| Objective: Observe whether the Staphylococcus epidermidis isolated from the secretions of ostiomeatal complex in normal human and “stagnated heat of gallbladder” type nasosinusitis patients can form bacterial biofilms.The effect of Biyuanshu oral liquid on the biofilm formation of Staphylococcus epidermidis in patients with chronic rhinosinusitis before and after treatment was studied and the formation of Staphylococcus epidermidis biofilms was observed by laser confocal microscopy.Methods: 1.A tall of 15 patients with chronic nasal-sinusitis(stagnated heat of gallbladder)and 10 cases of healthy subject.And give the patients Biyuanshu oral liquid(Huashenjituan),3 times a day,one at a time,one each time for 2 weeks of treatment.while healthy people do no treatment.2.The secretions of the chronic rhinosinusitis patients treated with Biyuanshu oral liquid before and after were collected,and then normal culture was performed.The isolated Staphylococcus epidermidis samples were preserved and frozen.3.The Congo red medium was used to screen polysaccharide adhesin(PIA)positive expression strains.4.The 96-well enzyme labeled crystal violet method was used to determine the biofilm-producing ability and growth of Staphylococcus epidermidis,a and the changes of the epithelial membrane of Staphylococcus epidermidis before and after the treatment of patients with chronic rhinosinusitis were compared.5.The Confocal microscopy was used to observe the bacterial biofilm formation in Staphylococcus epidermidis.Results: 1.The qualitative test of Congo red bacterium culture: among the 15 strains before treatment,10 were PIA+ strains;in the 15 strains after treatment,6 were PIA+ strains;3 strains of PIA+ in the 10 strains of control group.2.The quatitative test of 96-well enzyme labeled crystal violet method: Before treatment of CRS patients,the membrane-forming strains accounted for 73% in 15 strains of staphylococcal epidermis of the nasal-sinus sinus;and 15 cases of nasal sinus Staphylococcus epidermidis after treatment,40% of which were membrane-producing strains;10 strains of Staphylococcus epidermidis in normal human rhinasanasal sinus,of which 30% were biofilm producing strains.There was no significant difference in the growth rate of free bacteria between group before treatment of PIA+ and group after treatment of PIA-at the time of 0h,1h,2h,18h(P>0.05).In 4h,8h and 24 h,there was a significant difference in the growth of free bacteria between the PIA+ group before treatment and the PIA-group after treatment(P<0.05).There was significant difference between the two groups of bacteria growth in group PIA+ group before treatment and the PIA-group after treatment at 8h,18 h and 24h(P<0.05).At 24 h,there was significant difference of growth in adherent bacteria between the PIA+ group before treatment and the group of PIA-after treatment(P<0.01).The growth of PIA+ group before treatment was not significant difference compared with that of PIA-group after treatment at 0h,1h,2h,and 4h(P>0.05).3.Observation by Confocal Laser Scanning Microscopy: In 4h,Staphylococcus epidermidis can see the scattered bacteria marked as red on the cover glass,after 4h,the number of bacteria gradually increased,and Green fluorescence surrounds the bacteria,which indicates that the BF begins to form.Bacteria are more concentrated in 20 h and gradually formed an initial biofilm.In 24-48 h,large-scale biofilms are formed,and a lot of polysaccharides are formed around the bacteria,which are inlaid closely,suggesting that the biofilm has been differentiated and mature at this time.After 72 h,the amount of bacteria adhered to the slide and the extracellular polysaccharide of bacteria decreased,and the biofilm structure gradually disintegrated,and then entered the depolymerization and re colonization period.Conclusion: 1.The Staphylococcus epidermidis isolated from patients with chronic rhinosinusitis and healthy people,have the ability to form bacterial biofilm,and the rate of membrane production of CRS patients was higher than that of the healthy people.The expression rate of PIA decreased after treatment compared with before treatment.There was no significant difference between qualitative and quantitative tests in judging the ability of producing membrane.2.Biyuan Shu oral liquid inhibits the formation of Staphylococcus epidermidis biofilm.3.The dynamic process of Staphylococcus epidermidis can be observed under confocal laser scanning microscope. |
PDF Full Text Request