Special Issue "H5N1 Influenza Virus"

Guest Editor
Prof. Dr. Daniel R. Perez
Associate Professor, Department of Veterinary Medicine, Room 1215, 8075 Greenmead Drive, University of Maryland, College Park, College Park, MD 20742, USA
Website: http://avianflu.umd.edu
E-Mail: avianflu@me.com
Phone: +(301) 314 6811
Fax: +(301) 314 6855
Interests: Interspecies Transmission of H9N2 avian influenza viruses; Bullet Pathogenesis and Transmission of 2009 H1N1pdm influenza strains; BulletUniversal vaccine approaches for animals and humans; BulletCross-protective immunity during H5N1 infections; BulletVirus-host interactions - Influenza receptors in avian and mammalian hosts

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• H5N1
• influenza virus

Type of Paper: Review
Title: Avian influenza H5N1 Zoonosis from Poultry to Humans: Why has Vaccination Failed?
Author: Cassandra Berry
Affiliation: School of Veterinary and Biomedical Sciences, Murdoch University, South Street, Murdoch, Perth 6150, Western Australia, Australia.
Abstract: Highly pathogenic avian influenza virus H5N1 strains have emerged as zoonotic viral pathogens over the last decade and have eluded our serious attempts of control by vaccination, with numerous countries having endemic regions for H5N1 virus. Although the biology and genomics of H5N1 influenza viruses are well characterised, having been extensively researched since 2003, viral outbreaks still occur in domestic poultry, posing a dangerous threat for human transmission. Here we review our futile attempts to control H5N1 viral outbreaks in domestic poultry flocks and question our capacity for future achievements of eradicating zoonotic H5N1 influenza. Vaccination procedures regarding domestic poultry rather than wild migratory birds and other species will be discussed with particular reference to control of potentially deadly but silent infection in vaccinated poultry.

Type of Paper: Review
Title: A Review of Molecular Mechanisms Involved in Cross-species Transmission of the H5N1 Avian Influenza A Virus
Authors: Hong Zhang¹, Ke Xu and Bing Sun
Affiliation: Key Laboratory of Molecular Virology & Immunology, Institut Pasteur of Shanghai, Shanghai institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, 200025, China
^* Address correspondence to Bing Sun, bsun@sibs.ac.cn, or Ke Xu, kxu@sibs.ac.cn
Abstract: Since the breakout of human infection with highly pathogenic avian influenza H5N1 virus in Hong Kong in the year 1997, people realize that avian influenza viruses are able to be transmitted from birds to humans leading to cross-species infection. Thus, it raises a great concern on the possibility of a human pandemic caused by either an avian virus or a reassortant between human flu viruses and avian flu viruses, which could be more virulent, easily-transmissible and lethal to humans. Understanding the mechanisms involved in cross-species transmission of the H5N1 virus is therefore mandatory for preventing and controlling such a possible pandemic. It is believed that two host cellular barriers mainly contribute to the restriction of a certain influenza virus infection, the cell membrane where the binding and enter of the virus occurs and the nuclear envelop where the vRNP components need to cross enabling the replication and transcription of viral genome in the nucleus. Many viral and host factors have been already known to be involved in these two events, and more unrevealed factors are demanded to be discovered. In this review, we will summarize the current knowledge of molecular mechanisms in H5N1 cross-species transmission, including viral and host factors that determine the ability of the virus to cross cell membrane and nuclear envelop to replicate in a new host.

