| Antibacterial peptides (ABPs) are important resources from which it may be able to develop novel therapeutic agents against the ever growing number of antibiotics resistant microorganisms. The small intestine of pig may possess one or more antimicrobial peptides that could be developed as excellent therapeutic agents. After the fresh small intestines of pig were treated by grinding, ultrasonic treatment, heating and 5% acetic acid extraction, antibacterial peptides was isolated from small intestines of pig by Sephadex G-100 , Sephadex G-25 gel filtration chromatography and ultrafiltration. The fractions obtained were tested against Escherichia coli O78. The fractions showing the antibacterial activity underwent further characterization. The purity and molecular weight of the antibacterial peptide were estimated using tricine polyacrylamide gel electrophoresis (Tricine SDS-PAGE). The results showed that an electrophoresis-grade antibacterial peptide was purified from small intestine of pig and its molecular weight was estimated to be about 5972 daltons.Using agarose diffusion assay and live bacteria counting assay, its antibacterial activity against 11 bacteria strains {Salmonella pullorum C79-13, Escherichia coli O1 C83845, Staphylococcus aureus ATCC25923, Escherichia coli O2, Escherichia coli O-j%, Pseudomonas aeruginosa ATCC27853, porcine pathogenic Escherichia coli isolate, chicken pathogenic Escherichia coli isolate, porcine pathogenic Salmonella typhisuis isolate, Staphylococcus albus and fish pathogenic Aeromonas hydrophila isolate ) was revealed. The results showed that the small intestine antibacterial peptide was strongly active against all the bacteria strains above except E.coli O2 and porcine pathogenic S.typhisuis isolate, and the bacteria-killing rates varied from 59.5% to 98.5%. The results preliminarily indicated that the small intestine antibacterial peptide was strongly active against bacteria and possessed broad antibacterial or bactericidal spectrum.Morphological observation of S.pullorum C79-13, E.coli Ox C83845 and S.aureus ATCC25923 treated with the small intestine antibacterial peptide, respectively, at 37.0C for 15 minutes under scanning electron microscopy (SEM) showed that the surfaces of the tested bacteria above became considerably rough, crimpled, shrivelled, distorted and blebbed. The results provided conclusive evidence that the small intestine antibacterial peptide was membrane active and how the antibacterial peptide killed bacteria.Infectious bronchitis virus (IBV) was treated with the small intestine antibacterial peptide at room temperature for 15 minutes and the small intestine ABP-treated IBV was inoculated into allantoic cavity of 9-day-old chicken embryos. The hemagglutination titer of IBV in chicken embryo allantoic fluids was determined and the pathological changes of chicken embryos were observed. The results showed that the hemagglutination titer of IBV was lower and the pathological changes were less serious than the chicken embryos inoculated with the ABP-untreated IBV. SPF chicken were challenged by Newcastle disease virus (NDV) and infectious bursal disease virus (IBDV) treated with the small intestine antibacterial peptide, respectively, and the results showed that the small intestine antibacterial peptidecould reduce the mortality and incidence of chickens caused by ND and IBD.The small intestine antibacterial peptide was administered intramuscularly into chicken at 10ug per chicken dose weekly since 7 days old. The immune organ index, the phagocytic percentage and phagocytic index of macrophage in the abdominal cavity of chicken, the serum antibody titers against NDV and IBDV, the growth index, the villus length, mucosa thickness and V/C value of duodenum, jejunum and ileum were determined at the age of 7, 21, 35, 49 days, respectively. The number of intraepithelial lymphocytes (IEL), mast cell and goblet cell of duodenum, jejunum, ileum and the IgA positive substances were also counted at above age, respectively. The results showed that the s... |
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