Assessing resistance to the echinocandin antifungal drug caspofungin in Candida albicans by profiling mutations in FKS1
Assessing resistance to the echinocandin antifungal drug caspofungin in Candida albicans by profiling mutations in FKS1
复制标题
DOI：
10.1128/aac.01653-05
复制
发表时间：
2006-06-01
影响因子：
4.9
通讯作者：
Perlin, David S.
中科院分区：
文献类型：
--
作者：
Balashov, Sergey V.;Park, Steven;Perlin, David S.
白色念珠菌临床分离株对棘白菌素药物卡泊芬净的耐药性正在缓慢出现，并与FKS 1基因中短保守区域的突变有关。最突出的变化发生在丝氨酸645位的Fks 1 p与脯氨酸，酪氨酸和苯丙氨酸的取代。建立了一种针对FKS 1基因突变的等位基因特异性实时荧光PCR分子信标检测方法，用于快速鉴定耐药。改变丝氨酸645的突变在杂合和纯合状态下都被可靠地鉴定。用分子信标试验评价了两个大的收集自不同菌株的自发突变株。对卡泊芬净耐药的白色念珠菌（MIC> 16 μ g/ml），目的是了解FKS 1突变与棘白菌素耐药之间的关系。在回收的85株耐药分离株中，所有分离株均被鉴定为FKSI突变; 93%在Ser 645处显示变化，其中62%显示特征性S645 P置换，表示为FKS 1中的纯合或杂合突变。另外两个显著的氨基酸置换S645 Y和S645 F的频率分别为22%和8%。还鉴定了三个新的突变：T1922 C、G1932 T和C1934 G，分别编码F641 S、L 644 F和S645 C取代。一个菌株具有L 644 F和S645 C的双氨基酸取代。等位基因特异性探针在多重测定中组合，用于可靠地筛选已知的FKSI突变。这些数据支持FKS 1 p取代在棘白菌素抗性中的重要性，并证明了将分子筛选应用于常规抗性评估的可行性。
Resistance of clinical isolates of Candida albicans to the echinocandin drug caspofungin is slowly emerging and is linked to mutations in short conserved regions in the FKS1 gene. The most prominent changes occurred at the serine 645 position in Fks1p with substitutions of proline, tyrosine, and phenylalanine. An allele-specific real-time PCR molecular-beacon assay was developed for rapid identification of drug resistance by targeting FKS1 mutations. Mutations altering serine 645 were reliably identified in both heterozygous and homozygous states. The molecular-beacon assay was used to evaluate two large collections of spontaneous mutants from separate strains of C. albicans with resistance (MICs, > 16 mu g/ml) to caspofungin with the goal of understanding the relationship between FKS1 mutations and echinocandin resistance. Of 85 resistant isolates recovered, all were identified with mutations in FKSI; 93% showed changes at Ser645, with 62% displaying a characteristic S645P substitution expressed as either a homozygous or a heterozygous mutation in FKS1 Two other prominent amino acid substitutions, S645Y and S645F, were found at frequencies of 22% and 8%, respectively. Three new mutations were also identified: T1922C, G1932T, and C1934G, encoding F641S, L644F, and S645C substitutions, respectively. One strain had the double amino acid substitution L644F and S645C. Allele-specific probes were combined in a multiplex assay for reliable screening of known FKSI mutations. These data support the importance of FKS1p substitutions in echinocandin resistance and demonstrate the feasibility of applying molecular screening for routine resistance assessment.