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H7N9 subtype avian influenza viruses (AIVs) cause 1567 human infections and have high mortality, posing a significant threat to public health. Previously, we reported that two avian-derived H7N9 isolates (A/chicken/Eastern China/JTC4/2013 and A/chicken/Eastern China/JTC11/2013) exhibit different pathogenicities in mice. To understand the genetic basis for the differences in virulence, we constructed a series of mutant viruses based on reverse genetics. We found that the PB2-E627K mutation alone was not sufficient to increase the virulence of H7N9 in mice, despite its ability to enhance polymerase activity in mammalian cells. However, combinations with PB1-V719M and/or PA-N444D mutations significantly enhanced H7N9 virulence. Additionally, these combined mutations augmented polymerase activity, thereby intensifying virus replication, inflammatory cytokine expression, and lung injury, ultimately increasing pathogenicity in mice. Overall, this study revealed that virulence in H7N9 is a polygenic trait and identified novel virulence-related residues (PB2-627K combined with PB1-719M and/or PA-444D) in viral ribonucleoprotein (vRNP) complexes. These findings provide new insights into the molecular mechanisms underlying AIV pathogenesis in mammals, with implications for pandemic preparedness and intervention strategies. [ABSTRACT FROM AUTHOR]

Abstract H7N9 subtype avian influenza viruses (AIVs) cause 1567 human infections and have high mortality, posing a significant threat to public health. Previously, we reported that two avian-derived H7N9 isolates (A/chicken/Eastern China/JTC4/2013 and A/chicken/Eastern China/JTC11/2013) exhibit different pathogenicities in mice. To understand the genetic basis for the differences in virulence, we constructed a series of mutant viruses based on reverse genetics. We found that the PB2-E627K mutation alone was not sufficient to increase the virulence of H7N9 in mice, despite its ability to enhance polymerase activity in mammalian cells. However, combinations with PB1-V719M and/or PA-N444D mutations significantly enhanced H7N9 virulence. Additionally, these combined mutations augmented polymerase activity, thereby intensifying virus replication, inflammatory cytokine expression, and lung injury, ultimately increasing pathogenicity in mice. Overall, this study revealed that virulence in H7N9 is a polygenic trait and identified novel virulence-related residues (PB2-627K combined with PB1-719M and/or PA-444D) in viral ribonucleoprotein (vRNP) complexes. These findings provide new insights into the molecular mechanisms underlying AIV pathogenesis in mammals, with implications for pandemic preparedness and intervention strategies.

We characterized the evolution and molecular characteristics of avian influenza A(H7N9) viruses isolated in China during 2021–2023. We systematically analyzed the 10-year evolution of the hemagglutinin gene to determine the evolutionary branch. Our results showed recent antigenic drift, providing crucial clues for updating the H7N9 vaccine and disease prevention and control.

Numerous studies have reported a correlation between gut microbiota and influenza A virus (IAV) infection and disease severity. However, the causal relationship between these factors remains inadequately explored. This investigation aimed to assess the influence of gut microbiota on susceptibility to human infection with H7N9 avian IAV and the severity of influenza A (H1N1)pdm09 infection. A two-sample Mendelian randomization analysis was conducted, integrating our in-house genome-wide association study (GWAS) on H7N9 susceptibility and H1N1pdm09 severity with a metagenomics GWAS dataset from a Chinese population. Twelve and fifteen gut microbiotas were causally associated with H7N9 susceptibility or H1N1pdm09 severity, separately. Notably, Clostridium hylemonae and Faecalibacterium prausnitzii were negative associated with H7N9 susceptibility and H1N1pdm09 severity, respectively. Moreover, Streptococcus peroris and Streptococcus sanguinis were associated with H7N9 susceptibility, while Streptococcus parasanguini and Streptococcus suis were correlated with H1N1pdm09 severity. These results provide novel insights into the interplay between gut microbiota and IAV pathogenesis as well as new clues for mechanism research regarding therapeutic interventions or IAV infections. Future studies should concentrate on clarifying the regulatory mechanisms of gut microbiota and developing efficacious approaches to reduce the incidence of IAV infections, which could improve strategy for preventing and treating IAV infection worldwide. [ABSTRACT FROM AUTHOR]

Influenzavirus is among the most relevant candidates for a next pandemic. We review here the phylogeny of former influenza pandemics, and discuss candidate lineages. After briefly reviewing the other existing antiviral options, we discuss in detail the evidences supporting the efficacy of passive immunotherapies against influenzavirus, with a focus on convalescent plasma. [ABSTRACT FROM AUTHOR]

Vaccination is crucial for the prevention and mitigation of avian influenza infections in China. The inactivated H7N9 vaccine, when administered to poultry, significantly lowers the risk of infection among both poultry and humans, while also markedly decreasing the prevalence of H7N9 detections. Highly pathogenic (HP) H7N9 viruses occasionally appear, whereas their low pathogenicity (LP) counterparts have been scarcely detected since 2018. However, these contributing factors remain poorly understood. We conducted an exploratory investigation of the mechanics via the application of comprehensive bioinformatic approaches. We delineated the Yangtze River Delta (YRD) H7N9 lineage into 5 clades (YRD-A to E). Our findings highlight the emergence and peak occurrence of the LP H7N9-containing YRD-E clade during the 5th epidemic wave in China’s primary poultry farming areas. A more effective control of LP H7N9 through vaccination was observed compared to that of its HP H7N9 counterpart. YRD-E exhibited a tardy evolutionary trajectory, denoted by the conservation of its genetic and antigenic variation. Our analysis of YRD-E revealed only minimal amino acid substitutions along its phylogenetic tree and a few selective sweep mutations since 2016. In terms of epidemic fitness, the YRD-E was measured to be lower than that of the HP variants. Collectively, these findings underscore the conserved evolutionary patterns distinguishing the YRD-E. Given the conservation presented in its evolutionary patterns, the YRD-E LP H7N9 is hypothesized to be associated with a reduction following the mass vaccination in a relatively short period owing to its lower probability of antigenic variation that might affect vaccine efficiency.

ABSTRACTHuman infections with the H7N9 influenza virus have been eliminated in China through vaccination of poultry; however, the H7N9 virus has not yet been eradicated from poultry. Carefully analysis of H7N9 viruses in poultry that have sub-optimal immunity may provide a unique opportunity to witness the evolution of highly pathogenic avian influenza virus in the context of vaccination. Between January 2020 and June 2023, we isolated 16 H7N9 viruses from samples we collected during surveillance and samples that were sent to us for disease diagnosis. Genetic analysis indicated that these viruses belonged to a single genotype previously detected in poultry. Antigenic analysis indicated that 12 of the 16 viruses were antigenically close to the H7-Re4 vaccine virus that has been used since January 2022, and the other four viruses showed reduced reactivity with the vaccine. Animal studies indicated that all 16 viruses were nonlethal in mice, and four of six viruses showed reduced virulence in chickens upon intranasally inoculation. Importantly, the H7N9 viruses detected in this study exclusively bound to the avian-type receptors, having lost the capacity to bind to human-type receptors. Our study shows that vaccination slows the evolution of H7N9 virus by preventing its reassortment with other viruses and eliminates a harmful characteristic of H7N9 virus, namely its ability to bind to human-type receptors.

