scholarly journals Detailed Evolutionary Analyses of the F Gene in the Respiratory Syncytial Virus Subgroup A

Viruses ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2525
Author(s):  
Mariko Saito ◽  
Hiroyuki Tsukagoshi ◽  
Mitsuru Sada ◽  
Soyoka Sunagawa ◽  
Tatsuya Shirai ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Binding Sites ◽  
3D Structure ◽  
Neutralizing Antibody ◽  
Antibody Binding ◽  
Phylogenetic Distance ◽  
F Protein ◽  
F Gene ◽  
Conformational Epitopes ◽  
Syncytial Virus

We performed evolution, phylodynamics, and reinfection-related antigenicity analyses of respiratory syncytial virus subgroup A (RSV-A) fusion (F) gene in globally collected strains (1465 strains) using authentic bioinformatics methods. The time-scaled evolutionary tree using the Bayesian Markov chain Monte Carlo method estimated that a common ancestor of the RSV-A, RSV-B, and bovine-RSV diverged at around 450 years ago, and RSV-A and RSV-B diverged around 250 years ago. Finally, the RSV-A F gene formed eight genotypes (GA1‑GA7 and NA1) over the last 80 years. Phylodynamics of RSV-A F gene, including all genotype strains, increased twice in the 1990s and 2010s, while patterns of each RSV-A genotype were different. Phylogenetic distance analysis suggested that the genetic distances of the strains were relatively short (less than 0.05). No positive selection sites were estimated, while many negative selection sites were found. Moreover, the F protein 3D structure mapping and conformational epitope analysis implied that the conformational epitopes did not correspond to the neutralizing antibody binding sites of the F protein. These results suggested that the RSV-A F gene is relatively conserved, and mismatches between conformational epitopes and neutralizing antibody binding sites of the F protein are responsible for the virus reinfection.

scholarly journals Relationship between the loss of neutralizing antibody binding and fusion activity of the F protein of human respiratory syncytial virus

2007 ◽  
Vol 4 (1) ◽  
pp. 71 ◽  
Author(s):  
Changbao Liu ◽  
Nicole D Day ◽  
Patrick J Branigan ◽  
Lester L Gutshall ◽  
Robert T Sarisky ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibody ◽  
Antibody Binding ◽  
Human Respiratory Syncytial Virus ◽  
Fusion Activity ◽  
F Protein ◽  
Syncytial Virus

scholarly journals Characterization of Pre-F-GCN4t, a Modified Human Respiratory Syncytial Virus Fusion Protein Stabilized in a Noncleaved Prefusion Conformation

2017 ◽  
Vol 91 (13) ◽  
Author(s):  
Normand Blais ◽  
Martin Gagné ◽  
Yoshitomo Hamuro ◽  
Patrick Rheault ◽  
Martine Boyer ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Antibody Response ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Human Respiratory Syncytial Virus ◽  
Vaccine Antigen ◽  
Significant Advance ◽  
F Protein ◽  
Syncytial Virus

ABSTRACT The human respiratory syncytial virus (hRSV) fusion (F) protein is considered a major target of the neutralizing antibody response to hRSV. This glycoprotein undergoes a major structural shift from the prefusion (pre-F) to the postfusion (post-F) state at the time of virus-host cell membrane fusion. Recent evidences suggest that the pre-F state is a superior target for neutralizing antibodies compared to the post-F state. Therefore, for vaccine purposes, we have designed and characterized a recombinant hRSV F protein, called Pre-F-GCN4t, stabilized in a pre-F conformation. To show that Pre-F-GCN4t does not switch to a post-F conformation, it was compared with a recombinant post-F molecule, called Post-F-XC. Pre-F-GCN4t was glycosylated and trimeric and displayed a conformational stability different from that of Post-F-XC, as shown by chemical denaturation. Electron microscopy analysis suggested that Pre-F-GCN4t adopts a lollipop-like structure. In contrast, Post-F-XC had a typical elongated conical shape. Hydrogen/deuterium exchange mass spectrometry demonstrated that the two molecules had common rigid folding core and dynamic regions and provided structural insight for their biophysical and biochemical properties and reactivity. Pre-F-GCN4t was shown to deplete hRSV-neutralizing antibodies from human serum more efficiently than Post-F-XC. Importantly, Pre-F-GCN4t was also shown to bind D25, a highly potent monoclonal antibody specific for the pre-F conformation. In conclusion, this construct presents several pre-F characteristics, does not switch to the post-F conformation, and presents antigenic features required for a protective neutralizing antibody response. Therefore, Pre-F-GCN4t can be considered a promising candidate vaccine antigen. IMPORTANCE Human respiratory syncytial virus (RSV) is a global leading cause of infant mortality and adult morbidity. The development of a safe and efficacious RSV vaccine remains an important goal. The RSV class I fusion (F) glycoprotein is considered one of the most promising vaccine candidates, and recent evidences suggest that the prefusion (pre-F) state is a superior target for neutralizing antibodies. Our study presents the physicochemical characterization of Pre-F-GCN4t, a molecule designed to be stabilized in the pre-F conformation. To confirm its pre-F conformation, Pre-F-GCN4t was analyzed in parallel with Post-F-XC, a molecule in the post-F conformation. Our results show that Pre-F-GCN4t presents characteristics of a stabilized pre-F conformation and support its use as an RSV vaccine antigen. Such an antigen may represent a significant advance in the development of an RSV vaccine.

