title: Construction ofBacillus subtilisstrain engineered for expression of porcine β-defensin-2/cecropin P1 fusion antimicrobial peptides and its growth-promoting effect and antimicrobial activity.
abstract: OBJECTIVE
  To generate recombinantBacillus subtilis(B. subtilis) engineered for expression of porcine β-defensin-2 (pBD-2) and cecropin P1 (CP1) fusion antimicrobial peptide and investigate their anti-bacterial activityin vitroand their growth-promoting and disease resisting activityin vivo.
  
  
  METHODS
  The pBD-2 and CP1 fused gene was synthesized using the main codons ofB. subtilisand inserted into plasmid pMK4 vector to construct their expression vector. The fusion peptide-expressingB. subtiliswas constructed by transformation with the vector. The expressed fusion peptide was detected with Western blot. The antimicrobial activity of the expressed fusion peptide and the recovered pBD-2 and CP1 by enterokinase digestionin vitrowas analyzed by the bacterial growth-inhibitory activity assay. To analyze the engineeredB. subtilison growth promotion and disease resistance, the weaned piglets were fed with basic diet supplemented with the recombinant B. subtilis. Then the piglets were challenged by enteropathogenicEscherichia coli(E. coli). The weight gain and diarrhea incidence of piglets were measured after challenge.
  
  
  RESULTS
  The recombinantB. subtilisengineered for expression of pBD-2/CP1 fusion peptide was successfully constructed using the main codons of theB. subtilis. Both expressed pBD-2/CP1 fusion peptide and their individual peptides recovered from parental fusion peptide by enterokinase digestion possessed the antimicrobial activities to a variety of the bacteria, including gram-negative bacteria (E. coli,Salmonella typhimurium, andHaemophilus parasuis) and gram-positive bacteria (Staphylococcus aureus). Supplementing the engineeredB. subtilisto the pig feed could significantly promote the piglet growth and reduced diarrhea incidence of the piglets.
  
  
  CONCLUSIONS
  The generatedB. subtilisstrain can efficiently express pBD-2/CP1 fusion antimicrobial peptide, the recovered pBD-2 and CP1 peptides possess potent antimicrobial activities to a variety of bacterial speciesin vitro. Supplementation of the engineeredB. subtilisin pig feed obviously promote piglet growth and resistance to the colibacillosis.
authors.affiliation: Department of Biotechnology, College of Environmental and Chemical Engineering, Yanshan University, Qinhuangdao 066004, China.|Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Jian
authors.initials: J
authors.lastname: Xu
authors.affiliation: Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Fei
authors.initials: F
authors.lastname: Zhong
authors.affiliation: Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Yonghong
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Jianlou
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Shanshan
authors.initials: S
authors.lastname: Huo
authors.affiliation: Department of Biotechnology, College of Environmental and Chemical Engineering, Yanshan University, Qinhuangdao 066004, China.
authors.forename: Hongyu
authors.initials: H
authors.lastname: Lin
authors.affiliation: Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Liyue
authors.initials: L
authors.lastname: Wang
authors.affiliation: Laboratory of Molecular Virology and Immunology, College of Veterinary Medicine, Agricultural University of Hebei, Hebei Engineering and Technology Research Center of Veterinary Biotechnology, Baoding 071000, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Cui
authors.affiliation: Department of Biotechnology, College of Environmental and Chemical Engineering, Yanshan University, Qinhuangdao 066004, China.
authors.forename: Xiujin
authors.initials: X
authors.lastname: Li
keywords: Antimicrobial Activity; Cecropin P1; Engineered Bacillus subtilis; Piglets; Porcine β-defensin-2