Since the first human infection reported in 2013, H7N9 avian influenza virus (AIV) has been regarded as a serious threat to human health. In this study, we sought to identify the virulence determinant of the H7N9 virus in mammalian hosts. By comparing the virulence of the SH/4664 H7N9 virus, a non-virulent H9N2 virus, and various H7N9-H9N2 hybrid viruses in infected mice, we first pinpointed PB2 as the primary viral factor accounting for the difference between H7N9 and H9N2 in mammalian virulence. We further analyzed the in vivo effects of individually mutating H7N9 PB2 residues different from the closely related H9N2 virus and consequently found residue 473, alongside the well-known residue 627, to be critical for the virulence of the H7N9 virus in mice and the activity of its reconstituted viral polymerase in mammalian cells. The importance of PB2-473 was further strengthened by studying reverse H7N9 substitutions in the H9N2 background. Finally, we surprisingly found that species-specific usage of ANP32A, a family member of host factors connecting with the PB2-627 polymorphism, mediates the contribution of PB2 473 residue to the mammalian adaption of AIV polymerase, as the attenuating effect of PB2 M473T on the viral polymerase activity and viral growth of the H7N9 virus could be efficiently complemented by co-expression of chicken ANP32A but not mouse ANP32A and ANP32B. Together, our studies uncovered the PB2 473 residue as a novel viral host range determinant of AIVs via s pecies-specific co-opting of the ANP32 host factor to support viral polymerase activity. [ABSTRACT FROM AUTHOR]

The adaptation of egg-derived H7N9 candidate vaccine virus (CVV) in the mammalian cell line is an approach to developing a high-growth virus strain for the mass production of vaccine manufacturing. The adaptive mutations that occur in hemagglutinin (HA) are critical to the activity and potency of the vaccine virus. Previously, we identified a new mutation of A169S in the HA protein of an MDCK-adapted H7N9 vaccine virus (A/Anhui/2013, RG268); however, whether and how this mutation affects vaccine potency remain to be investigated. In this study, we serially passaged RG268 in MDCK cells and found that the HA titer and the TCID50 of the passaged virus RG268-M5 were 4-fold (HA units/50 μL) and 3.5-fold (log10 TCID50/mL) higher than those of the original CVV. By inspecting tandem MS spectra, we identified a new glycosylation site at N167 near the receptor binding site of the HA protein of RG268-M5. Flow cytometry results revealed that RG268-M5 could efficiently infect MDCK cells and initiate viral protein replication as well as that of RG268. Though the new glycosylation site is in the antigenic epitope of viral HA protein, the HI assay result indicated that the antigenicity of RG268-M5 was similar to RG268. Additionally, immunizing mice with RG268-M5 mixed aluminum hydroxide could induce potent antibody responses against the homologous and heterologous H7N9 viruses in vitro whereas the titers were comparable with those from the RG268 group. These results provide in-depth structural information regarding the effects of site-specific glycosylation on virus properties, which have implications for novel avian influenza vaccine development. [ABSTRACT FROM AUTHOR]

Avian influenza (AI) viruses cause infection in birds and humans. Several H5N1 and H7N9 variants are highly pathogenic avian influenza (HPAI) viruses. H5N1 is a highly infectious bird virus infecting primarily poultry, but unlike other AIs, H5N1 also infects mammals and transmits to humans with a case fatality rate above 40%. Similarly, H7N9 can infect humans, with a case fatality rate of over 40%. Since 1996, there have been several HPAI outbreaks affecting humans, emphasizing the need for safe and effective antivirals. We show that probenecid potently inhibits H5N1 and H7N9 replication in prophylactically or therapeutically treated A549 cells and normal human broncho-epithelial (NHBE) cells, and H5N1 replication in VeroE6 cells and mice. [ABSTRACT FROM AUTHOR]

Simple Summary: Avian influenza viruses have the ability to breach species barriers and infect mammals, posing a significant threat to public health. The H7N9 subtype of avian influenza virus that emerged in China in 2013 resulted in 1568 human infections, with an alarming mortality rate of nearly 40%. While human infections caused by avian influenza viruses typically occur sporadically, the reasons behind the widespread impact of H7N9 remain unclear. During surveillance for avian influenza in live poultry markets in eastern China in 2013, we isolated two H7N9 avian influenza viruses from seemingly healthy chickens. One of these chicken-derived viruses demonstrated the ability to bind to mammalian receptors and naturally harbored the mammalian molecular marker PB2-627K. Furthermore, this virus proved to be highly pathogenic to mice. In summary, our findings suggest that the presence of a mammal-adapted H7N9 virus in poultry in 2013 could be a significant factor explaining the unusually high number of human infections during that period. Avian influenza viruses can cross species barriers and adapt to mammals. The H7N9 subtype AIV that emerged in China in 2013 caused 1568 human infections, with a mortality rate of nearly 40%. We conducted a retrospective analysis of H7N9 viruses that were isolated in live poultry markets in 2013. We found that two avian-origin H7N9 isolates, A/chicken/Eastern China/JTC4/2013 and A/chicken/Eastern China/JTC11/2013, have a similar genetic background but exhibit different pathogenicity in mice. Whole-genome alignment of the two H7N9 viruses was carried out, and only six amino acid differences mapped in five genes, including the well-known virulence molecular marker PB2-E627K. Our retrospective analysis highlighted the importance of monitoring the adaptive mutations in avian influenza viruses with zoonotic potential. [ABSTRACT FROM AUTHOR]

Avian influenza A viruses (IAVs) that cross the species barrier to infect humans have the potential to initiate a new pandemic. However, the host factors influencing avian IAV infection remain poorly understood. To address this knowledge gap, we conducted a two‐sample Mendelian randomization (MR) analysis by integrating our in‐house genome‐wide association study (GWAS) of avian IAV H7N9 susceptibility (with 217 cases and 116 controls) with the largest GWAS of serum IgA levels to date (sample size 41 263). Using the inverse‐variance weighted (IVW) method, we discovered that genetically decreased serum IgA levels were associated with an increased risk of H7N9 infection (β = −2.528, 95% confidence interval [CI]: −4.572 to −0.484; p = 0.015). Consistent results were obtained from three other MR methods, including robust IVW estimation (β = −2.506, 95% CI: −4.109 to −0.902; p = 0.002), generalized summary‐data‐based MR (GSMR) (β = −2.238, 95% CI: −4.106 to −0.602; p = 0.019), and MR‐pleiotropy residual sum and outlier (MR‐PRESSO) (β = −2.528, 95% CI: −4.396 to −0.892; p = 0.026). In conclusion, our analysis provided compelling evidence support a causal relationship between genetically predicted serum IgA levels and avian IAV H7N9 susceptibility. [ABSTRACT FROM AUTHOR]

Numerous studies have reported a correlation between gut microbiota and influenza A virus (IAV) infection and disease severity. However, the causal relationship between these factors remains inadequately explored. This investigation aimed to assess the influence of gut microbiota on susceptibility to human infection with H7N9 avian IAV and the severity of influenza A (H1N1)pdm09 infection. A two-sample Mendelian randomization analysis was conducted, integrating our in-house genome-wide association study (GWAS) on H7N9 susceptibility and H1N1pdm09 severity with a metagenomics GWAS dataset from a Chinese population. Twelve and fifteen gut microbiotas were causally associated with H7N9 susceptibility or H1N1pdm09 severity, separately. Notably, Clostridium hylemonae and Faecalibacterium prausnitzii were negative associated with H7N9 susceptibility and H1N1pdm09 severity, respectively. Moreover, Streptococcus peroris and Streptococcus sanguinis were associated with H7N9 susceptibility, while Streptococcus parasanguini and Streptococcus suis were correlated with H1N1pdm09 severity. These results provide novel insights into the interplay between gut microbiota and IAV pathogenesis as well as new clues for mechanism research regarding therapeutic interventions or IAV infections. Future studies should concentrate on clarifying the regulatory mechanisms of gut microbiota and developing efficacious approaches to reduce the incidence of IAV infections, which could improve strategy for preventing and treating IAV infection worldwide.