scholarly journals Characterization of the epitope for anti-human respiratory syncytial virus F protein monoclonal antibody 101F using synthetic peptides and genetic approaches

2007 ◽  
Vol 88 (10) ◽  
pp. 2719-2723 ◽  
Author(s):  
Sheng-Jiun Wu ◽  
Albert Schmidt ◽  
Eric J. Beil ◽  
Nicole D. Day ◽  
Patrick J. Branigan ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Antigenic Site ◽  
Neutralizing Antibody ◽  
Human Respiratory Syncytial Virus ◽  
Peptide Sequence ◽  
F Protein ◽  
Syncytial Virus ◽  
A Minor ◽  
Key Residues

Chimeric 101F (ch101F) is a mouse–human chimeric anti-human respiratory syncytial virus (HRSV) neutralizing antibody that recognizes residues within antigenic site IV, V, VI of the fusion (F) glycoprotein. The binding of ch101F to a series of peptides overlapping aa 422–438 spanning antigenic site IV, V, VI was analysed. Residues 423–436 comprise the minimal peptide sequence for ch101F binding. Substitution analysis revealed that R429 and K433 are critical for ch101F binding, whilst K427 makes a minor contribution. Binding of ch101F to a series of single mutations at positions 427, 429 and 433 in the F protein expressed recombinantly on the cell surface confirmed the peptide results. Sequence analysis of viruses selected for resistance to neutralization by ch101F indicated that a single change (K433T) in the F protein allowed ch101F escape. The results confirm that ch101F and palivizumab have different epitope specificity and define key residues for ch101F recognition.

scholarly journals Immunoprotective Activity and Safety of a RespiratorySyncytial Virus Vaccine: Mucosal Delivery of Fusion Glycoprotein with aCpG OligodeoxynucleotideAdjuvant

2003 ◽  
Vol 77 (24) ◽  
pp. 13156-13160 ◽  
Author(s):  
Gregory A. Prince ◽  
James J. Mond ◽  
David D. Porter ◽  
Kevin C. Yim ◽  
Steve J. Lan ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibody ◽  
Live Virus ◽  
Cotton Rat ◽  
F Protein ◽  
Virus Challenge ◽  
Cpg Oligodeoxynucleotides ◽  
Cotton Rats ◽  
Syncytial Virus

ABSTRACT CpG oligodeoxynucleotides (ODN) were identified that stimulated immunoglobulin production and cell proliferation in cotton rat cells in vitro. Three of these ODN were used as a mucosal adjuvant in the noses of cotton rats immunized via this route with respiratory syncytial virus fusion (F) protein. The CpG ODN markedly increased the cotton rat humoral neutralizing-antibody response to respiratory syncytial virus. Such immunized animals had a marked reduction in the production of infectious virus after a live-virus challenge. Animals immunized with the combination of F protein and CpG developed enhanced pulmonary pathology consisting of alveolitis and interstitial pneumonitis after a live-virus challenge. Similar enhanced disease has been seen in cotton rats and children immunized with formalin-inactivated respiratory syncytial virus.

scholarly journals The Optimal Concentration of Formaldehyde is Key to Stabilizing the Pre-Fusion Conformation of Respiratory Syncytial Virus Fusion Protein

Viruses ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 628 ◽  
Author(s):  
Wei Zhang ◽  
Lu-Jing Zhang ◽  
Lu-Ting Zhan ◽  
Min Zhao ◽  
Guang-Hua Wu ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Fusion Protein ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Mean Value ◽  
Optimal Concentration ◽  
F Protein ◽  
Rsv Infection ◽  
Antibody Titers ◽  
Syncytial Virus

