Gyrase mutations in laboratory-selected, fluoroquinolone-resistant mutants of Mycobacterium tuberculosis H37Ra

To characterize mechanisms of resistance to fluoroquinolones by Mycobacterium tuberculosis, mutants of strain H37Ra were selected in vitro with ofloxacin. Their quinolone resistance-determining regions for gyrA and gyrB were amplified and sequenced to identify mutations in gyrase A or B. Three types of mutants were obtained: (i) one mutant (TKp1) had no mutations in gyrA or gyrB; (ii) mutants that had single missense mutations in gyrA, and (iii) mutants that had two missense mutations resulting in either two altered gyrase A residues or an altered residue in both gyrases A and B. The TKp1 mutant had slightly reduced levels of uptake of [14C]norfloxacin, which was associated with two- to fourfold increases in the MICs of ofloxacin, ciprofloxacin, and sparfloxacin. Gyrase mutations caused a much greater increase in the MICs of fluoroquinolones. For mutants with single gyrA mutations, the increases in the MICs were 4- to 16-fold, and for mutants with double gyrase mutations, the MICs were increased 32-fold or more compared with those for the parent. A gyrA mutation in TKp1 secondary mutants was associated with 32- to 128-fold increases in the MICs of ofloxacin and ciprofloxacin compared with the MICs for H37Ra and an eight-fold increase in the MIC of sparfloxacin. Sparfloxacin was the most active fluoroquinolone tested. No sparfloxacin-resistant single-step mutants were selected at concentrations of > 2.5 micrograms/ml, and high-level resistance (i.e., MIC, > and = 5 micrograms/ml) was associated with two gyrase mutations. Mutations in gyrB and possibly altered levels of intracellular accumulation of drug are two additional mechanisms that may be used by M. tuberculosis in the development of fluoroquinolone resistance. Because sparfloxacin is more active in vitro and selection of resistance appears to be less likely to occur, it may have important advantage over ofloxacin or ciprofloxacin for the treatment of tuberculosis.     

Sparfloxacin resistance in clinical isolates of Streptococcus pneumoniae: involvement of multiple mutations in gyrA and parC genes

Antimicrobial susceptibility testing revealed among 150 clinical isolates of Streptococcus pneumoniae 4 pneumococcal isolates with resistance to fluoroquinolones (MIC of ciprofloxacin, >/=32 microgram/ml; MIC of sparfloxacin, >/=16 microgram/ml). Gene amplification and sequencing analysis of gyrA and parC revealed nucleotide changes leading to amino acid substitutions in both GyrA and ParC of all four fluoroquinolone-resistant isolates. In the case of strains 182 and 674 for which sparfloxacin MICs were 16 and 64 microgram/ml, respectively, nucleotide changes were detected at codon 81 in gyrA and codon 79 in parC; these changes led to an Ser-->Phe substitution in GyrA and an Ser-->Phe substitution in ParC. Strains 354 and 252, for which sparfloxacin MICs were 128 microgram/ml, revealed multiple mutations in both gyrA and parC. These strains exhibited nucleotide changes at codon 85 leading to a Glu-->Lys substitution in GyrA, in addition to Ser-79-->Tyr and Lys-137-->Asn substitutions in ParC. Moreover, strain 252 showed additional nucleotide changes at codon 93, which led to a Trp-->Arg substitution in GyrA. These results suggest that sparfloxacin resistance could be due to the multiple mutations in GyrA and ParC. However, it is possible that other yet unidentified mutations may also be involved in the high-level resistance to fluoroquinolones in S. pneumoniae.     

Contribution of mutations in gyrA and parC genes to fluoroquinolone resistance of mutants of Streptococcus pneumoniae obtained in vivo and in vitro