Antigen sparing is an important strategy for pandemic vaccine development because of the limitation of worldwide vaccine production during disease outbreaks. However, several clinical studies have demonstrated that the current aluminum (Alum)-adjuvanted influenza vaccines fail to sufficiently enhance immune responses to meet licensing criteria. Here, we used pandemic H7N9 as a model virus to demonstrate that a 10-fold lower amount of vaccine antigen combined with Alum and TLR9 agonist can provide stronger protective effects than using Alum as the sole adjuvant. We found that the Alum/CpG 1018 combination adjuvant could induce more robust virus-specific humoral immune responses, including higher total IgG production, hemagglutination-inhibiting antibody activity, and neutralizing antibody titres, than the Alum-adjuvanted formulation. Moreover, this combination adjuvant shifted the immune response toward a Th1-biased immune response. Importantly, the Alum/CpG 1018-formulated vaccine could confer better protective immunity against H7N9 challenge than that adjuvanted with Alum alone. Notably, the addition of CpG 1018 to the Alum-adjuvanted H7N9 whole-virion vaccine exhibited an antigen-sparing effect without compromising vaccine efficacy. These findings have significant implications for improving Alum-adjuvanted influenza vaccines using the approved adjuvant CpG 1018 for pandemic preparedness.

• Recombinant hemagglutinin protein vaccine produced in insect cells provides protection against H7N9 infection. • H7N9 vaccine immunogenicity was dependent on inclusion of Advax or Advax-CpG adjuvants. • Whereas Advax was more Th2 biased, Advax-CpG was more Th1 biased. • Recombinant hemagglutinin vaccine protected better than inactivated virus vaccine. • Advax adjuvant was able to overcome neonatal H7N9 vaccine hypo-responsiveness. There is a major unmet need for strategies to improve the immunogenicity and effectiveness of pandemic influenza vaccines, particularly in poor responder populations such as neonates. Recombinant protein approaches to pandemic influenza offer advantages over more traditional inactivated virus approaches, as they are free of problems such as egg adaptation or need for high level biosecurity containment for manufacture. However, a weakness of recombinant proteins is their low immunogenicity. We asked whether the use of an inulin polysaccharide adjuvant (Advax) alone or combined with a TLR9 agonist (CpG55.2) would enhance the immunogenicity and protection of a recombinant hemagglutinin vaccine against H7N9 influenza (rH7HA), including in neonatal mice. Advax adjuvant induced predominantly IgG1 responses against H7HA, whereas Advax-CpG55.2 adjuvant also induced IgG2a, IgG2b and IgG3 responses, consistent with the TLR9 agonist component inducing a Th1 bias. Advax-CpG55.2 adjuvanted rH7HA induced high serum neutralizing antibody titers in adult mice. In newborns it similarly overcame immune hypo-responsiveness and enhanced serum anti-rH7HA IgG levels in 7-day-old BALB/C and C57BL/6 mice. Immunized adult mice were protected against a lethal H7N9 virus challenge. When formulated with Advax-CpG55.2 adjuvant, greater protection was seen with rH7HA than with inactivated H7 whole virus antigen. Advax-CpG55.2 adjuvanted rH7HA represents a promising influenza vaccine platform for further development. [ABSTRACT FROM AUTHOR]

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The adaptation of egg-derived H7N9 candidate vaccine virus (CVV) in the mammalian cell line is an approach to developing a high-growth virus strain for the mass production of vaccine manufacturing. The adaptive mutations that occur in hemagglutinin (HA) are critical to the activity and potency of the vaccine virus. Previously, we identified a new mutation of A169S in the HA protein of an MDCK-adapted H7N9 vaccine virus (A/Anhui/2013, RG268); however, whether and how this mutation affects vaccine potency remain to be investigated. In this study, we serially passaged RG268 in MDCK cells and found that the HA titer and the TCID50 of the passaged virus RG268-M5 were 4-fold (HA units/50 μL) and 3.5-fold (log10 TCID50/mL) higher than those of the original CVV. By inspecting tandem MS spectra, we identified a new glycosylation site at N167 near the receptor binding site of the HA protein of RG268-M5. Flow cytometry results revealed that RG268-M5 could efficiently infect MDCK cells and initiate viral protein replication as well as that of RG268. Though the new glycosylation site is in the antigenic epitope of viral HA protein, the HI assay result indicated that the antigenicity of RG268-M5 was similar to RG268. Additionally, immunizing mice with RG268-M5 mixed aluminum hydroxide could induce potent antibody responses against the homologous and heterologous H7N9 viruses in vitro whereas the titers were comparable with those from the RG268 group. These results provide in-depth structural information regarding the effects of site-specific glycosylation on virus properties, which have implications for novel avian influenza vaccine development.

Avian influenza (AI) viruses cause infection in birds and humans. Several H5N1 and H7N9 variants are highly pathogenic avian influenza (HPAI) viruses. H5N1 is a highly infectious bird virus infecting primarily poultry, but unlike other AIs, H5N1 also infects mammals and transmits to humans with a case fatality rate above 40%. Similarly, H7N9 can infect humans, with a case fatality rate of over 40%. Since 1996, there have been several HPAI outbreaks affecting humans, emphasizing the need for safe and effective antivirals. We show that probenecid potently inhibits H5N1 and H7N9 replication in prophylactically or therapeutically treated A549 cells and normal human broncho-epithelial (NHBE) cells, and H5N1 replication in VeroE6 cells and mice.

Highly pathogenic (HP) avian influenza A H7N9 virus has emerged in China since 2016. In recent years, it has been most prevalent in northern China. However, several strains of HP H7N9 reappeared in southwestern China (Yunnan Province) in 2021. As a result, we are wondering if these viruses have re-emerged in situ or been reintroduced. Here, we present phylogenetic evidence that the HP H7N9 viruses isolated in Yunnan emigrated from northern to southwestern China in 2020. The northern subregion of China has become a novel epicenter in HP H7N9 dissemination. Meanwhile, a cleavage motif re-emerged due to the T341I mutation, implying a parallel evolution. This cross-region transmission, which originated in non-adjacent provinces and traveled a great geographic distance in an unknown way, indicates that HP H7N9 dissemination did not halt in 2020, even under the shadow of the COVID-19 pandemic. Additional surveillance studies in poultry are required to determine the HP H7N9 virus's geographic distribution and spread. [ABSTRACT FROM AUTHOR]

Influenza A viruses are a diverse group of pathogens that have been responsible for millions of human and avian deaths throughout history. Here, we illustrate the transmission potential of H7N9 influenza A virus between Coturnix quail (Coturnix sp.), domestic ducks (Anas platyrhynchos domesticus), chickens (Gallus gallus domesticus), and house sparrows (Passer domesticus) co-housed in an artificial barnyard setting. In each of four replicates, individuals from a single species were infected with the virus. Quail shed virus orally and were a source of infection for both chickens and ducks. Infected chickens transmitted the virus to quail but not to ducks or house sparrows. Infected ducks transmitted to chickens, resulting in seroconversion without viral shedding. House sparrows did not shed virus sufficiently to transmit to other species. These results demonstrate that onward transmission varies by index species, and that gallinaceous birds are more likely to maintain H7N9 than ducks or passerines. • Quail and chickens transmit H7N9 influenza virus in a mixed-barnyard setting. • Onward transmission is dependent on index species and level of viral shedding. • House sparrows are insufficient hosts of H7N9 influenza virus to establish interspecies transmission. • Some avian influenza viruses are more likely to be maintained in wild birds, while H7N9 is more likely to be found in poultry. [ABSTRACT FROM AUTHOR]

