The effect of silicon metabolism on genetic transformation in Bacillus subtilis

Germanium dioxide is found to increase the frequencies of the genetical transformation in Bacillus subtilis 30-40 fold. The increased frequency of transformation was registered in Sil- mutant in contrast to Sil+ strain having the decreased one. Bacillus megatherium strain KU-2 and Bacillus oligonitrophilus KU-1 were isolated from soil. These strains possess better ability to utilize the orthoclase and biotite. Germanium dioxide did not induce the transformation frequencies increase in these strains. Sil mutant of Bacillus oligonitrophilus demonstrated no competence to transformation.     

The effect of chemical fixatives on cell walls of Bacillus subtilis.

Cell walls of Bacillus subtilis were treated with several chemical fixatives which are commonly used preparatory to electron microscopy; i.e., osmium tetroxide, formaldehyde, acrolein, crotonaldehyde, and glutaraldehyde. Dimensional analysis was performed on thin sections of fixed walls from plastic embeddings and, by means of the statistical technique of multiple comparisons, significant differences were found between wall thicknesses from the various fixations. These differences varied with the fixation time and the type of fixative used in the reaction. When compared to embedded walls which had been stained before fixation, the overall effect was a reduction in wall thickness which was attributed to fixative action and not to the embedding or staining processes. The reduction of wall thickness was even more apparent when dimensions of fixed walls were compared to published dimensions of both frozen sections and freeze-etch profiles. Since these fixatives bind to reactive sites within the wall fabric, a change in electrochemical charge density is effected which can be monitored in terms of heavy-metal-binding capacity. Most monoaldehyde fixatives and osmium tetroxide render the wall as reactive, or less reactive, to uranyl acetate as unfixed walls, whereas glutaraldehyde can significantly increase the binding capacity.     

A study of the mechanisms of probiotic effect of Bacillus subtilis 8130 strain

The wild-type Bacillus subtilis strain 8130 secreted metabolites that stimulated two to three times the growth of the test cultures of lactic acid bacteria. It exhibited endoglucanase activity that depended on the composition of nutrient medium. The addition of the product of two-stage culturing of B. subtilis 8130 to the diet of pigs (0.2% of fodder weight) made it possible to increase the daily weight gain by 19% and decrease the consumption of mixed fodder by 10%. Digestion of protein, fat, and other organic compounds increased by 3-4% and cellulose by 12%. It was shown that B. subtilis 8130 is a probiotic with targeted action stimulating digestion (primarily the digestion of cellulose). The enrichment of a dry-beer pellet with the product of solid-phase fermentation by bacillus (1 x 10(8) cells per gram dry pellet) allowed the pellet to entered into the diet of a calf (6% of the weight of fodder with probiotic), causing additional weight gain by 12% and a 10% economy of fodder consumption.     

A study of the mechanisms of probiotic effect of Bacillus subtilis strain 8130

The wild-type Bacillus subtilis strain 8130 secreted metabolites that stimulated two to three times the growth of the test cultures of lactic acid bacteria. It exhibited endoglucanase activity that depended on the composition of nutrient medium. The addition of the product of two-stage culturing of B. subtilis 8130 to the diet of pigs (0.2% of fodder weight) made it possible to increase the daily weight gain by 19% and decrease the consumption of mixed fodder by 10%. Digestion of protein, fat, and other organic compounds increased by 3–4% and cellulose by 12%. It was shown that B. subtilis 8130 is a probiotic with targeted action stimulating digestion (primarily the digestion of cellulose). The enrichment of a dry-beer pellet with the product of solid-phase fermentation by bacillus (1 × 10⁸ cells per gram dry pellet) allowed the pellet to entered into the diet of a calf (6% of the weight of fodder with probiotic), causing additional weight gain by 12% and a 10% economy of fodder consumption.     

The essential YycFG two-component system controls cell wall metabolism in Bacillus subtilis

Adaptation of bacteria to the prevailing environmental and nutritional conditions is often mediated by two-component signal transduction systems (TCS). The Bacillus subtilis YycFG TCS has attracted special attention as it is essential for viability and its regulon is poorly defined. Here we show that YycFG is a regulator of cell wall metabolism. We have identified five new members of the YycFG regulon: YycF activates expression of yvcE, lytE and ydjM and represses expression of yoeB and yjeA. YvcE(CwlO) and LytE encode endopeptidase-type autolysins that participate in peptidoglycan synthesis and turnover respectively. We show that a yvcE lytE double mutant strain is not viable and that cells lacking LytE and depleted for YvcE exhibit defects in lateral cell wall synthesis and cell elongation. YjeA encodes a peptidoglycan deacetylase that modifies peptidoglycan thereby altering its susceptibility to lysozyme digestion and YdjM is also predicted to have a role in cell wall metabolism. A genetic analysis shows that YycFG essentiality is polygenic in nature, being a manifestation of disrupted cell wall metabolism caused by aberrant expression of a number of YycFG regulon genes.     

