微波杀菌过程中大肠杆菌与金黄色葡萄球菌细胞膜通透性的改变

Alterations of membrane permeability in Escherichia coli and Staphylococcus aureus under microwave

摘要:

对细胞膜通透性变化的研究是认识微波杀菌机理的途径之一。用荧光探针检测微波处理后细胞内Ca²⁺浓度的变化,可以精确地表征细胞膜通透性的改变。选用二乙酸荧光素(FDA)和Fluo-3/AM两种荧光染料,对大肠杆菌(Escherichiacoli)和金黄色葡萄球菌(Staphylococcus aureus)经微波处理后的酯酶活性及细胞膜通透性进行研究,结果表明大肠杆菌与金黄色葡萄球菌的胞内非特异性酯酶(NSE)活性及细胞膜通透性的变化情形有所不同。在50℃、55℃、60℃和65℃微波处理条件下,大肠杆菌细胞膜通透性分别增加了20.7%、28.1%、74.8%、89.8%,而金黄色葡萄球的增加不显著,分别比对照组提高了4.1%、6.0%、21.9%和19.7%。细胞膜通透性的改变与微生物致死率有一定的相关性,也可能是微波杀菌非热效应的表现之一。

Abstract:

Alteration of cell membrane permeability is speculated to be one of the mechanisms by which microwave kills microorganisms. It has been reported that permeability alteration may be reflected by cell shape changes observed under electron microscopy, or detected by measuring the leakage of intracellular protein and DNA using spectrophotometry. These methods, however, suffer from accuracy and sensitivity. Calcium is an important cell signaling molecule. Its level is tightly regulated with an intracellular to extracellular differential of approximately 1 to 10,000. Damage of cells will lead to alterations in membrane permeability and consequently influx of extracellular Ca²⁺. In the present study two probes, fluorescein diacetate (FDA) and fluo-3/AM, were used to quantify membrane permeability of E.coli and S.aureus after microwave treatment. These chemical probes, after metabolized by intracellular esterases and binding to Ca²⁺, emit strong fluorescence. Our data showed 20.7%, 28.1%, 74.8% and 89.8% increases in cel membrane permeability of E.coli after 50, 55, 60 and 65℃ microwave treatment, respectively, compared to untreated controls. Modest membrane permeability increases of 4.1%, 6.0%, 21.9% and 19.7% were seen for S.aureus. The permeability levels correlate with the fatality rates of microbes. These results suggest that alteration in cell membrane permeability contributes, in part, to the nonthermal-effect of cell killing by microwave.