中国兽药杂志2025，Vol.59Issue(9)：16-23,8.DOI:10.11751/ISSN.1002-1280.2025.9.03
猪肺炎支原体实时荧光定量PCR方法的建立及在病原筛查中的应用
Development and Application of Real-Time Quantitative PCR for Detection of Mycoplasma hyopneumoniae in Pathogen Screening
摘要
Abstract
To develop a rapid and accurate Real-time quantitative PCR(qPCR)method for detecting Mycoplasma hyopneumoniae(Mhp),primers and a probe were designed within the highly conserved region of the 16S rRNA gene following primer design principles.The method was established through systematic optimization,and its specificity,sensitivity,and repeatability were evaluated.This method was subsequently applied to detect 50 clinical samples.The results demonstrated that this qPCR method specifically detected Mhp without cross-reactivity to nine non-target bacterial/viral strains.The limit of detection was determined to be 20 copies/μL.In repeatability tests using eight different standard concentrations across five replicates,the coefficient of variation for Ct values remained below 1%.When applied to 50 clinical nasal and pharyngeal swab samples,pharyngeal swabs exhibited higher detection rates and nucleic acid concentrations compared to nasal swabs.This study successfully established a qPCR method for Mhp,which can be applied to screen experimental pigs or clinically assess infection status by detecting Mhp pathogen nucleic acid in pharyngeal swab samples.关键词
猪肺炎支原体/实时荧光定量PCR/DNA标准品/鼻拭子/咽拭子Key words
Mycoplasma hyopneumoniae/Real-time quantitative PCR/DNA standard/nasal swab/pharyngeal swab分类
农业科技引用本文复制引用
候凤,贺笋,张慧敏,师小潇,王鹏江,高扬毅,李佳,武天增,李媛,韩瑞锴..猪肺炎支原体实时荧光定量PCR方法的建立及在病原筛查中的应用[J].中国兽药杂志,2025,59(9):16-23,8.基金项目
新疆维吾尔自治区"天山英才"培养计划青年科技拔尖人才项目(2022TSYCJC0018) （2022TSYCJC0018）
新疆生产建设兵团中央引导地方科技发展资金科技成果转移转化项目(2024YD019) （2024YD019）
