• Biomedical Science Letters
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• v.10 no.4
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• pp.507-514
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• 2004

The Pseudomonas aeruginosa isolated from the clinical specimens has a mutation on the QRDR (quinolone resistance determining region). There were obvious mutations in both gyrA and parC gene which are major targets of quinolone. Simultaneous mutations were found two sites or more on these genes in all of ten strains. GyrB or parE gene had only silent mutation without converted amino acids. We confirmed that P. aeruginosa from clinical specimens exhibited decreased sensitivity to fluroquiolone due to changed Thr-83→lle and Asp-87→Asn types on gyrA and altered Ser-87→Leu type on parC. This is the first finding that a new Met-93→Thr type on parC as well as mutations on gyrB or parE genes differed from existing patterns. This study showed more mutations of gyrA rather than parC, suggesting that change of Type Ⅳ topoisomerase is more serious than that of type Ⅱ (DNA gyrase).

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.70.2-71
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• 2003

Quinolone resistance in Streptococcus pneumoniae is related to mutations in the DNA gyrase and topoisomerase IV genes. DW-286a displayed potent activity against S. pneumoniae C9211 (MIC, 0.015 ${\mu}$g/ml) compared with gemifloxacin (MIC, 0.06 ${\mu}$g/ml). This study was performed to analyze the ability of DW-286a to cause resistance development in S. pneumoniae and to establish whether DNA gyrase or topoisomerase IV is primary target. DW-286a resistant mutants of S. pneumoniae C9211 were generated by stepwise selection at increasing drug concentration. (omitted)

• YAKHAK HOEJI
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• v.52 no.3
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• pp.219-224
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• 2008

Clinically isolated methicillin-resistant Staphylococcus aureus strains were exposed to subinhibitory concentration of DW286, DW-224a, gemifloxacin, trovafloxacin, sparfloxacin and ciprofloxacin during 26- to 39-days period. Subculturing led to resistance development, and most of the selected mutants were above susceptible breakpoints. Selected mutants had broad cross resistance to other quinolone antibiotics and only one mutant was completely susceptible to all fluoroquinolones. Twenty five among 42 mutants revealed mutations on DNA gyrase and topoisomerase IV by sequencing. Also 16 mutants had fluoroquinolones MICs that were 4-32 times lower in the presence of reserpine. In conclusion, alterations in DNA gyrase or topoisomerase IV and action of efflux pumping out system are the resistance mechanisms of DW-224a.

• Korean Journal of Clinical Laboratory Science
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• v.48 no.2
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• pp.74-81
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• 2016

Ureaplasma species can normally colonize in the bodies of healthy individuals. Their colonization is associated with various diseases including non-gonococcal urethritis, chorioamnionitis, neonatal meningitis, and prematurity. In 2012, the sum of the resistant and intermediate resistant rates of Ureaplasma spp. to ofloxacin and ciprofloxacin was 66.08% and 92.69%, respectively. DNA point mutations in the genes encoding DNA gyrase (topoisomerase II) and topoisomerase IV are commonly responsible for fluoroquinolone resistance. Each enzyme is composed of two subunits encoded by gyrA and gyrB genes for DNA gyrase and parC and parE genes for topoisomerase IV. In the current study, these genes were sequenced in order to determine the role of amino acid substitutions in Ureaplasma spp. clinical isolates. From December 2012 to May 2013, we examined mutation patterns of the quinolone resistance-determining region (QRDR) in Ureaplasma spp. DNA sequences in the QRDR region of Ureaplasma clinical isolates were compared with those of reference strains including U. urealyticum serovar 8 (ATCC 27618) and U. parvum serovar 3 (ATCC 27815). Mutations were detected in all ofloxacin- and ciprofloxacin-resistant isolates, however no mutations were detected in drug-susceptible isolates. Most of the mutations related to fluoroquinolone resistance occurred in the parC gene, causing amino acid substitutions. Newly found amino acid substitutions in this study were Asn481Ser in GyrB; Phe149Leu, Asp150Met, Asp151Ile, and Ser152Val in ParC; and Pro446Ser and Arg448Lys in ParE. Continuous monitoring and accumulation of mutation data in fluoroquinolone-resistant Ureaplasma clinical isolates are essential to determining the tendency and to understanding the mechanisms underlying antimicrobial resistance.

• Journal of Microbiology
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• v.45 no.2
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• pp.168-170
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• 2007

In this research 26 Shigella isolates were examined by PCR and direct nucleotide sequencing for genetic alterations in the quinolone-resistance determining regions (QRDRs). We tested for the presence of qnr genes by PCR in 91 strains, but no qnr genes were found. The results did show, however, some novel mutations at codon 83 of gyrA ($Ser{\rightarrow}Ile$) and codon 64 of parC ($Ala64{\rightarrow}Cys,\;Ala64{\rightarrow}Asp$), which were related to fluroquinolone resistance.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.47 no.3
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• pp.247-254
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• 2014

To investigate the acquisition of quinolone resistance, we examined mutations in the quinolone resistance-determining region (QRDR) of type II topoisomerase genes in ciprofloxacin (CIP)-resistant clinical isolates and in vitro mutants of Streptococcus parauberis. The CIP-resistant clinical isolates had one base change responsible for a Ser-79${\rightarrow}$Thr in the QRDR of parC. However, the CIP-resistant in vitro mutants had an altered QRDR of parC (Ser-79${\rightarrow}$Ile) that differed from that of the isolates. None of the CIP-resistant S. parauberis clinical isolates or in vitro mutants exhibited amino acid changes in gyrA or gyrB. However, even though involvement in the increased resistance was not clear, an Arg-449${\rightarrow}$Ser mutation outside of the QRDR of parE was detected in CIP-resistant mutant 2P1. These results suggest that the topoisomerase IV gene, parC (and possibly parE, as well), is the primary ciprofloxacin target in S. parauberis. Additionally we established a high-resolution melting (HRM) assay capable of detecting the dominant mutation in four type II topoisomerase genes conferring ciprofloxacin resistance. These rapid and reliable assays may provide a convenient method of surveillance for genetic mutations conferring antibiotic resistance.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1997.04a
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• pp.40-47
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• 1997