We have analyzed by gene amplification and sequencing mutations in the quinolone resistance-determining regions of the gyrA, gyrB, and parC genes of fluoroquinolone-resistant Streptococcus pneumoniae mutants obtained during therapy or in vitro. Mutations leading to substitutions in ParC were detected in the two mutants obtained in vivo, BM4203-R (substitution of a histidine for an aspartate at position 84 [Asp-84-->His]; Staphylococcus aureus coordinates) and BM4204-R (Ser-80-->Phe), and in two mutants obtained in vitro (Ser-80-->Tyr). An additional mutant obtained in vitro, BM4205-R3, displayed a higher level of fluoroquinolone resistance and had a mutation in gyrA leading to a Ser-84-->Phe change. We could not detect any mutation in the three remaining mutants obtained in vitro. Total DNA from BM4203-R, BM4204-R, and BM4205-R3 was used to transform S. pneumoniae CP1000 by selection on fluoroquinolones. For the parC mutants, transformants with phenotypes indistinguishable from those of the donors were obtained at frequencies (5 x 10(-3) to 8 x 10(-3)) compatible with monogenic transformation. By contrast, transformants were obtained at a low frequency (4 x 10(-5)), compatible with the transformation of two independent genes, for the gyrA mutant. Resistant transformants of CP1000 were also obtained with an amplified fragment of parC from BM4203-R and BM4204-R but not with a gyrA fragment from BM4205-R3. All transformants had mutations identical to those in the donors. These data strongly suggest that ParC is the primary target for fluoroquinolones in S. pneumoniae and that BM4205-R3 is resistant to higher levels of the drugs following the acquisition of two mutations, including one in gyrA.     

Interferons alpha/beta inhibit IL-7-induced proliferation of CD4- CD8- CD3- CD44+ CD25+ thymocytes, but do not inhibit that of CD4- CD8- CD3- CD44- CD25- thymocytes

We have determined partial sequences of the gyrA and parC genes of Enterobacter cloacae type strain including the regions analogous to the quinolone resistance-determining region of the Escherichia coli gyrA gene. The deduced 65- and 49-amino acid sequences of the determined regions of the E. cloacae gyrA and parC genes were identical to the corresponding regions of the E. coli GyrA and ParC proteins, respectively. We examined 40 clinical strains of E. cloacae isolated from patients with urinary tract infection for susceptibilities to nalidixic acid and ciprofloxacin. Based on the nalidixic acid and ciprofloxacin MICs, these isolates were divided into 19 quinolone-susceptible strains (MICs of nalidixic acid, 3.13-25 mg/L; MICs of ciprofloxacin, < or = 0.025 mg/L) and 21 quinolone-resistant strains (MICs of nalidixic acid, 400 to > 800 mg/L; MICs of ciprofloxacin, 0.39-100 mg/L). We analysed five quinolone-susceptible and 21 quinolone-resistant strains for alterations in GyrA and ParC. The five quinolone-susceptible strains had amino acid sequences in GyrA and ParC identical to those of type strain. Of the 21 quinolone-resistant isolates, three (MICs of nalidixic acid, 400 to > 800 mg/L; MICs of ciprofloxacin, 0.39-3.13 mg/L) had a single amino acid change at the position equivalent to Ser-83 in the E. coli GyrA protein and no alterations in ParC; one (MIC of nalidixic acid, > 800 mg/L; MIC of ciprofloxacin, 3.13 mg/L) had a single amino acid change at Ser-83 in GyrA and a single amino acid change at the position equivalent to Glu-84 in the E. coli ParC protein; two (MIC of nalidixic acid, > 800 mg/L; MIC of ciprofloxacin, 25 mg/L) had double amino acid changes at Ser-83 and Asp-87 in GyrA and no alterations in ParC; and 15 (MICs of nalidixic acid, > 800 mg/L; MICs of ciprofloxacin, 25-100 mg/L) had double amino acid changes at Ser-83 and Asp-87 in GyrA and a single amino acid change at Ser-80 or Glu-84 in ParC. This study suggests, that in clinical isolates of E. cloacae, DNA gyrase is a primary target of quinolones, that only a single amino acid change at Ser-83 in GyrA is sufficient to generate high-level resistance to nalidixic acid and to decrease susceptibility to ciprofloxacin, and that the accumulation of amino acid changes in GyrA and the simultaneous presence of the ParC alterations play a central role in developing high-level resistance to ciprofloxacin.     

