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The COVID-19 pandemic continues to wreak havoc as worldwide SARS-CoV-2 infection, hospitalization, and death rates climb unabated. Effective vaccines remain the most promising approach to counter SARS-CoV-2. Yet, while promising results are emerging from COVID-19 vaccine trials, the need for multiple doses and the challenges associated with the widespread distribution and administration of vaccines remain concerns. Here, we engineered the coat protein of the MS2 bacteriophage and generated nanoparticles displaying multiple copies of the SARS-CoV-2 spike (S) protein. The use of these nanoparticles as vaccines generated high neutralizing antibody titers and protected Syrian hamsters from a challenge with SARS-CoV-2 after a single immunization with no infectious virus detected in the lungs. This nanoparticle-based vaccine platform thus provides protection after a single immunization and may be broadly applicable for protecting against SARS-CoV-2 and future pathogens with pandemic potential.

Guidelines from the CDC and the WHO recommend the wearing of face masks to prevent the spread of coronavirus (CoV) disease 2019 (COVID-19); however, the protective efficiency of such masks against airborne transmission of infectious severe acute respiratory syndrome CoV-2 (SARS-CoV-2) droplets/aerosols is unknown. Here, we developed an airborne transmission simulator of infectious SARS-CoV-2-containing droplets/aerosols produced by human respiration and coughs and assessed the transmissibility of the infectious droplets/aerosols and the ability of various types of face masks to block the transmission. We found that cotton masks, surgical masks, and N95 masks all have a protective effect with respect to the transmission of infective droplets/aerosols of SARS-CoV-2 and that the protective efficiency was higher when masks were worn by a virus spreader. Importantly, medical masks (surgical masks and even N95 masks) were not able to completely block the transmission of virus droplets/aerosols even when completely sealed. Our data will help medical workers understand the proper use and performance of masks and determine whether they need additional equipment to protect themselves from infected patients. IMPORTANCE Airborne simulation experiments showed that cotton masks, surgical masks, and N95 masks provide some protection from the transmission of infective SARS-CoV-2 droplets/aerosols; however, medical masks (surgical masks and even N95 masks) could not completely block the transmission of virus droplets/aerosols even when sealed., (Copyright © 2020 Ueki et al.)

Worldwide outbreaks of influenza (pandemics) are caused by influenza A viruses to which persons lack protective immune responses. Currently, we are unable to predict which influenza virus strains may cause a pandemic. In this article, we summarize some of the information that will be needed to better assess the pandemic potential of influenza viruses, and we discuss our current gaps in knowledge., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Animal influenza viruses pose a clear threat to public health. Transmissibility among humans is a prerequisite for a novel influenza virus to cause a human pandemic. A novel reassortant swine influenza virus acquired sustained human-to-human transmissibility and caused the 2009 influenza pandemic. However, the molecular aspects of influenza virus transmission remain poorly understood. Here, we show that an amino acid in hemagglutinin (HA) is important for the 2009 H1N1 influenza pandemic virus (2009/H1N1) to bind to human virus receptors and confer respiratory droplet transmissibility in mammals. We found that the change from glutamine (Q) to arginine (R) at position 226 of HA, which causes a switch in receptor-binding preference from human α-2,6 to avian α-2,3 sialic acid, resulted in a virus incapable of respiratory droplet transmission in guinea pigs and reduced the virus's ability to replicate in the lungs of ferrets. The change from alanine (A) to threonine (T) at position 271 of PB2 also abolished the virus's respiratory droplet transmission in guinea pigs, and this mutation, together with the HA Q226R mutation, abolished the virus's respiratory droplet transmission in ferrets. Furthermore, we found that amino acid 271A of PB2 plays a key role in virus acquisition of the mutation at position 226 of HA that confers human receptor recognition. Our results highlight the importance of both the PB2 and HA genes on the adaptation and transmission of influenza viruses in humans and provide important insights for monitoring and evaluating the pandemic potential of field influenza viruses.

