Tripartite resistance-nodulation-division (RND) efflux pumps confer multidrug resistance (MDR) in Gram-negative bacteria and are critical for many physiological functions including virulence and biofilm formation. The common laboratory strain ofE. coli,K-12 MG1655 has six recognised RND transporters participating in tripartite pump formation (AcrB, AcrD, AcrF, CusA, MdtBC, and MdtF). However, by studying >20,000E. coligenomes we show thatE. colibelonging to phylogroups B2, D, E, F and G, which are commonly associated with infection, possess an additional, seventh RND transporter, EefB. It is found in a five gene operon,eefRABCD,which also encodes a TetR family transcription factor, a periplasmic adapter protein, an outer membrane factor and major facilitator superfamily pump. In contrast,E. colifrom phylogroups A, B1 and C, generally containing environmental and commensal strains, do not encode the operon and instead encode an uncharacterised ORF,ycjD. In phylogroups where theeefRABCDoperon is present it was very highly conserved. In fact, conservation levels were comparable to that of the majorE. coliRND efflux system AcrAB-TolC, suggesting a critical biological function. Protein modelling shows that this pump is highly divergent from endogenousE. coliRND systems with unique structural features, while showing similarities to efflux systems found inPseudomonas aeruginosa. However, unlike other major RND efflux systems, EefABC does not appear to transport antimicrobials and instead may be important for infection or survival in the host environment.
Importance
Efflux pumps are molecular machines that export molecules out of bacterial cells. The efflux pumps belonging to the RND family are particularly important as they export antibiotics out of Gram-negative bacterial cells, contributing to antibiotic resistance. The important human pathogen,E. coli, has been previously reported to have six RND pumps. However, we show that phylogroups ofE. colicommonly associated with infection encode a seventh RND pump, EefABC which is highly conserved, suggesting an important biological function. While the function of EefABC inE. coliremains to be resolved, it does not seem to transport antimicrobial compounds. These findings are important because they reveal a new RND pump, potentially involved in virulence and survival in the host, that could represent a new therapeutic target. Additionally, it again shows that laboratory type strains of common bacterial pathogens are not representative of those that are infection causing.