Sally Furniss
Well-known member
<!-- #BeginEditable "content_area_2" -->[FONT=Arial, Helvetica, sans-serif]Recombination Resulting in Virulence Shift in Avian Influenza Outbreak, Chile[/FONT]
David L. Suarez,*
Dennis A. Senne,? Jill Banks,? Ian H. Brown,? Steve C. Essen,? Chang-Won Lee,* Ruth J. Manvell,? Christian Mathieu-Benson,? Valentina Moreno,? Janice C. Pedersen,? Brundaban Panigrahy,? Hern?n Rojas,? Erica Spackman,* and Dennis J. Alexander?
*Southeast Poultry Research Laboratory, Athens, Georgia, USA; ?National Veterinary Services Laboratories, Ames, Iowa, USA; ?Veterinary Laboratories Agency?Weybridge, New Haw, Addlestone, Surrey, United Kingdom; and ?Ministerio de Agricultura, Santiago, Chile
Influenza A viruses infecting chickens and turkeys are usually at one of two extremes of virulence. Highly pathogenic avian influenza (HPAI) viruses cause a systemic disease with rapid death in chickens and turkeys, which often approaches 100%. Low pathogenic avian influenza (LPAI) viruses cause a localized infection with little or no disease unless exacerbated by other organisms or poor environmental conditions. To date, all HPAI isolates have been of the H5 or H7 subtypes, although not all H5 or H7 subtype viruses cause HPAI.
Although the virulence of AI viruses for birds is a polygenic trait, one virulence factor is correlated with the hemagglutinin cleavage site. For all influenza A viruses, the hemagglutinin glycoprotein is produced as a precursor, HA0, which requires posttranslational cleavage by host proteases before it is functional and virus particles are infectious (1). All HPAI viruses examined to date have had a motif with multiple basic amino acids (arginine and lysine) at the HA0 cleavage site. In contrast, the cleavage motifs of LPAI viruses typically have only two basic amino acids, at positions ?1 and ?4 from the cleavage site for the H5 and at positions ?1 and ?3 for the H7 subtype (2). This difference appears to have a direct influence on viral virulence as LPAI viruses are limited to cleavage by host proteases such as trypsin-like enzymes and are thus restricted to replication at sites in the host where such enzymes are found, i.e., the respiratory and intestinal tracts. Whereas the multiple basic amino acids at the HA0 cleavage sites of HPAI viruses, either as a result of insertion or substitution (2?4), allows the HA0 precursor to be cleavable by ubiquitous host proteases (5). As a result the HPAI viruses are able to replicate systemically, damaging vital organs and tissues, which results in severe disease and death (1).
Viruses of the H5 or H7 subtype isolated from free-living birds are almost invariably of low pathogenicity for poultry. With the exception of a large die-off of terns in South Africa in 1961 (6), from which A/tern/South Africa/61 (H5N3) was isolated, HPAI virus isolations from free-living birds have been associated with contact with infected poultry, usually as a result of surveillance of birds trapped or found dead on infected poultry farms. In addition, results of phylogenetic studies of H7 subtype viruses indicate that HPAI viruses do not constitute a separate phylogenetic lineage or lineages but appear to arise from low pathogenic strains (7?9). This finding is supported by the in vitro selection of mutants virulent for chickens from an avirulent H7 virus (10) and the emergence of HPAI virus from an LPAI virus isolated from swans after repeated passage in chickens (11).
These findings conform to the theories of the molecular basis for the mutation of avian influenza subtype H5 and H7 viruses from low to high virulence in poultry put forward by Garcia et al. (12) and Perdue et al. (13). Essentially they propose that spontaneous duplication of purine triplets results in the insertion of basic amino acids at the HA0 cleavage site and that this occurs due to a transcription fault by the polymerase complex. As pointed out by Perdue et al. (13) this mechanism is clearly not the only means by which HPAI viruses arise, as some appear to result from nucleotide substitution rather than insertion, while others have insertions without repeating nucleotides. Attempts to assess the minimum requirements to confer virulence for H7 avian influenza viruses have used site-directed mutagenesis of the cleavage site of cloned HA followed by in vitro expression to examine the effect that progressive amino acid changes at each position of the cleavage site would have upon cleavability (3,14,15). For some H7 influenza viruses, the tetrapeptide motif R-X-R/K-R at the cleavage site leading up to the cleavage point is sufficient for cleavage by furin-like proteases, and variants with cleavage sites that do not conform to this motif are not pathogenic for chickens (3). However, motifs that differ from this standard have been observed from natural outbreaks of H7 HPAI (16).