Type of Paper: Article
Title: Magnetic Electrochemical Bar Code Array for Point Mutation Detection in H5N1 Neuraminidase Gene
Authors: Ludmila Krejcova ¹, David Hynek ^1,2, Pavel Kopel ^1,2, Miguel Angel Merlos Rodrigo ^1,2, Vojtech Adam ^1,2, Jaromir Hubalek ^2,3, Libuse Trnkova ^1,2 and Rene Kizek ^1,2
Affiliations: ¹ Department of Chemistry and Biochemistry, Faculty of Agronomy, Mendel University in Brno, Zemedelska 1, CZ-613 00 Brno, Czech Republic, European Union;  ² Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, CZ-616 00 Brno, Czech Republic, European Union; ³ Department of Microelectronics, Faculty of Electrical Engineering and  Communication, Brno University of Technology, Udolni 53, CZ-602 00 Brno, Czech Republic, European Union; kizek@sci.muni.cz
Abstract: Since its first official detection in the Guangdong province of China in 1996, highly pathogenic avian influenza virus of H5N1 subtype (HPAI H5N1) has reportedly been the cause of outbreaks in birds in more than 60 countries, 24 of which were European. The main issue is still development of effective antiviral drugs. In this case, there was observed single point mutation in the neuraminidase gene, which causes resistance to antiviral drug and is, therefore, subjected to many studies including ours. In this study, we developed magnetic electrochemical bar code array for detection single point mutations (mismatches in up to four nucleotides) in H5N1 neuraminidase gene. Magnetic particles with covalently bound oligo dT were used as a tool for isolation of complementary H5N1 chains (H5N1 Zhejin, China and Aichi). There were tested chains of two lengths as 17 or 33 bp. These chains were hybridized with yield higher than 80 %. For detection of H5N1 chains, oligonucleotide chains of lengths of 12 or 28 bp labelled with quantum dots (CdS, ZnS and/or PbS). Individual probes hybridized to target molecules specifically with efficiency higher than 60 %. The obtained signals identified mutations present in the sequence. Suggested experimental procedure allows obtaining further information from the redox signals of nucleic acids. Moreover, the used biosensor exhibits sequence specificity and low limits of detection in subnanogram quantities of target nucleic acids.
Keywords: voltammetry; highly pathogenic influenza; antiviral resistance; paramagnetic particles; hybridization; quantum dots; automated separation; electrochemistry

Type of Paper: Commentary
Author: Michalann Harthill
Affiliation: Geochemistry and Health International, Inc., P.O. Box 3523, Frederick, MD 21701 USA; E-Mail: mharthill@gmail.com (M.H.); Tel.: +1-301-639-9765
Abstract: Previous research determined that host nutritional deficiency in the antioxidant selenium impaired immunocompetence against and increased virulence of several RNA viruses.  To scope the influence of poor soil Se bioavailability to subsistence foodcrops on the geographic occurrence of RNA viral infectious diseases (VIDs), the literature was searched for correlations between selenium blood values and origins of any VIDS, including outbreak of H5N1 influenza A.  Though polygenic in its pathogenicity, recounted here are associations of the H5N1 influenza A virulent proteins M1, PB2-627Lys, and avirulent PB2-627Glu with host Se-nutrition status.
Keywords: H5N1 influenza Avirus; host selenium-nutrition status; M1 protein; PB2-627Glu; PB2-627Lys; selenium deficiency

Type of Paper: Article
Title: Nanotechnology Based Microarray Assay for Detection and Differentiation of Pandemic Influenza Virus (2009 A/H1N1)
from other major Seasonal Influenza Viruses
Authors: Jiangqin Zhao, Shixing Tang, Xue Wang and Indira Hewlett.
Affiliation: Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892, USA
Abstract: Since pandemic strains of swine flu (2009 A/H1N1) and avian influenza virus (H5N1) have spread in the world, rapid and sensitive diagnostic tools for the detection of these influenza viruses are crucial for early detection, appropriate treatment, epidemiologic investigations, and timely responses to a pandemic threat. We previously developed a rapid and simple gold nanoparticle (NP)-based genomic microarray assay for identification of avian flu H5N1 and discrimination from major influenza A virus strains (H1N1, H3N2). In this study, we expanded the platform to detect swine flu. Multiple specific capture and intermediate oligonucleotides were designed for the matrix(M), hemaglutinin(HA) and neuraminidase(NA) genes of swine flu 2009(A/H1N1) virus. Microarray was printed in the same format for HA, NA and M genes for seasonal H1N1, H3N2, H5N1 and 2009(A/H1N1). Viral RNA was tested using capture-target-intermediate oligonucleotide (oligo) hybridization and gold NP-mediated silver staining. The signals for 4 capture oligos designed for HA and NA genes of 2009(A/H1N1) virus were specifically detected without cross hybridization with seasonal flu H1N1, H3N2 and H5N1 viral RNA. All of 3 M capture oligos showed strong signals with swine flu viral RNA but 2 out of 3 M capture oligos showed cross hybridization with seasonal H1N1, H3N2 and H5N1. The current microarray assay was able to detect influenza A viruses (H1N1, H3N2 and H5N1) and distinguish 2009(A/H1N1) virus from other major influenza A viruses. The new method can be useful for simultaneous detection and subtyping of major influenza A viruses and the platform can be rapidly modified to detect other emerging influenza strains in public health settings.
Biography: Jiangqin Zhao has completed his Ph.D in 2001 from University of Glasgow and postdoctoral studies from University of Glasgow and University of Leicester in UK. Since 2006, Dr Zhao was invited to work in the Laboratory of Molecular Virology, Center for Biologics Evaluation and Research (CBER) of the U.S. Food and Drug Administration (FDA) to join its research efforts in the area of viral infections diseases. He has published more than 30 papers in reputed journals

