Detection of ERG11 point mutations in Iranian fluconazoleresistant Candida albicans isolates

Background and Purpose: Candidiasis is referred to a group of superficial anddeep-tissue fungal infections often caused by Candida albicans. The superficialinfections affect the oral, oropharynx, esophagus, and vaginal mucosa. The treatmentof choice for these infections is the use of azoles, such as fluconazole. However, theincreased use of these antifungal agents has led to the emergence of azole-resistantisolates of C. albicans. Different mechanisms have been suggested for thedevelopment of drug resistance, such as mutations in the encoding gene ERG11.Mutations in ERG11 result in changes in the ERG11p spatial construction and reducethe affinity between the protein and azole. This study aimed to determine thesusceptibility profile of C. albicans clinical isolates to fluconazole usingmicrodilution method. The present research was also targeted toward the detection ofmutations that might be related to fluconazole resistance by the amplification andsequencing of ERG11 gene.Materials and Methods: This study was conducted on a total of 216 clinical isolatesobtained from Mashhad, Isfahan, and Tehran cities in Iran, during 2016-2018. Theclinical isolates were identified using molecular techniques. Furthermore, minimuminhibitory concentration (MICs) was determined according to the clinical and laboratorystandards institute M27-A3 and M27-S4 documents. The concentration range forfluconazole was obtained as 0.063-64 μg/ml. In the resistant strains, ERG11 genes wereamplified by specific primers. Subsequently, cycle sequencing reactions were performedon purified polymerase chain reaction (PCR) products in forward and reverse directions.Finally, the results were analyzed by MEGA (version 7) and Gene Runner software(version 6.5.30).Results: Out of 216 strains, 100 (46.3%) species were identified as C. albicans. TheMIC values for fluconazole had a range of 0.125-16 μg/ml with the MIC50 and MIC90values of 0.5 and 1 μg/ml, respectively. Totally, 41 nucleotide changes were detectedamong 4 resistant isolates. In this regard, 4 out of 41 mutations in codons caused changesin ERG11p; however, these mutations did not lead to fluconazole resistance.Conclusion: Fluconazole resistance among clinical isolates is not merely due to thechanges in ERG11p. This resistance may be also related to some other mechanisms, suchas the prevention of the intracellular accumulation of the antifungal agent and alterationof the target enzyme to diminish drug binding.