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Purpose: Meniscal injuries are common in knee surgery and often require preservation techniques to prevent secondary osteoarthritis. Despite advancements in repair techniques, some patients undergo partial meniscectomy, which can lead to postmeniscectomy syndrome. To address these challenges, meniscal substitution techniques like scaffolds have been developed. However, a comprehensive synthesis of the existing evidence through an umbrella review is lacking. Methods: A comprehensive search was conducted in the MEDLINE, Embase and Scopus databases to identify relevant systematic reviews and meta-analyses. Studies were screened based on predefined inclusion and exclusion criteria. The quality of included studies was assessed using the AMSTAR-2 tool. Results: A total of 17 studies met the inclusion criteria and were included in the review. Most studies focused on the use of collagen-based scaffolds, with fewer studies evaluating synthetic scaffolds. The majority of studies (52.9%) were rated as having 'Critically Low' overall confidence, with only one study (5.9%) rated as 'High' confidence and most studies exhibiting methodological limitations, such as small sample sizes and lack of long-term follow-up. Despite these limitations, the majority of studies reported positive short-term outcomes, including pain relief and functional improvement, following scaffold implantation. However, some studies noted a relatively high failure rate. Radiographically, outcomes also varied, with some studies reporting morphological deterioration of the implant seen on MRI, while others noted possible chondroprotective effects. Conclusions: Meniscal scaffold-based approaches show promise in the management of meniscal deficiency; however, the current evidence is limited by methodological shortcomings. One notable gap in the literature is the lack of clear guidelines for patient selection and surgical technique. Future research should focus on conducting well-designed randomized controlled trials with long-term follow-up to further elucidate the benefits and indications of these techniques in clinical practice. Additionally, efforts should be made to develop consensus guidelines to standardize the use of meniscal scaffolds and improve patient outcomes. Despite limited availability, synthesizing the literature on meniscal scaffold-based approaches is crucial for understanding research, guiding clinical decisions and informing future directions. Level of Evidence: Level IV.
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Chickens genetically resistant to avian influenza could prevent future outbreaks. In chickens, influenza A virus (IAV) relies on host protein ANP32A. Here we use CRISPR/Cas9 to generate homozygous gene edited (GE) chickens containing two ANP32A amino acid substitutions that prevent viral polymerase interaction. After IAV challenge, 9/10 edited chickens remain uninfected. Challenge with a higher dose, however, led to breakthrough infections. Breakthrough IAV virus contained IAV polymerase gene mutations that conferred adaptation to the edited chicken ANP32A. Unexpectedly, this virus also replicated in chicken embryos edited to remove the entire ANP32A gene and instead co-opted alternative ANP32 protein family members, chicken ANP32B and ANP32E. Additional genome editing for removal of ANP32B and ANP32E eliminated all viral growth in chicken cells. Our data illustrate a first proof of concept step to generate IAV-resistant chickens and show that multiple genetic modifications will be required to curtail viral escape.
