Cryptic Species and Azole Resistance in the Aspergillus niger Complex

Abstract

ABSTRACTAspergillus nigeris a common clinical isolate. Multiple species comprise theAspergillussectionNigriand are separable using sequence data. The antifungal susceptibility of these cryptic species is not known. We determined the azole MICs of 50 black aspergilli, 45 from clinical specimens, using modified EUCAST (mEUCAST) and Etest methods. Phylogenetic trees were prepared using the internal transcribed spacer, beta-tubulin, and calmodulin sequences to identify strains to species level and the results were compared with those obtained withcyp51Asequences. We attempted to correlatecyp51Amutations with azole resistance. Etest MICs were significantly different from mEUCAST MICs (P< 0.001), with geometric means of 0.77 and 2.79 mg/liter, respectively. Twenty-six of 50 (52%) isolates were itraconazole resistant by mEUCAST (MICs > 8 mg/liter), with limited cross-resistance to other azoles. Using combined beta-tubulin/calmodulin sequences, the 45 clinical isolates grouped into 5 clades,A. awamori(55.6%),A. tubingensis(17.8%),A. niger(13.3%),A. acidus(6.7%), and an unknown group (6.7%), none of which were morphologically distinguishable. Itraconazole resistance was found in 36% of the isolates in theA. awamorigroup, 90% of theA. tubingensisgroup, 33% of theA. nigergroup, 100% of theA. acidusgroup, and 67% of the unknown group. These data suggest thatcyp51Amutations in sectionNigrimay not play as important a role in azole resistance as inA. fumigatus, although some mutations (G427S, K97T) warrant further study. Numerous cryptic species are found in clinical isolates of theAspergillussectionNigriand are best reported as “A. nigercomplex” by clinical laboratories. Itraconazole resistance was common in this data set, but azole cross-resistance was unusual. The mechanism of resistance remains obscure.