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During the Omicron wave, previous variants such as BA.2, BA.4, and BA.5 were replaced by newer variants with additional mutations in the spike protein. These variants, BA.4.6, BQ.1.1, and XBB, have spread in different countries with different degrees of success. Here, we evaluated the replicative ability and pathogenicity of BA.4.6, BQ1.1, and XBB clinical isolates in male Syrian hamsters. Although we found no substantial differences in weight change among hamsters infected with these Omicron subvariants, the replicative ability of BQ.1.1 and XBB in lung tissue was higher than that of BA.4.6 and BA.5. Of note, BQ.1.1 was lethal in both male and female transgenic human ACE2 hamsters. In competition assays, XBB replicated better than BQ.1.1 in the nasal turbinate tissues of female hamsters previously infected with Omicron BA.2. These results suggest that newer Omicron subvariants in the XBB family are still evolving and should be closely monitored. Omicron variants BQ.1.1 and XBB increased pathogenicity in wild-type hamsters, with BQ.1.1 showing higher mortality in human ACE2 transgenic hamsters compared to earlier Omicron variants.. [ABSTRACT FROM AUTHOR]

Prolonged treatment of immunocompromised influenza patients with viral neuraminidase (NA) inhibitors is required, because the immune system of such patients fails to eradicate the viruses. Here, we attempted to eradicate influenza virus from the respiratory organs of nude mice, which is a model of immunocompromised hosts, by using combination therapy of the viral polymerase inhibitor favipiravir and monoclonal antibodies (mAbs) against the receptor-binding site (RBS) and stem of viral hemagglutinin (HA). Although monotherapy or combination therapy of two antivirals (two mAbs or favipiravir plus a mAb) suppressed virus replication, they failed to eradicate viruses from nude mice. In contrast, the triple combination therapy of favipiravir plus anti-Stem and anti-RBS mAbs completely stopped virus replication in nude mice, resulting in virus clearance. Triple combination approaches should be considered for the treatment of human immunocompromised patients with severe influenza. [ABSTRACT FROM AUTHOR]

Coronavirus disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) remains a global pandemic. Although several vaccines targeting SARS-CoV-2 spike proteins protect against COVID-19 infection, mutations affecting virus transmissibility and immune evasion potential have reduced their efficacy, leading to the need for a more efficient strategy. Available clinical evidence regarding COVID-19 suggests that endothelial dysfunction with thrombosis is a central pathogenesis of progression to systemic disease, in which overexpression of plasminogen activator inhibitor-1 (PAI-1) may be important. Here we developed a novel peptide vaccine against PAI-1 and evaluated its effect on lipopolysaccharide (LPS)-induced sepsis and SARS-CoV-2 infection in mice. Administration of LPS and mouse-adapted SARS-CoV-2 increased serum PAI-1 levels, although the latter showed smaller levels. In an LPS-induced sepsis model, mice immunized with PAI-1 vaccine showed reduced organ damage and microvascular thrombosis and improved survival compared with vehicle-treated mice. In plasma clot lysis assays, vaccination-induced serum IgG antibodies were fibrinolytic. However, in a SARS-CoV-2 infection model, survival and symptom severity (i.e., body weight reduction) did not differ between vaccine- and vehicle-treated groups. These results indicate that although PAI-1 may promote the severity of sepsis by increasing thrombus formation, it might not be a major contributor to COVID-19 exacerbation. [ABSTRACT FROM AUTHOR]

The Madin-Darby canine kidney (MDCK) cell line is an in vitro model for influenza A virus (IAV) infection and propagation. MDCK-SIAT1 (SIAT1) and humanized MDCK (hCK) cell lines are engineered MDCK cells that express N-glycans with elevated levels of sialic acid (Sia) in α2,6-linkage (α2,6-Sia) that are recognized by many human IAVs. To characterize the N-glycan structures in these cells and the potential changes compared to the parental MDCK cell line resulting from engineering, we analyzed the N-glycans from these cells at different passages, using both mass spectrometry and specific lectin and antibody binding. We observed significant differences between the three cell lines in overall complex N-glycans and terminal galactose modifications. MDCK cells express core fucosylated, bisected complex-type N-glycans at all passage stages, in addition to expressing α2,6-Sia on short N-glycans and α2,3-Sia on larger N-glycans. By contrast, SIAT1 cells predominantly express α2,6-Sia glycans and greatly reduced level of α2,3-Sia glycans. Additionally, they express bisected, sialylated N-glycans that are scant in MDCK cells. The hCK cells exclusively express α2,6-Sia glycans. Unexpectedly, hCK glycoproteins bound robustly to the plant lectin MAL-1, indicating α2,3-Sia glycans, but such binding was not Sia-dependent and closely mirrored that of an antibody that recognizes glycans with terminal 3-O-sulfate galactose (3-O-SGal). The 3-O-SGal epitope is highly expressed in N-glycans on multiple hCK glycoproteins. These results indicate vastly different N-glycomes between MDCK cells and the engineered clones that could relate to IAV infectivity. [ABSTRACT FROM AUTHOR]