DC-SIGN and Galectin-3 are two different lectins and have been reported to participate in regulation of several virus infections. WHO has pointed that H5N1 and H7N9 avian influenza viruses (AIVs) play continuous threats to global health. AIV hemagglutinin (HA) protein—a highly glycosylated protein—mediates influenza infection and was proposed to have DC-SIGN and Gal3 interactive domains. This study aims to address the individual and collaborative roles of DC-SIGN and Gal3 toward AIVs infection. Firstly, A549 cells with DC-SIGN expression or Gal3-knockdown, via lentiviral vector-mediated CD209 gene expression or LGALS-3 gene knockdown, respectively were generated. Quantitative reverse transcription PCR (qRT-PCR) results indicated that DC-SIGN expression and Gal3 knockdown in A549 cells significantly promoted and ameliorated HA or NP gene expression, respectively after H5N1 and H7N9-reverse genetics (RG) virus postinfections (P  < 0.05). Similar results observed in immunoblotting, indicating that DC-SIGN expression significantly facilitated H5N1-RG and H7N9-RG infections (P  < 0.05), whereas Gal3 knockdown significantly reduced both viral infections (P  < 0.05). Furthermore, we found that DC-SIGN and Gal3 co-expression significantly enhanced infectivity of both H5N1-RG and H7N9-RG viruses (P  < 0.01) and higher regulatory capabilities by DC-SIGN and Gal3 in H5N1-RG than H7N9-RG were noted. The promoting effect mainly relied on exogenous Gal3 and DC-SIGN directly interacting with the HA protein of H5N1 or H7N9 AIVs, subsequently enhancing virus infection. This study sheds light on two different lectins individually and collaboratively regulating H5N1 and H7N9 AIVs infection and suggests that inhibitors against DC-SIGN and Gal3 interacting with HA could be utilized as alternative antiviral strategies. [ABSTRACT FROM AUTHOR]

A differentiating infected from vaccinated animals (DIVA) vaccine is an ideal strategy for viral eradication in poultry. Here, according to the emerging highly pathogenic H7N9 avian influenza virus (AIV), a DIVA vaccine strain, named rGD4HALo-mH3-TX, was successfully developed, based on a substituted 12 peptide of H3 virus located at HA2. In order to meet with the safety requirement of vaccine production, the multi-basic amino acid located at the HA cleavage site was modified. Meanwhile, six inner viral genes from a H9N2 AIV TX strainwere introduced for increasing viral production. The rGD4HALo-mH3-TX strain displayed a similar reproductive ability with rGD4 and low pathogenicity in chickens, suggesting a good productivity and safety. In immuned chickens, rGD4HALo-mH3-TX induced a similar antibody level with rGD4 and provided 100% clinical protection and 90% shedding protection against highly pathogenic virus challenge. rGD4HALo-mH3-TX strain also produced a good cross-protection against low pathogenic AIV JD/17. Moreover, serological DIVA characteristics were evaluated by a successfully established competitive inhibition ELISA based on a 3G10 monoclonal antibody, and the result showed a strong reactivity with antisera of chickens vaccinated with H7 subtype strains but not rGD4HALo-mH3-TX. Collectedly, rGD4HALo-mH3-TX is a promising DIVA vaccine candidate against both high and low pathogenic H7N9 subtype AIV.

Objectives: We aimed to compared the clinical features of acute respiratory distress syndrome (ARDS) induced by COVID-19 and H7N9 virus infections. Methods: Clinical data of 100 patients with COVID-19 and 46 patients with H7N9 were retrospectively analyzed. Results: Elevated inflammatory indices and coagulation disorders were more common in COVID-19-ARDS group than in the H7N9-ARDS group. The median interval from illness onset to ARDS development was shorter in H7N9-ARDS. The PaO2/FiO2 level was lower in H7N9-ARDS, whereas the Sepsis-related Organ Failure Assessment score was higher in COVID-19-ARDS. The proportion of patients with disseminated intravascular coagulation and liver injury in COVID-19-ARDS and H7N9-ARDS was 45.5% versus 3.1% and 28.8% versus 50%, respectively (P

Since 2013, H7N9 avian influenza viruses (AIVs) have caused more than 1,500 human infections and the culling of millions of poultry. Despite large-scale poultry vaccination, H7N9 AIVs continue to circulate among poultry in China and pose a threat to human health. Previously, we isolated and generated four monoclonal antibodies (mAbs) derived from humans naturally infected with H7N9 AIV. Here, we investigated the hemagglutinin (HA) epitopes of H7N9 AIV targeted by these mAbs (L3A44, K9B-122, L4A-14, and L4B-18) using immune escape studies. Our results revealed four key antigenic epitopes at HA amino acid positions 125, 133, 149, and 217. The mutant H7N9 viruses representing escape mutations containing an alanine-to-threonine substitution at residue 125 (A125T), a glycine-to-glutamic acid substitution at residue 133 (G133E), an asparagine-to-aspartic acid substitution at residue 149 (N149D), or a leucine-to-glutamine substitution at residue 217 (L217Q) showed reduced or completely abolished cross-reactivity with the mAbs, as measured by a hemagglutination inhibition (HI) assay. We further assessed the potential risk of these mutants to humans should they emerge following mAb treatment by measuring the impact of these HA mutations on virus fitness and evasion of host adaptive immunity. Here, we showed that the L4A-14 mAb had broad neutralizing capabilities, and its escape mutant N149D had reduced viral stability and human receptor binding and could be neutralized by both postinfection and antigen-induced sera. Therefore, the L4A-14 mAb could be a therapeutic candidate for H7N9 AIV infection in humans and warrants further investigation for therapeutic applications. IMPORTANCE Avian influenza virus (AIV) H7N9 continues to circulate and evolve in birds, posing a credible threat to humans. Antiviral drugs have proven useful for the treatment of severe influenza infections in humans; however, concerns have been raised as antiviral-resistant mutants have emerged. Monoclonal antibodies (mAbs) have been studied for both prophylactic and therapeutic applications in infectious disease control and have demonstrated great potential. For example, mAb treatment has significantly reduced the risk of people developing severe disease with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In addition to the protection efficiency, we should also consider the potential risk of the escape mutants generated by mAb treatment to public health by assessing their viral fitness and potential to compromise host adaptive immunity. Considering these parameters, we assessed four human mAbs derived from humans naturally infected with H7N9 AIV and showed that the mAb L4A-14 displayed potential as a therapeutic candidate. [ABSTRACT FROM AUTHOR]

Reassortment can introduce one or more gene segments of influenza A viruses (IAVs) into another, resulting in novel subtypes. Since 2013, a new outbreak of human highly pathogenic avian influenza has emerged in the Yangtze River Delta (YRD) and South‐Central regions of China. In this study, using Anhui province as an example, we discuss the possible impact of H7N9 IAVs on future influenza epidemics through a series of gene reassortment events. Sixty‐one human H7N9 isolates were obtained from five outbreaks in Anhui province from 2013 to 2019. Bioinformatics analyses revealed that all of them were characterized by low pathogenicity and high human or mammalian tropism and had introduced novel avian influenza A virus (AIV) subtypes such as H7N2, H7N6, H9N9, H5N6, H6N6, and H10N6 through gene reassortment. In reassortment events, Anhui isolates may donate one or more segments of HA, NA, and the six internal protein‐coding genes for the novel subtype AIVs. Our study revealed that H7N9, H9N2, and H5N1 can serve as stable and persistent gene pools for AIVs in the YRD and South‐Central regions of China. Novel AIV subtypes might be generated continuously by reassortment. These AIVs may have obtained human‐type receptor‐binding abilities from their donors and prefer binding to them, which can cause human epidemics through accidental spillover infections. Facing the continual threat of emerging avian influenza, constant monitoring of AIVs should be conducted closely for agricultural and public health. [ABSTRACT FROM AUTHOR]

Avian influenza viruses can cross species barriers and adapt to mammals. The H7N9 subtype AIV that emerged in China in 2013 caused 1568 human infections, with a mortality rate of nearly 40%. We conducted a retrospective analysis of H7N9 viruses that were isolated in live poultry markets in 2013. We found that two avian-origin H7N9 isolates, A/chicken/Eastern China/JTC4/2013 and A/chicken/Eastern China/JTC11/2013, have a similar genetic background but exhibit different pathogenicity in mice. Whole-genome alignment of the two H7N9 viruses was carried out, and only six amino acid differences mapped in five genes, including the well-known virulence molecular marker PB2-E627K. Our retrospective analysis highlighted the importance of monitoring the adaptive mutations in avian influenza viruses with zoonotic potential.

