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目的探讨巢式PCR法与实时荧光PCR法在恙虫病诊断中的作用。方法采集2010年-2014年北京市404例恙虫病疑似患者的全血样本,分别以巢式PCR法与实时荧光PCR法检测恙虫病东方体的gro EL基因与47 k D蛋白基因,以血清特异性Ig M检测结果为诊断恙虫病的金标准,研究2种PCR方法的灵敏度。结果以急性期血清Ig M阳性或恢复期血清Ig M转阳作为确诊标准,374例患者确诊为恙虫病。其中巢式PCR法检测gro EL基因阳性者323例,灵敏度为86.36%;实时荧光PCR法检测47 k D蛋白基因阳性者328例,灵敏度为87.70%。而单份急性期血清Ig M抗体检测的灵敏度为87.17%。结论实时荧光PCR法的检测时间短,适合用于快速诊断,但对于检测阴性的样本应考虑进行血清Ig M或多个基因的分子生物学检测以明确诊断。
Objective To explore the role of nested PCR and real-time fluorescence PCR in the diagnosis of scrub typhus. Methods A total of 404 blood samples were collected from 404 patients with suspected tsutsugamushi disease in Beijing from 2010 to 2014. The gro EL gene and 47 kD protein gene of tsutsugamushi were detected by nested PCR and real-time fluorescence PCR, respectively. Serum specificity IgM test results for the diagnosis of tsutsugamushi gold standard, the sensitivity of the two PCR methods. Results Acute serum Ig M positive or convalescent serum Ig M as the standard of diagnosis, 374 patients diagnosed as tsutsugamushi disease. Among them, 323 cases with positive gro EL gene were detected by nested PCR, and the sensitivity was 86.36%. The real-time fluorescent PCR assay detected the positive of 47 kD gene in 328 cases, the sensitivity was 87.70%. However, the single-phase serum IgM antibody detection sensitivity was 87.17%. Conclusion The real-time fluorescent PCR method has short detection time and is suitable for rapid diagnosis. However, for the negative samples, molecular biological tests of serum Ig M or multiple genes should be considered in order to confirm the diagnosis.