肽核酸荧光原位杂交技术快速检测食品中的李斯特菌属及单增李斯特菌

Determination of Listeria genus and Listeria monocytogenes in foods by peptide nucleic acid-fluorescence in situ hybridization

目的建立利用肽核酸荧光原位杂交技术(PNA-FISH)快速检测食品中李斯特菌属及单增李斯特菌的方法。方法针对李斯特菌属、单增李斯特菌分别设计合成2份PNA探针lis-16S-1、lm-16S-2,并建立荧光原位杂交技术,优化杂交条件,对选取的13株李斯特菌和其他9株非李斯特菌进行检测,验证探针的特异性和灵敏度,并对118份食品样品用LB肉汤2次增菌培养后进行PNA-FISH检测。结果探针灵敏度和特异性均为100%,从118份食品中检出14株李斯特菌和8株单增李斯特菌,结果与API方法和VITEK方法鉴定结果一致。结论 PNA-FISH方法可靠易行,对从食品中检测致病性单增李斯特菌有较高的实用性。

Objective To develop a method of fluorescence in situ hybridization（FISH） using peptide nucleic acid（PNA） probes for the rapid detection of Listeria genus and Listeria monocytogenes in foodstuffs. Method Two PNA probes of lis-16S-1 and lm-16S-2 were synthesized for specific identification of Listeria genus and Listeria monocytogenes respectively, the PNA-FISH method was developed, and then the optimal reaction conditions were definitely explored. Thirteen strains of Listeria and 9 other bacterial species were detected to evaluate the sensitivity and specificity of the method, and 118 foodstuffs were detected using PNA-FISH after cultured by LB broth for 2 times. Results The sensitivity and specificity of PNA probes were both 100%. Fourteen strains of Listeria and 8 strains of Listeria monocytogenes were detected from 118 copies of foodstuffs, in coincidence with the methods of API and Vitek. Conclusion The method of PNA-FISH was reliable and suitable for the rapid identification of Listeria genus and Listeria monocytogenes in foodstuffs.