Dataset Information

The role of chemotaxis and efflux pumps on nitrate reduction in the toxic regions of a ciprofloxacin concentration gradient.

ABSTRACT: Spatial concentration gradients of antibiotics are prevalent in the natural environment. Yet, the microbial response in these heterogeneous systems remains poorly understood. We used a microfluidic reactor to create an artificial microscopic ecosystem that generates diffusive gradients of solutes across interconnected microenvironments. With this reactor, we showed that chemotaxis toward a soluble electron acceptor (nitrate) allowed Shewanella oneidensis MR-1 to inhabit and sustain metabolic activity in highly toxic regions of the antibiotic ciprofloxacin (>80× minimum inhibitory concentration, MIC). Acquired antibiotic resistance was not observed for cells extracted from the reactor, so we explored the role of transient adaptive resistance by probing multidrug resistance (MDR) efflux pumps, ancient elements that are important for bacterial physiology and virulence. Accordingly, we constructed an efflux pump deficient mutant (∆mexF) and used resistance-nodulation-division (RND) efflux pump inhibitors (EPIs). While batch results showed the importance of RND efflux pumps for microbial survival, microfluidic studies indicated that these pumps were not necessary for survival in antibiotic gradients. Our work contributes to an emerging body of knowledge deciphering the effects of antibiotic spatial heterogeneity on microorganisms and highlights differences of microbial response in these systems versus well-mixed batch conditions.

SUBMITTER: Alcalde RE

PROVIDER: S-EPMC8443623 | biostudies-literature | 2021 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The role of chemotaxis and efflux pumps on nitrate reduction in the toxic regions of a ciprofloxacin concentration gradient.

Alcalde Reinaldo E RE Dundas Christopher M CM Dong Yiran Y Sanford Robert A RA Keitz Benjamin Keith BK Fouke Bruce W BW Werth Charles J CJ

The ISME journal 20210429 10

Spatial concentration gradients of antibiotics are prevalent in the natural environment. Yet, the microbial response in these heterogeneous systems remains poorly understood. We used a microfluidic reactor to create an artificial microscopic ecosystem that generates diffusive gradients of solutes across interconnected microenvironments. With this reactor, we showed that chemotaxis toward a soluble electron acceptor (nitrate) allowed Shewanella oneidensis MR-1 to inhabit and sustain metabolic act ...[more]

PMID: 33927341

Similar Datasets

Project description:RND (Resistance-Nodulation-Division) family transporters are widespread especially among Gram-negative bacteria, and catalyze the active efflux of many antibiotics and chemotherapeutic agents. They have very large periplasmic domains, and form tripartite complexes with outer membrane channels and periplasmic adaptor proteins. AcrAB-TolC complex of Escherichia coli, which pumps out a very wide range of drugs, has been studied most intensively. Early studies showed that the transporter captures even those substrates that cannot permeate across the cytoplasmic membrane, such as dianionic beta-lactams, suggesting that the capture can occur from the periplasm. It was also suggested that the capture occurs from the cytoplasmic membrane/periplasm interface, because most substrates contain a sizable hydrophobic domain; however, this may simply be a reflection of the nature of the binding site within AcrB. Genetic studies of chimeric transporters showed that much of the substrate specificity is determined by their periplasmic domains. Biochemical studies with intact cells recently led to the determination of the kinetic constants of AcrB for some beta-lactams, and the result confirms the old prediction that AcrB is a rather slow pump. Reconstitution of purified AcrB and its relatives showed that the pump is a drug/proton antiporter, that AcrA strongly stimulates the activity of the pump, and that AcrB seems to have a highest affinity for conjugated bile salts. Structural study with mutants of the network of charged residues in the transmembrane domain showed that protonation here produced a far-reaching conformational change, which was found to be present in one of the protomers in the asymmetric crystal structure of the wild-type AcrB. The functional rotatory hypothesis then predicts that the drug bound in the periplasmic domain is extruded through this conformational change initiated by the protonation of one of the residues in the aforementioned network, an idea that was recently supported by disulfide cross-linking as well as by the behavior of linked AcrB protomers.

Project description:Multidrug resistance (MDR) efflux pumps are involved in bacterial intrinsic resistance to multiple antimicrobials. Expression of MDR efflux pumps can be either constitutive or transiently induced by various environmental signals, which are typically perceived by bacterial two-component systems (TCSs) and relayed to the bacterial nucleoid, where gene expression is modulated for niche adaptation. Here, we demonstrate that RstA/RstB, a TCS previously shown to control acid-induced and biofilm-related genes in Escherichia coli, confers resistance to multiple antibiotics in Pseudomonas fluorescens by directly regulating the MDR efflux pumps EmhABC and MexCD-OprJ. Moreover, we show that phosphorylation of the conserved Asp52 residue in RstA greatly enhances RstA-DNA interaction, and regulation of the multidrug resistance by RstA/RstB is dependent on the phosphorylation of the RstA Asp52 residue by RstB. Proteome analysis reveals RstA/RstB also positively regulates the efflux pump MexEF-OprN and enzymes involved in anaerobic nitrate respiration and pyoverdine biosynthesis. Our results suggest that, by coupling the expression of multiple efflux pumps and anaerobic nitrate respiration, RstA/RstB could play a role in defense against nitrosative stress caused by anaerobic nitrate respiration. IMPORTANCE Microenvironmental hypoxia typically increases bacterial multidrug resistance by elevating expression of multidrug efflux pumps, but the precise mechanism is currently not well understood. Here, we showed that the two-component system RstA/RstB not only positively regulated expression of several efflux pumps involved in multidrug resistance, but also promoted expression of enzymes involved in anaerobic nitrate respiration and pyoverdine biosynthesis. These results suggested that, by upregulating expression of efflux pumps and pyoverdine biosynthesis-related enzymes, RstA/RstB could play a role in promoting bacterial tolerance to hypoxia by providing protection against nitrosative stress.

Dataset Information

The role of chemotaxis and efflux pumps on nitrate reduction in the toxic regions of a ciprofloxacin concentration gradient.

Publications

The role of chemotaxis and efflux pumps on nitrate reduction in the toxic regions of a ciprofloxacin concentration gradient.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets