Lethality to ferrets of H5N1 influenza viruses isolated from humans and poultry in 2004 - PubMed (original) (raw)

doi: 10.1128/JVI.79.4.2191-2198.2005.

Jerold E Rehg, Scott Krauss, Hui-Ling Yen, Yi Guan, Malik Peiris, Tien D Nguyen, Thi H Hanh, Pilipan Puthavathana, Hoang T Long, Chantanee Buranathai, Wilina Lim, Robert G Webster, Erich Hoffmann

Affiliations

Lethality to ferrets of H5N1 influenza viruses isolated from humans and poultry in 2004

Elena A Govorkova et al. J Virol. 2005 Feb.

Erratum in

Abstract

The 2004 outbreaks of H5N1 influenza viruses in Vietnam and Thailand were highly lethal to humans and to poultry; therefore, newly emerging avian influenza A viruses pose a continued threat, not only to avian species but also to humans. We studied the pathogenicity of four human and nine avian H5N1/04 influenza viruses in ferrets (an excellent model for influenza studies). All four human isolates were fatal to intranasally inoculated ferrets. The human isolate A/Vietnam/1203/04 (H5N1) was the most pathogenic isolate; the severity of disease was associated with a broad tissue tropism and high virus titers in multiple organs, including the brain. High fever, weight loss, anorexia, extreme lethargy, and diarrhea were observed. Two avian H5N1/04 isolates were as pathogenic as the human viruses, causing lethal systemic infections in ferrets. Seven of nine H5N1/04 viruses isolated from avian species caused mild infections, with virus replication restricted to the upper respiratory tract. All chicken isolates were nonlethal to ferrets. A sequence analysis revealed polybasic amino acids in the hemagglutinin connecting peptides of all H5N1/04 viruses, indicating that multiple molecular differences in other genes are important for a high level of virulence. Interestingly, the human A/Vietnam/1203/04 isolate had a lysine substitution at position 627 of PB2 and had one to eight amino acid changes in all gene products except that of the M1 gene, unlike the A/chicken/Vietnam/C58/04 and A/quail/Vietnam/36/04 viruses. Our results indicate that viruses that are lethal to mammals are circulating among birds in Asia and suggest that pathogenicity in ferrets, and perhaps humans, reflects a complex combination of different residues rather than a single amino acid difference.

PubMed Disclaimer

Figures

FIG. 1.

FIG. 1.

Clinical signs of infection with H5N1/04 influenza viruses in ferrets. (A) Changes in body temperatures of ferrets infected with H5N1/04 influenza viruses. Ferrets were inoculated with 106 EID50 of the A/Vietnam/1203/04, A/quail/Vietnam/36/04, A/chicken/Vietnam/C58/04, or A/PR/8/34 (H1N1) influenza virus, and body temperatures were monitored daily by the use of subcutaneous implantable temperature transponders for 12 days postinfection. Each data point represents the mean value ± SD for two to four ferrets. (B) Changes in weights of ferrets infected with H5N1/04 influenza viruses. The weights of ferrets were measured on days 0, 2, 4, 6, 8, 10, and 12 after inoculation. The loss or gain of weight was calculated for each ferret as the percent change in the initial mean starting weight on day 0. Values are the averages ± SD for two to four ferrets for each group. The clinical signs of infection with A/Vietnam/1203/04 (C) or A/quail/Vietnam/36/04 (D) virus were noted daily. The numbers on the bars represent the numbers of ferrets showing signs/total number of ferrets. The ferrets infected with the A/quail/Vietnam/36/04 influenza virus showed clinical signs of severe infection and were sacrificed on day 7.

FIG. 2.

FIG. 2.

