WorldWideScience

Sample records for influenza virus h5

  1. Sensitization with vaccinia virus encoding H5N1 hemagglutinin restores immune potential against H5N1 influenza virus.

    Science.gov (United States)

    Yasui, Fumihiko; Itoh, Yasushi; Ikejiri, Ai; Kitabatake, Masahiro; Sakaguchi, Nobuo; Munekata, Keisuke; Shichinohe, Shintaro; Hayashi, Yukiko; Ishigaki, Hirohito; Nakayama, Misako; Sakoda, Yoshihiro; Kida, Hiroshi; Ogasawara, Kazumasa; Kohara, Michinori

    2016-11-28

    H5N1 highly pathogenic avian influenza (H5N1 HPAI) virus causes elevated mortality compared with seasonal influenza viruses like H1N1 pandemic influenza (H1N1 pdm) virus. We identified a mechanism associated with the severe symptoms seen with H5N1 HPAI virus infection. H5N1 HPAI virus infection induced a decrease of dendritic cell number in the splenic extrafollicular T-cell zone and impaired formation of the outer layers of B-cell follicles, resulting in insufficient levels of antibody production after infection. However, in animals vaccinated with a live recombinant vaccinia virus expressing the H5 hemagglutinin, infection with H5N1 HPAI virus induced parafollicular dendritic cell accumulation and efficient antibody production. These results indicate that a recombinant vaccinia encoding H5 hemagglutinin gene does not impair dendritic cell recruitment and can be a useful vaccine candidate.

  2. Newcastle disease virus-based H5 influenza vaccine protects chickens from lethal challenge with a highly pathogenic H5N2 avian influenza virus

    OpenAIRE

    Ma, Jingjiao; Lee, Jinhwa; Liu, Haixia; Mena, Ignacio; Davis, A. Sally; Sunwoo, Sun Young; Lang, Yuekun; Duff, Michael; Morozov, Igor; Li, Yuhao; Yang, Jianmei; García-Sastre, Adolfo; Richt, Juergen A.; Ma, Wenjun

    2017-01-01

    Since December 2014, Eurasian-origin, highly pathogenic avian influenza H5 viruses including H5N1, H5N2, and H5N8 subtypes (called H5Nx viruses), which belong to the H5 clade 2.3.4.4, have been detected in U.S. wild birds. Subsequently, highly pathogenic H5N2 and H5N8 viruses have caused outbreaks in U.S. domestic poultry. Vaccination is one of the most effective ways to control influenza outbreaks and protect animal and public health. Newcastle disease virus (NDV)-based influenza vaccines ha...

  3. Rapid detection of the avian influenza virus H5N1 subtype in Egypt

    African Journals Online (AJOL)

    Dr

    highly pathogenic avian influenza virus subtype H5N1 in Egypt is threatening poultry and ... Key words: Avian influenza virus, H5N1, fluorescent antibody enzyme-linked immunosorbent assay (ELISA) ..... poultry and is potentially zoonotic.

  4. Avian Influenza Virus (H5N1): a Threat to Human Health

    OpenAIRE

    Peiris, J. S. Malik; de Jong, Menno D.; Guan, Yi

    2007-01-01

    Pandemic influenza virus has its origins in avian influenza viruses. The highly pathogenic avian influenza virus subtype H5N1 is already panzootic in poultry, with attendant economic consequences. It continues to cross species barriers to infect humans and other mammals, often with fatal outcomes. Therefore, H5N1 virus has rightly received attention as a potential pandemic threat. However, it is noted that the pandemics of 1957 and 1968 did not arise from highly pathogenic influenza viruses, ...

  5. Avian Influenza Virus A (H5N1), Detected through Routine Surveillance, in Child, Bangladesh

    Science.gov (United States)

    Alamgir, A.S.M.; Sultana, Rebecca; Islam, M. Saiful; Rahman, Mustafizur; Fry, Alicia M.; Shu, Bo; Lindstrom, Stephen; Nahar, Kamrun; Goswami, Doli; Haider, M. Sabbir; Nahar, Sharifun; Butler, Ebonee; Hancock, Kathy; Donis, Ruben O.; Davis, Charles T.; Zaman, Rashid Uz; Luby, Stephen P.; Uyeki, Timothy M.; Rahman, Mahmudur

    2009-01-01

    We identified avian influenza virus A (H5N1) infection in a child in Bangladesh in 2008 by routine influenza surveillance. The virus was of the same clade and phylogenetic subgroup as that circulating among poultry during the period. This case illustrates the value of routine surveillance for detection of novel influenza virus. PMID:19751601

  6. Avian influenza virus (H5N1): a threat to human health

    NARCIS (Netherlands)

    Peiris, J. S. Malik; de Jong, Menno D.; Guan, Yi

    2007-01-01

    Pandemic influenza virus has its origins in avian influenza viruses. The highly pathogenic avian influenza virus subtype H5N1 is already panzootic in poultry, with attendant economic consequences. It continues to cross species barriers to infect humans and other mammals, often with fatal outcomes.

  7. Novel Eurasian highly pathogenic influenza A H5 viruses in wild birds, Washington, USA

    Science.gov (United States)

    Ip, Hon S.; Kim Torchetti, Mia; Crespo, Rocio; Kohrs, Paul; DeBruyn, Paul; Mansfield, Kristin G.; Baszler, Timothy; Badcoe, Lyndon; Bodenstein, Barbara L.; Shearn-Bochsler, Valerie I.; Killian, Mary Lea; Pederson, Janice C.; Hines, Nichole; Gidlewski, Thomas; DeLiberto, Thomas; Sleeman, Jonathan M.

    2015-01-01

    Novel Eurasian lineage avian influenza A(H5N8) virus has spread rapidly and globally since January 2014. In December 2014, H5N8 and reassortant H5N2 viruses were detected in wild birds in Washington, USA, and subsequently in backyard birds. When they infect commercial poultry, these highly pathogenic viruses pose substantial trade issues.

  8. Novel Eurasian highly pathogenic avian influenza A H5 viruses in wild birds, Washington, USA, 2014.

    Science.gov (United States)

    Ip, Hon S; Torchetti, Mia Kim; Crespo, Rocio; Kohrs, Paul; DeBruyn, Paul; Mansfield, Kristin G; Baszler, Timothy; Badcoe, Lyndon; Bodenstein, Barbara; Shearn-Bochsler, Valerie; Killian, Mary Lea; Pedersen, Janice C; Hines, Nichole; Gidlewski, Thomas; DeLiberto, Thomas; Sleeman, Jonathan M

    2015-05-01

    Novel Eurasian lineage avian influenza A(H5N8) virus has spread rapidly and globally since January 2014. In December 2014, H5N8 and reassortant H5N2 viruses were detected in wild birds in Washington, USA, and subsequently in backyard birds. When they infect commercial poultry, these highly pathogenic viruses pose substantial trade issues.

  9. Rapid detection of the avian influenza virus H5N1 subtype in Egypt ...

    African Journals Online (AJOL)

    The unprecedented spread of highly pathogenic avian influenza virus subtype H5N1 in Egypt ... Effective diagnosis and control management are needed to control the disease. ... Reconstituted clinical samples consisting of H5 AIVs mixed with ...

  10. Establishment of the cross-clade antigen detection system for H5 subtype influenza viruses using peptide monoclonal antibodies specific for influenza virus H5 hemagglutinin.

    Science.gov (United States)

    Takahashi, Hitoshi; Nagata, Shiho; Odagiri, Takato; Kageyama, Tsutomu

    2018-04-15

    The H5 subtype of highly pathogenic avian influenza (H5 HPAI) viruses is a threat to both animal and human public health and has the potential to cause a serious future pandemic in humans. Thus, specific and rapid detection of H5 HPAI viruses is required for infection control in humans. To develop a simple and rapid diagnostic system to detect H5 HPAI viruses with high specificity and sensitivity, we attempted to prepare monoclonal antibodies (mAbs) that specifically recognize linear epitopes in hemagglutinin (HA) of H5 subtype viruses. Nine mAb clones were obtained from mice immunized with a synthetic partial peptide of H5 HA molecules conserved among various H5 HPAI viruses. The antigen-capture enzyme-linked immunosorbent assay using the most suitable combination of these mAbs, which bound specifically to lysed H5 HA under an optimized detergent condition, was specific for H5 viruses and could broadly detect H5 viruses in multiple different clades. Taken together, these peptide mAbs, which recognize linear epitopes in a highly conserved region of H5 HA, may be useful for specific and highly sensitive detection of H5 HPAI viruses and can help in the rapid diagnosis of human, avian, and animal H5 virus infections. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Newcastle disease virus-based H5 influenza vaccine protects chickens from lethal challenge with a highly pathogenic H5N2 avian influenza virus.

    Science.gov (United States)

    Ma, Jingjiao; Lee, Jinhwa; Liu, Haixia; Mena, Ignacio; Davis, A Sally; Sunwoo, Sun Young; Lang, Yuekun; Duff, Michael; Morozov, Igor; Li, Yuhao; Yang, Jianmei; García-Sastre, Adolfo; Richt, Juergen A; Ma, Wenjun

    2017-01-01

    Since December 2014, Eurasian-origin, highly pathogenic avian influenza H5 viruses including H5N1, H5N2, and H5N8 subtypes (called H5N x viruses), which belong to the H5 clade 2.3.4.4, have been detected in U.S. wild birds. Subsequently, highly pathogenic H5N2 and H5N8 viruses have caused outbreaks in U.S. domestic poultry. Vaccination is one of the most effective ways to control influenza outbreaks and protect animal and public health. Newcastle disease virus (NDV)-based influenza vaccines have been demonstrated to be efficacious and safe in poultry. Herein, we developed an NDV-based H5 vaccine (NDV-H5) that expresses a codon-optimized ectodomain of the hemagglutinin from the A/chicken/Iowa/04-20/2015 (H5N2) virus and evaluated its efficacy in chickens. Results showed that both live and inactivated NDV-H5 vaccines induced hemagglutinin inhibition antibody titers against the H5N2 virus in immunized chickens after prime and booster, and both NDV-H5 vaccines completely protected chickens from lethal challenge with the highly pathogenic H5N2 A/turkey/Minnesota/9845-4/2015 virus. No clinical signs and only minimal virus shedding was observed in both vaccinated groups. In contrast, all mock-vaccinated, H5N2-infected chickens shed virus and died within 5 days post challenge. Furthermore, one dose of the live NDV-H5 vaccine also provided protection of 90% chickens immunized by coarse spraying; after exposure to H5N2 challenge, sera from vaccinated surviving chickens neutralized both highly pathogenic H5N1 and H5N8 viruses. Taken together, our results suggest that the NDV-based H5 vaccine is able to protect chickens against intercontinental highly pathogenic H5N x viruses and can be used by mass application to protect the poultry industry.

  12. Recombinant Parainfluenza Virus 5 Expressing Hemagglutinin of Influenza A Virus H5N1 Protected Mice against Lethal Highly Pathogenic Avian Influenza Virus H5N1 Challenge

    Science.gov (United States)

    Li, Zhuo; Mooney, Alaina J.; Gabbard, Jon D.; Gao, Xiudan; Xu, Pei; Place, Ryan J.; Hogan, Robert J.; Tompkins, S. Mark

    2013-01-01

    A safe and effective vaccine is the best way to prevent large-scale highly pathogenic avian influenza virus (HPAI) H5N1 outbreaks in the human population. The current FDA-approved H5N1 vaccine has serious limitations. A more efficacious H5N1 vaccine is urgently needed. Parainfluenza virus 5 (PIV5), a paramyxovirus, is not known to cause any illness in humans. PIV5 is an attractive vaccine vector. In our studies, a single dose of a live recombinant PIV5 expressing a hemagglutinin (HA) gene of H5N1 (rPIV5-H5) from the H5N1 subtype provided sterilizing immunity against lethal doses of HPAI H5N1 infection in mice. Furthermore, we have examined the effect of insertion of H5N1 HA at different locations within the PIV5 genome on the efficacy of a PIV5-based vaccine. Interestingly, insertion of H5N1 HA between the leader sequence, the de facto promoter of PIV5, and the first viral gene, nucleoprotein (NP), did not lead to a viable virus. Insertion of H5N1 HA between NP and the next gene, V/phosphorprotein (V/P), led to a virus that was defective in growth. We have found that insertion of H5N1 HA at the junction between the small hydrophobic (SH) gene and the hemagglutinin-neuraminidase (HN) gene gave the best immunity against HPAI H5N1 challenge: a dose as low as 1,000 PFU was sufficient to protect against lethal HPAI H5N1 challenge in mice. The work suggests that recombinant PIV5 expressing H5N1 HA has great potential as an HPAI H5N1 vaccine. PMID:23077314

  13. Subclinical avian influenza A(H5N1) virus infection in human, Vietnam

    NARCIS (Netherlands)

    Le, Mai Quynh; Horby, Peter; Fox, Annette; Nguyen, Hien Tran; Le Nguyen, Hang Khanh; Hoang, Phuong Mai Vu; Nguyen, Khanh Cong; de Jong, Menno D.; Jeeninga, Rienk E.; Rogier van Doorn, H.; Farrar, Jeremy; Wertheim, Heiman F. L.

    2013-01-01

    Laboratory-confirmed cases of subclinical infection with avian influenza A(H5N1) virus in humans are rare, and the true number of these cases is unknown. We describe the identification of a laboratory-confirmed subclinical case in a woman during an influenza A(H5N1) contact investigation in northern

  14. Inefficient Transmission of H5N1 Influenza Viruses in a Ferret Contact Model▿

    OpenAIRE

    Yen, Hui-Ling; Lipatov, Aleksandr S.; Ilyushina, Natalia A.; Govorkova, Elena A.; Franks, John; Yilmaz, Neziha; Douglas, Alan; Hay, Alan; Krauss, Scott; Rehg, Jerold E.; Hoffmann, Erich; Webster, Robert G.

    2007-01-01

    The abilities to infect and transmit efficiently among humans are essential for a novel influenza A virus to cause a pandemic. To evaluate the pandemic potential of widely disseminated H5N1 influenza viruses, a ferret contact model using experimental groups comprised of one inoculated ferret and two contact ferrets was used to study the transmissibility of four human H5N1 viruses isolated from 2003 to 2006. The effects of viral pathogenicity and receptor binding specificity (affinity to synth...

  15. Distinct Pathogenesis of Hong Kong-Origin H5N1 Viruses in Mice Compared to That of Other Highly Pathogenic H5 Avian Influenza Viruses

    OpenAIRE

    Dybing, Jody K.; Schultz-Cherry, Stacey; Swayne, David E.; Suarez, David L.; Perdue, Michael L.

    2000-01-01

    In 1997, an outbreak of virulent H5N1 avian influenza virus occurred in poultry in Hong Kong (HK) and was linked to a direct transmission to humans. The factors associated with transmission of avian influenza virus to mammals are not fully understood, and the potential risk of other highly virulent avian influenza A viruses infecting and causing disease in mammals is not known. In this study, two avian and one human HK-origin H5N1 virus along with four additional highly pathogenic H5 avian in...

  16. Highly pathogenic avian influenza virus (H5N1) in experimentally infected adult mute swans.

    Science.gov (United States)

    Kalthoff, Donata; Breithaupt, Angele; Teifke, Jens P; Globig, Anja; Harder, Timm; Mettenleiter, Thomas C; Beer, Martin

    2008-08-01

    Adult, healthy mute swans were experimentally infected with highly pathogenic avian influenza virus A/Cygnus cygnus/Germany/R65/2006 subtype H5N1. Immunologically naive birds died, whereas animals with preexisting, naturally acquired avian influenza virus-specific antibodies became infected asymptomatically and shed virus. Adult mute swans are highly susceptible, excrete virus, and can be clinically protected by preexposure immunity.

  17. Pathogenicity and Transmission of H5 and H7 Highly Pathogenic Avian Influenza Viruses in Mallards

    Science.gov (United States)

    Costa-Hurtado, Mar; Shepherd, Eric; DeJesus, Eric; Smith, Diane; Spackman, Erica; Kapczynski, Darrell R.; Suarez, David L.; Stallknecht, David E.; Swayne, David E.

    2016-01-01

    ABSTRACT Wild aquatic birds have been associated with the intercontinental spread of H5 subtype highly pathogenic avian influenza (HPAI) viruses of the A/goose/Guangdong/1/96 (Gs/GD) lineage during 2005, 2010, and 2014, but dispersion by wild waterfowl has not been implicated with spread of other HPAI viruses. To better understand why Gs/GD H5 HPAI viruses infect and transmit more efficiently in waterfowl than other HPAI viruses, groups of mallard ducks were challenged with one of 14 different H5 and H7 HPAI viruses, including a Gs/GD lineage H5N1 (clade 2.2) virus from Mongolia, part of the 2005 dispersion, and the H5N8 and H5N2 index HPAI viruses (clade 2.3.4.4) from the United States, part of the 2014 dispersion. All virus-inoculated ducks and contact exposed ducks became infected and shed moderate to high titers of the viruses, with the exception that mallards were resistant to Ck/Pennsylvania/83 and Ck/Queretaro/95 H5N2 HPAI virus infection. Clinical signs were only observed in ducks challenged with the H5N1 2005 virus, which all died, and with the H5N8 and H5N2 2014 viruses, which had decreased weight gain and fever. These three viruses were also shed in higher titers by the ducks, which could facilitate virus transmission and spread. This study highlights the possible role of wild waterfowl in the spread of HPAI viruses. IMPORTANCE The spread of H5 subtype highly pathogenic avian influenza (HPAI) viruses of the Gs/GD lineage by migratory waterfowl is a serious concern for animal and public health. H5 and H7 HPAI viruses are considered to be adapted to gallinaceous species (chickens, turkeys, quail, etc.) and less likely to infect and transmit in wild ducks. In order to understand why this is different with certain Gs/GD lineage H5 HPAI viruses, we compared the pathogenicity and transmission of several H5 and H7 HPAI viruses from previous poultry outbreaks to Gs/GD lineage H5 viruses, including H5N1 (clade 2.2), H5N8 and H5N2 (clade 2.3.4.4) viruses, in

  18. Avian influenza A virus subtype H5N2 in a red-lored Amazon parrot.

    Science.gov (United States)

    Hawkins, Michelle G; Crossley, Beate M; Osofsky, Anna; Webby, Richard J; Lee, Chang-Won; Suarez, David L; Hietala, Sharon K

    2006-01-15

    A 3-month-old red-lored Amazon parrot (Amazona autumnalis autumnalis) was evaluated for severe lethargy. Avian influenza virus hemagglutinin subtype H5N2 with low pathogenicity was characterized by virus isolation, real-time reverse transcriptase PCR assay, chicken intravenous pathogenicity index, and reference sera. The virus was also determined to be closely related to a virus lineage that had been reported only in Mexico and Central America. The chick was admitted to the hospital and placed in quarantine. Supportive care treatment was administered. Although detection of H5 avian influenza virus in birds in the United States typically results in euthanasia of infected birds, an alternative strategy with strict quarantine measures and repeated diagnostic testing was used. The chick recovered from the initial clinical signs after 4 days and was released from quarantine 9 weeks after initial evaluation after 2 consecutive negative virus isolation and real-time reverse transcriptase PCR assay results. To the authors' knowledge, this is the first report of H5N2 avian influenza A virus isolated from a psittacine bird and represents the first introduction of this virus into the United States, most likely by illegal importation of psittacine birds. Avian influenza A virus should be considered as a differential diagnosis for clinical signs of gastrointestinal tract disease in psittacine birds, especially in birds with an unknown history of origin. Although infection with avian influenza virus subtype H5 is reportable, destruction of birds is not always required.

  19. Reduced experimental infectivity and transmissibility of intercontinental H5 (H5N8 and H5N2) compared to Eurasian H5N1 highly pathogenic avian influenza viruses for chickens, turkeys, and Japanese quail

    Science.gov (United States)

    H5N1 high pathogenicity avian influenza (HPAI) virus (HPAIV) emerged in 1996 in Guangdong China and has since spread to infect and cause deaths in wild birds, poultry and humans in over 63 countries in Asia, Europe and Africa; and more recently a reassortant H5N8 clade 2.3.4.4 HPAI virus has spread ...

  20. H5N1 avian influenza virus: human cases reported in southern China.

    NARCIS (Netherlands)

    Crofts, J.; Paget, J.; Karcher, F.

    2003-01-01

    Two cases of confirmed influenza due to the avian influenza A H5N1 virus were reported last week in Hong Kong (1). The cases occurred in a Hong Kong family who had recently visited Fujian province in southern China. The daughter, aged 8 years, died following a respiratory illness. The cause of her

  1. Prevalence of Avian Origin H5 and H7 Influenza Virus Antibodies in ...

    African Journals Online (AJOL)

    As part of ongoing influenza surveillance efforts in livestock and companion animals in Nigeria, a study was conducted to investigate the prevalence of avian H5 and H7 influenza virus antibodies in exotic and Nigerian village dogs in Ibadan and Sagamu, two cities in Oyo and Ogun states respectively. One hundred and ...

  2. Molecular Characterizations of Surface Proteins Hemagglutinin and Neuraminidase from Recent H5Nx Avian Influenza Viruses

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Hua; Carney, Paul J.; Mishin, Vasiliy P.; Guo, Zhu; Chang, Jessie C.; Wentworth, David E.; Gubareva, Larisa V.; Stevens, James; Schultz-Cherry, S.

    2016-04-06

    ABSTRACT

    During 2014, a subclade 2.3.4.4 highly pathogenic avian influenza (HPAI) A(H5N8) virus caused poultry outbreaks around the world. In late 2014/early 2015, the virus was detected in wild birds in Canada and the United States, and these viruses also gave rise to reassortant progeny, composed of viral RNA segments (vRNAs) from both Eurasian and North American lineages. In particular, viruses were found with N1, N2, and N8 neuraminidase vRNAs, and these are collectively referred to as H5Nx viruses. In the United States, more than 48 million domestic birds have been affected. Here we present a detailed structural and biochemical analysis of the surface antigens of H5N1, H5N2, and H5N8 viruses in addition to those of a recent human H5N6 virus. Our results with recombinant hemagglutinin reveal that these viruses have a strict avian receptor binding preference, while recombinantly expressed neuraminidases are sensitive to FDA-approved and investigational antivirals. Although H5Nx viruses currently pose a low risk to humans, it is important to maintain surveillance of these circulating viruses and to continually assess future changes that may increase their pandemic potential.

    IMPORTANCEThe H5Nx viruses emerging in North America, Europe, and Asia pose a great public health concern. Here we report a molecular and structural study of the major surface proteins of several H5Nx influenza viruses. Our results improve the understanding of these new viruses and provide important information on their receptor preferences and susceptibilities to antivirals, which are central to pandemic risk assessment.

  3. Typing of Poultry Influenza Virus (H5 and H7 by Reverse Transcription- Polymerase Chain Reaction

    Directory of Open Access Journals (Sweden)

    Cesare Bonacina

    2010-01-01

    Full Text Available The ability of the influenza Orthomixovirus to undergo to continually antigenically changes that can affect its pathogenicity and its diffusion, explains the growing seriousness of this disease and the recent epizoozies in various parts of the world. There have been 15 HA and 9 NA type A sub-types of the influenza virus identified all of which are present in birds. Until now the very virulent avian influenza viruses identified were all included to the H5 and H7 sub-types. We here show that is possible to identify the H5 and H7 sub-types with reverse transcription-polymerase chain reaction (RT-PCR by using a set of specific primers for each HA sub-type. The RT-PCR is a quick and sensitive method of identifying the HA sub-types of the influenza virus directly from homogenised organs.

  4. Estimation of transmission parameters of H5N1 avian influenza virus in chickens.

    Directory of Open Access Journals (Sweden)

    Annemarie Bouma

    2009-01-01

    Full Text Available Despite considerable research efforts, little is yet known about key epidemiological parameters of H5N1 highly pathogenic influenza viruses in their avian hosts. Here we show how these parameters can be estimated using a limited number of birds in experimental transmission studies. Our quantitative estimates, based on Bayesian methods of inference, reveal that (i the period of latency of H5N1 influenza virus in unvaccinated chickens is short (mean: 0.24 days; 95% credible interval: 0.099-0.48 days; (ii the infectious period of H5N1 virus in unvaccinated chickens is approximately 2 days (mean: 2.1 days; 95%CI: 1.8-2.3 days; (iii the reproduction number of H5N1 virus in unvaccinated chickens need not be high (mean: 1.6; 95%CI: 0.90-2.5, although the virus is expected to spread rapidly because it has a short generation interval in unvaccinated chickens (mean: 1.3 days; 95%CI: 1.0-1.5 days; and (iv vaccination with genetically and antigenically distant H5N2 vaccines can effectively halt transmission. Simulations based on the estimated parameters indicate that herd immunity may be obtained if at least 80% of chickens in a flock are vaccinated. We discuss the implications for the control of H5N1 avian influenza virus in areas where it is endemic.

  5. Features of pathology in mice experimentally infected with highly pathogenic H5N1 influenza virus

    International Nuclear Information System (INIS)

    Ryabchikova, E. I.; Taranov, O. S.; Malkova, E. M.; Gritsyk, O. B.; Demina, O. K.

    2009-01-01

    Avian influenza became a new threat and has set people thinking about possibility of new influenza pandemic which may be caused by highly pathogenic H5N1 influenza virus. The virus could acquire ability of fast spreading between the humans and new pandemics could kill millions. Influenza virus H5N1 exhibited its deadly essence by taking out many millions of birds in nature and aviculture; other millions of chicks and ducks were killed to prevent spread of the epizootic. The strains isolated in Russia belong to Qinghai group of H5N1 influenza virus, and were imported to Russia by migratory birds. We examined time-course changes in mice blood and lungs after intranasal infection with strains A /Chicken/ Kurgan/ 05/2005, A/ Duck/ Kurgan/08/ 2005 and A/ Chicken/ Suzdalka/ Nov-11/2005 differing in virulence for this animal species. Development of leucopenia and severe damage of hemopoiesis were found in mice infected with all H5N1 influenza virus strains. Pathological changes in mice lungs during the infection with above mentioned strains, and strain-specific features have been examined. Main characteristics of lung pathology in all mice were focal nature of the alterations, severe damage of bronchial epithelium and pronounced alteration of lung vasculature. Strain A/Chicken/Suzdalka/Nov-11/2005 induced massive apoptosis of infected bronchial cells which may be a part of mechanism responsible for avirulent properties of this strain. The most interesting finding was absence of serious direct virus damage of the lung evidencing for principal role of the host humoral mechanisms in pathogenesis of H5N1 influenza in mice.(author)

  6. Molecular Evolution and Emergence of H5N6 Avian Influenza Virus in Central China.

    Science.gov (United States)

    Du, Yingying; Chen, Mingyue; Yang, Jiayun; Jia, Yane; Han, Shufang; Holmes, Edward C; Cui, Jie

    2017-06-15

    H5N6 avian influenza virus (AIV) has posed a potential threat to public health since its emergence in China in 2013. To understand the evolution and emergence of H5N6 AIV in the avian population, we performed molecular surveillance of live poultry markets (LPMs) in Wugang Prefecture, Hunan Province, in central China, during 2014 and 2015. Wugang Prefecture is located on the Eastern Asian-Australian migratory bird flyway, and a human death due to an H5N6 virus was reported in the prefecture on 21 November 2016. In total, we sampled and sequenced the complete genomes of 175 H5N6 AIVs. Notably, our analysis revealed that H5N6 AIVs contain at least six genotypes arising from segment reassortment, including a rare variant that possesses an HA gene derived from H5N1 clade 2.3.2 and a novel NP gene that has its origins with H7N3 viruses. In addition, phylogenetic analysis revealed that genetically similar H5N6 AIVs tend to cluster according to their geographic regions of origin. These results help to reveal the evolutionary behavior of influenza viruses prior to their emergence in humans. IMPORTANCE The newly emerged H5N6 influenza A virus has caused more than 10 human deaths in China since 2013. In November 2016, a human death due to an H5N6 virus, in Wugang Prefecture, Hunan Province, was confirmed by the WHO. To better understand the evolution and emergence of H5N6 viruses, we surveyed live poultry markets (LPMs) in Wugang Prefecture before the reported human death, with a focus on revealing the diversity and genomic origins of H5N6 in birds during 2014 and 2015. In general, H5N6 viruses in this region were most closely related to H5N1 clade 2.3.4.4, with the exception of one virus with an HA gene derived from clade 2.3.2 such that it represents a novel reassortant. Clearly, the ongoing surveillance of LPMs is central to monitoring the emergence of pathogenic influenza viruses. Copyright © 2017 American Society for Microbiology.

  7. Novel Highly Pathogenic Avian Influenza A(H5N6) Virus in the Netherlands, December 2017.

    Science.gov (United States)

    Beerens, Nancy; Koch, Guus; Heutink, Rene; Harders, Frank; Vries, D P Edwin; Ho, Cynthia; Bossers, Alex; Elbers, Armin

    2018-04-17

    A novel highly pathogenic avian influenza A(H5N6) virus affecting wild birds and commercial poultry was detected in the Netherlands in December 2017. Phylogenetic analysis demonstrated that the virus is a reassortant of H5N8 clade 2.3.4.4 viruses and not related to the Asian H5N6 viruses that caused human infections.

  8. Epidemiology of avian influenza H5N1 virus in Egypt and its zoonotic potential

    Directory of Open Access Journals (Sweden)

    Nahed Hamed Ghoneim

    2014-09-01

    Full Text Available Objective: To investigate the epidemiology of avian influenza H5N1 virus in domestic poultry and its zoonotic potential in Egypt. Methods: Tracheal swabs were collected from two hundred and forty three domestic poultry (chickens, ducks and geese from commercial farms and backyards, and thirty two blood samples from unvaccinated chickens. Fifty two throat swabs and twenty blood samples were collected from persons who are in contact with diseased and/or infected birds. Tracheal and throat swabs were examined for the presence of avian influenza virus H5N1 genome by real-time RT-PCR whereas blood samples were tested by competitive ELISA for the presence of avian influenza virus H5 antibodies. Results: The overall prevalence of H5N1 in the examined birds was 5.3% while the prevalence rates among different poultry species were 9%, 4.7% and 0% for ducks, chicken and geese respectively. Moreover, we detected H5 antibodies in 12.5% of the examined backyard chickens. All examined humans were negative for both viral RNA and antibodies. Conclusions: Our findings highlight the broad circulation of H5N1 virus among poultry in Egypt whereas it still has a limited zoonotic potential so far.

  9. Isolation and characterization of highly pathogenic avian influenza virus subtype H5N1 from donkeys

    Science.gov (United States)

    2010-01-01

    Background The highly pathogenic H5N1 is a major avian pathogen that crosses species barriers and seriously affects humans as well as some mammals. It mutates in an intensified manner and is considered a potential candidate for the possible next pandemic with all the catastrophic consequences. Methods Nasal swabs were collected from donkeys suffered from respiratory distress. The virus was isolated from the pooled nasal swabs in specific pathogen free embryonated chicken eggs (SPF-ECE). Reverse transcriptase polymerase chain reaction (RT-PCR) and sequencing of both haemagglutingin and neuraminidase were performed. H5 seroconversion was screened using haemagglutination inhibition (HI) assay on 105 donkey serum samples. Results We demonstrated that H5N1 jumped from poultry to another mammalian host; donkeys. Phylogenetic analysis showed that the virus clustered within the lineage of H5N1 from Egypt, closely related to 2009 isolates. It harboured few genetic changes compared to the closely related viruses from avian and humans. The neuraminidase lacks oseltamivir resistant mutations. Interestingly, HI screening for antibodies to H5 haemagglutinins in donkeys revealed high exposure rate. Conclusions These findings extend the host range of the H5N1 influenza virus, possess implications for influenza virus epidemiology and highlight the need for the systematic surveillance of H5N1 in animals in the vicinity of backyard poultry units especially in endemic areas. PMID:20398268

  10. Isolation and characterization of highly pathogenic avian influenza virus subtype H5N1 from donkeys

    Directory of Open Access Journals (Sweden)

    Abdel-Ghany Ahmad E

    2010-04-01

    Full Text Available Abstract Background The highly pathogenic H5N1 is a major avian pathogen that crosses species barriers and seriously affects humans as well as some mammals. It mutates in an intensified manner and is considered a potential candidate for the possible next pandemic with all the catastrophic consequences. Methods Nasal swabs were collected from donkeys suffered from respiratory distress. The virus was isolated from the pooled nasal swabs in specific pathogen free embryonated chicken eggs (SPF-ECE. Reverse transcriptase polymerase chain reaction (RT-PCR and sequencing of both haemagglutingin and neuraminidase were performed. H5 seroconversion was screened using haemagglutination inhibition (HI assay on 105 donkey serum samples. Results We demonstrated that H5N1 jumped from poultry to another mammalian host; donkeys. Phylogenetic analysis showed that the virus clustered within the lineage of H5N1 from Egypt, closely related to 2009 isolates. It harboured few genetic changes compared to the closely related viruses from avian and humans. The neuraminidase lacks oseltamivir resistant mutations. Interestingly, HI screening for antibodies to H5 haemagglutinins in donkeys revealed high exposure rate. Conclusions These findings extend the host range of the H5N1 influenza virus, possess implications for influenza virus epidemiology and highlight the need for the systematic surveillance of H5N1 in animals in the vicinity of backyard poultry units especially in endemic areas.

  11. Influence of virus strain and antigen mass on efficacy of H5 avian influenza inactivated vaccines.

    Science.gov (United States)

    Swayne, D E; Beck, J R; Garcia, M; Stone, H D

    1999-06-01

    The influence of vaccine strain and antigen mass on the ability of inactivated avian influenza (AI) viruses to protect chicks from a lethal, highly pathogenic (HP) AI virus challenge was studied. Groups of 4-week-old chickens were immunized with inactivated vaccines containing one of 10 haemagglutinin subtype H5 AI viruses, one heterologous H7 AI virus or normal allantoic fluid (sham), and challenged 3 weeks later by intra-nasal inoculation with a HP H5 chicken-origin AI virus. All 10 H5 vaccines provided good protection from clinical signs and death, and produced positive serological reactions on agar gel immunodiffusion and haemagglutination inhibition tests. In experiment 1, challenge virus was recovered from the oropharynx of 80% of chickens in the H5 vaccine group. In five H5 vaccine groups, challenge virus was not recovered from the cloaca of chickens. In the other five H5 vaccine groups, the number of chickens with detection of challenge virus from the cloaca was lower than in the sham group (P turkey/Wisconsin/68 (H5N9) was the best vaccine candidate of the H5 strains tested (PD50= 0.006 μg AI antigen). These data demonstrate that chickens vaccinated with inactivated H5 whole virus AI vaccines were protected from clinical signs and death, but usage of vaccine generally did not prevent infection by the challenge virus, as indicated by recovery of virus from the oropharynx. Vaccine use reduced cloacal detection rates, and quantity of virus shed from the cloaca and oropharynx in some vaccine groups, which would potentially reduce environmental contamination and disease transmission in the field.

  12. Novel Eurasian Highly Pathogenic Influenza A H5 Viruses in Wild Birds, Washington, USA, 2014

    Centers for Disease Control (CDC) Podcasts

    2015-03-24

    Sarah Gregory reads an abridged version of the article, Novel Eurasian Highly Pathogenic Influenza A H5 Viruses in Wild Birds, Washington, USA, 2014.  Created: 3/24/2015 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 4/13/2015.

  13. Highly pathogenic avian influenza virus (H5N1) isolated from whooper swans, Japan.

    Science.gov (United States)

    Uchida, Yuko; Mase, Masaji; Yoneda, Kumiko; Kimura, Atsumu; Obara, Tsuyoshi; Kumagai, Seikou; Saito, Takehiko; Yamamoto, Yu; Nakamura, Kikuyasu; Tsukamoto, Kenji; Yamaguchi, Shigeo

    2008-09-01

    On April 21, 2008, four whooper swans were found dead at Lake Towada, Akita prefecture, Japan. Highly pathogenic avian influenza virus of the H5N1 subtype was isolated from specimens of the affected birds. The hemagglutinin (HA) gene of the isolate belongs to clade 2.3.2 in the HA phylogenetic tree.

  14. Silent spread of highly pathogenic Avian Influenza H5N1 virus amongst vaccinated commercial layers

    NARCIS (Netherlands)

    Poetri, O.N.; Boven, M.; Claassen, I.J.T.M.; Koch, G.; Wibawan, I.W.; Stegeman, A.; Broek, van den J.; Bouma, A.

    2014-01-01

    The aim of this study was to determine whether a single vaccination of commercial layer type chickens with an inactivated vaccine containing highly pathogenic avian influenza virus strain H5N1 A/chicken/Legok/2003, carried out on the farm, was sufficient to protect against infection with the

  15. Human infection with highly pathogenic H5N1 influenza virus

    NARCIS (Netherlands)

    Gambotto, Andrea; Barratt-Boyes, Simon M.; de Jong, Menno D.; Neumann, Gabriele; Kawaoka, Yoshihiro

    2008-01-01

    Highly pathogenic H5N1 influenza A viruses have spread relentlessly across the globe since 2003, and they are associated with widespread death in poultry, substantial economic loss to farmers, and reported infections of more than 300 people with a mortality rate of 60%. The high pathogenicity of

  16. Comparing introduction to Europe of highly pathogenic avian influenza viruses A(H5N8) in 2014 and A(H5N1) in 2005

    NARCIS (Netherlands)

    Adlhoch, C.; Gossner, C.; Koch, G.; Brown, I.; Bouwstra, R.J.; Verdonck, F.; Penttinen, P.; Harder, T.

    2014-01-01

    Since the beginning of November 2014, nine outbreaks of highly pathogenic avian influenza virus (HPAIV) A(H5N8) in poultry have been detected in four European countries. In this report, similarities and differences between the modes of introduction of HPAIV A(H5N1) and A(H5N8) into Europe are

  17. Influenza A virus H5-specific antibodies in mute swans (Cygnus olor) in the USA.

    Science.gov (United States)

    Kistler, Whitney M; Stallknecht, David E; Lebarbenchon, Camille; Pedersen, Kerri; Marks, David R; Mickley, Randy; DeLiberto, Thomas J; Yabsley, Michael J

    2015-04-01

    The use of serologic assays for influenza A virus (IAV) surveillance in wild birds has increased because of the availability of commercial enzyme-linked immunosorbent assays (ELISAs). Recently, an H5-specific blocking ELISA (bELISA) was shown to reliably detect H5-specific antibodies to low- and high-pathogenic H5 viruses in experimentally infected waterfowl. Mute Swans (Cygnus olor) were frequently associated with highly pathogenic H5N1 outbreaks in Europe and may have a similar role if highly pathogenic H5N1 is introduced into North America. We measured the prevalence of antibodies to the nucleoprotein and H5 protein in Mute Swans using three serologic assays. We collected 340 serum samples from Mute Swans in Michigan, New Jersey, New York, and Rhode Island, US. We detected antibodies to the IAV nucleoprotein in 66.2% (225/340) of the samples. We detected H5-specific antibodies in 62.9% (214/340) and 18.8% (64/340) using a modified H5 bELISA protocol and hemagglutination inhibition (HI) assay, respectively. The modified H5 bELISA protocol detected significantly more positive samples than did the manufacturer's protocol. We also tested 46 samples using virus neutralization. Neutralization results had high agreement with the modified H5 bELISA protocol and detected a higher prevalence than did the HI assay. These results indicate that North American Mute Swans have high nucleoprotein and H5 antibody prevalences.

  18. Infectivity, transmission and pathogenicity of H5 highly pathogenic avian influenza clade 2.3.4.4 (H5N8 and H5N2) United States index viruses in Pekin ducks and Chinese geese

    Science.gov (United States)

    In late 2014, a H5N8 highly pathogenic avian influenza (HPAI) virus, clade 2.3.4.4, spread by migratory birds into North America mixing with low pathogenicity AI viruses to produce a H5N2 HPAI virus. The H5N8 and H5N2 HPAI viruses were detected initially in wild waterfowl and backyard birds, and lat...

  19. Viremia associated with fatal outcomes in ferrets infected with avian H5N1 influenza virus.

    Directory of Open Access Journals (Sweden)

    Xue Wang

    Full Text Available Avian H5N1 influenza viruses cause severe disease and high mortality in infected humans. However, tissue tropism and underlying pathogenesis of H5N1 virus infection in humans needs further investigation. The objective of this work was to study viremia, tissue tropism and disease pathogenesis of H5N1 virus infection in the susceptible ferret animal model. To evaluate the relationship of morbidity and mortality with virus loads, we performed studies in ferrets infected with the H5N1 strain A/VN/1203/04 to assess clinical signs after infection and virus load in lung, brain, ileum, nasal turbinate, nasal wash, and blood. We observed that H5N1 infection in ferrets is characterized by high virus load in the brain and and low levels in the ileum using real-time PCR. In addition, viral RNA was frequently detected in blood one or two days before death and associated with symptoms of diarrhea. Our observations further substantiate pathogenicity of H5N1 and further indicate that viremia may be a bio-marker for fatal outcomes in H5N1 infection.

  20. Highly pathogenic influenza A(H5N1 virus survival in complex artificial aquatic biotopes.

    Directory of Open Access Journals (Sweden)

    Viseth Srey Horm

    Full Text Available BACKGROUND: Very little is known regarding the persistence of Highly Pathogenic Avian Influenza (HPAI H5N1 viruses in aquatic environments in tropical countries, although environmental materials have been suggested to play a role as reservoirs and sources of transmission for H5N1 viruses. METHODOLOGY/PRINCIPAL FINDINGS: The survival of HPAI H5N1 viruses in experimental aquatic biotopes (water, mud, aquatic flora and fauna relevant to field conditions in Cambodia was investigated. Artificial aquatic biotopes, including simple ones containing only mud and water, and complex biotopes involving the presence of aquatic flora and fauna, were set up. They were experimentally contaminated with H5N1 virus. The persistence of HPAI H5N1 virus (local avian and human isolates was determined by virus isolation in embryonated chicken eggs and by real-time reverse-polymerase chain reaction. Persistence of infectious virus did not exceed 4 days, and was only identified in rain water. No infectious virus particles were detected in pond and lake water or mud even when high inoculum doses were used. However, viral RNA persisted up to 20 days in rain water and 7 days in pond or lake water. Viral RNA was also detected in mud samples, up to 14 days post-contamination in several cases. Infectious virus and viral RNA was detected in few cases in the aquatic fauna and flora, especially in bivalves and labyrinth fish, although these organisms seemed to be mostly passive carriers of the virus rather than host allowing virus replication. CONCLUSIONS/SIGNIFICANCE: Although several factors for the survival and persistence of HPAI viruses in the environment are still to be elucidated, and are particularly hard to control in laboratory conditions, our results, along with previous data, support the idea that environmental surveillance is of major relevance for avian influenza control programs.

  1. Rapid and highly informative diagnostic assay for H5N1 influenza viruses.

    Directory of Open Access Journals (Sweden)

    Nader Pourmand

    Full Text Available A highly discriminative and information-rich diagnostic assay for H5N1 avian influenza would meet immediate patient care needs and provide valuable information for public health interventions, e.g., tracking of new and more dangerous variants by geographic area as well as avian-to-human or human-to-human transmission. In the present study, we have designed a rapid assay based on multilocus nucleic acid sequencing that focuses on the biologically significant regions of the H5N1 hemagglutinin gene. This allows the prediction of viral strain, clade, receptor binding properties, low- or high-pathogenicity cleavage site and glycosylation status. H5 HA genes were selected from nine known high-pathogenicity avian influenza subtype H5N1 viruses, based on their diversity in biologically significant regions of hemagglutinin and/or their ability to cause infection in humans. We devised a consensus pre-programmed pyrosequencing strategy, which may be used as a faster, more accurate alternative to de novo sequencing. The available data suggest that the assay described here is a reliable, rapid, information-rich and cost-effective approach for definitive diagnosis of H5N1 avian influenza. Knowledge of the predicted functional sequences of the HA will enhance H5N1 avian influenza surveillance efforts.

  2. Amino Acid Substitutions Associated with Avian H5N6 Influenza A Virus Adaptation to Mice

    Directory of Open Access Journals (Sweden)

    Chunmao Zhang

    2017-09-01

    Full Text Available At least 15 cases of human beings infected with H5N6 have been reported since 2014, of which at least nine were fatal. The highly pathogenic avian H5N6 influenza virus may pose a serious threat to both public health and the poultry industry. However, the molecular features promoting the adaptation of avian H5N6 influenza viruses to mammalian hosts is not well understood. Here, we sequentially passaged an avian H5N6 influenza A virus (A/Northern Shoveler/Ningxia/488-53/2015 10 times in mice to identify the adaptive amino acid substitutions that confer enhanced virulence to H5N6 in mammals. The 1st and 10th passages of the mouse-adapted H5N6 viruses were named P1 and P10, respectively. P1 and P10 displayed higher pathogenicity in mice than their parent strain. P10 showed significantly higher replication capability in vivo and could be detected in the brains of mice, whereas P1 displayed higher replication efficiency in their lungs but was not detectable in the brain. Similar to its parent strain, P10 remained no transmissible between guinea pigs. Using genome sequencing and alignment, multiple amino acid substitutions, including PB2 E627K, PB2 T23I, PA T97I, and HA R239H, were found in the adaptation of H5N6 to mice. In summary, we identified amino acid changes that are associated with H5N6 adaptation to mice.

  3. Physician's knowledge, attitudes, and practices regarding seasonal influenza, pandemic influenza, and highly pathogenic avian influenza A (H5N1) virus infections of humans in Indonesia

    OpenAIRE

    Mangiri, Amalya; Iuliano, A. Danielle; Wahyuningrum, Yunita; Praptiningsih, Catharina Y.; Lafond, Kathryn E.; Storms, Aaron D.; Samaan, Gina; Ariawan, Iwan; Soeharno, Nugroho; Kreslake, Jennifer M.; Storey, J. Douglas; Uyeki, Timothy M.

    2016-01-01

    Indonesia has reported highest number of fatal human cases of highly pathogenic avian influenza (HPAI) A (H5N1) virus infection worldwide since 2005. There are limited data available on seasonal and pandemic influenza in Indonesia. During 2012, we conducted a survey of clinicians in two districts in western Java, Indonesia, to assess knowledge, attitudes, and practices (KAP) of clinical diagnosis, testing, and treatment of patients with seasonal influenza, pandemic influenza, or HPAI H5N1 vir...

  4. New avian influenza A virus subtype combination H5N7 identified in Danish mallard ducks

    DEFF Research Database (Denmark)

    Bragstad, K.; Jørgensen, Poul Henrik; Handberg, Kurt

    2005-01-01

    sequence was most closely related to the HPAIV A/Chicken/Netheriancts/01/03 (H7N7) that infected chickens and humans in the Netherlands in 2003. Ten persons with direct or indirect contact with the Danish mallard ducks showed signs Of influenza-like illness 2-3 clays following the killing of the ducks......During the past years increasing incidences of influenza A zoonosis have made it of uppermost importance to possess methods for rapid and precise identification and characterisation of influenza A Viruses. We present here a convenient one-step RT-PCR method that will amplify full......-length haemagglutinin (HA) and neuraminidase (NA) directly from clinical samples and from all known subtypes of influenza A. We applied the method on samples collected in September 2003 from a Danish flock of mallards with general health problems and by this a previously undescribed influenza A subtype combination, H5N...

  5. [Influenza A/H5N1 virus outbreaks and prepardness to avert flu pandemic].

    Science.gov (United States)

    Haque, A; Lucas, B; Hober, D

    2007-01-01

    This review emphasizes the need to improve the knowledge of the biology of H5N1 virus, a candidate for causing the next influenza pandemic. In-depth knowledge of mode of infection, mechanisms of pathogenesis and immune response will help in devising an efficient and practical control strategy against this flu virus. We have discussed limitations of currently available vaccines and proposed novel approaches for making better vaccines against H5N1 influenza virus. They include cell-culture system, reverse genetics, adjuvant development. Our review has also underscored the concept of therapeutic vaccine (anti-disease vaccine), which is aimed at diminishing 'cytokine storm' seen in acute respiratory distress syndrome and/or hemophagocytosis.

  6. Genetic characterization of highly pathogenic avian influenza A H5N8 viruses isolated from wild birds in Egypt.

    Science.gov (United States)

    Kandeil, Ahmed; Kayed, Ahmed; Moatasim, Yassmin; Webby, Richard J; McKenzie, Pamela P; Kayali, Ghazi; Ali, Mohamed A

    2017-07-01

    A newly emerged H5N8 influenza virus was isolated from green-winged teal in Egypt during December 2016. In this study, we provide a detailed characterization of full genomes of Egyptian H5N8 viruses and some virological features. Genetic analysis demonstrated that the Egyptian H5N8 viruses are highly pathogenic avian influenza viruses. Phylogenetic analysis revealed that the genome of the Egyptian H5N8 viruses was related to recently characterized reassortant H5N8 viruses of clade 2.3.4.4 isolated from different Eurasian countries. Multiple peculiar mutations were characterized in the Egyptian H5N8 viruses, which probably permits transmission and virulence of these viruses in mammals. The Egyptian H5N8 viruses preferentially bound to avian-like receptors rather than human-like receptors. Also, the Egyptian H5N8 viruses were fully sensitive to amantadine and neuraminidase inhibitors. Chicken sera raised against commercial inactivated avian influenza-H5 vaccines showed no or very low reactivity with the currently characterized H5N8 viruses in agreement with the genetic dissimilarity. Surveillance of avian influenza in waterfowl provides early warning of specific threats to poultry and human health and hence should be continued.

  7. Avian influenza virus (H5N1; effects of physico-chemical factors on its survival

    Directory of Open Access Journals (Sweden)

    Hameed Sajid

    2009-03-01

    Full Text Available Abstract Present study was performed to determine the effects of physical and chemical agents on infective potential of highly pathogenic avian influenza (HPAI H5N1 (local strain virus recently isolated in Pakistan during 2006 outbreak. H5N1 virus having titer 108.3 ELD50/ml was mixed with sterilized peptone water to get final dilution of 4HA units and then exposed to physical (temperature, pH and ultraviolet light and chemical (formalin, phenol crystals, iodine crystals, CID 20, virkon®-S, zeptin 10%, KEPCIDE 300, KEPCIDE 400, lifebuoy, surf excel and caustic soda agents. Harvested amnio-allantoic fluid (AAF from embryonated chicken eggs inoculated with H5N1 treated virus (0.2 ml/egg was subjected to haemagglutination (HA and haemagglutination inhibition (HI tests. H5N1 virus lost infectivity after 30 min at 56°C, after 1 day at 28°C but remained viable for more than 100 days at 4°C. Acidic pH (1, 3 and basic pH (11, 13 were virucidal after 6 h contact time; however virus retained infectivity at pH 5 (18 h, 7 and 9 (more than 24 h. UV light was proved ineffectual in inactivating virus completely even after 60 min. Soap (lifebuoy®, detergent (surf excel® and alkali (caustic soda destroyed infectivity after 5 min at 0.1, 0.2 and 0.3% dilution. All commercially available disinfectants inactivated virus at recommended concentrations. Results of present study would be helpful in implementing bio-security measures at farms/hatcheries levels in the wake of avian influenza virus (AIV outbreak.

  8. Detection of H5 Avian Influenza Viruses by Antigen-Capture Enzyme-Linked Immunosorbent Assay Using H5-Specific Monoclonal Antibody▿

    OpenAIRE

    He, Qigai; Velumani, Sumathy; Du, Qingyun; Lim, Chee Wee; Ng, Fook Kheong; Donis, Ruben; Kwang, Jimmy

    2007-01-01

    The unprecedented spread of highly pathogenic avian influenza virus subtype H5N1 in Asia and Europe is threatening animals and public health systems. Effective diagnosis and control management are needed to control the disease. To this end, we developed a panel of monoclonal antibodies (MAbs) against the H5N1 avian influenza virus (AIV) and implemented an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) to detect the H5 viral antigen. Mice immunized with denatured hemagglutinin (H...

  9. Experimental assessment of the pathogenicity of two avian influenza A H5 viruses in ostrich chicks (Struthio camelus) and chickens

    DEFF Research Database (Denmark)

    Manvell, R.J.; Jørgensen, Poul Henrik; Nielsen, O.L.

    1998-01-01

    Virus excretion, immune response, and, for chickens, deaths were recorded in 3-week-old ostriches and chickens inoculated by either the intramuscular or intranasal route with one of two influenza A viruses of subtype H5, One of the viruses, A/turkey/England/50-92/91 (H5N1) (50/92), was highly...

  10. Risk factors for exposure to influenza a viruses, including subtype H5 viruses, in Thai free-grazing ducks.

    Science.gov (United States)

    Beaudoin, A L; Kitikoon, P; Schreiner, P J; Singer, R S; Sasipreeyajan, J; Amonsin, A; Gramer, M R; Pakinsee, S; Bender, J B

    2014-08-01

    Free-grazing ducks (FGD) have been associated with highly pathogenic avian influenza (HPAI) H5N1 outbreaks and may be a viral reservoir. In July-August 2010, we assessed influenza exposure of Thai FGD and risk factors thereof. Serum from 6254 ducks was analysed with enzyme-linked immunosorbent assay (ELISA) to detect antibodies to influenza A nucleoprotein (NP), and haemagglutinin H5 protein. Eighty-five per cent (5305 ducks) were seropositive for influenza A. Of the NP-seropositive sera tested with H5 assays (n = 1423), 553 (39%) were H5 ELISA positive and 57 (4%) suspect. Twelve per cent (74 of 610) of H5 ELISA-positive/suspect ducks had H5 titres ≥ 1 : 20 by haemagglutination inhibition. Risk factors for influenza A seropositivity include older age, poultry contact, flock visitors and older purchase age. Study flocks had H5 virus exposure as recently as March 2010, but no HPAI H5N1 outbreaks have been identified in Thailand since 2008, highlighting a need for rigorous FGD surveillance. © 2012 Blackwell Verlag GmbH.

  11. Acid Stability of the Hemagglutinin Protein Regulates H5N1 Influenza Virus Pathogenicity

    Energy Technology Data Exchange (ETDEWEB)

    DuBois, Rebecca M.; Zaraket, Hassan; Reddivari, Muralidhar; Heath, Richard J.; White, Stephen W.; Russell, Charles J. (Tennessee-HSC); (SJCH)

    2012-12-10

    Highly pathogenic avian influenza viruses of the H5N1 subtype continue to threaten agriculture and human health. Here, we use biochemistry and x-ray crystallography to reveal how amino-acid variations in the hemagglutinin (HA) protein contribute to the pathogenicity of H5N1 influenza virus in chickens. HA proteins from highly pathogenic (HP) A/chicken/Hong Kong/YU562/2001 and moderately pathogenic (MP) A/goose/Hong Kong/437-10/1999 isolates of H5N1 were found to be expressed and cleaved in similar amounts, and both proteins had similar receptor-binding properties. However, amino-acid variations at positions 104 and 115 in the vestigial esterase sub-domain of the HA1 receptor-binding domain (RBD) were found to modulate the pH of HA activation such that the HP and MP HA proteins are activated for membrane fusion at pH 5.7 and 5.3, respectively. In general, an increase in H5N1 pathogenicity in chickens was found to correlate with an increase in the pH of HA activation for mutant and chimeric HA proteins in the observed range of pH 5.2 to 6.0. We determined a crystal structure of the MP HA protein at 2.50 {angstrom} resolution and two structures of HP HA at 2.95 and 3.10 {angstrom} resolution. Residues 104 and 115 that modulate the acid stability of the HA protein are situated at the N- and C-termini of the 110-helix in the vestigial esterase sub-domain, which interacts with the B loop of the HA2 stalk domain. Interactions between the 110-helix and the stalk domain appear to be important in regulating HA protein acid stability, which in turn modulates influenza virus replication and pathogenesis. Overall, an optimal activation pH of the HA protein is found to be necessary for high pathogenicity by H5N1 influenza virus in avian species.

  12. Susceptibility of swine to H5 and H7 low pathogenic avian influenza viruses.

    Science.gov (United States)

    Balzli, Charles; Lager, Kelly; Vincent, Amy; Gauger, Phillip; Brockmeier, Susan; Miller, Laura; Richt, Juergen A; Ma, Wenjun; Suarez, David; Swayne, David E

    2016-07-01

    The ability of pigs to become infected with low pathogenic avian influenza (LPAI) viruses and then generate mammalian adaptable influenza A viruses is difficult to determine. Yet, it is an important link to understanding any relationship between LPAI virus ecology and possible epidemics among swine and/or humans. Assess susceptibility of pigs to LPAI viruses found within the United States and their direct contact transmission potential. Pigs were inoculated with one of ten H5 or H7 LPAI viruses selected from seven different bird species to test infectivity, virulence, pathogenesis, and potential to transmit virus to contact pigs through histological, RRT-PCR and seroconversion data. Although pigs were susceptible to infection with each of the LPAI viruses, no clinical disease was recognized in any pig. During the acute phase of the infection, minor pulmonary lesions were found in some pigs and one or more pigs in each group were RRT-PCR-positive in the lower respiratory tract, but no virus was detected in upper respiratory tract (negative nasal swabs). Except for one group, one or more pigs in each LPAI group developed antibody. No LPAI viruses transmitted to contact pigs. LPAI strains from various bird populations within the United States are capable of infecting pigs. Although adaptability and transmission of individual strains seem unlikely, the subclinical nature of the infections demonstrates the need to improve sampling and testing methods to more accurately measure incidence of LPAI virus infection in pigs, and their potential role in human-zoonotic LPAI virus dynamics. © 2016 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

  13. Influenza H5N1 virus infection of polarized human alveolar epithelial cells and lung microvascular endothelial cells

    Directory of Open Access Journals (Sweden)

    Yuen Kit M

    2009-10-01

    Full Text Available Abstract Background Highly pathogenic avian influenza (HPAI H5N1 virus is entrenched in poultry in Asia and Africa and continues to infect humans zoonotically causing acute respiratory disease syndrome and death. There is evidence that the virus may sometimes spread beyond respiratory tract to cause disseminated infection. The primary target cell for HPAI H5N1 virus in human lung is the alveolar epithelial cell. Alveolar epithelium and its adjacent lung microvascular endothelium form host barriers to the initiation of infection and dissemination of influenza H5N1 infection in humans. These are polarized cells and the polarity of influenza virus entry and egress as well as the secretion of cytokines and chemokines from the virus infected cells are likely to be central to the pathogenesis of human H5N1 disease. Aim To study influenza A (H5N1 virus replication and host innate immune responses in polarized primary human alveolar epithelial cells and lung microvascular endothelial cells and its relevance to the pathogenesis of human H5N1 disease. Methods We use an in vitro model of polarized primary human alveolar epithelial cells and lung microvascular endothelial cells grown in transwell culture inserts to compare infection with influenza A subtype H1N1 and H5N1 viruses via the apical or basolateral surfaces. Results We demonstrate that both influenza H1N1 and H5N1 viruses efficiently infect alveolar epithelial cells from both apical and basolateral surface of the epithelium but release of newly formed virus is mainly from the apical side of the epithelium. In contrast, influenza H5N1 virus, but not H1N1 virus, efficiently infected polarized microvascular endothelial cells from both apical and basolateral aspects. This provides a mechanistic explanation for how H5N1 virus may infect the lung from systemic circulation. Epidemiological evidence has implicated ingestion of virus-contaminated foods as the source of infection in some instances and our

  14. Isolation of avian influenza H5N1 virus from vaccinated commercial layer flock in Egypt

    Directory of Open Access Journals (Sweden)

    El-Zoghby Elham F

    2012-11-01

    Full Text Available Abstract Background Uninterrupted transmission of highly pathogenic avian influenza virus (HPAIV H5N1 of clade 2.2.1 in Egypt since 2006 resulted in establishment of two main genetic clusters. The 2.2.1/C group where all recent human and majority of backyard origin viruses clustered together, meanwhile the majority of viruses derived from vaccinated poultry in commercial farms grouped in 2.2.1.1 clade. Findings In the present investigation, an HPAIV H5N1 was isolated from twenty weeks old layers chickens that were vaccinated with a homologous H5N1 vaccine at 1, 7 and 16 weeks old. At twenty weeks of age, birds showed cyanosis of comb and wattle, decrease in egg production and up to 27% mortality. Examined serum samples showed low antibody titer in HI test (Log2 3.2± 4.2. The hemagglutinin (HA and neuraminidase (NA genes of the isolated virus were closely related to viruses in 2.2.1/C group isolated from poultry in live bird market (LBM and backyards or from infected people. Conspicuous mutations in the HA and NA genes including a deletion within the receptor binding domain in the HA globular head region were observed. Conclusions Despite repeated vaccination of layer chickens using a homologous H5N1 vaccine, infection with HPAIV H5N1 resulted in significant morbidity and mortality. In endemic countries like Egypt, rigorous control measures including enforcement of biosecurity, culling of infected birds and constant update of vaccine virus strains are highly required to prevent circulation of HPAIV H5N1 between backyard birds, commercial poultry, LBM and humans.

  15. Broad-spectrum detection of H5 subtype influenza A viruses with a new fluorescent immunochromatography system.

    Directory of Open Access Journals (Sweden)

    Akira Sakurai

    Full Text Available Immunochromatography (IC is an antigen-detection assay that plays an important role in the rapid diagnosis of influenza virus because the protocol is short time and easy to use. Despite the usability of IC, the sensitivity is approximately 10(3 pfu per reaction. In addition, antigen-antibody interaction-based method cannot be used for the detection of influenza viruses with major antigenic change. In this study, we established the use of fluorescent immunochromatography (FLIC to detect a broad spectrum of H5 subtype influenza A viruses. This method has improved sensitivity 10-100 fold higher than traditional IC because of the use of fluorescent conjugated beads. Our Type-E FLIC kit detected all of the H5 subtype influenza viruses that were examined, as well as recombinant hemagglutinin (HA proteins (rHAs belonging to the Eurasian H5 subtype viruses and the Type-N diagnosed North American H5 subtype influenza A viruses. Thus, this kit has the improved potential to detect H5 subtype influenza viruses of different clades with both Type-E and Type-N FLIC kits. Compared with PCR-based diagnosis, FLIC has a strong advantage in usability, because the sample preparation required for FLIC is only mix-and-drop without any additional steps such as RNA extraction. Our results can provide new strategies against the spread and transmission of HPAI H5N1 viruses in birds and mammals including humans.

  16. An emerging avian influenza A virus H5N7 is a genetic reassortant of highly pathogenic genes

    DEFF Research Database (Denmark)

    Bragstad, K.; Jørgensen, Poul Henrik; Handberg, Kurt

    2006-01-01

    We full genome characterised the newly discovered avian influenza virus H5N7 subtype combination isolated from a stock of Danish game ducks to investigate the composition of the genome and possible features of high pathogenicity. It was found that the haemagglutinin and the acidic polymerase gene...... low pathogenic avian influenza A viruses. (c) 2006 Elsevier Ltd. All rights reserved....

  17. Effect of receptor binding domain mutations on receptor binding and transmissibility of avian influenza H5N1 viruses

    DEFF Research Database (Denmark)

    Maines, Taronna R; Chen, Li-Mei; Van Hoeven, Neal

    2011-01-01

    Although H5N1 influenza viruses have been responsible for hundreds of human infections, these avian influenza viruses have not fully adapted to the human host. The lack of sustained transmission in humans may be due, in part, to their avian-like receptor preference. Here, we have introduced recep...

  18. Genome characterisation of the newly discovered avian influenza A H5N7 virus subtype combination

    DEFF Research Database (Denmark)

    Bragstad, K.; Jørgensen, Poul Henrik; Handberg, K.J.

    2007-01-01

    In Denmark, in 2003, a previously unknown subtype combination of avian influenza A virus, H5N7 (A/Mallard/Denmark/64650/03), was isolated from a flock of 12,000 mallards. The H5N7 subtype combination might be a reassortant between recent European avian influenza A H5, H7, and a third subtype......) and the human-fatal A/Netherlands/219/03 (H7N7), respectively. The basic polymerase 1 and 2 genes were phylogenetically equidistant to both A/Duck/Denmark/65047/04 (H5N2) and A/Chicken/Netherlands/1/03 (H7N7). The nucleoprotein and matrix gene had highest nucleotide sequence similarity to the H6 subtypes A....../Duck/Hong Kong/3096/99 (H6N2) and A/WDk/ST/1737/2000 (H6N8), respectively. All genes of the H5N7 strain were of avian origin, and no further evidence of pathogenicity to humans has been found....

  19. Widespread detection of highly pathogenic H5 influenza viruses in wild birds from the Pacific Flyway of the United States.

    Science.gov (United States)

    Bevins, S N; Dusek, R J; White, C L; Gidlewski, T; Bodenstein, B; Mansfield, K G; DeBruyn, P; Kraege, D; Rowan, E; Gillin, C; Thomas, B; Chandler, S; Baroch, J; Schmit, B; Grady, M J; Miller, R S; Drew, M L; Stopak, S; Zscheile, B; Bennett, J; Sengl, J; Brady, Caroline; Ip, H S; Spackman, E; Killian, M L; Torchetti, M K; Sleeman, J M; Deliberto, T J

    2016-07-06

    A novel highly pathogenic avian influenza virus belonging to the H5 clade 2.3.4.4 variant viruses was detected in North America in late 2014. Motivated by the identification of these viruses in domestic poultry in Canada, an intensive study was initiated to conduct highly pathogenic avian influenza surveillance in wild birds in the Pacific Flyway of the United States. A total of 4,729 hunter-harvested wild birds were sampled and highly pathogenic avian influenza virus was detected in 1.3% (n = 63). Three H5 clade 2.3.4.4 subtypes were isolated from wild birds, H5N2, H5N8, and H5N1, representing the wholly Eurasian lineage H5N8 and two novel reassortant viruses. Testing of 150 additional wild birds during avian morbidity and mortality investigations in Washington yielded 10 (6.7%) additional highly pathogenic avian influenza isolates (H5N8 = 3 and H5N2 = 7). The geographically widespread detection of these viruses in apparently healthy wild waterfowl suggest that the H5 clade 2.3.4.4 variant viruses may behave similarly in this taxonomic group whereby many waterfowl species are susceptible to infection but do not demonstrate obvious clinical disease. Despite these findings in wild waterfowl, mortality has been documented for some wild bird species and losses in US domestic poultry during the first half of 2015 were unprecedented.

  20. The antigenic property of the H5N1 avian influenza viruses isolated in central China

    Directory of Open Access Journals (Sweden)

    Zou Wei

    2012-08-01

    Full Text Available Abstract Background Three influenza pandemics outbroke in the last century accompanied the viral antigen shift and drift, resulting in the change of antigenic property and the low cross protective ability of the existed antibody to the newly emerged pandemic virus, and eventually the death of millions of people. The antigenic characterizations of the viruses isolated in central China in 2004 and 2006–2007 were investigated in the present study. Results Hemagglutinin inhibition assay and neutralization assay displayed differential antigenic characteristics of the viruses isolated in central China in two periods (2004 and 2006–2007. HA genes of the viruses mainly located in two branches in phylogeny analysis. 53 mutations of the deduced amino acids of the HA genes were divided into 4 patterns. Mutations in pattern 2 and 3 showed the main difference between viruses isolated in 2004 and 2006–2007. Meanwhile, most amino acids in pattern 2 and 3 located in the globular head of the HA protein, and some of the mutations evenly distributed at the epitope sites. Conclusions The study demonstrated that a major antigenic drift had occurred in the viruses isolated in central China. And monitoring the antigenic property should be the priority in preventing the potential pandemic of H5N1 avian influenza virus.

  1. Purification of neuraminidase from Influenza virus subtype H5N1

    Directory of Open Access Journals (Sweden)

    Simson Tariga

    2009-03-01

    Full Text Available Influenza-virus neuraminidase plays vital role in the survival of the organisms. Vaccination of animals with this glycoprotein confers immune responses so that enable it to protect the animals from incoming infection. Supplementation of conventional vaccines with this glycoprotein increases the protection and longevity of the vaccine. Purified neuraminidase can also be used to develop serological tests for differentiation of serologically positive animals due to infection or to vaccination. In this study purification of neuraminidase from influenza virus subtype H5N1 was described. Triton x-100 and Octyl β-D-glucopyranoside were used to extract and diluted the glycoprotein membrane. The enzymatic activity of the neuraminidase was assayed using a fluorochrome substrate, 4-methylumbelliferyl-a-D-N-acetyl neuraminic acid, which was found to be simple, sensitive and suitable for the purification purpose. The neuraminidase was absorbed selectively on an oxamic-acid agarose column. The purity of neuraminidase eluted from this affinity column was high. A higher purity of the neuraminidase was obtained by further separation with gel filtration on Superdex-200. The purified neuraminidase was enzymatically active and did not contain any detectable haemagglutinin, either by haemagglutination assay or by monospecific antibodies raised against H5N1 hemagglutinin. The purified neuraminidase was recognized strongly by antibodies raised against an internal but only weakly by that against C-terminal regions of the neuraminidase protein of H5N1-influenza virus. The purified neuraminidase was in tetrameric forms but dissociated into monomeric form on reducing condition, or mostly dimeric form on non-reducing SDS-PAGE.

  2. Avian Influenza A (H5N1)

    Centers for Disease Control (CDC) Podcasts

    In this podcast, CDC's Dr. Tim Uyeki discusses H5N1, a subtype of influenza A virus. This highly pathogenic H5N1 virus doesn't usually infect people, although some rare infections with H5N1 viruses have occurred in humans. We need to use a comprehensive strategy to prevent the spread of H5N1 virus among birds, including having human health and animal health work closely together.

  3. Inefficient transmission of H5N1 influenza viruses in a ferret contact model.

    Science.gov (United States)

    Yen, Hui-Ling; Lipatov, Aleksandr S; Ilyushina, Natalia A; Govorkova, Elena A; Franks, John; Yilmaz, Neziha; Douglas, Alan; Hay, Alan; Krauss, Scott; Rehg, Jerold E; Hoffmann, Erich; Webster, Robert G

    2007-07-01

    The abilities to infect and transmit efficiently among humans are essential for a novel influenza A virus to cause a pandemic. To evaluate the pandemic potential of widely disseminated H5N1 influenza viruses, a ferret contact model using experimental groups comprised of one inoculated ferret and two contact ferrets was used to study the transmissibility of four human H5N1 viruses isolated from 2003 to 2006. The effects of viral pathogenicity and receptor binding specificity (affinity to synthetic sialosaccharides with alpha2,3 or alpha2,6 linkages) on transmissibility were assessed. A/Vietnam/1203/04 and A/Vietnam/JP36-2/05 viruses, which possess "avian-like" alpha2,3-linked sialic acid (SA) receptor specificity, caused neurological symptoms and death in ferrets inoculated with 10(3) 50% tissue culture infectious doses. A/Hong Kong/213/03 and A/Turkey/65-596/06 viruses, which show binding affinity for "human-like" alpha2,6-linked SA receptors in addition to their affinity for alpha2,3-linked SA receptors, caused mild clinical symptoms and were not lethal to the ferrets. No transmission of A/Vietnam/1203/04 or A/Turkey/65-596/06 virus was detected. One contact ferret developed neutralizing antibodies to A/Hong Kong/213/03 but did not exhibit any clinical signs or detectable virus shedding. In two groups, one of two naïve contact ferrets had detectable virus after 6 to 8 days when housed together with the A/Vietnam/JP36-2/05 virus-inoculated ferrets. Infected contact ferrets showed severe clinical signs, although little or no virus was detected in nasal washes. This limited virus shedding explained the absence of secondary transmission from the infected contact ferret to the other naïve ferret that were housed together. Our results suggest that despite their receptor binding affinity, circulating H5N1 viruses retain molecular determinants that restrict their spread among mammalian species.