Background: Annually, influenza A viruses circulate the world causing wide-spread sickness, economic loss, and death. One way to better defend against influenza virus-induced disease may be to develop novel host-based therapies, targeted at mitigating viral pathogenesis through the management of virus-dysregulated host functions. However, mechanisms that govern aberrant host responses to influenza virus infection remain incompletely understood. We previously showed that the pandemic H1N1 virus influenza A/California/04/2009 (H1N1; CA04) has enhanced pathogenicity in the lungs of cynomolgus macaques relative to a seasonal influenza virus isolate (A/Kawasaki/UTK-4/2009 (H1N1; KUTK4))., Results: Here, we used microarrays to identify host gene sequences that were highly differentially expressed (DE) in CA04-infected macaque lungs, and we employed a novel strategy - combining functional and pathway enrichment analyses, transcription factor binding site enrichment analysis and protein-protein interaction data - to create a CA04 differentially regulated host response network. This network describes enhanced viral RNA sensing, immune cell signaling and cell cycle arrest in CA04-infected lungs, and highlights a novel, putative role for the MYC-associated zinc finger (MAZ) transcription factor in regulating these processes., Conclusions: Our findings suggest that the enhanced pathology is the result of a prolonged immune response, despite successful virus clearance. Most interesting, we identify a mechanism which normally suppresses immune cell signaling and inflammation is ineffective in the pH1N1 virus infection; a dyregulatory event also associated with arthritis. This dysregulation offers several opportunities for developing strain-independent, immunomodulatory therapies to protect against future pandemics.

We provide a brief introduction into the genome organization, life cycle, pathogenicity, and host range of influenza A viruses. We also briefly summarize influenza pandemics and currently available measures to control influenza virus outbreaks, including vaccines and antiviral compounds to influenza viruses.

Background: In 2009, a pandemic (pdm) influenza A(H1N1) virus infection quickly circulated globally resulting in about 18,000 deaths around the world. In Japan, infected patients accounted for 16% of the total population. The possibility of human-to-human transmission of highly pathogenic novel influenza viruses is becoming a fear for human health and society., Methodology: To address the clinical need for rapid diagnosis, we have developed a new method, the "RT-SmartAmp assay", to rapidly detect the 2009 pandemic influenza A(H1N1) virus from patient swab samples. The RT-SmartAmp assay comprises both reverse transcriptase (RT) and isothermal DNA amplification reactions in one step, where RNA extraction and PCR reaction are not required. We used an exciton-controlled hybridization-sensitive fluorescent primer to specifically detect the HA segment of the 2009 pdm influenza A(H1N1) virus within 40 minutes without cross-reacting with the seasonal A(H1N1), A(H3N2), or B-type (Victoria) viruses., Results and Conclusions: We evaluated the RT-SmartAmp method in clinical research carried out in Japan during a pandemic period of October 2009 to January 2010. A total of 255 swab samples were collected from outpatients with influenza-like illness at three hospitals and eleven clinics located in the Tokyo and Chiba areas in Japan. The 2009 pdm influenza A(H1N1) virus was detected by the RT-SmartAmp assay, and the detection results were subsequently compared with data of current influenza diagnostic tests (lateral flow immuno-chromatographic tests) and viral genome sequence analysis. In conclusion, by the RT-SmartAmp assay we could detect the 2009 pdm influenza A(H1N1) virus in patients' swab samples even in early stages after the initial onset of influenza symptoms. Thus, the RT-SmartAmp assay is considered to provide a simple and practical tool to rapidly detect the 2009 pdm influenza A(H1N1) virus.

Highly pathogenic H5N1 avian influenza viruses have caused infection in humans, with a high mortality rate, since 1997. While the pathogenesis of this infection is not completely understood, hypercytokinaemia and alveolar macrophages are thought to play a role. To gain further insight into the cytokine-mediated pathogenesis of this infection in humans, we measured various cytokines produced by primary human macrophages infected with H5N1, pandemic H1N1 or seasonal influenza viruses. We found that many cytokines were produced at higher levels on infection with the H5N1 strains tested compared with seasonal influenza viruses. Interestingly, the extent of cytokine induction varied among the H5N1 strains and did not correlate with replicative ability in macrophages. Further, a pandemic H1N1 virus induced higher levels of several cytokines compared with seasonal viruses and some H5N1 strains. Our results demonstrate that high cytokine induction is not a universal feature of all H5N1 viruses.