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Vírus da Influenza A , Influenza Aviária , Embrião de Galinha , Animais , Influenza Aviária/genética , Edição de Genes , RNA Polimerase Dependente de RNA/genética , RNA Polimerase Dependente de RNA/metabolismo , Galinhas/genética , Vírus da Influenza A/genética , Vírus da Influenza A/metabolismoRESUMO
ANP32 proteins, which act as influenza polymerase cofactors, vary between birds and mammals. In mammals, ANP32A and ANP32B have been reported to serve essential but redundant roles to support influenza polymerase activity. The well-known mammalian adaptation PB2-E627K enables influenza polymerase to use mammalian ANP32 proteins. However, some mammalian-adapted influenza viruses do not harbor this substitution. Here, we show that alternative PB2 adaptations, Q591R and D701N, also allow influenza polymerase to use mammalian ANP32 proteins, whereas other PB2 mutations, G158E, T271A, and D740N, increase polymerase activity in the presence of avian ANP32 proteins as well. Furthermore, PB2-E627K strongly favors use of mammalian ANP32B proteins, whereas D701N shows no such bias. Accordingly, PB2-E627K adaptation emerges in species with strong pro-viral ANP32B proteins, such as humans and mice, while D701N is more commonly seen in isolates from swine, dogs, and horses, where ANP32A proteins are the preferred cofactor. Using an experimental evolution approach, we show that the passage of viruses containing avian polymerases in human cells drove acquisition of PB2-E627K, but not in the absence of ANP32B. Finally, we show that the strong pro-viral support of ANP32B for PB2-E627K maps to the low-complexity acidic region (LCAR) tail of ANP32B. IMPORTANCE Influenza viruses naturally reside in wild aquatic birds. However, the high mutation rate of influenza viruses allows them to rapidly and frequently adapt to new hosts, including mammals. Viruses that succeed in these zoonotic jumps pose a pandemic threat whereby the virus adapts sufficiently to efficiently transmit human-to-human. The influenza virus polymerase is central to viral replication and restriction of polymerase activity is a major barrier to species jumps. ANP32 proteins are essential for influenza polymerase activity. In this study, we describe how avian influenza viruses can adapt in several different ways to use mammalian ANP32 proteins. We further show that differences between mammalian ANP32 proteins can select different adaptive changes and are responsible for some of the typical mutations that arise in mammalian-adapted influenza polymerases. These different adaptive mutations may determine the relative zoonotic potential of influenza viruses and thus help assess their pandemic risk.
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Vírus da Influenza A , Influenza Aviária , Influenza Humana , Proteínas Nucleares , Animais , Cães , Humanos , Camundongos , Proteínas de Ciclo Celular/metabolismo , Cavalos , Vírus da Influenza A/genética , Vírus da Influenza A/metabolismo , Influenza Aviária/genética , Influenza Humana/genética , Mamíferos , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Nucleotidiltransferases/metabolismo , Proteínas de Ligação a RNA/metabolismo , RNA Polimerase Dependente de RNA/genética , RNA Polimerase Dependente de RNA/metabolismo , Suínos , Proteínas Virais/genética , Proteínas Virais/metabolismo , Replicação ViralRESUMO
Perioperative management for patients undergoing shoulder arthroplasty has evolved significantly over the years to reduce overt complications and improve patient outcomes. The groundwork for perioperative care encompasses initial patient selection and education strategies for achieving successful outcome. Multimodal pain management strategies have advanced patient care with the increased use of new regional/local anesthetics. In addition, complications resulting from blood loss and transfusions have been curtailed with the use of synthetic antifibrinolytic agents. It remains critical for shoulder arthroplasty surgeons to optimize patients during the perioperative period through various modalities to maximize functional progression, outcomes, and patient's satisfaction following shoulder arthroplasty.
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Antifibrinolíticos , Artroplastia do Ombro , Anestésicos Locais , Artroplastia do Ombro/efeitos adversos , Humanos , Manejo da Dor/métodos , Dor Pós-Operatória/etiologiaRESUMO
PURPOSE: To assess the national trends in arthroscopic and open rotator cuff repair surgery and the associated demographics, complications, and risk factors specific to each procedure. METHODS: A retrospective cohort study was performed using the National Surgical Quality Improvement Program (NSQIP) dataset between the years 2007 and 2018. Patients were identified using Common Procedural Terminology codes for open and arthroscopic rotator cuff repair. Variables collected including basic demographics, procedural, and outcome specific variables as available through the NSQIP repository. Appropriate statistical measures were used to compare the groups, with the χ2 test used for categorical variables and t test for continuous variables. RESULTS: The arthroscopic cohort comprised 39,013 patients; the open group consisted of 8,664. Reported arthroscopic and open cases increased significantly between 2007 and 2018 from 135 to 7,269 and 65 to 1,168, respectively. Average operative time for arthroscopic procedure was 89 minutes and 76 minutes for open. The open group consisted of a slightly greater percentage of smokers, 18.3% versus 15.2%, and patients with diabetes, 18.2% versus 15.9%, both of which were statistically significant (P < .001). Open cases had an odds ratio of 3.05 for superficial infections and 7.40 for deep infections, both of which were statistically significant (P < .001). The open cohort also had an odds ratio of 1.71 for unplanned readmissions when compared with the arthroscopic cohort, which was also statistically significant (P < .001). CONCLUSIONS: According to the NSQIP database, the increase in arthroscopic procedures is significantly outpacing the increase in open procedures during this study period, which matches the trends seen in previous studies. Patients with diabetes and who smoke also represent a greater risk group for postoperative complications when undergoing open surgery. These findings suggest that perhaps the decision to pursue one technique over the other may be influenced both by provider preference and patient-related factors. LEVEL OF EVIDENCE: III, retrospective comparative trial.
