Pathogenic Influenza Virus Research Articles

Usp25-Erlin1/2 activity limits cholesterol flux to restrict virus infection

Reprogramming lipid metabolic pathways is a critical feature of activating immune responses to infection. However, how these reconfigurations occur is poorly understood. Our previous screen to identify cellular deubiquitylases (DUBs) activated during influenza virus infection revealed Usp25 as a prominent hit. Here, we show that Usp25-deleted human lung epithelial A549 cells display a >10-fold increase in pathogenic influenza virus production, which was rescued upon reconstitution with the wild type but not the catalytically deficient (C178S) variant. Proteomic analysis of Usp25 interactors revealed a strong association with Erlin1/2, which we confirmed as its substrate. Newly synthesized Erlin1/2 were degraded in Usp25-/- or Usp25C178S cells, activating Srebp2, with increased cholesterol flux and attenuated TLR3-dependent responses. Our study therefore defines the function of a deubiquitylase that serves to restrict a range of viruses by reprogramming lipid biosynthetic flux to install appropriate inflammatory responses.

Genetic characteristics of the first human infection with the G4 genotype eurasian avian-like H1N1 swine influenza virus in Shaanxi Province,China

Objective: To analyze the genetic characteristics of the first human infection with the G4 genotype of Eurasian avian H1N1 swine influenza virus (EA H1N1 SIV) in Shaanxi Province. Methods: The patient's throat swab samples were collected, and MDCK cells were inoculated for virus isolation to obtain the virus strain. The whole genome deep sequencing method was used to obtain the eight gene segments of the isolated strain. The nucleotide homology analysis was conducted through the Blast program in the GenBank database, and a phylogenetic tree was constructed to analyze the genetic characteristics of the virus. Results: The throat swab specimens of the case were confirmed as EA H1N1 SIV in the laboratory, and the isolated strain was named A/Shaanxi-Weicheng/1351/2022(H1N1v). Homology analysis found that the PB2, NP, HA, NA, and M genes of this isolate had the highest nucleotide homology with A/swing/Beijing/0301/2018 (H1N1), about 98.29%, 98.73%, 97.41%, 97.52%, and 99.08%, respectively. The phylogenetic tree showed that the isolate belonged to G4 genotype EA H1N1 SIV, with PB2, PB1, PA, NP and M genes from pdm/09 H1N1, HA and NA genes from EA H1N1, and NS gene from Triple-reassortant H1N1. The cleavage site of the HA protein was IPSIQSR↓G, which was the molecular characteristic of the low pathogenic influenza virus. No amino acid mutations associated with neuraminidase inhibitors were found in the NA protein. PB2 protein 701N mutation, PA protein P224S mutation, NP protein Q357K mutation, M protein P41A mutation, and NS protein 92D all indicated its enhanced adaptability to mammals. Conclusion: The patient is the first human infection with G4 genotype EA H1N1 SIV in Shaanxi province. The virus is low pathogenic, but its adaptability to mammals is enhanced. Therefore, it is necessary to strengthen the monitoring of such SIVs.

Pathogenicity of H5N8 avian influenza virus in chickens and in duck breeds and the role of MX1 and IFN-α in infection outcome and transmission to contact birds

This study examined the pathogenicity, immunogenicity, and transmission potential of the H5N8 HPAI clade 2.3.4.4b virus in three breeds of ducks and in broiler chickens. Chickens, Muscovy, Pekin, and Mallard ducks (n = 10) received a dose of 6 log10 EID50 of HPAIV H5N8 directly. Nine contact chickens were introduced to each group on the day of infection. All infected chickens died, with MDT of 7.6 days. Muscovy and Pekin ducks died by 11.1% and 10%, respectively, with MDTs of 7 and 6 days. No Mallards died but showed more severe clinical disease than Pekin ducks. Mallards had the highest MX1 gene expression in the lung and spleen and IFN-α in the spleen. MX1 expression levels were lower in the spleen and lung of Pekin ducks, in the spleen of chickens and in the lung of Muscovy ducks than in noninfected controls. However, viral shedding was higher in ducks than in chickens and was highest in Mallards. 66.7% of chickens placed in contact with infected chickens died and 77.8% of in-contact chickens to infected three duck breeds died. In conclusion, there was a diversity in sensitivity and immunogenicity for HPAIV H5N8 among duck breeds, resulting in diverse infection outcomes and transmissibility to contacts. This study provides duck/chicken interface models for HPAIV transmission to poultry.

