- A Naturally Occurring Deletion in the Effector Domain of H5N1 Swine Influenza Virus Nonstructural Protein 1 Regulates Viral Fitness and Host Innate Immunity. [Journal Article]
- JVJ Virol 2018 Mar 21
- Nonstructural protein 1 (NS1) of influenza A virus regulates innate immune responses via various mechanisms. We previously showed that a naturally occurring deletion (the EALQR motif) in the NS1 effe...
Nonstructural protein 1 (NS1) of influenza A virus regulates innate immune responses via various mechanisms. We previously showed that a naturally occurring deletion (the EALQR motif) in the NS1 effector domain of an H5N1 swine-origin avian influenza virus impairs the inhibition of type I interferon (IFN) in chicken fibroblasts and attenuates virulence in chickens. Here, we found that the virus bearing this deletion in its NS1 effector domain showed diminished inhibition of IFN-related cytokine expression and attenuated virulence in mice. We further show that deletion of the EALQR motif disrupts NS1 dimerization, impairing double-stranded RNA (dsRNA) sequestration and competitive binding with RIG-I. In addition, the EALQR-deleted NS1 could not bind to TRIM25, unlike full-length NS1, and was less able to block TRIM25 oligomerization and self-ubiquitination, further impairing the inhibition of TRIM25-mediated RIG-I ubiquitination compared with full-length NS1. Our data demonstrate that the EALQR deletion prevents NS1 from blocking RIG-I-mediated IFN induction via a novel mechanism to attenuate the viral replication and virulence in mammalian cells and animals.IMPORTANCEH5 highly pathogenic avian influenza viruses have infected more than 800 individuals with an overall 53% case fatality rate across 16 countries. Among viral proteins, nonstructural protein 1 (NS1) of influenza virus is considered a key determinant for type I interferon (IFN) antagonism, pathogenicity, and host range. However, that how NS1 precisely modulates virus-host interaction facilitating virus survival is not fully understood. Here, we report that a naturally occurring deletion (the EALQR motif) in the NS1 effector domain of an H5N1 swine-origin avian influenza virus disrupted the NS1 dimerization, which diminished the blockade of IFN induction via the RIG-I signaling pathway, thereby impairing virus replication and virulence in the host. Our study demonstrates the EALQR motif of NS1 regulates virus fitness to attain a virus-host compromise state in animals, and identifies this critical motif as a potential target for the future development of small molecular drugs and attenuated vaccines.
- Nucleosides for the treatment of respiratory RNA virus infections. [Journal Article]
- ACAntivir Chem Chemother 2018 Jan-Dec; 26:2040206618764483
- Influenza virus, respiratory syncytial virus, human metapneumovirus, parainfluenza virus, coronaviruses, and rhinoviruses are among the most common viruses causing mild seasonal colds. These RNA viru...
Influenza virus, respiratory syncytial virus, human metapneumovirus, parainfluenza virus, coronaviruses, and rhinoviruses are among the most common viruses causing mild seasonal colds. These RNA viruses can also cause lower respiratory tract infections leading to bronchiolitis and pneumonia. Young children, the elderly, and patients with compromised cardiac, pulmonary, or immune systems are at greatest risk for serious disease associated with these RNA virus respiratory infections. In addition, swine and avian influenza viruses, together with severe acute respiratory syndrome-associated and Middle Eastern respiratory syndrome coronaviruses, represent significant pandemic threats to the general population. In this review, we describe the current medical need resulting from respiratory infections caused by RNA viruses, which justifies drug discovery efforts to identify new therapeutic agents. The RNA polymerase of respiratory viruses represents an attractive target for nucleoside and nucleotide analogs acting as inhibitors of RNA chain synthesis. Here, we present the molecular, biochemical, and structural fundamentals of the polymerase of the four major families of RNA respiratory viruses: Orthomyxoviridae, Pneumoviridae/Paramyxoviridae, Coronaviridae, and Picornaviridae. We summarize past and current efforts to develop nucleoside and nucleotide analogs as antiviral agents against respiratory virus infections. This includes molecules with very broad antiviral spectrum such as ribavirin and T-705 (favipiravir), and others targeting more specifically one or a few virus families. Recent advances in our understanding of the structure(s) and function(s) of respiratory virus polymerases will likely support the discovery and development of novel nucleoside analogs.
- Survival of viral pathogens in animal feed ingredients under transboundary shipping models. [Journal Article]
- PlosPLoS One 2018; 13(3):e0194509
- The goal of this study was to evaluate survival of important viral pathogens of livestock in animal feed ingredients imported daily into the United States under simulated transboundary conditions. El...
