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目的 鉴定山东地区恙虫病东方体的基因型。方法 采用聚合酶链反应 /限制性片段长度多态性分析 (PCR/RFLP)技术对恙虫病东方体山东分离株、小盾纤恙螨匀浆及恙虫病病人全血中提取的DNASta5 6基因进行分析研究 ,并与NestedPCR基因分型的结果进行比较。结果 山东地区 35份标本 ( 2 3株恙虫病东方体山东分离株、2份采自黑线姬鼠体外的小盾纤恙螨匀浆及 10份恙虫病人全血 )提取的DNA群引物扩增产物经HinfⅠ、HhaⅠ、SnaBⅠ酶切后呈现 2种不同的RFLP图谱 ,经与已知序列的Ot内切酶图谱相比 :33份Ot内切酶图谱与文献报告的日本地方株—Kawasaki株具有相似的酶切图谱 ,但缺乏HhaⅠ的酶切位点 ;另 2份与Karp参考株具有相同的酶切图谱。该结果与NestedPCR基因分型的结果相一致。结论 山东地区恙虫病东方体流行株主要基因型与日本地方株Kawasaki型类似 ,但与日本地方株存在遗传差异 ;同时还有Karp型Ot存在。采用PCR/RFLP方法可以准确对Ot-Sta5 6基因分型。
Objective To identify the genotype of Orientia tsutsugamushi in Shandong Province. Methods The DNASta5 6 gene extracted from the isolates of Shandong tsutsugamushi, the homozygotes of chigger sibiricum and the whole blood of patients with tsutsugamushi by PCR-RFLP Analytical studies and comparison with NestedPCR genotyping results. Results DNA group primers extracted from 35 specimens (23 Shandong strains of Orientia tsutsugamushi, 2 copies of homozygous chigger sibiricum and 10 pieces of whole blood of tsutsugamushi) were collected from Shandong Province The products were digested with HinfⅠ, HhaⅠand SnaBⅠrespectively, and showed two different RFLP maps. Compared with the Ot-endonuclease map of the known sequence, 33 Ot-endonuclease maps were compared with those reported in the literature (Kawasaki strain, Japan) Similar map of the enzyme digestion, but the lack of Hha Ⅰ restriction sites; the other two and Karp reference strains have the same restriction map. This result is consistent with the results of NestedPCR genotyping. Conclusion The major genotypes of oriental tsutsugamushi in endemic tsutsugamushi in Shandong Province are similar to those of the Japanese endemic strain Kawasaki, but there are genetic differences with the Japanese local isolates. There is also Karp-type Ot present. Ot-Sta5 6 genotyping can be accurately performed using the PCR / RFLP method.