  4. Genetic Characterization of Continually Evolving Highly Pathogenic H5N6 Influenza Viruses in China, 2012–2016

    Science.gov (United States)

    Li, Meng; Zhao, Na; Luo, Jing; Li, Yuan; Chen, Lin; Ma, Jiajun; Zhao, Lin; Yuan, Guohui; Wang, Chengmin; Wang, Yutian; Liu, Yanhua; He, Hongxuan

    2017-01-01

    H5N6 is a highly pathogenic avian influenza (HPAI) and a zoonotic disease that causes recurring endemics in East Asia. At least 155 H5N6 outbreaks, including 15 human infections, have been reported in China. These repeated outbreaks have increased concern that the H5N6 virus may cross over to humans and cause a pandemic. In February, 2016, peafowls in a breeding farm exhibited a highly contagious disease. Post-mortem examinations, including RT-PCR, and virus isolation, confirmed that the highly pathogenic H5N6 influenza virus was the causative agent, and the strain was named A/Pavo Cristatus/Jiangxi/JA1/2016. In animal experiments, it exhibited high pathogenicity in chickens and an estimated median lethal dose in mice of ~104.3 TCID50. A phylogenetic analysis showed that JA1/2016 was clustered in H5 clade 2.3.4.4. FG594-like H5N6 virus from Guangdong Province was the probable predecessor of JA1/2016, and the estimated divergence time was June 2014. Furthermore, we found that H5N6 influenza viruses can be classified into the two following groups: Group 1 and Group 2. Group 2 influenza viruses have not been detected since the end of 2014, whereas Group 1 influenza viruses have continually evolved and reassorted with the “gene pool” circulating in south China, resulting in the rise of novel subtypes of this influenza virus. An increase in the number of its identified hosts, the expanding range of its distribution, and the continual evolution of H5N6 AIVs enhance the risk that an H5N6 virus may spread to other continents and cause a pandemic. PMID:28293218

  5. Genetic Characterization of Continually Evolving Highly Pathogenic H5N6 Influenza Viruses in China, 2012-2016.

    Science.gov (United States)

    Li, Meng; Zhao, Na; Luo, Jing; Li, Yuan; Chen, Lin; Ma, Jiajun; Zhao, Lin; Yuan, Guohui; Wang, Chengmin; Wang, Yutian; Liu, Yanhua; He, Hongxuan

    2017-01-01

    H5N6 is a highly pathogenic avian influenza (HPAI) and a zoonotic disease that causes recurring endemics in East Asia. At least 155 H5N6 outbreaks, including 15 human infections, have been reported in China. These repeated outbreaks have increased concern that the H5N6 virus may cross over to humans and cause a pandemic. In February, 2016, peafowls in a breeding farm exhibited a highly contagious disease. Post-mortem examinations, including RT-PCR, and virus isolation, confirmed that the highly pathogenic H5N6 influenza virus was the causative agent, and the strain was named A/ Pavo Cristatus /Jiangxi/JA1/2016. In animal experiments, it exhibited high pathogenicity in chickens and an estimated median lethal dose in mice of ~10 4.3 TCID 50 . A phylogenetic analysis showed that JA1/2016 was clustered in H5 clade 2.3.4.4. FG594-like H5N6 virus from Guangdong Province was the probable predecessor of JA1/2016, and the estimated divergence time was June 2014. Furthermore, we found that H5N6 influenza viruses can be classified into the two following groups: Group 1 and Group 2. Group 2 influenza viruses have not been detected since the end of 2014, whereas Group 1 influenza viruses have continually evolved and reassorted with the "gene pool" circulating in south China, resulting in the rise of novel subtypes of this influenza virus. An increase in the number of its identified hosts, the expanding range of its distribution, and the continual evolution of H5N6 AIVs enhance the risk that an H5N6 virus may spread to other continents and cause a pandemic.

  6. Comparing introduction to Europe of highly pathogenic avian influenza viruses A(H5N8) in 2014 and A(H5N1) in 2005.

    Science.gov (United States)

    Adlhoch, C; Gossner, C; Koch, G; Brown, I; Bouwstra, R; Verdonck, F; Penttinen, P; Harder, T

    2014-12-18

    Since the beginning of November 2014, nine outbreaks of highly pathogenic avian influenza virus (HPAIV) A(H5N8) in poultry have been detected in four European countries. In this report, similarities and differences between the modes of introduction of HPAIV A(H5N1) and A(H5N8) into Europe are described. Experiences from outbreaks of A(H5N1) in Europe demonstrated that early detection to control HPAIV in poultry has proven pivotal to minimise the risk of zoonotic transmission and prevention of human cases.

  7. Fatal H5N6 Avian Influenza Virus Infection in a Domestic Cat and Wild Birds in China.

    Science.gov (United States)

    Yu, Zhijun; Gao, Xiaolong; Wang, Tiecheng; Li, Yanbing; Li, Yongcheng; Xu, Yu; Chu, Dong; Sun, Heting; Wu, Changjiang; Li, Shengnan; Wang, Haijun; Li, Yuanguo; Xia, Zhiping; Lin, Weishi; Qian, Jun; Chen, Hualan; Xia, Xianzhu; Gao, Yuwei

    2015-06-02

    H5N6 avian influenza viruses (AIVs) may pose a potential human risk as suggested by the first documented naturally-acquired human H5N6 virus infection in 2014. Here, we report the first cases of fatal H5N6 avian influenza virus (AIV) infection in a domestic cat and wild birds. These cases followed human H5N6 infections in China and preceded an H5N6 outbreak in chickens. The extensive migration routes of wild birds may contribute to the geographic spread of H5N6 AIVs and pose a risk to humans and susceptible domesticated animals, and the H5N6 AIVs may spread from southern China to northern China by wild birds. Additional surveillance is required to better understand the threat of zoonotic transmission of AIVs.

  8. Transmission of highly pathogenic avian influenza H5N1 virus in Pekin ducks is significantly reduced by a genetically distant H5N2 vaccine

    NARCIS (Netherlands)

    Goot, van der J.A.; Boven, van M.; Stegeman, A.; Water, van de S.G.P.; Jong, de M.C.M.; Koch, G.

    2008-01-01

    Domestic ducks play an important role in the epidemiology of H5N1 avian influenza. Although it is known that vaccines that have a high homology with the challenge virus are able to prevent infection in ducks, little is yet known about the ability of genetically more distant vaccines in preventing

  9. New Orf virus (Parapoxvirus) recombinant expressing H5 hemagglutinin protects mice against H5N1 and H1N1 influenza A virus.

    Science.gov (United States)

    Rohde, Jörg; Amann, Ralf; Rziha, Hanns-Joachim

    2013-01-01

    Previously we demonstrated the versatile utility of the Parapoxvirus Orf virus (ORFV) as a vector platform for the development of potent recombinant vaccines. In this study we present the generation of new ORFV recombinants expressing the hemagglutinin (HA) or nucleoprotein (NP) of the highly pathogenic avian influenza virus (HPAIV) H5N1. Correct foreign gene expression was examined in vitro by immunofluorescence, Western blotting and flow cytometry. The protective potential of both recombinants was evaluated in the mouse challenge model. Despite adequate expression of NP, the recombinant D1701-V-NPh5 completely failed to protect mice from lethal challenge. However, the H5 HA-expressing recombinant D1701-V-HAh5n mediated solid protection in a dose-dependent manner. Two intramuscular (i.m.) injections of the HA-expressing recombinant protected all animals from lethal HPAIV infection without loss of body weight. Notably, the immunized mice resisted cross-clade H5N1 and heterologous H1N1 (strain PR8) influenza virus challenge. In vivo antibody-mediated depletion of CD4-positive and/or CD8-posititve T-cell subpopulations during immunization and/or challenge infection implicated the relevance of CD4-positive T-cells for induction of protective immunity by D1701-V-HAh5n, whereas the absence of CD8-positive T-cells did not significantly influence protection. In summary, this study validates the potential of the ORFV vectored vaccines also to combat HPAIV.

  10. Adaptation of high-growth influenza H5N1 vaccine virus in Vero cells: implications for pandemic preparedness.

    Directory of Open Access Journals (Sweden)

    Yu-Fen Tseng

    Full Text Available Current egg-based influenza vaccine production technology can't promptly meet the global demand during an influenza pandemic as shown in the 2009 H1N1 pandemic. Moreover, its manufacturing capacity would be vulnerable during pandemics caused by highly pathogenic avian influenza viruses. Therefore, vaccine production using mammalian cell technology is becoming attractive. Current influenza H5N1 vaccine strain (NIBRG-14, a reassortant virus between A/Vietnam/1194/2004 (H5N1 virus and egg-adapted high-growth A/PR/8/1934 virus, could grow efficiently in eggs and MDCK cells but not Vero cells which is the most popular cell line for manufacturing human vaccines. After serial passages and plaque purifications of the NIBRG-14 vaccine virus in Vero cells, one high-growth virus strain (Vero-15 was generated and can grow over 10(8 TCID(50/ml. In conclusion, one high-growth H5N1 vaccine virus was generated in Vero cells, which can be used to manufacture influenza H5N1 vaccines and prepare reassortant vaccine viruses for other influenza A subtypes.

  11. Evolution of highly pathogenic avian H5N1 influenza viruses

    Energy Technology Data Exchange (ETDEWEB)

    Macken, Catherine A [Los Alamos National Laboratory; Green, Margaret A [Los Alamos National Laboratory

    2009-01-01

    Highly pathogenic avian H5N1 viruses have circulated in Southeast Asia for more than a decade, are now endemic in parts of this region, and have also spread to more than 60 countries on three continents. The evolution of these viruses is characterized by frequent reassortment events that have created a significant number of different genotypes, both transient and longer lasting. However, fundamental questions remain about the generation and perpetuation of this substantial genetic diversity. These gaps in understanding may, in part, be due to the difficulties of genotyping closely related viruses, and limitations in the size of the data sets used in analysis. Using our recently published novel genotyping procedure ('two-time test'), which is amenable to high throughput analysis and provides an increased level of resolution relative to previous analyses, we propose a detailed model for the evolution and diversification of avian H5N1 viruses. Our analysis suggests that (i) all current H5N1 genotypes are derived from a single, clearly defined sequence of initial reassortment events; (ii) reassortment of the polymerase and NP genes may have played an important role in avian H5N1 virus evolution; (iii) the current genotype Z viruses have diverged into three distinguishable sub-genotypes in the absence of reassortment; (iv) some potentially significant molecular changes appear to be correlated with particular genotypes (for example, reassortment of the internal genes is often paralleled by a change in the HA clade); and (v) as noted in earlier studies of avian influenza A virus evolution, novel segments are typically derived from different donors (i.e., there is no obvious pattern of gene linkage in reassortment). The model of avian H5N1 viral evolution by reassortment and mutation that emerges from our study provides a context within which significant amino acid changes may be revealed; it also may help in predicting the 'success' of newly emerging

  12. A duplex real-time RT-PCR assay for detecting H5N1 avian influenza virus and pandemic H1N1 influenza virus

    OpenAIRE

    Kang, Xiao-ping; Jiang, Tao; Li, Yong-qiang; Lin, Fang; Liu, Hong; Chang, Guo-hui; Zhu, Qing-yu; Qin, E-de; Qin, Cheng-feng; Yang, Yin-hui

    2010-01-01

    Abstract A duplex real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay was improved for simultaneous detection of highly pathogenic H5N1 avian influenza virus and pandemic H1N1 (2009) influenza virus, which is suitable for early diagnosis of influenza-like patients and for epidemiological surveillance. The sensitivity of this duplex real-time RT-PCR assay was 0.02 TCID50 (50% tissue culture infective dose) for H5N1 and 0.2 TCID50 for the pandemic H1N1, which was the same a...

  13. Antibodies to H5 subtype avian influenza virus and Japanese encephalitis virus in northern pintails (Anas acuta) sampled in Japan

    Science.gov (United States)

    Ramey, Andy M.; Spackman, Erica; Yeh, Jung-Yong; Fujita, Go; Konishi, Kan; Reed, John A.; Wilcox, Benjamin R.; Brown, Justin D.; Stallknecht, David E.

    2013-01-01

    Blood samples from 105 northern pintails (Anas acuta) captured on Hokkaido, Japan were tested for antibodies to avian influenza virus (AIV), Japanese encephalitis virus (JEV), and West Nile virus (WNV) to assess possible involvement of this species in the spread of economically important and potentially zoonotic pathogens. Antibodies to AIV were detected in 64 of 105 samples (61%). Of the 64 positives, 95% and 81% inhibited agglutination of two different H5 AIV antigens (H5N1 and H5N9), respectively. Antibodies to JEV and WNV were detected in five (5%) and none of the samples, respectively. Results provide evidence for prior exposure of migrating northern pintails to H5 AIV which couldhave implications for viral shedding and disease occurrence. Results also provide evidence for limited involvement of this species in the transmission and spread of flaviviruses during spring migration.

  14. Deaths among wild birds during highly pathogenic avian influenza A(H5N8) virus outbreak, the Netherlands

    NARCIS (Netherlands)

    Kleyheeg, Erik; Slaterus, Roy; Bodewes, Rogier; Rijks, Jolianne M.; Spierenburg, Marcel A.H.; Beerens, Nancy; Kelder, Leon; Poen, Marjolein J.; Stegeman, Jan A.; Fouchier, Ron A.M.; Kuiken, Thijs; Jeugd, van der Henk P.

    2017-01-01

    During autumn–winter 2016–2017, highly pathogenic avian influenza A(H5N8) viruses caused mass die-offs among wild birds in the Netherlands. Among the ≈13,600 birds reported dead, most were tufted ducks (Aythya fuligula) and Eurasian wigeons (Anas penelope). Recurrence of avian influenza outbreaks

  15. Deaths among Wild Birds during Highly Pathogenic Avian Influenza A(H5N8) Virus Outbreak, the Netherlands

    NARCIS (Netherlands)

    Kleyheeg, Erik; Slaterus, Roy; Bodewes, Rogier; Rijks, Jolianne M.; Spierenburg, Marcel A.H.; Beerens, Nancy; Kelder, Leon; Poen, Marjolein J.; Stegeman, Jan A.; Fouchier, Ron A. M.; Kuiken, Thijs; Jeugd, Henk P. van der

    2017-01-01

    During autumn–winter 2016–2017, highly pathogenic avian influenza A(H5N8) viruses caused mass die-offs among wild birds in the Netherlands. Among the ≈13,600 birds reported dead, most were tufted ducks (Aythya fuligula) and Eurasian wigeons (Anas penelope). Recurrence of avian influenza outbreaks

  16. Characterizing wild bird contact and seropositivity to highly pathogenic avian influenza A (H5N1) virus in Alaskan residents.

    Science.gov (United States)

    Reed, Carrie; Bruden, Dana; Byrd, Kathy K; Veguilla, Vic; Bruce, Michael; Hurlburt, Debby; Wang, David; Holiday, Crystal; Hancock, Kathy; Ortiz, Justin R; Klejka, Joe; Katz, Jacqueline M; Uyeki, Timothy M

    2014-09-01

    Highly pathogenic avian influenza A (HPAI) H5N1 viruses have infected poultry and wild birds on three continents with more than 600 reported human cases (59% mortality) since 2003. Wild aquatic birds are the natural reservoir for avian influenza A viruses, and migratory birds have been documented with HPAI H5N1 virus infection. Since 2005, clade 2.2 HPAI H5N1 viruses have spread from Asia to many countries. We conducted a cross-sectional seroepidemiological survey in Anchorage and western Alaska to identify possible behaviors associated with migratory bird exposure and measure seropositivity to HPAI H5N1. We enrolled rural subsistence bird hunters and their families, urban sport hunters, wildlife biologists, and a comparison group without bird contact. We interviewed participants regarding their exposures to wild birds and collected blood to perform serologic testing for antibodies against a clade 2.2 HPAI H5N1 virus strain. Hunters and wildlife biologists reported exposures to wild migratory birds that may confer risk of infection with avian influenza A viruses, although none of the 916 participants had evidence of seropositivity to HPAI H5N1. We characterized wild bird contact among Alaskans and behaviors that may influence risk of infection with avian influenza A viruses. Such knowledge can inform surveillance and risk communication surrounding HPAI H5N1 and other influenza viruses in a population with exposure to wild birds at a crossroads of intercontinental migratory flyways. © 2014 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

  17. Fluorescence biosensor based on CdTe quantum dots for specific detection of H5N1 avian influenza virus

    Science.gov (United States)

    Hoa Nguyen, Thi; Dieu Thuy Ung, Thi; Hien Vu, Thi; Tran, Thi Kim Chi; Quyen Dong, Van; Khang Dinh, Duy; Liem Nguyen, Quang

    2012-09-01

    This report highlights the fabrication of fluorescence biosensors based on CdTe quantum dots (QDs) for specific detection of H5N1 avian influenza virus. The core biosensor was composed of (i) the highly luminescent CdTe/CdS QDs, (ii) chromatophores extracted from bacteria Rhodospirillum rubrum, and (iii) the antibody of β-subunit. This core part was linked to the peripheral part of the biosensor via a biotin-streptavidin-biotin bridge and finally connected to the H5N1 antibody to make it ready for detecting H5N1 avian influenza virus. Detailed studies of each constituent were performed showing the image of QDs-labeled chromatophores under optical microscope, proper photoluminescence (PL) spectra of CdTe/CdS QDs, chromatophores and the H5N1 avian influenza viruses.

  18. Isolation and molecular characterization of an H5N1 swine influenza virus in China in 2015.

    Science.gov (United States)

    Wu, Haibo; Yang, Fan; Lu, Rufeng; Xu, Lihua; Liu, Fumin; Peng, Xiuming; Wu, Nanping

    2018-03-01

    In 2015, an H5N1 influenza virus was isolated from a pig in Zhejiang Province, Eastern China. This strain was characterized by whole-genome sequencing with subsequent phylogenetic analysis. Phylogenetic analysis showed that all segments from this strain belonged to clade 2.3.2 and that it had received its genes from poultry influenza viruses in China. A Glu627Lys mutation associated with pathogenicity was observed in the PB2 protein. This strain was moderately pathogenic in mice and was able to replicate without prior adaptation. These results suggest that active surveillance of swine influenza should be used as an early warning system for influenza outbreaks in mammals.

  19. Induction of long-term protective immune responses by influenza H5N1 virus-like particles.

    Directory of Open Access Journals (Sweden)

    Sang-Moo Kang

    Full Text Available Recurrent outbreaks of highly pathogenic H5N1 avian influenza virus pose a threat of eventually causing a pandemic. Early vaccination of the population would be the single most effective measure for the control of an emerging influenza pandemic.Influenza virus-like particles (VLPs produced in insect cell-culture substrates do not depend on the availability of fertile eggs for vaccine manufacturing. We produced VLPs containing influenza A/Viet Nam1203/04 (H5N1 hemagglutinin, neuraminidase, and matrix proteins, and investigated their preclinical immunogenicity and protective efficacy. Mice immunized intranasally with H5N1 VLPs developed high levels of H5N1 specific antibodies and were 100% protected against a high dose of homologous H5N1 virus infection at 30 weeks after immunization. Protection is likely to be correlated with humoral and cellular immunologic memory at systemic and mucosal sites as evidenced by rapid anamnestic responses to re-stimulation with viral antigen in vivo and in vitro.These results provide support for clinical evaluation of H5N1 VLP vaccination as a public health intervention to mitigate a possible pandemic of H5N1 influenza.

  20. Highly pathogenic avian influenza virus subtype H5N1 in mute swans (Cygnus olor) in Central Bosnia.

    Science.gov (United States)

    Goletić, Teufik; Gagić, Abdulah; Residbegović, Emina; Kustura, Aida; Kavazović, Aida; Savić, Vladimir; Harder, Timm; Starick, Elke; Prasović, Senad

    2010-03-01

    In order to determine the actual prevalence of avian influenza viruses (AIVs) in wild birds in Bosnia and Herzegovina, extensive surveillance was carried out between October 2005 and April 2006. A total of 394 samples representing 41 bird species were examined for the presence of influenza A virus using virus isolation in embryonated chicken eggs, PCR, and nucleotide sequencing. AIV subtype H5N1 was detected in two mute swans (Cygnus olor). The isolates were determined to be highly pathogenic avian influenza (HPAI) virus and the hemagglutinin sequence was closely similar to A/Cygnus olor/Astrakhan/ Ast05-2-10/2005 (H5N1). This is the first report of HPAI subtype H5N1 in Bosnia and Herzegovina.

  1. Evidence for common ancestry among viruses isolated from wild birds in Beringia and highly pathogenic intercontinental reassortant H5N1 and H5N2 influenza A viruses

    Science.gov (United States)

    Ramey, Andy M.; Reeves, Andrew; Teslaa, Joshua L.; Nashold, Sean W.; Donnelly, Tyrone F.; Bahl, Justin; Hall, Jeffrey S.

    2016-01-01

    Highly pathogenic clade 2.3.4.4 H5N8, H5N2, and H5N1 influenza A viruses were first detected in wild, captive, and domestic birds in North America in November–December 2014. In this study, we used wild waterbird samples collected in Alaska prior to the initial detection of clade 2.3.4.4 H5 influenza A viruses in North America to assess the evidence for: (1) dispersal of highly pathogenic influenza A viruses from East Asia to North America by migratory birds via Alaska and (2) ancestral origins of clade 2.3.4.4 H5 reassortant viruses in Beringia. Although we did not detect highly pathogenic influenza A viruses in our sample collection from western Alaska, we did identify viruses that contained gene segments sharing recent common ancestry with intercontinental reassortant H5N2 and H5N1 viruses. Results of phylogenetic analyses and estimates for times of most recent common ancestry support migratory birds sampled in Beringia as maintaining viral diversity closely related to novel highly pathogenic influenza A virus genotypes detected in North America. Although our results do not elucidate the route by which highly pathogenic influenza A viruses were introduced into North America, genetic evidence is consistent with the hypothesized trans-Beringian route of introduction via migratory birds.

  2. High antiviral effects of hibiscus tea extract on the H5 subtypes of low and highly pathogenic avian influenza viruses.

    Science.gov (United States)

    Baatartsogt, Tugsbaatar; Bui, Vuong N; Trinh, Dai Q; Yamaguchi, Emi; Gronsang, Dulyatad; Thampaisarn, Rapeewan; Ogawa, Haruko; Imai, Kunitoshi

    2016-10-01

    Viral neuraminidase inhibitors are widely used as synthetic anti-influenza drugs for the prevention and treatment of influenza. However, drug-resistant influenza A virus variants, including H5N1 highly pathogenic avian influenza viruses (HPAIVs), have been reported. Therefore, the discovery of novel and effective antiviral agents is warranted. We screened the antiviral effects of 11 herbal tea extracts (hibiscus, black tea, tencha, rosehip tea, burdock tea, green tea, jasmine tea, ginger tea, lavender tea, rose tea and oak tea) against the H5N1 HPAIV in vitro. Among the tested extracts, only the hibiscus extract and its fractionated extract (frHibis) highly and rapidly reduced the titers of all H5 HPAIVs and low pathogenic AIVs (LPAIVs) used in the pre-treatment tests of Madin-Darby canine kidney (MDCK) cells that were inoculated with a mixture of the virus and the extract. Immunogold electron microscopy showed that anti-H5 monoclonal antibodies could not bind to the deformed H5 virus particles pretreated with frHibis. In post-treatment tests of MDCK cells cultured in the presence of frHibis after infection with H5N1 HPAIV, the frHibis inhibited viral replication and the expression of viral antigens and genes. Among the plants tested, hibiscus showed the most prominent antiviral effects against both H5 HPAIV and LPAIV.

  3. Mucosal immunity induced by adenovirus-based H5N1 HPAI vaccine confers protection against a lethal H5N2 avian influenza virus challenge

    International Nuclear Information System (INIS)

    Park, Ki Seok; Lee, Jiyeung; Ahn, So Shin; Byun, Young-Ho; Seong, Baik Lin; Baek, Yun Hee; Song, Min-Suk; Choi, Young Ki; Na, Yun Jeong; Hwang, Inhwan; Sung, Young Chul; Lee, Chang Geun

    2009-01-01

    Development of effective vaccines against highly pathogenic avian influenza (HPAI) H5N1 viruses is a global public health priority. Considering the difficulty in predicting HPAI H5N1 pandemic strains, one strategy used in their design includes the development of formulations with the capacity of eliciting broad cross-protective immunity against multiple viral antigens. To this end we constructed a replication-defective recombinant adenovirus-based avian influenza virus vaccine (rAdv-AI) expressing the codon-optimized M2eX-HA-hCD40L and the M1-M2 fusion genes from HPAI H5N1 human isolate. Although there were no significant differences in the systemic immune responses observed between the intramuscular prime-intramuscular boost regimen (IM/IM) and the intranasal prime-intramuscular boost regimen (IN/IM), IN/IM induced more potent CD8 + T cell and antibody responses at mucosal sites than the IM/IM vaccination, resulting in more effective protection against lethal H5N2 avian influenza (AI) virus challenge. These findings suggest that the strategies used to induce multi-antigen-targeted mucosal immunity, such as IN/IM delivery of rAdv-AI, may be a promising approach for developing broad protective vaccines that may be more effective against the new HPAI pandemic strains.

  4. Dinamika Seroprevalensi Virus Avian Influenza H5 pada Itik di Pasar Unggas Beringkit dan Galiran

    Directory of Open Access Journals (Sweden)

    I Gusti Ngurah Narendra Putra

    2013-11-01

    Full Text Available Normal 0 false false false EN-US X-NONE X-NONE Live Bird Market (LBM has a high potential for spreading Avian Influenza Virus (AIV between fowls or from fowl to human. Up to now, a dinamic of avian flue incidents at many LBMs in Bali has not been reported. This research aimed to reveal a dynamic of seroprevalences of avian influenza in ducks at Beringkit (Badung and Galiran (Kelungkung LBMs. A total of 35 duck blood samples was collected from each of LBMs. Sampling was conducted monthly from March to August, 2012 . AIV antibody of duck serum was measured using Rapid Hemagglutination Inhibition (Rapid HI test. Seroprevalence differences were analyzes with Chi-square (?2 Nonparametric statistical test. The results showed that seroprevalences of AIV H5 in ducks at Beringkit and Galiran LBMs were very high, ranged from 68.6% to 100% and 65.7% to 97.1% respectively. A Dynamic of AIV H5 seroprevalences in ducks at Beringkit and Galiran LBM had a similar pattern, except in July 2012. This indicates that VAI H5 has been circulating for a long time and has been to be an endemic virus infection in ducks at LBMs in Bali. It can be suggested that an Avian Influenza Virus monitoring should be done continuously over a long period. /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin:0in; mso-para-margin-bottom:.0001pt; text-align:justify; line-height:150%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin;}

  5. Viral and Host Factors Required for Avian H5N1 Influenza A Virus Replication in Mammalian Cells

    Directory of Open Access Journals (Sweden)

    Hong Zhang

    2013-06-01

    Full Text Available Following the initial and sporadic emergence into humans of highly pathogenic avian H5N1 influenza A viruses in Hong Kong in 1997, we have come to realize the potential for avian influenza A viruses to be transmitted directly from birds to humans. Understanding the basic viral and cellular mechanisms that contribute to infection of mammalian species with avian influenza viruses is essential for developing prevention and control measures against possible future human pandemics. Multiple physical and functional cellular barriers can restrict influenza A virus infection in a new host species, including the cell membrane, the nuclear envelope, the nuclear environment, and innate antiviral responses. In this review, we summarize current knowledge on viral and host factors required for avian H5N1 influenza A viruses to successfully establish infections in mammalian cells. We focus on the molecular mechanisms underpinning mammalian host restrictions, as well as the adaptive mutations that are necessary for an avian influenza virus to overcome them. It is likely that many more viral and host determinants remain to be discovered, and future research in this area should provide novel and translational insights into the biology of influenza virus-host interactions.

  6. The special neuraminidase stalk-motif responsible for increased virulence and pathogenesis of H5N1 influenza A virus.

    Directory of Open Access Journals (Sweden)

    Hongbo Zhou

    Full Text Available The variation of highly pathogenic avian influenza H5N1 virus results in gradually increased virulence in poultry, and human cases continue to accumulate. The neuraminidase (NA stalk region of influenza virus varies considerably and may associate with its virulence. The NA stalk region of all N1 subtype influenza A viruses can be divided into six different stalk-motifs, H5N1/2004-like (NA-wt, WSN-like, H5N1/97-like, PR/8-like, H7N1/99-like and H5N1/96-like. The NA-wt is a special NA stalk-motif which was first observed in H5N1 influenza virus in 2000, with a 20-amino acid deletion in the 49(th to 68(th positions of the stalk region. Here we show that there is a gradual increase of the special NA stalk-motif in H5N1 isolates from 2000 to 2007, and notably, the special stalk-motif is observed in all 173 H5N1 human isolates from 2004 to 2007. The recombinant H5N1 virus with the special stalk-motif possesses the highest virulence and pathogenicity in chicken and mice, while the recombinant viruses with the other stalk-motifs display attenuated phenotype. This indicates that the special stalk-motif has contributed to the high virulence and pathogenicity of H5N1 isolates since 2000. The gradually increasing emergence of the special NA stalk-motif in H5N1 isolates, especially in human isolates, deserves attention by all.

  7. New Orf virus (Parapoxvirus recombinant expressing H5 hemagglutinin protects mice against H5N1 and H1N1 influenza A virus.

    Directory of Open Access Journals (Sweden)

    Jörg Rohde

    Full Text Available Previously we demonstrated the versatile utility of the Parapoxvirus Orf virus (ORFV as a vector platform for the development of potent recombinant vaccines. In this study we present the generation of new ORFV recombinants expressing the hemagglutinin (HA or nucleoprotein (NP of the highly pathogenic avian influenza virus (HPAIV H5N1. Correct foreign gene expression was examined in vitro by immunofluorescence, Western blotting and flow cytometry. The protective potential of both recombinants was evaluated in the mouse challenge model. Despite adequate expression of NP, the recombinant D1701-V-NPh5 completely failed to protect mice from lethal challenge. However, the H5 HA-expressing recombinant D1701-V-HAh5n mediated solid protection in a dose-dependent manner. Two intramuscular (i.m. injections of the HA-expressing recombinant protected all animals from lethal HPAIV infection without loss of body weight. Notably, the immunized mice resisted cross-clade H5N1 and heterologous H1N1 (strain PR8 influenza virus challenge. In vivo antibody-mediated depletion of CD4-positive and/or CD8-posititve T-cell subpopulations during immunization and/or challenge infection implicated the relevance of CD4-positive T-cells for induction of protective immunity by D1701-V-HAh5n, whereas the absence of CD8-positive T-cells did not significantly influence protection. In summary, this study validates the potential of the ORFV vectored vaccines also to combat HPAIV.

  8. Quantitative transmission characteristics of different H5 low pathogenic avian influenza viruses in Muscovy ducks.

    Science.gov (United States)

    Niqueux, Éric; Picault, Jean-Paul; Amelot, Michel; Allée, Chantal; Lamandé, Josiane; Guillemoto, Carole; Pierre, Isabelle; Massin, Pascale; Blot, Guillaume; Briand, François-Xavier; Rose, Nicolas; Jestin, Véronique

    2014-01-10

    EU annual serosurveillance programs show that domestic duck flocks have the highest seroprevalence of H5 antibodies, demonstrating the circulation of notifiable avian influenza virus (AIV) according to OIE, likely low pathogenic (LP). Therefore, transmission characteristics of LPAIV within these flocks can help to understand virus circulation and possible risk of propagation. This study aimed at estimating transmission parameters of four H5 LPAIV (three field strains from French poultry and decoy ducks, and one clonal reverse-genetics strain derived from one of the former), using a SIR model to analyze data from experimental infections in SPF Muscovy ducks. The design was set up to accommodate rearing on wood shavings with a low density of 1.6 ducks/m(2): 10 inoculated ducks were housed together with 15 contact-exposed ducks. Infection was monitored by RNA detection on oropharyngeal and cloacal swabs using real-time RT-PCR with a cutoff corresponding to 2-7 EID50. Depending on the strain, the basic reproduction number (R0) varied from 5.5 to 42.7, confirming LPAIV could easily be transmitted to susceptible Muscovy ducks. The lowest R0 estimate was obtained for a H5N3 field strain, due to lower values of transmission rate and duration of infectious period, whereas reverse-genetics derived H5N1 strain had the highest R0. Frequency and intensity of clinical signs were also variable between strains, but apparently not associated with longer infectious periods. Further comparisons of quantitative transmission parameters may help to identify relevant viral genetic markers for early detection of potentially more virulent strains during surveillance of LPAIV. Copyright © 2013 Elsevier B.V. All rights reserved.

  9. Novel Reassortant Influenza A(H5N8) Viruses among Inoculated Domestic and Wild Ducks, South Korea, 2014

    Science.gov (United States)

    Kang, Hyun-Mi; Lee, Eun-Kyoung; Song, Byung-Min; Jeong, Jipseol; Choi, Jun-Gu; Jeong, Joojin; Moon, Oun-Kyong; Yoon, Hachung; Cho, Youngmi; Kang, Young-Myong; Lee, Hee-Soo

    2015-01-01

    An outbreak of highly pathogenic avian influenza, caused by a novel reassortant influenza A (H5N8) virus, occurred among poultry and wild birds in South Korea in 2014. The aim of this study was to evaluate the pathogenesis in and mode of transmission of this virus among domestic and wild ducks. Three of the viruses had similar pathogenicity among infected domestic ducks: the H5N8 viruses were moderately pathogenic (0%–20% mortality rate); in wild mallard ducks, the H5N8 and H5N1 viruses did not cause severe illness or death; viral replication and shedding were greater in H5N8-infected mallards than in H5N1-infected mallards. Identification of H5N8 viruses in birds exposed to infected domestic ducks and mallards indicated that the viruses could spread by contact. We propose active surveillance to support prevention of the spread of this virus among wild birds and poultry, especially domestic ducks. PMID:25625281

  10. Highly Pathogenic H5N1 Avian Influenza Viruses Exhibit Few Barriers to Gene Flow in Vietnam

    Science.gov (United States)

    Carrel, Margaret; Wan, Xiu-Feng; Nguyen, Tung; Emch, Michael

    2013-01-01

    Locating areas where genetic change is inhibited can illuminate underlying processes that drive evolution of pathogens. The persistence of highly pathogenic H5N1 avian influenza in Vietnam since 2003, and the continuous molecular evolution of Vietnamese avian influenza viruses, indicates that local environmental factors are supportive not only of incidence but also of viral adaptation. This article explores whether gene flow is constant across Vietnam, or whether there exist boundary areas where gene flow exhibits discontinuity. Using a dataset of 125 highly pathogenic H5N1 avian influenza viruses, principal components analysis and wombling analysis are used to indicate the location, magnitude, and statistical significance of genetic boundaries. Results show that a small number of geographically minor boundaries to gene flow in highly pathogenic H5N1 avian influenza viruses exist in Vietnam, but that overall there is little division in genetic exchange. This suggests that differences in genetic characteristics of viruses from one region to another are not the result of barriers to H5N1 viral exchange in Vietnam, and that H5N1 avian influenza is able to spread relatively unimpeded across the country. PMID:22350419

  11. Experimental infection of highly pathogenic avian influenza virus H5N1 in black-headed gulls (Chroicocephalus ridibundus)

    OpenAIRE

    Ramis , Antonio; van Amerongen , Geert; van de Bildt , Marco; Leijten , Loneke; Vanderstichel , Raphael; Osterhaus , Albert; Kuiken , Thijs

    2014-01-01

    Historically, highly pathogenic avian influenza viruses (HPAIV) rarely resulted in infection or clinical disease in wild birds. However, since 2002, disease and mortality from natural HPAIV H5N1 infection have been observed in wild birds including gulls. We performed an experimental HPAIV H5N1 infection of black-headed gulls (Chroicocephalus ridibundus) to determine their susceptibility to infection and disease from this virus, pattern of viral shedding, clinical signs, pathological changes a...

  12. Ferrets develop fatal influenza after inhaling small particle aerosols of highly pathogenic avian influenza virus A/Vietnam/1203/2004 (H5N1

    Directory of Open Access Journals (Sweden)

    Sosna William A

    2010-09-01

    Full Text Available Abstract Background There is limited knowledge about the potential routes for H5N1 influenza virus transmission to and between humans, and it is not clear whether humans can be infected through inhalation of aerosolized H5N1 virus particles. Ferrets are often used as a animal model for humans in influenza pathogenicity and transmissibility studies. In this manuscript, a nose-only bioaerosol inhalation exposure system that was recently developed and validated was used in an inhalation exposure study of aerosolized A/Vietnam/1203/2004 (H5N1 virus in ferrets. The clinical spectrum of influenza resulting from exposure to A/Vietnam/1203/2004 (H5N1 through intranasal verses inhalation routes was analyzed. Results Ferrets were successfully infected through intranasal instillation or through inhalation of small particle aerosols with four different doses of Influenza virus A/Vietnam/1203/2004 (H5N1. The animals developed severe influenza encephalomyelitis following intranasal or inhalation exposure to 101, 102, 103, or 104 infectious virus particles per ferret. Conclusions Aerosolized Influenza virus A/Vietnam/1203/2004 (H5N1 is highly infectious and lethal in ferrets. Clinical signs appeared earlier in animals infected through inhalation of aerosolized virus compared to those infected through intranasal instillation.

  13. Virulence of H5N1 Influenza Virus in Cattle Egrets (Bubulcus Ibis)

    DEFF Research Database (Denmark)

    Phuong, Do Quy; Dung, Nguyen Tien; Jørgensen, Poul Henrik

    2011-01-01

    for insect control in households. In this study, six Cattle Egrets were experimentally infected intranasally with highly pathogenic avian influenza (AI) A/duck/Vietnam/40D/04 (H5N1) to investigate a possible epidemiologic role for Cattle Egrets in outbreaks of H5N1 AI in Vietnam. The Cattle Egrets were...

  14. Isolation and identification of highly pathogenic avian influenza virus subtype H5N1 in peafowl (Pavo cristatus).

    Science.gov (United States)

    Ismail, Mahmoud Moussa; Khan, Owais Ahmed; Cattoli, Giovanni; Lu, Huaguang

    2010-03-01

    An outbreak of highly pathogenic avian influenza (HPAI) virus subtype H5N1 was first diagnosed in a "backyard" flock of peafowl (Pavo cristatus) raised on palace premises in the Kingdom of Saudi Arabia in December 3, 2007. The flock consisted of 40 peafowl, and their ages ranged from 3 to 5 years old. Affected birds suffered from depression, anorexia, and white diarrhea. Four dead birds were submitted for HPAI diagnosis at the Central Veterinary Diagnostic Laboratory in Riyadh. Brain and liver tissues and tracheal and cloacal swabs were taken from the dead birds and processed for a real-time reverse transcriptase (RT)-PCR test and virus isolation in specific-pathogen-free embryonating chicken eggs. The H5N1 subtype of avian influenza virus was isolated from the four dead birds and identified by a real-time RT-PCR before and after egg inoculation. The virus isolates were characterized as HPAI H5N1 virus by sequencing analysis. Phylogenetic comparisons revealed that the H5N1 viruses isolated from peafowl belong to the genetic clade 2.2 according to the World Health Organization nomenclature. The peafowl H5N1 virus falls into 2.2.2 sublineage II and clusters with the H5N1 viruses isolated from poultry in Saudi Arabia in 2007-08.

  15. Influenza A H5N1 clade 2.3.4 virus with a different antiviral susceptibility profile replaced clade 1 virus in humans in northern Vietnam

    NARCIS (Netherlands)

    Le, Mai T. Q.; Wertheim, Heiman F. L.; Nguyen, Hien D.; Taylor, Walter; Hoang, Phuong V. M.; Vuong, Cuong D.; Nguyen, Hang L. K.; Nguyen, Ha H.; Nguyen, Thai Q.; Nguyen, Trung V.; van, Trang D.; Ngoc, Bich T.; Bui, Thinh N.; Nguyen, Binh G.; Nguyen, Liem T.; Luong, San T.; Phan, Phuc H.; Pham, Hung V.; Nguyen, Tung; Fox, Annette; Nguyen, Cam V.; Do, Ha Q.; Crusat, Martin; Farrar, Jeremy; Nguyen, Hien T.; de Jong, Menno D.; Horby, Peter

    2008-01-01

    BACKGROUND: Prior to 2007, highly pathogenic avian influenza (HPAI) H5N1 viruses isolated from poultry and humans in Vietnam were consistently reported to be clade 1 viruses, susceptible to oseltamivir but resistant to amantadine. Here we describe the re-emergence of human HPAI H5N1 virus infections

  16. Evolution of highly pathogenic H5N1 avian influenza viruses in Vietnam between 2001 and 2007.

    Directory of Open Access Journals (Sweden)

    Xiu-Feng Wan

    Full Text Available Highly pathogenic avian influenza (HPAI H5N1 viruses have caused dramatic economic losses to the poultry industry of Vietnam and continue to pose a serious threat to public health. As of June 2008, Vietnam had reported nearly one third of worldwide laboratory confirmed human H5N1 infections. To better understand the emergence, spread and evolution of H5N1 in Vietnam we studied over 300 H5N1 avian influenza viruses isolated from Vietnam since their first detection in 2001. Our phylogenetic analyses indicated that six genetically distinct H5N1 viruses were introduced into Vietnam during the past seven years. The H5N1 lineage that evolved following the introduction in 2003 of the A/duck/Hong Kong/821/2002-like viruses, with clade 1 hemagglutinin (HA, continued to predominate in southern Vietnam as of May 2007. A virus with a clade 2.3.4 HA newly introduced into northern Vietnam in 2007, reassorted with pre-existing clade 1 viruses, resulting in the emergence of novel genotypes with neuraminidase (NA and/or internal gene segments from clade 1 viruses. A total of nine distinct genotypes have been present in Vietnam since 2001, including five that were circulating in 2007. At least four of these genotypes appear to have originated in Vietnam and represent novel H5N1 viruses not reported elsewhere. Geographic and temporal analyses of H5N1 infection dynamics in poultry suggest that the majority of viruses containing new genes were first detected in northern Vietnam and subsequently spread to southern Vietnam after reassorting with pre-existing local viruses in northern Vietnam. Although the routes of entry and spread of H5N1 in Vietnam remain speculative, enhanced poultry import controls and virologic surveillance efforts may help curb the entry and spread of new HPAI viral genes.

  17. Evaluation of twenty rapid antigen tests for the detection of human influenza A H5N1, H3N2, H1N1, and B viruses.

    Science.gov (United States)

    Taylor, Janette; McPhie, Kenneth; Druce, Julian; Birch, Chris; Dwyer, Dominic E

    2009-11-01

    Twenty rapid antigen assays were compared for their ability to detect influenza using dilutions of virus culture supernatants from human isolates of influenza A H5N1 (clade 1 and 2 strains), H3N2 and H1N1 viruses, and influenza B. There was variation amongst the rapid antigen assays in their ability to detect different influenza viruses. Six of the 12 assays labeled as distinguishing between influenza A and B had comparable analytical sensitivities for detecting both influenza A H5N1 strains, although their ability to detect influenza A H3N2 and H1N1 strains varied. The two assays claiming H5 specificity did not detect either influenza A H5N1 strains, and the two avian influenza-specific assays detected influenza A H5N1, but missed some influenza A H3N2 virus supernatants. Clinical trials of rapid antigen tests for influenza A H5N1 are limited. For use in a pandemic where novel influenza strains are circulating (such as the current novel influenza A H1N1 09 virus), rapid antigen tests should ideally have comparable sensitivity and specificity for the new strains as for co-circulating seasonal influenza strains.

  18. In vitro evolution of H5N1 avian influenza virus toward human-type receptor specificity

    DEFF Research Database (Denmark)

    Chen, Li-Mei; Blixt, Klas Ola; Stevens, James

    2012-01-01

    Acquisition of a2-6 sialoside receptor specificity by a2-3 specific highly-pathogenic avian influenza viruses (H5N1) is thought to be a prerequisite for efficient transmission in humans. By in vitro selection for binding a2-6 sialosides, we identified four variant viruses with amino acid....... Unlike the wild type H5N1, this mutant virus was transmitted by direct contact in the ferret model although not by airborne respiratory droplets. However, a reassortant virus with the mutant hemagglutinin, a human N2 neuraminidase and internal genes from an H5N1 virus was partially transmitted via...... respiratory droplets. The complex changes required for airborne transmissibility in ferrets suggest that extensive evolution is needed for H5N1 transmissibility in humans....

  19. Influence of Novel Highly Pathogenic Avian Influenza A (H5N1 Virus Infection on Migrating Whooper Swans Fecal Microbiota

    Directory of Open Access Journals (Sweden)

    Na Zhao

    2018-02-01

    Full Text Available The migration of wild birds plays an important role in the transmission and spread of H5 highly pathogenic avian influenza (HPAI virus, posing a severe risk to animal and human health. Substantial evidence suggests that altered gut microbial community is implicated in the infection of respiratory influenza virus. However, the influence of H5N1 infection in gut microbiota of migratory birds remains unknown. In January 2015, a novel recombinant H5N1 virus emerged and killed about 100 migratory birds, mainly including whooper swans in Sanmenxia Reservoir Area of China. Here, we describe the first fecal microbiome diversity study of H5N1-infected migratory birds. By investigating the influence of H5N1 infection on fecal bacterial communities in infected and uninfected individuals, we found that H5N1 infection shaped the gut microbiota composition by a difference in the dominance of some genera, such as Aeromonas and Lactobacillus. We also found a decreased α diversity and increased β diversity in infectious individuals. Our results highlight that increases in changes in pathogen-containing gut communities occur when individuals become infected with H5N1. Our study may provide the first evidence that there are statistical association among H5N1 presence and fecal microbiota compositional shifts, and properties of the fecal microbiota may serve as the risk of gut-linked disease in migrates with H5N1 and further aggravate the disease transmission.

  20. Influence of Novel Highly Pathogenic Avian Influenza A (H5N1) Virus Infection on Migrating Whooper Swans Fecal Microbiota.

    Science.gov (United States)

    Zhao, Na; Wang, Supen; Li, Hongyi; Liu, Shelan; Li, Meng; Luo, Jing; Su, Wen; He, Hongxuan

    2018-01-01

    The migration of wild birds plays an important role in the transmission and spread of H5 highly pathogenic avian influenza (HPAI) virus, posing a severe risk to animal and human health. Substantial evidence suggests that altered gut microbial community is implicated in the infection of respiratory influenza virus. However, the influence of H5N1 infection in gut microbiota of migratory birds remains unknown. In January 2015, a novel recombinant H5N1 virus emerged and killed about 100 migratory birds, mainly including whooper swans in Sanmenxia Reservoir Area of China. Here, we describe the first fecal microbiome diversity study of H5N1-infected migratory birds. By investigating the influence of H5N1 infection on fecal bacterial communities in infected and uninfected individuals, we found that H5N1 infection shaped the gut microbiota composition by a difference in the dominance of some genera, such as Aeromonas and Lactobacillus . We also found a decreased α diversity and increased β diversity in infectious individuals. Our results highlight that increases in changes in pathogen-containing gut communities occur when individuals become infected with H5N1. Our study may provide the first evidence that there are statistical association among H5N1 presence and fecal microbiota compositional shifts, and properties of the fecal microbiota may serve as the risk of gut-linked disease in migrates with H5N1 and further aggravate the disease transmission.

  1. Avian Influenza A (H5N1)

    Centers for Disease Control (CDC) Podcasts

    2009-05-27

    In this podcast, CDC's Dr. Tim Uyeki discusses H5N1, a subtype of influenza A virus. This highly pathogenic H5N1 virus doesn't usually infect people, although some rare infections with H5N1 viruses have occurred in humans. We need to use a comprehensive strategy to prevent the spread of H5N1 virus among birds, including having human health and animal health work closely together.  Created: 5/27/2009 by Emerging Infectious Diseases.   Date Released: 5/27/2009.

  2. Protective Efficacy of Recombinant Turkey Herpes Virus (rHVT-H5) and Inactivated H5N1 Vaccines in Commercial Mulard Ducks against the Highly Pathogenic Avian Influenza (HPAI) H5N1 Clade 2.2.1 Virus.

    Science.gov (United States)

    Kilany, Walid H; Safwat, Marwa; Mohammed, Samy M; Salim, Abdullah; Fasina, Folorunso Oludayo; Fasanmi, Olubunmi G; Shalaby, Azhar G; Dauphin, Gwenaelle; Hassan, Mohammed K; Lubroth, Juan; Jobre, Yilma M

    2016-01-01

    In Egypt, ducks kept for commercial purposes constitute the second highest poultry population, at 150 million ducks/year. Hence, ducks play an important role in the introduction and transmission of avian influenza (AI) in the Egyptian poultry population. Attempts to control outbreaks include the use of vaccines, which have varying levels of efficacy and failure. To date, the effects of vaccine efficacy has rarely been determined in ducks. In this study, we evaluated the protective efficacy of a live recombinant vector vaccine based on a turkey Herpes Virus (HVT) expressing the H5 gene from a clade 2.2 H5N1 HPAIV strain (A/Swan/Hungary/499/2006) (rHVT-H5) and a bivalent inactivated H5N1 vaccine prepared from clade 2.2.1 and 2.2.1.1 H5N1 seeds in Mulard ducks. A 0.3ml/dose subcutaneous injection of rHVT-H5 vaccine was administered to one-day-old ducklings (D1) and another 0.5ml/dose subcutaneous injection of the inactivated MEFLUVAC was administered at 7 days (D7). Four separate challenge experiments were conducted at Days 21, 28, 35 and 42, in which all the vaccinated ducks were challenged with 106EID50/duck of H5N1 HPAI virus (A/chicken/Egypt/128s/2012(H5N1) (clade 2.2.1) via intranasal inoculation. Maternal-derived antibody regression and post-vaccination antibody immune responses were monitored weekly. Ducks vaccinated at 21, 28, 35 and 42 days with the rHVT-H5 and MEFLUVAC vaccines were protected against mortality (80%, 80%, 90% and 90%) and (50%, 70%, 80% and 90%) respectively, against challenges with the H5N1 HPAI virus. The amount of viral shedding and shedding rates were lower in the rHVT-H5 vaccine groups than in the MEFLUVAC groups only in the first two challenge experiments. However, the non-vaccinated groups shed significantly more of the virus than the vaccinated groups. Both rHVT-H5 and MEFLUVAC provide early protection, and rHVT-H5 vaccine in particular provides protection against HPAI challenge.

  3. Spatiotemporal Structure of Molecular Evolution of H5N1 Highly Pathogenic Avian Influenza Viruses in Vietnam

    OpenAIRE

    Carrel, Margaret A.; Emch, Michael; Jobe, R. Todd; Moody, Aaron; Wan, Xiu-Feng

    2010-01-01

    Background Vietnam is one of the countries most affected by outbreaks of H5N1 highly pathogenic avian influenza viruses. First identified in Vietnam in poultry in 2001 and in humans in 2004, the virus has since caused 111 cases and 56 deaths in humans. In 2003/2004 H5N1 outbreaks, nearly the entire poultry population of Vietnam was culled. Our earlier study (Wan et al., 2008, PLoS ONE, 3(10): e3462) demonstrated that there have been at least six independent H5N1 introductions into Vietnam and...

  4. Highly Pathogenic Avian Influenza A(H5N1) Virus Struck Migratory Birds in China in 2015.

    Science.gov (United States)

    Bi, Yuhai; Zhang, Zhenjie; Liu, Wenjun; Yin, Yanbo; Hong, Jianmin; Li, Xiangdong; Wang, Haiming; Wong, Gary; Chen, Jianjun; Li, Yunfeng; Ru, Wendong; Gao, Ruyi; Liu, Di; Liu, Yingxia; Zhou, Boping; Gao, George F; Shi, Weifeng; Lei, Fumin

    2015-08-11

    Approximately 100 migratory birds, including whooper swans and pochards, were found dead in the Sanmenxia Reservoir Area of China during January 2015. The causative agent behind this outbreak was identified as H5N1 highly pathogenic avian influenza virus (HPAIV). Genetic and phylogenetic analyses revealed that this Sanmenxia H5N1 virus was a novel reassortant, possessing a Clade 2.3.2.1c HA gene and a H9N2-derived PB2 gene. Sanmenxia Clade 2.3.2.1c-like H5N1 viruses possess the closest genetic identity to A/Alberta/01/2014 (H5N1), which recently caused a fatal respiratory infection in Canada with signs of meningoencephalitis, a highly unusual symptom with influenza infections in humans. Furthermore, this virus was shown to be highly pathogenic to both birds and mammals, and demonstrate tropism for the nervous system. Due to the geographical location of Sanmenxia, these novel H5N1 viruses also have the potential to be imported to other regions through the migration of wild birds, similar to the H5N1 outbreak amongst migratory birds in Qinghai Lake during 2005. Therefore, further investigation and monitoring is required to prevent this novel reassortant virus from becoming a new threat to public health.

  5. Identifying antigenicity associated sites in highly pathogenic H5N1 influenza virus hemagglutinin by using sparse learning

    OpenAIRE

    Cai, Zhipeng; Ducatez, Mariette F.; Yang, Jialiang; Zhang, Tong; Long, Li-Ping; Boon, Adrianus C.; Webby, Richard J.; Wan, Xiu-Feng

    2012-01-01

    Since the isolation of A/goose/Guangdong/1/1996 (H5N1) in farmed geese in southern China, highly pathogenic H5N1 avian influenza viruses have posed a continuous threat to both public and animal health. The non-synonymous mutation of the H5 hemagglutinin gene has resulted in antigenic drift, leading to difficulties in both clinical diagnosis and vaccine strain selection. Characterizing H5N1’s antigenic profiles would help resolve these problems. In this study, a novel sparse learning method wa...

  6. Identifying antigenicity-associated sites in highly pathogenic H5N1 influenza virus hemagglutinin by using sparse learning.

    OpenAIRE

    Cai, Zhipeng; Yang, Jialiang; Zhang, Tong; Long, Li-Ping; Boon, Adrianus C; Webby, Richard J; Wan, Xiu-Feng

    2012-01-01

    Since the isolation of A/goose/Guangdong/1/1996 (H5N1) in farmed geese in southern China, highly pathogenic H5N1 avian influenza viruses have posed a continuous threat to both public and animal health. The non-synonymous mutation of the H5 hemagglutinin (HA) gene has resulted in antigenic drift, leading to difficulties in both clinical diagnosis and vaccine strain selection. Characterizing H5N1's antigenic profiles would help resolve these problems. In this study, a novel sparse learning meth...

  7. Vaccination with recombinant RNA replicon particles protects chickens from H5N1 highly pathogenic avian influenza virus.

    Directory of Open Access Journals (Sweden)

    Stefan J Halbherr

    Full Text Available Highly pathogenic avian influenza viruses (HPAIV of subtype H5N1 not only cause a devastating disease in domestic chickens and turkeys but also pose a continuous threat to public health. In some countries, H5N1 viruses continue to circulate and evolve into new clades and subclades. The rapid evolution of these viruses represents a problem for virus diagnosis and control. In this work, recombinant vesicular stomatitis virus (VSV vectors expressing HA of subtype H5 were generated. To comply with biosafety issues the G gene was deleted from the VSV genome. The resulting vaccine vector VSV*ΔG(HA was propagated on helper cells providing the VSV G protein in trans. Vaccination of chickens with a single intramuscular dose of 2×10⁸ infectious replicon particles without adjuvant conferred complete protection from lethal H5N1 infection. Subsequent application of the same vaccine strongly boosted the humoral immune response and completely prevented shedding of challenge virus and transmission to sentinel birds. The vaccine allowed serological differentiation of infected from vaccinated animals (DIVA by employing a commercially available ELISA. Immunized chickens produced antibodies with neutralizing activity against multiple H5 viruses representing clades 1, 2.2, 2.5, and low-pathogenic avian influenza viruses (classical clade. Studies using chimeric H1/H5 hemagglutinins showed that the neutralizing activity was predominantly directed against the globular head domain. In summary, these results suggest that VSV replicon particles are safe and potent DIVA vaccines that may help to control avian influenza viruses in domestic poultry.

  8. Highly pathogenic avian influenza A (H5N1) virus in wildlife: diagnostics, epidemiology and molecular characteristics

    NARCIS (Netherlands)

    Keawcharoen, J.

    2010-01-01

    Since 2003, highly pathogenic avian influenza virus subtype H5N1 outbreaks have been reported in Southeast Asia causing high mortality in poultry and have also been found to cross the species barrier infecting human and other mammalian species. Thailand is one of the countries severely affected by

  9. Exploring contacts facilitating transmission of influenza A(H5N1) virus between poultry farms in West Java, Indonesia

    NARCIS (Netherlands)

    Wibawa, Hendra; Karo-Karo, Desniwaty; Pribadi, Eko Sugeng; Bouma, Annemarie; Bodewes, Rogier; Vernooij, Hans; Diyantoro,; Sugama, Agus; Muljono, David H.; Koch, Guus; Tjatur Rasa, Fadjar Sumping; Stegeman, Arjan

    2018-01-01

    Highly pathogenic avian influenza virus (HPAIV) H5N1 has been reported in Asia, including Indonesia since 2003. Although several risk factors related to the HPAIV outbreaks in poultry in Indonesia have been identified, little is known of the contact structure of farms of different poultry production

  10. Experimental infection of highly pathogenic avian influenza virus H5N1 in black-headed gulls (Chroicocephalus ridibundus)

    NARCIS (Netherlands)

    A. Ramis (Antonio); G. van Amerongen (Geert); M.W.G. van de Bildt (Marco); L.M.E. Leijten (Lonneke); R. Vanderstichel (R.); A.D.M.E. Osterhaus (Albert); T. Kuiken (Thijs)

    2014-01-01

    textabstractHistorically, highly pathogenic avian influenza viruses (HPAIV) rarely resulted in infection or clinical disease in wild birds. However, since 2002, disease and mortality from natural HPAIV H5N1 infection have been observed in wild birds including gulls. We performed an experimental

  11. Experimental infection of swans and geese with highly pathogenic avian influenza virus (H5N1) of Asian lineage.

    Science.gov (United States)

    Brown, Justin D; Stallknecht, David E; Swayne, David E

    2008-01-01

    The role of wild birds in the epidemiology of the Asian lineage highly pathogenic avian influenza (HPAI) virus subtype H5N1 epizootic and their contribution to the spread of the responsible viruses in Eurasia and Africa are unclear. To better understand the potential role of swans and geese in the epidemiology of this virus, we infected 4 species of swans and 2 species of geese with an HPAI virus of Asian lineage recovered from a whooper swan in Mongolia in 2005, A/whooper swan/Mongolia/244/2005 (H5N1). The highest mortality rates were observed in swans, and species-related differences in clinical illness and viral shedding were evident. These results suggest that the potential for HPAI (H5N1) viral shedding and the movement of infected birds may be species-dependent and can help explain observed deaths associated with HPAI (H5N1) infection in anseriforms in Eurasia.

  12. A duplex real-time RT-PCR assay for detecting H5N1 avian influenza virus and pandemic H1N1 influenza virus

    Directory of Open Access Journals (Sweden)

    Qin E-de

    2010-06-01

    Full Text Available Abstract A duplex real-time reverse transcriptase polymerase chain reaction (RT-PCR assay was improved for simultaneous detection of highly pathogenic H5N1 avian influenza virus and pandemic H1N1 (2009 influenza virus, which is suitable for early diagnosis of influenza-like patients and for epidemiological surveillance. The sensitivity of this duplex real-time RT-PCR assay was 0.02 TCID50 (50% tissue culture infective dose for H5N1 and 0.2 TCID50 for the pandemic H1N1, which was the same as that of each single-target RT-PCR for pandemic H1N1 and even more sensitive for H5N1 with the same primers and probes. No cross reactivity of detecting other subtype influenza viruses or respiratory tract viruses was observed. Two hundred and thirty-six clinical specimens were tested by comparing with single real-time RT-PCR and result from the duplex assay was 100% consistent with the results of single real-time RT-PCR and sequence analysis.

  13. Novel Polymerase Gene Mutations for Human Adaptation in Clinical Isolates of Avian H5N1 Influenza Viruses.

    Directory of Open Access Journals (Sweden)

    Yasuha Arai

    2016-04-01

    Full Text Available A major determinant in the change of the avian influenza virus host range to humans is the E627K substitution in the PB2 polymerase protein. However, the polymerase activity of avian influenza viruses with a single PB2-E627K mutation is still lower than that of seasonal human influenza viruses, implying that avian viruses require polymerase mutations in addition to PB2-627K for human adaptation. Here, we used a database search of H5N1 clade 2.2.1 virus sequences with the PB2-627K mutation to identify other polymerase adaptation mutations that have been selected in infected patients. Several of the mutations identified acted cooperatively with PB2-627K to increase viral growth in human airway epithelial cells and mouse lungs. These mutations were in multiple domains of the polymerase complex other than the PB2-627 domain, highlighting a complicated avian-to-human adaptation pathway of avian influenza viruses. Thus, H5N1 viruses could rapidly acquire multiple polymerase mutations that function cooperatively with PB2-627K in infected patients for optimal human adaptation.

  14. Full-Genome Analysis of Avian Influenza A(H5N1) Virus from a Human, North America, 2013

    Science.gov (United States)

    Pabbaraju, Kanti; Tellier, Raymond; Wong, Sallene; Li, Yan; Bastien, Nathalie; Tang, Julian W.; Drews, Steven J.; Jang, Yunho; Davis, C. Todd; Tipples, Graham A.

    2014-01-01

    Full-genome analysis was conducted on the first isolate of a highly pathogenic avian influenza A(H5N1) virus from a human in North America. The virus has a hemagglutinin gene of clade 2.3.2.1c and is a reassortant with an H9N2 subtype lineage polymerase basic 2 gene. No mutations conferring resistance to adamantanes or neuraminidase inhibitors were found. PMID:24755439

  15. A low pathogenic H5N2 influenza virus isolated in Taiwan acquired high pathogenicity by consecutive passages in chickens.

    OpenAIRE

    Soda, Kosuke; Cheng, Ming-Chu; Yoshida, Hiromi; Endo, Mayumi; Lee, Shu-Hwae; Okamatsu, Masatoshi; Sakoda, Yoshihiro; Wang, Ching-Ho; Kida, Hiroshi

    2011-01-01

    H5N2 viruses were isolated from cloacal swab samples of apparently healthy chickens in Taiwan in 2003 and 2008 during surveillance of avian influenza. Each of the viruses was eradicated by stamping out. The official diagnosis report indicated that the Intravenous Pathogenicity Indexes (IVPIs) of the isolates were 0.00 and 0.89, respectively, indicating that these were low pathogenic strains, although the hemagglutinin of the strain isolated in 2008 (Taiwan08) had multibasic amino acid residue...

  16. An enzyme-linked immunosorbent assay for detection of avian influenza virus subtypes H5 and H7 antibodies

    DEFF Research Database (Denmark)

    Jensen, Trine Hammer; Ajjouri, Gitte; Handberg, Kurt

    2013-01-01

    subtypes H5 and H7. The ELISAs were evaluated with polyclonal chicken anti-AIV antibodies against AIV subtypes: H1N2, H5N2, H5N7, H7N1, H7N7, H9N9, H10N4 and H16N3. RESULTS: Both the H5 and H7 ELISA proved to have a high sensitivity and specificity and the ELISAs detected H5 and H7 antibodies earlier......BACKGROUND: Avian influenza virus (AIV) subtypes H5 and H7 attracts particular attention because of the risk of their potential pathogenicity in poultry. The haemagglutination inhibition (HI) test is widely used as subtype specific test for serological diagnostics despite the laborious nature...

  17. Identification of Two novel reassortant avian influenza a (H5N6) viruses in whooper swans in Korea, 2016.

    Science.gov (United States)

    Jeong, Jipseol; Woo, Chanjin; Ip, Hon S; An, Injung; Kim, Youngsik; Lee, Kwanghee; Jo, Seong-Deok; Son, Kidong; Lee, Saemi; Oem, Jae-Ku; Wang, Seung-Jun; Kim, Yongkwan; Shin, Jeonghwa; Sleeman, Jonathan; Jheong, Weonhwa

    2017-03-21

    On November 20, 2016 two novel strains of H5N6 highly pathogenic avian influenza virus (HPAIVs) were isolated from three whooper swans (Cygnus cygnus) at Gangjin Bay in South Jeolla province, South Korea. Identification of HPAIVs in wild birds is significant as there is a potential risk of transmission of these viruses to poultry and humans. Phylogenetic analysis revealed that Gangjin H5N6 viruses classified into Asian H5 clade 2.3.4.4 lineage and were distinguishable from H5N8 and H5N1 HPAIVs previously isolated in Korea. With the exception of the polymerase acidic (PA) gene, the viruses were most closely related to A/duck/Guangdong/01.01SZSGXJK005-Y/2016 (H5N6) (98.90 ~ 99.74%). The PA genes of the two novel Gangjin H5N6 viruses were most closely related to AIV isolates previously characterized from Korea, A/hooded crane/Korea/1176/2016 (H1N1) (99.16%) and A/environment/Korea/W133/2006 (H7N7) (98.65%). The lack of more recent viruses to A/environment/Korea/W133/2006 (H7N7) indicates the need for analysis of recent wild bird AIVs isolated in Korea because they might provide further clues as to the origin of these novel reassortant H5N6 viruses. Although research on the origins and epidemiology of these infections is ongoing, the most likely route of infection for the whooper swans was through direct or indirect contact with reassortant viruses shed by migratory wild birds in Korea. As H5N6 HPAIVs can potentially be transmitted to poultry and humans, continuous monitoring of AIVs among wild birds will help to mitigate this risk.

  18. Spatiotemporal structure of molecular evolution of H5N1 highly pathogenic avian influenza viruses in Vietnam.

    Science.gov (United States)

    Carrel, Margaret A; Emch, Michael; Jobe, R Todd; Moody, Aaron; Wan, Xiu-Feng

    2010-01-08

    Vietnam is one of the countries most affected by outbreaks of H5N1 highly pathogenic avian influenza viruses. First identified in Vietnam in poultry in 2001 and in humans in 2004, the virus has since caused 111 cases and 56 deaths in humans. In 2003/2004 H5N1 outbreaks, nearly the entire poultry population of Vietnam was culled. Our earlier study (Wan et al., 2008, PLoS ONE, 3(10): e3462) demonstrated that there have been at least six independent H5N1 introductions into Vietnam and there were nine newly emerged reassortants from 2001 to 2007 in Vietnam. H5N1 viruses in Vietnam cluster distinctly around Hanoi and Ho Chi Minh City. However, the nature of the relationship between genetic divergence and geographic patterns is still unclear. In this study, we hypothesized that genetic distances between H5N1 viruses in Vietnam are correlated with geographic distances, as the result of distinct population and environment patterns along Vietnam's long north to south longitudinal extent. Based on this hypothesis, we combined spatial statistical methods with genetic analytic techniques and explicitly used geographic space to explore genetic evolution of H5N1 highly pathogenic avian influenza viruses at the sub-national scale in Vietnam. Our dataset consisted of 125 influenza viruses (with whole genome sets) isolated in Vietnam from 2003 to 2007. Our results document the significant effect of space and time on genetic evolution and the rise of two regional centers of genetic mixing by 2007. These findings give insight into processes underlying viral evolution and suggest that genetic differentiation is associated with the distance between concentrations of human and poultry populations around Hanoi and Ho Chi Minh City. The results show that genetic evolution of H5N1 viruses in Vietnamese domestic poultry is highly correlated with the location and spread of those viruses in geographic space. This correlation varies by scale, time, and gene, though a classic isolation by

  19. Spatiotemporal structure of molecular evolution of H5N1 highly pathogenic avian influenza viruses in Vietnam.

    Directory of Open Access Journals (Sweden)

    Margaret A Carrel

    2010-01-01

    Full Text Available Vietnam is one of the countries most affected by outbreaks of H5N1 highly pathogenic avian influenza viruses. First identified in Vietnam in poultry in 2001 and in humans in 2004, the virus has since caused 111 cases and 56 deaths in humans. In 2003/2004 H5N1 outbreaks, nearly the entire poultry population of Vietnam was culled. Our earlier study (Wan et al., 2008, PLoS ONE, 3(10: e3462 demonstrated that there have been at least six independent H5N1 introductions into Vietnam and there were nine newly emerged reassortants from 2001 to 2007 in Vietnam. H5N1 viruses in Vietnam cluster distinctly around Hanoi and Ho Chi Minh City. However, the nature of the relationship between genetic divergence and geographic patterns is still unclear.In this study, we hypothesized that genetic distances between H5N1 viruses in Vietnam are correlated with geographic distances, as the result of distinct population and environment patterns along Vietnam's long north to south longitudinal extent. Based on this hypothesis, we combined spatial statistical methods with genetic analytic techniques and explicitly used geographic space to explore genetic evolution of H5N1 highly pathogenic avian influenza viruses at the sub-national scale in Vietnam. Our dataset consisted of 125 influenza viruses (with whole genome sets isolated in Vietnam from 2003 to 2007. Our results document the significant effect of space and time on genetic evolution and the rise of two regional centers of genetic mixing by 2007. These findings give insight into processes underlying viral evolution and suggest that genetic differentiation is associated with the distance between concentrations of human and poultry populations around Hanoi and Ho Chi Minh City.The results show that genetic evolution of H5N1 viruses in Vietnamese domestic poultry is highly correlated with the location and spread of those viruses in geographic space. This correlation varies by scale, time, and gene, though a classic

  20. Contact variables for exposure to avian influenza H5N1 virus at the human-animal interface.

    Science.gov (United States)

    Rabinowitz, P; Perdue, M; Mumford, E

    2010-06-01

    Although the highly pathogenic avian influenza H5N1 virus continues to cause infections in both avian and human populations, the specific zoonotic risk factors remain poorly understood. This review summarizes available evidence regarding types of contact associated with transmission of H5N1 virus at the human-animal interface. A systematic search of the published literature revealed five analytical studies and 15 case reports describing avian influenza transmission from animals to humans for further review. Risk factors identified in analytical studies were compared, and World Health Organization-confirmed cases, identified in case reports, were classified according to type of contact reported using a standardized algorithm. Although cases were primarily associated with direct contact with sick/unexpectedly dead birds, some cases reported only indirect contact with birds or contaminated environments or contact with apparently healthy birds. Specific types of contacts or activities leading to exposure could not be determined from data available in the publications reviewed. These results support previous reports that direct contact with sick birds is not the only means of human exposure to avian influenza H5N1 virus. To target public health measures and disease awareness messaging for reducing the risk of zoonotic infection with avian influenza H5N1 virus, the specific types of contacts and activities leading to transmission need to be further understood. The role of environmental virus persistence, shedding of virus by asymptomatic poultry and disease pathophysiology in different avian species relative to human zoonotic risk, as well as specific modes of zoonotic transmission, should be determined.

