Analysis of gyrA mutations in quinolone-resistant and -susceptible Campylobacter jejuni isolates from retail poultry and human clinical isolates by non-radioactive single-strand conformation polymorphism analysis and DNA sequencing

AIMS: The aims of this study were to characterize the molecular variations in the quinolone resistance-determining region (QRDR) of gyrA among quinolone-resistant and -susceptible Campylobacter jejuni isolates originating from foods of animal origin and human infections and to evaluate the suitability of the single-strand conformation polymorphism (SSCP) method as a screening method for molecular characterization of fluoroquinolone resistance. METHODS AND RESULTS: Alterations in QRDR of gyrA from 182 C. jejuni isolates were determined by nonradioisotopic SSCP analysis and direct sequencing. A total of 13 types of nucleic acid sequence combinations within the QRDR of the gyrA gene resulted in 11 different SSCP patterns. All nalidixic acid resistant strains possessed nucleotide substitution at either codon Thr-86 or Asp-90. Silent mutations were detected additionally. Thr-86 to Ile mutation was detected in all 139 ciprofloxacin resistant strains, which showed cross-resistance to nalidixic acid. CONCLUSIONS: The SSCP method is suitable for a molecular screening of quinolone resistant C. jejuni isolates and in combination with DNA sequencing suitable to detect genetic variations of the QRDR of gyrA. SIGNIFICANCE AND IMPACT OF STUDY: This study provides data of the genetic variations of the QRDR of gyrA from C. jejuni isolates of foods and human beings.     

Two distinct mutations in gyrA lead to ciprofloxacin and nalidixic acid resistance in Campylobacter coli and Campylobacter jejuni isolated from chickens and beef cattle

AIMS: The aim of this study was to identify point mutations in the gyrA quinolone resistance determining region (QRDR) of Campylobacter coli (n = 27) and Campylobacter jejuni (n = 26) that confer nalidixic acid (NAL) resistance without conferring resistance to ciprofloxacin (CIP). METHODS AND RESULTS: Point mutations in the QRDR of gyrA from C. coli and C. jejuni isolates were identified by direct sequencing. All isolates (n = 14) with minimum inhibitory concentrations (MICs) >or=4 microg ml(-1) for CIP and >or=32 microg ml(-1) for NAL possessed a missense mutation leading to substitution of Ile for Thr at codon 86. Three isolates with a missense mutation leading to a Thr86Ala substitution had MICs <4 mug ml(-1) for CIP and >or=32 microg ml(-1) for NAL. CONCLUSIONS: These data confirm previous findings that Thr86Ile mutations confer resistance to both CIP and NAL. However, resistance to NAL alone was conferred by a single Thr86Ala mutation. SIGNIFICANCE AND IMPACT OF THE STUDY: Resistance to NAL alone arises independently from CIP resistance. In addition, the role of other previously described point mutations in quinolone resistance is discussed.     

Fluoroquinolone resistance in Helicobacter pylori: role of mutations at position 87 and 91 of GyrA on the level of resistance and identification of a resistance conferring mutation in GyrB

Background and Aims: Fluoroquinolone‐containing regimens have been suggested as an alternate to standard triple therapy for the treatment of Helicobacter pylori infections. To determine the relationship between fluoroquinolone resistance and mutations of GyrA and GyrB in H. pylori, we exchanged the mutations at positions 87and 91 of GyrA among fluoroquinolone‐resistant clinical isolates. GyrB of a strain with no mutations in GyrA was also analyzed to identify mechanisms of resistance to norfloxacin. Materials & Methods: Natural transformation was performed using the amplified fragment of the gyrA and gyrB gene as donor DNA. The amino acid sequences of GyrA and GyrB were determined by DNA sequencing of the gyrA and gyrB genes. Results: Norfloxacin‐resistant strains which had mutations at position 87 and 91 became susceptible when the mutations were converted to the wild type. When the mutation from Asp to Asn at position 91 was exchanged to the mutation from Asn to Lys at position 87, the MIC to levofloxacin, gatifloxacin, and sitafloxacin increased. Norfloxacin‐resistant strain TS132 with no mutations in GyrA but had a mutation at position 463 in GyrB. Transformants obtained by natural transformation using gyrB DNA of TS132 had a mutation at position 463 of GyrB and revealed resistant to norfloxacin and levofloxacin. Conclusion: Mutation from Asn to Lys at position 87 of GyrA confers higher resistance to levofloxacin and gatifloxacin than does mutation from Asp to Asn at position 91. We propose that mutation at position 463 in GyrB as a novel mechanism of fluoroquinolone resistance in H. pylori.     

