RND efflux pumps in Gram-negative bacteria; regulation, structure and role in antibiotic resistance.

Rresistance-nodulation-division (RND) efflux pumps in Gram-negative bacteria remove multiple, structurally distinct classes of antimicrobials from inside bacterial cells therefore directly contributing to multidrug resistance. There is also emerging evidence that many other mechanisms of antibiotic resistance rely on the intrinsic resistance conferred by RND efflux. In addition to their role in antibiotic resistance, new information has become available about the natural role of RND pumps including their established role in virulence of many Gram-negative organisms.

Beyond Antimicrobial Resistance: Evidence for a Distinct Role of the AcrD Efflux Pump in Salmonella Biology.

Unlabelled: For over 20 years, bacterial multidrug resistance (MDR) efflux pumps have been studied because of their impact on resistance to antimicrobials. However, critical questions remain, including why produce efflux pumps under non-antimicrobial treatment conditions, and why have multiple pumps if their only purpose is antimicrobial efflux? Salmonella spp. possess five efflux pump families, including the resistance-nodulation-division (RND) efflux pumps.

How to Measure Export via Bacterial Multidrug Resistance Efflux Pumps.

Bacterial multidrug resistance (MDR) efflux pumps are an important mechanism of antibiotic resistance and are required for many pathogens to cause infection. They are also being harnessed to improve microbial biotechnological processes, including biofuel production. Therefore, scientists of many specialties must be able to accurately measure efflux activity.

AcrB drug-binding pocket substitution confers clinically relevant resistance and altered substrate specificity.

The incidence of multidrug-resistant bacterial infections is increasing globally and the need to understand the underlying mechanisms is paramount to discover new therapeutics. The efflux pumps of Gram-negative bacteria have a broad substrate range and transport antibiotics out of the bacterium, conferring intrinsic multidrug resistance (MDR). The genomes of pre- and posttherapy MDR clinical isolates of Salmonella Typhimurium from a patient that failed antibacterial therapy and died were sequenced.

Molecular mechanisms of antibiotic resistance.

Antibiotic-resistant bacteria that are difficult or impossible to treat are becoming increasingly common and are causing a global health crisis. Antibiotic resistance is encoded by several genes, many of which can transfer between bacteria. New resistance mechanisms are constantly being described, and new genes and vectors of transmission are identified on a regular basis.

Multidrug efflux pumps in Gram-negative bacteria and their role in antibiotic resistance.

Gram-negative bacteria express a plethora of efflux pumps that are capable of transporting structurally varied molecules, including antibiotics, out of the bacterial cell. This efflux lowers the intracellular antibiotic concentration, allowing bacteria to survive at higher antibiotic concentrations. Overexpression of some efflux pumps can cause clinically relevant levels of antibiotic resistance in Gram-negative pathogens.

Expression of homologous RND efflux pump genes is dependent upon AcrB expression: implications for efflux and virulence inhibitor design.

Objectives: Enterobacteriaceae have multiple efflux pumps that confer intrinsic resistance to antibiotics. AcrB mediates clinically relevant multidrug resistance and is required for virulence and biofilm formation, making it an attractive target for the design of inhibitors. The aim of this study was to assess the viability of single transporters as a target for efflux inhibition using Salmonella Typhimurium as the model pathogen.

Redundancy in the periplasmic adaptor proteins AcrA and AcrE provides resilience and an ability to export substrates of multidrug efflux.

Objectives: The components of the AcrAB-TolC efflux pump function as a tripartite efflux system conferring resistance to multiple antibiotics and the individual components can also function in conjunction with other efflux pumps. This study aimed to establish whether redundancy exists between the homologous periplasmic adaptor proteins (PAPs) AcrA and AcrE and to measure the impact of this redundancy on antimicrobial resistance and the potential efficacy of inhibitor molecules.

Methods: The acrE gene was inactivated in Salmonella enterica serovar Typhimurium SL1344 and a ΔacrA mutant by insertion of the aph gene.

RamA, which controls expression of the MDR efflux pump AcrAB-TolC, is regulated by the Lon protease.

Objectives: RamA regulates the AcrAB-TolC multidrug efflux system. Using Salmonella Typhimurium, we investigated the stability of RamA and its impact on antibiotic resistance.

Methods: To detect RamA, we introduced ramA::3XFLAG::aph into plasmid pACYC184 and transformed this into Salmonella Typhimurium SL1344ramA::cat and lon::aph mutants.

Choice of bacterial growth medium alters the transcriptome and phenotype of Salmonella enterica Serovar Typhimurium.

The type of bacterial culture medium is an important consideration during design of any experimental protocol. The aim of this study was to understand the impact of medium choice on bacterial gene expression and physiology by comparing the transcriptome of Salmonella enterica SL1344 after growth in the widely used LB broth or the rationally designed MOPS minimal medium. Transcriptomics showed that after growth in MOPS minimal media, compared to LB, there was increased expression of 42 genes involved in amino acid synthesis and 23 genes coding for ABC transporters.

SadA, a trimeric autotransporter from Salmonella enterica serovar Typhimurium, can promote biofilm formation and provides limited protection against infection.

Salmonella enterica is a major cause of morbidity worldwide and mortality in children and immunocompromised individuals in sub-Saharan Africa. Outer membrane proteins of Salmonella are of significance because they are at the interface between the pathogen and the host, they can contribute to adherence, colonization, and virulence, and they are frequently targets of antibody-mediated immunity. In this study, the properties of SadA, a purported trimeric autotransporter adhesin of Salmonella enterica serovar Typhimurium, were examined.

Periplasmic adaptor protein AcrA has a distinct role in the antibiotic resistance and virulence of Salmonella enterica serovar Typhimurium.

Objectives: AcrA can function as the periplasmic adaptor protein (PAP) in several RND tripartite efflux pumps, of which AcrAB-TolC is considered the most important. This system confers innate multiple antibiotic resistance. Disruption of acrB or tolC impairs the ability of Salmonella Typhimurium to colonize and persist in the host.

Structure, function and inhibition of RND efflux pumps in Gram-negative bacteria: an update.

Resistance nodulation division efflux systems have a major role in both intrinsic and acquired multi-drug resistance in Gram-negative bacteria. The recent structure of an assembled tripartite system, AcrAB-TolC, revealed that AcrB is docked onto TolC, which remains in an open state once part of the assembled complex and three AcrA molecules complete the structure. This is in contrast to data for the MexAB-OprM system of P.

Epidemiological evidence for the role of the hemoglobin receptor, hmbR, in meningococcal virulence.

The distribution of the hemoglobin receptor gene (hmbR) was investigated among disease and carriage Neisseria meningitidis isolates, revealing that the gene was detected at a significantly higher frequency among disease isolates than among carriage isolates. In isolates without hmbR, the locus was occupied by the cassettes exl2 or exl3 or by a "pseudo hmbR" gene, designated exl4. The hmbR locus exhibited characteristics of a pathogenicity island in published genomes of N.

The global consequence of disruption of the AcrAB-TolC efflux pump in Salmonella enterica includes reduced expression of SPI-1 and other attributes required to infect the host.

The mechanisms by which RND pumps contribute to pathogenicity are currently not understood. Using the AcrAB-TolC system as a paradigm multidrug-resistant efflux pump and Salmonella enterica serovar Typhimurium as a model pathogen, we have demonstrated that AcrA, AcrB, and TolC are each required for efficient adhesion to and invasion of epithelial cells and macrophages by Salmonella in vitro. In addition, AcrB and TolC are necessary for Salmonella to colonize poultry.