The use of human intestinal enteroid cell cultures for detection of multiple gastroenteric viruses.

Human norovirus (HuNoV) and human astrovirus (HAstV) are viral enteric pathogens and known causative agents of acute gastroenteritis. Identifying the presence of these viruses in environmental samples such as irrigation water, or foods exposed to virus contaminated water (e.g.

Coinfection of Influenza A and B and Human OC43 Coronavirus in Normal Human Bronchial Epithelial Cells.

Background: Influenza viruses and seasonal coronaviruses are pathogens transmitted via an airborne route that can cause respiratory diseases in humans that have similar symptoms such as fever, cough, and pneumonia. These two viruses can infect similar human tissues, such as the respiratory tract and nasal, bronchial, and alveolar epithelial cells. Influenza virus and seasonal coronavirus coinfections are poorly understood.

Genomic Analysis of Influenza A and B Viruses Carrying Baloxavir Resistance-Associated Substitutions Serially Passaged in Human Epithelial Cells.

Baloxavir marboxil (baloxavir) is an FDA-approved inhibitor of the influenza virus polymerase acidic (PA) protein. Here, we used next-generation sequencing to compare the genomic mutational profiles of IAV H1N1 and H3N2, and IBV wild type (WT) and mutants (MUT) viruses carrying baloxavir resistance-associated substitutions (H1N1-PA I38L, I38T, and E199D; H3N2-PA I38T; and IBV-PA I38T) during passaging in normal human bronchial epithelial (NHBE) cells. We determined the ratio of nonsynonymous to synonymous nucleotide mutations (d/d) and identified the location and type of amino acid (AA) substitutions that occurred at a frequency of ≥30%.

Interchangeability of the Assays Used to Assess the Activity of Anti-SARS-CoV-2 Monoclonal Antibodies.

The recent global COVID-19 pandemic caused by SARS-CoV-2 lasted for over three years. A key measure in combatting this pandemic involved the measurement of the monoclonal antibody (mAb)-mediated inhibition of binding between the spike receptor-binding domain (RBD) and hACE2 receptor. Potency assessments of therapeutic anti-SARS-CoV-2 mAbs typically include binding or cell-based neutralization assays.

Impact of Baloxavir Resistance-Associated Substitutions on Influenza Virus Growth and Drug Susceptibility.

Baloxavir marboxil (baloxavir) is a recently FDA-approved influenza virus polymerase acidic (PA) endonuclease inhibitor. Several PA substitutions have been demonstrated to confer reduced susceptibility to baloxavir; however, their impacts on measurements of antiviral drug susceptibility and replication capacity when present as a fraction of the viral population have not been established. We generated recombinant A/California/04/09 (H1N1)-like viruses (IAV) with PA I38L, I38T, or E199D substitutions and B/Victoria/504/2000-like virus (IBV) with PA I38T.

Comparison of the Antiviral Activity of Remdesivir, Chloroquine, and Interferon-β as Single or Dual Agents Against the Human Beta-Coronavirus OC43.

The human beta-coronavirus strain, OC43, provides a useful model for testing the antiviral activity of various agents. We compared the activity of several antiviral drugs against OC43, including remdesivir, chloroquine, interferon (IFN)-β, IFN-λ1, and IFN-λ4, in two distinct cell types: human colorectal carcinoma cell line (HCT-8 cells) and normal human bronchial epithelial (NHBE) cells. We also tested whether these agents mediate additive, synergistic, or antagonistic activity against OC43 infection when used in combination.

Generation and characterization of interferon-beta-resistant H1N1 influenza A virus.

Interferons (IFNs) mediate innate antiviral activity against many types of viruses, including influenza viruses. In light of their potential use as anti-influenza agents, we examined whether resistance to these host antiviral proteins can develop. We generated IFN-β-resistant variants of the A/California/04/09 (H1N1) virus by serial passage in a human airway epithelial cell line, Calu-3, under IFN-β selective pressure.

Adaptation of influenza B virus by serial passage in human airway epithelial cells.