In the spring of 2009, a novel influenza A virus of the H1N1 subtype emerged that transmitted efficiently among humans; by June of 2009, the outbreak reached pandemic status. The pandemic virus possesses six viral RNA segments from so-called triple reassortant swine viruses that emerged in North American pig populations in the late 1990s and two viral RNA segments from Eurasian avian-like swine influenza viruses. Most human infections with the virus have been mild; however, severe and fatal infections occurred among certain risk groups, but also among those without any known risk factors. Here, we summarize the evolutionary, epidemiological, clinical, and molecular findings on the pandemic virus. We also discuss the arsenal of antiviral compounds and vaccines available to prevent and treat infections with the virus., (© 2011 Blackwell Publishing Ltd.)

The global emergence of many severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants jeopardizes the protective antiviral immunity induced after infection or vaccination. To address the public health threat caused by the increasing SARS-CoV-2 genomic diversity, the National Institute of Allergy and Infectious Diseases within the National Institutes of Health established the SARS-CoV-2 Assessment of Viral Evolution (SAVE) programme. This effort was designed to provide a real-time risk assessment of SARS-CoV-2 variants that could potentially affect the transmission, virulence, and resistance to infection- and vaccine-induced immunity. The SAVE programme is a critical data-generating component of the US Government SARS-CoV-2 Interagency Group to assess implications of SARS-CoV-2 variants on diagnostics, vaccines and therapeutics, and for communicating public health risk. Here we describe the coordinated approach used to identify and curate data about emerging variants, their impact on immunity and effects on vaccine protection using animal models. We report the development of reagents, methodologies, models and notable findings facilitated by this collaborative approach and identify future challenges. This programme is a template for the response to rapidly evolving pathogens with pandemic potential by monitoring viral evolution in the human population to identify variants that could reduce the effectiveness of countermeasures.

The COVID-19 pandemic has served as a stark reminder that outbreaks of novel pathogens (i.e., those not previously encountered by humans) have always plagued mankind and will continue to do so. The COVID-19 pandemic has also taught us that a single exposure to a novel pathogen is typically not sufficient to build robust population immunity that exists against common respiratory viruses. Robust population-level immunity can be achieved through repeated natural infection (typically at the cost of high mortality and overwhelmed public health resources) and/or repeated vaccination (which may be limited by vaccine availability, a country's economic resources, and/or vaccine hesitancy). Here, we suggest that the broader use of antivirals could be a mitigation strategy to limit severe disease and the burden on healthcare systems during widespread virus circulation while allowing the buildup of population immunity. [ABSTRACT FROM AUTHOR]

One of the most pressing and consequential problems in infectious disease research is to better understand the potential of viruses to cause a pandemic, or, in simple terms, determine which virus will cause the next pandemic. We here define pandemics as WHO-declared pandemics, or disease outbreaks commonly referred to as pandemics that predate the WHO pandemic framework. Despite extensive research in the field of infectious diseases in recent decades, all pandemics have found us unprepared, with enormous losses of human lives, tremendous costs for public health, and vast and potentially long-lasting economic losses. Here, we discuss viruses that may cause a pandemic in the future. [ABSTRACT FROM AUTHOR]

Background: Coronavirus disease (COVID‐19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), was first detected in Japan in January 2020 and has spread throughout the country. Previous studies have reported that viral interference among influenza virus, rhinovirus, and other respiratory viruses can affect viral infections at the host and population level. Methods: To investigate the impact of COVID‐19 on influenza and other respiratory virus infections, we analyzed clinical specimens collected from 2244 patients in Japan with respiratory diseases between January 2018 and September 2020. Results: The frequency of influenza and other respiratory viruses (coxsackievirus A and B; echovirus; enterovirus; human coronavirus 229E, HKU1, NL63, and OC43; human metapneumovirus; human parainfluenza virus 1, 2, 3, and 4; human parechovirus; human respiratory syncytial virus; human adenovirus; human bocavirus; human parvovirus B19; herpes simplex virus type 1; and varicella‐zoster virus) was appreciably reduced among all patients during the COVID‐19 pandemic except for that of rhinovirus in children younger than 10 years, which was appreciably increased. COVID‐19 has not spread among this age group, suggesting an increased risk of rhinovirus infection in children. Conclusions: Rhinovirus infections should be continuously monitored to understand their increased risk during the COVID‐19 pandemic and viral interference with SARS‐CoV‐2. [ABSTRACT FROM AUTHOR]

