Chromid mrsa agar plates

To determine transmission to HCW, all HCW on the outpatient departments were sampled periodically. This screening was performed before the implementation of the adjustment (0-measurement) and two times after the implementation, in 2008 and 2009. The target was a compliance of 90 percent of all HCW. Samples were directly plated on chromID S. aureus and chromID MRSA agar plates (bioMérieux, La Balme, France), and subsequently placed in a Mueller-Hinton (MH) broth supplemented with 6.5% sodium chloride. The overnight MH broth was subcultured onto both chromID S. aureus and chromID MRSA agar plates [12 ].
Consequences of eventual MRSA positive HCW were predefined. A history would be taken to identify known risk factors for MRSA carriage [1 ]. Also, it would be checked whether a MRSA positive patient with the same Spa type had been on the department in the period preceding the positive sample. Furthermore, the observed prevalence in the periodic screening will be compared with the prevalence of MRSA in HCW found by Wulf et al., being 0.15 percent [13 (link)]. If the percentage of MRSA positive HCW during periodic screening rounds will not exceed this percentage and no contact with MRSA positive patients with the same Spa type can be demonstrated, the adjustment in the S&D policy for outpatients would be considered successful.

In the laboratory, 100 mL of water samples were first pre-filtered using a sterilized gauze (PZN: 04046708, FESMED Verbandmittel GmbH, Frankenberg/Sa., Germany) to remove the fine particles. The pre-filtered water was free from any macro-particles, sediment and most fine particles. Thereafter, each sample was filtered using the EZ-Fit filtration system with 0.45 µm pore size filter membranes (merckmillipore, Darmstadt, Germany). After filtration, filter membranes were transferred to 10 mL TSB (Carl Roth GmbH, Karlsruhe, Germany) containing 2 µg/mL cefotaxime (VWR International, Darmstadt, Germany) followed by overnight incubation at 37 °C and shaking at 200 rpm. Depending on the turbidity level, dilution of the overnight cultures followed (up to 10,000 fold dilution). For each sample, 100 µL of the dilutions were plated on chromogenic media CHROMID CARB/OXA, CHROMID ESBL, CHROMID Colistin, and CHROMID MRSA agar plates (bioMérieux, Nürtingen, Germany), and incubated overnight at 37 °C. Putative antibiotic-resistant colonies of E. coli (red-purple colonies) and KEC (Klebsiella spp., Enterobacter spp., Citrobacter spp. (blue colonies)) were subcultivated until pure cultures were achieved. Single pure colonies were picked for further verification and characterization.

The extended laboratory procedure is described elsewhere [8 (link)]. In short, quantitative cultures were performed by serially diluting ESwab—medium and incubating on chromID S. aureus and chromID MRSA agar plates (BioMérieux, La Balme Les Grottes, France). The number of CFU was counted on both plates. Semi-quantitative cultures were performed with dry swabs on the same media, directly plated and after enrichment. Wet wipe samples were enriched in two consecutive media and subsequently cultured on blood and Brilliance MRSA agar plates (Oxoid, Basingstoke, UK).
All S. aureus strains were defined by green colonies on selective S. aureus agar in combination with a positive coagulase slide and DNase test. Methicillin susceptibility was tested using the cefoxitin disk diffusion method according to EUCAST standards [13 ], followed by a duplex PCR for the nuc and mecA genes as described previously [14 (link)].