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Genetic incompatibility among influenza A viruses
International Congress Series 1219 (2001) 1019 – 1021
Genetic incompatibility among inf luenza A viruses Masato Hattaa, Peter Halfmanna, Krisna Wellsa, Yoshihiro Kawaokaa,b,* a
Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, WI 53706, USA b Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai, Minato, Tokyo 108-8639, Japan
Abstract Although influenza A viruses occasionally transmit from one species to another, host range restriction exists among these viruses. Current human influenza A viruses are thought to originate from avian viruses. However, human influenza A viruses do not replicate in duck intestine, a major replication site of avian viruses. Although previous studies identified that the HA and NA genes restrict human virus replication in ducks, the contribution of the other genes remains unknown. To determine the genetic basis for host range restriction in ducks, we first established a reverse genetics system for generating A/Memphis/8/88 (H3N2) and A/Mallard/NY/6750/78 (H2N2) viruses from cloned cDNA. Using this system, we attempted to generate reassortant viruses with various combinations of genes. However, reassortant viruses with some of the gene combinations were not generated, suggesting that there may be incompatibility between the genes of avian and human strains. D 2001 Elsevier Science B.V. All rights reserved. Keywords: Host range; Reassortant; Reverse genetics
1. Introduction Influenza A viruses infect swine, horses, seals, and a large variety of birds as well as humans. Phylogenetic studies have revealed species-specific lineages of viral genes and have demonstrated that the prevalence of interspecies transmission depends on the host animal species. Aquatic birds are thought to be the source of all influenza A viruses in *
Corresponding author. Department of Microbiology and Immunology, Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai, Minato, Tokyo 108-8639, Japan. Tel.: +81-3-5449-5310; fax: +81-35449-5408. E-mail address: [email protected] (Y. Kawaoka). 0531-5131/01/$ – see front matter D 2001 Elsevier Science B.V. All rights reserved. PII: S 0 5 3 1 - 5 1 3 1 ( 0 1 ) 0 0 4 0 0 - 9
1020
M. Hatta et al. / International Congress Series 1219 (2001) 1019–1021
other animal species. There have been three human influenza pandemics in this century. Avian influenza A viruses are thought to contribute to all of these pandemic strains [1,2]. However, avian influenza viruses replicate poorly in humans, and similarly, human viruses do not replicate efficiently in birds, demonstrating host range restriction among these viruses [3– 6]. The HA and NA genes restrict human virus replication in duck intestinal tracts. However, the contribution of the other genes remains unknown. As an initial step in determining the genetic basis for host range restriction in ducks, we attempted to generate reassortant viruses with various combinations of genes using a reverse genetics system [7,8].
2. Generation of reassortant viruses using reverse genetics We attempted to generate the human influenza A/Memphis/8/88 (H3N2) and avian influenza A/Mallard/NY/6750/78 (H2N2) viruses entirely from cloned cDNA using plasmid-driven reverse genetics [8]. To this end, the cDNAs encoding all eight segments of these viruses were cloned between the human RNA polymerase I promoter and mouse RNA polymerase I terminator. Transfectant viruses were generated as reported earlier [8]. To determine which of the genes are important for host range restriction in ducks, we next generated single- or multiple-gene reassortant viruses possessing only one or some of the genes from A/Memphis/8/88 and the rest from A/Mallard/NY/6750/78 virus. Singlegene reassortant viruses possessing the PB2 or NP gene of A/Memphis/8/88 virus and the rest from A/Mallard/NY/6750/78 virus were obtained. Other single-gene reassortant viruses were not generated. Also, multiple-gene reassortant viruses possessing the HA and NA; PB1, PB2, and PA; or PB1, PB2, PA and NP of A/Memphis/8/88 virus and the rest from A/Mallard/NY/6750/78 virus were obtained. These results indicate that there may be incompatibility between the genes of avian and human strains.
3. Discussion High-frequency reassortment is characteristic of influenza A viruses. However, genetic variation of the HA/NA subtype combinations is limited. In this study, we demonstrate that there may be some restriction on generating reassortant viruses between avian and human species because multiple-gene reassortant viruses were generated easier than single-gene reassortants. Our findings provide a molecular basis for the mechanisms of generating reassortant viruses among influenza A viruses.
Acknowledgements We thank Martha McGregor, Nicole Poznik, and Ryo Kawaoka for excellent technical assistance. Automated sequencing was performed at the University of WisconsinMadison, Biotechnology Center.
