Zhang Yang, Ke Lihui, Sun Tao, Liu Yang, Wei Bo, Du Minghua
Comprehensive Technical Service Center of Xuzhou Customs, Xuzhou Customs, Xuzhou 221000, China.
Department of Thoracic Surgery, Beijing Tiantan Hospital, Capital Medical University, Beijing 100070, China.
Micromachines (Basel). 2023 Oct 9;14(10):1917. doi: 10.3390/mi14101917.
Malaria is listed as one of the three most hazardous infectious diseases worldwide. Travelers and migrants passing through exit and entry ports are important sources of malaria pandemics globally. Developing accurate and rapid detection technology for malaria is important. Here, a novel hairpin-mediated amplification (HMA) technique was proposed for the detection of four Plasmodium species, including , , , and . Based on the conserved nucleotide sequence of Plasmodium, specific primers and probes were designed for the HMA process, and the amplicon can be detected using lateral flow detection (LFD); the results can be read visually without specialized equipment. The specificity of HMA-LFD was evaluated using nucleic acids extracted from four different Plasmodium species and two virus species. The sensitivity of HMA-LFD was valued using 10× serial dilutions of plasmid containing the template sequence. Moreover, 78 blood samples were collected to compare HMA-LFD and qPCR. The HMA-LFD results were all positive for four different Plasmodium species and negative for the other two virus species. The sensitivity of HMA-LFD was tested to be near five copies/μL. The analysis of clinical samples indicated that the consistency of HMA-LFD and qPCR was approximately 96.15%. Based on these results, the HMA-LFD assay was demonstrated to be a rapid, sensitive, and specific technique for the detection of Plasmodium and has great advantages for on-site detection in low-resource areas and exit and entry ports.
疟疾被列为全球三大最危险的传染病之一。出入境口岸的旅行者和移民是全球疟疾大流行的重要源头。开发准确、快速的疟疾检测技术至关重要。在此，提出了一种新型发夹介导扩增（HMA）技术用于检测四种疟原虫物种，包括 、 、 和 。基于疟原虫的保守核苷酸序列，为HMA过程设计了特异性引物和探针，扩增产物可使用侧向流动检测（LFD）进行检测；无需专门设备即可直观读取结果。使用从四种不同疟原虫物种和两种病毒物种中提取的核酸评估HMA-LFD的特异性。使用含模板序列的质粒进行10倍系列稀释来评估HMA-LFD的灵敏度。此外，收集了78份血样以比较HMA-LFD和qPCR。对于四种不同疟原虫物种，HMA-LFD结果均为阳性，而对于其他两种病毒物种则为阴性。经测试，HMA-LFD的灵敏度接近5个拷贝/μL。临床样本分析表明，HMA-LFD与qPCR的一致性约为96.15%。基于这些结果，HMA-LFD检测方法被证明是一种检测疟原虫的快速、灵敏且特异的技术，在资源匮乏地区以及出入境口岸的现场检测方面具有巨大优势。
