Multilocus microsatellite typing for Rhizopus oryzae.

Area of Science:

• Medical Mycology
• Molecular Biology
• Epidemiology

Background:

• Rhizopus oryzae is a primary cause of zygomycosis, a serious infection in immunocompromised individuals.
• The epidemiology of zygomycosis is poorly understood due to a lack of standardized molecular typing methods for causative agents.
• Effective strain differentiation is crucial for tracking disease outbreaks and understanding transmission.

Purpose of the Study:

• To develop and validate a novel multilocus microsatellite typing (MLMT) method for Rhizopus oryzae.
• To assess the discriminatory power and stability of the developed MLMT method for fungal strain typing.
• To explore the applicability of the MLMT method for related species like Rhizopus delemar.

Main Methods:

• Bioinformatic screening of R. oryzae genome sequences to identify microsatellite loci using Repeat Masker.
• Selection and validation of three polymorphic microsatellite loci (RO3, RO4, RO8) based on amplification and size variation in clinical isolates.
• Strain differentiation of 30 clinical isolates using automated capillary electrophoresis with fluorescently labeled primers.

Main Results:

• The developed MLMT method demonstrated high discriminatory power (combined D=0.986) for differentiating R. oryzae strains.
• The selected microsatellite loci showed good stability across seven subcultures.
• The MLMT method was applicable to both R. oryzae and R. delemar, though unable to distinguish between them.

Conclusions:

• The novel MLMT method provides a powerful and reliable tool for the molecular typing of Rhizopus oryzae strains.
• This method is expected to significantly advance epidemiological investigations of zygomycosis.
• The MLMT system holds promise for routine use in clinical and research settings for fungal strain identification.