Background: To date, there is no licensed vaccine available to prevent respiratory syncytial virus (RSV) infection. The valuable pre-fusion conformation of the fusion protein (pre-F) is prone to lose high neutralizing antigenic sites. The goals of this study were to stabilize pre-F protein by fixatives and try to find the possibility of developing an inactivated RSV vaccine. Methods: The screen of the optimal fixative condition was performed with flow cytometry. BALB/c mice were immunized intramuscularly with different immunogens. The serum neutralizing antibody titers of immunized mice were determined by neutralization assay. The protection and safety of these immunogens were assessed. Results: Fixation in an optimal concentration of formaldehyde (0.0244%–0.0977%) or paraformaldehyde (0.0625%–1%) was able to stabilize pre-F. Additionally, BALB/c mice inoculated with optimally stabilized pre-F protein (opti-fixed) induced a higher anti-RSV neutralization (9.7 log2, mean value of dilution rate) than those inoculated with unstable (unfixed, 8.91 log2, p < 0.01) or excessively fixed (exce-fixed, 7.28 log2, p < 0.01) pre-F protein. Furthermore, the opti-fixed immunogen did not induce enhanced RSV disease. Conclusions: Only the proper concentration of fixatives could stabilize pre-F and the optimal formaldehyde condition provides a potential reference for development of an inactivated RSV vaccine.

scholarly journals Murine Immune Responses to Virus-Like Particle-Associated Pre- and Postfusion Forms of the Respiratory Syncytial Virus F Protein

2015 ◽  
Vol 89 (13) ◽  
pp. 6835-6847 ◽  
Author(s):  
Lori McGinnes Cullen ◽  
Madelyn R. Schmidt ◽  
Sarah A. Kenward ◽  
Robert T. Woodland ◽  
Trudy G. Morrison
Keyword(s):  
Respiratory Syncytial Virus ◽  
Neutralizing Antibodies ◽  
Vaccine Candidate ◽  
Neutralizing Antibody ◽  
Enzyme Linked Immunosorbent Assay ◽  
Chimeric Proteins ◽  
Wild Type ◽  
F Protein ◽  
Core Proteins ◽  
Syncytial Virus

ABSTRACTVirus-like particles (VLPs) built on the Newcastle disease virus (NDV) core proteins, NP and M, and containing two chimeric proteins, F/F and H/G, composed of respiratory syncytial virus (RSV) fusion protein (F) and glycoprotein (G) ectodomains fused to the transmembrane and cytoplasmic domains of the NDV F and HN proteins, respectively, stimulate durable, protective RSV neutralizing antibodies in mice. Here, we report the properties of VLPs constructed to contain mutant RSV F protein ectodomains stabilized in prefusion (pre-F/F) or postfusion (post-F/F) configurations. The structures of the chimeric proteins assembled into VLPs were verified immunologically by their reactivities with a conformationally restricted anti-F protein monoclonal antibody. Following immunization of mice, without adjuvant, pre-F/F-containing VLPs induced significantly higher neutralizing antibody titers than the post-F/F-containing VLPs or the wild-type F/F-containing VLPs after a single immunization but not after prime and boost immunization. The specificities of anti-F IgG induced by the two mutant VLPs were assessed by enzyme-linked immunosorbent assay (ELISA) using soluble forms of the prefusion and postfusion forms of the F protein as targets. While both types of VLPs stimulated similar levels of IgG specific for the soluble postfusion F protein, titers of IgG specific for prefusion F induced by the pre-F/F-containing VLPs were higher than those induced by post-F/F-containing VLPs. Thus, VLPs containing a stabilized prefusion form of the RSV F protein represent a promising RSV vaccine candidate.IMPORTANCEThe development of vaccines for respiratory syncytial virus has been hampered by a lack of understanding of the requirements for eliciting high titers of neutralizing antibodies. The results of this study suggest that particle-associated RSV F protein containing mutations that stabilize the structure in a prefusion conformation may stimulate higher titers of protective antibodies than particles containing F protein in a wild-type or postfusion conformation. These findings indicate that the prefusion F protein assembled into VLPs has the potential to produce a successful RSV vaccine candidate.

scholarly journals Sites responsible for infectivity and antigenicity on nervous necrosis virus (NNV) appear to be distinct

