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Enterohemorrhagic Escherichia coli O157:H7 antigens produced in transgenic lettuce effective as an oral vaccine in mice

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Abstract

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Transgene with recombination sites to address biosafety concerns engineered into lettuce to produce EspB and γ-intimin C280 for oral vaccination against EHEC O157:H7.

Abstract

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a food-borne pathogen where ruminant farm animals, mainly bovine, serve as reservoirs. Bovine vaccination has been used to prevent disease outbreaks, and the current method relies on vaccines subcutaneously injected three times per year. Since EHEC O157:H7 colonizes mucosal surfaces, an oral vaccine that produces an IgA response could be more convenient. Here, we report on oral vaccination against EHEC O157:H7 in mice orally gavaged with transgenic lettuce that produces EHEC O157:H7 antigens EspB and γ-intimin C280. Younger leaves accumulated a higher concentration of antigens; and in unexpanded leaves of 30-day-old T2 plants, EspB and γ-intimin C280 were up to 32 and 51 μg/g fresh weight, respectively. Mice orally gavaged with lettuce powders containing < 3 µg antigens for 6 days showed a mucosal immune response with reduced colonization of EHEC O157:H7. This suggests that the transgenic lettuce has potential to be used for bovine vaccination. To promote the biosafety of crop plants producing medically relevant proteins, recombination sites were built into our transgenic lines that would permit optional marker removal by Cre-lox recombination, as well as transgene deletion in pollen by CinH-RS2 recombination. The ability to upgrade the transgenic lettuce by stacking additional antigen genes or replacing older genes with newer versions would also be possible through the combined use of Bxb-att and Cre-lox recombination systems.

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Data availability

The data supporting the findings of this study are available upon request from D.W. Ow.

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Acknowledgements

We thank the following individuals: Dr. Han Zhiguo, Dr. Li Ruyu, Dr. Li Yongqing of the South China National Botanical Garden for advice on vector construction, transformation or Southern blot analysis; Dr. Xu Zengfu in Xishuangbanna Tropical Botanical Garden, and Dr. Zhao Lingxia, Shanghai Jiao Tong University for suggestions on lettuce transformation and cultivation; Dr. Kyoko Hiwasa-Tanase, University of Tsukuba, and Dr. Kei-ichiro Mishiba, Osaka Prefecture University for advice on cloning ubiquitin promoter and terminator of lettuce and Southern blot analysis of transgenic lettuce, respectively: Dr. Fan Hongying and Dr. Kong Jinjin from Southern Medical University for advice on preliminary mice experiments. Dr. Marisa Lopez Bilbao and Dr. Laura Radonic from Instituto de Agrobiotecnología y Biología Molecular (IABIMO) INTA-CONICET for greenhouse space and assistance in growing transgenic lettuce in Argentina.

Funding

This work was supported by Guangdong Basic and Applied Basic Research Foundation (Grant No. 2020A1515110765), the National Key R&D Project (Grant No. 2016YFD0101904) from the Ministry of Science and Technology of China, and the Key Research Program of Frontier Sciences CAS, China (Grant No. QYZDY-SSW-SMC010) and Argentina grant from National Science and Technology Promotion Agency (Grant No. PICT-2018-02280).

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Authors and Affiliations

  1. Plant Gene Engineering Center; Key Laboratory of South China Agricultural Plant Molecular Analysis and Genetic Improvement; Guangdong Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, 510650, People’s Republic of China

    Yun-Lin Su, Huan Song, Tianfang Cheng, Yufang Wang & David W. Ow

  2. Instituto de Agrobiotecnología y Biología Molecular (IABIMO) INTA-CONICET, Hurlingham, Argentina

    Mariano Larzábal, Libia Yael Smith & Angel Adrian Cataldi

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  1. Yun-Lin Su
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  2. Mariano Larzábal
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  3. Huan Song
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Contributions

YS, DWO, ML and AAC designed the experiments; YS, ML, HS, TC, YW and LS conducted the experiments; YS, DWO, ML and AAC wrote the paper.

Corresponding author

Correspondence to David W. Ow.

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All authors declared no conflict of interest.

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The authors declare that the experiments comply with the current laws of the country in which they were performed. The Institutional Committee for the Care and Use of Experimental Animals CICUAE INTA CICVyA approved the conduct of the mice experiments of this study (number of reference 20/2022).

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Communicated by Ian D. Godwin.

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Su, YL., Larzábal, M., Song, H. et al. Enterohemorrhagic Escherichia coli O157:H7 antigens produced in transgenic lettuce effective as an oral vaccine in mice. Theor Appl Genet 136, 214 (2023). https://doi.org/10.1007/s00122-023-04460-5

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