肺炎克雷伯菌gyrA基因与parC基因的突变研究

目的探讨肺炎克雷伯菌对环丙沙星和左氧氟沙星的药物敏感性,及对喹诺酮敏感和耐药菌株中gyrA与parC基因的突变情况。方法收集肺炎克雷伯菌临床分离株231株,采用K-B纸片法测定肺炎克雷伯菌对环丙沙星和左氧氟沙星的敏感性,随机选取对环丙沙星和左氧氟沙星均耐药菌株4株和均敏感的菌株3株,分别PCR扩增gyrA基因和parC基因的耐药决定区,扩增片段长度分别为625、319bp,PCR扩增产物经纯化后测序并做序列分析。结果肺炎克雷伯菌对环丙沙星和左氧氟沙星的耐药率分别为51.1%（118/231）和45.9%（106/231）;gyrA和parC基因经序列分析显示,耐药株均有gyrA基因的突变,其中1株出现第83、87和27位氨基酸的改变,2株出现第83位氨基酸的改变,1株出现第47位点的改变;环丙沙星敏感株中未出现gyrA基因的突变。4株耐药株均有parC基因的突变,引起相应氨基酸Ser80→Arg的改变,2株环丙沙星敏感株也发生了同样的改变。结论哈尔滨地区肺炎克雷伯菌对环丙沙星和左氧氟沙星的耐药性显著,在喹诺酮耐药株中有gyrA和parC基因的同时突变,在敏感株中也发现了parC基因的突变。     

Mutations in the gyrA and parC genes in ciprofloxacin-resistant clinical isolates of Acinetobacter baumannii in Korea

Mutation with Ser-83-->Leu in gyrA gene was associated with the principal mutation for ciprofloxacin resistance in clinical isolates of Acinetobacter baumannii. Double mutation, Ser-83-->Leu in gyrA gene and Ser-80-->Leu in parC gene, was the most frequently detected among ciprofloxacin-resistant isolates. A novel mutation with Ser-80-->Trp in parC gene, in addition to mutation in gyrA gene, was associated with a high-level ciprofloxacin resistance. These results suggested that the presence of an additional mutation in the parC gene contributed to a higher-level of ciprofloxacin resistance than a single mutation in the gyrA gene (geometric mean MICs of ciprofloxacin, 44.1 versus 16 microg/ml, P < 0.05).     

Molecular characterization of mutation associated with rifampicin and isoniazid resistance in Mycobacterium tuberculosis isolates

Nucleotide changes in catalase peroxidase (Kat G) gene and gene encoding the beta subunit of RNA polymerase (rpo B), responsible for isoniazid and rifampicin drug resistance were determined in the clinical isolates of Mycobacterium tuberculosis by PCR-RFLP, Line probe assay and DNA sequencing. PCR-RFLP test was performed by HapII cleavage of an amplified fragment of Kat G gene to detect the transversion 315AGC-->ACC(Ser-->Thr) which is associated with INH drug resistance. The Line probe assay kit was evaluated to detect the mutation in 81bp RMP resistance determining region of rpo B gene associated with RMP drug resistance. These results were validated by DNA sequencing and drug susceptibility test. Kat G S 315 T mutation was found in 74.19% strains of M. tuberculosis from Delhi. This mutation was not found in any of the susceptible strains tested. The line probe assay kit and DNA sequencing identified 18 isolates as RMP resistant with specific mutation, while one of the RMP resistant strain was identified as RMP susceptible, with a concordance of 94.73% with the phenotypic drug susceptibility result. Majority (8 of 19, 42.1%) of resistant isolates involved base changes at codon 531 of rpo B gene. Both PCR-RFLP and Line probe assay test can be used in many of the clinical microbiology laboratories for early detection of isoniazid and rifampicin drug resistance in clinical isolates of M. tuberculosis.     

