Inhibitory Actions of Biofilm-degrading Enzymes against the Biofilm Formed by Vibrio parahaemolyticus on 304 Stainless Steel
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Abstract:
Vibrio parahaemolyticus biofilm matrix was used as the target, and cellulase, lipase and protease K were screened and selected as single enzymes by the crystal violet staining assay using 96-well plates. These single enzymes were combined to form composite enzymes. On the basis of single factor experiments, response surface methodology was used to optimize the concentration of the combined enzymes. The inhibitory actions of the combined enzymes against Vibrio parahaemolyticus biofilm on 304 stainless steel were evaluated by the crystal violet staining assay, colony counting and scanning electron microscopy. The composite enzymes obtained by response surface optimization with 1.00 mg/mL of cellulase, 1.10 mg/mL of lipase and 1.50 μg/mL of protease K exhibited the highest inhibition rate (89.73%). The rate of inhibition for the composite enzyme against the Vibrio parahaemolyticus biofilm significantly increased compared with the single enzyme, which was generally in accordance with the predicted value. These results indicated that the model fitted well and the concentration of each enzyme in the composite enzyme was accurate and feasible. The optimized composite enzymes inhibited effectively the growth of Vibrio parahaemolyticus biofilm on 304 stainless steel with the inhibition rate as 59.35%. Examinations by scanning electron microscopy also confirmed the inhibitory effect of the composite enzymes. The composite enzymes can effectively increase the rate of inhibition against biofilm, which provides new ideas for the degradation and elimination of pathogenic bacterial biofilms.
