Sporulation in Bacillis subtilis. Biochemical changes

1. During the course of growth and sporulation of Bacillus subtilis in chemically defined media, measurements were made of 16 different parameters, including the specific activities of nine intracellular enzymes. 2. Towards the end of exponential growth, proteolytic activity increased and reached a maximum soon after growth ceased. 3. In the presence of an excess of phosphate the specific activity of alkaline phosphatase increased fivefold at the end of exponential growth. 4. The specific activity of malate dehydrogenase remained at a high constant level throughout sporulation, but the specific activity of fumarase showed a two- to three-fold increase 5–9hr. after the end of exponential growth. 5. Aconitase activity was barely detectable during exponential growth in a glucose–glutamate medium, but increased rapidly when glutamate was replaced by citrate or when the glucose in the medium was exhausted. 6. The specific activity of alanine dehydrogenase increased threefold 1–5hr. after the end of exponential growth. 7. The specific activity of soluble NADH oxidase doubled 4–6hr. after the end of exponential growth. 8. Glucose dehydrogenase was undetectable until 4hr. after the end of exponential growth, but its specific activity increased 20-fold over the next 3–4hr. 9. The onset of refractility, the synthesis of 2,6-dipicolinic acid and the appearance of heat-resistance occurred in this order some 6–12hr. after the end of exponential growth. 10. The significance of these changes is discussed in relation to the morphological development of the spore.     

Kinetics of petite mutation and thermal death in Saccharomyces cerevisiae growing at superoptimal temperatures.

Mass formation of petite mutants took place in a strain of Saccharomyces cerevisiae when grown at superoptimal temperatures. After an initial period of exponential growth, a second period followed during which exponential death and net exponential petite mutation concurred with exponential growth. The specific rates of the three exponential processes were of the same order of magnitude and varied with the temperature. Net exponential petite mutation did not occur during the deathless first period of growth at superoptimal temperatures nor at any time during growth at suboptimal temperatures. Mitochondria are discussed as possible targets of thermal death in mesophilic yeasts.     

Changing proportions of DNA components in Euglena gracilis

The ratios of satellite deoxyribonucleic acid components to chromosomal deoxyribonucleic acid in Euglena gracilis Z were measured by analytical density gradient ultracentrifugation. Chloroplast deoxyribonucleic acid with a buoyant density of 1.685 g/cm³ exhibited a constant ratio to chromosomal deoxyribonucleic acid during exponential growth and increased twofold as the culture reached the end of the exponential growth phase. The quantity of a satellite deoxyribonucleic acid with a buoyant density of 1.691 g/cm³ was not sufficient to measure the ratio to chromosomal deoxyribonucleic acid during exponential growth but increased to approximately equal the quantity of chloroplast deoxyribonucleic acid as the culture approached the end of the exponential growth phase. The quantity of a deoxyribonucleic acid component with a buoyant density of 1.700 g/cm³ was not sufficient to measure the ratio to chromosomal deoxyribonucleic acid during exponential growth but represented approximately one-third of the total deoxyribonucleic acid as the culture entered the stationary phase of growth.     

Non-exponential growth by mammalian cells in culture

The concept of exponential growth by mammalian cells in culture is based upon the apparent linearity of semilogarithmic data plots. This method of graphical analysis is known to be an unreliable test of the exponential hypothesis. We have re-examined the question of growth exponentiality using the more sensitive method of Smith plots, in which specific growth rate is plotted against either time or density on transformed graphical coordinates which linearize the mathematical expression of the growth hypothesis being tested. With exponential growth, data points fall on a horizontal straight line when specific growth rate is plotted against time or density. Using both our own and literature data, we have performed Smith plot analyses on the growth of 125 different mammalian and avian cell lines. Of these, only eleven exhibited an exponential phase. The remaining cell lines all had non-exponential growth patterns. The most common of these consisted of an initial period of growth acceleration followed by a later phase of deceleratory growth. A smaller number of lines exhibited deceleratory kinetics at all times after plating. We conclude that mammalian cell growth in culture is predominantly non-exponential, and that the apparent exponentiality of semilogarithmic data plots is usually a methodological artifact.     

