Article

Improved enzyme-linked immunosorbent assay using C-terminal truncated recombinant antigens of Babesia bovis rhoptry-associated protein-1 for detection of specific antibodies.

National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
Journal of Clinical Microbiology (Impact Factor: 4.07). 05/2004; 42(4):1601-4. DOI: 10.1128/JCM.42.4.1601-1604.2004
Source: PubMed

ABSTRACT An enzyme-linked immunosorbent assay (ELISA) based on a recombinant rhoptry-associated protein-1 (RAP-1) of Babesia bovis has been previously developed, but it was imperfect because some cross-reactions were still present in Babesia bigemina-infected bovine sera. To improve its accuracy for the specific detection of the antibodies to B. bovis, we constructed three C-terminal truncated recombinant antigens of the RAP-1-rCT1 (amino acids [aa] 301 to 408), rCT2 (aa 388 to 490), and rCT3 (aa 466 to 565)-by using a baculovirus expression system and evaluated their diagnostic potentials using ELISA. rCT1 and rCT2 were better diagnostic antigens in their sensitivities and diagnostic efficiencies than rCT3, although none of the recombinant antigens showed any cross-reactivity to B. bigemina-infected bovine sera. These results confirmed that the N-terminal 300-aa region caused cross-reactivity of the entire RAP-1 antigen, and the C-terminal truncated recombinant antigens were shown to be useful reagents for species-specific serodiagnosis.

0 Bookmarks
 · 
52 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Incidence of bovine babesiosis in Portugal is currently unknown. In this study, a first survey of Babesia bovis and Babesia bigemina infection in cattle was carried out using blood samples from 406 clinically healthy individuals from different districts from Central and Southern regions of Portugal and analyzed by indirect enzyme linked immunosorbent assay (iELISA) and nested polymerase chain reaction (nPCR). Overall, serological testing revealed that 79% and 52% of cattle were positive for B. bovis and B. bigemina antibodies, respectively, whereas nPCR testing detected 71% and 34% cattle infected with B. bovis and B. bigemina protozoan, respectively. This is the first report of the prevalence of B. bovis and B. bigemina in cattle obtained by serological and DNA analysis studies in Central and Southern regions of Portugal. These data suggests high incidence of Babesia sp. infection in Portugal and can be used for designing adequate control programs.
    Veterinary Parasitology 12/2009; · 2.38 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The present study was conducted to demonstrate the epidemiological distribution of bovine babesiosis in the northern regions of Thailand. A total of 700 serum samples of dairy cows in the northern provinces (Chiang Rai, Chiang Mai, Lumpang, and Mae Hong Sorn) were tested for antibodies against Babesia bovis and B. bigemina. Species-specific enzyme-linked (rRAP-1/CTs) were performed. According to the results, 517 (73.8%) and 484 (69.1%) were positive for B. bovis and B. bigemina, respectively. In addition, 370 (52.9%) were positive for mixed infections by both ELISAs. On the other hand, all samples were also examined by the indirect fluorescent-antibody test (IFAT) with B. bovis- and B. bigemina-infected blood smears. According to the IFAT, 482 (68.8%) and 531 (75.8%) were positive for these infections, respectively. The overall concordances between the ELISA and IFAT techniques were 93.6% and 90.7% for B. bovis and B. bigemina infections, respectively. These results indicated that babesia infections are widespread in the northern parts of Thailand. To our knowledge, this is the first report describing the epidemiology of Babesia infections using rRAP-1/CT-based ELISAs in these areas.
    Veterinary Parasitology 03/2010; 170(3-4):193-6. · 2.38 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Five Babesia bovis recombinant proteins, including merozoite surface antigen 2c (BbMSA-2c), C-terminal rhoptry-associated protein 1 (BbRAP-1/CT), truncated thrombospondin-related anonymous protein (BbTRAP-T), spherical body protein 1 (BbSBP-1), and spherical body protein 4 (BbSBP-4), were evaluated as diagnostic antigens to detect the infection in cattle. The recombinant proteins were highly antigenic when tested with experimentally B. bovis-infected bovine serum in Western blot analysis. Furthermore, five antisera that had been raised against each of the recombinant proteins reacted specifically with the corresponding authentic protein, as determined in Western blot analysis. Next, enzyme-linked immunosorbent assays (ELISAs) using these recombinant proteins were evaluated for diagnostic use, and the sensitivity and specificity of each protein were demonstrated with a series of serum samples from experimentally B. bovis-infected cattle. Furthermore, a total of 669 field serum samples collected from cattle in regions of B. bovis endemicity in seven countries were tested with the ELISAs, and the results were compared to those of an indirect fluorescent antibody test (IFAT), as a reference. Among five recombinant antigens, recombinant BbSBP-4 (rBbSBP-4) had the highest concordance rate (85.3%) and kappa value (0.705), indicating its reliability in the detection of specific antibodies to B. bovis in cattle, even in different geographical regions. Overall, we have successfully developed an ELISA based on rBbSBP-4 as a new serological antigen for a practical and sensitive test which will be applicable for epidemiologic survey and control programs in the future.
    Clinical and vaccine Immunology: CVI 02/2011; 18(2):337-42. · 2.60 Impact Factor

Full-text

View
0 Downloads
Available from