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Antigenic characterisation of yeast-expressed lyssavirus nucleoproteins

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Abstract

In Europe, three genotypes of the genus Lyssavirus, family Rhabdoviridae, are present, classical rabies virus (RABV, genotype 1), European bat lyssavirus type 1 (EBLV-1, genotype 5) and European bat lyssavirus type 2 (EBLV-2, genotype 6). The entire authentic nucleoprotein (N protein) encoding sequences of RABV (challenge virus standard, CVS, strain), EBLV-1 and EBLV-2 were expressed in yeast Saccharomyces cerevisiae at high level. Purification of recombinant N proteins by caesium chloride gradient centrifugation resulted in yields between 14–17, 25–29 and 18–20 mg/l of induced yeast culture for RABV-CVS, EBLV-1 and EBLV-2, respectively. The purified N proteins were evaluated by negative staining electron microscopy, which revealed the formation of nucleocapsid-like structures. The antigenic conformation of the N proteins was investigated for their reactivity with monoclonal antibodies (mAbs) directed against different lyssaviruses. The reactivity pattern of each mAb was virtually identical between immunofluorescence assay with virus-infected cells, and ELISA and dot blot assay using the corresponding recombinant N proteins. These observations lead us to conclude that yeast-expressed lyssavirus N proteins share antigenic properties with naturally expressed virus protein. These recombinant proteins have the potential for use as components of serological assays for lyssaviruses.

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Acknowledgements

The authors thank Roland Riebe for cultivation of hybridomas and provision of cell culture supernatants, Susanne Granzow and Marc Mertens for their help and Jeanette Kliemt for excellent technical assistance. The experiments were supported by grant of Lithuanian Science and Study Foundation, Grant No. B11. The lyssavirus N protein-reactive mAbs and EBLV-1 and EBLV-2 strains were kindly provided by James H. Cox (Tübingen). The Neospora caninum-specific mAbs 5.2.15 and 1.19.1 were a generous gift of Gereon Schares (Wusterhausen). N Johnson and AR Fooks were funded by Defra (UK) Grant SE0524.

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Author notes

  1. Rainer G. Ulrich

    Present address: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute for Novel and Emerging Infectious Diseases, Boddenblick 5a, 17493, Greifswald – Insel Riems, Germany

Authors and Affiliations

  1. Institute of Biotechnology, V. Graičiūno 8, 02241, Vilnius, Lithuania

    Indre Kucinskaite, Mindaugas Juozapaitis, Andrius Serva, Aurelija Zvirbliene & Kestutis Sasnauskas

  2. Veterinary Laboratories Agency – Weybridge, Woodham Lane, Surrey, KT15 3NB, UK

    Nicholas Johnson & Anthony R. Fooks

  3. Institute of Botany, Zaliuju ezeru 49, 08406, Vilnius, Lithuania

    Juozas Staniulis

  4. Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Epidemiology, Seestraße 55, 16868, Wusterhausen, Germany

    Thomas Müller & Rainer G. Ulrich

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  1. Indre Kucinskaite
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Correspondence to Rainer G. Ulrich.

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Kucinskaite, I., Juozapaitis, M., Serva, A. et al. Antigenic characterisation of yeast-expressed lyssavirus nucleoproteins. Virus Genes 35, 521–529 (2007). https://doi.org/10.1007/s11262-007-0127-7

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  • DOI: https://doi.org/10.1007/s11262-007-0127-7

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