Influenza A viruses cause respiratory infections that range from asymptomatic to deadly in humans. Widespread outbreaks (pandemics) are attributable to ‘novel’ viruses that possess a viral hemagglutinin (HA) gene to which humans lack immunity. After a pandemic, these novel viruses form stable virus lineages in humans and circulate until they are replaced by other novel viruses. The factors and mechanisms that facilitate virus transmission among hosts and the establishment of novel lineages are not completely understood, but the HA and basic polymerase 2 (PB2) proteins are thought to play essential roles in these processes by enabling avian influenza viruses to infect mammals and replicate efficiently in their new host. Here, we summarize our current knowledge of the contributions of HA, PB2, and other viral components to virus transmission and the formation of new virus lineages. [ABSTRACT FROM AUTHOR]

An influenza A virus that originated in pigs caused a pandemic in 2009. The sialidase activity of the neuraminidase (NA) of previous pandemic influenza A viruses are stable at low pH (≤5). Here, we identified the amino acids responsible for this property. We found differences in low-pH stability at pH 5.0 among pandemic (H1N1) 2009 viruses, which enhanced the replication of these viruses. Low-pH-stable NA enhancement of virus replication may have contributed to the rapid worldwide spread and adaptation to humans of pandemic (H1N1) 2009 viruses during the early stages of the 2009 pandemic. [ABSTRACT FROM AUTHOR]

Despite the high prevalence of highly pathogenic H5N1 influenza A viruses in Indonesia, epidemiology information on seasonal human influenza is lacking. The present authors, therefore, conducted virologic surveillance in Surabaya, East Java from October 2008 to March 2010. Influenza viruses, including pandemic (H1N1) 2009 viruses, were isolated from 71 of 635 individuals tested. Seasonal influenza peaked in the rainy season. Compared with seasonal influenza viruses, pandemic 2009 viruses were isolated from younger patients with milder symptoms. Given the high prevalence of H5N1 infections in humans, continued influenza surveillance is essential for pandemic preparedness. [ABSTRACT FROM AUTHOR]

Since the beginning of the 20th century, humans have experienced four influenza pandemics, including the devastating 1918 'Spanish influenza'. Moreover, H5N1 highly pathogenic avian influenza (HPAI) viruses are currently spreading worldwide, although they are not yet efficiently transmitted among humans. While the threat of a global pandemic involving a highly pathogenic influenza virus strain looms large, our mechanisms to address such a catastrophe remain limited. Here, we show that pre-stimulation of Toll-like receptors (TLRs) 2 and 4 increased resistance against influenza viruses known to induce high pathogenicity in animal models. Our data emphasize the complexity of the host response against different influenza viruses, and suggest that TLR agonists might be utilized to protect against lethality associated with highly pathogenic influenza virus infection in humans. [ABSTRACT FROM AUTHOR]

The spread of avian H5N1 influenza viruses around the globe has become a worldwide public health concern. To evaluate the pathogenic potential of reassortant viruses between currently cocirculating avian H5N1 and human H3N2 influenza viruses, we generated all the 254 combinations of reassortant viruses between A/chicken/South Kalimantan/UT6028/06 (5K06, H5N1) and Al Tokyo/Ut-Sk-1/07 (Tok07. H3N2) influenza viruses by reverse genetics. We found that the presence of Tok07 PB2 protein in the ribonucleoprotein (RNP) complex allowed efficient viral RNA tran- scription in a minigenome assay and that RNP activity played an essential role in the viability and replicative ability of the jeassortant viruses. When the pathogenicity of 75 reassortant H5 viruses was tested in mice, 22 were more pathogenic than the parental 51(06 virus, and three were extremely virulent. Strikingly, all 22 of these viruses obtained their PB2 segment from Tok07 virus. Further analysis showed that Tok07 PB 1 alone lacked the ability to enhance the pathogenicity of the reassortant viruses but could do so by cooper- ating with Tok07 PB2. Our data demonstrate that reassortment between an avian H5N1virus with low pathogenicity in mice and a human virus could result in highly pathogenic viruses and that the human virus PB2 segment functions in the background of an avian H5N1 virus, enhancing its virulence. Our findings highlight the importance of surveillance programs to monitor the emergence of human H5 reassortant viruses, especially those containing a PB2 segment of human origin. [ABSTRACT FROM AUTHOR]

