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Title: Comparative evaluation of immunofluorescent antibody and new immunoblot tests for the specific detection of antibodies against Besnoitia besnoiti tachyzoites and bradyzoites in bovine sera

Author
item SCHARES, G - Institute For Animal Health - Germany
item BASSO, W - National University Of Laplata
item MAJZOUB, M - Ludwig-Maximilians University
item ROSTAHER, A - Ludwig-Maximilians University
item SCHARR, J - Ludwig-Maximilians University
item LANGENMAYER, M - Ludwig-Maximilians University
item SELMAIR, J - Ludwig-Maximilians University
item Dubey, Jitender
item CORTES, H - Universidade De Evora
item CONRATHS, F - Wusterhausen
item GOLLNICK, N - Ludwig-Maximilians University

Submitted to: Veterinary Parasitology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/1/2010
Publication Date: 5/10/2010
Citation: Schares, G., Basso, W., Majzoub, M., Rostaher, A., Scharr, J.C., Langenmayer, M.C., Selmair, J., Dubey, J.P., Cortes, H.C., Conraths, F.J., Gollnick, N.S. 2010. Comparative evaluation of immunofluorescent antibody and new immunoblot tests for the specific detection of antibodies against Besnoitia besnoiti tachyzoites and bradyzoites in bovine sera. Veterinary Parasitology. 171:32-40.

Interpretive Summary: Besnoitia species are closely related to Toxoplasma gondii and Neospora caninum that cause abortion and mortality in livestock. These parasites have common antigens that pose problems in diagnosis. In the present paper, researchers describe immunological tests to differentially diagnose these parasitic infections. The results will be of interest to biologists, parasitologists, and veterinarians.

Technical Abstract: Besnoitia besnoiti, an economically important disease in cattle in many countries of Africa and Asia has started to spread in Europe. Serological identification of subclinically infected cattle is important because introduction of these animals into naive herds seems to play a major role in the transmission of the parasite. We report new, simplified immunoblot-based serological tests for the detection of B. besnoiti specific antibodies. Antigens were used under non-reducing conditions in the immunoblots, because reduction of the antigen with ß-mercaptoethanol diminished the antigenicity in both, tachyzoites and bradyzoites. Ten B. besnoiti tachyzoite and ten bradyzoite antigens of 15-45 kDa molecular weight were recognized by B. besnoiti infected cattle, but not or only weakly detected by cattle infected with related protozoan parasites, Neospora caninum, Toxoplasma gondii, Sarcocystis cruzi, Sarcocystis hominis, or Sarcocystis hirsuta. The sensitivity and specificity of B. besnoiti immunoblots were determined with sera from 62 German cattle with clinically confirmed besnoitiosis and 404 sera from unexposed German cattle including 214 sera from animals with a N. caninum-specific antibody response. Using a new scoring system, the highest specificity (100%) and sensitivity (90%) of the immunoblots were observed when reactivity to at least four of the ten selected tachyzoite or bradyzoite antigens was considered as positive. When a cut-off based on this scoring system was applied to both the tachyzoite- and the bradyzoite-based immunoblots, there was an almost perfect agreement with the indirect fluorescent antibody test with a titre of 200 as the positive cut-off. We identified and partially characterized 10 tachyzoite and 10 bradyzoite B. besnoiti antigens which may help to develop new specific and sensitive serological tests based on individual antigens and in the identification of possible vaccine candidates.