda também foi menor nos grupos CLI e AZI. Os valores de Velocidade Curvilinear dos grupos SDT, SMT e CLA foram menores. Os grupos SMT, ERI e AZI apresentaram valores mais baixos de Motilidade Total e Motilidade Progressiva. Quanto à citometria de fluxo, os grupos SDT, SMT e AZI apresentaram percentagens maiores de reação acrossomal e membrana plasmática lesada. Os antibióticos que podem ser adicionados ao diluente de sêmen bovino foram clindamicina, claritromicina e trimetoprim.

B ackground : Acute lymphoblastic leukemia (ALL) is a malignant neoplasia in which there is proliferation of lymphoid progenitor cells in the bone marrow, blood, and extramedullary sites. This disorder has a fast and progressive development; in dogs, cases of infiltration of ALL cells in the central nervous system (CNS) are uncommon and rare. Diagnosis can be achieved with the help of the clinical history and physical, radiographic, hematological, myelographic, and cerebrospinal fluid (CSF) tests in patients with or without neurological clinical signs. The present report aims to describe a case of ALL and the presence of lymphoblasts in the CSF of a dog with neurological clinical signs. Case : An 8-year-old Lhasa Apso dog was examined at the Veterinary Hospital of Universidade Federal do Parana, Curitiba campus. At the physical examination, the animal exhibited apathy and paralysis of pelvic limbs, which progressed to tetraplegia. Abdominal palpation revealed presence of hepatosplenomegaly and absence of lymphadenomegaly. No alterations were observed in radiographs of the cervical, thoracic or lumbar spine. A complete blood count revealed presence of non-regenerative anemia (hematocrit = 22%), extreme lymphocytosis (185,229 cells/µL), lymphoblasts at a level of 72% (133,364 cells/µL), and thrombocytopenia (66,000 platelets/µL). The biochemical tests revealed increased alkaline phosphatase (859 IU/L). The levels of alanine aminotransferase, creatinine, urea, total protein, albumin, and globulin were normal. The diagnosis of ALL was achieved with the help of a myelogram. The myelogram findings included 39% of mature lymphocytes and 59% of lymphoblasts exhibiting large size, spherical shape, poorly delimited borders, with a high nucleus/cytoplasm ratio, marked cytoplasmic basophilia, and 2 to 3 evident nucleoli; metarubricytes (1%) and promyelocytes (0.6%) were also observed. The CSF contained an increased number of nucleated cells (27 cells/µL) comprising lymphocytes (43%), macrophages (33%), and segmented neutrophils (24%). Of the 11.6 lymphocytes per µL of CSF, 8.1 were lymphoblasts, which indicates infiltration of ALL cells in the CNS. The animal died one day after collection of bone barrow and CSF. Discussion : Relevant alterations observed in this case included the neurological signs caused by the infiltration of neoplastic cells in the CNS, severe leukocytosis and lymphocytosis, with large amounts of lymphoblasts in the blood and predominance of lymphoblasts in the bone marrow, which are alterations typically found in ALL. The animal also exhibited non-regenerative anemia and thrombocytopenia, which were secondary to infiltration of leukemic cells in the bone marrow. The CSF exhibited pleocytosis (27 cells/ µL), and 30% of the cells observed were lymphoblasts. Lymphoblast infiltration in the CNS of leukemic dogs is rare, and other studies have reported absence of neurological signs or neurological signs different from those observed in the present study. CSF analysis in indicated in cases of leukemia to assess leukemic cell infiltration in the CNS. In the case reported here, the plasma level of alkaline phosphatase was increased (859 IU/L) as a consequence of hepatomegaly and hepatic cholestasis. ALL is a very aggressive, proliferative neoplasia, and the resulting lymphoblasts infiltrated the CNS of the animal. In cases of ALL, performing complete blood count, myelogram, and CSF analysis is indicated whether the patients exhibit neurological signs or not.

