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目的建立并应用环介导等温扩增法(LAMP)检测肠道门诊感染性腹泻中的沙门菌。方法通过在线软件Primer Explorer V5设计针对沙门菌的保守区引物,建立LAMP检测方法,采用荧光定量PCR法、LAMP法和分离培养法,对本院2015年肠道门诊137例患者的粪便标本进行检测。对建立的方法进行特异度、灵敏度试验,与荧光定量PCR法和培养法进行比对。结果 137份标本中,荧光定量PCR法检出28份阳性,LAMP法检出28份阳性,分离培养法检出23份阳性。3种方法的检测结果差异无统计学意义(P>0.05)。LAMP对6株沙门菌属扩增的结果均为阳性,而对志贺菌等5株致病菌株扩增的结果均为阴性。LAMP检测方法具有良好的灵敏度和特异度。结论 LAMP检测方法灵敏特异,简单快速,比荧光定量PCR法更适用于基层疾病预防控制机构,有着较为广泛的发展前景。
Objective To establish and apply loop-mediated isothermal amplification assay (LAMP) for the detection of Salmonella in infectious gut diarrhea patients. Methods Primers designed by Primer Explorer V5 were used to design primers for conservative region of Salmonella. A LAMP assay was established. The stool samples of 137 patients in the enteral clinic in 2015 were detected by fluorescence quantitative PCR, LAMP assay and isolation culture. . Specificity and sensitivity test of the established method were compared with those of fluorescence quantitative PCR and culture method. Results Of the 137 samples, 28 were positive by fluorescence quantitative PCR, 28 were positive by LAMP method and 23 were positive by isolation culture method. There was no significant difference between the three methods (P> 0.05). The results of amplification of LAMP against 6 strains of Salmonella were all positive, but none of 5 strains of Shigella spp. LAMP detection method has good sensitivity and specificity. Conclusion The LAMP detection method is sensitive and specific, simple and rapid, and is more suitable than the fluorescence quantitative PCR method for prevention and control of grassroots disease. It has a wide range of development prospects.