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Title:   Author  item
                                            Bass, Kristinitem
                                            Nonnecke, Brianitem
                                            Palmer, Mitchellitem
                                            Thacker, Tyleritem
                                            HARDEGGER, R - Prionics Agitem
                                            SCHROEDER, B - Prionics Agitem
                                            RAEBER, A - Prionics Agitem
                                            Waters, Wade
  Submitted to: Clinical and Vaccine Immunology
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 10/9/2013
                                    Publication  Date: 12/1/2013
                                    Publication URL: https://handle.nal.usda.gov/10113/58113Citation: Bass, K.E., Nonnecke, B.J., Palmer, M.V., Thacker, T.C., Hardegger, R., Schroeder, B., Raeber, A.J., Waters, W.R. 2013. Clinical and diagnostic developments of a gamma interferon release assay for use in bovine tuberculosis control programs. Clinical and Vaccine Immunology. 20(12):1827-1835.Interpretive Summary: Bovine tuberculosis (bTB), caused by Mycobacterium bovis, accounts for up to 10% of human TB cases in developing countries and is increasing in cattle in the US and UK. Control of bTB is hindered by the presence of numerous wildlife reservoirs such as white-tailed deer, European badgers, and brush-tailed possums. Reasons for the failure to eradicate the disease are multi-factorial. Limitations in the sensitivity and specificity of diagnostic tests are factors contributing to the persistence of bTB. Test interpretation and accuracy are confounded by variations in the frequency of tuberculin testing, ambiguities in test result interpretation, and the use of varying tuberculin preparations The BovigamTM assay is approved for use within the United States as a supplemental test for bovine tuberculosis. This assay relies on a specific response to the tuberculosis agent. In the present study, reagents used in the test and test applications were evaluated to improve test specificity and sensitivity. Knowledge obtained from this study may lead to an improved test for tuberculosis, thereby, advancing the national tuberculosis eradication program.
                                Technical Abstract:  Currently the Bovigam assay is used as an official supplemental test within the bovine tuberculosis eradication program. This assay measures interferon-gamma (IFN-gamma) produced by lymphocytes in response to specific antigens. The objectives of the present study were to evaluate two Mycobacterium bovis specific peptide cocktails, purified protein derivatives (PPDs) from two sources, liquid and lyophilized antigen preparations, and a second generation IFN-gamma release assay (Bovigam, B2G, Prionics AG). Three strains of M. bovis were used for experimental challenge: M. bovis 95-1315, M. bovis Ravenel, and M. bovis 10-7428. Additionally, samples from tuberculosis-affected herds (i.e. natural infection) were evaluated. Robust responses to both peptide cocktails HP (PC-HP) and ESAT6/CFP10 (PC-EC), as well as PPDs were elicited as early as three weeks after challenge. Only minor differences in responses to Commonwealth Serum Laboratories (CSL) and Lelystad PPDs were detected with samples from experimentally infected animals. For instance, responses to Lelystad M. avium derived PPD (PPDa) exceeded respective response to CSL PPDa in M. bovis Ravenel infected and control animals. However, 1:4 dilution of stimulated plasma demonstrated greater separation of PPDb from PPDa responses (i.e., PPDb – PPDa) with use of Lelystad PPDs, suggesting that Lelystad PPDs provide greater diagnostic sensitivity than CSL PPDs. With samples from tuberculosis-affected herds, responses to Lelystad PPDs generally exceeded respective responses to CSL PPDs. Responses to lyophilized and liquid antigen preparations did not differ. Responses detected with first (B1G) and second (B2G) generation IFN-gamma release assay kits (Bovigam) did not differ throughout the study. In conclusion, antigens may be stored in a lyophilized state without a loss in potency, PC-HP and PC-EC are dependable biomarkers for aiding diagnosis of bovine tuberculosis, and second generation Bovigam kits have sensitivity/specificity comparable to current kits.


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