AUTHOR=Daneshnia Farnaz , Hilmioğlu Polat Süleyha , Ilkit Macit , Shor Erika , de Almeida Júnior João Nobrega , Favarello Larissa M. , Colombo Arnaldo Lopes , Arastehfar Amir , Perlin David S. 

TITLE=Determinants of fluconazole resistance and the efficacy of fluconazole and milbemycin oxim combination against Candida parapsilosis clinical isolates from Brazil and Turkey

JOURNAL=Frontiers in Fungal Biology

VOLUME=3

YEAR=2022

URL=https://www.frontiersin.org/journals/fungal-biology/articles/10.3389/ffunb.2022.906681

DOI=10.3389/ffunb.2022.906681

ISSN=2673-6128

ABSTRACT=<p>Fluconazole-resistant <italic>Candida parapsilosis</italic> (FLZR-CP) outbreaks are a growing public health concern and have been reported in numerous countries. Patients infected with FLZR-CP isolates show fluconazole therapeutic failure and have a significantly increased mortality rate. Because fluconazole is the most widely used antifungal agent in most regions with outbreaks, it is paramount to restore its antifungal activity. Milbemycin oxim (MOX), a well-known canine endectocide, is a potent efflux pump inhibitor that significantly potentiates the activity of fluconazole against FLZR <italic>C</italic>. <italic>glabrata</italic> and <italic>C</italic>. <italic>albicans.</italic> However, the FLZ-MOX combination has not been tested against FLZR-CP isolates, nor is it known whether MOX may also potentiate the activity of echinocandins, a different class of antifungal drugs. Furthermore, the extent of involvement of efflux pumps <italic>CDR1</italic> and <italic>MDR1</italic> and ergosterol biosynthesis enzyme <italic>ERG11</italic> and their link with gain-of-function (GOF) mutations in their transcription regulators (<italic>TAC1</italic>, <italic>MRR1</italic>, and <italic>UPC2</italic>) are poorly characterized among FLZR-CP isolates. We analyzed 25 C. <italic>parapsilosis</italic> isolates collected from outbreaks in Turkey and Brazil by determining the expression levels of <italic>CDR1</italic>, <italic>MDR1</italic>, and <italic>ERG11</italic>, examining the presence of potential GOF mutations in their transcriptional regulators, and assessing the antifungal activity of FLZ-MOX and micafungin-MOX against FLZR and multidrug-resistant (MDR) <italic>C</italic>. <italic>parapsilosis</italic> isolates. <italic>ERG11</italic> was found to be universally induced by fluconazole in all isolates, while expression of <italic>MDR1</italic> was unchanged. Whereas mutations in <italic>MRR1</italic> and <italic>UPC2</italic> were not detected, <italic>CDR1</italic> was overexpressed in three Brazilian FLZR-CP isolates, which also carried a novel <italic>TAC1<sup>L518F</sup></italic> mutation. Of these three isolates, one showed increased basal expression of <italic>CDR1</italic>, while the other two overexpressed <italic>CDR1</italic> only in the presence of fluconazole. Interestingly, MOX showed promising antifungal activity against FLZR isolates, reducing the FLZ MIC 8- to 32-fold. However, the MOX and micafungin combination did not exert activity against an MDR <italic>C</italic>. <italic>parapsilosis</italic> isolate. Collectively, our study documents that the mechanisms underpinning FLZR are region specific, where <italic>ERG11</italic> mutations were the sole mechanism of FLZR in Turkish FLZR-CP isolates, while simultaneous overexpression of <italic>CDR1</italic> was observed in some Brazilian counterparts. Moreover, MOX and fluconazole showed potent synergistic activity, while the MOX-micafungin combination showed no synergy.</p>