Rationale: Respiratory syncytial virus (RSV) and Streptococcus pneumoniae are major respiratory pathogens. Coinfection with RSV and S. pneumoniae is associated with severe and often fatal pneumonia but the molecular basis for this remains unclear.Objectives: To determine if interaction between RSV and pneumococci enhances pneumococcal virulence.Methods: We used confocal microscopy and Western blot to identify the receptors involved in direct binding of RSV and pneumococci, the effects of which were studied in both in vivo and in vitro models of infection. Human ciliated respiratory epithelial cell cultures were infected with RSV for 72 hours and then challenged with pneumococci. Pneumococci were collected after 2 hours exposure and changes in gene expression determined using quantitative real-time polymerase chain reaction.Measurements and Main Results: Following incubation with RSV or purified G protein, pneumococci demonstrated a significant increase in the inflammatory response and bacterial adherence to human ciliated epithelial cultures and markedly increased virulence in a pneumonia model in mice. This was associated with extensive changes in the pneumococcal transcriptome and significant up-regulation in the expression of key pneumococcal virulence genes, including the gene for the pneumococcal toxin, pneumolysin. We show that mechanistically this is caused by RSV G glycoprotein binding penicillin binding protein 1a.Conclusions: The direct interaction between a respiratory virus protein and the pneumococcus resulting in increased bacterial virulence and worsening disease outcome is a new paradigm in respiratory infection.Keywords: respiratory syncytial virus; pneumococcus; cilia; virulence; G protein Respiratory syncytial virus (RSV) and Streptococcus pneumoniae, also known as the pneumococcus, are major respiratory pathogens causing a huge global healthcare burden, predominantly in young children and the elderly (1). Global RSV disease burden is estimated at 64 million cases and 160,000 deaths every year (2). Pneumococcal pneumonia results in more than 1 million deaths each year worldwide with approximately half a million in children younger than 5 years (3). There is increasing evidence that RSV and the pneumococcus interact to increase the severity of respiratory disease (4-6). Seasonal increases in infections This article has an online supplement, which is accessible from this issue's table of contents at www.atsjournals.org This article has embedded videos, which play in place from both the online PDF or HTML versions. If you cannot view Flash videos on your device, please try playing the videos in the online PDF, or access the original uncompressed videos at http://www.atsjournals.org/doi/suppl/10.1164/rccm.201311-2110OC. To play, mouse over the image and click on the arrow that will appear in the center or on the lower left.

Respiratory syncytial virus is a major cause of respiratory disease. There are conflicting accounts of the response of human epithelial cells to respiratory syncytial virus and a lack of data on its effect on ciliary function. Our aim was to study the early stages of respiratory syncytial virus infection of primary human basal and ciliated cultures.Using high speed videomicroscopy, we found that ciliary beat frequency was unaffected by respiratory syncytial virus infection over 72 h; however, ciliary dyskinesia significantly increased within 24 h of infection (p,0.05). Transmission electron microscopy revealed that ultrastructural abnormalities were confined to ciliated cells, including increased cilia loss and mitochondrial damage. Confocal immunofluorescence microscopy showed that respiratory syncytial virus antigen gradually spread from the cell surface to the ciliary tip of infected cells over 3 days. Interestingly, ciliated cultures secreted fewer viruses than basal (progenitor) cell cultures and produced a chemokine response focused on recruitment of neutrophils.This study highlights differences in infection models and underscores the need to explore further the role of ciliated cells in the establishment of respiratory syncytial virus infection.Increased ciliary dyskinesia combined with ciliary loss and epithelial damage is likely to result in reduced mucociliary clearance early in the infective process. @ERSpublications Increased ciliary dyskinesia with ciliary loss and epithelial damage can result in reduced mucociliary clearance

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