Critical factors influencing growth of L forms of Proteus mirabilis.

The observations, primarily by Dienes and associates, of L colonies arising spontaneously or by penicillin induction from species of Proteus or Salmonella led to the description of two morphologically distinct colonial forms, designated as the 3B type and the 3A type. It has been reported further that the viable elements of these two types exhibit markedly dissimilar properties such as size, nutritional requirements, and ability to return to the bacilliform state (Dienes and Weinberger, 1951). Biochemical and enzymological analyses of 3B L forms have supported the concept that these elements are essentially spheroplasts and possess the capabilities of the bacillary cells from which they are derived (Kandler, Zehender, and Miller, 1956). In contrast, however, 3A type L forms are deficient in many of the capabilities displayed by the parent cells. The manifold uncertainties concerning the genetic, morphological, and biochemical characteristics of L forms and pleuropneumonia-like organisms (PPLO) have been extensively detailed in a recent symposium (Liebermeister, 1960; Sharp, 1960; Medill-Brown, Hutchinson, and Cocklin, 1960). Recent investigations (Landman, Altenbern, and Ginoza, 1958) have shown that a soft agar medium can be devised upon which every Proteus mirabilis cell plated would be converted to and would propagate as the 3B type of L form. There are no definitive data concerning conversion of bacillary cells to 3A type L forms. Preliminary qualitative experiments in this laboratory to determine conditions yielding the maximal number of 3A type L colonies from an inoculum of Proteus cells indicated that low penicillin concentration, low tonicity, and an incubation temperature of 30 C were favorable. Widely fluctuating quantitative results from seemingly identical experiments prompted a comprehensive examination of many factors possibly influencing growth of 3A type L colonies from an inoculum of Proteus cells. The data in the present paper establish the crucial importance of several unsuspected factors, which may partially account for the often encountered erratic nature of growth of L forms.