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Evaluation of a toxoid fusion protein vaccine produced in plants to protect poultry against necrotic enteritis

, , , , , , ,
1 Center for Immunotherapy, Vaccines and Virotherapy, The Biodesign Institute, Arizona State University, Tempe, AZ, United States
2 School of Life Sciences, Arizona State University, Tempe, AZ, United States
DOI
10.7287/peerj.preprints.27390v1
Published
Accepted
Subject Areas
Agricultural Science, Biotechnology, Microbiology, Veterinary Medicine
Keywords
Clostridium perfringens, plant-based antigen production, Necrotic enteritis vaccine, poultry vaccine
Copyright
© 2018 Hunter et al.
Licence
This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ Preprints) and either DOI or URL of the article must be cited.
Cite this article
Hunter JG, Wilde S, Tafoya AM, Horsman J, Yousif M, Diamos AG, Roland KL, Mason HS. 2018. Evaluation of a toxoid fusion protein vaccine produced in plants to protect poultry against necrotic enteritis. PeerJ Preprints 6:e27390v1

Abstract

Background. Necrotic enteritis (NE) is caused by type A strains of the bacterium Clostridium perfringens. Total global economic losses to the poultry industry due to NE is estimated to be over 2 billion dollars annually. Traditionally, NE has been effectively controlled by inclusion of antibiotics in the diet of poultry. However, recent concerns regarding the impact of this practice on increasing antibiotic resistance in human pathogens have led us to consider alternative approaches, such as vaccination, for controlling this disease. NE strains of C. perfringensproduce two major toxins, a-toxin and NetB. Immune responses against either toxin can provide partial protection against NE.

Methods. We have developed a fusion protein combining a non-toxic carboxyl-terminal domain of a-toxin (PlcC) and an attenuated, mutant form of NetB (NetB-W262A) for use as a vaccine antigen to immunize poultry against NE. We utilized a DNA sequence that was codon-optimized for Nicotiana benthamianato enable high levels of expression. The 6-His tagged PlcC-NetB fusion protein was synthesized in N. benthamianausing a geminiviral replicon transient expression system, purified by metal affinity chromatography, and used to immunize broiler birds.

Results. Immunized birds produced a strong serum IgY response against both the plant produced PlcC-NetB protein and against bacterially produced His-PlcC and His-NetB. Immunized birds were significantly protected against a subsequent in-feed challenge with virulent C. perfringenswhen treated with the fusion protein. These results indicate that a plant-produced PlcC-NetB toxoid is a promising vaccine candidate for controlling NE in poultry.

Author Comment

This is a submission to PeerJ for review.

Supplemental Information

DNA nucleotides sequences for PlcC-NetB fusion protein genes

DNA sequences and encoded amino acid sequences for the 6His-PlcC-NetB fusion protein either with or without ER-targeting signal peptide.

DOI: 10.7287/peerj.preprints.27390v1/supp-3

Additional Information

Competing Interests

The authors declare that they have no competing interests.

Author Contributions

Joseph G Hunter conceived and designed the experiments, performed the experiments, analyzed the data, prepared figures and/or tables, authored or reviewed drafts of the paper.

Shyra Wilde performed the experiments, analyzed the data, prepared figures and/or tables.

Amanda M Tafoya performed the experiments.

Jamie Horsman performed the experiments.

Miranda Yousif performed the experiments.

Andrew G Diamos conceived and designed the experiments, performed the experiments, analyzed the data, contributed reagents/materials/analysis tools, authored or reviewed drafts of the paper.

Kenneth L Roland conceived and designed the experiments, analyzed the data, contributed reagents/materials/analysis tools, prepared figures and/or tables, authored or reviewed drafts of the paper, approved the final draft.

Hugh S Mason conceived and designed the experiments, analyzed the data, contributed reagents/materials/analysis tools, prepared figures and/or tables, authored or reviewed drafts of the paper, approved the final draft.

Animal Ethics

The following information was supplied relating to ethical approvals (i.e., approving body and any reference numbers):

Arizona State University Institutional Animal Care and Use Committee approved this research.

Patent Disclosures

The following patent dependencies were disclosed by the authors:

PCT/US18/40632, VACCINE FOR PREVENTION OF NECROTIC ENTERITIS IN POULTRY

US 13/060,414, A DNA replicon system for high-level rapid production of vaccines and monoclonal antibody therapeutics in plants

DNA Deposition

The following information was supplied regarding the deposition of DNA sequences:

The fusion protein nucleotide sequences are in GenBank accession numbers MH883808 and MH883809.

Data Deposition

The following information was supplied regarding data availability:

Original Western blot images for Fig. 2 and Fig. 3 are provided.

Funding

This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2016-67016-24947 from the USDA National Institute of Food and Agriculture and startup funds from ASU to Ken Roland, and by support from the Biodesign Institute at ASU for Hugh Mason. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Evaluation of a toxoid fusion protein vaccine produced in plants to protect poultry against necrotic enteritis
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