[ODP-079] The emergence of novobiocin-resistant mutants in actinomycetes and their characteristics
Novobiocin (NOV) is an aminocoumarin antibiotic that inhibits bacterial DNA synthesis. In a previous study, we found that in actinomycetes, NOV-resistant (NOVr) mutants could occur at a high frequency in the presence of NOV. The aim of this study is to verify this phenomenon and to elucidate the underlying mechanism.
Streptomyces coelicolor A3 (2), S. lividans 66, and S. curacoi NBRC 12761 were used in this study. Spores (1 × 105 to 1 × 108) were inoculated onto an agar medium containing NOV at the concentrations of 2- to 8-fold the minimum inhibitory concentration (MIC) and were incubated at 30°C for 7 days. Colonies that grew well on the NOV-containing agar medium were counted, and that number was used to calculate the frequency of NOVr mutants. In S. coelicolor, NOVr mutants occurred at the extremely high frequency of 5 × 10-5 when spores (1 × 105) were inoculated and incubated on an agar medium containing NOV at a concentration of 2-fold the MIC. Interestingly, the appearance frequency of NOVr mutants were reduced inversely to the increasing amount of inoculated spores. These results suggest that NOV can induce NOVr mutations in S. coelicolor. Incidentally, in S. lividans and S. curacoi, NOVr mutants also occurred at a high frequency. We are aiming to identify the mutated genes for understanding the mechanisms of NOV induction of NOVr mutations in actinomycetes.
Streptomyces coelicolor A3 (2), S. lividans 66, and S. curacoi NBRC 12761 were used in this study. Spores (1 × 105 to 1 × 108) were inoculated onto an agar medium containing NOV at the concentrations of 2- to 8-fold the minimum inhibitory concentration (MIC) and were incubated at 30°C for 7 days. Colonies that grew well on the NOV-containing agar medium were counted, and that number was used to calculate the frequency of NOVr mutants. In S. coelicolor, NOVr mutants occurred at the extremely high frequency of 5 × 10-5 when spores (1 × 105) were inoculated and incubated on an agar medium containing NOV at a concentration of 2-fold the MIC. Interestingly, the appearance frequency of NOVr mutants were reduced inversely to the increasing amount of inoculated spores. These results suggest that NOV can induce NOVr mutations in S. coelicolor. Incidentally, in S. lividans and S. curacoi, NOVr mutants also occurred at a high frequency. We are aiming to identify the mutated genes for understanding the mechanisms of NOV induction of NOVr mutations in actinomycetes.