  1. Isolation and characterization of virus of highly pathogenic avian influenza H5 subtype of chicken from outbreaks in Indonesia

    Directory of Open Access Journals (Sweden)

    Agus Wiyono

    2004-03-01

    Full Text Available A study on the isolation and characterization of Highly Pathogenic Avian Influenza of chicken from outbreaks in Indonesia was conducted at Indonesian Research Institute for Veterinary Science. Outbreaks of avian disease had been reported in Indonesia since August 2003 affecting commercial layer, broiler, quail, and ostrich and also native chicken with showing clinical signs such as cyanosis of wattle and comb, nasal discharges and hypersalivation, subcutaneous ptechiae on foot and leg, diarre and sudden high mortality. The aim of this study is to isolate and characterize the causal agent of the disease. Samples of serum, feather follicle, tracheal swab, as well as organs of proventriculus, intestine, caecal tonsil, trachea and lungs were collected from infected animals. Serum samples were tested haemaglutination/haemaglutination inhibition to Newcastle Disease and Egg Drop Syndrome viruses. Isolation of virus of the causal agent of the outbreak was conducted from samples of feather follicle, tracheal swab, and organs using 11 days old specific pathogen free (SPF embryonated eggs. The isolated viruses were then characterised by agar gel precipitation test using swine influenza reference antisera, by haemaglutination inhibition using H1 to H15 reference antisera, and by electron microscope examination. The pathogenicity of the viruses was confirmed by intravenous pathogenicity index test and its culture in Chicken Embryo Fibroblast primary cell culture without addition of trypsin. The study revealed that the causative agent of the outbreaks of avian disease in Indonesia was avian influenza H5 subtype virus based upon serological tests, virus isolation and characterization using swine influenza reference antisera, and electron microscope examination. While subtyping of the viruses using H1 to H15 reference antisera suggested that the virus is very likely to be an avian influenza H5N1 subtype virus. The pathogenicity test confirmed that the viruses

  2. An induced pocket for the binding of potent fusion inhibitor CL-385319 with H5N1 influenza virus hemagglutinin.

    Directory of Open Access Journals (Sweden)

    Runming Li

    Full Text Available The influenza glycoprotein hemagglutinin (HA plays crucial roles in the early stage of virus infection, including receptor binding and membrane fusion. Therefore, HA is a potential target for developing anti-influenza drugs. Recently, we characterized a novel inhibitor of highly pathogenic H5N1 influenza virus, CL-385319, which specifically inhibits HA-mediated viral entry. Studies presented here identified the critical binding residues for CL-385319, which clustered in the stem region of the HA trimer by site-directed mutagenesis. Extensive computational simulations, including molecular docking, molecular dynamics simulations, molecular mechanics generalized Born surface area (MM_GBSA calculations, charge density and Laplacian calculations, have been carried out to uncover the detailed molecular mechanism that underlies the binding of CL-385319 to H5N1 influenza virus HA. It was found that the recognition and binding of CL-385319 to HA proceeds by a process of "induced fit" whereby the binding pocket is formed during their interaction. Occupation of this pocket by CL-385319 stabilizes the neutral pH structure of hemagglutinin, thus inhibiting the conformational rearrangements required for membrane fusion. This "induced fit" pocket may be a target for structure-based design of more potent influenza fusion inhibitors.

  3. Comparative pathogenesis of an avian H5N2 and a swine H1N1 influenza virus in pigs.

    Directory of Open Access Journals (Sweden)

    Annebel De Vleeschauwer

    2009-08-01

    Full Text Available Pigs are considered intermediate hosts for the transmission of avian influenza viruses (AIVs to humans but the basic organ pathogenesis of AIVs in pigs has been barely studied. We have used 42 four-week-old influenza naive pigs and two different inoculation routes (intranasal and intratracheal to compare the pathogenesis of a low pathogenic (LP H5N2 AIV with that of an H1N1 swine influenza virus. The respiratory tract and selected extra-respiratory tissues were examined for virus replication by titration, immunofluorescence and RT-PCR throughout the course of infection. Both viruses caused a productive infection of the entire respiratory tract and epithelial cells in the lungs were the major target. Compared to the swine virus, the AIV produced lower virus titers and fewer antigen positive cells at all levels of the respiratory tract. The respiratory part of the nasal mucosa in particular showed only rare AIV positive cells and this was associated with reduced nasal shedding of the avian compared to the swine virus. The titers and distribution of the AIV varied extremely between individual pigs and were strongly affected by the route of inoculation. Gross lung lesions and clinical signs were milder with the avian than with the swine virus, corresponding with lower viral loads in the lungs. The brainstem was the single extra-respiratory tissue found positive for virus and viral RNA with both viruses. Our data do not reject the theory of the pig as an intermediate host for AIVs, but they suggest that AIVs need to undergo genetic changes to establish full replication potential in pigs. From a biomedical perspective, experimental LP H5 AIV infection of pigs may be useful to examine heterologous protection provided by H5 vaccines or other immunization strategies, as well as for further studies on the molecular pathogenesis and neurotropism of AIVs in mammals.

  4. Purification and production of monospecific antibody to the hemagglutinin from Subtype H5N1 influenza virus

    Directory of Open Access Journals (Sweden)

    Simson Tarigan

    2010-12-01

    Full Text Available The purpose of this study was to purify the hemagglutinin from H5N1 virus and to generate monospecific antibody appropriate for production of sensitive and specific immunoassay for H5N1 avian influenza. For this purpose, a local isolate H5N1 virus (A/Ck/West Java/Hamd/2006 was propagated in chicken embryos. The viral pellet was dissolved in a Triton-X-100 solution, undissolved viral particles were pelleted by ultracentrifuge, and the supernatant containing viral surface glycoproteins (Hemagglutinin and neuraminidase was collected. The neuraminidase in the supernatant was absorbed by passing the supernatant through an Oxamic-acid-superose column. After dialyzing extensively, the filtrate was further fractionated with an anion exchange chromatography (Q-sepharose column. Proteins adsorbed by the column were eluted stepwisely with 0.10, 0.25, 0.25 and 0.75 M NaCl in 20 mM Tris, ph 8. Hemagglutinin (H5 was found to be eluted from the column with the 0.5 M NaCl elution buffer. The purified H5 was free from other viral proteins based on immunoassays using commercial antibodies to H5N1 nucleoprotein and neuraminidase. When used as ELISA’s coating antigen, the purified H5 proved to be sensitive and specific for hemagglutinin H5. Cross reactions with other type-A-influenza virus, H6, H7 dan H9, were negligibly low. For the production of monospecific antiserum, the purified H5 was separated with SDS-PAGE, the band containing the H5 monomer was cut out , homogenised and injected into rabbits. The antiserum was capable of detecting the presence of inactivated H5N1 virus in a very dilute suspension, with a detection limit of 0.04 heagglutination (HA unit. The purified hemagglutinin and the serum raised against it should be useful for developing specific, sensitive and affordable immunoassay for H5N1 avian influenza.

  5. Birds and influenza H5N1 virus movement to and within North America

    Czech Academy of Sciences Publication Activity Database

    Rappole, J. H.; Hubálek, Zdeněk

    2006-01-01

    Roč. 12, č. 10 (2006), s. 1486-1492 ISSN 1080-6040 Institutional research plan: CEZ:AV0Z60930519 Keywords : West Nile virus * Avian influenza * waterfowl Subject RIV: FN - Epidemiology, Contagious Diseases ; Clinical Immunology Impact factor: 5.094, year: 2006 http://www.cdc.gov/ncidod/EID/vol12no10/05-1577.htm

  6. Human infection with a highly pathogenic avian influenza A (H5N6) virus in Yunnan province, China.

    Science.gov (United States)

    Xu, Wen; Li, Hong; Jiang, Li

    2016-01-01

    Highly pathogenic avian influenza A H5N6 virus has caused four human infections in China. This study reports the preliminary findings of the first known human case of H5N6 in Yunnan province. The patient initially developed symptoms of sore throat and coughing on 27 January 2015. The disease rapidly progressed to severe pneumonia, multiple organ dysfunctions and acute respiratory distress syndrome and the patient died on 6 February. Virological analysis determined that the virus belonged to H5 clade 2.3.4.4 and it has obtained partial ability for mammalian adaptation and amantadine resistance. Environmental investigation found H5 in 63% of the samples including poultry faeces, tissues, cage surface swabs and sewage from local live poultry markets by real-time RT-PCR. These findings suggest that the expanding and enhancing of surveillance in both avian and humans are necessary to monitor the evolution of H5 influenza virus and to facilitate early detection of suspected cases.

  7. In vitro assessment of attachment pattern and replication efficiency of H5N1 influenza A viruses with altered receptor specificity

    NARCIS (Netherlands)

    S. Chutinimitkul (Salin); D.A.J. van Riel (Debby); V.J. Munster (Vincent); J.M.A. van den Brand (Judith); G.F. Rimmelzwaan (Guus); T. Kuiken (Thijs); A.D.M.E. Osterhaus (Albert); R.A.M. Fouchier (Ron); E. de Wit (Emmie)

    2010-01-01

    textabstractThe continuous circulation of the highly pathogenic avian influenza (HPAI) H5N1 virus has been a cause of great concern. The possibility of this virus acquiring specificity for the human influenza A virus receptor, α2,6-linked sialic acids (SA), and being able to transmit efficiently

  8. Histopathological evaluation of the diversity of cells susceptible to H5N1 virulent avian influenza virus.

    Science.gov (United States)

    Ogiwara, Haru; Yasui, Fumihiko; Munekata, Keisuke; Takagi-Kamiya, Asako; Munakata, Tsubasa; Nomura, Namiko; Shibasaki, Futoshi; Kuwahara, Kazuhiko; Sakaguchi, Nobuo; Sakoda, Yoshihiro; Kida, Hiroshi; Kohara, Michinori

    2014-01-01

    Patients infected with highly pathogenic avian influenza A H5N1 viruses (H5N1 HPAIV) show diffuse alveolar damage. However, the temporal progression of tissue damage and repair after viral infection remains poorly defined. Therefore, we assessed the sequential histopathological characteristics of mouse lung after intranasal infection with H5N1 HPAIV or H1N1 2009 pandemic influenza virus (H1N1 pdm). We determined the amount and localization of virus in the lung through IHC staining and in situ hybridization. IHC used antibodies raised against the virus protein and antibodies specific for macrophages, type II pneumocytes, or proliferating cell nuclear antigen. In situ hybridization used RNA probes against both viral RNA and mRNA encoding the nucleoprotein and the hemagglutinin protein. H5N1 HPAIV infection and replication were observed in multiple lung cell types and might result in rapid progression of lung injury. Both type II pneumocytes and macrophages proliferated after H5N1 HPAIV infection. However, the abundant macrophages failed to block the viral attack, and proliferation of type II pneumocytes failed to restore the damaged alveoli. In contrast, mice infected with H1N1 pdm exhibited modest proliferation of type II pneumocytes and macrophages and slight alveolar damage. These results suggest that the virulence of H5N1 HPAIV results from the wide range of cell tropism of the virus, excessive virus replication, and rapid development of diffuse alveolar damage. Copyright © 2014 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  9. Corneal Opacity in Domestic Ducks Experimentally Infected With H5N1 Highly Pathogenic Avian Influenza Virus.

    Science.gov (United States)

    Yamamoto, Y; Nakamura, K; Yamada, M; Mase, M

    2016-01-01

    Domestic ducks can be a key factor in the regional spread of H5N1 highly pathogenic avian influenza (HPAI) virus in Asia. The authors performed experimental infections to examine the relationship between corneal opacity and H5N1 HPAI virus infection in domestic ducks (Anas platyrhyncha var domestica). A total of 99 domestic ducks, including 3 control birds, were used in the study. In experiment 1, when domestic ducks were inoculated intranasally with 2 H5N1 HPAI viruses, corneal opacity appeared more frequently than neurologic signs and mortality. Corneal ulceration and exophthalmos were rare findings. Histopathologic examinations of the eyes of domestic ducks in experiment 2 revealed that corneal opacity was due to the loss of corneal endothelial cells and subsequent keratitis with edema. Influenza viral antigen was detected in corneal endothelial cells and some other ocular cells by immunohistochemistry. Results suggest that corneal opacity is a characteristic and frequent finding in domestic ducks infected with the H5N1 HPAI virus. Confirming this ocular change may improve the detection rate of infected domestic ducks in the field. © The Author(s) 2015.

  10. Seroprevalence of antibodies against highly pathogenic avian influenza A (H5N1 virus among poultry workers in Bangladesh, 2009.

    Directory of Open Access Journals (Sweden)

    Sharifa Nasreen

    Full Text Available We conducted a cross-sectional study in 2009 to determine the seroprevalence and risk factors for highly pathogenic avian influenza A (H5N1 [HPAI H5N1] virus antibodies among poultry workers at farms and live bird markets with confirmed/suspected poultry outbreaks during 2009 in Bangladesh. We tested sera by microneutralization assay using A/Bangladesh/207095/2008 (H5N1; clade 2.2.2 virus with confirmation by horse red blood cell hemagglutination inhibition and H5-specific Western blot assays. We enrolled 212 workers from 87 farms and 210 workers from three live bird markets. One hundred and two farm workers (48% culled poultry. One hundred and ninety-three farm workers (91% and 178 market workers (85% reported direct contact with poultry that died during a laboratory confirmed HPAI H5N1 poultry farm outbreak or market poultry die-offs from suspected HPAI H5N1. Despite exposure to sick poultry, no farm or market poultry workers were seropositive for HPAI H5N1 virus antibodies (95% confidence interval 0-1%.

  11. Pathogenicity of the Korean H5N8 highly pathogenic avian influenza virus in commercial domestic poultry species.

    Science.gov (United States)

    Lee, Dong-Hun; Kwon, Jung-Hoon; Noh, Jin-Yong; Park, Jae-Keun; Yuk, Seong-Su; Erdene-Ochir, Tseren-Ochir; Lee, Joong-Bok; Park, Seung-Yong; Choi, In-Soo; Lee, Sang-Won; Song, Chang-Seon

    2016-01-01

    In 2014, the highly pathogenic avian influenza (HPAI) virus H5N8 triggered outbreaks in wild birds and poultry farms in South Korea. In the present study, we investigated the pathogenicity of the H5N8 HPAI virus, belonging to the clade 2.3.4.4, in different species of poultry. For this, we examined clinical signs and viral shedding levels following intranasal inoculation of the virus in 3-week-old commercial layer chickens and quails, 10-week-old Korean native chickens, and 8-week-old Muscovy ducks. Intranasal inoculation with 10(6.0) viruses at 50% egg-infective dose resulted in 100% mortality in the layer chickens (8/8) and quails (4/4), but 60% and 0% deaths in the Korean native chickens (3/5) and Muscovy ducks (0/4), respectively. In addition, transmission of the inoculated virus to contact-exposed birds was evident in all the species used in this study. Based on our results, we conclude that the H5N8 HPAI virus has lower pathogenicity and transmissibility in poultry species compared with previously reported H5N1 HPAI viruses.

  12. Pathobiology of highly pathogenic avian influenza virus (H5N1) infection in mute swans (Cygnus olor).

    Science.gov (United States)

    Pálmai, Nimród; Erdélyi, Károly; Bálint, Adám; Márton, Lázár; Dán, Adám; Deim, Zoltán; Ursu, Krisztina; Löndt, Brandon Z; Brown, Ian H; Glávits, Róbert

    2007-06-01

    The results of pathological, virological and polymerase chain reaction examinations carried out on 35 mute swans (Cygnus olor) that succumbed to a highly pathogenic avian influenza virus (H5N1) infection during an outbreak in Southern Hungary are reported. The most frequently observed macroscopic lesions included: haemorrhages under the epicardium, in the proventricular and duodenal mucosa and pancreas; focal necrosis in the pancreas; myocardial degeneration; acute mucous enteritis; congestion of the spleen and lung, and the accumulation of sero-mucinous exudate in the body cavity. Histopathological lesions comprised: lymphocytic meningo-encephalomyelitis accompanied by gliosis and occasional perivascular haemorrhages; multi-focal myocardial necrosis with lympho-histiocytic infiltration; pancreatitis with focal necrosis; acute desquamative mucous enteritis; lung congestion and oedema; oedema of the tracheal mucosa and, in young birds, the atrophy of the bursa of Fabricius as a result of lymphocyte depletion and apoptosis. The observed lesions and the moderate to good body conditions were compatible with findings in acute highly pathogenic avian influenza infections of other bird species reported in the literature. Skin lesions and lesions typical for infections caused by strains of lower pathogenicity (low pathogenic avian influenza virus) such as emaciation or fibrinous changes in the reproductive and respiratory organs, sinuses and airsacs were not observed. The H5N1 subtype avian influenza virus was isolated in embryonated fowl eggs from all cases and it was identified by classical and molecular virological methods.

  13. Centralized Consensus Hemagglutinin Genes Induce Protective Immunity against H1, H3 and H5 Influenza Viruses.

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    Richard J Webby

    Full Text Available With the exception of the live attenuated influenza vaccine there have been no substantial changes in influenza vaccine strategies since the 1940's. Here we report an alternative vaccine approach that uses Adenovirus-vectored centralized hemagglutinin (HA genes as vaccine antigens. Consensus H1-Con, H3-Con and H5-Con HA genes were computationally derived. Mice were immunized with Ad vaccines expressing the centralized genes individually. Groups of mice were vaccinated with 1 X 1010, 5 X 107 and 1 X 107 virus particles per mouse to represent high, intermediate and low doses, respectively. 100% of the mice that were vaccinated with the high dose vaccine were protected from heterologous lethal challenges within each subtype. In addition to 100% survival, there were no signs of weight loss and disease in 7 out of 8 groups of high dose vaccinated mice. Lower doses of vaccine showed a reduction of protection in a dose-dependent manner. However, even the lowest dose of vaccine provided significant levels of protection against the divergent influenza strains, especially considering the stringency of the challenge virus. In addition, we found that all doses of H5-Con vaccine were capable of providing complete protection against mortality when challenged with lethal doses of all 3 H5N1 influenza strains. This data demonstrates that centralized H1-Con, H3-Con and H5-Con genes can be effectively used to completely protect mice against many diverse strains of influenza. Therefore, we believe that these Ad-vectored centralized genes could be easily translated into new human vaccines.

  14. Influenza A H5N1 clade 2.3.4 virus with a different antiviral susceptibility profile replaced clade 1 virus in humans in northern Vietnam.

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    Mai T Q Le

    2008-10-01

    Full Text Available Prior to 2007, highly pathogenic avian influenza (HPAI H5N1 viruses isolated from poultry and humans in Vietnam were consistently reported to be clade 1 viruses, susceptible to oseltamivir but resistant to amantadine. Here we describe the re-emergence of human HPAI H5N1 virus infections in Vietnam in 2007 and the characteristics of the isolated viruses.Respiratory specimens from patients suspected to be infected with avian influenza in 2007 were screened by influenza and H5 subtype specific polymerase chain reaction. Isolated H5N1 strains were further characterized by genome sequencing and drug susceptibility testing. Eleven poultry outbreak isolates from 2007 were included in the sequence analysis. Eight patients, all of them from northern Vietnam, were diagnosed with H5N1 in 2007 and five of them died. Phylogenetic analysis of H5N1 viruses isolated from humans and poultry in 2007 showed that clade 2.3.4 H5N1 viruses replaced clade 1 viruses in northern Vietnam. Four human H5N1 strains had eight-fold reduced in-vitro susceptibility to oseltamivir as compared to clade 1 viruses. In two poultry isolates the I117V mutation was found in the neuraminidase gene, which is associated with reduced susceptibility to oseltamivir. No mutations in the M2 gene conferring amantadine resistance were found.In 2007, H5N1 clade 2.3.4 viruses replaced clade 1 viruses in northern Vietnam and were susceptible to amantadine but showed reduced susceptibility to oseltamivir. Combination antiviral therapy with oseltamivir and amantadine for human cases in Vietnam is recommended.

  15. Seroprevalence of avian influenza A (H5N1 virus among poultry workers in Jiangsu Province, China: an observational study

    Directory of Open Access Journals (Sweden)

    Huo Xiang

    2012-04-01

    Full Text Available Abstract Background Since 2003 to 06 Jan 2012, the number of laboratory confirmed human cases of infection with avian influenza in China was 41 and 27 were fatal. However, the official estimate of the H5N1 case-fatality rate has been described by some as an over estimation since there may be numerous undetected asymptomatic/mild cases of H5N1 infection. This study was conducted to better understand the real infection rate and evaluate the potential risk factors for the zoonotic spread of H5N1 viruses to humans. Methods A seroepidemiological survey was conducted in poultry workers, a group expected to have the highest level of exposure to H5N1-infected birds, from 3 counties with habitat lakes of wildfowl in Jiangsu province, China. Serum specimens were collected from 306 participants for H5N1 serological test. All participants were interviewed to collect information about poultry exposures. Results The overall seropositive rate was 2.61% for H5N1 antibodies. The poultry number was found associated with a 2.39-fold significantly increased subclinical infection risk after adjusted with age and gender. Conclusions Avian-to -human transmission of avian H5N1 virus remained low. Workers associated with raising larger poultry flocks have a higher risk on seroconversion.

  16. Pulmonary immunization of chickens using non-adjuvanted spray-freeze dried whole inactivated virus vaccine completely protects against highly pathogenic H5N1 avian influenza virus

    NARCIS (Netherlands)

    Peeters, Ben; Tonnis, Wouter F.; Murugappan, Senthil; Rottier, Peter; Koch, Guus; Frijlink, Henderik W.; Huckriede, Anke; Hinrichs, Wouter L. J.

    2014-01-01

    Highly pathogenic avian influenza (HPAI) H5N1 virus is a major threat to public health as well as to the global poultry industry. Most fatal human infections are caused by contact with infected poultry. Therefore, preventing the virus from entering the poultry population is a priority. This is,

  17. Avian influenza A (H5N1)

    NARCIS (Netherlands)

    de Jong, Menno D.; Hien, Tran Tinh

    2006-01-01

    Since their reemergence in 2003, highly pathogenic avian influenza A (H5N1) viruses have reached endemic levels among poultry in several southeast Asian countries and have caused a still increasing number of more than 100 reported human infections with high mortality. These developments have ignited

  18. Pathology of whooper swans (Cygnus cygnus) infected with H5N1 avian influenza virus in Akita, Japan, in 2008.

    Science.gov (United States)

    Ogawa, Shuji; Yamamoto, Yu; Yamada, Manabu; Mase, Masaji; Nakamura, Kikuyasu

    2009-10-01

    Two (1 adult and 1 young bird) of 4 H5N1-highly-pathogenic-avian-influenza (HPAI)-virus-infected whooper swans in Akita, Japan, in 2008 were investigated pathologically. Macroscopically, white spots with hemorrhages were scattered in the pancreas in the adult bird. Histologically, the adult bird had severe necrotizing pancreatitis and mild nonpurulent encephalitis. The young bird had severe nonpurulent encephalitis and nonpurulent enteric ganglionitis, and intestinal venous wall thickening. Virus antigens were detected in the lesions of pancreatitis in the adult bird and of encephalitis in adult and young birds. These findings suggest that the swans died or became moribund due to neurological disorders and necrotizing pancreatitis caused by H5N1 HPAI virus infection.

  19. Molecular epidemiology of circulating highly pathogenic avian influenza (H5N1) virus in chickens, in Bangladesh, 2007-2010

    DEFF Research Database (Denmark)

    Ahmed, Syed Sayeem Uddin; Themudo, Goncalo Espregueira Cruz; Christensen, Jens Peter

    2012-01-01

    Bangladesh has been severely hit by highly pathogenic avian influenza H5N1 (HPAI-H5N1). However, little is known about the genetic diversity and the evolution of the circulating viruses in Bangladesh. In the present study, we analyzed the hemagglutinin gene of 30 Bangladeshi chicken isolates from...... several amino acid substitutions, but they are not indicative of adaptation toward human infection. The Mantel correlation test confirmed significant correlation between genetic distances and temporal distances between the viruses. The Bayesian tree shows that isolates from waves 3 and 4 derived from...... virus in Bangladesh. Furthermore, the formation of a subclade capable of transmission to humans cannot be ruled out. The findings of this study might provide valuable information for future surveillance, prevention and control programme....

  20. Experimental infection of macaques with a wild water bird-derived highly pathogenic avian influenza virus (H5N1.

    Directory of Open Access Journals (Sweden)

    Tomoko Fujiyuki

    Full Text Available Highly pathogenic avian influenza virus (HPAIV continues to threaten human health. Non-human primate infection models of human influenza are desired. To establish an animal infection model with more natural transmission and to determine the pathogenicity of HPAIV isolated from a wild water bird in primates, we administered a Japanese isolate of HPAIV (A/whooper swan/Hokkaido/1/2008, H5N1 clade 2.3.2.1 to rhesus and cynomolgus monkeys, in droplet form, via the intratracheal route. Infection of the lower and upper respiratory tracts and viral shedding were observed in both macaques. Inoculation of rhesus monkeys with higher doses of the isolate resulted in stronger clinical symptoms of influenza. Our results demonstrate that HPAIV isolated from a water bird in Japan is pathogenic in monkeys by experimental inoculation, and provide a new method for HPAIV infection of non-human primate hosts, a good animal model for investigation of HPAIV pathogenicity.

  1. A highly pathogenic avian influenza virus H5N1 with 2009 pandemic H1N1 internal genes demonstrated increased replication and transmission in pigs

    Science.gov (United States)

    This study investigated the pathogenicity and transmissibility of a reverse-genetics derived highly pathogenic avian influenza (HPAI) H5N1 influenza A virus (IAV), A/Iraq/775/06, and a reassortant virus comprised of the HA and NA from A/Iraq/775/06 and the internal genes of a 2009 pandemic H1N1, A/N...

  2. Experimental assessment of the pathogenicity of two avian influenza A H5 viruses in ostrich chicks (Struthio camelus) and chickens.

    Science.gov (United States)

    Manvell, R J; Jørgensen, P H; Nielsen, O L; Alexander, D J

    1998-01-01

    Virus excretion, immune response, and, for chickens, deaths were recorded in 3-week-old ostriches and chickens inoculated by either the intramuscular or intranasal route with one of two influenza A viruses of subtype H5. One of the viruses, A/turkey/England/50-92/91 (H5N1) (50/92), was highly pathogenic for chickens causing 5/5 deaths by each route of inoculation. The other virus, A/ostrich/Denmark-Q/72420/96 (H5N2) (72420/96), isolated from ostriches in quarantine in Denmark during 1996, was of low pathogenicity for chickens, causing no clinical signs by either route of inoculation. No significant clinical signs were seen in any of the ostriches infected with either of the viruses by either route of infection. Both viruses were recoverable from both species up to 12 days post-infection, and low serological responses were detected in surviving infected ostriches and chickens at 21 days after inoculation.

  3. Expression of H5 hemagglutinin vaccine antigen in common duckweed (Lemna minor) protects against H5N1 high pathogenicity avian influenza virus challenge in immunized chickens.

    Science.gov (United States)

    Bertran, Kateri; Thomas, Colleen; Guo, Xuan; Bublot, Michel; Pritchard, Nikki; Regan, Jeffrey T; Cox, Kevin M; Gasdaska, John R; Dickey, Lynn F; Kapczynski, Darrell R; Swayne, David E

    2015-07-09

    A synthetic hemagglutinin (HA) gene from the highly pathogenic avian influenza (HPAI) virus A/chicken/Indonesia/7/2003 (H5N1) (Indo/03) was expressed in aquatic plant Lemna minor (rLemna-HA). In Experiment 1, efficacy of rLemna-HA was tested on birds immunized with 0.2μg or 2.3 μg HA and challenged with 10(6) mean chicken embryo infectious doses (EID50) of homologous virus strain. Both dosages of rLemna-HA conferred clinical protection and dramatically reduced viral shedding. Almost all the birds immunized with either dosage of rLemna-HA elicited HA antibody titers against Indo/03 antigen, suggesting an association between levels of anti-Indo/03 antibodies and protection. In Experiment 2, efficacy of rLemna-HA was tested on birds immunized with 0.9 μg or 2.2 μg HA and challenged with 10(6) EID50 of heterologous H5N1 virus strains A/chicken/Vietnam/NCVD-421/2010 (VN/10) or A/chicken/West Java/PWT-WIJ/2006 (PWT/06). Birds challenged with VN/10 exhibited 100% survival regardless of immunization dosage, while birds challenged with PWT/06 had 50% and 30% mortality at 0.9 μg HA and 2.2 μg HA, respectively. For each challenge virus, viral shedding titers from 2.2 μg HA vaccinated birds were significantly lower than those from 0.9μg HA vaccinated birds, and titers from both immunized groups were in turn significantly lower than those from sham vaccinated birds. Even if immunized birds elicited HA titers against the vaccine antigen Indo/03, only the groups challenged with VN/10 developed humoral immunity against the challenge antigen. None (rLemna-HA 0.9 μg HA) and 40% (rLemna-HA 2.2 μg HA) of the immunized birds challenged with PWT/06 elicited pre-challenge antibody titers, respectively. In conclusion, Lemna-expressed HA demonstrated complete protective immunity against homologous challenge and suboptimal protection against heterologous challenge, the latter being similar to results from inactivated whole virus vaccines. Transgenic duckweed-derived HA could be a

  4. Penerapan Metode Diagnosis Cepat Virus Avian Influenza H5N1 dengan Metode Single Step Multiplex RT-PCR

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    Aris Haryanto

    2010-12-01

    Full Text Available Avian influenza (AI virus is a segmented single stranded (ss RNA virus with negative polarity andbelong to the Orthomyxoviridae family. Diagnose of AI virus can be performed using conventional methodsbut it has low sensitivity and specificity. The objective of the research was to apply rapid, precise, andaccurate diagnostic method for AI virus and also to determine its type and subtype based on the SingleStep Multiplex Reverse Transcriptase-Polymerase Chain Reaction targeting M, H5, and N1 genes. In thismethod M, H5 and NI genes were simultaneously amplified in one PCR tube. The steps of this researchconsist of collecting viral RNAs from 10 different AI samples originated from Maros Disease InvestigationCenter during 2007. DNA Amplification was conducted by Simplex RT-PCR using M primer set. Then, bysingle step multiplex RT-PCR were conducted simultaneously using M, H5 and N1 primers set. The RTPCRproducts were then separated on 1.5% agarose gel, stained by ethidum bromide and visualized underUV transilluminator. Results showed that 8 of 10 RNA virus samples could be amplified by Simplex RTPCRfor M gene which generating a DNA fragment of 276 bp. Amplification using multiplex RT-PCRmethod showed two of 10 samples were AI positive using multiplex RT-PCR, three DNA fragments weregenerated consisting of 276 bp for M gene, 189 bp for H5 gene, and 131 bp for N1. In this study, rapid andeffective diagnosis method for AI virus can be conducted by using simultaneous Single Step Multiplex RTPCR.By this technique type and subtype of AI virus, can also be determined, especially H5N1.

  5. Host-specific exposure and fatal neurologic disease in wild raptors from highly pathogenic avian influenza virus H5N1 during the 2006 outbreak in Germany

    NARCIS (Netherlands)

    J.M.A. van den Brand (Judith); O. Krone (Oliver); P.U. Wolf (Peter U.); M.W.G. van de Bildt (Marco); G. van Amerongen (Geert); A.D.M.E. Osterhaus (Albert); T. Kuiken (Thijs)

    2015-01-01

    textabstractRaptors may contract highly pathogenic avian influenza virus H5N1 by hunting or scavenging infected prey. However, natural H5N1 infection in raptors is rarely reported. Therefore, we tested raptors found dead during an H5N1 outbreak in wild waterbirds in Mecklenburg-Western Pomerania,

  6. Pathogenesis of H5N1 influenza virus infections in mice and ferret models differ between respiratory and digestive system exposure

    Science.gov (United States)

    Background. Epidemiological, clinical and laboratory data suggests H5N1 influenza viruses are transmitted through and predominantly affect the respiratory system of mammals. Some data suggests digestive system involvement. However, direct evidence of alimentary transmission and infection in mammal...

  7. Protection of chickens against H5N1 highly pathogenic avian influenza virus infection by live vaccination with infectious laryngotracheitis virus recombinants expressing H5 hemagglutinin and N1 neuraminidase.

    Science.gov (United States)

    Pavlova, Sophia P; Veits, Jutta; Keil, Günther M; Mettenleiter, Thomas C; Fuchs, Walter

    2009-01-29

    Attenuated vaccine strains of the alphaherpesvirus causing infectious laryngotracheitis of chickens (ILTV, gallid herpesvirus 1) can be used for mass application. Previously, we showed that live virus vaccination with recombinant ILTV expressing hemagglutinin of highly pathogenic avian influenza viruses (HPAIV) protected chickens against ILT and fowl plague caused by HPAIV carrying the corresponding hemagglutinin subtypes [Lüschow D, Werner O, Mettenleiter TC, Fuchs W. Protection of chickens from lethal avian influenza A virus infection by live-virus vaccination with infectious laryngotracheitis virus recombinants expressing the hemagglutinin (H5) gene. Vaccine 2001;19(30):4249-59; Veits J, Lüschow D, Kindermann K, Werner O, Teifke JP, Mettenleiter TC, et al. Deletion of the non-essential UL0 gene of infectious laryngotracheitis (ILT) virus leads to attenuation in chickens, and UL0 mutants expressing influenza virus haemagglutinin (H7) protect against ILT and fowl plague. J Gen Virol 2003;84(12):3343-52]. However, protection against H5N1 HPAIV was not satisfactory. Therefore, a newly designed dUTPase-negative ILTV vector was used for rapid insertion of the H5-hemagglutinin, or N1-neuraminidase genes of a recent H5N1 HPAIV isolate. Compared to our previous constructs, protein expression was considerably enhanced by insertion of synthetic introns downstream of the human cytomegalovirus immediate-early promoter within the 5'-nontranslated region of the transgenes. Deletion of the viral dUTPase gene did not affect in vitro replication of the ILTV recombinants, but led to sufficient attenuation in vivo. After a single ocular immunization, all chickens developed H5- or N1-specific serum antibodies. Nevertheless, animals immunized with N1-ILTV died after subsequent H5N1 HPAIV challenge, although survival times were prolonged compared to non-vaccinated controls. In contrast, all chickens vaccinated with either H5-ILTV alone, or H5- and N1-ILTV simultaneously, survived

  8. Adaption of wild-bird origin H5Nx highly pathogenic avian influenza virus Clade 2.3.4.4 in vaccinated poultry

    Science.gov (United States)

    The 2014-2015 incursion of H5Nx clade 2.3.4.4 high pathogenicity avian influenza (HPAI) virus caused the largest animal health emergency in U.S. history and renewed interest in developing vaccines against these newly emergent viruses. Our previous research demonstrated several H5 vaccines with varyi...

  9. Immune Responses of Chickens Infected with Wild Bird-Origin H5N6 Avian Influenza Virus

    Directory of Open Access Journals (Sweden)

    Shimin Gao

    2017-06-01

    Full Text Available Since April 2014, new infections of H5N6 avian influenza virus (AIV in humans and domestic poultry have caused considerable economic losses in the poultry industry and posed an enormous threat to human health worldwide. In previous research using gene sequence and phylogenetic analysis, we reported that H5N6 AIV isolated in February 2015 (ZH283 in Pallas’s sandgrouse was highly similar to that isolated in a human in December 2015 (A/Guangdong/ZQ874/2015, whereas a virus (i.e., SW8 isolated in oriental magpie-robin in 2014 was highly similar to that of A/chicken/Dongguan/2690/2013 (H5N6. However, the pathogenicity, transmissibility, and host immune-related response of chickens infected by those wild bird-origin H5N6 AIVs remain unknown. In response, we examined the viral distribution and mRNA expression profiles of immune-related genes in chickens infected with both viruses. Results showed that the H5N6 AIVs were highly pathogenic to chickens and caused not only systemic infection in multiple tissues, but also 100% mortality within 3–5 days post-infection. Additionally, ZH283 efficiently replicated in all tested tissues and transmitted among chickens more rapidly than SW8. Moreover, quantitative real-time polymerase chain reaction analysis showed that following infection with H5N6, AIVs immune-related genes remained active in a tissue-dependent manner, as well as that ZH283 induced mRNA expression profiles such as TLR3, TLR7, IL-6, TNF-α, IL-1β, IL-10, IL-8, and MHC-II to a greater extent than SW8 in the tested tissues of infected chickens. Altogether, our findings help to illuminate the pathogenesis and immunologic mechanisms of H5N6 AIVs in chickens.

  10. MERS-CoV and H5N1 influenza virus antagonize antigen presentation by altering the epigenetic landscape

    Energy Technology Data Exchange (ETDEWEB)

    Menachery, Vineet D.; Schafer, Alexandra; Burnum-Johnson, Kristin E.; Mitchell, Hugh D.; Eisfeld-Fenney, Amie J.; Walters, Kevin B.; Nicora, Carrie D.; Purvine, Samuel O.; Casey, Cameron P.; Monroe, Matthew E.; Weitz, Karl K.; Stratton, Kelly G.; Webb-Robertson, Bobbie-Jo M.; Gralinski, Lisa; Metz, Thomas O.; Smith, Richard D.; Waters, Katrina M.; Sims, Amy C.; Kawaoka, Yoshihiro; Baric, Ralph

    2018-01-16

    Convergent evolution dictates that diverse groups of viruses will target both similar and distinct host pathways in order to manipulate the immune response and improve infection. In this study, we sought to leverage this uneven viral antagonism to identify critical host factors that govern disease outcome. Utilizing a systems based approach, we examined differential regulation of IFNγ dependent genes following infection with highly pathogenic viruses including influenza (H5N1-VN1203, H1N1-CA04) and coronaviruses (SARS-CoV, MERS-CoV). Categorizing by function, we observed down regulation of genes associated with antigen presentation following both H5N1-VN1203 and MERS-CoV infection. Further examination revealed global down regulation of antigen presentation genes and was confirmed by proteomics for both H5N1-VN1203 and MERS-CoV infection. Importantly, epigenetic analysis suggested that DNA methylation rather than histone modification plays a crucial role in MERS-CoV mediated antagonism of antigen presentation genes; in contrast, H5N1-VN1203 likely utilizes a combination of epigenetic mechanisms to target antigen presentation. Together, the results indicate a common approach utilized by H5N1-VN1203 and MERS-CoV to modulate antigen presentation and the host adaptive immune response.

  11. An Impedance Aptasensor with Microfluidic Chips for Specific Detection of H5N1 Avian Influenza Virus

    Directory of Open Access Journals (Sweden)

    Jacob Lum

    2015-07-01

    Full Text Available In this research a DNA aptamer, which was selected through SELEX (systematic evolution of ligands by exponential enrichment to be specific against the H5N1 subtype of the avian influenza virus (AIV, was used as an alternative reagent to monoclonal antibodies in an impedance biosensor utilizing a microfluidics flow cell and an interdigitated microelectrode for the specific detection of H5N1 AIV. The gold surface of the interdigitated microelectrode embedded in a microfluidics flow cell was modified using streptavidin. The biotinylated aptamer against H5N1 was then immobilized on the electrode surface using biotin–streptavidin binding. The target virus was captured on the microelectrode surface, causing an increase in impedance magnitude. The aptasensor had a detection time of 30 min with a detection limit of 0.0128 hemagglutinin units (HAU. Scanning electron microscopy confirmed the binding of the target virus onto the electrode surface. The DNA aptamer was specific to H5N1 and had no cross-reaction to other subtypes of AIV (e.g., H1N1, H2N2, H7N2. The newly developed aptasensor offers a portable, rapid, low-cost alternative to current methods with the same sensitivity and specificity.

  12. The creation of a contagious H5N1 influenza virus: implications for the education of life scientists

    Directory of Open Access Journals (Sweden)

    T. Novossiolova

    2014-12-01

    Full Text Available The paper contends that the ongoing controversy surrounding the creation of a contagious H5N1 influenza virus has already exposed the severe limitations of the possibility of preventing the hostile misuse of the life scien­ces by dint of oversight of proposals and publications. It further argues that in order to prevent the potential wholesale militarisation of the life sciences, it is essential that life scientists become aware of their responsibilities within the context of the Biological and Toxin Weapons Convention (BTWC and actively contribute their expertise to strengthening the biological weapons non-proliferation regime.

  13. Infection of mice with a human influenza A/H3N2 virus induces protective immunity against lethal infection with influenza A/H5N1 virus.

    Science.gov (United States)

    Kreijtz, J H C M; Bodewes, R; van den Brand, J M A; de Mutsert, G; Baas, C; van Amerongen, G; Fouchier, R A M; Osterhaus, A D M E; Rimmelzwaan, G F

    2009-08-06

    The transmission of highly pathogenic avian influenza (HPAI) A viruses of the H5N1 subtype from poultry to man and the high case fatality rate fuels the fear for a pandemic outbreak caused by these viruses. However, prior infections with seasonal influenza A/H1N1 and A/H3N2 viruses induce heterosubtypic immunity that could afford a certain degree of protection against infection with the HPAI A/H5N1 viruses, which are distantly related to the human influenza A viruses. To assess the protective efficacy of such heterosubtypic immunity mice were infected with human influenza virus A/Hong Kong/2/68 (H3N2) 4 weeks prior to a lethal infection with HPAI virus A/Indonesia/5/05 (H5N1). Prior infection with influenza virus A/Hong Kong/2/68 reduced clinical signs, body weight loss, mortality and virus replication in the lungs as compared to naive mice infected with HPAI virus A/Indonesia/5/05. Priming by infection with respiratory syncytial virus, a non-related virus did not have a beneficial effect on the outcome of A/H5N1 infections, indicating that adaptive immune responses were responsible for the protective effect. In mice primed by infection with influenza A/H3N2 virus cytotoxic T lymphocytes (CTL) specific for NP(366-374) epitope ASNENMDAM and PA(224-232) SCLENFRAYV were observed. A small proportion of these CTL was cross-reactive with the peptide variant derived from the influenza A/H5N1 virus (ASNENMEVM and SSLENFRAYV respectively) and upon challenge infection with the influenza A/H5N1 virus cross-reactive CTL were selectively expanded. These CTL, in addition to those directed to conserved epitopes, shared by the influenza A/H3N2 and A/H5N1 viruses, most likely contributed to accelerated clearance of the influenza A/H5N1 virus infection. Although also other arms of the adaptive immune response may contribute to heterosubtypic immunity, the induction of virus-specific CTL may be an attractive target for development of broad protective vaccines. Furthermore the

  14. Experimental and Field Results Regarding Immunity Induced by a Recombinant Turkey Herpesvirus H5 Vector Vaccine Against H5N1 and Other H5 Highly Pathogenic Avian Influenza Virus Challenges.

    Science.gov (United States)

    Gardin, Yannick; Palya, Vilmos; Dorsey, Kristi Moore; El-Attrache, John; Bonfante, Francesco; Wit, Sjaak de; Kapczynski, Darrell; Kilany, Walid Hamdy; Rauw, Fabienne; Steensels, Mieke; Soejoedono, Retno D

    2016-05-01

    Vaccination against H5N1 highly pathogenic avian influenza (AI) virus (HPAIV) is one of the possible complementary means available for affected countries to control AI when the disease has become, or with a high risk of becoming, endemic. Efficacy of the vaccination against AI relies essentially, but not exclusively, on the capacity of the vaccine to induce immunity against the targeted virus (which is prone to undergo antigenic variations), as well as its capacity to overcome interference with maternal immunity transmitted by immunized breeding hens to their progeny. This property of the vaccine is a prerequisite for its administration at the hatchery, which assures higher and more reliable vaccine coverage of the populations than vaccination at the farm. A recombinant vector vaccine (Vectormune® AI), based on turkey herpesvirus expressing the hemagglutinin gene of an H5N1 HPAIV as an insert, has been used in several experiments conducted in different research laboratories, as well as in controlled field trials. The results have demonstrated a high degree of homologous and cross protection against different genetic clades of the H5N1 HPAIV. Furthermore, vaccine-induced immunity was not impaired by the presence of passive immunity, but on the contrary, cumulated with it for improved early protection. The demonstrated levels of protection against the different challenge viruses exhibited variations in terms of postchallenge mortality, as well as challenge virus shedding. The data presented here highlight the advantages of this vaccine as a useful and reliable tool to complement biosecurity and sanitary policies for better controlling the disease due to HPAIV of H5 subtypes, when the vaccination is applied as a control measure.

  15. Internal Gene Cassette from a Genotype S H9N2 Avian Influenza Virus Attenuates the Pathogenicity of H5 Viruses in Chickens and Mice

    Directory of Open Access Journals (Sweden)

    Xiaoli Hao

    2017-10-01

    Full Text Available H9N2 avian influenza virus (AIV of genotype S frequently donate internal genes to facilitate the generation of novel reassortants such as H7N9, H10N8, H5N2 and H5N6 AIVs, posing an enormous threat to both human health and poultry industry. However, the pathogenicity and transmission of reassortant H5 viruses with internal gene cassette of genotype S H9N2-origin in chickens and mice remain unknown. In this study, four H5 reassortants carrying the HA and NA genes from different clades of H5 viruses and the remaining internal genes from an H9N2 virus of the predominant genotype S were generated by reverse genetics. We found that all four H5 reassortant viruses showed attenuated virulence in both chickens and mice, thus leading to increased the mean death times compared to the corresponding parental viruses. Consistently, the polymerase activity and replication ability in mammalian and avian cells, and the cytokine responses in the lungs of chickens and mice were also decreased when compared to their respective parental viruses. Moreover, these reassortants transmitted from birds to birds by direct contact but not by an airborne route. Our data indicate that the internal genes as a whole cassette from genotype S H9N2 viruses play important roles in reducing the pathogenicity of the H5 recombinants in chickens and mice, and might contribute to the circulation in avian or mammalian hosts.

  16. An epidemiological study of avian influenza A (H5) virus in nomadic ducks and their raising practices in northeastern Bangladesh, 2011-2012.

    Science.gov (United States)

    Sarkar, Shamim; Khan, Salah Uddin; Mikolon, Andrea; Rahman, Mohammad Ziaur; Abedin, Jaynal; Zeidner, Nord; Sturm-Ramirez, Katherine; Luby, Stephen P

    2017-05-01

    In Bangladesh, nomadic duck flocks are groups of domestic ducks reared for egg production that are moved to access feeding sites beyond their owners' village boundaries and are housed overnight in portable enclosures in scavenging areas. The objectives of this study were to measure the prevalence of influenza A virus RNA and H5-specific antibodies in nomadic ducks and to characterize nomadic duck raising practices in northeastern Bangladesh. We tested duck egg yolk specimens by competitive ELISA to detect antibodies against avian influenza A (H5) and environmental fecal samples by real-time reverse-transcription polymerase chain reaction (rRT-PCR) to detect influenza A virus RNA and H5 subtype. The median age of the ducks was 24 months (range: 8-36 months) and the median flock size was 300 ducks (range: 105-1100). Of 1860 egg yolk samples, 556 (30%, 95% confidence interval (CI): 28-32) were positive for antibodies against H5 and 58 flocks (94%) had at least one egg with H5-specific antibodies. Of 496 fecal samples, 121 (24%, 95% CI: 22-29) had detectable influenza A RNA. Thirty-three flocks (53%) had at least one fecal sample positive for influenza A RNA. Nomadic ducks in Bangladesh are commonly infected with avian influenza A (H5) virus and may serve as a bridging host for transmission of avian influenza A (H5) virus or other avian influenza A viruses subtypes between wild waterfowl, backyard poultry, and humans in Bangladesh. © 2016 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

  17. Multiple reassorted viruses as cause of highly pathogenic avian influenza A(H5N8) virus epidemic, the Netherlands, 2016

    NARCIS (Netherlands)

    Beerens, Nancy; Heutink, Rene; Bergervoet, Saskia A.; Harders, Frank; Bossers, Alex; Koch, Guus

    2017-01-01

    In 2016, an epidemic of highly pathogenic avian influenza A virus subtype H5N8 in the Netherlands caused mass deaths among wild birds, and several commercial poultry farms and captive bird holdings were affected. We performed complete genome sequencing to study the relationship between the wild bird

  18. Virus-like particles displaying H5, H7, H9 hemagglutinins and N1 neuraminidase elicit protective immunity to heterologous avian influenza viruses in chickens

    International Nuclear Information System (INIS)

    Pushko, Peter; Tretyakova, Irina; Hidajat, Rachmat; Zsak, Aniko; Chrzastek, Klaudia; Tumpey, Terrence M.; Kapczynski, Darrell R.

    2017-01-01

    Avian influenza (AI) viruses circulating in wild birds pose a serious threat to public health. Human and veterinary vaccines against AI subtypes are needed. Here we prepared triple-subtype VLPs that co-localized H5, H7 and H9 antigens derived from H5N1, H7N3 and H9N2 viruses. VLPs also contained influenza N1 neuraminidase and retroviral gag protein. The H5/H7/H9/N1/gag VLPs were prepared using baculovirus expression. Biochemical, functional and antigenic characteristics were determined including hemagglutination and neuraminidase enzyme activities. VLPs were further evaluated in a chicken AI challenge model for safety, immunogenicity and protective efficacy against heterologous AI viruses including H5N2, H7N3 and H9N2 subtypes. All vaccinated birds survived challenges with H5N2 and H7N3 highly pathogenic AI (HPAI) viruses, while all controls died. Immune response was also detectable after challenge with low pathogenicity AI (LPAI) H9N2 virus suggesting that H5/H7/H9/N1/gag VLPs represent a promising approach for the development of broadly protective AI vaccine. - Highlights: •VLPs were prepared that co-localized H5, H7 and H9 subtypes in a VLP envelope. •VLPs were characterized including electron microscopy, HA assay and NA enzyme activity. •Experimental VLP vaccine was evaluated in an avian influenza challenge model. •VLPs induced immune responses against heterologous H5, H7 and H9 virus challenges.

  19. Virus-like particles displaying H5, H7, H9 hemagglutinins and N1 neuraminidase elicit protective immunity to heterologous avian influenza viruses in chickens

    Energy Technology Data Exchange (ETDEWEB)

    Pushko, Peter, E-mail: ppushko@medigen-usa.com [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States); Tretyakova, Irina; Hidajat, Rachmat [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States); Zsak, Aniko; Chrzastek, Klaudia [USDA SEPRL, 934 College Station Rd, Athens, GA (United States); Tumpey, Terrence M. [Influenza Division, CDC,1600 Clifton Road N.E., Atlanta, GA (United States); Kapczynski, Darrell R. [USDA SEPRL, 934 College Station Rd, Athens, GA (United States)

    2017-01-15

    Avian influenza (AI) viruses circulating in wild birds pose a serious threat to public health. Human and veterinary vaccines against AI subtypes are needed. Here we prepared triple-subtype VLPs that co-localized H5, H7 and H9 antigens derived from H5N1, H7N3 and H9N2 viruses. VLPs also contained influenza N1 neuraminidase and retroviral gag protein. The H5/H7/H9/N1/gag VLPs were prepared using baculovirus expression. Biochemical, functional and antigenic characteristics were determined including hemagglutination and neuraminidase enzyme activities. VLPs were further evaluated in a chicken AI challenge model for safety, immunogenicity and protective efficacy against heterologous AI viruses including H5N2, H7N3 and H9N2 subtypes. All vaccinated birds survived challenges with H5N2 and H7N3 highly pathogenic AI (HPAI) viruses, while all controls died. Immune response was also detectable after challenge with low pathogenicity AI (LPAI) H9N2 virus suggesting that H5/H7/H9/N1/gag VLPs represent a promising approach for the development of broadly protective AI vaccine. - Highlights: •VLPs were prepared that co-localized H5, H7 and H9 subtypes in a VLP envelope. •VLPs were characterized including electron microscopy, HA assay and NA enzyme activity. •Experimental VLP vaccine was evaluated in an avian influenza challenge model. •VLPs induced immune responses against heterologous H5, H7 and H9 virus challenges.

  20. Tropism and Infectivity of Influenza Virus, Including Highly Pathogenic Avian H5N1 Virus, in Ferret Tracheal Differentiated Primary Epithelial Cell Cultures

    Science.gov (United States)

    Zeng, Hui; Goldsmith, Cynthia S.; Maines, Taronna R.; Belser, Jessica A.; Gustin, Kortney M.; Pekosz, Andrew; Zaki, Sherif R.; Katz, Jacqueline M.

    2013-01-01

    Tropism and adaptation of influenza viruses to new hosts is partly dependent on the distribution of the sialic acid (SA) receptors to which the viral hemagglutinin (HA) binds. Ferrets have been established as a valuable in vivo model of influenza virus pathogenesis and transmission because of similarities to humans in the distribution of HA receptors and in clinical signs of infection. In this study, we developed a ferret tracheal differentiated primary epithelial cell culture model that consisted of a layered epithelium structure with ciliated and nonciliated cells on its apical surface. We found that human-like (α2,6-linked) receptors predominated on ciliated cells, whereas avian-like (α2,3-linked) receptors, which were less abundant, were presented on nonciliated cells. When we compared the tropism and infectivity of three human (H1 and H3) and two avian (H1 and H5) influenza viruses, we observed that the human influenza viruses primarily infected ciliated cells and replicated efficiently, whereas a highly pathogenic avian H5N1 virus (A/Vietnam/1203/2004) replicated efficiently within nonciliated cells despite a low initial infection rate. Furthermore, compared to other influenza viruses tested, VN/1203 virus replicated more efficiently in cells isolated from the lower trachea and at a higher temperature (37°C) compared to a lower temperature (33°C). VN/1203 virus infection also induced higher levels of immune mediator genes and cell death, and virus was recovered from the basolateral side of the cell monolayer. This ferret tracheal differentiated primary epithelial cell culture system provides a valuable in vitro model for studying cellular tropism, infectivity, and the pathogenesis of influenza viruses. PMID:23255802

  1. Three-dimensional printed magnetophoretic system for the continuous flow separation of avian influenza H5N1 viruses.

    Science.gov (United States)

    Wang, Yuhe; Li, Yanbin; Wang, Ronghui; Wang, Maohua; Lin, Jianhan

    2017-04-01

    As a result of the low concentration of avian influenza viruses in samples for routine screening, the separation and concentration of these viruses are vital for their sensitive detection. We present a novel three-dimensional printed magnetophoretic system for the continuous flow separation of the viruses using aptamer-modified magnetic nanoparticles, a magnetophoretic chip, a magnetic field, and a fluidic controller. The magnetic field was designed based on finite element magnetic simulation and developed using neodymium magnets with a maximum intensity of 0.65 T and a gradient of 32 T/m for dragging the nanoparticle-virus complexes. The magnetophoretic chip was designed by SOLIDWORKS and fabricated by a three-dimensional printer with a magnetophoretic channel for the continuous flow separation of the viruses using phosphate-buffered saline as carrier flow. The fluidic controller was developed using a microcontroller and peristaltic pumps to inject the carrier flow and the viruses. The trajectory of the virus-nanoparticle complexes was simulated using COMSOL for optimization of the carrier flow and the magnetic field, respectively. The results showed that the H5N1 viruses could be captured, separated, and concentrated using the proposed magnetophoretic system with the separation efficiency up to 88% in a continuous flow separation time of 2 min for a sample volume of 200 μL. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Prolonged excretion of a low-pathogenicity H5N2 avian influenza virus strain in the Pekin duck

    Science.gov (United States)

    Carranza-Flores, José Manuel; Padilla-Noriega, Luis; Loza-Rubio, Elizabeth

    2013-01-01

    H5N2 strains of low-pathogenicity avian influenza virus (LPAIV) have been circulating for at least 17 years in some Mexican chicken farms. We measured the rate and duration of viral excretion from Pekin ducks that were experimentally inoculated with an H5N2 LPAIV that causes death in embryonated chicken eggs (A/chicken/Mexico/2007). Leghorn chickens were used as susceptible host controls. The degree of viral excretion was evaluated with real-time reverse transcriptase-polymerase chain reaction (RRT-PCR) using samples from oropharyngeal and cloacal swabs. We observed prolonged excretion from both species of birds lasting for at least 21 days. Prolonged excretion of LPAIV A/chicken/Mexico/2007 is atypical. PMID:23820212

  3. Genetic and phylogenetic characterizations of a novel genotype of highly pathogenic avian influenza (HPAI) H5N8 viruses in 2016/2017 in South Korea.

    Science.gov (United States)

    Kim, Young-Il; Park, Su-Jin; Kwon, Hyeok-Il; Kim, Eun-Ha; Si, Young-Jae; Jeong, Ju-Hwan; Lee, In-Won; Nguyen, Hiep Dinh; Kwon, Jin-Jung; Choi, Won Suk; Song, Min-Suk; Kim, Chul-Joong; Choi, Young-Ki

    2017-09-01

    During the outbreaks of highly pathogenic avian influenza (HPAI) H5N6 viruses in 2016 in South Korea, novel H5N8 viruses were also isolated from migratory birds. Phylogenetic analysis revealed that the HA gene of these H5N8 viruses belonged to clade 2.3.4.4, similarly to recent H5Nx viruses, and originated from A/Brk/Korea/Gochang1/14(H5N8), a minor lineage of H5N8 that appeared in 2014 and then disappeared. At least four reassortment events occurred with different subtypes (H5N8, H7N7, H3N8 and H10N7) and a chicken challenge study revealed that they were classified as HPAI viruses according to OIE criteria. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Genomic and Phylogenetic Characterization of Novel, Recombinant H5N2 Avian Influenza Virus Strains Isolated from Vaccinated Chickens with Clinical Symptoms in China

    Directory of Open Access Journals (Sweden)

    Huaiying Xu

    2015-02-01

    Full Text Available Infection of poultry with diverse lineages of H5N2 avian influenza viruses has been documented for over three decades in different parts of the world, with limited outbreaks caused by this highly pathogenic avian influenza virus. In the present study, three avian H5N2 influenza viruses, A/chicken/Shijiazhuang/1209/2013, A/chicken/Chiping/0321/2014, and A/chicken/Laiwu/0313/2014, were isolated from chickens with clinical symptoms of avian influenza. Complete genomic and phylogenetic analyses demonstrated that all three isolates are novel recombinant viruses with hemagglutinin (HA and matrix (M genes derived from H5N1, and remaining genes derived from H9N2-like viruses. The HA cleavage motif in all three strains (PQIEGRRRKR/GL is characteristic of a highly pathogenic avian influenza virus strain. These results indicate the occurrence of H5N2 recombination and highlight the importance of continued surveillance of the H5N2 subtype virus and reformulation of vaccine strains.

  5. Experimental infection of highly pathogenic avian influenza virus H5N1 in black-headed gulls (Chroicocephalus ridibundus).

    Science.gov (United States)

    Ramis, Antonio; van Amerongen, Geert; van de Bildt, Marco; Leijten, Loneke; Vanderstichel, Raphael; Osterhaus, Albert; Kuiken, Thijs

    2014-08-19

    Historically, highly pathogenic avian influenza viruses (HPAIV) rarely resulted in infection or clinical disease in wild birds. However, since 2002, disease and mortality from natural HPAIV H5N1 infection have been observed in wild birds including gulls. We performed an experimental HPAIV H5N1 infection of black-headed gulls (Chroicocephalus ridibundus) to determine their susceptibility to infection and disease from this virus, pattern of viral shedding, clinical signs, pathological changes and viral tissue distribution. We inoculated sixteen black-headed gulls with 1 × 10(4) median tissue culture infectious dose HPAIV H5N1 (A/turkey/Turkey/1/2005) intratracheally and intraesophageally. Birds were monitored daily until 12 days post inoculation (dpi). Oropharyngeal and cloacal swabs were collected daily to detect viral shedding. Necropsies from birds were performed at 2, 4, 5, 6, 7, and 12 dpi. Sampling from selected tissues was done for histopathology, immunohistochemical detection of viral antigen, PCR, and viral isolation. Our study shows that all inoculated birds were productively infected, developed systemic disease, and had a high morbidity and mortality rate. Virus was detected mainly in the respiratory tract on the first days after inoculation, and then concentrated more in pancreas and central nervous system from 4 dpi onwards. Birds shed infectious virus until 7 dpi from the pharynx and 6 dpi from the cloaca. We conclude that black-headed gulls are highly susceptible to disease with a high mortality rate and are thus more likely to act as sentinel species for the presence of the virus than as long-distance carriers of the virus to new geographical areas.

  6. Comprehensive global amino acid sequence analysis of PB1F2 protein of influenza A H5N1 viruses and the influenza A virus subtypes responsible for the 20th‐century pandemics

    Science.gov (United States)

    Pasricha, Gunisha; Mishra, Akhilesh C.; Chakrabarti, Alok K.

    2012-01-01

    Please cite this paper as: Pasricha et al. (2012) Comprehensive global amino acid sequence analysis of PB1F2 protein of influenza A H5N1 viruses and the Influenza A virus subtypes responsible for the 20th‐century pandemics. Influenza and Other Respiratory Viruses 7(4), 497–505. Background  PB1F2 is the 11th protein of influenza A virus translated from +1 alternate reading frame of PB1 gene. Since the discovery, varying sizes and functions of the PB1F2 protein of influenza A viruses have been reported. Selection of PB1 gene segment in the pandemics, variable size and pleiotropic effect of PB1F2 intrigued us to analyze amino acid sequences of this protein in various influenza A viruses. Methods  Amino acid sequences for PB1F2 protein of influenza A H5N1, H1N1, H2N2, and H3N2 subtypes were obtained from Influenza Research Database. Multiple sequence alignments of the PB1F2 protein sequences of the aforementioned subtypes were used to determine the size, variable and conserved domains and to perform mutational analysis. Results  Analysis showed that 96·4% of the H5N1 influenza viruses harbored full‐length PB1F2 protein. Except for the 2009 pandemic H1N1 virus, all the subtypes of the 20th‐century pandemic influenza viruses contained full‐length PB1F2 protein. Through the years, PB1F2 protein of the H1N1 and H3N2 viruses has undergone much variation. PB1F2 protein sequences of H5N1 viruses showed both human‐ and avian host‐specific conserved domains. Global database of PB1F2 protein revealed that N66S mutation was present only in 3·8% of the H5N1 strains. We found a novel mutation, N84S in the PB1F2 protein of 9·35% of the highly pathogenic avian influenza H5N1 influenza viruses. Conclusions  Varying sizes and mutations of the PB1F2 protein in different influenza A virus subtypes with pandemic potential were obtained. There was genetic divergence of the protein in various hosts which highlighted the host‐specific evolution of the virus

  7. Experimental infection of clade 1.1.2 (H5N1), clade 2.3.2.1c (H5N1) and clade 2.3.4.4 (H5N6) highly pathogenic avian influenza viruses in dogs.

    Science.gov (United States)

    Lyoo, K S; Na, W; Phan, L V; Yoon, S W; Yeom, M; Song, D; Jeong, D G

    2017-12-01

    Since the emergence of highly pathogenic avian influenza (HPAI) H5N1 in Asia, the haemagglutinin (HA) gene of this virus lineage has continued to evolve in avian populations, and H5N1 lineage viruses now circulate concurrently worldwide. Dogs may act as an intermediate host, increasing the potential for zoonotic transmission of influenza viruses. Virus transmission and pathologic changes in HPAI clade 1.1.2 (H5N1)-, 2.3.2.1c (H5N1)- and 2.3.4.4 (H5N6)-infected dogs were investigated. Mild respiratory signs and antibody response were shown in dogs intranasally infected with the viruses. Lung histopathology showed lesions that were associated with moderate interstitial pneumonia in the infected dogs. In this study, HPAI H5N6 virus replication in dogs was demonstrated for the first time. Dogs have been suspected as a "mixing vessel" for reassortments between avian and human influenza viruses to occur. The replication of these three subtypes of the H5 lineage of HPAI viruses in dogs suggests that dogs could serve as intermediate hosts for avian-human influenza virus reassortment if they are also co-infected with human influenza viruses. © 2017 Blackwell Verlag GmbH.

  8. Seroprevalence of Antibodies to Avian Influenza A (H5) and A (H9) Viruses among Market Poultry Workers, Hanoi, Vietnam, 2001

    OpenAIRE

    Uyeki, Timothy M.; Nguyen, Doan C.; Rowe, Thomas; Lu, Xiuhua; Hu-Primmer, Jean; Huynh, Lien P.; Hang, Nguyen L. K.; Katz, Jacqueline M.

    2012-01-01

    BACKGROUND: The frequency of avian influenza A virus infections among poultry workers is not well understood. METHODS: A seroprevalence study of market poultry workers and persons without occupational poultry exposure was conducted during 2001 in Hanoi, Vietnam. Sera were tested for avian influenza H5 and H9 antibodies by microneutralization and Western blot assays. RESULTS: Seroprevalence of H5 and H9 antibodies was 4% and 3% in poultry workers and 1% and 3.5% in non-poultry workers, respect...

  9. Persistence of highly pathogenic avian influenza H5N1 virus defined by agro-ecological niche

    Science.gov (United States)

    Hogerwerf, Lenny; Wallace, Rob G.; Ottaviani, Daniela; Slingenbergh, Jan; Prosser, Diann; Bergmann, Luc; Gilbert, Marius

    2010-01-01

    The highly pathogenic avian influenza (HPAI) H5N1 virus has spread across Eurasia and into Africa. Its persistence in a number of countries continues to disrupt poultry production, impairs smallholder livelihoods, and raises the risk a genotype adapted to human-to-human transmission may emerge. While previous studies identified domestic duck reservoirs as a primary risk factor associated with HPAI H5N1 persistence in poultry in Southeast Asia, little is known of such factors in countries with different agro-ecological conditions, and no study has investigated the impact of such conditions on HPAI H5N1 epidemiology at the global scale. This study explores the patterns of HPAI H5N1 persistence worldwide, and for China, Indonesia, and India includes individual provinces that have reported HPAI H5N1 presence during the 2004–2008 period. Multivariate analysis of a set of 14 agricultural, environmental, climatic, and socio-economic factors demonstrates in quantitative terms that a combination of six variables discriminates the areas with human cases and persistence: agricultural population density, duck density, duck by chicken density, chicken density, the product of agricultural population density and chicken output/input ratio, and purchasing power per capita. The analysis identifies five agro-ecological clusters, or niches, representing varying degrees of disease persistence. The agro-ecological distances of all study areas to the medoid of the niche with the greatest number of human cases are used to map HPAI H5N1 risk globally. The results indicate that few countries remain where HPAI H5N1 would likely persist should it be introduced.

  10. Reoccurrence of H5Nx clade 2.3.4.4 highly pathogenic avian influenza viruses in wild birds during 2016

    Science.gov (United States)

    The Asian-origin H5N1 A/goose/Guangdong/1/1996 (Gs/GD) lineage of high pathogenicity avian influenza viruses (HPAIV) has become widespread across four continents, affecting poultry, wild birds and humans. H5N1 HPAIV has evolved into multiple hemagglutinin (HA) genetic clades and reassorting with dif...

  11. Local amplification of highly pathogenic avian influenza H5N8 viruses in wild birds in the Netherlands, 2016 to 2017

    NARCIS (Netherlands)

    Poen, Marjolein J.; Bestebroer, Theo M.; Vuong, Oanh; Scheuer, Rachel D.; Jeugd, van der Henk P.; Kleyheeg, Erik; Eggink, Dirk; Lexmond, Pascal; Brand, van den Judith M.A.; Begeman, Lineke; Vliet, van der Stefan; Müskens, Gerhard J.D.M.; Majoor, Frank A.; Koopmans, Marion P.G.; Kuiken, Thijs; Fouchier, Ron A.M.

    2018-01-01

    Introduction: Highly pathogenic avian influenza (HPAI) viruses of subtype H5N8 were re-introduced into the Netherlands by late 2016, after detections in southeast Asia and Russia. This second H5N8 wave resulted in a large number of outbreaks in poultry farms and the deaths of large numbers of wild

  12. Local amplification of highly pathogenic avian influenza H5N8 viruses in wild birds in the Netherlands, 2016 to 2017

    NARCIS (Netherlands)

    Poen, Marjolein J; Bestebroer, Theo M; Vuong, Oanh; Scheuer, Rachel D; van der Jeugd, Henk P; Kleyheeg, Erik; Eggink, Dirk; Lexmond, Pascal; van den Brand, Judith M A; Begeman, Lineke; van der Vliet, Stefan; Müskens, Gerhard J D M; Majoor, Frank A; Koopmans, Marion P G; Kuiken, Thijs; Fouchier, Ron A M

    IntroductionHighly pathogenic avian influenza (HPAI) viruses of subtype H5N8 were re-introduced into the Netherlands by late 2016, after detections in south-east Asia and Russia. This second H5N8 wave resulted in a large number of outbreaks in poultry farms and the deaths of large numbers of wild

  13. Effect of Neuraminidase Inhibitor–Resistant Mutations on Pathogenicity of Clade 2.2 A/Turkey/15/06 (H5N1) Influenza Virus in Ferrets

    OpenAIRE

    Ilyushina, Natalia A.; Seiler, Jon P.; Rehg, Jerold E.; Webster, Robert G.; Govorkova, Elena A.

    2010-01-01

    The acquisition of neuraminidase (NA) inhibitor resistance by H5N1 influenza viruses has serious clinical implications, as this class of drugs can be an essential component of pandemic control measures. The continuous evolution of the highly pathogenic H5N1 influenza viruses results in the emergence of natural NA gene variations whose impact on viral fitness and NA inhibitor susceptibility are poorly defined. We generated seven genetically stable recombinant clade 2.2 A/Turkey/15/06-like (H5N...

