The influenza virus hemagglutinin molecule possesses a globular head domain that mediates receptor binding and a stalk domain at the membrane-proximal region. We generated functional influenza ...viruses expressing chimeric hemagglutinins encompassing a variety of globular head and stalk combinations, not only from different hemagglutinin subtypes but also from different hemagglutinin phylogenetic groups. These chimeric recombinant viruses possess growth properties similar to those of wild-type influenza viruses and can be used as reagents to measure domain-specific antibodies in virological and immunological assays.
Background:
Variants of concern (VOCs) of SARS-CoV-2 have caused resurging waves of infections worldwide. In the Netherlands, the Alpha, Beta, Gamma, and Delta VOCs circulated widely between ...September 2020 and August 2021. We sought to elucidate how various control measures, including targeted flight restrictions, had impacted the introduction and spread of these VOCs in the Netherlands.
Methods:
We performed phylogenetic analyses on 39,844 SARS-CoV-2 genomes collected under the Dutch national surveillance program.
Results:
We found that all four VOCs were introduced before targeted flight restrictions were imposed on countries where the VOCs first emerged. Importantly, foreign introductions, predominantly from other European countries, continued during these restrictions. After their respective introductions into the Netherlands, the Alpha and Delta VOCs largely circulated within more populous regions of the country with international connections before asymmetric bidirectional transmissions occurred with the rest of the country and the VOC became the dominant circulating lineage.
Conclusions:
Our findings show that flight restrictions had limited effectiveness in deterring VOC introductions due to the strength of regional land travel importation risks. As countries consider scaling down SARS-CoV-2 surveillance efforts in the post-crisis phase of the pandemic, our results highlight that robust surveillance in regions of early spread is important for providing timely information for variant detection and outbreak control.
Funding:
None.
After the emergence of pandemic influenza viruses in 1957, 1968, and 2009, existing seasonal viruses were observed to be replaced in the human population by the novel pandemic strains. We have ...previously hypothesized that the replacement of seasonal strains was mediated, in part, by a population-scale boost in antibodies specific for conserved regions of the hemagglutinin stalk and the viral neuraminidase. Numerous recent studies have shown the role of stalk-specific antibodies in neutralization of influenza viruses; the finding that stalk antibodies can effectively neutralize virus alters the existing dogma that influenza virus neutralization is mediated solely by antibodies that react with the globular head of the viral hemagglutinin. The present study explores the possibility that stalk-specific antibodies were boosted by infection with the 2009 H1N1 pandemic virus and that those antibodies could have contributed to the disappearance of existing seasonal H1N1 influenza virus strains. To study stalk-specific antibodies, we have developed chimeric hemagglutinin constructs that enable the measurement of antibodies that bind the hemagglutinin protein and neutralize virus but do not have hemagglutination inhibition activity. Using these chimeric hemagglutinin reagents, we show that infection with the 2009 pandemic H1N1 virus elicited a boost in titer of virus-neutralizing antibodies directed against the hemagglutinin stalk. In addition, we describe assays that can be used to measure influenza virus-neutralizing antibodies that are not detected in the traditional hemagglutination inhibition assay.
Immunity induced by vaccination and infection, referred to as hybrid immunity, provides better protection against SARS-CoV-2 infections compared to immunity induced by vaccinations alone. To assess ...the development of hybrid immunity we investigated the induction of Nucleoprotein-specific antibodies in PCR-confirmed infections by Delta or Omicron in vaccinated individuals (n = 520). Eighty-two percent of the participants with a breakthrough infection reached N-seropositivity. N-seropositivity was accompanied by Spike S1 antibody boosting, and independent of vaccination status or virus variant. Following the infection relatively more antibodies to the infecting virus variant were detected. In conclusion, these data show that hybrid immunity through breakthrough infections is hallmarked by Nucleoprotein antibodies and broadening of the Spike antibody repertoire. Exposure to future SARS-CoV-2 variants may therefore continue to maintain and broaden vaccine-induced population immunity.
