scholarly journals Influence of theyjiL-mdtMGene Cluster on the Antibacterial Activity of Proline-Rich Antimicrobial Peptides Overcoming Escherichia coli Resistance Induced by the Missing SbmA Transporter System

2015 ◽  
Vol 59 (10) ◽  
pp. 5992-5998 ◽  
Author(s):  
Andor Krizsan ◽  
Daniel Knappe ◽  
Ralf Hoffmann
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Peptides ◽  
Random Mutagenesis ◽  
Major Facilitator Superfamily ◽  
Multiresistant Pathogens ◽  
Content Type ◽  
E Coli ◽  
Cytotoxic Compounds ◽  
Major Facilitator ◽  
Mfs Transporters

ABSTRACTIn view of increasing health threats from multiresistant pathogens, antimicrobial peptides (AMPs) and, specifically, proline-rich AMPs (PrAMPs) have been investigated in animal models. PrAMPs enter bacteria via the ABC transporter SbmA and inhibit intracellular targets. We used phage transduction (Tn10insertion) to screen by random mutagenesis for alternative uptake mechanisms for analogs of apidaecin 1b, a honeybee-derived PrAMP. All 24 apidaecin-resistant mutants had the Tn10insertion in thesbmAgene. ThesesbmA::Tn10insertion mutants and theEscherichia coliBW25113 ΔsbmA(JW0368) strain were still susceptible to the bactenecin PrAMP Bac7(1-35) and oncocin PrAMPs Onc18 and Onc112, as well as to Chex1-Arg20, despite significantly reduced internalizations. In a second round of random mutagenesis, the remaining susceptibility was linked to theyjiL-mdtMgene cluster.E. coliBW25113 and its ΔyjiLnull mutant (JW5785) were equally susceptible to all PrAMPs tested, whereas the BW25113 ΔmdtMmutant was less susceptible to oncocins. The JW0368yjiL::Tn10transposon mutant (BS2) was resistant to all short PrAMPs and susceptible only to full-length Bac7 and A3-APO. Interestingly, PrAMPs appear to enter bacteria via MdtM, a multidrug resistance transporter (drug/H+antiporter) of the major facilitator superfamily (MFS) that can efflux antibiotics, biocides, and bile salts. In conclusion, PrAMPs enter bacteria via ABC and MFS transporters that efflux antibiotics and cytotoxic compounds from the cytoplasm to the periplasm.

scholarly journals Induction of the Cpx Envelope Stress Pathway Contributes to Escherichia coli Tolerance to Antimicrobial Peptides

2013 ◽  
Vol 79 (24) ◽  
pp. 7770-7779 ◽  
Author(s):  
Bianca Audrain ◽  
Lionel Ferrières ◽  
Amira Zairi ◽  
Guillaume Soubigou ◽  
Curtis Dobson ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antimicrobial Peptides ◽  
Antimicrobial Peptide ◽  
Polymyxin B ◽  
Genetic Response ◽  
Content Type ◽  
E Coli ◽  
Multiresistant Bacteria ◽  
Envelope Stress ◽  
Stress Pathway

ABSTRACTAntimicrobial peptides produced by multicellular organisms as part of their innate system of defense against microorganisms are currently considered potential alternatives to conventional antibiotics in case of infection by multiresistant bacteria. However, while the mode of action of antimicrobial peptides is relatively well described, resistance mechanisms potentially induced or selected by these peptides are still poorly understood. In this work, we studied the mechanisms of action and resistance potentially induced by ApoEdpL-W, a new antimicrobial peptide derived from human apolipoprotein E. Investigation of the genetic response ofEscherichia coliupon exposure to sublethal concentrations of ApoEdpL-W revealed that this antimicrobial peptide triggers activation of RcsCDB, CpxAR, and σEenvelope stress pathways. This genetic response is not restricted to ApoEdpL-W, since several other antimicrobial peptides, including polymyxin B, melittin, LL-37, and modified S4dermaseptin, also activate severalE. colienvelope stress pathways. Finally, we demonstrate that induction of the CpxAR two-component system directly contributes toE. colitolerance toward ApoEdpL-W, polymyxin B, and melittin. These results therefore show thatE. colisenses and responds to different antimicrobial peptides by activation of the CpxAR pathway. While this study further extends the understanding of the array of peptide-induced stress signaling systems, it also provides insight into the contribution of Cpx envelope stress pathway toE. colitolerance to antimicrobial peptides.

scholarly journals No Single Irreplaceable Acidic Residues in the Escherichia coli Secondary Multidrug Transporter MdfA

