Surface-associated, motile populations of Pseudomonas aeruginosa gene regulation by Staphylococcus aureus exoproducts

Pseudomonas aeruginosa and Staphylococcus aureus are often co-isolated in persistent infections. The goal of this study was to determine how secreted products from S. aureus affect gene expression in surface-associated P. aerguinosa undergoing emergent motility. Therefore, media salts control or S. aureus supernatant was added to agar plates at 25% total volume. P. aeruginosa was inoculated on the agar and gene expression was measured from the leading edge after 17 h incubation using RNA-seq. P. aeruginosa moving on the agar containing S. aureus supernatant had an upregulation in genes involved in intermediate metabolite uptake and a downregulation in heme biosynthesis, response to heat, Type III Secretion System, and aerobic respiration pathways. We collected RNA-seq samples from P. aeruginosa collected from the leading edge on 0.5% or 1.5% (w/v) agar plates containing 25% (v/v) of either media salts control or S. aureus supernatant. RNAprotect (Qiagen) was added to each sample before purification using the Total RNA Purification Plus Kit (Norgen). Genomic DNA was removed using Turbo DNase (Invitrogen) and rRNA was removed using riboPOOLs (siTOOLs Biotech). The cDNA libraries were generated using NEBNext Ultra II Directional Library Kit and amplified with NEBNext index primers (New England Biolabs). Sequencing was performed at the National Cancer Institute Center for Cancer Research Genomics Core.