Phage adsorption and productive lysis in stable protoplast type L-forms of Bacillus subtilis and Streptomyces hygroscopicus.

Transferable productive lysis in stable protoplast type L-form cells of Bacillus subtilis was produced by 6 phages out of 14 strains virulent for the parent B. subtilis 170 and 1997. Most of these phages lytic for L-forms show the phi 29 morphology characteristic for the smallest B. subtilis phages containing double-stranded DNA. Among 31 actinophages, 23 of which were virulent for Streptomyces hygroscopicus, only SLE 109 and phi c 31 gave productive infection of the stable protoplast type L-form of S. hygroscopicus NG 33--354. Electron microscopic investigation and treatment by DNAse demonstrated that infection of L-form cells is an adsorption-injection process, and that it is not caused by transfection of free phage DNA or endocytotic uptake of phage particles. Because in both stable L-forms cell wall biosynthesis is blocked irreversibly the results allow the conclusion that specific receptors must be localized in the cytoplasmic membrane for those phages producing transferable lysis in protoplast type L-forms. Localization of receptors for certain phages in the cytoplasmic membrane seems to occur in many Gram-positive bacteria, but not in Gram-negative bacteria.

Properties of Streptomyces phage SLE111.

Phage SLE111 infecting Streptomyces levoris 1331 was morphological different from most actinophages described and yielded very high titres (10(11) p.f.u/ml) after lytic growth. Typical morphological changes of infected hyphae were observed by phase contrast and electron microscopy of ultrathin sections. The double-stranded, linear DNA (44.2 +/- 1.3 kb) was characterized, according to electron microscopic analysis, by absence of cohesive ends, of terminal redundance, and circular permutation. A stem-loop structure containing inverted repeats of about 200 bp was identified by electron microscopy in two alternative positions in the SLE111 genome.

Inverted duplication in the genome of the temperate Streptomyces phage SH3.

The DNA of the temperate Streptomyces phage SH3 contains 100 base-pair long inverted repeats separated by a 940 base-pair long segment of DNA as revealed by electronmicroscopic analysis of snapback structures formed after rapid intrastrand reannealing of denatured DNA. The inverted repeat structure was found preferentially at map unit 22 of the circular physical map, in rare cases also in other positions, suggesting a movable character of this genetic element.

Interaction of bacteriophage O2 with strains of the genus Oerskovia.

Bacteriophage O2 multiplies normally on Oerskovia turbata IMET 47 153. It has a burst size of about 100 p.f.u. per infected cell and a latent period of 100 min at 30 degrees C. On Oerskovia xanthineolytica IMET 47 383 clear spots were formed after addition of high phage concentrations onto agar top layers. By phase contrast observation, and measurement of the optical density of infected cultures, it was found that the clearing effect on strain IMET 47 383 was due to lysis-from-without. Phage O2 adsorbs and injects its DNA into cells of strain IMET 47 383 but phage multiplication does not occur, and the phage DNA becomes degraded. Inhibition of phage DNA injection by the combined action of xanthotoxin -- u.v. irradiation abolished the clearing activity of phage lysates. Therefore, both adsorption and DNA injection seem to be prerequisites for the release of a lytic activity out of the phage particle, which is responsible for the clearing effect on strain IMET 47 383.

Characterization of a stable spheroplast type L-form of Proteus mirabilis D 52 as cell envelope mutant. II. Electronmicroscopic investigations.

The ultrastructural analysis of a stable spheroplast type L-form of Proteus mirabilis D 52 revealed characteristic alterations in the organization of the cell envelope including defective changes in the cell envelope structure as for instance the loss of a coherent murein layer, the loss of some components in the outer cell wall layer, the formation of small membraneous vesicles at the tips of loose extensions of the cell wall, a decrease in associations and bindings between wall and membrane, an extension of the periplasmic space, an increase in membrane defects, as well as a disturbed cell division causing unusual modes of multiplication and the formation of various intracellular structures like membrane complexes, characteristic sheet-like membraneous bodies, typical inclusion bodies, and defective phage structures, which all could not be observed in normal rod-shaped cells. The results of these investigations and of those given in a previous paper (Gumpert and Taubeneck 1975) show, that the stable spheroplast type L-form LD 52 B must be considered as a true cell envelope mutant in which the biosynthesis and structure of the cell envelope is altered genetically by one or several mutations whereas the main biochemical activities are the same like those of the parent bacterium. The profound alterations in the cell envelope system, however, lead to some changes in the whole cell organization, which apparently in turn cause disorders even in metabolic and biosynthetic processes not directly involved in the biosynthesis of the cell envelope.

Characterization of a stable spheroplast type L-form of Proteus mirabilis D 52 as cell envelope mutant. I. Isolation, growth characteristics, biochemical activities, and sensitivity to bacteriophages.

A stable spheroplast type L-form could be isolated by transferring 627 single colonies and 195 agar blocks with several colonies of unstable L-forms of Proteus mirabilis D 52 on agar media without supplements of penicillin. The L-form grows well on complex and synthetic agar media, however, it failed to grow in any of the liquid media which have been proved. With one exception (formation of acid from maltose) the L-form shows the same bioche mical activities like the parent rod-shaped bacterium. However, the insensitivity for various phages and the failure of DAP in the envelopes demonstrate that there are profound alterations in the biosynthesis and structure of the murein and of the outer wall layers. The results of these investigations and an ultrastructural analysis (Gumpert and Taubeneck 1975) show that the stable spheroplast type L-form LD 52 B of Proteus mirabilis must be considered as a true cell envelope mutant.