Serodiagnostic potential of Mycobacterium avium MAV2054 and MAV5183 proteins

Abstract

The Mycobacterium avium-M. intracellulare complex (MAC) causes a pulmonary disease (PD) similar to tuberculosis (TB). Diagnosis of MAC-PD is complicated and time-consuming. In this study, the serodiagnostic potential of the newly identified MAV2054 and MAV5183 proteins was evaluated in subjects with MAC-PD, pulmonary TB, or latent TB and in noninfected healthy controls (HC), together with HspX and the 38-kDa antigen, well-known serodiagnostic M. tuberculosis antigens. All four antigens evoked significantly higher IgG responses in MAC-PD and active TB than in latent TB and HC subjects. Among the antigens, MAV2054 elicited the highest antibody responses in pulmonary TB and MAC-PD patients. IgG titers against MAV2054 and MAV5183 were significantly higher in MAC-PD than in pulmonary TB subjects. In addition, the levels of IgG against all antigens in the M. intracellulare and fibrocavitary forms were higher than those in the M. avium and nodular bronchiectatic forms, respectively. Based on sensitivity and receiver operator characteristic curve analysis, the best candidates for detection of MAC-PD and pulmonary TB were MAV2054 and the 38-kDa antigen, respectively. In total, 76.0% of MAC-PD and 65.0% of active TB patients were reactive to at least two antigens. In contrast, only 2.8% of HC subjects were reactive with two or more antigens. Our findings suggest that an enzyme-linked immunosorbent assay (ELISA) using the four antigens would be valuable for screening for mycobacterial lung disease, including MAC-PD and pulmonary TB, although it does not provide good discrimination of the disease-causing pathogens.

Fig 1

Identification of strong MAV2054 and MAV5183 seroreactivity in MAC-PD patients. An ammonium sulfate precipitate of M. avium CFP was fractionated by hydrophobic interaction chromatography using phenyl Sepharose. Each of the primary fractions was further fractionated by hydroxyapaptite chromatography. The third fractionation was conducted by DEAE ion-exchange chromatography. IgG titers in all fractions of sera from MAC-PD, pulmonary TB, and HC subjects were determined by ELISA. Two fractions, F15 (A) and F30 (B), were selected, subjected to SDS-PAGE, and stained with Coomassie brilliant blue.

Fig 2

SDS-PAGE analysis of purified recombinant MAV2054 (A) and MAV5183 (B) proteins. The proteins were overexpressed in E. coli, purified by Ni-NTA affinity chromatography, and analyzed by SDS-PAGE with Coomassie brilliant blue staining. Each lane was loaded with 20 μg protein. Bars indicate molecular mass.