Ji Wenxiang, Yuan Lufei, Hu Tingting, Jiang Min, Shen Jilu
Department of Clinical Laboratory, the First Affiliated Hospital of Anhui Medical University, Hefei, 230022, People's Republic of China.
Department of Clinical Laboratory, Anhui Public Health Clinical Center, Hefei, 230012, People's Republic of China.
Arch Microbiol. 2025 Aug 18;207(9):226. doi: 10.1007/s00203-025-04438-5.
Whooping cough (pertussis) is an acute respiratory infectious disease caused by Bordetella pertussis (BP). It poses a risk to infants and young children. This investigation aimed to construct a simple, rapid, and accurate diagnostic protocol for BP detection that does not depend on complex equipment or large-scale instruments. This study combines Recombinase Polymerase Amplification (RPA) technology with the CRISPR/Cas12a system, utilizing immunochromatographic lateral flow strips (ILFS) test and fluorescence curves to observe data. This diagnostic strategy does not require complex equipment used in traditional diagnostic approaches (such as bacterial culture, pathogen detection, and molecular biology techniques), which has increased its accessibility and ease of use. The validation data indicate that the RPA-CRISPR/Cas12a-ILFS and RPA-CRISPR/Cas12a fluorescence detection analyses had a lower detection threshold of 10 copies/µL and did not cross-react with other prevalent infections. Furthermore, 40 clinical samples were evaluated and compared via qPCR, which revealed that the RPA-CRISPR/Cas12a method has 100% sensitivity and specificity. In addition, the RPA-CRISPR/Cas12a diagnostic platform showed significant potential for clinical application, specifically when resources are limited, enabling point-of-care testing. This platform's simplicity, accuracy, and efficiency make it a powerful tool for pertussis diagnosis, which can improve patient care and public health outcomes.
百日咳是由百日咳博德特氏菌（BP）引起的急性呼吸道传染病。它对婴幼儿构成风险。本研究旨在构建一种简单、快速且准确的BP检测诊断方案，该方案不依赖复杂设备或大型仪器。本研究将重组酶聚合酶扩增（RPA）技术与CRISPR/Cas12a系统相结合，利用免疫层析侧流试纸条（ILFS）检测和荧光曲线来观察数据。这种诊断策略不需要传统诊断方法中使用的复杂设备（如细菌培养、病原体检测和分子生物学技术），这提高了其可及性和易用性。验证数据表明，RPA-CRISPR/Cas12a-ILFS和RPA-CRISPR/Cas12a荧光检测分析的检测下限为10拷贝/微升，且不与其他常见感染发生交叉反应。此外, 通过qPCR对40份临床样本进行评估和比较，结果显示RPA-CRISPR/Cas12a方法具有100%的敏感性和特异性。此外，RPA-CRISPR/Cas12a诊断平台在临床应用中显示出巨大潜力，特别是在资源有限的情况下，可实现即时检测。该平台的简单性、准确性和效率使其成为百日咳诊断的有力工具，可改善患者护理和公共卫生结果。