Influenza A viral ribonucleoprotein (vRNP) complexes comprise the eight genomic negative-sense RNAs, each of which is bound to multiple copies of the vRNP and a trimeric viral polymerase complex. The influenza virus life cycle centres on the vRNPs, which in turn rely on host cellular processes to carry out functions that are necessary for the successful completion of the virus life cycle. In this Review, we discuss our current knowledge about vRNP trafficking within host cells and the function of these complexes in the context of the virus life cycle, highlighting how structure contributes to function and the crucial interactions with host cell pathways, as well as on the information gaps that remain. An improved understanding of how vRNPs use host cell pathways is essential to identify mechanisms of virus pathogenicity, host adaptation and, ultimately, new targets for antiviral intervention. [ABSTRACT FROM AUTHOR]

The COVID-19 pandemic continues to wreak havoc as worldwide SARS-CoV-2 infection, hospitalization, and death rates climb unabated. Effective vaccines remain the most promising approach to counter SARS-CoV-2. Yet, while promising results are emerging from COVID-19 vaccine trials, the need for multiple doses and the challenges associated with the widespread distribution and administration of vaccines remain concerns. Here, we engineered the coat protein of the MS2 bacteriophage and generated nanoparticles displaying multiple copies of the SARS-CoV-2 spike (S) protein. The use of these nanoparticles as vaccines generated high neutralizing antibody titers and protected Syrian hamsters from a challenge with SARS-CoV-2 after a single immunization with no infectious virus detected in the lungs. This nanoparticle-based vaccine platform thus provides protection after a single immunization and may be broadly applicable for protecting against SARS-CoV-2 and future pathogens with pandemic potential. Chiba et al. describe the use of MS2 bacteriophage coat proteins to develop nanocarriers that display the SARS-CoV-2 spike proteins multivalently for vaccine applications. The vaccine elicited high neutralizing antibody titers and protected Syrian hamsters from virus infection after a single immunization. [ABSTRACT FROM AUTHOR]

Equine influenza virus is an important pathogen for the horse industry because of its economic impact, and vaccination is a key control measure. Our previous work suggested that a mutation at position 144 in the hemagglutinin of Florida sublineage clade 2 viruses reduces the cross-neutralizing activity of antiserum against a former vaccine strain. To confirm this suggestion, here, we generated viruses by reverse genetics. Antibody titers against the mutated viruses were one-tenth to one-sixteenth of those against the former vaccine strain. Our findings confirm that this single amino acid substitution reduces the cross-reactivity of antiserum against this former Japanese vaccine. [ABSTRACT FROM AUTHOR]

The domestic ferret (Mustela putorius furo) is an important animal model for multiple human respiratory diseases. It is considered the 'gold standard' for modeling human influenza virus infection and transmission. Here we describe the 2.41 Gb draft genome assembly of the domestic ferret, constituting 2.28 Gb of sequence plus gaps. We annotated 19,910 protein-coding genes on this assembly using RNA-seq data from 21 ferret tissues. We characterized the ferret host response to two influenza virus infections by RNA-seq analysis of 42 ferret samples from influenza time-course data and showed distinct signatures in ferret trachea and lung tissues specific to 1918 or 2009 human pandemic influenza virus infections. Using microarray data from 16 ferret samples reflecting cystic fibrosis disease progression, we showed that transcriptional changes in the CFTR-knockout ferret lung reflect pathways of early disease that cannot be readily studied in human infants with cystic fibrosis disease. [ABSTRACT FROM AUTHOR]

There is large demand for a quantitative method for rapid and ultra-sensitive detection of the influenza virus. Here, we established a digital influenza virus counting (DIViC) method that can detect a single virion without antibody. In the assay, a virion is stochastically entrapped inside a femtoliter reactor array device for the fluorogenic assay of neuraminidase, and incubated for minutes. By analyzing 600,000 reactors, the practical limit of detection reached the order of 103 (PFU)/mL, only 10-times less sensitive than RT-PCR and more than 1000-times sensitive than commercial rapid test kits (RIDTs). Interestingly, neuraminidase activity differed among virions. The coefficient of variance was 30-40%, evidently broader than that of alkaline phosphatase measured as a model enzyme for comparison, suggesting the heterogeneity in size and integrity among influenza virus particles. Sensitivity to oseltamivir also differed between virions. We also tested DIViC using clinical gargle samples that imposes less burden for sampling while with less virus titre. The comparison with RIDTs showed that DIViC was largely superior to RIDTs in the sensitivity with the clinical samples although a few false-positive signals were observed in some clinical samples that remains as a technical challenge. [ABSTRACT FROM AUTHOR]

We tested the biological significance of two amino acid mutations in the PB2 protein (glutamic acid to lysine at position 627 and aspartic acid to asparagine at position 701) of A(H7N9) viruses for mammalian adaptation. Mutants were assessed for their viral polymerase activities, growth kinetics in mammalian and avian cells, and pathogenicity in mice. We found that lysine at position 627 and asparagine at position 701 in PB2 are essential for mammalian adaptation of A(H7N9) viruses. [ABSTRACT FROM AUTHOR]