• Biomedical Science Letters
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• v.13 no.2
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• pp.141-148
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• 2007

We determined the sequences of the quinolone resistance-determining region (QRDR) of gyrA and gyrB for 21 clinical strains of Enterobacteriaceae resistant to ciprofloxacin, norfloxacin and levofloxacin. The clinical strains were isolated from the specimens of three general hospitals in Busan. In the present study, we found mutations in type II topoisomerase (DNA gyrase) genes for all strains. We confirmed that some genera of Enterobacteriaceae of clinical specimen exhibited decreased sensitivity to fluroquinolone due to changes in Ser-83$\rightarrow$Leu and Asp-87$\rightarrow$Asn types on gyrA and alterations in Glu-465$\rightarrow$Arg and Ser-492$\rightarrow$Asn type on gyrB. All the twenty-one strains had a missense mutation in gyrA (codon 83 and 87). Three of them had an additional mutation in gyrB (codon 465 or 492), but one of them had an additional mutation in gyrB (codon 426, 427, 491, 495 and 496). The strains which had two mutations in type II topoisomerase genes (gyrA and gyrB) were significantly more resistant to fluoroquinolones than those with a single mutation in gyrA (mean MICs of ciprofloxacin: $\geq8\mu$g/ml, mean MICs of levofloxacin: $\geq16\mu$g/ml). Interestingly, the examination of silent nucleotide changes n the gyrA and gyrB genes revealed six different patterns of DNA polymorphism, respectively. Fifteen strains of the twenty-one strains bearing the gyrase A mutation shared the same polymorphism and eleven strains of the twenty-one strains bearing the gyrase B mutation shared the same polymorphism.

• Journal of Microbiology and Biotechnology
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• v.16 no.12
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• pp.1841-1848
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• 2006

Insights into gene expression have the potential for improvement of antibiotic yield and the development of robust production hosts for use in recombinant biomolecule production. $Cubicin^{TM}$ (daptomycin for injection) is a recently approved antibiotic active against many Gram(+) pathogens, including those resistant to methicillin, vancomycin, and fluoroquinolones. Daptomycin is produced as a secondary metabolite by Streptomyces roseosporus. A 128 kb region of DNA including the daptomycin biosynthetic gene cluster (dpt) has been cloned. and sequenced. Using a selected array of nucleic acid probes representing this region, we compared the expression levels of the dpt genes between S. roseosporus wild-type (WT) and derived S. roseosporus high-producer of daptomycin (HP). We observed that the majority of the biosynthetic genes were upregulated in HP compared with WT; a total of 12 genes, including those encoding daptomycin synthetase, showed consistently and significantly higher expression levels, at least 5-fold, in HP compared with WT. In contrast, some genes, flanking the dpt cluster, were expressed at higher levels in the WT strain. The expression of housekeeping genes such as S. roseosporus rpsL, rpsG, and 16S (positive controls) and presumptive intergenic regions in the dpt cluster (negative control) were identical in the two strains. In addition, we compared transcription during the early, mid-log, and early-stationary phases of growth in the HP strain. The same set of genes was upregulated and downregulated under all conditions examined; housekeeping genes showed no relative change in expression level over the periods of growth tested. Analyses of this type would be of value in studies of strain improvement and also for the identification of gene regulation processes that are important for secondary metabolite production.

• 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.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.41 no.6
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• pp.440-445
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• 2008

Ciprofloxacin is the most commonly used fluoroquinolones for treating bacterial disease in olive flounder (Paralichthys olivaceus) farming, but its withdrawal time for industrial-scale farming has not been established. The depletion of ciprofloxacin was investigated in the olive flounder under field conditions. Fish were kept in an inland fish farm and fed a commercial diet containing 5 mg/kg of ciprofloxacin for 13 days. Seven fishes per sampling time were examined during and after the treatment. Ciprofloxacin and its major metabolite, enrofloxacin, were analyzed using HPLC with a fluorescence detector. The concentrations of ciprofloxacin and enrofloxacin in muscle increased during the medication period, and then decreased rapidly. The ciprofloxacin and enrofloxacin concentration in the olive flounder peaked on days 11 and 13, respectively, with maximum concentrations in muscle of 0.58 and 0.73 mg/kg. Residual ciprofloxacin and enrofloxacin were eliminated rapidly; 6 days after treatment, the respective levels in muscle were 0.04 and 0.10 mg/kg and neither was detected 15 days post treatment. The level of ciprofloxacin accumulation at the beginning of oral administration was variable according to the farming conditions, but the overall exhaustion time was similar. In conclusion, an adequate withdrawal period for enrofloxacin is 15 days in the case of oral ciprofloxacin administration.

