Although quinolone resistance usually results from chromosomal mutations, recent studies indicate that quinolone resistance can also be plasmid mediated. The gene responsible, qnr , is distinct from the known quinolone resistance genes and in previous studies seemed to be restricted to Klebsiella pneumoniae and Escherichia coli isolates from the University of Alabama in Birmingham, where this resistance was discovered. In Shanghai, the frequency of ciprofloxacin resistance in E. coli has exceeded 50% since 1993. Seventy-eight unique ciprofloxacin-resistant clinical isolates of E. coli from Shanghai hospitals were screened for the qnr gene by colony blotting and Southern hybridization of plasmid DNA. Conjugation experiments were done with azide-resistant E. coli J53 as a recipient with selection for plasmid-encoded antimicrobial resistance (chloramphenicol, gentamicin, or tetracycline) and azide counterselection. qnr genes were sequenced, and the structure of the plasmid DNA adjacent to qnr was analyzed by primer walking with a sequential series of outward-facing sequencing primers with plasmid DNA templates purified from transconjugants. Six (7.7%) of 78 strains gave a reproducible hybridization signal with a qnr gene probe on colony blots and yielded strong signals on plasmid DNA preparations. Quinolone resistance was transferred from all six probe-positive strains. Transconjugants had 16- to 250-fold increases in the MICs of ciprofloxacin relative to that of the recipient. All six strains contained qnr with a nucleotide sequence identical to that originally reported, except for a single nucleotide change (CT A →CT G at position 537) encoding the same amino acid. qnr was located in complex In4 family class 1 integrons. Two completely sequenced integrons were designated In36 and In37. Transferable plasmid-mediated quinolone resistance associated with qnr is thus prevalent in quinolone-resistant clinical strains of E. coli from Shanghai and may contribute to the rapid increase in bacterial resistance to quinolones in China.

The bacterial resistance surveillance system is relatively well established at the national, provincial, and hospital levels in China. Two representative national surveillance networks for bacterial resistance are the China Antimicrobial Resistance Surveillance System (CARSS) and the China Antimicrobial Surveillance Network (CHINET), both established in 2005. CARSS data show the different bacterial resistance rates among different provinces and autonomous regions for each specific bacterium. CHINET data mainly represent the bacterial resistance profiles of teaching hospitals and show the changing trends of bacterial resistance in China. For clinical isolates, the ratio of gram-negative bacilli to gram-positive cocci is approximately 7 to 3. In general, gram-negative bacilli have higher antimicrobial resistance profiles in China. Regarding different bacterial species, antimicrobial resistance is multifaceted. The prevalence of extended-spectrum β-lactamases is high; Acinetobacter baumannii has a high antimicrobial resistance profile; and, notably, the prevalence of CRKP has been showing a marked increase since 2005. In addition, the prevalence of vancomycin-resistant Enterococcus is low, and the prevalence of methicillin-resistant Staphylococcus aureus and antimicrobial resistance in Pseudomonas aeruginosa showed decreasing trends from 2005 to 2017.

Since the discovery of qnrA in 1998, two additional qnr genes, qnrB and qnrS , have been described. These three plasmid-mediated genes contribute to quinolone resistance in gram-negative pathogens worldwide. A clinical strain of Proteus mirabilis was isolated from an outpatient with a urinary tract infection and was susceptible to most antimicrobials but resistant to ampicillin, sulfamethoxazole, and trimethoprim. Plasmid pHS10, harbored by this strain, was transferred to azide-resistant Escherichia coli J53 by conjugation. A transconjugant with pHS10 had low-level quinolone resistance but was negative by PCR for the known qnr genes, aac(6′)-Ib-cr and qepA . The ciprofloxacin MIC for the clinical strain and a J53/pHS10 transconjugant was 0.25 μg/ml, representing an increase of 32-fold relative to that for the recipient, J53. The plasmid was digested with HindIII, and a 4.4-kb DNA fragment containing the new gene was cloned into pUC18 and transformed into E. coli TOP10. Sequencing showed that the responsible 666-bp gene, designated qnrC , encoded a 221-amino-acid protein, QnrC, which shared 64%, 42%, 59%, and 43% amino acid identity with QnrA1, QnrB1, QnrS1, and QnrD, respectively. Upstream of qnrC there existed a new IS 3 family insertion sequence, IS Pmi1 , which encoded a frameshifted transposase. qnrC could not be detected by PCR, however, in 2,020 strains of Enterobacteriaceae . A new quinolone resistance gene, qnrC , was thus characterized from plasmid pHS10 carried by a clinical isolate of P . mirabilis .

