Dissemination of the blaCTX-M-15 gene among Enterobacteriaceae via outer membrane vesicles.

Background: Bacterial outer membrane vesicles (OMVs) are an emerging source of antibiotic resistance transfer but their role in the spread of the bla(CTX-M-15) gene encoding the most frequent CTX-M ESBL in Enterobacteriaceae is unknown. Objectives: To determine the presence of bla(CTX-M-15) and other antibiotic resistance genes in OMVs of the CTX-M-15-producing MDR Escherichia coli O104:H4 outbreak strain and the ability of these OMVs to spread these genes among Enterobacteriaceae under different conditions. Methods: OMV-borne antibiotic resistance genes were detected by PCR; OMV-mediated transfer of bla(CTX-M-15) and the associated bla(TEM-1) was quantified under laboratory conditions, simulated intraintestinal conditions and under ciprofloxacin stress; resistance to antibiotics and the ESBL phenotype were determined by the CLSI disc diffusion methods and the presence of pESBL by plasmid profiling and Southern blot hybridization. Results: E. coli O104:H4 OMVs carried bla(CTX-M-15) and bla(TEM-1) located on the pESBL plasmid, but not chromosomal antibiotic resistance genes. The OMVs transferred bla(CTX-M-15), bla(TEM-1) and the associated pESBL into Enterobacteriaceae of different species. The frequencies of the OMV-mediated transfer were significantly increased under simulated intraintestinal conditions and under ciprofloxacin stress when compared with laboratory conditions. The 'vesiculants' (i.e. recipients that received the bla(CTX-M-15)-and bla(TEM-1)-harbouring pESBL via OMVs) acquired resistance to cefotaxime, ceftazidime and cefpodoxime and expressed the ESBL phenotype. They were able to further spread pESBL and the bla(CTX-M-15) and bla(TEM-1) genes via OMVs. Conclusions: OMVs are efficient vehicles for dissemination of the bla(CTX-M-15) gene among Enterobacteriaceae and may contribute to bla(CTX-M-15) transfer in the human intestine.