Whole genome sequencing distinguishes between relapse and reinfection in recurrent leprosy cases

Abstract

Background Since leprosy is both treated and controlled by multidrug therapy (MDT) it is important to monitor recurrent cases for drug resistance and to distinguish between relapse and reinfec tion as a means of assessing therapeutic efficacy. All three objectives can be reached with single nucleotide resolution using next generation sequencing and bioinformatics analysis of Mycobacterium leprae DNA present in human skin. Methodology DNA was isolated by means of optimized extraction and enrichment methods from samples from three recurrent cases in leprosy patients participating in an open-label, randomized, controlled clinical trial of uniform MDT in Brazil (U-MDT/CT-BR). Genome-wide sequencing of M. leprae was performed and the resultant sequence assemblies analyzed in silico. Principal findings In all three cases, no mutations responsible for resistance to rifampicin, dapsone and ofloxa cin were found, thus eliminating drug resistance as a possible cause of disease recurrence However, sequence differences were detected between the strains from the first and second disease episodes in all three patients. In one case, clear evidence was obtained for reinfec tion with an unrelated strain whereas in the other two cases, relapse appeared more probable. Conclusions/Significance This is the first report of using M. leprae whole genome sequencing to reveal that treated and cured leprosy patients who remain in endemic areas can be reinfected by another strain. Next generation sequencing can be applied reliably to M. leprae DNA extracted from biopsies to discriminate between cases of relapse and reinfection, thereby providing a pow erful tool for evaluating different outcomes of therapeutic regimens and for following disease transmission.