M. Hatta et al. / International Congress Series 1219 (2001) 1019–1021
1021
References [1] W.G. Laver, R.G. Webster, Studies on the origin of pandemic inf luenza: III. Evidence implicating duck and equine inf luenza viruses as possible progenitors of the Hong Kong strain of human inf luenza, Virology 51 (1973) 383 – 391. [2] O.T. Gorman, W.J. Bean, Y. Kawaoka, I. Donatelli, Y.J. Guo, R.G. Webster, Evolution of inf luenza A virus nucleoprotein genes: implications for the origins of H1N1 human and classical swine viruses, J. Virol. 65 (1991) 3704 – 3714. [3] R.G. Webster, M. Yakhno, V.S. Hinshaw, W.J. Bean, K.G. Murti, Intestinal inf luenza: replication and characterization of inf luenza viruses in ducks, Virology 84 (1978) 268 – 278. [4] V.S. Hinshaw, R.G. Webster, C.W. Naeve, B.R. Murphy, Altered tissue tropism of human – avian reassortant inf luenza viruses, Virology 128 (1983) 260 – 263. [5] A.S. Beare, R.G. Webster, Replication of avian inf luenza viruses in humans, Arch. Virol. 119 (1991) 37 – 42. [6] B.R. Murphy, A.J. Buckler-White, W.T. London, J. Harper, E.L. Tierney, N.T. Miller, L.J. Reck, R.M. Chanock, V.S. Hinshaw, Avian – human reassortant inf luenza A viruses derived by mating avian and human inf luenza A viruses, J. Infect. Dis. 150 (1984) 841 – 850. [7] E. Fodor, L. Devenish, O.G. Engelhardt, P. Palese, G.G. Brownlee, A. Garcia-Sastre, Rescue of inf luenza A virus from recombinant DNA, J. Virol. 73 (1999) 9679 – 9682. [8] G. Neumann, T. Watanabe, H. Ito, S. Watanabe, H. Goto, P. Gao, M. Hughes, D.R. Perez, R. Donis, E. Hoffmann, G. Hobom, Y. Kawaoka, Generation of inf luenza A viruses entirely from cloned cDNAs, Proc. Natl. Acad. Sci. U. S. A. 96 (1999) 9345 – 9350.
Genetic incompatibility among inf luenza A viruses Masato Hattaa, Peter Halfmanna, Krisna Wellsa, Yoshihiro Kawaokaa,b,* a
Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, Madison, WI 53706, USA b Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai, Minato, Tokyo 108-8639, Japan
Abstract Although influenza A viruses occasionally transmit from one species to another, host range restriction exists among these viruses. Current human influenza A viruses are thought to originate from avian viruses. However, human influenza A viruses do not replicate in duck intestine, a major replication site of avian viruses. Although previous studies identified that the HA and NA genes restrict human virus replication in ducks, the contribution of the other genes remains unknown. To determine the genetic basis for host range restriction in ducks, we first established a reverse genetics system for generating A/Memphis/8/88 (H3N2) and A/Mallard/NY/6750/78 (H2N2) viruses from cloned cDNA. Using this system, we attempted to generate reassortant viruses with various combinations of genes. However, reassortant viruses with some of the gene combinations were not generated, suggesting that there may be incompatibility between the genes of avian and human strains. D 2001 Elsevier Science B.V. All rights reserved. Keywords: Host range; Reassortant; Reverse genetics
1. Introduction Influenza A viruses infect swine, horses, seals, and a large variety of birds as well as humans. Phylogenetic studies have revealed species-specific lineages of viral genes and have demonstrated that the prevalence of interspecies transmission depends on the host animal species. Aquatic birds are thought to be the source of all influenza A viruses in *
Corresponding author. Department of Microbiology and Immunology, Institute of Medical Science, University of Tokyo, 4-6-1, Shirokanedai, Minato, Tokyo 108-8639, Japan. Tel.: +81-3-5449-5310; fax: +81-35449-5408. E-mail address: [email protected] (Y. Kawaoka). 0531-5131/01/$ – see front matter D 2001 Elsevier Science B.V. All rights reserved. PII: S 0 5 3 1 - 5 1 3 1 ( 0 1 ) 0 0 4 0 0 - 9
1020
M. Hatta et al. / International Congress Series 1219 (2001) 1019–1021
other animal species. There have been three human influenza pandemics in this century. Avian influenza A viruses are thought to contribute to all of these pandemic strains [1,2]. However, avian influenza viruses replicate poorly in humans, and similarly, human viruses do not replicate efficiently in birds, demonstrating host range restriction among these viruses [3– 6]. The HA and NA genes restrict human virus replication in duck intestinal tracts. However, the contribution of the other genes remains unknown. As an initial step in determining the genetic basis for host range restriction in ducks, we attempted to generate reassortant viruses with various combinations of genes using a reverse genetics system [7,8].
