A Short-Tandem-Repeat Assay ( <i>Mmy</i> STR) for Studying Genetic Variation in <i>Madurella mycetomatis</i>

Abstract

<i>Madurella mycetomatis</i> is the major causative agent of eumycetoma, a neglected tropical infection characterized by painless subcutaneous lesions, inflammation, and grains draining from multiple sinuses. To study the epidemiology of mycetoma, a robust discriminatory typing technique is needed. We describe the use of a short-tandem-repeat assay (<i>Mmy</i>STR) for genotyping of <i>M. mycetomatis</i> isolates predominantly from Sudan. Eleven microsatellite markers (3 dinucleotides, 4 trinucleotide repeats, and 4 tetranucleotide repeats) were selected from the <i>M. mycetomatis</i> MM55 genome using the Tandem Repeats Finder software. PCR amplification primers were designed for each microsatellite marker using primer3 software and amplified in a multicolor multiplex PCR approach. To establish the extent of genetic variation within the population, a collection of 120 clinical isolates from different regions was genotyped with this assay. The 11 selected <i>Mmy</i>STR markers showed a large genotypic heterogeneity. From a collection of 120 isolates, 108 different genotypes were obtained. Simpson's diversity index (D) value for individual markers ranged from 0.081 to 0.881, and the combined panel displayed an overall D value of 0.997. The <i>Mmy</i>STR assay demonstrated high stability, reproducibility, and specificity. The <i>Mmy</i>STR assay is a promising new typing technique that can be used to genotype isolates of <i>M. mycetomatis</i> Apart from the possible contribution of host factors, the genetic diversity observed among this group of isolates might contribute to the different clinical manifestations of mycetoma. We recommend that the <i>Mmy</i>STR assay be used to establish a global reference database for future study of <i>M. mycetomatis</i> isolates.