Luis Fernando Pita Gondim

Isolation of Neospora caninum from the brain of a naturally infected dog, and production of encysted bradyzoites in gerbils

Neospora caninum was isolated from the brain of an adult dog in Brazil. Cerebral tissue from the dog was inoculated into Mongolian gerbils. Gerbils were euthanized 3-4 months later and bradyzoite-containing tissue cysts were observed in their brains. N. caninum (designated NC-Bahia) was isolated in cell culture after inoculation with tissue cysts from the gerbils. The identity of the parasite was confirmed by immunohistochemical examination and polymerase chain reaction (PCR). Gerbils may be a useful alternative to immunosuppressed mice for isolation of N. caninum and for production of encysted bradyzoites.

Neosporosis as a cause of abortion in dairy cattle in Rio Grande do Sul, southern Brazil

Forty-six aborted bovine fetuses submitted to the Faculty of Veterinary, Department of Clinical Pathology, Universidade Federal do Rio Grande do Sul, were examined histopathologically. Non-suppurative inflammation was observed mainly in the brain and heart of 22 fetuses. Brain lesions consisted primarily of mononuclear inflammatory cell infiltrates with occasional foci of necrosis. Protozoa that reacted with Neospora caninum antisera were seen in 18 of the 22 (81.8%) brain specimens from fetuses with encephalitis. Blood samples collected from 223 Holstein cows on five dairy herds were tested for N. caninum antibodies by an immunofluorescent antibody technique. These samples were obtained from aborting cattle and normally calving cattle (control group). Overall, 11.2% of cows sampled had N. caninum antibodies at a dilution of 1:200. Seroprevalence was higher (P = 0.0053) in aborting (23.3%) than in non-aborting cows (8.3%). Association between seropositivity to N. caninum and abortion was found, with seropositive cows being 3.3 times more likely to abort than seronegative cows (OR = 3.33; 95% CI: 1.38, 8.062). Additionally, N. caninum antibodies were detected in sera from seven cows that had aborted fetuses with lesions suggestive of protozoal infection. These results suggest that N. caninum is an important cause of abortion in dairy cattle in Rio Grande do Sul State, Brazil.

Seroprevalence of Neospora caninum in dairy cattle in Bahia, Brazil

Sera collected from 447 dairy cattle on 14 dairy farms were tested for Neospora caninum antibodies by use of an immunofluorescent antibody technique. Positive reactions with titres > or =1:200 were found in 63 (14.09%) of animals. Neospora positive sera were also tested for Toxoplasma gondii antibodies by using a commercial latex agglutination test. Antibodies to T. gondii were detected in 3 (4.76%) of 63 N. caninum positive sera. These results indicate that N. caninum infection is widespread among dairy cattle in Bahia state.

VARIATION OF THE INTERNAL TRANSCRIBED SPACER 1 SEQUENCE WITHIN INDIVIDUAL STRAINS AND AMONG DIFFERENT STRAINS OF NEOSPORA CANINUM

Small differences have been reported in the internal transcribed spacer 1 (ITS1) region among strains of Neospora caninum. We compared ITS1 sequences among 6 N. caninum strains analyzed in our laboratory, including 2 strains that have not been examined previously (NC-Illinois and NC-Bahia). Five sequences showed 100% similarity and also were identical to 7 of 11 sequences that were previously reported by others. In contrast, initial attempts to sequence the ITS1 of NC-Bahia generated 12 nucleotide differences compared with the other 5 strains, and several ambiguous bases. However, the single band containing the ITS1 region, as observed after electrophoresis on a 2% agarose gel, became divided into 2 distinct bands when reanalyzed using 5 or 10% polyacrylamide gel electrophoresis (PAGE), and the ITS1 within these separate bands were sequenced without ambiguity. The other 5 N. caninum strains were also reexamined using PAGE, and in each strain 2 distinct bands were discovered. In comparison, 2 strains of Toxoplasma gondii continued to show only 1 band when examined using PAGE. The ITS1 sequence of NC-Bahia, from Brazil, differs in several base pairs from those of North American and European strains of N. caninum. Intrastrain variation of the ITS1 region appears to be common in N. caninum, in contrast to T. gondii.

