Rapid detection of ERG11 polymorphism associated azole resistance in Candida tropicalis.
Rapid detection of ERG11 polymorphism associated azole resistance in Candida tropicalis.
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DOI：
10.1371/journal.pone.0245160
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发表时间：
2021
期刊：
影响因子：
3.7
通讯作者：
Ghosh AK
中科院分区：
文献类型：
--
作者：
Paul S;Dadwal R;Singh S;Shaw D;Chakrabarti A;Rudramurthy SM;Ghosh AK
热带假丝酵母对唑类药物耐药的报道日益增多，迫切需要快速耐药检测技术的发展。羊毛甾醇C14 α-脱甲基酶（ERG 11）基因的非同义突变是引起念珠菌唑类耐药的主要机制之一。热带植物本研究评价了四引物扩增耐药突变系统-PCR（T-ARMS-PCR）、限制性酶切位点突变（RSM）和高分辨率熔解曲线（HRM）分析方法用于快速检测耐药念珠菌的ERG 11基因多态性。热带植物12株唑类耐药株和19株敏感株。包括热带。对分离株进行DNA测序，以检查耐药和敏感分离株之间的ERG 11多态性状态。对T-ARMS-PCR、RSM和HRM三种方法进行了评价和验证，用于快速检测ERG 11突变。氟康唑对12株耐药菌株和19株敏感菌株的MIC分别为32-256 mg/L和0.5 ~ 1 mg/L。耐药菌株在ERG 11基因中存在A339 T和C461 T突变。T-ARMS-PCR和RSM方法区分了所有的抗性和敏感菌株，而HRM分析区分了所有，除了一个敏感菌株。这三种方法的灵敏度、特异性、分析灵敏度、分析时间和分析成本表明，这三种方法可用于快速检测C.热带植物此外，注意到所有三种方法与DNA测序具有极好的一致性。T-ARMS-PCR、RSM和HRM方法是一种快速、灵敏、廉价的检测唑类药物耐药的方法。Tropicalis并可在临床设置中实施，以更好地进行患者管理。
Increasing reports of azole resistance in Candida tropicalis, highlight the development of rapid resistance detection techniques. Nonsynonymous mutations in the lanosterol C14 alpha-demethylase (ERG11) gene is one of the predominant mechanisms of azole resistance in C. tropicalis. We evaluated the tetra primer-amplification refractory mutation system-PCR (T-ARMS-PCR), restriction site mutation (RSM), and high-resolution melt (HRM) analysis methods for rapid resistance detection based on ERG11 polymorphism in C. tropicalis. Twelve azole-resistant and 19 susceptible isolates of C. tropicalis were included. DNA sequencing of the isolates was performed to check the ERG11 polymorphism status among resistant and susceptible isolates. Three approaches T-ARMS-PCR, RSM, and HRM were evaluated and validated for the rapid detection of ERG11 mutation. The fluconazole MICs for the 12 resistant and 19 susceptible isolates were 32–256 mg/L and 0.5–1 mg/L, respectively. The resistant isolates showed A339T and C461T mutations in the ERG11 gene. The T-ARMS-PCR and RSM approaches discriminated all the resistant and susceptible isolates, whereas HRM analysis differentiated all except one susceptible isolate. The sensitivity, specificity, analytical sensitivity, time, and cost of analysis suggests that these three methods can be utilized for the rapid detection of ERG11 mutations in C. tropicalis. Additionally, an excellent concordance with DNA sequencing was noted for all three methods. The rapid, sensitive, and inexpensive T-ARMS-PCR, RSM, and HRM approaches are suitable for the detection of azole resistance based on ERG11 polymorphism in C. tropicalis and can be implemented in clinical setups for batter patient management.
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