scholarly journals Identification of Residues in the Human Respiratory Syncytial Virus Fusion Protein That Modulate Fusion Activity and Pathogenesis

2014 ◽  
Vol 89 (1) ◽  
pp. 512-522 ◽  
Author(s):  
Anne L. Hotard ◽  
Sujin Lee ◽  
Michael G. Currier ◽  
James E. Crowe ◽  
Kaori Sakamoto ◽  
...  
Keyword(s):  
Amino Acids ◽  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Lower Respiratory Tract ◽  
Human Respiratory Syncytial Virus ◽  
Fusion Activity ◽  
Viral Loads ◽  
Mucin Expression ◽  
Syncytial Virus ◽  
Airway Mucin

ABSTRACTHuman respiratory syncytial virus (RSV) lower respiratory tract infection can result in inflammation and mucus plugging of airways. RSV strain A2-line19F induces relatively high viral load and mucus in mice. The line 19 fusion (F) protein harbors five unique residues compared to the non-mucus-inducing strains A2 and Long, at positions 79, 191, 357, 371, and 557. We hypothesized that differential fusion activity is a determinant of pathogenesis. In a cell-cell fusion assay, line 19 F was more fusogenic than Long F. We changed the residues unique to line 19 F to the corresponding residues in Long F and identified residues 79 and 191 together as responsible for high fusion activity. Surprisingly, mutation of residues 357 or 357 with 371 resulted in gain of fusion activity. Thus, we generated RSV F mutants with a range of defined fusion activity and engineered these into recombinant viruses. We found a clear, positive correlation between fusion activity and early viral load in mice; however, we did not detect a correlation between viral loads and levels of airway mucin expression. The F mutant with the highest fusion activity, A2-line19F-K357T/Y371N, induced high viral loads, severe lung histopathology, and weight loss but did not induce high levels of airway mucin expression. We defined residues 79/191 as critical for line 19 F fusion activity and 357/371 as playing a role in A2-line19F mucus induction. Defining the molecular basis of the role of RSV F in pathogenesis may aid vaccine and therapeutic strategies aimed at this protein.IMPORTANCEHuman respiratory syncytial virus (RSV) is the most important lower respiratory tract pathogen of infants for which there is no vaccine. Elucidating mechanisms of RSV pathogenesis is important for rational vaccine and drug design. We defined specific amino acids in the fusion (F) protein of RSV strain line 19 critical for fusion activity and elucidated a correlation between fusion activity and viral load in mice. Further, we identified two distinct amino acids in F as contributing to the mucogenic phenotype of the A2-line19F virus. Taken together, these results illustrate a role for RSV F in virulence.

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 High Resolution Analysis of Respiratory Syncytial Virus Infection In Vivo

Viruses ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 926 ◽  
Author(s):  
Waleed Aljabr ◽  
Stuart Armstrong ◽  
Natasha Y. Rickett ◽  
Georgios Pollakis ◽  
Olivier Touzelet ◽  
...  
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
The Elderly ◽  
High Viral Load ◽  
Label Free ◽  
Respiratory Problems ◽  
Viral Loads ◽  
High Resolution Analysis ◽  
Syncytial Virus

Human respiratory syncytial virus (HRSV) is a major cause of pediatric infection and also causes disease in the elderly and those with underlying respiratory problems. There is no vaccine for HRSV and anti-viral therapeutics are not broadly applicable. To investigate the effect of HRSV biology in children, nasopharyngeal aspirates were taken from children with different viral loads and a combined high throughput RNAseq and label free quantitative proteomics approach was used to characterize the nucleic acid and proteins in these samples. HRSV proteins were identified in the nasopharyngeal aspirates from infected children, and their abundance correlated with viral load (Ct value), confirming HRSV infection. Analysis of the HRSV genome indicated that the children were infected with sub-group A virus and that minor variants in nucleotide frequency occurred in discrete clusters along the HRSV genome, and within a patient clustered distinctly within the glycoprotein gene. Data from the samples were binned into four groups; no-HRSV infection (control), high viral load (Ct < 20), medium viral load (Ct = 20–25), and low viral load (Ct > 25). Cellular proteins associated with the anti-viral response (e.g., ISG15) were identified in the nasopharyngeal aspirates and their abundance was correlated with viral load. These combined approaches have not been used before to study HRSV biology in vivo and can be readily applied to the study the variation of virus host interactions.