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Lesões do Manguito Rotador , Manguito Rotador , Artroscopia/efeitos adversos , Artroscopia/métodos , Demografia , Humanos , Reoperação , Estudos Retrospectivos , Fatores de Risco , Manguito Rotador/cirurgia , Lesões do Manguito Rotador/etiologia , Lesões do Manguito Rotador/cirurgia , Resultado do TratamentoRESUMO
Highly pathogenic H5N1 avian influenza viruses cause devastating outbreaks in farmed poultry with serious consequences for animal welfare and economic losses. Zoonotic infection of humans through close contact with H5N1 infected birds is often severe and fatal. England experienced an outbreak of H5N1 in turkeys in 1991 that led to thousands of farmed bird mortalities. Isolation of clonal populations of one such virus from this outbreak uncovered amino acid differences in the virus haemagglutinin (HA) gene whereby the different genotypes could be associated with distinct pathogenic outcomes in chickens; both low pathogenic (LP) and high pathogenic (HP) phenotypes could be observed despite all containing a multi-basic cleavage site (MBCS) in the HA gene. Using reverse genetics, three amino acid substitutions in HA were examined for their ability to affect pathogenesis in the chicken. Restoration of amino acid polymorphisms close to the receptor binding site that are commonly found in H5 viruses only partially improved viral fitness in vitro and in vivo. A third novel substitution in the fusion peptide, HA2G4R, enabled the HP phenotype. HA2G4R decreased the pH stability of HA and increased the pH of HA fusion. The substitutions close to the receptor binding site optimised receptor binding while modulating the pH of HA fusion. Importantly, this study revealed pathogenic determinants beyond the MBCS.
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Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/patogenicidade , Influenza Aviária/virologia , Doenças das Aves Domésticas/virologia , Receptores Virais/metabolismo , Substituição de Aminoácidos , Animais , Fusão Celular , Galinhas , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Concentração de Íons de Hidrogênio , Virus da Influenza A Subtipo H5N1/isolamento & purificação , Virus da Influenza A Subtipo H5N1/metabolismo , Influenza Aviária/genética , Influenza Aviária/metabolismo , Doenças das Aves Domésticas/genética , Doenças das Aves Domésticas/metabolismo , Ligação Proteica , Receptores Virais/genética , VirulênciaRESUMO
Avian influenza viruses occasionally infect and adapt to mammals, including humans. Swine are often described as "mixing vessels," being susceptible to both avian- and human-origin viruses, which allows the emergence of novel reassortants, such as the precursor to the 2009 H1N1 pandemic. ANP32 proteins are host factors that act as influenza virus polymerase cofactors. In this study, we describe how swine ANP32A, uniquely among the mammalian ANP32 proteins tested, supports the activity of avian-origin influenza virus polymerases and avian influenza virus replication. We further show that after the swine-origin influenza virus emerged in humans and caused the 2009 pandemic, it evolved polymerase gene mutations that enabled it to more efficiently use human ANP32 proteins. We map the enhanced proviral activity of swine ANP32A to a pair of amino acids, 106 and 156, in the leucine-rich repeat and central domains and show these mutations enhance binding to influenza virus trimeric polymerase. These findings help elucidate the molecular basis for the mixing vessel trait of swine and further our understanding of the evolution and ecology of viruses in this host.IMPORTANCE Avian influenza viruses can jump from wild birds and poultry into mammalian species such as humans or swine, but they only continue to transmit if they accumulate mammalian adapting mutations. Pigs appear uniquely susceptible to both avian and human strains of influenza and are often described as virus "mixing vessels." In this study, we describe how a host factor responsible for regulating virus replication, ANP32A, is different between swine and humans. Swine ANP32A allows a greater range of influenza viruses, specifically those from birds, to replicate. It does this by binding the virus polymerase more tightly than the human version of the protein. This work helps to explain the unique properties of swine as mixing vessels.
