Effect of osmolarity on sporulation in the spheroplasts of Saccharomycescerevisiae was examined. Spheroplasts were prepared from large cell preparation isolated from the stationary-phase cells by high-density centrifugation. The sporulation in the large cell preparation was characterized by its high synchrony. The sporulation in spheroplasts was induced with high synchrony and frequency under the osmotic condition near the lower limit for stabilizing spheroplasts, and with increasing the osmolarity the sporulation was delayed and proceeded asynchronously. A similar but rather more intense effect of the osmolarity on the sporulation was observed in whole cells. The sensitivity of spheroplasts to the inhibitor of DNA or protein synthesis synchronously changed during the sporulation culture at low osmolarity. The synchronous sporulation in spheroplasts may provide a system convenient for the investigation of the control mechanism of sporulation.
When Escherichia coli, K-10, was anaerobically grown on lactate in the presence of nitrate, the products formed were acetate (recovery in C atoms, 25%), formate (17%) and small amounts of pyruvate and ethanol. Nitrate was quantitatively reduced to nitrite. 14CO2 was formed from [2- 14C] lactate, whereas 14CO2 formation from [2-14C] glucose was almost nil. Citrate synthase, isocitrate dehydrogenase, 2-oxoglutarate dehydrogenase, succinyl-CoA synthetase, and malate dehydrogenase were twice as active in extracts from cells grown on lactate and nitrate compared with extracts from cells grown aerobically on glucose and 4-12 times as acitve with extracts from cells grown on glucose and nitrate. Activities of aconitate hydratase, succinate dehydrogenase, and fumarate hydratase were present in cells grown on lactate and nitrate. These enzymes showed maxima during the exponential phase of growth and activities of aconitate hydratase as well as succinate dehydrogenase fell to almost zero during the stationary phase. Essentially no activities of these three enzymes were found in cells grown on glucose and nitrate. Oxidation of lactate to carbon dioxide during nitrate respiration seems to be due to the operation of the tricarboxylic acid cycle in lactate-grown cells. Glucose- grown cells seem to lack this cycle.
The influence of several conditions on the attachment of rumen bacteria to cellulose powder was examined. The attachment of bacteria to cellulose occurred shortly after the addition of the cellulose, and maximum attachment was attained after 5min further incubation at 38°. The amount of bacteria attached to cellulose was approximately proportional to the quantity of cellulose powder until 2% in weight per volume. Reaction temperature had a very profound effect on the attachment of bacteria to cellulose. The amount of bacteria attached to cellulose increased as the reaction temperature was raised. There existed bacteria capable of attaching to cellulose even at 4°, though the amount of the bacteria attached was small. The specific amylase, urease and protease activities of bacteria attached to cellulose were remarkably low, compared with those of non-attachable bacteria. Conversely, the specific β-glucosidase activitiy of bacteria attached to cellulose was significantly higher than that of non-attachable bacteria. A trial was made to elute bacteria attached to cellulose powder first with an anaerobic salt solution and subsequently with 0.1% aqueous solution of methylcellulose. The constitution of bacteria eluted from cellulose with the methylcellulose solution varied in detail with samples. The most predominant group among bacteria eluted with the methylcellulose solution from cellulose were gram-negative rods and they accounted for 60.1 to 80.1% of the total bacteria eluted. The results of the specific amylase, β-glucosidase and urease activities of bacteria eluted with the methylcellulose solution from cellulose were similar to those of bacteria attached to cellulose.
More than 50% of the total aconitase activity in Escherichia coli was lost when cell-free extracts were prepared under air by a French press, by osmotic rupture of spheroplasts or by sonic treatment, as shown by the fact that the enzyme activity of the extracts prepared by sonic treatment under N2 was more than twice that of the extracts prepared by the other methods. When the extracts prepared by sonic treatment under N2 were gently shaken under air at 25°, about 70 to 80% of the original activity was lost over 30min, but a constant level of the enzyme activity remained even after prolonged incubation. The remaining aconitase activity after a long aerobic incubation was the same whether the extracts were prepared by sonic treatment under air or N2. Aconitase was stable when cell-free extracts were incubated under anaerobic conditions or when fluorocitrate, in concentrations less than 1μM, was added during aerobic incubation. Citrate was less effective than fluorocitrate in preventing enzyme inactivation. Aconitase in the extracts, which had been incubated under air with fluorocitrate, was stable against oxygen inactivation and showed different Vmax and Km values from those of the aconitase in the extracts that had been incubated under air without the addition of flourocitrate.
Responses of oligotrophic bacteria at microsites of soil to substrates at various concentrations were studied by means of capillary tubes. As a measure of dominance of oligotrophic bacteria, an index δ was proposed as follows; δ=(DNBcount-NBcount)/DNBcount where NB and DNB count are plate count on the conventional nutrient broth (NB) and its 100-fold dilution (DNB) respectively. The value of the index varied widely with the site and most frequently observed between 0 and 0.1 or 0.9 and 1.0, which is interpreted by an assumption that a population developed in a capillary tube consists of offspring of one or few of the original cells. The assumption is consistent with the J-shaped cell density histogram which is approximated by the Poisson distribution. Bacterial response to substrate is expressed by the mean δ value, three types of responses being observed; glucose and succinate were effective for the enrichment of oligotrophic bacteria, glutamate for zymogenic ones, and nutrient broth affected favorably either the former or the latter depending on its concentration. Baesd on these results a probabilistic view is proposed on bacterial behavior at microsites of soil.
Quinone systems, DNA base composition, cellular fatty acid composition, and phenotypic characteristics of 41 strains of named Flavobacterium species, 10 strains of named Cytophaga species, and 16 isolates belonging to the Flavobacterium-Cytophaga complex were investigated. These strains were divided into two major groups on the basis of the quinone systems and DNA base composition: The ubiquinone system (Q-8 or Q-10) and high values in DNA base composition (64.1 to 69.2%) were found in one group. The strains in this group were divided into three clusters based on the cellular fatty acid composition and phenotypic characteristics. In this respect they were related to Pseudomonas or Alcaligenes species. The other group was characterized by the presence of the menaquinone system (MK-6 or MK-7), low values in DNA base composition (30.6 to 44.8%), and the presence of iso- and anteiso-acids in the cellular fatty acids. The strains of this group were separated into four clusters based on cellular fatty acid composition. All the named strains of Cytophaga species were included in this group. Cytophaga species were recognized by chemotaxonomic characteristics, and the accommodation of Flavobacteriumaquatile strain Taylor F36 (=ATCC 11947=NCIB 8694), the type strain of the type species of the genus Flavobacterium, was discussed.