Background: The indiscriminate use of drugs is an issue in Veterinary Medicine, as it has serious consequences for the animals. Many drugs are myelotoxic and cause a decrease in the production of blood cells, which may be irreversible in some cases. The present work reports a case of pancytopenia induced by the concomitant use of myelotoxic drugs (estrogen, metamizole and phenobarbital) in a dog and describes findings on myelotoxicity, hematological alterations and treatment success.Case: A 7-year-old Lhasa Apso bitch was referred to the Veterinary Hospital of Federal University of Paraná, Curitiba campus, with hematuria and a history of treatment with phenobarbital [2 mg/kg twice a day (bis in die, BID)], metamizole [25 mg/kg three times a day (ter in die, TID)], and use of estrogen hormone (estradiol cypionate). At physical examination, the animal was normohydrated and exhibited normal palpable lymph nodes, pale mucous membranes, galactorrhea, and a body temperature of 36°C. A complete blood count including reticulocyte count and a total plasma protein (TPP) exam were requested. The results revealed pancytopenia (18% hematocrit, 1,400 total leucocytes/µL, and 22,000 reticulocytes/µL). An abdominal ultrasound exam did not detect any relevant alterations. In view of the results obtained, medullary aplasia was suspected. A bone marrow aspiration was performed. A myelogram revealed a decrease in cellularity (erythrocytic and granulocytic hypoplasia), with presence of rare erythroid and granulocytic precursors. The diagnosis was medullary aplasia. The animal was treated, and the evolution of the hematological alterations was monitored. The treatment consisted of administration of erythropoietin (100UI/kg subcutaneously every 48 h), prednisone (2 mg/kg BID), Leucogen (3 mg/kg BID), interferon (0.2 IU/kg BID) and Eritrós Dog Tabs [1 tablet once a day (semel in die, SID)]. After five days of treatment, the patient’s clinical picture improved (30% hematocrit, 5,300 total leukocytes/µL, 84,000 platelets/µL, and 195,000 reticulocytes/µL), and the hematological alterations were resolved after 25 days of treatment (43% hematocrit, 5,100 total leukocytes/µL, and 333,000 platelets/µL). The bitch was discharged after 89 days of treatment.Discussion: The hematological alterations found in the patient were consistent with pancytopenia, and the myelogram allowed the establishment of a diagnosis of medullary aplasia. There are various causes of pancytopenia in dogs; in this case, it was caused by medications, as the drugs administered to the patient (estrogens, metamizole, and phenobarbital) are myelotoxic. Canine bone marrow is susceptible to suppression by estrogens, which can induce medullary aplasia even with a single dose. No reports on hematological alterations caused by dipyrone (metamizole) in dogs were found; however, in humans, development of aplastic anemia, agranulocytosis, nephrotoxicity, and allergic reactions have been attributed to the use of this drug. Phenobarbital can cause adverse reactions that lead to anemia, leukopenia, and thrombocytopenia. Evaluating the bone marrow of animals with pancytopenia is important because this procedure allows the establishment of a diagnosis that may prompt treatment while hematopoietic precursors are still present in the bone marrow. In this case, a treatment using hematopoietic stimulants was employed owing to the presence of erythrocytic and myelocytic precursors in the patient’s bone marrow. The treatment instituted was efficacious, as only five days of therapy already improved the hematological condition of the patient, who was discharged after 89 days of treatment.

