Formation of Spheroplasts from Bacillus anthracis

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Chatterjee, B. R. (Baylor University College of Medicine, Houston, Tex.), and Robert P. Williams. Formation of spheroplasts from Bacillus anthracis. J. Bacteriol. 89:1128–1133. 1965.—Spheroplasts were prepared from Bacillus anthracis by combined treatment with lysozyme and glycine. Glycine, at a final concentration of 3%, was added to cultures of B. anthracis in nutrient broth that had grown at 37 C for 16 to 18 hr under 50% CO2. After additional incubation under CO2 for 2 hr, lysozyme, at the appropriate concentration (50 to 100 μg/ml), and sucrose, to a concentration of 15%, were added, and incubation was continued for 2 to 6 hr in CO2. At the end of this period, incubation in CO2 was discontinued. Spheroplasts formed after incubation in air for 6 to 12 hr. Lysozyme alone exhibited the same effect when added at much higher concentrations (500 to 2,000 μg/ml) to cultures growing under CO2. No spheroplasts formed when cultures were treated with glycine alone. Treatment with lysozyme was more effective on smooth strains than rough. Cells from young cultures were more susceptible to lysozyme than older cells. CO2 apparently was essential for formation of spheroplasts from B. anthracis.

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