Research On Germicidal Efficacy Of The Different PH Electrolyzed Oxidizing Water On Multi-drug Resistant Bacteria And Biofilms

Objectives1、To observe germicidal efficacy of the different pH EOW on MRSA and Multiple drug resistance of pseudomonas aeruginosa which were separated from clinical specimens.2、Choosing the silicone membrane as the carrier to eatablish bacterial biofilm in vitro, in order to explore the formation time of biofilms.3、To observe germicidal efficacy of the different pH Electrolyzed Oxidizing Water on the biofilms of MRSA and Multiple drug resistance of pseudomonas aeruginosa separated from clinical specimens. Methods1、Collected ten strains of Methicillin-resistant Staphylococcus aureus and Multiple drug resistance of pseudomonas aeruginosa separated from clinical specimens, and prepared bacterial suspension. According to the different pH, the EOW was divided into 3 groups, pH < 3.0, pH between 4.0 and 5.0, pH between 6.0 and 7.0, adding the control benzalkonium bromide group and saline group. The bacterial suspension was treated with EOW of different pH for 1 min, 3 min and 5 min respectively. Then, the mixtures were neutralized for 10 min. The final mixtures were cultured to determine the colony forming units of residual viable bacteria and to contrast the sterilizing rates between the 3 groups.2、Biofilms were setted up in vitro. Collected ten strains of MRSA and Multiple drug esistance of pseudomonas aeruginosa separated from clinical specimens, and prepared bacterial suspension. The silicone membrane was autoclaved and putted into 24-well plates filled with the bacterial suspension and TSB, cultivating for 1 day and 3 day. Then biofilm formation was observed by sliver staining and scanning electron microscope.3、Putting the biofilm model into the three groups, and incubating for 5 min, 10 min and 20 min respectively. The mixtures were neutralized for 10 min after incubation. The final mixtures were cultured to determine the colony forming units of residual viable bacteria and to contrast the sterilizing rates between the 3 groups. The morphology of the biofilm was observed by scanning electron microscope and fluorescence microscope. Results1、The average killing rates of MDR-PA in suspension exposed to the EOW of pH < 3.0, pH between 4.0 and 5.0, pH between 6.0 and 7.0 for 1min were 98.51%, 99.53% and 99.74% respectively. Multi-drug resistant P.Aeruginosa in suspension could be killed entirely after exposed to EOW of three groups for 3 min and 5 min. There was no statistically significant difference between the three groups(P > 0.05). When multi-drug resistant P.Aeruginosa in suspension exposed to benzalkonium for 1 min, 3 min and 5 min, the killing rates were 48.71%, 70.75% and 84.51% respectively. There was statistically significant difference compared with the three groups of EOW(P < 0.05). MRSA in suspension could be killed entirely within 1 min, 3 min and 5 min when exposed to the three kinds of EOW and benzalkonium and the killing rates were 100%. There was no statistically significant difference between the four groups(P > 0.05).2、After the biofilms were cultivated for 1 day, silver staining found carrier surface has formed black spots, and scanning electron microscopy showed that silicone membrane surface with scattered colonies gather, which show no formation of biofilms. After cultivated for 3 days, silver staining found carrier surface has formed a sheet of black floc and scanning electron microscopy showed that the surface of the carrier has formed sheet colonies, indicating that the boifilms have formed.3、The average killing rates of multi-drug resistant P.Aeruginosa biofilms exposed to the EOW of the different pH for 5 min and 10 min were above 99%. Multi-drug resistant P.Aeruginosa biofilms could be killed entirely within 20 min when exposed to the EOW of three groups. There was no statistically significant difference between the three groups(P > 0.05). The average killing rates of MRSA biofilms exposed to the EOW of the different pH for 5 min were above 99%, and were above 99.99% for 10 min and 20 min. There was no statistically significant difference between the three groups(P > 0.05).4、After biofilms were processed by EOW, the change of boifilms morphology were observed by Scanning electron microscope, bacteria began to wrinkle slightly 5 min later and wrinkled obviously with rupture individually after 10 min. There were a lot of bacteria ruptured 20 min later. Staining results showed that compared with the control group, after 5 min, 10 min and 20 min, the number of dead bacteria increased significantly, which was consistent with viable counts. Conclusions1、The effect of three pH EOW on MDR-PA sterilization is same and superior to the benzalkonium in the germicidal efficacy remarkably. Although the germicidal efficacy of the three pH EOW and benzalkonium bromide shows no difference on multiple drug-resistat bacteria, the sterilization spectrum of benzalkonium bromide is narrow, so EOW was recommended as the disinfection of multiple drug-resistant bacteria infection wound and object surface.2、Bacterial biofim was formed on silicone membrane after 3 days of culturing.3、EOW of 3 different p H hold the same germicidal effect on the biofilm of MDR-PA and MRSA.The EOWof pH between 6.0 and 7.0 is recommended as disinfectant of multiple drug-resistant bacteria infection or biofilm formation wound, due to non-corrosivity and stable available chlorine concentration.