Modifiable Risk Factors for the Emergence of Resistance to Pseudomonas aeruginosa

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Project Summary Carbapenem-resistant Pseudomonas aeruginosa (CR-P. aeruginosa) contributes to significant mortality in hospitalized patients. Only four β-lactam agents with activity against CR-P. aeruginosa are available in the US, of which ceftolozane-tazobactam (C-T) is the most frequently used to treat CR-P. aeruginosa. Pre-clinical investigations indicate C-T had activity against >90% of P. aeruginosa isolates; after clinical use, reports of P. aeruginosa that initially tested susceptible to C-T but became resistant during therapy emerged. Our preliminary data indicate that amongst 28 consecutive patients infected with CR-P. aeruginosa with isolates initially susceptible to C-T, 50% of patients had subsequent P. aeruginosa isolates with > 4-fold increase in C-T minimum inhibitory concentrations (MICs) up to 30 days after C-T exposure. We identified genetic markers contributing to resistance through whole genome sequencing (WGS) – notably mutations in the ampC-ampR region and in PBP3. Alarmingly, 86% of P. aeruginosa isolates initially susceptible to another novel antibiotic, ceftazidime-avibactam (C-A), exhibited resistance to C-A after C-T exposure - in the absence of C-A exposure. This is concerning as the emergence of resistance to one novel agent could eliminate the few remaining treatment options. We explored modifiable risk factors that could reduce the frequency of C-T resistance and found that 30% of patients who received C-T over 1 hour had subsequent C-T resistant isolates, whereas no patients who received the drug over 3 hours developed resistant isolates. We would like to repeat this exploratory work and include experimental validation studies in a cohort of 260 patients across seven hospitals who have paired P. aeruginosa clinical isolates stored at -80°C and were prescribed C-T. In Aim 1, WGS will identify the genetic loci associated with C-T resistance comparing paired CR-P. aeruginosa isolates from the same patient before and up to 30 days after C-T exposure. Validation will occur through cloning and site-directed mutagenesis work. Broth microdilution will determine susceptibility testing results for the three other β-lactams with activity against CR-P. aeruginosa to quantify the emergence of cross-resistance. In Aim 2, we will capitalize on diverse C-T administration strategies at participating sites and develop a machine learning derived decision tree to inform clinicians how to prescribe C-T most effectively to reduce the likelihood of acquired resistance. The following are some risk factors that will be investigated for potential inclusion: (1) infusion of C-T over 3 hours, (2) high-dose C-T, (3) combination antibiotic therapy, (4) dialysis dependency, (5) source of infection and source control, (6) number of days of C-T exposure. This work will (1) inform future drug development by identifying areas of the bacterial chromosome highly prone to resistance and in need of additional reinforcement and (2) identify modifiable risk factors that can be used by clinicians to enable novel antibiotics to remain effective treatment options for extended periods of time.