scholarly journals Glycosylation generates an efficacious and immunogenic vaccine against H7N9 influenza virus

PLoS Biology ◽  
2020 ◽  
Vol 18 (12) ◽  
pp. e3001024
Author(s):  
Jin Il Kim ◽  
Sehee Park ◽  
Joon-Yong Bae ◽  
Sunmi Lee ◽  
Jeonghun Kim ◽  
...  
Keyword(s):  
Avian Influenza ◽  
Viral Replication ◽  
Protective Efficacy ◽  
Case Fatality ◽  
Vaccine Virus ◽  
Human Infection ◽  
Influenza Viruses ◽  
Evolutionary Patterns ◽  
Viral Hemagglutinin ◽  
Ha Protein

Zoonotic avian influenza viruses pose severe health threats to humans. Of several viral subtypes reported, the low pathogenic avian influenza H7N9 virus has since February 2013 caused more than 1,500 cases of human infection with an almost 40% case-fatality rate. Vaccination of poultry appears to reduce human infections. However, the emergence of highly pathogenic strains has increased concerns about H7N9 pandemics. To develop an efficacious H7N9 human vaccine, we designed vaccine viruses by changing the patterns of N-linked glycosylation (NLG) on the viral hemagglutinin (HA) protein based on evolutionary patterns of H7 HA NLG changes. Notably, a virus in which 2 NLG modifications were added to HA showed higher growth rates in cell culture and elicited more cross-reactive antibodies than did other vaccine viruses with no change in the viral antigenicity. Developed into an inactivated vaccine formulation, the vaccine virus with 2 HA NLG additions exhibited much better protective efficacy against lethal viral challenge in mice than did a vaccine candidate with wild-type (WT) HA by reducing viral replication in the lungs. In a ferret model, the 2 NLG-added vaccine viruses also induced hemagglutination-inhibiting antibodies and significantly suppressed viral replication in the upper and lower respiratory tracts compared with the WT HA vaccines. In a mode of action study, the HA NLG modification appeared to increase HA protein contents incorporated into viral particles, which would be successfully translated to improve vaccine efficacy. These results suggest the strong potential of HA NLG modifications in designing avian influenza vaccines.

scholarly journals Genetic and biological properties of H7N9 avian influenza viruses detected after application of the H7N9 poultry vaccine in China

2021 ◽  
Vol 17 (4) ◽  
pp. e1009561
Author(s):  
Xin Yin ◽  
Guohua Deng ◽  
Xianying Zeng ◽  
Pengfei Cui ◽  
Yujie Hou ◽  
...  
Keyword(s):  
Avian Influenza ◽  
Animal Studies ◽  
Biological Properties ◽  
Human Infection ◽  
Influenza Viruses ◽  
Glycosylation Sites ◽  
Amino Acid Mutations ◽  
Poultry Vaccine ◽  
Ha Protein ◽  
And Control

The H7N9 avian influenza virus (AIV) that emerged in China have caused five waves of human infection. Further human cases have been successfully prevented since September 2017 through the use of an H7N9 vaccine in poultry. However, the H7N9 AIV has not been eradicated from poultry in China, and its evolution remains largely unexplored. In this study, we isolated 19 H7N9 AIVs during surveillance and diagnosis from February 2018 to December 2019, and genetic analysis showed that these viruses have formed two different genotypes. Animal studies indicated that the H7N9 viruses are highly lethal to chicken, cause mild infection in ducks, but have distinct pathotypes in mice. The viruses bound to avian-type receptors with high affinity, but gradually lost their ability to bind to human-type receptors. Importantly, we found that H7N9 AIVs isolated in 2019 were antigenically different from the H7N9 vaccine strain that was used for H7N9 influenza control in poultry, and that replication of these viruses cannot, therefore, be completely prevented in vaccinated chickens. We further revealed that two amino acid mutations at positions 135 and 160 in the HA protein added two glycosylation sites and facilitated the escape of the H7N9 viruses from the vaccine-induced immunity. Our study provides important insights into H7N9 virus evolution and control.