scholarly journals A41 Deep sequencing of respiratory syncytial virus links viral diversity to disease severity

2019 ◽  
Vol 5 (Supplement_1) ◽  
Author(s):  
Inne Nauwelaers ◽  
Tiina Talts ◽  
Monica Galiano ◽  
Peter Openshaw
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Disease Severity ◽  
Illumina Sequencing ◽  
Likelihood Method ◽  
Evolutionary Analysis ◽  
Viral Loads ◽  
Ct Value ◽  
A Genome ◽  
Syncytial Virus

Abstract Respiratory syncytial virus (RSV) is a common virus that can cause bronchiolitis in infants and pneumonia in immunocompromised and elderly people. RSV belongs to the Pneumoviridae family and consists of a genome of 15 kb. Its genome contains ten genes that code for eleven proteins, with M2 coding for two different proteins in overlapping open reading frames. It is unclear why some infected children have severe disease and others have mild or asymptomatic disease. In this project, methods for complete genome sequencing of RSV via Sanger and Illumina MiSeq platforms were optimized. One hundred and twenty-four community samples (59 RSV A and 65 RSV B) from 2014 to 2018 were collected (in collaboration with the Royal College of General Practitioners) and sequenced. Samples were selected based on viral load (e.g. Ct values had to be < 30). The genotype of each sample was determined by constructing phylogenetic trees with reference sequences from all genotypes. Trees were reconstructed using the maximum likelihood method. Furthermore, Illumina sequencing was used to deep sequence seven community samples and four hospital samples that were spatiotemporally matched (obtained via Imperial College NHS Trust hospitals). Variants were studied to investigate if certain variants influence disease severity (e.g. cause mild (community samples) or severe infection (hospital samples)). Analysis so far showed that ON1 (with a seventy-two nucleotide duplication in attachment protein G) is the most common genotype in both community and hospitalized samples (90% and 75% of samples, respectively), with GA2 (without duplication) as the next most common genotype for RSV A subtypes (7% and 25%). Three per cent of community samples were of the GA5 genotype. Samples from the RSV B subgroup all belong to the BA genotypes with a 60-nucleotide duplication in G. Samples that were selected for Illumina sequencing had a Ct value between 19.0 and 29.1, while hospital samples had a Ct value of 18.3 to 29.1. Viral load, therefore, did not explain disease severity in these selected samples. The Shannon entropy from Illumina sequenced samples averaged at 22.78 in community samples (ranges from 15 to 28) and 38.78 in hospitalized samples (ranges from 31 to 57). This indicated that diversity of the virus pool might influence disease severity; however, more samples need to be analyzed. There are no specific variants that could explain disease severity. Diversity of the virus pool could explain the link between higher viral loads and disease severity, which is sometimes found but cannot always be confirmed. Higher viral loads can harbor more diverse viral particles compared to lower viral loads. Future work will focus on more in-depth variation and diversity analysis and on evolutionary analysis of both community and hospital samples. We will also investigate intra-host evolution of RSV in acute infections using consecutive samples and its possible implications on the host response.

scholarly journals 998. Forward and Reverse Translational Approaches to Predict Efficacy of the Neutralizing Respiratory Syncytial Virus (RSV) Antibody MK-1654

2021 ◽  
Vol 8 (Supplement_1) ◽  
pp. S590-S590
Author(s):  
Brian M Maas ◽  
Jos Lommerse ◽  
Nele Plock ◽  
Radha Railkar ◽  
S Y Amy Cheung ◽  
...  
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Tract Infection ◽  
Lower Respiratory Tract ◽  
Incidence Rates ◽  
Independent Contractor ◽  
Phase 3 ◽  
Rsv Infection ◽  
Syncytial Virus ◽  
The Relationship