2006 ◽  
Vol 188 (15) ◽  
pp. 5635-5639 ◽  
Author(s):  
Nadejda Sigal ◽  
Shahar Molshanski-Mor ◽  
Eitan Bibi
Keyword(s):  
Escherichia Coli ◽  
Critical Role ◽  
Energy Coupling ◽  
Major Facilitator Superfamily ◽  
Multidrug Transporter ◽  
Coupling Mechanism ◽  
Acidic Residues ◽  
Major Facilitator ◽  
Mfs Transporters ◽  
Solute Transporters

ABSTRACT The largest family of solute transporters (major facilitator superfamily [MFS]) includes proton-motive-force-driven secondary transporters. Several characterized MFS transporters utilize essential acidic residues that play a critical role in the energy-coupling mechanism during transport. Surprisingly, we show here that no single acidic residue plays an irreplaceable role in the Escherichia coli secondary multidrug transporter MdfA.

scholarly journals Mdt(A), a New Efflux Protein Conferring Multiple Antibiotic Resistance in Lactococcus lactis andEscherichia coli

2001 ◽  
Vol 45 (4) ◽  
pp. 1109-1114 ◽  
Author(s):  
Vincent Perreten ◽  
Franziska V. Schwarz ◽  
Michael Teuber ◽  
Stuart B. Levy
Keyword(s):  
Escherichia Coli ◽  
Lactococcus Lactis ◽  
Major Facilitator Superfamily ◽  
Multiple Drug ◽  
Drug Transporter ◽  
Multiple Antibiotic Resistance ◽  
Atp Binding Site ◽  
E Coli ◽  
Transmembrane Segments ◽  
Major Facilitator

ABSTRACT The mdt(A) gene, previously designatedmef214, from Lactococcus lactis subsp.lactis plasmid pK214 encodes a protein [Mdt(A) (multiple drug transporter)] with 12 putative transmembrane segments (TMS) that contain typical motifs conserved among the efflux proteins of the major facilitator superfamily. However, it also has two C-motifs (conserved in the fifth TMS of the antiporters) and a putative ATP-binding site. Expression of the cloned mdt(A) gene decreased susceptibility to macrolides, lincosamides, streptogramins, and tetracyclines in L. lactis and Escherichia coli, but not in Enterococcus faecalis or inStaphylococcus aureus. Glucose-dependent efflux of erythromycin and tetracycline was demonstrated in L. lactisand in E. coli.

scholarly journals ThephtC-phtDLocus Equips Legionella pneumophila for Thymidine Salvage and Replication in Macrophages

2013 ◽  
Vol 82 (2) ◽  
pp. 720-730 ◽  
Author(s):  
Maris V. Fonseca ◽  
John-Demian Sauer ◽  
Sebastien Crepin ◽  
Brenda Byrne ◽  
Michele S. Swanson
Keyword(s):  
Escherichia Coli ◽  
Life Cycle ◽  
Thymidylate Synthase ◽  
Legionella Pneumophila ◽  
Opportunistic Pathogen ◽  
Nucleoside Transporters ◽  
Major Facilitator Superfamily ◽  
Pyrimidine Nucleoside ◽  
Content Type ◽  
Major Facilitator

ABSTRACTThe phagosomal transporter (Pht) family of the major facilitator superfamily (MFS) is encoded by phylogenetically related intracellular gammaproteobacteria, including the opportunistic pathogenLegionella pneumophila. The location of thephtgenes between the putative thymidine kinase (tdk) and phosphopentomutase (deoB) genes suggested that thephtCandphtDloci contribute to thymidine salvage inL. pneumophila. Indeed, aphtC+allele intransrestored pyrimidine uptake to anEscherichia colimutant that lacked all known nucleoside transporters, whereas aphtD+allele did not. The results of phenotypic analyses ofL. pneumophilastrains lackingphtCorphtDstrongly indicate thatL. pneumophilarequires PhtC and PhtD function under conditions where sustained dTMP synthesis is compromised. First, in broth cultures that mimicked thymidine limitation or starvation,L. pneumophilaexhibited a marked requirement for PhtC function. Conversely, mutation ofphtDconferred a survival advantage. Second, in medium that lacked thymidine, multicopyphtC+orphtD+alleles enhanced the survival ofL. pneumophilathymidylate synthase (thyA)-deficient strains, which cannot synthesize dTMP endogenously. Third, under conditions in which transport of the pyrimidine nucleoside analog 5-fluorodeoxyuridine (FUdR) would inhibit growth, PhtC and PhtD conferred a growth advantage toL. pneumophilathyA+strains. Finally, when cultured in macrophages,L. pneumophilarequired thephtC-phtDlocus to replicate. Accordingly, we propose that PhtC and PhtD contribute to protectL. pneumophilafrom dTMP starvation during its intracellular life cycle.