Occurrence of antibodies to Neospora caninum and Toxoplasma gondii in dairy cattle from the northern region of the Paraná State, Brazil

Three-hundred and eighty-five serum samples were taken from dairy cows on 90 farms in 12 counties from the northern region of the Parana State, Brazil. The samples were analyzed by IFAT for the detection of anti-Neospora caninum and anti-Toxoplasma gondii IgG antibodies. Forty-five (12%) samples were seropositive to N. caninum, while 102 (26%) samples were seropositive to T. gondii. Only four animals were seropositive to both coccidia. No significant difference was observed between the N. caninum serology and any of the variables studied, such as dairy cattle management, milk production, reproductive problems, feeding, and presence of dogs, cats and rodents. These data suggest that neosporosis is present among dairy cattle in the studied geographic region and the simultaneous detection of serum positive animals to both types of coccidian parasite demonstrates the independent occurrence of these coccidia in dairy cows.

Investigation of Neospora caninum, Hammondia sp., and Toxoplasma gondii in tissues from slaughtered beef cattle in Bahia, Brazil

Neospora caninum, Hammondia sp., and Toxoplasma gondii are parasites with morphological and genetic similarities. N. caninum and T. gondii are important abortive agents of cattle and sheep, respectively, and may infect numerous animal species. Hammondia sp. is not known to induce disease in animals, but may cause confusion in the identification of closely related coccidia. The aim of this study was to investigate infection rates caused by N. caninum, Hammondia sp., and T. gondii in beef cattle using a nested PCR for Toxoplasmatinae rDNA, followed by sequencing of the PCR products. Antibodies to N. caninum and T. gondii were also investigated in the tested animals. Brains and hearts were obtained from 100 beef cattle in a slaughterhouse in Bahia. Seven samples from brain tested positive for Toxoplasmatinae DNA. No positive reactions were found in heart tissues. After sequencing of the PCR products from all positive tissues, five sequences matched with N. caninum and two matched with T. gondii. Antibodies to N. caninum and T. gondii were found in 20% and 26% of the animals, respectively. The confirmation of N. caninum and the absence of Hammondia heydorni in the tested animals is suggestive that cattle are not efficient intermediate hosts of H. heydorni; however further studies need to be performed using a greater variety of tissues and a higher sample size. The detection of T. gondii DNA in bovine tissues reinforces the potential risk of transmission of this parasite to humans and other animals through the consumption of bovine meat.

Prevalence and risk factors associated to infection by Toxoplasma gondii in ovine in the State of Alagoas, Brazil

The aim of this study was to evaluate the prevalence of anti-Toxoplasma gondii antibodies and to identify risk factors associated to the infection in the three meso-regions of the State of Alagoas, Brazil. A total count of 23 towns and 27 meat sheep farms were visited where blood samples were collected in order to perform the indirect immunofluorescence test to evaluate the antibodies presence. Questionnaires exploring the production system and nutritional, sanitary, and reproduction handling were handed out. The prevalence rate was 32.9% and the number of foci was 100%. In the multivariate statistical analysis, there was a significant association for the following variables: age (OR = 4.01; C.I. 2.03–7.94), size of the property (or the farm; OR = 0.48; C.I. 0.26–0.90), semi-intensive rearing system (OR = 3.17; C.I. 1.24–8.13), running water source (OR = 3.13; C.I.–1.66–5.87), and presence of cats (OR = 1.72; C.I. 1.08–2.75). It is concluded that sheep of the three meso-regions of the State of Alagoas are exposed to the infection caused by T. gondii with high prevalence. Control and prophylactic measures must be adopted seeking the improvement of the rearing system and the implantation of health promoting programs in cooperation with sheep farmers in order to elucidate the transmission means of this disease.