Continued Circulation of Highly Pathogenic H5 Influenza Viruses in Vietnamese Live Bird Markets in 2018-2021.

We isolated 77 highly pathogenic avian influenza viruses during routine surveillance in live poultry markets in northern provinces of Vietnam from 2018 to 2021. These viruses are of the H5N6 subtype and belong to HA clades 2.3.4.4g and 2.3.4.4h. Interestingly, we did not detect viruses of clade 2.3.4.4b, which in recent years have dominated in different parts of the world. The viruses isolated in this current study do not encode major determinants of mammalian adaptation (e.g., PB2-E627K or PB1-D701N) but possess amino acid substitutions that may affect viral receptor-binding, replication, or the responses to human antiviral factors. Several of the highly pathogenic H5N6 virus samples contained other influenza viruses, providing an opportunity for reassortment. Collectively, our study demonstrates that the highly pathogenic H5 viruses circulating in Vietnam in 2018-2021 were different from those in other parts of the world, and that the Vietnamese H5 viruses continue to evolve through mutations and reassortment.

The SUMO-interacting motif in NS2 promotes adaptation of avian influenza virus to mammals.

Species differences in the host factor ANP32A/B result in the restriction of avian influenza virus polymerase (vPol) in mammalian cells. Efficient replication of avian influenza viruses in mammalian cells often requires adaptive mutations, such as PB2-E627K, to enable the virus to use mammalian ANP32A/B. However, the molecular basis for the productive replication of avian influenza viruses without prior adaptation in mammals remains poorly understood. We show that avian influenza virus NS2 protein help to overcome mammalian ANP32A/B-mediated restriction to avian vPol activity by promoting avian vRNP assembly and enhancing mammalian ANP32A/B-vRNP interactions. A conserved SUMO-interacting motif (SIM) in NS2 is required for its avian polymerase-enhancing properties. We also demonstrate that disrupting SIM integrity in NS2 impairs avian influenza virus replication and pathogenicity in mammalian hosts, but not in avian hosts. Our results identify NS2 as a cofactor in the adaptation process of avian influenza virus to mammals.

Highly Pathogenic H5 Influenza Viruses Isolated between 2016 and 2017 in Vietnamese Live Bird Markets.

Routine surveillance in live poultry markets in the northern regions of Vietnam from 2016 to 2017 resulted in the isolation of 27 highly pathogenic avian H5N1 and H5N6 viruses of 3 different clades (2.3.2.1c, 2.3.4.4f, and 2.3.4.4g). Sequence and phylogenetic analysis of these viruses revealed reassortment with various subtypes of low pathogenic avian influenza viruses. Deep-sequencing identified minor viral subpopulations encoding variants that may affect pathogenicity and sensitivity to antiviral drugs. Interestingly, mice infected with two different clade 2.3.2.1c viruses lost body weight rapidly and succumbed to virus infection, whereas mice infected with clade 2.3.4.4f or 2.3.4.4g viruses experienced non-lethal infections.

