Evaluation and Use of Three PCR-Based Typing Methods for Aspergillus fumigatus Isolates


Author address: 

Centers for Disease Control and Prevention, Atlanta, GA


Background: A. fumigatus is an opportunistic fungal pathogen responsible for most cases of human aspergillosis. Typing of isolates may lead to better prevention measures and improved guidelines for patient management due to a greater understanding of the potential sources of infections. Three PCR-based molecular typing methods were evaluated to determine their epidemiologic utility for use in outbreak and surveillance investigations. Methods: 49 Aspergillus fumigatus isolates from four nosocomial outbreaks were evaluated with respect to discriminatory power (D), reproducibility, typability, ease of use, and ease of interpretation. The 3 typing methods were: random amplified polymorphic DNA (RAPD) using 4 primers (R108, R151, UBC90 and NS2), sequence-specific polymorphic DNA (SSPD) using 5 primer pairs, and 4 polymorphic microsatellite markers (PMM). Results: Using RAPD 35 types were observed. PMM detected 36 types but SSPD detected only 14 types. 49 types were obtainable by a combination of all three methods. All three methods demonstrated 100% typability. PMM and RAPD had comparable high degrees of discriminatory power, D=0.988 and D=0.982, respectively, whereas SSPD had a lower discriminatory power (D=0.889). SSPD was the easiest method to interpret and perform, followed by PMM, which was easier to interpret and use than RAPD. Reproducibility was high for both SSPD and PMM. Bands differing in staining intensity were observed by RAPD making interpretation more difficult. Conclusions: The results demonstrated a high degree of genetic variation within clinical the population of isolates analyzed. A general concordance between methods was observed but not always apparent suggesting differences in stability of markers among methods or strain diversity may be due to recombination or spontaneous mutations. Despite comparable discrimination, PMM was more reproducible and easier to interpret than RAPD. This investigation will aid in planning epidemiologic and surveillance studies of A. fumigatus.

abstract No: 


Full conference title: 

ICAAC 41st
    • ICAAC 41st