Inflammation of the Nasal Mucosa is Associated with Susceptibility to Experimental Pneumococcal Challenge in Older Adults

Streptococcus pneumoniae colonization in the upper respiratory tract is linked to pneumococcal disease development, predominantly affecting young children and older adults. As the global population ages and comorbidities increase, there is a heightened concern about this infection. We investigated the immunological responses of older adults to pneumococcal controlled human infection by analysing the cellular composition and gene expression in the nasal mucosa. Our comparative analysis with data from a concurrent study in younger adults revealed distinct gene expression patterns in older individuals susceptible to colonization, highlighted by neutrophil activation and elevated levels of CXCL9 and CXCL10. Unlike younger adults challenged with pneumococcus, older adults did not show recruitment of monocytes into the nasal mucosa following nasal colonization. However, older adults who were protected from colonization showed increased degranulation of CD8+ T cells, both before and after pneumococcal challenge. These findings suggest age-associated cellular changes, in particular enhanced mucosal inflammation, that may predispose older adults to pneumococcal colonization. Healthy adults aged 50-84 were inoculated with an estimated 80,000 CFU per nostril of Streptococcus pneumoniae serotype 6B (strain BHN418, GenBank accession number ASHP00000000.1). Key eligibility criteria included capacity to give informed consent, no immunocompromised state or contact with susceptible individuals or children, no pneumococcal or influenza vaccine or infection in the last 2 years and not having taken part in EHPC studies in the past 3 years. All volunteers gave written informed consent, and research was conducted in compliance with all relevant ethical regulations. Participants who carried non-experimental pneumococcal strains at baseline (day -5) or had a viral infection were excluded from all analyses. Ethical approval was provided by the East Liverpool National Health Service Research and Ethics Committee (reference numbers 15/NW/0146 and 14/NW/1460) and Human Tissue Authority licensing number 12548. RNA sequencing: Nasal cells were collected in RLT (QIAGEN) and stored at -80°C until extraction. RNA extraction (RNeasy; QIAGEN), sample integrity assessment (Bioanalyzer; Agilent 2100), library preparation and RNA sequencing (BGISEQ-500RS) were performed at the Beijing Genome Institute. Quality control of raw sequencing data was done using FastQC tool. Mapping to a human reference genome assembly (GRCh38) was done using STAR 2.5.0a​. Read counts from the resulting binary alignment map (BAM) files were obtained with featureCounts​​ using a general transfer format (GTF) gene annotation from the Ensembl database​. The R/Bioconductor package DESeq2 (version 1.34.0) was used to identify differentially expressed genes among the samples after removing absent features (zero counts in more than 75% of samples)​. Genes with a p-value of < 0.05 and an absolute fold-change of > 2.0 were identified as differentially expressed. ***Submitters state that raw data have been lost and therefore are not provided.