Review on the Epizootiological Situation on Highly Pathogenic Avian Influenza around the World and in Russia in 2022

This paper describes the current situation on highly pathogenic avian influenza virus in 2022 and provides forecast of the possible further spread of avian influenza in Russia. In 2022, the circulation of a wide variety of highly pathogenic avian influenza virus subtypes, which have epizootiological and epidemiological significance, was recorded in the world. Outbreaks caused by highly pathogenic avian influenza virus were reported in over 60 countries. In addition, human infections with influenza viruses of the A(H5Nx) and A(H9N2) subtypes were registered. There was a large-scale epizootic which affected more than 10 regions of the European part of Russia and the Russian Far East in 2022. Outbreaks among wild birds and poultry were caused by the highly pathogenic influenza virus A(H5N1) of the clade 2.3.4.4b, at the same time genetic and antigenic diversity was observed among viruses circulating in Russia. Thus, an essential geographical role of the territory of Russia in the global spread of avian influenza virus has been shown once again, which highlights the importance of continuous avian influenza virus surveillance in the country.

4'-Fluorouridine mitigates lethal infection with pandemic human and highly pathogenic avian influenza viruses.

Influenza outbreaks are associated with substantial morbidity, mortality and economic burden. Next generation antivirals are needed to treat seasonal infections and prepare against zoonotic spillover of avian influenza viruses with pandemic potential. Having previously identified oral efficacy of the nucleoside analog 4'-Fluorouridine (4'-FlU, EIDD-2749) against SARS-CoV-2 and respiratory syncytial virus (RSV), we explored activity of the compound against seasonal and highly pathogenic influenza (HPAI) viruses in cell culture, human airway epithelium (HAE) models, and/or two animal models, ferrets and mice, that assess IAV transmission and lethal viral pneumonia, respectively. 4'-FlU inhibited a panel of relevant influenza A and B viruses with nanomolar to sub-micromolar potency in HAE cells. In vitro polymerase assays revealed immediate chain termination of IAV polymerase after 4'-FlU incorporation, in contrast to delayed chain termination of SARS-CoV-2 and RSV polymerase. Once-daily oral treatment of ferrets with 2 mg/kg 4'-FlU initiated 12 hours after infection rapidly stopped virus shedding and prevented transmission to untreated sentinels. Treatment of mice infected with a lethal inoculum of pandemic A/CA/07/2009 (H1N1)pdm09 (pdmCa09) with 4'-FlU alleviated pneumonia. Three doses mediated complete survival when treatment was initiated up to 60 hours after infection, indicating a broad time window for effective intervention. Therapeutic oral 4'-FlU ensured survival of animals infected with HPAI A/VN/12/2003 (H5N1) and of immunocompromised mice infected with pdmCa09. Recoverees were protected against homologous reinfection. This study defines the mechanistic foundation for high sensitivity of influenza viruses to 4'-FlU and supports 4'-FlU as developmental candidate for the treatment of seasonal and pandemic influenza.

Influenza viral matrix 1 protein aggravates viral pathogenicity by inducing TLR4-mediated reactive oxygen species production and apoptotic cell death

Influenza virus is one of the most challenging viruses threating human health. Since infection with influenza virus triggers inflammatory responses and induces cell death, the molecular and cellular mechanisms by which the virus-infected cells undergo apoptotic and necrotic cell death have been widely studied. However, most of the studies have focused on the molecular events occurring in the cytosol and there is limited information on the physiological correlation between virus-induced cell death and the viral pathogenesis in vivo. In this study, we demonstrate that the influenza virus matrix 1 (M1) protein is released from virus-infected cells and triggers apoptotic cell death of lung epithelial and pulmonary immune cells, through the activation of Toll-like receptor 4 (TLR4) signaling. Treatment with M1 protein led to robust cellular inflammatory responses, such as the production of proinflammatory cytokines and cellular reactive oxygen species (ROS), and induction of cell death. When M1 protein was administered in vivo, it induced the activation of inflammatory responses and cell death in the lungs. Furthermore, the administration of M1 aggravated lung pathology and mortality of the virus-infected mice in a TLR4-dependent manner. These results demonstrate that M1 is an important pathogenic factor contributing to influenza virus pathogenicity by enhancing cell death in the lungs, thereby expanding our understanding of the molecular mechanism of influenza virus-induced cell death through the interaction with an innate immune receptor.

