Several physical properties of spores of Bacillus megaterium, such as vapor pressure isotherm, complex dielectric constant, nuclear magnetic resonance absorption spectrum, as well as water sorption and desorption rates, were studied in comparison with those of vegetative cells of the same organism with respect to content and form of the cell water. Spores showed larger water content as compared with vegetative cells in the humidity range of less than 78% (R. H.) at 30°, while they showed smaller water content over more humid ranges up to 100% (R. H.). Spores showed a smaller dielectric loss at a microwave frequency of 10Gc and a broader NMR absorption pattern than vegetative cells. These facts are discussed briefly in terms of molecular motion of the cell water in spores and vegetative cells.
Synthetic activities of macromolecules by temperature-sensitive mutants of Pseudomonas-P were examined at elevated temperatures. One of the strains, Tms-7, exhibited unusually high synthetic activities of protein and nucleic acids under such conditions. Nucleic acid synthesis of this strain continued even in the absence of histidine, which is an amino acid required for growth. No appreciable protein synthesis occurred under these conditions. Analysis of nucleic acids synthesized in a histidine-free medium revealed that radioactive phosphate was incorporated into DNA, soluble and ribosomal RNAs, as well as into other heterogeneous RNAs. The synthesized RNAs were highly susceptible to enzymatic hydrolysis. The relaxed control of nucleic acid synthesis, however, appears to be unrelated to the temperature sensitivity.
The observations made on the relationships existing between various microorganisms isolated from natural sources have been described. The species of Chaetomium were found to be antagonistic towards some other fungi. Competition among microorganisms was noticed in many instances. Mutualistic symbiosis was found to occur between a sterile strain of Thielavia setosa, which formed cleistothecia only in the presence of other fungi like Aspergillus nidulans, A. variecolor, Thielavia sepedonium, and T. terricola.
Of various kinds of mycological character, the following characteristics were found to be the most common in aflatoxin-producing strains: Color of conidial heads green even in old culture; the reverse side of colony wrinkled and colored; presences of sclerotia, globose to subglobose vesicles, biseriate sterigmata and conidiophore with roughened walls; high productivities of pigment, total acid and kojic acid; low abilities of browning rice koji and of production of deferriferrichromes. Sixteen strains were thus selected from 160 industrial strains as mycologically somewhat similar ones to the so-called aflatoxin strains, and their aflatoxin productivity, together with that of the aflatoxin strains, was investigated by the same methods as those described in Parts I and II. It was found that all of the industrial strains and four of the aflatoxin strains did not produce aflatoxin. Thus, it was concluded that the mycological characters described above are the ones necessary to a fungus for the production of aflatoxin but that the fungus having these characters may not always produce aflatoxin.
Using Aspergillus niger 1617 as a test organism, the intracellular distribution of acid and alkaline phosphatases in growing hyphae and fruiting bodies was studied cytochemically and biochemically. Microscopical examination of the thin-layer cultures, devised specially for the present purpose, enabled distinction of younger from older hyphae. In younger hyphae, alkaline phosphatase was localized mainly in the nuclei, whereas in older ones it became dispersed in cytoplasm. Acid phosphatase in younger hyphae was distributed to surface structure, mitochondria, microsome, and cell sap, and that in older hyphae showed a similar distribution pattern. Both enzymes were highly active in rudimentary fruiting bodies. DEAE-cellulose column chromatography and acrylamide-gel electrophoresis revealed that both phosphatases were composed of at least two components and the relative balance of these components changed characteristically during the course of cultural development.
An extremely UV-resistant stage was found to appear only temporarily in the early phase of germination of B. subtilis spores. Kinetic analysis of the course of germination with special reference to ultraviolet and heat resistance revealed that the germinating spores remain at this transient stage for one to six minutes depending on the condition of germination, and then they are converted into germinated form with the loss of UV resistance. The effect of heat activation, germination temperature, pH, and composition of germination medium on the duration of this transient stage was studied. From the effect of metal ions, added to the germination medium, the hydrolytic degradation of spore mucopeptide catalyzed by an enzyme similar to STRANGE-DARK'S "cell-wall lytic enzyme" was considered to be responsible for the conversion of physiological characteristics of spores from the UV-resistant transient stage to germinated form.
Some general structural features of dormant and germinating spores of a thermophilic actinomycete, Micromonospora vulgaris, are described. The structure of these spores differs considerably from that in other actinomycetes so far reported. The mature dormant spore of Micromonospora vulgaris is covered with multilayered integuments consisting of outer spore coat, intermediate spore coat, inner multilayered coat and cortex-like coat. The spore core is separated from the cortex-like coat by a boundary region. A net-like texture is sometimes visible in the spore core. On germination the spore core swells. In the boundary region a plasma membrane and cell wall of the emerging cell seem to be synthesized. Ribosome granules of cytoplasm, extensive intracytoplasmic membrane systems, nuclear region and an unidentified fine granular structures appear to be visible in the germinating cell. Spore integuments show signs of degradation in the process of germination, this process being markedly in progress at the stage of emergence.
CCytological studies on the cellular components of streptomycin-sensitive strain B3 of Bacillus subtilis and its two resistant strains 2R2B3 and 2R3B3 reveal that development of streptomycin resistance is accompanied by an increase in both DNA and RNA content of cells. The resistant strains are able to concentrate in higher amounts 13 different amino acids in the free state in the internal environment. The levels of all the amino acids and amino sugars of cell walls are higher in the resistant strains.
Studies on the utilization of amino acids by B. subtilis during growth in the presence of streptomycin indicate that the antibiotic greatly influences the rate of amino acid uptake by the organism. There is, however, no selective action in the ultimate utilization of amino acids. The variation in the rate of assimilation of amino acids from the broth as induced by streptomycin may account for the difference in the rate of growth of bacterial cells. The antibiotic causes the accumulation of lesser number of free amino acids as compared to those accumulated in the streptomycin- untreated cells.