Comparison of genotyping methods for Cunninghamella bertholletiae

Background: Invasive fungal infections caused by filamentous fungi of the order Mucorales are serious complications in immunocompromised patients and often associated with fatal outcome. As a member of this order, Cunninghamella bertholletiae is a saprophytic fungus with naturally exhibited high minimum inhibitory concentrations against common antifungal drugs and with the potential for outbreaks in clinical settings.

Objectives and methods: In a proof-of-principle study, we evaluated the performance of microsatellite markers for the discrimination of thirteen C. bertholletiae isolates from various sources in comparison with a repetitive sequence-based PCR (rep-PCR) and random amplification of polymorphic DNA (RAPD). Based on the higher discriminatory power of the microsatellite PCR with five separate primer pairs (Simpson's index of 1 vs 0 [RAPD] and 0 [rep-PCR]), the novel method was applied to eight additional isolates, including four well-characterised isolates from a cluster of infections in a next step.

Results: In total, microsatellite PCR identified 21 separate genotypes. A probable epidemiological association of the cluster isolates could be demonstrated by microsatellite genotyping.

Conclusion: In conclusion, our findings demonstrate the value of microsatellite PCR in genotyping Cunninghamella bertholletiae and its potential for future applications with other species of the order Mucorales.