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Title: Resolving problems associated with cell-culture adaptation of influenza A(H3N2) viruses
Author: Brown, Jonathan
ISNI:       0000 0004 7963 8110
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2019
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Vaccination is the most effective public health intervention for preventing infection with seasonal influenza. However, due to the rapid mutation of influenza viruses circulating in the human population, vaccines must be regularly reformulated to keep pace with emerging antigenic variants. Failure to accurately determine which variants will be responsible for the majority of infections in the upcoming flu season results in reduced vaccine effectiveness and increased disease burden. Serological assays such as the haemagglutination inhibition (HAI) assay are used to determine the antigenic relationships between circulating influenza viruses and are central to selection of representative strains for the vaccine. The HAI assay relies on binding and agglutination of red blood cells (RBC) by the viral haemagglutinin (HA). However, modern influenza A(H3N2) isolates exhibit poor HA-mediated agglutination and the ability to agglutinate RBC through their neuraminidase (NA) surface protein making them problematic to characterise. Here the NA-mediated binding phenotype is confirmed to not occur in H3N2 clinical isolates but arises during passage of isolates in cell culture. Using a reverse genetics model of an H3N2 isolate (3C.2a subclade), A/Sydney/71/2014, mutations T148I/K and/or D151G associated with NA-mediated binding arose spontaneously during passage in MDCK as well as MDCK-SIAT1 cell culture. NA-mediated binding variants were detected in a wider panel of cell-passaged H3N2 isolates by next-generation sequencing and a single passage in a human airway epithelial (HAE) cell line, was sufficient to remove these variants in the majority of passage replicates (54%, n=24). Removal of artefactual NA-mediated binding is likely due to HAE cells presenting a subset of receptors which modern H3N2 viruses use but which are poorly represented on current culture cells. Here modification of cellular receptors is explored as a means to match environmental evolution of H3N2 receptor binding properties and avoid cell passage adaptations which threaten accurate H3N2 antigenic characterisation.
Supervisor: Harvey, Ruth ; Barclay, Wendy Sponsor: National Institute for Biological Standards and Control
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral