Murai K, Tachibana N, Okayama A, Shishime E, Tsuda K, Oshikawa T
Second Department of Medicine, Miyazaki Medical School, Japan.
Microbiol Immunol. 1992;36(11):1145-53. doi: 10.1111/j.1348-0421.1992.tb02118.x.
We developed a nested polymerase chain reaction (PCR) method to detect Rickettsia tsutsugamushi (R. tsutsugamushi) DNA and determined its sensitivity. Primers were selected from the DNA sequence of the 58-kDa group-specific antigen gene of the Karp strain. The target sequence of rickettsial DNA was detectable as the band corresponding to 88 bp in 1.0 microgram of the DNA extracted from BS-C-1 cells infected with R. tsutsugamushi. Rickettsia-specific bands were observed not only for the homologous Karp strain, but also for four heterologous strains: two other reference strains (Gilliam and Kato) and two prototype strains prevalent in Miyazaki district (Irie and Hirano). The minimum copy number detectable by this method was estimated to be five rickettsiae. All of nine peripheral blood mononuclear cell samples from patients with tsutsugamushi disease who were seen 2-11 days after disease onset tested positive for rickettsial DNA. The PCR assay method presented here could be a specific diagnostic tool for tsutsugamushi disease, especially in its early acute stage.
我们开发了一种巢式聚合酶链反应（PCR）方法来检测恙虫病东方体（R. tsutsugamushi）DNA，并确定其敏感性。引物是从Karp株58-kDa群特异性抗原基因的DNA序列中选择的。从感染恙虫病东方体的BS-C-1细胞中提取的1.0微克DNA中，立克次体DNA的目标序列可检测为对应于88 bp的条带。不仅在同源的Karp株中观察到立克次体特异性条带，而且在四种异源株中也观察到：另外两种参考株（Gilliam和Kato）以及在宫崎地区流行的两种原型株（Irie和Hirano）。该方法可检测到的最小拷贝数估计为五个恙虫病东方体。在发病后2-11天就诊的恙虫病患者的所有九个外周血单核细胞样本的立克次体DNA检测均呈阳性。本文介绍的PCR检测方法可能是恙虫病的一种特异性诊断工具，尤其是在其早期急性期。
