Screening And Analysis Mti-Pseudomonas Aeruginosa Active Substance

The rates of infection from the Pseudomonas aeruginosa,one of the three primaryopportunistic pathogens and stronger pathogenicity to immunodeficiency people,graduallyrose along with the overprescription of immunosuppressant and broad-spectrumantibiotic,chemotherapy and various invasive therapies.Currently,treatment ofPseudomonas aeruginosa infection mainly relies on multiple antibiotics, however, a largenumber of Pseudomonas aeruginosa multidrug-resistant strains continue to emerge inrecent years,making treatment increasingly difficult after infection.Antibacterial peptides(or antibacterial protein),a widespread cationic amphiphilic peptides, founded in all livingthings, with a broad antibacterial spectrum, faster and stronger onset of action, difficult toproduce resistance, has drawn extensive attention and has great development potential tobe new anti-infective agents.However, there are few studies of antibacterial peptides(orantibacterial protein) for Pseudomonas aeruginosa have been reported.The main purposeof this project was to screen strains, have antagonism effect on Pseudomonas aeruginosa,and isolated and identified the active ingredient to lay a foundation for the study of a newanti-Pseudomonas aeruginosa drug.Research contents:1.Screen strains, have antagonism effect to Pseudomonas aeruginosa, from differentsoil samples and then identified according to themorphological features and16SrDNAsequence analysis.2.Different factors were investigated such as the ingredients of medium, pH,temperature, fermentation time to optimize expression conditions for the active ingredientof the antagonism of Pseudomonas aeruginosa.3.Using several methods, including ammonium sulfate precipitation, ion exchangeand molecular sieve, of isolation and purification, we purified antibacterial peptide fromfermentation liquor.4.To analyze the characteristic of anti-Pseudomonas areruginosa active ingredientincluding pH stability, proteinase K sensitivity, high temperature tolerance, molecularweight, isoelectric point etc.Result:1.5strains，antagonism Pseudomonas aeruginosa,from soil samples were screenedout.Colony morphology, Gram staining and morphologica observation,16S rDNA amplification,sequencing and RDP sequence analysis,and clear the screening of strainsbelonging to Pseudomonad aceae and Acinetobacter.2.In this study, eleven different medium and the other (initial pH value, fermentationtemperature and fermentation time) the impact of antimicrobial substances, to obtain theoptimum fermentation conditions.Optimal results were yeast extract3%, tryptone5%,glucose3%, NaCl5%.The best culture conditions were pH7.0,37℃,250rpm, shakingculture for48h.3.The antibacterial substance,the purity is80%, can be isolated from other groups bysalting out with50%(NH4)2SO4or isoelectric precipitation.4.The biological features of antibacterial substance showed that the bacteriocin isnot resistant to protease K and high temperature.The molecular weight is35kD and pI is6.1.