Preparation of Spheroplasts from Vibrio Comma.

Chatterjee, B. R. (Baylor University College of Medicine, Houston, Texas), and Robert P. Williams. Preparation of spheroplasts from Vibrio comma. J. Bacteriol. 85:838-841. 1963.-Spheroplasts were prepared from several strains of Vibrio comma by lysozyme treatment combined with freezing and thawing of the organisms. The optimal concentration of lysozyme was 50 mug/ml, although some spheroplasts formed at a concentration of 10 mug/ml. Higher concentrations (200 mug/ml) caused lysis of cells along with spheroplast formation. Treatment was carried out in broth cultures containing 15% sucrose, and if the osmotic tension was lowered the spheroplasts lysed. Some motile, spherical cells were present in every preparation. Addition of 3% glycine to broth cultures resulted in rapid transformation of the vibrios into large, spherical bodies. However, these were actively motile, and were not sensitive to a lower osmotic tension. Therefore, they could not be considered as spheroplasts.