PCR技术快速检测痢疾志贺菌的研究

Rapid detection of Shigella dysenteriae by PCR assay

目的快速、准确检测食品中的痢疾志贺菌(Shigella dysenteriae)感染。方法针对痢疾志贺菌的侵袭性质粒抗原H基因(ipaH)设计引物,通过对PCR扩增体系中dNTP浓度、Mg2+浓度、引物浓度、酶用量、退火温度和循环数进行优化,建立痢疾志贺菌PCR快速检测方法,并对其特异性、敏感性进行分析。结果该方法检测的特异性好,痢疾志贺菌纯培养物和基因组DNA的灵敏度分别为1.06×102cfu/ml和106.34pg/PCR反应体系;直接检测模拟食品中的痢疾志贺菌,其检出限为3.21×104cfu/ml。结论该方法操作简单、检验周期短、特异性和灵敏度高,适用于快速、准确检测食品中致病菌的污染。

Based on the invasive plasmid antigen H gene（ipaH） of S.dysenteriae,one pair of specific primers was designed for PCR assays in this study.The concentrations of dNTP,Mg2 ＋ and primer,dosage of Taq DNA polymerase,annealing temperature and circulating parameter in the PCR amplification system were optimized.In this way,a rapid and stable method of PCR assay for the detection of S.dysenteriae was established.The specificity and sensitivity of PCR were also analyzed.The detection limits of pure culture and genomic DNA in the PCR assay were 1.06×102 cfu/ml and 106.34pg /PCR system,respectively.The detection limit for S.dysenteriae in artificially contaminated food samples was 3.21×104 cfu /ml.These results indicated that the PCR method for S.dysenteriae detection was simple,rapid,high in specificity and sensitivity and suitable for the detection of pathogens in foods caused by Shigella dysenteriae.