  14. Genetic and biological characterization of three poultry-origin H5N6 avian influenza viruses with all internal genes from genotype S H9N2 viruses.

    Science.gov (United States)

    Liu, Kaituo; Gu, Min; Hu, Shunlin; Gao, Ruyi; Li, Juan; Shi, Liwei; Sun, Wenqi; Liu, Dong; Gao, Zhao; Xu, Xiulong; Hu, Jiao; Wang, Xiaoquan; Liu, Xiaowen; Chen, Sujuan; Peng, Daxin; Jiao, Xinan; Liu, Xiufan

    2018-04-01

    During surveillance for avian influenza viruses, three H5N6 viruses were isolated in chickens obtained from live bird markets in eastern China, between January 2015 and April 2016. Sequence analysis revealed a high genomic homology between these poultry isolates and recent human H5N6 variants whose internal genes were derived from genotype S H9N2 avian influenza viruses. Glycan binding assays revealed that all avian H5N6 viruses were capable of binding to both human-type SAα-2,6Gal receptors and avian-type SAα-2,3Gal receptors. Their biological characteristics were further studied in BALB/c mice, specific-pathogen-free chickens, and mallard ducks. All three isolates had low pathogenicity in mice but were highly pathogenic to chickens, as evidenced by 100% mortality 36-120 hours post infection at a low dose of 10 3.0 EID 50 and through effective contact transmission. Moreover, all three poultry H5N6 isolates caused asymptomatic infections in ducks, which may serve as a reservoir host for their maintenance and dissemination; these migrating waterfowl could cause a potential global pandemic. Our study suggests that continuous epidemiological surveillance in poultry should be implemented for the early prevention of future influenza outbreaks.

  15. Multiplex Reverse Transcription-Polymerase Chain Reaction untuk Deteksi Cepat Virus Flu Burung H5N1 (MULTIPLEX REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION FOR RAPID DETECTION OF H5N1 AVIAN INFLUENZA VIRUS

    Directory of Open Access Journals (Sweden)

    Raden Wasito

    2015-05-01

    Full Text Available Avian influenza virus subtype H5N1 (AIV H5N1 is highly pathogenic and fatal in poultry. The virusis still endemic with low virulence rate, although it may play a critical role in causing high morbidity andmortality rates in poultry in Indonesia. In general, diagnostic approach for AIV H5N1 is based onconventional serological and viral isolation methods that have the potential to produce consumings oftime and relatively expensive cost within the laboratory without compromising test utility. Thus, amolecular approach of multiplex reverse transcription-polymerase chain reaction (mRT-PCR was developedand applied for the detection of matrix gene type A influenza viruses, AIV subtype subtype H5hemagglutinin gene with simultaneous detection of N1 nucleoprotein gene. Thirty sera specimens fromthe diseased commercial chickens that were specifically amplified positive-RT-PCR for AIV H5N1 wereselected for mRT-PCR. The mRT-PCR products were visualized by agarose gel electrophoresis and consistedof DNA fragments of AIV of 245 bp, 545 bp and 343 bp for M, H5 and N1 genes, respectively. Thus, themRT-PCR that can rapidly differentiate simultaneously between these genes is very important for thecontrol and even eradication of AIV transmission in poultry in Indonesia.

  16. Biosecurity and Circulation of Influenza A (H5N1) Virus in Live-Bird Markets in Bangladesh, 2012.

    Science.gov (United States)

    Biswas, P K; Giasuddin, M; Nath, B K; Islam, M Z; Debnath, N C; Yamage, M

    2017-06-01

    Bangladesh has been considered as one of the five countries endemic with highly pathogenic avian influenza A subtype H5N1 (HPAI H5N1). Live-bird markets (LBMs) in south Asian countries are believed to play important roles in the transmission of HPAI H5N1 and others due to its central location as a hub of the poultry trading. Food and Agriculture Organization (FAO) of the United Nations has been promoting improved biosecurity in LBMs in Bangladesh. In 2012, by enrolling 32 large LBMs: 10 with FAO interventions and 22 without assistance, we assessed the virus circulation in the selected LBMs by applying standard procedures to investigate market floors, poultry stall floors, poultry-holding cases and slaughter areas and the overall biosecurity using a questionnaire-based survey. Relative risk (RR) was examined to compare the prevalence of HPAI H5N1 in the intervened and non-intervened LBMs. The measures practised in significantly more of the FAO-intervened LBMs included keeping of slaughter remnants in a closed container; decontamination of poultry vehicles at market place; prevention of crows' access to LBM, market/floor cleaning by market committee; wet cleaning; disinfection of floor/poultry stall after cleaning; and good supply of clean water at market (P Bangladesh regardless of interventions, albeit at lower levels than in other endemic countries. © 2015 Blackwell Verlag GmbH.

  17. Generation, characterization and epitope mapping of two neutralizing and protective human recombinant antibodies against influenza A H5N1 viruses.

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    Lina Sun

    Full Text Available BACKGROUND: The development of new therapeutic targets and strategies to control highly pathogenic avian influenza (HPAI H5N1 virus infection in humans is urgently needed. Broadly cross-neutralizing recombinant human antibodies obtained from the survivors of H5N1 avian influenza provide an important role in immunotherapy for human H5N1 virus infection and definition of the critical epitopes for vaccine development. METHODOLOGY/PRINCIPAL FINDINGS: We have characterized two recombinant baculovirus-expressed human antibodies (rhAbs, AVFluIgG01 and AVFluIgG03, generated by screening a Fab antibody phage library derived from a patient recovered from infection with a highly pathogenic avian influenza A H5N1 clade 2.3 virus. AVFluIgG01 cross-neutralized the most of clade 0, clade 1, and clade 2 viruses tested, in contrast, AVFluIgG03 only neutralized clade 2 viruses. Passive immunization of mice with either AVFluIgG01 or AVFluIgG03 antibody resulted in protection from a lethal H5N1 clade 2.3 virus infection. Furthermore, through epitope mapping, we identify two distinct epitopes on H5 HA molecule recognized by these rhAbs and demonstrate their potential to protect against a lethal H5N1 virus infection in a mouse model. CONCLUSIONS/SIGNIFICANCE: Importantly, localization of the epitopes recognized by these two neutralizing and protective antibodies has provided, for the first time, insight into the human antibody responses to H5N1 viruses which contribute to the H5 immunity in the recovered patient. These results highlight the potential of a rhAbs treatment strategy for human H5N1 virus infection and provide new insight for the development of effective H5N1 pandemic vaccines.

  18. Airborne Detection of H5N8 Highly Pathogenic Avian Influenza Virus Genome in Poultry Farms, France.

    Science.gov (United States)

    Scoizec, Axelle; Niqueux, Eric; Thomas, Rodolphe; Daniel, Patrick; Schmitz, Audrey; Le Bouquin, Sophie

    2018-01-01

    In southwestern France, during the winter of 2016-2017, the rapid spread of highly pathogenic avian influenza H5N8 outbreaks despite the implementation of routine control measures, raised the question about the potential role of airborne transmission in viral spread. As a first step to investigate the plausibility of that transmission, air samples were collected inside, outside and downwind from infected duck and chicken facilities. H5 avian influenza virus RNA was detected in all samples collected inside poultry houses, at external exhaust fans and at 5 m distance from poultry houses. For three of the five flocks studied, in the sample collected at 50-110 m distance, viral genomic RNA was detected. The measured viral air concentrations ranged between 4.3 and 6.4 log 10 RNA copies per m 3 , and their geometric mean decreased from external exhaust fans to the downwind measurement point. These findings are in accordance with the possibility of airborne transmission and question the procedures for outbreak depopulation.

  19. Airborne Detection of H5N8 Highly Pathogenic Avian Influenza Virus Genome in Poultry Farms, France

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    Axelle Scoizec

    2018-02-01

    Full Text Available In southwestern France, during the winter of 2016–2017, the rapid spread of highly pathogenic avian influenza H5N8 outbreaks despite the implementation of routine control measures, raised the question about the potential role of airborne transmission in viral spread. As a first step to investigate the plausibility of that transmission, air samples were collected inside, outside and downwind from infected duck and chicken facilities. H5 avian influenza virus RNA was detected in all samples collected inside poultry houses, at external exhaust fans and at 5 m distance from poultry houses. For three of the five flocks studied, in the sample collected at 50–110 m distance, viral genomic RNA was detected. The measured viral air concentrations ranged between 4.3 and 6.4 log10 RNA copies per m3, and their geometric mean decreased from external exhaust fans to the downwind measurement point. These findings are in accordance with the possibility of airborne transmission and question the procedures for outbreak depopulation.

  20. Protection of White Leghorn chickens by U.S. emergency H5 vaccination against clade 2.3.4.4 H5N2 high pathogenicity avian influenza virus.

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    Bertran, Kateri; Balzli, Charles; Lee, Dong-Hun; Suarez, David L; Kapczynski, Darrell R; Swayne, David E

    2017-11-01

    During December 2014-June 2015, the U.S. experienced a high pathogenicity avian influenza (HPAI) outbreak caused by clade 2.3.4.4 H5Nx Goose/Guangdong lineage viruses with devastating consequences for the poultry industry. Three vaccines, developed based on updating existing registered vaccines or currently licensed technologies, were evaluated for possible use: an inactivated reverse genetics H5N1 vaccine (rgH5N1) and an RNA particle vaccine (RP-H5), both containing the hemagglutinin gene of clade 2.3.4.4 strain, and a recombinant herpesvirus turkey vectored vaccine (rHVT-H5) containing the hemagglutinin gene of clade 2.2 strain. The efficacy of the three vaccines, alone or in combination, was assessed in White Leghorn chickens against clade 2.3.4.4 H5N2 HPAI virus challenge. In Study 1, single (rHVT-H5) and prime-boost (rHVT-H5+rgH5N1 or rHVT-H5+RP-H5) vaccination strategies protected chickens with high levels of protective immunity and significantly reduced virus shedding. In Study 2, single vaccination with either rgH5N1 or RP-H5 vaccines provided clinical protection in adult chickens and significantly reduced virus shedding. In Study 3, double rgH5N1 vaccination protected adult chickens from clinical signs and mortality when challenged 20weeks post-boost, with high levels of long-lasting protective immunity and significantly reduced virus shedding. These studies support the use of genetically related vaccines, possibly in combination with a broad protective priming vaccine, for emergency vaccination programs against clade 2.3.4.4 H5Nx HPAI virus in young and adult layer chickens. Published by Elsevier Ltd.

  1. Preparasi Imunoglobulin G Kelinci sebagai Antigen Penginduksi Antibodi Spesifik Terhadap Virus Avian Influenza H5N1 Strain Legok

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    Ketut Karuni Nyanakumari Natih

    2010-06-01

    Full Text Available The aim of this research was to prepare rabbit Immunoglobulin G as anti-idiotype antibody (Ab2 ofAvian Influenza Virus (AIV H5N1. A polyclonal antibody was collected from guinea pigs immunized withinactivated AI vaccine H5N1of Legok strain. Antibody of H5N1 AI in serum was detected by Agar gelprecipitation test (AGPT and an Inhibition Hemmaglutination test (IHT. The highest titre of antibodywas obtained one week after the third immunization. Serum of guinea pigs containing IgG was purifiedusing the Montage Antibody purification kit & spin column with Prosep A media (Millipore. The AI H5N1IgG concentration was 8 mg/ml. AI H5N1 IgG, was then digested with pepsin to obtain F(ab2 fraction andwas called Ab1. The concentration of IgG and F(ab2 and purity of IgG were determined by UVspectrophotometer which showed Ab1 concentration 1 mg/ml. Molecular weight was estimated by sodiumdodecyl sulfate- polyacrilamide gel electrophoresis (SDS-PAGE. Ab2 was produced by immunization ofrabbit with Ab1. The first immunization was carried out by subcutaneous injection with 500 ?g of Ab1emulsified in Complete Freund Adjuvant. The immunization was repeated with the same dose of Ab1emulsified in Incomplete Freund Adjuvan at 1 week intervals. One week after the second immunization,rabbit’s serum was harvested and IgG was purified using the Montage Antibody purification kit & spincolumn with Prosep A media (Millipore. The rabbit IgG, called Ab2, was an anti-idiotypic antibody againstAIV-H5N1. In AGPT, a precipitation line appeared between Ab1 and Ab2. A partial reaction appearedbetween Ab2 and the AI H5N1 antigen was also detected. The results indicated that Ab2 is a possiblecandidate of imunogen for protection against an AI virus H5N1 infection.

  2. High-yield production of a stable Vero cell-based vaccine candidate against the highly pathogenic avian influenza virus H5N1

    International Nuclear Information System (INIS)

    Zhou, Fangye; Zhou, Jian; Ma, Lei; Song, Shaohui; Zhang, Xinwen; Li, Weidong; Jiang, Shude; Wang, Yue; Liao, Guoyang

    2012-01-01

    Highlights: ► Vero cell-based HPAI H5N1 vaccine with stable high yield. ► Stable high yield derived from the YNVa H3N2 backbone. ► H5N1/YNVa has a similar safety and immunogenicity to H5N1delta. -- Abstract: Highly pathogenic avian influenza (HPAI) viruses pose a global pandemic threat, for which rapid large-scale vaccine production technology is critical for prevention and control. Because chickens are highly susceptible to HPAI viruses, the supply of chicken embryos for vaccine production might be depleted during a virus outbreak. Therefore, developing HPAI virus vaccines using other technologies is critical. Meeting vaccine demand using the Vero cell-based fermentation process has been hindered by low stability and yield. In this study, a Vero cell-based HPAI H5N1 vaccine candidate (H5N1/YNVa) with stable high yield was achieved by reassortment of the Vero-adapted (Va) high growth A/Yunnan/1/2005(H3N2) (YNVa) virus with the A/Anhui/1/2005(H5N1) attenuated influenza vaccine strain (H5N1delta) using the 6/2 method. The reassorted H5N1/YNVa vaccine maintained a high hemagglutination (HA) titer of 1024. Furthermore, H5N1/YNVa displayed low pathogenicity and uniform immunogenicity compared to that of the parent virus.

  3. High-yield production of a stable Vero cell-based vaccine candidate against the highly pathogenic avian influenza virus H5N1

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    Zhou, Fangye; Zhou, Jian; Ma, Lei; Song, Shaohui; Zhang, Xinwen; Li, Weidong; Jiang, Shude [No. 5, Department of Bioproducts, Institute of Medical Biology, Chinese Academy of Medical Science and Pecking Union Medical College, Jiaoling Avenue 935, Kunming, Yunnan Province 650102, People' s Republic of China (China); Wang, Yue [National Institute for Viral Disease Control and Prevention, China Center for Disease Control and Prevention, Yingxin Lane 100, Xicheng District, Beijing 100052, People' s Republic of China (China); Liao, Guoyang [No. 5, Department of Bioproducts, Institute of Medical Biology, Chinese Academy of Medical Science and Pecking Union Medical College, Jiaoling Avenue 935, Kunming, Yunnan Province 650102, People' s Republic of China (China)

    2012-05-18

    Highlights: Black-Right-Pointing-Pointer Vero cell-based HPAI H5N1 vaccine with stable high yield. Black-Right-Pointing-Pointer Stable high yield derived from the YNVa H3N2 backbone. Black-Right-Pointing-Pointer H5N1/YNVa has a similar safety and immunogenicity to H5N1delta. -- Abstract: Highly pathogenic avian influenza (HPAI) viruses pose a global pandemic threat, for which rapid large-scale vaccine production technology is critical for prevention and control. Because chickens are highly susceptible to HPAI viruses, the supply of chicken embryos for vaccine production might be depleted during a virus outbreak. Therefore, developing HPAI virus vaccines using other technologies is critical. Meeting vaccine demand using the Vero cell-based fermentation process has been hindered by low stability and yield. In this study, a Vero cell-based HPAI H5N1 vaccine candidate (H5N1/YNVa) with stable high yield was achieved by reassortment of the Vero-adapted (Va) high growth A/Yunnan/1/2005(H3N2) (YNVa) virus with the A/Anhui/1/2005(H5N1) attenuated influenza vaccine strain (H5N1delta) using the 6/2 method. The reassorted H5N1/YNVa vaccine maintained a high hemagglutination (HA) titer of 1024. Furthermore, H5N1/YNVa displayed low pathogenicity and uniform immunogenicity compared to that of the parent virus.

  4. Influenza H5N1 and H1N1 virus replication and innate immune responses in bronchial epithelial cells are influenced by the state of differentiation.

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    Renee W Y Chan

    Full Text Available Influenza H5N1 virus continues to be enzootic in poultry and transmits zoonotically to humans. Although a swine-origin H1N1 virus has emerged to become pandemic, its virulence for humans remains modest in comparison to that seen in zoonotic H5N1 disease. As human respiratory epithelium is the primary target cells for influenza viruses, elucidating the viral tropism and host innate immune responses of influenza H5N1 virus in human bronchial epithelium may help to understand the pathogenesis. Here we established primary culture of undifferentiated and well differentiated normal human bronchial epithelial (NHBE cells and infected with highly pathogenic influenza H5N1 virus (A/Vietnam/3046/2004 and a seasonal influenza H1N1 virus (A/Hong Kong/54/1998, the viral replication kinetics and cytokine and chemokine responses were compared by qPCR and ELISA. We found that the in vitro culture of the well differentiated NHBE cells acquired the physiological properties of normal human bronchi tissue which express high level of alpha2-6-linked sialic acid receptors and human airway trypsin-like (HAT protease, in contrast to the low expression in the non-differentiated NHBE cells. When compared to H1N1 virus, the H5N1 virus replicated more efficiently and induced a stronger type I interferon response in the undifferentiated NHBE cells. In contrast, in well differentiated cultures, H5N1 virus replication was less efficient and elicited a lower interferon-beta response in comparison with H1N1 virus. Our data suggest that the differentiation of bronchial epithelial cells has a major influence in cells' permissiveness to human H1N1 and avian H5N1 viruses and the host innate immune responses. The reduced virus replication efficiency partially accounts for the lower interferon-beta responses in influenza H5N1 virus infected well differentiated NHBE cells. Since influenza infection in the bronchial epithelium will lead to tissue damage and associate with the

  5. Novel H5N8 clade 2.3.4.4 highly pathogenic avian influenza virus in wild awuatic birds, Russia, 2016

    Science.gov (United States)

    H5N1 high pathogenicity avian influenza virus (HPAIV) emerged in 1996 in Guangdong China (Gs/GD) and has evolved into multiple genetic clades. Since 2008, HPAIV H5 clade 2.3.4 with N2, N5 and N8 neuraminidase subtypes have been identified in mainland China and outbreak of HPAIV H5N8 clade 2.3.4.4 ou...

  6. Passive immunotherapy for influenza A H5N1 virus infection with equine hyperimmune globulin F(ab'2 in mice

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    Li Yanbin

    2006-03-01

    Full Text Available Abstract Background Avian influenza virus H5N1 has demonstrated considerable pandemic potential. Currently, no effective vaccines for H5N1 infection are available, so passive immunotherapy may be an alternative strategy. To investigate the possible therapeutic effect of antibody against highly pathogenic H5N1 virus on a mammal host, we prepared specific equine anti-H5N1 IgGs from horses vaccinated with inactivated H5N1 virus, and then obtained the F(ab'2 fragments by pepsin digestion of IgGs. Methods The horses were vaccinated with inactivated H5N1 vaccine to prepare anti-H5N1 IgGs. The F(ab'2 fragments were purified from anti-H5N1 hyperimmune sera by a protocol for 'enhanced pepsin digestion'. The protective effect of the F(ab'2 fragments against H5N1 virus infection was determined in cultured MDCK cells by cytopathic effect (CPE assay and in a BALB/c mouse model by survival rate assay. Results By the protocol for 'enhanced pepsin digestion', total 16 g F(ab'2 fragments were finally obtained from one liter equine antisera with the purity of over 90%. The H5N1-specific F(ab'2 fragments had a HI titer of 1:1024, and the neutralization titre of F(ab'2 reached 1: 2048. The in vivo assay showed that 100 μg of the F(ab'2 fragments could protect BALB/c mice infected with a lethal dose of influenza H5N1 virus. Conclusion The availability of highly purified H5N1-specific F(ab'2 fragments may be promising for treatment of influenza H5N1 infection. Our work has provided experimental support for the application of the therapeutic equine immunoglobulin in future large primate or human trials.

  7. Effect of species, breed and route of virus inoculation on the pathogenicity of H5N1 highly pathogenic influenza (HPAI) viruses in domestic ducks.

    Science.gov (United States)

    Pantin-Jackwood, Mary; Swayne, David E; Smith, Diane; Shepherd, Eric

    2013-07-22

    H5N1 highly pathogenic avian influenza (HPAI) viruses continue to be a threat to poultry in many regions of the world. Domestic ducks have been recognized as one of the primary factors in the spread of H5N1 HPAI. In this study we examined the pathogenicity of H5N1 HPAI viruses in different species and breeds of domestic ducks and the effect of route of virus inoculation on the outcome of infection. We determined that the pathogenicity of H5N1 HPAI viruses varies between the two common farmed duck species, with Muscovy ducks (Cairina moschata) presenting more severe disease than various breeds of Anas platyrhynchos var. domestica ducks including Pekin, Mallard-type, Black Runners, Rouen, and Khaki Campbell ducks. We also found that Pekin and Muscovy ducks inoculated with two H5N1 HPAI viruses of different virulence, given by any one of three routes (intranasal, intracloacal, or intraocular), became infected with the viruses. Regardless of the route of inoculation, the outcome of infection was similar for each species but depended on the virulence of the virus used. Muscovy ducks showed more severe clinical signs and higher mortality than the Pekin ducks. In conclusion, domestic ducks are susceptible to H5N1 HPAI virus infection by different routes of exposure, but the presentation of the disease varied by virus strain and duck species. This information helps support the planning and implementation of H5N1 HPAI surveillance and control measures in countries with large domestic duck populations.

  8. Isolation and Characterization of Avian Influenza Viruses, Including Highly Pathogenic H5N1, from Poultry in Live Bird Markets in Hanoi, Vietnam, in 2001

    Science.gov (United States)

    Nguyen, Doan C.; Uyeki, Timothy M.; Jadhao, Samadhan; Maines, Taronna; Shaw, Michael; Matsuoka, Yumiko; Smith, Catherine; Rowe, Thomas; Lu, Xiuhua; Hall, Henrietta; Xu, Xiyan; Balish, Amanda; Klimov, Alexander; Tumpey, Terrence M.; Swayne, David E.; Huynh, Lien P. T.; Nghiem, Ha K.; Nguyen, Hanh H. T.; Hoang, Long T.; Cox, Nancy J.; Katz, Jacqueline M.

    2005-01-01

    Since 1997, outbreaks of highly pathogenic (HP) H5N1 and circulation of H9N2 viruses among domestic poultry in Asia have posed a threat to public health. To better understand the extent of transmission of avian influenza viruses (AIV) to humans in Asia, we conducted a cross-sectional virologic study in live bird markets (LBM) in Hanoi, Vietnam, in October 2001. Specimens from 189 birds and 18 environmental samples were collected at 10 LBM. Four influenza A viruses of the H4N6 (n = 1), H5N2 (n = 1), and H9N3 (n = 2) subtypes were isolated from healthy ducks for an isolation frequency of over 30% from this species. Two H5N1 viruses were isolated from healthy geese. The hemagglutinin (HA) genes of these H5N1 viruses possessed multiple basic amino acid motifs at the cleavage site, were HP for experimentally infected chickens, and were thus characterized as HP AIV. These HA genes shared high amino acid identities with genes of other H5N1 viruses isolated in Asia during this period, but they were genetically distinct from those of H5N1 viruses isolated from poultry and humans in Vietnam during the early 2004 outbreaks. These viruses were not highly virulent for experimentally infected ducks, mice, or ferrets. These results establish that HP H5N1 viruses with properties similar to viruses isolated in Hong Kong and mainland China circulated in Vietnam as early as 2001, suggest a common source for H5N1 viruses circulating in these Asian countries, and provide a framework to better understand the recent widespread emergence of HP H5N1 viruses in Asia. PMID:15767421

  9. Potency of whole virus particle and split virion vaccines using dissolving microneedle against challenges of H1N1 and H5N1 influenza viruses in mice.

    Science.gov (United States)

    Nakatsukasa, Akihiro; Kuruma, Koji; Okamatsu, Masatoshi; Hiono, Takahiro; Suzuki, Mizuho; Matsuno, Keita; Kida, Hiroshi; Oyamada, Takayoshi; Sakoda, Yoshihiro

    2017-05-15

    Transdermal vaccination using a microneedle (MN) confers enhanced immunity compared with subcutaneous (SC) vaccination. Here we developed a novel dissolving MN patch for the influenza vaccine. The potencies of split virion and whole virus particle (WVP) vaccines prepared from A/Puerto Rico/8/1934 (H1N1) and A/duck/Hokkaido/Vac-3/2007 (H5N1), respectively, were evaluated. MN vaccination induced higher neutralizing antibody responses than SC vaccination in mice. Moreover, MN vaccination with a lower dose of antigens conferred protective immunity against lethal challenges of influenza viruses than SC vaccination in mice. These results suggest that the WVP vaccines administered using MN are an effective combination for influenza vaccine to be further validated in humans. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. TEST KIT FOR THE DETECTION AND GENOTYPING OF HIGHLY PATHOGENIC INFLUENZA VIRUS A H5N1 BY REAL-TIME POLYMERASE CHAIN REACTION

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    S. V. Stepaniuk

    2014-06-01

    Full Text Available Results of the annual monitoring of epizooties indicate that highly pathogenic HPAI/H5N1 avian influenza widely circulated in Eurasian region. Over a period of 2010–2013 years more than 165 cases of outbreaks in 14 countries were found out. Ukraine became one of the first countries in Europe where in Autonomous Republic of Crimea in October 2005 outbreak of avian epizootic with HPAI/H5N1 was documented and until February 2008 more than 236,000 poultry were killed. Since then the question of monitoring of infected both migrating birds and poultry in places of cross contact in Ukraine remains of high priority. The test system is developed for identification and genotyping A H5N1 on three genes (M, H5 and N1 HPAI/H5N1 in real-time mode for polymerase chain reaction. Test kit capacity to detect HPAI/h5n1avian influenza virus and differentiate it from the other viral infection agents of birds and animals were studied by testing of HPAI/H5N1 virus isolated during mass infection outbreak in Crimea in 2005 and cultural specimens of other viral pathogens. It was established that the «DIA Real Avian Influenza» test kit was capable to detect RNA influenza A virus of high pathogenic H5N1 strains having high sensitivity (100% while RNA of the Crimean HPAI/H5N1 isolate studying and specificity (100% while RNA viruses of Newcastle birds disease, fowl powershift, syndrome of drop in egg production and horse influenza studying.

  11. Seroprevalence of antibodies to avian influenza A (H5 and A (H9 viruses among market poultry workers, Hanoi, Vietnam, 2001.

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    Timothy M Uyeki

    Full Text Available BACKGROUND: The frequency of avian influenza A virus infections among poultry workers is not well understood. METHODS: A seroprevalence study of market poultry workers and persons without occupational poultry exposure was conducted during 2001 in Hanoi, Vietnam. Sera were tested for avian influenza H5 and H9 antibodies by microneutralization and Western blot assays. RESULTS: Seroprevalence of H5 and H9 antibodies was 4% and 3% in poultry workers and 1% and 3.5% in non-poultry workers, respectively. CONCLUSIONS: Seroprevalence of H5 and H9 antibodies was low among Hanoi market poultry workers in 2001, but can serve as a baseline for additional studies.

  12. Seroprevalence of Antibodies to Avian Influenza A (H5) and A (H9) Viruses among Market Poultry Workers, Hanoi, Vietnam, 2001

    Science.gov (United States)

    Rowe, Thomas; Lu, Xiuhua; Hu-Primmer, Jean; Huynh, Lien P.; Hang, Nguyen L. K.; Katz, Jacqueline M.

    2012-01-01

    Background The frequency of avian influenza A virus infections among poultry workers is not well understood. Methods A seroprevalence study of market poultry workers and persons without occupational poultry exposure was conducted during 2001 in Hanoi, Vietnam. Sera were tested for avian influenza H5 and H9 antibodies by microneutralization and Western blot assays. Results Seroprevalence of H5 and H9 antibodies was 4% and 3% in poultry workers and 1% and 3.5% in non-poultry workers, respectively. Conclusions Seroprevalence of H5 and H9 antibodies was low among Hanoi market poultry workers in 2001, but can serve as a baseline for additional studies. PMID:22928049

  13. Immunity to current H5 highly pathogenic avian influenza viruses: From vaccines to adaptive immunity in wild birds

    Science.gov (United States)

    Following the 2014-2015 outbreaks of H5N2 and H5N8 highly pathogenic avian influenza (HPAI) in the U.S., studies were performed to assess the immunity required for protection against future outbreaks should they occur. We assessed the ability of vaccines to induce protection of chickens and turkeys...

  14. Could Changes in the Agricultural Landscape of Northeastern China Have Influenced the Long-Distance Transmission of Highly Pathogenic Avian Influenza H5Nx Viruses?

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    Marius Gilbert

    2017-12-01

    Full Text Available In the last few years, several reassortant subtypes of highly pathogenic avian influenza viruses (HPAI H5Nx have emerged in East Asia. These new viruses, mostly of subtype H5N1, H5N2, H5N6, and H5N8 belonging to clade 2.3.4.4, have been found in several Asian countries and have caused outbreaks in poultry in China, South Korea, and Vietnam. HPAI H5Nx also have spread over considerable distances with the introduction of viruses belonging to the same 2.3.4.4 clade in the U.S. (2014–2015 and in Europe (2014–2015 and 2016–2017. In this paper, we examine the emergence and spread of these new viruses in Asia in relation to published datasets on HPAI H5Nx distribution, movement of migratory waterfowl, avian influenza risk models, and land-use change analyses. More specifically, we show that between 2000 and 2015, vast areas of northeast China have been newly planted with rice paddy fields (3.21 million ha in Heilongjiang, Jilin, and Liaoning in areas connected to other parts of Asia through migratory pathways of wild waterfowl. We hypothesize that recent land use changes in northeast China have affected the spatial distribution of wild waterfowl, their stopover areas, and the wild-domestic interface, thereby altering transmission dynamics of avian influenza viruses across flyways. Detailed studies of the habitat use by wild migratory birds, of the extent of the wild–domestic interface, and of the circulation of avian influenza viruses in those new planted areas may help to shed more light on this hypothesis, and on the possible impact of those changes on the long-distance patterns of avian influenza transmission.

  15. An H5N1-based matrix protein 2 ectodomain tetrameric peptide vaccine provides cross-protection against lethal infection with H7N9 influenza virus.

    Science.gov (United States)

    Leung, Ho-Chuen; Chan, Chris Chung-Sing; Poon, Vincent Kwok-Man; Zhao, Han-Jun; Cheung, Chung-Yan; Ng, Fai; Huang, Jian-Dong; Zheng, Bo-Jian

    2015-04-01

    In March 2013, a patient infected with a novel avian influenza A H7N9 virus was reported in China. Since then, there have been 458 confirmed infection cases and 177 deaths. The virus contains several human-adapted markers, indicating that H7N9 has pandemic potential. The outbreak of this new influenza virus highlighted the need for the development of universal influenza vaccines. Previously, we demonstrated that a tetrameric peptide vaccine based on the matrix protein 2 ectodomain (M2e) of the H5N1 virus (H5N1-M2e) could protect mice from lethal infection with different clades of H5N1 and 2009 pandemic H1N1 influenza viruses. In this study, we investigated the cross-protection of H5N1-M2e against lethal infection with the new H7N9 virus. Although five amino acid differences existed at positions 13, 14, 18, 20, and 21 between M2e of H5N1 and H7N9, H5N1-M2e vaccination with either Freund's adjuvant or the Sigma adjuvant system (SAS) induced a high level of anti-M2e antibody, which cross-reacted with H7N9-M2e peptide. A mouse-adapted H7N9 strain, A/Anhui/01/2013m, was used for lethal challenge in animal experiments. H5N1-M2e vaccination provided potent cross-protection against lethal challenge of the H7N9 virus. Reduced viral replication and histopathological damage of mouse lungs were also observed in the vaccinated mice. Our results suggest that the tetrameric H5N1-M2e peptide vaccine could protect against different subtypes of influenza virus infections. Therefore, this vaccine may be an ideal candidate for developing a universal vaccine to prevent the reemergence of avian influenza A H7N9 virus and the emergence of potential novel reassortants of influenza virus.

  16. Acquisition of human-type receptor binding specificity by new H5N1 influenza virus sublineages during their emergence in birds in Egypt.

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    Yohei Watanabe

    2011-05-01

    Full Text Available Highly pathogenic avian influenza A virus subtype H5N1 is currently widespread in Asia, Europe, and Africa, with 60% mortality in humans. In particular, since 2009 Egypt has unexpectedly had the highest number of human cases of H5N1 virus infection, with more than 50% of the cases worldwide, but the basis for this high incidence has not been elucidated. A change in receptor binding affinity of the viral hemagglutinin (HA from α2,3- to α2,6-linked sialic acid (SA is thought to be necessary for H5N1 virus to become pandemic. In this study, we conducted a phylogenetic analysis of H5N1 viruses isolated between 2006 and 2009 in Egypt. The phylogenetic results showed that recent human isolates clustered disproportionally into several new H5 sublineages suggesting that their HAs have changed their receptor specificity. Using reverse genetics, we found that these H5 sublineages have acquired an enhanced binding affinity for α2,6 SA in combination with residual affinity for α2,3 SA, and identified the amino acid mutations that produced this new receptor specificity. Recombinant H5N1 viruses with a single mutation at HA residue 192 or a double mutation at HA residues 129 and 151 had increased attachment to and infectivity in the human lower respiratory tract but not in the larynx. These findings correlated with enhanced virulence of the mutant viruses in mice. Interestingly, these H5 viruses, with increased affinity to α2,6 SA, emerged during viral diversification in bird populations and subsequently spread to humans. Our findings suggested that emergence of new H5 sublineages with α2,6 SA specificity caused a subsequent increase in human H5N1 influenza virus infections in Egypt, and provided data for understanding the virus's pandemic potential.

  17. Novel H7N2 and H5N6 Avian Influenza A Viruses in Sentinel Chickens: A Sentinel Chicken Surveillance Study

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    Teng Zhao

    2016-11-01

    Full Text Available In 2014, surveillance of sentinel chicken for avian influenza virus was conducted in aquatic bird habitat near Wuxi City, Jiangsu Province, China. Two H7N2, one H5N6, and two H9N2 viruses were isolated. Sequence analysis revealed that the H7N2 virus is a novel reassortant of H7N9 and H9N2 viruses and H5N6 virus is a reassortant of H5N1 clade 2.3.4 and H6N6 viruses. Substitutions V186 and L226 (H3 numbering in the hemagglutinin (HA gene protein was found in two H7N2 viruses but not in the H5N6 virus. Two A138 and A160 mutations were identified in the HA gene protein of all three viruses but a P128 mutation was only in the H5N6 virus. A deletion of three and eleven amino acids in the neuraminidase stalk region was found in two H7N2 and H5N6 viruses, respectively. Moreover, a mutation of N31 in M2 protein was observed in both two H7N2 viruses. High similarity of these isolated viruses to viruses previously identified among poultry and humans, suggests that peridomestic aquatic birds may play a role in sustaining novel virus transmission. Therefore, continued surveillance is needed to monitor these avian influenza viruses in wild bird and domestic poultry that may pose a threat to poultry and human health.

  18. In vitro assessment of attachment pattern and replication efficiency of H5N1 influenza A viruses with altered receptor specificity.

    Science.gov (United States)

    Chutinimitkul, Salin; van Riel, Debby; Munster, Vincent J; van den Brand, Judith M A; Rimmelzwaan, Guus F; Kuiken, Thijs; Osterhaus, Albert D M E; Fouchier, Ron A M; de Wit, Emmie

    2010-07-01

    The continuous circulation of the highly pathogenic avian influenza (HPAI) H5N1 virus has been a cause of great concern. The possibility of this virus acquiring specificity for the human influenza A virus receptor, alpha2,6-linked sialic acids (SA), and being able to transmit efficiently among humans is a constant threat to human health. Different studies have described amino acid substitutions in hemagglutinin (HA) of clinical HPAI H5N1 isolates or that were introduced experimentally that resulted in an increased, but not exclusive, binding of these virus strains to alpha2,6-linked SA. We introduced all previously described amino acid substitutions and combinations thereof into a single genetic background, influenza virus A/Indonesia/5/05 HA, and tested the receptor specificity of these 27 mutant viruses. The attachment pattern to ferret and human tissues of the upper and lower respiratory tract of viruses with alpha2,6-linked SA receptor preference was then determined and compared to the attachment pattern of a human influenza A virus (H3N2). At least three mutant viruses showed an attachment pattern to the human respiratory tract similar to that of the human H3N2 virus. Next, the replication efficiencies of these mutant viruses and the effects of three different neuraminidases on virus replication were determined. These data show that influenza virus A/Indonesia/5/05 potentially requires only a single amino acid substitution to acquire human receptor specificity, while at the same time remaining replication competent, thus suggesting that the pandemic threat posed by HPAI H5N1 is far from diminished.

  19. In Vitro Assessment of Attachment Pattern and Replication Efficiency of H5N1 Influenza A Viruses with Altered Receptor Specificity▿

    Science.gov (United States)

    Chutinimitkul, Salin; van Riel, Debby; Munster, Vincent J.; van den Brand, Judith M. A.; Rimmelzwaan, Guus F.; Kuiken, Thijs; Osterhaus, Albert D. M. E.; Fouchier, Ron A. M.; de Wit, Emmie

    2010-01-01

    The continuous circulation of the highly pathogenic avian influenza (HPAI) H5N1 virus has been a cause of great concern. The possibility of this virus acquiring specificity for the human influenza A virus receptor, α2,6-linked sialic acids (SA), and being able to transmit efficiently among humans is a constant threat to human health. Different studies have described amino acid substitutions in hemagglutinin (HA) of clinical HPAI H5N1 isolates or that were introduced experimentally that resulted in an increased, but not exclusive, binding of these virus strains to α2,6-linked SA. We introduced all previously described amino acid substitutions and combinations thereof into a single genetic background, influenza virus A/Indonesia/5/05 HA, and tested the receptor specificity of these 27 mutant viruses. The attachment pattern to ferret and human tissues of the upper and lower respiratory tract of viruses with α2,6-linked SA receptor preference was then determined and compared to the attachment pattern of a human influenza A virus (H3N2). At least three mutant viruses showed an attachment pattern to the human respiratory tract similar to that of the human H3N2 virus. Next, the replication efficiencies of these mutant viruses and the effects of three different neuraminidases on virus replication were determined. These data show that influenza virus A/Indonesia/5/05 potentially requires only a single amino acid substitution to acquire human receptor specificity, while at the same time remaining replication competent, thus suggesting that the pandemic threat posed by HPAI H5N1 is far from diminished. PMID:20392847

  20. Protection against H5N1 Highly Pathogenic Avian and Pandemic (H1N1) 2009 Influenza Virus Infection in Cynomolgus Monkeys by an Inactivated H5N1 Whole Particle Vaccine

    Science.gov (United States)

    Nakayama, Misako; Shichinohe, Shintaro; Itoh, Yasushi; Ishigaki, Hirohito; Kitano, Mitsutaka; Arikata, Masahiko; Pham, Van Loi; Ishida, Hideaki; Kitagawa, Naoko; Okamatsu, Masatoshi; Sakoda, Yoshihiro; Ichikawa, Takaya; Tsuchiya, Hideaki; Nakamura, Shinichiro; Le, Quynh Mai; Ito, Mutsumi; Kawaoka, Yoshihiro; Kida, Hiroshi; Ogasawara, Kazumasa

    2013-01-01

    H5N1 highly pathogenic avian influenza virus (HPAIV) infection has been reported in poultry and humans with expanding clade designations. Therefore, a vaccine that induces immunity against a broad spectrum of H5N1 viruses is preferable for pandemic preparedness. We established a second H5N1 vaccine candidate, A/duck/Hokkaido/Vac-3/2007 (Vac-3), in our virus library and examined the efficacy of inactivated whole particles of this strain against two clades of H5N1 HPAIV strains that caused severe morbidity in cynomolgus macaques. Virus propagation in vaccinated macaques infected with either of the H5N1 HPAIV strains was prevented compared with that in unvaccinated macaques. This vaccine also prevented propagation of a pandemic (H1N1) 2009 virus in macaques. In the vaccinated macaques, neutralization activity, which was mainly shown by anti-hemagglutinin antibody, against H5N1 HPAIVs in plasma was detected, but that against H1N1 virus was not detected. However, neuraminidase inhibition activity in plasma and T-lymphocyte responses in lymph nodes against H1N1 virus were detected. Therefore, cross-clade and heterosubtypic protective immunity in macaques consisted of humoral and cellular immunity induced by vaccination with Vac-3. PMID:24376571

  1. Protection against H5N1 highly pathogenic avian and pandemic (H1N1 2009 influenza virus infection in cynomolgus monkeys by an inactivated H5N1 whole particle vaccine.

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    Misako Nakayama

    Full Text Available H5N1 highly pathogenic avian influenza virus (HPAIV infection has been reported in poultry and humans with expanding clade designations. Therefore, a vaccine that induces immunity against a broad spectrum of H5N1 viruses is preferable for pandemic preparedness. We established a second H5N1 vaccine candidate, A/duck/Hokkaido/Vac-3/2007 (Vac-3, in our virus library and examined the efficacy of inactivated whole particles of this strain against two clades of H5N1 HPAIV strains that caused severe morbidity in cynomolgus macaques. Virus propagation in vaccinated macaques infected with either of the H5N1 HPAIV strains was prevented compared with that in unvaccinated macaques. This vaccine also prevented propagation of a pandemic (H1N1 2009 virus in macaques. In the vaccinated macaques, neutralization activity, which was mainly shown by anti-hemagglutinin antibody, against H5N1 HPAIVs in plasma was detected, but that against H1N1 virus was not detected. However, neuraminidase inhibition activity in plasma and T-lymphocyte responses in lymph nodes against H1N1 virus were detected. Therefore, cross-clade and heterosubtypic protective immunity in macaques consisted of humoral and cellular immunity induced by vaccination with Vac-3.

  2. Comprehensive global amino acid sequence analysis of PB1F2 protein of influenza A H5N1 viruses and the influenza A virus subtypes responsible for the 20th-century pandemics.

    Science.gov (United States)

    Pasricha, Gunisha; Mishra, Akhilesh C; Chakrabarti, Alok K

    2013-07-01

    PB1F2 is the 11th protein of influenza A virus translated from +1 alternate reading frame of PB1 gene. Since the discovery, varying sizes and functions of the PB1F2 protein of influenza A viruses have been reported. Selection of PB1 gene segment in the pandemics, variable size and pleiotropic effect of PB1F2 intrigued us to analyze amino acid sequences of this protein in various influenza A viruses. Amino acid sequences for PB1F2 protein of influenza A H5N1, H1N1, H2N2, and H3N2 subtypes were obtained from Influenza Research Database. Multiple sequence alignments of the PB1F2 protein sequences of the aforementioned subtypes were used to determine the size, variable and conserved domains and to perform mutational analysis. Analysis showed that 96·4% of the H5N1 influenza viruses harbored full-length PB1F2 protein. Except for the 2009 pandemic H1N1 virus, all the subtypes of the 20th-century pandemic influenza viruses contained full-length PB1F2 protein. Through the years, PB1F2 protein of the H1N1 and H3N2 viruses has undergone much variation. PB1F2 protein sequences of H5N1 viruses showed both human- and avian host-specific conserved domains. Global database of PB1F2 protein revealed that N66S mutation was present only in 3·8% of the H5N1 strains. We found a novel mutation, N84S in the PB1F2 protein of 9·35% of the highly pathogenic avian influenza H5N1 influenza viruses. Varying sizes and mutations of the PB1F2 protein in different influenza A virus subtypes with pandemic potential were obtained. There was genetic divergence of the protein in various hosts which highlighted the host-specific evolution of the virus. However, studies are required to correlate this sequence variability with the virulence and pathogenicity. © 2012 John Wiley & Sons Ltd.

  3. MVA-based H5N1 vaccine affords cross-clade protection in mice against influenza A/H5N1 viruses at low doses and after single immunization.

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    Joost H C M Kreijtz

    Full Text Available Human infections with highly pathogenic avian influenza viruses of the H5N1 subtype, frequently reported since 2003, result in high morbidity and mortality. It is feared that these viruses become pandemic, therefore the development of safe and effective vaccines is desirable. MVA-based H5N1 vaccines already proved to be effective when two immunizations with high doses were used. Dose-sparing strategies would increase the number of people that can be vaccinated when the amount of vaccine preparations that can be produced is limited. Furthermore, protective immunity is induced ideally after a single immunization. Therefore the minimal requirements for induction of protective immunity with a MVA-based H5N1 vaccine were assessed in mice. To this end, mice were vaccinated once or twice with descending doses of a recombinant MVA expressing the HA gene of influenza virus A/Vietnam/1194/04. The protective efficacy was determined after challenge infection with the homologous clade 1 virus and a heterologous virus derived from clade 2.1, A/Indonesia/5/05 by assessing weight loss, virus replication and histopathological changes. It was concluded that MVA-based vaccines allowed significant dose-sparing and afford cross-clade protection, also after a single immunization, which are favorable properties for an H5N1 vaccine candidate.

  4. The effect of age on the pathogenesis of a highly pathogenic avian influenza (HPAI) H5N1 virus in Pekin ducks (Anas platyrhynchos) infected experimentally

    OpenAIRE

    L?ndt, Brandon Z.; N??ez, Alejandro.; Banks, Jill; Alexander, Dennis J.; Russell, Christine; Richard? L?ndt, Angela C.; Brown, Ian H.

    2009-01-01

    Background? Highly pathogenic avian influenza (HPAI) H5N1 viruses have recently displayed increased virulence for wild waterfowl. Objectives? To study the effect of host age on the shedding and tissue dissemination of a HPAI H5N1 virus in infected Pekin ducks. Methods? Pekin ducks in two age?matched groups (n?=?18), 8 and 12?weeks old (wo) were each infected with 106 EID50/0?1?ml of HPAI A/turkey/Turkey/1/05 (H5N1, clade 2?2). Each day for 5?days, birds were monitored clinically, and cloacal ...

  5. Detection of distribution of avian influenza H5N1 virus by immunohistochemistry, chromogenic in situ hybridization and real-time PCR techniques in experimentally infected chickens.

    Science.gov (United States)

    Chamnanpood, Chanpen; Sanguansermsri, Donruedee; Pongcharoen, Sutatip; Sanguansermsri, Phanchana

    2011-03-01

    Ten specific pathogen free (SPF) chickens were inoculated intranasally with avian influenza virus subtype H5N1. Evaluation revealed distribution of the virus in twelve organs: liver, intestine, bursa, lung, trachea, thymus, heart, pancreas, brain, spleen, kidney, and esophagus. Immunohistochemistry (IHC), chromogenic in situ hybridization (CISH), and real-time polymerase chain reaction (PCR) were developed and compared for detection of the virus from the organs. The distribution of avian influenza H5N1 in chickens varied by animal and detecting technique. The heart, kidneys, intestines, lungs, and pancreas were positive with all three techniques, while the others varied by techique. The three techniques can be used to detect avian influenza effectively, but the pros and cons of each technique need to be determined. The decision of which technique to use depends on the objective of the examination, budget, type and quality of samples, laboratory facilities and technician skills.

  6. Comparative Pathogenesis of an Avian H5N2 and a Swine H1N1 Influenza Virus in Pigs

    DEFF Research Database (Denmark)

    De Vleeschauwer, Annebel; Atanasova, Kalina; Van Borm, Steven

    2009-01-01

    Pigs are considered intermediate hosts for the transmission of avian influenza viruses (AIVs) to humans but the basic organ pathogenesis of AIVs in pigs has been barely studied. We have used 42 four-week-old influenza naive pigs and two different inoculation routes (intranasal and intratracheal) ...

  7. Pathogenicity of Highly Pathogenic Avian Influenza Virus H5N1 in Naturally Infected Poultry in Egypt.

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    Ibrahim Thabet Hagag

    Full Text Available Highly pathogenic avian influenza virus (HPAIV H5N1 has been endemic in Egypt since 2006, and there is increasing concern for its potential to become highly transmissible among humans. Infection by HPAIV H5N1 has been described in experimentally challenged birds. However, the pathogenicity of the H5N1 isolated in Egypt has never been reported in naturally infected chickens and ducks. Here we report a 2013 outbreak of HPAIV H5N1 in commercial poultry farms and backyards in Sharkia Province, Egypt. The main symptoms were ecchymosis on the shanks and feet, cyanosis of the comb and wattles, subcutaneous edema of the head and neck for chickens, and nervous signs (torticollis for ducks. Within 48-72 hrs of the onset of illness, the average mortality rates were 22.8-30% and 28.5-40% in vaccinated chickens and non-vaccinated ducks, respectively. Tissue samples of chickens and ducks were collected for analyses with cross-section immunohistochemistry and real-time RT-PCR for specific viral RNA transcripts. While viral RNA was detected in nearly all tissues and sera collected, viral nucleoprotein was detected almost ubiquitously in all tissues, including testis. Interestingly, viral antigen was also observed in endothelial cells of most organs in chickens, and clearly detected in the trachea and brain in particular. Viral nucleoprotein was also detected in mononuclear cells of various organs, especially pulmonary tissue. We performed phylogenetic analyses and compared the genomic sequences of the hemagglutinin (HA and nonstructural proteins (NS among the isolated viruses, the HPAIV circulated in Egypt in the past and currently, and some available vaccine strains. Further analysis of deduced amino acids of both HA and NS1 revealed that our isolates carried molecular determinants of HPAIV, including the multibasic amino acids (PQGERRRK/KR*GLF in the cleavage site in HA and glutamate at position 92 (D92E in NS1. This is the first report of the pathogenicity

  8. Prevalence and distribution of avian influenza a(H5N1) virus clade variants in live bird markets of Vietnam, 2011-2013.

    Science.gov (United States)

    Nguyen, Diep T; Bryant, Juliet E; Davis, C Todd; Nguyen, Long V; Pham, Long T; Loth, Leo; Inui, Ken; Nguyen, Tung; Jang, Yunho; To, Thanh L; Nguyen, Tho D; Hoang, Diep T; Do, Hoa T; Nguyen, Trang T; Newman, Scott; Jennifer Siembieda; Pham, Dong V

    2014-12-01

    Active surveillance for avian influenza (Al) viruses in poultry sold at live bird markets (LBMs) was conducted in 44 of 63 provinces throughout Vietnam over two periods from September 2011 to February 2012 and October 2012 to June 2013. The study objectives were to assess the prevalence of avian influenza type A, H5, and H5N1 subtype viruses and characterize the geographical and temporal distribution of H5N1 virus genetic variants across the country. Monthly sampling was conducted in 394 LBMs located in 372 communes. A total of 9790 oropharyngeal swabs from poultry were screened for influenza A virus by real-time reverse-transcriptase PCR Virus isolation was attempted on all positive samples in embryonated chicken eggs, and the HA1 region of each H5 virus isolate was sequenced. Market prevalence of H5 subtype virus was 32.2% (127/394) over the cumulative 15 mo of surveillance. Phylogenetic analyses indicated that clade 1.1 viruses persisted in the south, whereas three genetically distinct subgroups of dade 2.3.2.1 were found simultaneously in northern, central, and southern Vietnam. Clade 2.3.2.1c viruses first appeared in July 2012 and spread rapidly to the center and south of Vietnam in late 2012, where they were predominant among clade 2.3.2.1 viruses and were detected in both active LBM surveillance and poultry outbreaks. Given the overlapping geographic distribution of dade variants and the antigenic divergence previously described for these dades, current AI poultry vaccines used in Vietnam may require bivalent formulations containing representatives of both dade 1.1 and dade 2.3.2.1 viruses.

  9. Protective Efficacy of Newcastle Disease Virus Expressing Soluble Trimeric Hemagglutinin against Highly Pathogenic H5N1 Influenza in Chickens and Mice

    NARCIS (Netherlands)

    Cornelissen, A.H.M.; Leeuw, de O.S.; Tacken, M.G.J.; Klos, H.C.; Vries, de R.P.; Boer-Luijtze, de E.A.; Zoelen-Bos, van D.J.; Rigter, A.; Rottier, P.J.M.; Moormann, R.J.M.; Haan, de C.A.M.

    2012-01-01

    Background: Highly pathogenic avian influenza virus (HPAIV) causes a highly contagious often fatal disease in poultry, resulting in significant economic losses in the poultry industry. HPAIV H5N1 also poses a major public health threat as it can be transmitted directly from infected poultry to

  10. Reassortant clade 2.3.4.4 Avian Influenza A(H5N6) Virus in a wild Mandarin Duck, South Korea, 2016

    Science.gov (United States)

    Highly pathogenic avian influenza viruses (HPAIV) have caused significant economic losses in the poultry industries and represents a serious threat to public health. H5N1 HPAIV was first detected in 1996 from a domestic goose in Guangdong China (Gs/GD) and has subsequently evolved into 10 geneticall...

  11. Efficacy of a Recombinant Turkey Herpesvirus H5 Vaccine Against Challenge With H5N1 Clades 1.1.2 and 2.3.2.1 Highly Pathogenic Avian Influenza Viruses in Domestic Ducks (Anas platyrhynchos domesticus).

    Science.gov (United States)

    Pantin-Jackwood, Mary J; Kapczynski, Darrell R; DeJesus, Eric; Costa-Hurtado, Mar; Dauphin, Gwenaelle; Tripodi, Astrid; Dunn, John R; Swayne, David E

    2016-03-01

    Domestic ducks are the second most abundant poultry species in many Asian countries and have played a critical role in the epizootiology of H5N1 highly pathogenic avian influenza (HPAI).In this study, the protective efficacy of a live recombinant vector vaccine based on a turkey herpesvirus (HVT) expressing the H5 gene from a clade 2.2 H5N1 HPAI strain (A/Swan/Hungary/4999/ 2006) (rHVT-H5/2.2), given at 3 days of age, was examined in Pekin ducks (Anas platyrhynchos domesticus). The vaccine was given alone or in combination with an inactivated H5N1 clade 2.3.2.1 reverse genetic (rgGD/2.3.2.1) vaccine given at 16 days of age, either as a single vaccination or in a prime-boost regime. At 30 days of age, ducks were challenged with one of two H5N1 HPAI viruses: A/duck/Vietnam/NCVD-2721/2013 (clade 1.1.2) or A/duck/Vietnam/NCVD-1584/2012 (clade 2.3.2.1.C). These viruses produced 100% mortality in less than 5 days in nonvaccinated control ducks. Ducks vaccinated with the rgGD/2.3.2.1 vaccine, with or without the rHVT-H5/2.2 vaccine, were 90%-100% protected against mortality after challenge with either of the two H5N1 HPAI viruses. The rHVT-H5/2.2 vaccine alone, however, conferred only 30% protection against mortality after challenge with either H5N1 HPAI virus; the surviving ducks from these groups shed higher amount of virus and for longer than the single-vaccinated rgGD/2.3.2.1 group. Despite low protection, ducks vaccinated with the rHVT-H5/2.2 vaccine and challenged with the clade 1.1.2 Vietnam virus had a longer mean death time than nonvaccinated controls (P = 0.02). A booster effect was found on reduction of virus shedding when using both vaccines, with lower oropharyngeal viral titers at 4 days after challenge with either HPAI virus (P study demonstrates the suboptimal protection with the rHVT-H5/2.2 vaccine given alone in Pekin ducks against H5N1 HPAI viruses and only a minor additive effect on virus shedding reduction when used with an inactivated vaccine in a

  12. H5N1 Influenza A Virus PB1-F2 Relieves HAX-1-Mediated Restriction of Avian Virus Polymerase PA in Human Lung Cells.

    Science.gov (United States)

    Mazel-Sanchez, B; Boal-Carvalho, I; Silva, F; Dijkman, R; Schmolke, M

    2018-06-01

    Highly pathogenic influenza A viruses (IAV) from avian hosts were first reported to directly infect humans 20 years ago. However, such infections are rare events, and our understanding of factors promoting or restricting zoonotic transmission is still limited. One accessory protein of IAV, PB1-F2, was associated with pathogenicity of pandemic and zoonotic IAV. This short (90-amino-acid) peptide does not harbor an enzymatic function. We thus identified host factors interacting with H5N1 PB1-F2, which could explain its importance for virulence. PB1-F2 binds to HCLS1-associated protein X1 (HAX-1), a recently identified host restriction factor of the PA subunit of IAV polymerase complexes. We demonstrate that the PA of a mammal-adapted H1N1 IAV is resistant to HAX-1 imposed restriction, while the PA of an avian-origin H5N1 IAV remains sensitive. We also showed HAX-1 sensitivity for PAs of A/Brevig Mission/1/1918 (H1N1) and A/Shanghai/1/2013 (H7N9), two avian-origin zoonotic IAV. Inhibition of H5N1 polymerase by HAX-1 can be alleviated by its PB1-F2 through direct competition. Accordingly, replication of PB1-F2-deficient H5N1 IAV is attenuated in the presence of large amounts of HAX-1. Mammal-adapted H1N1 and H3N2 viruses do not display this dependence on PB1-F2 for efficient replication in the presence of HAX-1. We propose that PB1-F2 plays a key role in zoonotic transmission of avian H5N1 IAV into humans. IMPORTANCE Aquatic and shore birds are the natural reservoir of influenza A viruses from which the virus can jump into a variety of bird and mammal host species, including humans. H5N1 influenza viruses are a good model for this process. They pose an ongoing threat to human and animal health due to their high mortality rates. However, it is currently unclear what restricts these interspecies jumps on the host side or what promotes them on the virus side. Here we show that a short viral peptide, PB1-F2, helps H5N1 bird influenza viruses to overcome a human restriction

  13. Single Dose of Consensus Hemagglutinin-Based Virus-Like Particles Vaccine Protects Chickens against Divergent H5 Subtype Influenza Viruses

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    Peipei Wu

    2017-11-01

    Full Text Available The H5 subtype highly pathogenic avian influenza (HPAI virus is one of the greatest threats to global poultry industry. To develop broadly protective H5 subunit vaccine, a recombinant consensus HA sequence (rHA was constructed and expressed in virus-like particles (rHA VLPs in the baculovirus-insect cell system. The efficacy of the rHA VLPs vaccine with or without immunopotentiator (CVCVA5 was assessed in chickens. Compared to the commercial Re6 or Re6-CVCVA5 vaccines, single dose immunization of chickens with rHA VLPs or rHA-CVCVA5 vaccines induced higher levels of serum hemagglutinin inhibition titers and neutralization titers, mucosal antibodies, IFN-γ and IL-4 cytokines in sera, and cytotoxic T lymphocyte responses. The rHA VLPs vaccine was superior to the commercial Re6 vaccine in conferring cross-protection against different clades of H5 subtype viruses. This study reports that H5 subtype consensus HA VLP single dose vaccination provides broad protection against HPAI virus in chickens.

  14. Effect of neuraminidase inhibitor-resistant mutations on pathogenicity of clade 2.2 A/Turkey/15/06 (H5N1 influenza virus in ferrets.

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    Natalia A Ilyushina

    2010-05-01

    Full Text Available The acquisition of neuraminidase (NA inhibitor resistance by H5N1 influenza viruses has serious clinical implications, as this class of drugs can be an essential component of pandemic control measures. The continuous evolution of the highly pathogenic H5N1 influenza viruses results in the emergence of natural NA gene variations whose impact on viral fitness and NA inhibitor susceptibility are poorly defined. We generated seven genetically stable recombinant clade 2.2 A/Turkey/15/06-like (H5N1 influenza viruses carrying NA mutations located either in the framework residues (E119A, H274Y, N294S or in close proximity to the NA enzyme active site (V116A, I117V, K150N, Y252H. NA enzyme inhibition assays showed that NA mutations at positions 116, 117, 274, and 294 reduced susceptibility to oseltamivir carboxylate (IC(50s increased 5- to 940-fold. Importantly, the E119A NA mutation (previously reported to confer resistance in the N2 NA subtype was stable in the clade 2.2 H5N1 virus background and induced cross-resistance to oseltamivir carboxylate and zanamivir. We demonstrated that Y252H NA mutation contributed for decreased susceptibility of clade 2.2 H5N1 viruses to oseltamivir carboxylate as compared to clade 1 viruses. The enzyme kinetic parameters (V(max, K(m and K(i of the avian-like N1 NA glycoproteins were highly consistent with their IC(50 values. None of the recombinant H5N1 viruses had attenuated virulence in ferrets inoculated with 10(6 EID(50 dose. Most infected ferrets showed mild clinical disease signs that differed in duration. However, H5N1 viruses carrying the E119A or the N294S NA mutation were lethal to 1 of 3 inoculated animals and were associated with significantly higher virus titers (P<0.01 and inflammation in the lungs compared to the wild-type virus. Our results suggest that highly pathogenic H5N1 variants carrying mutations within the NA active site that decrease susceptibility to NA inhibitors may possess increased

  15. D701N mutation in the PB2 protein contributes to the pathogenicity of H5N1 avian influenza viruses but not transmissibility in guinea pigs

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    Peirong eJiao

    2014-11-01

    Full Text Available H5N1 highly pathogenic avian influenza virus (HPAIV of clade 2.3.2 has been circulating in waterfowl in Southern China since 2003. Our previous studies showed that certain H5N1 HPAIV isolates within clade 2.3.2 from Southern China had high pathogenicity in different birds. Guinea pigs have been successfully used as models to evaluate the transmissibility of AIVs and other species of influenza viruses in mammalian hosts. However, few studies have reported pathogenicity and transmissibility of H5N1 HPAIVs of this clade in guinea pigs. In this study, we selected an H5N1 HPAIV isolate, A/duck/Guangdong/357/2008, to investigate the pathogenicity and transmissibility of the virus in guinea pigs. The virus had high pathogenicity in mice; additionally, it only replicated in some tissues of the guinea pigs without production of clinical signs, but was transmissible among guinea pigs. Interestingly, virus isolates from co-caged guinea pigs had the D701N mutation in the PB2 protein. These mutant viruses showed higher pathogenicity in mice and higher replication capability in guinea pigs but did not demonstrate enhanced the transmissibility among guinea pigs. These findings indicate the transmission of the H5N1 virus between mammals could induce virus mutations, and the mutant viruses might have higher pathogenicity in mammals without higher transmissibility. Therefore, the continued evaluation of the pathogenicity and transmissibility of avian influenza virus (AIVs in mammals is critical to the understanding of the evolutionary characteristics of AIVs and the emergence of potential pandemic strains.

  16. An M2e-based multiple antigenic peptide vaccine protects mice from lethal challenge with divergent H5N1 influenza viruses

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    Chan Chris CS

    2010-01-01

    Full Text Available Abstract Background A growing concern has raised regarding the pandemic potential of the highly pathogenic avian influenza (HPAI H5N1 viruses. Consequently, there is an urgent need to develop an effective and safe vaccine against the divergent H5N1 influenza viruses. In the present study, we designed a tetra-branched multiple antigenic peptide (MAP-based vaccine, designated M2e-MAP, which contains the sequence overlapping the highly conserved extracellular domain of matrix protein 2 (M2e of a HPAI H5N1 virus, and investigated its immune responses and cross-protection against different clades of H5N1 viruses. Results Our results showed that M2e-MAP vaccine induced strong M2e-specific IgG antibody responses following 3-dose immunization of mice with M2e-MAP in the presence of Freunds' or aluminium (alum adjuvant. M2e-MAP vaccination limited viral replication and attenuated histopathological damage in the challenged mouse lungs. The M2e-MAP-based vaccine protected immunized mice against both clade1: VN/1194 and clade2.3.4: SZ/406H H5N1 virus challenge, being able to counteract weight lost and elevate survival rate following lethal challenge of H5N1 viruses. Conclusions These results suggest that M2e-MAP presenting M2e of H5N1 virus has a great potential to be developed into an effective subunit vaccine for the prevention of infection by a broad spectrum of HPAI H5N1 viruses.

  17. Migratory birds, the H5N1 influenza virus and the scientific method

    Directory of Open Access Journals (Sweden)

    Stilianakis Nikolaos I

    2008-05-01

    Full Text Available Abstract Background The role of migratory birds and of poultry trade in the dispersal of highly pathogenic H5N1 is still the topic of intense and controversial debate. In a recent contribution to this journal, Flint argues that the strict application of the scientific method can help to resolve this issue. Discussion We argue that Flint's identification of the scientific method with null hypothesis testing is misleading and counterproductive. There is far more to science than the testing of hypotheses; not only the justification, bur also the discovery of hypotheses belong to science. We also show why null hypothesis testing is weak and that Bayesian methods are a preferable approach to statistical inference. Furthermore, we criticize the analogy put forward by Flint between involuntary transport of poultry and long-distance migration. Summary To expect ultimate answers and unequivocal policy guidance from null hypothesis testing puts unrealistic expectations on a flawed approach to statistical inference and on science in general.

  18. Antigenic, genetic, and pathogenic characterization of H5N1 highly pathogenic avian influenza viruses isolated from dead whooper swans (Cygnus cygnus) found in northern Japan in 2008.

    Science.gov (United States)

    Okamatsu, Masatoshi; Tanaka, Tomohisa; Yamamoto, Naoki; Sakoda, Yoshihiro; Sasaki, Takashi; Tsuda, Yoshimi; Isoda, Norikazu; Kokumai, Norihide; Takada, Ayato; Umemura, Takashi; Kida, Hiroshi

    2010-12-01

    In April and May 2008, whooper swans (Cygnus cygnus) were found dead in Hokkaido in Japan. In this study, an adult whooper swan found dead beside Lake Saroma was pathologically examined and the identified H5N1 influenza virus isolates were genetically and antigenically analyzed. Pathological findings indicate that the swan died of severe congestive edema in the lungs. Phylogenetic analysis of the HA genes of the isolates revealed that they are the progeny viruses of isolates from poultry and wild birds in China, Russia, Korea, and Hong Kong. Antigenic analyses indicated that the viruses are distinguished from the H5N1 viruses isolated from wild birds and poultry before 2007. The chickens vaccinated with A/duck/Hokkaido/Vac-1/2004 (H5N1) survived for 14 days after challenge with A/whooper swan/Hokkaido/1/2008 (H5N1), although a small amount of the challenge virus was recovered from the tissues of the birds. These findings indicate that H5N1 highly pathogenic avian influenza viruses are circulating in wild birds in addition to domestic poultry in Asia and exhibit antigenic variation that may be due to vaccination.

  19. Neuraminidase stalk length and additional glycosylation of the hemagglutinin influence the virulence of influenza H5N1 viruses for mice.

    Science.gov (United States)

    Matsuoka, Yumiko; Swayne, David E; Thomas, Colleen; Rameix-Welti, Marie-Anne; Naffakh, Nadia; Warnes, Christine; Altholtz, Melanie; Donis, Ruben; Subbarao, Kanta

    2009-05-01

    Following circulation of avian influenza H5 and H7 viruses in poultry, the hemagglutinin (HA) can acquire additional glycosylation sites, and the neuraminidase (NA) stalk becomes shorter. We investigated whether these features play a role in the pathogenesis of infection in mammalian hosts. From 1996 to 2007, H5N1 viruses with a short NA stalk have become widespread in several avian species. Compared to viruses with a long-stalk NA, viruses with a short-stalk NA showed a decreased capacity to elute from red blood cells and an increased virulence in mice, but not in chickens. The presence of additional HA glycosylation sites had less of an effect on virulence than did NA stalk length. The short-stalk NA of H5N1 viruses circulating in Asia may contribute to virulence in humans.

  20. Neuraminidase Stalk Length and Additional Glycosylation of the Hemagglutinin Influence the Virulence of Influenza H5N1 Viruses for Mice▿

    Science.gov (United States)

    Matsuoka, Yumiko; Swayne, David E.; Thomas, Colleen; Rameix-Welti, Marie-Anne; Naffakh, Nadia; Warnes, Christine; Altholtz, Melanie; Donis, Ruben; Subbarao, Kanta

    2009-01-01

    Following circulation of avian influenza H5 and H7 viruses in poultry, the hemagglutinin (HA) can acquire additional glycosylation sites, and the neuraminidase (NA) stalk becomes shorter. We investigated whether these features play a role in the pathogenesis of infection in mammalian hosts. From 1996 to 2007, H5N1 viruses with a short NA stalk have become widespread in several avian species. Compared to viruses with a long-stalk NA, viruses with a short-stalk NA showed a decreased capacity to elute from red blood cells and an increased virulence in mice, but not in chickens. The presence of additional HA glycosylation sites had less of an effect on virulence than did NA stalk length. The short-stalk NA of H5N1 viruses circulating in Asia may contribute to virulence in humans. PMID:19225004

  1. Role of domestic ducks in the emergence of a new genotype of highly pathogenic H5N1 avian influenza A viruses in Bangladesh.

    Science.gov (United States)

    Barman, Subrata; Marinova-Petkova, Atanaska; Hasan, M Kamrul; Akhtar, Sharmin; El-Shesheny, Rabeh; Turner, Jasmine Cm; Franks, John; Walker, David; Seiler, Jon; Friedman, Kimberly; Kercher, Lisa; Jeevan, Trushar; Darnell, Daniel; Kayali, Ghazi; Jones-Engel, Lisa; McKenzie, Pamela; Krauss, Scott; Webby, Richard J; Webster, Robert G; Feeroz, Mohammed M

    2017-08-09

    Highly pathogenic avian influenza H5N1 viruses were first isolated in Bangladesh in February 2007. Subsequently, clades 2.2.2, 2.3.4.2 and 2.3.2.1a were identified in Bangladesh, and our previous surveillance data revealed that by the end of 2014, the circulating viruses exclusively comprised clade 2.3.2.1a. We recently determined the status of circulating avian influenza viruses in Bangladesh by conducting surveillance of live poultry markets and waterfowl in wetland areas from February 2015 through February 2016. Until April 2015, clade 2.3.2.1a persisted without any change in genotype. However, in June 2015, we identified a new genotype of H5N1 viruses, clade 2.3.2.1a, which quickly became predominant. These newly emerged H5N1 viruses contained the hemagglutinin, neuraminidase and matrix genes of circulating 2.3.2.1a Bangladeshi H5N1 viruses and five other genes of low pathogenic Eurasian-lineage avian influenza A viruses. Some of these internal genes were closely related to those of low pathogenic viruses isolated from ducks in free-range farms and wild birds in a wetland region of northeastern Bangladesh, where commercially raised domestic ducks have frequent contact with migratory birds. These findings indicate that migratory birds of the Central Asian flyway and domestic ducks in the free-range farms in Tanguar haor-like wetlands played an important role in the emergence of this novel genotype of highly pathogenic H5N1 viruses.

