基于PMA-qPCR分析的电子束辐照杀灭微生物机制研究

Study on the disinfection mechanism of microorganisms by electron beam irradiation based on Pl qPCR analysis

本研究通过优化PMA实时荧光定量PCR条件获得大肠杆菌等3种指示微生物分析的最佳PMA条件,并设置1~5 kGy梯度剂量进行电子束辐照后,应用PMA-qPCR测试并使用SPSS软件对RQ值进行方差分析,推断辐照破坏细胞膜渗透性的作用机制。试验结果表明,大肠杆菌、金黄色葡萄球菌、枯草芽孢杆菌黑色变种的最佳PMA浓度均为50μmol/L,最适光解时间分别为30 min、15 min和60 min。辐照剂量、PMA处理以及两者的交互作用均显著(P≤0.001)。3种微生物PMA处理组的RQ值均低于未经PMA处理组的RQ值(除去枯草芽孢杆菌的3~5 kGy处理),并且随着辐照剂量的增加,经PMA处理后的RQ值在3种微生物中总体呈现下降趋势,据此推测电子束辐照可能破坏了3种微生物的细胞膜。推荐以上3种微生物的电子束辐照灭活剂量为5 kGy。

In this study,the PMA-qPCR conditions for three kinds of indicating microorganisms were obtained and optimized,a gradient dose of 1 to 5 kGy was set to conduct electron beam irradiation,PMA-qPCR test and SPSS software were used to analyze the variance of RQ value,and the mechanism of radiation damage to cell membrane permeability was inferred.The results showed that the optimum PMA concentration of Escherichia coli,Staphylococcus aureus and Bacillus subtilis var.niger was 50μmol/L,and the optimum photolysis time was 30 min,15 min and 60 min,respectively.The irradiation dose(0~5 kGy),PMA treatment and the interaction between them were significant(P≤0.001).The RQ values of the PMA-treated groups for the three microorganisms were all lower than those of the untreated groups(except for the 3~5 kGy treatments of Bacillus subtilis).Additionally,with the increase in irradiation dose,the RQ values after PMA treatment generally showed a decreasing trend among the three microorganisms.It was inferred that the electron beam irradiation is likely to destroying the cell membrane of three microorganisms.The recommended dose for inactivation of the three microorganisms is 5 kGy.