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Vírus da Influenza A Subtipo H1N1/genética , Proteínas Nucleares/genética , Infecções por Orthomyxoviridae/genética , Proteínas de Ligação a RNA/genética , RNA Polimerase Dependente de RNA/genética , Proteínas Virais/genética , Animais , Sítios de Ligação , Linhagem Celular , Galinhas , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Regulação da Expressão Gênica , Especificidade de Hospedeiro , Humanos , Vírus da Influenza A Subtipo H1N1/metabolismo , Modelos Moleculares , Mutação , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologia , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , RNA Polimerase Dependente de RNA/química , RNA Polimerase Dependente de RNA/metabolismo , Transdução de Sinais , Suínos , Proteínas Virais/química , Proteínas Virais/metabolismo , Replicação ViralRESUMO
The avian-origin influenza A virus polymerase is restricted in human cells. This restriction has been associated with species differences in host factor ANP32A. Avian ANP32A supports the activity of an avian-origin polymerase. However, the avian-origin polymerase is incompatible with human ANP32A. Avian ANP32A proteins harbor an additional 33 amino acids compared to human ANP32A proteins, which are crucial for their ability to support the avian-origin influenza virus polymerase. Here, we elucidate the interactions between ANP32A proteins and the influenza A virus polymerase using split luciferase complementation assays, coimmunoprecipitation, and in situ split Venus interaction assays. We show greater interaction of chicken ANP32A than human ANP32A with the viral polymerase and visualize these interactions in situ in the cell nucleus. We demonstrate that the 33 amino acids of chicken ANP32A and the PB2 627 domain of viral polymerase complex both contribute to this enhanced interaction. Finally, we show how these interactions are affected by the presence of viral RNA and the processivity of the polymerase, giving insights into the way that ANP32A might act during virus infection.IMPORTANCE Successful zoonotic transmission of influenza A virus into humans can lead to pandemics in an immunologically naive population. Host-encoded ANP32A proteins are required to support influenza A virus polymerase activity, and species differences in ANP32A can restrict the host range of influenza virus. Understanding how ANP32A proteins support the viral polymerase and how differences in ANP32A affect the ability of the polymerase to coopt these proteins will enhance our understanding of viral replication and species restriction as well as suggesting targeted antiviral approaches to treat influenza virus infection.
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Especificidade de Hospedeiro , Proteínas Nucleares/metabolismo , Proteínas de Ligação a RNA/metabolismo , RNA Polimerase Dependente de RNA/metabolismo , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Animais , Proteínas Aviárias/química , Proteínas Aviárias/genética , Proteínas Aviárias/metabolismo , Galinhas/genética , Galinhas/virologia , Células HEK293 , Humanos , Imunoprecipitação , Vírus da Influenza A/enzimologia , Influenza Aviária/virologia , Influenza Humana/virologia , Luciferases , Proteínas Nucleares/genética , Infecções por Orthomyxoviridae/virologia , RNA Viral , Proteínas de Ligação a RNA/genética , Replicação ViralRESUMO
ANP32 proteins have been implicated in supporting influenza virus replication, but most of the work to date has focused on the ability of avian Anp32 proteins to overcome restriction of avian influenza polymerases in human cells. Using a CRISPR approach, we show that the human acidic nuclear phosphoproteins (ANPs) ANP32A and ANP32B are functionally redundant but essential host factors for mammalian-adapted influenza A virus (IAV) and influenza B virus (IBV) replication in human cells. When both proteins are absent from human cells, influenza polymerases are unable to replicate the viral genome, and infectious virus cannot propagate. Provision of exogenous ANP32A or ANP32B recovers polymerase activity and virus growth. We demonstrate that this redundancy is absent in the murine Anp32 orthologues; murine Anp32A is incapable of recovering IAV polymerase activity, while murine Anp32B can do so. Intriguingly, IBV polymerase is able to use murine Anp32A. We show, using a domain swap and point mutations, that the leucine-rich repeat (LRR) 5 region comprises an important functional domain for mammalian ANP32 proteins. Our approach has identified a pair of essential host factors for influenza virus replication and can be harnessed to inform future interventions.IMPORTANCE Influenza virus is the etiological agent behind some of the most devastating infectious disease pandemics to date, and influenza outbreaks still pose a major threat to public health. Influenza virus polymerase, the molecule that copies the viral RNA genome, hijacks cellular proteins to support its replication. Current anti-influenza drugs are aimed against viral proteins, including the polymerase, but RNA viruses like influenza tend to become resistant to such drugs very rapidly. An alternative strategy is to design therapeutics that target the host proteins that are necessary for virus propagation. Here, we show that the human proteins ANP32A and ANP32B are essential for influenza A and B virus replication, such that in their absence cells become impervious to the virus. We map the proviral activity of ANP32 proteins to one region in particular, which could inform future intervention.