Pathogenicity, transmissibility, and immunogenicity of recombinant H9N2 avian influenza viruses based on representative viruses of Southeast China

H9N2 is currently the main subtype of avian influenza in China. In order to use reverse genetics to rapid preparation of seed strains for vaccine production, and intend to prevent and control the H9N2 subtype epidemic strains of avian influenza virus (AIV). In this study, we successfully rescued 2 H9N2 recombinant viruses based on the representative viruses of Southeast China and confirmed by RT-PCR and sequencing. Genetic stability, pathogenicity, transmissibility, and antigenicity of 2 recombinant viruses were evaluated. Compared to the FZ1, the growth kinetics of H9N2(HA+NA)/PR8 showed no significant difference, H9N2(HA+NA+M+PB1)/PR8 was slightly lower. Our study also confirmed 2 recombinant viruses had good genetic stability after 10 passages but possessed lower pathogenicity than FZ1. Although both recombinant viruses led to seroconversion in all inoculated birds on 14 dpi, they complete loss of viral transmission of the virus to contact birds. In addition, birds were immunized via hypodermic route by inactivated vaccines of H9N2(HA+NA)/PR8, H9N2(HA+NA+M+PB1)/PR8 and wild-type virus with a single dose, and the results showed that the hemagglutination inhibition titers on 21 dpv were 10.5, 9.6, and 10.5 log2, respectively. And recombinant viruses both provided a certain protection against wild-type virus challenge. In conclusion, these data indicated that 2 recombinant viruses will be expected to be used as inactivated vaccines to controlling the spread of H9N2 subtype AIV even have potential application for attenuated viral vaccines, which provides a reference for the prevention and control of influenza virus pandemics.

Protective Potential of Saussurea costus (Falc.) Lipsch. Roots against Cyclophosphamide-Induced Pulmonary Injury in Rats and Its In Vitro Antiviral Effect.

Diseases and infections of the respiratory tract are common global causes of morbidity and mortality. Our study attempts to elucidate a novel remedy for respiratory ailments, in addition to identifying and quantifying the metabolites of Saussurea costus root extract (SCRE) using HPLC. Then, in vitro antiviral and in vivo lung protective effects were elucidated. The in vitro antiviral potential of SCRE was analyzed via plaque assay against the low pathogenic human coronavirus (HCoV-229E) and human influenza virus (H1N1). The value of the half maximal inhibitory concentrations (IC50) of SCRE against HCoV-229E and H1N1 influenza virus were 23.21 ± 1.1 and 47.6 ± 2.3 µg/mL, respectively. SCRE showed a histological improvement, namely a decrease in inducible nitric oxide synthase (iNOS) and caspase-3 immunoexpression in in vivo cyclophosphamide (CP)-induced acute lung injury (ALI). Moreover, there was a considerable decline in microRNA-let-7a gene expression and a significant rise in heme oxygenase-1 (HO-1) gene expression, with a marked decrease in the malondialdehyde (MDA) level. Molecular docking studies revealed that the major constituents of SCRE have a good affinity for caspase-3, HO-1, and iNOS proteins. In conclusion, a traditional plant SCRE could be a promising source of novel therapeutic agents for treating and protecting respiratory tract diseases. More future investigations should be carried out to reveal its efficacy clinically.

Experimental Pathogenicity of H9N2 Avian Influenza Viruses Harboring a Tri-Basic Hemagglutinin Cleavage Site in Sonali and Broiler Chickens.

