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Immunogenicity of EIT chimeric protein expressed in transplastomic tobacco plants towards development of an oral vaccine against Escherichia coli O157:H7

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Abstract

Chloroplast genetic engineering offers an opportunity for high level expression and cost-effective recombinant protein production. Escherichia coli O157:H7 is one of the most important zoonotic pathogens causing hemorrhagic colitis (HC) and the life-threatening hemolytic-uremic syndrome in humans worldwide. The occurrence of zoonotic E. coli O157:H7 outbreaks in recent years has led to increased efforts in the development of safe and cost-effective immunogenic antigens against E. coli O157:H7. EspA and Tir/Intimin proteins are the important virulence factors which are encoded by the LEE locus of enterohemorrhagic E. coli. In this study, we hypothesized that the high level expression of the chimeric form of these effectors in chloroplasts and using tobacco transplastomic plants as an oral delivery system for the development of an edible-base vaccine would induce an immune response for the prevention of E. coli 0157:H7 attachment and colonization in animal model mice. The prokaryotic codon-optimized EIT protein was expressed in plastid genome via chloroplast transformation. Putative transplastomic plants were analyzed by PCR, and Southern blot analysis confirming chloroplast integration and homoplasmy in the T1 progeny. Immunoblotting and ELISA assays demonstrated that the EIT protein was expressed in chloroplasts and accumulated up to 1.4 % of total soluble protein in leaf tissue. In mice orally immunized with transplastomic tobacco plant leaves, high immunological responses (IgG and IgA specific antibodies) were detected in serum and feces. Finally, the challenging assay with E. coli O157:H7 in immunized mice showed reduced bacterial shedding.

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Acknowledgments

The authors thank Prof.V.S. Reddy (ICGEB, New Delhi, India) for generously providing the plasmid transformation vector pVSR326. We also thank Dr J. Amani for critical reading of this manuscript. This work was supported by funds from the National Institute of Genetic Engineering and Biotechnology (NIGEB).

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Authors and Affiliations

  1. National Institute of Genetic Engineering and Biotechnology, P.O. Box 14155-6343, Tehran, Iran

    Farrokh Karimi, Amir Mousavi & Ali Hatef Salmanian

  2. Department of Plant Breeding, Faculty of Agriculture, University of Tehran, P.O. Box 2824809, Karaj, Iran

    Houshang Alizadeh

  3. Molecular Immunology and Vaccine Research Lab, Pasteur Institute of Iran, Tehran, Iran

    Sima Rafati

  4. Department of Biotechnology, Faculty of Science, University of Maragheh, Maragheh, Iran

    Farrokh Karimi

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  1. Farrokh Karimi
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  2. Amir Mousavi
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  4. Houshang Alizadeh
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Correspondence to Amir Mousavi or Ali Hatef Salmanian.

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Karimi, F., Mousavi, A., Salmanian, A.H. et al. Immunogenicity of EIT chimeric protein expressed in transplastomic tobacco plants towards development of an oral vaccine against Escherichia coli O157:H7. Plant Biotechnol Rep 7, 535–546 (2013). https://doi.org/10.1007/s11816-013-0296-x

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