vanden Bossche H, Marichal P, Odds F C, Le Jeune L, Coene M C
Division of Medicinal Chemistry and Pharmacology, Janssen Research Foundation, Beerse, Belgium.
Antimicrob Agents Chemother. 1992 Dec;36(12):2602-10. doi: 10.1128/AAC.36.12.2602.
A Candida (Torulopsis) glabrata strain (B57149) became resistant to fluconazole after a patient carrying the organism was treated with the drug at 400 mg once daily for 9 days. Growth of the pretreatment isolate (B57148) was inhibited by 50% with 0.67 microM ketoconazole, 1.0 microM itraconazole, and 43 microM fluconazole, whereas growth of B57149 was inhibited slightly by 10 microM ketoconazole but was unaffected by 10 microM itraconazole or 100 microM fluconazole. This indicates cross-resistance to all three azole antifungal agents. The cellular fluconazole content of B57149 was from 1.5- to 3-fold lower than that of B57148, suggesting a difference in drug uptake between the strains. However, this difference was smaller than the measured difference in susceptibility and, therefore, cannot fully explain the fluconazole resistance of B57149. Moreover, the intracellular contents of ketoconazole and itraconazole differed by less than twofold between the strains, so that uptake differences did not account for the azole cross-resistance of B57149. The microsomal cytochrome P-450 content of B57149 was about twice that of B57148, a difference quantitatively similar to the increased subcellular ergosterol synthesis from mevalonate or lanosterol. These results indicate that the level of P-450-dependent 14 alpha-demethylation of lanosterol is higher in B57149. Increased ergosterol synthesis was also seen in intact B57149 cells, and this coincided with a decreased susceptibility of B57149 toward all three azoles and amphotericin B. B57149 also had higher squalene epoxidase activity, and thus, more terbinafine was needed to inhibit the synthesis of 2,3-oxidosqualene from squalene. P-450 content and ergosterol synthesis both decreased when isolate B57149 was subcultured repeatedly on drug-free medium. This repeated subculture also fully restored the strain's itraconazole susceptibility, but only partly increased its susceptibility to fluconazole. The results suggest that both lower fluconazole uptake and increased P-450-dependent ergosterol synthesis are involved in the mechanism of fluconazole resistance but that only the increased ergosterol synthesis contributes to itraconazole cross-resistance.
一名携带光滑念珠菌（球拟酵母菌）菌株（B57149）的患者每天服用400毫克氟康唑，持续9天进行治疗后，该菌株对氟康唑产生了耐药性。治疗前分离出的菌株（B57148），其生长受到0.67微摩尔酮康唑、1.0微摩尔伊曲康唑和43微摩尔氟康唑的抑制，抑制率达50%；而B57149的生长仅受到10微摩尔酮康唑的轻微抑制，不受10微摩尔伊曲康唑或100微摩尔氟康唑的影响。这表明该菌株对所有三种唑类抗真菌药物均产生了交叉耐药性。B57149的细胞氟康唑含量比B57148低1.5至3倍，提示这两种菌株在药物摄取方面存在差异。然而，这种差异小于所测得的药敏差异，因此无法完全解释B57149对氟康唑的耐药性。此外，两种菌株间酮康唑和伊曲康唑的细胞内含量差异不到两倍，所以摄取差异并不能解释B57149的唑类交叉耐药性。B57149微粒体细胞色素P - 450的含量约为B57148的两倍，这一差异在数量上与甲羟戊酸或羊毛甾醇亚细胞麦角固醇合成增加相似。这些结果表明，B57149中羊毛甾醇P - 450依赖性14α - 去甲基化水平较高。完整的B57149细胞中也观察到麦角固醇合成增加，这与B57149对所有三种唑类药物及两性霉素B的敏感性降低相吻合。B57149还具有较高的角鲨烯环氧化酶活性，因此，需要更多的特比萘芬才能抑制角鲨烯合成2,3 - 氧化角鲨烯。当将分离菌株B57149在无药培养基上反复传代培养时，P - 450含量和麦角固醇合成均降低。这种反复传代培养也使该菌株对伊曲康唑的敏感性完全恢复，但仅部分提高了其对氟康唑的敏感性。结果表明，氟康唑摄取降低和P - 450依赖性麦角固醇合成增加均参与了氟康唑耐药机制，但只有麦角固醇合成增加导致了伊曲康唑交叉耐药性。