The epidemiology of H5N1 and H7N9 avian viruses of humans infected in China differs despite both viruses being avian reassortants that have inherited six internal genes from a common ancestor, H9N2. The median age of infected populations is substantially younger for H5N1 virus (26 years) compared with H7N9 virus (63 years). Population susceptibility to infection with seasonal influenza is understood to be influenced by cross-reactive CD8+ T cells directed towards immunogenic peptides derived from internal viral proteins which may provide some level of protection against further influenza infection. Prior exposure to seasonal influenza peptides may influence the age-related infection patterns observed for H5N1 and H7N9 viruses. A comparison of relatedness of immunogenic peptides between historical human strains and the two avian emerged viruses was undertaken for a possible explanation in the differences in age incidence observed. There appeared to be some relationship between past exposure to related peptides and the lower number of H5N1 virus cases in older populations, however the relationship between prior exposure and older populations among H7N9 virus patients was less clear.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and Clustered Regularly Interspaced Short Palindromic Repeats-Associated Proteins (CRISPR-Cas) have promising prospects in the field of nucleic acid molecular diagnostics. However, Clustered Regularly Interspaced Short Palindromic Repeats-based fluorescence detection technology is mainly hindered by proteins with conjugated double bonds and autofluorescence, resulting in high fluorescence background, low sensitivity and incompatible reaction systems, which are not conducive to automatic clinical testing. Chemiluminescence (CL) detection technology has been applied mainly owing to its greatly high sensitivity, as well as low background and rapid response. Therefore, we developed a rapid, ultrasensitive and economical detection system based on Clustered Regularly Interspaced Short Palindromic Repeats-Clustered Regularly Interspaced Short Palindromic Repeats-Associated Proteins 13a combined with magnetic beads (MBs) and chemiluminescence (CL) (Cas13a-MB-CL) to detect Influenza A (H7N9), an acute respiratory tract infectious disease. The carboxyl functionalized magnetic beads (MBs-COOH) were covalently coupled with aminated RNA probe while the other end of the RNA probe was modified with biotin. Alkaline phosphatase labeled streptavidin (SA-ALP) binds with biotin to form magnetic beads composites. In presence of target RNA, the collateral cleavage activity of Cas13a was activated to degrade the RNA probes on MBs and released Alkaline phosphatase from the composites. The composites were then magnetically separated followed by addition of ALP substrate Disodium 2-chloro-5-{4-methoxyspiro [1,2-dioxetane-3,2′-(5′-chloro) tricyclo (3.3.1.13,7) decan]-4-yl}-1-phenyl phosphate (CDP-star), to generate the chemiluminescence signal. The activity of Associated Proteins 13a and presence of target RNA was quantified by measuring the chemiluminescence intensity. The proposed method accomplished the detection of H7N9 within 30 min at 25°C. When combined with Reverse Transcription- Recombinase Aides Amplification (RT-RAA), the low detection limit limit of detection was as low as 19.7 fM (3S/N). Our proposed MB-Associated Proteins 13a-chemiluminescence was further evaluated to test H7N9 clinical samples, showing superior sensitivity and specificity.

The zoonotic H7N9 avian influenza virus emerged with the H9N2-origin internal gene cassette. Previous studies have reported that genetic reassortments with H9N2 were common in the first five human H7N9 epidemic waves. However, our latest work found that the circulating high pathogenicity H7N9 virus has established a dominant internal gene cassette and has decreased the frequency of reassortment with H9N2 since 2018. This dominant cassette of H7N9 was distinct from the cocirculating H9N2, although they shared a common ancestor. As a result, we suppose that this dominant cassette may benefit the viral population fitness and promote its continuous circulation in chickens. [ABSTRACT FROM AUTHOR]

Unadjuvanted A/H7N9 vaccines are poorly immunogenic. The immune response is improved with the addition of MF59, an oil-in-water adjuvant. However, the cellular immunologic responses of MF59-adjuvanted A/H7N9 vaccine are not fully understood. 37 participants were vaccinated with 2 doses of 2013 influenza A/H7N9 vaccine (at Days 1 and 21) with or without MF59 and enrolled in an immunology substudy. Responses were assessed at multiple timepoints (Days 0, 8, 21, 29, and 42) for hemagglutination inhibition (HAI) and neutralizing antibody (Neut) assays, memory B cell responses by enzyme-linked ImmunoSpot; circulating follicular helper T cells (cT FH) and CD4 + T cells by intracellular cytokine staining. MF59-adjuvanted influenza A/H7N9 vaccine induced significantly higher hemagglutination inhibition (HAI) and neutralizing antibody (Neut) responses when compared to unadjuvanted vaccine. The adjuvanted vaccine elicited significantly higher levels of Inducible T-cell Co-Stimulator (ICOS) expression by CXCR3+CXCR5+CD4+ cT FH cells, compared to unadjuvanted vaccine. The magnitude of increase in cT FH cells (from baseline to Day 8) and in IL-21 expressing CD154+CD4+ T cells (from baseline to Days 8 and 21) correlated with HAI (at Day 29) and Neut antibody (at Days 8 and 29) titers. The increase in frequency of IL-21 expressing CD154+CD4+T cells (from baseline to Day 21) correlated with memory B cell frequency (at Day 42). cT FH activation is associated with HAI and Neut responses in recipients of MF59-adjuvanted influenza A/H7N9 vaccine relative to unadjuvanted vaccine. Future studies should focus on optimizing the cT FH response and use cT FH as an early biomarker of serological response to vaccination. This trial was registered at clinicaltrials.gov , trial number NCT01938742. [ABSTRACT FROM AUTHOR]

H9N2 avian influenza viruses (AIVs) have donated internal gene segments during the emergence of zoonotic AIVs, including H7N9. We used reverse genetics to generate A/Anhui/1/13 (H7N9) and three reassortant viruses (2:6 H7N9) which contained the hemagglutinin and neuraminidase from Anhui/13 (H7N9) and the six internal gene segments from H9N2 AIVs belonging to (i) G1 subgroup 2, (ii) G1 subgroup 3, or (iii) BJ94 lineages, enzootic in different regions throughout Asia. Infection of chickens with the 2:6 H7N9 containing G1-like H9N2 internal genes conferred attenuation in vivo, with reduced shedding and transmission to contact chickens. However, possession of BJ94-like H9N2 internal genes resulted in more rapid transmission and significantly elevated cloacal shedding compared to the parental Anhui/13 H7N9. In vitro analysis showed that the 2:6 H7N9 with BJ94-like internal genes had significantly increased replication compared to the Anhui/13 H7N9 in chicken cells. In vivo coinfection experiments followed, where chickens were coinfected with pairs of Anhui/13 H7N9 and a 2:6 H7N9 reassortant. During ensuing transmission events, the Anhui/13 H7N9 virus outcompeted 2:6 H7N9 AIVs with internal gene segments of BJ94-like or G1-like H9N2 viruses. Coinfection did lead to the emergence of novel reassortant genotypes that were transmitted to contact chickens. Some of the reassortant viruses had a greater replication in chicken and human cells compared to the progenitors. We demonstrated that the internal gene cassette determines the transmission fitness of H7N9 viruses in chickens, and the reassortment events can generate novel H7N9 genotypes with increased virulence in chickens and enhanced zoonotic potential. [ABSTRACT FROM AUTHOR]

