Department of Microbiology, Mahatma Gandhi Medical College & Research Institute, Sri Balaji Vidyapeeth, Puducherry, India.
Department of Genomics and Proteomics, Central Interdisciplinary Research Facility, Mahatma Gandhi Medical College and Research Institute, Puducherry - 607 403, India.
J Vector Borne Dis. 2020 Apr-Jun;57(2):147-152. doi: 10.4103/0972-9062.310866.
BACKGROUND & OBJECTIVES: Scrub typhus (ST), an important zoonosis caused by Orientia tsutsugamushi, is now prevalent throughout India. While demonstration of IgM antibody by Indirect Immunofluorescence Assay (IFA) is the gold standard serological test, IgM ELISA is an alternative. Demonstration of O. tsutsugamushi DNA in the blood or eschar confirms infection in the early febrile period.
Scrub typhus nested PCR (n-PCR) for 56 kDa, 47 kDa and groEL genes and ST IgM ELISA were performed for 210 clinically suspected ST patients. As healthy controls, 70 voluntary blood donors were included. Statistical analysis was performed for laboratory parameters using Fisher exact test/chi-square test. Ninety-five PCR products of n-PCR positive samples were purified and submitted for gene sequencing.
PCR was positive for one or more gene targets in 75.71% of IgM ELISA positive patients and 10% of antibody negative patients. All voluntary blood donors were negative for both antibodies and DNA. Gene sequences of 95 n-PCR positive products confirmed the presence of Orientia tsutsugamushi DNA in the samples and NCBI database accession numbers MG601875 to MG601969 were obtained.
INTERPRETATION & CONCLUSION: Compared to IgM ELISA, sensitivity of three PCRs was 30, 51.43 and 61.43% for 56 kDa, 47 kDa and groEL targets, respectively. Since IgM ELISA positivity can persist up to one year, PCR confirms ST diagnosis in the acute phase of the illness, in the presence of IgM and even before IgM appears. Inclusion of all three genes - 56 kDa, 47 kDa and groEL, instead of a single 56 kDa target, identifies and confirms maximum number of ST patients.
恙虫病（ST）是一种由恙虫东方体引起的重要动物源性传染病，目前在印度广泛流行。间接免疫荧光法（IFA）检测 IgM 抗体是血清学检测的金标准，但 IgM ELISA 是一种替代方法。在发热早期，血液或焦痂中检测到恙虫东方体 DNA 可确认感染。
对 210 例临床疑似 ST 患者进行恙虫病巢式 PCR（n-PCR）检测 56 kDa、47 kDa 和 groEL 基因及 ST IgM ELISA。选择 70 名自愿献血者作为健康对照。采用 Fisher 确切检验/卡方检验对实验室参数进行统计学分析。对 n-PCR 阳性样本的 95 个 PCR 产物进行纯化并提交基因测序。
IgM ELISA 阳性患者中，1 个或多个基因靶标阳性的患者占 75.71%，抗体阴性患者中占 10%。所有自愿献血者均对抗体和 DNA 检测均为阴性。95 个 n-PCR 阳性产物的基因序列证实了样本中存在恙虫东方体 DNA，获得的 NCBI 数据库登录号为 MG601875 至 MG601969。
与 IgM ELISA 相比，56 kDa、47 kDa 和 groEL 目标的三种 PCR 的敏感性分别为 30%、51.43%和 61.43%。由于 IgM ELISA 阳性可持续长达 1 年，PCR 可在 IgM 存在时甚至在 IgM 出现之前，在疾病的急性阶段确认 ST 诊断。包含 56 kDa、47 kDa 和 groEL 三个基因，而不是单个 56 kDa 靶标，可以识别并确认最多数量的 ST 患者。
