Ocorrência de anticorpos anti-Neospora spp. em jumentos (Equus asinus) no estado de Sucre - Colômbia / Occurrence of anti-Neospora spp. Antibodies in donkeys (Equus asinus) in the state of Sucre - Colombia

A neosporose tem grande importância em bovinos, e sua patogênese neste hospedeiro, em termos gerais, está esclarecida, porém, em equídeos é pobremente conhecida. Na Colômbia ainda não foram reportados anticorpos Anti-Neospora spp. em equídeos, sendo assim objetivou-se no presente estudo avaliar a ocorrência do parasito em jumentos (Equus asinus) de fazendas que apresentavam risco da doença nestes animais. Foram utilizados 56 animais no estado de Sucre (Colômbia), escolhidos aleatoriamente dentro das fazendas selecionadas. Utilizou-se um peptídeo recombinante originado de Neospora caninum (NcGRA1) para o diagnóstico por Dot-ELISA, e o soro foi diluído em 1:200. Este estudo reporta, pela primeira vez no estado de Sucre e na Colômbia, a presença de anticorpos anti-Neospora spp. na espécie Equus asinus, com uma ocorrência de 19,7% (11/56) dos animais amostrados...

The neosporosis has great importance in cattle, and its pathogenesis in this host has been generally clarified, however, in horses, neosporosis is poorly known, and in Colombia anti-Neospora spp antibodies have not been reported. Therefore, the main objective in the present study was to evaluate the occurrence of this parasite in donkeys (Equus asinus) from farms that presented a risk of disease in these animals, as well as no health plan for them. Were used 56 animals randomly chosen inside selected farms in the state of Sucre (Colombia). A recombinant peptide originated from Neospora caninum (NcGRA1) was used for the diagnosis with Dot-ELISA and serum was diluted 1:200. This study is the first to report the presence of anti-Neospora spp. in donkeys (Equus asinus) in the state of Sucre, and in Colombia. The occurrence was in 19.7% of the animals sampled (11/56)...

Immune response of bovines stimulated by synthetic vaccine SBm7462 against Rhipicephalus (Boophilus) microplus.

Ten-month-old calves Bos taurus taurus were immunized with three doses of SBm7462 with saponin as an adjuvant at 30-day intervals and were evaluated for IgG isotypes, phenotype circulating lymphocytes and changes in the lymph nodes (LN). SBm7462 stimulated the production of predominantly IgG1-isotype IgG antibodies. The lymph nodes exhibited activation at the seventh day after the first immunization, with areas of paracortical and interfollicular hyperplasia and the early formation of germinal centers (GC). Fifteen days after the first immunization, the GC exhibited compartmentalization of cellular populations, a light zone (LZ), a dark zone (DZ) and a mantle. At the same time, hyperplasia of the medullary cords was observed with cells associating with DC cells. Seven days after the first immunization, apoptosis in the DZ and in the paracortical region became evident. By day 15, there was an increase in the medullary cords, which became more numerous at days 35 and 42. PAP-positive cells were found in the paracortical region, medullary cords and GC 7 days after the first immunization. At day 35, there were further strongly PAP-positive cells in the medullary cords. By comparison, none of these changes were observed in the lymph nodes of control groups at any of the days analyzed. The number of CD21(+) lymphocytes increased in the immunized groups after the first inoculation, with a maximum number observed at 15 and 10 days after the first and third immunizations, respectively. Compared to pre-immunization counts, the percentage of WC1(+) gammadelta T-lymphocytes displayed more variation, increasing 5 days after the second immunization but decreasing over the following days. According to the results, the synthetic anti Rhipicephalus microplus vaccine elicits a complete immune response being T-dependant.

Synthetic vaccine (SBm7462) against the cattle tick Rhipicephalus (Boophilus) microplus: preservation of immunogenic determinants in different strains from South America.

The synthetic vaccine SBm7462 is based on three immunogenic epitopes (4822, 4823 and 4824) contained within protein Bm86 derived from the Australian Yeerongpilly strain of the tick Rhipicephalus (Boophilus) microplus. Twenty strains of the tick originating from Brazil, Argentina, Colombia and Uruguay were analysed in order to identify differences compared with sequences present in components of vaccine SBm7462. For each parasite population, three cDNA fragments containing the nucleotides coding for the epitopes 4822, 4824 and 4823 were sequenced, and the amino acid sequences were deduced and compared with those of the homologous bm86 gene. The results indicate that the epitope sequences of vaccine SBm7462 are conserved in the South American populations of the tick. The conservation of such sequences is very important for the immunological response of different populations of R. (B.) microplus.

