Formation And Regulated Mechanism Of Biofilm In Streptococcus Agalactiae Isolated From A Dairy Cow With Mastitis

The mastitis is a frequently diseasein the dairy farm,it has a great impact on the economic benefits of dairy farm.Streptococcus agalactiae also known as Group B streptococcus(GBS),is one of the cow mastitis pathogenic bacteria,which belongs to streptococcus and gram positive bacteria.The pathogenic of Streptococcus agalactiae is closely related to the virulence factors it producted,and the types and quantities of virulence factors are differences between bacterial strains which come from different host origins.Now it is widely believed that some virulence factors of Streptococcus agalactiae play an important role in adhering and invading the body cells and they are easy to form biofilm on the surface of bovine mammary.Bacterial biofilms interferethe normal immune function of organisms and cause diseases.Chinese medicine contains many alkaloids,boots,polysaccharides,essential oil,soap,organic acids,vitamins,nutrilite,mineral substance and etc.The function of these substances is regulating the body’s immune system,directly effects on the microbial pathogens and repairs the damaged tissues and organs.Furthermore,the Chinese medicine also has these characteristics of low toxicity,low residue and low drug resistance,so it has wide prospect in the treatment of bacterial biofilm disease.In this study,through the research of formation and regulated mechanism of biofilmof Streptococcus agalactiaeprovide experimental evidencefor thetreatment of dairy cows with mastitis.1.Reserach the most suitable biofilm formation of Streptococcus agalactiae To discoverthe most suitable biofilm formation of Streptococcus agalactiae,in this study,we contrast several different culture medium.The results showedthat TSB + 1%Glucose medium was more suitable for generating biofilm of Streptococcus agalactiae.After that,using TSB + 1% Glucose(add 1%milk)medium to culture gbs,we found that the milk enhance biofilm formation of Streptococcus agalactiae.The most suitable culture of biofilm formation of Streptococcus agalactiae is TSB + 1%Glucose(add 1%milk).2.The effect of the water extract and ethanol extract of the two Chinese medicine on Streptococcus agalactiae Through the water extract and ethanol extract of cnidium and solanum nigrum,add the extracts to the culture plate of Streptococcus agalactiae.We found that the water extract of solanum nigrum inhibit the growth of Streptococcus agalactiae,cnidium extracts had no effect on the growth of Streptococcus agalactiae.3.The effect of water extract of Solanum nigrum on biofilmof Streptococcus agalactiae The 15 mg / m Lwater extract of Solanum nigrum was added to thebroth of Streptococcus agalactiaeunder the TSB + 1% Glucose culture medium with milk,it will not form biofilm.In the TSB + 1% Glucose medium without milk,the concentration is7.5mg/m L.4.Search for theregμLate biofilm genes ofStreptococcus agalactiae After a review of relevant literature,we foundthat the regulate biofilm genes of Streptococcus agalactiae arecps E and neu A,obtaining the gene sequences of cps E and neu Aby experiment.5.The regμLatory mechanisms of water extract of solanum nigrum to the biofilm formation of Streptococcus agalactiae Abstract the RNA of Streptococcus agalactiae that add the water extract of solanum nigrum,reverse transcription and real-time PCR.The results showed that the water extract ofsolanum nigrum inhibited the gene expression of cps E andneu A.To study the biofilm forming condition of Streptococcus agalactiae,we confirmed the regulatory genes of biofilm are cps E and neu A.Observing the effect of Chinese medicineon Streptococcus agalactiae growth and biofilm formation,we found that the water extract of Solanum nigrum has a great impact on the biofilm formationof Streptococcus agalactiae.From the result of real-time PCR,we found that the water extract of Solanum nigrumcan influence the expression of cps E and neu A,and then inhibit the biofilm formation of Streptococcus agalactiae.