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目的:建立一种快速检测患者血液标本中恙虫病东方体的方法,用于恙虫病的早期实验室诊断。方法:收集患者血液标本,提取全基因组DNA;根据恙虫病东方体56 kD蛋白编码基因特异基因引物,采用巢式PCR方法对其进行检测,扩增的基因片段进行序列分析。结果:132份标本中,34份得到预期片段大小的PCR扩增产物,随机选取3份扩增产物TA克隆后测序分析,证实PCR扩增产物为恙虫病东方体基因片段。结论:该方法灵敏、特异、快速,可直接用于患者血液标本的恙虫病东方体特异性基因的检测,可为恙虫病东方体感染的早期诊断提供实验室依据。
Objective: To establish a rapid method for the detection of Orientia tsutsugamushi in the blood samples of patients for the early laboratory diagnosis of tsutsugamushi disease. Methods: Blood samples of patients were collected and the whole genome DNA was extracted. Based on the 56 kD protein-coding gene specific primers of Orien tsutsugamushi, the nested PCR method was used to detect and amplify the gene fragments. Results: Of the 132 samples, 34 were obtained PCR amplification products of the expected fragment size, randomly selected 3 copies of amplification product TA cloning and sequencing analysis confirmed that the PCR amplification products for the tsutsugamushi oriental gene fragment. CONCLUSION: The method is sensitive, specific and rapid and can be directly applied to the detection of oriental specific genes of tsutsugamushi in patients with blood samples. It can provide a laboratory basis for the early diagnosis of oriental tsutsugamushi infection.