Bactericidal Effect Of Blue Light On Salmonella And Effect Of PagL Expression On Germicidal Efficiency

Salmonella is an important food-borne pathogen in the world.The outbreak of S.enteritidis contamination in poultry eggs poses a severe challenge to human health,so it is urgent to develop effective sterilization technology.Blue light is a new sterilization technology developed in recent years,which hardly affects food quality while sterilizing.However,there is no report on the sterilization of S.enteritidis in poultry eggs by blue light.Therefore,this paper applies blue light to sterilization and preservation of S.enteritidis in poultry eggs for the first time,and systematically studies the sterilization efficiency and quality of poultry eggs.In addition,the germicidal mechanism of blue light has just started,and which structural macromolecules and related genes of bacteria affect the sensitivity to blue light need to be explored.In the early stage of this experiment,it was found that the modification of bacterial lipopolysaccharide(LPS)had a great influence on the germicidal efficiency of blue light.Therefore,this paper also studied the change of blue light sensitivity after overexpression of pag L(fatty acid chain modification gene on LPS lipid A),and explored its mechanism.S.enteritidis was inoculated on the surface of eggs,and then irradiated by blue light,so as to achieve the sterilization of S.enteritidis on the surface of eggs.The inactivated eggs were stored,and samples were taken every 7 days.The colony count of egg shells and the index of eggs were measured,and then the preservation effect of blue light on eggs was observed.On the other hand,the changes of sensitivity of S.typhimurium to blue light,and the changes of cell membrane permeability and lipids of wild-type and mutant strains were studied after pag L expression.The results are as follows:(1)Blue light has a good inactivation effect on S.enteritidis.Firstly,aiming at the bacterial suspension,the dose of blue light irradiation was 0-20 J·cm^-2,the density of bacteria did not change obviously,but began to decrease when it exceeded 20 J·cm^-2.When the irradiation dose was 240 J·cm^-2,the bacterial biomass decreased by more than 2 log₁₀ CFU·ml^-1,and the mortality rate reached 99%.When the irradiation dose reached 360 J·cm^-2,the bacterial density of S.enteritidis decreased rapidly,with a decrease of 5.19 log₁₀ CFU·ml^-1.Blue light also had good inactivated efficiency for S.enteritidis in eggs.The blue light irradiation dose of 54.6 J·cm^-2 reduced the number of colonies on eggshell surface by 3.73 log₁₀ CFU·egg-1,and prolonged the storage period of eggs to more than 5weeks.(2)Compared with the control group,the effect of blue light irradiation on egg quality during sterilization include:1)Reduce weight loss,and in the fifth week,the weight loss of eggstreated with blue light was 3.09±0.23%,while that of control was 3.88±0.13%,which was significantly higher than that of eggs treated with blue light by 25.6%;2)The yolk was fresher.The egg yolk index of the eggs in the two groups showed the same trend in the first 4 weeks.After 5 weeks,the yolk of blue light treated eggs was complete,while the yolk of control group was scattered;3)Slow down Huugh unit reduction.The Haugh unit of eggs treated with blue light decreased from 86.27±5.89 to 45.66±2.64,while that of control group decreased to 0.4)The change of albumen p H of eggs had little difference,showing a trend of increasing at first and then decreasing,but it was always higher than the initial value.5)There are few plaques on the surface of eggs,and the plaques were smaller.However,the plaques on the surface of eggs in the control group are larger and more;6)Slow down the decrease rate of amino acid level in eggs.In the blue light group,it decreased from 84.58±0.35g·(100 g)^-1 to 76.4±0.32 g·(100 g)^-1,and in the control group it decreased to 72.9±0.27 g·(100 g)^-1.These results indicated that blue light was helpful to improve eggs quality.(3)The mass spectrum results of LPS lipid A showed that the m/z value of wild-type strain S.typhimurium SL1344 was 1955,while the m/z values of S.typhimurium SL1344/p WSK29-pag L included 1728 and 1955.the former showed that Pag L catalyzed the removal of C14 fatty acid chain at position 3,while the latter showed that some lipid A was not acted by Pag L.Compared with SL1344,SL1344/p WSK29-pag L had higher sensitivity to blue light.At the irradiation dose of 120 J·cm^-2,the bacterial biomass of S.typhimurium SL1344/p WSK29-pag L decreased by 2.107 log₁₀ CFU·ml^-1,with a mortality rate of over 99%,while that of S.typhimurium SL1344 decreased by 1.679 log₁₀ CFU·ml^-1,with a mortality rate of 97.9%.The hydrophobicity of SL1344/p WSK29-pag L was 4 times higher than that of wild type.The membrane permeability of mutant SL1344/p WSK29-pag L was 1.39 times higher than that of wild type.The lipidomic data showed that the mutant SL1344/p WSK29-pag L contained cardiolipin(CL),phosphatidylethanolamines(PE),phosphatidylserine(PS),monoacylglycerol(MG),ceramide,(Cer)and monogalactosyl diglycerids(MGDG)were higher,and the wild strain SL1344 did not contain digalactosy diglycerids(DGDG).It shows that the change of lipid group may be one of the key reasons for the increase of blue light sensitivity.