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Acquisition of human-type receptor binding specificity by new H5N1 influenza virus sublineages during their emergence in birds in Egypt.

Watanabe Y, Ibrahim MS, Ellakany HF, Kawashita N, Mizuike R, Hiramatsu H, Sriwilaijaroen N, Takagi T, Suzuki Y, Ikuta K - PLoS Pathog. (2011)

Bottom Line: The phylogenetic results showed that recent human isolates clustered disproportionally into several new H5 sublineages suggesting that their HAs have changed their receptor specificity.Using reverse genetics, we found that these H5 sublineages have acquired an enhanced binding affinity for α2,6 SA in combination with residual affinity for α2,3 SA, and identified the amino acid mutations that produced this new receptor specificity.Interestingly, these H5 viruses, with increased affinity to α2,6 SA, emerged during viral diversification in bird populations and subsequently spread to humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Virology, Research Institute for Microbial Diseases (BIKEN), Osaka University, Osaka, Japan. nabe@biken.osaka-u.ac.jp

ABSTRACT
Highly pathogenic avian influenza A virus subtype H5N1 is currently widespread in Asia, Europe, and Africa, with 60% mortality in humans. In particular, since 2009 Egypt has unexpectedly had the highest number of human cases of H5N1 virus infection, with more than 50% of the cases worldwide, but the basis for this high incidence has not been elucidated. A change in receptor binding affinity of the viral hemagglutinin (HA) from α2,3- to α2,6-linked sialic acid (SA) is thought to be necessary for H5N1 virus to become pandemic. In this study, we conducted a phylogenetic analysis of H5N1 viruses isolated between 2006 and 2009 in Egypt. The phylogenetic results showed that recent human isolates clustered disproportionally into several new H5 sublineages suggesting that their HAs have changed their receptor specificity. Using reverse genetics, we found that these H5 sublineages have acquired an enhanced binding affinity for α2,6 SA in combination with residual affinity for α2,3 SA, and identified the amino acid mutations that produced this new receptor specificity. Recombinant H5N1 viruses with a single mutation at HA residue 192 or a double mutation at HA residues 129 and 151 had increased attachment to and infectivity in the human lower respiratory tract but not in the larynx. These findings correlated with enhanced virulence of the mutant viruses in mice. Interestingly, these H5 viruses, with increased affinity to α2,6 SA, emerged during viral diversification in bird populations and subsequently spread to humans. Our findings suggested that emergence of new H5 sublineages with α2,6 SA specificity caused a subsequent increase in human H5N1 influenza virus infections in Egypt, and provided data for understanding the virus's pandemic potential.

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Effect of HA mutations in sublineage A viruses on receptor specificity of EG/D1 virus HA.(A) The two mutations found in the HAs of sublineage A viruses were introduced into the HA of EG/D1 virus as single and double mutations. (B) The reverse mutations were introduced into the HA of EG/12 virus. Direct binding to sialylglycopolymers containing either α2,3-linked (blue) or α2,6-linked (red) sialic acid was assayed. Mutations are indicated by subscripts. Each data point is the mean ± SD of triplicate experiments.
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ppat-1002068-g003: Effect of HA mutations in sublineage A viruses on receptor specificity of EG/D1 virus HA.(A) The two mutations found in the HAs of sublineage A viruses were introduced into the HA of EG/D1 virus as single and double mutations. (B) The reverse mutations were introduced into the HA of EG/12 virus. Direct binding to sialylglycopolymers containing either α2,3-linked (blue) or α2,6-linked (red) sialic acid was assayed. Mutations are indicated by subscripts. Each data point is the mean ± SD of triplicate experiments.

Mentions: To identify mutations enabling α2,6 SA binding, we focused on viruses in sublineages A and B, to which most human isolates belonged. Comparison of 6 HA sequences of sublineage A viruses with 100 HA sequences of other H5 viruses isolated in Egypt identified two amino acid changes in the sublineage A virus HAs (Table 1): Q192H and S235P (H5 HA numbering). Introduction of the Q192H mutation into EG/D1 HA (denoted rEG/D1Q192H) markedly increased viral binding to α2,6 SA (Figure 3A). However, introduction of the S235P mutation into EG/D1 HA (denoted rEG/D1S235P) only slightly increased α2,6 SA binding. There was no synergistic effect with both mutations: the double mutant had similar α2,6 SA binding to that of the single Q192H mutant. In contrast, the H192Q mutation, but not the P235S mutation, in HAs of EG/12 (denoted rEG/D1-EG/12 HAH192Q) and EG/4822 decreased α2,6 SA binding (Figures 3B and S3A). These findings suggested that the Q192H mutation in H5N1 avian viruses increased the binding affinity of HA for the human receptor.