Despite the annual public health burden of seasonal influenza and the continuing threat of a global pandemic posed by the emergence of highly pathogenic/pandemic strains, conventional influenza vaccines do not provide universal protection, and exhibit suboptimal efficacy rates, even when they are well matched to circulating strains. To address the need for a highly effective universal influenza vaccine, we have developed a novel M2-deficient single replication vaccine virus (M2SR) that induces strong cross-protective immunity against multiple influenza strains in mice. M2SR is able to infect cells and expresses all viral proteins except M2, but is unable to generate progeny virus. M2SR generated from influenza A/Puerto Rico/8/34 (H1N1) protected mice against lethal challenge with influenza A/Puerto Rico/8/34 (H1N1, homosubtypic) and influenza A/Aichi/2/1968 (H3N2, heterosubtypic). The vaccine induced strong systemic and mucosal antibody responses of both IgA and IgG classes. Strong virus-specific T cell responses were also induced. Following heterologous challenge, significant numbers of IFN-γ-producing CD8 T cells, with effector or effector/memory phenotypes and specific for conserved viral epitopes, were observed in the lungs of vaccinated mice. A substantial proportion of the CD8 T cells expressed Granzyme B, suggesting that they were capable of killing virus-infected cells. Thus, our data suggest that M2-deficient influenza viruses represent a promising new approach for developing a universal influenza vaccine. [ABSTRACT FROM AUTHOR]

Abstract: Currently available influenza vaccines provide suboptimal protection. In order to improve the quality of protective immune responses elicited following vaccination, we developed an oil-in-water nanoemulsion (NE)-based adjuvant for an intranasally-delivered inactivated influenza vaccine. Using a prime-boost vaccination regimen, we show that intranasal vaccines containing the W805EC NE elicited higher titers of serum hemagglutination inhibiting (HAI) antibody and influenza-specific IgG and IgA titers compared to vaccines that did not contain the NE. Similarly, vaccines containing the W805EC NE resulted in higher influenza-specific IgA levels in the bronchoalveolar lavage (BAL) fluid and nasal wash when compared to vaccines formulated without NE. The higher antibody titers in mice immunized with the NE-containing vaccines correlated with reduced viral loads in the lungs and nasal turbinates following a high dose viral challenge. Mice immunized with vaccines containing the W805EC NE also showed a reduction in body weight loss following challenge compared to mice immunized with equivalent vaccines produced without NE. Taken together, our results show that the W805EC NE substantially improves the magnitude of protective influenza-specific antibody responses and is a promising mucosal adjuvant for influenza vaccines and vaccines against other mucosal pathogens. [Copyright &y& Elsevier]

Avian A/H5N1 influenza viruses pose a pandemic threat. As few as five amino acid substitutions, or four with reassortment, might be sufficient for mammal-to-mammal transmission through respiratory droplets. From surveillance data, we found that two of these substitutions are common in A/H5N1 viruses, and thus, some viruses might require only three additional substitutions to become transmissible via respiratory droplets between mammals. We used a mathematical model of within-host virus evolution to study factors that could increase and decrease the probability of the remaining substitutions evolving after the virus has infected a mammalian host. These factors, combined with the presence of some of these substitutions in circulating strains, make a virus evolving in nature a potentially serious threat. These results highlight critical areas in which more data are needed for assessing, and potentially averting, this threat. [ABSTRACT FROM AUTHOR]