• Bulletin of the Korean Chemical Society
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• v.35 no.11
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• pp.3275-3279
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• 2014

A mass balance method for purity assessment of thermolabile organic reference materials was established by combining several techniques, including liquid chromatography with UV/VIS detector (LC-UV), Karl-Fischer (K-F) Coulometry, and thermal gravimetric analysis (TGA). This method was applied to three fluoroquinolones like enrofloxacin, norfloxacin and ciprofloxacin. LC-UV was used to analyze structurally related organic impurities based on UV/VIS absorbance spectra obtained in combination with LC separation. For all three organic reference materials, the UV/VIS spectra of the separated impurities were similar to that of the major component of the corresponding materials. This indicates that the impurities are structurally related to the respective reference material sharing common chromophores. Impurities could be quantified by comparing their absorbances at the wavelength of maximum absorbance (${\lambda}_{max}$). The water contents of the reference materials were measured by K-F Coulometry by an oven-drying method. The total inorganic impurities contents were assayed from ash residues in TGA analysis with using air as a reagent gas. The final purities estimated from results of those analytical techniques were assigned as ($99.91{\pm}0.06$), ($97.09{\pm}0.17$) and ($91.85{\pm}0.17$)% (kg/kg) for enrofloxacin, norfloxacin and ciprofloxacin, respectively. The assigned final purities would be applied to the reference materials which will be used as calibrators for the certification of those compounds in matrix CRMs as starting points for the traceability of their certified values to SI units.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.39 no.2
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• pp.72-77
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• 2006

Enrofloxacin (ENRO) is one of the most commonly used fluoroquinolones for treating bacterial disease in olive flounder (Paralichthys olivaceus) farming, but its withdrawal time for industrial-scale farming has not been well established. Withdrawal times of ENRO following oral administration were evaluated in olive flounder under field conditions. Fish were held in an inland fish tank and fed a commercial mediated diet containing 5 mg/kg of ENRO for 9 days. Seven fish per sampling point were examined during and after treatment. ENRO and its major metabolite, ciprofloxacin (CIP), were analyzed using high-performance liquid chromatography with a fluorescence detector. The concentration of ENRO and CIP in muscle increased during the medication period, and then decreased rapidly The sum of ENRO and CIP concentration in olive flounder peaked on day 6, with a maximal concentration in muscle of 4.30 mg/kg. ENRO residues were eliminated rapidly; at 10 days post treatment, the level in muscle was 0.10 mg/kg, but it took about 50 days to be reduced to below 0.1 mg/kg. After 60 days, the residual concentration was below 0.1 mg/kg in all samples. The level of ENRO accumulation at the beginning of oral administration was variable, according to the farming conditions, but the overall exhaustion time was almost the same. We concluded that an adequate withdrawal period of enrofloxacin is 60 days in the case of oral administration.

• Journal of Microbiology and Biotechnology
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• v.22 no.4
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• pp.567-570
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• 2012

In vitro activities of 13 antibiotics were assessed against 85 Brucella abortus isolates from naturally infected cattle in the Republic of Korea during 1998-2006, using broth microdilution test. Tetracyclines showed the most excellent activity against B. abortus, displaying MIC values of 0.5 ${\mu}g/ml$ or below. In particular, minocycline showed the lowest $MIC_{50/90}$ values (0.125/0.125 ${\mu}g/ml$) in this study. Among four fluoroquinolones tested, ciprofloxacin ($MIC_{50/90}$, 0.5/1 ${\mu}g/ml$) and norfloxacin ($MIC_{50/90}$, 8/8 ${\mu}g/ml$) had the most and the least activities, respectively. Gentamicin ($MIC_{50/90}$, 1/1 ${\mu}g/ml$) was more effective than streptomycin, erythromycin, rifampin, and chloramphenicol ($MIC_{50/90}$, 2/2 ${\mu}g/ml$).

