Detection of antigen 85B protein level produced by Mycobacterium tuberculosis for development of first-line anti-tuberculosis drugs susceptibility

Abstract

Tuberculosis (TB) is a worldwide public health problem and the leading cause of death in the world in patients’ infectious disease. Mycobacterium tuberculosis is the major cause of TB in humans. Patients with tuberculosis require rapid, accurate and consistent antibiotic therapy to be treated effectively. It has been reported that the antigen 85 (Ag85) complex proteins can be secreted incessantly by live mycobacteria during mycobacteria culture in liquid media; thus, Ag85 could be targeted for the detection of M. tuberculosis and mycobacterial drug susceptibility. The aims of this study were: 1) to determine the levels of Ag85B protein produced by M. tuberculosis under conditions of treatment with and without various anti-tuberculosis drugs; and 2) to develop an Ag85B assessment technique for drug susceptibility of M. tuberculosis to isoniazid, rifampicin and ethambutol. A sandwich ELISA assay using anti-Ag85B monoclonal antibody was developed to detect Ag85B protein and determine the susceptibility of mycobacterial to anti-TB drugs. The result shows that an increase in Ag85B protein was detected in M. tuberculosis growth. Conversely, in the presence of anti-TB drugs, if M. tuberculosis is inhibited or killed by the drug, The increase in Ag85B protein is not observed. Therefore, anti-tuberculosis drugs susceptibility can be detected using the difference in the level of Ag85B between two conditions, testing M. tuberculosis with and without antituberculosis drug. This novel technique for drug susceptibility testing could report the result of mono-drug resistant M. tuberculosis within 9 days after culturing in liquid media with and without anti-TB drug. The sensitivity and specificity for the detection of monodrug resistant M. tuberculosis is 90% and 100%, respectively. Therefore, it is an innovative new technique developed to determine mycobacterial drug susceptibility by measuring Ag85B released by live mycobacteria. The advantages of this new technique are that it can perform and obtain results in a shorter time than the standard proportion method and can be developed further for use in the medical laboratory.