Infection of cells by respiratory syncytial virus induces the formation of cytoplasmic inclusion bodies (IBs) where all the components of the viral RNA polymerase complex are concentrated. However, ...the exact organization and function of these IBs remain unclear. In this study, we use conventional and super-resolution imaging to dissect the internal structure of IBs. We observe that newly synthetized viral mRNA and the viral transcription anti-terminator M2-1 concentrate in IB sub-compartments, which we term "IB-associated granules" (IBAGs). In contrast, viral genomic RNA, the nucleoprotein, the L polymerase and its cofactor P are excluded from IBAGs. Live imaging reveals that IBAGs are highly dynamic structures. Our data show that IBs are the main site of viral RNA synthesis. They further suggest that shortly after synthesis in IBs, viral mRNAs and M2-1 transiently concentrate in IBAGs before reaching the cytosol and suggest a novel post-transcriptional function for M2-1.Respiratory syncytial virus (RSV) induces formation of inclusion bodies (IBs) sheltering viral RNA synthesis. Here, Rincheval et al. identify highly dynamic IB-associated granules (IBAGs) that accumulate newly synthetized viral mRNA and the viral M2-1 protein but exclude viral genomic RNA and RNA polymerase complexes.
(1) described the predictors of nonseroconversion after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection (36.1% of cases), where nonresponders had significant higher cycle ...threshold (C{subscriptt) and were younger. In April 2020, a 55-year-old female nursing manager had mild SARS-CoV-2 pneumonia diagnosed that did not require admission, confirmed by weakly positive genes E and RNA-dependent RNA polymerase PCR testing (both C{subscriptt >33, near the limit of detection using homemade techniques). All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.
•Respiratory syncytial virus can cause severe infection in adults not only in children.•Coinfection increases the burden of respiratory disease.•Molecular testing is associated with an increased ...prevalence of coinfections.
We present the case of an 81-year-old man, who was immunocompetent, who was admitted to the hospital with symptoms of fever and dyspnea suspected to be caused by COVID-19. Further examination revealed a triple coinfection, as determined by multiplex polymerase chain reaction testing, caused by the respiratory syncytial virus, human coronavirus OC43, and rhinovirus. Upon auscultation, diffuse wheezing without crackles was detected. After ruling out the possibility of acute heart failure with pulmonary edema, the patient was treated with nebulization of terbutaline for a period of 72 hours. This case serves to demonstrate the potential dangers of lifting barrier measures, such as mandatory face masks in high-risk areas, during the fall-winter season. In addition, it highlights the challenges that may arise in the post-COVID-19 era because reliance on flu vaccinations alone may not be sufficient.
Respiratory syncytial virus (RSV) is the most important cause of severe lower-respiratory tract disease in calves and young children, yet no human vaccine nor efficient curative treatments are ...available. Here we describe a recombinant human RSV reverse genetics system in which the red fluorescent protein (mCherry) or the firefly luciferase (Luc) genes are inserted into the RSV genome. Expression of mCherry and Luc are correlated with infection rate, allowing the monitoring of RSV multiplication in cell culture. Replication of the Luc-encoding virus in living mice can be visualized by bioluminescent imaging, bioluminescence being detected in the snout and lungs of infected mice after nasal inoculation. We propose that these recombinant viruses are convenient and valuable tools for screening of compounds active against RSV, and can be used as an extremely sensitive readout for studying effects of antiviral therapeutics in living mice.
Respiratory syncytial virus (RSV) is the primary cause of severe respiratory infection in infants worldwide. Replication of RSV genomic RNA occurs in cytoplasmic inclusions generating viral ...ribonucleoprotein complexes (vRNPs). vRNPs then reach assembly and budding sites at the plasma membrane. However, mechanisms ensuring vRNPs transportation are unknown. We generated a recombinant RSV harboring fluorescent RNPs allowing us to visualize moving vRNPs in living infected cells and developed an automated imaging pipeline to characterize the movements of vRNPs at a high throughput. Automatic tracking of vRNPs revealed that around 10% of the RNPs exhibit fast and directed motion compatible with transport along the microtubules. Visualization of vRNPs moving along labeled microtubules and restriction of their movements by microtubule depolymerization further support microtubules involvement in vRNPs trafficking. Approximately 30% of vRNPs colocalize with Rab11a protein, a marker of the endosome recycling (ER) pathway and we observed vRNPs and Rab11-labeled vesicles moving together. Transient inhibition of Rab11a expression significantly reduces vRNPs movements demonstrating Rab11 involvement in RNPs trafficking. Finally, Rab11a is specifically immunoprecipitated with vRNPs in infected cells suggesting an interaction between Rab11 and the vRNPs. Altogether, our results strongly suggest that RSV RNPs move on microtubules by hijacking the ER pathway.