The interferon-induced myxovirus resistance protein A (MxA) is a potent restriction factor that prevents zoonotic infection from influenza A virus (IAV) subtype H7N9. Individuals expressing antivirally inactive MxA variants are highly susceptible to these infections. However, human-adapted IAVs have acquired specific mutations in the viral nucleoprotein (NP) that allow escape from MxA-mediated restriction but that have not been observed in MxA-sensitive, human H7N9 isolates. To date, it is unknown whether H7N9 can adapt to escape MxA-mediated restriction. To study this, we infected Rag2-knockout (Rag2−/−) mice with a defect in T and B cell maturation carrying a human MxA transgene (MxAtg/−Rag2−/−). In these mice, the virus could replicate for several weeks facilitating host adaptation. In MxAtg/−Rag2−/−, but not in Rag2−/− mice, the well-described mammalian adaptation E627K in the viral polymerase subunit PB2 was acquired, but no variants with MxA escape mutations in NP were detected. Utilizing reverse genetics, we could show that acquisition of PB2 E627K allowed partial evasion from MxA restriction in MxAtg/tg mice. However, pretreatment with type I interferon decreased viral replication in these mice, suggesting that PB2 E627K is not a true MxA escape mutation. Based on these results, we speculate that it might be difficult for H7N9 to acquire MxA escape mutations in the viral NP. This is consistent with previous findings showing that MxA escape mutations cause severe attenuation of IAVs of avian origin. [ABSTRACT FROM AUTHOR]

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The zoonotic H7N9 avian influenza (AI) virus first emerged in 2013 as a low pathogenic (LPAI) strain, and has repeatedly caused human infection resulting in severe respiratory illness and a mortality of ~39% (>600 deaths) across five epidemic waves. This virus has circulated in poultry with little to no discernible clinical signs, making detection and control difficult. Contrary to published data, our group has observed a subset of specific pathogen free chickens infected with the H7N9 virus succumb to disease, showing clinical signs consistent with highly pathogenic AI (HPAI). Viral genome sequencing revealed two key mutations had occurred following infection in the haemagglutinin (HA 226 L>Q) and nucleoprotein (NP 373 A>T) proteins. We further investigated the impact of the NP mutation and demonstrated that only chickens bearing a single nucleotide polymorphism (SNP) in their IFITM1 gene were susceptible to the H7N9 virus. Susceptible chickens demonstrated a distinct loss of CD8+ T cells from the periphery as well as a dysregulation of IFNγ that was not observed for resistant chickens, suggesting a role for the NP mutation in altered T cell activation. Alternatively, it is possible that this mutation led to altered polymerase activity, as the mutation occurs in the NP 360-373 loop which has been previously show to be important in RNA binding. These data have broad ramifications for our understanding of the pathobiology of AI in chickens and humans and provide an excellent model for investigating the role of antiviral genes in a natural host species. [ABSTRACT FROM AUTHOR]

Avian influenza remains a threat to human wellbeing. Hypochlorite derivatives are commonly used as disinfectants to prevent the spread of the disease. The World Health Organization (WHO) has listed chlorine dioxide (ClO2) as an A1-level, safe, and efficient disinfectant. In this study, we tested the efficacy of ClO2, in aqueous solution and gas forms, against avian influenza A (H7N9) virus. The virus suspension was mixed with ClO2 aqueous solutions of various concentrations and for various time intervals. Aliquots of the mixture were then serially diluted, and the 50% tissue culture infective dose (TCID50) was measured with a hemagglutination test on MDCK cells. ClO2 gas produced from generators was introduced in a chamber containing the virus suspension in a Petri dish. The infective activity of the surviving virus was measured by the hemagglutination test. An aqueous solution of ClO2 at 126 µg/mL for 15 s was effective given that no surviving virus was detected with the hemagglutination test. ClO2 gas at >5 µL/L sustained for 1 h inactivated the virus effectively, while at 2.5 µL/L for 1 h, it only partially inactivated the virus. ClO2 as gas or aqueous solution at a certain concentration is effective in inactivating the H7N9 virus, and can be applied for the decontamination and disinfection of environments.

The H7 subtype avian influenza viruses are circulating widely worldwide, causing significant economic losses to the poultry industry and posing a serious threat to human health. In 2019, H7N2 and H7N9 co-circulated in Chinese poultry, yet the risk of H7N2 remained unclear. We isolated and sequenced four H7N2 viruses from chickens, revealing them as novel reassortants with H7N9-derived HA, M, NS genes and H9N2-derived PB2, PB1, PA,NP, NA genes. To further explore the key segment of pathogenicity, H7N2-H7N9NA and H7N2-H9N2HA single-substitution were constructed. Pathogenicity study showed H7N2 isolates to be highly pathogenic in chickens, with H7N2-H7N9NA slightly weaker than H7N2-Wild type. Transcriptomic analysis suggested that H7N9-derived HA genes primarily drove the high pathogenicity of H7N2 isolates, eliciting a strong inflammatory response. These findings underscored the increased threat posed by reassorted H7N2 viruses to chickens, emphasizing the necessity of long-term monitoring of H7 subtype avian influenza viruses. • Four highly pathogenic H7N2 avian influenza viruses were isolated in China during 2019. • H7N2 isolates were reassortants with H7N9-derived HA genes and H9N2-derived NA genes. • H7N9-derived HA genes might be a critical virulence contributor to novel H7 avian influenza viruses. [ABSTRACT FROM AUTHOR]

Strategies that improve influenza vaccine immunogenicity are critical for the development of vaccines for pandemic preparedness. Hemagglutinin (HA)-specific CD4+ T cell epitopes support protective B cell responses against seasonal influenza. However, in the case of avian H7N9, which poses a pandemic threat, HA elicits only weak neutralizing antibody responses in infection and vaccination without adjuvant. We hypothesized that an immune-engineered H7N9 HA incorporating a broadly reactive H3N2 HA-specific memory CD4+ T cell epitope that replaces a regulatory T cell-inducing epitope at the corresponding position in H7N9 HA could harness preexisting influenza T cell immunity to increase CD4+ T cells that are needed for protective antibody development. We designed and produced a virus-like particle (VLP) vaccine that carries the epitope augmented H7N9 HA (OPT1) and immunized HLA-DR3 transgenic mice with established H3N2 immunity. OPT1-VLPs stimulated higher stem cell, central, and effector memory CD4+ T cell levels over wild type VLP immunization. In addition, activated, IL-21-producing follicular helper T cell frequencies were enhanced. This novel immunogen design strategy illustrates that site-specific modifications aimed to augment T cell epitope content enhance CD4+ T cell responses among critical subpopulations capable of aiding protective immune responses upon antigen re-encounter and that mobilization of immune memory can be used to overcome the poor immunogenicity of avian influenza viruses.

ObjectiveThe outbreak of COVID-19 in 2020 is reminiscent of the H7N9 outbreak in 2013, which poses a huge threat to human health. We aim to compare clinical features and survival factors in fatal cases of COVID-19 and H7N9.MethodsData on confirmed COVID-19 and H7N9 fatal cases identified in mainland China were analyzed to compare demographic characteristics and clinical severity. Survival curves were estimated by the Kaplan–Meier method and compared using log-rank tests and a restricted mean survival time model. A Cox regression model was used to identify survival factors in fatal cases of COVID-19 and H7N9.ResultsSimilar demographic characteristics were observed in fatal cases of COVID-19 and H7N9. The proportion of fatal cases of H7N9 receiving antibiotics, antiviral drugs, and oxygen treatment was higher than that of COVID-19. The potential protective factors for fatal COVID-19 cases were receiving antibiotics (HR: 0.37, 95% CI: 0.22–0.61), oxygen treatment (HR: 0.66, 95% CI: 0.44–0.99), and corticosteroids (HR: 0.46, 95% CI: 0.35–0.62). In contrast, antiviral drugs (HR: 0.21, 95% CI: 0.08–0.56) and corticosteroids (HR: 0.45, 95% CI: 0.29–0.69) were the protective factors for H7N9 fatal cases.ConclusionThe proportion of males, those having one or more underlying medical condition, and older age was high in COVID-19 and H7N9 fatal cases. Offering antibiotics, oxygen treatment, and corticosteroids to COVID-19 cases extended the survival time. Continued global surveillance remains an essential component of pandemic preparedness.

