菌落P CR改良技术快速鉴定阿萨希毛孢子菌的实验研究

摘要

目的：探索快速鉴定阿萨希毛孢子菌（Trichosporon asahii，T．asahii）的简便方法。方法分别应用直接法、直接离心法、酶解法、酶解改良法4种方法进行样本预处理后的菌落PCR技术检测16株T．asahii，评价上述4种方法的敏感性和微量样本阳性率，同时与DNA常规提取法PCR结果进行比较。结果酶解改良法、直接法、常规提取法敏感性均为102 CFU／mL，阳性率分别为93．75％、75％、50％；直接离心法敏感性为104 CFU／mL，阳性率为43．75％；酶解法结果为阴性。结论酶解改良法是一种简便快速的PCR模板获取方法，适用于菌落PCR技术对T．asahii进行快速鉴定时的样本预处理。%Objective To explore a direct colony PCR method for rapid identification of T.asahii.Methods Four methods inclu⁃ding direct colony,direct colony with centrifugation,enzymolysis,and improved enzymolysis method were used to preliminarily sample treatment.The sensitivity and the positive rate of colony PCR based on four methods for isolates of T.asahii were evaluated and com⁃pared with the PCR results of the traditional DNA extraction.Results The sensitivity of the improved enzymolysis method,direct col⁃ony method and the traditional DNA extraction were both 102 CFU/mL,while the positive rates were 93.75%,75% and 50%,respec⁃tively.The direct colony with centrifugation presented the sensitivity of 104 CFU/mL and the positive rate of 43.75%.The enzymolysis method presented total negative results in the detection of both sensitivity and positive rate. Conclusion The improved enzymolysis method which presented the advantages of high sensitivity and positive rate of trace samples,could be applied for the rapid identifica⁃tion of T.asahii.