Influenza B viruses cause seasonal epidemics and are a considerable burden to public health. To understand their adaptation capability, we examined the genetic changes that occurred following 15 serial passages of two influenza B viruses, B/Brisbane/60/2008 and B/Victoria/504/2000, in human epithelial cells. Thirteen distinct amino acid mutations were found in the PB1, PA, hemagglutinin (HA), neuraminidase (NA), and M proteins after serial passage in the human lung epithelial cell line, Calu-3, and normal human bronchial epithelial (NHBE) cells.

Laninamivir-Interferon Lambda 1 Combination Treatment Promotes Resistance by Influenza A Virus More Rapidly than Laninamivir Alone.

Each year, 5% to 20% of the population of the United States becomes infected with influenza A virus. Combination therapy with two or more antiviral agents has been considered a potential treatment option for influenza virus infection. However, the clinical results derived from combination treatment with two or more antiviral drugs have been variable.

Influenza A virus hemagglutinin mutations associated with use of neuraminidase inhibitors correlate with decreased inhibition by anti-influenza antibodies.

Background: Vaccination and the use of neuraminidase inhibitors (NAIs) are currently the front lines of defense against seasonal influenza. The activity of influenza vaccines and antivirals drugs such as the NAIs can be affected by mutations in the influenza hemagglutinin (HA) protein. Numerous HA substitutions have been identified in nonclinical NAI resistance-selection experiments as well as in clinical specimens from NAI treatment or surveillance studies.

Influenza H1 Mosaic Hemagglutinin Vaccine Induces Broad Immunity and Protection in Mice.

Annually, influenza A virus (IAV) infects ~5-10% of adults and 20-30% of children worldwide. The primary resource to protect against infection is by vaccination. However, vaccination only induces strain-specific and transient immunity.

Effect of influenza H1N1 neuraminidase V116A and I117V mutations on NA activity and sensitivity to NA inhibitors.

Neuraminidase inhibitors (NAIs) play a key role in the management of influenza. Given the limited number of FDA-approved anti-influenza drugs, evaluation of potential drug-resistant variants is of high priority. Two NA mutations, V116A and I117V, are found in ∼0.

Identification and quantification of defective virus genomes in high throughput sequencing data using DVG-profiler, a novel post-sequence alignment processing algorithm.

Most viruses are known to spontaneously generate defective viral genomes (DVG) due to errors during replication. These DVGs are subgenomic and contain deletions that render them unable to complete a full replication cycle in the absence of a co-infecting, non-defective helper virus. DVGs, especially of the copyback type, frequently observed with paramyxoviruses, have been recognized to be important triggers of the antiviral innate immune response.

A comparison of interferon gene expression induced by influenza A virus infection of human airway epithelial cells from two different donors.

Influenza is an acute respiratory disease that can cause local annual epidemics and worldwide pandemics of different morbidity and mortality. Our understanding of host factors that modulate the frequency and severity of influenza virus infections is less than complete. In this study, we examined the inter-individual variations in the innate immune responses to H1N1 and H3N2 influenza A viruses (IAV) using primary cultures of normal human bronchial epithelial (NHBE) cells derived from two different donors (D1 and D2).

Immunogenicity and Viral Shedding of Russian-Backbone, Seasonal, Trivalent, Live, Attenuated Influenza Vaccine in a Phase II, Randomized, Placebo-Controlled Trial Among Preschool-Aged Children in Urban Bangladesh.

Background: We evaluated a Russian-backbone, live, attenuated influenza vaccine (LAIV) for immunogenicity and viral shedding in a randomized, placebo-controlled trial among Bangladeshi children.

Methods: Healthy children received a single, intranasal dose of LAIV containing the 2011-2012 recommended formulation or placebo. Nasopharyngeal wash (NPW) specimens were collected on days 0, 2, 4, and 7.

Anti-neuraminidase antibodies against pandemic A/H1N1 influenza viruses in healthy and influenza-infected individuals.

The main objective of the study was to evaluate neuraminidase inhibiting (NI) antibodies against A/H1N1pdm09 influenza viruses in the community as a whole and after infection. We evaluated NI serum antibodies against A/California/07/09(H1N1)pdm and A/South Africa/3626/2013(H1N1)pdm in 134 blood donors of different ages using enzyme-linked lectin assay and in 15 paired sera from convalescents with laboratory confirmed influenza. The neuraminidase (NA) proteins of both A/H1N1pdm09 viruses had minimal genetic divergence, but demonstrated different enzymatic and antigenic properties.

