Synergistic antifungal activity of statin-azole associations as witnessed by Saccharomyces cerevisiae- and Candida utilis-bioassays and ergosterol quantification.
BACKGROUND
Frequent opportunist fungal infections and the resistance to available antifungal drugs promoted the development of new alternatives for treatment, like antifungal drug combinations.


OBJECTIVE
This work aimed to detect the antifungal synergism between statins and azoles by means of an agar-well diffusion bioassay with Saccharomyces cerevisiae ATCC 32051 and Candida utilis Pr(1-2) as test strains.


METHODS
Synergistic antifungal effects were tested by simultaneously adding a sub inhibitory concentration (SIC) of statin (atorvastatin, lovastatin, pravastatin, rosuvastatin or simvastatin) plus a minimal inhibitory concentration (MIC) of azole (clotrimazole, fluconazole, itraconazole, ketoconazole or miconazole) to yeast-embedded YNB agar plates, and a positive result corresponded to a yeast growth inhibition halo higher than that produced by the MIC of the azole alone. Yeast cell ergosterol quantification by RP-HPLC was used to confirm statin-azole synergism, and ergosterol rescue bioassays were performed for evaluating statin-induced ergosterol synthesis blockage.


RESULTS
Growth inhibition was significantly increased when clotrimazole, fluconazole, itraconazole, ketoconazole and miconazole were combined with atorvastatin, lovastatin, rosuvastatin and simvastatin. Highest growth inhibition increments were observed on S. cerevisiae (77.5%) and C. utilis (43.2%) with a SIC of simvastatin plus a MIC of miconazole, i.e. 4 + 2.4 μg/ml or 20 + 4.8 μg/ml, respectively. Pravastatin showed almost no significant effects (0-7.6% inhibition increase). Highest interaction ratios between antifungal agents corresponded to simvastatin-miconazole combinations and were indicative of synergism. Synergism was also confirmed by the increased reduction in cellular ergosterol levels (S. cerevisiae, 40% and C. utilis, 22%). Statin-induced ergosterol synthesis blockage was corroborated by means of ergosterol rescue bioassays, pravastatin being the most easily abolished inhibition whilst rosuvastatin being the most ergosterol-refractory.


CONCLUSIONS
Selected statin-azole combinations might be viable alternatives for the therapeutic management of mycosis at lower administration doses or with a higher efficiency.
PROIMI-CONICET (Planta Piloto de Procesos Industriales Microbiológicos), Tucumán, Argentina.
María Eugenia
ME
Cabral
Lucía I C
LI
Figueroa
Julia I
JI
Fariña
D000935:Antifungal Agents / Q000494:pharmacology*; D002175:Candida / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism; D052978:Disk Diffusion Antimicrobial Tests; D025141:Drug Resistance, Fungal; D004357:Drug Synergism; D004875:Ergosterol / Q000096:biosynthesis; D006801:Humans; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology*; D066298:In Vitro Techniques; D008826:Microbial Sensitivity Tests; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism; D013045:Species Specificity; D014230:Triazoles / Q000494:pharmacology*
D000935:Antifungal Agents; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D014230:Triazoles; D004875:Ergosterol