| Pulsed electric field(PEF)technology,an emerging non-thermal technique,possesses the ability to inactivate micro-organisms.Although there have been large quantities of studies on PEF,study on the mechanism of micro-organisms inactivated by PEF as well as the effect of cytoplasmic membrane modified by stress behavior on PEF resistance of micro-organisms is not fully understood.As a kind of common foodbone pathogenic bacteria,Salmonella Typhimurium,its adaptation to stressful environment has been focused on by researchers.Therefore,S.Typhimurium was selected as a mode in this work to investigate the effect of growth temperature(10,20,30,37 and 45?)on the phospholipid,fatty acid composition of cytoplasmic membrane and cytoplasmic membrane fluidity,which was aiming to explore the relationship between the efficacy of PEF and the structure and properties of cytoplasmic membrane.In addition,in this paper,the effect of PEF on lipids and oxidative injury of S.Typhimurium was investigated to provide more information on the mechanism of micro-organisms inactivated by PEF.1.The results of study on differences in PEF resistance of S.Typhimurium at different growth temperature and growth phase showed that the PEF sensitivity of S.Typhimurium cells at the exponential phase was higher than those at stationary phase.During the stationary phase,an increase in growth temperature,resulted in a lower reduction of Log10 cycles.In addition,three models(first-order kinetics,Weibull and log-logistic model)were conducted to fit the survival curves of S.Typhimurium and Weibull model produced the best fit to the survival curves.2.The methods of gas chromatography and fluorescence anisotropy were used to investigate the effect of growth temperature on fatty acid composition of cytoplasmic membrane and cytoplasmic membrane fluidity S.Typhimurium cells at the stationary phase.Results indicated that increase of growth temperature resulted in UFA/SFA and cytoplasmic membrane fluidity.In addition,phospholipids phase transition temperature was analyzed by differential scanning calorimetry,which revealed that the increment of growth temperature leaded to increase in phase transition temperature of phospholipids.Thus membrane fluidity was the dominant factor for PEF resistance of S.Typhimurium.3.Quantitative real-time PCR(qPCR)analysis was conducted to investigate the transcription levels of fatty acid biosynthesis-associated genes and alternative sigma factors,aiming to explore the mechanism of S.Typhimurium inactivated by PEF.Results indicated that cfa and fabD genes were down-regulated,while the transcription of fabA was up-regulated when cells were cultivated at relatively lower temperature,which altered the lipids composition,contributing to the increase of membrane fluidity.Despite the fact that alternative sigma factors(rpo S,rpoE and rpoH)showed an obvious relationship with growth temperature,these genes were not essential for S.Typhimurium to develop PEF resistance.4.The objective of research was S.Typhimurium before and after PEF treatment.Results showed that PEF treatment decreased membrane fluidity,possibly due to the down-regulation of cfa and fabA gene expressions and the up-regulation of fabD gene expression,which was characterized by the composition of membrane lipid(decrease in UFA/SFA).Results of phospholipid conformation of S.Typhimurium by Raman spectra indicated that after PEF treatment,both of the intensity ratio of I1139/I1082 and I2874/I2962 increased,which also revealed decrease in membrane fluidity.In addition,oxidative injury induced by PEF treatment was associated with an increase in the content of MDA.The up-regulation of cytochrome bo oxidase gene expressions(cyoA,cyoB and cyoC)showed that membrane damage was induced by PEF treatment which was related to the repairing mechanism of alleviating the oxidative injury caused by PEF treatment. |
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