Low-pathogenic avian influenza (LPAI) H9N2 virus is endemic in Bangladesh, causing huge economic losses in the poultry industry. Although a considerable number of Bangladeshi LPAI H9N2 viruses have been molecularly characterized, there is inadequate information on the pathogenicity of H9N2 viruses in commercial poultry. In this study, circulating LPAI H9N2 viruses from recent field outbreaks were characterized, and their pathogenicity in commercial Sonali (crossbred) and broiler chickens was assessed. Phylogenetic analysis of currently circulating field viruses based on the hemagglutinin (HA) and neuraminidase (NA) gene sequences revealed continuous circulation of G1 lineages containing the tri-basic hemagglutinin cleavage site (HACS) motif (PAKSKR*GLF) at the HA protein. Both the LPAI susceptible Sonali and broiler chickens were infected with selected H9N2 isolates A/chicken/Bangladesh/2458-LT2/2020 or A/chicken/Bangladesh/2465-LT56/2021 using intranasal (100 µL) and intraocular (100 µL) routes with a dose of 106 EID50/mL. Infected groups (LT_2-So1 and LT_56-So2; LT_2-Br1 and LT_56-Br2) revealed no mortality or clinical signs. However, at gross and histopathological investigation, the trachea, lungs, and intestine of the LT_2-So1 and LT_56-So2 groups displayed mild to moderate hemorrhages, congestion, and inflammation at different dpi. The LT 2-Br1 and LT 56-Br2 broiler groups showed nearly identical changes in the trachea, lungs, and intestine at various dpi, indicating no influence on pathogenicity in the two commercial bird species under study. Overall, the prominent lesions were observed up to 7 dpi and started to disappear at 10 dpi. The H9N2 viruses predominantly replicated in the respiratory tract, and higher titers of virus were shed through the oropharyngeal route than the cloacal route. Finally, this study demonstrated the continuous evolution of tri-basic HACS containing H9N2 viruses in Bangladesh with a low-pathogenic phenotype causing mild to moderate tracheitis, pneumonia, and enteritis in Sonali and commercial broiler chickens.

Comprehensive analysis of lipid metabolism in influenza virus infection.

Influenza A virus (IAV) exploits host metabolic pathways to support its replication. To improve the understanding of lipid metabolic changes that could occur upon IAV infection, a comprehensive analysis of lipid metabolites in A549 cells infected with the avian H9N2 virus at the different time points was performed. It was found that H9N2 infection could largely promote the level of lipid metabolites. Further, these metabolites were mainly included in glycerophospholipids (GP), sphingolipid (SP), glycerolipids (GL), fatty acid (FA), sterollipids (ST), triglyceride (TG), and prenol lipids (PR). Specifically, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that these metabolites were mainly associated with the glycerphospholipid metabolism, glycosylphosphatidylinositol (GPI)-anchor biosynthesis, and autophagy. Furthermore, it is interesting to note that these metabolites, including FFA(19:1), PE(P-17:0_20:3), PE(P-18:1_20:2), LPC(14:0/0:0), PE(O-18:0_20:3), and MGDG(16:0_18:1), are upregulated and shared in the top 10 at 12 h, 24 h, 36 h, and 48 h after H9N2 infection, indicative of the possibility of acting as biomarkers for the diagnosis in the lung infected with influenza virus. These pathways and altered metabolites could provide new understandings about biological characteristics and pathogenicity of influenza virus and have the potential to serve as biomarkers for influenza.

Phytoestrogen β-Sitosterol Exhibits Potent In Vitro Antiviral Activity against Influenza A Viruses.

