Trans-envelope multidrug efflux pumps of Gram-negative bacteria and their synergism with the outer membrane barrier

Zgurskaya, Helen I.; Rybenkov, Valentin V.; Krishnamoorthy, Ganesh; Leus, Inga V.

doi:10.1016/j.resmic.2018.02.002

Cited by 51 publications

(36 citation statements)

References 74 publications

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“…Where on the AcrB protein the EPMs bind remains unknown. EPM35 and EPM43 potentiate multiple AcrB substrates, suggesting they bind in the hydrophobic trap [ 77 , 78 ]. Alternatively, the EPMs may bind outside of the substrate pocket and, for instance, disrupt AcrB folding, localization or interactions with AcrA.…”

Section: Discussionmentioning

confidence: 99%

A cell-based infection assay identifies efflux pump modulators that reduce bacterial intracellular load

Reens

Crooks

et al. 2018

PLoS Pathog

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Bacterial efflux pumps transport small molecules from the cytoplasm or periplasm outside the cell. Efflux pump activity is typically increased in multi-drug resistant (MDR) pathogens; chemicals that inhibit efflux pumps may have potential for antibiotic development. Using an in-cell screen, we identified three efflux pump modulators (EPMs) from a drug diversity library. The screening platform uses macrophages infected with the human Gram-negative pathogen Salmonella enterica (Salmonella) to identify small molecules that prevent bacterial replication or survival within the host environment. A secondary screen for hit compounds that increase the accumulation of an efflux pump substrate, Hoechst 33342, identified three small molecules with activity comparable to the known efflux pump inhibitor PAβN (Phe-Arg β-naphthylamide). The three putative EPMs demonstrated significant antibacterial activity against Salmonella within primary and cell culture macrophages and within a human epithelial cell line. Unlike traditional antibiotics, the three compounds did not inhibit bacterial growth in standard microbiological media. The three compounds prevented energy-dependent efflux pump activity in Salmonella and bound the AcrB subunit of the AcrAB-TolC efflux system with KDs in the micromolar range. Moreover, the EPMs display antibacterial synergy with antimicrobial peptides, a class of host innate immune defense molecules present in body fluids and cells. The EPMs also had synergistic activity with antibiotics exported by AcrAB-TolC in broth and in macrophages and inhibited efflux pump activity in MDR Gram-negative ESKAPE clinical isolates. Thus, an in-cell screening approach identified EPMs that synergize with innate immunity to kill bacteria and have potential for development as adjuvants to antibiotics.

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Section: Discussionmentioning

confidence: 99%

A cell-based infection assay identifies efflux pump modulators that reduce bacterial intracellular load

Reens

Crooks

et al. 2018

PLoS Pathog

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“…In contrast to these strains, clinical MDR isolates evolve from diverse genetic backgrounds, acquire specific antibiotic-resistance determinants, upregulate various efflux pumps, and diminish outer membrane permeability. All these changes, as well as additional poorly characterized physiological differences, can influence the impact of efflux [[10], [11], [12]].…”

Section: Introductionmentioning

confidence: 99%

Quantitative contribution of efflux to multi-drug resistance of clinical Escherichia coli and Pseudomonas aeruginosa strains

