Construction of Strain Engineered for Expression of Porcine β-defensin-2/cecropin P1 Fusion Antimicrobial Peptides and Its Growth-promoting Effect and Antimicrobial Activity

Overview

Journal Asian-Australas J Anim Sci

Specialty Veterinary Medicine

Date 2016 Jul 8

PMID 27383796

Citations 15

Authors

Jian Xu

Fei Zhong

Yonghong Zhang

Jianlou Zhang

Shanshan Huo

Hongyu Lin

Liyue Wang

Dan Cui

Xiujin Li

Affiliations

Soon will be listed here.

Abstract

Objective: To generate recombinant () engineered for expression of porcine β-defensin-2 (pBD-2) and cecropin P1 (CP1) fusion antimicrobial peptide and investigate their anti-bacterial activity and their growth-promoting and disease resisting activity .

Methods: The pBD-2 and CP1 fused gene was synthesized using the main codons of and inserted into plasmid pMK4 vector to construct their expression vector. The fusion peptide-expressing was 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 digestion was analyzed by the bacterial growth-inhibitory activity assay. To analyze the engineered on 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 enteropathogenic (). The weight gain and diarrhea incidence of piglets were measured after challenge.

Results: The recombinant engineered for expression of pBD-2/CP1 fusion peptide was successfully constructed using the main codons of the . 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 (, , and ) and gram-positive bacteria (). Supplementing the engineered to the pig feed could significantly promote the piglet growth and reduced diarrhea incidence of the piglets.

Conclusion: The generated strain 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 species . Supplementation of the engineered in pig feed obviously promote piglet growth and resistance to the colibacillosis.

Citing Articles

Expression of recombination antimicrobial protein PIL22-PBD-2 in Pichia pastoris and verification of its biological function in vitro.

Li X, Qiu P, Yue M, Zhang Y, Lei C, Wang J Vet Res. 2025; 56(1):52.

PMID: 40055823 PMC: 11889930. DOI: 10.1186/s13567-024-01428-1.

Engineered Bacillus subtilis WB600/ZD prevents Salmonella Infantis-induced intestinal inflammation and alters the colon microbiota in a mouse model.

Li W, Wang X, Chen K, Zhu Y, Yang G, Jin Y Vet Res. 2025; 56(1):35.

PMID: 39920770 PMC: 11806837. DOI: 10.1186/s13567-024-01438-z.

Microbial production systems and optimization strategies of antimicrobial peptides: a review.

Lou M, Ji S, Wu R, Zhu Y, Wu J, Zhang J World J Microbiol Biotechnol. 2025; 41(2):66.

PMID: 39920500 DOI: 10.1007/s11274-025-04278-x.

spp. as potential probiotics: promoting piglet growth by improving intestinal health.

Tang X, Zeng Y, Xiong K, Zhong J Front Vet Sci. 2024; 11:1429233.

PMID: 39132437 PMC: 11310147. DOI: 10.3389/fvets.2024.1429233.

Protective role of Cecropin AD against LPS-induced intestinal mucosal injury in chickens.

Zhi Y, Li T, Li Y, Zhang T, Du M, Zhang Q Front Immunol. 2024; 14:1290182.

PMID: 38162646 PMC: 10757607. DOI: 10.3389/fimmu.2023.1290182.

References

Baker A, Davis E, Spencer J, Moser R, Rehberger T . The effect of a Bacillus-based direct-fed microbial supplemented to sows on the gastrointestinal microbiota of their neonatal piglets. J Anim Sci. 2013; 91(7):3390-9. DOI: 10.2527/jas.2012-5821. View

Wang X, Li X, Zhang Z, Shen X, Zhong F . Codon optimization enhances secretory expression of Pseudomonas aeruginosa exotoxin A in E. coli. Protein Expr Purif. 2010; 72(1):101-6. DOI: 10.1016/j.pep.2010.02.011. View

Arcidiacono S, Soares J, Meehan A, Marek P, Kirby R . Membrane permeability and antimicrobial kinetics of cecropin P1 against Escherichia coli. J Pept Sci. 2009; 15(6):398-403. DOI: 10.1002/psc.1125. View