Influenza is a contagious infection in humans that is caused frequently by low pathogenic seasonal influenza viruses and occasionally by pathogenic avian influenza viruses (AIV) of H5, H7, and H9 subtypes. Recently, the clinical sector in poultry and humans has been confronted with many challenges, including the limited number of antiviral drugs and the rapid evolution of drug-resistant variants. Herein, the anti-influenza activities of various plant-derived phytochemicals were investigated against highly pathogenic avian influenza A/H5N1 virus (HPAIV H5N1) and seasonal low pathogenic human influenza A/H1N1 virus (LPHIV H1N1). Out of the 22 tested phytochemicals, the steroid compounds β-sitosterol and β-sitosterol-O-glucoside have very potent activity against the predefined influenza A viruses (IAV). Both steroids could induce such activity by affecting multiple stages during IAV replication cycles, including viral adsorption and replication with a major and significant impact on the virus directly in a cell-free status "viricidal effect". On a molecular level, several molecular docking studies suggested that β-sitosterol and β-sitosterol-O-glucoside exhibited viricidal effects through blocking active binding sites of the hemagglutinin surface protein, as well as showing inhibitory effects against replication through the binding with influenza neuraminidase activity and blocking the active sites of the M2 proton channel activity. The phytoestrogen β-sitosterol has structural similarity with the active form of the female sex hormone estradiol, and this similarity is likely one of the molecular determinants that enables the phytoestrogen β-sitosterol and its derivative to control IAV infection in vitro. This promising anti-influenza activity of β-sitosterol and its O-glycoside derivative, according to both in vitro and cheminformatics studies, recommend both phytochemicals for further studies going through preclinical and clinical phases as efficient anti-influenza drug candidates.

Pathogenicity and pathogenesis of a recent highly pathogenic avian influenza subtype H5N8 in mule ducklings in Egypt.

In late 2017, an H5N8 highly pathogenic avian influenza (HPAI) virus, clade 2.3.4.4, was isolated from domestic ducks in Egypt, which was associated with high morbidity and low mortality. The pathogenicity increased due to the continuous circulation of virus in ducks. Thus, this study aimed to monitor the pathogenesis and pathogenicity of new H5N8 Avian influenza (AI) virus in mule ducklings. The lethal dose 50 (LD50) for this new local HPAI H5N8 isolate was calculated. Twenty ducklings were inoculated with 0.1 mL of dilution containing 10 LD50 HPAI per duck. The clinical signs and mortalities were recorded until 30 days post-infection (DPI) to confirm viral pathogenesis. Reverse transcription polymerase chain reaction was used to detect viral shedding from collected cloacal swabs after 3rd, 5th, 7th, 10th, 14th, 21st, and 30th DPI. The main histopathological lesions associated with the presence of HPAI virus were also recorded on the 3rd and 14th DPI. The result showed that the LD50 of the new HPAI H5N8 was 104 log10. Clinical signs were observed after 2nd DPI, but it was clinically severe on 3rd, 4th, and 5th DPI in the form of respiratory and gastric disorders, forming 90% of all diseased ducklings, whereas 30% of the infected ducks only showed nervous signs. The mortality rate peaked on 4th and 5th DPI with a cumulative mortality rate of 60% for the inoculated ducks, whereas no mortality was recorded after 6th DPI. Dead ducks showed typical postmortem lesions of AI disease. Necrosis and ecchymotic or petechial hemorrhages on the heart, pancreas, liver, and spleen were observed, whereas the lung showed pneumonia. With regard to viral shedding, infected ducklings shed the virus from its gut until 7th DPI, but the number of duck shedders gradually decreased until 14th DPI after viral shedding. The histopathological findings indicated that the spleen and thymus showed necrosis and hemorrhages, whereas the brain showed multifocal malacic foci and spread meningitis. Moreover, the lung had intrabronchial hyaline degeneration and fibrinous pneumonia on 3rd DPI. Furthermore, the liver showed multifocal necrotic foci and subcapsular hemorrhage, whereas the kidney showed remarkable tubular degeneration, mostly within the collecting tubules. Furthermore, the heart showed marked myocardiolysis of the cardiac muscle fibers. On 14th DPI, all histopathological lesions of the examined organs were restored to normal. The currently circulating HPAI H5N8 virus strain has high virulence, particularly for imported mule ducks that originated from non-vaccinated breeder ducks. Therefore, vaccination and quarantine measures must be applied on imported 1-day-old mule ducklings. Moreover, the pathogenesis must be reviewed and monitored for updating circulating AI strains caused by the continuous and rapid evolution of AI viruses.

