Pollution Controlling Of Enterobacter Sakazakii In Infant Milk Powders And The Rapid Detetion

Enterobacter sakazakii was an important conditional pathogenic bacteria that can cause life-threatening meningitis,fatal enterocolitis and sepsis and other diseases in neonates and infants.Mortality rates caused by Enterobacter sakazakii had been reported to be higher than 50%,so the Enterobacter sakazakii was the key control index of infant formula milk powder.At present,Enterobacter sakazakii had gradually been widely concern in the world,but the domestic research was still relatively few.The pollution control of Enterobacter sakazakii had not been perfect.Based on the biological characteristics of Enterobacter sakazakii,the heat resistance and germicidal efficacy of disinfectant were analyzed and experiented in this paper.Three different Enterobacter sakazakii strains of 3305373standard,3305373 and 3205373 were choosed in the course of the study.The bacteria was studied for 12min～24min at the temperature 63℃,65℃ and 67℃ respectively,and 10 s～30 s at the temperature 87℃and 93℃.The experimental results showed that three kinds of Enterobacter sakazakii were killed by 24min,21min,and 15min at the temperature of 63℃,65℃,67℃,and 25 s and 15s at the temperature of 87℃ and 93℃,respectively.Several types of disinfectants in dairy production process were used in this study,such as sodium hydroxide,green pioneer,benzalkonium bromide,bleaching powder,alcohol.The Enterobacter sakazakii was soaked,inoculated and cultivated in the disinfectant of different concentrations,The results showed that sodium hydroxide had best disinfection effect compared with alcohol,benzalkonium bromide,bleaching powder The bactericidal concentration of benzalkonium bromide and ethanol required was longerThe fluorescence quantitative PCR and an advanced way called immune sensor were established to detecte the Enterobacter sakazakii in this experiment.As the universal primers,conserved sequences of 16S rDNA and 23 S rDNA from bacteria was used to establish the fluorescence quantitative PCR and their specificity and sensitivity were tested also.Different brands of milk powder were studie as actual case.Five strains of bacteria was tested to Verify the specificity of the primer.The nano-specific compound was immobilized on glassy carbon electrode surface.Then,the immune senser was prepared by using a way of physical or bovine serum albumin-glutaraldehyde cross-Linking method,in which case horseradish peroxidase was immobilized on the antibody of the bacteria..The optimized results were 0.5 mmol/L hydrogen peroxide,PBS solution of pH 7.0 and 20 min incubation time.In addition,the linear relationship between concentration and response current were determined by differential pulse voltammetry and the linear relationship were ΔIpc(μA)=1.182 lg[C/cfu.mL-1]-1.723,R2=0.998 9,With the detection limit was 5.5×101 cfu/mL(S/N).The results showed that the immune sensor method was agreet with these national standard method.Therefore,the immune sensor method can be successfully applied in the detection of Enterobacter sakazakii.Compared with the national standard method,these two test methods presented many advantages,such as short period,high sensitivity and simple operation and can be applied on the detection of food safety as new detection technology.