Research On Screening And Bacteriostatic Effect Of Antibacterial Active Substance Of Chicken Escherichia Coli In Vitro

Chicken-derived colibacillosis is an infectious disease caused by pathogenic E.coli.According to the combination of bacterial antigen(O),surface antigen(K)and flagella antigen(H)of E.coli,it can be divided into different serotypes.There are about thousands of serotypes known so far.According to research,chicken E.coli caused serious harm to chickens through different pathogenic pathways.The cause of the disease was mostly poor sanitation in the poultry house,which caused the cloaca of the chicken to be contaminated by E.coli and invaded the oviduct,which would affect The function of the fallopian tube,dysplasia of the follicle,and problems in the protein secretion process will ultimately affect the egg quality and egg production rate.Chicken salpingitis will not only reduce the laying rate of laying hens,but also cause economic losses to farmers.E.coli from chicken is the most common pathogen in veterinary clinical infectious diseases.Its serotypes are numerous and the immune protection between bacteria is poor,which leads to the dependence of the disease on antibiotic treatment.However,with the unreasonable use of antibiotics in veterinary clinics,The chicken E.coli gradually appeared drug-resistant strains,which brought challenges to clinical treatment and aggravated the residue of antibiotics in poultry products,which seriously endangered animal food safety.Therefore,screening new active substances to replace antibiotics has become a research focus.This experiment uses bacterial isolation and culture and PCR technology to isolate and identify chicken E.coli from the oviduct of chickens,and use this as an indicator bacteria to study the in vitro antibacterial effects of probiotics,polysaccharides and traditional Chinese medicines.Select Bacillus subtilis and Lactobacillus plantarum with the best antibacterial effect,use high performance liquid chromatography and mass spectrometry to separate and identify antibacterial peptides,and determine the antibacterial activity and minimum inhibitory concentration of their crude extracts respectively.Study its stability to temperature,pH and light.The results of the study showed that the E.coli RCB592 strain of chicken origin was isolated and identified from the oviduct of chickens.The results of the in vitro antibacterial test showed that the antibacterial effect of a single probiotic is Bacillus subtilis and Lactobacillus plantarum.The diameter of the bacterial circle was(20.18±0.14)mm and(17.55±0.57)mm,which were significantly higher than the diameter of the inhibition zone of the compound probiotics(P<0.05).Chitooligosaccharides and Astragalus polysaccharides had no antibacterial effect(P>0.05).In vitro antibacterial test of Chinese medicine in Coptis and Poria,Coptis and Red Peony,Coptis and Honeysuckle,Coptis and Houttuynia,Combination of Coptis and Cinnamomi,Coptis and Poria and Red Peony,Coptis and Poria and Houttuynia,Coptis and Red Peony The antibacterial effect of the combination with honeysuckle was the best(P<0.05).Enterobactin was isolated and identified from extracts of Bacillus subtilis and Lactobacillus plantarum.The contents of enterobactin were 27.68 mg/g and 11.45 mg/g analyzed by high performance liquid chromatography.After analysis by liquid chromatography and mass spectrometry,the 6 amino acid sequences of Bacillus subtilis enterobactin belong to the SdpB family of antibacterial peptides and antibacterial peptide system proteins of various species;the 2 amino acid sequences of Lactobacillus enterobacterin belong to Ⅱc Cyclic bacteriocins phytocyclins and V bacteriocins.The crude extracts of Bacillus subtilis and Lactobacillus plantarum had good antibacterial activity against E.coli(P<0.05),and the minimum inhibitory concentrations were 6.25 μg/mL and 12.5 μg/mL,respectively.The antimicrobial peptide stability test results show that the antibacterial activity of the crude extract of Bacillus subtilis has good thermal stability,acid-base stability and light stability,and the antibacterial activity of the crude extract of Lactobacillus plantarum has good thermal stability and Light stability.It can be seen that the in vitro antibacterial effect of Bacillus subtilis and Lactobacillus plantarum on E.coli RCB592 strain of chicken origin is better than that of polysaccharides and traditional Chinese medicine.The main antibacterial active substance is enterobactin in antibacterial peptides,and it is clear that enterobactin The amino acid sequence of the antibacterial activity has good thermal stability,acid-base stability and light stability.So as to provide a basis for clinical exploration of antibiotic substitutes and further study of the antibacterial mechanism of probiotics.