多重荧光定量PCR法快速鉴定MRSA的临床研究

目的 构建mecA、nuc、内标基因3种基因联合检测的单管多通道Taqman探针荧光定量PCR法,用于鉴定耐甲氧西林金黄色葡萄球菌(MRSA),并与传统细菌培养和鉴定方法进行比较,评估其临床应用价值.方法 以169例临床标本和经常规细菌培养和VITEK2 Compact全自动微生物分析仪鉴定的金黄色葡萄球菌(SA)15...

Clinical study on multiple fluorescence quantitative PCR for the rapid detection of MRSA

Objective To establish the method of identifying MRSA with Taqman-fluorescence quantitative PCR based on the detection for mecA,nuc,endogenous reference gene,which is a three-gene combined detection in a single tube multiple channel,and compare with the traditional bacterial culture and identification method to evaluate its clinical application value for the rapid detection of MRSA.Methods A total of 169 clinical samples and 156 SA strains were collected in the study.The SA strains were isolated from clinical samples and confirmed by VITEK2 Compact microbial analyzer.MecA,nuc,endogenous reference gene PCR primers and Taqman fluorescent probes were designed by using Primer premier 5.0 and Beacon Designer 7.0.FAM,Cy5 and VIC markers were used to label the fluorescent probe at 5′end,and 3′end was labeled with MGB,and the detection was carried out in the fluorescent quantitative PCR instrument,while the 169 cases of clinical samples were also cultured and tested by the traditional method.Results The traditional method confirmed 48 cases of MRSA,and the detection rate was 28.40%,while the multiple fluorescent quantitative PCR detected 53 cases of MRSA,and the detection rate was 31.36%.There was no significant difference between traditional method and Taqman-fluorescence quantitative PCR(P>0.05).In 156 SA strains,the traditional methord confirmed 106 strains of MRSA,and the detection rate was 67.95%,while the multiple fluorescence quantitative PCR detected 108 strains of MRSA,and the detection rate was 69.23%.There is also no significant difference between traditional method and Taqman-fluorescence quantitative PCR(P>0.05).Conclusion Taqman-fluorescence quantitative PCR improved positive detection rate and reduced the risk of missing detection of MRSA,it is much more time-saving than tradition method,which can be used in rapid clinical identification of MRSA.