Diversity of moxifloxacin resistance during a nosocomial outbreak of a predominantly ribotype ARU 027 Clostridium difficile diarrhea

To characterize the extent and diversity of moxifloxacin resistance among Clostridium difficile isolates recovered during a predominantly Anaerobe Reference Unit (ARU) ribotype 027-associated nosocomial outbreak of antibiotic associated diarrhea we measured the susceptibility of 34 field isolates and 6 laboratory strains of C. difficile to moxifloxacin. We ribotyped the isolates as well as assaying them by PCR for the metabolic gene, gdh, and the virulence genes, tcdA, tcdB, tcdC, cdtA and cdtB. All the laboratory isolates, including the historical ARU 027 isolate Cd196, were susceptible to moxifloxacin (≤2 μg/mL). 13 field isolates were susceptible to ≤2 μg/mL. Five were resistant to from 4 to 12 μg/mL (moderate resistance); 16 were resistant to ≥16 μg/mL (high resistance). We sequenced the quinolone resistance determining regions of gyrA (position 71-460) and gyrB (position 1059-1448) from two susceptible laboratory strains, all five isolates with moderate resistance and two highly resistant isolates. Two highly resistant isolates (Pitt 40, ribotype ARU 027 and Pitt 33, ribotype ARU 001) had the same C245T (Thr₈₂ΔIle) mutation. No other changes were seen. Amplification with primer pairs specific for the C245T mutant gyrA and for the wild type gene respectively confirmed all 16 highly resistant ARU 027 isolates, as well as the highly resistant isolates from other ribotypes, had the C245T mutation and that the mutation was absent from all other isolates. Among the five isolates with moderate resistance we found combinations of mutations within gyrA (T128A, Val₄₃ΔAsp and G349T, Ala₁₁₇ΔSer) and gyrB (G1276A, Arg₄₂₆ΔAsn). The G1396A (Glu₄₆₆ΔLys) mutation was not associated with increased resistance.     

CIP耐药的铜绿假单胞菌两种分子耐药机制关系的研究

目的探讨环丙沙星（CIP）耐药的铜绿假单胞菌临床分离株主动外排药物与gyrA、parC基因突变的关系。方法联合碳酰氰基-对-氯苯腙（CCCP）和CIP对CIP耐药的铜绿假单胞菌株进行主动外排阳性株和阴性株的筛选,并对这些菌株的gyrA,parC基因进行聚合酶链式反应-限制性片段长度多态性分析（PCR—RFLP）。结果57％（55／97）的CIP耐药菌株最小抑菌浓度（MIC）可被逆转,gyrA单基因突变率为65％,gyrA和pa-C双基因突变率为35％,未发现parC单基因突变的菌株。主动外排阳性组与阴性组gyrA、parC基因突变情况差异无显著性。结论在本地区铜绿假单胞菌对CIP的耐药机制中,主动外排系统表达上调与抗菌药物作用靶位的改变均占有重要的地位,两者可能是并存的两种相对独立的机制。     

Serotypes, antimicrobial susceptibility, and gyr A gene mutation of Campylobacter jejuni isolates from humans and chickens in Thailand

In Thailand, 51% (36/70) Campylobacter jejuni isolates from humans and 68% (47/69) isolates from poultry were classified into 10 Penner serotypes (serotype B, C, R, E, G, A, K, D, I, and L) and 9 serotypes (serotype A, C, I, K, B, E, S, D, and L), respectively. The rate of antimicrobial drug resistance to nalidixic acid, ciprofloxacin, ampicillin, tetracycline, and erythromycin shown by human isolates were 96%, 96%, 29%, 57%, and 14%, while that shown by poultry isolates were 77%, 77%, 22%, 26%, and 17%, respectively. All quinolone-resistant strains contained a mutation in the gyrA gene (T(86)-->I(86)), suggesting that the strains were already widespread in Thailand.     

Construction of a yrA plasmid for genetic characterization of fluoroquinolone-resistant Staphylococcus aureus

Abstract We have constructed a gyrA trans -complementation plasmid, pAT512, by cloning the wild-type gyrA gene of Staphylococcus aurues into expression vector pAT392. Introduction by electrotransformation of pAT512 into a high-level fluoroquinolone resistant mutant of S. aureus (ciprofloxacin MIC= 16 μ g ml⁻¹) having a gyrA mutation which results in a Ser-84 to Leu substitution, reduced the MICs of norfloxacin and ciprofloxacin for the host of four- and eight-fold, respectively.     

