Optimizing a culture-based approach to detect intestinal carriage of carbapenem-resistant Pseudomonas aeruginosa in humans.

INTRODUCTION AND OBJECTIVE: Carbapenem-resistant Pseudomonas aeruginosa (CRPA) causes severe and potentially life-threatening infections in hospitalized patients with mortality rates exceeding 40%. To detect intestinal CRPA carriage in humans for surveillance purposes or to prevent spread and outbreaks in hospitals, a highly-sensitive culture method is needed. We aimed to develop such a highly-sensitive method, that would be feasible in laboratories with limited resources as well. METHODS: This study used seven well-defined CRPA strains belonging to high-risk clones, including six strains with a carbapenemase gene and one OprD-deficient strain. We applied a stepwise approach that 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 using human stools. RESULTS: The two most sensitive enrichment broths were tryptic soy broth (TSB) with 2 mg/L vancomycin and TSB with 2 mg/L vancomycin and 2 mg/L imipenem and the most sensitive selective agar plates were Pseudomonas isolation agar (PIA; Becton Dickinson), PIA (Sigma-Aldrich), and M-PA-C (Becton Dickinson). After the spiking experiment, the best method for detecting CRPA based on sensitivity and selectivity was the combination of TSB with 2 mg/L vancomycin and 2 mg/L imipenem as an enrichment broth for overnight incubation, followed by subculturing the broth on M-PA-C agar plate. CONCLUSION: We have thus developed a highly-sensitive selective method to detect intestinal CRPA carriage in humans that can be applied in limited-resource laboratories. This may contribute to overall efforts to control CRPA.