The zoonotic H7N9 avian influenza (AI) virus first emerged in 2013 as a low pathogenic (LPAI) strain, and has repeatedly caused human infection resulting in severe respiratory illness and a mortality of ~39% (>600 deaths) across five epidemic waves. This virus has circulated in poultry with little to no discernible clinical signs, making detection and control difficult. Contrary to published data, our group has observed a subset of specific pathogen free chickens infected with the H7N9 virus succumb to disease, showing clinical signs consistent with highly pathogenic AI (HPAI). Viral genome sequencing revealed two key mutations had occurred following infection in the haemagglutinin (HA 226 L>Q) and nucleoprotein (NP 373 A>T) proteins. We further investigated the impact of the NP mutation and demonstrated that only chickens bearing a single nucleotide polymorphism (SNP) in their IFITM1 gene were susceptible to the H7N9 virus. Susceptible chickens demonstrated a distinct loss of CD8+ T cells from the periphery as well as a dysregulation of IFNγ that was not observed for resistant chickens, suggesting a role for the NP mutation in altered T cell activation. Alternatively, it is possible that this mutation led to altered polymerase activity, as the mutation occurs in the NP 360-373 loop which has been previously show to be important in RNA binding. These data have broad ramifications for our understanding of the pathobiology of AI in chickens and humans and provide an excellent model for investigating the role of antiviral genes in a natural host species.

The influenza A (H7N9) virus has been seriously concerned for its potential to cause an influenza pandemic. To understand the spread and evolution process of the virus, a spatial and temporal Bayesian evolutionary analysis was conducted on 2,052 H7N9 viruses isolated during 2013 and 2018. It revealed that the H7N9 virus was probably emerged in a border area of Anhui Province in August 2012, approximately 6 months earlier than the first human case reported. Two major epicenters had been developed in the Yangtze River Delta and Peral River Delta regions by the end of 2013, and from where the viruses have also spread to other regions at an average speed of 6.57 km/d. At least 24 genotypes showing have been developed and each of them showed a distinct spatio-temporal distribution pattern. Furthermore, A random forest algorithm-based model has been developed to predict the occurrence risk of H7N9 virus. The model has a high overall forecasting precision (> 97%) and the monthly H7N9 occurrence risk for each county of China was predicted. These findings provide new insights for a comprehensive understanding of the origin, evolution, and occurrence risk of H7N9 virus. Moreover, our study also lays a theoretical basis for conducting risk-based surveillance and prevention of the disease. [ABSTRACT FROM AUTHOR]

• Patients with H7N9- acute respiratory distress syndrome (ARDS) showed a more acute disease course and severity. • Disseminated intravascular coagulation was more common in patients with COVID-19-ARDS. • The main causes of death between H7N9-ARDS and COVID-19-ARDS were different. • This is the first study to compare clinical features between the two diseases. We aimed to compared the clinical features of acute respiratory distress syndrome (ARDS) induced by COVID-19 and H7N9 virus infections. Clinical data of 100 patients with COVID-19 and 46 patients with H7N9 were retrospectively analyzed. Elevated inflammatory indices and coagulation disorders were more common in COVID-19-ARDS group than in the H7N9-ARDS group. The median interval from illness onset to ARDS development was shorter in H7N9-ARDS. The PaO 2 /FiO 2 level was lower in H7N9-ARDS, whereas the Sepsis-related Organ Failure Assessment score was higher in COVID-19-ARDS. The proportion of patients with disseminated intravascular coagulation and liver injury in COVID-19-ARDS and H7N9-ARDS was 45.5% versus 3.1% and 28.8% versu s 50%, respectively (P <0.05). The mean interval from illness onset to death was shorter in H7N9-ARDS. A total of 59.1% patients with H7N9-ARDS died of refractory hypoxemia compared with 28.9% with COVID-19-ARDS (P = 0.014). Patients with COVID-19-ARDS were more likely to die of septic shock and multiple organ dysfunction compared with H7N9-ARDS (71.2% vs 36.4%, P = 0.005). Patients with H7N9 were more susceptible to develop severe ARDS and showed a more acute disease course. COVID-19-ARDS was associated with severe inflammatory response and coagulation dysfunction, whereas liver injury was more common in H7N9-ARDS. The main causes of death between patients with the two diseases were different. [ABSTRACT FROM AUTHOR]

The outcome of infection with influenza A virus is determined by a complex virus-host interaction. A new H7N9 virus of avian origin crossed the species barrier to infect humans, causing high mortality and emerged as a potential pandemic threat. The mechanisms underlying the virulence and pathogenicity of H7N9 virus remains elusive. H7N9 virus originated from a genetic assortment that involved the avian H9N2 virus, which was the donor of the six internal genes. Unlike the H7N9 virus, the H9N2 virus caused only mild phenotype in infected mice. In this study, we used the mouse infection model to dissect the difference in the host response between the H7N9 and H9N2 viruses. Through analyzing transcriptomics of infected lungs, we surprisingly found that the H9N2 infection elicited an earlier induction of innate immunity than H7N9 infection. This finding was further corroborated by an immunohistochemical study demonstrating earlier recruitment of macrophage to the H9N2-infected lung than the H7N9-infected lung, which could occur as early as 6 hours post infection. In contrast, H7N9 infection was characterized by a late, strong lung CD8+ T cell response that is more robust than H9N2 infection. The different pattern of immune response may underlie more severe lung pathology caused by H7N9 infection compared to H9N2 infection. Finally, we could show that co-infection of the H9N2 virus protected mice from the challenge of both H7N9 and PR8 viruses, thereby strengthening the importance of the induction of an early innate immunity in the host's defense against influenza infection. Collectively, our study unraveled a previously unidentified difference in host response between H7N9 and H9N2 infection and shed new insight on how virus-host interaction shapes the in vivo outcome of influenza infection. [ABSTRACT FROM AUTHOR]

Influenza virus infections continue to be a significant and recurrent public health problem. Although vaccine efficacy varies, regular immunisation is the most effective method for suppressing the influenza virus. Antiviral drugs are available for influenza, although two of the four FDA-approved antiviral treatments have resulted in significant drug resistance. Therefore, new treatments are being sought to reduce the burden of flu-related illness. The time-consuming development of treatments for new and re-emerging diseases such as influenza and the high failure rate are increasing concerns. In this context, we used an in silico-based drug repurposing method to repurpose FDA-approved drugs as potential therapies against the H7N9 virus. To find potential inhibitors, a total of 2568 drugs were screened. Promacta, tucatinib, and lurasidone were identified as promising hits in the DrugBank database. According to the calculations of MM-GBSA, tucatinib (−54.11 kcal/mol) and Promacta (−56.20 kcal/mol) occupied the active site of neuraminidase with a higher binding affinity than the standard drug peramivir (−49.09 kcal/mol). Molecular dynamics (MD) simulation studies showed that the C-α atom backbones of the complexes of tucatinib and Promacta neuraminidase were stable throughout the simulation period. According to ADME analysis, the hit compounds have a high gastrointestinal absorption (GI) and do not exhibit properties that allow them to cross the blood–brain barrier (BBB). According to the in silico toxicity prediction, Promacta is not cardiotoxic, while lurasidone and tucatinib show only weak inhibition. Therefore, we propose to test these compounds experimentally against the influenza H7N9 virus. The investigation and validation of these potential H7N9 inhibitors would be beneficial in order to bring these compounds into clinical settings. [ABSTRACT FROM AUTHOR]

