Artículo

Este artículo es de Acceso Abierto y puede ser descargado en su versión final desde nuestro repositorio
Consulte el artículo en la página del editor
Consulte la política de Acceso Abierto del editor

Abstract:

The RND-type efflux pumps are responsible for the multidrug resistance phenotype observed in many clinically relevant species. Also, RND pumps have been implicated in physiological processes, with roles in the virulence mechanisms of several pathogenic bacteria. We have previously shown that the BepC outer membrane factor of Brucella suis is involved in the efflux of diverse drugs, probably as part of a tripartite complex with an inner membrane translocase. In the present work, we characterize two membrane fusion protein-RND translocases of B. suis encoded by the bepDE and bepFG loci. MIC assays showed that the B. suis AbepE mutant was more sensitive to deoxycholate (DOC), ethidium bromide, and crystal violet. Furthermore, multicopy bepDE increased resistance to DOC and crystal violet and also to other drugs, including ampicillin, norfloxacin, ciprofloxacin, tetracycline, and doxycycline. In contrast to the ΔbepE mutant, the resistance profile of B. suis remained unaltered when the other RND gene (bepG) was deleted. However, the ΔbepE ΔbepG double mutant showed a more severe phenotype than the ΔbepE mutant, indicating that BepFG also contributes to drug resistance. An open reading frame (bepR) coding for a putative regulatory protein of the TetR family was found upstream of the bepDE locus. BepR strongly repressed the activity of the bepDE promoter, but DOC released the repression mediated by BepR. A clear induction of the bepFG promoter activity was observed only in the BepDE-defective mutant, indicating a regulatory interplay between the two RND efflux pumps. Although only the BepFG-defective mutant showed a moderate attenuation in model cells, the activities of both bepDE and bepFG promoters were induced in the intracellular environment of HeLa cells. Our results show that B. suis harbors two functional RND efflux pumps that may contribute to virulence. Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Registro:

Documento: Artículo
Título:Interplay between two RND systems mediating antimicrobial resistance in brucella suis
Autor:Martin, F.A.; Posadas, D.M.; Carrica, M.C.; Cravero, S.L.; O'Callaghan, D.; Zorreguieta, A.
Filiación:Fundación Instituto Leloir, IIBBA CONICET and FCEyN, Universidad de Buenos Aires, Patricias Argentinas 435, (C1405BWE) Buenos Aires, Argentina
Instituto de Biotecnología, CICVyA, INTA-Castelar, Las Cabañas y Los Reseros s/n (B1712WAA) Castelar, Buenos Aires, Argentina
Institut National de la Santé et de la Recherche Médicale, ESPRI26, UFR de Médecine, CS 83021, Avenue Kennedy, 30908 NIMES Cedex 02, France
Palabras clave:ampicillin; carrier protein; ciprofloxacin; crystal violet; deoxycholic acid; doxycycline; ethidium bromide; norfloxacin; tetracycline; antiinfective agent; bacterial protein; carrier protein; crystal violet; deoxycholic acid; ethidium; repressor protein; antibiotic resistance; article; Brucella suis; controlled study; gene repression; nonhuman; phenotype; priority journal; promoter region; amino acid sequence; drug effect; epithelium cell; gene deletion; gene dosage; gene expression regulation; genetics; HeLa cell; human; metabolism; microbiological examination; microbiology; pathogenicity; physiology; sequence alignment; virulence; Brucella melitensis biovar Suis; Amino Acid Sequence; Anti-Bacterial Agents; Bacterial Proteins; Brucella suis; Deoxycholic Acid; Drug Resistance, Bacterial; Epithelial Cells; Ethidium; Gene Deletion; Gene Dosage; Gene Expression Regulation, Bacterial; Gentian Violet; Hela Cells; Humans; Membrane Transport Proteins; Microbial Sensitivity Tests; Repressor Proteins; Sequence Alignment; Virulence
Año:2009
Volumen:191
Número:8
Página de inicio:2530
Página de fin:2540
DOI: http://dx.doi.org/10.1128/JB.01198-08
Título revista:Journal of Bacteriology
Título revista abreviado:J. Bacteriol.