The use of plant lectins to regulate H1N1 influenza A virus receptor binding activity.

We applied an in vitro selection approach using two different plant lectins that bind to α2,3- or α2,6-linked sialic acids to determine which genetic changes of the A/California/04/09 (H1N1) virus alter hemagglutinin (HA) receptor binding toward α2,3- or α2,6-linked glycans. Consecutive passages of the A/California/04/09 virus with or without lectins in human lung epithelial Calu-3 cells led to development of three HA1 amino acid substitutions, N129D, G155E, and S183P, and one mutation in the neuraminidase (NA), G201E. The S183P mutation significantly increased binding to several α2,6 SA-linked glycans, including YDS, 6'SL(N), and 6-Su-6'SLN, compared to the wild-type virus (↑3.

Rapid one-step biotinylation of biological and non-biological surfaces.

We describe a rapid one-step method to biotinylate virtually any biological or non-biological surface. Contacting a solution of biotin-spacer-lipid constructs with a surface will form a coating within seconds on non-biological surfaces or within minutes on most biological membranes including membrane viruses. The resultant biotinylated surface can then be used to interact with avidinylated conjugates, beads, vesicles, surfaces or cells.

Influenza virus NS1 protein mutations at position 171 impact innate interferon responses by respiratory epithelial cells.

The influenza virus NS1 protein interacts with a wide range of proteins to suppress the host cell immune response and facilitate virus replication. The amino acid sequence of the 2009 pandemic virus NS1 protein differed from sequences of earlier related viruses. The functional impact of these differences has not been fully defined.

Generation and characterization of interferon-lambda 1-resistant H1N1 influenza A viruses.

Influenza A viruses pose a constant potential threat to human health. In view of the innate antiviral activity of interferons (IFNs) and their potential use as anti-influenza agents, it is important to know whether viral resistance to these antiviral proteins can arise. To examine the likelihood of emergence of IFN-λ1-resistant H1N1 variants, we serially passaged the A/California/04/09 (H1N1) strain in a human lung epithelial cell line (Calu-3) in the presence of increasing concentrations of recombinant IFN-λ1 protein.

Impact of Influenza A Virus Infection on the Proteomes of Human Bronchoepithelial Cells from Different Donors.

Susceptibility to influenza A virus is determined by a balance of viral and host factors. The genetic background of the host contributes to the severity of disease, but the influenza-related proteomes of cells from different individuals have not been compared. We used high-resolution mass spectrometry to identify proteins in normal human bronchial epithelial (NHBE) cells isolated from three different donors.

Amino Acids in Hemagglutinin Antigenic Site B Determine Antigenic and Receptor Binding Differences between A(H3N2)v and Ancestral Seasonal H3N2 Influenza Viruses.

Unlabelled: Influenza A H3N2 variant [A(H3N2)v] viruses, which have caused human infections in the United States in recent years, originated from human seasonal H3N2 viruses that were introduced into North American swine in the mid-1990s, but they are antigenically distinct from both the ancestral and current circulating H3N2 strains. A reference A(H3N2)v virus, A/Minnesota/11/2010 (MN/10), and a seasonal H3N2 strain, A/Beijing/32/1992 (BJ/92), were chosen to determine the molecular basis for the antigenic difference between A(H3N2)v and the ancestral viruses. Viruses containing wild-type and mutant MN/10 or BJ/92 hemagglutinins (HAs) were constructed and probed for reactivity with ferret antisera against MN/10 and BJ/92 in hemagglutination inhibition assays.

Effects of hemagglutinin amino acid substitutions in H9 influenza A virus escape mutants.

We assessed the pH optimum of fusion, HA thermostability, and in vitro replication kinetics of previously obtained influenza H9 escape mutants. The N198S mutation significantly increased the optimum pH of fusion. Four HA changes, S133N, T189A, N198D, and L226Q, were associated with a significant increase in HA thermostability compared to the wild-type virus.