Crude extracts of recombinant baculovirus expressing rabbit hemorrhagic disease virus 2 VLPs from both insect and rabbit cells protect rabbits from rabbit hemorrhagic disease caused by RHDV2
Vaccines against viral pathogens are often composed of recombinant proteins expressed in different systems. Such proteins expressed by recombinant baculoviruses have been proven to be effective for vaccination. Especially, after codon usage optimization high amounts of recombinant viral proteins can be obtained which can assemble to virus like particles (VLPs) spontaneously. In this study we compared two different codon usages of RHDV2-VP1 to improve the expression of recombinant VP1 of RHDV2 by recombinant baculoviruses after infection of insect SF9 cells or transduction of mammalian RK13 cells in order to gain high protein yields. Also the influence on the auto-assembly of RHDV2-VP1 to VLPs was investigated. Finally, the immunogenic potential of such recombinant vaccines against RHDV2 to induce a protective immune response in rabbits against RHDV2 should be characterized. There was no influence of different codon usages on RHDV2-VP1 gene expression in the respective cell lines detected. However, in insect cell line SF9 higher rates of recombinant VP1 were measured in comparison to the transduction of mammalian cells RK13. Auto-assembly of RHDV2-VP1 to VLPs was observed in both cell systems by electron microscopy. Finally, both RHDV-VP1 VLPs derived from mammalian and insect cells were able to induce a protective humoral immune response in rabbits against RHDV2.
The author’s thank Dr. Günther Keil and Dr. Horst Schirrmeier for their methodological help. The excellent technical assistance of Katrin Giesow, Bianka Hillmann, Petra Meyer, Mandy Jörn and Sabine Weber is gratefully acknowledged.
This study was funded by IDT Biologika (Riems), Greifswald-Insel Riems, Germany for C.M.
Compliance with ethical standards
Conflict of interest
The authors Claudia Müller, Reiner Ulrich, Kati Franzke, Marcus Müller, and Bernd Köllner declare that they have no conflict of interest.
All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. Prior to the start of all trials the approval from the Federal state Ethical Committee for Animal Experimentation (LALLF-7221.3-1-025/15) was given and was performed following the acquirements of the EU directive 2010/63 and the EG recommendation 2007/526/.
Human and animal rights
This article does not contain any studies with human participants or animals performed by any of the authors.
- 4.Bárcena J, Morales M, Vázquez B, Boga JA, Parra F, Lucientes J, Pagès-Manté A, Sánchez-Vizcaíno JM, Blasco R, Torres JM (2000) Horizontal transmissible protection against myxomatosis and rabbit hemorrhagic disease by using a recombinant myxoma virus. J Virol 74(3):1114–1123CrossRefPubMedPubMedCentralGoogle Scholar
- 6.Bertagnoli S, Gelfi J, Le Gall G, Boilletot E, Vautherot JF, Rasschaert D, Laurent S, Petit F, Boucraut-Baralon C, Milon A (1996) Protection against myxomatosis and rabbit viral hemorrhagic disease with recombinant myxoma viruses expressing rabbit hemorrhagic disease virus capsid protein. J Virol 70(8):5061–5066PubMedPubMedCentralGoogle Scholar
- 9.Camarda A, Pugliese N, Cavadini P, Circella E, Capucci L, Caroli A, Legretto M, Mallia E, Lavazza A (2014) Detection of the new emerging rabbit haemorrhagic disease type 2 virus (RHDV2) in Sicily from rabbit (Oryctolagus cuniculus) and Italian hare (Lepus corsicanus). Res Vet Sci 97(3):642–645CrossRefPubMedGoogle Scholar
- 12.Da Silva Junior HC (2012) da Silva e Mouta Junior S, de Mendonça MCL, de Souza Pereira MC, da Rocha Nogueira A, de Azevedo MLB, Leite JPG, de Moraes MTB: Comparison of two eukaryotic systems for the expression of VP6 protein of rotavirus specie A: transient gene expression in HEK293-T cells and insect cell-baculovirus system. Biotechnol Lett 34(9):1623–1627CrossRefPubMedGoogle Scholar
