Development and validation of a microsatellite marker-based method for tracing infections by Microsporum canis

Abstract Background Microsporum canis is a dermatophyte fungus harbored by cats and dogs and is frequently transmitted to humans. Molecular tools able to discriminate fungal isolates at the strain level would prove extremely useful for confirming the route of infection, thus contributing to optimization of prophylaxis and hygienic regimens. Objective To develop and validate a microsatellite marker-based method for use in tracking infections by M. canis. Methods Primers were designed against sequences flanking the microsatellites individuated by a BLAST search using the nucleotide sequence information assembled by the M. canis CBS 113480 genome project. The PCR conditions were standardized and fragment analysis was performed using a genetic analyzer. The resolving power of the markers was investigated on 26 unrelated M. canis strains while the reproducibility of the technique and the stability of the markers were evaluated on a single strain subcultured in time as well as on 36 strains isolated from nine outbreak episodes. Results Eight markers were recognized as being the most polymorphic within the set of M. canis strains isolated from unrelated distant hosts, with a total of 22 multilocus genotypes, which corresponded to a genotypic diversity of 97%. Repeated tests on subcultures of M. canis reference strain CBS 113480 always yielded the same results. Identical multilocus genotypes were obtained for all the isolates from each outbreak episode. Conclusion The high resolving power and reproducibility of the markers that were identified support the potential of these tools to detect sources and routes of infection by M. canis.