The goal of this study was to evaluate survival of important viral pathogens of livestock in animal feed ingredients imported daily into the United States under simulated transboundary conditions. Eleven viruses were selected based on global significance and impact to the livestock industry, including Foot and Mouth Disease Virus (FMDV), Classical Swine Fever Virus (CSFV), African Swine Fever Virus (ASFV), Influenza A Virus of Swine (IAV-S), Pseudorabies virus (PRV), Nipah Virus (NiV), Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), Swine Vesicular Disease Virus (SVDV), Vesicular Stomatitis Virus (VSV), Porcine Circovirus Type 2 (PCV2) and Vesicular Exanthema of Swine Virus (VESV). Surrogate viruses with similar genetic and physical properties were used for 6 viruses. Surrogates belonged to the same virus families as target pathogens, and included Senecavirus A (SVA) for FMDV, Bovine Viral Diarrhea Virus (BVDV) for CSFV, Bovine Herpesvirus Type 1 (BHV-1) for PRV, Canine Distemper Virus (CDV) for NiV, Porcine Sapelovirus (PSV) for SVDV and Feline Calicivirus (FCV) for VESV. For the remaining target viruses, actual pathogens were used. Virus survival was evaluated using Trans-Pacific or Trans-Atlantic transboundary models involving representative feed ingredients, transport times and environmental conditions, with samples tested by PCR, VI and/or swine bioassay. SVA (representing FMDV), FCV (representing VESV), BHV-1 (representing PRV), PRRSV, PSV (representing SVDV), ASFV and PCV2 maintained infectivity during transport, while BVDV (representing CSFV), VSV, CDV (representing NiV) and IAV-S did not. Notably, more viruses survived in conventional soybean meal, lysine hydrochloride, choline chloride, vitamin D and pork sausage casings. These results support published data on transboundary risk of PEDV in feed, demonstrate survival of certain viruses in specific feed ingredients ("high-risk combinations") under conditions simulating transport between continents and provide further evidence that contaminated feed ingredients may represent a risk for transport of pathogens at domestic and global levels.
- Swine Flu: Knowledge, Attitude, and Practices Survey of Medical and Dental Students of Karachi. [Journal Article]
- CCureus 2018 Jan 09; 10(1):e2048
- Introduction Pakistan is extremely susceptible to an influenza outbreak, as it shares borders with the most affected countries, namely China and India. The medical and dental students come into direc...
Introduction Pakistan is extremely susceptible to an influenza outbreak, as it shares borders with the most affected countries, namely China and India. The medical and dental students come into direct contact with the affected population and should be aware of the risk factors and signs and symptoms pertaining to swine influenza virus (SIV). Hence, this survey was conducted to assess the knowledge, perceptions and self-care practices of the medical and dental students with regards to this pandemic. Methods A descriptive, cross-sectional study was conducted to evaluate the swine flu-related knowledge, attitudes and practices of the medical and dental students at various institutions in Karachi, Pakistan. We approached 613 students that were available on the dates of this survey, keeping a medical to dental student ratio of 75:25. All students from first to final year comprised of the study population, and no internists or medical personnel were included. The questionnaire was divided into three sections, namely knowledge, attitudes and, practices. All questions were based on a multiple choice format. The data were entered and interpreted using the IBM Statistical Package for the Social Sciences 23.0 (IBM Corp., Armonk, New York). Results The majority of the students were aware that the swine flu is a transmittable disease (n=485, 80.8%). Most students identified the signs and symptoms correctly; however, diarrhea (15.5%) and vomiting (32.2%) were the least correct answers (n=93, n=193 respectively). Most of the preventative measures were reported accurately by the participants. Despite this, only 15.5% students (n=93) reported the use of a facemask when suffering from fever, cough and a runny nose. Conclusion There is a dire need for the routine integration of the awareness and management programs in the medical and dental schools. There exists a gap between the policy and practice, and it is high time we bridge the divide. The students should also be vaccinated annually for influenza A.
- Antigenically diverse swine-origin H1N1 variant influenza viruses exhibit differential ferret pathogenesis and transmission phenotypes. [Journal Article]
- JVJ Virol 2018 Mar 14
- Influenza A(H1) viruses circulating in swine represent an emerging virus threat as zoonotic infections occur sporadically following exposure to swine. A fatal infection caused by an H1N1 variant (H1N...
