Article

Gene Expression Profile and Immunological Evaluation of Unique Hypothetical Unknown Proteins of Mycobacterium leprae by Using Quantitative Real-Time PCR

Department of Microbiology, Immunology and Pathology. Colorado State University, Fort Collins, Colorado 80523-2025.
Clinical and vaccine Immunology: CVI (Impact Factor: 2.47). 12/2012; 20(2). DOI: 10.1128/CVI.00419-12
Source: PubMed

ABSTRACT

The cell-mediated immunity (CMI)-based in vitro gamma interferon release assay (IGRA) of Mycobacterium leprae-specific antigens has potential as a promising diagnostic means to detect those individuals in the early stages of M. leprae infection. Diagnosis of leprosy is a major obstacle toward ultimate disease control and has been compromised in the past
by the lack of specific markers. Comparative bioinformatic analysis among mycobacterial genomes identified potential M. leprae-specific proteins called “hypothetical unknowns.” Due to massive gene decay and the prevalence of pseudogenes, it is unclear
whether any of these proteins are expressed or are immunologically relevant. In this study, we performed cDNA-based quantitative
real-time PCR to investigate the expression status of 131 putative open reading frames (ORFs) encoding hypothetical unknowns.
Twenty-six of the M. leprae-specific antigen candidates showed significant levels of gene expression compared to that of ESAT-6 (ML0049), which is an
important T cell antigen of low abundance in M. leprae. Fifteen of 26 selected antigen candidates were expressed and purified in Escherichia coli. The seroreactivity to these proteins of pooled sera from lepromatous leprosy patients and cavitary tuberculosis patients
revealed that 9 of 15 recombinant hypothetical unknowns elicited M. leprae-specific immune responses. These nine proteins may be good diagnostic reagents to improve both the sensitivity and specificity
of detection of individuals with asymptomatic leprosy.

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