Seigo Suzue

Mutations producing resistance to norfloxacin in Pseudomonas aeruginosa.

Two genetically distinct classes of norfloxacin-resistant Pseudomonas aeruginosa PAO4009 mutants were isolated spontaneously. Two norfloxacin resistance genes, nfxA and nfxB, were mapped hex-9001 and leu-9005 and between pro-9031 and ilv-9023, respectively, on the P. aeruginosa PAO chromosome. The nfxA gene was shown to be an allele of nalA by transductional analysis with bacteriophage F116L. The nfxB mutant showed a 16-fold increase in resistance to norfloxacin and a slight increase in resistance to nalidixic acid. The nfxB mutant was unique in that it showed hypersusceptibility to beta-lactam and aminoglycoside antibiotics. This mutant had about a threefold-lower rate of norfloxacin uptake than that of the wild-type strain or nfxA mutant. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of outer membrane proteins demonstrated the appearance of a 54,000-dalton protein in the nfxB mutant. These findings suggested that the norfloxacin resistance mechanism in the nfxB mutant might be an alteration in outer membrane permeability to norfloxacin. Images

In Vitro Antibacterial Activity of AM-715, a New Nalidixic Acid Analog

AM-715 [1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(1-piperazinyl)-3-quinolinecarboxylic acid] is a new nalidixic acid analog. AM-715 has a broad spectrum of antibacterial activity against gram-positive and gram-negative bacteria. The antibacterial activity of AM-715 was greater than those of pipemidic acid and nalidixic acid. AM-715 had higher antibacterial activity against Pseudomonas aeruginosa than did gentamicin. Most nalidixic acid-resistant bacteria were susceptible to AM-715, and cross-resistance was not observed between AM-715 and various antibiotics. The minimal concentration of AM-715 required to inhibit the growth of 75% of the total number of clinical isolates was as follows: Escherichia coli, 0.04 μg/ml; Klebsiella pneumoniae, 0.1 μg/ml; Serratia marcescens, 0.88 μg/ml; Enterobacter spp., 0.076 μg/ml; Staphylococcus aureus, 1.10 μg/ml; P. aeruginosa, 0.38 μg/ml; and nalidixic acid-resistant strains of gram-negative bacteria, 0.62 μg/ml. AM-715 at minimal inhibitory concentrations or at slightly higher concentrations had bactericidal activity against various species of bacteria. The effect of inoculum sizes on minimal inhibitory concentrations and minimal bactericidal concentrations of AM-715 against gram-negative bacteria was smaller than on those of pipemidic acid and nalidixic acid. The dose-response curve of AM-715 indicated a steep gradient, and the 50% inhibited doses of AM-715 were 0.014 μg/ml against E. coli ML4707 and 0.21 μg/ml against P. aeruginosa NC-5.

Isolation and characterization of norfloxacin-resistant mutants of Escherichia coli K-12.

We isolated spontaneous mutants from Escherichia coli K-12 with low-level resistance to norfloxacin. These mutants were classified into the following three types on the basis of their properties: (i) NorA appeared to result for mutation in the gyrA locus for the A subunit of DNA gyrase; (ii) NorB showed low-level resistance to quinolones and other antimicrobial agents (e.g., cefoxitin, chloramphenicol, and tetracycline), and the norB gene was considered to map at about 34 min on the E. coli K-12 chromosome; (iii) NorC was less susceptible to norfloxacin and ciprofloxacin but was hypersusceptible to hydrophobic quinolones such as nalidixic acid and rosoxacin, hydrophobic antibiotics, dyes, and detergents. Susceptibility to bacteriophages and the hydrophobicity of the NorC cell surface also differed from that of the parent strain. The norC gene was located near the lac locus at 8 min on the E. coli K-12 chromosome. Both NorB and NorC mutants had a lower rate of norfloxacin uptake, and it was found that the NorB mutant was altered in OmpF porin and that the NorC mutant was altered in both OmpF porin and apparently in the lipopolysaccharide structure of the outer membrane.

In vitro and in vivo antibacterial activity of AM-833, a new quinolone derivative.

AM-833 showed potent activity against members of the family Enterobacteriaceae, Neisseria spp., and Haemophilus influenzae and good activity against staphylococci, Pseudomonas aeruginosa, and Branhamella catarrhalis. Against these bacteria, its activity was roughly comparable to that of norfloxacin and ofloxacin but was slightly less potent than that of ciprofloxacin. This compound also showed good activity against drug-resistant strains such as methicillin-resistant Staphylococcus aureus and gentamicin-resistant Pseudomonas aeruginosa. The protective effects of a single oral dose of AM-833 on systemic bacterial infections in mice were greater than those of norfloxacin. AM-833 was as effective as ofloxacin and ciprofloxacin against systemic infections with Escherichia coli and Pseudomonas aeruginosa, and it showed somewhat higher activity against staphylococcal infections than did the other quinolones. AM-833 exhibited good prophylactic activity against E. coli infections. AM-833 also proved effective against localized infections such acute pneumonia and ascending urinary tract infections in mice. The excellent therapeutic efficacy of AM-833 against these systemic and local infections may be a result of its good oral absorption and high levels in tissues.

Comparative Activities of AM-715 and Pipemidic and Nalidixic Acids Against Experimentally Induced Systemic and Urinary Tract Infections

Dose for dose, AM-715 was at least five times as active as pipemidic acid and nalidixic acid against systemic and urinary bladder-kidney infections in mice.