Use of biodegradable PLGA microspheres as a slow release delivery system for the Boophilus microplus synthetic vaccine SBm7462.

The synthetic anti-Boophilus microplus vaccine SBm7462 derived from the tick intestinal protein, Bm86, induced a protective immune response when emulsified in saponin and used in cattle. Using a mice model, and with the objective of improving the vaccine by continual peptide release, it was encapsulated in PLGA 50:50 microspheres and inoculated in BALB/c mice to assess the immunological response by detection of anti-peptide IgGs. Comparative studies were made with the peptide emulsified in saponin and with another synthetic vaccine, and the microsphere/peptide was characterized for efficiency of encapsulation, in vitro release profile, morphology, size, peptide integrity after encapsulation and stability in different pHs. The findings showed that saponin enhances a better immune response from SBm7462 and that the PLGA 50:50 microspheres are suitable for use with this peptide.

Immunization of cattle with synthetic peptides derived from the Boophilus microplus gut protein (Bm86).

Three synthetic peptides (SBm4912, SBm7462 and SBm19733), derived from the Bm86 glycoprotein from Boophilus microplus gut, were constructed and used to immunize cattle from a tick-free area. The immunized animals received three subcutaneous doses of the peptides, with saponin as adjuvant, at 30-day intervals. The immune response was evaluated by IgG elicited against the peptides by the detection of anti-Bm86 specific antibodies in situ and by Western blotting analysis. After tick challenge, reduction in the number, weight and oviposition capacity of engorged females was observed in the tick population that had fed on immunized animals. The results pointed a high efficacy (81.05%) for the SBm7462 synthetic peptide in relation to the others (p<0.01), demonstrating the efficiency of the immune response elicited by synthetic peptides to control the cattle tick B. microplus.

Ultrastructure of Babesia bigemina gametes obtained in "in vitro" erythrocyte cultures.

Propagation of Babesia bigemina in "in vitro" erythrocyte cultures resulted in the appearance of the sexual stage of the parasite, characterized by cytoplasmic projections and microtubules. These morphologically distinct parasites were found to adhere to each other, with later fusion of their membranes. No nuclear fusion was observed. The ultrastructure of the sexual stages was described.

Toxoplasmosis in naturally infected deer from Brazil.

Serum samples from 107 cervids were examined for Toxoplasma gondii antibodies using indirect hemagglutination (IHA), indirect immunofluorescence (IFA), enzyme linked immunosorbent assay (ELISA) and Dot-ELISA. Samples were obtained from 66 marsh deer (Blastocerus dichotomus) in the State of São Paulo (Brazil) and from 41 pampas deer (Ozotocerus bezoarticus) in the State of Goiás (Brazil). Antibodies to T. gondii were found in 23 (22%) of the deer, with 18 and 5 positive samples, respectively, for B. dichotomus and O. bezoarticus. The highest prevalence of T. gondii antibodies were young adults (32%), following by adults (27%) and fawns (13%). Only one serum sample (8%) from a newborn fawn was positive in the serological tests. The convenience of the Dot-ELISA test is obvious when compared with other serological tests for both laboratory or field surveys, mainly due to its features of practicability and reagent stability.

Exoantigens of an attenuated strain of Babesia bovis used as a vaccine against bovine babesiosis.