scholarly journals Vaccination with Consensus H7 Elicits Broadly Reactive and Protective Antibodies against Eurasian and North American Lineage H7 Viruses

Vaccines ◽  
2020 ◽  
Vol 8 (1) ◽  
pp. 143
Author(s):  
Gendeal M. Fadlallah ◽  
Fuying Ma ◽  
Zherui Zhang ◽  
Mengchan Hao ◽  
Juefu Hu ◽  
...  
Keyword(s):  
Avian Influenza ◽  
North American ◽  
Human Infection ◽  
Influenza Viruses ◽  
Avian Influenza Viruses ◽  
Lethal Challenge ◽  
Protective Antibodies ◽  
H7 Subtype ◽  
H7n3 Virus

H7 subtype avian influenza viruses have caused outbreaks in poultry, and even human infection, for decades in both Eurasia and North America. Although effective vaccines offer the best protection against avian influenza viruses, antigenically distinct Eurasian and North American lineage subtype H7 viruses require the development of cross-protective vaccine candidates. In this study, a methodology called computationally optimized broadly reactive antigen (COBRA) was used to develop four consensus H7 antigens (CH7-22, CH7-24, CH7-26, and CH7-28). In vitro experiments confirmed the binding of monoclonal antibodies to the head and stem domains of cell surface-expressed consensus HAs, indicating display of their antigenicity. Immunization with DNA vaccines encoding the four antigens was evaluated in a mouse model. Broadly reactive antibodies against H7 viruses from Eurasian and North American lineages were elicited and detected by binding, inhibition, and neutralizing analyses. Further infection with Eurasian H7N9 and North American H7N3 virus strains confirmed that CH7-22 and CH7-24 conferred the most effective protection against hetero-lethal challenge. Our data showed that the consensus H7 vaccines elicit a broadly reactive, protective response against Eurasian and North American lineage H7 viruses, which are suitable for development against other zoonotic influenza viruses.

scholarly journals Development of a duplex TaqMan real-time RT-PCR assay for simultaneous detection of newly emerged H5N6 influenza viruses

2019 ◽  
Vol 16 (1) ◽  
Author(s):  
Lin Liu ◽  
Ying Zhang ◽  
Pengfei Cui ◽  
Congcong Wang ◽  
Xianying Zeng ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Simultaneous Detection ◽  
Reaction System ◽  
Taxonomic Status ◽  
Human Infection ◽  
Influenza Viruses ◽  
Rt Pcr ◽  
Avian Influenza Viruses ◽  
Pcr Assay

Abstract Background In 2017–2018, a new highly pathogenic H5N6 avian influenza virus (AIV) variant appeared in poultry and wild birds in Asian and European countries and caused multiple outbreaks. These variant strains are different from the H5N6 virus associated with human infection in previous years, and their genetic taxonomic status and antigenicity have changed. Therefore, revision of the primers and probes of fluorescent RT-PCR is important to detect the new H5N6 subtype AIV in poultry and reduce the risk of an epidemic in birds or humans. Methods In this study, the primers and probes including three groups of HA and four groups of NA for H5N6 influenza virus were evaluated. Then a set of ideal primer and probes were selected to further optimize the reaction system and established a method of double rRT-PCR assay. The specificity of this method was determined by using H1~H16 subtype AIV. Results The results showed that fluorescence signals were obtained for H5 virus in FAM channel and N6 virus in VIC channel, and no fluorescent signal was observed in other subtypes of avian influenza viruses. The detection limit of this assay was 69 copies for H5 and 83 copies for N6 gene. And, the variability tests of intra- and inter-assay showed excellent reproducibility. Moreover, this assay showed 100% agreement with virus isolation method in detecting samples from poultry. Conclusion The duplex rRT-PCR assay presented here has high specificity, sensitivity and reproducibility, and can be used for laboratory surveillance and rapid diagnosis of newly emerged H5N6 subtype avian influenza viruses.

