Identification, Diagnostic Potential, and Natural Expression of Immunodominant Seroreactive Peptides Encoded by Five Mycobacterium Tuberculosis-specific Genomic Regions

Comparative genomic studies have identified several Mycobacterium tuberculosis-specific genomic regions of difference (RDs) which are absent in the vaccine strains of Mycobacterium bovis BCG and which may be useful in the specific diagnosis of tuberculosis (TB). In this study, a total of 775 synthetic peptides covering the sequences of 39 open reading frame (ORF) proteins encoded by genes predicted in five RDs of M. tuberculosis, i.e., RD1, RD4, RD5, RD6, and RD7, were tested by enzyme-linked immunosorbent assays for antibody reactivity with sera from HIV-negative pulmonary TB patients (n = 100) and M. bovis BCG-vaccinated healthy subjects (n = 100). The results identified three immunodominant peptides reactive with TB sera, i.e., amino acids (aa) 346 to 370 of RD1ORF Rv3876, aa 241 to 265 of RD6ORF Rv1508c, and aa 325 to 336 of RD6ORF Rv1516c. These peptides had significantly stronger antibody reactivity with sera from TB patients than with sera from healthy subjects (P < 0.05) and significantly higher rates of positivity with TB sera (positives = 66 to 93%) than sera from healthy subjects (positives = 10 to 28%). Antipeptide antibodies were raised in rabbits after immunization with pools of 11 peptides corresponding to each protein. Probing of culture filtrates and whole-cell lysates of M. tuberculosis with antipeptide antibodies suggested the natural expression of Rv1516c in whole-cell lysates of M. tuberculosis. The results suggest the potential of the identified immunodominant RD peptides in the serodiagnosis of TB.