O Neospora caninum é um protozoário intracelular identificado pela primeira vez como causador de encefalomielite e miosite em cães e desde então foi detectado em ampla variedade de animais domésticos. O cão e os canídeos selvagens são os hospedeiros definitivos do protozoário e eliminam oocistos nas fezes e se infectam pela ingestão de cistos em tecidos infectados. Embora a transmissão vertical seja altamente eficiente em vacas, a transmissão congênita de Neospora caninum por infecção natural em cães é pouco relatada (DUBEY & SCHARES, 2011). O objetivo do presente estudo foi relatar a presença de DNA de Neospora caninum em amostra de fígado de um filhote de uma ninhada de uma cadela soropositiva. Uma cadela da raça Bulldog Francês de 3 anos de idade, matriz reprodutora de um criatório de cães e assintomática, apresentava histórico de uma primeira gestação de 3 de 5 filhotes que desenvolveram sinais neurológicos e vieram a óbito nas primeiras semanas de vida. Essa cadela apresentou soropositividade para Neospora caninum na RIFI com titulação 1:100. Amostras de tecidos desses filhotes necropsiados foram testados para N. caninum no PCR e foram negativos. Ela foi inseminada artificialmente novamente com sêmen de um macho soronegativo para uma segunda gestação. Nessa gestação nasceram cinco filhotes, sendo que quatro apresentaram sinais clínicos, como letargia, dispnéia, dificuldade para mamar, choro persistente, ataxia e opacidade da córnea, que se iniciaram aos cinco aos setes dias de vida. Foram tratados com clindamicina e sulfa com trimetropim, mas vieram a óbito no décimo dia. Os filhotes foram necropsiados e amostras de tecidos (Cérebro, coração, fígado, rim, pulmão e timo) foram coletadas para a realização do diagnóstico molecular e análise histopatológicade. A extração de DNA foi realizada com kit comercial. Para a amplificação de DNA N. caninum foi empregada a reação de cadeia da polimerase nested-PCR com os primers NP6 plus, NP21 plus, NP7 e NP6, que amplificam a região NC5. Foi detectado DNA de N. caninum na amostra do fígado. Na análise histopatológica os tecidos não apresentaram alterações macroscópicas e microscópicas sugestivas de neosporose. A transmissão transplacentária em cadelas não é eficiente, cerca de 80% dos filhotes de cadelas soropositivas não são infectados, mas cadelas com infecção subclínica pode transmitir o protozoário em gestações sucessivas (BARBER & TREES., 1998). A maioria dos relatos são de infecções experimentais, nem sempre é observada a transmissão vertical e a ocorrência pode variar entre as gestações. Os casos de neosporose mais graves geralmente acometem filhotes infectados congenitamente e os sinais clínicos se iniciam a partir das três semanas após o nascimento. O presente relato ressalta a possibilidade de transmissão congênita de N. caninum em cadelas em gestações sucessivas, a importância de realizar sorologia para N. caninum nas matrizes reprodutoras dos criatórios e retirá-las da reprodução em caso de soropositividade.

A pardela-preta ( Procellaria aequinoctialis ) e uma ave procellariiforme, possui distribuicao circumpolar ao longo de todo o oceano austral e migra para o hemisferio sul durante o inverno. E considerada uma especie globalmente vulneravel pela Uniao Mundial para a Conservacao da Natureza (IUCN, 2020), devido a mortalidade acidental da especie na pesca com espinhel e a degradacao do seu habitat natural causando rapido declinio populacional. Neospora caninum e um protozoario intracelular formador de cisto que infecta aves e mamiferos. Os canideos domesticos e silvestres sao os hospedeiros definitivos e eliminam os oocistos nas fezes. As aves se infectam pela ingestao de alimentos e agua contaminados com oocistos esporulados e por meio da ingestao de cistos em tecidos infectados (DONAHOE et al.,2015). O objetivo do presente estudo foi relatar a presenca de DNA de Neospora caninum no coracao de uma ave pardela-preta ( P. aequinoctialis) , femea e adulta. Amostras de tecidos (musculatura peitoral, cerebro e coracao) foram coletadas para a realizacao do diagnostico molecular de N. caninum e acondicionadas e fixadas em formalina 10% tamponada para analise histopatologica. A extracao de DNA foi realizada com kit comercial. Para a amplificacao de DNA N. caninum foi empregada a reacao de cadeia da polimerase nested -PCR com os primers JS4, CT2c, JS4b e CT2b, que amplificam a regiao ITS1. Foi detectado DNA de N. caninum na amostra do coracao. O produto da PCR foi submetido a purificacao e sequenciamento. Foi obtida sequencia nucleotidica (MW023246) que apresentou identidade de 99% com isolados de N. caninum depositados no Genbank . Na analise histopatologica os tecidos nao apresentavam alteracao macroscopica e a analise microscopica revelou congestao no miocardio. O presente relato demonstra que a pardela-preta ( P. aequinoctialis ) pode ser naturalmente infectada por N. caninum e ate o momento e primeira descricao nesta especie de ave. Existem poucos estudos sobre a ocorrencia de N. caninum na fauna marinha e as possiveis fontes de infeccao ainda nao foram bem elucidadas. Estudos demonstraram que mamiferos marinhos (lontras, morsas e golfinhos) estao expostos ao protozoario indicando uma possivel contaminacao das aguas marinhas por oocistos O achado nesse relato sugere que as aves marinhas podem ser potenciais hospedeiros intermediarios de N. caninum , atuando como sentinela e indicador de contaminacao do ambiente marinho, alem da sua participacao no ciclo silvestre do parasito.