Bovine babesiosis caused by Babesia bovis remains a significant constraint to beef and milk cattle production throughout the world. Exoantigens released by the parasites in culture supernatants are a potential source of antigen to induce protective immunity. An attenuated strain of B. bovis from Brazil, catalogued as BbUFV1, was maintained in vitro by the MASP method, and exoantigen-containing supernatant fluids were collected daily to form a pool representing a 72-h culture cycle for preparation of the vaccine. Exoantigen concentration was estimated using a two-site EIA. Three groups of susceptible non-splenectomised male Bos taurus cattle, 14 months old, were used. Group A (vaccinated) received two subcutaneous immunizations with a 21-day interval of B. bovis supernatant, content 6500 EIA units of exoantigens plus 1.5 mg saponin, and Group B (adjuvant control) received two injections of adjuvant alone. Four weeks after the second immunization, Groups A, B and C (control) were challenged intravenously with 10(8) virulent parasites of a heterologous B. bovis strain. The results showed that exoantigens present in in vitro cultures can induce a high degree of protection against virulent heterologous challenge exposure. In Group A only one animal showed discrete parasitaemia; all developed a fever and slight decreases in PCV, with a rapid return to normal values. One animal of Group B died; the survivors showed fever, anaemia and parasitaemia. All animals of Group C died between 7 and 13 days after challenge. Vaccination elicited both humoral and cell-mediated immune responses. In Group A, after the challenge, the maximum antibody titer was 12,800. When vaccinated, cattle were tested at the moment of challenge for B. bovis-specific cell-mediated immunity by the monocytemigration inhibition test. A mean inhibition index of 60 +/- 0.33 was observed. Preliminary Western blot analysis of the immunogen revealed at least four proteins of molecular weight ranging between 30 and 160 kDa.

Initial interaction between Haemonchus Placei infective larvae and different Arthrobotrys isolates

Um estudo da interaçäo inicial entre larvas infectantes de Haemonchus placei e diferentes isolados do fungo predador Arthrobotrys das espécies A. conoides, A. robusta e A. musiformis foi realizado pelo uso da microscopia eletrônica de transmissäo. A captura dos nematóides por estes fungos ocorreu em estruturas do complexo hifal (Armadilhas) e foi efetuada pela cobertura adesiva presente sobre estas armadilhas. Esta similar interaçäo foi observada entre os isolados B (A. robusta) e (A. robusta), 3 (A. musiformis) e larvas infectantes de H. placei as 9 horas depois da adiçäo dos nematóidies. As 6 h de interaçäo esta adesäo foi somente observada pela utilizaçäo do isolado E. Isto poderia explicar a variaçäo existente na capacidade predatória de diferentes isolados

Antigenic profile of a pure isolate of Anaplasma marginale of Brazilian origin, using a western blot technique.

Anaplasma marginale initial bodies of the Brazilian isolate AUFV1 were purified from infected erythrocytes using a combination of lysis, ultrasonic disruption and differential centrifugation. Initial bodies were solubilised with a buffer containing protease inhibitors and non-ionic detergents. Immunochemical analysis by the Western blot technique revealed at least five proteins with apparent molecular weights (MW) of 105, 100, 97, 87 and 38 kDa when homologous sera were used as primary antibodies. Sera from cattle from Mato Grosso do Sul State in Brazil revealed five proteins of 105, 100, 87, 38 and 25 kDa; other heterologous sera obtained from Illinois, USA, bound to four antigens with MW of 105, 100, 87 and 38 kDa, the latter being stronger and broader than the others. No bands were observed in the non-infected erythrocyte control when the different A. marginale sera or antibodies against Babesia bovis and Babesia bigemina were used. Antibodies from cattle infected with the A. marginale isolates bound to proteins of 105, 100, 87 and 38 kDa, indicating that there are at least four peptides common to the isolates. The major surface proteins, designated MSP-1, MSP-2 and MSP-3, are also present in the Brazilian isolate AUFV1 and the sample from the National Research Centre of Beef Cattle in Mato Grosso do Sul State. One practical consideration of this study is the possibility of cross-protection between different Anaplasma isolates including some from Brazil.

Babesia bovis: infectivity of an attenuated strain of Brazilian origin for the tick vector, Boophilus microplus.

In order to verify the infectivity and transovarian transmission of one strain of Babesia bovis of Brazilian origin, attenuated by repeated passages in splenectomized calves to the biological tick vector, Boophilus microplus, an inoculation of that strain was performed in hemoparasite-free calves infested with hemoparasite-free ticks. Subsequently, engorged female ticks were collected to examine hemolymph and progeny for presence of developing stages of Babesia bovis. Using an optical microscope, intrahemocytic merozoites were observed in female hemolymph on the second and fourth days, respectively, after ticks dropped from calves. By histological observation of F1 larvae, after more than 24 h of adherence on calves, sporokinetes and small merozoites were present in the samples of hemolymph. These results confirmed the maintenance of infectivity by the vaccine strain attenuated after 26 passages for tick Boophilus microplus.