ISSN:00219193
CODEN:JOBAA
CAS:ampicillin, 69-52-3, 69-53-4, 7177-48-2, 74083-13-9, 94586-58-0; carrier protein, 80700-39-6; ciprofloxacin, 85721-33-1; crystal violet, 467-63-0, 548-62-9; deoxycholic acid, 83-44-3; doxycycline, 10592-13-9, 17086-28-1, 564-25-0; ethidium bromide, 1239-45-8; norfloxacin, 70458-96-7; tetracycline, 23843-90-5, 60-54-8, 64-75-5; ethidium, 3546-21-2; Anti-Bacterial Agents; Bacterial Proteins; Deoxycholic Acid, 83-44-3; Ethidium, 3546-21-2; Gentian Violet, 548-62-9; Membrane Transport Proteins; Repressor Proteins
PDF:https://bibliotecadigital.exactas.uba.ar/download/paper/paper_00219193_v191_n8_p2530_Martin.pdf
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00219193_v191_n8_p2530_Martin

Referencias:

  • Aires, J.R., Nikaido, H., Aminoglycosides are captured from both periplasm and cytoplasm by the AcrD multidrug efflux transporter of Escherichia coli- (2005) J. Bacteriol, 187, pp. 1923-1929
  • Akama, H., Matsuura, T., Kashiwagi, S., Yoneyama, H., Narita, S.-I., Tsuki-hara, T., Nakagawa, A., Nakae, T., Crystal structure of the membrane fusion protein, MexA, of the multidrug transporter in Pseudomonas aeruginosa (2004) J. Biol. Chem, 279, pp. 25939-25942
  • Altschul, S.F., Gish, W., Miller, W., Myers, E.W., Lipman, D.J., Basic local alignment search tool (1990) J. Mol. Biol, 215, pp. 403-410
  • Andersen, C., Channel-tunnels: Outer membrane components of type I secretion systems and multidrug efflux pumps of Gram-negative bacteria (2003) Rev. Physiol. Biochem. Pharmacol, 147, pp. 122-165
  • Ariza, J., Bosilkovski, M., Cascio, A., Colmenero, J.D., Corbel, M.J., Falagas, M.E., Memish, Z.A., Pappas, G., Perspectives for the treatment of brucellosis in the 21st century: The Ioannina recommendations (2007) PLoS Med, 4, pp. e317
  • Avila-Sakar, A.J., Misaghi, S., Wilson-Kubalek, E.M., Downing, K.H., Zgurskaya, H., Nikaido, H., Nogales, E., Lipid-layer crystallization and preliminary three-dimensional structural analysis of AcrA, the periplasmic component of a bacterial multidrug efflux pump (2001) J. Struct. Biol, 136, pp. 81-88
  • Batut, J., Andersson, S.G.E., O'Callaghan, D., The evolution of chronic infection strategies in the alpha-proteobacteria (2004) Nat. Rev. Microbiol, 2, pp. 933-945
  • Begley, M., Gahan, C.G.M., Hill, C., The interaction between bacteria and bile (2005) FEMS Microbiol. Rev, 29, pp. 625-651
  • Boschiroli, M.L., Ouahrani-Bettache, S., Foulongne, V., Michaux-Chara- chon, S., Bourg, G., Allardet-Servent, A., Cazevieille, C., O'Callaghan, D., The Brucella suis virB operon is induced intracellularly in macrophages (2002) Proc. Natl. Acad. Sci. USA, 99, pp. 1544-1549
  • Celli, J., Gorvel, J.-P., Organelle robbery: Brucella interactions with the endoplasmic reticulum (2004) Curr. Opin. Microbiol, 7, pp. 93-97
  • Chan, Y.Y., Chua, K.L., The Burkholderia pseudomallei BpeAB- OprB efflux pump: Expression and impact on quorum sensing and virulence (2005) J. Bacteriol, 187, pp. 4707-4719
  • Chan, Y.Y., Tan, T.M.C., Ong, Y.M., Chua, K.L., BpeAB-OprB, a multidrug efflux pump in Burkholderia pseudomallei (2004) Antimicrob. Agents Chemother, 48, pp. 1128-1135
  • Comerci, D.J., Martinez-Lorenzo, M.J., Sieira, R., Gorvel, J.P., Ugalde, R.A., Essential role of the VirB machinery in the maturation of the Brucella abortus-containing vacuole (2001) Cell Microbiol, 3, pp. 159-168
