Curing of an R factor from Escherichia coli by hydroxyurea and cytosine arabinoside

Hydroxyurea (HU) and cytosine arabinoside (Ara-C) eliminate R factor R46 from Escherichia coli strain J5-3. The highest frequency of elimination for both drugs occurred at concentrations and times that produced the lowest survivor levels. 5% of cells were antibiotic-sensitive after 5 h incubation in 5 mg/ml Ara-C, whilst 6 and 7% of the survivors had lost the R factor after 8 and 24 h incubation, respectively in 20 mg/ml HU. The number of survivors began to increase after 5 h incubation in Ara-C, probably due to the inactivation of the drug in the phosphate-buffered medium. All four antibiotic resistances (ampicillin, streptomycin, sulphonamide and tetracycline) mediated by R46 were lost simultaneously and elimination of the whole plasmid was confirmed by conjugation experiments in which the 'cured' cells was shown to be as efficient recipients of R46 as the control R- strain in crosses with E. coli strain J6-2 (R46). No covalently closed circular plasmid DNA was demonstrable in cured cell lines. R+ and R- strains had similar sensitivities to HU but J5-3 R+ was much more sensitive to Ara-C than the isogenic R- strain. It may be concluded that HU eliminates R46 actively whereas although Ara-C may produce some active elimination it acts mainly by selection of R- cells present in the culture.