Study On The Effect Of Maggot Antibacterial Peptide On Diarrhea In Mice With Escherichia Coli

Object:calf diarrhea is one of the most common and infectious diseases in cattle breeding.Currently,it is mainly controlled by antibiotics.However,due to the long-term and large-scale use of antibiotics,the emergence of drug-resistant strains not only reduces antibiotics.The therapeutic effect also brings problems such as drug residues,which indirectly harm people's health.Therefore,researchers have been searching for related alternative or auxiliary drugs,and the research on antimicrobial peptides(AMPs)has received wide attention from researchers.However,since calf is a large-scale economic animal,there are many problems in directly using calf as a test subject.However,mice are often used as an important model animal in the medical field because of their small size and short reproductive cycle.Therefore,mice can be used as sputum.Replacement model animals for studies of cattle diarrhea infection,morbidity and treatment.Antibacterial peptide is a kind of small molecule polypeptide with antibacterial effect produced by the organism against external pathogen invasion.Its antibacterial mechanism is different from that of antibiotics,it does not produce drug residues,and it is not easy to produce drug resistance.The large number of easily-producing maggots in the biological world are rich in antibacterial peptides,which can grow healthily in harsh environments such as rot,feces and garbage containing large amounts of bacteria.Therefore,using Musca domestica larvae and maggots as experimental materials,the pathogenic Escherichia coli isolated from the faeces of calf diarrhea was used to induce the high-expression of the maggot antibacterial peptide from the maggots,in order to study the replacement or supplement of the maggot antibacterial peptides.The therapeutic effect of antibiotics on calf diarrhea caused by pathogenic Escherichia coli.In this experiment,pathogenic Escherichia coli was isolated from the feces of calf diarrhea,and the isolated Escherichia coli was administered to mice to establish a pathological model of Escherichia coli-induced diarrhea in mice.Mice treated with maggot antibacterial peptide in the treatment of diarrhea,observe the changes of clinical symptoms in the treatment of mice,detect the tumor necrosis factor(TNF-?)content in the duodenum and the number of E.coli and lactobacilli colonies The changes,as well as the content of IL-6 in the serum of mice and the cure rate of diarrhea in mice,provide a theoretical basis for the future replacement of muscarinic antibacterial Method:Test 1: Isolation and Identification of E.coli Caused by calf DiarrheaThe feces of diarrhea in 10-day-old calf were collected aseptically,and the bacteria were isolated and cultured.The isolates were stained and microscopically examined,PCR-identified,and the isolates were tested for pathogenicity in mice.Test 2: Study on antibacterial activity of mycelium antibacterial peptide against E.coli caused by calf diarrhea(1)crude reference purification of maggot antibacterial peptide;(2)determination of antibacterial peptide concentration of maggots by Coomassie brilliant blue method;(3)determination of minimum inhibitory concentration of antibacterial peptides of maggots;(4)KB paper method Sensitivity tests for E.coli for muscarinic peptides and enrofloxacin.Trial 3: Establishment of diarrhea model of E.coli mice with calf diarrheaThe pathogenic Escherichia coli isolated from the faeces of calf diarrhea was prepared and injected into mice with different concentrations of E.coli suspension.The clinical symptoms of the mice,the changes in body weight of the mice and the determination of the large intestine in the feces of diarrhea mice were recorded.The number of bacilli,mouse mortality and diarrhea rate were used to construct an E.coli mouse diarrhea model.Test 4: Therapeutic effect of maggot antibacterial peptide on diarrhea in Escherichia coli mice(1)The test mice were divided into 4 groups: blank control group,normal saline group,antibiotic(ennofloxacin)group and maggot antibacterial peptide treatment group.The mice in each experimental group were collected at treatment for 1 day,3 days and 5 days.Duodenal tissue samples were observed for histopathological changes and therapeutic effects;(2)The content of TNF-? in the duodenum and the number of E.coli and lactobacilli colonies were detected by ELISA.(3)The content of IL-6 in mouse serum was determined by ELIS.Result:1.The isolates from the calf feces were isolated and cultured,stained microscopically,PCR-identified,and pathogenicity of mice were obtained.Pathogenic Escherichia coli caused by calf diarrhea was obtained.2.The results of the drug sensitivity test of the maggot antibacterial peptide against calf diarrhea in Escherichia coli showed that the antibacterial peptide of the maggot antibacterial peptide against the calf diarrhea E.coli was(12.72�0.69mm)lower than the antibiotic(ennofloxacin).The inhibition zone(19.87�0.51mm);the minimum inhibitory concentration of mycelium antibacterial peptide and antibiotic(ennofloxacin)induced by E.coli induced by calf diarrhea in Escherichia coli was 2.19 ug./mL,1.56ug/mL.3.The mice were intragastrically administrated with 1�108 CFU/mL,2�108 CFU/mL,3�108 CFU/mL,4�108 CFU/mL,5�108 CFU/mL E.coli suspension,and the clinical symptoms of the mice were changed.The decrease in body weight and the increase in the number of E.coli in the feces of diarrhea mice,the increase in diarrhea rate and mortality,and the concentration of E.coli induced by diarrhea in mice were 4�108 CFU/mL,and the pathological model of mouse diarrhea was successfully constructed.4.Mice diarrhea peptide was used to treat mouse diarrhea model.The duodenum tissues of mice were collected at 1d,3d and 5d,respectively.The TNF-? and mouse serum IL-6 in the duodenum were detected.The content and the number of E.coli and lactobacilli in the duodenum of mice and the cure rate of diarrhea in mice.(1)The TNF-? content in the duodenum of mice was detected by ELISA.The results showed that compared with the saline group,the mice were treated with the antibacterial peptide treatment group on the 1st,3rd,and 5th day.The content of TNF-? in the intestine decreased significantly(P<0.01).(2)The mice with diarrhea were treated with maggot antibacterial peptide,and the viable counts of E.coli and lactobacilli in the duodenum of mice were found.During the treatment,the number of E.coli in the duodenum of the mice treated with the antibacterial peptides of the maggots in 1d,3d and 5d was significantly decreased(P<0.01),and the number of lactobacilli in the duodenum of mice was extremely high.Significantly increased trend(P <0.01).(3)After treatment,the serum IL-6 content in mice was detected by ELISA.The results showed that the levels of IL-6 in the antibiotic treatment group and the antimicrobial peptide treatment group were significantly lower than those in the saline group(P<0.05).(4)The cure rate of mice was inferred based on the number of E.coli in the duodenum of mice: the cure rate of the mice treated with saline was 27%,and that of the antimicrobial peptide group was 46%.Antibiotics(ennofloxacin)The treatment group was 60%Conclusion: The maggot antibacterial peptide has a certain therapeutic effect on calf diarrhea in mice with Escherichia coli diarrhea.Although the cure rate of the antibacterial peptide of maggots is lower than that of the antibiotic(ennofloxacin)group,the antibacterial peptides and antibiotics of maggots are also small for diarrhea.The mouse has a therapeutic effect,indicating that the maggot antibacterial peptide can be used to treat calf diarrhea.