一种半巢式重组聚合酶扩增检测法，用于检测临床样本中的立克次体病原体。
A seminested recombinase polymerase amplification assay to detect rickettsial pathogens in clinical samples.
机构信息
Center for Disease Control and Prevention of Xilingol League, Xilinhaote, Inner Mongolia, China; State Key Laboratory of Infectious Diseases Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing, China.
Center for Disease Control and Prevention of Inner Mongolia, Hohhot, Inner Mongolia, China.
出版信息
Diagn Microbiol Infect Dis. 2023 Dec;107(4):116067. doi: 10.1016/j.diagmicrobio.2023.116067. Epub 2023 Sep 9.
Treatment at the early stage of onset is vital for the prognosis of rickettsioses. But the absence of specific clinical symptoms complicates the diagnosis of this condition. Herein we established a seminested recombinase polymerase amplification assay (snRPA-nfo) that enables quick detection and differentiation of rickettsial pathogens in clinical samples with high sensitivity and specificity. The conserved 17-kDa protein gene of Rickettsia sibirica and the 47-kDa protein gene of Orientia tsutsugamushi were targeted for the duplex RPA-nfo assay. The snRPA-nfo assay exhibited an increased LOD in spiked blood samples, up to 1000-fold in comparison to standard RPA-nfo, and a better detection rate (83.3%, 5/6) than TaqMan PCR (16.6%, 1/6, Ct ≤ 35) in clinically confirmed patient blood samples. Thus, snRPA-nfo assay represents a promising alternative to TaqMan PCR in the early diagnosis of rickettsioses for point-of-care testing as well as in resource-limited settings.
在疾病发作的早期进行治疗对于立克次氏体病的预后至关重要。但缺乏特定的临床症状使得这种疾病的诊断变得复杂。在此，我们建立了一种半巢式重组聚合酶扩增检测法（snRPA-nfo），该方法能够快速检测和区分临床样本中的立克次体病原体，具有高灵敏度和特异性。针对西伯利亚立克次体的 17 kDa 蛋白基因和恙虫东方体的 47 kDa 蛋白基因，我们设计了双重组酶聚合酶扩增检测法（RPA-nfo）。与标准 RPA-nfo 相比，snRPA-nfo 检测法在加标血样中的 LOD 提高了 1000 倍，在临床确诊的患者血样中的检测率（83.3%，5/6）也优于 TaqMan PCR（16.6%，1/6，Ct ≤ 35）。因此，snRPA-nfo 检测法有望替代 TaqMan PCR 用于即时检测，以进行立克次氏体病的早期诊断，尤其是在资源有限的环境下。
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