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A Recombinant Probiotic, Lactobacillus casei, Expressing the Clostridium perfringens α-toxoid, as an Orally Vaccine Candidate Against Gas Gangrene and Necrotic Enteritis

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Abstract

The alpha-toxin is one of the virulence factors of Clostridium perfringens for gas gangrene in humans and animals or necrotic enteritis in poultry. The C-terminal domain of this toxin ( cpa 247-370 ) was synthesized and cloned into pT1NX vector to construct the pT1NX-alpha plasmid. This surface-expressing plasmid was electroporated into Lactobacillus casei ATCC 393, generating the recombinant L. casei strain expressing alpha-toxoid (LC-α strain). Expression of this modified alpha-toxoid was confirmed by SDS-PAGE, immunoblotting, and direct immunofluorescence microscopy. BALB/c mice, immunized orally by the recombinant LC-α strain, elicited mucosal and significantly humoral immune responses (p < 0.05) and developed a protection against 900 MLD/mL of the standard alpha-toxin. This study showed that this recombinant LC-α strain could be a promising vaccine candidate against gas gangrene and necrotic enteritis.

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References

  1. Naylor CE, Jepson M, Crane DT, Titball RW, Miller J, Basak AK, Bolgiano B (1999) Characterisation of the calcium-binding C-terminal domain of Clostridium perfringens alpha-toxin. J Mol Biol 294(3):757–770

    Article  CAS  PubMed  Google Scholar 

  2. Titball RW, Naylor CE, Basak AK (1999) The Clostridium perfringens α-toxin. Anaerobe 5(2):51–64

    Article  CAS  PubMed  Google Scholar 

  3. Urbina P, Collado MI, Alonso A, Goñi FM, Flores-Díaz M, Alape-Girón A, Ruysschaert J-M, Lensink MF (2011) Unexpected wide substrate specificity of C. perfringens α-toxin phospholipase C. BBA-Biomembranes 1808(10):2618–2627

    Article  CAS  PubMed  Google Scholar 

  4. TITBALL R, BASAK A (1999) Structure function studies of Clostridium perfringens alpha-toxin; a gas gangrene causing protein. Jour Chem Soc Pak Vol 21(3):239

    Google Scholar 

  5. Titball RW (1993) Bacterial phospholipases C. Microbiol Rev 57(2):347–366

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Naylor CE, Eaton JT, Howells A, Justin N, Moss DS, Titball RW, Basak AK (1998) Structure of the key toxin in gas gangrene. Nat Struct Mol Biol 5(8):738–746

    Article  CAS  Google Scholar 

  7. Titball RW, Fearn AM, Williamson ED (1993) Biochemical and immunological properties of the C-terminal domain of the alpha-toxin of Clostridium perfringens. FEMS Microbiol Lett 110(1):45–50

    Article  CAS  PubMed  Google Scholar 

  8. Williamson E, Titball R (1993) A genetically engineered vaccine against the alpha-toxin of Clostridium perfringens protects mice against experimental gas gangrene. Vaccine 11(12):1253–1258

    Article  CAS  PubMed  Google Scholar 

  9. Qin J, Wang X, Kong J, Ma C, Xu P (2014) Construction of a food-grade cell surface display system for Lactobacillus casei. Microbiol Res 169(9):733–740

    Article  CAS  PubMed  Google Scholar 

  10. Salvarani FM, Conceicão FR, Cunha CE, Moreira GM, Pires PS, Silva RO, Alves GG, Lobato FC (2013) Vaccination with recombinant Clostridium perfringens toxoids α and β promotes elevated antepartum and passive humoral immunity in swine. Vaccine 31(38):4152–4155

    Article  CAS  PubMed  Google Scholar 

  11. Schotte L, Steidler L, Vandekerckhove J, Remaut E (2000) Secretion of biologically active murine interleukin-10 by Lactococcus lactis. Enzym Microb Technol 27(10):761–765

    Article  CAS  Google Scholar 

  12. Waterfield N, Le Page R, Wilson P, Wells J (1995) The isolation of lactococcal promoters and their use in investigating bacterial luciferase synthesis in Lactococcus lactis. Gene 165(1):9–15

    Article  CAS  PubMed  Google Scholar 

  13. Mohamadzadeh M, Duong T, Sandwick S, Hoover T, Klaenhammer T (2009) Dendritic cell targeting of Bacillus anthracis protective antigen expressed by Lactobacillus acidophilus protects mice from lethal challenge. Proc Natl Acad Sci 106(11):4331–4336

    Article  PubMed  Google Scholar 

  14. Alimolaei M, Golchin M, Daneshvar H (2016) Oral immunization of mice against Clostridium perfringens epsilon toxin with a Lactobacillus casei vector vaccine expressing epsilon toxoid. Infect Genet Evol 40:282–287