Epidemiology of bovine anaplasmosis and babesiosis in Latin America and the Caribbean.

The various parameters which interact in the epizootiology of babesiosis and anaplasmosis and which it is necessary to obtain in a survey are analysed and the two diseases compared. Where no data are available from Latin America and the Caribbean, the references from Australia and the United States of America are discussed. The two principal objectives of a study on the epidemiology of babesiosis and anaplasmosis are: to determine the risk of occurrence to understand the relevant factors leading to outbreaks. The first objective is relatively simple to achieve for both diseases by calculating the animal inoculation rate (h) at a determined age and the critical value of h for enzootic stability. The second objective requires exhaustive and complex studies. For babesiosis epidemiology, studies of the tick infection rate, cattle infestation rate, genetic composition of cattle, acaricides, stocking rate, climatic data and other factors are necessary. Anaplasmosis epidemiology cannot be satisfactorily explained by considering it purely as a tick-borne disease. Difficulties to be overcome include the great diversity of haematophagous Diptera in Latin America, and the scarcity of data on the capacity of these insects as anaplasmosis vectors and their feed preferences on cattle.

Epidemiology of bovine anaplasmosis and babesiosis in Latin America and the Caribbean

The various parameters which interact in the epizootiology of babesiosis and anaplasmosis and which it is necessary to obtain in a survey are analysed and the two diseases compared. Where no data are available from Latin America and the Caribbean, the references from Australia and the United States of America are discussed. The two principal objectives of a study on the epidemiology of babesiosis and anaplasmosis are: to determine the risk of occurrence to understand the relevant factors leading to outbreaks. The first objective is relatively simple to achieve for both diseases by calculating the animal inoculation rate (h) at a determined age and the critical value of h for enzootic stability. The second objective requires exhaustive and complex studies. For babesiosis epidemiology, studies of the tick infection rate, cattle infestation rate, genetic composition of cattle, acaricides, stocking rate, climatic data and other factors are necessary. Anaplasmosis epidemiology cannot be satisfactorily explained by considering it purely as a tick-borne disease. Difficulties to be overcome include the great diversity of haematophagous Diptera in Latin America, and the scarcity of data on the capacity of these insects as anaplasmosis vectors and their feed preferences on cattle. (AU)

Description of lesions in cattle in a natural outbreak of Babesia bovis infection in Brazil.

This paper describes the pathologic features of a natural infection by Babesia bovis (B. argentina) in Brazil. Microscopic examination of cerebrum, midbrain, cerebellum, liver, kidney, heart and spleen of five fatal cases revealed variable degrees of congestion, particularly in the brain, liver and kidney. The packing of erythrocytes, the majority of which were parasitized, was most marked in the capillaries of brain, kidney and less in liver. Lymphocytic glomerulonephritis was observed. Variable degrees of fatty degeneration were noted in the liver, distention of hepatic canaliculi and biliary retention was marked. A strong activation of the mononuclear phagocytic system was evident in all the subjects studied.

Susceptibility of Brazilian samples of Boophilus microplus to organophosphorus acaricides.

This paper describes the susceptibility of various samples of Boophilus microplus from the southern region of Minas Gerais State in Brazil to the 4 commonest organophosphorus acaricides employed in the area. All samples showed some degree of resistance to each of the compounds used. The resistance factor varied from 1.42 to 132.90, being less in the case of dicrotophos fenthion than with chlorpyrifos and coumaphos.

Epidemiology of cattle anaplasmosis in Colombia: I. prevalence and distribution of agglutinating antibodies.

A survey to detect antibodies against Anaplasma marginale, using the Card Test, was conducted on serum samples collected from cattle of Cordoba Department and north-eastern region of the Department of Antioquia in the Republic of Colombia. The samples were from native cattle of Spanish descent (Bos taurus), Zebu (Bos indicus) and various Zebu-native crosses. The age range was between three and eight year olds. The results revealed that 90.3% of the samples were positive. All breeds showed a high prevalence of antibodies.