Marcy l’Etoile, France) and these were incubated twice overnight at 35°C. Colonies were identified using the MALDI-TOF and antibiotic susceptibility was determined with the VITEK®2. To determine the presence of highly resistant Acinetobacter calcoaceticus-baumannii complex (A. baumannii), the swab was placed in a TSB with 2 mg/L ceftazidime and 50 mg/L vancomycin, and incubated overnight at 35°C. From the broth, a MacConkey agar and a ChromID CarbaSmart agar were inoculated, and incubated twice overnight at 35°C. Suspected colonies were identified using the MALDI-TOF and antibiotic susceptibility was determined with the VITEK®2. To confirm the presence of highly resistant A. baumannii, an Etest (usually for meropenem and imipenem) or disk diffusion was performed, which was decided by the supervising clinical microbiologist. Universal screening samples Samples were taken with flocked swabs (Copan). Nasal samples were screened for MRSA, and perianal samples were screened for VRE, highly resistant Pseudomonas aeruginosa, highly resistant A. baumannii, carbapenemase-producing Enterobacterales (CPE), and ESBLE. Nasal samples were placed in the accompanying 2mL 2.5% NaCl TSB medium (Copan). Of the TSB medium, 800 µL was pipetted in a 6.5% NaCl TSB and incubated for 24 hours at 35°C. A nuc gene PCR was performed to identify the presence of S. aureus using established procedures. When the PCR was positive, a blood agar (BD diagnostics, Sparks, USA) was inoculated and incubated twice overnight at 35°C. Colonies were identified using the MALDI-TOF. To determine beta-lactam antibiotic resistance, a cefoxitin disk diffusion (30 µg; Oxoid, Basingstoke, UK) was performed. A growth inhibition zone of <22mm after 18 to 24 hours was considered resistant. For cefoxitin-resistant isolates, a multiplex PCR to detect mecA and mecC genes was performed using established procedures. All MRSA strains were stored in -80°C. Perianal samples were placed in the accompanying 1 mL Amies medium ((e-Swabs (Copan)). Of the Amies medium, 250µL was pipetted in an Enterococcosel broth with 8 mg/L amoxicillin, and 250µL in a TSB with 50 mg/L vancomycin. From the amoxicillin broth, a BrillianceTM VRE was inoculated and incubated twice overnight at 35°C to screen for VRE. From the vancomycin broth, a ChromID CarbaSmart plate was inoculated on both sides and incubated twice overnight at 35°C to screen for CPE, highly resistant P. aeruginosa, and highly resistant A. baumannii. Additionally, a BrillianceTM ESBL agar plate was inoculated from the vancomycin broth, to screen for ESBL-E, highly resistant P. aeruginosa, and highly resistant A. baumannii. All colonies were identified to species level using the MALDI-TOF. For suspected VRE and ESBL-E, based on growth on the BrillianceTM VRE or ESBL agar plate, respectively, antibiotic susceptibility was determined with the VITEK®2. For suspected carbapenemase-producing bacteria, based on growth on the ChromID Carba Smart, a PCR was performed to detect blaVIM, blaIMP, blaNDM, blaKPC and blaOXA-48-like genes using established procedures. For isolates that were negative for these carbapenemase genes, a CIM test was performed All identified colonies were stored at -80°C. 2 93 Comparing universal screening and universal risk-assessment for MDRO
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