Removal of Pseudomonas putida biofilm and associated extracellular polymeric substances from stainless steel using simulated clean-in-place system
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Pseudomonas putida is a common food contact surface biofilm producer, producing high amounts of extracellular polymeric substances. The purpose of this study was to develop a method for observing Ps. putida biofilm and associated EPS remaining after clean-in-place treatment and determine the ability of CIP to remove EPS from stainless steel. Stainless steel coupons were soiled with 3-day Ps. putida biofilm growth. Samples were cleaned using a simulated CIP system. DNA staining using Hoescht 33258 and EPS staining using Lectin PNA were compared on cleaned coupons cleaned using 1.28%, 2.0%, 4.0%, and 6.0% sodium hydroxide at 66°C for 3 min followed by a sterile water rinse, neutralizing in phosphate buffer and viewed under a epifluoerescent microscope. The effectiveness of cleaning for removing EPS was also determined using 1.5% and 2.0% sodium hydroxide at 68°C and 70°C. Sodium hydroxide concentration of 1.28% at 66°C is sufficient to remove Hoescht but not Lectin binding material. A minimum of 2.5% sodium hydroxide at 66°C was sufficient to remove Hoescht and Lectin binding material.