Isolation and characterization of a dibenzofuran-degrading yeast: identification of oxidation and ring cleavage products

PURPOSE: To investigate the development of fluoroquinolone resistance among Neisseria gonorrhoeae isolates in Japan and the frequency and patterns of mutations involving the GyrA and ParC proteins, which confer quinolone resistance to the bacteria, in isolates. MATERIALS AND METHODS: Antimicrobial susceptibilities of 145 gonococcal isolates, including 79 isolated from February 1992 through February 1993 and 66 isolated from February 1995 through February 1996, to six fluoroquinolones and several other antibiotics were compared with those of 27 isolates obtained from 1981 through 1984. To identify mutations in gyrA and parC genes of the isolates, the quinolone resistance-determining regions of the gyrA and parC genes were PCR-amplified and the PCR products were directly sequenced. RESULTS: The minimum inhibitory concentration for 90% of strains (MIC90) values of norfloxacin for the isolates from 1992 to 93 (4 microg./ml.) and 1995 to 96 (8 microg./ml.) were 16- and 32-fold, respectively, higher than those for isolates from 1981 to 84 (0.25 microg./ml.). The MIC90 values of ciprofloxacin for isolates from 1992 to 93 (0.5 microg./ml.) and 1995 to 96 (1 microg./ml.) showed increase of 8- and 16-fold, respectively, in comparison with those from 1981 to 84 (0.063 microg./ml.). The isolates from 1992 to 93 and 1995 to 96 were also less susceptible to newer fluoroquinolones including levofloxacin, sparfloxacin, DU-6859a and AM-1155, as compared with those from 1981 to 84. In 46 (67.6%) and 16 (23.5%) of the 68 gonococcal strains sequenced, GyrA and ParC mutations were identified, respectively. No ParC substitutions were identified in any isolates without co-existence of the GyrA mutation. A Ser-91 to Phe mutation, which was detected in 30 (65.2%) of the 46 isolates with GyrA mutations, was the most common GyrA mutation. Mutants with the single Ser-91 to Phe substitution in GyrA were 12-fold and at least 13-fold, respectively, less susceptible to norfloxacin and ciprofloxacin than the wild type. CONCLUSIONS: The results obtained in this study suggest that a high prevalence of gonococcal isolates with the Ser-91 to Phe mutation in GyrA has reduced the susceptibility of this organism to fluoroquinolones in Japan.     

Role of mutations in DNA gyrase genes in ciprofloxacin resistance of Pseudomonas aeruginosa susceptible or resistant to imipenem

In Pseudomonas aeruginosa, resistance to imipenem is mainly related to a lack of protein OprD and resistance to fluoroquinolones is mainly related to alterations in DNA gyrase. However, strains cross resistant to fluoroquinolones and imipenem have been selected in vitro and in vivo with fluoroquinolones. We investigated the mechanisms of resistance to fluoroquinolones in 30 clinical strains of P. aeruginosa resistant to ciprofloxacin (mean MIC, >8 micrograms/ml), 20 of which were also resistant to imipenem (mean MIC, >16 micrograms/ml). By immunoblotting, OprD levels were markedly decreased in all of the imipenem-resistant strains. Plasmids carrying the wild-type gyrA gene (pPAW207) or gyrB gene (pPBW801) of Escherichia coli were introduced into each strain by transformation. MICs of imipenem did not change after transformation, whereas those of ciprofloxacin and sparfloxacin dramatically decreased (25- to 70-fold) for all of the strains. For 28 of them (8 susceptible and 20 resistant to imipenem), complementation was obtained with pPAW207 but not with pPBW801. After complementation, the geometric mean MICs of ciprofloxacin and sparfloxacin (MICs of 0.3 microgram/ml and 0.5 microgram/ml, respectively) were as low as those for wild-type strains. Complementation was obtained only with pPBW801 for one strain and with pPAW207 and pPBW801 for one strain highly resistant to fluoroquinolones. These results demonstrate that in clinical practice, gyrA mutations are the major mechanism of resistance to fluoroquinolones even in the strains of P. aeruginosa resistant to imipenem and lacking OprD, concomitant resistance to these drugs being the result of the addition of at least two independent mechanisms.     

