title: Fluconazole treatment is effective against a Candida albicans erg3/erg3 mutant in vivo despite in vitro resistance.
abstract: Candida albicans ERG3 encodes a sterol C5,6-desaturase which is essential for synthesis of ergosterol. Defective sterol C5,6 desaturation has been considered to be one of the azole resistance mechanisms in this species. However, the clinical relevance of this resistance mechanism is still unclear. In this study, we created a C. albicans erg3/erg3 mutant by the "Ura-blaster" method and confirmed the expected azole resistance using standard in vitro testing and the presence of ergosta-7,22-dien-3beta-ol instead of ergosterol. For in vivo studies, a wild-type URA3 was placed back into its native locus in the erg3 homozygote to avoid positional effects on URA3 expression. Defective hyphal formation of the erg3 homozygote was observed not only in vitro but in kidney tissues. A marked attenuation of virulence was shown by the longer survival and the lower kidney burdens of mice inoculated with the reconstituted Ura+ erg3 homozygote relative to the control. To assess fluconazole efficacy in a murine model of disseminated candidiasis, inoculum sizes of the control and the erg3 homozygote were chosen which provided a similar organ burden. Under these conditions, fluconazole was highly effective in reducing the organ burden in both groups. This study demonstrates that an ERG3 mutation causing inactivation of sterol C5,6-desaturase cannot confer fluconazole resistance in vivo by itself regardless of resistance measured by standard in vitro testing. The finding questions the clinical significance of this resistance mechanism.
authors.affiliation: Second Department of Internal Medicine, Nagasaki University School of Medicine, 1-7-1 Sakamoto, Nagasaki 852-8501, Japan.
authors.forename: Taiga
authors.initials: T
authors.lastname: Miyazaki
authors.forename: Yoshitsugu
authors.initials: Y
authors.lastname: Miyazaki
authors.forename: Koichi
authors.initials: K
authors.lastname: Izumikawa
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kakeya
authors.forename: Shunichi
authors.initials: S
authors.lastname: Miyakoshi
authors.forename: John E
authors.initials: JE
authors.lastname: Bennett
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kohno
mesh_terms: D000818:Animals; D000935:Antifungal Agents / Q000627:therapeutic use*; D002176:Candida albicans / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology / Q000472:pathogenicity; D002177:Candidiasis / Q000188:drug therapy* / Q000473:pathology; D025141:Drug Resistance, Fungal; D005260:Female; D015725:Fluconazole / Q000627:therapeutic use*; D051379:Mice; D008807:Mice, Inbred BALB C; D008826:Microbial Sensitivity Tests; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics / Q000502:physiology*; D013261:Sterols / Q000032:analysis; D014774:Virulence
chemical_list: D000935:Antifungal Agents; D013261:Sterols; D015725:Fluconazole; D010088:Oxidoreductases; C039306:sterol delta-5 desaturase