scholarly journals A selC-Associated Genomic Island of the Extraintestinal Avian Pathogenic Escherichia coli Strain BEN2908 Is Involved in Carbohydrate Uptake and Virulence

2006 ◽  
Vol 188 (3) ◽  
pp. 977-987 ◽  
Author(s):  
Iman Chouikha ◽  
Pierre Germon ◽  
Annie Brée ◽  
Philippe Gilot ◽  
Maryvonne Moulin-Schouleur ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genomic Island ◽  
Bacterial Species ◽  
Coli Strain ◽  
Major Facilitator Superfamily ◽  
Integrase Gene ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Major Facilitator Superfamily Transporter ◽  
Major Facilitator

ABSTRACT The complete nucleotide sequence and genetic organization of a new genomic island (AGI-3) isolated from the extraintestinal avian pathogenic Escherichia coli strain BEN2908 is reported. This 49,600-bp island is inserted at the selC locus and contains putative mobile genetic elements such as a phage-related integrase gene, transposase genes, and direct repeats. AGI-3 shows a mosaic structure of five modules. Some of these modules are present in other E. coli strains and in other pathogenic bacterial species. The gene cluster aec-35 to aec-37 of module 1 encodes proteins associated with carbohydrates assimilation such as a major facilitator superfamily transporter (Aec-36), a glycosidase (Aec-37), and a putative transcriptional regulator of the LacI family (Aec-35). The aec-35 to aec-37 cluster was found in 11.6% of the tested pathogenic and nonpathogenic E. coli strains. When present, the aec-35 to aec-37 cluster is strongly associated with the selC locus (97%). Deletion of the aec-35-aec-37 region affects the assimilation of seven carbohydrates, decreases the growth rate of the strain in minimal medium containing galacturonate or trehalose, and attenuates the virulence of E. coli BEN2908 for chickens.

scholarly journals Trichophyton rubrum Azole Resistance Mediated by a New ABC Transporter, TruMDR3

2019 ◽  
Vol 63 (11) ◽  
Author(s):  
Michel Monod ◽  
Marc Feuermann ◽  
Karine Salamin ◽  
Marina Fratti ◽  
Maya Makino ◽  
...  
Keyword(s):  
Trichophyton Rubrum ◽  
Heterologous Gene Expression ◽  
Efflux Pumps ◽  
Major Facilitator Superfamily ◽  
Azole Resistance ◽  
Multidrug Efflux ◽  
Content Type ◽  
Milbemycin Oxime ◽  
Major Facilitator ◽  
Mfs Transporters

ABSTRACT The mechanisms of terbinafine resistance in a set of clinical isolates of Trichophyton rubrum have been studied recently. Of these isolates, TIMM20092 also showed reduced sensitivity to azoles. The azole resistance of TIMM20092 could be inhibited by milbemycin oxime, prompting us to examine the potential of T. rubrum to develop resistance through multidrug efflux transporters. The introduction of a T. rubrum cDNA library into Saccharomyces cerevisiae allowed the isolation of one transporter of the major facilitator superfamily (MFS) conferring resistance to azoles (TruMFS1). To identify more azole efflux pumps among 39 ABC and 170 MFS transporters present within the T. rubrum genome, we performed a BLASTp analysis of Aspergillus fumigatus, Candida albicans, and Candida glabrata on transporters that were previously shown to confer azole resistance. The identified candidates were further tested by heterologous gene expression in S. cerevisiae. Four ABC transporters (TruMDR1, TruMDR2, TruMDR3, and TruMDR5) and a second MFS transporter (TruMFS2) proved to be able to operate as azole efflux pumps. Milbemycin oxime inhibited only TruMDR3. Expression analysis showed that both TruMDR3 and TruMDR2 were significantly upregulated in TIMM20092. TruMDR3 transports voriconazole (VRC) and itraconazole (ITC), while TruMDR2 transports only ITC. Disruption of TruMDR3 in TIMM20092 abolished its resistance to VRC and reduced its resistance to ITC. Our study highlights TruMDR3, a newly identified transporter of the ABC family in T. rubrum, which can confer azole resistance if overexpressed. Finally, inhibition of TruMDR3 by milbemycin suggests that milbemycin analogs could be interesting compounds to treat dermatophyte infections in cases of azole resistance.