The closely related apicomplexa protozoa, Toxoplasma gondii, Neospora spp., and Sarcocystis neurona, have all been associated with neurological and reproductive diseases in horses. However, there is limited data regarding the presence of these three parasites in equine placental tissues and amniotic fluid. The aim of the present report was to investigate the presence of the DNA of T. gondii, Neospora spp. and S. neurona in placentas and amniotic fluid in mares. Anti-T. gondii, anti- S. neurona and anti- Neospora spp. antibody titers were evaluated in 31 mares in the final third of pregnancy by indirect fluorescent antibody test (IFAT). The presence of parasite DNA in placentas and amniotic fluid was evaluated by polymerase chain reaction (PCR), using two target loci (ITS1 and Nc5). No antibodies to were identified nor was any T. gondii DNA detected in any mare. Antibodies to Neospora spp. were found in 6 mares (19.35 %) and DNA from this protozoan was detected in four placentas (12.9 %) and in five amniotic fluid samples (16.6 %). Antibodies to Sarcocystis spp. were detected in nine mares and S. neurona DNA was found in only one placenta (3.23 %). Our results suggest that the transplacental route may be a potential source of Neospora caninum infection in mares. Further studies are needed to understand the role of transplacental transmission in the epidemiology of these protozoa.

We aimed to verify whether age influences the biochemical profile of healthy lambs during the first four months of life by characterizing the dynamics of its parameters and verifying whether there are differences between the profiles of growing animals and the reference values established for adults. Blood samples of 34 ½ White Dorper × ½ Suffolk female lambs were collected at 30, 60, 90, and 120 days of age, and the serum concentrations of total proteins, total globulins, albumin, urea, creatinine, bilirubins (total, conjugated, and unconjugated), total cholesterol, triglycerides, glucose, gamma-glutamyltransferase, aspartate aminotransferase, and creatine kinase were determined. Descriptive statistics of these parameters were performed to compare them with the reference intervals used for adult sheep. Age influenced the dynamics of all the biochemical parameters between 30 and 120 days of age. Moreover, except for the total, conjugated and unconjugated bilirubins, total cholesterol, and aspartate aminotransferase means, the values of the lambs differed from the reference intervals established for adults. In conclusion, there is an influence of age on biochemical parameters in healthy female lambs in the first four months of life, and often the biochemical parameters of young animals differ from those of adults. Therefore, the interpretation of tests performed on growing animals should be made based on specific reference intervals for this age group.

Raptors are carnivorous birds with great hunting ability. Toxoplasma gondii, Neospora caninum and Sarcocystis spp. are intracellular Apicomplexan protozoans which infect a wide range of intermediate hosts, including birds. The aims of this study were to evaluate the serological reactivity of captive raptors serum to T. gondii, N. caninum and S. neurona antigens and identify possible risk factors associated with the infection. From August 2014 to September 2015, blood samples from 72 raptors were collected and serum samples were tested by immunofluorescence antibody test (IFAT). Antigen slides were prepared using tachyzoites of T. gondii and N. caninum and using merozoites of S. neurona. Serum samples were tested at the following cut-off dilutions: 1:16 for T. gondii and 1:50 for N. caninum and S. neurona. An anti-chicken IgY antibody conjugated with FITC was used as a secondary antibody at 1:50 dilution. Out of the 72 raptors serum tested by IFAT, 2.7% reacted to N. caninum, 8.3% to T. gondii and 11.1% to S. neurona antigens. The region in which the sample was collected, the reason the raptors were kept in captivity and diet were statistically associated with seropositivity to T. gondii and the use of the birds and diet were statistically associated with seropositivity to N. caninum and S. neurona (p ≤ 0.05). We highlight the occurrence of these protozoans in birds of prey and the importance of good hygiene and feeding management of these birds in captivity to reduce the risk of protozoal infections.