Current vaccines against influenza virus infection rely on the induction of neutralizing antibodies targeting the globular head of the viral hemagglutinin (HA). Protection against seasonal antigenic ...drift or sporadic pandemic outbreaks requires further vaccine development to induce cross-protective humoral responses, potentially to the more conserved HA stalk region. Here, we present a novel viral vaccine adjuvant comprised of two synthetic ligands for Toll-like receptor 4 (TLR4) and TLR7. 1Z105 is a substituted pyrimido5,4-bindole specific for the TLR4-MD2 complex, and 1V270 is a phospholipid-conjugated TLR7 agonist. Separately, 1Z105 induces rapid Th2-associated IgG1 responses, and 1V270 potently generates Th1 cellular immunity. 1Z105 and 1V270 in combination with recombinant HA from the A/Puerto Rico/8/1934 strain (rPR/8 HA) effectively induces rapid and sustained humoral immunity that is protective against lethal challenge with a homologous virus. More importantly, immunization with the combined adjuvant and rPR/8 HA, a commercially available split vaccine, or chimeric rHA antigens significantly improves protection against both heterologous and heterosubtypic challenge viruses. Heterosubtypic protection is associated with broadly reactive antibodies to HA stalk epitopes. Histological examination and cytokine profiling reveal that intramuscular (i.m.) administration of 1Z105 and 1V270 is less reactogenic than a squalene-based adjuvant, AddaVax. In summary, the combination of 1Z105 and 1V270 with a recombinant HA induces rapid, long-lasting, and balanced Th1- and Th2-type immunity; demonstrates efficacy in a variety of murine influenza virus vaccine models assaying homologous, heterologous, and heterosubtypic challenge viruses; and has an excellent safety profile.
Novel adjuvants are needed to enhance immunogenicity and increase the protective breadth of influenza virus vaccines to reduce the seasonal disease burden and ensure pandemic preparedness. We show here that the combination of synthetic Toll-like receptor 4 (TLR4) and TLR7 ligands is a potent adjuvant for recombinant influenza virus hemagglutinin, inducing rapid and sustained immunity that is protective against influenza viruses in homologous, heterologous, and heterosubtypic challenge models. Combining TLR4 and TLR7 ligands balances Th1- and Th2-type immune responses for long-lived cellular and neutralizing humoral immunity against the viral hemagglutinin. The combined adjuvant has an attractive safety profile and the potential to augment seasonal-vaccine breadth, contribute to a broadly neutralizing universal vaccine formulation, and improve response time in an emerging pandemic.
A species barrier for the influenza A virus is the differential expression of sialic acid, which can either be α2,3-linked for avians or α2,6-linked for human viruses. The influenza A virus hosts ...also express other species-specific sialic acid derivatives. One major modification at C-5 is N-glycolyl (NeuGc), instead of N-acetyl (NeuAc). N-glycolyl is mammalian specific and expressed in pigs and horses, but not in humans, ferrets, seals, or dogs. Hemagglutinin (HA) adaptation to either N-acetyl or N-glycolyl is analyzed on a sialoside microarray containing both α2,3- and α2,6-linkage modifications on biologically relevant N-glycans. Binding studies reveal that avian, human, and equine HAs bind either N-glycolyl or N-acetyl. Structural data on N-glycolyl binding HA proteins of both H5 and H7 origin describe this specificity. Neuraminidases can cleave N-glycolyl efficiently, and tissue-binding studies reveal strict species specificity. The exclusive manner in which influenza A viruses differentiate between N-glycolyl and N-acetyl is indicative of selection.
Tick-borne encephalitis is a vaccine-preventable disease of concern for public health in large parts of Europe, with EU notification rates increasing since 2018. It is caused by the orthoflavivirus ...tick-borne encephalitis virus (TBEV) and a diagnosis of infection is mainly based on serology due to its short viremic phase, often before symptom onset. The interpretation of TBEV serology is hampered by a history of orthoflavivirus vaccination and by previous infections with related orthoflaviviruses. Here, we sought to improve TBEV sero-diagnostics using an antigen combination of in-house expressed NS1 and EDIII in a multiplex, low-specimen-volume set-up for the detection of immune responses to TBEV and other clinically important orthoflaviviruses (i.e., West Nile virus, dengue virus, Japanese encephalitis virus, Usutu virus and Zika virus). We show that the combined use of NS1 and EDIII results in both a specific and sensitive test for the detection of TBEV IgG for patient diagnostics, vaccination responses and in seroprevalence studies. This novel approach potentially allows for a low volume-based, simultaneous analysis of IgG responses to a range of orthoflaviviruses with overlapping geographic circulations and clinical manifestations.
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