In recent years, the prevalence of tuberculosis worldwide has increased, and with it, the number of drug-resistant tuberculosis strains. This has brought new challenges towards prevention and control of the disease. Therefore, it is urgent to find reliable and rapid diagnostic methods for tuberculosis in general, and for the drug-resistant forms of the disease. To this aim, we assessed 17 tuberculosis-specific protein candidates for the detection of tuberculosis-specific antibodies. First, we established an indirect ELISA method to detect anti-Mycobacterium tuberculosis IgM and IgG. We tested 453 sera and analyzed the efficacy of the protein candidates for diagnosis of tuberculosis. Next, we screened antigens rich in T cell epitopes for their ability to induce high levels of IFN-γ, in order to define their suitability does develop detection tests based on IFN-γ release assay (IGRAs). The antigens CFP-10, PPE57, 38kDa, and Rv3807 showed higher diagnostic potential for the detection of anti-tuberculosis IgM, whereas PPE57, Ag85B, CFP-10, Rv0220, and 38kDa antigens performed better for anti-tuberculosis IgG detection. Worth noting is that CFP-10, 38kDa, and PPE57 detected efficiently both IgM and IgG. Rv1987, Rv3807, PPE57, Rv0220, and MPT64 proteins alone and combinations of Rv1987 + Rv3807, 16kDa + Rv0220, and MPT64 + Rv1986 tested in IGRAs displayed a good correlation with the positive control constituted by a cocktail of ESAT-6 + CFP-10 + TB7.7 (ECT), known for their stimulating properties (r > 0.5, p < 0.01). Among these antigen candidates, Rv0220 and Rv1987 + Rv3807 were the most potent. We discovered CFP-10, 38kDa, and PPE57 for the detection of anti-M. tuberculosis IgM and IgG, and Rv0220 alone or the combination Rv1987 + Rv3807 as the strongest stimulators in IGRAs. These antigens provide new references for the screening of tuberculosis-specific antibodies and effective stimulation in IGRAs.
This is a preview of subscription content, log in to check access.
Buy single article
Instant access to the full article PDF.
Price includes VAT for USA
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
This is the net price. Taxes to be calculated in checkout.
Vasilyeva I, Belilovsky E, Borisov S, Sterlikov S (2017) Global tuberculosis reports by WHO, compilation and interpretation. Tuber Lung Dis 95(5):7–16
Chao W, Li-xian W (2013) Advances in the study of the prevalence of co-infection with AIDS and TB and the mechanisms by which AIDS and TB interact. J Path Biol 8(8):756–758
Casadevall A (2017) Antibodies to Mycobacterium tuberculosis. N Engl J Med 376(3):283
Andersen P, Andersen AB, Sørensen AL, Nagai S (1995) Recall of long-lived immunity to Mycobacterium tuberculosis infection in mice. J Immunol 154(7):3359
Mcmanus DP, Loukas A (2008) Current status of vaccines for schistosomiasis. Clin Microbiol Rev 21(1):225
Baumann R, Kaempfer S, Chegou NN, Oehlmann W, Spallek R, Loxton AG, van Helden PD, Black GF, Singh M, Walzl G (2015) A subgroup of latently Mycobacterium tuberculosis infected individuals is characterized by consistently elevated IgA responses to several mycobacterial antigens. Mediat Inflamm 2015(7):364758
Chen T, Lin J, Wang W, Joy F, Chen L, Wang Y, Li H, Guo H, Zhou J, Chen X (2015) Cytokine and antibody based diagnostic algorithms for sputum culture-positive pulmonary tuberculosis. PLoS One 10(12):e0144705
S-j LI, Z-g Z, Qin Y, Luo Q, Wang L, X-q LI, M-c LI, H-c L, Wan K-l (2018) Application value of detecting Mycobacterium tuberculosis specific IgG/IgM antibodies for tuberculosis diagnosis with colloidal gold immunochromatography assay. Chin J Zoonoses 34(2):139–143,149
