Abstract
The swine-origin pandemic A(H1N1)2009 virus, A(H1N1)pdm09, is still circulating in parts of the human population. To monitor variants that may escape from vaccination specificity, antigenic characterization of circulating viruses is important. In this study, a hybridoma clone producing human monoclonal antibody against A(H1N1)pdm09, designated 5E4, was prepared using peripheral lymphocytes from a vaccinated volunteer. The 5E4 showed viral neutralization activity and inhibited hemagglutination. 5E4 escape mutants harbored amino acid substitutions (A189T and D190E) in the hemagglutinin (HA) protein, suggesting that 5E4 recognized the antigenic site Sb in the HA protein. To study the diversity of Sb in A(H1N1)pdm09, 58 viral isolates were obtained during the 2009/10 and 2010/11 winter seasons in Osaka, Japan. Hemagglutination-inhibition titers were significantly reduced against 5E4 in the 2010/11 compared with the 2009/10 samples. Viral neutralizing titers were also significantly decreased in the 2010/11 samples. By contrast, isolated samples reacted well to ferret anti-A(H1N1)pdm09 serum from both seasons. Nonsynonymous substitution rates revealed that the variant Sb and Ca2 sequences were being positively selected between 2009/10 and 2010/11. In 7,415 HA protein sequences derived from GenBank, variants in the antigenic sites Sa and Sb increased significantly worldwide from 2009 to 2013. These results indicate that the antigenic variants in Sb are likely to be in global circulation currently.
Highlights
In April 2009, the swine-origin pandemic A(H1N1)2009 virus, A(H1N1)pdm09, emerged, originating from the swine H1 virus in North America and the avian-like swine virus in Europe [1,2]
Establishment of human MAbs (HuMAbs) against A(H1N1)pdm09 peripheral blood mononuclear cells (PBMCs) were prepared from a healthy volunteer 2 weeks after vaccination with the HA split vaccine, which included the A/California/7/2009 strain, and were used for the fusion with SPYMEG cells
A hybridoma clone was established that produced a HuMAb, designated 5E4, against A(H1N1)pdm09, which did not react with seasonal influenza A (A/New Caledonia/20/1999 and A/Brisbane/ 59/2007 for H1N1 and A/Hiroshima/52/2005 and A/Uruguay/ 716/2007 for H3N2) or influenza B viruses (B/Florida/4/2006 and B/Malaysia/2506/2004), according to immunofluorescence assay (IFA)
Summary
In April 2009, the swine-origin pandemic A(H1N1)2009 virus, A(H1N1)pdm, emerged, originating from the swine H1 virus in North America and the avian-like swine virus in Europe [1,2]. A(H1N1)pdm spread rapidly across the world and is still circulating among humans. A(H1N1)pdm has remained closely related to one of the earliest viruses isolated, A/California/ 7/2009, with little change in genetic makeup even in the most variable genes, hemagglutinin (HA) and neuraminidase (NA) [4,5]. The lack of significant antigenic change was reflected in the WHO vaccine formulation decision to recommend the use of an A/California/7/2009-like strain for developing northern hemisphere 2013/14 influenza vaccines [6]. Continued surveillance and antigenic characterization of circulating viruses are crucial to the identification of emerging variants that show significant evolution and that may require the selection of alternative viruses for developing a future vaccine
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