Fungal pneumonia has been a differential diagnosis in dogs and cats with pulmonary clinical signs, posting also a public health risk to respective owners. Since a common-source environmental exposure may result in infection, dogs and cats may also act as sentinels for both animal and human disease. Accordingly, the present study has aimed to evaluate the occurrence of fungal pneumonia in dogs and cats with pulmonary clinical signs from May 2013 to February 2015 in southern Brazil. Thoracic radiographs, blood work and non-bronchoscopic Bronchoalveolar Lavage (BAL), followed by cytology and microbiologic analysis were performed in all animals. One dog was positive in the BAL fluid culture for a pathogenic fungus (Cryptococcus neoformans) and another dog to an opportunistic fungus (Candida parapsilosis). Both dogs have presented cough as the prior clinical sign and showed thoracic radiographic changes. In conclusion, the occurrence of fungal pneumonia in southern Brazil in dogs and cats through of this study was relatively low. The BAL fluid culture may allow diagnosis of fungal pneumonia caused by opportunistic or pathogenic fungi and this diagnosis should be a concern in dogs and cats with pulmonary clinical signs. RESUMO: Pneumonia fúngica tem sido um diagnóstico diferencial em cães e gatos com sinais clínicos pulmonares, sendo também um risco de saúde pública para seus respectivos proprietários. Uma vez que uma fonte comum de exposição ambiental pode resultar em infecção, cães e gatos podem agir como sentinelas da doença para ambos, pessoas e animais. Assim, o presente estudo teve como objetivo avaliar a ocorrência de pneumonia fúngica em cães e gatos com sinais clínicos pulmonares, de maio de 2013 a fevereiro 2015 no sul do Brasil. Radiografias torácicas, exame de sangue e lavado broncoalveolar (LBA) não broncoscópico seguido de análise citológica e microbiológica foram realizados em todos os animais. Um cão foi positivo na cultura do fluido do LBA para um fungo patogênico (Cryptococcus neoformans) e outro cão para um fungo oportunista (Candida parapsilosis). Ambos os cães apresentaram tosse como sinal clínico principal e alterações radiológicas torácicas. Em conclusão, a ocorrência de pneumonia fúngica no sul do Brasil em cães e gatos por meio deste estudo foi relativamente baixa. A cultura do LBA pode possibilitar o diagnóstico de pneumonia fúngica causada por fungos oportunistas ou patogênicos e este diagnóstico diferencial deve ser uma preocupação em cães e gatos com sinais clínicos pulmonares.

Bovine neosporosis has become a disease of international concern as it is among the main causes of abortion in cattle. Viable N. caninum has been isolated from brains of fetuses and neonatal calves, and there is no report of isolation of tachyzoites from kidney. Also, detailed information about the genetic diversity of N. caninum is scarce. N. caninum tachyzoites were isolated from the kidney and the brain of an aborted 4-month-old bovine foetus. The parasite was confirmed to be N. caninum by PCR. The tachyzoites of the new isolate, named BNC-PR4, were propagated in Vero cell cultures. Pathogenicity of the parasite was examined in BALB/c mice. Mice inoculated intraperitoneally with BNC-PR4 failed to yield clinical signs of disease and did not induce severe brain lesions, suggesting a bovine isolate with low virulence. The N. caninum-positive DNA sample was further analyzed by multilocus microsatellite (MS) genotyping for MS4, MS5, MS6A, MS6B, MS7, MS8, MS10, MS12, and MS21. Multilocus-microsatellite genotyping revealed a unique genetic profile that differed from previously reported isolates.

Toxoplasma gondii, Neospora caninum and Sarcocystis neurona are obligate intracellular parasites within the phylum Apicomplexa. The red-tailed Amazon parrot (Amazona brasiliensis) is a near-threatened species of psittacine that is endemic to the Atlantic Forest of Brazil and has been designated as a bioindicator because of its sensitivity to environmental qualitative status and changes. The aim of this study was to evaluate the presence of antibodies against T. gondii, N. caninum and S. neurona in wild red-tailed Amazon parrot nestlings on Rasa Island, Brazil. Blood samples were collected from 51 parrots and plasma samples were stored at – 20 °C until immunofluorescence antibody tests (IFAT) were performed. Antigen slides were prepared using tachyzoites of T. gondii (RH strain) and, N. caninum (NC-1 strain) and using merozoites of S. neurona (SNR37 strain). Plasma samples were tested at initial dilutions of 1:16 for T. gondii, 1:50 for N. caninum and 1:5 for S. neurona. An anti-chicken antibody conjugated with FITC was used as a secondary antibody at 1:50 dilution. No antibodies for any of these three protozoa were found, thus suggesting that these wild red-tailed Amazon parrot nestlings had not been exposed to these parasites.

