Development of a highly-sensitive method to detect the carriage of carbapenem-resistant Pseudomonas aeruginosa in humans

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) causes severe and potentially life-threatening infections in hospitalized patients with mortality rates of more than 40%. To detect CRPA carriage in humans for surveillance purposes or to prevent spread and outbreaks in hospitals, a highly-sensitive culture method for CRPA carriage in humans is needed. We aimed to develop such a highly-sensitive method, that would be feasible in laboratories with limited resources. In this study, seven well-defined CRPA strains belonging to high-risk clones were used, including one CRPA without a carbapenemase gene and six carbapenem-resistant isolates with carbapenemase genes. We applied a stepwise approach wherein we included four enrichment broths and eight Pseudomonas aeruginosa-selective culture media. Spiking experiments were performed to further evaluate the combination of the most sensitive enrichment broths and selective agar plates in human samples. The two most sensitive enrichments broths were TSB-vancomycin and TSB-vancomycin with 2 mg/L imipenem and the most sensitive selective agar plates were Pseudomonas isolation agar Becton Dickinson, Pseudomonas isolation agar Sigma-Aldrich, and M-PA-C (Becton Dickinson). After the spiking experiment, the best method for detecting CRPA based on the sensitivity and the selectivity was the combination of TSB-vancomycin with 2 mg/L imipenem as an enrichment broth for overnight incubation, followed by subculturing the broth on M-PA-C agar plate. We have thus developed a highly-sensitive selective method to detect CRPA carriage in humans, which can also be applied in limited-resource laboratories. This may contribute to an overall effort to control CRPA.