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Betainfluenzavirus/fisiologia , Proteínas de Ciclo Celular/genética , Vírus da Influenza A/fisiologia , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/genética , Proteínas de Ligação a RNA/genética , RNA Polimerase Dependente de RNA/metabolismo , Animais , Sistemas CRISPR-Cas , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Modelos Animais de Doenças , Humanos , Camundongos , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Mutação Puntual , Domínios Proteicos , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Proteínas Virais/metabolismo , Replicação ViralRESUMO
Influenza A viruses (IAV) are subject to species barriers that prevent frequent zoonotic transmission and pandemics. One of these barriers is the poor activity of avian IAV polymerases in human cells. Differences between avian and mammalian ANP32 proteins underlie this host range barrier. Human ANP32A and ANP32B homologues both support function of human-adapted influenza polymerase but do not support efficient activity of avian IAV polymerase which requires avian ANP32A. We show here that the gene currently designated as avian ANP32B is evolutionarily distinct from mammalian ANP32B, and that chicken ANP32B does not support IAV polymerase activity even of human-adapted viruses. Consequently, IAV relies solely on chicken ANP32A to support its replication in chicken cells. Amino acids 129I and 130N, accounted for the inactivity of chicken ANP32B. Transfer of these residues to chicken ANP32A abolished support of IAV polymerase. Understanding ANP32 function will help develop antiviral strategies and aid the design of influenza virus resilient genome edited chickens.
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Especificidade de Hospedeiro , Interações Hospedeiro-Patógeno , Vírus da Influenza A/crescimento & desenvolvimento , Proteínas Nucleares/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Linhagem Celular , Galinhas , Humanos , Vírus da Influenza A/enzimologia , RNA Polimerase Dependente de RNA/metabolismo , Replicação ViralRESUMO
In Figure 4, seasonal influenza virus was erroneously indicated as having "HA α2-3 SA preference" instead of "HA drift from population immunity" to represent ongoing evolution of seasonal influenza virus. This has now been corrected in all versions of the Review. The publisher apologizes to the authors and to readers for this error.
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Influenza A viruses cause pandemics when they cross between species and an antigenically novel virus acquires the ability to infect and transmit between these new hosts. The timing of pandemics is currently unpredictable but depends on ecological and virological factors. The host range of an influenza A virus is determined by species-specific interactions between virus and host cell factors. These include the ability to bind and enter cells, to replicate the viral RNA genome within the host cell nucleus, to evade host restriction factors and innate immune responses and to transmit between individuals. In this Review, we examine the host barriers that influenza A viruses of animals, especially birds, must overcome to initiate a pandemic in humans and describe how, on crossing the species barrier, the virus mutates to establish new interactions with the human host. This knowledge is used to inform risk assessments for future pandemics and to identify virus-host interactions that could be targeted by novel intervention strategies.