Efflux pump-mediated quinolone resistance in Staphylococcus aureus strains wild type for gyrA, gyrB, grlA, and norA

Fluoroquinolone efflux was studied in 47 Staphylococcus aureus clinical strains with MICs of ciprofloxacin (CFX) of < or = 2 micrograms/ml. Forty-three strains were wild type for gyrA, gyrB, and grlA quinolone resistance-determining regions and for norA and its promoter region. Forty of these strains (MICs of CFX, 0.1 to 0.2 microgram/ml) did not show efflux of fluoroquinolones. Three strains (MICs of CFX, 1 to 2 micrograms/ml) showed efflux. These results suggest that efflux can appear in S. aureus clinical strains in the absence of mutations in norA and its promoter.     

Detection of mutations in parC in quinolone-resistant clinical isolates of Escherichia coli

The gene parC encodes the A subunit of topoisomerase IV of Escherichia coli. Mutations in the parC region analogous to those in the quinolone resistance-determining region of gyrA were investigated in 27 clinical isolates of E. coli for which ciprofloxacin MICs were 0.0007 to 128 micrograms/ml. Of 15 isolates for which ciprofloxacin MICs were > or = 1 microgram/ml, 8 showed a change in the serine residue at position 80 (Ser-80), 4 showed a change in Glu-84, and 3 showed changes in both amino acids. No mutations were detected in 12 clinical isolates for which ciprofloxacin MICs were < or = 0.25 micrograms/ml. These findings suggest that ParC from E. coli may be another target for quinolones and that mutations at residues Ser-80 and Glu-84 may contribute to decreased fluoroquinolone susceptibility.     

Antibacterial activity of gatifloxacin (AM-1155, CG5501, BMS-206584), a newly developed fluoroquinolone, against sequentially acquired quinolone-resistant mutants and the norA transformant of Staphylococcus aureus

Alternate mutations in the grlA and gyrA genes were observed through the first- to fourth-step mutants which were obtained from four Staphylococcus aureus strains by sequential selection with several fluoroquinolones. The increases in the MICs of gatifloxacin accompanying those mutational steps suggest that primary targets of gatifloxacin in the wild type and the first-, second-, and third-step mutants are wild-type topoisomerase IV (topo IV), wild-type DNA gyrase, singly mutated topo IV, and singly mutated DNA gyrase, respectively. Gatifloxacin had activity equal to that of tosufloxacin and activity more potent than those of norfloxacin, ofloxacin, ciprofloxacin, and sparfloxacin against the second-step mutants (grlA gyrA; gatifloxacin MIC range, 1.56 to 3.13 microg/ml) and had the most potent activity against the third-step mutants (grlA gyrA grlA; gatifloxacin MIC range, 1.56 to 6.25 microg/ml), suggesting that gatifloxacin possesses the most potent inhibitory activity against singly mutated topo IV and singly mutated DNA gyrase among the quinolones tested. Moreover, gatifloxacin selected resistant mutants from wild-type and the second-step mutants at a low frequency. Gatifloxacin possessed potent activity (MIC, 0.39 microg/ml) against the NorA-overproducing strain S. aureus NY12, the norA transformant, which was slightly lower than that against the parent strain SA113. The increases in the MICs of the quinolones tested against NY12 were negatively correlated with the hydrophobicity of the quinolones (correlation coefficient, -0.93; P < 0.01). Therefore, this slight decrease in the activity of gatifloxacin is attributable to its high hydrophobicity. Those properties of gatifloxacin likely explain its good activity against quinolone-resistant clinical isolates of S. aureus harboring the grlA, gyrA, and/or norA mutations.     

Anomalous patterns of callosal connections develop in visual cortex of monocularly enucleated hamsters

Six multiply resistant isolates of Salmonella typhimurium var. copenhagen with high-level resistance to fluoroquinolones (e.g., MIC of ciprofloxacin: 32 micrograms/ml) were isolated from human patients (n = 3) and from cattle (n = 3). The isolates were examined by complementation tests using a set of broad-host-range plasmids, which carry either the gyrA+ or the gyrB+ genes or a combination of both from Escherichia coli K-12. The results indicated a combination of gyrA and gyrB mutations in all isolates. Subsequent direct sequencing of PCR-generated internal DNA fragments of gyrA revealed an identical double mutation in all six isolates (Ser-83-->Ala and Asp-87-->Asn). In addition, the results of phenotypic (i.e., phagetype, biotype, serotype) and genotypic characterization [i.e., ribotyping and polymerase chain reaction fingerprinting (PCR-fingerprinting)] were identical for all six isolates and were distinguishable from a quinolone-susceptible strain of the same serovar and an unrelated isolate of S. typhimurium. These data indicate the clonal identity of the fluoroquinolone-resistant strains of S. typhimurium isolated from men and cattle in Germany.     