The minimum requirement for an active RNA-dependent RNA polymerase of respiratory syncytial virus (RSV) is a complex made of two viral proteins, the polymerase large protein (L) and the ...phosphoprotein (P). Here we have investigated the domain on P that is responsible for this critical P-L interaction. By use of recombinant proteins and serial deletions, an L binding site was mapped in the C-terminal region of P, just upstream of the N-RNA binding site. The role of this molecular recognition element of about 30 amino acid residues in the L-P interaction and RNA polymerase activity was evaluated in cellula using an RSV minigenome system and site-directed mutagenesis. The results highlighted the critical role of hydrophobic residues located in this region.
Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract illness in infants. Since no vaccine and no good antivirals against RSV are available, it is essential to better understand how the viral machinery functions in order to develop new antiviral strategies. Like all negative-strand RNA viruses, RSV codes for its own machinery to replicate and transcribe its genome. The core of this machinery is composed of two proteins, the phosphoprotein (P) and the large protein (L). Here, using recombinant proteins, we have mapped and characterized the P domain responsible for this L-P interaction and the formation of an active L-P complex. These findings extend our understanding of the mechanism of action of RSV RNA polymerase and allow us to define a new target for the development of drugs against RSV.
•Outcome of viral respiratory tract infections is closely related to superinfections.•Better use of antibiotics requires routine molecular testing.•Extending vaccination coverage in patients at risk ...is essential to limit mortality.
Following a study of predictors of superinfection in viral respiratory tract infections (VRTIs), this study analyzes the predictors of the outcome.
Multicenter retrospective study conducted among adults who tested positive for VRTIs with reverse-transcription polymerase chain reaction. We compared characteristics between influenza virus, Paramyxoviridae, and Pneumoviridae and identified predictors of favorable short-term outcome, admission to the intensive care unit (ICU), and mortality.
A total of 590 patients had VRTI, including 347 (59%) influenza infections. Mean (SD) patient age was 71.0 (18.3) years, with a sex ratio of 0.91. In multivariate analyses, predictors of favorable short-term outcome were age ≤75 years (adjusted odds ratio aOR 5.38 95% confidence interval, 1.59-18.2), absence of respiratory disease (4.94 1.01-24.37), and absence of superinfection (aOR 3.91 1.37-11.13). The predictors of ICU admission were age ≤75 years (aOR 3.28 1.71-6.25), chronic respiratory disease (aOR 2.49 1.20-5.19), and procalcitonin level >0.25 ng/mL (aOR 4.25 1.55-11.67). Predictors of mortality were use of inhaled corticosteroids (2.49 1.10-5.63), influenza infection (2.73 1.27-5.85), Charlson score ≥5 (5.35 1.90-15.05), superinfection (2.54 1.05-6.18), and eosinophil count <50/µL (4.39 1.19-16.2). Certainty of superinfection was significantly associated with mortality (2.23 1.15-4.3).
Our study revealed that superinfection was significantly related to the outcome, and that virus species affects mortality. These findings emphasize the need for improving the tools used in daily practice to confirm certainty of superinfection and for broader implementation of vaccination of individuals at risk of VRTIs.
We report evaluation of 30 assays’ (17 rapid tests (RDTs) and 13 automated/manual ELISA/CLIA assay (IAs)) clinical performances with 2594 sera collected from symptomatic patients with positive ...SARS-CoV-2 rRT-PCR on a respiratory sample, and 1996 pre-epidemic serum samples expected to be negative. Only 4 RDT and 3 IAs fitted both specificity (> 98%) and sensitivity (> 90%) criteria according to French recommendations. Serology may offer valuable information during COVID-19 pandemic, but inconsistent performances observed among the 30 commercial assays evaluated, which underlines the importance of independent evaluation before clinical implementation.
Acquired infections in hospitalized elderly people are a growing concern. In long-term care facilities with multiple staff and visitor contacts, virus outbreaks are a common challenge for infection ...prevention teams. Although several studies have reported nosocomial RSV outbreaks in long term care facilities, molecular epidemiology data are scarce.
RSV RNA was detected in respiratory samples from 19 patients in a long-term care hospital for elderly in Paris in March 2019 over a 3 weeks period. Genotyping was performed using nucleotide sequencing. Sociodemographic and clinical characteristics of cases part of a unique cluster, were retrospectively reviewed.
Molecular investigation of theses RSV cases, revealed a unique cluster of 12 nosocomial cases in 2 adjacent wards. Mean age of these outbreak's cases was 89. All patients had underlying medical conditions. Seven exhibited lower respiratory symptoms and three experienced decompensation of underlying chronic heart condition. Two patients died.
This case report highlights the importance of RSV in causing substantial disease in elderly in case of nosocomial outbreak and the contributions of molecular epidemiology in investigation and management of such outbreak.
PDF