Replication of H5N1/04 influenza viruses in upper respiratory tracts (A) and internal organs (B) of ferrets. (A) Virus titers were determined at 3, 5, and 7 days postinfection in nasal washes of ferrets infected with 106 EID50 of the A/Vietnam/1203/04, A/quail/Vietnam/36/04, A/chicken/Vietnam/C58/04, or A/PR/8/34 (H1N1) virus. Values (log10 EID50/ml) are the mean virus titers ± SD of all live ferrets on the indicated days. The lower limit of virus detection was <1 log10 EID50 per 1.0 ml of nasal wash or tissue homogenate. For calculations of the means, influenza virus-positive samples with virus titers of <1 log10 EID50/ml were assigned a value of 0.5 log10 EID50/ml. (B) Virus titers were determined for the lungs, brains, olfactory bulbs, spleens, and intestines of ferrets at days 3 and 6 postinfection for animals infected with A/Vietnam/1203/04 and at day 3 postinfection for animals infected with A/quail/Vietnam/36/04. Values (log10 EID50/ml) are the means ± SD for two to four animals.

FIG. 3.

FIG. 3.

Histologic changes in brains, lungs, and livers of ferrets infected with H5N1/04 viruses. The images shown are hematoxylin-and-eosin-stained sections of tissue from ferrets inoculated with A/Vietnam/1203/04 (isolated from a human) (A, C, and E) and A/chicken/Vietnam/C58/04 (isolated from a chicken) (B, D, and F) obtained on day 6 after inoculation. (A**)** Hypercellular brain tissue showing infiltrate of mononuclear inflammatory cells, neuronal degeneration, neuronophagia (white arrows), and perivascular mononuclear infiltrate (black arrow). (C) Lung showing bronchiole (b) with epithelial necrosis, intraluminal debris, and inflammatory cells. Alveoli (a) are lined with proliferating hypertrophic pneumocytes (white arrows) and are filled with a mononuclear cell infiltrate. (E**)** Liver showing portal tract (P) with biliary duct necrosis and inflammatory cell infiltration. The tissue in this area is necrotic and hemorrhagic, unlike normal liver tissue (white arrow). The brain, lung, and liver (B, D, and F) of a ferret inoculated with A/chicken/Vietnam/C58/04 were normal. Magnification, ×20.

FIG. 4.

FIG. 4.

Location of amino acid changes in three-dimensional structure of HA. A schematic representation of the HA monomer shows residues that differ between the A/Vietnam/1203/04, A/chicken/Vietnam/C58/04, and A/quail/Vietnam/36/04 viruses, based on the three-dimensional structure of H5 from A/duck/Singapore/3/97 (H5N3).

Similar articles

Cited by

References

    1. Bosch, F. X., W. Garten, H. D. Klenk, and R. Rott. 1981. Proteolytic cleavage of influenza virus hemagglutinins: primary structure of the connecting peptide between HA1 and HA2 determines proteolytic cleavability and pathogenicity of avian influenza viruses. Virology 113:725-735. - PubMed
    1. Cauthen, A. N., D. E. Swayne, S. Schultz-Cherry, M. L. Perdue, and D. L. Suarez. 2000. Continued circulation in China of highly pathogenic avian influenza viruses encoding the hemagglutinin gene associated with the 1997 H5N1 outbreak in poultry and humans. J. Virol. 74:6592-6599. - PMC - PubMed
    1. Chen, H., G. Deng, Z. Li, G. Tian, Y. Li, P. Jiao, L. Zhang, Z. Liu, R. G. Webster, and K. Yu. 2004. The evolution of H5N1 influenza viruses in ducks in southern China. Proc. Natl. Acad. Sci. USA 101:10452-10457. - PMC - PubMed
    1. Cheung, C. Y., L. L. Poon, A. S. Lau, W. Luk, Y. L. Lau, K. F. Shortridge, S. Gordon, Y. Guan, and J. S. Peiris. 2002. Induction of proinflammatory cytokines in human macrophages by influenza A (H5N1) viruses: a mechanism for the unusual severity of human disease? Lancet 360:1831-1837. - PubMed
    1. Claas, E. C., J. C. de Jong, R. van Beek, G. F. Rimmelzwaan, and A. D. Osterhaus. 1998. Human influenza virus A/HongKong/156/97 (H5N1) infection. Vaccine 16:977-978. - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources