Submitted to: Journal of Clinical Microbiology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 7/1/2005
Publication Date: 9/1/2005
Citation: Koo, H.C., Park, Y.H., Ahn, J., Waters, W.R., Palmer, M.V., Hamilton, M.J., Barrington, G., Abdelaziz, M.A., Cho, S., Shin, S. 2005. Use of rMPB70 Protein and ESAT-6 Peptide as Antigens for Comparison of the Enzyme-linked Immunosorbent, Immunochromatographic, and Latex Bead Agglutination Assays for Serodiagnosis of Bovine Tuberculosis. Journal of Clinical Microbiology. 43(9):4498-4506. Interpretive Summary: Despite highly successful eradication efforts in several countries, tuberculosis of cattle remains a serious health concern worldwide. Recent outbreaks of tuberculosis in Michigan, California, Texas, and New Mexico demonstrate that the disease is far from eliminated from the United States. Improved techniques are needed for detection of infected cattle. In this study, three new test were developed, tested, and compared for the specific diagnosis of tuberculosis infection of cattle. Responses by cattle infected with the tubercle bacillus were distinguishable from responses by cattle infected with closely related bacteria. Knowledge obtained from this study will enable more accurate detection of cattle with tuberculosis, thereby, enhancing the tuberculosis eradication program.
Technical Abstract: ABSTRACT: Current assays used to detect Mycobacterium bovis (Mbv) and/or M. avium subsp. paratuberculosis (Mptb) infection of cattle or deer lack sensitivity and specificity or are impractical for rapid field diagnostic applications. Serologic assays using Mbv- or Mptb-specific antigens may be used to overcome these limitations. To test this strategy, a synthetic peptide derived from Mbv early secretory antigenic target-6 (ESAT6-p) and a recombinant major secreted immunogenic protein of Mbv (rMPB70) were used in an enzyme-linked immunoabsorbent assay (EIA), an immunochromatographic assay (ICGA, dipstick), and a latex bead agglutination assay (LBAA). Sera from non-infected, Mbv-infected (natural and experimental routes), and Mptb-infected (natural and experimental routes) were evaluated. Receiver operating characteristic (ROC) analysis of EIA results and comparison with bacterial culture established suitable cut-off values for assessing sensitivity and specificity. Assays with ESAT6-p had sensitivity of 94.3 and 96.1 % and specificity of 94.7, 100 %, and kappa (K) values of 0.93, 0.94 with the EIA and LBAA, respectively. Assays with rMPB70 had a sensitivity of 91.4, 75.3 and 81.8 % and specificity of 96.1, 100, 100 % and K values of 0.89, 0.84, 0.8 for EIA, ICGA and LBAA, respectively. Examination of serial samples of sera collected from experimentally infected cattle and deer revealed ESAT-6p specific responses developed early after infection whereas responses to rMPB70 developed later in the course of disease. Advantages of the LBAA and ICGA assays for initial testing is that results can be obtained in two to three hours by LBAA or 20 minutes by ICGA under field conditions and both assays can be used with multiple species as species specific secondary antibodies are not required.