Development of an enzyme-linked immunosorbent assay for detection of antibodies to Babesia bigemina in cattle

Abstract

Monoclonal antibodies, directed against a 58-kDa Babesia bigemina merozoite antigen that reacted strongly with immune sera from experimentally and naturally infected cattle in Western blots, were used to develop a competitive-inhibition enzyme-linked immunosorbent assay (ELISA). As based on the testing of 70 antibody-positive sera from experimentally infected cattle and 166 antibody-negative sera collected in non-endemic areas of Australia, the sensitivity and specificity of the ELISA were 95.7% and 97.0%, respectively. In sequential sera collected from six calves during the course of experimental B. bigemina infections the ELISA detected seroconversion at about 10 days post-inoculation. The specificity of the ELISA was not affected by the presence of antibodies to B. bovis, Anaplasma marginale or Theileria buffeli. In 42 sera from cattle experimentally infected with B. bovis but negative for B. bigemina the specificity of the ELISA was 95.2%. The use of a competitive-inhibition ELISA format detecting only antibody directed against a single epitope on the 58-kDa antigen appears to have overcome many of the specificity problems that have plagued serological tests for B. bigemina in the past. The test should be useful for epidemiology studies, particularly in areas where B. bovis and B. bigemina have overlapping distributions.

Avaliação de um ELISA para detecção de anticorpos contra Babesia bigemina em bovinos e sua aplicação em um inquérito sorológico no Brasil

“…The test was also highly sensitive (98.0%) in detecting B. bigemina antibodies. Such results were comparable to those obtained by Molloy et al (1998), using a competitive ELISA based on monoclonal antibodies directed against a 58 kDa B. bigemina merozoite antigen, which showed a sensitivity and specificity of 95.7% and 97.0%, respectively.…”

“…Recently, a highly specific competitive ELISA, based on monoclonal antibodies to a single epitope (p58), was developed for B. bigemina (Molloy et al 1998). In spite of the high specificity of this test, the techniques employed for its development require well-equipped laboratories, often not available in developing countries.…”

Comparison between a soluble antigen-based ELISA and IFAT in detecting antibodies against Babesia canis in dogs

“…The results in this study confirmed that a soluble antigen prepared from infected erythrocytes could be used in a sensitive ELISA to detect antibodies to B. canis. As well as this, other studies with another species of Babesia have used the experimental infection on splenectomized animals to investigate antibodies by ELISA (WEILAND, 1986;MACHADO et al, 1994;ARAÚJO et al, 1998;MOLLOY et al, 1998;KUMAR et al, 2003).…”

Serological survey of Babesia bovis, Babesia bigemina, and Anaplasma marginale antibodies in cattle from the semi-arid region of the state of Bahia, Brazil, by enzyme-linked immunosorbent assays

“…More recently, efforts have been made to improve these serological tests for detection of antibodies against tick-borne diseases agents, especially A. marginale. One of these advances is the use of major surface protein-5 (MSP-5) as recombinant antigen (Knowles et al 1996, Molloy et al 1998, Reynna-Bello et al 1998, although ELISA tests with A. marginale recombinant MSP1a and MSP2 antigens with high sensitivity and specificity have been recently described (Araújo et al 2005, in press). This protein is highly conserved among isolates of A. marginale and elicits a strong humoral response, which can be detected by serological tests for long periods (Knowles et al 1996).…”

“…Various ELISA tests to detect antibodies against Babesia spp. and A. marginale with crude antigens were developed and evaluated in the last years (Molloy et al 1998, Madruga et al 2000a). More recently, efforts have been made to improve these serological tests for detection of antibodies against tick-borne diseases agents, especially A. marginale.…”