Synergistic effects of inhaled aztreonam plus tobramycin on hypermutable cystic fibrosis Pseudomonas aeruginosa isolates in a dynamic biofilm model evaluated by mechanism-based modelling and whole genome sequencing

Hypermutable Pseudomonas aeruginosa strains are highly prevalent in chronic lung infections of patients with cystic fibrosis (CF). Acute exacerbations of these infections have limited treatment options. This study aimed to investigate inhaled aztreonam and tobramycin against clinical hypermutable P. aeruginosa strains using the CDC dynamic in vitro biofilm reactor (CBR), mechanism-based mathematical modelling (MBM) and genomic studies. Two CF multidrug-resistant strains were investigated in a 168 h CBR (n = 2 biological replicates). Regimens were inhaled aztreonam (75 mg 8-hourly) and tobramycin (300 mg 12-hourly) in monotherapies and combination. The simulated pharmacokinetic profiles of aztreonam and tobramycin (t 1/2 = 3 h) were based on published lung fluid concentrations in patients with CF. Total viable and resistant counts were determined for planktonic and biofilm bacteria. MBM of total and resistant bacterial counts and whole genome sequencing were completed. Both isolates showed reproducible bacterial regrowth and resistance amplification for the monotherapies by 168 h. The combination performed synergistically, with minimal resistant subpopulations compared to the respective monotherapies at 168 h. Mechanistic synergy appropriately described the antibacterial effects of the combination regimen in the MBM. Genomic analysis of colonies recovered from monotherapy regimens indicated noncanonical resistance mechanisms were likely responsible for treatment failure. The combination of aztreonam and tobramycin was required to suppress the regrowth and resistance of planktonic and biofilm bacteria in all biological replicates of both hypermutable multidrug-resistant P. aeruginosa CF isolates. The developed MBM could be utilised for future investigations of this promising inhaled combination.