Abstract Background MK-1654 is a respiratory syncytial virus (RSV) F glycoprotein neutralizing monoclonal antibody (mAb) with an extended half-life in late development to prevent RSV infection in infants. Neutralizing mAbs, like MK-1654, have great potential for prophylaxis against viral infection. However, well-validated approaches for clinical dose and efficacy predictions are lacking. Methods Summary-level literature data from RSV prevention studies were used in a model-based meta-analysis (MBMA) to describe the relationship between RSV incidence rates and serum neutralizing antibody (SNA) titer. The model was validated using viral challenge experiments in cotton rats and phase 3 RSV-A efficacy results in infants for an anti-RSV F mAb, REGN-2222. A phase 2b human RSV challenge study (HCS) in adults was also conducted with MK-1654. Participants (N=70) received 100, 200, 300, or 900 mg of MK-1564 or placebo and were challenged intranasally with RSV 29 days later. RSV viral load and symptomatic infection were monitored. Data from the HCS were compared to model predictions. The MBMA was used to predict efficacy of MK-1654 in a virtual population of pre- and full- term infants. Results The relationship between SNA titer and RSV incidence rate defined using the viral load data from the cotton rat approximated the relationship identified for infants from the clinical MBMA. The MBMA was quantitatively consistent with the phase 3 efficacy results against RSV A for REGN-2222. In the HCS, RSV nasal viral load measured by RT-qPCR and quantitative culture as well as symptomatic infections were decreased in MK-1654 recipients compared to placebo. Incidence rates of RSV infection in the HCS were also consistent with MBMA predictions. The model-based clinical trial simulations for MK-1654 indicated a high probability of substantial efficacy against RSV-associated medically attended lower respiratory tract infection ( &gt;75% for 5 months) for doses ≥75 mg. Conclusion Our MBMA successfully quantified the relationship between RSV SNA and clinically relevant endpoints, including lower respiratory tract infection in infants. MBMA-based efficacy predictions support continued development of the MK-1654 antibody for the prevention of RSV in infants. Disclosures Brian M. Maas, PharmD, Merck & Co., Inc. (Employee, Shareholder) Jos Lommerse, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Nele Plock, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Radha Railkar, PhD, Merck & Co., Inc. (Employee, Shareholder) S. Y. Amy Cheung, PhD, Certara (Employee, Shareholder) Luzelena Caro, PhD, Merck & Co., Inc. (Employee, Shareholder) Jingxian Chen, PhD, Merck & Co., Inc. (Employee, Shareholder) Wen Liu, MPH, Merck & Co., Inc. (Employee, Shareholder) Ying Zhang, PhD, Merck & Co., Inc. (Employee, Shareholder) Qinlei Huang, MS, Merck & Co., Inc. (Employee, Shareholder) Wei Gao, PhD, Merck & Co., Inc. (Employee, Shareholder) Li Qin, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Jie Meng, MSc, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Han Witjes, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Emilie Schindler, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Benjamin Guiastrennec, PharmD, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Francesco Bellanti, PhD, Certara (Employee, Shareholder)Merck & Co., Inc. (Independent Contractor) Daniel Spellman, PhD, Merck & Co., Inc. (Employee, Shareholder) Brad Roadcap, MS, Merck & Co., Inc. (Employee, Shareholder) Amy Espeseth, PhD, Merck & Co., Inc. (Employee, Shareholder) S. Aubrey Stoch, MD, Merck & Co., Inc. (Employee, Shareholder) Eseng Lai, MD, PhD, Merck & Co., Inc. (Employee, Shareholder) Kalpit A. Vora, PhD, Merck & Co., Inc. (Employee, Shareholder) Antonios O. Aliprantis, MD, PhD, Merck & Co., Inc. (Employee, Shareholder) Jeffrey R. Sachs, PhD, Merck & Co., Inc. (Employee, Shareholder)

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
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