Abstract: Vaccination is a key strategy for preventing influenza virus infections. Here, we generated a reassortant virus (SC/AAca) containing the hemagglutinin and neuraminidase genes from a 2009 pandemic influenza virus A/Sichuan/1/2009 (H1N1) (SC/09) and six internal genes from the cold-adapted virus A/Ann Arbor/6/60 (H2N2) (AAca). The SC/AAca reassortant induced a sound humoral immune response and complete protection against homologous SC/09 virus challenge in mice after intranasal administration of an at least 106 50% egg infectious dose (EID50) of SC/AAca. SC/AAca inoculation also induced significant CD4+ and CD8+ T cell responses and provided solid protection against heterologous H1N1 and H5N1 virus challenge. Our results suggest that this 2009 H1N1 live vaccine will provide protection against both 2009 pandemic and seasonal H1N1 virus infection and might reduce the severity of H5N1 virus infection in humans. The induction of cross-reactive virus-specific T cell responses may be an effective approach to develop universal influenza vaccines. [Copyright &y& Elsevier]

Abstract: The 2009 influenza pandemic brought home the importance of vaccines in infection control. Previously, we demonstrated an M2 cytoplasmic tail mutant H5N1 influenza virus could serve as a live-attenuated vaccine. Here, we adapted that strategy, generating a mutant pandemic (H1N1) 2009 virus that grew well in cell culture, but replicated less well in mice than did wild-type virus. The mutant virus elicited sterile immunity in mice, completely protecting them from challenge with a pandemic (H1N1) 2009 virus. Our results indicate that M2 cytoplasmic tail mutants are suitable for live-attenuated vaccines against pandemic viruses. [Copyright &y& Elsevier]

Abstract: Outbreaks of highly pathogenic avian influenza viruses (HPAIVs) would cause disasters worldwide. Various strategies against HPAIVs are required to control damage. It is thought that the use of non-pathogenic avian influenza viruses as live vaccines will be effective in an emergency, even though there might be some adverse effects, because small amounts of live vaccines will confer immunity to protect against HPAIV infection. Therefore, live vaccines have the advantage of being able to be distributed worldwide soon after an outbreak. In the present study, we found that intranasal administration of a live H5N1 subtype non-pathogenic virus induced antibody and cytotoxic T lymphocyte responses and protected mice against H5N1 HPAIV infection. In addition, it was found that a small amount (100PFU) of the live vaccine was as effective as 100μg (approximately 1010–11 PFU of virus particles) of the inactivated whole particle vaccine in mice. Consequently, the use of live virus vaccines might be one strategy for preventing pandemics of HPAIVs in an emergency. [Copyright &y& Elsevier]

Reassortment is an important driving force for influenza virus evolution, and a better understanding of the factors that affect this process could improve our ability to respond to future influenza pandemics and epidemics. To identify factors that restrict the generation of reassortant viruses, we cotransfected human embryonic kidney cells with plasmids for the synthesis of viral RNAs of both A/equine/Prague/1/56 (Prague; H7N7) and A/Yokohama/2017/03 (Yokohama; H3N2) viruses together with the supporting protein expression plasmids. Of the possible 256 genotypes, we identified 29 genotypes in 120 randomly plaque-picked reassortants examined. Analyses of these reassortants suggested that the formation of functional ribonucleoprotein (RNP) complexes was a restricting factor, a finding that correlated with the activities of RNP complexes composed of different combinations of the proteins from the two viruses, as measured in a minigenome assay. For at least one nonfunctional RNP complex (i.e., Prague PB2, Prague PB1, Yokohama PA, and Prague NP), the lack of activity was due to the inability of the three polymerase subunit proteins to form a heterotrimer. Adaptation of viruses possessing a gene encoding a chimera of the PA proteins of the two viruses and the remaining genes from Prague virus resulted in compensatory mutations in the PB2 and/or PA protein. These results indicate substantial incompatibility among the gene products of the two test viruses, a critical role for the RNP complex in the generation of reassortant viruses, and a functional interaction of PB2 and PA. [ABSTRACT FROM AUTHOR]

