Use of Pseudotyped viruses for the production of reference materials as part of emerging viral outbreak preparedness

Abstract

Purpose: The recent public health emergencies of international concern (PHEIC), caused by Ebola and Zika virus, have highlighted the lack of prophylactic treatments and need for effective diagnostics for emerging viruses. In response to this, we established International Reference Reagents for antibody detection, which enables the comparison of results from laboratories worldwide undertaking treatment/vaccine efficacy clinical trials. The requirement for high containment facilities to handle viruses with outbreak potential, such as BSL4 for Ebola virus, is a constraining challenge. We have developed serological assays using replication defective pseudotyped viruses (PV) for the evaluation of reference material to avert high containment requirements. Methods & Materials: The preferred candidate material is a pool of plasma or sera from convalescent patients as this provides a commutable standard. For the reference material for Ebola antibody, several donations were received at NIBSC and solvent-detergent treated to ensure inactivation of virus. Characterisation is performed in-house using both PV neutralisation assays and ELISA platforms. PV is produced via plasmid transfection of 293T cells, with plasmids encoding a lentiviral core component, a reporter gene which is packaged within the core and the sequence of the envelope protein for the virus of interest. The ability of candidate material to block PV entry into target cells is measured via reporter signal reduction. Results: Following a WHO-sponsored International Collaborative study, the 1st WHO International Standard for Ebola virus antibodies was established in 2017. Participants in the collaborative study evaluated the candidate materials using assays available in their laboratories. Data from neutralisation assays employing PV with either a lentiviral or vesiculoviral core component was correlated with the wildtype assay and showed a better correlation coefficient when the vesiculovirus PV system was used. Conclusion: To support preparedness activities, antibody reference materials are now being established for other WHO priority pathogens such as Lassa and Nipah virus. To characterise the serological material at a low containment level, we have established assays using PV generated with both the lentiviral and vesiculovirus platforms. The two systems are currently being investigated to determine the most appropriate for each high containment virus.