Influenza A(H1) viruses circulating in swine represent an emerging virus threat as zoonotic infections occur sporadically following exposure to swine. A fatal infection caused by an H1N1 variant (H1N1v) virus was detected in a patient with reported exposure to swine and who presented with pneumonia, respiratory failure, and cardiac arrest. To understand the genetic and phenotypic characteristics of the virus, genome sequence analysis, antigenic characterization, and ferret pathogenesis and transmissibility experiments were performed. Antigenic analysis of the virus isolated from the fatal case, A/Ohio/09/2015, demonstrated significant antigenic drift away from classical swine H1N1 variant viruses and H1N1 pandemic 2009 viruses. A substitution in the H1 hemagglutinin (G155E) was identified that likely impacted antigenicity, and reverse genetics was employed to understand the molecular mechanism of antibody escape. Reversion of the substitution to 155G, in a reverse genetics A/Ohio/09/2015 virus, showed that this residue was central to the loss of hemagglutination inhibition by ferret antisera raised against a prototypical H1N1 pandemic 2009 virus (A/California/07/2009), as well as gamma lineage classical swine H1N1 viruses, demonstrating the importance of this residue for antibody recognition of this H1 lineage. When analyzed in the ferret model, A/Ohio/09/2015 and another H1N1v virus (A/Iowa/39/2015), as well as A/California/07/2009, replicated efficiently in the respiratory tract of ferrets. The two H1N1v viruses transmitted efficiently among cohoused ferrets, but respiratory droplet transmission studies showed that A/California/07/2009 transmitted through the air more efficiently. Pre-existing immunity to A/California/07/2009 did not fully protect ferrets from challenge with A/Ohio/09/2015.IMPORTANCEHuman infections with classical swine influenza A(H1N1) viruses that circulate in pigs continue to occur in the United States following exposure to swine. To understand the genetic and virologic characteristics of a virus (A/Ohio/09/2015) associated with a fatal infection and a virus associated with a non-fatal infection (A/Iowa/39/2015), we performed genome sequence analysis, antigenic testing, and pathogenicity and transmission studies in a ferret model. Reverse genetics was employed to identify a single antigenic site substitution (HA G155E) responsible for antigenic variation of A/Ohio/09/2015 compared to related classical swine influenza A(H1N1) viruses. Ferrets with pre-existing immunity to the pandemic A(H1N1) virus were challenged with A/Ohio/09/2015 demonstrating decreased protection. This data illustrates the potential for currently circulating swine influenza viruses to infect and cause illness in humans with pre-existing immunity to H1N1 pandemic 2009 viruses and a need for ongoing risk assessment and development of candidate vaccine viruses for improved pandemic preparedness.
- Maternally-derived antibodies do not inhibit swine influenza virus replication in piglets but decrease excreted virus infectivity and impair post-infectious immune responses. [Journal Article]
- VMVet Microbiol 2018; 216:142-152
- Maternally-derived antibodies (MDA) reduce piglet susceptibility to swine influenza A virus, but interfere with post-infectious immune responses, raising questions about protection after waning of pa...
Maternally-derived antibodies (MDA) reduce piglet susceptibility to swine influenza A virus, but interfere with post-infectious immune responses, raising questions about protection after waning of passive immunity. We therefore analysed the impact of different levels of residual MDA on virus excretion and immune responses in piglets born to vaccinated sows (MDA+) and infected with H1N1 at 5, 7 or 11 weeks of age, in comparison to piglets born to unvaccinated sows (MDA-). Subsequent protection against a second homologous infection occurring 4 weeks after the primo-infection was also investigated. MDA- pigs showed clinical signs, shed the virus, and developed specific immune responses despite some age-dependent differences: 7-week-old pigs were less affected clinically, showed a 2-day delayed excretion peak and excreted less virus than younger pigs. In MDA+ animals, clinical signs increased together with the decrease of MDA levels related to the age at infection-time. Virus shedding was not prevented and genome quantification profiles were similar to those obtained in MDA- piglets. However, viral particles excreted by 5-week-old MDA+ piglets appeared to be less infectious than those shed by MDA- piglets at the same age. Humoral response was affected by MDA as illustrated by the absence of HI and neutralizing response regardless the infection age, but anti-NP/M responses were less affected. Proliferative T cell responses were slightly delayed by high MDA levels. Nevertheless, MDA+ animals were all protected from a second infection, like MDA- piglets. In conclusion, responses of pigs to H1N1 were affected by both the physiological development of animals at infection and the MDA level.
- Appearance of reassortant European avian-origin H1 influenza A viruses of swine in Vietnam. [Journal Article]
- TETransbound Emerg Dis 2018 Mar 06
- Three subtypes-H1N1, H1N2 and H3N2-of influenza A viruses of swine (IAVs-S) are currently endemic in swine worldwide, but there is considerable genotypic diversity among each subtype and limited geog...
Three subtypes-H1N1, H1N2 and H3N2-of influenza A viruses of swine (IAVs-S) are currently endemic in swine worldwide, but there is considerable genotypic diversity among each subtype and limited geographical distribution. Through IAVs-S monitoring in Vietnam, two H1N2 influenza A viruses were isolated from healthy pigs in Ba Ria-Vung Tau Province, Southern Vietnam, on 2 December 2016. BLAST and phylogenetic analyses revealed that their HA and NA genes were derived from those of European avian-like H1N2 IAVs-S that contained avian-origin H1 and human-like N2 genes, and were particularly closely related to those of IAVs-S circulating in the Netherlands, Germany or Denmark. In addition, the internal genes of these Vietnamese isolates were derived from human A(H1N1)pdm09 viruses, suggesting that the Vietnamese H1N2 IAVs-S are reassortants between European H1N2 IAVs-S and human A(H1N1)pdm09v. The appearance of European avian-like H1N2 IAVs-S in Vietnam marks their first transmission outside Europe. Our results and statistical analyses of the number of live pigs imported into Vietnam suggest that the European avian-like H1N2 IAVs-S may have been introduced into Vietnam with their hosts through international trade. These findings highlight the importance of quarantining imported pigs to impede the introduction of new IAVs-S.