MAMA-PCR assay for the detection of point mutations associated with high-level erythromycin resistance in Campylobacter jejuni and Campylobacter coli strains

SYNOPSIS: Twenty Campylobacter jejuni and 16 Campylobacter coli strains isolated from humans and food/animals, including 17 isolates resistant to erythromycin, were analyzed. A combined mismatch amplification mutation assay-PCR technique was developed to detect the mutations A 2074 C and A 2075 G in the 23S rRNA gene associated with erythromycin resistance. All high-level erythromycin-resistant strains examined by DNA sequencing carried the transition mutation A 2075 G, whereas no isolate carried the A 2074 C mutation. No mutations were found among the susceptible and low-level erythromycin-resistant strains.     

Novel gyrase mutations and characterization of ciprofloxacin-resistant clinical strains of Enterococcus faecalis isolated in Poland

The activity of ciprofloxacin, sparfloxacin and moxifloxacin was determined for 205 Enterococcus faecalis isolates from patients of five hospitals (Warsaw, Poland; collected from 2000 to 2002). Ciprofloxacin resistant and intermediate isolates were numerous (53.7%). Among them, highly resistant (MIC > or = 16 mg/l) isolates predominated (98%). Isolates resistant to ciprofloxacin were also resistant to sparfloxacin and moxifloxacin. The parC and gyrA QRDRs (quinolone-resistance-determining region) of 11 isolates with ciprofloxacin MICs from 1 to 256 mg/l were analysed by DNA sequencing. In ParC one kind of amino acid substitution (of Ser-85 to Ile) in 9 E. faecalis strains with MICs from 16 to 256 mg/l was observed. In GyrA Ser-84 was changed to one of four different amino acids: Arg, Ile, Cys or Tyr, however no association between the amino acid type and MIC value was found. The last two substitutions have not been reported to date for E. faecalis. Moreover, our results may suggest that mutations within parC and gyrA are associated with development of a high-level of ciprofloxacin resistance.     

Frequency and Geographic Distribution of gyrA and gyrB Mutations Associated with Fluoroquinolone Resistance in Clinical Mycobacterium Tuberculosis Isolates: A Systematic Review

Background

The detection of mutations in the gyrA and gyrB genes in the Mycobacterium tuberculosis genome that have been demonstrated to confer phenotypic resistance to fluoroquinolones is the most promising technology for rapid diagnosis of fluoroquinolone resistance.

Methods

In order to characterize the diversity and frequency of gyrA and gyrB mutations and to describe the global distribution of these mutations, we conducted a systematic review, from May 1996 to April 2013, of all published studies evaluating Mycobacterium tuberculosis mutations associated with resistance to fluoroquinolones. The overall goal of the study was to determine the potential utility and reliability of these mutations as diagnostic markers to detect phenotypic fluoroquinolone resistance in Mycobacterium tuberculosis and to describe their geographic distribution.

Results

Forty-six studies, covering four continents and 18 countries, provided mutation data for 3,846 unique clinical isolates with phenotypic resistance profiles to fluoroquinolones. The gyrA mutations occurring most frequently in fluoroquinolone-resistant isolates, ranged from 21–32% for D94G and 13–20% for A90V, by drug. Eighty seven percent of all strains that were phenotypically resistant to moxifloxacin and 83% of ofloxacin resistant isolates contained mutations in gyrA. Additionally we found that 83% and 80% of moxifloxacin and ofloxacin resistant strains respectively, were observed to have mutations in the gyrA codons interrogated by the existing MTBDRsl line probe assay. In China and Russia, 83% and 84% of fluoroquinolone resistant strains respectively, were observed to have gyrA mutations in the gene regions covered by the MTBDRsl assay.

Conclusions

Molecular diagnostics, specifically the Genotype MTBDRsl assay, focusing on codons 88–94 should have moderate to high sensitivity in most countries. While we did observe geographic differences in the frequencies of single gyrA mutations across countries, molecular diagnostics based on detection of all gyrA mutations demonstrated to confer resistance should have broad and global utility.     

