A recombinant plasmid containing gyr A encoding wild-type Escherichia coli quinolone susceptible DNA gyrase A subunits has been used as a broad host range gene probe. Strains expressing gyr A-mediated quinolone resistance become susceptible to quinolones upon insertion of the plasmid, whereas the plasmid without gyrA (pLA2917, vector) has no effect. Fifteen highly ciprofloxacin-resistant E. coli and three Klebsiella pneumoniae (MICs 2-64 mg/L) were isolated from clinical specimens in the Hospital de la Princesa, Madrid, Spain. Plasmid pNJR3-2 and pLA2917 were introduced into the clinical isolates by conjugation, and transconjugants selected with tetracycline or kanamycin (for which the plasmids encode resistance). Ten transconjugants from each mating, the original isolates, the gene probe and vector control were screened for susceptibility to nalidixic acid, ciprofloxacin, ofloxacin, norfloxacin, tetracycline, chloramphenicol, cefoxitin and trimethoprim. Lower MICs of quinolones were seen for the transconjugants of two K. pneumoniae isolates in the presence of the gene probe, suggesting that these isolates harboured mutations in gyr A. Plasmid profiles confirmed the presence of the probe. The susceptibility of the third K. pneumoniae strain and all E. coli isolates were unaffected by insertion of the plasmid, suggesting another mechanism was responsible for quinolone resistance.