scholarly journals Multiple Natural Substitutions in Avian Influenza A Virus PB2 Facilitate Efficient Replication in Human Cells

2016 ◽  
Vol 90 (13) ◽  
pp. 5928-5938 ◽  
Author(s):  
Benjamin Mänz ◽  
Miranda de Graaf ◽  
Ramona Mögling ◽  
Mathilde Richard ◽  
Theo M. Bestebroer ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Avian Influenza ◽  
Mammalian Cells ◽  
Influenza A ◽  
Polymerase Activity ◽  
Host Adaptation ◽  
Influenza Viruses ◽  
Amino Acid Substitutions ◽  
Avian Influenza A Virus

ABSTRACTA strong restriction of the avian influenza A virus polymerase in mammalian cells generally limits viral host-range switching. Although substitutions like E627K in the PB2 polymerase subunit can facilitate polymerase activity to allow replication in mammals, many human H5N1 and H7N9 viruses lack this adaptive substitution. Here, several previously unknown, naturally occurring, adaptive substitutions in PB2 were identified by bioinformatics, and their enhancing activity was verified usingin vitroassays. Adaptive substitutions enhanced polymerase activity and virus replication in mammalian cells for avian H5N1 and H7N9 viruses but not for a partially human-adapted H5N1 virus. Adaptive substitutions toward basic amino acids were frequent and were mostly clustered in a putative RNA exit channel in a polymerase crystal structure. Phylogenetic analysis demonstrated divergent dependency of influenza viruses on adaptive substitutions. The novel adaptive substitutions found in this study increase basic understanding of influenza virus host adaptation and will help in surveillance efforts.IMPORTANCEInfluenza viruses from birds jump the species barrier into humans relatively frequently. Such influenza virus zoonoses may pose public health risks if the virus adapts to humans and becomes a pandemic threat. Relatively few amino acid substitutions—most notably in the receptor binding site of hemagglutinin and at positions 591 and 627 in the polymerase protein PB2—have been identified in pandemic influenza virus strains as determinants of host adaptation, to facilitate efficient virus replication and transmission in humans. Here, we show that substantial numbers of amino acid substitutions are functionally compensating for the lack of the above-mentioned mutations in PB2 and could facilitate influenza virus emergence in humans.

scholarly journals Characterization of H9N2 influenza A viruses isolated from chicken products imported into Japan from China

2006 ◽  
Vol 135 (3) ◽  
pp. 386-391 ◽  
Author(s):  
M. MASE ◽  
M. ETO ◽  
K. IMAI ◽  
K. TSUKAMOTO ◽  
S. YAMAGUCHI
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Avian Influenza ◽  
Avian Influenza Virus ◽  
Influenza A ◽  
H9n2 Virus ◽  
Amino Acid Substitutions ◽  
Influenza A Viruses ◽  
Potential Sources

We characterized eleven H9N2 influenza A viruses isolated from chicken products imported from China. Genetically they were classified into six distinct genotypes, including five already known genotypes and one novel genotype. This suggested that such multiple genotypes of the H9N2 virus have possibly already become widespread and endemic in China. Two isolates have amino-acid substitutions that confer resistance to amantadine in the M2 region, and this supported the evidence that this mutation might be a result of the wide application of amantadine for avian influenza treatment in China. These findings emphasize the importance of surveillance for avian influenza virus in this region, and of quarantining imported chicken products as potential sources for the introduction of influenza virus.

scholarly journals Evolution and Adaptation of the Avian H7N9 Virus into the Human Host

2020 ◽  
Vol 8 (5) ◽  
pp. 778
Author(s):  
Andrew T. Bisset ◽  
Gerard F. Hoyne
Keyword(s):  
Amino Acid ◽  
Avian Influenza ◽  
Influenza A ◽  
Amino Acid Sequences ◽  
Influenza Viruses ◽  
Amino Acid Substitutions ◽  
Upper Respiratory Tract ◽  
Viral Polymerase ◽  
Evolutionary Changes ◽  
Avian Origin

