Mutant prevention concentrations and phenotypic and genomic profiling of first-step resistance mechanisms to classical and novel β-lactams in <i>Pseudomonas aeruginosa</i>

A growing number of novel antipseudomonal β-lactams have been introduced in recent years, but the emergence of resistance is still a major concern in the treatment of <i>Pseudomonas aeruginosa</i> infections. Here, we compared the mutant prevention concentrations (MPCs) and the nature of first-step resistant mutants to classical and novel β-lactams in <i>P. aeruginosa</i>. MPCs were determined in duplicate experiments for ceftazidime, ceftazidime/avibactam, ceftolozane/tazobactam, imipenem, imipenem/relebactam, meropenem, meropenem/vaborbactam, aztreonam, aztreonam/avibactam, and cefiderocol in PAO1, PAOMS (Δ<i>mutS</i>), and three extensively drug-resistant (XDR) clinical strains belonging to high-risk clones ST111, ST175, and ST235. Four mutants per strain and antibiotic, obtained from the highest concentration showing growth, were characterized through the determination of the susceptibility profiles and whole genome sequencing. Imipenem/relebactam presented the lowest MPC values, followed by ceftolozane/tazobactam. Overall, the MICs of the mutants were consistent with the antibiotic selection concentration, except for cefiderocol, which were much lower. MPCs were lower for ceftazidime/avibactam and imipenem/relebactam than those of the corresponding β-lactam alone. In contrast, MPCs of meropenem ± vaborbactam and aztreonam ± avibactam were identical in most strains. Ceftolozane/tazobactam and ceftazidime/avibactam derivatives presented mutations in <i>ampC</i>, <i>galU</i>, <i>cpxRS,</i> and/or in <i>bla_OXA-2</i> when present in the parent strain (ST235). Cefiderocol mutants were mainly defective in iron-uptake systems, particularly PiuA/DC. All carbapenems had oprD as the first-step mechanism. Imipenem/relebactam, meropenem ± vaborbactam, and aztreonam ± avibactam selected mutations frequently included efflux pumps and regulators. Imipenem ± relebactam also selected <i>aroB</i> mutations. This work first describes the MPCs and first-step resistance mechanisms for classical and novel β-lactams in <i>P. aeruginosa</i>. The identified shared and differential resistance development patterns between the available classical and novel β-lactams should be helpful to guide treatment strategies for XDR <i>P. aeruginosa</i> infections.