应用重组酶聚合酶扩增检测法快速分群解脲支原体的探讨

Rapid clustering detection method of ureaplasma urealyticum by recombinant polymerase amplification detection

目的探讨应用重组酶聚合酶扩增(RPA)技术对解脲支原体(UU)快速分群的检测方法,并与PCR-反向点杂交法进行比较,建立UU快速分群的床旁检测(POCT)方法。方法采用UU标准菌株、收集的临床标本及阴性质控菌株,经引物设计和菌株、临床标本DNA提取,对UU标准菌株、收集临床标本及阴性质控菌株进行RPA法、PCR-反向点杂交法分群检测,通过两种方法的结果比较,评价RPA法检测UU快速分群试验的可行性。结果RPA法成功将UU标准菌株分群;113例临床标本中检测发现39例UU感染,其中12例UU为生物Ⅰ群,27例为生物Ⅱ群;阴性质控菌株未扩增;以上UU标准菌株、临床标本、阴性质控菌株经过PCR-反向点杂交法分群,结果与RPA法一致,成功建立UU RPA分群法。结论RPA法与PCR-反向点杂交法对UU进行分群结果一致,应用RPA法可以简捷、快速、高效、灵敏地完成UU的分群,获得致病型UU的诊断结果。

Objective To investigate the method of fast clustering detection method of ureaplasma urealyticum(UU)by recombinase polymerase amplification(RPA)and compare it with PCR reverse dot hybridization,to establish a method of rapid clustering of UU by bedside detection(POCT).Methods The standard strains of UU,the clinical samples and the negative control strains were collected.Then the primers were designed,and the DNA extraction from the above strains and clinical samples were obtained for subsequent analysis.All of these 3 kinds of samples were collected and clustered by RPA method and PCR-reverse dot hybridization.The results were compared to evaluate the feasibility of RPA method for detecting UU fast clustering experiment.Results The standard strain of UU was successfully clustered by RPA method.Among the 113 clinical samples,39 cases of UU infection were detected,among which 12 cases were biologicalⅠand 27 cases were biologicalⅡ.The negative strain was not amplified.The results of these strains by PCR-reverse dot hybridization were consistent with RPA method.UU RPA clustering method was established successfully.Conclusion The results of RPA and PCR-reverse dot hybridization for UU are consistent.Using RPA method can be a simple,rapid,efficient and sensitive to UU clustering and obtain the diagnostic results of pathogenic UU.