Isolation Procedure for CP E. coli from Caeca Samples under Review towards an Increased Sensitivity

Abstract

Due to the increasing reports of carbapenemase-producing Enterobacteriaceae (CPE) from livestock in recent years, the European Reference Laboratory for Antimicrobial Resistances (EURLAR) provided a protocol for their recovery from caecum and meat samples. This procedure exhibited limitations for the detection of CPE with low carbapenem MIC values. Therefore, it was modified by a second, selective enrichment in lysogeny broth with cefotaxime (CTX 1 mg/L) and with meropenem (MEM 0.125 mg/L) at 37 C under microaerophilic conditions. By Real-time PCR, these enrichments are pre-screened for the most common carbapenemase genes. Another adaptation was the use of in-house prepared MacConkey agar with MEM and MEM+CTX instead of commercial selective agar. According to the EURL-method, we achieved 100% sensitivity and specificity using the in-house media instead of commercial agar, which decreased the sensitivity to ~75%. Comparing the method with and without the second enrichment, no substantial influence on sensitivity and specificity was detected. Nevertheless, this enrichment has simplified the CPE-isolation regarding the accompanying microbiota and the separation of putative colonies. In conclusion, the sensitivity of the method can be increased with slight modifications. ; This work was financially supported by a grant of the German Federal Institute for Risk Assessment (43-001 and 43-002). The work of Natalie Pauly was supported by the European Joint Project (EJP) IMPART funded by the European Union's Horizon 2020 research and innovation programme under Grant Agreement No 773830. The work of Stefan Schwarz is supported by the German Federal Ministry of Education and Research (BMBF) under project number and 01KI2009D as part of the Research Network Zoonotic Infectious Diseases.