Direct Binding of Respiratory Syncytial Virus to Pneumococci: A Phenomenon That Enhances Both Pneumococcal Adherence to Human Epithelial Cells and Pneumococcal Invasiveness in a Murine Model

Abstract

In a previous study we showed that pneumococcal adherence to epithelial cells was enhanced by a preceding respiratory syncytial virus (RSV) infection. RSV-glycoproteins, expressed on the infected cell surface, may play a role in this enhanced pneumococcal binding, by acting as bacterial receptors. In the current study, it was attempted to analyze the capacity of pneumococci to interact directly with RSV virions. By flowcytometry, a direct interaction between RSV and pneumococci could be detected. Heparin, an inhibitor of RSV infectivity that interacts with RSV protein-G, blocked RSV-pneumococcal binding, indicating that the latter interaction is indeed mediated by protein-G. RSV-pneumococcal complexes showed enhanced adherence to uninfected human epithelial cells, compared with pneumococcal adherence without bound RSV, and this enhancement was also blocked by heparin. In addition, the significance of these findings in vitro was explored in vivo in a murine model. Both mice that were pretreated with RSV at day 4 before pneumococcal challenge and mice infected with both agents simultaneously showed significantly higher levels of bacteraemia than controls. Simultaneous infection with both agents enhanced the development of pneumococcal bacteraemia most strongly. It was hypothesized that direct viral binding is another mechanism by which RSV can induce enhanced pneumococcal binding to epithelial cells, a phenomenon that is translated in vivo by a higher invasiveness of pneumococci when administered simultaneously with RSV to mice. Apparently, RSV acts in this process as a direct coupling particle between bacteria and uninfected epithelial cells, thereby increasing colonization by and enhancing invasiveness of pneumococci. Clinical and epidemiologic data suggest that respiratory syncytial virus (RSV) infections in humans can be complicated by bacterial superinfection e.g., with Streptococcus pneumoniae, leading to increased morbidity (1-5). Mechanisms underlying bacterial superinfection include virus-induced local destruction of the epithelium, compromising the host's physiologic barrier, and virus-induced modulation of the immune response (6,7).In addition, enhanced bacterial adherence to virus-infected cells is considered an important factor increasing the risk of bacterial superinfection (8 -11). In a previous study in vitro, we obtained evidence for such a mechanism. A preceding RSV infection of human respiratory tract epithelium led to significantly enhanced adherence of S. pneumoniae (12).The basis of RSV-enhanced pneumococcal adherence is not known. RSV infection both leads to expression of viral glycoproteins and up-regulation of cellular molecules on the hostcell membrane. Both could possibly serve as bacterial receptors, as has been described for Neisseria meningitidis: viral glycoprotein G as well as cellular molecules, like CD 14 and CD18, are involved in enhanced binding of N. meningitidis to RSV-infected cells (13,14

Viral–bacterial interactions in the respiratory tract

“…Next to increased expression of host receptors, viral proteins displayed on the cell surface can enhance adherence and internalization of bacteria. The RSV attachment glycoprotein (G) protein, present on the surface of either RSV virions or infected cells, can serve as a binding structure for non-typeable H. influenzae (Avadhanula et al, 2007), S. pneumoniae (Avadhanula et al, 2007;Hament et al, 2005) and Pseudomonas aeruginosa (Van Ewijk et al, 2007). Similarly, integration of influenza virus haemagglutinin into the cell membrane of infected cells facilitates attachment and invasive disease of group A streptococci in mice (Okamoto et al, 2003).…”

Viral/bacterial respiratory tract co-infections in children

“…The RSV G glycoprotein has previously been shown to act as a receptor for S. pneumoniae adherence, thus promoting pneumococcal colonisation of the airways (128,136,137). RSV has also been shown to enhance pneumococcal virulence in animal models, with co-infection resulting in increased inflammation (128,137,262).…”

“…Evidence from previous studies suggests that RSV may trigger an outgrowth of S. pneumoniae from the URT flora through promotion of bacterial adherence and disruption of bacterial clearance mechanisms (128,138,140,262). However, we have previously hypothesised that, since RSV virion and S. pneumoniae complexes have been shown to form (128,136,137), these complexes may be co-transmitted in respiratory secretions, particularly in settings of rapid transmission such as childcare centres (6). Longitudinal studies with samples taken before, during and after an RSV infection within the same participant may represent the best way to differentiate between these hypotheses, however, to date such longitudinal studies have focused on characterising the nasopharyngeal microbiota, and have only shown increases in abundance of the Streptococcus genus during viral ARI, without species-level resolution and without reporting changes in abundances surrounding an ARI episode within individuals (5, 18).…”

“…Viral infection of airway epithelial cells has been found to aid bacterial adherence to the URT by a number of mechanisms, although the effect differs between viruses, bacterial strains and experimental models. Free RSV virions have been shown to bind directly to H. influenzae and S. pneumoniae (128,136,137), which could through proximity augment each pathogen's binding to their respective host cell receptors ( Figure 1.2A). There is also some evidence that the RSV G glycoprotein, localised at the host cell membrane of virus infected cells, can act as a receptor for bacteria (Figure 1.2B) (136,137).…”

“…The diversity of S. pneumoniae strains observed in the study described in Chapter 3, and the known diversity of RSV strains in the paediatric population (188,236) led to the question investigated in the final two research chapters of this thesis: do specific RSV and S. pneumoniae strains interact preferentially with each other? There is evidence that RSV and S. pneumoniae can synergistically interact in cell culture and animal models through binding interactions between the bacteria and the RSV G glycoprotein, the extent of which appears in part dependent on the pneumococcal strain (128,136,137,140). However, previous binding studies of RSV and S. pneumoniae have been performed with a small range of pneumococcal isolates and cell culture-adapted RSV strains, which may not closely resemble those circulating and potentially interacting in the paediatric population.…”