oqxAB Encoding a Multidrug Efflux Pump in Human Clinical Isolates of Enterobacteriaceae

Abstract

The genes for multidrug efflux pump OqxAB, which is active on fluoroquinolones, were found in human clinical isolates on a plasmid in Escherichia coli and on the chromosome of Klebsiella pneumoniae. IS26-like sequences flanked the plasmid-mediated oqxAB genes, suggesting that they had been mobilized as part of a composite transposon.Plasmid-borne genes conferring quinolone resistance have been increasingly recognized (7,10). Recently a plasmid-encoded efflux pump, OqxAB, conferring resistance to the quinoxaline-di-N-oxide olaquindox, which has been used as a growth promoter in pigs, was discovered in Escherichia coli isolates of porcine origin in Denmark and Sweden (4-6). OqxAB was encoded by the genes oqxA and oqxB located on a 52-kb conjugative plasmid designated pOLA52 and conferred resistance to multiple agents, including fluoroquinolones (4, 9). We have investigated the prevalence of this plasmid-encoded multidrug efflux pump in clinical isolates of Enterobacteriaceae and have for the first time identified an oqxAB-encoding plasmid in an E. coli isolate of human origin.Isolates were from the collection of blood isolates from Seoul National University Hospital collected from 1998 to 2006. The same set of isolates was previously surveyed for other plasmid-mediated quinolone resistance (PMQR) genes (8). A total of 461 clinical isolates were screened by PCR for the oqxA gene. Isolates positive for oqxA were also tested for oqxB, and strains positive for both genes were confirmed by sequencing of the PCR products. The primers used are shown in Table 1.One (0.4%) of 261 E. coli isolates, 3 (4.6%) of 65 Enterobacter cloacae isolates, and 100 (74.1%) of 135 Klebsiella pneumoniae isolates were provisionally classified as positive for both oqxA and oqxB. The oqxAB-positive E. coli was isolated from the blood of a patient in 1999. A BLAST search of the nucleotide sequence similarity of the oqxB PCR products obtained from the three E. cloacae isolates gave identities of only 88% (399/453) with pOLA52 (GenBank accession number EU370913) and 86% (394/454) with the hydrophobe/amphiphile efflux-1 (HAE1) family transporter of Enterobacter sp. strain 638 (GenBank accession number CP000653). There was, however, substantial similarity between the complete nucleotide sequences of the tandem oqxA and oqxB genes from E. coli 1-12 (GenBank accession number GQ120634; 99.5% and 99.0%, respectively), K. pneumoniae 4-39 (GenBank accession number FJ975560; 98.2% and 99.0%, respectively), and K. pneumoniae 5-80 (GenBank accession number FJ975561; 99.4% and 98.9%, respectively) relative to those of pOLA52 and the chromosomal genes in K. pneumoniae MGH78578 (GenBank accession number NC009648). Since the oqxAB genes appear to be chromosomal in K. pneumoniae (3, 9), E. coli 1-12 was the most likely candidate to contain an oqxAB-encoding plasmid (Table 2).To test for the plasmid location of oqxAB, plasmid DNAs were obtained using a plasmid midi kit (Qiagen, Valencia, CA) and hybridized with a horseradish peroxidase-labeled oqxB probe as prev...

Identification of strain harboring both aac(6')-Ib and aac(6')-Ib-cr variant simultaneously in Escherichia coli and Klebsiella pneumoniae

“…Among these, mutations at Ser-83 and Asp-87 have been found with a higher frequency in quinolone resistant E. coli clinical isolates than in susceptible isolates (19). The genes for multidrug efflux pump oqxAB, which is active on fluoroquinolones, were detected in human clinical isolates on a plasmid in E. coli and on the chromosome of K. pneumoniae (20). Therefore, in order to find the other resistance mechanisms against fluoroquinolones, we performed tests to characterize genetic mutation in QRDR of gyrA gene and oqxAB gene against used isolates in this study.…”

Plasmid-Mediated Quinolone Resistance

“…Linkage of oqxAB with genes for CTX-M-14 and other plasmid-mediated CTX-M alleles has been noted (160). It is common (usually 75% or more) on the chromosome of K. pneumoniae isolates, where up to 20-fold variation in expression implies the presence of regulatory control (73,203,206,208,209). In K. pneumoniae overexpression of the nearby rarA gene is associated with increased oqxAB expression, while increased expression of adjacent oqxR gene downregulates OqxAB production (210,211).…”

“…It has a wide substrate specificity including chloramphenicol, trimethoprim, and quinolones such as ciprofloxacin, flumequin, norfloxacin, and nalidixic acid (13). oqxAB has been found on plasmids in clinical isolates of E. coli and K. pneumoniae and in the chromosome and on plasmids of S. enteritis flanked in both locations by IS26-like elements (202)(203)(204)(205)(206)(207). In E. coli isolates from farms in China where olaquindox was in use, oqxAB was found on transmissible plasmids in 39% of isolates from animals and 30% of isolates from farm workers (204).…”

The Challenge of Efflux-Mediated Antibiotic Resistance in Gram-Negative Bacteria

“…In addition to MDR mediated by the chromosomally encoded OqxAB pump (302)(303)(304), overexpression of the newly identified RND-type KpgABC pump was involved in tigecycline nonsusceptibility due to an IS element insertion in the promoter region of kpgABC (305). The KexD RND pump expressed by cloning provides MDR with the requirement of AcrA accessory and KocC OM proteins (306).…”