Investigating alternative strategies for the prevention of pneumococcal colonisation and disease

© 2017 Dr. Jayne ManningStreptococcus pneumoniae (the pneumococcus) is an important respiratory pathogen that colonises the human nasopharynx. While generally asymptomatic, colonisation is a prerequisite for disease. Infants are exposed to pneumococci soon after birth, carry the bacterium more often compared to older children and adults, and suffer the greatest burden of pneumococcal disease. Pneumococcal conjugate vaccines (PCVs) target the bacterium’s polysaccharide capsule. PCV use has dramatically reduced pneumococcal carriage and invasive disease caused by 10 – 13 of the > 90 antigenically-distinct pneumococcal capsule types (serotypes). However, this reduction has been coupled with an increased prevalence of non-vaccine-type pneumococci within vaccinated populations. Consequently, overall pneumococcal carriage rates have remained largely unchanged. In addition, PCV efficacy against pneumococcal pneumonia and otitis media (OM) is not comparable to that observed for invasive disease. This thesis aims to investigate how two alternative strategies to PCVs – probiotics and a pneumococcal whole cell vaccine – affect pneumococcal colonisation and disease. The probiotic bacterium Streptococcus salivarius is a commensal of the human respiratory tract that can inhibit pneumococcal adherence to epithelial cells in vitro. Here, a pharyngeal epithelial cell model was used to investigate how pre- administration of S. salivarius inhibits pneumococcal adherence. The megaplasmids of S. salivarius strains K12 and M18, which harbour known bacteriocins, were found not to be required to inhibit pneumococcal adherence in vitro, despite mediating the inhibition of pneumococcal growth on solid agar. Further experiments testing K12 and two pneumococcal isolates (serotypes 19F and 6A) suggest that K12 employs more than one mechanism to inhibit pneumococcal adherence. A significant inverse correlation was observed between K12 adherence and 19F adherence, and no inhibition was observed when K12-epithelial cell contact was prevented using a transwell system, suggesting some form of direct competitive inhibition by intact bacteria was taking place. In contrast, K12 inhibited 6A in the absence of epithelial cell contact, and no correlation was observed between K12 and 6A adherence, suggesting other, possibly secreted factor mechanisms were associated with inhibition of 6A adherence. The pneumococcal whole cell vaccine (WCV) consists of killed, unencapsulated pneumococci. In mice, WCV induces serotype-independent protection against colonisation and invasive pneumococcal disease via TH17 and antibody-mediated immunity, respectively. This study used infant mouse models to investigate the effect of WCV in two clinically relevant settings: influenza A virus (IAV)-induced OM, and in the presence of established pneumococcal colonisation (“therapeutic” vaccination). A single sub-cutaneous dose of WCV did not prevent the development of IAV-induced pneumococcal OM, but did significantly reduce the density of pneumococci in the middle ears of mice six days after IAV co-infection. Experiments performed in immune-deficient mice suggest that both CD4+ T cells and antibodies were required for this effect. When administered to colonised mice, a single dose of therapeutic WCV significantly reduced pneumococcal colonisation density in what was apparently an antibody-mediated manner. Prior colonisation enhanced the systemic WCV- specific IgG response, but not splenic IL-17A responses. Significant reductions in pneumococcal density were also observed in the nasopharynx and middle ears of therapeutically-vaccinated mice after IAV co-infection. Overall, this thesis contributes to our knowledge of probiotic mechanisms inhibiting pneumococcal adherence, and provides the first evidence for the therapeutic use of WCV. Therapeutic strategies to reduce disease severity would be in particular useful in low-income settings where infants more often experience early, high-density pneumococcal carriage and the highest burden of pneumococcal disease