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Hyun Jung Gye ◽  
Toyohiko Nishizawa
Keyword(s):  
Receptor Binding ◽  
Binding Sites ◽  
Conformational Stability ◽  
Neutralizing Antibody ◽  
Cell Receptor ◽  
Antibody Binding ◽  
Nervous Necrosis Virus ◽  
Necrosis Virus ◽  
Conformational Epitopes ◽  
Rate Of Decline

AbstractNervous necrosis virus (NNV) is a pathogenic fish-virus belonging to the genus Betanodavirus (Nodaviridae). Surface protrusions on NNV particles play a crucial role in both antigenicity and infectivity. We exposed purified NNV particles to different physicochemical conditions to investigate the effects on antigenicity and infectivity, in order to reveal information regarding the conformational stability and spatial relationships of NNV neutralizing-antibody binding sites and cell receptor binding sites. Treatment with PBS at 37 °C, drastically reduced NNV antigenicity by 66–79% on day one, whereas its infectivity declined gradually from 107.6 to 105.8 TCID50/ml over 10 days. When NNV was treated with carbonate/bicarbonate buffers at different pHs, both antigenicity and infectivity of NNV declined due to higher pH. However, the rate of decline with respect to antigenicity was more moderate than for infectivity. NNV antigenicity declined 75–84% after treatment with 2.0 M urea, however, there was no reduction observed in infectivity. The antibodies used in antigenicity experiments have high NNV-neutralizing titers and recognize conformational epitopes on surface protrusions. The maintenance of NNV infectivity means that receptor binding sites are functionally preserved. Therefore, it seems highly likely that NNV neutralizing-antibody binding sites and receptor binding sites are independently located on surface protrusions.

scholarly journals A Chimeric A2 Strain of Respiratory Syncytial Virus (RSV) with the Fusion Protein of RSV Strain Line 19 Exhibits Enhanced Viral Load, Mucus, and Airway Dysfunction

2009 ◽  
Vol 83 (9) ◽  
pp. 4185-4194 ◽  
Author(s):  
Martin L. Moore ◽  
Michael H. Chi ◽  
Cindy Luongo ◽  
Nicholas W. Lukacs ◽  
Vasiliy V. Polosukhin ◽  
...  
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Airway Hyperresponsiveness ◽  
Laboratory Strain ◽  
F Protein ◽  
F Gene ◽  
Mucin Expression ◽  
Rsv Infection ◽  
Syncytial Virus ◽  
Airway Mucin

ABSTRACT Respiratory syncytial virus (RSV) is the leading cause of respiratory failure and viral death in infants. Abundant airway mucus contributes to airway obstruction in RSV disease. Interleukin-13 (IL-13) is a mediator of pulmonary mucus secretion. It has been shown that infection of BALB/c mice with the RSV line 19 strain but not with the RSV A2 laboratory strain results in lung IL-13 and mucus expression. Here, we sequenced the RSV line 19 genome and compared it to the commonly used A2 and Long strains. There were six amino acid differences between the line 19 strain and both the A2 and Long RSV strains, five of which are in the fusion (F) protein. The Long strain, like the A2 strain, did not induce lung IL-13 and mucus expression in BALB/c mice. We hypothesized that the F protein of RSV line 19 is more mucogenic than the F proteins of A2 and Long. We generated recombinant, F-chimeric RSVs by replacing the F gene of A2 with the F gene of either line 19 or Long. Infection of BALB/c mice with RSV rA2 line 19F resulted in lower alpha interferon lung levels 24 h postinfection, higher lung viral load, higher lung IL-13 levels, greater airway mucin expression levels, and greater airway hyperresponsiveness than infection with rA2-A2F or rA2-LongF. We identified the F protein of RSV line 19 as a factor that plays a role in pulmonary mucin expression in the setting of RSV infection.

scholarly journals Alternative Virus-Like Particle-Associated Prefusion F Proteins as Maternal Vaccines for Respiratory Syncytial Virus

2019 ◽  
Vol 93 (23) ◽  
Author(s):  
Jorge C. G. Blanco ◽  
Lurds R. Fernando ◽  
Wei Zhang ◽  
Arash Kamali ◽  
Marina S. Boukhvalova ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Young Children ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
F Protein ◽  
Maternal Immunization ◽  
Maternal Vaccination ◽  
Virus Like Particle ◽  
Cotton Rats ◽  
Syncytial Virus

