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The optimisation of Pseudotyped viruses for the characterisation of immune responses to equine influenza virus

Scott, Simon D.; Kinsley, Rebecca; Temperton, Nigel; Daly, Janet M.

Authors

Simon D. Scott

Rebecca Kinsley

Nigel Temperton

Janet M. Daly



Abstract

Pseudotyped viruses (PVs) produced by co-transfecting cells with plasmids expressing lentiviral core proteins and viral envelope proteins are potentially powerful tools for studying various aspects of equine influenza virus (EIV) biology. The aim of this study was to optimise production of equine influenza PVs. Co-transfection of the HAT protease to activate the haemagglutinin (HA) yielded a higher titre PV than TMPRSS2 with the HA from A/equine/Richmond/1/2007 (H3N8), whereas for A/equine/Newmarket/79 (H3N8), both proteases resulted in equivalent titres. TMPRSS4 was ineffective with the HA of either strain. There was also an inverse relationship between the amount of protease-expression plasmids and the PV titre obtained. Interestingly, the PV titre obtained by co-transfection of a plasmid encoding the cognate N8 NA was not as high as that generated by the addition of exogenous neuraminidase (NA) from Clostridium perfringens to allow the release of nascent PV particles. Finally, initial characterisation of the reliability of PV neutralisation tests (PVNTs) demonstrated good intra-laboratory repeatability. In conclusion, we have demonstrated that equine influenza PV production can be readily optimised to provide a flexible tool for studying EIV.

Citation

Scott, S. D., Kinsley, R., Temperton, N., & Daly, J. M. (2016). The optimisation of Pseudotyped viruses for the characterisation of immune responses to equine influenza virus. Pathogens, 5(4), https://doi.org/10.3390/pathogens5040068

Journal Article Type Article
Acceptance Date Dec 4, 2016
Publication Date Dec 15, 2016
Deposit Date Jan 9, 2017
Publicly Available Date Jan 9, 2017
Journal Pathogens
Electronic ISSN 2076-0817
Publisher MDPI
Peer Reviewed Peer Reviewed
Volume 5
Issue 4
Article Number 68
DOI https://doi.org/10.3390/pathogens5040068
Keywords equine influenza; pseudotyped virus; neutralisation assay
Public URL http://eprints.nottingham.ac.uk/id/eprint/39687
Publisher URL http://www.mdpi.com/2076-0817/5/4/68
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0





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