Abstract
Influenza A(H3N2) viruses evade human immunity primarily by acquiring antigenic changes in the haemagglutinin (HA). HA receptor-binding features of contemporary A(H3N2) viruses hinder traditional antigenic characterization using haemagglutination inhibition and promote selection of HA mutants. Thus, alternative approaches are needed to reliably assess antigenic relatedness between circulating viruses and vaccines. We developed a high content imaging-based neutralization test (HINT) to reduce antigenic mischaracterization resulting from virus adaptation to cell culture. Ferret reference antisera were raised using clinical specimens containing viruses representing recent vaccine strains. Analysis of viruses circulating during 2011–2018 showed that gain of an N158-linked glycosylation in HA was a molecular determinant of antigenic distancing between A/Hong Kong/4801/2014-like (clade 3C.2a) and A/Texas/50/2012-like viruses (clade 3C.1), while multiple evolutionary HA F193S substitution were linked to antigenic distancing from A/Switzerland/97152963/2013-like (clade 3C.3a) and further antigenic distancing from A/Texas/50/2012-like viruses. Additionally, a few viruses carrying HA T135K and/or I192T showed reduced neutralization by A/Hong Kong/4801/2014-like antiserum. Notably, this technique elucidated the antigenic characteristics of clinical specimens, enabling direct characterization of viruses produced in vivo, and eliminating in vitro culture, which rapidly alters the genotype/phenotype. HINT is a valuable new antigenic analysis tool for vaccine strain selection.
Highlights
Influenza A viruses of the H3N2 antigenic subtype are important respiratory pathogens causing annual outbreaks of human illness since their emergence as a pandemic virus in 1968
high content imaging-based neutralization test (HINT) was carried out in SIAT1 cell cultures, due to their higher expression level of α-2,6-linked sialic acid receptors compared to regular MDCK cells, which makes them the preferred cell line to culture contemporary seasonal A(H3N2) viruses21
HINT titres were determined by calculating the reciprocal dilution of the antiserum needed to reduce the infected cell population (ICP) by 50% (IC50) compared to the control wells with no serum (100% infection) by curve fitting analysis
Summary
Influenza A viruses of the H3N2 antigenic subtype are important respiratory pathogens causing annual outbreaks of human illness since their emergence as a pandemic virus in 1968. WHO CCs present their data at the bi-annual vaccine selection consultation meeting where decisions are made regarding the need for updating one or more vaccine components These decisions require scientific evidence of antigenic drift and depend on availability of suitable candidate vaccine viruses. Forecasting the major antigenic groups of influenza viruses that are most likely to dominate in the season and producing suitable egg-propagated vaccine viruses is a daunting task, and various degrees of antigenic divergence (mismatch) have occurred over the years. This was the case for the Northern Hemisphere (NH) 2014–15 influenza season. Special attention has been paid to the development and application of new assays to better characterize circulating influenza viruses
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