The production of anti-pili monoclonal antibodies for the development of a point of care diagnostic test for rapid detection of M. tuberculosis

The production of anti-pili monoclonal antibodies for the development of a point of care diagnostic test for rapid detection of M. tuberculosis

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Email Address: 
pillay2@gmail.com
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+27 (0) 71 482-9504

Mycobacterium tuberculosis is the second deadliest infectious disease in the developing world after HIV. Pili, hair-like outgrowths present on the surface of Mycobacterium tuberculosis (MTB) bacilli, have been reported to extend far beyond the surface of the bacterial cell. Therefore the use of these appendages as TB diagnostic markers may be advantageous. In addition, M. tuberculosis pili (MTP) represent novel putative diagnostic targets for use on paper-based lateral flow assays. The use of cheaply available paper-based microfluidics in the design of point-of-care tests would assist diagnosis of major infectious diseases in low-income countries as these don’t require extensive training or electricity to be implemented. In the past 5 years, Naidoo’s research group has gathered evidence to substantiate the potential role of MTP for TB diagnosis. BLAST analysis of the mtp gene and the MTP amino acid sequence on publically available complete genomes and amplicon sequencing of the mtp gene in all M. tuberculosis complex (MTBC) strains, non-tuberculous mycobacteria (NTM) and respiratory pathogens and commensals showed that the gene is unique to and conserved within the genome of MTBC clinical isolates. MTP were shown to be essential for pili production and biofilm formation. Real Time PCR was used to assess mtp gene expression in aerobically and anaerobically grown planktonic and biofilm-derived cultures of M. tuberculosis strains and confirmed the exclusive mtp gene expression in MTBC strains. A synthetic MTP peptide was purchased and evaluated in a slot blot assay for the detection of anti-MTP IgG antibodies. The slot blot assay yielded a high sensitivity (100 %) and specificity (92.5 %), positive predictive value (95.24%) and negative predictive value (100%), confirming that MTP is a valuable TB diagnostic biomarker that should be further evaluated in larger population based studies. This is further strongly supported by the positive response of the HIV co-infected patients, and further testing on smear negative HIV co-infected patients and children may show that MTP can potentially detect antibodies in these problematic samples. However, screening for antibodies in blood serum faces a few challenges, including the detection of past infections and disease episodes and antibodies elicited due to BCG vaccination. Naidoo’s study is based on the generation of anti-MTP monoclonal antibodies for the design of a lateral flow point of care test to detect pili antigen in sputum samples which can be a potentially more accurate test to be used at the patient’s bedside.