The Ability of a Non-Egg Adapted (Cell-Like) A(H1N1)pdm09 Virus to Egg-Adapt at HA Loci Other than 222 and 223 and Its Effect on the Yield of Viral Protein

Abstract

Previous studies on influenza A(H1N1)pdm09 candidate vaccine viruses (CVVs) that had adapted to growth in embryonated chicken eggs by the acquisition of amino acid substitutions at HA positions 222 or 223 showed that improved protein yield could be conferred by additional amino acid substitutions in the haemagglutinin (HA) that arose naturally during passaging of the virus in eggs. In this study we investigated, by means of reverse genetics, the ability of a non-egg adapted (cell-like) A(H1N1)pdm09 virus to egg-adapt at HA loci other than 222/223, introducing amino acid substitutions previously identified as egg adaptations in pre-H1N1pdm09 H1N1 viruses and assessing their effect on protein yield and antigenicity. We also investigated the effect on the protein yield of these substitutions in viruses that had A(H1N1)pdm09 internal genes rather than the traditional PR8 internal genes of a CVV. The data show that a cell-like A/Christchurch/16/2010 can be egg-adapted via amino acid substitutions in at least three alternative HA loci (187, 190 and 216), in viruses with either PR8 or A/California/7/2009 internal genes, but that the effects on protein yield vary depending on the amino acid substitution and the internal genes of the virus. Since CVVs need to produce high protein yields to be suitable for vaccine manufacture, the findings of this study will assist in the future characterisation of both wild type viruses and lab-derived CVVs for vaccine use.

Assessment of the antigenic evolution of a clade 6B.1 human H1N1pdm influenza virus revealed differences between ferret and human convalescent sera

“… 10 A virus with the D187A substitution (which became dominant in clade 6B.1A.5a.1) could not be rescued by Nicolson et al. 10 and did not emerge in our study, suggesting that this substitution may have had a disadvantage in the genetic background of 2015–2016 H1N1pdm viruses, while the D187A substitution may have conferred an advantage in the context of H1N1pdm viruses circulating from 2018 onwards. HA position 187 is located on the ‘inner rim’ of the receptor-binding pocket.…”

“… 9 Moreover, passages of a cell-grown H1N1pdm influenza virus in embryonated chicken eggs resulted in mutations at HA positions 187, 190, and 216. 10 At position 187, three different substitutions (D187N, D187V, and D187A) emerged during egg passages, but only the D187V substitutions (detected most frequently in our study, which was carried out in eukaryotic cells) yielded genetically stable virus. 10 A virus with the D187A substitution (which became dominant in clade 6B.1A.5a.1) could not be rescued by Nicolson et al.…”

“… 10 At position 187, three different substitutions (D187N, D187V, and D187A) emerged during egg passages, but only the D187V substitutions (detected most frequently in our study, which was carried out in eukaryotic cells) yielded genetically stable virus. 10 A virus with the D187A substitution (which became dominant in clade 6B.1A.5a.1) could not be rescued by Nicolson et al. 10 and did not emerge in our study, suggesting that this substitution may have had a disadvantage in the genetic background of 2015–2016 H1N1pdm viruses, while the D187A substitution may have conferred an advantage in the context of H1N1pdm viruses circulating from 2018 onwards.…”

The Effect of Mice Adaptation Process on the Pathogenicity of Influenza A/South Africa/3626/2013 (H1N1)pdm09 Model Strain

“…In addition, the R variant is usually a result of egg adaptation. The mechanism of egg adaptation was illustrated early [21,22]. It was shown that the S190R variant of the A/Hunan/26/2016 (H1N1)pdm09 virus had increased pathogenicity in mice [21].…”