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两种 DNA 提取方法检测粪类圆线虫感染的评价。

Evaluation of Two DNA Extraction Methods for Detection of Strongyloides stercoralis Infection.

机构信息

Swiss Tropical and Public Health Institute, Basel, Switzerland.

University of Basel, Basel, Switzerland.

出版信息

J Clin Microbiol. 2018 Mar 26;56(4). doi: 10.1128/JCM.01941-17. Print 2018 Apr.

Abstract

is present worldwide, but its prevalence is still uncertain, mainly due to the lack of sensitivity of diagnostic methods. Molecular techniques are under development, but a standardized protocol is still unavailable. We compared the sensitivity of real-time PCR, using two extraction protocols, with that of the Baermann technique. Samples were collected in the framework of the baseline screening of a randomized clinical trial evaluating moxidectin against in Lao People's Democratic Republic. Two stool samples from each participant were processed by the Baermann method, and one subsample was processed by PCR. DNA was extracted using the QIAamp DNA stool minikit based on the standard protocol for the QIAamp DNA minikit (QIA) and using a modification of the QIA procedure (POL). Subsequently, all extracted samples were analyzed by real-time PCR. Overall, 95 samples were analyzed by the three diagnostic methods. Sixty-nine (72.6%) samples were positive according to the Baermann method, 25 (26.3%) by the QIA method, and 62 (65.3%) by the POL method. The sensitivities were 86% (95% confidence interval [CI], 76.7 to 92.9), 31.0% (95% CI, 21.3 to 42.6), and 78.0% (95% CI, 66.8 to 86.1) for the Baermann, QIA, and POL methods, respectively. The sensitivities calculated for each day of the Baermann method separately were 60% (48.4 to 70.8%) and 64% (52.2 to 74.2%) for days 1 and 2, respectively. In conclusion, the POL method revealed a good performance and was comparable to the Baermann test performed on two stool samples and superior to the Baermann method performed on one stool sample. Additional studies are needed to standardize a PCR protocol for diagnosis.

摘要

目前全球均有隐孢子虫病存在,但该病的流行情况仍不确定,主要原因是诊断方法的敏感性不足。目前正在开发分子技术,但仍缺乏标准化的方案。我们比较了实时 PCR 法的敏感性,该方法使用了两种提取方案,与巴氏法进行比较。这些样本是在老挝人民民主共和国开展的一项随机临床试验的基线筛查中收集的,该试验评估了莫昔克丁对隐孢子虫的疗效。每位参与者的两份粪便样本用巴氏法处理,一份粪便样本用 PCR 法处理。使用 QIAamp DNA 粪便试剂盒(基于 QIAamp DNA 小试剂盒的标准方案)和修改后的 QIA 方案(POL)提取 DNA。随后,所有提取的样本均通过实时 PCR 法进行分析。总共分析了 95 个样本,采用了三种诊断方法。巴氏法检测阳性的样本有 69 个(72.6%),QIA 法检测阳性的样本有 25 个(26.3%),POL 法检测阳性的样本有 62 个(65.3%)。巴氏法、QIA 法和 POL 法的敏感性分别为 86%(95%置信区间 [CI],76.7 至 92.9)、31.0%(95% CI,21.3 至 42.6)和 78.0%(95% CI,66.8 至 86.1)。巴氏法每天分别检测一个样本时,第 1 天和第 2 天的敏感性分别为 60%(48.4 至 70.8%)和 64%(52.2 至 74.2%)。综上,POL 法具有良好的性能,与巴氏法检测两份粪便样本的效果相当,优于巴氏法检测一份粪便样本的效果。需要进一步的研究来标准化隐孢子虫诊断的 PCR 方案。

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