  2. Experimental challenge and pathology of highly pathogenic avian influenza virus H5N1 in dunlin (Calidris alpina), an intercontinental migrant shorebird species

    Science.gov (United States)

    Hall, Jeffrey S.; Franson, J. Christian; Gill, Robert E.; Meteyer, Carol U.; TeSlaa, Joshua L.; Nashold, Sean W.; Dusek, Robert J.; Ip, Hon S.

    2011-01-01

    Background Shorebirds (Charadriiformes) are considered one of the primary reservoirs of avian influenza. Because these species are highly migratory, there is concern that infected shorebirds may be a mechanism by which highly pathogenic avian influenza virus (HPAIV) H5N1 could be introduced into North America from Asia. Large numbers of dunlin (Calidris alpina) migrate from wintering areas in central and eastern Asia, where HPAIV H5N1 is endemic, across the Bering Sea to breeding areas in Alaska. Low pathogenic avian influenza virus has been previously detected in dunlin, and thus, dunlin represent a potential risk to transport HPAIV to North America. To date no experimental challenge studies have been performed in shorebirds.

  3. Isolation of an H5N8 Highly Pathogenic Avian Influenza Virus Strain from Wild Birds in Seoul, a Highly Urbanized Area in South Korea.

    Science.gov (United States)

    Kwon, Jung-Hoon; Lee, Dong-Hun; Jeong, Jei-Hyun; Yuk, Seong-Su; Erdene-Ochir, Tseren-Ochir; Noh, Jin-Yong; Hong, Woo-Tack; Jeong, Sol; Gwon, Gyeong-Bin; Lee, Sang-Won; Choi, In-Soo; Song, Chang-Seon

    2017-07-01

    Asian-lineage H5 highly pathogenic avian influenza viruses (HPAIV) have caused recurrent outbreaks in poultry and wild birds. In January 2014, H5N8 HPAIV caused outbreaks in South Korea and subsequently spread to East Asia, Europe, and North America. We report the isolation of an H5N8 HPAIV strain from wild birds in Seoul, the most-developed city in South Korea. We analyzed the complete genome sequence of this isolate and estimated its origin using a phylogenetic analysis. The Seoul H5N8 isolate clustered phylogenetically with strains isolated from migratory wild birds but was distinct from Korean poultry isolates. This H5N8 virus was likely introduced into the urbanized city by migratory wild birds. Therefore, wild bird habitats in urbanized areas should be carefully monitored for HPAIV.

  4. Prophylactic and therapeutic efficacy of avian antibodies against influenza virus H5N1 and H1N1 in mice.

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    Huan H Nguyen

    Full Text Available BACKGROUND: Pandemic influenza poses a serious threat to global health and the world economy. While vaccines are currently under development, passive immunization could offer an alternative strategy to prevent and treat influenza virus infection. Attempts to develop monoclonal antibodies (mAbs have been made. However, passive immunization based on mAbs may require a cocktail of mAbs with broader specificity in order to provide full protection since mAbs are generally specific for single epitopes. Chicken immunoglobulins (IgY found in egg yolk have been used mainly for treatment of infectious diseases of the gastrointestinal tract. Because the recent epidemic of highly pathogenic avian influenza virus (HPAIV strain H5N1 has resulted in serious economic losses to the poultry industry, many countries including Vietnam have introduced mass vaccination of poultry with H5N1 virus vaccines. We reasoned that IgY from consumable eggs available in supermarkets in Vietnam could provide protection against infections with HPAIV H5N1. METHODS AND FINDINGS: We found that H5N1-specific IgY that are prepared from eggs available in supermarkets in Vietnam by a rapid and simple water dilution method cross-protect against infections with HPAIV H5N1 and related H5N2 strains in mice. When administered intranasally before or after lethal infection, the IgY prevent the infection or significantly reduce viral replication resulting in complete recovery from the disease, respectively. We further generated H1N1 virus-specific IgY by immunization of hens with inactivated H1N1 A/PR/8/34 as a model virus for the current pandemic H1N1/09 and found that such H1N1-specific IgY protect mice from lethal influenza virus infection. CONCLUSIONS: The findings suggest that readily available H5N1-specific IgY offer an enormous source of valuable biological material to combat a potential H5N1 pandemic. In addition, our study provides a proof-of-concept for the approach using virus

  5. Evaluation of the zoonotic potential of multiple subgroups of clade 2.3.4.4 influenza A (H5N8) virus.

    Science.gov (United States)

    Lee, Yu-Na; Lee, Eun-Kyoung; Song, Byung-Min; Heo, Gyeong-Beom; Woo, Sang-Hee; Cheon, Sun-Ha; Lee, Youn-Jeong

    2018-03-01

    Clade 2.3.4.4 H5N8 highly pathogenic avian influenza viruses (HPAIVs) have spread worldwide. Phylogenetic analysis identified two genetic groups of the H5N8 HPAIVs in South Korea; group A evolved further into four subgroups. Here, we examined the zoonotic potential, both in vivo and in vitro, of genetically distinct subgroups of H5N8 HPAIVs isolated in South Korea. When compared with other subgroups, A/mallard/Korea/H2102/2015 (H2102) virus caused relatively severe disease in mice at high doses. In ferrets, all H5N8 viruses replicated restrictively in the respiratory tract and did not induce significant clinical signs of influenza infection. In vitro studies, all viruses displayed a hemagglutinin phenotype that was poorly adapted for infection of mammals, although the H2102 virus exhibited higher replication kinetics at 33°C than the others. Although H5N8 HPAIVs have not yet acquired all the characteristics required for adaptation to mammals, their ability to evolve continuously underscores the need for timely risk assessment. Copyright © 2018 Elsevier Inc. All rights reserved.

  6. Protective efficacy of Newcastle disease virus expressing soluble trimeric hemagglutinin against highly pathogenic H5N1 influenza in chickens and mice.

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    Lisette A H M Cornelissen

    Full Text Available BACKGROUND: Highly pathogenic avian influenza virus (HPAIV causes a highly contagious often fatal disease in poultry, resulting in significant economic losses in the poultry industry. HPAIV H5N1 also poses a major public health threat as it can be transmitted directly from infected poultry to humans. One effective way to combat avian influenza with pandemic potential is through the vaccination of poultry. Several live vaccines based on attenuated Newcastle disease virus (NDV that express influenza hemagglutinin (HA have been developed to protect chickens or mammalian species against HPAIV. However, the zoonotic potential of NDV raises safety concerns regarding the use of live NDV recombinants, as the incorporation of a heterologous attachment protein may result in the generation of NDV with altered tropism and/or pathogenicity. METHODOLOGY/PRINCIPAL FINDINGS: In the present study we generated recombinant NDVs expressing either full length, membrane-anchored HA of the H5 subtype (NDV-H5 or a soluble trimeric form thereof (NDV-sH5(3. A single intramuscular immunization with NDV-sH5(3 or NDV-H5 fully protected chickens against disease after a lethal challenge with H5N1 and reduced levels of virus shedding in tracheal and cloacal swabs. NDV-sH5(3 was less protective than NDV-H5 (50% vs 80% protection when administered via the respiratory tract. The NDV-sH5(3 was ineffective in mice, regardless of whether administered oculonasally or intramuscularly. In this species, NDV-H5 induced protective immunity against HPAIV H5N1, but only after oculonasal administration, despite the poor H5-specific serum antibody response it elicited. CONCLUSIONS/SIGNIFICANCE: Although NDV expressing membrane anchored H5 in general provided better protection than its counterpart expressing soluble H5, chickens could be fully protected against a lethal challenge with H5N1 by using the latter NDV vector. This study thus provides proof of concept for the use of recombinant

  7. Experimental assessment of the pathogenicity of eight avian influenza A viruses of H5 subtype for chickens, turkeys, ducks and quail.

    Science.gov (United States)

    Alexander, D J; Parsons, G; Manvell, R J

    1986-01-01

    Clinical signs, death, virus excretion and immune response were measured in 2-week-old chickens, turkeys, quail and ducks infected by intramuscular, intranasal and contact routes with eight influenza viruses of H5 subtype. Six of the viruses: A/chicken/Scotland/59 (H5N1), ck/Scot; A/tern/South Africa/61 (H5N3), tern/SA; A/turkey/Ontario/ 7732/66 (H5N9); ty/Ont; A/chicken/Pennsylvania/1370/83 (H5N2); Pa/1370; A/turkey/Ireland/83 (H5N8); ty/Ireland, and A/duck/Ireland/ 113/84 (HSN8); dk/Ireland, were highly pathogenic for chickens and turkeys. Two viruses, A/chicken/Pennsylvania/1/83 (H5N2), Pa/1 and A/turkey/Italy/ZA/80 (H5N2), ty/Italy, were of low pathogenicity. Ck/Scot was more pathogenic for chickens than turkeys while ty/Ont was more pathogenic for turkeys than chickens. Other viruses showed little difference in their pathogenicity for these two hosts. No clinical signs or deaths were seen in any of the infected ducks. Only two viruses, dk/Ireland and ty/Ireland, produced consistent serological responses in ducks, although intramuscular infection with tern/SA and ty/Italy resulted in some ducks with positive HI titres. These four were the only viruses reisolated from ducks. Quail showed some resistance to viruses which were highly pathogenic for chickens and turkeys, most notably to ck/Scot and ty/Ont and to a lesser extent tern/SA and Pa/1370. Transmission of virus from intranasally infected birds to birds placed in contact varied considerably with both host and infecting virus and the various combinations of these.

  8. Experimentally infected domestic ducks show efficient transmission of Indonesian H5N1 highly pathogenic avian influenza virus, but lack persistent viral shedding.

    Science.gov (United States)

    Wibawa, Hendra; Bingham, John; Nuradji, Harimurti; Lowther, Sue; Payne, Jean; Harper, Jenni; Junaidi, Akhmad; Middleton, Deborah; Meers, Joanne

    2014-01-01

    Ducks are important maintenance hosts for avian influenza, including H5N1 highly pathogenic avian influenza viruses. A previous study indicated that persistence of H5N1 viruses in ducks after the development of humoral immunity may drive viral evolution following immune selection. As H5N1 HPAI is endemic in Indonesia, this mechanism may be important in understanding H5N1 evolution in that region. To determine the capability of domestic ducks to maintain prolonged shedding of Indonesian clade 2.1 H5N1 virus, two groups of Pekin ducks were inoculated through the eyes, nostrils and oropharynx and viral shedding and transmission investigated. Inoculated ducks (n = 15), which were mostly asymptomatic, shed infectious virus from the oral route from 1 to 8 days post inoculation, and from the cloacal route from 2-8 dpi. Viral ribonucleic acid was detected from 1-15 days post inoculation from the oral route and 1-24 days post inoculation from the cloacal route (cycle threshold ducks seroconverted in a range of serological tests by 15 days post inoculation. Virus was efficiently transmitted during acute infection (5 inoculation-infected to all 5 contact ducks). However, no evidence for transmission, as determined by seroconversion and viral shedding, was found between an inoculation-infected group (n = 10) and contact ducks (n = 9) when the two groups only had contact after 10 days post inoculation. Clinical disease was more frequent and more severe in contact-infected (2 of 5) than inoculation-infected ducks (1 of 15). We conclude that Indonesian clade 2.1 H5N1 highly pathogenic avian influenza virus does not persist in individual ducks after acute infection.

  9. Development of a dual-protective live attenuated vaccine against H5N1 and H9N2 avian influenza viruses by modifying the NS1 gene.

    Science.gov (United States)

    Choi, Eun-hye; Song, Min-Suk; Park, Su-Jin; Pascua, Philippe Noriel Q; Baek, Yun Hee; Kwon, Hyeok-il; Kim, Eun-Ha; Kim, Semi; Jang, Hyung-Kwan; Poo, Haryoung; Kim, Chul-Joong; Choi, Young Ki

    2015-07-01

    An increasing number of outbreaks of avian influenza H5N1 and H9N2 viruses in poultry have caused serious economic losses and raised concerns for human health due to the risk of zoonotic transmission. However, licensed H5N1 and H9N2 vaccines for animals and humans have not been developed. Thus, to develop a dual H5N1 and H9N2 live-attenuated influenza vaccine (LAIV), the HA and NA genes from a virulent mouse-adapted avian H5N2 (A/WB/Korea/ma81/06) virus and a recently isolated chicken H9N2 (A/CK/Korea/116/06) virus, respectively, were introduced into the A/Puerto Rico/8/34 backbone expressing truncated NS1 proteins (NS1-73, NS1-86, NS1-101, NS1-122) but still possessing a full-length NS gene. Two H5N2/NS1-LAIV viruses (H5N2/NS1-86 and H5N2/NS1-101) were highly attenuated compared with the full-length and remaining H5N2/NS-LAIV viruses in a mouse model. Furthermore, viruses containing NS1 modifications were found to induce more IFN-β activation than viruses with full-length NS1 proteins and were correspondingly attenuated in mice. Intranasal vaccination with a single dose (10(4.0) PFU/ml) of these viruses completely protected mice from a lethal challenge with the homologous A/WB/Korea/ma81/06 (H5N2), heterologous highly pathogenic A/EM/Korea/W149/06 (H5N1), and heterosubtypic highly virulent mouse-adapted H9N2 viruses. This study clearly demonstrates that the modified H5N2/NS1-LAIV viruses attenuated through the introduction of mutations in the NS1 coding region display characteristics that are desirable for live attenuated vaccines and hold potential as vaccine candidates for mammalian hosts.

  10. Reassortant H9N2 influenza viruses containing H5N1-like PB1 genes isolated from black-billed magpies in Southern China.

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    Guoying Dong

    Full Text Available H9N2 influenza A viruses have become endemic in different types of terrestrial poultry and wild birds in Asia, and are occasionally transmitted to humans and pigs. To evaluate the role of black-billed magpies (Pica pica in the evolution of influenza A virus, we conducted two epidemic surveys on avian influenza viruses in wild black-billed magpies in Guangxi, China in 2005 and characterized three isolated black-billed magpie H9N2 viruses (BbM viruses. Phylogenetic analysis indicated that three BbM viruses were almost identical with 99.7 to 100% nucleotide homology in their whole genomes, and were reassortants containing BJ94-like (Ck/BJ/1/94 HA, NA, M, and NS genes, SH/F/98-like (Ck/SH/F/98 PB2, PA, and NP genes, and H5N1-like (Ck/YN/1252/03, clade 1 PB1 genes. Genetic analysis showed that BbM viruses were most likely the result of multiple reassortments between co-circulating H9N2-like and H5N1-like viruses, and were genetically different from other H9N2 viruses because of the existence of H5N1-like PB1 genes. Genotypical analysis revealed that BbM viruses evolved from diverse sources and belonged to a novel genotype (B46 discovered in our recent study. Molecular analysis suggested that BbM viruses were likely low pathogenic reassortants. However, results of our pathogenicity study demonstrated that BbM viruses replicated efficiently in chickens and a mammalian mouse model but were not lethal for infected chickens and mice. Antigenic analysis showed that BbM viruses were antigenic heterologous with the H9N2 vaccine strain. Our study is probably the first report to document and characterize H9N2 influenza viruses isolated from black-billed magpies in southern China. Our results suggest that black-billed magpies were susceptible to H9N2 influenza viruses, which raise concerns over possible transmissions of reassortant H9N2 viruses among poultry and wild birds.

  11. Prediction and identification of T cell epitopes in the H5N1 influenza virus nucleoprotein in chicken.

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    Yanxia Hou

    Full Text Available T cell epitopes can be used for the accurate monitoring of avian influenza virus (AIV immune responses and the rational design of vaccines. No T cell epitopes have been previously identified in the H5N1 AIV virus nucleoprotein (NP in chickens. For the first time, this study used homology modelling techniques to construct three-dimensional structures of the peptide-binding domains of chicken MHC class Ι molecules for four commonly encountered unique haplotypes, i.e., B4, B12, B15, and B19. H5N1 AIV NP was computationally parsed into octapeptides or nonapeptides according to the peptide-binding motifs of MHC class I molecules of the B4, B12, B15 and B19 haplotypes. Seventy-five peptide sequences were modelled and their MHC class I molecule-binding abilities were analysed by molecular docking. Twenty-five peptides (Ten for B4, six for B12, two for B15, and seven for B19 were predicted to be potential T cell epitopes in chicken. Nine of these peptides and one unrelated peptide were manually synthesized and their T cell responses were tested in vitro. Spleen lymphocytes were collected from SPF chickens that had been immunised with a NP-expression plasmid, pCAGGS-NP, and they were stimulated using the synthesized peptides. The secretion of chicken IFN-γ and the proliferation of CD8(+ T cells were tested using an ELISA kit and flow cytometry, respectively. The significant secretion of chicken IFN-γ and proliferation of CD8(+ T lymphocytes increased by 13.7% and 11.9% were monitored in cells stimulated with peptides NP(89-97 and NP(198-206, respectively. The results indicate that peptides NP(89-97 (PKKTGGPIY and NP(198-206 (KRGINDRNF are NP T cell epitopes in chicken of certain haplotypes. The method used in this investigation is applicable to predicting T cell epitopes for other antigens in chicken, while this study also extends our understanding of the mechanisms of the immune response to AIV in chicken.

  12. Highly pathogenic avian influenza virus subtype H5N1 in Africa: a comprehensive phylogenetic analysis and molecular characterization of isolates.

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    Giovanni Cattoli

    Full Text Available Highly pathogenic avian influenza virus A/H5N1 was first officially reported in Africa in early 2006. Since the first outbreak in Nigeria, this virus spread rapidly to other African countries. From its emergence to early 2008, 11 African countries experienced A/H5N1 outbreaks in poultry and human cases were also reported in three of these countries. At present, little is known of the epidemiology and molecular evolution of A/H5N1 viruses in Africa. We have generated 494 full gene sequences from 67 African isolates and applied molecular analysis tools to a total of 1,152 A/H5N1 sequences obtained from viruses isolated in Africa, Europe and the Middle East between 2006 and early 2008. Detailed phylogenetic analyses of the 8 gene viral segments confirmed that 3 distinct sublineages were introduced, which have persisted and spread across the continent over this 2-year period. Additionally, our molecular epidemiological studies highlighted the association between genetic clustering and area of origin in a majority of cases. Molecular signatures unique to strains isolated in selected areas also gave us a clearer picture of the spread of A/H5N1 viruses across the continent. Mutations described as typical of human influenza viruses in the genes coding for internal proteins or associated with host adaptation and increased resistance to antiviral drugs have also been detected in the genes coding for transmembrane proteins. These findings raise concern for the possible human health risk presented by viruses with these genetic properties and highlight the need for increased efforts to monitor the evolution of A/H5N1 viruses across the African continent. They further stress how imperative it is to implement sustainable control strategies to improve animal and public health at a global level.

  13. Characterization of a non-pathogenic H5N1 influenza virus isolated from a migratory duck flying from Siberia in Hokkaido, Japan, in October 2009

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    Okamatsu Masatoshi

    2011-02-01

    Full Text Available Abstract Background Infection with H5N1 highly pathogenic avian influenza viruses (HPAIVs of domestic poultry and wild birds has spread to more than 60 countries in Eurasia and Africa. It is concerned that HPAIVs may be perpetuated in the lakes in Siberia where migratory water birds nest in summer. To monitor whether HPAIVs circulate in migratory water birds, intensive surveillance of avian influenza has been performed in Mongolia and Japan in autumn each year. Until 2008, there had not been any H5N1 viruses isolated from migratory water birds that flew from their nesting lakes in Siberia. In autumn 2009, A/mallard/Hokkaido/24/09 (H5N1 (Mal/Hok/24/09 was isolated from a fecal sample of a mallard (Anas platyrhynchos that flew from Siberia to Hokkaido, Japan. The isolate was assessed for pathogenicity in chickens, domestic ducks, and quails and analyzed antigenically and phylogenetically. Results No clinical signs were observed in chickens inoculated intravenously with Mal/Hok/24/09 (H5N1. There was no viral replication in chickens inoculated intranasally with the isolate. None of the domestic ducks and quails inoculated intranasally with the isolate showed any clinical signs. There were no multiple basic amino acid residues at the cleavage site of the hemagglutinin (HA of the isolate. Each gene of Mal/Hok/24/09 (H5N1 is phylogenetically closely related to that of influenza viruses isolated from migratory water birds that flew from their nesting lakes in autumn. Additionally, the antigenicity of the HA of the isolate was similar to that of the viruses isolated from migratory water birds in Hokkaido that flew from their northern territory in autumn and different from those of HPAIVs isolated from birds found dead in China, Mongolia, and Japan on the way back to their northern territory in spring. Conclusion Mal/Hok/24/09 (H5N1 is a non-pathogenic avian influenza virus for chickens, domestic ducks, and quails, and is antigenically and genetically

  14. Experimental challenge and pathology of highly pathogenic avian influenza virus H5N1 in dunlin (Calidris alpina), an intercontinental migrant shorebird species.

    Science.gov (United States)

    Hall, Jeffrey S; Franson, J Christian; Gill, Robert E; Meteyer, Carol U; TeSlaa, Joshua L; Nashold, Sean; Dusek, Robert J; Ip, Hon S

    2011-09-01

    Shorebirds (Charadriiformes) are considered one of the primary reservoirs of avian influenza. Because these species are highly migratory, there is concern that infected shorebirds may be a mechanism by which highly pathogenic avian influenza virus (HPAIV) H5N1 could be introduced into North America from Asia. Large numbers of dunlin (Calidris alpina) migrate from wintering areas in central and eastern Asia, where HPAIV H5N1 is endemic, across the Bering Sea to breeding areas in Alaska. Low pathogenic avian influenza virus has been previously detected in dunlin, and thus, dunlin represent a potential risk to transport HPAIV to North America. To date no experimental challenge studies have been performed in shorebirds. Wild dunlin were inoculated intranasally and intrachoanally various doses of HPAIV H5N1. The birds were monitored daily for virus excretion, disease signs, morbidity, and mortality. The infectious dose of HPAIV H5N1 in dunlin was determined to be 10(1.7) EID(50)/100 μl and that the lethal dose was 10(1.83) EID(50)/100 μl. Clinical signs were consistent with neurotropic disease, and histochemical analyses revealed that infection was systemic with viral antigen and RNA most consistently found in brain tissues. Infected birds excreted relatively large amounts of virus orally (10(4) EID(50)) and smaller amounts cloacally. Dunlin are highly susceptible to infection with HPAIV H5N1. They become infected after exposure to relatively small doses of the virus and if they become infected, they are most likely to suffer mortality within 3-5 days. These results have important implications regarding the risks of transport and transmission of HPAIV H5N1 to North America by this species and raises questions for further investigation. Published 2011. This article is a US Government work and is in the public domain in the USA.

  15. Detection of H5N1 high-pathogenicity avian influenza virus in meat and tracheal samples from experimentally infected chickens.

    Science.gov (United States)

    Das, Amaresh; Spackman, Erica; Thomas, Colleen; Swayne, David E; Suarez, David L

    2008-03-01

    The Asian H5N1 highly pathogenic avian influenza (HPAI) virus causes a systemic disease with high mortality of poultry and is potentially zoonotic. In both chickens and ducks, the virus has been demonstrated to replicate in both cardiac and skeletal muscle cells. Experimentally, H5N1 HPAI virus has been transmitted to chickens through the consumption of raw infected meat. In this study, we investigated virus replication in cardiac and skeletal muscle and in the trachea of chickens after experimental intranasal inoculation with the H5N1 HPAI virus. The virus was detected in tissues by real-time reverse transcription-polymerase chain reaction (RRT-PCR) and virus isolation, and in the trachea by RRT-PCR and a commercial avian influenza (AI) viral antigen detection test. A modified RNA extraction protocol was developed for rapid detection of the virus in tissues by RRT-PCR. The H5N1 HPAI virus was sporadically detected in meat and the tracheas of infected birds without any clinical sign of disease as early as 6 hr postinfection (PI), and was detected in all samples tested at 24 hr PI and later. No differences in sensitivity were seen between virus isolation and RRT-PCR in meat samples. The AI viral antigen detection test on tracheal swabs was a useful method for identifying infected chickens when they were sick or dead, but was less sensitive in detecting infected birds when they were preclinical. This study provides data indicating that preslaughter tracheal swab testing can identify birds infected with HPAI among the daily mortality and prevent infected flocks from being sent to processing plants. In addition, the modified RNA extraction and RRT-PCR test on meat samples provide a rapid and sensitive method of identifying HPAI virus in illegal contraband or domestic meat samples.

  16. Emerging highly pathogenic H5 avian influenza viruses in France during winter 2015/16: phylogenetic analyses and markers for zoonotic potential.

    Science.gov (United States)

    Briand, François-Xavier; Schmitz, Audrey; Ogor, Katell; Le Prioux, Aurélie; Guillou-Cloarec, Cécile; Guillemoto, Carole; Allée, Chantal; Le Bras, Marie-Odile; Hirchaud, Edouard; Quenault, Hélène; Touzain, Fabrice; Cherbonnel-Pansart, Martine; Lemaitre, Evelyne; Courtillon, Céline; Gares, Hélène; Daniel, Patrick; Fediaevsky, Alexandre; Massin, Pascale; Blanchard, Yannick; Eterradossi, Nicolas; van der Werf, Sylvie; Jestin, Véronique; Niqueux, Eric

    2017-03-02

    Several new highly pathogenic (HP) H5 avian influenza virus (AIV) have been detected in poultry farms from south-western France since November 2015, among which an HP H5N1. The zoonotic potential and origin of these AIVs immediately became matters of concern. One virus of each subtype H5N1 (150169a), H5N2 (150233) and H5N9 (150236) was characterised. All proved highly pathogenic for poultry as demonstrated molecularly by the presence of a polybasic cleavage site in their HA protein - with a sequence (HQRRKR/GLF) previously unknown among avian H5 HPAI viruses - or experimentally by the in vivo demonstration of an intravenous pathogenicity index of 2.9 for the H5N1 HP isolate. Phylogenetic analyses based on the full genomes obtained by NGS confirmed that the eight viral segments of the three isolates were all part of avian Eurasian phylogenetic lineage but differed from the Gs/Gd/1/96-like lineage. The study of the genetic characteristics at specific amino acid positions relevant for modulating the adaptation to and the virulence for mammals showed that presently, these viruses possess most molecular features characteristic of AIV and lack some major characteristics required for efficient respiratory transmission to or between humans. The three isolates are therefore predicted to have no significant pandemic potential. This article is copyright of The Authors, 2017.

  17. Lethal infection by a novel reassortant H5N1 avian influenza A virus in a zoo-housed tiger.

    Science.gov (United States)

    He, Shang; Shi, Jianzhong; Qi, Xian; Huang, Guoqing; Chen, Hualan; Lu, Chengping

    2015-01-01

    In early 2013, a Bengal tiger (Panthera tigris) in a zoo died of respiratory distress. All specimens from the tiger were positive for HPAI H5N1, which were detected by real-time PCR, including nose swab, throat swab, tracheal swab, heart, liver, spleen, lung, kidney, aquae pericardii and cerebrospinal fluid. One stain of virus, A/Tiger/JS/1/2013, was isolated from the lung sample. Pathogenicity experiments showed that the isolate was able to replicate and cause death in mice. Phylogenetic analysis indicated that HA and NA of A/Tiger/JS/1/2013 clustered with A/duck/Vietnam/OIE-2202/2012 (H5N1), which belongs to clade 2.3.2.1. Interestingly, the gene segment PB2 shared 98% homology with A/wild duck/Korea/CSM-28/20/2010 (H4N6), which suggested that A/Tiger/JS/1/2013 is a novel reassortant H5N1 subtype virus. Immunohistochemical analysis also confirmed that the tiger was infected by this new reassortant HPAI H5N1 virus. Overall, our results showed that this Bengal tiger was infected by a novel reassortant H5N1, suggesting that the H5N1 virus can successfully cross species barriers from avian to mammal through reassortment. Copyright © 2014 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  18. Challenge for One Health: Co-Circulation of Zoonotic H5N1 and H9N2 Avian Influenza Viruses in Egypt.

    Science.gov (United States)

    Kim, Shin-Hee

    2018-03-09

    Highly pathogenic avian influenza (HPAI) H5N1 viruses are currently endemic in poultry in Egypt. Eradication of the viruses has been unsuccessful due to improper application of vaccine-based control strategies among other preventive measures. The viruses have evolved rapidly with increased bird-to-human transmission efficacy, thus affecting both animal and public health. Subsequent spread of potentially zoonotic low pathogenic avian influenza (LPAI) H9N2 in poultry has also hindered efficient control of avian influenza. The H5N1 viruses acquired enhanced bird-to-human transmissibility by (1) altering amino acids in hemagglutinin (HA) that enable binding affinity to human-type receptors, (2) loss of the glycosylation site and 130 loop in the HA protein and (3) mutation of E627K in the PB2 protein to enhance viral replication in mammalian hosts. The receptor binding site of HA of Egyptian H9N2 viruses has been shown to contain the Q234L substitution along with a H191 mutation, which can increase human-like receptor specificity. Therefore, co-circulation of H5N1 and H9N2 viruses in poultry farming and live bird markets has increased the risk of human exposure, resulting in complication of the epidemiological situation and raising a concern for potential emergence of a new influenza A virus pandemic. For efficient control of infection and transmission, the efficacy of vaccine and vaccination needs to be improved with a comprehensive control strategy, including enhanced biosecurity, education, surveillance, rapid diagnosis and culling of infected poultry.

  19. Challenge for One Health: Co-Circulation of Zoonotic H5N1 and H9N2 Avian Influenza Viruses in Egypt

    Directory of Open Access Journals (Sweden)

    Shin-Hee Kim

    2018-03-01

    Full Text Available Highly pathogenic avian influenza (HPAI H5N1 viruses are currently endemic in poultry in Egypt. Eradication of the viruses has been unsuccessful due to improper application of vaccine-based control strategies among other preventive measures. The viruses have evolved rapidly with increased bird-to-human transmission efficacy, thus affecting both animal and public health. Subsequent spread of potentially zoonotic low pathogenic avian influenza (LPAI H9N2 in poultry has also hindered efficient control of avian influenza. The H5N1 viruses acquired enhanced bird-to-human transmissibility by (1 altering amino acids in hemagglutinin (HA that enable binding affinity to human-type receptors, (2 loss of the glycosylation site and 130 loop in the HA protein and (3 mutation of E627K in the PB2 protein to enhance viral replication in mammalian hosts. The receptor binding site of HA of Egyptian H9N2 viruses has been shown to contain the Q234L substitution along with a H191 mutation, which can increase human-like receptor specificity. Therefore, co-circulation of H5N1 and H9N2 viruses in poultry farming and live bird markets has increased the risk of human exposure, resulting in complication of the epidemiological situation and raising a concern for potential emergence of a new influenza A virus pandemic. For efficient control of infection and transmission, the efficacy of vaccine and vaccination needs to be improved with a comprehensive control strategy, including enhanced biosecurity, education, surveillance, rapid diagnosis and culling of infected poultry.

  20. Characterization of H5N6 highly pathogenic avian influenza viruses isolated from wild and captive birds in the winter season of 2016-2017 in Northern Japan.

    Science.gov (United States)

    Hiono, Takahiro; Okamatsu, Masatoshi; Matsuno, Keita; Haga, Atsushi; Iwata, Ritsuko; Nguyen, Lam Thanh; Suzuki, Mizuho; Kikutani, Yuto; Kida, Hiroshi; Onuma, Manabu; Sakoda, Yoshihiro

    2017-09-01

    On 15 November 2016, a black swan that had died in a zoo in Akita prefecture, northern Japan, was strongly suspected to have highly pathogenic avian influenza (HPAI); an HPAI virus (HPAIV) belonging to the H5N6 subtype was isolated from specimens taken from the bird. After the initial report, 230 cases of HPAI caused by H5N6 viruses from wild birds, captive birds, and domestic poultry farms were reported throughout the country during the winter season. In the present study, 66 H5N6 HPAIVs isolated from northern Japan were further characterized. Phylogenetic analysis of the hemagglutinin gene showed that the H5N6 viruses isolated in northern Japan clustered into Group C of Clade 2.3.4.4 together with other isolates collected in Japan, Korea and Taiwan during the winter season of 2016-2017. The antigenicity of the Japanese H5N6 isolate differed slightly from that of HPAIVs isolated previously in Japan and China. The virus exhibited high pathogenicity and a high replication capacity in chickens, whereas virus growth was slightly lower in ducks compared with that of an H5N8 HPAIV isolate collected in Japan in 2014. Comprehensive analyses of Japanese isolates, including those from central, western, and southern Japan, as well as rapid publication of this information are essential for facilitating greater control of HPAIVs. © 2017 The Societies and John Wiley & Sons Australia, Ltd.

  1. Effect of neuraminidase inhibitor-resistant mutations on pathogenicity of clade 2.2 A/Turkey/15/06 (H5N1) influenza virus in ferrets.

    Science.gov (United States)

    Ilyushina, Natalia A; Seiler, Jon P; Rehg, Jerold E; Webster, Robert G; Govorkova, Elena A

    2010-05-27

    The acquisition of neuraminidase (NA) inhibitor resistance by H5N1 influenza viruses has serious clinical implications, as this class of drugs can be an essential component of pandemic control measures. The continuous evolution of the highly pathogenic H5N1 influenza viruses results in the emergence of natural NA gene variations whose impact on viral fitness and NA inhibitor susceptibility are poorly defined. We generated seven genetically stable recombinant clade 2.2 A/Turkey/15/06-like (H5N1) influenza viruses carrying NA mutations located either in the framework residues (E119A, H274Y, N294S) or in close proximity to the NA enzyme active site (V116A, I117V, K150N, Y252H). NA enzyme inhibition assays showed that NA mutations at positions 116, 117, 274, and 294 reduced susceptibility to oseltamivir carboxylate (IC(50)s increased 5- to 940-fold). Importantly, the E119A NA mutation (previously reported to confer resistance in the N2 NA subtype) was stable in the clade 2.2 H5N1 virus background and induced cross-resistance to oseltamivir carboxylate and zanamivir. We demonstrated that Y252H NA mutation contributed for decreased susceptibility of clade 2.2 H5N1 viruses to oseltamivir carboxylate as compared to clade 1 viruses. The enzyme kinetic parameters (V(max), K(m) and K(i)) of the avian-like N1 NA glycoproteins were highly consistent with their IC(50) values. None of the recombinant H5N1 viruses had attenuated virulence in ferrets inoculated with 10(6) EID(50) dose. Most infected ferrets showed mild clinical disease signs that differed in duration. However, H5N1 viruses carrying the E119A or the N294S NA mutation were lethal to 1 of 3 inoculated animals and were associated with significantly higher virus titers (Pinfluenza drugs that target different virus/host factors and can limit the emergence of resistance.

  2. Unique Infectious Strategy of H5N1 Avian Influenza Virus Is Governed by the Acid-Destabilized Property of Hemagglutinin.

    Science.gov (United States)

    Daidoji, Tomo; Watanabe, Yohei; Arai, Yasuha; Kajikawa, Junichi; Hirose, Ryohei; Nakaya, Takaaki

    Highly pathogenic avian influenza (HPAI) H5N1 virus emerged in 1997 as a zoonotic disease in Hong Kong. It has since spread to Asia and Europe and is a serious threat to both the poultry industry and human health. For effective surveillance and possible prevention/control of HPAI H5N1 viruses, it is necessary to understand the molecular mechanism underlying HPAI H5N1 pathogenesis. The hemagglutinin (HA) protein of influenza A viruses (IAVs) is one of the major determinants of host adaptation, transmissibility, and viral virulence. The main function of the HA protein is to facilitate viral entry and viral genome release within host cells before infection. To achieve viral infection, IAVs belonging to different subtypes or strains induce viral-cell membrane fusion at different endosomal pH levels after internalization through endocytosis. However, host-specific endosomal pH also affects induction of membrane fusion followed by infection. The HA protein of HPAI H5N1 has a higher pH threshold for membrane fusion than the HA protein of classical avian influenza viruses. Although this particular property of HA (which governs viral infection) is prone to deactivation in the avian intestine or in an ambient environment, it facilitates efficient infection of host cells, resulting in a broad host tropism, regardless of the pH in the host endosome. Accumulated knowledge, together with further research, about the HA-governed mechanism underlying HPAI H5N1 virulence (i.e., receptor tropism and pH-dependent viral-cell membrane fusion) will be helpful for developing effective surveillance strategies and for prevention/control of HPAI H5N1 infection.

  3. DC-SIGN mediates avian H5N1 influenza virus infection in cis and in trans

    International Nuclear Information System (INIS)

    Wang, S.-F.; Huang, Jason C.; Lee, Y.-M.; Liu, S.-J.; Chan, Yu-Jiun; Chau, Y.-P.; Chong, P.; Chen, Y.-M.A.

    2008-01-01

    DC-SIGN, a C-type lectin receptor expressed in dendritic cells (DCs), has been identified as a receptor for human immunodeficiency virus type 1, hepatitis C virus, Ebola virus, cytomegalovirus, dengue virus, and the SARS coronavirus. We used H5N1 pseudotyped and reverse-genetics (RG) virus particles to study their ability to bind with DC-SIGN. Electronic microscopy and functional assay results indicate that pseudotyped viruses containing both HA and NA proteins express hemagglutination and are capable of infecting cells expressing α-2,3-linked sialic acid receptors. Results from a capture assay show that DC-SIGN-expressing cells (including B-THP-1/DC-SIGN and T-THP-1/DC-SIGN) and peripheral blood dendritic cells are capable of transferring H5N1 pseudotyped and RG virus particles to target cells; this action can be blocked by anti-DC-SIGN monoclonal antibodies. In summary, (a) DC-SIGN acts as a capture or attachment molecule for avian H5N1 virus, and (b) DC-SIGN mediates infections in cis and in trans

  4. H5N2 Highly Pathogenic Avian Influenza Viruses from the US 2014-2015 outbreak have an unusually long pre-clinical period in turkeys

    OpenAIRE

    Spackman, Erica; Pantin-Jackwood, Mary J.; Kapczynski, Darrell R.; Swayne, David E.; Suarez, David L.

    2016-01-01

    Background From December 2014 through June 2015, the US experienced the most costly highly pathogenic avian influenza (HPAI) outbreak to date. Most cases in commercial poultry were caused by an H5N2 strain which was a reassortant with 5 Eurasian lineage genes, including a clade 2.3.4.4 goose/Guangdong/1996 lineage hemagglutinin, and 3 genes from North American wild waterfowl low pathogenicity avian influenza viruses. The outbreak primarily affected turkeys and table-egg layer type chickens. T...

  5. Broadly-reactive human monoclonal antibodies elicited following pandemic H1N1 influenza virus exposure protect mice from highly pathogenic H5N1 challenge.

    Science.gov (United States)

    Nachbagauer, Raffael; Shore, David; Yang, Hua; Johnson, Scott K; Gabbard, Jon D; Tompkins, S Mark; Wrammert, Jens; Wilson, Patrick C; Stevens, James; Ahmed, Rafi; Krammer, Florian; Ellebedy, Ali H

    2018-06-13

    Broadly cross-reactive antibodies that recognize conserved epitopes within the influenza virus hemagglutinin (HA) stalk domain are of particular interest for their potential use as therapeutic and prophylactic agents against multiple influenza virus subtypes including zoonotic virus strains. Here, we characterized four human HA stalk-reactive monoclonal antibodies (mAbs) for their binding breadth and affinity, in vitro neutralization capacity, and in vivo protective potential against an highly pathogenic avian influenza virus. The monoclonal antibodies were isolated from individuals shortly following infection with (70-1F02 and 1009-3B05) or vaccination against (05-2G02 and 09-3A01) A(H1N1)pdm09. Three of the mAbs bound HAs from multiple strains of group 1 viruses, and one mAb, 05-2G02, bound to both group 1 and group 2 influenza A HAs. All four antibodies prophylactically protected mice against a lethal challenge with the highly pathogenic A/Vietnam/1203/04 (H5N1) strain. Two mAbs, 70-1F02 and 09-3A01, were further tested for their therapeutic efficacy against the same strain and showed good efficacy in this setting as well. One mAb, 70-1F02, was co-crystallized with H5 HA and showed similar heavy chain only interactions as a the previously described anti-stalk antibody CR6261. Finally, we showed that antibodies that compete with these mAbs are prevalent in serum from an individual recently infected with A(H1N1)pdm09 virus. The antibodies described here can be developed into broad-spectrum antiviral therapeutics that could be used to combat infections with zoonotic or emerging pandemic influenza viruses. IMPORTANCE The rise in zoonotic infections of humans with emerging influenza viruses is a worldwide public health concern. The majority of recent zoonotic human influenza cases were caused by H7N9 and H5Nx viruses and were associated with high morbidity and mortality. In addition, seasonal influenza viruses are estimated to cause up to 650,000 deaths annually

  6. Live bird markets of Bangladesh: H9N2 viruses and the near absence of highly pathogenic H5N1 influenza.

    Directory of Open Access Journals (Sweden)

    Nicholas J Negovetich

    2011-04-01

    Full Text Available Avian influenza surveillance in Bangladesh has been passive, relying on poultry farmers to report suspected outbreaks of highly pathogenic H5N1 influenza. Here, the results of an active surveillance effort focusing on the live-bird markets are presented. Prevalence of influenza infection in the birds of the live bird markets is 23.0%, which is similar to that in poultry markets in other countries. Nearly all of the isolates (94% were of the non-pathogenic H9N2 subtype, but viruses of the H1N2, H1N3, H3N6, H4N2, H5N1, and H10N7 subtypes were also observed. The highly pathogenic H5N1-subtype virus was observed at extremely low prevalence in the surveillance samples (0.08%, and we suggest that the current risk of infection for humans in the retail poultry markets in Bangladesh is negligible. However, the high prevalence of the H9 subtype and its potential for interaction with the highly pathogenic H5N1-subtype, i.e., reassortment and attenuation of host morbidity, highlight the importance of active surveillance of the poultry markets.

  7. First introduction of highly pathogenic H5NI avian influenza A viruses in wild and domestic birds in Denmark, Northern Europe

    DEFF Research Database (Denmark)

    Bragstad, K.; Jørgensen, Poul Henrik; Handberg, Kurt

    2007-01-01

    Background: Since 2005 highly pathogenic ( HP) avian influenza A H5N1 viruses have spread from Asia to Africa and Europe infecting poultry, humans and wild birds. HP H5N1 virus was isolated in Denmark for the first time in March 2006. A total of 44 wild birds were found positive for the HP H5N1...... infection. In addition, one case was reported in a backyard poultry flock. Results: Full-genome characterisation of nine isolates revealed that the Danish H5N1 viruses were highly similar to German H5N1 isolates in all genes from the same time period. The haemagglutinin gene grouped phylogenetically in H5...... clade 2 subclade 2 and closest relatives besides the German isolates were isolates from Croatia in 2005, Nigeria and Niger in 2006 and isolates from Astrakhan in Russia 2006. The German and Danish isolates shared unique substitutions in the NA, PB1 and NS2 proteins. Conclusion: The first case of HP H5N1...

  8. Pathogenesis of the 1918 pandemic and H5N1 influenza virus infection in a guinea pig model: The antiviral potential of exogenous alpha-interferon to reduce virus shedding

    Science.gov (United States)

    Although highly pathogenic avian influenza H5N1 viruses have yet to acquire the ability to transmit efficiently among humans, the geographic expansion, and continued outbreaks in humans and avian species underscore the need for more effective influenza vaccines and antivirals. Additional small anim...

  9. Migration of whooper swans and outbreaks of highly pathogenic avian influenza H5N1 virus in eastern Asia.

    Directory of Open Access Journals (Sweden)

    Scott H Newman

    Full Text Available Evaluating the potential involvement of wild avifauna in the emergence of highly pathogenic avian influenza H5N1 (hereafter H5N1 requires detailed analyses of temporal and spatial relationships between wild bird movements and disease emergence. The death of wild swans (Cygnus spp. has been the first indicator of the presence of H5N1 in various Asian and European countries; however their role in the geographic spread of the disease remains poorly understood. We marked 10 whooper swans (Cygnus cygnus with GPS transmitters in northeastern Mongolia during autumn 2006 and tracked their migratory movements in relation to H5N1 outbreaks. The prevalence of H5N1 outbreaks among poultry in eastern Asia during 2003-2007 peaked during winter, concurrent with whooper swan movements into regions of high poultry density. However outbreaks involving poultry were detected year round, indicating disease perpetuation independent of migratory waterbird presence. In contrast, H5N1 outbreaks involving whooper swans, as well as other migratory waterbirds that succumbed to the disease in eastern Asia, tended to occur during seasons (late spring and summer and in habitats (areas of natural vegetation where their potential for contact with poultry is very low to nonexistent. Given what is known about the susceptibility of swans to H5N1, and on the basis of the chronology and rates of whooper swan migration movements, we conclude that although there is broad spatial overlap between whooper swan distributions and H5N1 outbreak locations in eastern Asia, the likelihood of direct transmission between these groups is extremely low. Thus, our data support the hypothesis that swans are best viewed as sentinel species, and moreover, that in eastern Asia, it is most likely that their infections occurred through contact with asymptomatic migratory hosts (e.g., wild ducks at or near their breeding grounds.

  10. Migration of whooper swans and outbreaks of highly pathogenic avian influenza H5N1 virus in eastern Asia.

    Science.gov (United States)

    Newman, Scott H; Iverson, Samuel A; Takekawa, John Y; Gilbert, Martin; Prosser, Diann J; Batbayar, Nyambyar; Natsagdorj, Tseveenmyadag; Douglas, David C

    2009-05-28

    Evaluating the potential involvement of wild avifauna in the emergence of highly pathogenic avian influenza H5N1 (hereafter H5N1) requires detailed analyses of temporal and spatial relationships between wild bird movements and disease emergence. The death of wild swans (Cygnus spp.) has been the first indicator of the presence of H5N1 in various Asian and European countries; however their role in the geographic spread of the disease remains poorly understood. We marked 10 whooper swans (Cygnus cygnus) with GPS transmitters in northeastern Mongolia during autumn 2006 and tracked their migratory movements in relation to H5N1 outbreaks. The prevalence of H5N1 outbreaks among poultry in eastern Asia during 2003-2007 peaked during winter, concurrent with whooper swan movements into regions of high poultry density. However outbreaks involving poultry were detected year round, indicating disease perpetuation independent of migratory waterbird presence. In contrast, H5N1 outbreaks involving whooper swans, as well as other migratory waterbirds that succumbed to the disease in eastern Asia, tended to occur during seasons (late spring and summer) and in habitats (areas of natural vegetation) where their potential for contact with poultry is very low to nonexistent. Given what is known about the susceptibility of swans to H5N1, and on the basis of the chronology and rates of whooper swan migration movements, we conclude that although there is broad spatial overlap between whooper swan distributions and H5N1 outbreak locations in eastern Asia, the likelihood of direct transmission between these groups is extremely low. Thus, our data support the hypothesis that swans are best viewed as sentinel species, and moreover, that in eastern Asia, it is most likely that their infections occurred through contact with asymptomatic migratory hosts (e.g., wild ducks) at or near their breeding grounds.

  11. Experimental infection with highly pathogenic H5N8 avian influenza viruses in the Mandarin duck (Aix galericulata) and domestic pigeon (Columba livia domestica).

    Science.gov (United States)

    Kwon, Jung-Hoon; Noh, Yun Kyung; Lee, Dong-Hun; Yuk, Seong-Su; Erdene-Ochir, Tseren-Ochir; Noh, Jin-Yong; Hong, Woo-Tack; Jeong, Jei-Hyun; Jeong, Sol; Gwon, Gyeong-Bin; Song, Chang-Seon; Nahm, Sang-Soep

    2017-05-01

    Wild birds play a major role in the evolution, maintenance, and dissemination of highly pathogenic avian influenza viruses (HPAIV). Sub-clinical infection with HPAI in resident wild birds could be a source of dissemination of HPAIV and continuous outbreaks. In this study, the pathogenicity and infectivity of two strains of H5N8 clade 2.3.4.4 virus were evaluated in the Mandarin duck (Aix galericulata) and domestic pigeon (Columba livia domestica). None of the birds experimentally infected with H5N8 viruses showed clinical signs or mortality. The H5N8 viruses efficiently replicated in the virus-inoculated Mandarin ducks and transmitted to co-housed Mandarin ducks. Although relatively high levels of viral shedding were noted in pigeons, viral shedding was not detected in some of the pigeons and the shedding period was relatively short. Furthermore, the infection was not transmitted to co-housed pigeons. Immunohistochemical examination revealed the presence of HPAIV in multiple organs of the infected birds. Histopathological evaluation showed the presence of inflammatory responses primarily in HPAIV-positive organs. Our results indicate that Mandarin ducks and pigeons can be infected with H5N8 HPAIV without exhibiting clinical signs; thus, they may be potential healthy reservoirs of the H5N8 HPAIV. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. A unique influenza A (H5N1 virus causing a focal poultry outbreak in 2007 in Manipur, India

    Directory of Open Access Journals (Sweden)

    Raut Satish

    2009-02-01

    Full Text Available Abstract Background A focal H5N1 outbreak in poultry was reported from Manipur, a north-eastern state, of India, in 2007. The aim of this study was to genetically characterize the Manipur isolate to understand the relationship with other H5N1 isolates and to trace the possible source of introduction of the virus into the country. Results Characterization of the complete genome revealed that the virus belonged to clade 2.2. It was distinctly different from viruses of the three EMA sublineages of clade 2.2 but related to isolates from wild migratory waterfowl from Russia, China and Mongolia. The HA gene, had the cleavage site GERRRRKR, earlier reported in whooper swan isolates from Mongolia in 2005. A stop codon at position 29 in the PB1-F2 protein could have implications on the replication efficiency. The acquisition of polymorphisms as seen in recent isolates of 2005–07 from distinct geographical regions suggests the possibility of transportation of H5N1 viruses through migratory birds. Conclusion Considering that all eight genes of the earlier Indian isolates belonged to the EMA3 sublineage and similar strains have not been reported from neighbouring countries of the subcontinent, it appears that the virus may have been introduced independently.

  13. Differential host determinants contribute to the pathogenesis of 2009 pandemic H1N1 and human H5N1 influenza A viruses in experimental mouse models.

    Science.gov (United States)

    Otte, Anna; Sauter, Martina; Alleva, Lisa; Baumgarte, Sigrid; Klingel, Karin; Gabriel, Gülsah

    2011-07-01

    Influenza viruses are responsible for high morbidities in humans and may, eventually, cause pandemics. Herein, we compared the pathogenesis and host innate immune responses of a seasonal H1N1, two 2009 pandemic H1N1, and a human H5N1 influenza virus in experimental BALB/c and C57BL/6J mouse models. We found that both 2009 pandemic H1N1 isolates studied (A/Hamburg/05/09 and A/Hamburg/NY1580/09) were low pathogenic in BALB/c mice [log mouse lethal dose 50 (MLD(50)) >6 plaque-forming units (PFU)] but displayed remarkable differences in virulence in C57BL/6J mice. A/Hamburg/NY1580/09 was more virulent (logMLD(50) = 3.5 PFU) than A/Hamburg/05/09 (logMLD(50) = 5.2 PFU) in C57BL/6J mice. In contrast, the H5N1 influenza virus was more virulent in BALB/c mice (logMLD(50) = 0.3 PFU) than in C57BL/6J mice (logMLD(50) = 1.8 PFU). Seasonal H1N1 influenza revealed marginal pathogenicity in BALB/c or C57BL/6J mice (logMLD(50) >6 PFU). Enhanced susceptibility of C57BL/6J mice to pandemic H1N1 correlated with a depressed cytokine response. In contrast, enhanced H5N1 virulence in BALB/c mice correlated with an elevated proinflammatory cytokine response. These findings highlight that host determinants responsible for the pathogenesis of 2009 pandemic H1N1 influenza viruses are different from those contributing to H5N1 pathogenesis. Our results show, for the first time to our knowledge, that the C57BL/6J mouse strain is more appropriate for the evaluation and identification of intrinsic pathogenicity markers of 2009 pandemic H1N1 influenza viruses that are "masked" in BALB/c mice. Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  14. Southward autumn migration of waterfowl facilitates cross-continental transmission of the highly pathogenic avian influenza H5N1 virus

    Science.gov (United States)

    Xu, Yanjie; Gong, Peng; Wielstra, Ben; Si, Yali

    2016-08-01

    The highly pathogenic avian influenza subtype H5N1 (HPAI H5N1) is a worldwide zoonotic infectious disease, threatening humans, poultry and wild birds. The role of wild birds in the spread of HPAI H5N1 has previously been investigated by comparing disease spread patterns with bird migration routes. However, the different roles that the southward autumn and northward spring migration might play in virus transmission have hardly been explored. Using direction analysis, we analyze HPAI H5N1 transmission directions and angular concentration of currently circulating viral clades, and compare these with waterfowl seasonal migration directions along major waterfowl flyways. Out of 22 HPAI H5N1 transmission directions, 18 had both a southward direction and a relatively high concentration. Differences between disease transmission and waterfowl migration directions were significantly smaller for autumn than for spring migration. The four northward transmission directions were found along Asian flyways, where the initial epicenter of the virus was located. We suggest waterfowl first picked up the virus from East Asia, then brought it to the north via spring migration, and then spread it to other parts of world mainly by autumn migration. We emphasize waterfowl autumn migration plays a relatively important role in HPAI H5N1 transmission compared to spring migration.

  15. The NS segment of H5N1 avian influenza viruses (AIV) enhances the virulence of an H7N1 AIV in chickens.

    Science.gov (United States)

    Vergara-Alert, Júlia; Busquets, Núria; Ballester, Maria; Chaves, Aida J; Rivas, Raquel; Dolz, Roser; Wang, Zhongfang; Pleschka, Stephan; Majó, Natàlia; Rodríguez, Fernando; Darji, Ayub

    2014-01-25

    Some outbreaks involving highly pathogenic avian influenza viruses (HPAIV) of subtypes H5 and H7 were caused by avian-to-human transmissions. In nature, different influenza A viruses can reassort leading to new viruses with new characteristics. We decided to investigate the impact that the NS-segment of H5 HPAIV would have on viral pathogenicity of a classical avian H7 HPAIV in poultry, a natural host. We focussed this study based on our previous work that demonstrated that single reassortment of the NS-segment from an H5 HPAIV into an H7 HPAIV changes the ability of the virus to replicate in mammalian hosts. Our present data show that two different H7-viruses containing an NS-segment from H5-types (FPV NS GD or FPV NS VN) show an overall highly pathogenic phenotype compared with the wild type H7-virus (FPV), as characterized by higher viral shedding and earlier manifestation of clinical signs. Correlating with the latter, higher amounts of IFN-β mRNA were detected in the blood of NS-reassortant infected birds, 48 h post-infection (pi). Although lymphopenia was detected in chickens from all AIV-infected groups, also 48 h pi those animals challenged with NS-reassortant viruses showed an increase of peripheral monocyte/macrophage-like cells expressing high levels of IL-1β, as determined by flow cytometry. Taken together, these findings highlight the importance of the NS-segment in viral pathogenicity which is directly involved in triggering antiviral and pro-inflammatory cytokines found during HPAIV pathogenesis in chickens.

  16. Pathology of natural infections by H5N1 highly pathogenic avian influenza virus in mute (Cygnus olor) and whooper (Cygnus cygnus) swans.

    Science.gov (United States)

    Teifke, J P; Klopfleisch, R; Globig, A; Starick, E; Hoffmann, B; Wolf, P U; Beer, M; Mettenleiter, T C; Harder, T C

    2007-03-01

    Mortality in wild aquatic birds due to infection with highly pathogenic avian influenza viruses (HPAIV) is a rare event. During the recent outbreak of highly pathogenic avian influenza in Germany, mortality due to H5N1 HPAIV was observed among mute and whooper swans as part of a rapid spread of this virus. In contrast to earlier reports, swans appeared to be highly susceptible and represented the mainly affected species. We report gross and histopathology and distribution of influenza virus antigen in mute and whooper swans that died after natural infection with H5N1 HPAIV. At necropsy, the most reliable lesions were multifocal hemorrhagic necrosis in the pancreas, pulmonary congestion and edema, and subepicardial hemorrhages. Major histologic lesions were acute pancreatic necrosis, multifocal necrotizing hepatitis, and lymphoplasmacytic encephalitis with neuronal necrosis. Adrenals displayed consistently scattered cortical and medullary necrosis. In spleen and Peyer's patches, mild lymphocyte necrosis was present. Immunohistochemical demonstration of HPAIV nucleoprotein in pancreas, adrenals, liver, and brain was strongly consistent with histologic lesions. In the brain, a large number of neurons and glial cells, especially Purkinje cells, showed immunostaining. Occasionally, ependymal cells of the spinal cord were also positive. In the lungs, influenza virus antigen was identified in a few endothelial cells but not within pneumocytes. The infection of the central nervous system supports the view that the neurotropism of H5N1 HPAIV leads to nervous disturbances with loss of orientation. More investigations are necessary to clarify the mechanisms of the final circulatory failure, lung edema, and rapid death of the swans.

  17. Surveillance plan for the early detection of H5N1 highly pathogenic avian influenza virus in migratory birds in the United States: surveillance year 2009

    Science.gov (United States)

    Brand, Christopher J.

    2009-01-01

    Executive Summary: This Surveillance Plan (Plan) describes plans for conducting surveillance of wild birds in the United States and its Territories and Freely-Associated States to provide for early detection of the introduction of the H5N1 Highly Pathogenic Avian Influenza (HPAI) subtype of the influenza A virus by migratory birds during the 2009 surveillance year, spanning the period of April 1, 2009 - March 31, 2010. The Plan represents a continuation of surveillance efforts begun in 2006 under the Interagency Strategic Plan for the Early Detection of H5N1 Highly Pathogenic Avian Influenza in Wild Migratory Birds (U.S. Department of Agriculture and U.S. Department of the Interior, 2006). The Plan sets forth sampling plans by: region, target species or species groups to be sampled, locations of sampling, sample sizes, and sampling approaches and methods. This Plan will be reviewed annually and modified as appropriate for subsequent surveillance years based on evaluation of information from previous years of surveillance, changing patterns and threats of H5N1 HPAI, and changes in funding availability for avian influenza surveillance. Specific sampling strategies will be developed accordingly within each of six regions, defined here as Alaska, Hawaiian/Pacific Islands, Lower Pacific Flyway (Washington, Oregon, California, Idaho, Nevada, Arizona), Central Flyway, Mississippi Flyway, and Atlantic Flyway.

  18. Indium-tin-oxide thin film transistor biosensors for label-free detection of avian influenza virus H5N1

    International Nuclear Information System (INIS)

    Guo, Di; Zhuo, Ming; Zhang, Xiaoai; Xu, Cheng; Jiang, Jie; Gao, Fu; Wan, Qing; Li, Qiuhong; Wang, Taihong

    2013-01-01

    Highlights: ► A highly selective label-free biosensor is established based on indium-tin-oxide thin-film transistors (ITO TFTs). ► AI H5N1 virus was successfully detected through shift in threshold voltage and field-effect mobility of ITO TFT. ► The ITO TFT is applied in biosensor for the first time and shows good reusability and stability. ► Fabrication of the platform is simple with low cost, which is suitable for mass commercial production. -- Abstract: As continuous outbreak of avian influenza (AI) has become a threat to human health, economic development and social stability, it is urgently necessary to detect the highly pathogenic avian influenza H5N1 virus quickly. In this study, we fabricated indium-tin-oxide thin-film transistors (ITO TFTs) on a glass substrate for the detecting of AI H5N1. The ITO TFT is fabricated by a one-shadow-mask process in which a channel layer can be simultaneously self-assembled between ITO source/drain electrodes during magnetron sputtering deposition. Monoclonal anti-H5N1 antibodies specific for AI H5N1 virus were covalently immobilized on the ITO channel by (3-glycidoxypropyl)trimethoxysilane. The introduction of target AI H5N1 virus affected the electronic properties of the ITO TFT, which caused a change in the resultant threshold voltage (V T ) and field-effect mobility. The changes of I D –V G curves were consistent with an n-type field effect transistor behavior affected by nearby negatively charged AI H5N1 viruses. The transistor based sensor demonstrated high selectivity and stability for AI H5N1 virus sensing. The sensor showed linear response to AI H5N1 in the concentrations range from 5 × 10 −9 g mL −1 to 5 × 10 −6 g mL −1 with a detection limit of 0.8 × 10 −10 g mL −1 . Moreover, the ITO TFT biosensors can be repeatedly used through the washing processes. With its excellent electric properties and the potential for mass commercial production, ITO TFTs can be promising candidates for the

  19. Indium-tin-oxide thin film transistor biosensors for label-free detection of avian influenza virus H5N1

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Di; Zhuo, Ming [Key Laboratory for Micro-Nano Optoelectronic Devices of Ministry of Education, State Key Laboratory for Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082 (China); Zhang, Xiaoai [State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing (China); Xu, Cheng; Jiang, Jie [Key Laboratory for Micro-Nano Optoelectronic Devices of Ministry of Education, State Key Laboratory for Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082 (China); Gao, Fu [State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing (China); Wan, Qing, E-mail: wanqing7686@hotmail.com [Key Laboratory for Micro-Nano Optoelectronic Devices of Ministry of Education, State Key Laboratory for Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082 (China); Li, Qiuhong, E-mail: liqiuhong2004@hotmail.com [Key Laboratory for Micro-Nano Optoelectronic Devices of Ministry of Education, State Key Laboratory for Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082 (China); Wang, Taihong, E-mail: thwang@hnu.cn [Key Laboratory for Micro-Nano Optoelectronic Devices of Ministry of Education, State Key Laboratory for Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082 (China)

    2013-04-22

    Highlights: ► A highly selective label-free biosensor is established based on indium-tin-oxide thin-film transistors (ITO TFTs). ► AI H5N1 virus was successfully detected through shift in threshold voltage and field-effect mobility of ITO TFT. ► The ITO TFT is applied in biosensor for the first time and shows good reusability and stability. ► Fabrication of the platform is simple with low cost, which is suitable for mass commercial production. -- Abstract: As continuous outbreak of avian influenza (AI) has become a threat to human health, economic development and social stability, it is urgently necessary to detect the highly pathogenic avian influenza H5N1 virus quickly. In this study, we fabricated indium-tin-oxide thin-film transistors (ITO TFTs) on a glass substrate for the detecting of AI H5N1. The ITO TFT is fabricated by a one-shadow-mask process in which a channel layer can be simultaneously self-assembled between ITO source/drain electrodes during magnetron sputtering deposition. Monoclonal anti-H5N1 antibodies specific for AI H5N1 virus were covalently immobilized on the ITO channel by (3-glycidoxypropyl)trimethoxysilane. The introduction of target AI H5N1 virus affected the electronic properties of the ITO TFT, which caused a change in the resultant threshold voltage (V{sub T}) and field-effect mobility. The changes of I{sub D}–V{sub G} curves were consistent with an n-type field effect transistor behavior affected by nearby negatively charged AI H5N1 viruses. The transistor based sensor demonstrated high selectivity and stability for AI H5N1 virus sensing. The sensor showed linear response to AI H5N1 in the concentrations range from 5 × 10{sup −9} g mL{sup −1} to 5 × 10{sup −6} g mL{sup −1} with a detection limit of 0.8 × 10{sup −10} g mL{sup −1}. Moreover, the ITO TFT biosensors can be repeatedly used through the washing processes. With its excellent electric properties and the potential for mass commercial production, ITO TFTs

  20. Highly Pathogenic Avian Influenza A(H5N1) Viruses at the Animal-Human Interface in Vietnam, 2003-2010.

    Science.gov (United States)

    Creanga, Adrian; Hang, Nguyen Le Khanh; Cuong, Vuong Duc; Nguyen, Ha T; Phuong, Hoang Vu Mai; Thanh, Le Thi; Thach, Nguyen Co; Hien, Pham Thi; Tung, Nguyen; Jang, Yunho; Balish, Amanda; Dang, Nguyen Hoang; Duong, Mai Thuy; Huong, Ngo Thu; Hoa, Do Ngoc; Tho, Nguyen Dang; Klimov, Alexander; Kapella, Bryan K; Gubareva, Larisa; Kile, James C; Hien, Nguyen Tran; Mai, Le Quynh; Davis, C Todd

    2017-09-15

    Mutation and reassortment of highly pathogenic avian influenza A(H5N1) viruses at the animal-human interface remain a major concern for emergence of viruses with pandemic potential. To understand the relationship of H5N1 viruses circulating in poultry and those isolated from humans, comprehensive phylogenetic and molecular analyses of viruses collected from both hosts in Vietnam between 2003 and 2010 were performed. We examined the temporal and spatial distribution of human cases relative to H5N1 poultry outbreaks and characterized the genetic lineages and amino acid substitutions in each gene segment identified in humans relative to closely related viruses from avian hosts. Six hemagglutinin clades and 8 genotypes were identified in humans, all of which were initially identified in poultry. Several amino acid mutations throughout the genomes of viruses isolated from humans were identified, indicating the potential for poultry viruses infecting humans to rapidly acquire molecular markers associated with mammalian adaptation and antiviral resistance. Published by Oxford University Press for the Infectious Diseases Society of America 2017. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  1. DNA microarray-based solid-phase RT-PCR for rapid detection and identification of influenza virus type A and subtypes H5 and H7

    DEFF Research Database (Denmark)

    Yi, Sun; Dhumpa, Raghuram; Bang, Dang Duong

    2011-01-01

    of RNA extract in the liquid phase with sequence-specific nested PCR on the solid phase. A simple ultraviolet cross-linking method was used to immobilize the DNA probes over an unmodified glass surface, which makes solid-phase PCR a convenient possibility for AIV screening. The testing of 33 avian fecal....... In this article, a DNA microarray-based solid-phase polymerase chain reaction (PCR) approach has been developed for rapid detection of influenza virus type A and for simultaneous identification of pathogenic virus subtypes H5 and H7. This solid-phase RT-PCR method combined reverse-transcription amplification...

  2. Study on Efficacy of Gamma Radiation on the Inactivation of Highly Pathogenic Avian Influenza Virus H5N1 (Thai isolate) in Chicken Meat and Chicken Feces

    International Nuclear Information System (INIS)

    Pinyochon, Wasana; Piadang, Nattayana; Mulika, Ladda; Parchariyanon, Sujira; Vitittheeranon, Arag; Damrongwatapokin, Sudarat

    2006-09-01

    A study on the efficacy of gamma radiation on the inactivation of a highly pathogenic avian influenza virus H5N1 subtype, Thai isolate was carried out. The virus was in the form frozen infected allantoic fluid frozen chicken meat and frozen chicken feces. The result indicated that 9 kilo grey of gamma radiation could completely inactivated 106.0 EID50/ml of AIV infected allantoic fluid and 22 kiel grey and 15 kilo grey of gamma radiation completely inactivate 106.0 EID50/10/ grams of chicken meat and 106.0 EID50/5 grams of chicken feces respectively.

  3. A molecular and antigenic survey of H5N1 highly pathogenic avian influenza virus isolates from smallholder duck farms in Central Java, Indonesia during 2007-2008

    Directory of Open Access Journals (Sweden)

    Junaidi Akhmad

    2011-09-01

    Full Text Available Abstract Background Indonesia is one of the countries most severely affected by H5N1 highly pathogenic avian influenza (HPAI virus in terms of poultry and human health. However, there is little information on the diversity of H5N1 viruses circulating in backyard farms, where chickens and ducks often intermingle. In this study, H5N1 virus infection occurring in 96 smallholder duck farms in central Java, Indonesia from 2007-2008 was investigated and the molecular and antigenic characteristics of H5N1 viruses isolated from these farms were analysed. Results All 84 characterised viruses belonged to H5N1 clade 2.1 with three virus sublineages being identified: clade 2.1.1 (1, clade 2.1.3 (80, and IDN/6/05-like viruses (3 that did not belong to any of the present clades. All three clades were found in ducks, while only clade 2.1.3 was isolated from chickens. There were no significant amino acid mutations of the hemagglutinin (HA and neuraminidase (NA sites of the viruses, including the receptor binding, glycosylation, antigenic and catalytic sites and NA inhibitor targets. All the viruses had polybasic amino acids at the HA cleavage site. No evidence of major antigenic variants was detected. Based on the HA gene, identical virus variants could be found on different farms across the study sites and multiple genetic variants could be isolated from HPAI outbreaks simultaneously or at different time points from single farms. HPAI virus was isolated from both ducks and chickens; however, the proportion of surviving duck cases was considerably higher than in chickens. Conclusions The 2.1.3 clade was the most common lineage found in this study. All the viruses had sequence characteristic of HPAI, but negligible variations in other recognized amino acids at the HA and NA proteins which determine virus phenotypes. Multiple genetic variants appeared to be circulating simultaneously within poultry communities. The high proportion of live duck cases compared to

  4. A molecular and antigenic survey of H5N1 highly pathogenic avian influenza virus isolates from smallholder duck farms in Central Java, Indonesia during 2007-2008.

