Molecular typing of Aspergillus fumigatus isolates collected from cystic fibrosis patients

Twenty-seven strains of Aspergillus fumigatus was isolated from the respiratory tract secretions of patients with cystic fibrosis (CF) together with P. aeruginosa and in some cases S. aureus. The potential pathogenic role of A. fumigatus in this patients category is controversial and it is supposed to be a colonizer. The aim of this work was to verify if C.F patients are colonized by a unique Aspergillus fumigatus genotype because literature data report that some genotypes are dominant and persistent in immunocompromised hosts. In order to verify genetic relatedness, molecular typing methods including random amplified polymorhic DNA (RAPD) with two different primers (NS3 and R108) and Afut I restriction fragment lenght polymorphism (RFLP), were performed. Use of combination of methods resulted in a greater degree of discriminatory power than that can be achieved by a single method. Our results suggest that different types of A. fumigatus exist in CF patients as colonizers. For comparative purposes, we also tested the in vitro susceptibility to various antifungals (voriconazole, itraconazole and amphotericin B). No significant differences were observed except for one strain with high MIC value for itraconazole (16 mg/ml). Furthermore there was no correlation between genotype and susceptibility patterns. The strain with high MIC for itraconazole (ITZ) was furtherly investigated for the full coding region sequences of 14a-sterol demethylase genes (cyp51A and cyp51B). Sequence analysis revealed no significant point mutation related to the different behaviour regarding ITZ. The expression of efflux pumps are now under evaluation.