Identification of N- and C-3-Modified Laudanosoline Derivatives as Novel Influenza PAN Endonuclease Inhibitors

Influenza PAN inhibitors are of particular importance in current efforts to develop a new generation of antiviral drugs due to the growing emergence of highly pathogenic influenza viruses and the resistance to existing antiviral inhibitors. Herein, we design and synthesize a set of 1,3-cis-N-substituted-1,2,3,4-tetrahydroisoquinoline derivatives to enhance their potency by further exploiting the pockets 3 and 4 in the PAN endonuclease based on the hit d,l-laudanosoline. Particularly, the lead compound 35 exhibited potent and broad anti-influenza virus effects with EC50 values ranging from 0.43 to 1.12 μM in vitro and good inhibitory activity in a mouse model. Mechanistic studies demonstrated that 35 could bind tightly to the PAN endonuclease of RNA-dependent RNA polymerase, thus blocking the viral replication to exert antiviral activity. Overall, our study might establish the importance of 1,2,3,4-tetrahydroisoquinoline-6,7-diol-based derivatives for the development of novel PAN inhibitors of influenza viruses.

Global dissemination of influenza A virus is driven by wild bird migration through arctic and subarctic zones.

Influenza A viruses (IAV) circulate endemically among many wild aquatic bird populations that seasonally migrate between wintering grounds in southern latitudes to breeding ranges along the perimeter of the circumpolar arctic. Arctic and subarctic zones are hypothesized to serve as ecologic drivers of the intercontinental movement and reassortment of IAVs due to high densities of disparate populations of long distance migratory and native bird species present during breeding seasons. Iceland is a staging ground that connects the East Atlantic and North Atlantic American flyways, providing a unique study system for characterizing viral flow between eastern and western hemispheres. Using Bayesian phylodynamic analyses, we sought to evaluate the viral connectivity of Iceland to proximal regions and how inter-species transmission and reassortment dynamics in this region influence the geographic spread of low and highly pathogenic IAVs. Findings demonstrate that IAV movement in the arctic and subarctic reflects wild bird migration around the perimeter of the circumpolar north, favouring short-distance flights between proximal regions rather than long distance flights over the polar interior. Iceland connects virus movement between mainland Europe and North America, consistent with the westward migration of wild birds from mainland Europe to Northeastern Canada and Greenland. Though virus diffusion rates were similar among avian taxonomic groups in Iceland, gulls play an outsized role as sinks of IAVs from other avian hosts prior to onward migration. These data identify patterns of virus movement in northern latitudes and inform future surveillance strategies related to seasonal and emergent IAVs with potential public health concern.

Transmission and pathogenicity of canine H3N2 influenza virus in dog and guinea pig models

BackgroundInfluenza A virus causes respiratory disease in many animal species as well as in humans. Due to the high human-animal interface, the monitoring of canine influenza in dogs and the study of the transmission and pathogenicity of canine influenza in animals are important.MethodsEight-week-old beagle dogs (Canis lupus familaris) (n = 13) were used for the intraspecies transmission model. The dogs were inoculated intranasally with 1 ml of 106 EID50 per ml of canine H3N2 influenza virus (A/canine/Thailand/CU-DC5299/2012) (CIV-H3N2). In addition, 4-week-old guinea pigs (Cavia porcellus) (n = 20) were used for the interspecies transmission model. The guinea pigs were inoculated intranasally with 300 µl of 106 EID50 per ml of CIV-H3N2.ResultsFor the Thai CIV-H3N2 challenged in the dog model, the incoculated and direct contact dogs developed respiratory signs at 2 dpi. The dogs shed the virus in the respiratory tract at 1 dpi and developed an H3-specific antibody against the virus at 10 dpi. Lung congestion and histopathological changes in the lung were observed. For the Thai CIV-H3N2 challenge in the guinea pig model, the incoculated, direct contact and aerosol-exposed guinea pigs developed fever at 1–2 dpi. The guinea pigs shed virus in the respiratory tract at 2 dpi and developed an H3-specific antibody against the virus at 7 dpi. Mild histopathological changes in the lung were observed.ConclusionThe result of this study demonstrated evidence of intraspecies and interspecies transmission of CIV-H3N2 in a mammalian model.