    Science.gov (United States)

    Wibawa, Hendra; Henning, Joerg; Wong, Frank; Selleck, Paul; Junaidi, Akhmad; Bingham, John; Daniels, Peter; Meers, Joanne

    2011-09-07

    Indonesia is one of the countries most severely affected by H5N1 highly pathogenic avian influenza (HPAI) virus in terms of poultry and human health. However, there is little information on the diversity of H5N1 viruses circulating in backyard farms, where chickens and ducks often intermingle. In this study, H5N1 virus infection occurring in 96 smallholder duck farms in central Java, Indonesia from 2007-2008 was investigated and the molecular and antigenic characteristics of H5N1 viruses isolated from these farms were analysed. All 84 characterised viruses belonged to H5N1 clade 2.1 with three virus sublineages being identified: clade 2.1.1 (1), clade 2.1.3 (80), and IDN/6/05-like viruses (3) that did not belong to any of the present clades. All three clades were found in ducks, while only clade 2.1.3 was isolated from chickens. There were no significant amino acid mutations of the hemagglutinin (HA) and neuraminidase (NA) sites of the viruses, including the receptor binding, glycosylation, antigenic and catalytic sites and NA inhibitor targets. All the viruses had polybasic amino acids at the HA cleavage site. No evidence of major antigenic variants was detected. Based on the HA gene, identical virus variants could be found on different farms across the study sites and multiple genetic variants could be isolated from HPAI outbreaks simultaneously or at different time points from single farms. HPAI virus was isolated from both ducks and chickens; however, the proportion of surviving duck cases was considerably higher than in chickens. The 2.1.3 clade was the most common lineage found in this study. All the viruses had sequence characteristic of HPAI, but negligible variations in other recognized amino acids at the HA and NA proteins which determine virus phenotypes. Multiple genetic variants appeared to be circulating simultaneously within poultry communities. The high proportion of live duck cases compared to chickens over the study period suggests that ducks are

  5. Potential Biological and Climatic Factors That Influence the Incidence and Persistence of Highly Pathogenic H5N1 Avian Influenza Virus in Egypt

    Directory of Open Access Journals (Sweden)

    Ahmed H. Salaheldin

    2018-03-01

    Full Text Available Highly pathogenic H5N1 avian influenza virus (A/H5N1 of clade 2.2.1 is endemic in poultry in Egypt where the highest number of human infections worldwide was reported. During the last 12 years the Egyptian A/H5N1 evolved into several genotypes. In 2007-2014 vaccinated poultry suffered from antigenic drift variants of clade 2.2.1.1 and in 2014/2015 an unprecedented upsurge of A/H5N1 clade 2.2.1.2 occurred in poultry and humans. Factors contributing to the endemicity or re-emergence of A/H5N1 in poultry in Egypt remain unclear. Here, three potential factors were studied: climatic factors (temperature, relative humidity, and wind speed, biological fitness in vitro, and pathogenicity in domestic Pekin and Muscovy ducks. Statistical analyses using negative binomial regression models indicated that ambient temperature in winter months influenced the spread of A/H5N1 in different geographic areas analyzed in this study. In vitro, at 4 and 56°C 2.2.1.1 and recent 2.2.1.2 viruses were more stable than other viruses used in this study. Further, Pekin ducks were more resistant than Muscovy ducks and the viruses were excreted for up to 2 weeks post-infection assuming a strong role as a reservoir. Taken together, ambient temperature in winter months potentially contributes to increasing outbreaks in some regions in Egypt. Heat stability of clade 2.2.1.1 and recent 2.2.1.2 viruses probably favors their persistence at elevated temperatures. Importantly, asymptomatically infected Pekin ducks may play an important role in the spread of avian and human-like A/H5N1 in Egypt. Therefore, control measures including targeted surveillance and culling of silently infected Pekin ducks should be considered.

  6. Potential Biological and Climatic Factors That Influence the Incidence and Persistence of Highly Pathogenic H5N1 Avian Influenza Virus in Egypt

    Science.gov (United States)

    Salaheldin, Ahmed H.; Kasbohm, Elisa; El-Naggar, Heba; Ulrich, Reiner; Scheibner, David; Gischke, Marcel; Hassan, Mohamed K.; Arafa, Abdel-Satar A.; Hassan, Wafaa M.; Abd El-Hamid, Hatem S.; Hafez, Hafez M.; Veits, Jutta; Mettenleiter, Thomas C.; Abdelwhab, Elsayed M.

    2018-01-01

    Highly pathogenic H5N1 avian influenza virus (A/H5N1) of clade 2.2.1 is endemic in poultry in Egypt where the highest number of human infections worldwide was reported. During the last 12 years the Egyptian A/H5N1 evolved into several genotypes. In 2007-2014 vaccinated poultry suffered from antigenic drift variants of clade 2.2.1.1 and in 2014/2015 an unprecedented upsurge of A/H5N1 clade 2.2.1.2 occurred in poultry and humans. Factors contributing to the endemicity or re-emergence of A/H5N1 in poultry in Egypt remain unclear. Here, three potential factors were studied: climatic factors (temperature, relative humidity, and wind speed), biological fitness in vitro, and pathogenicity in domestic Pekin and Muscovy ducks. Statistical analyses using negative binomial regression models indicated that ambient temperature in winter months influenced the spread of A/H5N1 in different geographic areas analyzed in this study. In vitro, at 4 and 56°C 2.2.1.1 and recent 2.2.1.2 viruses were more stable than other viruses used in this study. Further, Pekin ducks were more resistant than Muscovy ducks and the viruses were excreted for up to 2 weeks post-infection assuming a strong role as a reservoir. Taken together, ambient temperature in winter months potentially contributes to increasing outbreaks in some regions in Egypt. Heat stability of clade 2.2.1.1 and recent 2.2.1.2 viruses probably favors their persistence at elevated temperatures. Importantly, asymptomatically infected Pekin ducks may play an important role in the spread of avian and human-like A/H5N1 in Egypt. Therefore, control measures including targeted surveillance and culling of silently infected Pekin ducks should be considered. PMID:29636730

  7. Host-specific exposure and fatal neurologic disease in wild raptors from highly pathogenic avian influenza virus H5N1 during the 2006 outbreak in Germany.

    Science.gov (United States)

    van den Brand, Judith Ma; Krone, Oliver; Wolf, Peter U; van de Bildt, Marco W G; van Amerongen, Geert; Osterhaus, Albert D M E; Kuiken, Thijs

    2015-03-05

    Raptors may contract highly pathogenic avian influenza virus H5N1 by hunting or scavenging infected prey. However, natural H5N1 infection in raptors is rarely reported. Therefore, we tested raptors found dead during an H5N1 outbreak in wild waterbirds in Mecklenburg-Western Pomerania, Germany, in 2006 for H5N1-associated disease. We tested 624 raptors of nine species-common buzzard (385), Eurasian sparrowhawk (111), common kestrel (38), undetermined species of buzzard (36), white-tailed sea eagle (19), undetermined species of raptor (12), northern goshawk (10), peregrine falcon (6), red kite (3), rough-legged buzzard (3), and western marsh-harrier (1)-for H5N1 infection in tracheal or combined tracheal/cloacal swabs of all birds, and on major tissues of all white-tailed sea eagles. H5N1 infection was detected in two species: common buzzard (12 positive, 3.1%) and peregrine falcon (2 positive, 33.3%). In all necropsied birds (both peregrine falcons and the six freshest common buzzards), H5N1 was found most consistently and at the highest concentration in the brain, and the main H5N1-associated lesion was marked non-suppurative encephalitis. Other H5N1-associated lesions occurred in air sac, lung, oviduct, heart, pancreas, coelomic ganglion, and adrenal gland. Our results show that the main cause of death in H5N1-positive raptors was encephalitis. Our results imply that H5N1 outbreaks in wild waterbirds are more likely to lead to exposure to and mortality from H5N1 in raptors that hunt or scavenge medium-sized birds, such as common buzzards and peregrine falcons, than in raptors that hunt small birds and do not scavenge, such as Eurasian sparrowhawks and common kestrels.

  8. Suboptimal protection against H5N1 highly pathogenic avian influenza viruses from Vietnam in ducks vaccinated with commercial poultry vaccines.

    Science.gov (United States)

    Cha, Ra Mi; Smith, Diane; Shepherd, Eric; Davis, C Todd; Donis, Ruben; Nguyen, Tung; Nguyen, Hoang Dang; Do, Hoa Thi; Inui, Ken; Suarez, David L; Swayne, David E; Pantin-Jackwood, Mary

    2013-10-09

    Domestic ducks are the second most abundant poultry species in many Asian countries including Vietnam, and play a critical role in the epizootiology of H5N1 highly pathogenic avian influenza (HPAI) [FAO]. In this study, we examined the protective efficacy in ducks of two commercial H5N1 vaccines widely used in Vietnam; Re-1 containing A/goose/Guangdong/1/1996 hemagglutinin (HA) clade 0 antigens, and Re-5 containing A/duck/Anhui/1/2006 HA clade 2.3.4 antigens. Ducks received two doses of either vaccine at 7 and at 14 or 21 days of age followed by challenge at 30 days of age with viruses belonging to the HA clades 1.1, 2.3.4.3, 2.3.2.1.A and 2.3.2.1.B isolated between 2008 and 2011 in Vietnam. Ducks vaccinated with the Re-1 vaccine were protected after infection with the two H5N1 HPAI viruses isolated in 2008 (HA clades 1.1 and 2.3.4.3) showing no mortality and limited virus shedding. The Re-1 and Re-5 vaccines conferred 90-100% protection against mortality after challenge with the 2010 H5N1 HPAI viruses (HA clade 2.3.2.1.A); but vaccinated ducks shed virus for more than 7 days after challenge. Similarly, the Re-1 and Re-5 vaccines only showed partial protection against the 2011 H5N1 HPAI viruses (HA clade 2.3.2.1.A and 2.3.2.1.B), with a high proportion of vaccinated ducks shedding virus for more than 10 days. Furthermore, 50% mortality was observed in ducks vaccinated with Re-1 and challenged with the 2.3.2.1.B virus. The HA proteins of the 2011 challenge viruses had the greatest number of amino acid differences from the two vaccines as compared to the viruses from 2008 and 2009, which correlates with the lesser protection observed with these viruses. These studies demonstrate the suboptimal protection conferred by the Re-1 and Re-5 commercial vaccines in ducks against H5N1 HPAI clade 2.3.2.1 viruses, and underscore the importance of monitoring vaccine efficacy in the control of H5N1 HPAI in ducks. Published by Elsevier Ltd.

  9. Characterisation of a highly pathogenic H5N1 clade 2.3.2 influenza virus isolated from swans in Shanghai, China.

    Science.gov (United States)

    Zhao, Guo; Zhong, Lei; Lu, Xinlun; Hu, Jiao; Gu, Xiaobing; Kai, Yan; Song, Qingqing; Sun, Qing; Liu, Jinbao; Peng, Daxin; Wang, Xiaoquan; Liu, Xiaowen; Liu, Xiufan

    2012-02-01

    In spring 2009, one strain of H5N1 clade 2.3.2 virus was isolated from wild swans in Shanghai, indicating the importance of the wild swan in the ecology of this highly pathogenic avian influenza virus (HPAIV) in Eastern China. Pathogenicity experiments conducted in this study indicated that the virus was highly pathogenic for chickens but lowly pathogenic for mammalian hosts, as evidenced by reduced infection of mice. The analysis of complete genome sequences and genetic evolution showed that A/Swan/Shanghai/10/09 (SW/SH/09) may be derived from the strain A/silky chicken/Shantou/475/2004 (CK/ST/04), which is homologous to the influenza viruses isolated from chicken, duck, pika, little egret, swan, mandarin duck and bar-headed goose in China Hunan, China Qinghai, Mongolia, Russia, Japan, Korea, Laos and Hong Kong during 2007-2011, indicating that the virus has retro-infected diverse wild birds from chicken, and significant spread of the virus is still ongoing through overlapping migratory flyways. On the basis of the molecular analysis, we also found that there was a deletion of the glycosylation site (NSS) in amino acid 156 of the hemagglutinin (HA) protein when compared with that of the other Clade 2.3.2 viruses isolated between 2007 and 2011. More importantly, the sequence analysis of SW/SH/09 virus displayed the drug-resistant mutations on the matrix protein (M2) and neuraminidase (NA) genes.

  10. Structure-activity relationships of 3-O-β-chacotriosyl oleanic acid derivatives as entry inhibitors for highly pathogenic H5N1 influenza virus.

    Science.gov (United States)

    Li, Sumei; Jia, Xiuhua; Shen, Xintian; Wei, Zhuwen; Jiang, Zhiyan; Liao, Yixian; Guo, Yiming; Zheng, Xiaojun; Zhong, Guohua; Song, Gaopeng

    2017-08-15

    Highly pathogenic H5N1 virus (H5N1) entry is a key target for the development of novel anti-influenza agents with new mechanisms of action. In our continuing efforts to identify novel potential anti-H5N1 entry inhibitors, a series of 3-O-β-chacotriosyl oleanolic acid analogs have been designed, synthesized and evaluated as H5N1 entry inhibitors based on two small molecule inhibitors 1 and 2 previously discovered by us. The anti-H5N1 entry activities were determined based on HA/HIV and VSVG/HIV entry assays. Compound 15 displayed the most promising anti-H5N1 entry activities with average IC 50 values of 4.05μM and good selective index (22.9). Detailed structure-activity relationships (SARs) studies suggested that either the introduction of an additional oxo group to position 11 at OA or alteration of the C-3 configuration of OA from 3β- to 3α-forms can significantly enhance the selective index while maintaining their antiviral activities in vitro. Molecular simulation analysis confirmed that the compounds exert their inhibitory activity through binding tightly to hemagglutinin (HA2) protein near the fusion peptide and prevent virus entry. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. The effect of age on the pathogenesis of a highly pathogenic avian influenza (HPAI) H5N1 virus in Pekin ducks (Anas platyrhynchos) infected experimentally.

    Science.gov (United States)

    Löndt, Brandon Z; Núñez, Alejandro; Banks, Jill; Alexander, Dennis J; Russell, Christine; Richard-Löndt, Angela C; Brown, Ian H

    2010-01-01

    Highly pathogenic avian influenza (HPAI) H5N1 viruses have recently displayed increased virulence for wild waterfowl. To study the effect of host age on the shedding and tissue dissemination of a HPAI H5N1 virus in infected Pekin ducks. Pekin ducks in two age-matched groups (n = 18), 8 and 12 weeks old (wo) were each infected with 10(6) EID(50)/0.1 ml of HPAI A/turkey/Turkey/1/05 (H5N1, clade 2.2). Each day for 5 days, birds were monitored clinically, and cloacal and oropharyngeal swabs collected, before three birds from each group were selected randomly for post-mortem examination. Tissue samples were collected for examination by real-time RT-PCR, histopathology and immunohistochemistry (IHC). Severe clinical signs, including incoordination and torticollis were observed in the 8 wo group resulting in 100% mortality by 4 dpi. Mild clinical signs were observed in the 12 wo group with no mortality. Real-time RT-PCR and IHC results demonstrated the systemic spread of H5N1 virus in birds of both age groups. Higher levels of virus shedding were detected in oropharyngeal swabs than in cloacal swabs, with similar levels of shedding detected in both age groups. Variations in level and temporal dissemination of virus within tissues of older ducks, and the presence of the virus in brain and heart were observed, which coincided with the appearance of clinical signs preceding death in younger birds. These results are consistent with reports of natural infections of wild waterfowl and poultry possibly indicating an age-related association with dissemination and clinical outcome in ducks following infection with H5N1 HPAI virus.

  12. Detection of antibody responses by using haemagglutination inhibiton test and the protection titer of avian influenza virus H5N1 subtype

    Directory of Open Access Journals (Sweden)

    Risa Indriani

    2004-10-01

    Full Text Available Study on the detection of antibody responses using haemagglutination inhibition (HI test and the protection titer to Avian influenza (AI virus H5N1 subtype local isolate has been conducted at the Research Institute for Veterinary Science (RIVS. A total number of 50 village chicken (10 chicken served as un-injected controls and 30 quail were injected intramuscularly with inactivated virus of AI H5N1 subtype local isolate. Serum samples were collected 3 weeks after injection and were tested using haemagglutination inhibition tests. The correlation between antibody titer and its protection to AI virus H5N1 local isolate were measured by challenging the birds with AI virus H5N1 local isolate The HI test was then used to determine field serum samples. A total number of 48 village chicken from three (3 Districts (Bekasi, Tangerang and Bogor and 96 quails from two (2 farms in District of Sukabumi which were all vaccinated with commercial AI adjuvant vaccine were sampled. The study revealed that village chicken and quails showed antibody responses after 3 weeks vaccination and that titer of ≥ 3 log 2 was able to protect chicken and quails when they were challenged with local isolate virus. Based on this result, village chicken field samples from Districts of Tangerang, Bekasi and Bogor showed antibody titer which will protect 50, 100 and 85% of the flocks respectively. While quail field samples from Farm I and Farm II in District of Sukabumi showed antibody titer which will protect 60-100% and 0-80% of the flocks respectively. It is concluded that the study has successfully measured antibody titer to AI virus H5N1 subtype which protect village chicken and quails from local isolate virus challenge so that the results will be used to analyze field serum samples after vaccination program to eradicate AI from Indonesia.

  13. Evolutionary genetics of highly pathogenic H5N1 avian influenza viruses isolated from whooper swans in northern Japan in 2008.

    Science.gov (United States)

    Usui, Tatsufumi; Yamaguchi, Tsuyoshi; Ito, Hiroshi; Ozaki, Hiroichi; Murase, Toshiyuki; Ito, Toshihiro

    2009-12-01

    In April and May 2008, highly pathogenic avian influenza viruses subtype H5N1 were isolated from dead or moribund whooper swans in Aomori, Akita and Hokkaido prefectures in northern Japan. To trace the genetic lineage of the isolates, the nucleotide sequences of all eight genes were determined and phylogenetically analyzed. The Japanese strains were nearly identical to chicken viruses isolated in Russia in April 2008 and closely related to viruses isolated from dead wild birds in Hong Kong in 2007-2008. Their HA genes clustered in clade 2.3.2. On the other hand, NA and the other internal genes were closely related to those of clade 2.3.4 viruses (genotype V) whose NP genes originated from an HA clade 2.3.2 virus. In conclusion, the H5N1 viruses isolated in Japan, Russia and Hong Kong were derived from a common ancestor virus belonging to genotype V that was generated from genetic reassortment events between viruses of HA clades 2.3.2 and 2.3.4.

  14. Highly Pathogenic H5N1 Influenza A Virus Strains Provoke Heterogeneous IFN-α/β Responses That Distinctively Affect Viral Propagation in Human Cells

    Science.gov (United States)

    Matthaei, Markus; Budt, Matthias; Wolff, Thorsten

    2013-01-01

    The fatal transmissions of highly pathogenic avian influenza A viruses (IAV) of the H5N1 subtype to humans and high titer replication in the respiratory tract indicate that these pathogens can overcome the bird-to-human species barrier. While type I interferons (IFN-α/β) are well described to contribute to the species barrier of many zoonotic viruses, current data to the role of these antiviral cytokines during human H5N1 IAV infections is limited and contradictory. We hypothesized an important role for the IFN system in limiting productive infection of avian H5N1 strains in human cells. Hence, we examined IFN-α/β gene activation by different avian and human H5N1 isolates, if the IFN-α/β response restricts H5N1 growth and whether the different strains were equally capable to regulate the IFN-α/β system via their IFN-antagonistic NS1 proteins. Two human H5N1 isolates and a seasonal H3N2 strain propagated efficiently in human respiratory cells and induced little IFN-β, whereas three purely avian H5N1 strains were attenuated for replication and provoked higher IFN secretion. Replication of avian viruses was significantly enhanced on interferon-deficient cells, and exogenous IFN potently limited the growth of all strains in human cells. Moreover, IFN-α/β activation by all strains depended on retinoic acid-inducible gene I excluding principal differences in receptor activation between the different viruses. Interestingly, all H5N1 NS1 proteins suppressed IFN-α/β induction comparably well to the NS1 of seasonal IAV. Thus, our study shows that H5N1 strains are heterogeneous in their capacity to activate human cells in an NS1-independent manner. Our findings also suggest that H5N1 viruses need to acquire adaptive changes to circumvent strong IFN-α/β activation in human host cells. Since no single amino acid polymorphism could be associated with a respective high- or low induction phenotype we propose that the necessary adaptations to overcome the human IFN

  15. Highly pathogenic H5N1 influenza A virus strains provoke heterogeneous IFN-α/β responses that distinctively affect viral propagation in human cells.

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    Markus Matthaei

    Full Text Available The fatal transmissions of highly pathogenic avian influenza A viruses (IAV of the H5N1 subtype to humans and high titer replication in the respiratory tract indicate that these pathogens can overcome the bird-to-human species barrier. While type I interferons (IFN-α/β are well described to contribute to the species barrier of many zoonotic viruses, current data to the role of these antiviral cytokines during human H5N1 IAV infections is limited and contradictory. We hypothesized an important role for the IFN system in limiting productive infection of avian H5N1 strains in human cells. Hence, we examined IFN-α/β gene activation by different avian and human H5N1 isolates, if the IFN-α/β response restricts H5N1 growth and whether the different strains were equally capable to regulate the IFN-α/β system via their IFN-antagonistic NS1 proteins. Two human H5N1 isolates and a seasonal H3N2 strain propagated efficiently in human respiratory cells and induced little IFN-β, whereas three purely avian H5N1 strains were attenuated for replication and provoked higher IFN secretion. Replication of avian viruses was significantly enhanced on interferon-deficient cells, and exogenous IFN potently limited the growth of all strains in human cells. Moreover, IFN-α/β activation by all strains depended on retinoic acid-inducible gene I excluding principal differences in receptor activation between the different viruses. Interestingly, all H5N1 NS1 proteins suppressed IFN-α/β induction comparably well to the NS1 of seasonal IAV. Thus, our study shows that H5N1 strains are heterogeneous in their capacity to activate human cells in an NS1-independent manner. Our findings also suggest that H5N1 viruses need to acquire adaptive changes to circumvent strong IFN-α/β activation in human host cells. Since no single amino acid polymorphism could be associated with a respective high- or low induction phenotype we propose that the necessary adaptations to

  16. Characterization of a low pathogenic avian influenza H5N2 virus isolated from a turkey breeder flock in Manitoba, Canada.

    Science.gov (United States)

    Berhane, Y; Joseph, T; Kehler, H; Hisanaga, T; Embury-Hyatt, C; Diederich, S; McGreevy, K Hooper; Handel, K; Cottam-Birt, C; Pasick, J

    2014-03-01

    In November 2010, an outbreak of avian influenza (AI) due to the H5N2 subtype virus occurred in a turkey breeder farm in northern Manitoba, Canada. The only clinical signs observed were depression, decrease in food consumption, and loss of egg production. The hemagglutinin (HA) cleavage (HA(0)) site of the isolated H5N2 virus was PQRETR/GLF, consistent with low pathogenic AI viruses. The intravenous pathogenicity index of this virus was zero. Whole-genome sequencing of two isolates that originated from two different barns was performed, and both isolates had 100% identical protein sequence in PB2, HA, NP, M1, M2, NS1, and NS2. The remaining gene segments (PB1, PA, and NA) had a single amino-acid difference when compared with each other. The nucleotide and protein sequences of eight gene segments from both isolates showed 99 or greater identity with other AI viruses that have been circulating in free-living aquatic birds in Canada and the United States within the last 10 yr. Phylogenetic analysis of the HA and neuraminidase (NA) gene segments showed that these viruses are closely related to other H5 strains that have been isolated from Manitoba and other parts of Canada. Serologic testing of archived serum samples collected from these turkeys a week before the outbreak showed no evidence of AI infection. In addition, other farms that were located within 3 km radius from the infected farm and farms that had epidemiologic connection with the farm also tested negative for the presence of H5N2 AI virus or antibody. This indicates that the virus might have been introduced to the farm from wild aquatic birds only a short time before detection. Results of this study highlight the importance of early detection and the significance of ongoing Canada-wide surveillance of AI in domestic poultry as well as in wild aquatic birds/ducks.

  17. Highly (H5N1 and low (H7N2 pathogenic avian influenza virus infection in falcons via nasochoanal route and ingestion of experimentally infected prey.

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    Kateri Bertran

    Full Text Available An experimental infection with highly pathogenic avian influenza (HPAI and low pathogenic avian influenza (LPAI viruses was carried out on falcons in order to examine the effects of these viruses in terms of pathogenesis, viral distribution in tissues and viral shedding. The distribution pattern of influenza virus receptors was also assessed. Captive-reared gyr-saker (Falco rusticolus x Falco cherrug hybrid falcons were challenged with a HPAI H5N1 virus (A/Great crested grebe/Basque Country/06.03249/2006 or a LPAI H7N2 virus (A/Anas plathyrhynchos/Spain/1877/2009, both via the nasochoanal route and by ingestion of previously infected specific pathogen free chicks. Infected falcons exhibited similar infection dynamics despite the different routes of exposure, demonstrating the effectiveness of in vivo feeding route. H5N1 infected falcons died, or were euthanized, between 5-7 days post-infection (dpi after showing acute severe neurological signs. Presence of viral antigen in several tissues was confirmed by immunohistochemistry and real time RT-PCR (RRT-PCR, which were generally associated with significant microscopical lesions, mostly in the brain. Neither clinical signs, nor histopathological findings were observed in any of the H7N2 LPAI infected falcons, although all of them had seroconverted by 11 dpi. Avian receptors were strongly present in the upper respiratory tract of the falcons, in accordance with the consistent oral viral shedding detected by RRT-PCR in both H5N1 HPAI and H7N2 LPAI infected falcons. The present study demonstrates that gyr-saker hybrid falcons are highly susceptible to H5N1 HPAI virus infection, as previously observed, and that they may play a major role in the spreading of both HPAI and LPAI viruses. For the first time in raptors, natural infection by feeding on infected prey was successfully reproduced. The use of avian prey species in falconry husbandry and wildlife rehabilitation facilities could put valuable birds

  18. Highly (H5N1) and low (H7N2) pathogenic avian influenza virus infection in falcons via nasochoanal route and ingestion of experimentally infected prey.

    Science.gov (United States)

    Bertran, Kateri; Busquets, Núria; Abad, Francesc Xavier; García de la Fuente, Jorge; Solanes, David; Cordón, Iván; Costa, Taiana; Dolz, Roser; Majó, Natàlia

    2012-01-01

    An experimental infection with highly pathogenic avian influenza (HPAI) and low pathogenic avian influenza (LPAI) viruses was carried out on falcons in order to examine the effects of these viruses in terms of pathogenesis, viral distribution in tissues and viral shedding. The distribution pattern of influenza virus receptors was also assessed. Captive-reared gyr-saker (Falco rusticolus x Falco cherrug) hybrid falcons were challenged with a HPAI H5N1 virus (A/Great crested grebe/Basque Country/06.03249/2006) or a LPAI H7N2 virus (A/Anas plathyrhynchos/Spain/1877/2009), both via the nasochoanal route and by ingestion of previously infected specific pathogen free chicks. Infected falcons exhibited similar infection dynamics despite the different routes of exposure, demonstrating the effectiveness of in vivo feeding route. H5N1 infected falcons died, or were euthanized, between 5-7 days post-infection (dpi) after showing acute severe neurological signs. Presence of viral antigen in several tissues was confirmed by immunohistochemistry and real time RT-PCR (RRT-PCR), which were generally associated with significant microscopical lesions, mostly in the brain. Neither clinical signs, nor histopathological findings were observed in any of the H7N2 LPAI infected falcons, although all of them had seroconverted by 11 dpi. Avian receptors were strongly present in the upper respiratory tract of the falcons, in accordance with the consistent oral viral shedding detected by RRT-PCR in both H5N1 HPAI and H7N2 LPAI infected falcons. The present study demonstrates that gyr-saker hybrid falcons are highly susceptible to H5N1 HPAI virus infection, as previously observed, and that they may play a major role in the spreading of both HPAI and LPAI viruses. For the first time in raptors, natural infection by feeding on infected prey was successfully reproduced. The use of avian prey species in falconry husbandry and wildlife rehabilitation facilities could put valuable birds of prey and

  19. Experimental infection of a North American raptor, American Kestrel (Falco sparverius, with highly pathogenic avian influenza virus (H5N1.

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    Jeffrey S Hall

    2009-10-01

    Full Text Available Several species of wild raptors have been found in Eurasia infected with highly pathogenic avian influenza virus (HPAIV subtype H5N1. Should HPAIV (H5N1 reach North America in migratory birds, species of raptors are at risk not only from environmental exposure, but also from consuming infected birds and carcasses. In this study we used American kestrels as a representative species of a North American raptor to examine the effects of HPAIV (H5N1 infection in terms of dose response, viral shedding, pathology, and survival. Our data showed that kestrels are highly susceptible to HPAIV (H5N1. All birds typically died or were euthanized due to severe neurologic disease within 4-5 days of inoculation and shed significant amounts of virus both orally and cloacally, regardless of dose administered. The most consistent microscopic lesions were necrosis in the brain and pancreas. This is the first experimental study of HPAIV infection in a North American raptor and highlights the potential risks to birds of prey if HPAIV (H5N1 is introduced into North America.

  20. Experimental infection of a North American raptor, American kestrel (Falco sparverius), with highly pathogenic avian influenza virus (H5N1)

    Science.gov (United States)

    Hall, Jeffrey S.; Ip, Hon S.; Franson, J.C.; Meteyer, C.; Nashold, Sean W.; Teslaa, Joshua L.; French, J.; Redig, P.; Brand, C.

    2009-01-01

    Several species of wild raptors have been found in Eurasia infected with highly pathogenic avian influenza virus (HPAIV) subtype H5N1. Should HPAIV (H5N1) reach North America in migratory birds, species of raptors are at risk not only from environmental exposure, but also from consuming infected birds and carcasses. In this study we used American kestrels as a representative species of a North American raptor to examine the effects of HPAIV (H5N1) infection in terms of dose response, viral shedding, pathology, and survival. Our data showed that kestrels are highly susceptible to HPAIV (H5N1). All birds typically died or were euthanized due to severe neurologic disease within 4-5 days of inoculation and shed significant amounts of virus both orally and cloacally, regardless of dose administered. The most consistent microscopic lesions were necrosis in the brain and pancreas. This is the first experimental study of HPAIV infection in a North American raptor and highlights the potential risks to birds of prey if HPAIV (H5N1) is introduced into North America.

  1. PB2 mutations D701N and S714R promote adaptation of an influenza H5N1 virus to a mammalian host.

    Science.gov (United States)

    Czudai-Matwich, Volker; Otte, Anna; Matrosovich, Mikhail; Gabriel, Gülsah; Klenk, Hans-Dieter

    2014-08-01

    Mutation D701N in the PB2 protein is known to play a prominent role in the adaptation of avian influenza A viruses to mammalian hosts. In contrast, little is known about the nearby mutations S714I and S714R, which have been observed in some avian influenza viruses highly pathogenic for mammals. We have generated recombinant H5N1 viruses with PB2 displaying the avian signature 701D or the mammalian signature 701N and serine, isoleucine, and arginine at position 714 and compared them for polymerase activity and virus growth in avian and mammalian cells, as well as for pathogenicity in mice. Mutation D701N led to an increase in polymerase activity and replication efficiency in mammalian cells and in mouse pathogenicity, and this increase was significantly enhanced when mutation D701N was combined with mutation S714R. Stimulation by mutation S714I was less distinct. These observations indicate that PB2 mutation S714R, in combination with the mammalian signature at position 701, has the potential to promote the adaptation of an H5N1 virus to a mammalian host. Influenza A/H5N1 viruses are avian pathogens that have pandemic potential, since they are spread over large parts of Asia, Africa, and Europe and are occasionally transmitted to humans. It is therefore of high scientific interest to understand the mechanisms that determine the host specificity and pathogenicity of these viruses. It is well known that the PB2 subunit of the viral polymerase is an important host range determinant and that PB2 mutation D701N plays an important role in virus adaptation to mammalian cells. In the present study, we show that mutation S714R is also involved in adaptation and that it cooperates with D701N in exposing a nuclear localization signal that mediates importin-α binding and entry of PB2 into the nucleus, where virus replication and transcription take place. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  2. PB2 amino acid at position 627 affects replicative efficiency, but not cell tropism, of Hong Kong H5N1 influenza A viruses in mice

    International Nuclear Information System (INIS)

    Shinya, Kyoko; Hamm, Stefan; Hatta, Masato; Ito, Hiroshi; Ito, Toshihiro; Kawaoka, Yoshihiro

    2004-01-01

    A single amino acid substitution, from glutamic acid to lysine at position 627 of the PB2 protein, converts a nonlethal H5N1 influenza A virus isolated from a human to a lethal virus in mice. In contrast to the nonlethal virus, which replicates only in respiratory organs, the lethal isolate replicates in a variety of organs, producing systemic infection. Despite a clear difference in virulence and organ tropism between the two viruses, it remains unknown whether the dissimilarity is a result of differences in cell tropism or the reduced replicative ability of the nonlethal virus in mouse cells in general. To determine how this single amino acid change affects virulence and organ tropism in mice, we investigated the growth kinetics of the two H5N1 viruses both in vitro and in vivo. The identity of the PB2 amino acid at position 627 did not appreciably affect viral replicative efficiency in chicken embryo fibroblasts and a quail cell line; however, viruses with lysine at this position instead of glutamic acid grew better in the different mouse cells tested. When the effect of this substitution was investigated in mice, all of the test viruses showed the same cell tropism, but infection by viruses containing lysine at position 627 spread more rapidly than those viruses containing glutamic acid at this position. Further analysis showed a difference in local immune responses: neutrophil infiltration in lungs infected with viruses containing lysine at position 627 persisted longer than that associated with viruses lacking a glutamic acid substitution. Our data indicate that the amino acid at position 627 of the PB2 protein determines the efficiency of viral replication in mouse (not avian) cells, but not tropism among cells in different mouse organs. The presence of lysine leads to more aggressive viral replication, overwhelming the host's defense mechanisms and resulting in high mortality rates in mice

  3. Spatial modeling of wild bird risk factors to investigate highly pathogenic A(H5N1) avian influenza virus transmission

    Science.gov (United States)

    Prosser, Diann J.; Hungerford, Laura L.; Erwin, R. Michael; Ottinger, Mary Ann; Takekawa, John Y.; Newman, Scott H.; Xiao, Xianming; Ellis, Erie C.

    2016-01-01

    One of the longest-persisting avian influenza viruses in history, highly pathogenic avian influenza virus (HPAIV) A(H5N1), continues to evolve after 18 years, advancing the threat of a global pandemic. Wild waterfowl (family Anatidae), are reported as secondary transmitters of HPAIV, and primary reservoirs for low-pathogenic avian influenza viruses, yet spatial inputs for disease risk modeling for this group have been lacking. Using GIS and Monte Carlo simulations, we developed geospatial indices of waterfowl abundance at 1 and 30 km resolutions and for the breeding and wintering seasons for China, the epicenter of H5N1. Two spatial layers were developed: cumulative waterfowl abundance (WAB), a measure of predicted abundance across species, and cumulative abundance weighted by H5N1 prevalence (WPR), whereby abundance for each species was adjusted based on prevalence values then totaled across species. Spatial patterns of the model output differed between seasons, with higher WAB and WPR in the northern and western regions of China for the breeding season and in the southeast for the wintering season. Uncertainty measures indicated highest error in southeastern China for both WAB and WPR. We also explored the effect of resampling waterfowl layers from 1 km to 30 km resolution for multi-scale risk modeling. Results indicated low average difference (less than 0.16 and 0.01 standard deviations for WAB and WPR, respectively), with greatest differences in the north for the breeding season and southeast for the wintering season. This work provides the first geospatial models of waterfowl abundance available for China. The indices provide important inputs for modeling disease transmission risk at the interface of poultry and wild birds. These models are easily adaptable, have broad utility to both disease and conservation needs, and will be available to the scientific community for advanced modeling applications.

  4. Outbreaks of avian influenza A (H5N2), (H5N8), and (H5N1) among birds--United States, December 2014-January 2015.

    Science.gov (United States)

    Jhung, Michael A; Nelson, Deborah I

    2015-02-06

    During December 15, 2014-January 16, 2015, the U.S. Department of Agriculture received 14 reports of birds infected with Asian-origin, highly pathogenic avian influenza A (HPAI) (H5N2), (H5N8), and (H5N1) viruses. These reports represent the first reported infections with these viruses in U.S. wild or domestic birds. Although these viruses are not known to have caused disease in humans, their appearance in North America might increase the likelihood of human infection in the United States. Human infection with other avian influenza viruses, such as HPAI (H5N1) and (H5N6) viruses and (H7N9) virus, has been associated with severe, sometimes fatal, disease, usually following contact with poultry.

  5. Discordant correlation between serological assays observed when measuring heterosubtypic responses against avian influenza H5 and H7 viruses in unexposed individuals.

    Science.gov (United States)

    Molesti, Eleonora; Ferrara, Francesca; Lapini, Giulia; Montomoli, Emanuele; Temperton, Nigel

    2014-01-01

    The human population is constantly exposed to multiple influenza A subtypes due to zoonotic spillover and rapid viral evolution driven by intrinsic error-prone replication and immunological pressure. In this context, antibody responses directed against the HA protein are of importance since they have been shown to correlate with protective immunity. Serological techniques, detecting these responses, play a critical role for influenza surveillance, vaccine development, and assessment. As the recent human pandemics and avian influenza outbreaks have demonstrated, there is an urgent need to be better prepared to assess the contribution of the antibody response to protection against newly emerged viruses and to evaluate the extent of preexisting heterosubtypic immunity in populations. In this study, 68 serum samples collected from the Italian population between 1992 and 2007 were found to be positive for antibodies against H5N1 as determined by single radial hemolysis (SRH), but most were negative when evaluated using haemagglutination inhibition (HI) and microneutralisation (MN) assays. As a result of these discordant serological findings, the increased sensitivity of lentiviral pseudotypes was exploited in pseudotype-based neutralisation (pp-NT) assays and the results obtained provide further insight into the complex nature of humoral immunity against influenza A viruses.

  6. Discordant Correlation between Serological Assays Observed When Measuring Heterosubtypic Responses against Avian Influenza H5 and H7 Viruses in Unexposed Individuals

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    Eleonora Molesti

    2014-01-01

    Full Text Available The human population is constantly exposed to multiple influenza A subtypes due to zoonotic spillover and rapid viral evolution driven by intrinsic error-prone replication and immunological pressure. In this context, antibody responses directed against the HA protein are of importance since they have been shown to correlate with protective immunity. Serological techniques, detecting these responses, play a critical role for influenza surveillance, vaccine development, and assessment. As the recent human pandemics and avian influenza outbreaks have demonstrated, there is an urgent need to be better prepared to assess the contribution of the antibody response to protection against newly emerged viruses and to evaluate the extent of preexisting heterosubtypic immunity in populations. In this study, 68 serum samples collected from the Italian population between 1992 and 2007 were found to be positive for antibodies against H5N1 as determined by single radial hemolysis (SRH, but most were negative when evaluated using haemagglutination inhibition (HI and microneutralisation (MN assays. As a result of these discordant serological findings, the increased sensitivity of lentiviral pseudotypes was exploited in pseudotype-based neutralisation (pp-NT assays and the results obtained provide further insight into the complex nature of humoral immunity against influenza A viruses.

  7. Overview of incursions of Asian H5N1 subtype highly pathogenic avian influenza virus into Great Britain, 2005-2008.

    Science.gov (United States)

    Alexander, Dennis J; Manvell, Ruth J; Irvine, Richard; Londt, Brandon Z; Cox, Bill; Ceeraz, Vanessa; Banks, Jill; Browna, Ian H

    2010-03-01

    Since 2005 there have been five incursions into Great Britain of highly pathogenic avian influenza (HPAI) viruses of subtype H5N1 related to the ongoing global epizootic. The first incursion occurred in October 2005 in birds held in quarantine after importation from Taiwan. Two incursions related to wild birds: one involved a single dead whooper swan found in March 2006 in the sea off the east coast of Scotland, and the other involved 10 mute swans and a Canada goose found dead over the period extending from late December 2007 to late February 2008 on or close to a swannery on the south coast of England. The other two outbreaks occurred in commercial poultry in January 2007 and November 2007, both in the county of Suffolk. The first of these poultry outbreaks occurred on a large turkey farm, and there was no further spread. The second outbreak occurred on a free-range farm rearing turkeys, ducks, and geese and spread to birds on a second turkey farm that was culled as a dangerous contact. Viruses isolated from these five outbreaks were confirmed to be Asian H5N1 HPAI viruses; the quarantine outbreak was attributed to a clade 2.3 virus and the other four to clade 2.2 viruses. This article describes the outbreaks, their control, and the possible origins of the responsible viruses.

  8. Demographic and clinical predictors of mortality from highly pathogenic avian influenza A (H5N1 virus infection: CART analysis of international cases.

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    Rita B Patel

    Full Text Available Human infections with highly pathogenic avian influenza (HPAI A (H5N1 viruses have occurred in 15 countries, with high mortality to date. Determining risk factors for morbidity and mortality from HPAI H5N1 can inform preventive and therapeutic interventions.We included all cases of human HPAI H5N1 reported in World Health Organization Global Alert and Response updates and those identified through a systematic search of multiple databases (PubMed, Scopus, and Google Scholar, including articles in all languages. We abstracted predefined clinical and demographic predictors and mortality and used bivariate logistic regression analyses to examine the relationship of each candidate predictor with mortality. We developed and pruned a decision tree using nonparametric Classification and Regression Tree methods to create risk strata for mortality.We identified 617 human cases of HPAI H5N1 occurring between December 1997 and April 2013. The median age of subjects was 18 years (interquartile range 6-29 years and 54% were female. HPAI H5N1 case-fatality proportion was 59%. The final decision tree for mortality included age, country, per capita government health expenditure, and delay from symptom onset to hospitalization, with an area under the receiver operator characteristic (ROC curve of 0.81 (95% CI: 0.76-0.86.A model defined by four clinical and demographic predictors successfully estimated the probability of mortality from HPAI H5N1 illness. These parameters highlight the importance of early diagnosis and treatment and may enable early, targeted pharmaceutical therapy and supportive care for symptomatic patients with HPAI H5N1 virus infection.

  9. Vaccine protection of chickens against antigenically diverse H5 highly pathogenic avian influenza isolates with a live HVT vector vaccine expressing the influenza hemagglutinin gene derived from a clade 2.2 avian influenza virus.

    Science.gov (United States)

    Kapczynski, Darrell R; Esaki, Motoyuki; Dorsey, Kristi M; Jiang, Haijun; Jackwood, Mark; Moraes, Mauro; Gardin, Yannick

    2015-02-25

    Vaccination is an important tool in the protection of poultry against avian influenza (AI). For field use, the overwhelming majority of AI vaccines produced are inactivated whole virus formulated into an oil emulsion. However, recombinant vectored vaccines are gaining use for their ability to induce protection against heterologous isolates and ability to overcome maternal antibody interference. In these studies, we compared protection of chickens provided by a turkey herpesvirus (HVT) vector vaccine expressing the hemagglutinin (HA) gene from a clade 2.2 H5N1 strain (A/swan/Hungary/4999/2006) against homologous H5N1 as well as heterologous H5N1 and H5N2 highly pathogenic (HP) AI challenge. The results demonstrated all vaccinated birds were protected from clinical signs of disease and mortality following homologous challenge. In addition, oral and cloacal swabs taken from challenged birds demonstrated that vaccinated birds had lower incidence and titers of viral shedding compared to sham-vaccinated birds. Following heterologous H5N1 or H5N2 HPAI challenge, 80-95% of birds receiving the HVT vector AI vaccine at day of age survived challenge with fewer birds shedding virus after challenge than sham vaccinated birds. In vitro cytotoxicity analysis demonstrated that splenic T lymphocytes from HVT-vector-AI vaccinated chickens recognized MHC-matched target cells infected with H5, as well as H6, H7, or H9 AI virus. Taken together, these studies provide support for the use of HVT vector vaccines expressing HA to protect poultry against multiple lineages of HPAI, and that both humoral and cellular immunity induced by live vaccines likely contributes to protection. Published by Elsevier Ltd.

  10. Stable lentiviral transformation of CHO cells for the expression of the hemagglutinin H5 of avian influenza virus in suspension culture

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    Alaín González Pose

    2014-09-01

    Full Text Available Avian influenza virus H5N1 has caused extensive damage worldwide among poultry and humans. Effective expression systems are needed for the production of viral proteins required for monitoring this devastating disease. The present study deals with the establishment of a stable expression system for the hemagglutinin H5 (HAH5 of avian influenza virus using CHO cells in suspension culture transduced with a recombinant lentiviral vector. The synthetic gene coding the HAH5 protein was inserted in a lentiviral vector with the aim of performing a stable transduction of CHO cells. After the selection of recombinant clones, the one with the highest expression level was adapted to suspension culture and the HAH5 protein was purified by immunoaffinity chromatography from the culture supernatant. There were no significant differences when this protein, purified or direct from the culture supernatant of CHO or SiHa cells, was utilized in an immunologic assay using positive and negative sera as reference. It was also demonstrated that the HAH5 protein in its purified form is able to bind anti-HAH5 antibodies generated with proper and non-proper folded proteins. The results demonstrate that the CHO cell line stably transduced with a lentiviral vector coding the sequence of the HAH5 protein and cultured in suspension can be a suitable expression system to obtain this protein for diagnostic purpose in a consistent and reliable manner.

  11. Geographical and Historical Patterns in the Emergences of Novel Highly Pathogenic Avian Influenza (HPAI H5 and H7 Viruses in Poultry

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    Madhur S. Dhingra

    2018-06-01

    Full Text Available Over the years, the emergence of novel H5 and H7 highly pathogenic avian influenza viruses (HPAI has been taking place through two main mechanisms: first, the conversion of a low pathogenic into a highly pathogenic virus, and second, the reassortment between different genetic segments of low and highly pathogenic viruses already in circulation. We investigated and summarized the literature on emerging HPAI H5 and H7 viruses with the aim of building a spatio-temporal database of all these recorded conversions and reassortments events. We subsequently mapped the spatio-temporal distribution of known emergence events, as well as the species and production systems that they were associated with, the aim being to establish their main characteristics. From 1959 onwards, we identified a total of 39 independent H7 and H5 LPAI to HPAI conversion events. All but two of these events were reported in commercial poultry production systems, and a majority of these events took place in high-income countries. In contrast, a total of 127 reassortments have been reported from 1983 to 2015, which predominantly took place in countries with poultry production systems transitioning from backyard to intensive production systems. Those systems are characterized by several co-circulating viruses, multiple host species, regular contact points in live bird markets, limited biosecurity within value chains, and frequent vaccination campaigns that impose selection pressures for emergence of novel reassortants. We conclude that novel HPAI emergences by these two mechanisms occur in different ecological niches, with different viral, environmental and host associated factors, which has implications in early detection and management and mitigation of the risk of emergence of novel HPAI viruses.

  12. Gene expression profiles in primary duodenal chick cells following transfection with avian influenza virus H5 DNA plasmid encapsulated in silver nanoparticles

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    Jazayeri SD

    2013-02-01

    Full Text Available Seyed Davoud Jazayeri,1 Aini Ideris,1,2 Kamyar Shameli,3 Hassan Moeini,1 Abdul Rahman Omar1,21Institute of Bioscience, 2Faculty of Veterinary Medicine, 3Faculty of Science, Universiti Putra Malaysia, Serdang, Selangor, MalaysiaAbstract: In order to develop a systemically administered safe and effective nonviral gene delivery system against avian influenza virus (AIV that induced cytokine expression, the hemagglutinin (H5 gene of AIV, A/Ck/Malaysia/5858/04 (H5N1 and green fluorescent protein were cloned into a coexpression vector pIRES (pIREGFP-H5 and formulated using green synthesis of silver nanoparticles (AgNPs with poly(ethylene glycol and transfected into primary duodenal cells taken from 18-day-old specific-pathogen-free chick embryos. The AgNPs were prepared using moderated temperature and characterized for particle size, surface charge, ultraviolet-visible spectra, DNA loading, and stability. AgNPs and AgNP-pIREGFP-H5 were prepared in the size range of 13.9 nm and 25 nm with a positive charge of +78 ± 0.6 mV and +40 ± 6.2 mV, respectively. AgNPs with a positive surface charge could encapsulate pIREGFP-H5 efficiently. The ultraviolet-visible spectra for AgNP-pIREGFP-H5 treated with DNase I showed that the AgNPs were able to encapsulate pIREGFP-H5 efficiently. Polymerase chain reaction showed that AgNP-pIREGFP-H5 entered into primary duodenal cells rapidly, as early as one hour after transfection. Green fluorescent protein expression was observed after 36 hours, peaked at 48 hours, and remained stable for up to 60 hours. In addition, green fluorescent protein expression generally increased with increasing DNA concentration and time. Cells were transfected using Lipocurax in vitro transfection reagent as a positive control. A multiplex quantitative mRNA gene expression assay in the transfected primary duodenal cells via the transfection reagent and AgNPs with pIREGFP-H5 revealed expression of interleukin (IL-18, IL-15, and IL-12

  13. Emergence of multiple clade 2.3.2.1 influenza A (H5N1) virus subgroups in Vietnam and detection of novel reassortants.

    Science.gov (United States)

    Creanga, Adrian; Thi Nguyen, Diep; Gerloff, Nancy; Thi Do, Hoa; Balish, Amanda; Dang Nguyen, Hoang; Jang, Yunho; Thi Dam, Vui; Thor, Sharmi; Jones, Joyce; Simpson, Natosha; Shu, Bo; Emery, Shannon; Berman, LaShondra; Nguyen, Ha T; Bryant, Juliet E; Lindstrom, Steve; Klimov, Alexander; Donis, Ruben O; Davis, C Todd; Nguyen, Tung

    2013-09-01

    Phylogenetic analyses of 169 influenza A(H5N1) virus genomes were conducted for samples collected through active surveillance and outbreak responses in Vietnam between September 2010 and September 2012. While clade 1.1 viruses persisted in southern regions, three genetically distinct subgroups of clade 2.3.2.1 were found in northern and central Vietnam. The identification of each subgroup corresponded with detection of novel reassortants, likely due to their overlapping circulation throughout the country. While the previously identified clade 1.1 and A/Hubei/1/2010-like 2.3.2.1 genotypes remained the predominant viruses detected, four viruses were found to be reassortants between A/Hubei/1/2010-like (HA, NA, PB2, PB1, PA, NP) and A/duck/Vietnam/NCVD-885/2010-like (M, NS) viruses and one virus was identified as having A/duck/Vietnam/NCVD-885/2010-like HA, NA, PB1, and NP with A/Hubei/1/2010-like PB2 and PA genes. Additionally, clade 2.3.2.1 A/Hong Kong/6841/2010-like viruses, first detected in mid-2012, were identified as reassortants comprised of A/Hubei/1/2010-like PB2 and PA and A/duck/Vietnam/NCVD-885/2010-like PB1, NP, NA, M, NS genes. Published by Elsevier Inc.

  14. Prevalence of avian influenza virus in wild birds before and after the HPAI H5N8 outbreak in 2014 in South Korea.

    Science.gov (United States)

    Shin, Jeong-Hwa; Woo, Chanjin; Wang, Seung-Jun; Jeong, Jipseol; An, In-Jung; Hwang, Jong-Kyung; Jo, Seong-Deok; Yu, Seung Do; Choi, Kyunghee; Chung, Hyen-Mi; Suh, Jae-Hwa; Kim, Seol-Hee

    2015-07-01

    Since 2003, highly pathogenic avian influenza (HPAI) virus outbreaks have occurred five times in Korea, with four HPAI H5N1 outbreaks and one HPAI H5N8 outbreak. Migratory birds have been suggested to be the first source of HPAI in Korea. Here, we surveyed migratory wild birds for the presence of AI and compared regional AI prevalence in wild birds from September 2012 to April 2014 for birds having migratory pathways in South Korea. Finally, we investigated the prevalence of AI in migratory birds before and after HPAI H5N8 outbreaks. Overall, we captured 1617 migratory wild birds, while 18,817 feces samples and 74 dead birds were collected from major wild bird habitats. A total of 21 HPAI viruses were isolated from dead birds, and 86 low pathogenic AI (LPAI) viruses were isolated from captured birds and from feces samples. Spatiotemporal distribution analysis revealed that AI viruses were spread southward until December, but tended to shift north after January, consistent with the movement of migratory birds in South Korea. Furthermore, we found that LPAI virus prevalences within wild birds were notably higher in 2013-2014 than the previous prevalence during the northward migration season. The data from our study demonstrate the importance of the surveillance of AI in wild birds. Future studies including in-depth genetic analysis in combination with evaluation of the movement and ecology of migratory birds might help us to bridge the gaps in our knowledge and better explain, predict, and ultimately prevent future HPAI outbreaks.

  15. Southward autumn migration of waterfowl facilitates cross-continental transmission of the highly pathogenic avian influenza H5N1 virus

    NARCIS (Netherlands)

    Xu, Yanjie; Gong, Peng; Wielstra, Ben; Si, Yali

    2016-01-01

    The highly pathogenic avian influenza subtype H5N1 (HPAI H5N1) is a worldwide zoonotic infectious disease, threatening humans, poultry and wild birds. The role of wild birds in the spread of HPAI H5N1 has previously been investigated by comparing disease spread patterns with bird migration

  16. Signal Immune Reactions of Macrophages Differentiated from THP-1 Monocytes to Infection with Pandemic H1N1PDM09 Virus and H5N2 and H9N2 Avian Influenza A Virus.

    Science.gov (United States)

    Sokolova, T M; Poloskov, V V; Shuvalov, A N; Rudneva, I A; Timofeeva, T A

    2018-03-01

    In culture of THP-1 cells differentiated into macrophages with PMA (THP-PMA macrophages) infected with influenza viruses of subtypes H1, H5 and H9, we measured the expression of TLR7 and RIG1 receptor genes, sensors of viral RNA and ribonucleoprotein, and the levels of production of inflammatory cytokines IL-1β, TNFα, IL-10, and IFNα. The sensitivity and inflammatory response of THP-PMA macrophages to pandemic influenza A virus H1N1pdm09 and avian influenza H5N2 and H9N2 viruses correlate with the intracellular level of their viral RNA and activation of the RIG1 gene. Abortive infection is accompanied by intensive macrophage secretion of TNFα, IL-1β, and toxic factors inducing cell death. Activity of endosomal TLR7 receptor gene changed insignificantly in 24 h after infection and significantly decreased in 48 and 72 h under the action of H5N2 and H9N2, which correlated with manifestation of the cytopathogenic effect of these viruses. H5N2 and H9N2 avian viruses in THP-PMA macrophages are strong activators of the expression of the gene of the cytoplasmic RIG1 receptor 24 and 48 h after infection, and the pandemic virus H1N1pdm09 is a weak stimulator of RIG1 gene. Avian influenza H5N2 and H9N2 viruses are released by rapid induction of the inflammatory response in macrophages. At the late stages of infection, we observed a minor increase in IL-10 secretion in macrophages and, probably, the polarization of a part of the population in type M2. The studied influenza A viruses are weak inductors of IFN in THP-PMA macrophages. In the culture medium of THP-PMA macrophages infected with H9N2 and H5N2 viruses, MTT test revealed high levels of toxic factors causing the death of Caco-2 cells. In contrast to avian viruses, pandemic virus H1N1pdm09 did not induce production of toxic factors.

  17. Spatial transmission of H5N6 highly pathogenic avian influenza viruses among wild birds in Ibaraki Prefecture, Japan, 2016-2017.

    Science.gov (United States)

    Tsunekuni, Ryota; Yaguchi, Yuji; Kashima, Yuki; Yamashita, Kaoru; Takemae, Nobuhiro; Mine, Junki; Tanikawa, Taichiro; Uchida, Yuko; Saito, Takehiko

    2018-05-01

    From 29 November 2016 to 24 January 2017, sixty-three cases of H5N6 highly pathogenic avian influenza virus (HPAIV) infections were detected in wild birds in Ibaraki Prefecture, Japan. Here, we analyzed the genetic, temporal, and geographic correlations of these 63 HPAIVs to elucidate their dissemination throughout the prefecture. Full-genome sequence analysis of the Ibaraki isolates showed that 7 segments (PB2, PB1, PA, HA, NP, NA, NS) were derived from G1.1.9 strains while the M segment was from G1.1 strains; both groups of strains circulated in south China. Pathological studies revealed severe systemic infection in dead swans (the majority of dead birds and the only species necropsied), thus indicating high susceptibility to H5N6 HPAIVs. Coalescent phylogenetic analysis using the 7 G1.1.9-derived segments enabled detailed analysis of the short-term evolution of these highly homologous HPAIVs. This analysis revealed that the H5N6 HPAIVs isolated from wild birds in Ibaraki Prefecture were divided into 7 groups. Spatial analysis demonstrated that most of the cases concentrated around Senba Lake originated from a single source, and progeny viruses were transmitted to other locations after the infection expanded in mute swans. In contrast, within just a 5-km radius of the area in which cases were concentrated, three different intrusions of H5N6 HPAIVs were evident. Multi-segment analysis of short-term evolution showed that not only was the invading virus spread throughout Ibaraki Prefecture but also that, despite the small size of this region, multiple invasions had occurred during winter 2016-2017.

  18. MDA5 can be exploited as efficacious genetic adjuvant for DNA vaccination against lethal H5N1 influenza virus infection in chickens.

    Directory of Open Access Journals (Sweden)

    Matthias Liniger

    Full Text Available Chickens lack the retinoic acid-inducible gene I (RIG-I and sense avian influenza virus (AIV infections by means of the melanoma differentiation-associated gene 5 product (chMDA5. Plasmid-driven expression of the N-terminal half of chMDA5 containing the caspase activation and recruitment domains [chMDA5(1-483] triggers interferon-β responses in chicken cells. We hypothesized that mimicking virus infection by chMDA5(1-483 expression may enhance vaccine-induced adaptive immunity. In order to test this, the potential genetic adjuvant properties of chMDA5(1-483 were evaluated in vivo in combination with a suboptimal quantity of a plasmid DNA vaccine expressing haemagglutinin (HA of H5N1 AIV. Co-administration of the HA plasmid with plasmid DNA for chMDA5(1-483 expression resulted in approximately 10-fold higher HA-specific antibody responses than injection of the HA plasmid mixed with empty vector DNA as control. Accordingly, compared with HA DNA vaccination alone, the chMDA5(1-483-adjuvanted HA DNA vaccine mediated enhanced protection against a lethal H5N1 challenge infection in chickens, with reduced clinical signs and cloacal virus shedding. These data demonstrate that innate immune activation by expression of signaling domains of RIG-I-like receptors can be exploited to enhance vaccine efficacy.

  19. Co-administration of avian influenza virus H5 plasmid DNA with chicken IL-15 and IL-18 enhanced chickens immune responses.

    Science.gov (United States)

    Lim, Kian-Lam; Jazayeri, Seyed Davoud; Yeap, Swee Keong; Alitheen, Noorjahan Banu Mohamed; Bejo, Mohd Hair; Ideris, Aini; Omar, Abdul Rahman

    2012-08-06

    DNA vaccines offer several advantages over conventional vaccines in the development of effective vaccines against avian influenza virus (AIV). However, one of the limitations of the DNA vaccine in poultry is that it induces poor immune responses. In this study, chicken interleukin (IL) -15 and IL-18 were used as genetic adjuvants to improve the immune responses induced from the H5 DNA vaccination in chickens. The immunogenicity of the recombinant plasmid DNA was analyzed based on the antibody production, T cell responses and cytokine production, following inoculation in 1-day-old (Trial 1) and 14-day-old (Trial 2) specific-pathogen-free chickens. Hence, the purpose of the present study was to explore the role of chicken IL-15 and IL-18 as adjuvants following the vaccination of chickens with the H5 DNA vaccine. The overall HI antibody titer in chickens immunized with pDis/H5 + pDis/IL-15 was higher compared to chickens immunized with pDis/H5 (p chickens exhibited a shorter time to achieve the highest HI titer in comparison to the inoculation of the 1-day-old chickens. The cellular immunity was assessed by the flow cytometry analysis to enumerate CD4+ and CD8 + T cells in the peripheral blood. The chickens inoculated with pDis/H5 + pDis/IL-15 demonstrated the highest increase in CD4+ T cells population relative to the control chickens. However, this study revealed that pDis/H5 + pDis/IL-15 was not significant (P > 0.05) in inducing CD8+ T cells. Meanwhile, with the exception of Trial 1, the flow cytometry results for Trial 2 demonstrated that the pDis/H5 + pDis/IL-18 inoculated group was able to trigger a higher increase in CD4+ T cells than the pDis/H5 group (P 0.05) in modulating CD8+ T cells population in both trials. The pDis/H5 + pDis/IL-15 inoculated group showed the highest IL-15 gene expression in both trials compared to other inoculated groups (P chicken IL-15 and IL-18,with pDis/H5 + pDis/IL-15 being a better vaccine candidate

  20. Characterization of avian influenza H5N1 virosome

    Directory of Open Access Journals (Sweden)

    Chatchai Sarachai

    2014-04-01

    Full Text Available The purpose of this study was to prepare and characterize virosome containing envelope proteins of the avian influenza (H5N1 virus. The virosome was prepared by the solubilization of virus with octaethyleneglycol mono (n-dodecyl ether (C12E8 followed by detergent removal with SM2 Bio-Beads. Biochemical analysis by SDS-PAGE and western blotting, indicated that avian influenza H5N1 virosome had similar characteristics to the parent virus and contained both the hemagglutinin (HA, 60-75 kDa and neuraminidase (NA, 220 kDa protein, with preserved biological activity, such as hemagglutination activity. The virosome structure was analyzed by negative stained transmission electron microscope (TEM demonstrated that the spherical shapes of vesicles with surface glycoprotein spikes were harbored. In conclusion, the biophysical properties of the virosome were similar to the parent virus, and the use of octaethyleneglycol mono (n-dodecyl ether to solubilize viral membrane, followed by removal of detergent using polymer beads adsorption (Bio-Beads SM2 was the preferable method for obtaining avian influenza virosome. The outcome of this study might be useful for further development veterinary virus vaccines.

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

    Science.gov (United States)

    Matsuu, Aya; Kobayashi, Tomoko; Patchimasiri, Tuangthong; Shiina, Takashi; Suzuki, Shingo; Chaichoune, Kridsada; Ratanakorn, Parntep; Hiromoto, Yasuaki; Abe, Haruka; Parchariyanon, Sujira; Saito, Takehiko

    2016-01-01

    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. PMID:27078641

  2. H5N2 Highly Pathogenic Avian Influenza Viruses from the US 2014-2015 outbreak have an unusually long pre-clinical period in turkeys.

    Science.gov (United States)

    Spackman, Erica; Pantin-Jackwood, Mary J; Kapczynski, Darrell R; Swayne, David E; Suarez, David L

    2016-11-22

    From December 2014 through June 2015, the US experienced the most costly highly pathogenic avian influenza (HPAI) outbreak to date. Most cases in commercial poultry were caused by an H5N2 strain which was a reassortant with 5 Eurasian lineage genes, including a clade 2.3.4.4 goose/Guangdong/1996 lineage hemagglutinin, and 3 genes from North American wild waterfowl low pathogenicity avian influenza viruses. The outbreak primarily affected turkeys and table-egg layer type chickens. Three isolates were selected for characterization in turkeys: the US index isolate from December 2014 (A/northern pintail/WA/40964/2014), and two poultry isolates from April 2015 (A/chicken/IA/13388/2015 and A/turkey/MN/12528/2015). Four week old broad-breasted white turkeys were inoculated with one of three doses (10 2 , 10 4 or 10 6 50% egg infectious doses [EID 50 ] per bird) of each of the isolates to evaluate infectious dose and pathogenesis. The mean bird infectious dose of A/northern pintail/WA/40964/2014 and A/turkey/MN/12528/2015 was 10 5 EID 50 per bird, but was 10 3 EID 50 per bird for A/chicken/IA/13388/2015, suggesting the latter had greater adaptation to gallinaceous birds. All three isolates had unusually long mean death time of 5.3-5.9 days post challenge, and the primary clinical signs were severe lethargy and neurological signs which started no more than 24 h before death (the average pre-clinical period was 4 days). Infected turkeys also shed high levels of virus by both the oropharyngeal and cloacal routes. The unusually long mean death times, high levels of virus in feces, and increased adaptation of the later viruses may have contributed to the rapid spread of the virus during the peak of the outbreak.

  3. Expression and immunogenicity of M2e peptide of avian influenza virus H5N1 fused to ricin toxin B chain produced in duckweed plants

    Science.gov (United States)

    Firsov, Aleksey; Tarasenko, Irina; Mitiouchkina, Tatiana; Shaloiko, Lyubov; Kozlov, Oleg; Vinokurov, Leonid; Rasskazova, Ekaterina; Murashev, Arkadii; Vainstein, Alexander; Dolgov, Sergey

    2018-02-01

    The amino acid sequence of the extracellular domain of the virus-encoded M2 matrix protein (peptide M2e) is conserved among all subtypes of influenza A strains, enabling the development of a broad-range vaccine against them. We expressed M2e from avian influenza virus A/chicken/Kurgan/5/2005 (H5N1) in nuclear-transformed duckweed plants for further development of an avian influenza vaccine. The 30-amino acid N-terminal fragment of M2, including M2e (denoted M130), was selected for expression. The M2e DNA sequence fused in-frame to the 3’ end of ricin toxin B chain (RTB) was cloned under control of the CaMV 35S promoter into pBI121. The resulting plasmid was used for duckweed transformation, and 23 independent transgenic duckweed lines were obtained. Asialofetuin-binding ELISA of protein samples from the transgenic plants using polyclonal anti-RTB antibodies confirmed the expression of the RTB–M130 fusion protein in 20 lines. Quantitative ELISA of crude protein extracts from these lines showed RTB–M130 accumulation ranging from 0.25–2.5 µg/g fresh weight (0.0006–0.01% of total soluble protein). Affinity chromatography with immobilized asialofetuin and western blot analysis of protein samples from the transgenic plants showed expression of fusion protein RTB–M130 in the dimeric form with a molecular mass of about 70 kDa. Mice were immunized orally with a preparation of total soluble protein from transgenic plants, receiving four doses of 7 μg duckweed-derived RTB–M130 each, with no additional adjuvant. Specific IgG against M2e was detected in immunized mice, and the endpoint titer of anti-M2e IgG was 1024. It was confirmed that oral immunization with RTB-M2e induces production of specific antibodies against peptide M2e, one of the most conserved antigens of the influenza virus. These results may provide further information for the development of a duckweed-based expression system to produce a broad-range edible vaccine against avian influenza.

  4. Incidence of contamination of live bird markets in Bangladesh with influenza A virus and subtypes H5, H7 and H9.

    Science.gov (United States)

    Biswas, P K; Giasuddin, M; Chowdhury, P; Barua, H; Debnath, N C; Yamage, M

    2018-06-01

    In the absence of robust active surveillance of avian influenza viruses (AIV) affecting poultry in South Asian countries, monitoring of live bird markets (LBMs) can be an alternative. In a longitudinal study of 32 LBM, five environments were sampled as follows: market floor, stall floor, slaughter area, poultry holding cage and water used for meat processing. Samples were taken monthly for 5 months, September 2013-January 2014. Incidence rates (IRs) of LBM contamination with AIV and its subtypes H5, H7 and H9 were assessed. In 10 of the LBM selected, biosecurity measures had been implemented through FAO interventions: the other 22 were non-intervened. Standard procedures were applied to detect AIV and three subtypes in pooled samples (1:5). An LBM was considered positive for AIV or a subtype if at least one of the pooled samples tested positive. The incidence rates of LBM contamination with AIV, H5, H7 and H9 were 0.194 (95% confidence interval (CI) 0.136-0.276), 0.031 (95% CI 0.013-0.075), 0 and 0.175 (95% CI 0.12-0.253) per LBM-month at risk, respectively. The log IR ratio between the FAO-intervened and non-intervened LBM for contamination with AIV was -0.329 (95% CI -1.052 to -0.394, p = .372), 0.598 (95% CI -1.593 to 2.789, p = .593) with subtype H5 and -0.500 (95% CI -1.249 to 0.248, p = .190) with subtype H9, indicating no significant difference. The results obtained suggest that both H5 and H9 were circulating in LBM in Bangladesh in the second half of 2013. The incidence of contamination with H9 was much higher than with H5. © 2017 Blackwell Verlag GmbH.

  5. Avian Influenza A(H5N1) Virus Outbreak Investigation: Application of the FAO-OIE-WHO Four-way Linking Framework in Indonesia.

    Science.gov (United States)

    Setiawaty, V; Dharmayanti, N L P I; Misriyah; Pawestri, H A; Azhar, M; Tallis, G; Schoonman, L; Samaan, G

    2015-08-01

    WHO, FAO and OIE developed a 'four-way linking' framework to enhance the cross-sectoral sharing of epidemiological and virological information in responding to zoonotic disease outbreaks. In Indonesia, outbreak response challenges include completeness of data shared between human and animal health authorities. The four-way linking framework (human health laboratory/epidemiology and animal health laboratory/epidemiology) was applied in the investigation of the 193 rd human case of avian influenza A(H5N1) virus infection. As recommended by the framework, outbreak investigation and risk assessment findings were shared. On 18 June 2013, a hospital in West Java Province reported a suspect H5N1 case in a 2-year-old male. The case was laboratory-confirmed that evening, and the information was immediately shared with the Ministry of Agriculture. The human health epidemiology/laboratory team investigated the outbreak and conducted an initial risk assessment on 19 June. The likelihood of secondary cases was deemed low as none of the case contacts were sick. By 3 July, no secondary cases associated with the outbreak were identified. The animal health epidemiology/laboratory investigation was conducted on 19-25 June and found that a live bird market visited by the case was positive for H5N1 virus. Once both human and market virus isolates were sequenced, a second risk assessment was conducted jointly by the human health and animal health epidemiology/laboratory teams. This assessment concluded that the likelihood of additional human cases associated with this outbreak was low but that future sporadic human infections could not be ruled out because of challenges in controlling H5N1 virus contamination in markets. Findings from the outbreak investigation and risk assessments were shared with stakeholders at both Ministries. The four-way linking framework clarified the type of data to be shared. Both human health and animal health teams made ample data available, and there was

  6. Electrochemical detection of avian influenza virus H5N1 gene sequence using a DNA aptamer immobilized onto a hybrid nanomaterial-modified electrode

    International Nuclear Information System (INIS)

    Liu Xianggang; Cheng Ziqiang; Fan Hai; Ai Shiyun; Han Ruixia

    2011-01-01

    Highlights: → A sensitive electrochemical biosensor for the detection of gene sequence was developed. → The biosensor was assembled by MWNT, polypyrrole nanowires and gold nanoparticles. → The hybrid nanomaterials could provide a porous structure with good properties. → The biosensor has highly selectivity and sensitivity. → The design strategy is expected to have extensive applications in other biosensors - Abstract: A sensitive electrochemical method for the detection of avian influenza virus (AIV) H5N1 gene sequence using a DNA aptamer immobilized onto a hybrid nanomaterial-modified electrode was developed. To enhance the selectivity and sensitivity, the modified electrode was assembled with multi-wall carbon nanotubes (MWNT), polypyrrole nanowires (PPNWs) and gold nanoparticles (GNPs). This electrode offered a porous structure with a large effective surface area, highly electrocatalytic activities and electronic conductivity. Therefore, the amount of DNA aptamer immobilized onto the electrode was increased while the accessibility of the detection target was maintained. The biosensor is based on the hybridization and preferred orientation of a DNA aptamer immobilized onto a modified electrode surface with its target (H5N1 specific sequence) present in solution. It is selective for the H5N1 specific sequence, and the signal of the indicator was approximately linear to log(concentration) of the H5N1 specific sequence from 5.0 x 10 -12 to 1.0 x 10 -9 M (R = 0.9863) with a detection limit of 4.3 x 10 -13 M. These studies showed that the new hybrid nanomaterial (MWNT/PPNWs/GNPs) and the DNA aptamer could be used to fabricate an electrochemical biosensor for gene sequence detection. Furthermore, this design strategy is expected to have extensive applications in other biosensors.

