A subset of neutrophils in cystic fibrosis (CF) airways abundantly release the serine protease neutrophil elastase (NE), causing lung damage. Triggers of this subset are unknown, limiting intervention. Respiratory viral-bacterial co-infections may be a possible cause, but this has not been directly studied. To explore this, we developed a laboratory pipeline to characterise responses of airway tissue and neutrophils to virus and bacteria. Differentiated paediatric primary airway epithelial cultures were challenged individually or co-infected with rhinovirus and bacteria representative of pathogens and oral flora found in CF airways (Pseudomonas aeruginosa, Staphylococcus aureus, Neisseria lactamica, and Prevotella nigrescens). Cultures were then washed with medical saline to sample infection milieus. Washes were used to assess epithelial cytokine release and also applied to an in vitro model of neutrophil transmigration to the airways. Infection microenvironments were characterised by increased antiviral signals with exclusive rhinovirus infection, including MIG and IP-10, while P. aeruginosa uniquely increased pro-inflammatory cytokines IL-1α and β. Neutrophils migrating into washes primed by P. aeruginosa and P. nigrescens had elevated marker of NE release CD63 (7 fold, p<0.001), and reduced expression of phagocytic receptor CD16 (0.3 fold, p<0.001). These changes are reflective of neutrophils in CF airways. Our findings indicate that certain bacteria, as primary or secondary infections, trigger NE release in recruited neutrophils. Both pathogens and oral microbes may activate this subset in CF airways. Ongoing studies seek to characterise reprogramming mechanisms and identify potential therapeutic targets.

Aspergillus is increasingly associated with lung inflammation and mucus plugging in early cystic fibrosis (CF) disease during which conidia burden is low and strains appear to be highly diverse. It is unknown whether clinical Aspergillus strains vary in their capacity to induce epithelial inflammation and mucus production. We tested the hypothesis that individual colonising strains of Aspergillus fumigatus would induce different responses. Ten paediatric CF Aspergillus isolates were compared along with two systemically invasive clinical isolates and an ATCC reference strain. Isolates were first characterised by ITS gene sequencing and screened for antifungal susceptibility. Three clusters (A−C) of Aspergillus isolates were identified by ITS. Antifungal susceptibility was variable, particularly for itraconazole. Submerged CF and non-CF monolayers as well as differentiated primary airway epithelial cell cultures were incubated with conidia for 24 h to allow germination. None of the clinical isolates were found to significantly differ from one another in either IL-6 or IL-8 release or gene expression of secretory mucins. Clinical Aspergillus isolates appear to be largely homogenous in their mucostimulatory and immunostimulatory capacities and, therefore, only the antifungal resistance characteristics are likely to be clinically important.