| Presently,the fish diseases account for a significant portion,they are the largest percentage in them.Most of fish diseases are bacterial hemorrhage diseases caused by Aeromonas hydrophila.Previously, antibiotics is used as feed additives frequently to achieve the purposes of prevention and treatment.In recently years,the world's aquaculture industry has been plagued by drug-resistant microorganisms and the residues of antibiotics also bring potential threats to human health.There is an urgent need to find a new product to replace antibiotics. The laboratory maintained an antibacterial substance produced by Bacillus subtilis fmbJ mainly cyclic lipopeptide, the lipopeptide has a good thermal stability and the stability of some enzymes, anti-microbial activity was higher, it may be a green feed additive.Purpose of this experiment is to explore the inhibitory mechanism though lipopeptide produced by bacillus subtilis act on Aeromonas hydrophila cells;the same time researching the condition of the product process of lipopeptide.Finally, the lipopeptide products as a feed additive for fish growth,as well as immunization and anti-oxidation were measured as a lipopeptide as feed additives to determine the feasibility study. The major findings are as follows:1. Sensibility and inhibition effect of lipopeptide of Bacillus subtilis fmbJ to Aeromonas hydrophila was studied.The results showed that,Aeromonas hydrophila has high sensitivity.minimal inhibitory concentration is 20μg/mL;minimum bactericidal concentration is 32μg/mL; Determination Aeromonas hydrophila growth curve,through cell growth changes of the lipopeptide effect on bacterial in different periods,got its bacteriostatic and bactericidal effect;By ion experiments, and propidium iodide staining, determined the role of lipopeptide cell membrane permeability of Aeromonas hydrophila.The test Electron microscope observed more accurately the role of ion experiments,and propidium iodide staining and other tests,determination of lipopeptide right Aeromonas hydrophila bacterial cell membrane permeability; Electron microscope to observe the test more accurately the role of lipopeptide on cells of Aeromonas hydrophila, resulted the lack of bacterial cell wall, flagella lost and so on. 2. Studied the role of the lipid-peptide on Genomic DNA and protein of the Aeromonas hydrophila,the results showed that,lipopeptide is able to combine with Aeromonas hydrophila chromosomal DNA which led to changes in the maximum absorption peak and produced hypochromic effect and hyperchromic effect.At the same time lipopeptide can has some impact on extracellular protease synthesized by Aeromonas hydrophila,the extracellular protease is the main virulence factors of Aeromonas hydrophila, so this more clearly demonstrated lipopeptide has high-performance inhibitory and antibacterial effect on Aeromonas hydrophila.3. Studied the lipopeptide product preparation process conditions.after added excipient, product recovery rate of lipopeptide was optimized by orthogonal with different inlet temperatures,different wind speed and different speed peristaltic pump.The results showed that,the highest recovery rate of lipopeptide products is highest.In the peristaltic pump speed 50 ml/h,inlet air temperature of 210℃,wind speed 70 M3/h,the highest recovery rate reached 75%.4. Studied in the basis of allogynogenetic crucian carp diets supplemented with different amounts of lipopeptide products will gibel carp growth, changes in intestinal flora, immune and antioxidant effects, etc. The results showed that,growth, the changes in intestinal flora,immune and antioxidant capacity of allogynogenetic crucian carp are better than the control group after added lipopeptide.Weight gain rate and liver body weight ratio of allogynogenetic crucian carp have certain degree of increased added 4‰lipopeptide in the basal diet;harmful bacteria reduced in the intestinal flora;immunity and antioxidant capacity were significantly increased (p<0.05). |
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