A 23 bp cyp51A Promoter Deletion Associated With Voriconazole Resistance in Clinical and Environmental Isolates of Neocosmospora keratoplastica

Abstract

In the fungal pathogen Aspergillus fumigatus, resistance to azole antifungals is often linked to mutations in CYP51A, a gene that encodes the azole antifungal drug target lanosterol 14α-demethylase. The aim of this study was to investigate whether similar changes could be associated with azole resistance in a Malaysian Fusarium solani species complex (FSSC) isolate collection. Most (11 of 15) clinical FSSC isolates were Neocosmospora keratoplastica and the majority (6 of 10) of environmental isolates were Neocosmospora suttoniana strains. All 25 FSSC isolates had high minimum inhibitory concentrations (MICs) for itraconazole and posaconazole, low MICs for amphotericin B, and various (1 to >32 mg/l) voriconazole susceptibilities. There was a tight association between a 23 bp CYP51A promoter deletion and high (>32 mg/l) voriconazole MICs; of 19 FSSC strains sequenced, nine isolates had voriconazole MICs > 32 mg/l, and they all contained the 23 bp CYP51A promoter deletion, although it was absent in the ten remaining isolates with low (≤12 mg/l) voriconazole MICs. Surprisingly, this association between voriconazole resistance and the 23 bp CYP51A promoter deletion held true across species boundaries. It was randomly distributed within and across species boundaries and both types of FSSC isolates were found among environmental and clinical isolates. Three randomly selected N. keratoplastica isolates with low (≤8 mg/l) voriconazole MICs had significantly lower (1.3-7.5 times) CYP51A mRNA expression levels than three randomly selected N. keratoplastica isolates with high (>32 mg/l) voriconazole MICs. CYP51A expression levels, however, were equally strongly induced (∼6,500-fold) by voriconazole in two representative strains reaching levels, after 80 min of induction, that were comparable to those of CYP51B. Our results suggest that FSSC isolates with high voriconazole MICs have a 23 bp CYP51A promoter deletion that provides a potentially useful marker for voriconazole resistance in FSSC isolates. Early detection of possible voriconazole resistance is critical for choosing the correct treatment option for patients with invasive fusariosis.

PDR Transporter ABC1 Is Involved in the Innate Azole Resistance of the Human Fungal Pathogen Fusarium keratoplasticum

“…F. keratoplasticum 2781, a clinical isolate that is resistant (MIC >32 mg/L) to itraconazole (ITC), posaconazole (PSC) and voriconazole (VRC) (James et al, 2020), was grown on potato dextrose agar (PDA; Oxoid Ltd., Hampshire, United Kingdom) plates incubated at 28 • C for 4-7 days. S. cerevisiae AD (Sagatova et al, 2015) was grown on yeast extract peptone dextrose [YPD; 1% w/v yeast extract (Formedium Ltd., Norfolk, United Kingdom), 2% w/v peptone (Formedium), and 2% w/v glucose (Formedium)] agar plates incubated at 30 • C for ∼3 days.…”

“…CYP51A and CYP51B are sterol 14-α demethylases, but the function of CYP51C remains unknown (Fan et al, 2013;Zheng et al, 2019). We have recently reported a 23 bp CYP51A promoter deletion that was associated with increased VRC resistance in both clinical and environmental FSSC isolates (James et al, 2020).…”

“…Species of the Fusarium solani species complex (FSSC) are ubiquitous moulds often detected in soil or on plants (Nalim et al, 2011;James et al, 2020) and in indoor plumbing drains (Short et al, 2014). They can infect plants (Coleman et al, 2009), animals (O'Donnell et al, 2016), and humans (Horn et al, 2014).…”

“…Fusarium keratoplasticum and Fusarium falciforme, two terrestrial FSSC species, have caused fusariosis of endangered sea turtle eggs (Sarmiento-Ramirez et al, 2014;Gleason et al, 2020). F. keratoplasticum and Fusarium petroliphilum are the two FSSC species most frequently isolated in the clinic (Herkert et al, 2019;James et al, 2020). FSSC infections are among the most common opportunistic nosocomial mould infections after infections caused by Aspergilli (Alastruey- Izquierdo et al, 2013;Lass-Florl and Cuenca-Estrella, 2017).…”

“…Aspergilli have two CYP51 orthologues (CYP51A and CYP51B) (Mellado et al, 2001) whereas Fusarium spp. have three CYP51 orthologues (CYP51A, CYP51B and CYP51C) (Becher et al, 2011;Zheng et al, 2019;James et al, 2020). The CYP51C orthologue is a unique characteristic of this fungal genus.…”