scholarly journals Phenotypic Effects of Substitutions within the Receptor Binding Site of Highly Pathogenic Avian Influenza H5N1 Virus Observed during Human Infection

2020 ◽  
Vol 94 (13) ◽  
Author(s):  
Dirk Eggink ◽  
Monique Spronken ◽  
Roosmarijn van der Woude ◽  
Jocynthe Buzink ◽  
Frederik Broszeit ◽  
...  
Keyword(s):  
Amino Acid ◽  
Avian Influenza ◽  
Binding Site ◽  
Receptor Binding ◽  
Human Infection ◽  
Influenza Viruses ◽  
Receptor Specificity ◽  
Receptor Binding Site ◽  
Human Type ◽  
Human Infections

ABSTRACT Highly pathogenic avian influenza (HPAI) viruses are enzootic in wild birds and poultry and continue to cause human infections with high mortality. To date, more than 850 confirmed human cases of H5N1 virus infection have been reported, of which ∼60% were fatal. Global concern persists that these or similar avian influenza viruses will evolve into viruses that can transmit efficiently between humans, causing a severe influenza pandemic. It was shown previously that a change in receptor specificity is a hallmark for adaptation to humans and evolution toward a transmittable virus. Substantial genetic diversity was detected within the receptor binding site of hemagglutinin of HPAI A/H5N1 viruses, evolved during human infection, as detected by next-generation sequencing. Here, we investigated the functional impact of substitutions that were detected during these human infections. Upon rescue of 21 mutant viruses, most substitutions in the receptor binding site (RBS) resulted in viable virus, but virus replication, entry, and stability were often impeded. None of the tested substitutions individually resulted in a clear switch in receptor preference as measured with modified red blood cells and glycan arrays. Although several combinations of the substitutions can lead to human-type receptor specificity, accumulation of multiple amino acid substitutions within a single hemagglutinin during human infection is rare, thus reducing the risk of virus adaptation to humans. IMPORTANCE H5 viruses continue to be a threat for public health. Because these viruses are immunologically novel to humans, they could spark a pandemic when adapted to transmit between humans. Avian influenza viruses need several adaptive mutations to bind to human-type receptors, increase hemagglutinin (HA) stability, and replicate in human cells. However, knowledge on adaptive mutations during human infections is limited. A previous study showed substantial diversity within the receptor binding site of H5N1 during human infection. We therefore analyzed the observed amino acid changes phenotypically in a diverse set of assays, including virus replication, stability, and receptor specificity. None of the tested substitutions resulted in a clear step toward a human-adapted virus capable of aerosol transmission. It is notable that acquiring human-type receptor specificity needs multiple amino acid mutations, and that variability at key position 226 is not tolerated, reducing the risk of them being acquired naturally.

scholarly journals Immune Escape Adaptive Mutations in the H7N9 Avian Influenza Hemagglutinin Protein Increase Virus Replication Fitness and Decrease Pandemic Potential

2020 ◽  
Vol 94 (19) ◽  
Author(s):  
Pengxiang Chang ◽  
Joshua E. Sealy ◽  
Jean-Remy Sadeyen ◽  
Sushant Bhat ◽  
Deimante Lukosaityte ◽  
...  
Keyword(s):  
Thermal Stability ◽  
Avian Influenza ◽  
Virus Replication ◽  
Receptor Binding ◽  
Immune Escape ◽  
Influenza Viruses ◽  
Escape Mutant ◽  
Hemagglutinin Protein ◽  
Ha Protein ◽  
The Impact

