Evaluation of live attenuated Streptococcus pneumoniae vaccine strains on the epithelial response to colonisation using a human challenge model

Background: Streptococcus pneumoniae frequently colonises

the human nasopharynx but causes over 500,000 deaths each year

from Pneumonia, Sepsis and Meningitis. Nasopharyngeal carriage

is required for transmission and is a pre-requisite for disease.

The conjugate polysaccharide vaccines have proven effective in

decreasing disease. However, replacement of vaccine serotypes

with non-vaccine types in carriage threatens the future of the vaccines’

efficiency.

Using an Experimental Human Pneumococcal Challenge model

(EHPC) and epithelial cell culture models, we have previously

shown that pneumococcal colonisation involves both direct epithelial

association and micro-invasion, inducing innate immunity and

clearance without overt disease. Repeated challenge in the EHPC

with the same strain decreases subsequent carriage efficiency and

diminishes transmission potential and/or progression to disease,

suggesting active mucosal immunity in the nasopharynx.

Methods and materials: We have generated live attenuated

strains of 6B S. pneumoniae (AS1 and AS2) that have double virulence

deletions and cannot revert to cause disease. Here, we have

explored the hypothesis that despite their attenuation in a mouse

model of disease, these attenuated strains retain their ability to

invade the epithelium and induce epithelial-derived innate immunity

in humans.

Colonisation were measured by confocal microscopy and microbiology

density by CFU counts. Epithelial activation was measured

by flow cytometry, ELISA and RNAseq.

Results: We found that both mutants colonised the human

nasopharynx and formed epithelial associations with microinvasion

in the EHPC model. In vitro, both mutants adhered, invaded

and transmigrated across the epithelium 4-fold less than wild type.

However, both mutants still resulted in secretion of IL-8, IL-6 and

ICAM-1 secretion and barrier integrity was maintained. PCA analyses

revealed that epithelial transcriptomic responses between wild

type and the mutants generally overlapped, indicating overall similar

stimulation of signaling pathways following exposure.

Conclusion: The results reveal that attenuation of these pneumococcal

strains has not led to loss of their ability to elicit amucosal

immune/inflammatory response. This approach provides an exciting

new pipeline for the development and testing of novel vaccines.

The application of these attenuated strains in the EHPC also has the

potential to provide important new knowledge on the mechanisms

behind bacterial clearance, transmission and disease progression

during colonisation.