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      Pathogenicity of Genetically Similar, H5N1 Highly Pathogenic Avian Influenza Virus Strains in Chicken and the Differences in Sensitivity among Different Chicken Breeds

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          Abstract

          Differences in the pathogenicity of genetically closely related H5N1 highly pathogenic avian influenza viruses (HPAIVs) were evaluated in White Leghorn chickens. These viruses varied in the clinical symptoms they induced, including lethality, virus shedding, and replication in host tissues. A comparison of the host responses in the lung, brain, and spleen suggested that the differences in viral replication efficiency were related to the host cytokine response at the early phase of infection, especially variations in the proinflammatory cytokine IL-6. Based on these findings, we inoculated the virus that showed the mildest pathogenicity among the five tested, A/pigeon/Thailand/VSMU-7-NPT/2004, into four breeds of Thai indigenous chicken, Phadu-Hung-Dang (PHD), Chee, Dang, and Luang-Hung-Khao (LHK), to explore effects of genetic background on host response. Among these breeds, Chee, Dang, and LHK showed significantly longer survival times than White Leghorns. Virus shedding from dead Thai indigenous chickens was significantly lower than that from White Leghorns. Although polymorphisms were observed in the Mx and MHC class I genes, there was no significant association between the polymorphisms in these loci and resistance to HPAIV.

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          Characterization of an avian influenza A (H5N1) virus isolated from a child with a fatal respiratory illness.

          An avian H5N1 influenza A virus (A/Hong Kong/156/97) was isolated from a tracheal aspirate obtained from a 3-year-old child in Hong Kong with a fatal illness consistent with influenza. Serologic analysis indicated the presence of an H5 hemagglutinin. All eight RNA segments were derived from an avian influenza A virus. The hemagglutinin contained multiple basic amino acids adjacent to the cleavage site, a feature characteristic of highly pathogenic avian influenza A viruses. The virus caused 87.5 to 100 percent mortality in experimentally inoculated White Plymouth Rock and White Leghorn chickens. These results may have implications for global influenza surveillance and planning for pandemic influenza.
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            Influenza A viruses: new research developments.

            Influenza A viruses are zoonotic pathogens that continuously circulate and change in several animal hosts, including birds, pigs, horses and humans. The emergence of novel virus strains that are capable of causing human epidemics or pandemics is a serious possibility. Here, we discuss the value of surveillance and characterization of naturally occurring influenza viruses, and review the impact that new developments in the laboratory have had on our understanding of the host tropism and virulence of viruses. We also revise the lessons that have been learnt from the pandemic viruses of the past 100 years.
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              A single-amino-acid substitution in the NS1 protein changes the pathogenicity of H5N1 avian influenza viruses in mice.

              In this study, we explored the molecular basis determining the virulence of H5N1 avian influenza viruses in mammalian hosts by comparing two viruses, A/Duck/Guangxi/12/03 (DK/12) and A/Duck/Guangxi/27/03 (DK/27), which are genetically similar but differ in their pathogenicities in mice. To assess the genetic basis for this difference in virulence, we used reverse genetics to generate a series of reassortants and mutants of these two viruses. We found that a single-amino-acid substitution of serine for proline at position 42 (P42S) in the NS1 protein dramatically increased the virulence of the DK/12 virus in mice, whereas the substitution of proline for serine at the same position (S42P) completely attenuated the DK/27 virus. We further demonstrated that the amino acid S42 of NS1 is critical for the H5N1 influenza virus to antagonize host cell interferon induction and for the NS1 protein to prevent the double-stranded RNA-mediated activation of the NF-kappaB pathway and the IRF-3 pathway. Our results indicate that the NS1 protein is critical for the pathogenicity of H5N1 influenza viruses in mammalian hosts and that the amino acid S42 of NS1 plays a key role in undermining the antiviral immune response of the host cell.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                14 April 2016
                2016
                : 11
                : 4
                : e0153649
                Affiliations
                [1 ]Thailand-Japan Zoonotic Diseases Collaborating Center (ZDCC), Kasetlang, Chatuchak, Bangkok, Thailand
                [2 ]Research Team for Zoonotic Diseases, National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Kannondai, Tsukuba, Ibaraki, Japan
                [3 ]Transboundary Animal Diseases Research Center, Joint Faculty of Veterinary, Kagoshima University, Korimoto, Kagoshima, Japan
                [4 ]Department of Molecular Life Science, Tokai University School of Medicine, Shimokasuya Isehara, Kanagawa, Japan
                [5 ]The Monitoring and Surveillance Center for Zoonotic Diseases in Wildlife and Exotic Animals, Faculty of Veterinary Science, Mahidol University, Salaya, Phuttamonthon, Nakhon Pathom, Thailand
                [6 ]National Institute of Animal Health, Kasetklang, Chatuchak, Bangkok, Thailand
                [7 ]Influenza and Prion Disease Research Center, National Institute of Animal Health, National Agriculture and Food Research Organization (NARO), Kannnondai, Tsukuba, Ibaraki, Japan
                University of Edinburgh, UNITED KINGDOM
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                Conceived and designed the experiments: T. Saito. Performed the experiments: AM TK YH HA. Analyzed the data: AM T. Shiina SS. Contributed reagents/materials/analysis tools: TP KC PR SP. Wrote the paper: AM T. Saito.

                Article
                PONE-D-15-48378
                10.1371/journal.pone.0153649
                4841636
                27078641
                078c6e02-a7d4-48ad-9477-bd001b8b6b87
                © 2016 Matsuu et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 4 November 2015
                : 3 March 2016
                Page count
                Figures: 4, Tables: 2, Pages: 17
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/501100001700, Ministry of Education, Culture, Sports, Science, and Technology;
                Award ID: Founding Research Center for Emerging and Reemerging Infectious Diseases
                Award Recipient :
                This work was supported by a program of the Founding Research Center for Emerging and Reemerging Infectious Diseases, which was launched via a project commissioned by the Ministry of Education, Culture, Sports, Science, and Technology of Japan (Grant number: not applicable, http://www.amed.go.jp/program/list/01/06/jgrid02.html; in Japanese) to T. Saito. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Organisms
                Animals
                Vertebrates
                Amniotes
                Birds
                Fowl
                Gamefowl
                Chickens
                Biology and Life Sciences
                Organisms
                Animals
                Vertebrates
                Amniotes
                Birds
                Poultry
                Chickens
                Biology and Life Sciences
                Agriculture
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                Biology and life sciences
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                Viruses
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                Biology and life sciences
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                Medicine and Health Sciences
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                Zoonoses
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                People and Places
                Population Groupings
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                Medicine and Health Sciences
                Pathology and Laboratory Medicine
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                Molecular Biology
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                Custom metadata
                The nucleotide sequences of protein-coding regions in all eight gene segments of four viruses are available from GeneBank (Accession number: EPI568201-EPI568208, EPI568209-EPI568216, EPI568217-EPI568223, and EPI568225-EPI568232). The complete sequence data of Mx cDNA of White Leghorns and Thai indigenous chickens are available from GeneBank (Accession number: LC033614-LC033685). BF gene data of White Leghorns and Thai indigenous chickens are available from GeneBank (Accession number: LC033686-LC033783).

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