中国兽医科学2025，Vol.55Issue(11)：1470-1478,9.DOI:10.16656/j.issn.1673-4696.2025.0206
可视化禽偏肺病毒B型和C型二重RT-LAMP检测方法的建立
Establishment of visual duplex RT-LAMP method for detecting avian metapneumovirus type B and type C
摘要
Abstract
In order to establish a rapid and differential diagnostic method for avian metapneumo-virus type B(aMPV/B)and type C(aMPV/C),the primers and fluorescent probes were designed based on the conserved gene fragments of aMPV/B and aMPV/C.A duplex reverse transcription loop-mediated isothermal amplification(RT-LAMP)detection method for aMPV/B and aMPV/C was developed by optimizing the probe concentration and reaction temperature.The results showed that the optimal probe concentration for both aMPV/B and aMPV/C was 0.24 μmol/L,and the optimal reaction temperature was 64 ℃.Specificity tests indicated that the duplex RT-LAMP method could specifically detect aMPV/B and aMPV/C,while showing negative amplification for 12 common avian pathogens such as avian influenza virus(AIV)and Newcastle disease virus(NDV).Sensitivity and interference tests demonstrated that the duplex fluorescent RT-LAMP method was 10 times more sensitive than conventional RT-PCR for both aMPV/B and aMPV/C,and had good resistance to interference.Repeatability tests showed that the intra-assay and inter-assay coefficient of variation of the duplex fluorescent RT-LAMP method for aMPV/B and aMPV/C was less than 3％.Clinical sample testing results were highly consistent with those of RT-PCR.In conclusion,this study success-fully established a duplex visual RT-LAMP detection method,which can provide technical support for the clinical prevention and control of aMPV/B and aMPV/C.关键词
禽偏肺病毒/环介导等温扩增/可视化/鉴别诊断Key words
avian metapneumovirus/loop-mediated isothermal amplification/visualization/diffe-rential diagnosis分类
农业科技引用本文复制引用
尹文巧,李小凤,谢芝勋,张艳芳,范晴,谢志勤,罗思思,谢丽基,李孟,吴嫒琼..可视化禽偏肺病毒B型和C型二重RT-LAMP检测方法的建立[J].中国兽医科学,2025,55(11):1470-1478,9.基金项目
广西科技基地和人才专项(桂科AD17195083) （桂科AD17195083）
国家现代农业产业体系广西家禽产业创新团队项目(nycytxgx-cxtd-2024-19) （nycytxgx-cxtd-2024-19）
广西重点研发计划项目(桂科AB25069499) （桂科AB25069499）
广西八桂学者专项(2019A50) （2019A50）