  • Cserzo, M., Eisenhaber, F., Eisenhaber, B., Simon, I., TM or not TM: Transmembrane protein prediction with low false positive rate using DAS-TMfilter (2004) Bioinformatics, 20, pp. 136-137
  • Das, D., Xu, Q.S., Lee, J.Y., Ankoudinova, I., Huang, C., Lou, Y., DeGio- vanni, A., Kim, S.-H., Crystal structure of the multidrug efflux transporter AcrB at 3.1A resolution reveals the N-terminal region with conserved amino acids (2007) J. Struct. Biol, 158, pp. 494-502
  • Delpino, M.V., Marchesini, M.I., Estein, S.M., Comerci, D.J., Cassataro, J., Fossati, C.A., Baldi, P.C., A bile salt hydrolase of Brucella abortus contributes to the establishment of a successful infection through the oral route in mice (2007) Infect. Immun, 75, pp. 299-305
  • Dinh, T., I. T. Paulsen, and M. H. Saier, Jr. 1994. A family of extracytoplasmic proteins that allow transport of large molecules across the outer membranes of gram-negative bacteria. J. Bacteriol. 176:3825-3831; Duque, E., Segura, A., Mosqueda, G., Ramos, J.L., Global and cognate regulators control the expression of the organic solvent efflux pumps TtgABC and TtgDEF of Pseudomonas putida (2001) Mol. Microbiol, 39, pp. 1100-1106
  • Elkins, C.A., Nikaido, H., 3D structure of AcrB: The archetypal multidrug efflux transporter of Escherichia coli likely captures substrates from periplasm (2003) Drug Resist. Updat, 6, pp. 9-13
  • Gasteiger, E., Gattiker, A., Hoogland, C., Ivanyi, I., Appel, R.D., Bairoch, A., ExPASy: The proteomics server for in-depth protein knowledge and analysis (2003) Nucleic Acids Res, 31, pp. 3784-3788
  • Gee, J.M., Valderas, M.W., Kovach, M.E., Grippe, V.K., Robertson, G.T., Ng, W.-L., Richardson, J.M., Roop II, R.M., The Brucella abortus Cu, Zn superoxide dismutase is required for optimal resistance to oxidative killing by murine macrophages and wild-type virulence in experimentally infected mice (2005) Infect. Immun, 73, pp. 2873-2880
  • Grkovic, S., Brown, M.H., Hardie, K.M., Firth, N., Skurray, R.A., Stable low-copy-number Staphylococcus aureus shuttle vectors (2003) Microbiology, 149, pp. 785-794
  • Grkovic, S., Brown, M.H., Roberts, N.J., Paulsen, J.T., Skurray, R.A., QacR is a repressor protein that regulates expression of the Staphylococcus aureus multidrug efflux pump QacA (1998) J. Biol. Chem, 12, pp. 18665-18673
  • Guan, L., Nakae, T., Identification of essential charged residues in transmembrane segments of the multidrug transporter MexB of Pseudomonas aeruginosa (2001) J. Bacteriol, 183, pp. 1734-1739
  • Hillen, W., Berens, C., Mechanisms underlying expression of Tn10 encoded tetracycline resistance (1994) Annu. Rev. Microbiol, 48, pp. 345-369
  • Ko, J., Splitter, G.A., Molecular host-pathogen interaction in brucellosis: Current understanding and future approaches to vaccine development for mice and humans (2003) Clin. Microbiol. Rev, 16, pp. 65-78
  • Kohler, S., Foulongne, V., Ouahrani-Bettache, S., Bourg, G., Teyssier, J., Ramuz, M., Liautard, J.-P., The analysis of the intramacrophagic virulome of Brucella suis deciphers the environment encountered by the pathogen inside the macrophage host cell (2002) Proc. Natl. Acad. Sci. USA, 99, pp. 15711-15716