Influenza viruses arise from animal reservoirs, and have the potential to cause pandemics. In 2013, low pathogenic novel avian influenza A(H7N9) viruses emerged in China, resulting from the reassortment of avian-origin viruses. Following evolutionary changes, highly pathogenic strains of avian influenza A(H7N9) viruses emerged in late 2016. Changes in pathogenicity and virulence of H7N9 viruses have been linked to potential mutations in the viral glycoproteins hemagglutinin (HA) and neuraminidase (NA), as well as the viral polymerase basic protein 2 (PB2). Recognizing that effective viral transmission of the influenza A virus (IAV) between humans requires efficient attachment to the upper respiratory tract and replication through the viral polymerase complex, experimental evidence demonstrates the potential H7N9 has for increased binding affinity and replication, following specific amino acid substitutions in HA and PB2. Additionally, the deletion of extended amino acid sequences in the NA stalk length was shown to produce a significant increase in pathogenicity in mice. Research shows that significant changes in transmissibility, pathogenicity and virulence are possible after one or a few amino acid substitutions. This review aims to summarise key findings from that research. To date, all strains of H7N9 viruses remain restricted to avian reservoirs, with no evidence of sustained human-to-human transmission, although mutations in specific viral proteins reveal the efficacy with which these viruses could evolve into a highly virulent and infectious, human-to-human transmitted virus.

scholarly journals Plasticity of the Influenza Virus H5 HA Protein

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Huihui Kong ◽  
David F. Burke ◽  
Tiago Jose da Silva Lopes ◽  
Kosuke Takada ◽  
Masaki Imai ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Mammalian Cells ◽  
Influenza A ◽  
Viral Antigen ◽  
Influenza Viruses ◽  
Influenza A Viruses ◽  
Highly Pathogenic ◽  
Antigenic Properties ◽  
Ha Protein

ABSTRACT Since the emergence of highly pathogenic avian influenza viruses of the H5 subtype, the major viral antigen, hemagglutinin (HA), has undergone constant evolution, resulting in numerous genetic and antigenic (sub)clades. To explore the consequences of amino acid changes at sites that may affect the antigenicity of H5 viruses, we simultaneously mutated 17 amino acid positions of an H5 HA by using a synthetic gene library that, theoretically, encodes all combinations of the 20 amino acids at the 17 positions. All 251 mutant viruses sequenced possessed ≥13 amino acid substitutions in HA, demonstrating that the targeted sites can accommodate a substantial number of mutations. Selection with ferret sera raised against H5 viruses of different clades resulted in the isolation of 39 genotypes. Further analysis of seven variants demonstrated that they were antigenically different from the parental virus and replicated efficiently in mammalian cells. Our data demonstrate the substantial plasticity of the influenza virus H5 HA protein, which may lead to novel antigenic variants. IMPORTANCE The HA protein of influenza A viruses is the major viral antigen. In this study, we simultaneously introduced mutations at 17 amino acid positions of an H5 HA expected to affect antigenicity. Viruses with ≥13 amino acid changes in HA were viable, and some had altered antigenic properties. H5 HA can therefore accommodate many mutations in regions that affect antigenicity. The substantial plasticity of H5 HA may facilitate the emergence of novel antigenic variants.

scholarly journals Resurrected ancestral influenza A viruses recapitulate the avian-to-mammalian adaptation of European avian-like swine influenza virus

2021 ◽  
Author(s):  
Wen Su ◽  
Rhodri Harfoot ◽  
Yvonne Su ◽  
Jennifer DeBeauchamp ◽  
Udayan Joseph ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Influenza A ◽  
Swine Influenza Virus ◽  
Swine Influenza ◽  
Polymerase Activity ◽  
Influenza Viruses ◽  
Mammalian Host ◽  
Influenza A Viruses ◽  
Sequence Reconstruction

Abstract The emergence of a pandemic influenza virus may be better anticipated if we better understand the evolutionary steps taken by avian influenza viruses as they adapt to mammals. We used ancestral sequence reconstruction to resurrect viruses representing initial adaptive stages of the European avian-like H1N1 virus as it transitioned from avian to swine hosts. We demonstrate that efficient transmissibility in pigs was gained through stepwise adaptation after 1983. These time-dependent adaptations resulted in changes in hemagglutinin receptor binding specificity and increased viral polymerase activity. An NP-R351K mutation under strong positive selection increased the transmissibility of a reconstructed virus. The stepwise-adaptation of a wholly avian influenza virus to a mammalian host suggests a window where targeted intervention may have highest impact. Successful intervention will, however, require strategic coordination of surveillance and risk assessment activities to identify these adapting viruses and guide pandemic preparedness resources.

scholarly journals Genetic diversification of H5N1 highly pathogenic avian influenza A virus during replication in wild ducks

