Molecular mechanisms of acquired antifungal drug resistance in principal fungal pathogens and EUCAST guidance for their laboratory detection and clinical implications

Abstract

The increasing incidence and changing epidemiology of invasive fungal infections continue to present many challenges to their effective management. The repertoire of antifungal drugs available for treatment is still limited although there are new antifungals on the horizon. Successful treatment of invasive mycoses is dependent on a mix of pathogen-, host- and antifungal drug-related factors. Laboratories need to be adept at detection of fungal pathogens in clinical samples in order to effectively guide treatment by identifying isolates with acquired drug resistance. While there are international guidelines on how to conduct in vitro antifungal susceptibility testing, these are not performed as widely as for bacterial pathogens. Furthermore, fungi generally are recovered in cultures more slowly than bacteria, and often cannot be cultured in the laboratory. Therefore, non-culture-based methods, including molecular tests, to detect fungi in clinical specimens are increasingly important in patient management and are becoming more reliable as technology improves. Molecular methods can also be used for detection of target gene mutations or other mechanisms that predict antifungal drug resistance. This review addresses acquired antifungal drug resistance in the principal human fungal pathogens and describes known resistance mechanisms and what in-house and commercial tools are available for their detection. It is emphasized that this approach should be complementary to culture-based susceptibility testing, given the range of mutations, resistance mechanisms and target genes that may be present in clinical isolates, but may not be included in current molecular assays.

Figure 1.

Amino acid (AA) sequences of Fks1 and Fks2 in 10 WT Candida species. Amino acid codons associated with increased MIC are underlined and in bold font. In the online version a colour indication is applied to inform origin (naturally occurring or acquired) and impact on the MIC (strong, weak or silent). Red: ‘strong R’ mutation, subscript at codons involving a mutation or deletion; superscript at codon involving a mutation or stop codon. Yellow: ‘weak R’ mutation. Blue: inherent AA difference with proven or possible relation to intrinsic lower susceptibility. Grey: inherent AA difference of unknown importance. Green: inherent AA difference, probably with no effect. ^aOf note: combination of the following alterations outside the defined hotspots has also been confirmed as cause of echinocandin resistance: Fks1 W508stop combined with Fks2 E655K. ECOFFs indicated in () are estimated WT upper limits (peak MIC + 2 dilutions) based upon the MICs of Danish blood isolates. *Inaccurate annotation, sequencing of entire gene-sequence required. #The micafungin (but not anidulafungin) ECOFF for C. krusei is noticeably higher (0.25 mg/L) than for C. albicans (0.015 mg/L) and C. glabrata (0.03 mg/L). NA, not available. This figure appears in colour in the online version of JAC and in black and white in the print version of JAC.