Skin smears in leprosy: is reduction in number of sites justified?

BACKGROUND: Infections caused by Streptococcus pneumoniae continue to be a significant cause of mortality and morbidity in humans. Diseases caused by multi-resistant pneumococci are increasing rapidly worldwide. The fluoroquinolones have been widely used clinically to treat infectious diseases. The results of a study here on the five fluoroquinolones susceptibilities of S. pneumoniae are reported from the Taichung Veterans General Hospital. METHODS: Minimum inhibitory concentrations (MICs) of five quinolones (enoxacin, norfloxacin, ofloxacin, levofloxacin and ciprofloxacin) were determined for 106 strains of S. pneumoniae. All MICs were determined by the agar dilution method utilizing Mueller-Hinton agar supplemented with 5% sheep blood. RESULTS: MIC90 of levofloxacin was 1 microgram/ ml, and was unaffected by penicillin-susceptibility. MIC90 of ofloxacin and that of ciprofloxacin were 2 and 4 micrograms/ml, respectively, with 90.6% sensitive to ofloxacin. MIC90 of enoxacin and that of norfloxacin were higher than other compounds. CONCLUSIONS: The in vitro activity of levofloxacin is twice that of ofloxacin, 4-fold of ciprofloxacin, 16-fold of norfloxacin, and 64-fold of enoxacin. MICs of these five quinolones were unaffected by penicillin-susceptibility. The antibacterial activity of levofloxacin was better than that of ofloxacin and ciprofloxacin, norfloxacin, or enoxacin against S. pneumoniae.     

Association of mutations in grlA and gyrA topoisomerase genes with resistance to ciprofloxacin in epidemic and sporadic isolates of methicillin-resistant Staphylococcus aureus

The types of topoisomerase alterations in genomically diverse epidemic and sporadic strains of methicillin- and fluoroquinolone-resistant Staphylococcus aureus isolated from European hospitals between 1984 and 1994 were characterized. Convergent dual mutations in gyrA (codon 83, 84, or 88) and grlA (codon 79 and/or 80) were found in all strains exhibiting high-level resistance to ciprofloxacin (MIC, 16 to > or = 128 microg/ml). In some epidemic strains, the resistant phenotype and genotype appeared in the 1990s and persisted thereafter.     

Tumour size measured by transrectal ultrasonography in the staging of prostate cancer before radical prostatectomy

Fluoroquinolone-resistant mutants of Mycoplasma hominis were selected in vitro from the PG21 susceptible reference strain either by multistep selection on increasing concentrations of various fluoroquinolones or by one-step selection on agar medium with ofloxacin. The quinolone resistance-determining regions (QRDR) of the structural genes encoding the A and b subunits of DNA gyrase were amplified by PCR, and the nucleotide sequences of eight multistep-selected resistant strains were compared to those of susceptible strain PG21. Four high-level resistant mutants that were selected on norfloxacin or ofloxacin contained a C-to-T transition in the gyrA QRDR, leading to substitution of Ser-83 by Leu in the GyrA protein. Analysis of the sequence of the gyrB QRDR of the eight multistep-selected mutants did not reveal any difference compared to that of the gyrB QRDR of the reference strain M. hominis PG21. Similar analyses of eight one-step-selected mutants did not reveal any base change in the gyrA and gyrB QRDRs. These results suggest that in M. hominis, like in other bacterial species, a gyrA mutation at Ser-83 is associated with fluoroquinolone resistance.     

