title: Antibiotic multiresistance plasmid pRSB101 isolated from a wastewater treatment plant is related to plasmids residing in phytopathogenic bacteria and carries eight different resistance determinants including a multidrug transport system.
abstract: Ten different antibiotic resistance plasmids conferring high-level erythromycin resistance were isolated from an activated sludge bacterial community of a wastewater treatment plant by applying a transformation-based approach. One of these plasmids, designated pRSB101, mediates resistance to tetracycline, erythromycin, roxythromycin, sulfonamides, cephalosporins, spectinomycin, streptomycin, trimethoprim, nalidixic acid and low concentrations of norfloxacin. Plasmid pRSB101 was completely sequenced and annotated. Its size is 47 829 bp. Conserved synteny exists between the pRSB101 replication/partition (rep/par) module and the pXAC33-replicon from the phytopathogen Xanthomonas axonopodis pv. citri. The second pRSB101 backbone module encodes a three-Mob-protein type mobilization (mob) system with homology to that of IncQ-like plasmids. Plasmid pRSB101 is mobilizable with the help of the IncP-1alpha plasmid RP4 providing transfer functions in trans. A 20 kb resistance region on pRSB101 is located within an integron-containing Tn402-like transposon. The variable region of the class 1 integron carries the genes dhfr1 for a dihydrofolate reductase, aadA2 for a spectinomycin/streptomycin adenylyltransferase and bla(TLA-2) for a so far unknown Ambler class A extended spectrum beta-lactamase. The integron-specific 3'-segment (qacEDelta1-sul1-orf5Delta) is connected to a macrolide resistance operon consisting of the genes mph(A) (macrolide 2'-phosphotransferase I), mrx (hydrophobic protein of unknown function) and mphR(A) (regulatory protein). Finally, a putative mobile element with the tetracycline resistance genes tetA (tetracycline efflux pump) and tetR was identified upstream of the Tn402-specific transposase gene tniA. The second 'genetic load' region on pRSB101 harbours four distinct mobile genetic elements, another integron belonging to a new class and footprints of two more transposable elements. A tripartite multidrug (MDR) transporter consisting of an ATP-binding-cassette (ABC)-type ATPase and permease, and an efflux membrane fusion protein (MFP) of the RND-family is encoded between the replication/partition and the mobilization module. Homologues of the macrolide resistance genes mph(A), mrx and mphR(A) were detected on eight other erythromycin resistance-plasmids isolated from activated sludge bacteria. Plasmid pRSB101-like repA amplicons were also obtained from plasmid-DNA preparations of the final effluents of the wastewater treatment plant indicating that pRSB101-like plasmids are released with the final effluents into the environment.
authors.affiliation: Fakultät für Biologie, Lehrstuhl für Genetik, Universität Bielefeld, Postfach 100131, D-33501 Bielefeld, Germany.
authors.forename: Rafael
authors.initials: R
authors.lastname: Szczepanowski
authors.affiliation: Fakultät für Biologie, Lehrstuhl für Genetik, Universität Bielefeld, Postfach 100131, D-33501 Bielefeld, Germany.
authors.forename: Irene
authors.initials: I
authors.lastname: Krahn
authors.affiliation: Center for Biotechnology (CeBiTec), Bioinformatics Resource Facility, Universität Bielefeld, Universitätsstrasse 25, D-33615 Bielefeld, Germany.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Linke
authors.affiliation: Center for Biotechnology (CeBiTec), Bioinformatics Resource Facility, Universität Bielefeld, Universitätsstrasse 25, D-33615 Bielefeld, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Goesmann
authors.affiliation: Fakultät für Biologie, Lehrstuhl für Genetik, Universität Bielefeld, Postfach 100131, D-33501 Bielefeld, Germany.
authors.forename: Alfred
authors.initials: A
authors.lastname: Pühler
authors.affiliation: Fakultät für Biologie, Lehrstuhl für Genetik, Universität Bielefeld, Postfach 100131, D-33501 Bielefeld, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schlüter
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000235:genetics*; D018528:ATP-Binding Cassette Transporters / Q000235:genetics; D017920:Antiporters / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D002511:Cephalosporins / Q000494:pharmacology; D004251:DNA Transposable Elements / Q000235:genetics; D004269:DNA, Bacterial / Q000737:chemistry; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004917:Erythromycin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009268:Nalidixic Acid / Q000494:pharmacology; D009713:Nucleotidyltransferases / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000235:genetics; D011815:R Factors / Q000235:genetics*; D017422:Sequence Analysis, DNA; D012722:Sewage / Q000382:microbiology*; D000198:Spectinomycin / Q000494:pharmacology; D013307:Streptomycin / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D026801:Synteny; D013753:Tetracycline Resistance / Q000235:genetics; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics; D014296:Trimethoprim Resistance; D014974:Xanthomonas / Q000235:genetics; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D018528:ATP-Binding Cassette Transporters; D017920:Antiporters; D001426:Bacterial Proteins; D002511:Cephalosporins; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; C083180:Mob protein, Bacteria; D012722:Sewage; D013449:Sulfonamides; D014157:Transcription Factors; C079051:tetA protein, Bacteria; D009268:Nalidixic Acid; D004917:Erythromycin; D000198:Spectinomycin; D013762:Tetrahydrofolate Dehydrogenase; D017853:Phosphotransferases (Alcohol Group Acceptor); C061369:macrolide 2'-kinase; D009713:Nucleotidyltransferases; C022682:streptomycin-spectinomycin adenylyltransferase; D001618:beta-Lactamases; D013307:Streptomycin