Public Health Research Institute, New Jersey Medical School-UMDNJ, Newark, NJ 07103-3535, USA.
Antimicrob Agents Chemother. 2011 May;55(5):2245-55. doi: 10.1128/AAC.01350-10. Epub 2011 Mar 7.
Echinocandins are highly bound to serum proteins, altering their antifungal properties. The addition of 50% human serum to the MIC assay improves the identification of echinocandin-resistant Candida spp. harboring fks hot spot mutations. However, this modification cannot readily be applied to the method of the CLSI M27-A3 document due to safety and standardization difficulties. The aim of this study was to evaluate commercial bovine serum albumin (BSA) as a safe and standardized alternative to human serum. A collection of 28 echinocandin-susceptible strains, 10 Candida parapsilosis sensu lato strains (with naturally reduced echinocandin susceptibility), and 40 FKS hot spot mutants was used in this work. When RPMI 1640 was used for susceptibility testing, wild-type strains and fks mutants showed MIC range overlaps (-2, -1, and -3 2-fold-dilution steps separated these populations for anidulafungin, caspofungin, and micafungin, respectively). On the other hand, the addition of BSA to RPMI 1640 differentially increased echinocandin MIC values for these groups of strains, allowing better separation between populations, with no MIC range overlaps for any of the echinocandin drugs tested. Moreover, the use of RPMI-BSA reduced the number of fks hot spot mutant isolates for which MIC values were less than or equal to the upper limit for the wild type (very major errors) from 9, 2, and 7 with RPMI alone to 3, 0, and 3 for anidulafungin, caspofungin, and micafungin, respectively. When RPMI-BSA was used to study the susceptibility of C. parapsilosis sensu lato species to echinocandins, the strains behaved as anidulafungin- and micafungin-resistant isolates (MIC, ≥8 μg/ml). These data support the need for a revision of the CLSI protocol for in vitro testing of echinocandin susceptibility in order to identify all or most of the fks hot spot mutants. Also, caspofungin could be used as a surrogate marker of reduced susceptibility to echinocandins.
棘白菌素类药物与血清蛋白高度结合，改变其抗真菌特性。将 50%人血清加入 MIC 测定中，可以提高对携带 fks 热点突变的棘白菌素类耐药念珠菌的鉴定。然而，由于安全性和标准化方面的困难，这种改良不能轻易应用于 CLSI M27-A3 文件中的方法。本研究旨在评估商业牛血清白蛋白（BSA）作为一种安全且标准化的人血清替代物。本研究使用了 28 株棘白菌素类敏感株、10 株近平滑念珠菌（具有天然降低的棘白菌素类敏感性）和 40 株 FKS 热点突变株。当使用 RPMI 1640 进行药敏试验时，野生型菌株和 fks 突变株的 MIC 范围存在重叠（阿尼达霉素、卡泊芬净和米卡芬净分别用 2 倍稀释步骤区分了这两种群体）。另一方面，BSA 对 RPMI 1640 的添加，使这些菌株群体的棘白菌素类 MIC 值有差异增加，更好地分离了不同群体，在测试的任何一种棘白菌素类药物中，MIC 范围均无重叠。此外，使用 RPMI-BSA 可减少 MIC 值等于或低于野生型上限（非常大的错误）的棘白菌素类 FKS 热点突变株分离株的数量，从单独使用 RPMI 时的 9、2 和 7 株，分别减少到阿尼达霉素、卡泊芬净和米卡芬净的 3、0 和 3 株。当使用 RPMI-BSA 研究近平滑念珠菌对棘白菌素类药物的敏感性时，这些菌株表现为阿尼达霉素和米卡芬净耐药分离株（MIC≥8μg/ml）。这些数据支持需要修订 CLSI 体外棘白菌素类药敏试验协议，以鉴定所有或大多数 FKS 热点突变株。此外，卡泊芬净可作为棘白菌素类药物敏感性降低的替代标志物。
