‘I had a little bird, its name was Enza. I opened the window, and in-flu-enza.’ Children's rhyme c. 1918.
The recent emergence of a novel H1N1 ‘swine flu’ and the H5N1 ‘avian flu’ has thrown into sharp relief the importance of natural populations of influenza viruses in predicting pandemic influenza. The influenza A virus has the potential to cause high levels of mortality in humans, and in contrast to its less virulent relatives, influenza B and C, influenza A virus is repeatedly reintroduced into the human population from animal reservoirs.
Influenza A viruses possess an RNA genome that encodes 10 genes and are divided into eight segments. Segments might be viewed as individual chromosomes, each containing only one or two genes. Two of the gene products, haemagglutinin (HA) and neuroaminidase (NA), have antigenic potential (i.e. the ability to elicit an antibody response from its host). The name of an influenza subtype refers to the specific HA and NA alleles (e.g. H1N1 ‘1918 Spanish flu’, H3N2 ‘1968 Hong Kong flu’, H5N1 ‘avian flu’). Subtypes were traditionally distinguished by the inability of neutralizing antibodies to cross-react among them. So far, 16 HA and 9 NA antigenic subtypes have been identified (Dugan et al., 2008).
Influenza virus generates variation via two processes: mutation and reassortment. Mutation allows for escape from the host immune system and has produced a diverse array of subtypes, possibly over the last several thousand years (Chen and Holmes, 2006). This gradual change is called ‘antigenic drift’ (but note that this nomenclature is slightly misleading because antigenic drift can be the product of natural selection, rather than genetic drift).
Reassortment, a truly impressive evolutionary feature by which the influenza virus generates variation, is the exchange of genomic segments and can occur between distantly related lineages. This genetic exchange usually involves whole genomic segments; homologous recombination appears to be exceptionally rare within influenza gene segments (Boni et al., 2008). When two distantly related influenza subtypes, say H9N2 and H7N3, simultaneously infect a single host cell, they can reassort to produce novel antigenic combinations, such as H9N3 and H7N2; other segments not containing the HA and NA genes can also reassort, creating highly mosaic viral genomes. When reassortment occurs in NA and HA segments, it is termed ‘antigenic shift’ because it introduces a novel antigenic profile into a genetic background.
Although influenza A virus has been found in a diverse array of animals, including birds, swine, horses, cats, aquatic mammals such as seals and whales, and humans (Webby et al., 2007), the viral genomic segments from each of these species have one thing in common: they can all be traced back to influenza in waterfowl and shorebirds, who act as the primary reservoirs of the virus (Webster et al., 1992). It is from these viral populations that all influenza A viruses originate (Olsen et al., 2006).
In aquatic birds, influenza virus is generally asymptomatic and transmitted via the faecal-oral route. The virus remains viable in water for several days at ambient temperatures, providing an efficient mode of transmission (Webster et al., 1978). Moreover, influenza can spread over broad distances by the annual migrations of avian hosts (Olsen et al., 2006). These migrations result in both a greater range of the virus and its increased spread among various avian species. In fact, viruses isolated from multiple species at the same place and time are more likely to be closely related than viruses isolated from a single species at different places and/or times (Chen and Holmes, 2009).
Avian influenza virus was once assumed to be in a sort of evolutionary stasis, in which the virus had reached a fitness maximum within its natural hosts (Webster et al., 1992; 2007). Sequence analysis of avian subtypes suggests that the opposite may be true. A study of Canadian ducks revealed that genetic subtypes in wild populations are constantly undergoing reassortment, thereby producing segments with a myriad of evolutionary histories (Hatchette et al., 2004). Due to extensive reassortment, there is no single phylogenetic tree of an influenza subtype, only a history of its genes. In addition, the substitution rate of influenza virus in birds is remarkably rapid, and the virus evolves under similar selective constraints as human influenza virus (Chen and Holmes, 2006). Rather than being in evolutionary stasis, avian influenza genomes appear to be in a state of constant flux.
Although birds are the natural reservoir for influenza virus, swine also harbour a diverse array of viral subtypes (Van Reeth, 2007); but, unlike birds, swine transmit the virus via the respiratory route and are susceptible to pathogenic infection. American swine are infected with a ‘classic’ H1N1 subtype that is closely related to the ‘1918 Spanish flu’ as well as a different H1N1 subtype from ducks that emerged in 1979 (Van Reeth, 2007). In Europe, this more recent H1N1 has replaced the ‘classic’ H1N1 subtype. Swine are also infected with H1N2 and H3N2 subtypes and a ‘triple reassortant’ subtype, whose genome contains elements from avian, swine, and human influenza viruses (Zhou et al., 1999). The novel H1N1 ‘swine flu’ pandemic in humans appears to be the result of reassortment among H1N1, H1N2, H3N2 and the ‘triple reassortant’ subtypes (Smith et al., 2009).
Swine have long been thought to serve as the mixing vessel for human influenza epidemics and pandemics (Ito et al., 1998). Humans and birds have slightly altered versions of sialic acids that act as cell receptors for the influenza virus, whereas swine have both versions; this observation led to the hypothesis that swine can act as an intermediate host between birds and humans. Although swine clearly have the potential to serve as the source of influenza reassortment [a small outbreak of H3N2 in humans in 1993 (Claas et al., 1994) and the current H1N1 ‘swine flu’ pandemic (Fraser et al., 2009) appear to be evidence of this], it is unclear if this type of reassortment led to the 1918 human pandemic (Reid and Taubenberger, 2003). Therefore, the importance of intermediate hosts in producing pandemic influenza is still unclear.
Although all influenza A viruses can be traced back to birds, monitoring the virus in many species is important for public health. Surprisingly, surveillance of swine influenza appears to have missed the immediate ancestor of the novel H1N1 ‘swine flu’ pandemic virus (Smith et al., 2009). Tracking the diversity of naturally occurring influenza virus may be able to provide a warning of, and possibly even prevent, future pandemics.