Department of Neurosurgery, Zhongnan hospital of Wuhan university, China.
Wuhan Primbio Medical Laboratory, Wuhan, 430075, China.
Diagn Microbiol Infect Dis. 2025 Jan;111(1):116563. doi: 10.1016/j.diagmicrobio.2024.116563. Epub 2024 Oct 20.
Respiratory infections are one of the leading causes of death worldwide. Rapid and accurate detection of pathogens is crucial for timely treatment. This study established a detection method based on multiplex PCR combined with capillary electrophoresis, capable of simultaneously detecting 28 common respiratory pathogens. We used specially designed homo-tag primers, significantly reducing the formation of primer dimers and improving the specificity and sensitivity of amplification. After two rounds of PCR amplification, the products were subjected to fragment analysis using the ABI 3500XL Genetic Analyzer, enabling automated result interpretation. The detection limit of this method reaches 2.77 × 10 copies/ml, with some pathogens reaching as low as 2.77 × 10 copies/ml. The detection of 147 clinical specimens showed that the overall consistency of this method with culture, colloidal gold, fluorescent quantitative PCR, and NGS was 75.5 %. The multiplex PCR detection method established in this study is rapid, sensitive, and high-throughput, can be used for routine screening of common pathogens in respiratory infections, providing an important basis for clinical diagnosis and treatment.
呼吸道感染是全球主要死因之一。快速准确地检测病原体对于及时治疗至关重要。本研究建立了一种基于多重 PCR 结合毛细管电泳的检测方法，能够同时检测 28 种常见呼吸道病原体。我们使用专门设计的同型标签引物，显著减少引物二聚体的形成，提高扩增的特异性和灵敏度。经过两轮 PCR 扩增后，使用 ABI 3500XL 遗传分析仪对产物进行片段分析，实现自动结果解读。该方法的检测限达到 2.77×10 拷贝/ml，某些病原体的检测限低至 2.77×10 拷贝/ml。对 147 份临床标本的检测表明，该方法与培养、胶体金、荧光定量 PCR 和 NGS 的总体一致性为 75.5%。本研究建立的多重 PCR 检测方法快速、敏感、高通量，可用于常规筛查呼吸道感染中的常见病原体，为临床诊断和治疗提供重要依据。
