Visualizing the replication of respiratory syncytial virus in cells and in living mice

Abstract

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.

A Druggable Pocket at the Nucleocapsid/Phosphoprotein Interaction Site of Human Respiratory Syncytial Virus

“…Antiviral activities of BPdCs. To assess the relevance of the results obtained in vitro with BPdCs, we used a recombinant virus, rHRSV-mCherry (26), to monitor RSV multiplication by following the expression of the red fluorescent mCherry protein in cell cultures. Preliminary experiments with the highestaffinity compound, M76, did not show significant inhibitory activity.…”

“…The inhibition of RSV replication was measured in cell cultures using an assay based on a human RSV reverse-genetics system in which the red fluorescent protein mCherry was inserted into the RSV genome, recombinant HRSV-Cherry (rHRSVmCherry), as described previously (26). HEp-2 cells (ATCC number CCL-23) or BHK-21 cells maintained in Eagle's minimum essential medium (EMEM) or Dulbecco modified essential medium (DMEM), respectively, were seeded at 5 ϫ 10 4 cells per well in 96-well plates the day before and infected for 2 h with 500 PFU of rHRSV-mCherry or 1 h with 50 PFU of vesicular stomatitis virus (VSV)-green fluorescent protein (GFP) (27).…”

Animal models of respiratory syncytial virus infection

“…Following IN inoculation, hRSV replicates in the nasal passages and lungs (Table 2) [78], [79]. Although mice are susceptible to hRSV infection throughout life, older mice are more susceptible than younger animals [80], [81].…”

Kinésithérapie respiratoire dans la bronchiolite : stop ou encore ?

“…En l'absence de traitement curatif, les traitements médicaux sont limités et reposent sur la désobstruction rhinopharyngée, les antipyrétiques, le fractionnement de l'alimentation. La découverte récente d'une nouvelle méthode pour étudier in vivo le virus respiratoire syncitial (principal virus imputé dans la bronchiolite aiguë du nourrisson) sur des souris vivantes permet d'espérer la découverte d'un traitement anti-rétroviral, voire d'un vaccin [1].…”