Implications of amino acid substitutions in GyrA at position 83 in terms of oxolinic acid resistance in field isolates of Burkholderia glumae, a causal agent of bacterial seedling rot and grain rot of rice

Oxolinic acid (OA), a quinolone, inhibits the activity of DNA gyrase composed of GyrA and GyrB and shows antibacterial activity against Burkholderia glumae. Since B. glumae causes bacterial seedling rot and grain rot of rice, both of which are devastating diseases, the emergence of OA-resistant bacteria has important implications on rice cultivation in Japan. Based on the MIC of OA, 35 B. glumae field isolates isolated from rice seedlings grown from OA-treated seeds in Japan were divided into sensitive isolates (OSs; 0.5 microg/ml), moderately resistant isolates (MRs; 50 microg/ml), and highly resistant isolates (HRs; > or =100 microg/ml). Recombination with gyrA of an OS, Pg-10, led MRs and HRs to become OA susceptible, suggesting that gyrA mutations are involved in the OA resistance of field isolates. The amino acid at position 83 in the GyrA of all OSs was Ser, but in all MRs and HRs it was Arg and Ile, respectively. Ser83Arg and Ser83Ile substitutions in the GyrA of an OS, Pg-10, resulted in moderate and high OA resistance, respectively. Moreover, Arg83Ser and Ile83Ser substitutions in the GyrA of MRs and HRs, respectively, resulted in susceptibility to OA. These results suggest that Ser83Arg and Ser83Ile substitutions in GyrA are commonly responsible for resistance to OA in B. glumae field isolates.     

Rapid detection of fluoroquinolone resistance by isothermal chimeric primer-initiated amplification of nucleic acids from clinical isolates of Neisseria gonorrhoeae

To ensure a complete response to fluoroquinolone therapy against Neisseria gonorrhoeae infections, rapid susceptibility determinations are required. We assessed a new approach, an isothermal chimeric primer-initiated amplification of nucleic acids (ICAN)/hybrid-chromatography method to detect rapidly fluoroquinolone resistance in N. gonorrhoeae. Comparison of the amplification results with fluoroquinolone minimum inhibitory concentrations (MICs), which were determined by an agar dilution method, showed that the new method accurately determined fluoroquinolone resistance in all ciprofloxacin- and/or gatifloxacin-resistant isolates, but agreed with results based on MICs in only 6 of 8 (75.0%) ciprofloxacin-susceptible and 7 of 12 (58.3%) gatifloxacin-susceptible isolates. Our results suggest that this method can rapidly and reliably detect point mutations in the gyrA gene as well as fluoroquinolone resistance in resistant isolates of N. gonorrhoeae.     

Genetic diversity and quinolone resistance in Campylobacter jejuni isolates from poultry in Senegal

We used the multilocus sequence typing (MLST) method to evaluate the genetic diversity of 46 Campylobacter jejuni isolates from chickens and to determine the link between quinolone resistance and sequence type (ST). There were a total of 16 ST genotypes, and the majority of them belonged to seven clonal complexes previously identified by using isolates from human disease. The ST-353 complex was the most common complex, whereas the ST-21, ST-42, ST-52, and ST-257 complexes were less well represented. The resistance phenotype varied for each ST, and the Thr-86-Ile substitution in the GyrA protein was the predominant mechanism of resistance to quinolone. Nine of the 14 isolates having the Thr-86-Ile substitution belonged to the ST-353 complex. MLST showed that the emergence of quinolone resistance is not related to the diffusion of a unique clone and that there is no link between ST genotype and quinolone resistance. Based on silent mutations, different variants of the gyrA gene were shown to exist for the same ST. These data provide useful information for understanding the epidemiology of C. jejuni in Senegal.     

Role of gyrA mutation and loss of OprF in the multiple antibiotic resistance phenotype of Pseudomonas aeruginosa G49

Abstract A clinical isolate of Pseudomonas aeruginosa G48, became resistant during fluoroquinolone treatment giving rise to the post-therapy isolate, G49. To determine whether mutation in gyrA gave rise to fluoroquinolone resistance, G49 was transformed with a plasmid encoding gyrA (pNJR3-2); this reduced the MIC of fluoroquinolones for G49 two-fold. DNA sequencing of gyrA of G49 demonstrated a mutation at Thr-83, substituting with isoleucine. The outer membrane of G49 was shown to lack OprF, suggesting that loss of this protein may be involved in the multiple antibiotic resistance phenotype; however, when G49 was transformed with a plasmid encoding oprF (pRWS), expression of oprF was shown to have no effect upon the phenotype.     

