Effects Of L-Methionine On The Biofilm Formation Of Pseudomonas Aeruginosa Infected By Polyvinyl Chloride(PVC) Catheters

Objective:To explore the effect of L-methionine(L-Met)on Pseudomonas aeruginosa biofilm infected by PVC catheter in vitroMethods:1?Construction of Pseudomonas aeruginosa biofilm in vitro on PVC catheter surface:The formation process of biofilm was monitored by crystal violet assay technology.2?Pseudomonas aeruginosa was cultured in an untreated control group(O?M)and 0.125?M,0.25?M,0.5?M,1?M,2?M,4?M L-Methionine(L-Met)concentration medium for 72 hours,and PVC was added for drainage A catheter was used to construct an in vitro Pseudomonas aeruginosa biofilm model,and the growth kinetics of pathogenic bacteria was measured using XTT after 72 hours of culture;crystal violet was used to semi-quantitatively determine the thickness of biofilm formation at 6 hours,12 hours,24 hours,48 hours,and 72 hours,respectively.The distribution and structure of pathogenic bacteria on the surface of biofilm were observed by electron microscope.3?The mature biofilm was formed by adding PVC drainage catheter to TSB medium containing Pseudomonas aeruginosa for 72 hours,and then the untreated control group(0?M),mixed drug group(0.5?M L-Met+2.5p.g/mL ciprofloxacin),methionine group(0.5?M L-Met)and ciprofloxacin group(2.5?g/mL ciprofloxacin)were replaced for 24 hours,and XTT was used.Semi-quantitative determination of biofilm formation thickness by crystal violet;Scanning electron microscope was used to observe the distribution and structure of pathogens on biofilm surface.Results:1?Crystal violet detection during the construction of in vitro pseudomonas aeruginosa biofilm model,it was found that the curve of OD570 value from 6 to 48 hours presented an upward trend,and the curve of OD570 value from 48 to 72 hours presented a slight downward trend.2?XTT test showed that the OD450 value in the L-Methionine group of 0.125?M,0.25?M,0.5?M,1?M,2?M,and 4?M was not statistically significant compared with the untreated group.3?The crystal violet test showed that the OD570 value of only 0.5?M L-Methionine group showed an upward trend during the 6 to 24 hours culture period and was lower than that of the control group at each time period,and decreased during 24 to 72 hours.The ODs70 value at 72 hours was significantly lower than that of the control group,with statistics.Significance(P<0.05).The 0?M,0.125?M,0.2?M,1?M,2?M,and 4?M L-Methionine group OD570 value curve showed an upward trend at 6 to 48 hours,reached the plateau at 48 to 72 hours,and had no statistical significance with the control group.4?Scanning electron microscope observation found that only 0.5?M L-Methionine group of Pseudomonas aeruginosa bacteria aggregates significantly reduced,scattered distribution,visible bacteria in the floating state,the remaining groups of bacteria in the biofilm agglomerates,it is surrounded by thick cellulose-like mucus.5?After constructing the PVC catheter Pseudomonas aeruginosa biofilm in vitro model and replacing the culture medium for continuous culture for 24 hours,XTT detection found that OD450 in the mixed drug group was lower than that in the control group,with statistical significance(p<0.05).6?After constructing the PVC catheter Pseudomonas aeruginosa biofilm in vitro model and replacing the culture medium for continuous culture for 24 hours,crystal violet detection found that OD570 in methionine group and mixed drug group was lower than that in control group,with statistical significance(p<0.05).7?Scanning electron microscope showed that the bacteria in the biofilm of Pseudomonas aeruginosa in the mixed drug group were scattered,the bacteria in the floating state were significantly reduced,and the morphology of the bacteria was destroyed.It can be seen that some scattered debris-like substances,thick cellulose-like mucus substances around bacteria and mucus silk-like substances between bacteria disappeared,and the three-dimensional layered network structure also disappeared.Conclusions:1?Pseudomonas aeruginosa can form biofilm on the surface of PVC catheter in vitro.2?L-methionine can inhibit the formation of Pseudomonas aeruginosa biofilm on PVC catheter surface in vitro and decompose the formed biofilm,but L-methionine has no killing effect on Pseudomonas aeruginosa.3?L-Methionine(L-Met)can improve the sensiti vity of Pseudomonas aeruginosa to ciprofloxacin,so low concentration of ciprofloxacin combined with L-Methionine can be considered to treat Pseudomonas aeruginosa biofilm infection on PVC catheter surface.