In silico predicted conserved B-cell epitopes in the merozoite surface antigen-2 family of B. bovis are neutralization sensitive.

The merozoite surface antigens MSA-2 of Babesia bovis constitute a family of polymorphic GPI-anchored glycoproteins located at the parasite cell surface, that contain neutralization-sensitive B-cell epitopes. These are therefore putative vaccine candidates for bovine babesiosis. It was previously shown that (i) the MSA-2 antigens of the biologically cloned Mo7 strain are encoded by four tandemly organized genes: msa-2a(1), a(2), b and c, and (ii) at least one allele of each of these genes is present in the Argentine R1A strain with a moderate degree of polymorphism. The present work was aimed at defining neutralization-sensitive B-cell epitopes in the MSA-2 family, that are conserved among different B. bovis geographical isolates. To this end, msa-2a, b and c alleles from different isolates from Argentina, USA and Mexico were amplified by PCR, cloned and sequenced. Bioinformatic analysis by ClustalW alignments and B-cell epitope prediction algorithms performed on these sequences allowed the identification of several regions containing putative conserved B-cell epitopes. Four peptides representing these regions: (KDYKTMVKFCN from msa-2a(1); YYKKHIS, from msa-2b; and THDALKAVKQLIKT and ELLKLLIEA from msa-2c) were chemically synthesized, conjugated to keyhole limpet hemocyanin and used to inoculate mice to obtain immune sera. Anti-peptide antibodies recognized B. bovis merozoite extracts in all cases in ELISA tests. In addition, these sera reacted with the surface of merozoites of an Argentine and a Mexican B. bovis strains in immunofluorescence assays, and sera against two of the selected peptides inhibited invasion of erythrocytes by in vitro cultured merozoites. Taken together, the results show that the peptide sequences selected by bioinformatic analysis represent expressed and geographically conserved B. bovis B-cell epitopes that might be strong candidates for development of subunit vaccines.

Incidence and isolation of bluetongue virus infection in cattle of the Santo Tomé Department, Corrientes Province, Argentina.

Sentinel herds were monitored for the detection of bluetongue (BT)-specific antibodies and virus over two periods, namely: June 1999 to August 2000 and September 2000 to April 2001. Herds were located in Santo Tomé (Herds 1 and 2) where BTV activity was known to occur. From June 1999 to August 2000, the cumulative incidence (CI) of bluetongue virus (BTV) infection was 0% and 35% in Herds 1 and 2, respectively. In the second period, the CI of BTV infection was 10% and 97% in Herds 1 and 2, respectively. The virus was isolated from red blood cells of animals that seroconverted and was identified as serotype 4. Averages of the monthly maximal temperatures were always above 19 degrees C. However, averages of the monthly median temperatures were below 19 degrees C and averages of the monthly minimal temperatures were below 15 degrees C from May 2000 to August 2000. There was no viral activity detected at that time. Culicoides insignis was identified as the predominant potential vector species (99%) trapped near sentinel herds. Although clinical disease has never been reported in Argentina, viral activity was detected and the virus has been isolated in sentinel herds.

Serological evidence of Neospora caninum infections in beef bulls in six counties of the Corrientes province, Argentina.

The purpose of the present work was to identify Neospora caninum infections in beef bulls belonging to 19 herds from six counties located in the Corrientes province, Argentina. The presence of antibodies to N. caninum was evaluated in 305 serum samples of bulls (Bos taurus and Bos indicus). Age and breed were recorded. An indirect fluorescent antibody test was used to determine specific antibodies. The number of bulls with natural Neospora-infection was 15 of 305 (4.9%). No association between serologic status and breed (odds ratio (OR), 0.53; 95% CI, 0.18-1.53) was found. Neospora-infected beef bulls were identified in the present work. The bull role in bovine neosporosis and the risk of horizontal transmission for cows should be investigated.

[Bluetongue: isolation and characterization of the virus and identification of vectors in northeastern Argentina]. / Lengua Azul: Aislamiento y caracterización del virus e identificación de vectores en el noreste argentino.

To establish if BTV was circulating in Argentina, 94 bovines from the Santo Tomé and Ituzaingó Departments of Corrientes Province were sampled every 30-60 days during 14 months. Red blood cells from those animals that showed seroconvertion with a c-ELISA were processed for virus isolation by inoculation in embryonated chicken eggs and cell cultures. Cells with CPE were positive by direct and indirect immunofluorescence with BTV specific reagents. These samples examined by electron microscopy showed virus particles with BTV morphological characteristics. Blood samples and tissue culture supernantants were positive by RT-PCR technique with primers corresponding to the segment 3 of the BTV genome. Haematophagous insects were captured in one farm using light traps and Culicoides insignis Lutz was the predominant species detected. This is the first isolation of BTV in Argentina from northeastern bovines without any disease symptom.

Lengua Azul: Aislamiento y caracterización del virus e identificación de vectores en el noreste argentino / Bluetongue: isolation and characterization of the virus and identification of vectors in northeastern Argentina

To establish if BTV was circulating in Argentina, 94 bovines from the Santo TomÚ and Ituzaingó Departments of Corrientes Province were sampled every 30-60 days during 14 months. Red blood cells from those animals that showed seroconvertion with a c-ELISA were processed for virus isolation by inoculation in embryonated chicken eggs and cell cultures. Cells with CPE were positive by direct and indirect immunofluorescence with BTV specific reagents. These samples examined by electron microscopy showed virus particles with BTV morphological characteristics. Blood samples and tissue culture supernantants were positive by RT-PCR technique with primers corresponding to the segment 3 of the BTV genome. Haematophagous insects were captured in one farm using light traps and Culicoides insignis Lutz was the predominant species detected. This is the first isolation of BTV in Argentina from northeastern bovines without any disease symptom.

Detection of virus infection-associated antigen and 3D antibodies in cattle vaccinated against foot and mouth disease.

The analysis of sera obtained from animals vaccinated or revaccinated with inactivated vaccines against foot and mouth disease (FMD) virus showed that these vaccines induced antibodies against the virus infection-associated (VIA) antigen, detectable by agar gel immunodiffusion (AGID). The present study evaluates the antibody response to protein 3D and the VIA antigen (VIAA) of FMD virus induced by different vaccines in a group of 51 calves. This response was detected using AGID and a liquid-phase blocking sandwich enzyme-linked immunosorbent assay (ELISA) for anti-3D antibodies (ELISA-3D). No anti-VIAA or anti-3D antibodies were detected after the initial vaccination. Following revaccination, animals giving positive results were detected by both methods. This immune response disappeared 60-120 days post-revaccination (dprv) according to the AGID method, and 90-180 dprv when ELISA-3D was used. Samples of oesophageal-pharyngeal fluid obtained from animals that remained positive for anti-VIAA antibodies at 90-120 dprv gave negative results for viral isolation, indicating that the transitional antibody response induced by the vaccine was due to the presence of non-structural antigens in the vaccine and not to viral infection. These results indicate that the ELISA-3D method could be used as a complementary method for sero-epidemiological studies as an indirect indicator of viral activity, as long as the age and vaccination status of the animals being sampled are taken into consideration.