    Article  CAS  PubMed  Google Scholar 

  15. Klaenhammer TR (1984) A general method for plasmid isolation in lactobacilli. Curr Microbiol 10(1):23–28

    Article  CAS  Google Scholar 

  16. Alimolaei M, Ezatkhah M, Bafti MS (2014) Genetic and antigenic typing of Clostridium perfringens isolates from ostriches. Infect Genet Evol 28:210–213

    Article  PubMed  Google Scholar 

  17. Holmgren J, Adamsson J, Anjuère F, Clemens J, Czerkinsky C, Eriksson K, Flach C-F, George-Chandy A, Harandi AM, Lebens M (2005) Mucosal adjuvants and anti-infection and anti-immunopathology vaccines based on cholera toxin, cholera toxin B subunit and CpG DNA. Immunol Lett 97(2):181–188

    Article  CAS  PubMed  Google Scholar 

  18. Mohamadzadeh M, Olson S, Kalina WV, Ruthel G, Demmin GL, Warfield KL, Bavari S, Klaenhammer TR (2005) Lactobacilli activate human dendritic cells that skew T cells toward T helper 1 polarization. Proc Natl Acad Sci U S A 102(8):2880–2885

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Li X, Xia C, Li Y (2009) Induced expression of alpha-toxin gene of Clostridium perfringens in recombinant Lactobacillus casei and their immunoprotective in mice. Wei sheng wu xue bao = Acta microbiologica Sinica 49(8):1115–1120

    CAS  PubMed  Google Scholar 

  20. Wei M-Q, Rush CM, Norman JM, Hafner LM, Epping RJ, Timms P (1995) An improved method for the transformation of Lactobacillus strains using electroporation. J Microbiol Methods 21(1):97–109

    Article  Google Scholar 

  21. Hoang TH, Hong HA, Clark GC, Titball RW, Cutting SM (2008) Recombinant Bacillus subtilis expressing the Clostridium perfringens alpha toxoid is a candidate orally delivered vaccine against necrotic enteritis. Infect Immun 76(11):5257–5265

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Stevens DL, Titball RW, Jepson M, Bayer CR, Hayes-Schroer SM, Bryant AE (2004) Immunization with the C-domain of α-toxin prevents lethal infection, localizes tissue injury, and promotes host response to challenge with Clostridium perfringens. J Infect Dis 190(4):767–773

    Article  CAS  PubMed  Google Scholar 

  23. Lee J-S, Poo H, Han DP, Hong S-P, Kim K, Cho MW, Kim E, Sung M-H, Kim C-J (2006) Mucosal immunization with surface-displayed severe acute respiratory syndrome coronavirus spike protein on Lactobacillus casei induces neutralizing antibodies in mice. J Virol 80(8):4079–4087

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by a grant from Iran National Science Foundation (INSF) under grant number 920238680.

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Authors and Affiliations

  1. Department of Molecular Microbiology, Kerman Branch, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Kerman, Iran

    Mojtaba Alimolaei

  2. Department of Anaerobic Bacterial Vaccine Research and Production, Kerman Branch, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Kerman, Iran

    Mojtaba Alimolaei, Majid Ezatkhah & Mehrdad Shamsaddini Bafti

  3. Department of Pathobiology, Faculty of Veterinary Medicine, Shahid Bahonar University of Kerman, Post Box: 76169-133, Kerman, 7636161313, Iran

    Mojtaba Alimolaei & Mehdi Golchin

  4. Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahid Bahonar University of Kerman, Kerman, Iran

    Jalil Abshenas

Authors
  1. Mojtaba Alimolaei
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  2. Mehdi Golchin
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  3. Jalil Abshenas
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  4. Majid Ezatkhah
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  5. Mehrdad Shamsaddini Bafti
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Contributions

Mojtaba Alimolaei performed the entire experiments and data analysis and wrote the manuscript. Mehdi Golchin supervised experiments and manuscript writing. Jalil Abshenas collaborated in working with the laboratory animals. Mehrdad Shamsaddini-Bafti collaborated in the challenge test. Majid Ezatkhah contributed to the scientific discussion.

Corresponding author

Correspondence to Mehdi Golchin.

Ethics declarations

The study was approved by the Animal Experimentation Ethics Committee of Kerman University of Medical Sciences (Code: 92023868/92/11/07).

Conflict of Interest

The authors declare that they have no conflict of interest.

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Alimolaei, M., Golchin, M., Abshenas, J. et al. A Recombinant Probiotic, Lactobacillus casei, Expressing the Clostridium perfringens α-toxoid, as an Orally Vaccine Candidate Against Gas Gangrene and Necrotic Enteritis. Probiotics & Antimicro. Prot. 10, 251–257 (2018). https://doi.org/10.1007/s12602-017-9276-8

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  • DOI: https://doi.org/10.1007/s12602-017-9276-8

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