Reduced accumulation of drug in Candida krusei accounts for itraconazole resistance

Abstract

Due to intrinsic resistance Candida krusei is emerging as a systemic pathogen in AIDS patients undergoing fluconazole therapy, but acquired resistance to itraconazole has not been studied biochemically. We report here studies on the basis for azole resistance and sterol composition in C. krusei. An itraconazole-resistant isolate showed reduced susceptibility to azole drugs in in vitro growth inhibition studies. Accumulation of 14␣-methyl-3,6-diol under azole treatment was associated with growth arrest. In vitro ergosterol biosynthesis and type II binding studies suggested no alteration in the affinity to azole drugs of the target enzyme, the cytochrome P-450 sterol 14␣-demethylase, in the resistant isolate. Resistance was associated with a decreased intracellular content of drug in the resistant isolate.

Identification and expression of multidrug resistancerelated ABC transporter genes in Candida krusei

“…The relative insensitivity of C. krusei lanosterol demethylase to uconazole as reported by Orozco et al [9] may be suf cient to explain the intrinsic resistance of this organism. However, our data do indicate a potential role for C. krusei MDR upregulation in acquired azole resistance; e.g., the itraconazole resistance of a clinical isolate which was correlated with decreased accumulation [7]. It will be of interest to apply the sequences reported here to studies of ABC transporter expression in C. krusei clinical isolates with varying azole susceptibility.…”

“…Conversely, relatively large differences in intracellular accumulation of these azoles were noted, which correlate with their different effects on growth [4]. Reduced accumulation was similarly identi ed by Venkateswarlu et al [7] as the basis for itraconazole resistance in a clinical isolate. In contrast, Orozco et al [9] observed a large difference in the cellfree inhibition by uconazole of C. krusei and C. albicans lanosterol demethylase and no signi cant difference in uconazole accumulation.…”

“…Not surprisingly, the increased use in recent years of uconazole for therapy and prophylaxis has been associated with an increased incidence of C. krusei infection [6]. Itraconazole represents an alternative agent; however, itraconazole-resistant strains have been reported [7]. The basis for azole resistance in C. krusei has been examined in several studies, with somewhat contradictory results.…”

Regulation of multidrug resistance in pathogenic fungi

“…Several early investigations had shown that azole resistance in clinical Candida isolates often correlated with decreased intracellular drug accumulation, either by reduced uptake or by increased efflux (Clark et al, 1996;Hitchcock et al, 1993;Joseph-Horne et al, 1995;Lamb et al, 1997;Parkinson et al, 1995;Ryley et al, 1984;Venkateswarlu et al, 1995Venkateswarlu et al, , 1996. Sanglard and coworkers were the first to demonstrate that an increased expression of efflux pumps encoded by CDR1, CDR2, and MDR1 was associated with reduced intracellular fluconazole accumulation and azole resistance of clinical C. albicans isolates (Sanglard et al, 1995(Sanglard et al, , 1997.…”

“…Candida krusei is intrinsically resistant to fluconazole, due to the low affinity of its sterol 14a-demethylase for this drug (Orozco et al, 1998), and can acquire resistance to other azoles by reduced intracellular drug accumulation (Marichal et al, 1995;Venkateswarlu et al, 1996). Two partial genes encoding ABC transporters were cloned from C. krusei genomic DNA by PCR with degenerate primers and designated as ABC1 and ABC2 (Katiyar and Edlind, 2001).…”