  • Kohler, S., Ouahrani-Bettache, S., Layssac, M., Teyssier, J., Liautard, J.P., Constitutive and inducible expression of green fluorescent protein in Brucella suis (1999) Infect. Immun, 67, pp. 6695-6697
  • Kulakov, I.K., O'Callaghan, D., Ramuz, M., Liautard, J.P., Skavronskaia, A.G., A genetic and molecular biological study of the patho-genicity factors in Brucella (1996) Zh. Mikrobiol. Epidemiol. Immunobiol, 3, pp. 34-39
  • Larkin, M. A, G. Blackshields, N. P. Brown, R. Chenna, P. A. McGettigan, H. McWilliam, F. Valentin, I. M. Wallace, A. Wilm, R. Lopez, J. D. Thompson, T. J. Gibson, and D. G. Higgins. 2007. Clustal W and Clustal X version 2.0. Bioinformatics 23:2947-2948; Li, X.Z., Nikaido, H., Poole, K., Role of mexA-mexB-oprM in antibiotic efflux in Pseudomonas aeruginosa (1995) Antimicrob. Agents Chemother, 39, pp. 1948-1953
  • Lin, J., Akiba, M., Sahin, O., Zhang, Q., CmeR functions as a transcriptional repressor for the multidrug efflux pump CmeABC in Campy-lobacter jejuni (2005) Antimicrob. Agents Chemother, 49, pp. 1067-1075
  • Lin, J., Huang, S., Zhang, Q., Outer membrane proteins: Key players for bacterial adaptation in host niches (2002) Microbes Infect, 4, pp. 325-331
  • Lin, J., Michel, L.O., Zhang, Q., CmeABC functions as a multidrug efflux system in Campylobacter jee'uni (2002) Antimicrob. Agents Chemother, 46, pp. 2124-2131
  • Ma, D., Alberti, M., Lynch, C., Nikaido, H., Hearst, J.E., The local repressor AcrR plays a modulating role in the regulation of acrAB genes of Escherichia coli by global stress signals (1996) Mol. Microbiol, 19, pp. 101-112
  • Ma, D., Cook, D.N., Alberti, M., Pon, N.G., Nikaido, H., Hearst, J.E., Genes acrA and acrB encode a stress-induced efflux system of Escherichia coli (1995) Mol. Microbiol, 16, pp. 45-55
  • Ma, D., Cook, D.N., Hearst, J.E., Nikaido, H., Efflux pumps and drug resistance in gram-negative bacteria (1994) Trends Microbiol, 2, pp. 489-493
  • Martínez de Tejada, G., Moriyón, I., The outer membranes of Brucella spp. are not barriers to hydrophobic permeants (1993) J. Bacteriol, 175, pp. 5273-5275
  • Middlemiss, J.K., Poole, K., Differential impact of MexB mutations on substrate selectivity of the MexAB-OprM multidrug efflux pump of Pseudomonas aeruginosa (2004) J. Bacteriol, 186, pp. 1258-1269
  • Moriyon, I., Berman, D.T., Effects ofnonionic, ionic, and dipolar ionic detergents and EDTA on the Brucella cell envelope (1982) J. Bacteriol, 152, pp. 822-828
  • Moriyón, I., Gamazo, C., Díaz, R., Properties of the outer membrane of Brucella (1987) Ann. Inst. Pasteur Microbiol, 138, pp. 89-91
  • Murakami, S., Yamaguchi, A., Multidrug-exporting secondary transporters (2003) Curr. Opin. Struct. Biol, 13, pp. 443-452
  • Nicholas, K.B., Nicholas Jr., H.B., Deerfield II, D.W., GeneDoc: Analysis and visualization of genetic variation (1997) EMBNET News, 4, pp. 1-4
  • Notredame, C., Higgins, D.G., Heringa, J., T-Coffee: A novel method for fast and accurate multiple sequence alignment (2000) J. Mol. Biol, 302, pp. 205-217
  • O'Callaghan, D., Cazevieille, C., Allardet-Servent, A., Boschiroli, M.L., Bourg, G., Foulongne, V., Frutos, P., Ramuz, M., A homo- logue of the Agrobacterium tumefaciens VirB and Bordetella pertussis Ptl type IV secretion systems is essential for intracellular survival of Brucella suis (1999) Mol. Microbiol, 33, pp. 1210-1220