New quinolones generally have a broad antibacterial spectrum against gram-positive, gram-negative, glucose-nonfermenting and anaerobic bacteria. Some of newly developed quinolones have potent activities against S. aureus including MRSA, S.pneumoniae including PRSP, B. fragilis, chlamydiae, mycoplasmas and mycobacteria as well, and show good activities against various strains resistant to antibacterial agents of other classes. Quinolones display postantibiotic effects in vitro and are bactericidal at concentrations similar to or twice that of the minimum inhibitory concentrations (MICs) for susceptible pathogens. In experimental murine infection models including systemic infections with various pathogens such as S. aureus, S. pyogenes, S. pneumoniae, E. coli and P. aeruginosa, quinolones have shown good oral efficacy as well as parenteral efficacy. Good oral absorption and good tissue penetration of quinolones account for good therapeutic effects in clinical settings. The target of quinolones are two structurally related type II topoisomerases, DNA gyrase and DNA topoisomerase IV. Quinolones are shown to stabilize the ternary quinolone-gyrase-DNA complex and inhibit the religation of the cleaved double-stranded DNA. Bacteria can acquire resistance to quinolones by mutations of these target enzymes. Mutation sites and amino acid changes in DNA gyrase and DNA topoisomerase IV are similar in the organisms examined, suggesting that the mechanism of quinolone resistance in the target enzymes is essentially the same among various organisms. Quinolones act on both the target enzymes to different degrees depending on the organisms or agents tested, and bacteria become highly resistant to quinolones in a step-wise fashion. Incomplete cross-resistance among quinolones in some strains of E. coli and S. aureus suggests the possibility of finding quinolones active against quinolone-resistant strains which are prevailing now. To find such quinolones, the potency toward two target enzymes and the membrane permeability including influx and/or efflux systems should be taken into account.

• Environmental Mutagens and Carcinogens
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• v.9 no.1
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• pp.13-22
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• 1989

The effect of novobiocin (NOV), and inhibitor of topoisomerase II, on ethyl methanesulfonate (EMS)-or bleomycin (BLM)-induced DNA repair synthesis was examined during the cell cycle of Chinese hamster ovary (CHO)-K1 cells. Three assays were employed in this study: cell survival, alkaline elution and unscheduled DNA synthesis. EMS was effective at killing CHO cells in G1 phase, wheras BLM preferentially killed cells in G2 and S phases. EMS induced the much more amount of DNA damage in G1 phase, while BLM induced in G2 phase than the other phases. The both of pre- and post-treatment with BOV inhibitied EMS- or BLM-induced DNA repair synthesis in G1 and G2 phases, and pretreatment with NOV inhibited more effectively than the post-treated group. These results suggested that CHO cells exhibited a differential sensitivity to cell lethality and DNA damage in relation to cell cycle according to used chemical agents, and that DNA topoisomerase II participated in an initial stage of DNA repair.

• Molecules and Cells
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• v.20 no.3
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• pp.392-400
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• 2005

The genes encoding the DNA gyrase A (GyrA) and B subunits (GyrB) of Methylovorus sp. strain SS1 were cloned and sequenced. gyrA and gyrB coded for proteins of 846 and 799 amino acids with calculated molecular weights of 94,328 and 88,714, respectively, and complemented Escherichia coli gyrA and gyrB temperature sensitive (ts) mutants. To analyze the role of type II topoisomerases in the intrinsic quinolone resistance of methylotrophic bacteria, the sequences of the quinolone resistance-determining regions (QRDRs) in the A subunit of DNA gyrase and the C subunit (ParC) of topoisomerase IV (Topo IV) of Methylovorus sp. strain SS1, Methylobacterium extorquens AM1 NCIB 9133, Methylobacillus sp, strain SK1 DSM 8269, and Methylophilus methylotrophus NCIB 10515 were determined. The deduced amino acid sequences of the QRDRs of the ParCs in the four methylotrophic bacteria were identical to that of E. coli ParC. The sequences of the QRDR in GyrA were also identical to those in E. coli GyrA except for the amino acids at positions 83, 87, or 95. The $Ser^{83}$ to Thr substitution in Methylovorus sp. strain SS1, and the $Ser^{83}$ to Leu and $Asp^{87}$ to Asn substitutions in the three other methylotrophs, agreed well with the minimal inhibitory concentrations of quinolones in the four bacteria, suggesting that these residues play a role in the intrinsic susceptibility of methylotrophic bacteria to quinolones.

• Journal of Pharmaceutical Investigation
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• v.39 no.6
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• pp.431-435
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• 2009

The comparative superior in vitro activity of DW-224a was supported by the downward MIC distribution due to the weakened influence of alterations within target enzymes in ciprofloxacin-resistant Staphylococcus aureus. The MI$C_{50}$ for DW-224a was 4 $\mu$g/mL, similar to that of gemifloxacin, 8-fold less than that of sparfloxacin and 16-over-fold less than that of ciprofloxacin. We constructed combinations of amino acid changes, located at codon 80, 83 or 84 within GrlA and 84, 85 or 88 within GyrA, which were associated with MIC increase. The amino acid changes were less influential to the MIC of DW-224a compared to those of other fluoroquinolones, and it was verified from the requirement of a total of two GrlA- and two GyrA-alterations to reach the MIC of DW-224a over 32 $\mu$g/mL.