2023 Volume 92 Issue 2 Pages 47-52
Mycobacterium leprae is the major etiologic agent of leprosy, and its genotype can be divided into four single nucleotide polymorphism (SNP) types and 16 subtypes. Drug resistance and genotype of M. leprae are typically determined using PCR and Sanger DNA sequencing, however, this requires a great deal of effort. In this review, we present a rapid method that we developed to identify drug resistance and SNP genotype of M. leprae directly from clinical specimens by combining nested multiplex PCR with next generation sequence analysis.
We used this method to analyze clinical samples from two paucibacillary, nine multibacillary, and six type-undetermined leprosy patients. From these, we amplified drug resistance determining regions (DRDR) of folP1, rpoB, gyrA, and gyrB, and regions of 84 SNP-InDels in the M. leprae genome and their sequences were determined.
The results showed that seven samples were subtype 1A, three were 1D, and seven were 3K. Three samples of the subtype 3K had folp1 mutation. Of the three, two showed an A to G mutation at nucleotide 157 resulting in a Thr to Ala at amino acid 53(ACC→GCC) in folp1 and one had a C to T mutation at nucleotide 164 resulting in a Pro to Leu at amino acid 55(CCC→CTC) in folp1. No mutations were detected in the DRDRs of rpoB, gyrA, and gyrB in all the samples. The method may allow more rapid genetic analyses of M. leprae in clinical samples.