  • Pan, W., Spratt, B.G., Regulation of the permeability of the gonococcal cell envelope by the mtr system (1994) Mol. Microbiol, 11, pp. 769-775
  • Pearson, J.P., Van Delden, C., Iglewski, B.H., Active efflux and diffusion are involved in transport of Pseudomonas aeruginosa cell-to-cell signals (1999) J. Bacteriol, 181, pp. 1203-1210
  • Piddock, L.J.V., Clinically relevant chromosomally encoded multidrug resistance efflux pumps in bacteria (2006) Clin. Microbiol. Rev, 19, pp. 382-402
  • Piddock, L.J.V., Multidrug-resistance efflux pumps-not just for resistance (2006) Nat. Rev. Microbiol, 4, pp. 629-636
  • Pizarro-Cerdá, J., Moreno, E., Sanguedolce, V., Mege, J.L., Gorvel, J.P., Virulent Brucella abortus prevents lysosome fusion and is distributed within autophagosome-like compartments (1998) Infect. Immun, 66, pp. 2387-2392
  • Poole, K., Krebes, K., McNally, C., Neshat, S., Multiple antibiotic resistance in Pseudomonas aeruginosa: Evidence for involvement of an efflux operon (1993) J. Bacteriol, 175, pp. 7363-7372
  • Poole, K., Srikumar, R., Multidrug efflux in Pseudomonas aeruginosa: Components, mechanisms and clinical significance (2001) Curr. Top. Med. Chem, 1, pp. 59-71
  • Porte, F., Liautard, J.P., Köhler, S., Early acidification of phago- somes containing Brucella suis is essential for intracellular survival in murine macrophages (1999) Infect. Immun, 67, pp. 4041-4047
  • Posadas, D. M., F. A. Martín, J. V. Sabio y García, J. M. Spera, M. V. Delpino, P. Baldi, E. Campos, S. L. Cravero, and A. Zorreguieta. 2007. The TolC homologue of Brucella suis is involved in resistance to antimicrobial compounds and virulence. Infect. Immun. 75:379-389; Putman, M., van Veen, H.W., Konings, W.N., Molecular properties of bacterial multidrug transporters (2000) Microbiol. Mol. Biol. Rev, 64, pp. 672-693
  • Ramos, J.L., Duque, E., Godoy, P., Segura, A., Efflux pumps involved in toluene tolerance in Pseudomonas putida DOT-T1E (1998) J. Bacteriol, 180, pp. 3323-3329
  • Rosenberg, E.Y., Bertenthal, D., Nilles, M.L., Bertrand, K.P., Nikaido, H., Bile salts and fatty acids induce the expression of Escherichia coli AcrAB multidrug efflux pump through their interaction with Rob regulatory protein (2003) Mol. Microbiol, 48, pp. 1609-1619
  • Sambrook, J., Fritsch, E.F., Maniatis, T., (1989) Molecular cloning: A laboratory manual, , 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY
  • Schafer, A., Tauch, A., Jager, W., Kalinowski, J., Thierbach, G., Puhler, A., Small mobilizable multi-purpose cloning vectors derived from the Escherichia coli plasmids pK18 and pK19: Selection of defined deletions in the chromosome of Corynebacterium glutamicum (1994) Gene, 145, pp. 69-73
  • Schneiders,T.,S. G.Amyes,andS.B. Levy.2003. Role of AcrR and RamA in fluoroquinolone resistance in clinical Klebsiella pneumoniae isolates from Singapore. Antimicrob. Agents Chemother. 47:2831-2837; Sukchawalit, R., Vattanaviboon, P., Sallabhan, R., Mongkolsuk, S., Construction and characterization of regulated L-arabinose-inducible broad host range expression vectors in Xanthomonas (1999) FEMS Microbiol. Lett, 181, pp. 217-223
