Comparative proteomic analysis of sequential isolates of Mycobacterium tuberculosis from a patient with pulmonary tuberculosis turning from drug sensitive to multidrug resistant
Amit Singh1, Krishnamoorthy Gopinath2, Prashant Sharma3, Deepa Bisht3, Pawan Sharma4, Niti Singh5, Sarman Singh1
1 Division of Clinical Microbiology & Molecular Medicine, Department of Laboratory Medicine, All India Institute of Medical Sciences, New Delhi, India
2 Division of Clinical Microbiology & Molecular Medicine, Department of Laboratory Medicine, All India Institute of Medical Sciences, New Delhi India; Institute of Infectious Diseases & Molecular Medicine, University of Cape Town, South Africa
3 National JALMA Institute for Leprosy & other Mycobacterial Diseases (ICMR), Agra, India
4 Immunology Group, International Centre for Genetic Engineering & Biotechnology, New Delhi; North-East Region-BPMC, Department of Biotechnology, Govt. of India, New Delhi, India
5 Department of Microbiology, National Institute of Tuberculosis & Respiratory Diseases, New Delhi, India
Division of Clinical Microbiology & Molecular Medicine, Department of Laboratory Medicine, All India Institute of Medical Sciences, Ansari Nagar, New Delhi 110 029
Source of Support: None, Conflict of Interest: None
Background & objectives: Tuberculosis is a major health problem in India, and the emergence of multidrug resistant (MDR) and extensively drug resistant (XDR) strains of Mycobacterium tuberculosis (Mtb) has further complicated the situation. Though several studies characterizing drug sensitive and drug resistant strains are available in literature, almost all studies are done on unrelated strains. Therefore, the objective of this study was to compare the proteomic data of four sequential isolates of Mtb from a single patient who developed MDR-TB during the course of anti-tuberculosis therapy (ATT).
Methods: In this study, using two-dimensional (2D) gel electrophoresis and MALDI-TOF mass spectrometry, we compared and analyzed the cell lysate proteins of Mtb sequential clinical isolates from a patient undergoing anti-TB treatment. The mRNA expression levels of selected identified proteins were determined by quantitative real-time polymerase chain reaction (qRT-PCR).
Results: The genotypes of all four isolates remained homologous, indicating no re-infection. The initial isolate (before treatment) was sensitive to all first-line drugs, but the consecutive isolates were found to be resistant to isoniazid (INH) and rifampicin (RIF) and developed mutations in the katG, inhA and rpoB. the intensities of 27 protein spots were found to be consistently overexpressed in INH and RIF resistant isolates. The most prominent and overexpressed proteins found during the development of drug resistance were GarA (Rv1827), wag31 (Rv2145c), Rv1437 and Rv2970c.
Interpretation & conclusions: This preliminary proteomic study provides an insight about the proteins that are upregulated during drug resistance development. These upregulated proteins, identified here, could prove useful as immunodiagnostic and possibly drug resistant markers in future. However, more studies are required to confirm these findings.