Tatsuda D, Arimura H, Tokunaga H, Ishibashi M, Arakawa T, Tokunaga M . Expression and purification of cytokine receptor homology domain of human granulocyte-colony-stimulating factor receptor fusion protein in Escherichia coli. Protein Expr Purif. 2001; 21(1):87-91. DOI: 10.1006/prep.2000.1343. View

Sorokulova I, Pinchuk I, Denayrolles M, Osipova I, Huang J, Cutting S . The safety of two Bacillus probiotic strains for human use. Dig Dis Sci. 2007; 53(4):954-63. DOI: 10.1007/s10620-007-9959-1. View

Jukes T . Antibiotics in meat production. JAMA. 1975; 232(3):292-3. View

Fellermann K, Stange E . Defensins -- innate immunity at the epithelial frontier. Eur J Gastroenterol Hepatol. 2001; 13(7):771-6. DOI: 10.1097/00042737-200107000-00003. View

Marshall B, Levy S . Food animals and antimicrobials: impacts on human health. Clin Microbiol Rev. 2011; 24(4):718-33. PMC: 3194830. DOI: 10.1128/CMR.00002-11. View

Yang W, Feng J, Xiang F, Xie Z, Zhang G, Sabatier J . Endogenous animal toxin-like human β-defensin 2 inhibits own K(+) channels through interaction with channel extracellular pore region. Cell Mol Life Sci. 2014; 72(4):845-53. PMC: 11113244. DOI: 10.1007/s00018-014-1715-z. View

10.

Wu S, Zhang F, Huang Z, Liu H, Xie C, Zhang J . Effects of the antimicrobial peptide cecropin AD on performance and intestinal health in weaned piglets challenged with Escherichia coli. Peptides. 2012; 35(2):225-30. DOI: 10.1016/j.peptides.2012.03.030. View

11.

Stanton T . A call for antibiotic alternatives research. Trends Microbiol. 2013; 21(3):111-3. DOI: 10.1016/j.tim.2012.11.002. View

12.

Mukherjee S, Hooper L . Antimicrobial defense of the intestine. Immunity. 2015; 42(1):28-39. DOI: 10.1016/j.immuni.2014.12.028. View

13.

Gregory K, Mello C . Immobilization of Escherichia coli cells by use of the antimicrobial peptide cecropin P1. Appl Environ Microbiol. 2005; 71(3):1130-4. PMC: 1065138. DOI: 10.1128/AEM.71.3.1130-1134.2005. View

14.

Sang Y, Blecha F . Antimicrobial peptides and bacteriocins: alternatives to traditional antibiotics. Anim Health Res Rev. 2008; 9(2):227-35. DOI: 10.1017/S1466252308001497. View

15.

Castanon J . History of the use of antibiotic as growth promoters in European poultry feeds. Poult Sci. 2007; 86(11):2466-71. DOI: 10.3382/ps.2007-00249. View

16.

Wang N, Guo X, Xu J, Kong X, Gao S, Shan Z . Pollution characteristics and environmental risk assessment of typical veterinary antibiotics in livestock farms in Southeastern China. J Environ Sci Health B. 2014; 49(7):468-79. DOI: 10.1080/03601234.2014.896660. View

17.

Han F, Zhang H, Xia X, Xiong H, Song D, Zong X . Porcine β-defensin 2 attenuates inflammation and mucosal lesions in dextran sodium sulfate-induced colitis. J Immunol. 2015; 194(4):1882-93. DOI: 10.4049/jimmunol.1402300. View

18.

Li C, Zhao Y, Song X, Huang X, Zhao W . Molecular cloning, expression and characterization of the porcine β defensin 2 in E. coli. Protein Pept Lett. 2012; 20(6):715-23. DOI: 10.2174/0929866511320060010. View

19.

Bonavita R, Isticato R, Maurano F, Ricca E, Rossi M . Mucosal immunity induced by gliadin-presenting spores of Bacillus subtilis in HLA-DQ8-transgenic mice. Immunol Lett. 2015; 165(2):84-9. DOI: 10.1016/j.imlet.2015.04.005. View

20.

Ostaff M, Stange E, Wehkamp J . Antimicrobial peptides and gut microbiota in homeostasis and pathology. EMBO Mol Med. 2013; 5(10):1465-83. PMC: 3799574. DOI: 10.1002/emmm.201201773. View