  7. Electrochemical detection of avian influenza virus H5N1 gene sequence using a DNA aptamer immobilized onto a hybrid nanomaterial-modified electrode

    Energy Technology Data Exchange (ETDEWEB)

    Liu Xianggang [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); Cheng Ziqiang, E-mail: czqsd@126.com [College of Animal Science and Technology, Shandong Agricultural University, Taian 271018, Shandong (China); Fan Hai [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); Ai Shiyun, E-mail: ashy@sdau.edu.cn [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); Han Ruixia [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China)

    2011-07-15

    Highlights: > A sensitive electrochemical biosensor for the detection of gene sequence was developed. > The biosensor was assembled by MWNT, polypyrrole nanowires and gold nanoparticles. > The hybrid nanomaterials could provide a porous structure with good properties. > The biosensor has highly selectivity and sensitivity. > The design strategy is expected to have extensive applications in other biosensors - Abstract: A sensitive electrochemical method for the detection of avian influenza virus (AIV) H5N1 gene sequence using a DNA aptamer immobilized onto a hybrid nanomaterial-modified electrode was developed. To enhance the selectivity and sensitivity, the modified electrode was assembled with multi-wall carbon nanotubes (MWNT), polypyrrole nanowires (PPNWs) and gold nanoparticles (GNPs). This electrode offered a porous structure with a large effective surface area, highly electrocatalytic activities and electronic conductivity. Therefore, the amount of DNA aptamer immobilized onto the electrode was increased while the accessibility of the detection target was maintained. The biosensor is based on the hybridization and preferred orientation of a DNA aptamer immobilized onto a modified electrode surface with its target (H5N1 specific sequence) present in solution. It is selective for the H5N1 specific sequence, and the signal of the indicator was approximately linear to log(concentration) of the H5N1 specific sequence from 5.0 x 10{sup -12} to 1.0 x 10{sup -9} M (R = 0.9863) with a detection limit of 4.3 x 10{sup -13} M. These studies showed that the new hybrid nanomaterial (MWNT/PPNWs/GNPs) and the DNA aptamer could be used to fabricate an electrochemical biosensor for gene sequence detection. Furthermore, this design strategy is expected to have extensive applications in other biosensors.

  8. Natural Reassortants of Potentially Zoonotic Avian Influenza Viruses H5N1 and H9N2 from Egypt Display Distinct Pathogenic Phenotypes in Experimentally Infected Chickens and Ferrets.

    Science.gov (United States)

    Naguib, Mahmoud M; Ulrich, Reiner; Kasbohm, Elisa; Eng, Christine L P; Hoffmann, Donata; Grund, Christian; Beer, Martin; Harder, Timm C

    2017-12-01

    The cocirculation of zoonotic highly pathogenic avian influenza virus (HPAIV) of subtype H5N1 and avian influenza virus (AIV) of subtype H9N2 among poultry in Egypt for at least 6 years should render that country a hypothetical hot spot for the emergence of reassortant, phenotypically altered viruses, yet no reassortants have been detected in Egypt. The present investigations proved that reassortants of the Egyptian H5N1 clade 2.2.1.2 virus and H9N2 virus of the G1-B lineage can be generated by coamplification in embryonated chicken eggs. Reassortants were restricted to the H5N1 subtype and acquired between two and all six of the internal segments of the H9N2 virus. Five selected plaque-purified reassortant clones expressed a broad phenotypic spectrum both in vitro and in vivo Two groups of reassortants were characterized to have retarded growth characteristics in vitro compared to the H5N1 parent virus. One clone provoked reduced mortality in inoculated chickens, although the characteristics of a highly pathogenic phenotype were retained. Enhanced zoonotic properties were not predicted for any of these clones, and this prediction was confirmed by ferret inoculation experiments: neither the H5N1 parent virus nor two selected clones induced severe clinical symptoms or were transmitted to sentinel ferrets by contact. While the emergence of reassortants of Egyptian HPAIV of subtype H5N1 with internal gene segments of cocirculating H9N2 viruses is possible in principle, the spread of such viruses is expected to be governed by their fitness to outcompete the parental viruses in the field. The eventual spread of attenuated phenotypes, however, would negatively impact syndrome surveillance on poultry farms and might foster enzootic virus circulation. IMPORTANCE Despite almost 6 years of the continuous cocirculation of highly pathogenic avian influenza virus H5N1 and avian influenza virus H9N2 in poultry in Egypt, no reassortants of the two subtypes have been reported

  9. A study of analysis PB1-F2 protein of Influenza Viruses A/H1N1pdm09, A/ H3N2, and A/H5N1

    Directory of Open Access Journals (Sweden)

    Hana Apsari Pawestri

    2016-07-01

    Full Text Available Abstrak Tujuan. Protein PB1-F2 (polymerase basic 1-frame 2 adalah protein terbaru yang ditemukan pada virus Influenza dan telah terbukti berperan dalam induksi kematian sel dan patogenitas. Tujuan dari tulisan ini adalah untuk menganalisis protein PB1-F2 pada virus Influenza A/H5N1 dan A/H1N1pdm09. Metode. Kami melakukan pencarian data yang relevan yaitu sekuens gen virus Influenza A/H5N1 dan A/H1N1pdm09 dari Gen Bank National Center for Biotechnology Information (NCBI selama tahun 1997-2015. Data yang digunakan adalah data sekuens nukleotida gen PB1 (polymerase basic1 virus influenza A/H5N1 dan A/H1N1pdm09. Kemudian dilakukan analisis alignment untuk mengetahui variasi protein dan mutasi yang berhubungan dengan patogenitas dan virulensi. Hasil. Kami melakukan penelitian terhadap sekuens PB1-F2 sebanyak 3262 influenza A/H5N1 dan 2472 Influenza A/H1N1pdm09. Hasil analisis menunjukkan bahwa semua sekuens A/H5N1 memiliki panjang yang penuh sebanyak 90 asam amino, kecuali influenza pandemi 2009 hanya memiliki panjang 87 asam amino. Kemudian, ditemukan mutasi yang berhubungan dengan virulensi yang ditunjukan dengan perubahan asam amino Asparagin (N menjadi Serin (S. Mutasi tersebut terjadi pada Influenza A/H5N1 sebanyak 8.5% dan Influenza A/H1N1pdm09 sebanyak 0.5%. Kesimpulan. Ditemukan beberapa variasi panjang asam amino dan mutasi penting pada sekuens PB1-F2 dari subtipe yang berbeda yaitu influenza A/H5N1 dan A/H1N1pdm09  yang mengindikasikan seleksi spesifik karena introduksi dan adaptasi terhadap inang yang berbeda. Diperlukan penelitian lanjutan untuk lebih memahami variasi dan kontribusi protein PB1-F2 tersebut terhadap virulensi dan patogenitas virus Influenza. Kata kunci : Patogenesis, Virus Influenza, Protein  PB1-F2 Abstract Aim. Influenza virus PB1-F2 (polymerase basic 1-frame 2 protein is a novel protein previously shown to be involved in cell death induction and pathogenesis. Here we analysis the PB1-F2 protein of Influenza virus A/H

  10. A study of analysis PB1-F2 protein of Influenza Viruses A/H1N1pdm09, A/ H3N2, and A/H5N1

    Directory of Open Access Journals (Sweden)

    Hana Apsari Pawestri

    2016-07-01

    Full Text Available Abstrak Tujuan. Protein PB1-F2 (polymerase basic 1-frame 2 adalah protein terbaru yang ditemukan pada virus Influenza dan telah terbukti berperan dalam induksi kematian sel dan patogenitas. Tujuan dari tulisan ini adalah untuk menganalisis protein PB1-F2 pada virus Influenza A/H5N1 dan A/H1N1pdm09. Metode. Kami melakukan pencarian data yang relevan yaitu sekuens gen virus Influenza A/H5N1 dan A/H1N1pdm09 dari Gen Bank National Center for Biotechnology Information (NCBI selama tahun 1997-2015. Data yang digunakan adalah data sekuens nukleotida gen PB1 (polymerase basic1 virus influenza A/H5N1 dan A/H1N1pdm09. Kemudian dilakukan analisis alignment untuk mengetahui variasi protein dan mutasi yang berhubungan dengan patogenitas dan virulensi. Hasil. Kami melakukan penelitian terhadap sekuens PB1-F2 sebanyak 3262 influenza A/H5N1 dan 2472 Influenza A/H1N1pdm09. Hasil analisis menunjukkan bahwa semua sekuens A/H5N1 memiliki panjang yang penuh sebanyak 90 asam amino, kecuali influenza pandemi 2009 hanya memiliki panjang 87 asam amino. Kemudian, ditemukan mutasi yang berhubungan dengan virulensi yang ditunjukan dengan perubahan asam amino Asparagin (N menjadi Serin (S. Mutasi tersebut terjadi pada Influenza A/H5N1 sebanyak 8.5% dan Influenza A/H1N1pdm09 sebanyak 0.5%. Kesimpulan. Ditemukan beberapa variasi panjang asam amino dan mutasi penting pada sekuens PB1-F2 dari subtipe yang berbeda yaitu influenza A/H5N1 dan A/H1N1pdm09  yang mengindikasikan seleksi spesifik karena introduksi dan adaptasi terhadap inang yang berbeda. Diperlukan penelitian lanjutan untuk lebih memahami variasi dan kontribusi protein PB1-F2 tersebut terhadap virulensi dan patogenitas virus Influenza. Kata kunci : Patogenesis, Virus Influenza, Protein  PB1-F2 Abstract Aim. Influenza virus PB1-F2 (polymerase basic 1-frame 2 protein is a novel protein previously shown to be involved in cell death induction and pathogenesis. Here we analysis the PB1-F2 protein of Influenza virus A/H

  11. Characterization of the influenza A H5N1 viruses of the 2008-09 outbreaks in India reveals a third introduction and possible endemicity.

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    Alok K Chakrabarti

    Full Text Available Widespread infection of highly pathogenic avian influenza A H5N1 was reported from backyard and commercial poultry in West Bengal (WB, an eastern state of India in early 2008. Infection gradually spread to Tripura, Assam and Sikkim, the northeastern states, with 70 outbreaks reported between January 2008 and May 2009. Whole genome sequence analysis of three isolates from WB, one isolate from Tripura along with the analysis of hemagglutinin (HA and neuraminidase (NA genes of 17 other isolates was performed during this study. In the HA gene phylogenetic tree, all the 2008-09 Indian isolates belonged to EMA3 sublineage of clade 2.2. The closest phylogenetic relationship was found to be with the 2007-09 isolates from Bangladesh and not with the earlier 2006 and 2007 Indian isolates implying a third introduction into the country. The receptor-binding pocket of HA1 of two isolates from WB showed S221P mutation, one of the markers predicted to be associated with human receptor specificity. Two substitutions E119A (2 isolates of WB and N294S (2 other isolates of WB known to confer resistance to NA inhibitors were observed in the active site of neuraminidase. Several additional mutations were observed within the 2008-09 Indian isolates indicating genetic diversification. Overall, the study is indicative of a possible endemicity in the eastern and northeastern parts of the country, demanding active surveillance specifically in view of the critical mutations that have been observed in the influenza A H5N1 viruses.

  12. Human Clade 2.3.4.4 A/H5N6 Influenza Virus Lacks Mammalian Adaptation Markers and Does Not Transmit via the Airborne Route between Ferrets.

    Science.gov (United States)

    Herfst, Sander; Mok, Chris K P; van den Brand, Judith M A; van der Vliet, Stefan; Rosu, Miruna E; Spronken, Monique I; Yang, Zifeng; de Meulder, Dennis; Lexmond, Pascal; Bestebroer, Theo M; Peiris, J S Malik; Fouchier, Ron A M; Richard, Mathilde

    2018-01-01

    Since their emergence in 1997, A/H5N1 influenza viruses of the A/goose/Guangdong/1/96 lineage have diversified in multiple genetic and antigenic clades upon continued circulation in poultry in several countries in Eurasia and Africa. Since 2009, reassortant viruses carrying clade 2.3.4.4 hemagglutinin (HA) and internal and neuraminidase (NA) genes of influenza A viruses of different avian origin have been detected, yielding various HA-NA combinations, such as A/H5N1, A/H5N2, A/H5N3, A/H5N5, A/H5N6, and A/H5N8. Previous studies reported on the low pathogenicity and lack of airborne transmission of A/H5N2 and A/H5N8 viruses in the ferret model. However, although A/H5N6 viruses are the only clade 2.3.4.4 viruses that crossed the species barrier and infected humans, the risk they pose for human health remains poorly characterized. Here, the characterization of A/H5N6 A/Guangzhou/39715/2014 virus in vitro and in ferrets is described. This A/H5N6 virus possessed high polymerase activity, mediated by the E627K substitution in the PB2 protein, which corresponds to only one biological trait out of the three that were previously shown to confer airborne transmissibility to A/H5N1 viruses between ferrets. This might explain its lack of airborne transmission between ferrets. After intranasal inoculation, A/H5N6 virus replicated to high titers in the respiratory tracts of ferrets and was excreted for at least 6 days. Moreover, A/H5N6 virus caused severe pneumonia in ferrets upon intratracheal inoculation. Thus, A/H5N6 virus causes a more severe disease in ferrets than previously investigated clade 2.3.4.4 viruses, but our results demonstrate that the risk from airborne spread is currently low. IMPORTANCE Avian influenza A viruses are a threat to human health, as they cross the species barrier and infect humans occasionally, often with severe outcome. The antigenic and genetic diversity of A/H5 viruses from the A/goose/Guangdong/1/96 lineage is increasing, due to continued

  13. Hemagglutinin-based polyanhydride nanovaccines against H5N1 influenza elicit protective virus neutralizing titers and cell-mediated immunity

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    Ross KA

    2014-12-01

    Full Text Available Kathleen A Ross,1 Hyelee Loyd,2 Wuwei Wu,2 Lucas Huntimer,3 Shaheen Ahmed,4 Anthony Sambol,5 Scott Broderick,6 Zachary Flickinger,2 Krishna Rajan,6 Tatiana Bronich,4 Surya Mallapragada,1 Michael J Wannemuehler,3 Susan Carpenter,2 Balaji Narasimhan1 1Chemical and Biological Engineering, Iowa State University, Ames, IA, USA; 2Animal Science, Iowa State University, Ames, IA, USA; 3Veterinary Microbiology and Preventive Medicine, Iowa State University, Ames, IA, USA; 4Pharmaceutical Sciences, University of Nebraska Medical Center, Omaha, NE, USA; 5Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE, USA; 6Materials Science and Engineering, Iowa State University, Ames, IA, USA Abstract: H5N1 avian influenza is a significant global concern with the potential to become the next pandemic threat. Recombinant subunit vaccines are an attractive alternative for pandemic vaccines compared to traditional vaccine technologies. In particular, polyanhydride nanoparticles encapsulating subunit proteins have been shown to enhance humoral and cell-mediated immunity and provide protection upon lethal challenge. In this work, a recombinant H5 hemagglutinin trimer (H53 was produced and encapsulated into polyanhydride nanoparticles. The studies performed indicated that the recombinant H53 antigen was a robust immunogen. Immunizing mice with H53 encapsulated into polyanhydride nanoparticles induced high neutralizing antibody titers and enhanced CD4+ T cell recall responses in mice. Finally, the H53-based polyanhydride nanovaccine induced protective immunity against a low-pathogenic H5N1 viral challenge. Informatics analyses indicated that mice receiving the nanovaccine formulations and subsequently challenged with virus were similar to naïve mice that were not challenged. The current studies provide a basis to further exploit the advantages of polyanhydride nanovaccines in pandemic scenarios. Keywords: polymer, nanoparticle, vaccine, subunit

  14. Probable Tiger-to-Tiger Transmission of Avian Influenza H5N1

    Science.gov (United States)

    Thanawongnuwech, Roongroje; Amonsin, Alongkorn; Tantilertcharoen, Rachod; Damrongwatanapokin, Sudarat; Theamboonlers, Apiradee; Payungporn, Sunchai; Nanthapornphiphat, Kamonchart; Ratanamungklanon, Somchuan; Tunak, Eakchai; Songserm, Thaweesak; Vivatthanavanich, Veravit; Lekdumrongsak, Thawat; Kesdangsakonwut, Sawang; Tunhikorn, Schwann

    2005-01-01

    During the second outbreak of avian influenza H5N1 in Thailand, probable horizontal transmission among tigers was demonstrated in the tiger zoo. Sequencing and phylogenetic analysis of those viruses showed no differences from the first isolate obtained in January 2004. This finding has implications for influenza virus epidemiology and pathogenicity in mammals. PMID:15890122

  15. Efficacy of single dose of a bivalent vaccine containing inactivated Newcastle disease virus and reassortant highly pathogenic avian influenza H5N1 virus against lethal HPAI and NDV infection in chickens.

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    Dong-Hun Lee

    Full Text Available Highly pathogenic avian influenza (HPAI and Newcastle disease (ND are 2 devastating diseases of poultry, which cause great economic losses to the poultry industry. In the present study, we developed a bivalent vaccine containing antigens of inactivated ND and reassortant HPAI H5N1 viruses as a candidate poultry vaccine, and we evaluated its immunogenicity and protective efficacy in specific pathogen-free chickens. The 6:2 reassortant H5N1 vaccine strain containing the surface genes of the A/Chicken/Korea/ES/2003(H5N1 virus was successfully generated by reverse genetics. A polybasic cleavage site of the hemagglutinin segment was replaced by a monobasic cleavage site. We characterized the reverse genetics-derived reassortant HPAI H5N1 clade 2.5 vaccine strain by evaluating its growth kinetics in eggs, minimum effective dose in chickens, and cross-clade immunogenicity against HPAI clade 1 and 2. The bivalent vaccine was prepared by emulsifying inactivated ND (La Sota strain and reassortant HPAI viruses with Montanide ISA 70 adjuvant. A single immunization with this vaccine induced high levels of hemagglutination-inhibiting antibody titers and protected chickens against a lethal challenge with the wild-type HPAI and ND viruses. Our results demonstrate that the bivalent, inactivated vaccine developed in this study is a promising approach for the control of both HPAI H5N1 and ND viral infections.

  16. Spatial and temporal association of outbreaks of H5N1 influenza virus infection in wild birds with the 0 degrees C isotherm.

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    Leslie A Reperant

    2010-04-01

    Full Text Available Wild bird movements and aggregations following spells of cold weather may have resulted in the spread of highly pathogenic avian influenza virus (HPAIV H5N1 in Europe during the winter of 2005-2006. Waterbirds are constrained in winter to areas where bodies of water remain unfrozen in order to feed. On the one hand, waterbirds may choose to winter as close as possible to their breeding grounds in order to conserve energy for subsequent reproduction, and may be displaced by cold fronts. On the other hand, waterbirds may choose to winter in regions where adverse weather conditions are rare, and may be slowed by cold fronts upon their journey back to the breeding grounds, which typically starts before the end of winter. Waterbirds will thus tend to aggregate along cold fronts close to the 0 degrees C isotherm during winter, creating conditions that favour HPAIV H5N1 transmission and spread. We determined that the occurrence of outbreaks of HPAIV H5N1 infection in waterbirds in Europe during the winter of 2005-2006 was associated with temperatures close to 0 degrees C. The analysis suggests a significant spatial and temporal association of outbreaks caused by HPAIV H5N1 in wild birds with maximum surface air temperatures of 0 degrees C-2 degrees C on the day of the outbreaks and the two preceding days. At locations where waterbird census data have been collected since 1990, maximum mallard counts occurred when average and maximum surface air temperatures were 0 degrees C and 3 degrees C, respectively. Overall, the abundance of mallards (Anas platyrhynchos and common pochards (Aythya ferina was highest when surface air temperatures were lower than the mean temperatures of the region investigated. The analysis implies that waterbird movements associated with cold weather, and congregation of waterbirds along the 0 degrees C isotherm likely contributed to the spread and geographical distribution of outbreaks of HPAIV H5N1 infection in wild birds in

  17. Expression of Two N1 Clones with Single Amino Acid Dissimilarity of Avian Influenza H5N1 Virus

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    RISZA HARTAWAN

    2012-12-01

    Full Text Available Two clones of N1 gene derived from isolate A/Dk/Tangerang/Bbalitvet-ACIAR-TE11/2007 (H5N1 exhibit single mismatch of amino acid sequence at position 242 that is threonine and methionine for the clone #3 and #5, respectively. In order to evaluate the effect of the amino acid substitution, these clones were inserted into two different expression vectors that are pEGFP-C1 and pcDNA-3.3 TOPO® TA cloning. Subsequently, the respective recombinant clones were transfected into eukaryotic cells, including CEF, RK13 and VERO using Lipofectamine ‘plus’ reagent. As a result, the clone #3 retaining atypical sequence showed lower expression level rather than the clone #15 in both vectors and all type of cells. The 3D conformational modelling revealed that the mutation occurs in the inner part of glycoprotein embedded within envelope or matrix. Therefore, the missense mutation seems has no effect on the antigenic properties of neuraminidase but this substitution by any means causes lethal mutagenesis in the individual gene expression by reducing level of protein transcript.

  18. Detection and subtyping (H5 and H7) of avian type A influenza virus by reverse transcription-PCR and PCR-ELISA

    DEFF Research Database (Denmark)

    Munch, M.; Nielsen, L.P.; Handberg, Kurt

    2001-01-01

    A. A panel of reference influenza strains from various hosts including avian species, human, swine and horse were evaluated in a one tube RT-PCR using primers designed for the amplification of a 218 bp fragment of the NP gene. The PCR products were detected by PCR-ELISA by use of an internal......Avian influenza virus infections are a major cause of morbidity and rapid identification of the virus has important clinical, economical and epidemiological implications. We have developed a one-tube Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) for the rapid diagnosis of avian influenza...... catching probe confirming the NP influenza A origin. The PCR-ELISA was about 100 times more sensitive than detection of PCR products by agarose gel electrophoresis. RT-PCR and detection by PCR-ELISA is comparable in sensitivity to virus propagation in eggs. We also designed primers for the detection...

  19. The chest X-ray manifestations of children with highly pathogenic H5N1 avian influenza virus infection (a report of 1 final diagnosis case and 1 borderline case)

    International Nuclear Information System (INIS)

    Jin Ke; Chen Hua; Tan Lihua; Yuan Youhong; Xiao Enhua; Luo Ruping; Li Wanging; Xu Heping

    2006-01-01

    Objective: To describe the chest X-ray manifestations of children with highly pathogenic H5N1 avian influenza virus infection. Methods: The pulmonary X-ray findings in 1 patient was confirmed by the World Health Organization infected H5N1 avian influenza vires and 1 borderline patient was retrospectively analyzed. Results: Both sides of lung field showed the cloudy and massive infiltration in chest X-ray film. The lesions of lung distributed extensively and symmetrically. Radiological dynamic changes showed the variation of the lesions of lung was quick in a short time. It had a characteristic of roving around. The lesions of lung appeared fibrosis at the period of the end. Conclusion: There are some radiographic characteristics in children with H5N1 avian influenza vires infection. It will be helpful for its diagnosis when getting familiar with its X-ray manifestations, but the final diagnosis is dependent on the epidemiology history and laboratory results. (authors)

  20. Exploring contacts facilitating transmission of influenza A(H5N1) virus between poultry farms in West Java, Indonesia: A major role for backyard farms?

    Science.gov (United States)

    Wibawa, Hendra; Karo-Karo, Desniwaty; Pribadi, Eko Sugeng; Bouma, Annemarie; Bodewes, Rogier; Vernooij, Hans; Diyantoro; Sugama, Agus; Muljono, David H; Koch, Guus; Tjatur Rasa, Fadjar Sumping; Stegeman, Arjan

    2018-08-01

    Highly pathogenic avian influenza virus (HPAIV) H5N1 has been reported in Asia, including Indonesia since 2003. Although several risk factors related to the HPAIV outbreaks in poultry in Indonesia have been identified, little is known of the contact structure of farms of different poultry production types (backyard chickens, broilers, layers, and ducks). This study aims to quantify the contact rates associated with the movement of people, and movements of live birds and products and equipment that affect the risk of HPAIV H5N1 transmission between poultry farms in Indonesia. On 124 poultry farms in 6 districts in West Java, logbooks were distributed to record the movements of farmers/staff and visitors and their poultry contacts. Most movements in backyard chicken, commercial native chicken, broiler and duck farms were visits to and from other poultry farms, whilst in layer farms visits to and from poultry companies, visits to egg collection houses and visit from other poultry farms were most frequent. Over 75% of persons visiting backyard chicken and duck farms had previously visited other poultry farms on the same day. Visitors of backyard chicken farms had the highest average contact rate, either direct contact with poultry on other farms before the visits (1.35 contact/day) or contact during their visits in the farms (10.03 contact/day). These results suggest that backyard chicken farms are most at risk for transmission of HPAIV compared to farms of the other poultry production types. Since visits of farm-to-farm were high, backyard farms could also a potential source for HPAIV transmission to commercial poultry farms. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Thermal inactivation of H5N2 high-pathogenicity avian influenza virus in dried egg white with 7.5% moisture.

    Science.gov (United States)

    Thomas, Colleen; Swayne, David E

    2009-09-01

    High-pathogenicity avian influenza viruses (HPAIV) cause severe systemic disease with high mortality in chickens. Isolation of HPAIV from the internal contents of chicken eggs has been reported, and this is cause for concern because HPAIV can be spread by movement of poultry products during marketing and trade activity. This study presents thermal inactivation data for the HPAIV strain A/chicken/PA/1370/83 (H5N2) (PA/83) in dried egg white with a moisture content (7.5%) similar to that found in commercially available spray-dried egg white products. The 95% upper confidence limits for D-values calculated from linear regression of the survival curves at 54.4, 60.0, 65.5, and 71.1 degrees C were 475.4, 192.2, 141.0, and 50.1 min, respectively. The line equation y = [0.05494 x degrees C] + 5.5693 (root mean square error = 0.0711) was obtained by linear regression of experimental D-values versus temperature. Conservative predictions based on the thermal inactivation data suggest that standard industry pasteurization protocols would be very effective for HPAIV inactivation in dried egg white. For example, these calculations predict that a 7-log reduction would take only 2.6 days at 54.4 degrees C.

  2. Application and evaluation of RT-PCR-ELISA for the nucleoprotein and RT-PCR for detection of low-pathogenic H5 and H7 subtypes of avian influenza virus

    DEFF Research Database (Denmark)

    Dybkær, Karen; Munch, Mette; Handberg, Kurt J.

    2004-01-01

    Three 1-tube Reverse Transcriptase Polymerase Chain Reactions (RT-PCR) directed against the genes encoding the nucleoprotein (NP) and the H5 and H7 hemagglutinin (HA) gene, respectively, were used for detection of avian influenza virus (AIV) in various specimens. A total of 1,040 samples...... originating from chickens experimentally infected with 2 different low pathogenic avian influenza viruses, from domestic ducks and from wild aquatic birds were examined. The outcome of 1) the universal AIV RT-PCR including a PCR-enzyme-linked immunosorbent assay (ELISA) procedure directed against NP (NP RT...

  3. Gas-permeable ethylene bags for the small scale cultivation of highly pathogenic avian influenza H5N1 and other viruses in embryonated chicken eggs

    Directory of Open Access Journals (Sweden)

    McCurdy Kimberly S

    2010-01-01

    Full Text Available Abstract Background Embryonated chicken eggs (ECE are sometimes used for the primary isolation or passage of influenza viruses, other viruses, and certain bacteria. For small-scale experiments with pathogens that must be studied in biosafety level three (BSL3 facilities, inoculated ECE are sometimes manipulated and maintained in small egg incubators within a biosafety cabinet (BSC. To simplify the clean up and decontamination of an egg incubator in case of egg breakage, we explored whether ethylene breather bags could be used to encase ECE inoculated with pathogens. This concept was tested by determining embryo survival and examining virus yields in bagged ECE. Results Virus yields acceptable for many applications were attained when influenza-, alpha-, flavi-, canine distemper-, and mousepox viruses were propagated in ECE sealed within ethylene breather bags. Conclusions For many small-scale applications, ethylene breather bags can be used to encase ECE inoculated with various viruses.

  4. High rates of detection of Clade 2.3.4.4 Highly Pathogenic Avian Influenza H5 viruses in wild birds in the Pacific Northwest during the winter of 2014-2015

    Science.gov (United States)

    Ip, Hon S.; Dusek, Robert J.; Bodenstein, Barbara L.; Kim Torchetti, Mia; DeBruyn, Paul; Mansfield, Kristin G.; DeLiberto, Thomas; Sleeman, Jonathan M.

    2016-01-01

    In 2014, Clade 2.3.4.4 H5N8 highly pathogenic avian influenza (HPAI) viruses spread across the Republic of Korea and ultimately were reported in China, Japan, Russia and Europe. Mortality associated with a reassortant HPAI H5N2 virus was detected in poultry farms in Western Canada at the end of November. The same strain (with identical genetic structure) was then detected in free-living wild birds that had died prior to December 8 of unrelated causes in Whatcom County, Washington, USA in an area contiguous with the index Canadian location. A gyrfalcon (Falco rusticolus) that had hunted and fed on an American wigeon (Anas americana) on December 6 in the same area and died two days later, tested positive for the Eurasian origin HPAI H5N8. Subsequently, an Active Surveillance Program using hunter-harvest waterfowl in Washington and Oregon detected ten HPAI H5 viruses, of three different subtypes (four H5N2, three H5N8 and three H5N1) with 4 segments in common (HA, PB2, NP and MA). In addition, a mortality-based Passive Surveillance Program detected 18 HPAI (14 H5N2 and four H5N8) cases from Idaho, Kansas, Oregon, Minnesota, Montana, Washington and Wisconsin. Comparatively, mortality-based passive surveillance appears to be detecting these HPAI infections at a higher rate than active surveillance during the period following initial introduction into the US.

  5. Protection from pulmonary tissue damage associated with infection of cynomolgus macaques by highly pathogenic avian influenza virus (H5N1) by low dose natural human IFN-α administered to the buccal mucosa.

    Science.gov (United States)

    Strayer, David R; Carter, William A; Stouch, Bruce C; Stittelaar, Koert J; Thoolen, Robert J M M; Osterhaus, Albert D M E; Mitchell, William M

    2014-10-01

    Using an established nonhuman primate model for H5N1 highly pathogenic influenza virus infection in humans, we have been able to demonstrate the prophylactic mitigation of the pulmonary damage characteristic of human fatal cases from primary influenza virus pneumonia with a low dose oral formulation of a commercially available parenteral natural human interferon alpha (Alferon N Injection®). At the highest oral dose (62.5IU/kg body weight) used there was a marked reduction in the alveolar inflammatory response with minor evidence of alveolar and interstitial edema in contrast to the hemorrhage and inflammatory response observed in the alveoli of control animals. The mitigation of severe damage to the lower pulmonary airway was observed without a parallel reduction in viral titers. Clinical trial data will be necessary to establish its prophylactic human efficacy for highly pathogenic influenza viruses. Copyright © 2014. Published by Elsevier B.V.

  6. [Construction of a recombinant HVT virus expressing the HA gene of avian influenza virus H5N1 via Rde/ET recombination system].

    Science.gov (United States)

    Lan, Desong; Shi, Xingming; Wang, Yunfeng; Liu, Changjun; Wang, Mei; Cui, Hongyu; Tian, Guobin; Li, Jisong; Tong, Guangzhi

    2009-01-01

    In recent years,manipulation of large herpesvirus genomes has been facilitated by using bacterial artificial chromosome (BAC) vectors. We have previously reported the construction of the BAC clones (HVT BACs) of herpesvirus of turkey (HVT). With these BAC clones in hand,we manipulated the genome of HVT by utilizing Red/ET recombination system, and developed a biologically safe live vaccine based on the HVT BACs. In this two-step approach, we first transformed the plasmid pRedET into the DH10B competent cells that carried the HVT BACs,and added inducer L-arabinose into the cells. We prepared the cells into competent cells and electroporated the linear rpsL-neo counter-selection/selection cassette flanked by the 50 bp long homology arms into the cells. So the functional cassette was inserted into the U(S)2 locus. Only colonies carrying the modified BAC would survive Kanamycin selection on the agar plates. The successful integration of the rpsL-neo cassette was monitored by PCR and Streptomycin selection, for the insertion of rpsL-neo cassette cells will become Streptomycin sensitive. Secondly, in the same way, we replaced the rpsL-neo cassette with the hemagglutinin (HA) gene of (HPAIV) A/Goose/ Guangdong/1/96(H5N1) flanked by the same homology arms. Only colonies which lost the rpsL-neo cassette will grow on Streptomycin containing plates. Finally, we obtained many colonies of which the HA gene of the AIV was inserted into the U(S)2 locus to be modified of HVT. And we reconstituted one recombinant virus from transfecting one of these BAC clones DNA into chick embryo fibroblasts (CEFs). We achieved one rescued recombinant virus which designated as rHVT-HA3. The H5 subtype HA gene expression in this recombinant virus rHVT-HA3 was confirmed by immunofluorescence assay.

  7. Evaluation of a commercial competitive enzyme-linked immunosorbent assay for detection of avian influenza virus subtype H5 antibodies in zoo birds

    DEFF Research Database (Denmark)

    Jensen, Trine Hammer; Andersen, Jannie Holmegaard; Hjulsager, Charlotte Kristiane

    2017-01-01

    The hemagglutination inhibition (HI) test is the current gold standard for detecting antibodies to avian influenza virus (AIV). Enzyme-linked immunosorbent assays (ELISAs) have been explored for use in poultry and certain wild bird species because of high efficiency and lower cost. This study com...

  8. Role for migratory wild birds in the global spread of avian influenza H5N8

    Science.gov (United States)

    ,; Ip, Hon S.

    2016-01-01

    Avian influenza viruses affect both poultry production and public health. A subtype H5N8 (clade 2.3.4.4) virus, following an outbreak in poultry in South Korea in January 2014, rapidly spread worldwide in 2014–2015. Our analysis of H5N8 viral sequences, epidemiological investigations, waterfowl migration, and poultry trade showed that long-distance migratory birds can play a major role in the global spread of avian influenza viruses. Further, we found that the hemagglutinin of clade 2.3.4.4 virus was remarkably promiscuous, creating reassortants with multiple neuraminidase subtypes. Improving our understanding of the circumpolar circulation of avian influenza viruses in migratory waterfowl will help to provide early warning of threats from avian influenza to poultry, and potentially human, health.

  9. The PB2, PA, HA, NP, and NS genes of a highly pathogenic avian influenza virus A/whooper swan/Mongolia/3/2005 (H5N1 are responsible for pathogenicity in ducks

    Directory of Open Access Journals (Sweden)

    Kajihara Masahiro

    2013-02-01

    Full Text Available Abstract Background Wild ducks are the natural hosts of influenza A viruses. Duck influenza, therefore, has been believed inapparent infection with influenza A viruses, including highly pathogenic avian influenza viruses (HPAIVs in chickens. In fact, ducks experimentally infected with an HPAIV strain, A/Hong Kong/483/1997 (H5N1 (HK483, did not show any clinical signs. Another HPAIV strain, A/whooper swan/Mongolia/3/2005 (H5N1 (MON3 isolated from a dead swan, however, caused neurological dysfunction and death in ducks. Method To understand the mechanism whereby MON3 shows high pathogenicity in ducks, HK483, MON3, and twenty-four reassortants generated between these two H5N1 viruses were compared for their pathogenicity in domestic ducks. Results None of the ducks infected with MON3-based single-gene reassortants bearing the PB2, NP, or NS gene segment of HK483 died, and HK483-based single-gene reassortants bearing PB2, NP, or NS genes of MON3 were not pathogenic in ducks, suggesting that multiple gene segments contribute to the pathogenicity of MON3 in ducks. All the ducks infected with the reassortant bearing PB2, PA, HA, NP, and NS gene segments of MON3 died within five days post-inoculation, as did those infected with MON3. Each of the viruses was assessed for replication in ducks three days post-inoculation. MON3 and multi-gene reassortants pathogenic in ducks were recovered from all of the tissues examined and replicated with high titers in the brains and lungs. Conclusion The present results indicate that multigenic factors are responsible for efficient replication of MON3 in ducks. In particular, virus growth in the brain might correlate with neurological dysfunction and the disease severity.

  10. Isolation of H5N6, H7N9 and H9N2 avian influenza A viruses from air sampled at live poultry markets in China, 2014 and 2015.

    Science.gov (United States)

    Zhou, Jie; Wu, Jie; Zeng, Xianqiao; Huang, Guofeng; Zou, Lirong; Song, Yingchao; Gopinath, Divya; Zhang, Xin; Kang, Min; Lin, Jinyan; Cowling, Benjamin J; Lindsley, William G; Ke, Changwen; Peiris, Joseph Sriyal Malik; Yen, Hui-Ling

    2016-09-01

    Zoonotic infections by avian influenza viruses occur at the human-poultry interface, but the modes of transmission have not been fully investigated. We assessed the potential for airborne and fomite transmission at live poultry markets in Guangzhou city and in Hong Kong Special Administrative Region (SAR), China, during 2014 and 2015. Viral genome and infectious avian influenza A viruses of H5N6, H7N9, and H9N2 subtypes were detected predominantly from particles larger or equal to 1 μm in diameter in the air sampled with cyclone-based bioaerosol samplers at the live poultry markets in Guangzhou. Influenza A(H9N2) viruses were ubiquitously isolated every month during the study period from air and environmental swabs, and different lineages of H9N2 virus were isolated from markets where chickens and minor land-based poultry were sold. The use of de-feathering devices increased the quantity of virus-laden airborne particles while market closure reduced the amount of such particles. The results highlight the possibility of airborne transmission of avian influenza viruses among poultry or from poultry to humans within such settings. This may explain epidemiological observations in which some patients with H7N9 infection reported being in markets but no direct contact with live poultry or poultry stalls. This article is copyright of The Authors, 2016.

  11. An Adjuvanted A(H5N1) Subvirion Vaccine Elicits Virus-Specific Antibody Response and Improves Protection Against Lethal Influenza Viral Challenge in Mouse Model of Protein Energy Malnutrition.

    Science.gov (United States)

    Jones, Enitra N; Amoah, Samuel; Cao, Weiping; Sambhara, Suryaprakash; Gangappa, Shivaprakash

    2017-09-15

    Protein energy malnutrition (PEM) increases susceptibility to infectious diseases, including influenza infection, but no studies have addressed the potential influences of PEM on the immunogenicity and protective efficacy of avian influenza A(H5N1) vaccine. We investigated the role of PEM on vaccine-mediated protection after a lethal challenge with recombinant A(H5N1) virus using isocaloric diets providing either adequate protein (AP; 18% protein) or very low protein (VLP; 2% protein) in an established murine model of influenza vaccination. We demonstrated that mice maintained on a VLP diet succumb to lethal challenge at greater rates than mice maintained on an AP diet, despite comparable immunization regimens. Importantly, there was no virus-induced mortality in both VLP and AP groups of mice when either group was immunized with adjuvanted low-dose A(H5N1) subvirion vaccine. Our results suggest that adjuvanted vaccination in populations where PEM is endemic may be one strategy to boost vaccination-promoted immunity and improve outcomes associated with highly pathogenic A(H5N1). Published by Oxford University Press for the Infectious Diseases Society of America 2017. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  12. Host cytokine responses of pigeons infected with highly pathogenic Thai avian influenza viruses of subtype H5N1 isolated from wild birds.

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    Tsuyoshi Hayashi

    Full Text Available Highly pathogenic avian influenza virus (HPAIV of the H5N1 subtype has been reported to infect pigeons asymptomatically or induce mild symptoms. However, host immune responses of pigeons inoculated with HPAIVs have not been well documented. To assess host responses of pigeons against HPAIV infection, we compared lethality, viral distribution and mRNA expression of immune related genes of pigeons infected with two HPAIVs (A/Pigeon/Thailand/VSMU-7-NPT/2004; Pigeon04 and A/Tree sparrow/Ratchaburi/VSMU-16-RBR/2005; T.sparrow05 isolated from wild birds in Thailand. The survival experiment showed that 25% of pigeons died within 2 weeks after the inoculation of two HPAIVs or medium only, suggesting that these viruses did not cause lethal infection in pigeons. Pigeon04 replicated in the lungs more efficiently than T.sparrow05 and spread to multiple extrapulmonary organs such as the brain, spleen, liver, kidney and rectum on days 2, 5 and 9 post infection. No severe lesion was observed in the lungs infected with Pigeon04 as well as T.sparrow05 throughout the collection periods. Encephalitis was occasionally observed in Pigeon04- or T.sparrow05-infected brain, the severity, however was mostly mild. To analyze the expression of immune-related genes in the infected pigeons, we established a quantitative real-time PCR analysis for 14 genes of pigeons. On day 2 post infection, Pigeon04 induced mRNA expression of Mx1, PKR and OAS to a greater extent than T.sparrow05 in the lungs, however their expressions were not up-regulated concomitantly on day 5 post infection when the peak viral replication was observed. Expressions of TLR3, IFNα, IL6, IL8 and CCL5 in the lungs following infection with the two HPAIVs were low. In sum, Pigeon04 exhibited efficient replication in the lungs compared to T.sparrow05, but did not induce excessive host cytokine expressions. Our study has provided the first insight into host immune responses of pigeons against HPAIV infection.

  13. Single-dose mucosal immunization with a candidate universal influenza vaccine provides rapid protection from virulent H5N1, H3N2 and H1N1 viruses.

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    Graeme E Price

    2010-10-01

    Full Text Available The sudden emergence of novel influenza viruses is a global public health concern. Conventional influenza vaccines targeting the highly variable surface glycoproteins hemagglutinin and neuraminidase must antigenically match the emerging strain to be effective. In contrast, "universal" vaccines targeting conserved viral components could be used regardless of viral strain or subtype. Previous approaches to universal vaccination have required protracted multi-dose immunizations. Here we evaluate a single dose universal vaccine strategy using recombinant adenoviruses (rAd expressing the conserved influenza virus antigens matrix 2 and nucleoprotein.In BALB/c mice, administration of rAd via the intranasal route was superior to intramuscular immunization for induction of mucosal responses and for protection against highly virulent H1N1, H3N2, or H5N1 influenza virus challenge. Mucosally vaccinated mice not only survived, but had little morbidity and reduced lung virus titers. Protection was observed as early as 2 weeks post-immunization, and lasted at least 10 months, as did antibodies and lung T cells with activated phenotypes. Virus-specific IgA correlated with but was not essential for protection, as demonstrated in studies with IgA-deficient animals.Mucosal administration of NP and M2-expressing rAd vectors provided rapid and lasting protection from influenza viruses in a subtype-independent manner. Such vaccines could be used in the interval between emergence of a new virus strain and availability of strain-matched vaccines against it. This strikingly effective single-dose vaccination thus represents a candidate off-the-shelf vaccine for emergency use during an influenza pandemic.

  14. The pathogenecity of H5N1 highly pathogenic Avian Influenza (HPAI virus clade 2.3.2. in Indonesian indigenous chicken by contact tranmission with infected duck

    Directory of Open Access Journals (Sweden)

    R. Damayanti

    2017-05-01

    Full Text Available An experimental transmission study was conducted using nine healthy Indonesian indigenous chickens placed together with two 30 days old ducks which were experimentally infected with H5N1 HPAI clade 2.3.2 virus in the Biosafety Laboratory Level 3 (BSL-3 facilities. The aim of the study was to find out the pathogenicity of H5N1 HPAI virus clade 2.3.2 in Indonesian indigenous chickens. The study showed that within twenty four hours rearing, the chickens were exhibited mild clinical signs and by 48 hours, all of the chickens died, whereas the ducks survived but with severe clinical signs. The H5N1 HPAI virus has been successfully isolated from chickens and ducks swabs, confirming that those animals were infected by the virus. Histologically, the infected chicken encountered with severe inflammation reaction namely non suppuratives encephalitis, tracheitis, myocarditis, interstitial pneumonia, hepatitis, proventriculitis, enteritis, pancreatitis, nephritis and bursitis. Necrotizing spleen and pancreas were also prominent. Viral antigen was detected by immunohistochemistry staining in various affected visceral organs. This suggests that Indonesian indigenous chickens were susceptible to H5N1 HPAI virus clade 2.3.2 and it can be transmitted easily to Indonesian indigenous chickens by contact transmission with infected ducks.

  15. Protection patterns in duck and chicken after homo- or hetero-subtypic reinfections with H5 and H7 low pathogenicity avian influenza viruses: a comparative study.

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    Coralie Chaise

    Full Text Available Avian influenza viruses are circulating continuously in ducks, inducing a mostly asymptomatic infection, while chickens are accidental hosts highly susceptible to respiratory disease. This discrepancy might be due to a different host response to the virus between these two bird species and in particular to a different susceptibility to reinfection. In an attempt to address this question, we analyzed, in ducks and in chickens, the viral load in infected tissues and the humoral immune response after experimental primary and secondary challenge infections with either homologous or heterologous low pathogenicity avian influenza viruses (LPAIV. Following homologous reinfection, ducks were only partially protected against viral shedding in the lower intestine in conjunction with a moderate antibody response, whereas chickens were totally protected against viral shedding in the upper respiratory airways and developed a stronger antibody response. On the contrary, heterologous reinfection was not followed by a reduced viral excretion in the upper airways of chickens, while ducks were still partially protected from intestinal excretion of the virus, with no correlation to the antibody response. Our comparative study provides a comprehensive demonstration of the variation of viral tropism and control of the host humoral response to LPAIV between two different bird species with different degrees of susceptibility to avian influenza.

  16. A comparative evaluation of feathers, oropharyngeal swabs, and cloacal swabs for the detection of H5N1 highly pathogenic avian influenza virus infection in experimentally infected chickens and ducks.

    Science.gov (United States)

    Nuradji, Harimurti; Bingham, John; Lowther, Sue; Wibawa, Hendra; Colling, Axel; Long, Ngo Thanh; Meers, Joanne

    2015-11-01

    Oropharyngeal and cloacal swabs have been widely used for the detection of H5N1 highly pathogenic avian Influenza A virus (HPAI virus) in birds. Previous studies have shown that the feather calamus is a site of H5N1 virus replication and therefore has potential for diagnosis of avian influenza. However, studies characterizing the value of feathers for this purpose are not available, to our knowledge; herein we present a study investigating feathers for detection of H5N1 virus. Ducks and chickens were experimentally infected with H5N1 HPAI virus belonging to 1 of 3 clades (Indonesian clades 2.1.1 and 2.1.3, Vietnamese clade 1). Different types of feathers and oropharyngeal and cloacal swab samples were compared by virus isolation. In chickens, virus was detected from all sample types: oral and cloacal swabs, and immature pectorosternal, flight, and tail feathers. During clinical disease, the viral titers were higher in feathers than swabs. In ducks, the proportion of virus-positive samples was variable depending on viral strain and time from challenge; cloacal swabs and mature pectorosternal feathers were clearly inferior to oral swabs and immature pectorosternal, tail, and flight feathers. In ducks infected with Indonesian strains, in which most birds did not develop clinical signs, all sampling methods gave intermittent positive results; 3-23% of immature pectorosternal feathers were positive during the acute infection period; oropharyngeal swabs had slightly higher positivity during early infection, while feathers performed better during late infection. Our results indicate that immature feathers are an alternative sample for the diagnosis of HPAI in chickens and ducks. © 2015 The Author(s).

  17. Modeling the Association of Space, Time, and Host Species with Variation of the HA, NA, and NS Genes of H5N1 Highly Pathogenic Avian Influenza Viruses Isolated from Birds in Romania in 2005–2007

    Science.gov (United States)

    Alkhamis, Mohammad; Perez, Andres; Batey, Nicole; Howard, Wendy; Baillie, Greg; Watson, Simon; Franz, Stephanie; Focosi-Snyman, Raffaella; Onita, Iuliana; Cioranu, Raluca; Turcitu, Mihai; Kellam, Paul; Brown, Ian H.; Breed, Andrew C.

    2014-01-01

    SUMMARY Molecular characterization studies of a diverse collection of avian influenza viruses (AIVs) have demonstrated that AIVs’ greatest genetic variability lies in the HA, NA, and NS genes. The objective here was to quantify the association between geographical locations, periods of time, and host species and pairwise nucleotide variation in the HA, NA, and NS genes of 70 isolates of H5N1 highly pathogenic avian influenza virus (HPAIV) collected from October 2005 to December 2007 from birds in Romania. A mixed-binomial Bayesian regression model was used to quantify the probability of nucleotide variation between isolates and its association with space, time, and host species. As expected for the three target genes, a higher probability of nucleotide differences (odds ratios [ORs] > 1) was found between viruses sampled from places at greater geographical distances from each other, viruses sampled over greater periods of time, and viruses derived from different species. The modeling approach in the present study maybe useful in further understanding the molecular epidemiology of H5N1 HPAI virus in bird populations. The methodology presented here will be useful in predicting the most likely genetic distance for any of the three gene segments of viruses that have not yet been isolated or sequenced based on space, time, and host species during the course of an epidemic. PMID:24283126

  18. PA from an H5N1 highly pathogenic avian influenza virus activates viral transcription and replication and induces apoptosis and interferon expression at an early stage of infection

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    Wang Qiang

    2012-06-01

    Full Text Available Abstract Background Although gene exchange is not likely to occur freely, reassortment between the H5N1 highly pathogenic avian influenza virus (HPAIV and currently circulating human viruses is a serious concern. The PA polymerase subunit of H5N1 HPAIV was recently reported to activate the influenza replicon activity. Methods The replicon activities of PR8 and WSN strains (H1N1 of influenza containing PA from HPAIV A/Cambodia/P0322095/2005 (H5N1 and the activity of the chimeric RNA polymerase were analyzed. A reassortant WSN virus containing the H5N1 Cambodia PA (C-PA was then reconstituted and its growth in cells and pathogenicity in mice examined. The interferon promoter, TUNEL, and caspase 3, 8, and 9 activities of C-PA-infected cells were compared with those of WSN-infected cells. Results The activity of the chimeric RNA polymerase was slightly higher than that of WSN, and C-PA replicated better than WSN in cells. However, the multi-step growth of C-PA and its pathogenicity in mice were lower than those of WSN. The interferon promoter, TUNEL, and caspase 3, 8, and 9 activities were strongly induced in early infection in C-PA-infected cells but not in WSN-infected cells. Conclusions Apoptosis and interferon were strongly induced early in C-PA infection, which protected the uninfected cells from expansion of viral infection. In this case, these classical host-virus interactions contributed to the attenuation of this strongly replicating virus.

  19. Isolation of influenza A virus, subtype H5N2, and avian paramyxovirus type 1 from a flock of ostriches in Europe

    DEFF Research Database (Denmark)

    Jørgensen, Poul Henrik; Nielsen, O.L.; Hansen, C.

    1998-01-01

    A total of 146 of 506 ostriches (Struthio camelus) introduced into a quarantine in Denmark died within the first 23 days. The majority of deaths were in young birds up to 10 kg body weight. Avian influenza A viruses (AIVs) were isolated from 14 pools of organ tissues representing seven groups each......-Q-R-E-T-R*G-L-F- at the cleavage site of the haemagglutinin protein, typical of non-pathogenic AIVs. In addition, an avirulent avian paramyxovirus type 1 virus was isolated from one pool of kidney tissues. Bacteriological examination gave no significant results. The most characteristic pathological findings were impaction...

  20. In Silico Prediction and Experimental Confirmation of HA Residues Conferring Enhanced Human Receptor Specificity of H5N1 Influenza A Viruses

    Science.gov (United States)

    Schmier, Sonja; Mostafa, Ahmed; Haarmann, Thomas; Bannert, Norbert; Ziebuhr, John; Veljkovic, Veljko; Dietrich, Ursula; Pleschka, Stephan

    2015-06-01

    Newly emerging influenza A viruses (IAV) pose a major threat to human health by causing seasonal epidemics and/or pandemics, the latter often facilitated by the lack of pre-existing immunity in the general population. Early recognition of candidate pandemic influenza viruses (CPIV) is of crucial importance for restricting virus transmission and developing appropriate therapeutic and prophylactic strategies including effective vaccines. Often, the pandemic potential of newly emerging IAV is only fully recognized once the virus starts to spread efficiently causing serious disease in humans. Here, we used a novel phylogenetic algorithm based on the informational spectrum method (ISM) to identify potential CPIV by predicting mutations in the viral hemagglutinin (HA) gene that are likely to (differentially) affect critical interactions between the HA protein and target cells from bird and human origin, respectively. Predictions were subsequently validated by generating pseudotyped retrovirus particles and genetically engineered IAV containing these mutations and characterizing potential effects on virus entry and replication in cells expressing human and avian IAV receptors, respectively. Our data suggest that the ISM-based algorithm is suitable to identify CPIV among IAV strains that are circulating in animal hosts and thus may be a new tool for assessing pandemic risks associated with specific strains.

  1. Non-Attenuation Of Highly Pathogenic Avian Influenza H5N1 By ...

    African Journals Online (AJOL)

    Avian influenza H5N1 represents one of the most researched viruses in laboratories world-wide in recent times with regards to its epidemiology, ecology, biology and geography. The virus has caused 409 human cases and 256 human fatalities to date. Some laboratory activities and other lab related works predispose ...

  2. Novel genotypes of H9N2 influenza A viruses isolated from poultry in Pakistan containing NS genes similar to highly pathogenic H7N3 and H5N1 viruses.

    Directory of Open Access Journals (Sweden)

    Munir Iqbal

    2009-06-01

    Full Text Available The impact of avian influenza caused by H9N2 viruses in Pakistan is now significantly more severe than in previous years. Since all gene segments contribute towards the virulence of avian influenza virus, it was imperative to investigate the molecular features and genetic relationships of H9N2 viruses prevalent in this region. Analysis of the gene sequences of all eight RNA segments from 12 viruses isolated between 2005 and 2008 was undertaken. The hemagglutinin (HA sequences of all isolates were closely related to H9N2 viruses isolated from Iran between 2004 and 2007 and contained leucine instead of glutamine at position 226 in the receptor binding pocket, a recognised marker for the recognition of sialic acids linked alpha2-6 to galactose. The neuraminidase (NA of two isolates contained a unique five residue deletion in the stalk (from residues 80 to 84, a possible indication of greater adaptation of these viruses to the chicken host. The HA, NA, nucleoprotein (NP, and matrix (M genes showed close identity with H9N2 viruses isolated during 1999 in Pakistan and clustered in the A/Quail/Hong Kong/G1/97 virus lineage. In contrast, the polymerase genes clustered with H9N2 viruses from India, Iran and Dubai. The NS gene segment showed greater genetic diversity and shared a high level of similarity with NS genes from either H5 or H7 subtypes rather than with established H9N2 Eurasian lineages. These results indicate that during recent years the H9N2 viruses have undergone extensive genetic reassortment which has led to the generation of H9N2 viruses of novel genotypes in the Indian sub-continent. The novel genotypes of H9N2 viruses may play a role in the increased problems observed by H9N2 to poultry and reinforce the continued need to monitor H9N2 infections for their zoonotic potential.

  3. Identification of Molecular Markers Associated with Alteration of Receptor-Binding Specificity in a Novel Genotype of Highly Pathogenic Avian Influenza A(H5N1) Viruses Detected in Cambodia in 2013

    Science.gov (United States)

    Rith, Sareth; Davis, C. Todd; Duong, Veasna; Sar, Borann; Horm, Srey Viseth; Chin, Savuth; Ly, Sovann; Laurent, Denis; Richner, Beat; Oboho, Ikwo; Jang, Yunho; Davis, William; Thor, Sharmi; Balish, Amanda; Iuliano, A. Danielle; Sorn, San; Holl, Davun; Sok, Touch; Seng, Heng; Tarantola, Arnaud; Tsuyuoka, Reiko; Parry, Amy; Chea, Nora; Allal, Lotfi; Kitsutani, Paul; Warren, Dora; Prouty, Michael; Horwood, Paul; Widdowson, Marc-Alain; Lindstrom, Stephen; Villanueva, Julie; Donis, Ruben; Cox, Nancy

    2014-01-01

    Human infections with influenza A(H5N1) virus in Cambodia increased sharply during 2013. Molecular characterization of viruses detected in clinical specimens from human cases revealed the presence of mutations associated with the alteration of receptor-binding specificity (K189R, Q222L) and respiratory droplet transmission in ferrets (N220K with Q222L). Discovery of quasispecies at position 222 (Q/L), in addition to the absence of the mutations in poultry/environmental samples, suggested that the mutations occurred during human infection and did not transmit further. PMID:25210193

  4. Avian influenza H5N1 transmission in households, Indonesia.

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    Tjandra Y Aditama

    Full Text Available BACKGROUND: Disease transmission patterns are needed to inform public health interventions, but remain largely unknown for avian influenza H5N1 virus infections. A recent study on the 139 outbreaks detected in Indonesia between 2005 and 2009 found that the type of exposure to sources of H5N1 virus for both the index case and their household members impacted the risk of additional cases in the household. This study describes the disease transmission patterns in those outbreak households. METHODOLOGY/PRINCIPAL FINDINGS: We compared cases (n = 177 and contacts (n = 496 in the 113 sporadic and 26 cluster outbreaks detected between July 2005 and July 2009 to estimate attack rates and disease intervals. We used final size household models to fit transmission parameters to data on household size, cases and blood-related household contacts to assess the relative contribution of zoonotic and human-to-human transmission of the virus, as well as the reproduction number for human virus transmission. The overall household attack rate was 18.3% and secondary attack rate was 5.5%. Secondary attack rate remained stable as household size increased. The mean interval between onset of subsequent cases in outbreaks was 5.6 days. The transmission model found that human transmission was very rare, with a reproduction number between 0.1 and 0.25, and the upper confidence bounds below 0.4. Transmission model fit was best when the denominator population was restricted to blood-related household contacts of index cases. CONCLUSIONS/SIGNIFICANCE: The study only found strong support for human transmission of the virus when a single large cluster was included in the transmission model. The reproduction number was well below the threshold for sustained transmission. This study provides baseline information on the transmission dynamics for the current zoonotic virus and can be used to detect and define signatures of a virus with increasing capacity for human

  5. Influenza (Flu) Viruses

    Science.gov (United States)

    ... Types Seasonal Avian Swine Variant Pandemic Other Influenza (Flu) Viruses Language: English (US) Español Recommend on Facebook ... influenza circulate and cause illness. More Information about Flu Viruses Types of Influenza Viruses Influenza A and ...

  6. Comparison of temporal and spatial dynamics of seasonal H3N2, pandemic H1N1 and highly pathogenic avian influenza H5N1 virus infections in ferrets.

    Directory of Open Access Journals (Sweden)

    Judith M A van den Brand

    Full Text Available Humans may be infected by different influenza A viruses--seasonal, pandemic, and zoonotic--which differ in presentation from mild upper respiratory tract disease to severe and sometimes fatal pneumonia with extra-respiratory spread. Differences in spatial and temporal dynamics of these infections are poorly understood. Therefore, we inoculated ferrets with seasonal H3N2, pandemic H1N1 (pH1N1, and highly pathogenic avian H5N1 influenza virus and performed detailed virological and pathological analyses at time points from 0.5 to 14 days post inoculation (dpi, as well as describing clinical signs and hematological parameters. H3N2 infection was restricted to the nose and peaked at 1 dpi. pH1N1 infection also peaked at 1 dpi, but occurred at similar levels throughout the respiratory tract. H5N1 infection occurred predominantly in the alveoli, where it peaked for a longer period, from 1 to 3 dpi. The associated lesions followed the same spatial distribution as virus infection, but their severity peaked between 1 and 6 days later. Neutrophil and monocyte counts in peripheral blood correlated with inflammatory cell influx in the alveoli. Of the different parameters used to measure lower respiratory tract disease, relative lung weight and affected lung tissue allowed the best quantitative distinction between the virus groups. There was extra-respiratory spread to more tissues--including the central nervous system--for H5N1 infection than for pH1N1 infection, and to none for H3N2 infection. This study shows that seasonal, pandemic, and zoonotic influenza viruses differ strongly in the spatial and temporal dynamics of infection in the respiratory tract and extra-respiratory tissues of ferrets.

  7. Highly pathogenic avian influenza virus H5N1 controls type I IFN induction in chicken macrophage HD-11 cells: a polygenic trait that involves NS1 and the polymerase complex

    Science.gov (United States)

    2012-01-01

    Background Influenza A viruses are well characterized to antagonize type I IFN induction in infected mammalian cells. However, limited information is available for avian cells. It was hypothesised that avian influenza viruses (AIV) with distinct virulence may interact differently with the avian innate immune system. Therefore, the type I IFN responses induced by highly virulent and low virulent H5N1 AIV and reassortants thereof were analysed in chicken cells. Results The highly pathogenic (HP) AIV A/chicken/Yamaguchi/7/04 (H5N1) (Yama) did not induce type I IFN in infected chicken HD-11 macrophage-like cells. This contrasted with an NS1 mutant Yama virus (Yama-NS1A144V) and with the attenuated H5N1 AIV A/duck/Hokkaido/Vac-1/04 (Vac) carrying the haemagglutinin (HA) of the Yama virus (Vac-Yama/HA), that both induced type I IFN in these cells. The substitution of the NS segment from Yama with that from Vac in the Yama backbone resulted in induction of type I IFN secretion in HD-11 cells. However, vice versa, the Yama NS segment did not prevent type I IFN induction by the Vac-Yama/HA virus. This was different with the PB1/PB2/PA segment reassortant Yama and Vac-Yama/HA viruses. Whereas the Yama virus with the Vac PB1/PB2/PA segments induced type I IFN in HD-11 cells, the Vac-Yama/HA virus with the Yama PB1/PB2/PA segments did not. As reported for mammalian cells, the expression of H5N1 PB2 inhibited the activation of the IFN-β promoter in chicken DF-1 fibroblast cells. Importantly, the Yama PB2 was more potent at inhibiting the IFN-β promoter than the Vac PB2. Conclusions The present study demonstrates that the NS1 protein and the polymerase complex of the HPAIV Yama act in concert to antagonize chicken type I IFN secretion in HD-11 cells. PB2 alone can also exert a partial inhibitory effect on type I IFN induction. In conclusion, the control of type I IFN induction by H5N1 HPAIV represents a complex phenotype that involves a particular viral gene constellation

  8. Susceptibility of Muscovy (Cairina Moschata) and mallard ducks (Anas Platyrhynchos) to experimental infections by different genotypes of H5N1 avian influenza viruses

    DEFF Research Database (Denmark)

    Phuong, Do Quy; Dung, Nguyen Tien; Jørgensen, Poul Henrik

    2011-01-01

    exposed to infection with H5N1. To do this, an experimental study on infections with different genotypes of H5N1 in mallards and Muscovy ducks have been conducted, where it was found that the mortality of the inoculated Muscovy ducks was at least 80%, regardless of the virus strain employed. In contrast...... in the brain. Due to their high susceptibility, it is unlikely that Muscovy ducks act as a silent reservoir. The virus strains used in this study, to a certain degree, differed in their virulence properties to the bird species in question.......It is a fact that in Viet Nam, Muscovy ducks are raised in large populations (approximately 8 million), usually kept in small flocks together with mallards and chickens. As a result, it is a great concern for epidemiologists to elucidate possible differences in relation to these species being...

  9. Characterization of Clade 2.3.2.1 H5N1 Highly Pathogenic Avian Influenza Viruses Isolated from Wild Birds (Mandarin Duck and Eurasian Eagle Owl in 2010 in Korea

    Directory of Open Access Journals (Sweden)

    Youn-Jeong Lee

    2013-04-01

    Full Text Available Starting in late November 2010, the H5N1 highly pathogenic avian influenza (HPAI virus was isolated from many types of wild ducks and raptors and was subsequently isolated from poultry in Korea. We assessed the genetic and pathogenic properties of the HPAI viruses isolated from a fecal sample from a mandarin duck and a dead Eurasian eagle owl, the most affected wild bird species during the 2010/2011 HPAI outbreak in Korea. These viruses have similar genetic backgrounds and exhibited the highest genetic similarity with recent Eurasian clade 2.3.2.1 HPAI viruses. In animal inoculation experiments, regardless of their originating hosts, the two Korean isolates produced highly pathogenic characteristics in chickens, ducks and mice without pre-adaptation. These results raise concerns about veterinary and public health. Surveillance of wild birds could provide a good early warning signal for possible HPAI infection in poultry as well as in humans.

  10. Highly Pathogenic Avian Influenza H5N8 Clade 2.3.4.4 Virus: Equivocal Pathogenicity and Implications for Surveillance Following Natural Infection in Breeder Ducks in the United Kingdom.

    Science.gov (United States)

    Núñez, A; Brookes, S M; Reid, S M; Garcia-Rueda, C; Hicks, D J; Seekings, J M; Spencer, Y I; Brown, I H

    2016-02-01

    Since early 2014, several outbreaks involving novel reassortant highly pathogenic avian influenza (HPAI) A(H5N8) viruses have been detected in poultry and wild bird species in Asia, Europe and North America. These viruses have been detected in apparently healthy and dead wild migratory birds, as well as in domestic chickens, turkeys, geese and ducks. In this study, we describe the pathology of an outbreak of H5N8 HPAIV in breeder ducks in the UK. A holding with approximately 6000 breeder ducks, aged approximately 60 weeks, showed a gradual reduction in egg production and increased mortality over a 7-day period. Post-mortem examination revealed frequent fibrinous peritonitis, with severely haemorrhagic ovarian follicles and occasional splenic and pancreatic necrosis and high incidence of mycotic granulomas in the air sacs and lung. Low-to-moderate levels of HPAI H5N8 virus were detected mainly in respiratory and digestive tract, with minor involvement of other organs. Although histopathological examination confirmed the gross pathology findings, intralesional viral antigen detection by immunohistochemistry was not observed. Immunolabelled cells were rarely only present in inflamed air sacs and serosa, usually superficial to granulomatous inflammation. Abundant bacterial microcolonies were observed in haemorrhagic ovaries and oviduct. The limited viral tissue distribution and presence of inter-current fungal and bacterial infections suggest a minor role for HPAIV H5N8 in clinical disease in layer ducks. © 2015 Crown copyright.

  11. Genetic Reassortment Among the Influenza Viruses (Avian Influenza, Human Influenza and Swine Influenza in Pigs

    Directory of Open Access Journals (Sweden)

    Dyah Ayu Hewajuli

    2012-12-01

    Full Text Available Influenza A virus is a hazardous virus and harm to respiratory tract. The virus infect birds, pigs, horses, dogs, mammals and humans. Pigs are important hosts in ecology of the influenza virus because they have two receptors, namely NeuAc 2,3Gal and NeuAc 2,6Gal which make the pigs are sensitive to infection of influenza virus from birds and humans and genetic reassortment can be occurred. Classical swine influenza H1N1 viruses had been circulated in pigs in North America and other countries for 80 years. In 1998, triple reassortant H3N2 swine influenza viruses that contains genes of human influenza A virus (H3N2, swine influenza virus (H1N1 and avian influenza are reported as cause an outbreaks in pigs in North America. Furthermore, the circulation of triple reassortant H3N2 swine influenza virus resulting reassortant H1N1 swine influenza and reassortant H1N2 swine influenza viruses cause infection in humans. Humans who were infected by triple reassortant swine influenza A virus (H1N1 usually made direct contact with pigs. Although without any clinical symptoms, pigs that are infected by triple reassortant swine influenza A (H1N1 can transmit infection to the humans around them. In June 2009, WHO declared that pandemic influenza of reassortant H1N1 influenza A virus (novel H1N1 has reached phase 6. In Indonesia until 2009, there were 1005 people were infected by H1N1 influenza A and 5 of them died. Novel H1N1 and H5N1 viruses have been circulated in humans and pigs in Indonesia. H5N1 reassortant and H1N1 viruses or the seasonal flu may could arise because of genetic reassortment between avian influenza and humans influenza viruses that infect pigs together.

  12. Safety and immunogenicity of a modified-vaccinia-virus-Ankara-based influenza A H5N1 vaccine: a randomised, double-blind phase 1/2a clinical trial.

    Science.gov (United States)

    Kreijtz, Joost H C M; Goeijenbier, Marco; Moesker, Fleur M; van den Dries, Lennert; Goeijenbier, Simone; De Gruyter, Heidi L M; Lehmann, Michael H; Mutsert, Gerrie de; van de Vijver, David A M C; Volz, Asisa; Fouchier, Ron A M; van Gorp, Eric C M; Rimmelzwaan, Guus F; Sutter, Gerd; Osterhaus, Albert D M E

    2014-12-01

    Modified vaccinia virus Ankara (MVA) is a promising viral vector platform for the development of an H5N1 influenza vaccine. Preclinical assessment of MVA-based H5N1 vaccines showed their immunogenicity and safety in different animal models. We aimed to assess the safety and immunogenicity of the MVA-haemagglutinin-based H5N1 vaccine MVA-H5-sfMR in healthy individuals. In a single-centre, double-blind phase 1/2a study, young volunteers (aged 18-28 years) were randomly assigned with a computer-generated list in equal numbers to one of eight groups and were given one injection or two injections intramuscularly at an interval of 4 weeks of a standard dose (10(8) plaque forming units [pfu]) or a ten times lower dose (10(7) pfu) of the MVA-H5-sfMR (vector encoding the haemagglutinin gene of influenza A/Vietnam/1194/2004 virus [H5N1 subtype]) or MVA-F6-sfMR (empty vector) vaccine. Volunteers and physicians who examined and administered the vaccine were masked to vaccine assignment. Individuals who received the MVA-H5-sfMR vaccine were eligible for a booster immunisation 1 year after the first immunisation. Primary endpoint was safety. Secondary outcome was immunogenicity. The trial is registered with the Dutch Trial Register, number NTR3401. 79 of 80 individuals who were enrolled completed the study. No serious adverse events were identified. 11 individuals reported severe headache and lightheadedness, erythema nodosum, respiratory illness (accompanied by influenza-like symptoms), sore throat, or injection-site reaction. Most of the volunteers had one or more local (itch, pain, redness, and swelling) and systemic reactions (rise in body temperature, headache, myalgia, arthralgia, chills, malaise, and fatigue) after the first, second, and booster immunisations. Individuals who received the 10(7) dose had fewer systemic reactions. The MVA-H5-sfMR vaccine at 10(8) pfu induced significantly higher antibody responses after one and two immunisations than did 10(7) pfu when

  13. Comparative study of the hemagglutinin and neuraminidase genes of influenza A virus H3N2, H9N2, and H5N1 subtypes using bioinformatics techniques.

