Tuberculin Skin Testing Boosts Interferon Gamma Responses to DIVA Reagents in Mycobacterium bovis-Infected Cattle

BCG vaccination sensitises cattle to bovine tuberculin which compromises the use of the current bovine TB surveillance tests. Although the performance of a blood test (that utilises antigens expressed by Mycobacterium bovis but not by BCG) capable of discriminating infected from vaccinated animals (DIVA interferon-gamma test, DIT) has been evaluated in naturally infected TB field reactors, there is a need to perform similar analysis in a BCG vaccinated/ M. bovis infected population. Furthermore, we explored different scenarios under which a DIT may be implemented alongside BCG vaccination: (i) serial testing to resolve potential false positive skin test results; or (ii) standalone test to replace the single intradermal comparative cervical tuberculin skin test (SICCT). Our results demonstrated significantly better relative test sensitivity when the DIT was evaluated in a serial test scenario. Direct comparison of pre and post skin test blood samples revealed that the SICCT test induced significant boosting of the interferon-gamma response in M. bovis -infected animals to both the ESAT-6/CFP-10 and Rv3615c peptide cocktails that comprise the DIT, which persisted for the ESAT-6/CFP-10 reagent for at least 14 days. Importantly, no similar boosting effects were observed in non-infected BCG vaccinates, suggesting that DIVA blood testing after a recent skin test would have minimal impact on test specificity.