ABSTRACT H7N9 avian influenza viruses (AIVs) continue to evolve and remain a huge threat to human health and the poultry industry. Previously, serially passaging the H7N9 A/Anhui/1/2013 virus in the presence of homologous ferret antiserum resulted in immune escape viruses containing amino acid substitutions alanine to threonine at residues 125 (A125T) and 151 (A151T) and leucine to glutamine at residue 217 (L217Q) in the hemagglutinin (HA) protein. These HA mutations have also been found in field isolates in 2019. To investigate the potential threat of serum escape mutant viruses to humans and poultry, the impact of these HA substitutions, either individually or in combination, on receptor binding, pH of fusion, thermal stability, and virus replication were investigated. Our results showed the serum escape mutant formed large plaques in Madin-Darby canine kidney (MDCK) cells and grew robustly in vitro and in ovo. They had a lower pH of fusion and increased thermal stability. Of note, the serum escape mutant completely lost the ability to bind to human-like receptor analogues. Further analysis revealed that N-linked glycosylation, as a result of A125T or A151T substitutions in HA, resulted in reduced receptor-binding avidity toward both human and avian-like receptor analogues, and the A125T+A151T mutations completely abolished human-like receptor binding. The L217Q mutation enhanced the H7N9 acid and thermal stability while the A151T mutation dramatically decreased H7N9 HA thermal stability. To conclude, H7N9 AIVs that contain A125T+A151T+L217Q mutations in the HA protein may pose a reduced pandemic risk but remain a heightened threat for poultry. IMPORTANCE Avian influenza H7N9 viruses have been causing disease outbreaks in poultry and humans. We previously determined that propagation of H7N9 virus in virus-specific antiserum gives rise to mutant viruses carrying mutations A125T+A151T+L217Q in their hemagglutinin protein, enabling the virus to overcome vaccine-induced immunity. As predicted, these immune escape mutations were also observed in the field viruses that likely emerged in the immunized or naturally exposed birds. This study demonstrates that the immune escape mutants also (i) gained greater replication ability in cultured cells and in chicken embryos as well as (ii) increased acid and thermal stability but (iii) lost preferences for binding to human-type receptor while maintaining binding for the avian-like receptor. Therefore, they potentially pose reduced pandemic risk. However, the emergent virus variants containing the indicated mutations remain a significant risk to poultry due to antigenic drift and improved fitness for poultry.

scholarly journals Structural and Molecular Characterization of the Hemagglutinin from the Fifth-Epidemic-Wave A(H7N9) Influenza Viruses

2018 ◽  
Vol 92 (16) ◽  
Author(s):  
Hua Yang ◽  
Paul J. Carney ◽  
Jessie C. Chang ◽  
Zhu Guo ◽  
James Stevens
Keyword(s):  
Public Health ◽  
Avian Influenza ◽  
Influenza A ◽  
Human Infection ◽  
Influenza Viruses ◽  
Surface Proteins ◽  
Health Concern ◽  
Public Health Concern ◽  
H7n9 Influenza ◽  
Epidemic Wave

ABSTRACTThe avian influenza A(H7N9) virus continues to cause human infections in China and is a major ongoing public health concern. Five epidemic waves of A(H7N9) infection have occurred since 2013, and the recent fifth epidemic wave saw the emergence of two distinct lineages with elevated numbers of human infection cases and broader geographic distribution of viral diseases compared to the first four epidemic waves. Moreover, highly pathogenic avian influenza (HPAI) A(H7N9) viruses were also isolated during the fifth epidemic wave. Here, we present a detailed structural and biochemical analysis of the surface hemagglutinin (HA) antigen from viruses isolated during this recent epidemic wave. Results highlight that, compared to the 2013 virus HAs, the fifth-wave virus HAs remained a weak binder to human glycan receptor analogs. We also studied three mutations, V177K-K184T-G219S, that were recently reported to switch a 2013 A(H7N9) HA to human-type receptor specificity. Our results indicate that these mutations could also switch the H7 HA receptor preference to a predominantly human binding specificity for both fifth-wave H7 HAs analyzed in this study.IMPORTANCEThe A(H7N9) viruses circulating in China are of great public health concern. Here, we report a molecular and structural study of the major surface proteins from several recent A(H7N9) influenza viruses. Our results improve the understanding of these evolving viruses and provide important information on their receptor preference that is central to ongoing pandemic risk assessment.