O presente trabalho investigou a ocorrência e distribuição de anticorpos contra Sarcocystis neurona, Neospora caninume Toxoplasma gondii em cavalos, cães e gatos de Curitiba, estado do Paraná, Brasil. Amostras de soro de 100 cavalos,100 cães e 100 gatos da rotina do Laboratório de Patologia Clínica Veterinária do Hospital Veterinário da UniversidadeFederal do Paraná (UFPR) foram selecionadas. As 100 amostras de cães foram divididas em dois grupos: 35 amostrasde animais com sinal neurológico (convulsão) e 65 sem sinais neurológicos. Os animais eram adultos de diferentesraças, machos e fêmeas. As amostras foram analisadas pelo teste de reação de imunofluorescência indireta (RIFI) paraos protozoários S. neurona, N. caninum e T. gondii nas seguintes diluições de corte: cavalos: 1:50, 1:50 e 1:16; cães: 1:50,1:50 e 1:16; gatos: 1:50, 1:50 e 1:50, respectivamente. Os resultados obtidos foram 42% dos cavalos, 7% dos cães e 5% dosgatos soropositivos para S. neurona; 58% dos cavalos, 68% dos cães e 42% dos gatos soropositivos para N. caninum; e 36%dos cavalos, 20% dos cães e 21% dos gatos soropositivos para T. gondii. Entre os cães com sinal neurológico, 8.6%, 68.6%e 25.7% deles foram soropositivos para S. neurona, N. caninum e T. gondii, respectivamente. Entre os cães sem sinaisneurológicos, 6.2% 67.7% e 16.9% foram soropositivos para S. neurona, N. caninum e T. gondii, respectivamente. Não foiencontrada diferença estatística entre os grupos de cães soropositivos para os três protozoários com sinal neurológicoe sem sinais neurológicos. Coinfecção e altos títulos de anticorpos foram detectados. Os anticorpos contra Sarcocystisneurona, Neospora caninum e Toxoplasma gondii foram encontrados amplamente distribuídos entre cavalos, cães e gatosna região de Curitiba, estado do Paraná, Brasil. The occurrence and distribution of antibodies against Sarcocystis neurona, Neospora caninum and Toxoplasma gondii wasinvestigated in horses, dogs and cats from Curitiba, Paraná state, Brazil. Serum samples were selected from 100 horses,100 dogs and 100 cats from the routine of the Veterinary Clinical Pathology Laboratory in the Veterinary Hospital of theUniversity of Paraná (UFPR). The 100 dog samples were divided into two groups: 35 samples from dogs with neurologicalsign (convulsion) and 65 samples from dogs without neurological signs. The animals were adults of different breeds, malesand females. Samples were analyzed by indirect fluorescent antibody test (IFAT) for protozoa S. neurona, N. caninum andT. gondii at the following cut-off dilutions: horses: 1:50, 1:50 and 1:16; dogs: 1:50, 1:50 and 1:16; cats: 1:50, 1:50 and 1:50,respectively. The obtained results were 42% of horses, 7% of dogs and 5% of cats seropositive for S. neurona; 58% of horses,68% of dogs and 42% of cats seropositive to N. caninum, and 36% of horses, 20% of dogs and 21% of cats seropositive forT. gondii. Among the dogs with neurological signs, 8.6%, 68.6% and 25.7% were seropositive for S. neurona, N. caninumand T. gondii, respectively. Among the dogs without neurological signs, 6.2% 67.7% and 16.9% were seropositive forS. neurona, N. caninum and T. gondii, respectively. No statistical difference was found between groups of seropositivedogs for the three protozoa with neurological signs and without neurological signs. Co-infection and high antibodytiters were detected. The antibodies against Sarcocystis neurona, Neospora caninum and Toxoplasma gondii were foundwidely distributed among horses, dogs and cats in the region of Curitiba, state of Paraná, Brazil. &nbsp