Acquisition of human-type receptor binding specificity by new H5N1 influenza virus sublineages during their emergence in birds in Egypt.

Watanabe Y, Ibrahim MS, Ellakany HF, Kawashita N, Mizuike R, Hiramatsu H, Sriwilaijaroen N, Takagi T, Suzuki Y, Ikuta K - PLoS Pathog. (2011)

Effect of HA mutations in sublineage A viruses on receptor specificity of EG/D1 virus HA.(A) The two mutations found in the HAs of sublineage A viruses were introduced into the HA of EG/D1 virus as single and double mutations. (B) The reverse mutations were introduced into the HA of EG/12 virus. Direct binding to sialylglycopolymers containing either α2,3-linked (blue) or α2,6-linked (red) sialic acid was assayed. Mutations are indicated by subscripts. Each data point is the mean ± SD of triplicate experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3102706&req=5

ppat-1002068-g003: Effect of HA mutations in sublineage A viruses on receptor specificity of EG/D1 virus HA.(A) The two mutations found in the HAs of sublineage A viruses were introduced into the HA of EG/D1 virus as single and double mutations. (B) The reverse mutations were introduced into the HA of EG/12 virus. Direct binding to sialylglycopolymers containing either α2,3-linked (blue) or α2,6-linked (red) sialic acid was assayed. Mutations are indicated by subscripts. Each data point is the mean ± SD of triplicate experiments.
Mentions: To identify mutations enabling α2,6 SA binding, we focused on viruses in sublineages A and B, to which most human isolates belonged. Comparison of 6 HA sequences of sublineage A viruses with 100 HA sequences of other H5 viruses isolated in Egypt identified two amino acid changes in the sublineage A virus HAs (Table 1): Q192H and S235P (H5 HA numbering). Introduction of the Q192H mutation into EG/D1 HA (denoted rEG/D1Q192H) markedly increased viral binding to α2,6 SA (Figure 3A). However, introduction of the S235P mutation into EG/D1 HA (denoted rEG/D1S235P) only slightly increased α2,6 SA binding. There was no synergistic effect with both mutations: the double mutant had similar α2,6 SA binding to that of the single Q192H mutant. In contrast, the H192Q mutation, but not the P235S mutation, in HAs of EG/12 (denoted rEG/D1-EG/12 HAH192Q) and EG/4822 decreased α2,6 SA binding (Figures 3B and S3A). These findings suggested that the Q192H mutation in H5N1 avian viruses increased the binding affinity of HA for the human receptor.

Bottom Line: The phylogenetic results showed that recent human isolates clustered disproportionally into several new H5 sublineages suggesting that their HAs have changed their receptor specificity.Using reverse genetics, we found that these H5 sublineages have acquired an enhanced binding affinity for α2,6 SA in combination with residual affinity for α2,3 SA, and identified the amino acid mutations that produced this new receptor specificity.Interestingly, these H5 viruses, with increased affinity to α2,6 SA, emerged during viral diversification in bird populations and subsequently spread to humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Virology, Research Institute for Microbial Diseases (BIKEN), Osaka University, Osaka, Japan. nabe@biken.osaka-u.ac.jp

ABSTRACT
Highly pathogenic avian influenza A virus subtype H5N1 is currently widespread in Asia, Europe, and Africa, with 60% mortality in humans. In particular, since 2009 Egypt has unexpectedly had the highest number of human cases of H5N1 virus infection, with more than 50% of the cases worldwide, but the basis for this high incidence has not been elucidated. A change in receptor binding affinity of the viral hemagglutinin (HA) from α2,3- to α2,6-linked sialic acid (SA) is thought to be necessary for H5N1 virus to become pandemic. In this study, we conducted a phylogenetic analysis of H5N1 viruses isolated between 2006 and 2009 in Egypt. The phylogenetic results showed that recent human isolates clustered disproportionally into several new H5 sublineages suggesting that their HAs have changed their receptor specificity. Using reverse genetics, we found that these H5 sublineages have acquired an enhanced binding affinity for α2,6 SA in combination with residual affinity for α2,3 SA, and identified the amino acid mutations that produced this new receptor specificity. Recombinant H5N1 viruses with a single mutation at HA residue 192 or a double mutation at HA residues 129 and 151 had increased attachment to and infectivity in the human lower respiratory tract but not in the larynx. These findings correlated with enhanced virulence of the mutant viruses in mice. Interestingly, these H5 viruses, with increased affinity to α2,6 SA, emerged during viral diversification in bird populations and subsequently spread to humans. Our findings suggested that emergence of new H5 sublineages with α2,6 SA specificity caused a subsequent increase in human H5N1 influenza virus infections in Egypt, and provided data for understanding the virus's pandemic potential.

Show MeSH
Related in: MedlinePlus