Expression patterns of proliferating cell nuclear antigen in the small intestine of mice infected with Metagonimus yokogawai and Metagonimus Miyata type

The in vitro antimicrobial activities of AM-1155, a new fluoroquinolone, tosufloxacin and fleroxacin were tested against 55 clinical isolates of Neisseria gonorrhoeae using the agar dilution method. In our previous study, all the strains had been examined for mutations in the region corresponding to the quinolone-resistance determining region of the Escherichia coli gyrA gene and the analogous region of the parC gene, and tested for susceptibility to ciprofloxacin. In this study, the 55 isolates of N. gonorrhoeae were assigned to one of three categories based on the presence or absence of alterations in GyrA and ParC. In each category, the antimicrobial activity of AM-1155 against the isolates was compared with those of tosufloxacin and fleroxacin. The MICs of AM-1155 for 11 highly fluoroquinolone-resistant isolates with alterations in both GyrA and ParC ranged from 0.06 to 1.0 microgram/ml. The MICs inhibiting 50% (MIC50) and 90% (MIC90) of these isolates were 0.125 and 1.0 microgram/ml, respectively. The MICs of AM-1155 for 20 moderately fluoroquinolone-resistant isolates with alterations only in GyrA ranged from 0.03 to 0.25 microgram/ml (MIC50, 0.06 microgram/ml; MIC90m, 0.125 microgram/ml). The MICs of AM-1155 for 24 of the quinolone-susceptible isolates without alterations in either GyrA or ParC ranged from 0.004 to 0.03 microgram/ml (MIC50, 0.008 microgram/ml. MIC90, 0.015 microgram/ml). There were significant differences between the MIC distribution of AM-1155 and each corresponding MIC distribution of tosufloxacin and fleroxacin in these three categories to which the 55 isolates were assigned (p < 0.05). Based on the MIC90S of the tested fluoroquinolones, AM-1155 was two- and eightfold more active against the highly fluoroquinolone-resistant isolates than tosufloxacin and fleroxacin, respectively. Against the moderately fluoroquinolone-resistant isolates, AM-1155 was four- and sixteenfold more active than tosufloxacin and fleroxacin, respectively. Against the quinolone-susceptible strains, AM-1155 was also two- to fourfold more active than the other fluoroquinolones. Overall, AM-1155 exhibited more potent in vitro activity against both quinolone-resistant and quinolone-susceptible isolates of N. gonorrhoeae than tosufloxacin and fleroxacin. In ciprofloxacin treatment failures of gonorrhea at single doses of 500 mg. MICs for the causative organisms have ranged from 1.0 to 16.0 micrograms/ml. The MICs of AM-1155 for the isolates harboring quinolone resistance-associated genetic alterations, including strains exhibiting ciprofloxacin MICs of 2.0 and 8.0 micrograms/ml, still ranged from 0.03 to 1.0 microgram/mL A single-dose study in humans has demonstrated higher peak serum concentrations and longer half-lives of AM-1155, resulting in the AUC0-00 values of AM-1155, which are threefold greater than those of ciprofloxacin at the single doses of 400 and 600 mg. Because of its potent in vitro antimicrobial activity and advantageous pharmacokinetic behavior, AM-1155 may be a clinically useful agent for treating gonorrhea including that caused by quinolone-resistant strains.     

Emergence of in vitro resistance to fluoroquinolones in Neisseria gonorrhoeae isolated in Japan

To investigate emerging fluoroquinolone resistance in Neisseria gonorrhoeae isolated in Japan, we compared the in vitro antimicrobial susceptibilities of 79 gonococcal isolates from 1992 through 1993 to 14 fluoroquinolones and 14 other antibiotics with those of 27 isolates from between 1981 and 1984. The MICs at which 90% of the isolates were inhibited by nine fluroquinolones, including norfloxacin, enoxacin, ofloxacin, ciprofloxacin, tosufloxacin, lomefloxacin, fleroxacin, levofloxacin, and sparfloxacin, for isolates from 1992 to 1993 were 8- or 16-fold higher than those for isolates from 1981 to 1984. Furthermore, the MICs at which 90% of the isolates were inhibited by five fluroquinolones, including OPC-17116, T-3761, DU-6859a, AM-1155, and Q-35, that have recently been synthesized but have not yet been introduced for clinical use in Japan for isolates from 1992 to 1993 were also 2- to 16-fold higher than those for isolates from 1981 to 1984. The gonococcal isolates from 1992 to 1993 showed no significant decreases in susceptibility to beta-lactams, tetracyclines, macrolides, and spectinomycin, compared with those for isolates from 1981 to 1984. Our data indicate that the incidence of gonococcal strains with decreased susceptibilities to fluoroquinolones is increasing in Japan.     