The black-fronted piping-guan (

Background : Neospora caninum and Toxoplasma gondii are closely related cyst-forming apicomplexan parasites identified as important causes of reproductive failure in cattle. Moreover, abortion cases attributed to N. caninum and T. gondii infection have been occasionally reported in sheep. Due to the relatively scarce information on the molecular detection of N. caninum in the semen of naturally infected rams, this study aimed to detect parasitic DNA in fresh semen samples and in frozen extended semen straws from male sheep from artificial inseminations centers in Southern Brazil. Materials, Methods & Results : Semen samples of 38 rams from artificial insemination centers were evaluated. Eleven rams were naturally infected (seropositive for anti- N. caninum and/or anti- T. gondii IgG) and were selected for fresh semen collection. We tested all the samples for the closely related protozoan T. gondii to detect a possible cross-reaction and co-infection, due to the close similarity with N. caninum . The indirect fluorescent antibody test was used to detect IgG antibodies in the 11 serum samples from rams. Fresh semen samples were collected from 11 rams on days 1, 50, 55, and 58 using an artificial vagina and ewe in estrus. Other 27 rams had their frozen extended semen straws analyzed. A total of 20 fresh semen samples and 27 frozen extended semen straws samples were used to detect the presence of N. caninum and T. gondii DNA by polymerase chain reaction (PCR). Nc-5 and B1 genes were used as target regions to detect N. caninum and T. gondii DNA, respectively. The presence of N. caninum DNA was confirmed in the third collection of a fresh semen sample of one seropositive ram. T. gondii DNA was detected in a fresh semen sample of one seropositive ram. The DNA sequences of 186 bp from N. caninum (GenBank accession: MH806393) and 492 bp from T. gondii (GenBank accession: MH793503) were obtained by sequencing, and analysis revealed 99% and 100% identity, respectively, compared with other sequences deposited at GenBank. N. caninum and T. gondii DNAs were not detected in any of the 27 frozen extended semen straws used for artificial insemination. Discussion : This study demonstrated the presence of N. caninum and T. gondii DNA in fresh semen samples of naturally infected rams. The non-detection of N. caninum and T. gondii DNA in frozen semen samples of rams could be due to the dilution that was used to prepare the semen straws (GGL diluent and 5% glycerol), since fresh semen samples were not diluted prior to the test. Moreover, in our study, the volume of frozen semen samples (0.25 mL) used for PCR was lower than the volume of sediment obtained from fresh semen (0.5 mL), and the fresh semen centrifugation to obtain the sediment may have grouped the tachyzoites, increasing the sensitivity of the technique employed. No high IgG serological titers were detected in the rams at the time they were eliminating the parasite through fresh semen. The final titer of anti- N. caninum and anti -T. gondii IgGs in serum was 1:100, suggesting chronic infection. It is suggested that a new parasite elimination pathway is occurring among rams used for reproduction, due to the presence of N. caninum and T. gondii DNA in fresh semen samples from seropositive animals. Although the detection of genomic DNA of N. caninum and T. gondii in semen does not necessarily imply the presence of infectious stages of the parasites and does not determine their viability, these results demonstrate the need for further studies. Our study also indicates the need to reinforce preventive measures for sheep in artificial insemination centers until the risks are evaluated, by performing serological examinations with anti- N. caninum and anti- T. gondii antibodies, for instance, to select the rams that will be used for breeding.

Rangelia vitalii infects erythrocytes, leukocytes and endothelial cells of dogs. The present study aimed to report the molecular detection confirmed by sequencing of R. vitalii in the state of Paraná, as well as describe the clinical, hematological and biochemical alterations of the infected dogs. Three sick dogs from the metropolitan area of Curitiba, PR, Brazil, underwent a physical exam, and laboratory tests included hematology, biochemistry, polymerase chain reaction (PCR), and gene sequencing. Clinical signs included apathy, anorexia, and hemorrhage. Intra-erythrocytic and extracellular piroplasms were found on peripheral blood smears from all three dogs. Blood samples from these animals were positive for Babesia sp. by PCR targeting 18S rRNA. PCR products from all three dogs were sequenced, and BLAST analysis showed that the PCR-generated sequences were highly homologous with those of R. vitalii previously reported. Hematologic findings included severe anemia, shift of neutrophils to the regenerative left, and thrombocytopenia. Serum urea levels were increased in all three dogs, and direct bilirubin levels were elevated in one dog.