Cronobacter Sakazakii Biofilm Formation And Anti-microbial Photodynamic Technology With Toluidine Blue O

The Cronobacter sakazakii?C.sakazakii?could be isolated from a wide spectrum of food,like vegetables,cheese,meat and formula milk,causing infection of infant formula milk.In this study,the biofilm formation factors of C.sakazakii biofilm and its elimination by ultrasound treatment on glass surfaces were investigated through turbidimetric determination and plate colony counting method.The effect of anti-microbial photodynamic technology?APDT?against C.sakazakii biofilm was optimized using toluidine blue O?TBO?.The damage effect and morphological changes of APDT on C.sakazakii biofilm cells were observed by confocal laser scanning microscopy?CLSM?and scanning electron microscopy?SEM?.The influence of chitosan on the biofilm formation of C.sakazakii and its effect on APDT were also studied.The main result are as follows:1.Culture conditions have impact on the formation of C.sakazakii biofilm.Results showed that the optimal condition for C.sakazakii biofilm formation was 37?,at the pH 7.5 and the salinity 2.5%.CaCl₂?0.50%?and MgCl₂?1.50%?reached the most obvious promotion effect on C.sakazakii biofilm formation.Cu²⁺?1.50%?and EDTA?2.5%?have an inhibitory effect on the formation of C.sakazakii biofilm formation.2.Ultrasonic treatment has a significant effect on the elimination of C.sakazakii biofilm.C.sakazakii biofilm attached to the glass surface was eliminated by ultrasonic treatment,The elimination effect was the best when initial temperature at 30?and time at 14 min.The amount of living cells variety change of biofilm by ultrasound treatment was observed on the glass surface and elimination effect of ultrasonic treatment was proved and evaluated by CLSM.3.The efficacy of TBO-APDT against C.sakazakii biofilm was noticeable.The optimum culture condition obtained in this study were an incubation time 20.5 min,a PS concentration 56?g/mL,and an irradiation time 30.5 min,resulting in the theoretical value of log-transformed of CFU/cm²was 3.44.4.CLSM provided direct evidence for inactivation effect of APDT on C.sakazakii biofilm cells from the fluorescent color of the bacteria.When the irradiation time was longer,most cells showed red fluorescence,indicating that the inactivation effect was significant.SEM were employed to intuitively analyze external cell structure.The cells were damaged and the cell contents were dissolved.5.Chitosan significantly inhabited the C.sakazakii biofilm formation.Meanwhile,chitosan has a certain disruption effect on mature biofilm.Chitosan also has a promotion of TBO penetration for cells,increasing the inactivation effect.Therefore,The combination of APDT and chitosan may be widely used in the food industry to control biofilm.In summary,TBO-APDT can effectively control C.sakazakii biofilm,therefore,APDT has potential application in food safety particularly in the safety of dairy product processing..