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 Viral Load Dynamics and Clinical Disease Severity in Infants With Respiratory Syncytial Virus Infection

2018 ◽  
Vol 219 (8) ◽  
pp. 1207-1215 ◽  
Author(s):  
Cristina Garcia-Mauriño ◽  
Melissa Moore-Clingenpeel ◽  
Jessica Thomas ◽  
Sara Mertz ◽  
Daniel M Cohen ◽  
...  
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Virus Infection ◽  
Disease Severity ◽  
Respiratory Syncytial Virus Infection ◽  
Clinical Disease ◽  
Syncytial Virus

scholarly journals A Randomized, Placebo-Controlled, Respiratory Syncytial Virus Human Challenge Study of the Antiviral Efficacy, Safety, and Pharmacokinetics of RV521, an Inhibitor of the RSV-F Protein

2019 ◽  
Vol 64 (2) ◽  
Author(s):  
John DeVincenzo ◽  
Dereck Tait ◽  
John Efthimiou ◽  
Julie Mori ◽  
Young-In Kim ◽  
...  
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Adverse Events ◽  
Disease Severity ◽  
Controlled Trial ◽  
Primary Analysis ◽  
Double Blind ◽  
Challenge Virus ◽  
F Protein ◽  
Syncytial Virus

ABSTRACT Effective treatments for respiratory syncytial virus (RSV) infection are lacking. Here, we report a human proof-of-concept study for RV521, a small-molecule antiviral inhibitor of the RSV-F protein. In this randomized, double-blind, placebo-controlled trial, healthy adults were challenged with RSV-A Memphis-37b. After infection was confirmed (or 5 days after challenge virus inoculation), subjects received RV521 (350 mg or 200 mg) or placebo orally every 12 h for 5 days. The primary endpoint was area under the curve (AUC) for viral load, as assessed by reverse transcriptase quantitative PCR (RT-qPCR) of nasal wash samples. The primary efficacy analysis set included subjects successfully infected with RSV who received ≥1 dose of study drug. A total of 66 subjects were enrolled (n = 22 per group); 53 were included in the primary analysis set (RV521 350 mg: n = 16; 200 mg: n = 18; placebo: n = 19). The mean AUC of RT-qPCR-assessed RSV viral load (log10 PFU equivalents [PFUe]/ml · h) was significantly lower with RV521 350 mg (185.26; standard error [SE], 31.17; P = 0.002) and 200 mg (224.35; SE, 37.60; P = 0.007) versus placebo (501.39; SE, 86.57). Disease severity improved with RV521 350 mg and 200 mg versus placebo (P = 0.002 and P = 0.009, respectively, for AUC total symptom score [score × hours]). Daily nasal mucus weight was significantly reduced (P = 0.010 and P = 0.038 for RV521 350 mg and 200 mg, respectively, versus placebo). All treatment-emergent adverse events were grade 1 or 2. No subjects discontinued due to adverse events. There was no evidence of clinically significant viral resistance, and only three variants were detected. RV521 effectively reduced RSV viral load and disease severity in humans and was well tolerated. (This study has been registered at ClinicalTrials.gov under registration no. NCT03258502.)

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.

Association of Viral Load With Disease Severity in Outpatient Children With Respiratory Syncytial Virus Infection

2020 ◽  
Vol 222 (2) ◽  
pp. 298-304 ◽  
Author(s):  
Erika Uusitupa ◽  
Matti Waris ◽  
Terho Heikkinen
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Disease Severity ◽  
Respiratory Infections ◽  
Respiratory Illness ◽  
Lower Viral Load ◽  
Primary Analysis ◽  
Rsv Infection ◽  
Syncytial Virus ◽  
Outpatient Children

Abstract Background There are scarce data on whether viral load affects the severity of respiratory syncytial virus (RSV) disease in outpatient children. Methods We analyzed the association between viral load and disease severity among children who participated in a prospective cohort study of respiratory infections. The children were examined and nasal swabs for the detection of RSV were obtained during each respiratory illness. Quantification of RSV load was based on the cycle threshold (Ct) value. For the primary analysis, the children were divided into 2 groups: higher (Ct &lt; 27) and lower viral load (Ct ≥ 27). Results Among 201 episodes of RSV infection, children with higher viral load had significantly longer median durations of rhinitis (8 vs 6 days; P = .0008), cough (8 vs 6 days; P = .034), fever (2 vs 1 days; P = .018), and any symptom (10 vs 8 days; P = .024) than those with lower viral load. There were statistically significant negative correlations between the Ct values and durations of all measured symptoms. Conclusions Our findings support the concept that viral load drives the severity of RSV disease in children. Reducing the viral load by RSV antivirals might provide substantial benefits to outpatient children.

scholarly journals Relating plaque morphology to respiratory syncytial virus subgroup, viral load, and disease severity in children

2015 ◽  
Vol 78 (4) ◽  
pp. 380-388 ◽  
Author(s):  
Young-In Kim ◽  
Ryan Murphy ◽  
Sirshendu Majumdar ◽  
Lisa G. Harrison ◽  
Jody Aitken ◽  
...  
Keyword(s):  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Disease Severity ◽  
Plaque Morphology ◽  
Syncytial Virus

scholarly journals Seasonality, molecular epidemiology, and virulence of Respiratory Syncytial Virus (RSV): A perspective into the Brazilian Influenza Surveillance Program

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0251361
Author(s):  
Lucas A. Vianna ◽  
Marilda M. Siqueira ◽  
Lays P. B. Volpini ◽  
Iuri D. Louro ◽  
Paola C. Resende
Keyword(s):  
Genetic Diversity ◽  
Viral Load ◽  
Respiratory Syncytial Virus ◽  
Disease Severity ◽  
Clinical Severity ◽  
Influenza Surveillance ◽  
Surveillance Program ◽  
Inverse Association ◽  
Severe Acute Respiratory Infection ◽  
Syncytial Virus

Background Respiratory Syncytial Virus (RSV) is the main cause of pediatric morbidity and mortality. The complex evolution of RSV creates a need for worldwide surveillance, which may assist in the understanding of multiple viral aspects. Objectives This study aimed to investigate RSV features under the Brazilian Influenza Surveillance Program, evaluating the role of viral load and genetic diversity in disease severity and the influence of climatic factors in viral seasonality. Methodology We have investigated the prevalence of RSV in children up to 3 years of age with severe acute respiratory infection (SARI) in the state of Espirito Santo (ES), Brazil, from 2016 to 2018. RT-qPCR allowed for viral detection and viral load quantification, to evaluate association with clinical features and mapping of local viral seasonality. Gene G sequencing and phylogenetic reconstruction demonstrated local genetic diversity. Results Of 632 evaluated cases, 56% were caused by RSV, with both subtypes A and B co-circulating throughout the years. A discrete inverse association between average temperature and viral circulation was observed. No correlation between viral load and disease severity was observed, but children infected with RSV-A presented a higher clinical severity score (CSS), stayed longer in the hospital, and required intensive care, and ventilatory support more frequently than those infected by RSV-B. Regarding RSV diversity, some local genetic groups were observed within the main genotypes circulation RSV-A ON1 and RSV-B BA, with strains showing modifications in the G gene amino acid chain. Conclusion Local RSV studies using the Brazilian Influenza Surveillance Program are relevant as they can bring useful information to the global RSV surveillance. Understanding seasonality, virulence, and genetic diversity can aid in the development and suitability of antiviral drugs, vaccines, and assist in the administration of prophylactic strategies.

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.

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