• Korean Journal of Veterinary Research
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• v.60 no.3
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• pp.173-177
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• 2020

Edible offal is easily contaminated by Escherichia coli (E. coli) and fluoroquinolone (FQ)-resistant E. coli is considered a serious public health problem, thus, this study investigated the genetic characteristics of FQ-resistant E. coli from edible offal. A total of 22 FQ-resistant E. coli isolates were tested. A double mutation in each gyrA and parC led the highest MIC. Four (18.2%) isolates carried plasmid-mediated quinolone resistance genes. The fimH, eaeA, escV, astA, and iucC genes were confirmed. Seventeen isolates (77.3%) were positive for plasmid replicons. The isolates showed high genetic heterogeneity based on pulsed-field gel electrophoresis patterns.

• Bulletin of the Korean Chemical Society
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• v.25 no.3
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• pp.365-372
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• 2004

Two simple, rapid and sensitive spectrophotometric methods for the determination of three fluoroquinolones, namely levofloxacin, norfloxacin and ciprofloxacin have been performed either in pure form or in their tablets. In the first method, levofloxacin and norfloxacin are directly treated with bromocresol green (BCG) in dichloromethane while ciprofloxacin is allowed to react with the same dye in aqueous acidic buffer. Highly yellow colored complex species were formed instantaneously in case of levofloxacin and norfloxacin or after extraction into dichloromethane for ciprofloxacin. The formed complexes are quantified spectrophotometrically at their absorption maxima at 411 nm for levofloxacin and 412 nm for norfloxacin and ciprofloxacin. The second method involves the reaction of levofloxacin with ${\rho}$-chloranilic acid ( ${\rho}$-CA) and norfloxacin with tetracyanoethylene (TCNE) in acetonitrile to give complexes with maximum absorbance at 521 and 333 nm for the two drugs, respectively. Adopting the first procedure, calibration graphs were linear over the range 1- 20 ${\mu}g\;mL^{-1}$ with mean percentage recoveries of 100.41 ${\pm}$ 0.72, 99.99 ${\pm}$ 0.54 and 100.23 ${\pm}$ 0.91 for the theree drugs, respectively. For the second procedure, the concentration ranges were 15-250 ${\mu}g\;mL^{-1}$ for levofloxacin using ${\rho}$-CA and 0.8-16 ${\mu}g\;mL^{-1}$ for norfloxacin using TCNE with mean percentage recoveries of 99.88 ${\pm}$ 0.45 and 100.26 ${\pm}$ 0.68 for the two drugs, respectively. The proposed methods were successfully applied to determine these drugs in their tablet formulations and the results compared favorably to that of reference methods. The proposed methods are recommended for quality control and routine analysis.

• Journal of Microbiology and Biotechnology
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• v.18 no.11
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• pp.1853-1857
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• 2008

Salmonella contamination in chicken meat was studied with 100 chicken meat samples purchased from 55 shops located in various regions. A total of 21 isolates of Salmonella enterica were isolated from 21 chicken meat samples from four shops located at open markets, whereas there were none from supermarkets with well-equipped cold systems. Among these, 18 isolates were identified as Salmonella enterica serotype Haardt (S. Haardt) and three isolates were S. enterica serotype Muenchen. When the minimal inhibitory concentrations of the S. Haardt isolates were assayed with the agar dilution method to determine susceptibility to ampicillin, chloramphenicol, sulfisoxazole, tetracycline, and nalidixic acid, all 18 isolates were resistant to tetracycline and nalidixic acid and nine of these were resistant to ampicillin. These isolates showed reduced susceptibility to eight fluoroquinolones including ciprofloxacin, enrofloxacin, levofloxacin, gatifloxacin, gemifloxacin, moxifloxacin, norfloxacin, and ofloxacin. When quinolone resistance determining regions of gyrA and gyrB were sequenced, every isolate had the same missense mutation Ser83$\rightarrow$Tyr (TCC$\rightarrow$+TAC) in gyrA, whereas no mutation was found in gyrB. Pulsed-field gel electrophoresis with XbaI revealed a close relationship among these isolates, suggesting a contamination of raw chicken meat with clonal spread of nalidixic acid-resistant and quinolone-reduced susceptibility S. Haardt in chickens. Results in this study show the importance of a well-equipped cold system and the prudent use of fluoroquinolone in chickens to prevent the occurrence of quinolone-resistant isolates.