scholarly journals New Plasmid-Mediated Fluoroquinolone Efflux Pump, QepA, Found in an Escherichia coli Clinical Isolate

2007 ◽  
Vol 51 (9) ◽  
pp. 3354-3360 ◽  
Author(s):  
Kunikazu Yamane ◽  
Jun-ichi Wachino ◽  
Satowa Suzuki ◽  
Kouji Kimura ◽  
Naohiro Shibata ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Molecular Mechanisms ◽  
Efflux Pump ◽  
Resistance Mechanisms ◽  
Major Facilitator Superfamily ◽  
Gene Encoding ◽  
Resistance Level ◽  
E Coli ◽  
Susceptibility Tests ◽  
Major Facilitator

ABSTRACT Plasmid-mediated Qnr and AAC(6′)-Ib-cr have been recognized as new molecular mechanisms affecting fluoroquinolone (FQ) resistance. C316, an Escherichia coli strain demonstrating resistance to various FQs, was isolated in Japan. Resistance to FQs was augmented in an E. coli CSH2 transconjugant, but PCR failed to detect qnr genes, suggesting the presence of novel plasmid-mediated FQ resistance mechanisms. Susceptibility tests, DNA manipulation, and analyses of the gene and its product were performed to characterize the genetic determinant. A novel FQ-resistant gene, qepA, was identified in a plasmid, pHPA, of E. coli C316, and both qepA and rmtB genes were mediated by a probable transposable element flanked by two copies of IS26. Levels of resistance to norfloxacin, ciprofloxacin, and enrofloxacin were significantly elevated in E. coli transformants harboring qepA under AcrB-TolC-deficient conditions. QepA showed considerable similarities to transporters belonging to the 14-transmembrane-segment family of environmental actinomycetes. The effect of carbonyl cyanide m-chlorophenylhydrazone (CCCP) on accumulation of norfloxacin was assayed in a qepA-harboring E. coli transformant. The intracellular accumulation of norfloxacin was decreased in a qepA-expressing E. coli transformant, but this phenomenon was canceled by CCCP. The augmented FQ resistance level acquired by the probable intergeneric transfer of a gene encoding a major facilitator superfamily-type efflux pump from some environmental microbes to E. coli was first identified. Surveillance of the qepA-harboring clinical isolates should be encouraged to minimize further dissemination of the kind of plasmid-dependent FQ resistance determinants among pathogenic microbes.

scholarly journals GntP Is the Escherichia coli Fructuronic Acid Transporter and Belongs to the UxuR Regulon

2004 ◽  
Vol 186 (22) ◽  
pp. 7690-7696 ◽  
Author(s):  
Cristina Bates Utz ◽  
Ann B. Nguyen ◽  
Darren J. Smalley ◽  
April B. Anderson ◽  
Tyrrell Conway
Keyword(s):  
Escherichia Coli ◽  
Protein A ◽  
Physiological Role ◽  
Major Facilitator Superfamily ◽  
Receptor Protein ◽  
E Coli ◽  
The Subject ◽  
Camp Receptor ◽  
Major Facilitator

ABSTRACT Escherichia coli has four gluconate transporters, GntP, GntU, GntT, and IdnT, which are members of the major facilitator superfamily. The physiological function of GntP was previously unknown and is the subject of this study. GntP is not induced by gluconate, and despite being located adjacent to genes involved in glucuronate catabolism, gntP does not encode a glucuronate transporter. Here we identify gntP as the gene which encodes the fructuronate transporter. We show that gntP is induced by fructuronate and is a new member of the UxuR regulon: gntP is derepressed in an uxuR strain, UxuR binds in vitro specifically to an operator site that overlaps the gntP promoter, and UxuR binding is eliminated by fructuronate. Transcription of gntP requires activation by cyclic AMP (cAMP)-cAMP receptor protein. A gntP mutant cannot grow on fructuronate but grows normally on glucuronate and gluconate. Thus, the UxuR regulon is a module of sugar acid catabolism whose physiological role is for growth on fructuronate. Glucuronate, because it proceeds through a fructuronate intermediate, must induce the UxuR regulon and must also induce the ExuR regulon, which encodes the glucuronate transporter, ExuT, and the first step in its catabolism, UxaC. Thus, hexuronate catabolism in E. coli requires both the ExuR and UxuR regulons, while fructuronate catabolism requires only the UxuR regulon.