Between 1959 and the end of 2001, a total of 18 primary outbreaks (10 H7 and 8 H5) of HPAI in poultry were reported (17). However, the geographic distribution of these outbreaks was not uniform across the world (five were in the British Isles; five in Australia; three in areas in Europe other than British Isles; and one each in Pakistan, Hong Kong, Canada, United States, and Mexico). Until 2002, no influenza virus had been isolated in poultry or wild birds in the continent of South America. In May 2002, a LPAI virus of H7N3 subtype was isolated from a broiler breeder flock in Chile, and in June, an HPAI virus of the same subtype was obtained from the same flock (18). We describe the characterization of the LPAI and HPAI isolates obtained in Chile with particular reference to the HPAI virus, which, while having a 10?amino acid (aa) insert at the HA0 cleavage site, does not conform to the dogma that a -R-X-R/K-R*G-L-F- motif is a prerequisite for HPAI viruses. We also present evidence that the 10-aa insert present in the HPAI viruses is the result of recombination between the HA and the nucleoprotein genes.
http://www.cdc.gov/ncidod/EID/vol10no4/03-0396.htm
David L. Suarez,*
Dennis A. Senne,? Jill Banks,? Ian H. Brown,? Steve C. Essen,? Chang-Won Lee,* Ruth J. Manvell,? Christian Mathieu-Benson,? Valentina Moreno,? Janice C. Pedersen,? Brundaban Panigrahy,? Hern?n Rojas,? Erica Spackman,* and Dennis J. Alexander?*Southeast Poultry Research Laboratory, Athens, Georgia, USA; ?National Veterinary Services Laboratories, Ames, Iowa, USA; ?Veterinary Laboratories Agency?Weybridge, New Haw, Addlestone, Surrey, United Kingdom; and ?Ministerio de Agricultura, Santiago, Chile
[FONT=Arial, Helvetica, sans-serif]Suggested citation for this article:[/FONT][FONT=Arial, Helvetica, sans-serif] Suarez DL, Senne DA, Banks J, Brown IH, Essen SC, Lee C-W, et al. Recombination resulting in virulence shift in avian influenza outbreak, Chile. Emerg Infect Dis [serial online] 2004 Apr [date cited]. Available from: http://www.cdc.gov/ncidod/EID/vol10no4/03-0396.htm[/FONT]
<hr>
[FONT=Arial, Helvetica, sans-serif]Influenza A viruses occur worldwide in wild birds and are occasionally associated with outbreaks in commercial chickens and turkeys. However, avian influenza viruses have not been isolated from wild birds or poultry in South America. A recent outbreak in chickens of H7N3 low pathogenic avian influenza (LPAI) occurred in Chile. One month later, after a sudden increase in deaths, H7N3 highly pathogenic avian influenza (HPAI) virus was isolated. Sequence analysis of all eight genes of the LPAI virus and the HPAI viruses showed minor differences between the viruses except at the hemagglutinin (HA) cleavage site. The LPAI virus had a cleavage site similar to other low pathogenic H7 viruses, but the HPAI isolates had a 30-nucleotide insert. The insertion likely occurred by recombination between the HA and nucleoprotein genes of the LPAI virus, resulting in a virulence shift. Sequence comparison of all eight gene segments showed the Chilean viruses were also distinct from all other avian influenza viruses and represent a distinct South American clade.[/FONT]
Influenza viruses are segmented, negative-sense, single-stranded RNA viruses of the family Orthomyxoviridae and are divided into the genera Influenzavirus A, B and C. However, only type A influenza viruses have been known to cause natural infections of birds. Type A influenza viruses are further divided into subtypes based on antigenic relationships of the hemagglutinin (HA) and neuraminidase (NA) surface glycoproteins. To date, 15 unique HA subtypes (H1?H15) and nine unique NA subtypes (N1?N9) have been recognized. Each virus has one HA and one NA protein, potentially in any combination. Viruses of all HA and NA subtypes with most possible combinations of the HA and NA subtypes have been isolated from avian species.
Influenza A viruses infecting chickens and turkeys are usually at one of two extremes of virulence. Highly pathogenic avian influenza (HPAI) viruses cause a systemic disease with rapid death in chickens and turkeys, which often approaches 100%. Low pathogenic avian influenza (LPAI) viruses cause a localized infection with little or no disease unless exacerbated by other organisms or poor environmental conditions. To date, all HPAI isolates have been of the H5 or H7 subtypes, although not all H5 or H7 subtype viruses cause HPAI.