Effects of inhibitors on respiratory oscillations in synchronised Bacillus subtilis

Respiratory activity increased in a stepwise fashion during the cell cycle of Bacillus subtilis. This was true either for cells in growth medium or for those removed by filtration and supplied with an exogenous electron donor. KCN was used to probe terminal oxidase activity and cell cycle-dependent oscillations in the degree of inhibition were measured. The extent of stimulation by an uncoupling agent (CCCP) also varied during the cycle rising to two maxima at 0.14 and 0.69, intermediate to the step rises in oxygen uptake rates (0.4 and 0.8 of the cycle). However the net effect of the uncoupler was to convert the discontinuous pattern of oxygen uptake to a continuous one. These data are consistent with control of respiratory chain activity during the cell cycle of B. subtilis, and not of the amount of its components.     

Membrane synthesis in synchronous cultures of Bacillus subtilis 168

Synthesis of bacterial membranes has been investigated in Bacillus subtilis by examining incorporation of amino acids and glycerol into the protein and lipid of membranes of synchronous cultures. A simple reproducible fractionation scheme divides cellular proteins into three classes (i) truly cytoplasmic, (ii) loosely membrane bound, released by chelating agents, and (iii) tightly membrane bound. These comprise approximately 75, 10, and 15%, respectively, of cellular proteins in this organism. Incorporation of radioactivity into these fractions, using steady-state and pulse labeling has been followed during the cell cycle. Cytoplasmic proteins and the loosely membrane-bound proteins are labeled at an exponential rate throughout the cell cycle. The membrane fraction is labeled discontinuously in the cell cycle, with periods of rapid synthesis over the latter part of the cycle and a period with no net synthesis during the early part of the cycle. Pulse labeling indicates that synthesis of membrane occurs at a linear rate that doubles at a fixed time in each cycle, which coincides with the period of zero net synthesis. Rates of membrane synthesis measured by pulse labeling during the period of rapid membrane synthesis are significantly less than indicated by steady-state labeling. These discrepancies are consistent with the hypothesis that during the cell cycle certain proteins are added to the membrane from the cytoplasm and that during the period of zero net synthesis there is an efflux of proteins from the membrane. Evidence in favor of this has been presented. The activity of succinic dehydrogenase (a representative of class c) varies in a step-wise manner with periods of rapid increase, approximately coincident with bursts of membrane protein synthesis, alternating with periods without any increase in activity. The activities of malate dehydrogenase (class a) and reduced nicotinamide adenine dinucleotide dehydrogenase (class b) increased throughout the cell cycle. Phospholipid synthesis is continuous throughout the cell cycle.     

Protein secretion in phosphate-limited cultures of Bacillus subtilis 168

The secretion of proteins from Bacillus subtilis was studied under physiologically well-defined conditions in continuous cultures at a range of specific growth rates. The kinetics of secretion was analysed by using pulse-chase and immunoprecipitation techniques that allowed both processing and release to be monitored. Growth conditions were selected that were known to lead to significant changes in the anionic polymer composition of the cell wall. Under magnesium limitation only low levels of native proteins were released into the growth medium. In contrast, much higher amounts of released protein were observed under phosphate limitation. Although synthesis of native secretory proteins appeared to be highly regulated, only minor changes in the secretion of heterologous proteins were detected. Comparable kinetics of protein release of cells grown under different conditions indicated similar cell wall permeabilities. The large changes in the amounts of released proteins were not reflected in the production of chaperones and components required for protein secretion. The data suggest that the capacity of the secretion machinery is not a major limiting step in the export of native secretory proteins. Received: 23 September 1997 / Received revision: 10 November 1997 / Accepted: 16 November 1997     

Effects of ethanol and methanol on lipid metabolism in Bacillus subtilis

In Bacillus subtilis, the fatty acid moiety of the phospholipids was affected differently during growth in the presence of 1.1 M-methanol or 0.7 M-ethanol, though at these concentrations methanol and ethanol had the same effects on growth rate and completely inhibited sporulation. Synthesis of phosphatidylglycerol was also strongly inhibited and the amount of total cell phospholipids was reduced by 50% by both alcohols. The composition of fatty acids, especially the relative concentration of 12-methyltetradecanoic acid, was modified only by ethanol; in bacteria grown in the presence of methanol, changes in fatty acid composition were negligible. In non-sporulating mutants, synthesis of phosphatidylglycerol was much less affected than in the wild-type and synthesis of phosphatidylethanolamine was increased. In these strains, fatty acid composition was also modified by ethanol but unaffected by methanol.