Mycobacterium lepromatosis genome exhibits unusually high CpG dinucleotide content and selection is key force in shaping codon usage.

Mycobacterium lepromatosis was identified as a causative agent for leprosy in the year 2008 in the United States and later more cases were identified in Canada, Singapore, Brazil, and Myanmar. It is known to cause diffuse lepromatosis leprosy among humans. Since it is invasive, the mortality rates are higher in comparison to the M. leprae. At genomic level, there exists 90.9% similarity between M. lepromatosis and M. leprae. Codon usage analysis based on analyses of 228 coding sequences (CDSs) of M. lepromatosis, revealed that the genome is GC rich. Among the total 16 dinucleotides, CpG dinucleotide possesses the highest dinucleotide frequency in M. lepromatosis, that is strikingly an unobvious observation since higher CpG is associated with higher proinflammatory cytokine production and NF-κB activation that eventually leads to high pathogenicity. To evade immune response, CpG content is generally less in pathogens. The unusually high CpG content can be explained by the fact that the nucleotide composition of M. lepromatosis is CG rich. Various forces interplay to shape codon usage pattern of any organism including selection; mutation, nucleotide composition as well as GC biased gene conversion. To understand the interplay between various forces; neutrality, parity, Nc-GC3 (Effective number of codons-GC content at 3rd position of the codon), aromaticity (AROMO) and the general average hydropathicity score (GRAVY) analyses have been carried out. The analyses revealed that selection force is the major contributory force. Along with the selection; mutation, nucleotide composition as well as GC biased gene conversion also play role in shaping codon usage bias in M. lepromatosis. This is the first report on the codon usage in M. lepromatosis.

Effect of hsp65 DNA vaccination carrying immunostimulatory DNA sequences (CpG motifs) against Mycobacterium leprae multiplication in mice.

A DNA expressing hsp65 of Mycobacterium leprae (pACB/hsp65) was constructed by using a vector containing immunostimulatory DNA sequences (pACB). At 12 weeks post-immunization, spleen cells from BALB/cA mice immunized with pACB/hsp65, produced a significantly higher amount of IFN-gamma than mice immunized with pACB in the absence of any in vitro stimulation, and further enhanced its production upon secondary in vitro stimulation with M. leprae lysate and hsp65. On the other hand, while production of IL-12 was observed in mice immunized with pACB/hsp65 12 weeks before, the cytokine production was inhibited by in vitro secondary stimulation with M. leprae or hsp65. At 18 weeks post-immunization, the production of both IFN-gamma and IL-12 was apparently down-regulated, but that of IL-10 was up-regulated. IL-10 seemed to suppress the IFN-gamma and IL-12 productions, because their production was recovered by neutralization of IL-10 with anti-IL-10 mAb. Furthermore, when the efficiency of pACB/hsp65 as a vaccine against M. leprae was evaluated in vivo, the mice immunized with pACB/hsp65 suppressed the multiplication of subsequently challenged M. leprae. These results suggest that a DNA containing M. leprae-derived hsp65 and immunostimulatory sequences might be a potent vaccine candidate against M. leprae infection.