scholarly journals A New Generation of Modified Live-Attenuated Avian Influenza Viruses Using a Two-Strategy Combination as Potential Vaccine Candidates

2007 ◽  
Vol 81 (17) ◽  
pp. 9238-9248 ◽  
Author(s):  
Haichen Song ◽  
Gloria Ramirez Nieto ◽  
Daniel R. Perez
Keyword(s):  
Single Dose ◽  
Avian Influenza ◽  
H5n1 Virus ◽  
Vaccine Virus ◽  
Mass Vaccination ◽  
Influenza Viruses ◽  
Temperature Sensitive ◽  
Pathogenic Avian Influenza ◽  
Hpai H5n1 ◽  
H5n1 Vaccine

ABSTRACT In light of the recurrent outbreaks of low pathogenic avian influenza (LPAI) and highly pathogenic avian influenza (HPAI), there is a pressing need for the development of vaccines that allow rapid mass vaccination. In this study, we introduced by reverse genetics temperature-sensitive mutations in the PB1 and PB2 genes of an avian influenza virus, A/Guinea Fowl/Hong Kong/WF10/99 (H9N2) (WF10). Further genetic modifications were introduced into the PB1 gene to enhance the attenuated (att) phenotype of the virus in vivo. Using the att WF10 as a backbone, we substituted neuraminidase (NA) for hemagglutinin (HA) for vaccine purposes. In chickens, a vaccination scheme consisting of a single dose of an att H7N2 vaccine virus at 2 weeks of age and subsequent challenge with the wild-type H7N2 LPAI virus resulted in complete protection. We further extended our vaccination strategy against the HPAI H5N1. In this case, we reconstituted an att H5N1 vaccine virus, whose HA and NA genes were derived from an Asian H5N1 virus. A single-dose immunization in ovo with the att H5N1 vaccine virus in 18-day-old chicken embryos resulted in more than 60% protection for 4-week-old chickens and 100% protection for 9- to 12-week-old chickens. Boosting at 2 weeks posthatching provided 100% protection against challenge with the HPAI H5N1 virus for chickens as young as 4 weeks old, with undetectable virus shedding postchallenge. Our results highlight the potential of live att avian influenza vaccines for mass vaccination in poultry.

scholarly journals Influenza at the animal–human interface: a review of the literature for virological evidence of human infection with swine or avian influenza viruses other than A(H5N1)

2014 ◽  
Vol 19 (18) ◽  
Author(s):  
G S Freidl ◽  
A Meijer ◽  
E de Bruin ◽  
M de Nardi ◽  
O Munoz ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Influenza A ◽  
Swine Influenza Virus ◽  
Swine Influenza ◽  
Human Infection ◽  
Influenza Viruses ◽  
Diagnostic Methods ◽  
World Health ◽  
Review Of The Literature

Factors that trigger human infection with animal influenza virus progressing into a pandemic are poorly understood. Within a project developing an evidence-based risk assessment framework for influenza viruses in animals, we conducted a review of the literature for evidence of human infection with animal influenza viruses by diagnostic methods used. The review covering Medline, Embase, SciSearch and CabAbstracts yielded 6,955 articles, of which we retained 89; for influenza A(H5N1) and A(H7N9), the official case counts of the World Health Organization were used. An additional 30 studies were included by scanning the reference lists. Here, we present the findings for confirmed infections with virological evidence. We found reports of 1,419 naturally infected human cases, of which 648 were associated with avian influenza virus (AIV) A(H5N1), 375 with other AIV subtypes, and 396 with swine influenza virus (SIV). Human cases naturally infected with AIV spanned haemagglutinin subtypes H5, H6, H7, H9 and H10. SIV cases were associated with endemic SIV of H1 and H3 subtype descending from North American and Eurasian SIV lineages and various reassortants thereof. Direct exposure to birds or swine was the most likely source of infection for the cases with available information on exposure.

scholarly journals Phylogeography and Antigenic Diversity of Low-Pathogenic Avian Influenza H13 and H16 Viruses