2011 ◽  
Vol 92 (9) ◽  
pp. 2105-2110 ◽  
Author(s):  
Yohei Watanabe ◽  
Madiha S. Ibrahim ◽  
Hany F. Ellakany ◽  
Hatem S. Abd El-Hamid ◽  
Kazuyoshi Ikuta
Keyword(s):  
Amino Acid ◽  
Avian Influenza ◽  
Influenza A Virus ◽  
Influenza A ◽  
Highly Pathogenic Avian Influenza ◽  
Amino Acid Substitutions ◽  
Highly Pathogenic ◽  
Pathogenic Avian Influenza ◽  
Novel Variants ◽  
Avian Influenza A Virus

Highly pathogenic avian influenza A virus subtype H5N1 can potentially generate novel variants during replication of infected hosts. To determine which H5N1 variants predominate in wild birds, we determined the sequences of RT-PCR amplified viral genes from several organs of infected chickens and ducks from Egypt, where H5N1 outbreaks in birds are endemic. Comparison of the sequences in viruses from trachea, lung, brain and liver revealed diversification with different amino acid substitutions in different ducks, but no diversification in chickens. These specific amino acid substitutions were rare among viruses currently circulating in Egypt. In addition, the H5N1 variants showed distinct growth kinetics in duck, canine and human cells. Our findings suggested that ducks can generate H5N1 variants with novel amino acid substitutions that might serve as aetiological agents for new influenza virus outbreaks and epidemics.

scholarly journals Predominance of a Drifted Influenza A (H3N2) Clade and Its Association with Age-Specific Influenza Vaccine Effectiveness Variations, Influenza Season 2018–2019

Vaccines ◽  
2020 ◽  
Vol 8 (1) ◽  
pp. 78 ◽  
Author(s):  
Aharona Glatman-Freedman ◽  
Rakefet Pando ◽  
Hanna Sefty ◽  
Itay Omer ◽  
Alina Rosenberg ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Influenza A ◽  
Gene Mutations ◽  
Vaccine Virus ◽  
Vaccine Effectiveness ◽  
Influenza Viruses ◽  
Amino Acid Substitutions ◽  
Ha Gene ◽  
H3n2 Vaccine

Background: Influenza A (H3N2) clade 3C.3a was the predominant influenza virus in Israel throughout the 2018-2019 season, constituting a drift from the influenza A (H3N2) vaccine. We estimated the end-of season vaccine effectiveness (VE) by age, among community patients with influenza-like illness (ILI), considering the hemagglutinin (HA) gene mutations and amino acid substitutions of influenza A (H3N2) viruses detected. Methods: Nose-throat samples were analyzed for the presence of influenza virus, type/subtype, and HA gene sequence. HA gene sequences and amino acid substitutions were compared to the influenza A/Singapore/INFIMH-16-0019/2016 (H3N2)-like 2018-2019 vaccine virus, and a phylogenetic tree was generated. Influenza VE against influenza A (H3N2) was estimated using the test-negative design. VE was estimated by age group and by 15 year moving age intervals. Results: In total, 90% of the influenza A (H3N2) viruses belonged to the 3C.3a clade, constituting a unique situation in the northern hemisphere. Adjusted all-age influenza A (H3N2) VE was −3.5% (95% CI: −51.2 to 29.1). Although adjusted VEs were very low among infants, children, and young adults, a VE of 45% (95% CI: −19.2 to 74.6) was estimated among adults aged ≥45 years old. Conclusions: The higher VE point estimates among older adults may be related to previous exposure to similar influenza viruses.

scholarly journals S-Acylation of Proteins of Coronavirus and Influenza Virus: Conservation of Acylation Sites in Animal Viruses and DHHC Acyltransferases in Their Animal Reservoirs

Pathogens ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 669
Author(s):  
Dina A. Abdulrahman ◽  
Xiaorong Meng ◽  
Michael Veit
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Ion Channels ◽  
Substrate Specificity ◽  
Influenza A ◽  
3D Structure ◽  
Viral Fitness ◽  
Amino Acid Substitutions ◽  
Essential Function ◽  
Animal Reservoirs

Recent pandemics of zoonotic origin were caused by members of coronavirus (CoV) and influenza A (Flu A) viruses. Their glycoproteins (S in CoV, HA in Flu A) and ion channels (E in CoV, M2 in Flu A) are S-acylated. We show that viruses of all genera and from all hosts contain clusters of acylated cysteines in HA, S and E, consistent with the essential function of the modification. In contrast, some Flu viruses lost the acylated cysteine in M2 during evolution, suggesting that it does not affect viral fitness. Members of the DHHC family catalyze palmitoylation. Twenty-three DHHCs exist in humans, but the number varies between vertebrates. SARS-CoV-2 and Flu A proteins are acylated by an overlapping set of DHHCs in human cells. We show that these DHHC genes also exist in other virus hosts. Localization of amino acid substitutions in the 3D structure of DHHCs provided no evidence that their activity or substrate specificity is disturbed. We speculate that newly emerged CoVs or Flu viruses also depend on S-acylation for replication and will use the human DHHCs for that purpose. This feature makes these DHHCs attractive targets for pan-antiviral drugs.

