Project description:The spread of antimicrobial resistance (AMR), coupled with the decline in antibiotic development, has become a major public health concern. Recent studies estimate that around 700,000 people die each year from infections caused by multidrug-resistant (MDR) bacteria. This led the WHO to publish the ESKAPEE list of high priority pathogens for AMR, namely Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter spp. and Escherichia coli. Among these, Gram-negative bacteria (K. pneumoniae, A. baumannii, P. aeruginosa, Enterobacter spp., and E. coli) are particularly overrepresented. This is mainly due to their high propensity to develop multiple resistance mechanisms, in addition to their intrinsic resistance to many antimicrobials, which is due to their membrane composition and the expression of broad-spectrum efflux pumps. One strategy to combat such AMR is the use of drug enhancers that are able to restore the antibacterial activity of poorly active antibiotics. In this context, we demonstrated that the polyamino-isoprenyl enhancer, NV716, efficiently potentiates the antibacterial activity of two families of multi-target Ser/Cys-based enzyme inhibitors, namely the oxadiazolone derivatives (OX) and the Cyclipostins and Cyclophostin analogs (CyC), against Enterobacter cloacae, while remaining inactive against other Gram-negative bacteria. We confirmed that NV716 potentiates some OX & CyC compounds by permeabilizing the outer membrane and thus by increasing the inhibitor accumulation as shown by fluorescence confocal microscopy. By using bio-orthogonal click-chemistry activity-based protein profiling (CC-ABPP) approach coupled to proteomic analysis, we also identified the target proteins of the best OX & CyC inhibitors from E. cloacae lysate, thereby confirming their multi-target nature. Interestingly, 6 of the latter proteins were also captured via CC-ABPP in P. aeruginosa lysate, and are highly conserved in all Gram-negative bacteria. These results provide proof of concept that both OX & CyC, if successfully potentiated, could be used against a wide range of ESKAPEE Gram-negative bacteria.
Project description:Recently, we have reported on a highly drug-resistant carbapenemase-producing isolate of Enterobacter cloacae (Nepal et al., Virulence. 2018; 9: 1377-1389). In the present study, we asked the question whether and, if so, how this isolate responds to a sub-inhibitory challenge with the antibiotic imipenem. To answer this question, we applied a SILAC proteomics approach that allowed the quantification of changes in the relative abundance of bacterial protein in response to imipenem. The results show that the investigated E. cloacae isolate mounts a highly specific response to counteract the detrimental effects of imipenem.