- Molecular evolution of H1N1 swine influenza in Guangdong, China, 2016-2017. [Journal Article]
- IGInfect Genet Evol 2018 Feb 26; 60:103-108
- Swine are the main host of the H1N1 swine influenza virus (SIV), however, H1N1 can also infect humans and occasionally cause serious respiratory disease. To trace the evolution of the SIV in Guangdon...
Swine are the main host of the H1N1 swine influenza virus (SIV), however, H1N1 can also infect humans and occasionally cause serious respiratory disease. To trace the evolution of the SIV in Guangdong, China, we performed an epidemic investigation during the period of 2016-2017. Nine H1N1 influenza viruses were isolated from swine nasal swabs. Antigenic analysis revealed that these viruses belonged to two distinct antigenic groups, represented by A/Swine/Guangdong/101/2016 and A/Swine/Guangdong/52/2017. Additionally, three genotypes, known as GD52/17-like, GD493/17-like and GD101/16-like, were identified by phylogenetic analysis. Importantly, the genotypes including a minimum of 4 pdm/09-origin internal genes have become prevalent in China in recent years. A total of 2966 swine serum samples were used to perform hemagglutination inhibition (HI) tests, and the results showed that the seroprevalence values of SW/GD/101/16 (32.2% in 2016, 32.1% in 2017) were significantly higher than the seroprevalence values of SW/GD/52/17 (18.0% in 2016, 16.7% in 2017). Our study showed that the three reassortant genotypes of H1N1 SIV currently circulating in China are stable, but H1N1pdm09 poses challenges to human health by the introduction of internal genes into these reassortant genotypes. Strengthening SIV surveillance is therefore critical for SIV control and minimizing its potential threat to public health.
- Universal Vaccines and Vaccine Platforms to Protect against Influenza Viruses in Humans and Agriculture. [Review]
- FMFront Microbiol 2018; 9:123
- Influenza virus infections pose a significant threat to public health due to annual seasonal epidemics and occasional pandemics. Influenza is also associated with significant economic losses in anima...
Influenza virus infections pose a significant threat to public health due to annual seasonal epidemics and occasional pandemics. Influenza is also associated with significant economic losses in animal production. The most effective way to prevent influenza infections is through vaccination. Current vaccine programs rely heavily on the vaccine's ability to stimulate neutralizing antibody responses to the hemagglutinin (HA) protein. One of the biggest challenges to an effective vaccination program lies on the fact that influenza viruses are ever-changing, leading to antigenic drift that results in escape from earlier immune responses. Efforts toward overcoming these challenges aim at improving the strength and/or breadth of the immune response. Novel vaccine technologies, the so-called universal vaccines, focus on stimulating better cross-protection against many or all influenza strains. However, vaccine platforms or manufacturing technologies being tested to improve vaccine efficacy are heterogeneous between different species and/or either tailored for epidemic or pandemic influenza. Here, we discuss current vaccines to protect humans and animals against influenza, highlighting challenges faced to effective and uniform novel vaccination strategies and approaches.
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- Synonymous Mutations at the Beginning of the Influenza A Virus Hemagglutinin Gene Impact Experimental Fitness. [Journal Article]
- JMJ Mol Biol 2018 Feb 18
- The fitness effects of synonymous mutations can provide insights into biological and evolutionary mechanisms. We analyzed the experimental fitness effects of all single-nucleotide mutations, includin...
The fitness effects of synonymous mutations can provide insights into biological and evolutionary mechanisms. We analyzed the experimental fitness effects of all single-nucleotide mutations, including synonymous substitutions, at the beginning of the influenza A virus hemagglutinin (HA) gene. Many synonymous substitutions were deleterious both in bulk competition and for individually isolated clones. Investigating protein and RNA levels of a subset of individually expressed HA variants revealed that multiple biochemical properties contribute to the observed experimental fitness effects. Our results indicate that a structural element in the HA segment viral RNA may influence fitness. Examination of naturally evolved sequences in human hosts indicates a preference for the unfolded state of this structural element compared to that found in swine hosts. Our overall results reveal that synonymous mutations may have greater fitness consequences than indicated by simple models of sequence conservation, and we discuss the implications of this finding for commonly used evolutionary tests and analyses.