- 14.Duarte M, Carvalho C, Bernardo S, Barros SV, Benevides S, Flor L, Monteiro M, Marques I, Henriques M, Barros SC, Fagulha T, Ramos F, Luís T, Fevereiro M (2015) Rabbit haemorrhagic disease virus 2 (RHDV2) outbreak in Azores: Disclosure of common genetic markers and phylogenetic segregation within the European strains. Infect Genet Evol 35:163–171CrossRefPubMedGoogle Scholar
- 16.Farnós O, Boué O, Parra F, Martín-Alonso JM, Valdés O, Joglar M, Navea L, Naranjo P, Lleonart R (2005) High-level expression and immunogenic properties of the recombinant rabbit hemorrhagic disease virus VP60 capsid protein obtained in Pichia pastoris. J Biotechnol 117(3):215–224CrossRefPubMedGoogle Scholar
- 17.Fernández E, Toledo JR, Chiong M, Parra F, Rodríguez E, Montero C, Méndez L, Capucci L, Farnós O (2011) Single dose adenovirus vectored vaccine induces a potent and long-lasting immune response against rabbit hemorrhagic disease virus after parenteral or mucosal administration. Vet Immunol Immunopathol 142(3–4):179–188CrossRefPubMedGoogle Scholar
- 22.Granzow H, Schirrmeier H, Tews H (1989) Hämorrhagische Septikämie der Kaninchen—Erregernachweis und erste elektronenmikroskopische Charakterisierung. Mh Vet Med 4:379–380Google Scholar
- 23.Green KY, Lew JF, Jiang X, Kapikian AZ, Estes MK (1993) Comparison of the reactivities of baculovirus-expressed recombinant Norwalk virus capsid antigen with those of the native Norwalk virus antigen in serologic assays and some epidemiologic observations. J Clin Microbiol 31(8):2185–2191PubMedPubMedCentralGoogle Scholar
- 33.Keil GM, Pollin R, Müller C, Giesow K, Schirrmeier H (2016) BacMam Platform for vaccine antigen delivery. In: Brun A (eds) Methods in molecular biology vaccine technologies for veterinary viral diseases methods and protocols. Humana Press, New York, vol 1349, pp 105–19.Google Scholar
- 37.Le Gall-Reculé G, Lavazza A, Marchandeau S, Bertagnoli S, Zwingelstein F, Cavadini P, Martinelli N, Lombardi G, Guérin JL, Lemaitre E, Decors A, Boucher S, Le Normand B, Capucci L (2013) Emergence of a new lagovirus related to Rabbit Haemorrhagic Disease Virus. Vet Res 44:81CrossRefPubMedPubMedCentralGoogle Scholar
- 38.Liu SJ, Xue HP, Pu BQ, Qian NH (1984) A new viral disease in rabbits. Anim Husbandy Vet Med 16:253–255Google Scholar
- 40.Marin MS, Martín Alonso JM, Pérez Ordoyo García LI, Boga JA, Argüello-Villares JL, Casais R, Venugopal K, Jiang W, Gould EA, Parra F (1995) Immunogenic properties of rabbit haemorrhagic disease virus structural protein VP60 expressed by a recombinant baculovirus: an efficient vaccine. Virus Res 39(2–3):119–128CrossRefPubMedGoogle Scholar
- 41.Martin-Alonso A1, Martin-Carrillo N2, Garcia-Livia K3, Valladares B4, Foronda P5. Infect Genet Evol. 2016 44:46-50. https://doi.org/10.1016/j.meegid.2016.06.034. (Epub 2016 Jun 16. Emerging rabbit haemorrhagic disease virus 2 (RHDV2) at the gates of the African continent).
- 46.Ohlinger VF, Haas B, Ahl R, Weiland F (1989) Rabbit haemorrhagic disease—a contagious disease caused by a calicivirus. Tierärztliche Umschau (in german) 44:284–294Google Scholar
- 49.Puggioni G, Cavadini P, Maestrale C, Scivoli R, Botti G, Ligios C, Le Gall-Reculé G, Lavazza A, Capucci L (2013) The new French 2010 rabbit hemorrhagic disease virus causes an RHD-like disease in the Sardinian Cape hare (Lepus capensis mediterraneus). Vet Res 44:96CrossRefPubMedPubMedCentralGoogle Scholar
- 51.Sambrook J, Fritsch EF, Maniatis T (2000) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, Cold Spring HarborGoogle Scholar