Browsing by keyword "Tetracycline Resistance"

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    • Mutation spectrum in Escherichia coli DNA mismatch repair deficient (mutH) strain
      Rewinski, Caroline; Marinus, Martin G. (1987-10-26)
      The Dam-directed post-replicative mismatch repair system of Escherichia coli removes base pair mismatches from DNA. The products of the mutH, mutL and mutS genes, among others, are required for efficient mismatch repair. Absence of any of these gene products leads to persistence of mismatches in DNA with a resultant increase in spontaneous mutation rate. To determine the specificity of the mismatch repair system in vivo we have isolated and characterized 47 independent mutations from a mutH strain in the plasmid borne mnt repressor gene. The major class of mutations comprises AT to GC transitions that occur within six base pairs of the only two 5'-GATC-3' sequences in the mnt gene. In the wild type control strain, insertion of the IS1 element was the major spontaneous mutational event. A prediction of the Dam-directed mismatch repair model, that the mutation spectra of dam and mutH strains should be the same, was confirmed.

    • Mutations produced by DNA polymerase III holoenzyme of Escherichia coli after in vitro synthesis in the absence of single-strand binding protein
      Carraway, Margaretha; Rewinski, Caroline; Marinus, Martin G. (1990-10-01)
      Single-stranded plasmid DNA, containing the mnt gene, was replicated in vitro with DNA polymerase III holoenzyme. Escherichia coli mutH bacteria, defective in mismatch repair, were transformed with the products of in vitro synthesis. Mutations in mnt were readily identified and 33 out of 65 isolates were single base changes including transition, transversion and frameshift mutations. The remaining 32 isolates were deletions of apparently random length and substitutions (deletion/insertions). The intergenic deletions as well as the transition and frameshift mutations were identical to those previously isolated from mismatch repair-defective cells in vivo.

    • National collaborative study of the prevalence of antimicrobial resistance among clinical isolates of Haemophilus influenzae
      Doern, Gary V.; Jorgensen, James H.; Thornsberry, Clyde; Preston, David A.; Tubert, Tracy A.; Redding, Judith S.; Maher, Louise A. (1988-02-01)
      A total of 2,811 clinical isolates of Haemophilus influenzae were obtained during 1986 from 30 medical centers and one nationwide private independent laboratory in the United States. Among these, 757 (26.9%) were type b strains. The overall rate of beta-lactamase-mediated ampicillin resistance was 20.0%. Type b strains were approximately twice as likely as non-type b strains to produce beta-lactamase (31.7 versus 15.6%). The MICs of 12 antimicrobial agents were determined for all isolates. Ampicillin resistance among strains that lacked beta-lactamase activity was extremely uncommon (0.1%). Percentages of study isolates susceptible to cefamandole, cefaclor, cephalothin, and cephalexin were 98.7, 94.5, 87.3, and 43.3%, respectively. For 14 strains (0.5% of the total), chloramphenicol MICs were greater than or equal to 8.0 micrograms, and thus the strains were considered resistant. All of these resistant strains produced chloramphenicol acetyltransferase. In addition, all 14 strains were resistant to tetracycline; 11 produced beta-lactamase. The percentage of isolates susceptible to tetracycline was 97.7%. In contrast, erythromycin and sulfisoxazole were relatively inactive. The combination of erythromycin-sulfisoxazole (1/64) was more active than erythromycin alone but essentially equivalent in activity to sulfisoxazole alone. Finally, small numbers of clinical isolates of H. influenzae were resistant to trimethoprim-sulfamethoxazole and rifampin.

    • Role of plasmid multimers in mutation to tetracycline resistance
      Boe, Lars; Marinus, Martin G. (1991-10-01)
      As an additional system for analysing mutations that appear to be specifically induced or directed, we have used a plasmid that contains the mnt repressor gene inserted as an operon fusion with the tet gene of the plasmid pBR322. Thus, the mnt gene product acts as a negative transcriptional regulator of tet gene expression. Mutations inactivating the Mnt repressor are recessive while those destroying operator recognition (Oc) are dominant in conferring tetracycline resistance on the host. When resistance mutations were isolated on plates with high levels of tetracycline they were preferentially mnt- and the plasmids were monomers. Pre-exposure to low concentrations increased the frequency of resistant mutants by 100- to 1000-fold, and the mutations were now mostly Oc, located on one unit of a plasmid multimer. Recessive repressor mutations on one unit would not have been selected. We suggest that the high frequency of mutation in tandem multimeric plasmids may be caused by the formation of single-stranded and hence highly mutable regions by homologous pairing out of register. The role of tetracycline in promoting mutations is discussed.

    • Sequence specificity of streptozotocin-induced mutations
      Mack, Susan L.; Fram, Robert J.; Marinus, Martin G. (1988-10-25)
      The isolation and characterization of streptozotocin (STZ)-induced mutations in the phage P22 mnt repressor gene is described. Cells carrying the plasmid-borne mnt gene were exposed to STZ to give 10-20 percent survival and at least an eleven-fold increase in mutation frequency. DNA sequence analysis showed that 50 of 51 STZ-induced mutations were GC to AT transitions, and one was an AT to GC transition. We have also compared the STZ mutational spectrum to that for N-methyl-N'-nitro-N-nitroso-guanidine (MNNG). There are sites in the mnt gene which are mutated only by STZ; only by MNNG, or by both agents. Sites at which only STZ induced GC to AT transition mutations occur were in sequences that are pyrimidine rich 5' to the mutated site and purine rich 3' to the mutated site. Induction of mutations by both STZ and MNNG should be considered to maximize the number of mutable sites.