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Aves/virologia , Interações entre Hospedeiro e Microrganismos , Especificidade de Hospedeiro , Vírus da Influenza A/fisiologia , Influenza Aviária/virologia , Animais , Humanos , Imunidade Inata , Vírus da Influenza A/classificação , Influenza Humana/virologia , Pandemias , Especificidade da Espécie , Proteínas Virais/genéticaRESUMO
Influenza viruses use an RNA-dependent RNA polymerase (RdRp) to transcribe and replicate their segmented negative-stranded RNA genomes. The influenza A virus RdRp consists of a heterotrimeric complex of the proteins PB1, PB2, and PA. The RdRp is associated with the incoming influenza A viral RNA (vRNA) genome bound by the viral nucleoprotein (NP), in complexes called viral ribonucleoproteins, vRNPs. During the viral replication cycle, the RdRp snatches capped primers from nascent host mRNAs to carry out primary viral transcription. Viral mRNA translation produces new copies of the RdRp subunits and NP, which are required to stabilize and encapsidate complementary copies of the genome (cRNAs), forming cRNPs. These cRNPs then use the cRNAs to make new vRNAs, which are encapsidated into new vRNPs. Secondary transcription by new vRNPs results in further viral mRNAs and an increase of the viral protein load in the cell. The activities of the RdRp (mRNA, cRNA, and vRNA synthesis) in the influenza virus replication cycle can be measured on several levels, ranging from assessment of the accumulation of RNA products in virus-infected cells, through in situ reconstitution of the RdRp from cloned cDNAs, to in vitro biochemical assays that allow the dissection of individual functions of the RdRp enzyme. Here we describe these assays and point out the advantages and drawbacks of each.
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Ensaios Enzimáticos , Orthomyxoviridae/enzimologia , RNA Polimerase Dependente de RNA/metabolismo , Células Cultivadas , Ativação Enzimática , Expressão Gênica , Genes Reporter , Genoma Viral , Humanos , Orthomyxoviridae/fisiologia , RNA Viral , Transcrição Gênica , Replicação ViralRESUMO
The highly pathogenic avian influenza (HPAI) H5N1 influenza virus has been a public health concern for more than a decade because of its frequent zoonoses and the high case fatality rate associated with human infections. Severe disease following H5N1 influenza infection is often associated with dysregulated host innate immune response also known as cytokine storm but the virological and cellular basis of these responses has not been clearly described. We rescued a series of 6:2 reassortant viruses that combined a PR8 HA/NA pairing with the internal gene segments from human adapted H1N1, H3N2, or avian H5N1 viruses and found that mice infected with the virus with H5N1 internal genes suffered severe weight loss associated with increased lung cytokines but not high viral load. This phenotype did not map to the NS gene segment, and NS1 protein of H5N1 virus functioned as a type I IFN antagonist as efficient as NS1 of H1N1 or H3N2 viruses. Instead we discovered that the internal genes of H5N1 virus supported a much higher level of replication of viral RNAs in myeloid cells in vitro, but not in epithelial cells and that this was associated with high induction of type I IFN in myeloid cells. We also found that in vivo during H5N1 recombinant virus infection cells of haematopoetic origin were infected and produced type I IFN and proinflammatory cytokines. Taken together our data infer that human and avian influenza viruses are differently controlled by host factors in alternative cell types; internal gene segments of avian H5N1 virus uniquely drove high viral replication in myeloid cells, which triggered an excessive cytokine production, resulting in severe immunopathology.
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Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/fisiologia , Células Mieloides/virologia , Infecções por Orthomyxoviridae/genética , Replicação Viral/genética , Células A549 , Animais , Células Cultivadas , Cães , Feminino , Genes Virais/fisiologia , Células HEK293 , Humanos , Imunidade Inata/fisiologia , Virus da Influenza A Subtipo H5N1/imunologia , Virus da Influenza A Subtipo H5N1/patogenicidade , Influenza Humana/genética , Influenza Humana/imunologia , Influenza Humana/virologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Mieloides/imunologia , Células Mieloides/metabolismo , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/mortalidade , Infecções por Orthomyxoviridae/virologia , Índice de Gravidade de DoençaRESUMO
Influenza pandemics occur unpredictably when zoonotic influenza viruses with novel antigenicity acquire the ability to transmit amongst humans. Host range breaches are limited by incompatibilities between avian virus components and the human host. Barriers include receptor preference, virion stability and poor activity of the avian virus RNA-dependent RNA polymerase in human cells. Mutants of the heterotrimeric viral polymerase components, particularly PB2 protein, are selected during mammalian adaptation, but their mode of action is unknown. We show that a species-specific difference in host protein ANP32A accounts for the suboptimal function of avian virus polymerase in mammalian cells. Avian ANP32A possesses an additional 33 amino acids between the leucine-rich repeats and carboxy-terminal low-complexity acidic region domains. In mammalian cells, avian ANP32A rescued the suboptimal function of avian virus polymerase to levels similar to mammalian-adapted polymerase. Deletion of the avian-specific sequence from chicken ANP32A abrogated this activity, whereas its insertion into human ANP32A, or closely related ANP32B, supported avian virus polymerase function. Substitutions, such as PB2(E627K), were rapidly selected upon infection of humans with avian H5N1 or H7N9 influenza viruses, adapting the viral polymerase for the shorter mammalian ANP32A. Thus ANP32A represents an essential host partner co-opted to support influenza virus replication and is a candidate host target for novel antivirals.