In vitro susceptibility of Coxiella burnetii to trovafloxacin in comparison with susceptibilities to pefloxacin, ciprofloxacin, ofloxacin, doxycycline, and clarithromycin

The antibiotic susceptibilities of eight Greek isolates of Coxiella burnetii to trovafloxacin were determined by the shell vial assay. MICs of trovafloxacin and ofloxacin ranged from 1 to 2 microg/ml, those of pefloxacin ranged from 1 to 4 microg/ml, those of ciprofloxacin ranged from 4 to 8 microg/ml, those of doxycycline ranged from 1 to 2 microg/ml, and those of clarithromycin ranged from 2 to 4 microg/ml. Trovafloxacin exhibited no activity against C. burnetii at 4 microg/ml.     

Cervical stenosis and dystonic cerebral palsy

We have tested the in vitro activities of eight fluoroquinolones against 160 Brucella melitensis strains. The most active was sitafloxacin (MIC at which 90% of the isolates are inhibited [MIC90], 0.12 microg/ml). In decreasing order, the activities (MIC90s) of the rest of the tested fluoroquinolones were as follows: levofloxacin, 0.5 microg/ml; ciprofloxacin, trovafloxacin, and moxifloxacin, 1 microg/ml; and ofloxacin, grepafloxacin, and gatifloxacin, 2 microg/ml.     

Relationship between streptomycin susceptibility and rpsL mutations of Mycobacterium tuberculosis strains

The relationship between streptomycin (SM) susceptibility and rpsL mutations of Mycobacterium tuberculosis strains was studied. Of 18 clinically isolated SM-resistant M.tuberculosis strains, mutation was suspected in 9 strains (50%) with SM MICs of > or = 256 micrograms/ml by PCR-single strand conformation polymorphism targeting rpsL gene. On the other hand, using PCR-direct sequence method, amino acid substitution caused by single nucleotide point mutation in rpsL gene was demonstrated in 11 out of 18 strains (61%). The same amino acid substitution at codon 43 (Lys-->Arg) was observed in all 11 strains with SM MICs of > or = 256 micrograms/ml. In addition, PCR products obtained from these 11 strains could not be cut by a restriction enzyme, Mbo II, while H37Rv strain and the other 32 strains with SM MICs of < 256 micrograms/ml were cut into 2 fragments. In conclusion, our results suggest that highly SM-resistant M.tuberculosis strains with MICs of > or = 256 micrograms/ml could be rapidly and easily detected by the restriction enzymatic method.     

Purification and in vitro reconstitution of the essential protein components of an aromatic polyketide synthase

The in vitro activities of seven quinolones and the sequences of the quinolone resistance-determining regions (QRDR) in the A and B subunits of DNA gyrase were determined for 14 mycobacterial species. On the basis of quinolone activity, quinolones were arranged from that with the greatest to that with the least activity as follows: sparfloxacin, levofloxacin, ciprofloxacin, ofloxacin, pefloxacin, flumequine, and nalidixic acid. Based on MICs, the species could be organized into three groups: resistant (Mycobacterium avium, M. intracellulare, M. marinum, M. chelonae, M. abscessus [ofloxacin MICs, >/=8 microg/ml]), moderately susceptible (M. tuberculosis, M. bovis BCG, M. kansasii, M. leprae, M. fortuitum third biovariant, M. smegmatis [ofloxacin MICs, 0.5 to 1 microg/ml]), and susceptible (M. fortuitum, M. peregrinum, M. aurum [ofloxacin MICs, 相似文献   

Characterization of grlA, grlB, gyrA, and gyrB mutations in 116 unrelated isolates of Staphylococcus aureus and effects of mutations on ciprofloxacin MIC

One hundred sixteen unrelated clinical isolates of Staphylococcus aureus (70 ciprofloxacin resistant and 46 ciprofloxacin susceptible) from eight countries were studied for the presence of mutations in the grlA, grlB, gyrA, and gyrB gene loci. Two mutations within grlA (located at codons 80 and 84) and two mutations within gyrA (located at codons 84 and 88) were clearly associated with ciprofloxacin resistance, although other mutations detected within the four genes studied may also contribute to decreased susceptibility.