Bel Hadj Ali Insaf, Saadi-Ben Aoun Yusr, Hammami Zeineb, Rhouma Oumayma, Chakroun Ahmed Sahbi, Guizani Ikram
Laboratory of Molecular Epidemiology and Experimental Pathology-LR16IPT04, Institut Pasteur de Tunis, University of Tunis El Manar, Tunis 1002, Tunisia.
Pathogens. 2023 Oct 28;12(11):1292. doi: 10.3390/pathogens12111292.
Early and accurate detection of infectious diseases is a key step for surveillance, epidemiology and control, notably timely disease diagnosis, patient management and follow-up. In this study, we aimed to develop handheld ultra-fast duplex PCR assays coupled to amplicon detection by lateral flow (LF) immunoassay to deliver a rapid and simple molecular diagnostic test for concomitant detection and identification of the main parasites encountered in Tunisia. We selected two DNA targets to amplify / and / groups of species DNAs, respectively. We optimized the experimental conditions of a duplex ultra-fast PCR. The amplification is performed using a portable Palm convection PCR machine within 18 min, and the products are detected using an LF cassette within 10 min. The test allows the identification of the infecting species according to the position and number of test lines revealed. Tested on a selection of DNAs of representative strains of the three studied species ( = 37), the ultra-fast duplex PCR-LF showed consistent, stable and reproducible results. The analytical limit of detection of the test was 0.4 pg for , 4 pg for and 40 pg for .
传染病的早期准确检测是监测、流行病学和控制的关键步骤，尤其是疾病的及时诊断、患者管理和随访。在本研究中，我们旨在开发一种手持式超快速双重PCR检测方法，并结合侧向流动（LF）免疫分析进行扩增子检测，以提供一种快速简单的分子诊断测试，用于同时检测和鉴定突尼斯常见的主要寄生虫。我们分别选择了两个DNA靶标来扩增/和/物种DNA组。我们优化了双重超快速PCR的实验条件。使用便携式掌上对流PCR仪在18分钟内进行扩增，并在10分钟内使用LF检测卡检测产物。该测试可根据显示的测试线的位置和数量鉴定感染物种。在对三种研究物种的代表性菌株的DNA样本（=37）进行测试时，超快速双重PCR-LF显示出一致、稳定和可重复的结果。该测试的分析检测限对于为0.４pg，对于为4pg，对于为40pg。
