Identification of Mycobacterium tuberculosis adherence-mediating components: a review of key methods to confirm adhesin function

Document Type : Review Article


Medical Microbiology and Infection Control, University of KwaZulu-Natal, Durban, South Africa


Anti-adhesion therapy represents a potentially promising avenue for the treatment and prevention of tuberculosis in a post-antibiotic era. Adhesins are surface-exposed microbial structures or molecules that enable pathogenic organisms to adhere to host surfaces, a fundamental step towards host infection. Although several Mycobacterium tuberculosis adhesins have been identified, it is predicted that numerous additional adherence-mediating components contribute to the virulence and success of this pathogen. Significant further research to discern and characterize novel M. tuberculosis adhesins is, therefore, required to gain a holistic account of M. tuberculosis adhesion to the host. This would enable the identification of potential drug and vaccine targets for attenuating M. tuberculosis adherence and infectivity. Several methods have been successfully applied to the study and identification of M. tuberculosis adhesins. In this manuscript, we review these methods, which include adherence assays that utilize wild-type and gene knockout mutant strains, epitope masking and competitive inhibition analyses, extracellular matrix protein binding assays, microsphere adhesion assays, M. tuberculosis auto-aggregation assays, and in silico analyses.


1. World Health Organization. Global tuberculosis report 2014. (Online). Available from: (Accessed on 27th October, 2014).
2. Krachler AM, Orth K. Made to stick: anti-adhesion therapy for bacterial infections. Microbe 2013; 8:286-290.
3. Stones DH, Krachler AM. Fatal attraction: how bacterial adhesins affect host signaling and what we can learn from them. Int J Mol Sci 2015; 16:2626-2640.
4. Boland T, Latour RA, Stutzenberger FJ. Molecular basis of bacterial adhesion. In: An YH, Friedman RJ, editors. Handbook of bacterial adhesion: principles, methods, and applications. Totowa: Humana Press Inc; 2000. p. 29-41.
5. Ramsugit S, Pillay M. Pili of Mycobacterium tuberculosis: current knowledge and future prospects. Arch Microbiol 2015; 197:737-744.
6. Ribet D, Cossart P. How bacterial pathogens colonize their hosts and invade deeper tissues. Microbes Infect 2015; 17:173-183.
7. Soto GE, Hultgren SJ. Bacterial adhesins: common themes and variations in architecture and assembly. J Bacteriol 1999; 181:1059-1071.
8. Smith I. Mycobacterium tuberculosis pathogenesis and molecular determinants of virulence. Clin Microbiol Rev 2003; 16:463-496.
9. Govender VS, Ramsugit S, Pillay M. Mycobacterium tuberculosis adhesins: potential biomarkers as anti-tuberculosis therapeutic and diagnostic targets. Microbiology 2014; 160:1821-1831.
10. Elsinghorst EA. Measurement of invasion by gentamicin resistance. Methods Enzymol 1994; 236:405-420.
11. Brennan MJ, Delogu G, Chen Y, Bardarov S, Kriakov J, Alavi M, et al. Evidence that mycobacterial PE_PGRS proteins are cell surface constituents that influence interactions with other cells. Infect Immun 2001; 69:7326-7333.
12. Pethe K, Alonso S, Biet F, Delogu G, Brennan MJ, Locht C, et al. The heparin-binding haemagglutinin of M. tuberculosis is required for extrapulmonary dissemination. Nature 2001; 412:190-194.
13. Ramsugit S, Pillay M. Mycobacterium tuberculosis pili promote adhesion to and invasion of THP-1 macrophages. Jpn J Infect Dis 2014; 67:476-478.
14. Ramsugit S, Pillay B, Pillay M. Evaluation of the role of Mycobacterium tuberculosis pili (MTP) as an adhesin, invasin, and cytokine inducer of epithelial cells. Braz J Infect Dis 2016; 20:160-165.
15. Be NA, Bishai WR, Jain SK. Role of Mycobacterium tuberculosis pknD in the pathogenesis of central nervous system tuberculosis. BMC Microbiol 2012; 12:7.
16. Hickey TB, Thorson LM, Speert DP, Daffé M, Stokes RW. Mycobacterium tuberculosis Cpn60.2 and DnaK are located on the bacterial surface, where Cpn60.2 facilitates efficient bacterial association with macrophages. Infect Immun 2009; 77:3389-3401.
17. Abou-Zeid C, Ratliff TL, Wiker HG, Harboe M, Bennedsen J, Rook GA. Characterization of fibronectin-binding antigens released by Mycobacterium tuberculosis and Mycobacterium bovis BCG. Infect Immun 1988; 56:3046-3051.
