Introduction and Objectives Cystic fibrosis (CF) and non-cystic fibrosis bronchiectasis (BX) are chronic airway diseases with significant microbial involvement. The presence of mucoid Pseudomonas aeruginosa (mPA) is associated with worse outcomes in these patients. We hypothesised that the presence of this organism would also influence the microbial community structure. We used high throughput sequencing to identify microorganisms present in sputum from these patients, and to associate culture data of mPA with this analysis.
Methods Expectorated sputum was collected from 16 patients (9 CF, 7 BX) and DNA extracted using standard protocols. PCR of the bacterial 16S rRNA gene and subsequent sequencing using a Roche 454 GS Junior sequencer was compared with standard clinical culture. Non-parametric t test was used to identify significant differential abundance of taxa in the molecular data (Bonferonni-corrected).
Results In the pilot study, 35 000 high quality 16S rRNA sequences were generated and could be assigned to 240 different bacterial taxa. Both CF and BX samples were dominated by the γ-proteobacteria. The presence of culturable P aeruginosa in a sample had no significant effect on either community composition or structure. However, Principal Coordinate Analysis indicated that the presence of mPA in clinical culture was associated with a different community structure. Though there were no significant differences between the diversity of mPA positive samples (p>0.15), there were significant differences in the abundance of particular species; Achromobacter spp. and Pseudomonas spp. increased in relative abundance (p<0.001 in each case) and Haemophilus spp., Stenotrophomonas spp. and Staphylococcus spp. decreased in mPA positive vs mPA negative samples (p<0.001 in each case).
Conclusions Deep sequencing of sputum samples from CF and BX patients revealed a relationship between culture positivity for mPA and the presence of other known pathogens such as Achromobacter spp., suggesting an alternative mechanism for worse outcomes in these patients. This pilot has been extended to a larger cohort of 120 patients in order to confirm the result and data will be presented at the meeting.