Potential of novel Mycobacterium tuberculosis infection phase-dependent antigens in the diagnosis of TB disease in a high burden setting

Abstract

Background: Confirming tuberculosis (TB) disease in suspects in resource limited settings is challenging and calls for the development of more suitable diagnostic tools. Different Mycobacterium tuberculosis (M.tb) infection phase-dependent antigens may be differentially recognized in infected and diseased individuals and therefore useful as diagnostic tools for differentiating between M.tb infection states. In this study, we assessed the diagnostic potential of 118 different M.tb infection phase-dependent antigens in TB patients and household contacts (HHCs) in a high-burden setting.

Methods: Antigens were evaluated using the 7-day whole blood culture technique in 23 pulmonary TB patients and in 19 to 21 HHCs (total n = 101), who were recruited from a high-TB incidence community in Cape Town, South Africa. Interferon-gamma (IFN-γ) levels in culture supernatants were determined by ELISA.

Results: Eight classical TB vaccine candidate antigens, 51 DosR regulon encoded antigens, 23 TB reactivation antigens, 5 TB resuscitation promoting factors (rpfs), 6 starvation and 24 other stress response-associated TB antigens were evaluated in the study. The most promising antigens for ascertaining active TB were the rpfs (Rv0867c, Rv2389c, Rv2450c, Rv1009 and Rv1884c), with Areas under the receiver operating characteristics curves (AUCs) between 0.72 and 0.80. A combination of M.tb specific ESAT-6/CFP-10 fusion protein, Rv2624c and Rv0867c accurately predicted 73% of the TB patients and 80% of the non-TB cases after cross validation.

Conclusions: IFN-γ responses to TB rpfs show promise as TB diagnostic candidates and should be evaluated further for discrimination between M.tb infection states.

Figure 1

IFN-γ levels (pg/ml) elicited by classical TB antigens in 23 TB patients (TB) and 20 household contacts (HHC). Responses in TB cases are indicated by open symbols and those in household contacts by closed symbols. Error bars represent the median. Ag85A/B = Ag85A and Ag85B tested together, HSP = heat shock protein, PHA = Phytohaemagglutinin.

Figure 2

Receiver operating characteristic (ROC) curves showing the accuracies of the most promising DosR encoded antigens in the diagnosis of TB disease. Only ROC curves for antigens that differentiated between the TB cases and household contacts with AUCs above 0.71 are shown. ROC curves for all other DosR encoded antigens are shown in Additional file 9: Figure S1. AUC = Area under the curve.

Figure 3

Receiver operating characteristic curves showing the accuracies of the resuscitation promoting factors in the diagnosis of TB disease. AUC = Area under the curve.

Figure 4

Number of inclusions of antigens into the 10 general discriminant analysis models that most accurately predicted the presence or absence of TB disease. PHA was evaluated in all study participants as a positive control, and appeared in one of the models because IFN-γ data was analyzed in a blinded manner.

Figure 5

Number of inclusions of antigens in the 10 most accurate general discriminant analysis combinations that most accurately predicted the presence or absence of TB disease after supernatants were re-evaluated by the Quantiferon TB Gold ELISA.