scholarly journals Members of the Francisella tularensis Phagosomal Transporter Subfamily of Major Facilitator Superfamily Transporters Are Critical for Pathogenesis

2012 ◽  
Vol 80 (7) ◽  
pp. 2390-2401 ◽  
Author(s):  
Mark E. Marohn ◽  
Araceli E. Santiago ◽  
Kari Ann Shirey ◽  
Michael Lipsky ◽  
Stefanie N. Vogel ◽  
...  
Keyword(s):  
Francisella Tularensis ◽  
Vaccine Development ◽  
Cell Types ◽  
Major Facilitator Superfamily ◽  
Content Type ◽  
Infectious Dose ◽  
Mutant Strains ◽  
Major Facilitator ◽  
Mfs Transporters

ABSTRACTFrancisella tularensisis the causative agent of tularemia. Due to its aerosolizable nature and low infectious dose,F. tularensisis classified as a category A select agent and, therefore, is a priority for vaccine development. Survival and replication in macrophages and other cell types are critical toF. tularensispathogenesis, and impaired intracellular survival has been linked to a reduction in virulence. TheF. tularensisgenome is predicted to encode 31 major facilitator superfamily (MFS) transporters, and the nine-memberFrancisellaphagosomal transporter (Fpt) subfamily possesses homology with virulence factors in other intracellular pathogens. We hypothesized that these MFS transporters may play an important role inF. tularensispathogenesis and serve as good targets for attenuation and vaccine development. Here we show altered intracellular replication kinetics and attenuation of virulence in mice infected with three of the nine Fpt mutant strains compared with wild-type (WT)F. tularensisLVS. The vaccination of mice with these mutant strains was protective against a lethal intraperitoneal challenge. Additionally, we observed pronounced differences in cytokine profiles in the livers of mutant-infected mice, suggesting that alterations inin vivocytokine responses are a major contributor to the attenuation observed for these mutant strains. These results confirm that this subset of MFS transporters plays an important role in the pathogenesis ofF. tularensisand suggest that a focus on the development of attenuated Fpt subfamily MFS transporter mutants is a viable strategy toward the development of an efficacious vaccine.

scholarly journals Isolation of a Gene Responsible for the Oxidation oftrans-Anethole topara-Anisaldehyde by Pseudomonas putida JYR-1 and Its Expression in Escherichia coli

2012 ◽  
Vol 78 (15) ◽  
pp. 5238-5246 ◽  
Author(s):  
Dongfei Han ◽  
Ji-Young Ryu ◽  
Robert A. Kanaly ◽  
Hor-Gil Hur
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Putida ◽  
Hypothetical Protein ◽  
Aldehyde Group ◽  
Agrobacterium Vitis ◽  
T7 Promoter ◽  
Content Type ◽  
E Coli ◽  
Cell Extracts ◽  
Isoeugenol Monooxygenase

ABSTRACTA plasmid, pTA163, inEscherichia colicontained an approximately 34-kb gene fragment fromPseudomonas putidaJYR-1 that included the genes responsible for the metabolism oftrans-anethole to protocatechuic acid. Three Tn5-disrupted open reading frame 10 (ORF 10) mutants of plasmid pTA163 lost their abilities to catalyzetrans-anethole. Heterologously expressed ORF 10 (1,047 nucleotides [nt]) under a T7 promoter inE. colicatalyzed oxidative cleavage of a propenyl group oftrans-anethole to an aldehyde group, resulting in the production ofpara-anisaldehyde, and this gene was designatedtao(trans-anetholeoxygenase). The deduced amino acid sequence of TAO had the highest identity (34%) to a hypothetical protein ofAgrobacterium vitisS4 and likely contained a flavin-binding site. Preferred incorporation of an oxygen molecule from water intop-anisaldehyde using18O-labeling experiments indicated stereo preference of TAO for hydrolysis of the epoxide group. Interestingly, unlike the narrow substrate range of isoeugenol monooxygenase fromPseudomonas putidaIE27 andPseudomonas nitroreducensJin1, TAO fromP. putidaJYR-1 catalyzed isoeugenol,O-methyl isoeugenol, and isosafrole, all of which contain the 2-propenyl functional group on the aromatic ring structure. Addition of NAD(P)H to the ultrafiltered cell extracts ofE. coli(pTA163) increased the activity of TAO. Due to the relaxed substrate range of TAO, it may be utilized for the production of various fragrance compounds from plant phenylpropanoids in the future.

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