Grange J, Gibson J, Nassau E, Kardjito T (1980) Enzyme-linked immunosorbent assay (ELISA): a study of antibodies to Mycobacterium tuberculosis in the IgG, IgA and IgM classes in tuberculosis, sarcoidosis and Crohn’s disease. Tubercle 61(3):145–152
Devi KRU, Ramalingam B, Brennan PJ, Narayanan PR, Raja A (2001) Specific and early detection of IgG, IgA and IgM antibodies to Mycobacterium tuberculosis 38kDa antigen in pulmonary tuberculosis. Tuberculosis 81(3):249–253
Mahairas GG, Sabo PJ, Hickey MJ, Singh DC, Stover CK (1996) Molecular analysis of genetic differences between Mycobacterium bovis BCG and virulent M. bovis. J Bacteriol 178(5):1274–1282
Mustafa AS (2005) Mycobacterial gene cloning and expression, comparative genomics, bioinformatics and proteomics in relation to the development of new vaccines and diagnostic reagents. Med Princ Pract 14(Suppl.1):27–34
Mukherjee P, Dutta M, Datta P, Dasgupta A, Pradhan R, Pradhan M, Kundu M, Basu J, Chakrabarti P (2007) The RD1-encoded antigen Rv3872 of Mycobacterium tuberculosis as a potential candidate for serodiagnosis of tuberculosis. Clin Microbiol Infect 13(2):146–152
Rosenkrands I, Aagaard C, Weldingh K, Brock I, Dziegiel MH, Singh M, Hoff S, Ravn P, Andersen P (2008) Identification of Rv0222 from RD4 as a novel serodiagnostic target for tuberculosis. Tuberculosis 88(4):335–343
Baassi L, Sadki K, Seghrouchni F, Contini S, Cherki W, Nagelkerke N, Benjouad A, Saltini C, Colizzi V, El AR (2009) Evaluation of a multi-antigen test based on B-cell epitope peptides for the serodiagnosis of pulmonary tuberculosis. Int J Tuberc Lung Dis 13(7):848
Murthy MK, Parasa RRV, Deenadayalan A, Sharma P, Raja A (2007) Evaluation of the diagnostic potential of region of deletion-1–encoded antigen culture filtrate protein-10 in pulmonary tuberculosis. Diagn Microbiol Infect Dis 59(3):295–302
Rogan WJ, Gladen B (1978) Estimating prevalence from the results of a screening test. Am J Epidemiol 107(1):71–76
Betty RK, Jonathan ACS (2003) Essentials of medical statistics. Blackwell Science Ltd., Hoboken
Omrani MD, Ansari MHK, Agaverdizadae D (2009) PCR and Elisa methods (IgG and IgM): their comparison with conventional techniques for diagnosis of Mycobacterium tuberculosis. Pak J Biol Sci 12(4):373–377
Renshaw PS, Lightbody KL, Veverka V, Muskett FW, Kelly G, Frenkiel TA, Gordon SV, Hewinson RG, Burke B, Norman J (2005) Structure and function of the complex formed by the tuberculosis virulence factors CFP-10 and ESAT-6. EMBO J 24(14):2491–2498
Osadaoka M, Tateishi Y, Hirayama Y, Ozeki Y, Niki M, Kitada S, Maekura R, Tsujimura K, Koide Y, Ohara N (2013) Antigen 85A and mycobacterial DNA-binding protein 1 are targets of immunoglobulin G in individuals with past tuberculosis. Microbiol Immunol 57(1):30
Lin N, Qin YU, Liu YJ, Wei YF, Wan KL (2016) Application evaluation of Ag85A,Ag85B,16kDa and 38kDa in the serological diagnosis of tuberculosis. Chin J Health Lab Technol 12(28):2585–2591
Dong EJ, Zhang LX, Zhang CY (2011) Value of detection of recombinant Ag85A and Ag85B antigen of Mycobacterium tuberculosis in diagnosis of tuberculosis. China Tropical Medicine 2011(3):29–30
Bai Y, Xue Y, Gao H, Wang L, Ding T, Bai W, Fan A, Zhang J, An Q, Xu Z (2008) Expression and purification of Mycobacterium tuberculosis ESAT-6 and MPT64 fusion protein and its immunoprophylactic potential in mouse model. Protein Expr Purif 59(2):189–196
Azzurri A, Kanaujia GV, Sow OY, Bah B, Diallo A, Del PG, Gennaro ML (2006) Serological markers of pulmonary tuberculosis and of response to anti-tuberculosis treatment in a patient population in Guinea. Int J Immunopathol Pharmacol 19(1):199–208