Since glycopeptide-resistant enterococci (GRE) were reported in 1988, they have appeared in hospitals worldwide. Seven van gene cluster types ( vanA , vanB , vanC , vanD , vanE , vanG , and vanL ) are currently known. We investigated a clinical strain of Enterococcus faecium Efm-HS0661 that was isolated in 2006 from an inpatient with intra-abdominal infection in Shanghai. It was resistant to most antimicrobials, including vancomycin (MIC, >256 μg/ml) and teicoplanin (MIC, 96 μg/ml). Glycopeptide resistance could be transferred to E . faecium BM4105RF by conjugation. The donor and its transconjugant were negative by PCR for the known van genes. By cloning and primer walk sequencing, we discovered a novel van gene cluster, designated vanM . The vanM ligase gene was 1,032-bp in length and encoded a 343-amino-acid protein that shared 79.9, 70.8, 66.3, and 78.8% amino acid identity with VanA, VanB, VanD, and VanF, respectively. Although the vanM DNA sequence was closest to vanA , the organization of the vanM gene cluster was most similar to that of vanD . Upstream from the vanM cluster was an IS 1216- like element, which may play a role in the dissemination of this resistance determinant. Liquid chromatography-mass spectrometry analysis of peptidoglycan precursors extracted from the VanM-type strain Efm-HS0661 treated with vancomycin or teicoplanin revealed a modified precursor (UDP- N -acetylmuramic acid [MurNAc]-tetrapeptide- d -Lac), indicating that VanM, like VanA, confers glycopeptide resistance by the inducible synthesis of precursor ending in d -Ala- d -Lac.

Although quinolone resistance commonly results from chromosomal mutation, recent studies indicate that such resistance can also be transferred on plasmids carrying the gene responsible, qnr . One hundred ten ciprofloxacin-resistant clinical isolates of Klebsiella pneumoniae and Escherichia coli from the United States were screened for the qnr gene by PCR and Southern hybridization of plasmid DNA. Conjugation experiments were done with azide-resistant E. coli J53 as the recipient and selection with azide and sulfonamide, a resistance frequently linked to qnr . EcoRI and BamHI digests of qnr -hybridizing plasmids were subjected to electrophoresis on agarose gels and probed with qnr by Southern hybridization. qnr was detected in 8 (11.1%) of 72 K. pneumoniae strains. These eight positive strains were from six states in the United States. qnr was not found in any of the 38 E. coli strains tested. Quinolone resistance was transferred from seven of the eight probe-positive strains. Transconjugants with qnr -hybridizing plasmids had 32-fold increases in ciprofloxacin MICs relative to E. coli J53. For all eight strains, the sequence of qnr was identical to that originally reported. By size and restriction digests, four plasmids were related to the first-reported plasmid, pMG252, and three were different. Five new qnr plasmids encoded FOX-5 β-lactamase, as did pMG252, but two others produced SHV-7 extended-spectrum β-lactamase. Transferable plasmid-mediated quinolone resistance associated with qnr is now widely distributed in quinolone-resistant clinical strains of K. pneumoniae in the United States. Plasmid-determined quinolone resistance contributes to the increasing quinolone resistance of K. pneumoniae isolates and to the linkage previously observed between resistance to quinolones and the latest β-lactam antibiotics.