High Prevalence of Azole-Resistant Aspergillus fumigatus Among Iranian Cystic Fibrosis Patients: Should We Be Concerned?

Background: Cystic fibrosis (CF), an inherited autosomal recessive disorder, is linked with high morbidity and mortality rates due to bacteria, filamentous, yeast and black yeast-like fungi colonisation in the upper respiratory tract. Although Candida species are the most common fungi isolated from CF patients, azole-resistant Aspergillus fumigatus (ARAf) is a big concern for invasive aspergillosis. Notably, the exact prevalences of Aspergillus species and the prevalence of ARAf isolates among Iranian CF patients have yet to be previously reported and are unknown. We aimed to investigate the prevalence of ARAf isolates in CF patients among Iranian populations by focusing on molecular mechanisms of the mutations in the target gene.
Methods: The 1 year prospective study recovered 120 sputum samples from 103 CF patients. Of these, 55.1% (86/156) yielded Aspergillus species, screened for ARAf using plates containing itraconazole (4 mg/L) and voriconazole (1 mg/L). According to the CLSI-M38 guidelines, antifungal susceptibility testing was performed using the broth microdilution method. In all phenotypically resistant isolates, the target of azole agents, the cyp51A gene, was sequenced to detect any possible single nucleotide polymorphisms (SNP) mediating resistance.
Results: Of 120 samples, 101 (84.2%) were positive for filamentous fungi and yeast-like relatives, with 156 fungal isolates. The most common colonising fungi were Aspergillus species (55.1%, 86/156), followed by Candida species (39.8%, 62/156), Exophiala species (3.8%, 6/156) and Scedosporium species (1.3%, 2/156). Forty out of 86 (46.5%) were identified for section Fumigati, 36 (41.9%) for section Flavi, 6 (7%) for section Nigri and 4 (4.6%) for section Terrei. Fourteen out of 40 A. fumigatus isolates were phenotypically resistant. The overall proportion of ARAf in total fungal isolates was 9% (14/156). cyp51A gene analysis in resistant isolates revealed that 13 isolates harboured G448S, G432C, T289F, D255E, M220I, M172V, G138C, G54E and F46Y mutations and one isolate carried G448S, G432C, T289F, D255E, M220I, G138C, G54E and F46Y mutations. Additionally, this study detects two novel cyp51A single-nucleotide polymorphisms (I242V and D490E).
Conclusions: This study first investigated ARAf isolates in Iranian CF patients. Due to a resistance rate of up to 9%, it is recommended that susceptibility testing of Aspergillus isolates from CF patients receiving antifungal treatment be a part of the routine diagnostic workup. However, extensive multicentre studies with a high volume of CF patients are highly warranted to determine the impact of ARAf on CF patients.
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