Abstract: H5N1 highly pathogenic avian influenza viruses evolved into several clades, leading to appreciably distinct antigenicities of their hemagglutinins. As such, candidate H5N1 pre-pandemic vaccines for human use should be sought. Here, to evaluate fundamental immunogenic variations between H5N1 vaccines, we prepared four inactivated H5N1 test vaccines from different phylogenetic clades (clade 1, 2.1, 2.2, and 2.3.4) in accordance with the WHO recommendation, and tested their cross-clade immunity in a mouse model by vaccination followed by challenge with heterologous virulent viruses. All H5N1 vaccines tested provided full or partial cross-clade protective immunity, except one clade 2.2-based vaccine, which did not protect mice from clade 2.3.4 virus challenge. Among the test vaccines, a clade 2.1-based vaccine possessed the broadest-spectrum cross-immunity. These results suggest that currently stockpiled pre-pandemic vaccines, especially clade 2.1-based vaccines, will likely be useful as backup vaccines in a pandemic situation, even one involving antigenic-drifted viruses. [Copyright &y& Elsevier]

Viral pneumonia has been frequently reported during early stages of influenza virus pandemics and in many human cases of highly pathogenic avian influenza (HPAI) H5N1 virus infection. To better understand the pathogenesis of this disease, we produced nonlethal viral pneumonia in rhesus macaques by using an HPAI H5N1 virus (A/Anhui/2/2005; referred to as Anhui/2). Infected macaques were monitored for 14 days, and tissue samples were collected at 6 time points for virologic, histopathologic, and transcriptomic analyses. Anhui/2 efficiently replicated in the lung from 12 h to 3 days postinfection (p.i.) and caused temporal but severe pneumonia that began to resolve by day 14. Lung transcriptional changes were first observed at 6 h, and increased expression of vascular permeability regulators and neutrophil chemoattractants correlated with increased serum leakage and neutrophil infiltration in situ. Additional inflammatory, antiviral, and apoptotic genes were upregulated from 12 h, concurrent with viral antigen detection and increasing immune cell populations. A shift toward upregulation of acquired immunity was apparent after day 6. Expression levels of established immune cell molecular markers revealed remarkable similarity with pathological findings, indicating early and robust neutrophil infiltration, a slight delay in macrophage accumulation, and abundant late populations of T lymphocytes. We also characterized the putative mechanisms regulating a unique, pneumonia-associated biphasic fever pattern. Thus, this study is the first to use a comprehensive and integrative approach to delineate specific molecular mechanisms regulating influenza virus-induced pneumonia in nonhuman primates, an important first step toward better management of human influenza virus disease. [ABSTRACT FROM AUTHOR]

Outbreaks of highly pathogenic H5N1 influenza viruses in avian species began in Asia and have since spread to other continents. Concern regarding the pandemic potential of these viruses in humans is clearly warranted, and there is an urgent need to develop effective vaccines against them. Previously, we and others demonstrated that deletions of the M2 cytoplasmic tail caused a growth defect in A/WSN/33 (H1N1) influenza A virus in vitro (K. Iwatsuki-Horimoto, T. Horimoto, T. Noda, M. Kiso, J. Maeda, S. Watanabe, Y. Muramoto, K. Fujii, and Y. Kawaoka, J. Virol. 80:5233-5240, 2006; M. F. McCown and A. Pekosz, J. Virol. 79:3595-3605, 2005; M. F. McCown and A. Pekosz, J. Virol. 80:8178-8189, 2006). We therefore tested the feasibility of using M2 tail mutants as live attenuated vaccines against H5N1 virus. First we generated a series of highly pathogenic H5N1 (A/Vietnam/1203/04 [VN1203]) M2 cytoplasmic tail deletion mutants and examined their growth properties in vitro and in vivo. We found that one mutant, which contains an 11-amino-acid deletion from the C terminus (M2del11 virus), grew as well as the wild-type virus but replicated in mice less efficiently. We then generated a recombinant VN1203M2del11 virus whose hemagglutinin (HA) gene was modified by replacing sequences at the cleavage site with those of an avirulent type of HA (M2del11-HAavir virus). This M2del11-HAavir virus protected mice against challenge with lethal doses of homologous (VN1203; clade 1) and antigenically distinct heterologous (A/Indonesia/7/2005; clade 2) H5N1 viruses. Our results suggest that M2 cytoplasmic tail mutants have potential as live attenuated vaccines against H5N1 influenza viruses. [ABSTRACT FROM AUTHOR]