  • Taminiau, B., Daykin, M., Swift, S., Boschiroli, M.L., Tibor, A., Lestrate, P., De Bolle, X., Letesson, J.J., Identification of a quorum-sensing signal molecule in the facultative intracellular pathogen Brucella melitensis (2002) Infect. Immun, 70, pp. 3004-3011
  • Touzé, T., Eswaran, J., Bokma, E., Koronakis, E., Hughes, C., Koronakis, V., Interactions underlying assembly of the Escherichia coli AcrAB-TolC multidrug efflux system (2004) Mol. Microbiol, 53, pp. 697-706
  • Turkmani, A., Psaroulaki, A., Christidou, A., Samoilis, G., Mourad, T.A., Tabaa, D., Tselentis, Y., Uptake of ciprofloxacin and ofloxacin by 2 Brucella strains and their fluoroquinolone-resistant variants under different conditions. An in vitro study (2007) Diagn. Microbiol. Infect. Dis, 59, pp. 447-451
  • Velasco, J., Bengoechea, J.A., Brandenburg, K., Lindner, B., Seydel, U., Gonzalez, D., Zähringer, U., Moriyón, I., Brucella abortus and its closest phylogenetic relative, Ochrobactrum spp., differ in outer mem-brane permeability and cationic peptide resistance (2000) Infect. Immun, 68, pp. 3210-3218
  • Vergunst, A.C., Schrammeijer, B., den Dulk-Ras, A., de Vlaam, C.M., Regensburg-Tuink, T.J., Hooykaas, P.J., VirB/D4-dependent protein translocation from Agrobacterium into plant cells (2000) Science, 290, pp. 979-982
  • Wandersman, C., Delepelaire, P., TolC, an Escherichia coli outer membrane protein required for hemolysin secretion (1990) Proc. Natl. Acad. Sci. USA, 87, pp. 4776-4780
  • Wang, H., Dzink-Fox, J.L., Chen, M., Levy, S.B., Genetic characterization of highly fluoroquinolone-resistant clinical Escherichia coli strains from China: Role of acrR mutations (2001) Antimicrob. Agents Chemother, 45, pp. 1515-1521
  • Webber, M.A., Piddock, L.J., The importance of efflux pumps in bacterial antibiotic resistance (2003) J. Antimicrob. Chemother, 51, pp. 9-11
  • Yu, E.W., Aires, J.R., McDermott, G., Nikaido, H., A periplasmic drug-binding site of the AcrB multidrug efflux pump: A crystallographic and site-directed mutagenesis study (2005) J. Bacteriol, 187, pp. 6804-6815

Citas:

---------- APA ----------
Martin, F.A., Posadas, D.M., Carrica, M.C., Cravero, S.L., O'Callaghan, D. & Zorreguieta, A. (2009) . Interplay between two RND systems mediating antimicrobial resistance in brucella suis. Journal of Bacteriology, 191(8), 2530-2540.
http://dx.doi.org/10.1128/JB.01198-08
---------- CHICAGO ----------
Martin, F.A., Posadas, D.M., Carrica, M.C., Cravero, S.L., O'Callaghan, D., Zorreguieta, A. "Interplay between two RND systems mediating antimicrobial resistance in brucella suis" . Journal of Bacteriology 191, no. 8 (2009) : 2530-2540.
http://dx.doi.org/10.1128/JB.01198-08
---------- MLA ----------
Martin, F.A., Posadas, D.M., Carrica, M.C., Cravero, S.L., O'Callaghan, D., Zorreguieta, A. "Interplay between two RND systems mediating antimicrobial resistance in brucella suis" . Journal of Bacteriology, vol. 191, no. 8, 2009, pp. 2530-2540.
http://dx.doi.org/10.1128/JB.01198-08
---------- VANCOUVER ----------
Martin, F.A., Posadas, D.M., Carrica, M.C., Cravero, S.L., O'Callaghan, D., Zorreguieta, A. Interplay between two RND systems mediating antimicrobial resistance in brucella suis. J. Bacteriol. 2009;191(8):2530-2540.
http://dx.doi.org/10.1128/JB.01198-08