2. Generation of reassortant viruses using reverse genetics We attempted to generate the human influenza A/Memphis/8/88 (H3N2) and avian influenza A/Mallard/NY/6750/78 (H2N2) viruses entirely from cloned cDNA using plasmid-driven reverse genetics [8]. To this end, the cDNAs encoding all eight segments of these viruses were cloned between the human RNA polymerase I promoter and mouse RNA polymerase I terminator. Transfectant viruses were generated as reported earlier [8]. To determine which of the genes are important for host range restriction in ducks, we next generated single- or multiple-gene reassortant viruses possessing only one or some of the genes from A/Memphis/8/88 and the rest from A/Mallard/NY/6750/78 virus. Singlegene reassortant viruses possessing the PB2 or NP gene of A/Memphis/8/88 virus and the rest from A/Mallard/NY/6750/78 virus were obtained. Other single-gene reassortant viruses were not generated. Also, multiple-gene reassortant viruses possessing the HA and NA; PB1, PB2, and PA; or PB1, PB2, PA and NP of A/Memphis/8/88 virus and the rest from A/Mallard/NY/6750/78 virus were obtained. These results indicate that there may be incompatibility between the genes of avian and human strains.
3. Discussion High-frequency reassortment is characteristic of influenza A viruses. However, genetic variation of the HA/NA subtype combinations is limited. In this study, we demonstrate that there may be some restriction on generating reassortant viruses between avian and human species because multiple-gene reassortant viruses were generated easier than single-gene reassortants. Our findings provide a molecular basis for the mechanisms of generating reassortant viruses among influenza A viruses.
Acknowledgements We thank Martha McGregor, Nicole Poznik, and Ryo Kawaoka for excellent technical assistance. Automated sequencing was performed at the University of WisconsinMadison, Biotechnology Center.
M. Hatta et al. / International Congress Series 1219 (2001) 1019–1021
1021
References [1] W.G. Laver, R.G. Webster, Studies on the origin of pandemic inf luenza: III. Evidence implicating duck and equine inf luenza viruses as possible progenitors of the Hong Kong strain of human inf luenza, Virology 51 (1973) 383 – 391. [2] O.T. Gorman, W.J. Bean, Y. Kawaoka, I. Donatelli, Y.J. Guo, R.G. Webster, Evolution of inf luenza A virus nucleoprotein genes: implications for the origins of H1N1 human and classical swine viruses, J. Virol. 65 (1991) 3704 – 3714. [3] R.G. Webster, M. Yakhno, V.S. Hinshaw, W.J. Bean, K.G. Murti, Intestinal inf luenza: replication and characterization of inf luenza viruses in ducks, Virology 84 (1978) 268 – 278. [4] V.S. Hinshaw, R.G. Webster, C.W. Naeve, B.R. Murphy, Altered tissue tropism of human – avian reassortant inf luenza viruses, Virology 128 (1983) 260 – 263. [5] A.S. Beare, R.G. Webster, Replication of avian inf luenza viruses in humans, Arch. Virol. 119 (1991) 37 – 42. [6] B.R. Murphy, A.J. Buckler-White, W.T. London, J. Harper, E.L. Tierney, N.T. Miller, L.J. Reck, R.M. Chanock, V.S. Hinshaw, Avian – human reassortant inf luenza A viruses derived by mating avian and human inf luenza A viruses, J. Infect. Dis. 150 (1984) 841 – 850. [7] E. Fodor, L. Devenish, O.G. Engelhardt, P. Palese, G.G. Brownlee, A. Garcia-Sastre, Rescue of inf luenza A virus from recombinant DNA, J. Virol. 73 (1999) 9679 – 9682. [8] G. Neumann, T. Watanabe, H. Ito, S. Watanabe, H. Goto, P. Gao, M. Hughes, D.R. Perez, R. Donis, E. Hoffmann, G. Hobom, Y. Kawaoka, Generation of inf luenza A viruses entirely from cloned cDNAs, Proc. Natl. Acad. Sci. U. S. A. 96 (1999) 9345 – 9350.