Type of Paper: Review
Title: The Influence of Sequential Vaccination with Drifted Hemagglutinin Variants on H5N1 Immunity
Authors: Felix W. Santiago¹, David J. Topham¹, and John J. Treanor^1,2
Affiliations: ¹ New York Influenza Center of Excellence, David H. Smith Center for Vaccine Biology and Immunology, Aab Institute of Biomedical Sciences, and Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester NY; ² Division of Infectious Diseases, Department of Medicine, University of Rochester Medical Center, Rochester, NY
Abstract: Since their emergence in 1997, Highly Pathogenic Avian Influenza H5N1 viruses continue to cause occasional infections in humans.   The World Health Organization has identified H5N1 viruses as a potential pandemic threat and prototype viruses are selected yearly as possible vaccine candidates.  Pre-vaccination with currently stockpiled vaccines has been proposed as a pandemic control strategy. However, few studies have addressed the influence that pre-vaccination has on influenza-specific immunity following vaccination with a formulation containing a drifted H5 HA variant. In this review, we highlight key findings in this area of research and comment on the potential immunological benefits of establishing baseline H5N1-specific immunity.

Type of Paper: Review
Title: H5N1 Virus in Russia (2005-2012): Molecular Epidemiology, Ecology and Evolution. Or H5N1 Virus in Russia, 2005-2012
Authors: Kirill Sharshov, Vasiliy Marchenko, Ivan Susloparov, Alexander Alekseev, Alexander Durymanov, Alexander Shestopalov
Affiliation: Zoonotic Diseases and Influenza Department, State Research Centre of Virology and Biotechnology «VECTOR », Novosibirsk, Russia; Novosibirsk State University, Novosibirsk, Russia
Abstract: The paper focuses on some epidemiological, ecological and evolutionary aspects of H5N1 virus in Russia from the first detection till now. The first influenza A (H5N1) outbreak in Russia was reported in the summer of 2005 in the territory of Western Siberia (Lipatov etal., 2007). With migratory wild birds, virus spread westward across Eurasia and as far west as England and West Africa. Analysis of the nucleotide sequence of the hemagglutinin (HA) gene showed that strains isolated in Russia in 2005 were related phylogenetically to H5N1 viruses that had caused an outbreak among wild birds at Qinghai Lake in China in Spring 2005. Subsequently, the Qinghai-like (clade 2.2) HPAI H5N1 lineage was detected in wild birds and poultry in many countries. The source of these introductions, while still debated, is likely through bird migration. Interestingly enough numerous wild bird’s species were involved in Russia outbreaks (swans, gulls, grebes, spoonbills, sparrows and others). The first case of Fujian sub-clade 2.3.2 influenza virus (H5N1) lineage in the Russian Far East was recorded in April 2008. Prior to this, no HPAI H5N1 outbreaks of the Fujian lineage had been reported in Russia. In June 2009, an outbreak of HPAI was recorded in wild birds in Mongolia and on the Uvs-Nuur Lake in Russia. Phylogenetic analysis of HA gene showed that viruses belong to clade 2.3.2. These viruses were close to viruses isolated in Mongolia at the same time and in Japan (Hokkaido) in 2008. The spread of HPAI H5N1 west across the globe has caused serious debates on the roles of migratory birds in virus circulation. We suggest that wild birds brought the virus to Uvs-Nuur Lake from outside the country. Since prior to June 2009, the only case of new Fujian sub-clade 2.3.2 influenza virus (H5N1) lineage was in the Russian Far East, we believe that the virus isolated in 2009 from Uvs-Nuur Lake was probably introduced by wild birds that wintered in Southeast Asia.
We hypothesized that bodies of water like the Qinghai Lake and the Uvs Nuur Lake may play an important role in the circulation of avian influenza so we suggested enhancing of surveillance program in this area and therefore we continued to study new outbreaks thoroughly (Sharshov et al., 2010).
Our hypothesis was confirmed in June 2010. An outbreak of HPAI was recorded in wild birds at the Uvs Nuur Lake (OIE, 2010). It was shown a close antigenic relationship between the investigated strains and those isolated at the Uvs Nuur Lake in 2009 and in the Russian Far East in 2008 (both clade 2.3.2). Phylogenetic analysis of the hemagglutinin (HA) gene showed a close relation to strains isolated during outbreaks at the same location in 2009, at the Qinghai Lake in 2009 and in Mongolia in 2010 as all of them fall into clade 2.3.2. We can suppose that these strains originally appeared before or around 2009 at the Qinghai Lake and later in May 2010 caused an outbreak in Central Mongolia (Sakoda et al., 2010). In June 2010 dead birds were found at the Uvs Nuur Lake and viruses with 98-99% identity to Mongolian strains were isolated.
The paper contains detailed comparative virological, molecular, pathogenic characteristic H5N1viruses isolated in Russia (2005-2011). Our experience with the 2005-2007 outbreaks and their HPAI H5N1 (clade 2.2) viruses originally appearing around Qinghai lake and then spreading first to Russia and later to Europe and Africa (Hars et al., 2008; Alexander and Brown, 2009) leads us to the assumption of a similar scenario for clade 2.3.2 HPAI virus global spread. Recent detection of clade 2.3.2 H5N1 viruses during outbreaks in Romania and Bulgaria (Reid et al., 2010) confirm this hypothesis. Furthermore 2.3.2 viruses have been detected in wild birds in Japan (2011 Feb; WHO). So, further investigation of the global distribution of clade 2.3.2 and 2.2 HPAI viruses will prove invaluable to give us a better understanding of the Evolutionary Ecology of avian influenza viruses in natural host populations.
This work was supported by Russian Federal Program (grant 11.519.11.2014).

