Acute bronchiolitis, the most common lower respiratory tract infection and the leading cause of hospitalization in infants less than 12 months of age. It is caused by respiratory viruses, in particular Respiratory Syncytial Virus (RSV), Rhinovirus (RV) and Metapneumovirus (MPV) are frequently involved. It is still unknown why some full-term previously healthy infants may present a severe form of bronchiolitis, but the active interaction among host and viral factors is pivotal to determining the severity of respiratory symptoms. Many viral factors (virus type, subtype and genotype and viral load) may be implicated in bronchiolitis pathogenesis. Bronchiolitis can be viewed as a disease of defective (enhanced or depressed) immunoregulation. Th1 polarization is correlated with the efficient eradication of viral infection, whereas Th2 leads to an inefficient viral elimination and to a more severe disease. We propose a large study on infants hospitalized for bronchiolitis with the aim of: (i) characterizing the clinical phenotypes of bronchiolitis; (ii) establishing association of viral factors with patients' clinical features and disease severity; (iii) giving insights into the immunopathogenesis of viral bronchiolitis. The ultimate goal is the identification of severe bronchiolitis determinants with the aim of improving early diagnosis and developing targeted therapies.
This project will be mainly focused on studying in vivo immune response to respiratory viruses infection in infants hospitalized with bronchiolitis. Accordingly, as the main approach of the project, we plan to measure TLRs activation and the downstream pathways of the innate immune response directed to production of regulatory or inflammatory cytokines and to type I and/or III IFNs in infant's respiratory samples. We retain a central issue, not previously addressed in respiratory infections, to study whether and at which steps the activation of the innate immunity pathways in response to RSV and other viruses, could influence the clinical severity. In fact, TLR-driven, pro- or anti-inflammatory cytokine secretion and the extent of the IFNs induced genes (ISGs) activity act as a host defense response, but under certain conditions could be ineffective or detrimental to the host. It has recently proposed that RSV-related pathology is mediated by the host response which may contribute to disease progression via promotion of inappropriate Th-2 response. On the other hand, the influence of viral factors such as specific infecting strains and the relevance of elevated viral loads in determining bronchiolitis severity are not understood sufficiently. To this regard, some studies, including ours (Scagnolari 2012) revealed a significant association between disease severity and RSV load in nasopharyngeal secretion of infants with primary respiratory tract infection but other failed to found such a correlation. It is entirely possible that discrepancies were due to different strains of RSV circulating in different epidemic seasons. As a matter of fact, we detected for the first time in Italy a novel RSV-A genotype ON1, first identified in Ontario (Canada) in December 2010 (Pierangeli 2014). ON1 bears an insertion of 24 amino acids in the G glycoprotein as well as amino acid changes likely to change antigenicity. By early 2013, ON1 had spread so efficiently that replaced other RSV-A strains and it is now the only RSV-A circulating. Accordingly, we plan to measure ON1 viral loads in bronchiolitis cases to compare them to the previous circulating RSV-A and B. Moreover, it has been suggested that the clinical spectrum of bronchiolitis might reflect differences in the profile and extent of immunological and/or inflammatory mediators produced by RSV infected respiratory epithelial cells. Again, different RSV strains may give rise to different immune and/or inflammatory response. We believe that quantification of viral loads in sequential samples from the same patients, together with measurements of IFNrelated genes differential expression and cytokine production, will result in the identification of significant variation in important genes. These results may contribute to identify which innate immunity pathway is more relevant in the natural course of a specific viral respiratory infection. Whole-blood from patients will be collected to measure in vitro thire immune profile. An innovative technique (Frassanito, 2018) requiring a small amount of blood, a significant requisite in the case of pediatric patients, will be employed to detect CD4 and CD8 T lymphocytes producing IFNgamma, Il-4 or IL-17 (e.g. Type-1, Type-2 or Type-17 cells). The most innovative part of the research is aimed to correlate frequency and severity of RSV and other respiratory infections with clinical severity. Anyway, we believe that results from this project, may direct future studies. Increase knowledge on the pathogenesis of bronchiolitis may allow early detection of children at risk of developing severe forms of the disease. The proposed project will allow to identify the role played by viral and host factors to determine the severity of bronchiolitis, thus providing innovative tools for an early diagnosis of disease severity and harmful sequelae and developing targeted therapies.