Temporal proteomic analyses of human lung cells distinguish high pathogenicity influenza viruses and coronaviruses from low pathogenicity viruses.

Newly re-emerging viruses are of significant global concern. In late 2019, a new coronavirus, SARS-CoV-2, emerged in China and soon spread worldwide, causing the COVID-19 pandemic, which to date has caused >6 M deaths. There has been a wealth of studies on this new virus since its emergence. The coronaviruses consist of many animal and human pathogens, with some of the human coronavirus, such as strain OC43, normally causing only mild cold-like symptoms. Viruses usurp host cellular processes to successfully replicate. We used tandem mass tag mass spectrometry-based proteomic analyses of human lung MRC-5 cells infected with OC43 for various periods of time to delineate virus-induced host cell alterations. Numerous proteins involved in lipid metabolism, molecular transport, small molecule biochemistry, cell death and survival, humoral immune response, and inflammatory response were dysregulated. Comparison of our findings to previous studies that examined a range of differentially pathogenic influenza A viruses (IAV), and to SARS-CoV-2 data, revealed that proteins involved in the cell cycle, cytokine signaling, DNA replication, and anti-inflammatory responses were generally similarly affected by virtually all tested IAV and CoV. However, proteins involved in necrosis, protein metabolism, ECM regulation, and signal transduction were generally different. In addition, the more pathogenic CoV and IAV activated Rb-dependent repression of E2F-mediated transcription, whereas less pathogenic influenza and coronaviruses either inhibited or had no effect on this pathway.

Evaluation of Efficacy of Oil Adjuvanted H5N6 Inactivated Vaccine against Highly Pathogenic H5N6 and H5N1 Influenza Viruses Infected Chickens.

Over the last 20 years, circulating highly pathogenic (HP) Asian H5 subtype avian influenza viruses have caused global pandemics in poultry and sporadic infections in humans. Vaccines are a desirable solution to prevent viral infections in poultry and reduce transmission to humans. Herein, we investigated the efficacy of an oil-adjuvanted inactivated H5N6 vaccine against highly pathogenic H5N6 and H5N1 influenza virus infections in chickens. The polybasic amino acid cleavage site depleted HA gene and NA gene of A/Waterfowl/Korea/S57/2016 (clade 2.3.4.4) (H5N6) was assembled with the rest of the A/PR/8/34 (H1N1) genes to construct the vaccine virus. The vaccine virus was propagated in fertilized eggs, partially purified using a tangential flow filtration (TFF) system, and inactivated using formalin. The chickens were intramuscularly immunized with 384 HA, 192HA, and 96HA units of oil-adjuvanted inactivated H5N6 vaccine. Antibody titer, survival rate, and lung pathology were evaluated against the homologous H5N6: A/waterfowl/Korea/S57/2016 (clade 2.3.4.4) and heterologous H5N1: A/Hong Kong/213/2003 (clade 1) viruses 12 and 4 weeks post-vaccination (p.v.), respectively. Data were statistically analyzed using the Mann-Whitney U test. The 384HA (n = 10) and 192HA (n = 5) antigen-immunized chickens showed 100% survival after lethal infections with homologous H5N6, and no virus shedding was observed from tracheal and cloacal routes. All chickens that received the 384HA vaccine survived the challenge of heterologous H5N1 after 4 weeks of immunization. The chickens that received the 384HA vaccine showed mean HI titers of 60 and 240 after 12 and 4 weeks of vaccination, respectively, against HP H5N6, whereas a mean HI titer of 80 was observed in sera collected 4 weeks after vaccination against HP H5N1. Our findings indicate that one dose of 384HA oil-adjuvanted inactivated H5N6 vaccine can induce a long-lasting immune response against both homologous H5N6 and heterologous H5N1 infections in chickens.