2020 ◽  
Vol 94 (13) ◽  
Author(s):  
Josanne H. Verhagen ◽  
Marjolein Poen ◽  
David E. Stallknecht ◽  
Stefan van der Vliet ◽  
Pascal Lexmond ◽  
...  
Keyword(s):  
Gene Flow ◽  
Avian Influenza ◽  
Evolutionary History ◽  
Antigenic Variation ◽  
Wild Birds ◽  
Influenza Viruses ◽  
Avian Influenza Viruses ◽  
Pathogenic Avian Influenza ◽  
Low Pathogenic Avian Influenza ◽  
Ha Protein

ABSTRACT Low-pathogenic avian influenza viruses (LPAIVs) are genetically highly variable and have diversified into multiple evolutionary lineages that are primarily associated with wild-bird reservoirs. Antigenic variation has been described for mammalian influenza viruses and for highly pathogenic avian influenza viruses that circulate in poultry, but much less is known about antigenic variation of LPAIVs. In this study, we focused on H13 and H16 LPAIVs that circulate globally in gulls. We investigated the evolutionary history and intercontinental gene flow based on the hemagglutinin (HA) gene and used representative viruses from genetically distinct lineages to determine their antigenic properties by hemagglutination inhibition assays. For H13, at least three distinct genetic clades were evident, while for H16, at least two distinct genetic clades were evident. Twenty and ten events of intercontinental gene flow were identified for H13 and H16 viruses, respectively. At least two antigenic variants of H13 and at least one antigenic variant of H16 were identified. Amino acid positions in the HA protein that may be involved in the antigenic variation were inferred, and some of the positions were located near the receptor binding site of the HA protein, as they are in the HA protein of mammalian influenza A viruses. These findings suggest independent circulation of H13 and H16 subtypes in gull populations, as antigenic patterns do not overlap, and they contribute to the understanding of the genetic and antigenic variation of LPAIVs naturally circulating in wild birds. IMPORTANCE Wild birds play a major role in the epidemiology of low-pathogenic avian influenza viruses (LPAIVs), which are occasionally transmitted—directly or indirectly—from them to other species, including domestic animals, wild mammals, and humans, where they can cause subclinical to fatal disease. Despite a multitude of genetic studies, the antigenic variation of LPAIVs in wild birds is poorly understood. Here, we investigated the evolutionary history, intercontinental gene flow, and antigenic variation among H13 and H16 LPAIVs. The circulation of subtypes H13 and H16 seems to be maintained by a narrower host range, in particular gulls, than the majority of LPAIV subtypes and may therefore serve as a model for evolution and epidemiology of H1 to H12 LPAIVs in wild birds. The findings suggest that H13 and H16 LPAIVs circulate independently of each other and emphasize the need to investigate within-clade antigenic variation of LPAIVs in wild birds.

scholarly journals The Origin of Human-infecting Avian Influenza A H6N1 Virus

2013 ◽  
Author(s):  
Liangsheng Zhang ◽  
Zhenguo Zhang
Keyword(s):  
Phylogenetic Analysis ◽  
Avian Influenza ◽  
Influenza A ◽  
Human Infection ◽  
Influenza Viruses ◽  
Avian Influenza Viruses ◽  
Know How

In this study, we retraced the origin of the reported avian influenza A H6N1 virus infecting a 20-year-old woman in Taiwan. As we know, this is the first reported case of human infection by the H6N1 virus, because this subtype virus usually circulates in birds and poultry. Therefore it is crucial to know how this virus attained the ability to infect humans. Using phylogenetic analysis, we found that this virus was derived from reassortments of multiple lineages of H6N1 viruses and H5N2 viruses. The results deepen our understanding of how the new human-infecting virus originated and based on these we discussed possible explanations for the H6N1 infection of humans. Our results, together with recent studies of H7N9 viruses which result in severe disorders, suggest that reassortments among avian-type viruses are quite often, which may sometimes result in fatal infections in humans. Thus a close watch on the circulation of avian influenza viruses is pretty necessary.

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