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Proteínas Aviárias/química , Proteínas Aviárias/metabolismo , Especificidade de Hospedeiro , Vírus da Influenza A/enzimologia , Peptídeos e Proteínas de Sinalização Intracelular/química , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , RNA Polimerase Dependente de RNA/metabolismo , Proteínas Virais/metabolismo , Sequência de Aminoácidos , Animais , Proteínas Aviárias/deficiência , Linhagem Celular , Galinhas/virologia , Cricetinae , Cricetulus , Cães , Evolução Molecular , Regulação Viral da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Virus da Influenza A Subtipo H5N1/enzimologia , Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/fisiologia , Subtipo H7N9 do Vírus da Influenza A/enzimologia , Subtipo H7N9 do Vírus da Influenza A/genética , Subtipo H7N9 do Vírus da Influenza A/fisiologia , Vírus da Influenza A/genética , Vírus da Influenza A/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Proteínas Nucleares , Proteínas de Ligação a RNA , RNA Polimerase Dependente de RNA/genética , Especificidade da Espécie , Transcrição Gênica , Proteínas Virais/genética , Replicação ViralRESUMO
We hypothesise that some influenza virus adaptations to poultry may explain why the barrier for human-to-human transmission is not easily overcome once the virus has crossed from wild birds to chickens. Since the cluster of human infections with H5N1 influenza in Hong Kong in 1997, chickens have been recognized as the major source of avian influenza virus infection in humans. Although often severe, these infections have been limited in their subsequent human-to-human transmission, and the feared H5N1 pandemic has not yet occurred. Here we examine virus adaptations selected for during replication in chickens and other gallinaceous poultry. These include altered receptor binding and increased pH of fusion of the haemagglutinin as well as stalk deletions of the neuraminidase protein. This knowledge could aid the delivery of vaccines and increase our ability to prioritize research efforts on those viruses from the diverse array of avian influenza viruses that have greatest human pandemic potential. Also watch the Video Abstract.
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Virus da Influenza A Subtipo H5N1/patogenicidade , Subtipo H7N9 do Vírus da Influenza A/patogenicidade , Aves Domésticas/virologia , Animais , Humanos , Influenza Aviária/epidemiologia , Influenza Aviária/virologia , Influenza Humana/epidemiologia , Influenza Humana/virologia , PandemiasRESUMO
Typical avian influenza A viruses are restricted from replicating efficiently and causing disease in humans. However, an avian virus can become adapted to humans by mutating or recombining with currently circulating human viruses. These viruses have the potential to cause pandemics in an immunologically naïve human population. It is critical that we understand the molecular basis of host-range restriction and how this can be overcome. Here, we review our current understanding of the mechanisms by which influenza viruses adapt to replicate efficiently in a new host. We predominantly focus on the influenza polymerase, which remains one of the least understood host-range barriers.