scholarly journals Development of Neuraminidase Subtype-Specific Reference Antisera by Recombinant Protein Expressed in Baculovirus

2012 ◽  
Vol 20 (2) ◽  
pp. 140-145 ◽  
Author(s):  
Kyu-Jun Lee ◽  
Jun-Gu Choi ◽  
Hyun-Mi Kang ◽  
Kwang-Il Kim ◽  
Choi-Kyu Park ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Influenza A Virus ◽  
Diagnostic Tests ◽  
Influenza A ◽  
Recombinant Baculovirus ◽  
Cross Reactivity ◽  
Specific Reference ◽  
Simple Method ◽  
Avian Influenza A Virus

ABSTRACTOutbreaks of avian influenza A virus infection, particularly the H5N1 strains that have affected birds and some humans for the past 15 years, have highlighted the need for increased surveillance and disease control. Such measures require diagnostic tests to detect and characterize the different subtypes of influenza virus. In the current study, a simple method for producing reference avian influenza virus antisera to be used in diagnostic tests was developed. Antisera of nine avian influenza A virus neuraminidases (NA) used for NA subtyping were produced using a recombinant baculovirus. The recombinant NA (rNA) proteins were expressed in Sf9 insect cells and inoculated intramuscularly into specific-pathogen-free chickens with the ISA70 adjuvant. The NA inhibition antibody titers of the rNA antiserum were in the ranges of 5 to 8 and 6 to 9 log2units after the primary and boost immunizations, respectively. The antisera were subtype specific, showing low cross-reactivity against every other NA subtype using the conventional thiobarbituric acid NA inhibition assay. These results suggest that this simple method for producing reference NA antisera without purification may be useful for the diagnosis and surveillance of influenza virus.

scholarly journals Functional Analysis of PA Binding by Influenza A Virus PB1: Effects on Polymerase Activity and Viral Infectivity

2001 ◽  
Vol 75 (17) ◽  
pp. 8127-8136 ◽  
Author(s):  
Daniel R. Perez ◽  
Ruben O. Donis
Keyword(s):  
Amino Acids ◽  
Influenza Virus ◽  
Amino Acid ◽  
Viral Replication ◽  
Influenza A Virus ◽  
Influenza A ◽  
Amino Acid Sequences ◽  
Influenza Viruses ◽  
Virus Growth ◽  
Transcription And Replication

ABSTRACT Influenza A virus expresses three viral polymerase (P) subunits—PB1, PB2, and PA—all of which are essential for RNA and viral replication. The functions of P proteins in transcription and replication have been partially elucidated, yet some of these functions seem to be dependent on the formation of a heterotrimer for optimal viral RNA transcription and replication. Although it is conceivable that heterotrimer subunit interactions may allow a more efficient catalysis, direct evidence of their essentiality for viral replication is lacking. Biochemical studies addressing the molecular anatomy of the P complexes have revealed direct interactions between PB1 and PB2 as well as between PB1 and PA. Previous studies have shown that the N-terminal 48 amino acids of PB1, termed domain α, contain the residues required for binding PA. We report here the refined mapping of the amino acid sequences within this small region of PB1 that are indispensable for binding PA by deletion mutagenesis of PB1 in a two-hybrid assay. Subsequently, we used site-directed mutagenesis to identify the critical amino acid residues of PB1 for interaction with PA in vivo. The first 12 amino acids of PB1 were found to constitute the core of the interaction interface, thus narrowing the previous boundaries of domain α. The role of the minimal PB1 domain α in influenza virus gene expression and genome replication was subsequently analyzed by evaluating the activity of a set of PB1 mutants in a model reporter minigenome system. A strong correlation was observed between a functional PA binding site on PB1 and P activity. Influenza viruses bearing mutant PB1 genes were recovered using a plasmid-based influenza virus reverse genetics system. Interestingly, mutations that rendered PB1 unable to bind PA were either nonviable or severely growth impaired. These data are consistent with an essential role for the N terminus of PB1 in binding PA, P activity, and virus growth.

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