Summary: Background: Azole resistance in Aspergillus fumigatus is an emerging problem and reported from all continents. As triazole antifungals are the mainstay of therapy in the management of invasive aspergillosis, azole‐resistant A fumigatus has become a major medical concern and with complicated clinical management.Objective: Screening of environmental presence of azole‐resistant A fumigatus in Iran.Methods: Compost from Northern Iran, collected between 2017 and 2018, was screened for the presence of azole‐resistant A fumigatus with azole‐containing agar. Phenotypic MICs were obtained from selected, molecularly confirmed isolates. cyp51A gene sequencing and genotyping of azole‐resistant isolates were done.Results: Among 300 compost samples, three A fumigatus isolates had high voriconazole MICs (≥16 mg/L) and harboured the TR₄₆/Y121F/T289A mutation in the cyp51A gene. Microsatellite typing of these isolates showed that two strains had the same allele across all nine examined microsatellite loci and were genotypically related to Indian azole‐resistant strains. The other isolate had a different genotype.Conclusion: This is the first report of A fumigatus with TR₄₆/Y121F/T289A mutation from the region. Monitoring and surveillance of antifungal susceptibility of clinical A fumigatus is warranted in Iran and elsewhere in the region.

Azole-resistance in Aspergillus fumigatus is a worldwide medical concern complicating the management of aspergillosis (IA). Herein, we report the clonal spread of environmental triazole resistant A. fumigatus isolates in Iran. In this study, 63 A. fumigatus isolates were collected from 300 compost samples plated on Sabouraud dextrose agar supplemented with itraconazole (ITR) and voriconazole (VOR). Forty-four isolates had the TR₃₄/L98H mutation and three isolates a TR₄₆/Y121F/T289A resistance mechanism, while two isolates harbored a M172V substitution in cyp51A. Fourteen azole resistant isolates had no mutations in cyp51A. We found that 41 out of 44 A. fumigatus strains with the TR₃₄/L98H mutation, isolated from compost in 13 different Iranian cities, shared the same allele across all nine examined microsatellite loci. Clonal expansion of triazole resistant A. fumigatus in this study emphasizes the importance of establishing antifungal resistance surveillance studies to monitor clinical Aspergillus isolates in Iran, as well as screening for azole resistance in environmental A. fumigatus isolates.

Samples from a total of 67 stations, distributed amongst 32 cities along the Caspian Sea coastline, were collected during the summer of 2021 on sunny days. The samples were collected from each station, including both dry/wet sand and shoreline water. The grown samples were primarily analyzed for the macro/microscopic morphologic features of the fungi. Moreover, identification by PCR-RFLP was performed for yeasts, dermatophytes, and Aspergillus sp. strains. Antifungal susceptibility tests were performed for probable-isolated Aspergillus and Candida sp. A total of 268 samples were collected, from which 181 (67.54%) isolates were recovered. Yeast-like fungi and potential pathogenic black fungi were detected in 12 (6.6%) and 20 (11%) of the sand (dry/wet) samples. Potential pathogenic hyaline fungi were identified in 136 (75.1%) samples, in which Aspergillus sp. was the predominant genus and was detected in 76/136 (47.8%) samples as follows: A. section Flavi n = 44/76 (57.9%), A. section Nigri n = 19/76 (25%), A. section Nidulantes n = 9/76 (11.8%), and A. section Fumigati n = 4/76 (5.3%). The most effective azole antifungal agent was different per section: in A. section Fumigati, PSZ; in Aspergillus section Nigri, ITZ and ISZ; in A. section Flavi, EFZ; and in A. section Nidulantes, ISZ. Candida isolates were susceptible to the antifungals tested.

Background and Purpose: Because of the growing incidence of Aspergillus infection,typing methods of Aspergillus species are increasingly being used. Accordingly,studying the spread and population dynamics of strains isolating from clinical and environment, from a single host to large scale ecosystems is definitely needed. In the current study, we carried out a genetic analysis of nine microsatellite loci in isolates from different regions of Iran to compare and explore the genetic diversity between environmental and clinical A. fumigatus strains. Materials and Methods: Sixty-six clinical (n=43) and environmental (n= 23) isolates of A. fumigatus, have collected from six cities of Iran. All A. fumigatus isolates identified based on macroscopic and microscopic characters, the ability to grow at above 45°C, and confirmed using DNA sequencing of the partial b-tubulin gene.Sixty-six A. fumigatus isolates were subjected by microsatellite typing using three separate multiplex PCRs with a panel of nine short tandem repeats (STR) to evaluate the genetic relatedness. Results: The STR typing of 66 A. fumigatus isolates revealed 38 distinct genotypes distributed among environmental and clinical isolates. We identified 12 clones including 40 different isolates representing 60% of all isolates tested, which each clone included 2–7 isolates. Conclusion: The STR typing is considered as a valuable tool with excellent discriminatory power to study the molecular epidemiology and genotypic diversity of A.fumigatus isolates. These findings show that the high genetic diversity observed of Iranian A. fumigatus isolates with those outside Iran and formed a separate cluster.

Background: The overexpression of the efflux transporter genes is one of the important mechanisms of resistance in fungal pathogens such as Candida and Aspergillus species. Objective: Here, the expression alterations of drug efflux transporter genes were evaluated in non- Cyp51A voriconazole-resistant Aspergillus fumigatus isolates. Methods: Six A. fumigatus isolates including four voriconazole-resistant isolates with and without azole-resistance-related mutations in addition to two susceptible A. fumigatus isolates were selected from 300 previously characterized A. fumigatus clinical and environmental isolates, received during 2013-2015. In order to extract RNA, the minimum inhibitory concentrations (MICs) for the isolates were determined according to the broth microdilution protocol regarding the Clinical and Laboratory Standards Institute document M38-A2 (CLSI, 2008). Alteration in the expression of AfuMDR1, AfuMDR2, AfuMDR3, AfuMDR4, Cyp51A, and atrF was studied using the real-time polymerase chain reaction assay. Results: Based on REST® output, significant overexpression of atrF, AfuMDR1, AfuMDR3, and AfuMDR4/Cyp51A, atrF, AfuMDR2, AfuMDR4 genes was observed in the isolates without azoleresistance- related mutations, respectively. No significant overexpression was seen in the isolates with T34/L98H except for the AfuMDR3 and AfuMDR4(P<0.05). Conclusion: Our results support the hypothesis that efflux pump transporters can contribute to voriconazole resistance in A. fumigatus.