New insights into fluoroquinolone resistance in Mycobacterium tuberculosis: functional genetic analysis of gyrA and gyrB mutations

Fluoroquinolone antibiotics are among the most potent second-line drugs used for treatment of multidrug-resistant tuberculosis (MDR TB), and resistance to this class of antibiotics is one criterion for defining extensively drug resistant tuberculosis (XDR TB). Fluoroquinolone resistance in Mycobacterium tuberculosis has been associated with modification of the quinolone resistance determining region (QRDR) of gyrA. Recent studies suggest that amino acid substitutions in gyrB may also play a crucial role in resistance, but functional genetic studies of these mutations in M. tuberculosis are lacking. In this study, we examined twenty six mutations in gyrase genes gyrA (seven) and gyrB (nineteen) to determine the clinical relevance and role of these mutations in fluoroquinolone resistance. Transductants or clinical isolates harboring T80A, T80A+A90G, A90G, G247S and A384V gyrA mutations were susceptible to all fluoroquinolones tested. The A74S mutation conferred low-level resistance to moxifloxacin but susceptibility to ciprofloxacin, levofloxacin and ofloxacin, and the A74S+D94G double mutation conferred cross resistance to all the fluoroquinolones tested. Functional genetic analysis and structural modeling of gyrB suggest that M330I, V340L, R485C, D500A, D533A, A543T, A543V and T546M mutations are not sufficient to confer resistance as determined by agar proportion. Only three mutations, N538D, E540V and R485C+T539N, conferred resistance to all four fluoroquinolones in at least one genetic background. The D500H and D500N mutations conferred resistance only to levofloxacin and ofloxacin while N538K and E540D consistently conferred resistance to moxifloxacin only. Transductants and clinical isolates harboring T539N, T539P or N538T+T546M mutations exhibited low-level resistance to moxifloxacin only but not consistently. These findings indicate that certain mutations in gyrB confer fluoroquinolone resistance, but the level and pattern of resistance varies among the different mutations. The results from this study provide support for the inclusion of the QRDR of gyrB in molecular assays used to detect fluoroquinolone resistance in M. tuberculosis.     

Fluoroquinolone resistance and gyrA and parC mutations of Escherichia coli isolated from chicken

Escherichia coli is a common inhabitant of the intestinal tracts of animals and humans. The intestines of animals also represent an ideal environment for the selection and transfer of antimicrobial resistance genes. The aim of this study was to investigate the resistance of E. coli isolated from chicken fecal samples to fluoroquinolones and to analyze the characterization of mutations in its gyrA and parC gene related resistance. One hundred and twenty-eight E. coil isolates showed a high resistance to ciprofloxacin (CIP; 60.2%), enrofloxacin (ENO; 73.4%) and norfloxacin (NOR; 60.2%). Missense mutation in gyrA was only found in the amino acid codons of Ser-83 or Asp-87. A high percentage of isolates (60.2%) showed mutations at both amino acid codons. Missense mutation in parC was found in the amino acid codon of Ser-80 or Glu-84, and seven isolates showed mutations at both amino acid codons. Isolates with a single mutation in gyrA showed minimal inhibitory concentrations (MIC) for CIP (相似文献   

Role of an acrR mutation in multidrug resistance of in vitro-selected fluoroquinolone-resistant mutants of Salmonella enterica serovar Typhimurium

Quinolone resistance in Salmonella spp. is usually attributed to both active efflux and mutations leading to modification of the target enzymes DNA gyrase and topoisomerase IV. Here, we investigated the presence of mutations in the efflux regulatory genes of fluoroquinolone- and multidrug-resistant mutants of Salmonella enterica serovar Typhimurium (S. Typhimurium) selected in vitro with enrofloxacin that both carried a mutation in the target gene gyrA and overproduced the AcrAB efflux pump. No mutations were detected in the global regulatory loci marRAB and soxRS for the four strains studied. A mutation in acrR, the local repressor of acrAB, was found for two ciprofloxacin-resistant selected-mutants, leading to duplication of amino acids Ile75 and Glu76. Complementation experiments with wild-type acrR showed that the mutation identified in acrR partially contributed to the increase in resistance levels to several unrelated antibiotics. The acrR mutation also contributed to acrAB overexpression as shown by RT-PCR. Thus, this study underlines the role of an acrR mutation, in addition to the mutation in gyrA, in the fluoroquinolone and multidrug resistance phenotype of S. Typhimurium mutants, through overexpression of acrAB.