Casein Utilization by Streptococcus thermophilus Results in a Diauxic Growth in Milk

In milk, Streptococcus thermophilus displays two distinct exponential growth phases, separated by a nonexponential one, during which proteinase synthesis was initiated. During the second exponential phase, utilization of caseins as the source of amino acids resulted in a decrease in growth rate, presumably caused by a limiting peptide transport activity.     

Reproduction rate,feeding process,and leibich limitations in cell populations—Part 1. Feeding stochasticity and reproduction rate

A population of cells suspended in a liquid nutrient medium is considered. The process of growth, division and death of a cell is interpreted mathematically as the Bellman-Harris stochastic process governed by random meetings between the cell and nutrient particles. Growth of a cell is considered to be a result of two processes: mass inflow into and mass outflow from the cell. It is found that, in the absence of food limitations and inhibitors, population growth is not exponential. However, the exponential increase is approached asymptotically over time. Population net growth rate is a variable rather than a constant, but tends over time to a constant value which is the rate of exponential growth. The rate of exponential growth, the probabilities of cell division and death, and the life expectancy of a cell are expressed analytically via average rate of meetings between a cell and nutrient particles. The paper presents an independent phase in calculating mathematical relations between the rate of exponential growth and the concentration of food in a substrate.     

Growth and Enzyme Activity of Myxococcus virescens in Liquid Medium

The object of this work was to develop a method for determination of the growth of Myxocuccus virescens in liquid medium. The bacteria were grown in N III-B medium in 100 ml Kjeldahl flasks, which were fixed on a disc forming an angle of 50 degrees with the horizontal plane. The disc was rotated two full revolutions per minute. The total nitrogen content of the washed swarms, developing on the glass walls of the flasks, was used as an expression for the myxobacterial growth. After a lag the bacteria grown in total darkness had a growth phase, approximately exponential, of about 270 hours, which was followed by a steep phase of decline. When the bacteria were illuminated daily for a short period, a lag of 50–200 hours appeared in the middle of the exponential growth phase, after which a new exponential growth phase began. This second growth seemed to depend on variants insensitive to light induced lysis. The increase of enzymes, active on casein and autoclaved aerobacter cells, closely followed the first part of the growth curve. However both activities began to decrease before the growth maximum. No sign of proteolytic activity or lytic activity on autoclaved aerobacter cells could be detected after about 700 hours' incubation. In illuminated flasks it is shown that the production of yellow pigments in culture solution is sharply increased at the end of the exponential growth phase. The lytic enzymes of M. virescens seem to be extracellular, secreted during the exponential phase of growth. No activity was exhibited by washed cell swarms, even if they were sonically disintegrated.     

Biochemical changes accompanying brefeldin a formation inCurvularia lunata

Extracellular brefeldin A was detected in 4 % glucose-peptone-mineral salts cultures ofCurvularia lunata at the start of the exponential growth phase. Some fluctuations in brefeldin A levels occurred during the exponential growth phase followed by a significant reduction in level at the stationary growth phase. Broth glucose levels decreased according to a sigmoid relationship with time whereas broth pH remained fairly constant during the exponential growth phase followed by a gradual increase into the stationary growth phase. Mycelial brefeldin A levels were low throughout the various growth phases. The principal fatty acids present in decreasing order during the exponential growth phase were linoleic, oleic and palmitic acids. However, the content of linoleic acid was significantly reduced at the onset and during the stationary growth phase.     

