Optimization of a molecularly defined tuberculin formulation: recombinant fusion proteins and epitope surgery

Bovine tuberculosis is an infectious livestock disease of global economic and zoonotic importance. Surveillance programs are largely dependent on the skin test, which utilizes tuberculins. These are crude protein extracts of live bacterial cultures, which suffer from a number of limitations regarding their characterization, standardization, production, and performance. We aim to develop a skin test reagent composed of eight defined antigens that could replace the tuberculins. This study describes the refinement of this "molecularly defined tuberculin" (MDT) reagent into a fusion protein formulation comprising all eight antigens. The MDT is well defined, easily standardized, and delivers good test performance in experimentally infected, non-infected cattle and cattle sensitized to Johne's disease. The fusion protein formulation is an important step on the developmental pathway to a registered and marketable product.