Highly Sensitive Molecular Diagnostic Platform for Scrub Typhus Diagnosis Using O. tsutsugamushi Enrichment and Nucleic Acid Extraction.

IF 4.9 3区 工程技术 Q1 CHEMISTRY, ANALYTICAL Biosensors-Basel Pub Date : 2024-10-10 DOI:10.3390/bios14100493
Myoung Gyu Kim, Seulki Kim, Juho Jang, Jinkwan Lee, Namheon Kim, Yeji Yu, A Reum Kim, Seungjin Lim, Moonsuk Bae, Yong Shin
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Abstract

Scrub typhus is caused by the Gram-negative obligate intracellular bacterium Orientia tsutsugamushi, and this tick-borne disease is difficult to distinguish from other acute febrile illnesses as it typically presents with symptoms such as rash, crusting at the bite site, headache, myalgia, lymphadenopathy, and elevated liver transaminases. It can often be diagnosed clinically, but not all patients present with characteristic symptoms, so serological diagnosis and molecular techniques may be required. However, existing diagnostic tests often have low sensitivity and specificity, making early detection difficult. This study presents a nucleic acid extraction method using large volumes of plasma and buffy coat to increase sensitivity, as well as an improved detection method using two target genes. Using the I-PULL device, nucleic acids can be extracted from up to 4 mL of sample in 30 min, avoiding contamination. The extracted DNA detects two genes of O. tsutsugamushi, increasing sensitivity compared to single-gene detection. Clinical validation in 38 patient samples showed 100% specificity and 95.24% sensitivity for the single target gene, with specificity and sensitivity rising to 100% when both genes are analyzed. This molecular diagnostic platform can be useful for distinguishing scrub typhus from similar diseases.

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利用恙虫卵富集和核酸提取技术诊断恙虫病的高灵敏度分子诊断平台
恙虫病是由革兰氏阴性细胞内细菌恙虫病菌(Orientia tsutsugamushi)引起的,这种蜱媒疾病很难与其他急性发热性疾病区分开来,因为它通常会出现皮疹、叮咬部位结痂、头痛、肌痛、淋巴结病和肝转氨酶升高等症状。这种疾病通常可以通过临床诊断,但并非所有患者都表现出特征性症状,因此可能需要进行血清学诊断和分子技术检测。然而,现有诊断测试的灵敏度和特异性往往较低,因此难以早期发现。本研究提出了一种利用大量血浆和水溶液提取核酸的方法,以提高灵敏度,并利用两个目标基因改进了检测方法。使用 I-PULL 装置,可在 30 分钟内从多达 4 毫升的样本中提取核酸,避免了污染。提取的 DNA 可检测恙虫病的两个基因,与单基因检测相比,灵敏度更高。在 38 份患者样本中进行的临床验证显示,单个目标基因的特异性为 100%,灵敏度为 95.24%,当分析两个基因时,特异性和灵敏度均上升到 100%。该分子诊断平台可用于区分恙虫病和类似疾病。
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来源期刊
Biosensors-Basel
Biosensors-Basel Biochemistry, Genetics and Molecular Biology-Clinical Biochemistry
CiteScore
6.60
自引率
14.80%
发文量
983
审稿时长
11 weeks
期刊介绍: Biosensors (ISSN 2079-6374) provides an advanced forum for studies related to the science and technology of biosensors and biosensing. It publishes original research papers, comprehensive reviews and communications. Our aim is to encourage scientists to publish their experimental and theoretical results in as much detail as possible. There is no restriction on the length of the papers. The full experimental details must be provided so that the results can be reproduced. Electronic files and software regarding the full details of the calculation or experimental procedure, if unable to be published in a normal way, can be deposited as supplementary electronic material.
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