Differential delayed-type
hypersensitivity skin testing with
tuberculin purified
protein derivatives from Mycobacterium bovis and M. avium is the standard for diagnosing
bovine tuberculosis. However, improved tests based on defined, specific
antigens are urgently needed. In the present study, a combination of bioinformatics, molecular biology, and bovine models of
infection were used to screen mycobacterial
proteins for their potential as diagnostic
reagents which could be used in a whole-blood assay for diagnosis of
tuberculosis. Initial screening of 28
proteins selected in silico and expressed as recombinants in Escherichia coli indicated that CFP-10, ESAT-6, TB27.4, TB16.2, TB15.8, and TB10.4 induced strong
gamma interferon responses in experimentally infected cattle. A more thorough investigation over time in two groups of animals infected with a high (10(6) CFU) and a low (10(4) CFU) dose of M. bovis revealed that, for both groups, the strength of the in vitro response to individual
antigens varied greatly over time. However, combining the results for ESAT-6, CFP-10, and TB27.4, possibly supplemented with TB10.4, gave sensitivities at different
infection stages close to those obtained with M. bovis purified
protein derivative. Importantly, while responsiveness to ESAT-6 and CFP-10 correlated strongly for individual samples, the same was not the case for ESAT-6 and TB27.4 responsiveness. The results suggest that combinations of specific
antigens such as these have great potential in development of optimized diagnostic systems for
bovine tuberculosis.