title: Transfer of quinolone resistance gene qnrA1 to Escherichia coli through a 50 kb conjugative plasmid resulting from the splitting of a 300 kb plasmid.
abstract: OBJECTIVE
  To analyse the in vitro transfer of the qnrA1 gene by a 50 kb (pSZ50) self-transferable plasmid that derives from a 300 kb plasmid (pSZ300) and to determine the complete nucleotide sequence of plasmid pSZ50.
  
  
  METHODS
  Extended-spectrum β-lactamase (ESBL) and plasmid-mediated quinolone resistance (PMQR) genes of an Escherichia coli clinical isolate were analysed. Plasmid analysis included conjugation and selection on seven antibiotics examined by antimicrobial susceptibility testing, RFLP comparison, Southern hybridization, incompatibility group identification and shotgun sequencing.
  
  
  RESULTS
  The E. coli 5509 isolate carries the genes encoding the ESBL CTX-M-15 and the quinolone resistance determinants qnrA1, qnrB2 and aac(6')-Ib-cr on a 300 kb plasmid. Seven transfer resistances were analysed by conjugation under two conditions (30 and 37°C), leading to two distinct transconjugant phenotypes with different resistances. Transconjugants of phenotype A harboured a 300 kb plasmid named pSZ300 that conferred resistance to eight antibiotics and harboured the qnrA1, aac(6')-Ib-cr and bla(CTX-M-15) genes. Transconjugants of phenotype B were resistant to three antibiotics and they harboured the qnrA1 gene on an ≈ 50 kb plasmid named pSZ50. Both plasmids were self-transferable at a frequency of 1 × 10(-3). Plasmid pSZ300 was typed to be both an IncF and IncN plasmid, whereas pSZ50 corresponded only to type IncN. Fingerprinting and Southern hybridization showed that plasmid pSZ50 derived from pSZ300. The complete nucleotide sequence of plasmid pSZ50 was determined (51556 bp) and 55 open reading frames were predicted. The qnrA1 gene was identified in a tandem duplicate inside a sul1-type integron structure.
  
  
  CONCLUSIONS
  The plasmid pSZ300 represented a fusion of two replicons (IncF and IncN), and our observations suggest that the plasmid pSZ50 (IncN) may split and transfer antibiotic resistance determinants. This mechanism could be advantageous in the dissemination of antibiotic resistance genes.
authors.affiliation: Instituto Nacional de Salud Pública, CISEI, Cuernavaca, Morelos, Mexico.
authors.forename: Ulises
authors.initials: U
authors.lastname: Garza-Ramos
authors.forename: Humberto
authors.initials: H
authors.lastname: Barrios
authors.forename: Maria Jose
authors.initials: MJ
authors.lastname: Hernandez-Vargas
authors.forename: Teresa
authors.initials: T
authors.lastname: Rojas-Moreno
authors.forename: Fernando
authors.initials: F
authors.lastname: Reyna-Flores
authors.forename: Perla
authors.initials: P
authors.lastname: Tinoco
authors.forename: Victor
authors.initials: V
authors.lastname: Othon
authors.forename: Laurent
authors.initials: L
authors.lastname: Poirel
authors.forename: Patrice
authors.initials: P
authors.lastname: Nordmann
authors.forename: Vincent
authors.initials: V
authors.lastname: Cattoir
authors.forename: Guillermo
authors.initials: G
authors.lastname: Ruiz-Palacios
authors.forename: Jose Luis
authors.initials: JL
authors.lastname: Fernandez
authors.forename: Rosa Isela
authors.initials: RI
authors.lastname: Santamaria
authors.forename: Patricia
authors.initials: P
authors.lastname: Bustos
authors.forename: Natividad
authors.initials: N
authors.lastname: Castro
authors.forename: Jesus
authors.initials: J
authors.lastname: Silva-Sanchez
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D015139:Blotting, Southern; D003227:Conjugation, Genetic; D016172:DNA Fingerprinting; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D024881:Drug Resistance, Bacterial; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification; D004927:Escherichia coli Infections / Q000382:microbiology; D022761:Gene Transfer, Horizontal; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D012150:Polymorphism, Restriction Fragment Length; D015363:Quinolones / Q000494:pharmacology*; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D015363:Quinolones; D001618:beta-Lactamases