In acetate-limited chemostat cultures started with single-colony cultures of Thiobacilus versutus, a mutant appeared after approximately 85 volume changes. The inhomogeneity of the culture was detected by the development of two different types of colonies on agar plates. When a pure culture of the mutant was grown in a chemostat, parent colonies appeared after almost the same period of time. Electron micrographs of the mutant grown on butyrate showed the presence of fibrils surrounding the cells. The cells of the parent strain were bald when grown under the same conditions. The growth kinetics of the parent and the mutant were investigated in batch cultures with a variety of substrates and were found to be identical. Major differences between the two strains were observed during growth on mannitol; the mutant attained a lower yield and excreted large amounts of extracellular polysaccharides.The loss of homogeneity of a bacterial culture during cultivation in a rigidly controlled environment is not an unknown phenomenon. Calcott (3) recently summarized a number of studies in which parent-mutant transitions in chemostat cultures were observed. In continuous cultures, at nutrient concentrations much lower than the Ks, the outcome of the competition between parent and mutant depends on the pmaxIKs ratio. At higher substrate concentrations, the competition is always won by the organism with the lowest Ks, except for the situation in which the Monod curves cross over. In this situation, at substrate concentrations below the crossover point, K, is decisive, but at higher concentrations the organism with the highest growth rate is selected (16). Physiological causes of parent-mutant systems have been investigated in a number of cases (3). Although some of these studies have led to an understanding of the observed phenomenon, in some transitions take-over by a mutant without obvious reasons has been noticed.There are two possible ways to cope with the problem of the appearance of mutants in a pure culture. The first is to maintain a chemostat culture only for a short period of time, as suggested by Tempest (14); the second one is to wait until the most adapted organism has established itself (8).Here we report the appearance of a mutant with a different colonial morphology in a pure chemostat culture of Thiobacillus versutus. To identify the nature of this parentmutant transition, some growth kinetic and physiological parameters of both strains have been investigated in chemostat and batch cultures. MATERIALS AND METHODSOrganism, media, and cultivation. A T. versutus strain formerly called A2 (ATCC 25364) (5) was a gift from J. G. Kuenen (Delft, The Netherlands); in this paper this strain is called WS. The organism was maintained on thiosulfate agar slants stored at 4°C and subcultured every 2 months. The basal medium for growth in batch culture was that of Taylor and Hoare (13), containing 2 ml instead of 5 ml of trace element solution as described by Gottschal and Kuenen (4).For growth on methanol, formate, methylamine...
KEY WORDS Amicoacids Carbon dioxideevolution Casein Decomposition Glucose Humification Mineralization Respiration rate Phenolic compounds SUMMARYThe course of the CO 2 evolution rates of soil samples has been followed continuously in the absence and in the presence of various organic compounds. After an incubation period of 300 hours at 13 and 20~ the CO 2 evolution from pasture soil (containing 1.76~o soil organic carbon) amounted to 0.13 and 0.44 lag CO2-C.g soil -1.h -1, respectively. For arable soil (containing 1.20~ soil organic carbon) the rates amounted to 0.04 and 0.09 lag CO2-C.g soil-t.h 1, respectively.At 20~ larger amounts of the organic substrates added to the soil supplied with 20 lag NH4NO 3-N.g soil-1 were lost as CO2 than at 13~ indicating a higher efficiency of the growth of microorganisms at lower temperatures. In the absence of NH4NO3 the respiration rates were initially higher than in its presence, suggesting that a part of the soil microflora is inhibited by low concentrations of NH,NO3. The amounts of carbon lost were low for phenolcarboxylic acids with OH groups in the ortho position. The replacement of one of these groups by a methoxyl group resulted in a larger amount of the C lost as CO 2. The replacement of the COOH group by a C = C-COOH group had a decreasing effect on the decomposition of the phenolic acids tested. The decomposition of vanillic acid, p-hydroxybenzoic acid, and of the benzoic acids with OH groups in the meta position was as complete as that of glucose, amino acids or casein. The decomposition of bacterial cells to CO 2 was considerably less than that of glucose.No evidence could be obtained that the low percentage ofsubstrate converted to CO2 at the time of maximal respiration rate was due to the decreasing diffusion rate of substrate to the microbial colonies in the soil during the consumption of substrate.
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