The use of vaccinations and antiviral medications have gained popularity in the therapeutic management of avian influenza H7N9 virus lately. Antiviral medicines are more popular due to being readily available. The presence of the neuraminidase protein in the avian influenza H7N9 virus and its critical role in the cleavage of sialic acid have made it a target drug in the development of influenza virus drugs. Generally, the neuraminidase proteins have common conserved amino acid residues and any mutation that occurs around or within these conserved residues affects the susceptibility and replicability of the influenza H7N9 virus. Herein, we investigated the interatomic and intermolecular dynamic impacts of the experimentally reported E119V mutation on the oseltamivir resistance of the influenza H7N9 virus. We extensively employed molecular dynamic (MD) simulations and subsequent post-MD analyses to investigate the binding mechanisms of oseltamivir-neuraminidase wildtype and E119V mutant complexes. The results revealed that the oseltamivir-wildtype complex was more thermodynamically stable than the oseltamivir-E119V mutant complex. Oseltamivir exhibited a greater binding affinity for wildtype (−15.46 ± 0.23 kcal/mol) relative to the E119V mutant (−11.72 ± 0.21 kcal/mol). The decrease in binding affinity (−3.74 kcal/mol) was consistent with RMSD, RMSF, SASA, PCA, and hydrogen bonding profiles, confirming that the E119V mutation conferred lower conformational stability and weaker protein–ligand interactions. The findings of this oseltamivir-E119V mutation may further assist in the design of compounds to overcome E119V mutation in the treatment of influenza H7N9 virus patients. [ABSTRACT FROM AUTHOR]

Most Chinese provinces have a daily‐updated database of live animal movements; however, the data are not efficiently utilized to support interventions to control H7N9 and other avian influenzas. Based on official records, this study assessed the spatio‐temporal patterns of live broilers moved out of and within Guangxi in 2017. The yearly and monthly networks were analyzed for inter‐ and intra‐provincial movements, respectively. Approximately 200,000 movements occurred in 2017, involving the transport of 200 million live broilers from Guangxi. Although Guangxi exported to 24 out of 32 provinces of China, 95% of inter‐provincial movements occurred with three bordering provinces. Within Guangxi, counties were highly connected through the live broiler movements, creating conditions for rapid virus spreading throughout the province. Interestingly, a peak in movements during the Chinese Lunar New Year celebrations, late January in 2017, was not observed in this study, likely due to H7N9‐related control measures constraining live bird trading. Both intra‐ and inter‐provincial movements in March 2017 were significantly higher than in other months of that year, suggesting that dramatic price changes may influence the movement's network and reshape the risk pathways. However, despite these variations, the same small proportion of counties (less than 20%) exporting/importing more than 90% of inter‐ and intra‐provincial movements remains the same throughout the year. Interventions, particularly surveillance and improving biosecurity, targeted to those counties are thus likely to be more effective for avian influenza risk mitigation than implemented indiscriminately. Additionally, simulations further demonstrated that targeting counties according to their degree or betweenness in the movement network would be the most efficient way to limit disease transmission via broiler movements. The study findings provide evidence to support the design of risk‐based control interventions for H7N9 and all other avian influenza viruses in broiler value chains in Guangxi. [ABSTRACT FROM AUTHOR]

In response to a sudden increase in H7N9 human infections, China introduced an H5/H7 bivalent inactivated vaccine for poultry in Guangxi and Guangdong provinces in July 2017, which subsequently became integrated into the existing compulsory national H5N1 vaccination programme from September 2017. Although the vaccination programme effectively reduced H7N9 infections in humans and poultry, there are ongoing arguments against continuing this long‐term vaccination. These discussions have drawn policymakers to think about the possibility of stopping routine vaccination for H7N9 avian influenza viruses (AIVs) in China; however, they have not considered the poultry industry stakeholders' practices on and attitudes towards this vaccination. This study investigated H7N9 vaccination practices in the yellow broiler industry in Guangxi and stakeholders' attitudes on H7N9 vaccination, using a mixed methods design. The study found H7N9 vaccination was well adopted in the yellow broiler industry in Guangxi regardless of the source of the vaccines. Most stakeholders believed vaccination was the best measure to control H7N9 and H5N1 AIVs, and they showed a strong willingness to continue with vaccination even without government subsidies or freely provided vaccines. The motivations by stakeholders for using vaccines to control H7N9 and H5N1 were different due to the epidemiological differences between the two strains. Understanding poultry industry stakeholders' practices and attitudes on H7N9 vaccination has important practical implications in planning vaccination policies, particularly when considering the possibility of vaccination withdrawal. [ABSTRACT FROM AUTHOR]

Between 2013 and 2018, the novel A/Anhui/1/2013 (AH/13)-lineage H7N9 virus caused at least five waves of outbreaks in humans, totaling 1,567 confirmed human cases in China. Surveillance data indicated a disproportionate distribution of poultry infected with this AH/13-lineage virus, and laboratory experiments demonstrated that this virus can efficiently spread among chickens but not among Pekin ducks. The underlying mechanism of this selective transmission remains unclear. In this study, we demonstrated the absence of Neu5Gc expression in chickens across all respiratory and gastrointestinal tissues. However, Neu5Gc expression varied among different duck species and even within the tissues of the same species. The AH/13-lineage viruses exclusively bind to acetylneuraminic acid (Neu5Ac), in contrast to wild waterbird H7 viruses that bind both Neu5Ac and N-glycolylneuraminic acid (Neu5Gc). The level of Neu5Gc expression influences H7 virus replication and facilitates adaptive mutations in these viruses. In summary, our findings highlight the critical role of Neu5Gc in affecting the host range and interspecies transmission dynamics of H7 viruses among avian species.IMPORTANCEMigratory waterfowl, gulls, and shorebirds are natural reservoirs for influenza A viruses (IAVs) that can occasionally spill over to domestic poultry, and ultimately humans. This study showed wild-type H7 IAVs from waterbirds initially bind to glycan receptors terminated with N-acetylneuraminic acid (Neu5Ac) or N-glycolylneuraminic acid (Neu5Gc). However, after enzootic transmission in chickens, the viruses exclusively bind to Neu5Ac. The absence of Neu5Gc expression in gallinaceous poultry, particularly chickens, exerts selective pressure, shaping IAV populations, and promoting the acquisition of adaptive amino acid substitutions in the hemagglutinin protein. This results in the loss of Neu5Gc binding and an increase in virus transmissibility in gallinaceous poultry, particularly chickens. Consequently, the transmission capability of these poultry-adapted H7 IAVs in wild water birds decreases. Timely intervention, such as stamping out, may help reduce virus adaptation to domestic chicken populations and lower the risk of enzootic outbreaks, including those caused by IAVs exhibiting high pathogenicity.

Between 2013 and 2017, the A/Anhui/1/13-lineage (H7N9) low-pathogenicity avian influenza virus (LPAIV) was epizootic in chickens in China, causing mild disease, with 616 fatal human cases. Despite poultry vaccination, H7N9 has not been eradicated. Previously, we demonstrated increased pathogenesis in turkeys infected with H7N9, correlating with the emergence of the L217Q (L226Q H3 numbering) polymorphism in the haemagglutinin (HA) protein. A Q217-containing virus also arose and is now dominant in China following vaccination. We compared infection and transmission of this Q217-containing 'turkey-adapted' (ty-ad) isolate alongside the H7N9 (L217) wild-type ( wt ) virus in different poultry species and investigated the zoonotic potential in the ferret model. Both wt and ty-ad viruses demonstrated similar shedding and transmission in turkeys and chickens. However, the ty-ad virus was significantly more pathogenic than the wt virus in turkeys but not in chickens, causing 100 and 33% mortality in turkeys respectively. Expanded tissue tropism was seen for the ty-ad virus in turkeys but not in chickens, yet the viral cell receptor distribution was broadly similar in the visceral organs of both species. The ty-ad virus required exogenous trypsin for in vitro replication yet had increased replication in primary avian cells. Replication was comparable in mammalian cells, and the ty-ad virus replicated successfully in ferrets. The L217Q polymorphism also affected antigenicity. Therefore, H7N9 infection in turkeys can generate novel variants with increased risk through altered pathogenicity and potential HA antigenic escape. These findings emphasize the requirement for enhanced surveillance and understanding of A/Anhui/1/13-lineage viruses and their risk to different species.