ABSTRACT Maternal vaccination may be the most effective and safest approach to the protection of infants from respiratory syncytial virus (RSV) infection, a severe acute lower respiratory tract disease in infants and young children worldwide. We previously compared five different virus-like particle (VLP)-associated, mutation-stabilized prefusion F (pre-F) proteins, including the prototype DS-Cav1 F VLPs. We showed that alternative versions of prefusion F proteins have different conformations and induce different populations of anti-F protein antibodies. Two of these alternative pre-F VLPs, the UC-2 F and UC-3 F VLPs, stimulated in mice higher titers of neutralizing antibodies than DS-Cav1 F VLPs (M. L. Cullen, R. M. Schmidt, M. G. Torres, A. A. Capoferri, et al., Vaccines 7:21–41, 2019, https://doi.org/10.3390/vaccines7010021). Here we describe a comparison of these two pre-F VLPs with DS-Cav1 F VLPs as maternal vaccines in cotton rats and report that UC-3 F VLPs significantly increased the neutralizing antibody (NAb) titers in pregnant dams compared to DS-Cav1 F VLPs. The neutralizing antibody titers in the sera of the offspring of the dams immunized with UC-3 F VLPs were significantly higher than those in the sera of the offspring of dams immunized with DS-Cav1 VLPs. This increase in serum NAb titers translated to a 6- to 40-fold lower virus titer in the lungs of the RSV-challenged offspring of dams immunized with UC-3 F VLPs than in the lungs of the RSV-challenged offspring of dams immunized with DS-Cav1 F VLPs. Importantly, the offspring of UC-3 F VLP-immunized dams showed significant protection from lung pathology and from induction of inflammatory lung cytokine mRNA expression after RSV challenge. Immunization with UC-3 F VLPs also induced durable levels of high-titer neutralizing antibodies in dams. IMPORTANCE Respiratory syncytial virus (RSV) is a significant human pathogen severely impacting neonates and young children, but no vaccine exists to protect this vulnerable population. Furthermore, direct vaccination of neonates is likely ineffective due to the immaturity of their immune system, and neonate immunization is potentially unsafe. Maternal vaccination may be the best and safest approach to the protection of neonates through the passive transfer of maternal neutralizing antibodies in utero to the fetus after maternal immunization. Here we report that immunization of pregnant cotton rats, a surrogate model for human maternal immunization, with novel RSV virus-like particle (VLP) vaccine candidates containing stabilized prefusion RSV F proteins provides significant levels of protection of the offspring of immunized dams from RSV challenge. We also found that antibodies induced by VLPs containing different versions of the prefusion F protein varied by 40-fold in the extent of protection provided to the offspring of vaccinated dams upon RSV challenge.

scholarly journals Protective Efficacy and Immunogenicity of an Adenoviral Vector Vaccine Encoding the Codon-Optimized F Protein of Respiratory Syncytial Virus

2009 ◽  
Vol 83 (23) ◽  
pp. 12601-12610 ◽  
Author(s):  
Rebekka Kohlmann ◽  
Sarah Schwannecke ◽  
Bettina Tippler ◽  
Nicola Ternette ◽  
Vladimir V. Temchura ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Vaccine Development ◽  
Protective Efficacy ◽  
Neutralizing Antibody ◽  
Soluble Form ◽  
Booster Immunization ◽  
Vector Vaccine ◽  
F Gene ◽  
Viral Loads ◽  
Syncytial Virus

ABSTRACT Adenoviral vectors (AdV) have received considerable attention for vaccine development because of their high immunogenicity and efficacy. In previous studies, it was shown that DNA immunization of mice with codon-optimized expression plasmids encoding the fusion protein of respiratory syncytial virus (RSV F) resulted in enhanced protection against RSV challenge compared to immunization with plasmids carrying the wild-type cDNA sequence of RSV F. In this study, we constructed AdV carrying the codon-optimized full-length RSV F gene (AdV-F) or the soluble form of the RSV F gene (AdV-Fsol). BALB/c mice were immunized twice with AdV-F or AdV-Fsol and challenged with RSV intranasally. Substantial levels of antibody to RSV F were induced by both AdV vaccines, with peak neutralizing-antibody titers of 1:900. Consistently, the viral loads in lung homogenates and bronchoalveolar lavage fluids were significantly reduced by a factor of more than 60,000. The protection against viral challenge could be measured even 8 months after the booster immunization. AdV-F and AdV-Fsol induced similar levels of immunogenicity and protective efficacy. Therefore, these results encourage further development of AdV vaccines against RSV infection in humans.

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