SUMMARY: was grown in media containing various concentrations of benzylpenicillin; 0.05 μg./ml. was without effect and 0.40 μg./ml. inhibited growth. Intermediate concentrations in increasing steps of 0.05 μg./ml. were investigated. The Gram-positive cocci tended to become Gram-negative rods, the most effective concentration of penicillin for this effect was 0.15 ± 0.05 μg./ml. The rod forms frequently had surface vesicles which appeared to be membrane-bounded but were osmotically stable; the dry wt yield/ml. medium was diminished. Such forms contained up to 10 times more nisin/unit dry wt than the control. At penicillin 0.20 to 0.25 μg./ml. growth was followed by lysis which was followed by new growth after a long delay; morphologically these forms resembled the controls and the dry wt yield increased. Higher concentrations of penicillin (0.35 μg./ml.) decreased the dry wt yield/ml. medium and growth appeared only after a delay of 16 hr. Morphologically these forms tended to elongate again and the cell nisin/unit dry wt was 8 times higher than the control. Single-colony isolates obtained from the higher penicillin concentrations were grown in penicillin-free medium and when again challenged with penicillin no longer formed rods at 0.15 μg./ml. This more penicillin-resistant population occurred at a frequency of 1: 500 of the parent population and was distinguished from it by a number of physiological properties. These results suggest that penicillin acted by selecting resistant individuals already present in the parent population.


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