et al. 2019

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Background Efflux pumps mediate antimicrobial resistance in several WHO critical priority bacterial pathogens. However, most available data come from laboratory strains. The quantitative relevance of efflux in more relevant clinical isolates remains largely unknown. Methods We developed a versatile method for genetic engineering in multi-drug resistant (MDR) bacteria, and used this method to delete tolC and specific antibiotic-resistance genes in 18 representative MDR clinical E. coli isolates. We determined efflux activity and minimal inhibitory concentrations for a diverse set of clinically relevant antibiotics in these mutants. We also deleted oprM in MDR P. aeruginosa strains and determined the impact on antibiotic susceptibility. Findings tolC deletion abolished detectable efflux activity in 15 out of 18 tested E. coli strains, and modulated antibiotic susceptibility in many strains. However, all mutant strains retained MDR status, primarily because of other, antibiotic-specific resistance genes. Deletion of oprM altered antibiotic susceptibility in a fraction of clinical P. aeruginosa isolates. Interpretation Efflux modulates antibiotic resistance in clinical MDR isolates of E. coli and P. aeruginosa . However, when other antimicrobial-resistance mechanisms are present, inhibition of MDR efflux pumps alone is often not sufficient to restore full susceptibility even for antibiotics with a dramatic impact of efflux in laboratory strains. We propose that development of novel antibiotics should include target validation in clinical MDR isolates. Fund Innovative Medicines Initiative of European Union and EFPIA, Schweizerischer Nationalfonds, Swiss National Research Program 72, EU Marie Skłodowska-Curie program. The funders played no role in design, data collection, data analysis, interpretation, writing of the report, and in the decision to submit the paper for publication.

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“…Furthermore, sub-lethal concentrations of hydrogen peroxide did not induce transcription of macAB after short-term in vitro exposure ( Fig 2A ) or in time course assays with the 4/74 macAB :: lacZY transcriptional reporter strain ( S3B Fig ). Poly-specificity allows efflux pumps to take on various functional roles since net cellular resistance to toxic compounds results from redundancies in efflux pumps and dynamic properties of the outer membrane [ 77 ]. Given the poly-specificity of efflux channels like MacAB-TolC, it is likely that, in addition to antimicrobial peptides, some bacterially-generated molecules are exported to directly quench reactive oxygen species (ROS), as proposed by Bogomolnaya and colleagues [ 38 , 78 ].…”

Section: Discussionmentioning

confidence: 99%

Genetic variation in the MacAB-TolC efflux pump influences pathogenesis of invasive Salmonella isolates from Africa

Honeycutt

Wenner

et al. 2020

PLoS Pathog

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The various sub-species of Salmonella enterica cause a range of disease in human hosts. The human-adapted Salmonella enterica serovar Typhi enters the gastrointestinal tract and invades systemic sites to cause enteric (typhoid) fever. In contrast, most non-typhoidal serovars of Salmonella are primarily restricted to gut tissues. Across Africa, invasive non-typhoidal Salmonella (iNTS) have emerged with an ability to spread beyond the gastrointestinal tract and cause systemic bloodstream infections with increased morbidity and mortality. To investigate this evolution in pathogenesis, we compared the genomes of African iNTS isolates with other Salmonella enterica serovar Typhimurium and identified several macA and macB gene variants unique to African iNTS. MacAB forms a tripartite efflux pump with TolC and is implicated in Salmonella pathogenesis. We show that macAB transcription is upregulated during macrophage infection and after antimicrobial peptide exposure, with macAB transcription being supported by the PhoP/Q two-component system. Constitutive expression of macAB improves survival of Salmonella in the presence of the antimicrobial peptide C18G. Furthermore, these macAB variants affect replication in macrophages and influence fitness during colonization of the murine gastrointestinal tract. Importantly, the infection outcome resulting from these macAB variants depends upon both the Salmonella Typhimurium genetic background and the host gene Nramp1, an important determinant of innate resistance to intracellular bacterial infection. The variations we have identified in the MacAB-TolC efflux pump in African iNTS may reflect evolution within human host populations that are compromised in their ability to clear intracellular Salmonella infections.

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Trans-envelope multidrug efflux pumps of Gram-negative bacteria and their synergism with the outer membrane barrier

Cited by 51 publications

References 74 publications

A cell-based infection assay identifies efflux pump modulators that reduce bacterial intracellular load

A cell-based infection assay identifies efflux pump modulators that reduce bacterial intracellular load

Quantitative contribution of efflux to multi-drug resistance of clinical Escherichia coli and Pseudomonas aeruginosa strains

Genetic variation in the MacAB-TolC efflux pump influences pathogenesis of invasive Salmonella isolates from Africa

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