Brusasca PN, Colangeli R, Lyashchenko KP, Zhao X, Vogelstein M, Spencer JS, Mcmurray DN, Gennaro ML (2001) Immunological characterization of antigens encoded by the RD1 region of the Mycobacterium tuberculosis genome. Scand J Immunol 54(5):448–452
Chen JZ, Su XD, Zhang Y, Wang S, Shao LY, Wu J, Wang FF, Zhang S, Wang JL, Weng XH (2009) Novel recombinant RD2- and RD11-encoded Mycobacterium tuberculosis antigens are potential candidates for diagnosis of tuberculosis infections in BCG-vaccinated individuals. Microbes Infect 11(10–11):876
Kellar KL, Gehrke J, Weis SE, Mahmutovicmayhew A, Davila B, Zajdowicz MJ, Scarborough R, Lobue PA, Lardizabal AA, Daley CL (2011) Multiple cytokines are released when blood from patients with tuberculosis is stimulated with Mycobacterium tuberculosis antigens. PLoS One 6(11):e26545
Rangaka MX, Wilkinson KA, Glynn JR, Ling D, Menzies D, Mwansa-Kambafwile J, Fielding K, Wilkinson RJ, Pai M (2012) Predictive value of interferon-γ release assays for incident active tuberculosis: a systematic review and meta-analysis. Lancet Infect Dis 12(1):45
Kawamura LM, Grinsdale JA, Ho CS, Jereb JA (2012) Interferon-γ release assays for prediction of tuberculosis. Lancet Infect Dis 12(8):584–584
Park HJ, Shin JA, Kim HJ, Ahn CM, Chang YS (2014) Whole blood interferon-γ release assay is insufficient for the diagnosis of sputum smear negative pulmonary tuberculosis. Yonsei Med J 55(3):725
Basirudeen SAK, Balambal R, Aleyamma T, Venkatesan P, Alamelu R (2010) Role of QuantiFERON-TB gold, interferon gamma inducible protein-10 and tuberculin skin test in active tuberculosis diagnosis. PLoS One 5(2):e9051
Wiker HG, Harboe M (1992) The antigen 85 complex: a major secretion product of Mycobacterium tuberculosis. Microbiol Rev 56(4):648
Drowart A, Cambiaso CL, Huygen K, Serruys E, Portaels F, Jann E, Van Vooren JP (1997) Detection of rifampicin and isoniazid resistances of Mycobacterium tuberculosis strains by particle counting immunoassay (PACIA). Int J Tuberc Lung Dis 1(3):284
Launois P, Deleys R, Niang MN, Drowart A, Andrien M, Dierckx P, Cartel JL, Sarthou JL, Van Vooren JP, Huygen K (1994) T-cell-epitope mapping of the major secreted mycobacterial antigen Ag85A in tuberculosis and leprosy. Infect Immun 62(9):3679–3687
Deng MZ, Shi CW, Wang F, Rui-Ling FU, Wang C, Fang ZM, Fan XL (2010) Prokaryotic expression,purification and immunoreactivity of Mycobacterium tuberculosis Ag85A protein. Basic Clin Med 30(2):117–121
We thank LetPub (www.letpub.com) for its linguistic assistance during the preparation of this manuscript.
This research was supported by the National Key Research & Development Plan (2017YDF0500304) and National Natural Science Foundation of China (U1803236), and Major Scientific and Technological Projects of Corps (2017AA003).
Conflict of interest
The authors declare that they have no conflict of interest.
We have complied with the World Medical Association Declaration of the 1964 Helsinki regarding ethical conduct of research involving human subjects. The hospital based case–control study was approved by the Review Board of Kashgar Pulmonary Hospital (Xinjiang, China).
A written informed consent was obtained from all subjects who were recruited and interviewed for the study.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
About this article
Cite this article
Ma, Z., Ji, X., Yang, H. et al. Screening and evaluation of Mycobacterium tuberculosis diagnostic antigens. Eur J Clin Microbiol Infect Dis (2020). https://doi.org/10.1007/s10096-020-03951-3
- Mycobacterium tuberculosis
- Indirect ELISA
- IFN-γ release test (IGRAs)