关键词: 恙虫病东方体, 套式PCR, 恙螨, 血块及脾脏标本

Abstract: Objective: To detect Orientia tsutsugamushi in Chigger Mites and the specimens collected from the rodents to analyse the epidemic trend of scrub typhus. To evalute the sensitivity and specificity of nested polymerase chain reaction and promote the research in molecular epidemiology of O.tsutsugamushi in the country. Methods: NPCR was used to detect O.tsutsugamushi DNA in Chigger Mites and samples collected from artificial infected mice and field rodents. Two pairs of primer were synthesized on the basis of the DNA sequence from Sta 58kDa group specific antigen gene. Product of PCR amplification was analysed by 2% agarose gel electrophoresis. PCR products showed positive for O.tsutsugamushi DNA were further identified by 12% PAGE and PGEM-3Zf(+)/Hae Ⅲ DNA was used as nucleotide molecular weight marker. The result indicated the band is corresponding to a 88bp DNA fragment from Sta 58 kDa gene of O.tsutsugamushi . Result: One of 29 (3.45 %) blood clots and one of 112 (0.98 %) spleen tissue from field rats were positive for 88bp DNA fragment. Eight DNA extracts from 39 (20.5%) chigger mites produced the predicted DNA fragment.The specimens of both blood clots and spleen tissues from the mice at day 3 of postinfection showed negative to the specific PCR product. But positive at day 6 and 9. Conclusion: NPCR has high sensitivity and specificity. Hence. The PCR assay is a useful tool to investigate the natural epidemic foci of scrub typhus.