Canine and Bovine Neospora caninum Control Sera Examined for Cross-Reactivity Using Neospora caninum and Neospora hughesi Indirect Fluorescent Antibody Tests

Abstract Neospora caninum is a well known protozoan parasite of domestic and wild animals. Neospora hughesi is a closely related protozoan with an unknown life cycle, host range, and infection prevalence. Many serologic surveys of N. caninum have been performed without consideration of potential cross-reactions with N. hughesi, which could confound results. The aim of this study was to investigate whether postexposure sera from animals experimentally infected with N. caninum exhibit significant reactivity differences when tested using N. caninum and N. hughesi Immunofluorescent Antibody Tests (IFAT). Pre- and postinfection serum samples from 10 dogs, 20 calves, and 17 cows were tested by dual IFATs. All pre-exposure samples for N. caninum tested seronegative for both organisms. All postexposure samples that were seropositive for N. caninum were also positive for N. hughesi, although N. hughesi antibody titers were usually 1 dilution lower (P < 0.02). Serologic surveys for N. caninum may be confounded by cross-reacting titers with N. hughesi, but true positive N. caninum antibody titers are greater than, or equal to, cross-reacting N. hughesi antibody titers.

Occurrence of antibodies to Neospora caninum and Toxoplasma gondii in dogs from Pernambuco, Northeast Brazil

A serological survey was carried out to assess the occurrence of anti-Neospora caninum antibodies in dogs from the State of Pernambuco. A total of 625 serum samples of dogs (289 from Paulista, 168 from Amaraji and 168 from Garanhuns) were tested by an immunofluorescence antibody assay for the detection of anti-N. caninum antibodies. A total of 177 (28.3%; IC 95%, 24.9-32.1) samples were positive. The seropositivity rates found in Paulista, Amaraji and Garanhuns were 26% (IC 95%, 21-31.4), 26.2% (IC 95%, 19.7-33.5) and 34.5% (IC 95%, 27.4-42.2), respectively. Of the 177 serum samples positive to anti-N. caninum antibodies, 170 were additionally tested for the presence of anti-Toxoplasma gondii antibodies and out of these 57.6% (IC 95%, 49.8-65.2) were positive. The results indicate that dogs from Amaraji, Paulista and Garanhuns are exposed to both N. caninum and T. gondii infections. The presence of dogs infected by N. caninum in Pernambuco represents a potential risk factor for the occurrence of outbreaks of abortion in cattle and small ruminants in this state. This study is the largest serological survey on the presence of anti-N. caninum antibodies in dogs carried out in Brazil and reports for the first time the exposure to N. caninum and T. gondii in dogs from Pernambuco.

Freqüência de anticorpos IgG anti-Toxoplasma gondii em soros de cães errantes da cidade de Salvador-Bahia, Brasil

The Toxoplasma gondii is an obligate intestinal intracelular coccidian protozoan of felidae, being considered cosmopolitan, discovered in 1908 by Nicolle & Manceaux. The first report in the canine species happened in 1910, in Italy and, in Brazil in 1911. With the objective of evaluating the frequency of this parasite in the population of dogs of the city of Salvador-Ba, 225 samples of blood from animals of 10 sanitary districts were collected for the accomplishment of the reaction of indirect fluorescent antibody (IFA) for detection of IgG antibodies against-Toxoplasma gondii, using the strain AS28. 143 positive samples were detected, representing a frequency of 63.55.00%. The frequencies for sanitary districts were distributed as follow: Itapagipe 33.33%; Sao Caetano/Valeria 46.15.00%; Brotas 42.11%; Barra/Rio Vermelho 64.28%; Boca do Rio 80.00%; Itapua 65.38%; Cabula/Beiru 80.64%; Pau da Lima 73.91%; Cajazeiras 64.70.00% e Suburbio Ferroviario 73.33%. The serum titers found ranged from 1:16 to 1:16384, being 1:16 (28.67%), 1:64 (44.76%), 1:256 (21.68%), 1:1024 (4.20%) and 1:16384 (0.70.00%). Of the 123 males and 102 females, 67.48% and 58.82% were positive, respectively. Concerned to the age, of the 198 adults and 27 youngs, 70,20% and 14,80% were positive, respectively. The age and sanitary districts presented significant statistical result (p<0,05).