Influenza AH1N2 Viruses, United Kingdom, 2001-02 Influenza Season

Joanna S. Ellis, Adriana Alvarez-Aguero, Vicky Gregory, Yi Pu Lin, A. Hay, Maria C. Zambon


Emerging Infectious Diseases. 2003;9(2) 

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In the winter of 2001-02, which was a mild influenza season, influenza A H1N2 viruses were detected for the first time in the U.K. Analysis of the diversity of >200 H1N2 isolates from the U.K indicated that they were closely related antigenically and genetically and derived the HA gene from A/NewCaledonia/20/99-like H1N1 viruses, whereas the other seven genes originated from recently circulating H3N2 viruses. The H1N2 viruses were isolated throughout the influenza season and co-circulated with H3N2 viruses. In contrast to the previous winter's results, few influenza A H1N1 viruses were detected. Retrospective RT-PCR analysis of 61.4% of influenza AH1 viruses isolated during 2000-01 in the U.K. identified one H1N2 virus that had been isolated in March 2001, indicating that H1N2 viruses had not circulated widely in the U.K. before becoming established in autumn of 2001. H1N2 viruses have also been identified during 2001-02 from outbreaks of influenza in different countries in Africa, America, Asia, and Europe.[28] The earliest H1N2 viruses identified worldwide retrospectively were also isolated in March 2001 in Saudi Arabia.[23]

During the past decade, influenza A H1N1 viruses have circulated intermittently in the U.K.[29] Between October 2000 and April 2001, a season of low influenza activity in the U.K., influenza A H1N1 viruses were the predominate influenza A strain circulating. Most H1N1 viruses were antigenically closely related to the vaccine strain used in the 2000-01 influenza season, A/NewCaledonia/20/99. The circulation of H1N2 reassortant viruses appears to have displaced A/NewCaledonia/20/99-like H1N1 viruses, although circulation of H1N2 does not appear to be associated with the generation of viruses with antigenically different surface proteins since the HA and NA of H1N2 viruses are antigenically similar to those of recently circulating H1N1 and H3N2 viruses. The observation that H1N2 viruses were isolated mainly from children <15 years of age suggests a preexisting immunity to H1 and N2 in the population (Figure 2). Furthermore, a similar proportion of the H3N2 viruses isolated during 2001-02 (79%) were also isolated from children <15 years old, with A/Panama/2007/99-like H3N2 viruses circulating during the same period of time as the A/NewCaledonia/20/99-like H1N1 viruses in the U.K. This conclusion is supported by the observational disease data in which the age group most severely affected by the influenza viruses circulating was the 5-14 age group and suggests that disease associated with both H1N2 and H3N2 was mainly in children acquiring a primary infection. In contrast, during the 2000-01 winter season, when influenza A H1N1 and influenza B viruses co-circulated, the disease data demonstrated that the age groups most affected by influenza viruses were the 5-14 years of age group and 15-44 years of age group (available from: URL: The age distribution of the disease data correlates with the finding that most H1N1 viruses isolated during 2000-01 were from the 5-14 years of age and 15-44 years of age groups.

Our analysis of genetic variation in the HA gene showed that seven amino acid substitutions have accumulated in the HA1 gene of influenza A H1N1 viruses between the circulation of A/Beijing/252/95-like H1N1 viruses in the U.K. and the emergence of viruses similar to the new variant, A/NewCaledonia/20/99, in October 2000. Of these, four mutations (T136S, E156G, S186P, and I194L) are located in two of the five antigenic sites (A and B) of HA1. The emergence of A/NewCaledonia/20/99-like viruses correlates with the finding that new drift variants of epidemiologic importance typically have four or more amino acid substitutions located in two or more of the antigenic sites.[30] Additionally, the substitutions A193T and A218T are located in antigenic sites B and D, respectively, of HA1, indicating the potential for further drift in the HA gene of H1N2 viruses and the possible subsequent generation of a new epidemic variant.

Although H1N1 and H3N2 subtypes have co-circulated in the human population since 1977, reassortant combinations of HA and NA subtypes are rare. Reassortant H1N2 viruses have previously been isolated from sporadic cases in humans and were not maintained in circulation in humans.[9] The H1 HA-N2 NA combination may provide a better functional match between receptor-binding and receptor-destroying activities of HA and NA, respectively, perhaps providing H1N2 viruses with a fitness advantage over contemporary H1N1, H3N2, or both viruses. Postreassortment mutations in the HA gene located in the vicinity of the receptor-binding pocket have previously been shown to compensate for any imbalance between HA and NA activities and may be a factor in influenza virus evolution.[31,32] Analysis of the HA1 sequences of H1N1 and H1N2 viruses isolated between December 2000 and February 2002 in the U.K. showed three substitutions at positions 178, 193, and 218 that were present only in the HA gene of the H1N2 viruses analyzed; residue 193 situated near to the HA receptor binding pocket. As these three mutations were not observed in the HA1 of recent H1N1 viruses analyzed, the substitutions may have occurred in H1N2 viruses after reassortment.

The NA genes of the H1N2 viruses are closely related genetically to the NA genes of A/Moscow/10/99-like H3N2 viruses. Of the three substitutions observed in the NA genes of H1N2 viruses compared to those of A/Moscow/10/99, residues 199 and 431 are located in antigenic sites on surface of the NA gene.[33,34] Residue 199 has also been assigned to one of the 12 phylogenetically important regions of the NA gene.[35] All of the amino acid residues that form the sialic acid-binding site of the NA gene were conserved in the NA of H1N2 viruses. Although changes in receptor binding and the interaction between the HA and NA proteins of H1N2 reassortants may have facilitated the emergence of the H1N2 viruses, the contribution of the internal proteins to the replicative efficiency and transmission of these viruses and their interaction with the surface glycoproteins is unknown.

The early detection and characterization of newly emerging influenza variants are two of the primary aims of the World Health Organization global influenza surveillance network. Reassortment between circulating influenza viruses leading to the emergence of a novel subtype, such as H1N2, highlights the need for subtyping of the influenza A viruses isolated. Most current diagnostic tests rely on the detection and typing of the HA of influenza viruses. Only a few laboratories perform analysis of the NA gene (as NA inhibition assays are difficult to perform) or analyze the internal proteins and the genes encoding them. The use of molecular techniques provides a rapid means for the detection and subtyping of influenza viruses, although current PCR assays target only N1 and N2 subtypes.[22,36] In addition, heteroduplex mobility assays can be used to genetically characterize the HA and internal genes of influenza viruses.[20,37,38] However, although molecular methods aid the rapid detection and identification of influenza viruses, virus isolation by culture is still required for antigenic characterization of influenza viruses. How influenza H1N2 reassortants will evolve and whether H1N2 viruses will be maintained in circulation in humans remain to be seen.