    Science.gov (United States)

    Ahn, Insung; Son, Hyeon S

    2007-07-01

    To investigate the genomic patterns of influenza A virus subtypes, such as H3N2, H9N2, and H5N1, we collected 1842 sequences of the hemagglutinin and neuraminidase genes from the NCBI database and parsed them into 7 categories: accession number, host species, sampling year, country, subtype, gene name, and sequence. The sequences that were isolated from the human, avian, and swine populations were extracted and stored in a MySQL database for intensive analysis. The GC content and relative synonymous codon usage (RSCU) values were calculated using JAVA codes. As a result, correspondence analysis of the RSCU values yielded the unique codon usage pattern (CUP) of each subtype and revealed no extreme differences among the human, avian, and swine isolates. H5N1 subtype viruses exhibited little variation in CUPs compared with other subtypes, suggesting that the H5N1 CUP has not yet undergone significant changes within each host species. Moreover, some observations may be relevant to CUP variation that has occurred over time among the H3N2 subtype viruses isolated from humans. All the sequences were divided into 3 groups over time, and each group seemed to have preferred synonymous codon patterns for each amino acid, especially for arginine, glycine, leucine, and valine. The bioinformatics technique we introduce in this study may be useful in predicting the evolutionary patterns of pandemic viruses.

  14. The nuclear export protein of H5N1 influenza A viruses recruits Matrix 1 (M1) protein to the viral ribonucleoprotein to mediate nuclear export.

    Science.gov (United States)

    Brunotte, Linda; Flies, Joe; Bolte, Hardin; Reuther, Peter; Vreede, Frank; Schwemmle, Martin

    2014-07-18

    In influenza A virus-infected cells, replication and transcription of the viral genome occurs in the nucleus. To be packaged into viral particles at the plasma membrane, encapsidated viral genomes must be exported from the nucleus. Intriguingly, the nuclear export protein (NEP) is involved in both processes. Although NEP stimulates viral RNA synthesis by binding to the viral polymerase, its function during nuclear export implicates interaction with viral ribonucleoprotein (vRNP)-associated M1. The observation that both interactions are mediated by the C-terminal moiety of NEP raised the question whether these two features of NEP are linked functionally. Here we provide evidence that the interaction between M1 and the vRNP depends on the NEP C terminus and its polymerase activity-enhancing property for the nuclear export of vRNPs. This suggests that these features of NEP are linked functionally. Furthermore, our data suggest that the N-terminal domain of NEP interferes with the stability of the vRNP-M1-NEP nuclear export complex, probably mediated by its highly flexible intramolecular interaction with the NEP C terminus. On the basis of our data, we propose a new model for the assembly of the nuclear export complex of Influenza A vRNPs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. Comparative epidemiology of human infections with avian influenza A H7N9 and H5N1 viruses in China: a population-based study of laboratory-confirmed cases.

    Science.gov (United States)

    Cowling, Benjamin J; Jin, Lianmei; Lau, Eric H Y; Liao, Qiaohong; Wu, Peng; Jiang, Hui; Tsang, Tim K; Zheng, Jiandong; Fang, Vicky J; Chang, Zhaorui; Ni, Michael Y; Zhang, Qian; Ip, Dennis K M; Yu, Jianxing; Li, Yu; Wang, Liping; Tu, Wenxiao; Meng, Ling; Wu, Joseph T; Luo, Huiming; Li, Qun; Shu, Yuelong; Li, Zhongjie; Feng, Zijian; Yang, Weizhong; Wang, Yu; Leung, Gabriel M; Yu, Hongjie

    2013-07-13

    The novel influenza A H7N9 virus emerged recently in mainland China, whereas the influenza A H5N1 virus has infected people in China since 2003. Both infections are thought to be mainly zoonotic. We aimed to compare the epidemiological characteristics of the complete series of laboratory-confirmed cases of both viruses in mainland China so far. An integrated database was constructed with information about demographic, epidemiological, and clinical variables of laboratory-confirmed cases of H7N9 (130 patients) and H5N1 (43 patients) that were reported to the Chinese Centre for Disease Control and Prevention until May 24, 2013. We described disease occurrence by age, sex, and geography, and estimated key epidemiological variables. We used survival analysis techniques to estimate the following distributions: infection to onset, onset to admission, onset to laboratory confirmation, admission to death, and admission to discharge. The median age of the 130 individuals with confirmed infection with H7N9 was 62 years and of the 43 with H5N1 was 26 years. In urban areas, 74% of cases of both viruses were in men, whereas in rural areas the proportions of the viruses in men were 62% for H7N9 and 33% for H5N1. 75% of patients infected with H7N9 and 71% of those with H5N1 reported recent exposure to poultry. The mean incubation period of H7N9 was 3·1 days and of H5N1 was 3·3 days. On average, 21 contacts were traced for each case of H7N9 in urban areas and 18 in rural areas, compared with 90 and 63 for H5N1. The fatality risk on admission to hospital was 36% (95% CI 26-45) for H7N9 and 70% (56-83%) for H5N1. The sex ratios in urban compared with rural cases are consistent with exposure to poultry driving the risk of infection--a higher risk in men was only recorded in urban areas but not in rural areas, and the increased risk for men was of a similar magnitude for H7N9 and H5N1. However, the difference in susceptibility to serious illness with the two different viruses

  16. The Intersection of Care Seeking and Clinical Capacity for Patients With Highly Pathogenic Avian Influenza A (H5N1) Virus in Indonesia: Knowledge and Treatment Practices of the Public and Physicians.

    Science.gov (United States)

    Kreslake, Jennifer M; Wahyuningrum, Yunita; Iuliano, Angela D; Storms, Aaron D; Lafond, Kathryn E; Mangiri, Amalya; Praptiningsih, Catharina Y; Safi, Basil; Uyeki, Timothy M; Storey, J Douglas

    2016-12-01

    Indonesia has the highest human mortality from highly pathogenic avian influenza (HPAI) A (H5N1) virus infection in the world. A survey of households (N=2520) measured treatment sources and beliefs among symptomatic household members. A survey of physicians (N=554) in various types of health care facilities measured knowledge, assessment and testing behaviors, and perceived clinical capacity. Households reported confidence in health care system capacity but infrequently sought treatment for potential HPAI H5N1 signs/symptoms. More clinicians were confident in their knowledge of diagnosis and treatment than in the adequacy of related equipment and resources at their facilities. Physicians expressed awareness of the HPAI H5N1 suspect case definition, yet expressed only moderate knowledge in questioning symptomatic patients about exposures. Self-reported likelihood of testing for HPAI H5N1 virus was high after learning of certain exposures. Knowledge of antiviral treatment was moderate, but it was higher among clinicians in puskesmas. Physicians in private outpatient clinics, the most heavily used facilities, reported the lowest confidence in their diagnostic and treatment capabilities. Educational campaigns can encourage recall of possible poultry exposure when patients are experiencing signs/symptoms and can raise awareness of the effectiveness of antivirals to drive people to seek health care. Clinicians may benefit from training regarding exposure assessment and referral procedures, particularly in private clinics. (Disaster Med Public Health Preparedness. 2016;10:838-847).

  17. Immune response in domestic ducks following intradermal delivery of inactivated vaccine against H5N1 highly pathogenic avian influenza virus adjuvanted with oligodeoxynucleotides containing CpG motifs.

    Science.gov (United States)

    Yuk, Seong-Su; Lee, Dong-Hun; Park, Jae-Keun; To, Eredene-Ochir; Kwon, Jung-Hoon; Noh, Jin-Yong; Gomis, Susantha; Song, Chang-Seon

    2015-08-01

    Ducks are a natural reservoir for H5N1 highly pathogenic avian influenza (HPAI) viruses, which produces a range of clinical outcomes from asymptomatic infections to severe disease with mortality. Vaccination against HPAI is one of the few methods available for controlling avian influenza virus (AIV) infection in domestic ducks; therefore, it is necessary to improve vaccine efficacy against HPAI in domestic ducks. However, few studies have focused on enhancing the immune response by testing alternative administration routes and adjuvants. While attempting to maximize the efficacy of a vaccine, it is important to select an appropriate vaccine delivery route and adjuvant to elicit an enhanced immune response. Although several studies have indicated that the vaccination of ducks against HPAI viruses has offered protection against lethal virus challenge, the immunogenicity of the vaccine still requires improvement. In this study, we characterized the immune response following a novel vaccination strategy against H5N1 HPAI virus in domestic ducks. Our novel intradermal delivery system and the application of the cytosine-phosphodiester-guanine (CpG) oligodeoxynucleotide (ODN) adjuvant allowed us to obtain information regarding the sustained vaccine immunity. Compared with the intramuscular route of vaccination, the intradermal route resulted in higher antibody titer as well as lower antibody deviation following secondary vaccination. In addition, the use of a CpG-ODN adjuvant had a dose-sparing effect on antibody titer. Furthermore, when a high dose of antigen was used, the CpG-ODN-adjuvanted vaccine maintained a high mean antibody titer. This data demonstrates that intradermal immunization combined with administration of CpG-ODN as an adjuvant may be a promising strategy for improving vaccine efficacy in domestic ducks. © 2015 Poultry Science Association Inc.

  18. Human influenza A (H5N1): a brief review and recommendations for travelers.

    Science.gov (United States)

    Hurtado, Timothy R

    2006-01-01

    Although avian influenza A (H5N1) is common in birds worldwide, it has only recently led to disease in humans. Humans who are infected with the disease (referred to as human influenza A [H5N1]) have a greater than 50% mortality rate. Currently there has not been documented sustained human-to-human transmission; however, should the virus mutate and make this possible, the world could experience an influenza pandemic. Probable risk factors for infection include slaughtering, defeathering, and butchering fowl; close contact with wild birds or caged poultry; ingestion of undercooked poultry products; direct contact with surfaces contaminated with poultry feces; and close contact with infected humans. Possible risk factors include swimming in or ingesting water contaminated with bird feces or dead birds and the use of unprocessed poultry feces as fertilizer. Clinically, early human influenza A (H5N1) resembles typical influenza illnesses, with fever and a preponderance of lower respiratory tract symptoms. Often, patients develop rapidly progressive respiratory failure and require ventilatory support. Treatment is primarily supportive care with the addition of antiviral medications. Currently, travelers to countries with both human and avian influenza A (H5N1) have a low risk of developing the disease. There are no current recommended travel restrictions. Travelers are advised to avoid contact with all birds, especially poultry; avoid surfaces contaminated with poultry feces; and avoid undercooked poultry products. The use of prophylactic antiviral medications is not recommended.

  19. Influenza A H5N1 and HIV co-infection: case report

    Directory of Open Access Journals (Sweden)

    Simmons Cameron

    2010-06-01

    Full Text Available Abstract Background The role of adaptive immunity in severe influenza is poorly understood. The occurrence of influenza A/H5N1 in a patient with HIV provided a rare opportunity to investigate this. Case Presentation A 30-year-old male was admitted on day 4 of influenza-like-illness with tachycardia, tachypnea, hypoxemia and bilateral pulmonary infiltrates. Influenza A/H5N1 and HIV tests were positive and the patient was treated with Oseltamivir and broad-spectrum antibiotics. Initially his condition improved coinciding with virus clearance by day 6. He clinically deteriorated as of day 10 with fever recrudescence and increasing neutrophil counts and died on day 16. His admission CD4 count was 100/μl and decreased until virus was cleared. CD8 T cells shifted to a CD27+CD28- phenotype. Plasma chemokine and cytokine levels were similar to those found previously in fatal H5N1. Conclusions The course of H5N1 infection was not notably different from other cases. Virus was cleared despite profound CD4 T cell depletion and aberrant CD8 T cell activation but this may have increased susceptibility to a fatal secondary infection.

  20. Role of position 627 of PB2 and the multibasic cleavage site of the hemagglutinin in the virulence of H5N1 avian influenza virus in chickens and ducks.

    Directory of Open Access Journals (Sweden)

    Karel A Schat

    Full Text Available Highly pathogenic H5N1 avian influenza viruses have caused major disease outbreaks in domestic and free-living birds with transmission to humans resulting in 59% mortality amongst 564 cases. The mutation of the amino acid at position 627 of the viral polymerase basic-2 protein (PB2 from glutamic acid (E in avian isolates to lysine (K in human isolates is frequently found, but it is not known if this change affects the fitness and pathogenicity of the virus in birds. We show here that horizontal transmission of A/Vietnam/1203/2004 H5N1 (VN/1203 virus in chickens and ducks was not affected by the change of K to E at PB2-627. All chickens died between 21 to 48 hours post infection (pi, while 70% of the ducks survived infection. Virus replication was detected in chickens within 12 hours pi and reached peak titers in spleen, lung and brain between 18 to 24 hours for both viruses. Viral antigen in chickens was predominantly in the endothelium, while in ducks it was present in multiple cell types, including neurons, myocardium, skeletal muscle and connective tissues. Virus replicated to a high titer in chicken thrombocytes and caused upregulation of TLR3 and several cell adhesion molecules, which may explain the rapid virus dissemination and location of viral antigen in endothelium. Virus replication in ducks reached peak values between 2 and 4 days pi in spleen, lung and brain tissues and in contrast to infection in chickens, thrombocytes were not involved. In addition, infection of chickens with low pathogenic VN/1203 caused neuropathology, with E at position PB2-627 causing significantly higher infection rates than K, indicating that it enhances virulence in chickens.

  1. Avian influenza viruses in humans.

    Science.gov (United States)

    Malik Peiris, J S

    2009-04-01

    Past pandemics arose from low pathogenic avian influenza (LPAI) viruses. In more recent times, highly pathogenic avian influenza (HPAI) H5N1, LPAI H9N2 and both HPAI and LPAI H7 viruses have repeatedly caused zoonotic disease in humans. Such infections did not lead to sustained human-to-human transmission. Experimental infection of human volunteers and seroepidemiological studies suggest that avian influenza viruses of other subtypes may also infect humans. Viruses of the H7 subtype appear to have a predilection to cause conjunctivitis and influenza-like illness (ILI), although HPAI H7N7 virus has also caused fatal respiratory disease. Low pathogenic H9N2 viruses have caused mild ILI and its occurrence may be under-recognised for this reason. In contrast, contemporary HPAI H5N1 viruses are exceptional in their virulence for humans and differ from human seasonal influenza viruses in their pathogenesis. Patients have a primary viral pneumonia progressing to acute respiratory distress syndrome (ARDS) and multiple organ dysfunction syndrome. Over 380 human cases have been confirmed to date, with an overall case fatality of 63%. The zoonotic transmission of avian influenza is a rare occurrence, butthe greater public health concern is the adaptation of such viruses to efficient human transmission, which could lead to a pandemic. A better understanding of the ecology of avian influenza viruses and the biological determinants of transmissibility and pathogenicity in humans is important for pandemic preparedness.

  2. Differences in pathogenicity, response to vaccination, and innate immune responses in different types of ducks infected with a virulent H5N1 highly pathogenic avian influenza virus from Vietnam.

    Science.gov (United States)

    Cagle, Caran; Wasilenko, Jamie; Adams, Sean C; Cardona, Carol J; To, Thanh Long; Nguyen, Tung; Spackman, Erica; Suarez, David L; Smith, Diane; Shepherd, Eric; Roth, Jason; Pantin-Jackwood, Mary J

    2012-09-01

    In a previous study, we found clear differences in pathogenicity and response to vaccination against H5N1 highly pathogenic avian influenza (HPAI; HA dade 2.3.4) between Pekin (Anas platyrhynchos var. domestica) and Muscovy (Cairina moschata) ducks vaccinated using a commercial inactivated vaccine (Re-1). The objective of the present study was to further investigate the pathogenicity of H5N1 HPAI viruses in different species of ducks by examining clinical signs and innate immune responses to infection with a different strain of H5N1 HPAI virus (HA clade 1) in two domestic ducks, Pekin and Muscovy, and one wild-type duck, mallard (Anas platyrhynchos). Protection conferred by vaccination using the Re-1 vaccine against infection with this virus was also compared between Pekin and Muscovy ducks. Differences in pathogenicity were observed among the virus-infected ducks, as the Muscovy ducks died 2 days earlier than did the Pekin and mallard ducks, and they presented more-severe neurologic signs. Conversely, the Pekin and mallard ducks had significantly higher body temperatures at 2 days postinfection (dpi) than did the Muscovy ducks, indicating possible differences in innate immune responses. However, similar expression of innate immune-related genes was found in the spleens of virus-infected ducks at this time point. In all three duck species, there was up-regulation of IFN-alpha, IFN-gamma, IL-6, CCL19, RIG-I, and MHC class I and down-regulation of MHC class II, but variable expression of IL-18 and TLR7. As in our previous study, vaccinated Muscovy ducks showed less protection against virus infection than did Pekin ducks, as evidenced by the higher mortality and higher number of Muscovy ducks shedding virus when compared to Pekin ducks. In conclusion, infection with an H5N1 HPAI virus produced a systemic infection with high mortality in all three duck species; however, the disease was more severe in Muscovy ducks, which also had a poor response to vaccination. The

  3. In silico approach towards H5N1 virus protein and transcriptomics ...

    African Journals Online (AJOL)

    H5N1 influenza A virus is a serious threat to human population. With a considerable mortality rate, strategies for coping with the infection are being developed. Our research group and some others investigated the potential therapeutic and preventive measures for tackling H5N1 infections. Protein based and transcriptomics ...

  4. Dual Infection of Novel Influenza Viruses A/H1N1 and A/H3N2 in a Cluster of Cambodian Patients

    Science.gov (United States)

    2011-01-01

    influenza viruses as well as the avian influenza virus A/H5N1...on full genome sequencing. This incident confirms dual influenza virus infections and highlights the risk of zoonotic and seasonal influenza viruses ...North American swine influenza viruses , North American avian influenza viruses , human influenza viruses , and a Eurasian swine influenza virus . 18

  5. Advances and Future Challenges in Recombinant Adenoviral Vectored H5N1 Influenza Vaccines

    Directory of Open Access Journals (Sweden)

    Jianfeng Zhang

    2012-11-01

    Full Text Available The emergence of a highly pathogenic avian influenza virus H5N1 has increased the potential for a new pandemic to occur. This event highlights the necessity for developing a new generation of influenza vaccines to counteract influenza disease. These vaccines must be manufactured for mass immunization of humans in a timely manner. Poultry should be included in this policy, since persistent infected flocks are the major source of avian influenza for human infections. Recombinant adenoviral vectored H5N1 vaccines are an attractive alternative to the currently licensed influenza vaccines. This class of vaccines induces a broadly protective immunity against antigenically distinct H5N1, can be manufactured rapidly, and may allow mass immunization of human and poultry. Recombinant adenoviral vectors derived from both human and non-human adenoviruses are currently being investigated and appear promising both in nonclinical and clinical studies. This review will highlight the current status of various adenoviral vectored H5N1 vaccines and will outline novel approaches for the future.

  6. Pandemic influenza – including a risk assessment of H5N1

    Science.gov (United States)

    Taubenberger, J.K.; Morens, D.M.

    2009-01-01

    Summary Influenza pandemics and epidemics have apparently occurred since at least the Middle Ages. When pandemics appear, 50% or more of an affected population can be infected in a single year, and the number of deaths caused by influenza can dramatically exceed what is normally expected. Since 1500, there appear to have been 13 or more influenza pandemics. In the past 120 years there were undoubted pandemics in 1889, 1918, 1957, 1968, and 1977. Although most experts believe we will face another influenza pandemic, it is impossible to predict when it will appear, where it will originate, or how severe it will be. Nor is there agreement about the subtype of influenza virus most likely to cause the next pandemic. The continuing spread of H5N1 highly pathogenic avian influenza viruses has heightened interest in pandemic prediction. Despite uncertainties in the historical record of the pre-virology era, study of previous pandemics may help guide future pandemic planning and lead to a better understanding of the complex ecobiology underlying the formation of pandemic strains of influenza A viruses. PMID:19618626

  7. Pandemic influenza--including a risk assessment of H5N1.

    Science.gov (United States)

    Taubenberger, J K; Morens, D M

    2009-04-01

    Influenza pandemics and epidemics have apparently occurred since at least the Middle Ages. When pandemics appear, 50% or more of an affected population can be infected in a single year, and the number of deaths caused by influenza can dramatically exceed what is normally expected. Since 1500, there appear to have been 13 or more influenza pandemics. In the past 120 years there were undoubted pandemics in 1889, 1918, 1957, 1968, and 1977. Although most experts believe we will face another influenza pandemic, it is impossible to predict when it will appear, where it will originate, or how severe it will be. Nor is there agreement about the subtype of influenza virus most likely to cause the next pandemic. The continuing spread of H5N1 highly pathogenic avian influenza viruses has heightened interest in pandemic prediction. Despite uncertainties in the historical record of the pre-virology era, study of previous pandemics may help guide future pandemic planning and lead to a better understanding of the complex ecobiology underlying the formation of pandemic strains of influenza A viruses.

  8. First human case of avian influenza A (H5N6 in Yunnan province, China

    Directory of Open Access Journals (Sweden)

    Jibo He

    2015-08-01

    Full Text Available Objective: To report clinical, virological, and epidemiological features of the first death caused by a H5N6 avian influenza virus in Yunnan Province, China. Method: The case was described in clinical expression, chest radiography, blood test and treatment. Real-time RT-PCR was used to detect H5N6 virus RNA in clinical and environment samples. Epidemiological investigation was performed including case exposure history determinant, close contacts follow up, and environment sample collection. Results: The patient initially developed sore throat and coughs on 27 January 2015. The disease progressed to severe pneumonia, multiple organ dysfunction syndrome, and acute respiratory distress syndrome. And the patient died on 6 February. A highly pathogenic avian influenza A H5N6 virus was isolated from the tracheal aspirate specimen of the patient. The viral genome analyses revealed that the H5 hemmagglutinin gene belongs to 2.3.4.4 clade. Epidemiological investigation showed that the patient had exposure to wild bird. All close contacts of the patient did not present the same disease in seven consecutive days. A high H5 positive rate was detected in environmental samples from local live poultry markets. Conclusion: The findings suggest that studies on the source of the virus, transmission models, serologic investigations, vaccines, and enhancing surveillance in both humans and birds are necessary.

  9. Wild bird surveillance for highly pathogenic avian influenza H5 in North America

    Science.gov (United States)

    Flint, Paul L.; Pearce, John M.; Franson, J. Christian; Derksen, Dirk V.

    2015-01-01

    It is unknown how the current Asian origin highly pathogenic avian influenza H5 viruses arrived, but these viruses are now poised to become endemic in North America. Wild birds harbor these viruses and have dispersed them at regional scales. What is unclear is how the viruses may be moving from the wild bird reservoir into poultry holdings. Active surveillance of live wild birds is likely the best way to determine the true distribution of these viruses. We also suggest that sampling be focused on regions with the greatest risk for poultry losses and attempt to define the mechanisms of transfer to enhance biosecurity. Responding to the recent outbreaks of highly pathogenic avian influenza in North America requires an efficient plan with clear objectives and potential management outcomes.

  10. Anti-viral properties and mode of action of standardized Echinacea purpurea extract against highly pathogenic avian Influenza virus (H5N1, H7N7 and swine-origin H1N1 (S-OIV

    Directory of Open Access Journals (Sweden)

    Schoop Roland

    2009-11-01

    Full Text Available Abstract Background Influenza virus (IV infections are a major threat to human welfare and animal health worldwide. Anti-viral therapy includes vaccines and a few anti-viral drugs. However vaccines are not always available in time, as demonstrated by the emergence of the new 2009 H1N1-type pandemic strain of swine origin (S-OIV in April 2009, and the acquisition of resistance to neuraminidase inhibitors such as Tamiflu® (oseltamivir is a potential problem. Therefore the prospects for the control of IV by existing anti-viral drugs are limited. As an alternative approach to the common anti-virals we studied in more detail a commercial standardized extract of the widely used herb Echinacea purpurea (Echinaforce®, EF in order to elucidate the nature of its anti-IV activity. Results Human H1N1-type IV, highly pathogenic avian IV (HPAIV of the H5- and H7-types, as well as swine origin IV (S-OIV, H1N1, were all inactivated in cell culture assays by the EF preparation at concentrations ranging from the recommended dose for oral consumption to several orders of magnitude lower. Detailed studies with the H5N1 HPAIV strain indicated that direct contact between EF and virus was required, prior to infection, in order to obtain maximum inhibition in virus replication. Hemagglutination assays showed that the extract inhibited the receptor binding activity of the virus, suggesting that the extract interferes with the viral entry into cells. In sequential passage studies under treatment in cell culture with the H5N1 virus no EF-resistant variants emerged, in contrast to Tamiflu®, which produced resistant viruses upon passaging. Furthermore, the Tamiflu®-resistant virus was just as susceptible to EF as the wild type virus. Conclusion As a result of these investigations, we believe that this standard Echinacea preparation, used at the recommended dose for oral consumption, could be a useful, readily available and affordable addition to existing control options

  11. Unusually High Mortality in Waterfowl Caused by Highly Pathogenic Avian Influenza A(H5N1) in Bangladesh

    DEFF Research Database (Denmark)

    Haider, Najmul; Sturm-Ramirez, K.; Khan, S. U.

    2017-01-01

    a survey in three of these villages to identify suspected human influenza-like illness cases and collected nasopharyngeal and throat swabs. We tested all swabs by real-time RT-PCR, sequenced cultured viruses, and examined tissue samples by histopathology and immunohistochemistry to detect and characterize...... and immunohistochemistry staining of avian influenza viral antigens were recognized in the brain, pancreas and intestines of ducks and chickens. We identified ten human cases showing signs compatible with influenza-like illness; four were positive for influenza A/H3; however, none were positive for influenza A/H5......Mortality in ducks and geese caused by highly pathogenic avian influenza A(H5N1) infection had not been previously identified in Bangladesh. In June-July 2011, we investigated mortality in ducks, geese and chickens with suspected H5N1 infection in a north-eastern district of the country to identify...

  12. Prophylactic and therapeutic efficacy of human monoclonal antibodies against H5N1 influenza.

    Directory of Open Access Journals (Sweden)

    Cameron P Simmons

    2007-05-01

    Full Text Available New prophylactic and therapeutic strategies to combat human infections with highly pathogenic avian influenza (HPAI H5N1 viruses are needed. We generated neutralizing anti-H5N1 human monoclonal antibodies (mAbs and tested their efficacy for prophylaxis and therapy in a murine model of infection.Using Epstein-Barr virus we immortalized memory B cells from Vietnamese adults who had recovered from infections with HPAI H5N1 viruses. Supernatants from B cell lines were screened in a virus neutralization assay. B cell lines secreting neutralizing antibodies were cloned and the mAbs purified. The cross-reactivity of these antibodies for different strains of H5N1 was tested in vitro by neutralization assays, and their prophylactic and therapeutic efficacy in vivo was tested in mice. In vitro, mAbs FLA3.14 and FLD20.19 neutralized both Clade I and Clade II H5N1 viruses, whilst FLA5.10 and FLD21.140 neutralized Clade I viruses only. In vivo, FLA3.14 and FLA5.10 conferred protection from lethality in mice challenged with A/Vietnam/1203/04 (H5N1 in a dose-dependent manner. mAb prophylaxis provided a statistically significant reduction in pulmonary virus titer, reduced associated inflammation in the lungs, and restricted extrapulmonary dissemination of the virus. Therapeutic doses of FLA3.14, FLA5.10, FLD20.19, and FLD21.140 provided robust protection from lethality at least up to 72 h postinfection with A/Vietnam/1203/04 (H5N1. mAbs FLA3.14, FLD21.140 and FLD20.19, but not FLA5.10, were also therapeutically active in vivo against the Clade II virus A/Indonesia/5/2005 (H5N1.These studies provide proof of concept that fully human mAbs with neutralizing activity can be rapidly generated from the peripheral blood of convalescent patients and that these mAbs are effective for the prevention and treatment of H5N1 infection in a mouse model. A panel of neutralizing, cross-reactive mAbs might be useful for prophylaxis or adjunctive treatment of human cases of H5N1

  13. Efficacy of two H5N9-inactivated vaccines against challenge with a recent H5N1 highly pathogenic avian influenza isolate from a chicken in Thailand.

    Science.gov (United States)

    Bublot, Michel; Le Gros, François-Xavier; Nieddu, Daniela; Pritchard, Nikki; Mickle, Thomas R; Swayne, David E

    2007-03-01

    The objective of this study was to compare the efficacy of two avian influenza (AI) H5-inactivated vaccines containing either an American (A/turkey/Wisconsin/68 H5N9; H5N9-WI) or a Eurasian isolate (A/chicken/Italy/22A/98 H5N9; H5N9-It). Three-week-old specific pathogen-free chickens were vaccinated once and challenged 3 wk later with a H5N1 highly pathogenic AI (HPAI) virus isolated from a chicken in Thailand in 2004. All unvaccinated challenged birds died within 2 days, whereas 90% and 100% of chickens vaccinated with H5N9-WI and H5N9-It, respectively, were protected against morbidity and mortality. Both vaccines prevented cloacal shedding and significantly reduced oral shedding of the challenge HPAI virus. Additional chickens (vaccinated or unvaccinated) were placed in contact with the directly challenged birds 18 hr after challenge. All unvaccinated chickens in contact with unvaccinated challenged birds died within 3 days after contact, whereas unvaccinated chickens in contact with vaccinated challenged birds either showed a significantly delayed mortality or did not become infected. All vaccinated contacts were protected against clinical signs, and most chickens did not shed detectable amount of HPAI virus. Altogether, these data indicate that both vaccines protected very well against morbidity and mortality and reduced or prevented shedding induced by direct or contact exposure to Asian H5N1 HPAI virus.

  14. Tissue tropism of highly pathogenic avian influenza virus subtype H5N1 in naturally infected mute swans (Cygnus Olor ), domestic geese (Aser Anser var. domestica), pekin ducks (Anas platyrhynchos) and mulard ducks ( Cairina moschata x anas platyrhynchos).

    Science.gov (United States)

    Szeredi, Levente; Dán, Adám; Pálmai, Nimród; Ursu, Krisztina; Bálint, Adám; Szeleczky, Zsófia; Ivanics, Eva; Erdélyi, Károly; Rigó, Dóra; Tekes, Lajos; Glávits, Róbert

    2010-03-01

    The 2006 epidemic due to highly pathogenic avian influenza virus (HPAIV) subtype H5N1 in Hungary caused the most severe losses in waterfowl which were, according to the literature at the time, supposed to be the most resistant to this pathogen. The presence of pathological lesions and the amount of viral antigen were quantified by gross pathology, histopathology and immunohistochemistry (IHC) in the organs of four waterfowl species [mute swans (n = 10), domestic geese (n = 6), mulard ducks (n = 6) and Pekin ducks (n = 5)] collected during the epidemic. H5N1 subtype HPAIV was isolated from all birds examined. Quantitative real-time reverse transcriptase-polymerase chain reaction (qRRT-PCR) was also applied on a subset of samples [domestic geese (n = 3), mulard (n = 4) and Pekin duck (n = 4)] in order to compare its sensitivity with IHC. Viral antigen was detected by IHC in all cases. However, the overall presence of viral antigen in tissue samples was quite variable: virus antigen was present in 56/81 (69%) swan, 22/38 (58%) goose, 28/46 (61%) mulard duck and 5/43 (12%) Pekin duck tissue samples. HPAIV subtype H5N1 was detected by qRRT-PCR in all birds examined, in 19/19 (100%) goose, 7/28 (25%) mulard duck and 12/28 (43%) Pekin duck tissue samples. As compared to qRRTPCR, the IHC was less sensitive in geese and Pekin ducks but more sensitive in mulard ducks. The IHC was consistently positive above 4.31 log10 copies/reaction but it gave very variable results below that level. Neurotropism of the isolated virus strains was demonstrated by finding the largest amount of viral antigen and the highest average RNA load in the brain in all four waterfowl species examined.

  15. Using knowledge fusion to analyze avian influenza H5N1 in East and Southeast Asia.

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    Erjia Ge

    Full Text Available Highly pathogenic avian influenza (HPAI H5N1, a disease associated with high rates of mortality in infected human populations, poses a serious threat to public health in many parts of the world. This article reports findings from a study aimed at improving our understanding of the spatial pattern of the highly pathogenic avian influenza, H5N1, risk in East-Southeast Asia where the disease is both persistent and devastating. Though many disciplines have made important contributions to our understanding of H5N1, it remains a challenge to integrate knowledge from different disciplines. This study applies genetic analysis that identifies the evolution of the H5N1 virus in space and time, epidemiological analysis that determines socio-ecological factors associated with H5N1 occurrence, and statistical analysis that identifies outbreak clusters, and then applies a methodology to formally integrate the findings of the three sets of methodologies. The present study is novel in two respects. First it makes the initiative attempt to use genetic sequences and space-time data to create a space-time phylogenetic tree to estimate and map the virus' ability to spread. Second, by integrating the results we are able to generate insights into the space-time occurrence and spread of H5N1 that we believe have a higher level of corroboration than is possible when analysis is based on only one methodology. Our research identifies links between the occurrence of H5N1 by area and a set of socio-ecological factors including altitude, population density, poultry density, and the shortest path distances to inland water, coastlines, migrating routes, railways, and roads. This study seeks to lay a solid foundation for the interdisciplinary study of this and other influenza outbreaks. It will provide substantive information for containing H5N1 outbreaks.

  16. Using knowledge fusion to analyze avian influenza H5N1 in East and Southeast Asia.

    Science.gov (United States)

    Ge, Erjia; Haining, Robert; Li, Chi Pang; Yu, Zuguo; Waye, Miu Yee; Chu, Ka Hou; Leung, Yee

    2012-01-01

    Highly pathogenic avian influenza (HPAI) H5N1, a disease associated with high rates of mortality in infected human populations, poses a serious threat to public health in many parts of the world. This article reports findings from a study aimed at improving our understanding of the spatial pattern of the highly pathogenic avian influenza, H5N1, risk in East-Southeast Asia where the disease is both persistent and devastating. Though many disciplines have made important contributions to our understanding of H5N1, it remains a challenge to integrate knowledge from different disciplines. This study applies genetic analysis that identifies the evolution of the H5N1 virus in space and time, epidemiological analysis that determines socio-ecological factors associated with H5N1 occurrence, and statistical analysis that identifies outbreak clusters, and then applies a methodology to formally integrate the findings of the three sets of methodologies. The present study is novel in two respects. First it makes the initiative attempt to use genetic sequences and space-time data to create a space-time phylogenetic tree to estimate and map the virus' ability to spread. Second, by integrating the results we are able to generate insights into the space-time occurrence and spread of H5N1 that we believe have a higher level of corroboration than is possible when analysis is based on only one methodology. Our research identifies links between the occurrence of H5N1 by area and a set of socio-ecological factors including altitude, population density, poultry density, and the shortest path distances to inland water, coastlines, migrating routes, railways, and roads. This study seeks to lay a solid foundation for the interdisciplinary study of this and other influenza outbreaks. It will provide substantive information for containing H5N1 outbreaks.

  17. Cross-protection against lethal H5N1 challenge ferrets with an adjuvanted pandemic influenza vaccine

    NARCIS (Netherlands)

    B. Baras (Benoît); K.J. Stittelaar (Koert); J.H. Simon (James); R.J.M.M. Thoolen (Robert); S.P. Mossman (Sally); F.H. Pistoor (Frank); G. van Amerongen (Geert); M.A. Wettendorff (Martine); E. Hanon (Emmanuel); A.D.M.E. Osterhaus (Albert)

    2008-01-01

    textabstractBackground. Unprecedented spread between birds and mammals of highly pathogenic avian influenza viruses (HPAI) of the H5N1 subtype has resulted in hundreds of human infections with a high fatality rate. This has highlighted the urgent need for the development of H5N1 vaccines that can be

  18. Antibody titer has positive predictive value for vaccine protection against challenge with natural antigenic-drift variants of H5N1 high-pathogenicity avian influenza viruses from Indonesia

    Science.gov (United States)

    Beginning with Hong Kong in 2002, vaccines have been used as part of an integrated control strategy in 14 countries/regions to protect poultry against H5N1 high pathogenicity avian influenza (HPAI). H5N1 HPAI was first reported in Indonesia in 2003 and vaccination was initiated the following year. ...

  19. 2.1 Natural History of Highly Pathogenic Avian Influenza H5N1

    Science.gov (United States)

    Sonnberg, Stephanie; Webby, Richard J.; Webster, Robert G.

    2013-01-01

    The ecology of highly pathogenic avian influenza (HPAI) H5N1 has significantly changed from sporadic outbreaks in terrestrial poultry to persistent circulation in terrestrial and aquatic poultry and potentially in wild waterfowl. A novel genotype of HPAI H5N1 arose in 1996 in southern China and through ongoing mutation, reassortment, and natural selection, has diverged into distinct lineages and expanded into multiple reservoir hosts. The evolution of Goose/Guangdong-lineage highly pathogenic H5N1 viruses is ongoing: while stable interactions exist with some reservoir hosts, these viruses are continuing to evolve and adapt to others, and pose an un-calculable risk to sporadic hosts, including humans. PMID:23735535

  20. Pre-clinical evaluation of a replication-competent recombinant adenovirus serotype 4 vaccine expressing influenza H5 hemagglutinin.

    Science.gov (United States)

    Alexander, Jeff; Ward, Simone; Mendy, Jason; Manayani, Darly J; Farness, Peggy; Avanzini, Jenny B; Guenther, Ben; Garduno, Fermin; Jow, Lily; Snarsky, Victoria; Ishioka, Glenn; Dong, Xin; Vang, Lo; Newman, Mark J; Mayall, Tim

    2012-01-01

    Influenza virus remains a significant health and social concern in part because of newly emerging strains, such as avian H5N1 virus. We have developed a prototype H5N1 vaccine using a recombinant, replication-competent Adenovirus serotype 4 (Ad4) vector, derived from the U.S. military Ad4 vaccine strain, to express the hemagglutinin (HA) gene from A/Vietnam/1194/2004 influenza virus (Ad4-H5-Vtn). Our hypothesis is that a mucosally-delivered replicating Ad4-H5-Vtn recombinant vector will be safe and induce protective immunity against H5N1 influenza virus infection and disease pathogenesis. The Ad4-H5-Vtn vaccine was designed with a partial deletion of the E3 region of Ad4 to accommodate the influenza HA gene. Replication and growth kinetics of the vaccine virus in multiple human cell lines indicated that the vaccine virus is attenuated relative to the wild type virus. Expression of the HA transgene in infected cells was documented by flow cytometry, western blot analysis and induction of HA-specific antibody and cellular immune responses in mice. Of particular note, mice immunized intranasally with the Ad4-H5-Vtn vaccine were protected against lethal H5N1 reassortant viral challenge even in the presence of pre-existing immunity to the Ad4 wild type virus. Several non-clinical attributes of this vaccine including safety, induction of HA-specific humoral and cellular immunity, and efficacy were demonstrated using an animal model to support Phase 1 clinical trial evaluation of this new vaccine.

  1. Pre-clinical evaluation of a replication-competent recombinant adenovirus serotype 4 vaccine expressing influenza H5 hemagglutinin.

    Directory of Open Access Journals (Sweden)

    Jeff Alexander

    Full Text Available Influenza virus remains a significant health and social concern in part because of newly emerging strains, such as avian H5N1 virus. We have developed a prototype H5N1 vaccine using a recombinant, replication-competent Adenovirus serotype 4 (Ad4 vector, derived from the U.S. military Ad4 vaccine strain, to express the hemagglutinin (HA gene from A/Vietnam/1194/2004 influenza virus (Ad4-H5-Vtn. Our hypothesis is that a mucosally-delivered replicating Ad4-H5-Vtn recombinant vector will be safe and induce protective immunity against H5N1 influenza virus infection and disease pathogenesis.The Ad4-H5-Vtn vaccine was designed with a partial deletion of the E3 region of Ad4 to accommodate the influenza HA gene. Replication and growth kinetics of the vaccine virus in multiple human cell lines indicated that the vaccine virus is attenuated relative to the wild type virus. Expression of the HA transgene in infected cells was documented by flow cytometry, western blot analysis and induction of HA-specific antibody and cellular immune responses in mice. Of particular note, mice immunized intranasally with the Ad4-H5-Vtn vaccine were protected against lethal H5N1 reassortant viral challenge even in the presence of pre-existing immunity to the Ad4 wild type virus.Several non-clinical attributes of this vaccine including safety, induction of HA-specific humoral and cellular immunity, and efficacy were demonstrated using an animal model to support Phase 1 clinical trial evaluation of this new vaccine.

  2. Influenza A H5N1 immigration is filtered out at some international borders.

    Directory of Open Access Journals (Sweden)

    Robert G Wallace

    2008-02-01

    Full Text Available Geographic spread of highly pathogenic influenza A H5N1, the bird flu strain, appears a necessary condition for accelerating the evolution of a related human-to-human infection. As H5N1 spreads the virus diversifies in response to the variety of socioecological environments encountered, increasing the chance a human infection emerges. Genetic phylogenies have for the most part provided only qualitative evidence that localities differ in H5N1 diversity. For the first time H5N1 variation is quantified across geographic space.We constructed a statistical phylogeography of 481 H5N1 hemagglutinin genetic sequences from samples collected across 28 Eurasian and African localities through 2006. The MigraPhyla protocol showed southern China was a source of multiple H5N1 strains. Nested clade analysis indicated H5N1 was widely dispersed across southern China by both limited dispersal and long distance colonization. The UniFrac metric, a measure of shared phylogenetic history, grouped H5N1 from Indonesia, Japan, Thailand and Vietnam with those from southeastern Chinese provinces engaged in intensive international trade. Finally, H5N1's accumulative phylogenetic diversity was greatest in southern China and declined beyond. The gradient was interrupted by areas of greater and lesser phylogenetic dispersion, indicating H5N1 migration was restricted at some geopolitical borders. Thailand and Vietnam, just south of China, showed significant phylogenetic clustering, suggesting newly invasive H5N1 strains have been repeatedly filtered out at their northern borders even as both countries suffered recurring outbreaks of endemic strains. In contrast, Japan, while successful in controlling outbreaks, has been subjected to multiple introductions of the virus.The analysis demonstrates phylogenies can provide local health officials with more than hypotheses about relatedness. Pathogen dispersal, the functional relationships among disease ecologies across localities, and

  3. Construction and heterologous expression of a truncated Haemagglutinin (HA) protein from the avian influenza virus H5N1 in Escherichia coli.

    Science.gov (United States)

    Chee Wei, T; Nurul Wahida, A G; Shaharum, S

    2014-12-01

    Malaysia first reported H5N1 poultry case in 2004 and subsequently outbreak in poultry population in 2007. Here, a recombinant gene encoding of peptide epitopes, consisting fragments of HA1, HA2 and a polybasic cleavage site of H5N1 strain Malaysia, was amplified and cloned into pET-47b(+) bacterial expression vector. DNA sequencing and alignment analysis confirmed that the gene had no alteration and in-frame to the vector. Then, His-tagged truncated HA protein was expressed in Escherichia coli BL21 (DE3) under 1 mM IPTG induction. The protein expression was optimized under a time-course induction study and further purified using Ni-NTA agarose under reducing condition. Migration size of protein was detected at 15 kDa by Western blot using anti-His tag monoclonal antibody and demonstrated no discrepancy compared to its calculated molecular weight.

  4. Homosubtypic and heterosubtypic antibodies against highly pathogenic avian influenza H5N1 recombinant proteins in H5N1 survivors and non-H5N1 subjects.

    Science.gov (United States)

    Noisumdaeng, Pirom; Pooruk, Phisanu; Prasertsopon, Jarunee; Assanasen, Susan; Kitphati, Rungrueng; Auewarakul, Prasert; Puthavathana, Pilaipan

    2014-04-01

    Six recombinant vaccinia viruses containing HA, NA, NP, M or NS gene insert derived from a highly pathogenic avian influenza H5N1 virus, and the recombinant vaccinia virus harboring plasmid backbone as the virus control were constructed. The recombinant proteins were characterized for their expression and subcellular locations in TK(-) cells. Antibodies to the five recombinant proteins were detected in all 13 sequential serum samples collected from four H5N1 survivors during four years of follow-up; and those directed to rVac-H5 HA and rVac-NA proteins were found in higher titers than those directed to the internal proteins as revealed by indirect immunofluorescence assay. Although all 28 non-H5N1 subjects had no neutralizing antibodies against H5N1 virus, they did have cross-reactive antibodies to those five recombinant proteins. A significant increase in cross-reactive antibody titer to rVac-H5 HA and rVac-NA was found in paired blood samples from patients infected with the 2009 pandemic virus. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. Avian influenza H5N1 viral and bird migration networks in Asia

    Science.gov (United States)

    Tian, Huaivu; Zhou, Sen; Dong, Lu; Van Boeckel, Thomas P.; Cui, Yujun; Newman, Scott H.; Takekawa, John Y.; Prosser, Diann J.; Xiao, Xiangming; Wu, Yarong; Cazelles, Bernard; Huang, Shanqian; Yang, Ruifu; Grenfell, Bryan T.; Xu, Bing

    2015-01-01

    The spatial spread of the highly pathogenic avian influenza virus H5N1 and its long-term persistence in Asia have resulted in avian influenza panzootics and enormous economic losses in the poultry sector. However, an understanding of the regional long-distance transmission and seasonal patterns of the virus is still lacking. In this study, we present a phylogeographic approach to reconstruct the viral migration network. We show that within each wild fowl migratory flyway, the timing of H5N1 outbreaks and viral migrations are closely associated, but little viral transmission was observed between the flyways. The bird migration network is shown to better reflect the observed viral gene sequence data than other networks and contributes to seasonal H5N1 epidemics in local regions and its large-scale transmission along flyways. These findings have potentially far-reaching consequences, improving our understanding of how bird migration drives the periodic reemergence of H5N1 in Asia.

  6. Transforming growth factor-β: activation by neuraminidase and role in highly pathogenic H5N1 influenza pathogenesis.

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    Christina M Carlson

    2010-10-01

    Full Text Available Transforming growth factor-beta (TGF-β, a multifunctional cytokine regulating several immunologic processes, is expressed by virtually all cells as a biologically inactive molecule termed latent TGF-β (LTGF-β. We have previously shown that TGF-β activity increases during influenza virus infection in mice and suggested that the neuraminidase (NA protein mediates this activation. In the current study, we determined the mechanism of activation of LTGF-β by NA from the influenza virus A/Gray Teal/Australia/2/1979 by mobility shift and enzyme inhibition assays. We also investigated whether exogenous TGF-β administered via a replication-deficient adenovirus vector provides protection from H5N1 influenza pathogenesis and whether depletion of TGF-β during virus infection increases morbidity in mice. We found that both the influenza and bacterial NA activate LTGF-β by removing sialic acid motifs from LTGF-β, each NA being specific for the sialic acid linkages cleaved. Further, NA likely activates LTGF-β primarily via its enzymatic activity, but proteases might also play a role in this process. Several influenza A virus subtypes (H1N1, H1N2, H3N2, H5N9, H6N1, and H7N3 except the highly pathogenic H5N1 strains activated LTGF-β in vitro and in vivo. Addition of exogenous TGF-β to H5N1 influenza virus-infected mice delayed mortality and reduced viral titers whereas neutralization of TGF-β during H5N1 and pandemic 2009 H1N1 infection increased morbidity. Together, these data show that microbe-associated NAs can directly activate LTGF-β and that TGF-β plays a pivotal role protecting the host from influenza pathogenesis.

  7. Seroprevalence survey of avian influenza A (H5) in wild migratory birds in Yunnan Province, Southwestern China.

    Science.gov (United States)

    Chang, Hua; Dai, Feiyan; Liu, Zili; Yuan, Feizhou; Zhao, Siyue; Xiang, Xun; Zou, Fengcai; Zeng, Bangquan; Fan, Yating; Duan, Gang

    2014-02-03

    Highly pathogenic avian influenza virus (HPAIV) is a highly contagious disease which is a zoonotic pathogen of significant economic and public health concern. The outbreaks caused by HPAIV H5N1 of Asian origin have caused animal and human disease and mortality in several countries of Southeast Asia, such as Bangladesh, Cambodia, China, India, Indonesia, Laos, Myanmar, Thailand and Viet Nam. For the first time since 1961, this HPAIV has also caused extensive mortality in wild birds and has sparked debate of the role wild birds have played in the spread of this virus. Other than confirmed mortality events, little is known of this virus in wild birds. There is no report on the seroprevalence of avian influenza H5 infection in wild migratory birds in Yunnan Province. In this study we examined live wild birds in Yunnan Province for H5 specific antibody to better understand the occurrence of this disease in free living birds. Sera from 440 wild birds were collected from in Kunming and Northern Ailaoshan of Yunnan Province, Southwestern China, and assayed for H5 antibodies using the hemagglutination inhibition (HI) assays. The investigation revealed that the seroprevalence of avian influenza H5 was as following: Ciconiiformes 2.6%, Strigiformes 13.04%, Passeriformes 20%, Cuculiformes 21.74%, Gruiformes 0%, Columbiformes 0%, Charadriiformes 0% and Coraciiformes 0%. Statistical analyses showed that there was a significant difference of prevalence between the orders (P avian influenza H5 antibodies were detected in 23 of 440 (5.23%) sera. Mean HI titer 23 positive sera against H5 were 5.4 log₂. The results of the present survey indicated that the proportion of wild birds had previously infected AIV H5 at other times of the year. To our knowledge, this is the first seroprevalence report of avian influenza H5 infection in wild migratory birds in China' s southwestern Yunnan Province. The results of the present survey have significant public health concerns.

  8. Antigenic Variation in H5N1 clade 2.1 Viruses in Indonesia from 2005 to 2011

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    Vivi Setiawaty

    2013-01-01

    Full Text Available Influenza A (H5N1 virus, has spread to several countries in the world and has a high mortality rate. Meanwhile, the virus has evolved into several clades. The human influenza A (H5N1 virus circulating in Indonesia is a member of clade 2.1, which is different in antigenicity from other clades of influenza A (H5N1. An analysis of the antigenic variation in the H5 hemagglutinin gene (HA of the influenza A (H5N1 virus strains circulating in Indonesia has been undertaken. Several position of amino acid mutations, including mutations at positions 35, 53, 141, 145, 163, 174, 183, 184, 189, and 231, have been identified. The mutation Val-174-Iso appears to play an important role in immunogenicity and cross-reactivity with rabbit antisera. This study shows that the evolution of the H5HA antigenic variation of the influenza A (H5N1 virus circulating in Indonesia from 2005 to 2011 may affect the immunogenicity of the virus.

  9. Live, Attenuated Influenza A H5N1 Candidate Vaccines Provide Broad Cross-Protection in Mice and Ferrets

    Science.gov (United States)

    Mills, Kimberly L; Jin, Hong; Duke, Greg; Lu, Bin; Luke, Catherine J; Murphy, Brian; Swayne, David E; Kemble, George; Subbarao, Kanta

    2006-01-01

    Background Recent outbreaks of highly pathogenic influenza A H5N1 viruses in humans and avian species that began in Asia and have spread to other continents underscore an urgent need to develop vaccines that would protect the human population in the event of a pandemic. Methods and Findings Live, attenuated candidate vaccines possessing genes encoding a modified H5 hemagglutinin (HA) and a wild-type (wt) N1 neuraminidase from influenza A H5N1 viruses isolated in Hong Kong and Vietnam in 1997, 2003, and 2004, and remaining gene segments derived from the cold-adapted (ca) influenza A vaccine donor strain, influenza A/Ann Arbor/6/60 ca (H2N2), were generated by reverse genetics. The H5N1 ca vaccine viruses required trypsin for efficient growth in vitro, as predicted by the modification engineered in the gene encoding the HA, and possessed the temperature-sensitive and attenuation phenotypes specified by the internal protein genes of the ca vaccine donor strain. More importantly, the candidate vaccines were immunogenic in mice. Four weeks after receiving a single dose of 106 50% tissue culture infectious doses of intranasally administered vaccines, mice were fully protected from lethality following challenge with homologous and antigenically distinct heterologous wt H5N1 viruses from different genetic sublineages (clades 1, 2, and 3) that were isolated in Asia between 1997 and 2005. Four weeks after receiving two doses of the vaccines, mice and ferrets were fully protected against pulmonary replication of homologous and heterologous wt H5N1 viruses. Conclusions The promising findings in these preclinical studies of safety, immunogenicity, and efficacy of the H5N1 ca vaccines against antigenically diverse H5N1 vaccines provide support for their careful evaluation in Phase 1 clinical trials in humans. PMID:16968127

  10. Live, attenuated influenza A H5N1 candidate vaccines provide broad cross-protection in mice and ferrets.

    Directory of Open Access Journals (Sweden)

    Amorsolo L Suguitan

    2006-09-01

    Full Text Available Recent outbreaks of highly pathogenic influenza A H5N1 viruses in humans and avian species that began in Asia and have spread to other continents underscore an urgent need to develop vaccines that would protect the human population in the event of a pandemic.Live, attenuated candidate vaccines possessing genes encoding a modified H5 hemagglutinin (HA and a wild-type (wt N1 neuraminidase from influenza A H5N1 viruses isolated in Hong Kong and Vietnam in 1997, 2003, and 2004, and remaining gene segments derived from the cold-adapted (ca influenza A vaccine donor strain, influenza A/Ann Arbor/6/60 ca (H2N2, were generated by reverse genetics. The H5N1 ca vaccine viruses required trypsin for efficient growth in vitro, as predicted by the modification engineered in the gene encoding the HA, and possessed the temperature-sensitive and attenuation phenotypes specified by the internal protein genes of the ca vaccine donor strain. More importantly, the candidate vaccines were immunogenic in mice. Four weeks after receiving a single dose of 10(6 50% tissue culture infectious doses of intranasally administered vaccines, mice were fully protected from lethality following challenge with homologous and antigenically distinct heterologous wt H5N1 viruses from different genetic sublineages (clades 1, 2, and 3 that were isolated in Asia between 1997 and 2005. Four weeks after receiving two doses of the vaccines, mice and ferrets were fully protected against pulmonary replication of homologous and heterologous wt H5N1 viruses.The promising findings in these preclinical studies of safety, immunogenicity, and efficacy of the H5N1 ca vaccines against antigenically diverse H5N1 vaccines provide support for their careful evaluation in Phase 1 clinical trials in humans.

  11. Human mesenchymal stromal cells reduce influenza A H5N1-associated acute lung injury in vitro and in vivo.

    Science.gov (United States)

    Chan, Michael C W; Kuok, Denise I T; Leung, Connie Y H; Hui, Kenrie P Y; Valkenburg, Sophie A; Lau, Eric H Y; Nicholls, John M; Fang, Xiaohui; Guan, Yi; Lee, Jae W; Chan, Renee W Y; Webster, Robert G; Matthay, Michael A; Peiris, J S Malik

    2016-03-29

    Influenza can cause acute lung injury. Because immune responses often play a role, antivirals may not ensure a successful outcome. To identify pathogenic mechanisms and potential adjunctive therapeutic options, we compared the extent to which avian influenza A/H5N1 virus and seasonal influenza A/H1N1 virus impair alveolar fluid clearance and protein permeability in an in vitro model of acute lung injury, defined the role of virus-induced soluble mediators in these injury effects, and demonstrated that the effects are prevented or reduced by bone marrow-derived multipotent mesenchymal stromal cells. We verified the in vivo relevance of these findings in mice experimentally infected with influenza A/H5N1. We found that, in vitro, the alveolar epithelium's protein permeability and fluid clearance were dysregulated by soluble immune mediators released upon infection with avian (A/Hong Kong/483/97, H5N1) but not seasonal (A/Hong Kong/54/98, H1N1) influenza virus. The reduced alveolar fluid transport associated with down-regulation of sodium and chloride transporters was prevented or reduced by coculture with mesenchymal stromal cells. In vivo, treatment of aged H5N1-infected mice with mesenchymal stromal cells increased their likelihood of survival. We conclude that mesenchymal stromal cells significantly reduce the impairment of alveolar fluid clearance induced by A/H5N1 infection in vitro and prevent or reduce A/H5N1-associated acute lung injury in vivo. This potential adjunctive therapy for severe influenza-induced lung disease warrants rapid clinical investigation.

  12. New redox-active layer create via epoxy-amine reaction - The base of genosensor for the detection of specific DNA and RNA sequences of avian influenza virus H5N1.

    Science.gov (United States)

    Malecka, Kamila; Stachyra, Anna; Góra-Sochacka, Anna; Sirko, Agnieszka; Zagórski-Ostoja, Włodzimierz; Dehaen, Wim; Radecka, Hanna; Radecki, Jerzy

    2015-03-15

    This paper concerns the development of a redox-active monolayer and its application for the construction of an electrochemical genosensor designed for the detection of specific DNA and RNA oligonucleotide sequences related to the avian influenza virus (AIV) type H5N1. This new redox layer was created on a gold electrode surface step by step. Cyclic Voltammetry, Osteryoung Square-Wave Voltammetry and Differential Pulse Voltammetry were used for its characterization. This new redox-active layer was applied for the construction of the DNA biosensor. The NH2-NC3 probe (20-mer) was covalently attached to the gold electrode surface via a "click" reaction between the amine and an epoxide group. The hybridization process was monitored using the Osteryoung Square-Wave Voltammetry. The 20-mer DNA and ca. 280-mer RNA oligonucleotides were used as the targets. The constructed genosensor was capable to determine complementary oligonucleotide sequences with a detection limit in the pM range. It is able to distinguish the different position of the part RNA complementary to the DNA probe. The genosensor was very selective. The 20-mer DNA as well as the 280-mer RNA oligonucleotides without a complementary sequence generated a weak signal. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Detection of antibodies against H5 and H7 strains in birds: evaluation of influenza pseudovirus particle neutralization tests

    Directory of Open Access Journals (Sweden)

    Sofie Wallerström

    2014-01-01

    Full Text Available Introduction: Avian influenza viruses circulate in bird populations, and it is important to maintain and uphold our knowledge of the viral strains that are currently of interest in this context. Here, we describe the use of hemagglutinin-pseudotype retroviruses based on highly pathogenic influenza viruses for the screening of avian sera for influenza A antibodies. Our aim was also to determine whether the pseudovirus neutralization tests that we assessed were sensitive and simple to use compared to the traditional methods, including hemagglutination inhibition assays and microneutralization tests. Material and methods: H5 and H7 pseudovirus neutralization tests were evaluated by using serum from infected rabbits. Subsequently, the assays were further investigated using a panel of serum samples from avian species. The panel contained samples that were seropositive for five different hemagglutinin subtypes as well as influenza A seronegative samples. Results and discussion: The results suggest that the pseudovirus neutralization test is an alternative to hemagglutination inhibition assays, as we observed comparable titers to those of both standard microneutralizations assays as well as hemagglutinin inhibition assays. When evaluated by a panel of avian sera, the method also showed its capability to recognize antibodies directed toward low-pathogenic H5 and H7. Hence, we conclude that it is possible to use pseudoviruses based on highly pathogenic avian influenza viruses to screen avian sera for antibodies directed against influenza A subtypes H5 and H7.

  14. EVALUATION OF OIL BASED AVIAN INFLUENZA VACCINE (H5NI PREPARED WITH DIFFERENT CONCENTRATIONS OF ADJUVANT

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    M. IQBAL, M. NISAR, ANWARUL-HAQ, S. NOOR AND Z. J. GILL

    2008-12-01

    Full Text Available Bird flu vaccine from H5N1 strain of avian influenza virus was prepared with two concentrations of adjuvant (Montanide ISA 70MVG. Two vaccines (I and II were prepared containing 50 and 60% Montanide, respectively. Immune response of both the vaccines as single, as well as booster, dose was evaluated in layer birds through haemagglutination inhibition test. Single dose of both vaccines showed poor immune response, while booster dose gave better response with both the vaccines. However, the vaccine prepared with 60% Montanide provided better immune response compared with the vaccine containing 50% montanide.

  15. Unusually High Mortality in Waterfowl Caused by Highly Pathogenic Avian Influenza A(H5N1) in Bangladesh

    Science.gov (United States)

    Haider, N.; Sturm-Ramirez, K.; Khan, S. U.; Rahman, M. Z.; Sarkar, S.; Poh, M. K.; Shivaprasad, H. L.; Kalam, M. A.; Paul, S. K.; Karmakar, P. C.; Balish, A.; Chakraborty, A.; Mamun, A. A.; Mikolon, A. B.; Davis, C. T.; Rahman, M.; Donis, R. O.; Heffelfinger, J. D.; Luby, S. P.; Zeidner, N.

    2015-01-01

    Summary Mortality in ducks and geese caused by highly pathogenic avian influenza A (H5N1) infection had not been previously identified in Bangladesh. In June–July 2011, we investigated mortality in ducks, geese and chickens with suspected H5N1 infection in a north-eastern district of the country to identify the aetiologic agent and extent of the outbreak and identify possible associated human infections. We surveyed households and farms with affected poultry flocks in six villages in Netrokona district and collected cloacal and oropharyngeal swabs from sick birds and tissue samples from dead poultry. We conducted a survey in three of these villages to identify suspected human influenza-like illness cases and collected nasopharyngeal and throat swabs. We tested all swabs by real-time RT-PCR, sequenced cultured viruses, and examined tissue samples by histopathology and immunohistochemistry to detect and characterize influenza virus infection. In the six villages, among the 240 surveyed households and 11 small-scale farms, 61% (1789/2930) of chickens, 47% (4816/10 184) of ducks and 73% (358/493) of geese died within 14 days preceding the investigation. Of 70 sick poultry swabbed, 80% (56/70) had detectable RNA for influenza A/H5, including 89% (49/55) of ducks, 40% (2/5) of geese and 50% (5/10) of chickens. We isolated virus from six of 25 samples; sequence analysis of the hemagglutinin and neuraminidase gene of these six isolates indicated clade 2.3.2.1a of H5N1 virus. Histopathological changes and immunohistochemistry staining of avian influenza viral antigens were recognized in the brain, pancreas and intestines of ducks and chickens. We identified ten human cases showing signs compatible with influenza-like illness; four were positive for influenza A/H3; however, none were positive for influenza A/H5. The recently introduced H5N1 clade 2.3.2.1a virus caused unusually high mortality in ducks and geese. Heightened surveillance in poultry is warranted to guide

  16. Unusually High Mortality in Waterfowl Caused by Highly Pathogenic Avian Influenza A(H5N1) in Bangladesh.

    Science.gov (United States)

    Haider, N; Sturm-Ramirez, K; Khan, S U; Rahman, M Z; Sarkar, S; Poh, M K; Shivaprasad, H L; Kalam, M A; Paul, S K; Karmakar, P C; Balish, A; Chakraborty, A; Mamun, A A; Mikolon, A B; Davis, C T; Rahman, M; Donis, R O; Heffelfinger, J D; Luby, S P; Zeidner, N

    2017-02-01

    Mortality in ducks and geese caused by highly pathogenic avian influenza A(H5N1) infection had not been previously identified in Bangladesh. In June-July 2011, we investigated mortality in ducks, geese and chickens with suspected H5N1 infection in a north-eastern district of the country to identify the aetiologic agent and extent of the outbreak and identify possible associated human infections. We surveyed households and farms with affected poultry flocks in six villages in Netrokona district and collected cloacal and oropharyngeal swabs from sick birds and tissue samples from dead poultry. We conducted a survey in three of these villages to identify suspected human influenza-like illness cases and collected nasopharyngeal and throat swabs. We tested all swabs by real-time RT-PCR, sequenced cultured viruses, and examined tissue samples by histopathology and immunohistochemistry to detect and characterize influenza virus infection. In the six villages, among the 240 surveyed households and 11 small-scale farms, 61% (1789/2930) of chickens, 47% (4816/10 184) of ducks and 73% (358/493) of geese died within 14 days preceding the investigation. Of 70 sick poultry swabbed, 80% (56/70) had detectable RNA for influenza A/H5, including 89% (49/55) of ducks, 40% (2/5) of geese and 50% (5/10) of chickens. We isolated virus from six of 25 samples; sequence analysis of the hemagglutinin and neuraminidase gene of these six isolates indicated clade 2.3.2.1a of H5N1 virus. Histopathological changes and immunohistochemistry staining of avian influenza viral antigens were recognized in the brain, pancreas and intestines of ducks and chickens. We identified ten human cases showing signs compatible with influenza-like illness; four were positive for influenza A/H3; however, none were positive for influenza A/H5. The recently introduced H5N1 clade 2.3.2.1a virus caused unusually high mortality in ducks and geese. Heightened surveillance in poultry is warranted to guide appropriate

  17. Differences in the detection of highly pathogenic avian influenza H5N1 virus in feather samples from 4-week-old and 24-week-old infected Pekin ducks (Anas platyrhynchos var. domestica).

    Science.gov (United States)

    Aiello, Roberta; Beato, Maria Serena; Mancin, Marzia; Rigoni, Michela; Tejeda, Aurora Romero; Maniero, Silvia; Capua, Ilaria; Terregino, Calogero

    2013-08-30

    Previous studies have reported the detection of H5N1 HPAI virus in feathers from ducks naturally and experimentally infected and suggested that feather calami (FC) could be used as diagnostic samples for the early detection of H5N1 HPAI infections. Ducks are readily infected with H5N1 HPAI viruses although the development of clinical signs and deaths were reported as age-related with younger birds being more susceptible. The correlation between age and virus localisation in FC of infected ducks has not been studied to date. In the present study juvenile (4-week-old) and adult (24-week-old) Pekin ducks (Anas platyrhynchos var. domestica) were infected experimentally with a clade 2.2 H5N1 HPAI virus (A/duck/Nigeria/1071-23/2007). Tracheal (Tr) and cloacal (Cl) swabs and FC were collected at 3, 5, 7 and 10 days post infection and tested by RRT-PCR and a double antibody sandwich-ELISA (DAS-ELISA) developed in house. Virus was detected in swabs and FC of challenged ducks with a higher rate of detection in juvenile ducks. In this age group virus was detected over a longer period of time in FC compared to swabs. Our study showed that FC samples collected from young ducks are a valid diagnostic specimen for H5N1 HPAI virus detection. The DAS-ELISA on FC proved to be a suitable alternative diagnostic test when molecular and/or virus isolation techniques are not available therefore it could be useful in the diagnosis of H5N1 HPAI infections in under-resourced countries. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Swine Influenza/Variant Influenza Viruses

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    ... Address What's this? Submit What's this? Submit Button Influenza Types Seasonal Avian Swine Variant Pandemic Other Information on Swine Influenza/Variant Influenza Virus Language: English (US) Español Recommend ...

  19. On the role of vaccine dose and antigenic distance in the transmission dynamics of Highly Pathogenic Avian Influenza (HPAI) H5N1 virus and its selected mutants in vaccinated animals

    NARCIS (Netherlands)

    Sitaras, Ioannis

    2017-01-01

    Influenza virus infections can cause high morbidity and mortality rates among animals and humans, and result in staggering direct and indirect financial losses amounting to billions of US dollars. Ever since it emerged in 1996 in Guangdong province, People’s Republic of China, one particular

  20. Comparison of temporal and spatial dynamics of seasonal H3N2, pandemic H1N1 and highly pathogenic avian influenza H5N1 virus infections in ferrets

    NARCIS (Netherlands)

    J.M.A. van den Brand (Judith); K.J. Stittelaar (Koert); G. van Amerongen (Geert); L.A. Reperant (Leslie); L. de Waal (Leon); A.D.M.E. Osterhaus (Albert); T. Kuiken (Thijs)

    2012-01-01

    textabstractHumans may be infected by different influenza A viruses-seasonal, pandemic, and zoonotic-which differ in presentation from mild upper respiratory tract disease to severe and sometimes fatal pneumonia with extra-respiratory spread. Differences in spatial and temporal dynamics of these

  1. Public health concerns of highly pathogenic avian influenza H5N1 endemicity in Africa

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    Olubunmi Gabriel Fasanmi

    2017-10-01

    Full Text Available Highly pathogenic avian influenza virus (HPAIV H5N1 was first officially reported in Africa in 2006; thereafter this virus has spread rapidly from Nigeria to 11 other African countries. This study was aimed at utilizing data from confirmed laboratory reports to carry out a qualitative evaluation of the factors responsible for HPAI H5N1 persistence in Africa and the public health implications; and to suggest appropriate control measures. Relevant publications were sought from data banks and repositories of FAO, OIE, WHO, and Google scholars. Substantiated data on HPAI H5N1 outbreaks in poultry in Africa and in humans across the world were mined. HPAI H5N1 affects poultry and human populations, with Egypt having highest human cases (346 globally. Nigeria had a reinfection from 2014 to 2015, with outbreaks in Cote d'Ivoire, Ghana, Niger, Nigeria, and Burkina Faso throughout 2016 unabated. The persistence of this virus in Africa is attributed to the survivability of HPAIV, ability to evolve other subtypes through genetic reassortment, poor biosecurity compliance at the live bird markets and poultry farms, husbandry methods and multispecies livestock farming, poultry vaccinations, and continuous shedding of HPAIV, transboundary transmission of HPAIV through poultry trades; and transcontinental migratory birds. There is, therefore, the need for African nations to realistically reassess their status, through regular surveillance and be transparent with HPAI H5N1 outbreak data. Also, it is important to have an understanding of HPAIV migration dynamics which will be helpful in epidemiological modeling, disease prevention, control and eradication measures.

  2. Insight into Alternative Approaches for Control of Avian Influenza in Poultry, with Emphasis on Highly Pathogenic H5N1

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    Hafez M. Hafez

    2012-11-01

    Full Text Available Highly pathogenic avian influenza virus (HPAIV of subtype H5N1 causes a devastating disease in poultry but when it accidentally infects humans it can cause death. Therefore, decrease the incidence of H5N1 in humans needs to focus on prevention and control of poultry infections. Conventional control strategies in poultry based on surveillance, stamping out, movement restriction and enforcement of biosecurity measures did not prevent the virus spreading, particularly in developing countries. Several challenges limit efficiency of the vaccines to prevent outbreaks of HPAIV H5N1 in endemic countries. Alternative and complementary approaches to reduce the current burden of H5N1 epidemics in poultry should be encouraged. The use of antiviral chemotherapy and natural compounds, avian-cytokines, RNA interference, genetic breeding and/or development of transgenic poultry warrant further evaluation as integrated intervention strategies for control of HPAIV H5N1 in poultry.

  3. Identifikasi Secara Serologi Galur Virus Flu Burung Subtipe H5N1 Clade 2.1.3 dan Clade 2.3.2 pada Ayam Petelur (SEROLOGICAL IDENTIFICATION OF AVIAN INFLUENZA STRAIN