Assuntos
Vírus da Influenza A/patogenicidade , Adaptação Fisiológica , Animais , Aves , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/fisiologia , Genes Virais , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Glicoproteínas de Hemaglutininação de Vírus da Influenza/fisiologia , Especificidade de Hospedeiro/genética , Especificidade de Hospedeiro/imunologia , Especificidade de Hospedeiro/fisiologia , Humanos , Imunidade Inata , Vírus da Influenza A/genética , Vírus da Influenza A/fisiologia , Influenza Aviária/imunologia , Influenza Aviária/virologia , Influenza Humana/imunologia , Influenza Humana/virologia , Mutação , Neuraminidase/genética , Neuraminidase/fisiologia , Nucleoproteínas/genética , Nucleoproteínas/fisiologia , Proteínas Virais/genética , Proteínas Virais/fisiologiaRESUMO
Clade 2.2 Eurasian-lineage H5N1 highly pathogenic avian influenza viruses (HPAIVs) were first detected in Qinghai Lake, China, in 2005 and subsequently spread through Asia, Europe, and Africa. Importantly, these viruses carried a lysine at amino acid position 627 of the PB2 protein (PB2 627K), a known mammalian adaptation motif. Previous avian influenza virus isolates have carried glutamic acid in this position (PB2 627E), commonly described to restrict virus polymerase function in the mammalian host. We sought to examine the effect of PB2 627K on viral maintenance in the avian reservoir. Viruses constructed by reverse genetics were engineered to contain converse PB2 627K/E mutations in a Eurasian H5N1 virus (A/turkey/Turkey/5/2005 [Ty/05]) and, for comparison, a historical pre-Asian H5N1 HPAIV that naturally bears PB2 627E (A/turkey/England/50-92/1991 [50-92]). The 50-92 PB2 627K was genetically unstable during virus propagation, resulting in reversion to PB2 627E or the accumulation of the additional mutation PB2 628R and/or a synonymous mutation from an A to a G nucleotide at nucleotide position 1869 (PB2 A1869G). Intriguingly, PB2 628R and/or A1869G appeared to improve the genetic stability of 50-92 PB2 627K. However, the replication of 50-92 PB2 627K in conjunction with these stabilizing mutations was significantly restricted in experimentally infected chickens, where reversion to PB2 627E occurred. In contrast, no significant effects on viral fitness were observed for Ty/05 PB2 627E or 627K in in vitro or in vivo experiments. Our observations suggest that PB2 627K is supported in Eurasian-lineage viruses; in contrast, PB2 627K carries a significant fitness cost in the historical pre-Asian 50-92 virus.
Assuntos
Virus da Influenza A Subtipo H5N1/fisiologia , Mutação de Sentido Incorreto , RNA Polimerase Dependente de RNA/metabolismo , Proteínas Virais/metabolismo , Replicação Viral , Substituição de Aminoácidos , Animais , Ásia , Linhagem Celular , Galinhas , Europa (Continente) , Instabilidade Genômica , Ácido Glutâmico/genética , Humanos , Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/isolamento & purificação , Influenza Aviária/virologia , Lisina/genética , Mutagênese Sítio-Dirigida , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , RNA Polimerase Dependente de RNA/genética , Genética Reversa , Proteínas Virais/genéticaRESUMO
Reassortant influenza viruses with combinations of avian, human, and/or swine genomic segments have been detected frequently in pigs. As a consequence, pigs have been accused of being a "mixing vessel" for influenza viruses. This implies that pig cells support transcription and replication of avian influenza viruses, in contrast to human cells, in which most avian influenza virus polymerases display limited activity. Although influenza virus polymerase activity has been studied in human and avian cells for many years by use of a minigenome assay, similar investigations in pig cells have not been reported. We developed the first minigenome assay for pig cells and compared the activities of polymerases of avian or human influenza virus origin in pig, human, and avian cells. We also investigated in pig cells the consequences of some known mammalian host range determinants that enhance influenza virus polymerase activity in human cells, such as PB2 mutations E627K, D701N, G590S/Q591R, and T271A. The two typical avian influenza virus polymerases used in this study were poorly active in pig cells, similar to what is seen in human cells, and mutations that adapt the avian influenza virus polymerase for human cells also increased activity in pig cells. In contrast, a different pattern was observed in avian cells. Finally, highly pathogenic avian influenza virus H5N1 polymerase activity was tested because this subtype has been reported to replicate only poorly in pigs. H5N1 polymerase was active in swine cells, suggesting that other barriers restrict these viruses from becoming endemic in pigs.