18. Kumar S, Puniya BL, Parween S, Nahar P, Ramachandran S. Identification of novel adhesins of M. tuberculosis H37Rv using integrated approach of multiple computational algorithms and experimental analysis. PLoS One 2013; 8:e69790.
19. Alteri CJ, Xicohténcatl-Cortes J, Hess S, Caballero-Olín G, Girón JA, Friedman RL. Mycobacterium tuberculosis produces pili during human infection. Proc Natl Acad Sci U S A 2007; 104:5145-5150.
20. Pethe K, Puech V, Daffé M, Josenhans C, Drobecq H, Locht C, et al. Mycobacterium smegmatis laminin-binding glycoprotein shares epitopes with Mycobacterium tuberculosis heparin-binding haemagglutinin. Mol Microbiol 2001; 39:89-99.
21. Kinhikar AG, Vargas D, Li H, Mahaffey SB, Hinds L, Belisle JT, et al. Mycobacterium tuberculosis malate synthase is a laminin-binding adhesin. Mol Microbiol 2006; 60:999-1013.
22. Espitia C, Laclette JP, Mondragón-Palomino M, Amador A, Campuzano J, Martens A, et al. The PE-PGRS glycine-rich proteins of Mycobacterium tuberculosis: a new family of fibronectin-binding proteins? Microbiology 1999; 145:3487-3495.
23. Xolalpa W, Vallecillo AJ, Lara M, Mendoza-Hernandez G, Comini M, Spallek R, et al. Identification of novel bacterial plasminogen-binding proteins in the human pathogen Mycobacterium tuberculosis. Proteomics 2007; 7:3332-3341.
24. Esparza M, Palomares B, García T, Espinosa P, Zenteno E, Mancilla R. PstS-1, the 38-kDa Mycobacterium tuberculosis glycoprotein, is an adhesin, which binds the macrophage mannose receptor and promotes phagocytosis. Scand J Immunol 2015; 81:46-55.
25. Diaz-Silvestre H, Espinosa-Cueto P, Sanchez-
Gonzalez A, Esparza-Ceron MA, Pereira-Suarez AL, Bernal-Fernandez G, et al. The 19-kDa antigen of Mycobacterium tuberculosis is a major adhesin that binds the mannose receptor of THP-1 monocytic cells and promotes phagocytosis of mycobacteria. Microb Pathog 2005; 39:97-107.
26. Chitale S, Ehrt S, Kawamura I, Fujimura T, Shimono N, Anand N, et al. Recombinant Mycobacterium tuberculosis protein associated with mammalian cell entry. Cell Microbiol 2001; 3:247-254.
27. Menozzi FD, Rouse JH, Alavi M, Laude-Sharp M, Muller J, Bischoff R, et al. Identification of a heparin-binding hemagglutinin present in mycobacteria. J Exp Med 1996; 184:993-1001.
28. Ramsugit S, Guma S, Pillay B, Jain P, Larsen MH, Danaviah S, et al. Pili contribute to biofilm formation in vitro in Mycobacterium tuberculosis. Antonie Van Leeuwenhoek 2013; 104:725-735.
29. Sachdeva G, Kumar K, Jain P, Ramachandran S. SPAAN: a software program for prediction of adhesins and adhesin-like proteins using neural networks. Bioinformatics 2005; 21:483-491.
30. Nair R, Rost B. Mimicking cellular sorting improves prediction of subcellular localization. J Mol Biol 2005; 348:85-100.
31. Yu CS, Lin CJ, Hwang JK. Predicting subcellular localization of proteins for Gram-negative bacteria by support vector machines based on n-peptide compositions. Protein Sci 2004; 13:1402-1406.
32. Chen H, Huang N, Sun Z. SubLoc: a server/client suite for protein subcellular location based on SOAP. Bioinformatics 2006; 22:376-377.
33. Chaudhuri R, Kulshreshtha D, Raghunandanan MV, Ramachandran S. Integrative immunoinformatics for Mycobacterial diseases in R platform. Syst Synth Biol 2014; 8:27-39.
34. Yu NY, Wagner JR, Laird MR, Melli G, Rey S, Lo R, et al. PSORTb 3.0: improved protein subcellular localization prediction with refined localization subcategories and predictive capabilities for all prokaryotes. Bioinformatics 2010; 26:1608-1615.
35. Ragas A, Roussel L, Puzo G, Rivière M. The Mycobacterium tuberculosis cell-surface glycoprotein apa as a potential adhesin to colonize target cells via the innate immune system pulmonary C-type lectin surfactant protein A. J Biol Chem 2007; 282:5133-5142.
36. Hickey TB, Ziltener HJ, Speert DP, Stokes RW. Mycobacterium tuberculosis employs Cpn60.2 as an adhesin that binds CD43 on the macrophage surface. Cell Microbiol 2010; 12:1634-1647.
37. Kinhikar AG, Verma I, Chandra D, Singh KK, Weldingh K, Andersen P, et al. Potential role for ESAT6 in dissemination of M. tuberculosis via human lung epithelial cells. Mol Microbiol 2010; 75:92-106.
38. Pancholi V, Fischetti VA. A major surface protein on group A streptococci is a glyceraldehyde-3-phosphate-dehydrogenase with multiple binding activity. J Exp Med 1992; 176:415-426.
39. Alteri CJ. Novel pili of Mycobacterium tuberculosis. PhD Thesis, University of Arizona 2005.