Type of Paper: Review
Title: Proinflammatory Cytokine Responses to H5N1 Infections
Authors: Mark D. Wareing and Gregory A. Tannock
Affiliation: Center for Virology, Burnet Institute, 85 Commercial Road, Prahran, Victoria, Australia 3004
Abstract: Avian H5N1 influenza viruses as members of a new human influenza A subtype in Hong Kong in 1997 and highly pathogenic variants have persisted in South-East Asia since 2003.Human-human transmission is rare but mortality rates associated with highly pathogenic H5N1 viruses are significantly higher than for current epidemic strains. Pathology associated with severe human H5N1 infections is characterised by severe viral pneumonia, diffuse alveolar damage and acute respiratory distress syndrome (ARDS). A key feature of severe H5N1 infection is dysfunctional cytokine production resulting in cytokine storm. It is unclear what factors directly contribute to severe H5N1-induced cytokine disregulation. Factors associated with H5N1 disease severity include elevated virus replication and apoptosis, as well as other intrinsic properties of different H5N1 viruses and their capacity to inhibit the host cell interferon response. Here we review the cytokine response to influenza A infections and attempt to define the relationship between elements of cytokine storm and severe H5N1 pathology.

Type of Paper: Article
Title: Zoonotic Potential of North American Low Pathogenic Avian H5 Influenza A Viruses for Mammalian Hosts
Authors: Jennifer Humberd-Smith, Paula Brooks and Robert J. Hogan ^*
Affiliation: Department of Infectious Diseases, University of Georgia, College of Veterinary Medicine, Athens, GA 30602
Abstract: Avian influenza viruses have not only contributed genes to pandemic strains of influenza but avian influenza viruses are increasingly being transmitted directly to humans. Since 1997 highly pathogenic H5N1 avian influenza (HPAI) viruses have transmitted directly to humans with high incidence of mortality. Although human cases of infection with H5 HPAI have not been reported in North America, low pathogenic (LP) H5 influenza viruses have been increasingly isolated from wild bird populations and domestic poultry. In the present study we evaluated the pathogenicity of North American LP H5 isolates in mammalian models of influenza infection. H5 viruses isolated from domestic poultry replicated in mice without prior adaptation for up to 7 days post inoculation, however mice were refractory to infection with the tested wild bird isolate. In ferrets, all 3 LP H5 isolates replicated in the upper respiratory tract with mean peak titers on day 5 post inoculation (range, 10^3.7 to 10^5.6 EID₅₀/ml) and at a low level in the intestinal tract. Because LP H5 viruses may evolve into HPAI through adaptation to domestic poultry and LP H5 viruses are being isolated with increased frequency in North America, the replication of these isolates in mammalian models without prior adaptation highlights the importance of continued surveillance and characterization of LP H5 viruses.