Although the virulence of AI viruses for birds is a polygenic trait, one virulence factor is correlated with the hemagglutinin cleavage site. For all influenza A viruses, the hemagglutinin glycoprotein is produced as a precursor, HA0, which requires posttranslational cleavage by host proteases before it is functional and virus particles are infectious (1). All HPAI viruses examined to date have had a motif with multiple basic amino acids (arginine and lysine) at the HA0 cleavage site. In contrast, the cleavage motifs of LPAI viruses typically have only two basic amino acids, at positions ?1 and ?4 from the cleavage site for the H5 and at positions ?1 and ?3 for the H7 subtype (2). This difference appears to have a direct influence on viral virulence as LPAI viruses are limited to cleavage by host proteases such as trypsin-like enzymes and are thus restricted to replication at sites in the host where such enzymes are found, i.e., the respiratory and intestinal tracts. Whereas the multiple basic amino acids at the HA0 cleavage sites of HPAI viruses, either as a result of insertion or substitution (2?4), allows the HA0 precursor to be cleavable by ubiquitous host proteases (5). As a result the HPAI viruses are able to replicate systemically, damaging vital organs and tissues, which results in severe disease and death (1).
Viruses of the H5 or H7 subtype isolated from free-living birds are almost invariably of low pathogenicity for poultry. With the exception of a large die-off of terns in South Africa in 1961 (6), from which A/tern/South Africa/61 (H5N3) was isolated, HPAI virus isolations from free-living birds have been associated with contact with infected poultry, usually as a result of surveillance of birds trapped or found dead on infected poultry farms. In addition, results of phylogenetic studies of H7 subtype viruses indicate that HPAI viruses do not constitute a separate phylogenetic lineage or lineages but appear to arise from low pathogenic strains (7?9). This finding is supported by the in vitro selection of mutants virulent for chickens from an avirulent H7 virus (10) and the emergence of HPAI virus from an LPAI virus isolated from swans after repeated passage in chickens (11).
These findings conform to the theories of the molecular basis for the mutation of avian influenza subtype H5 and H7 viruses from low to high virulence in poultry put forward by Garcia et al. (12) and Perdue et al. (13). Essentially they propose that spontaneous duplication of purine triplets results in the insertion of basic amino acids at the HA0 cleavage site and that this occurs due to a transcription fault by the polymerase complex. As pointed out by Perdue et al. (13) this mechanism is clearly not the only means by which HPAI viruses arise, as some appear to result from nucleotide substitution rather than insertion, while others have insertions without repeating nucleotides. Attempts to assess the minimum requirements to confer virulence for H7 avian influenza viruses have used site-directed mutagenesis of the cleavage site of cloned HA followed by in vitro expression to examine the effect that progressive amino acid changes at each position of the cleavage site would have upon cleavability (3,14,15). For some H7 influenza viruses, the tetrapeptide motif R-X-R/K-R at the cleavage site leading up to the cleavage point is sufficient for cleavage by furin-like proteases, and variants with cleavage sites that do not conform to this motif are not pathogenic for chickens (3). However, motifs that differ from this standard have been observed from natural outbreaks of H7 HPAI (16).
Between 1959 and the end of 2001, a total of 18 primary outbreaks (10 H7 and 8 H5) of HPAI in poultry were reported (17). However, the geographic distribution of these outbreaks was not uniform across the world (five were in the British Isles; five in Australia; three in areas in Europe other than British Isles; and one each in Pakistan, Hong Kong, Canada, United States, and Mexico). Until 2002, no influenza virus had been isolated in poultry or wild birds in the continent of South America. In May 2002, a LPAI virus of H7N3 subtype was isolated from a broiler breeder flock in Chile, and in June, an HPAI virus of the same subtype was obtained from the same flock (18). We describe the characterization of the LPAI and HPAI isolates obtained in Chile with particular reference to the HPAI virus, which, while having a 10?amino acid (aa) insert at the HA0 cleavage site, does not conform to the dogma that a -R-X-R/K-R*G-L-F- motif is a prerequisite for HPAI viruses. We also present evidence that the 10-aa insert present in the HPAI viruses is the result of recombination between the HA and the nucleoprotein genes.
http://www.cdc.gov/ncidod/EID/vol10no4/03-0396.htm