| Plectasin is antimicrobial peptides isolated from P. nigrella and stongly inhibits the growth of gram-positive bacteria. To reduce the price for production, our lab has previously developed a recombinant yeast strain (PPle) expressing a multimer form of. In this study, optimization of the growth medium for the recombinant yeast strain was carried out by using a 30-L fermenter.. Moreover, the effects of antibacterial peptides on growth performance, gut health in weaned pigs were investigated. The mainly results were below:Experiment 1:optimized production of Recombinant Plectasin and research on antibacterial effectsThe study conducted liquid fermentation for the engineering bacteria in the 30-L fermentation tank using batch-fermentation process and comparing the expression of plectasin level with low salt BSM group, glycerol BSM group and soluble starch BSM group in three different medium. Taking the method of coomassie brilliant blue to detect the total protein in fermentation. To be specific, detected the way of expression of recombinant antimicrobial peptides and combined with flat bacte riostatic antimicrobial peptides for studying its stability and antiacterial effects. The results showed that:the wet weight of low salt group, glycerol basis group and soluble starch group reached the highest value after 114 hours fermentation, they are 450g/L,402g/L,277g/Lrespectively. The total protein concentration of the supernatant of the three groups are 0.375g/L,3.941g/L,5.63g/L respectively. Antibacterial results indicted that recombinant plectasin antimicrobial peptides still had bacteriostatc ring onphylococcus aureus after acid,pepsin and trypsin treatment. Summary:the expression level of recombinant antimicrobial peptides had been improved though the optimized medium. Also, the property of stability is good for the staphylococcus aureus.The best medium is glycerol BSM.Experiment 2:effects of recombinant plectasin on growth performance and intestinal health in weaned pigsThis test examined the recombinant Plectasin antimicrobial peptides on growth performance, immune function and intestinal health of biological efficacy of weaning pigs to understand the mechanism of antimicrobial peptides on weaning pigs.Choosed 30 healthy DLY piglets weaning at 24d of age, and randomly assigned to 5 dietary treatments. The treatments were basal diet (CON:without antibiotic feed), Sulfuric acid colistin (CS:0.3% sulfuric acid colistin), Antimicrobial peptides (AP:0.2% Recombinant Plectasin antimicrobial peptides), Probiotics (PB:0.1% probiotics), Joint Application (PPB:0.2% of recombinant plectasin antimicrobial peptides+0.1% probiotics). The experiment lasted for 21 days. The results were shown below:Compared with CON group, ADFI, ADG and F/G significantly increased in groups of CS and AP(P< 0.05). The rate of diarrhea had trend to be decreased in groups of CS and AP compare to the CON group (P=0.065). Moreover, the diarrhea index of CS (-91%, P< 0.01) and AP (-88%, P< 0.01) groups significantly decreased, respectively. Compared with the CON group, the other groups had tendency to increase the digestibilities of energy and dry matter (P<0.1). The digestibility of phosphorus of the PB group was higher than the rest four groups (P< 0.01). The duodenal enzymes of maltase and sucrose, the jejunal enzymes of maltase, sucrase and lactase have received a significant increase (P<0.01) in CS, AP and PPB groups than the CON group. The absorption of serum D-xylose was higher in AP and PPB than the other three groups (P< 0.01). Compared with other groups, the AP group significantly increased the content of Bifidobacteria (P< 0.05) in ileum. The content of lactobacilli of PB group was higher than the CON group (P< 0.05) while was similar with the other groups in ileum. The content of E. coli of PB group was higher than that in CS and AP group (P< 0.05), moreover, the content of E. coli of AP group was the lowest in caecum. Compared with other groups, the content of E. coli of the CS group had a trend to decrease in colon (P <0.1). Compared with CON group, the pH of AP. PB and PPB groups significantly decreased in the chyme of colon (P< 0.01). The other four groups significantly increased the duodenal villus height and VCR, the jejunal villus height and the ileal villus height and VCR while significantly decreased the ileal crypt depth (P< 0.01). Compared with the CON group, the levels of jejunal mucosa sIgA in CS, AP and PPB groups were significantly increased (P< 0.01). Compared with the rest four groups, the concentration of IgG was significant increased in the CON group (P< 0.01).In summary, the optimal fermentation medium for restructuring hyphae antibacterial peptide toxin glycerin fermentation basal salts medium, antibacterial peptides quantity reached 3.941 g/L; Basis diet with antibacterial peptide can significantly improve the production performance, improve nutrient digestibility, maintain the intestinal tissue morphological structure, strengthen the intestinal barrier function, maintain the intestinal microflora, improve intestinal health of the weaned pig. The results indicated that recombinant antimicrobial peptides can be used as a potential alternative for antibiotics in the feeding industry. |