Evaluation of growth hormone production from GH1 cells in vitro: Effect of culture media and time in culture

Summary Growth hormone production by a rat pituitary tumor cell line (GH1) was measured during lag, exponential, and plateau phases of growth in different culture media. Growth hormone secretion was low during lag and early exponential phase; it increased late in the exponential phase and continued to increase during the plateau phase. This biphasic pattern of growth hormone production was observed in all media and sera utilized. Both the doubling time and growth hormone production were influenced by the choice of media and sera. In addition, the length of time in culture affected the growth fraction with passage level 40 GH1 cells having a 79% growth fraction, whereas the growth fraction of passage level 100 cells was 95%. Using the population doubling time as a criterion for a choice of medium, F-10 medium supplemented with 20% fetal bovine serum consistently yielded the most rapid doubling time (32 hr), whereas Dulbecco's MEM supplemented with 15% horse serum and 2.5% fetal bovine serum yielded the greatest plateau cell density. Growth hormone secretion and the population doubling times were directly related to culture conditions including length of time in culture, choice of tissue culture media, choice of sera, and the phase of cell growth (lag, exponential or plateau).     

Cell Growth and Division: III. Conditions for Balanced Exponential Growth in a Mathematical Model

In a previous paper, we proposed a model in which the volume growth rate and probability of division of a cell were assumed to be determined by the cell's age and volume. Some further mathematical implications of the model are here explored. In particular we seek properties of the growth and division functions which are required for the balanced exponential growth of a cell population. Integral equations are derived which relate the distribution of birth volumes in successive generations and in which the existence of balanced exponential growth can be treated as an eigenvalue problem. The special case in which all cells divide at the same age is treated in some detail and conditions are derived for the existence of a balanced exponential solution and for its stability or instability. The special case of growth rate proportional to cell volume is seen to have neutral stability. More generally when the division probability depends on age only and growth rate is proportional to cell volume, there is no possibility of balanced exponential growth. Some comparisons are made with experimental results. It is noted that the model permits the appearance of differentiated cells. A generalization of the model is formulated in which cells may be described by many state variables instead of just age and volume.     

Alternative modes of population growth inhibition in a human lymphoid cell line growing in suspension

Using a human lymphoid cell line grown under continuous culture conditions, two distinct plateau states were induced, either by lack of sufficient medium-supplied nutrient, or by other unknown mechanisms dependent on cell density. Flow microfluorometric measurements show that growth arrest due to nutritional insufficiency results in an accumulation of cells with G1 DNA content. In contrast, growth arrest due to high cell density is not associated with an altered distribution of cells with respect to DNA content as the population progresses from exponential to plateau state growth. Cell size decreases with progression of the plateau state induced by either type of growth arrest. Cells in a plateau state induced by high cell density utilize glucose and incorporate exogenous amino acid into protein at approximately the same rate as exponential cells. Proliferating, high cell density, plateau state cells have cell cycle phase durations similar to exponential cells. The stable, plateau state cell density is maintained by cell loss. No stable, unbound growth inhibitory factor was found in the medium of density-inhibited plateau state cultures.     

Quantitation of bacillus subtilis L-form growth parameters in batch culture

Several methods were used to monitor the growth of a stable L-form in batch culture. The end of the exponential growth phase was determined with greatest accuracy by the amounts of deoxyribonucleic acid per milliliter of culture. Optical density and viable count data were not as reliable because the L-forms began to lyse at the end of exponential growth. Lysis was detected visually, by phase-contrast observations of wet mounts, and by release of ultraviolet-absorbing material into culture supernatant fluids.     

DNA synthesis rate and unlabeled cells with S-phase DNA content in P388 leukemic cells in vivo studied by flow cytometry and cell sorting

DNA synthesis kinetics of P388 leukemic cells growing in ascites form in BDF1 hybrid mice were investigated during the periods of exponential growth and growth restriction. Incorporation of tritiated thymidine, and in some instances tritiated uridine, was studied by autoradiography in cells sorted from S-phase fractions during DNA flow cytometry. During exponential growth continuous labeling with tritiated thymidine indicated a growth fraction of unity, whereas the growth fraction was about 30% during growth restriction. At this growth phase the majority of cells with S phase DNA content remained unlabeled after pulse labeling with tritiated thymidine or uridine, indicating that both the "salvage" and the "de novo" DNA synthesis pathways were blocked in most S-phase cells. After pulse labeling with tritiated thymidine the DNA synthesis rate pattern was investigated by sorting of consecutive fractions of cells throughout the S phase followed by quantitative autoradiography. With exception of a reduced rate in the middle of S phase, the DNA synthesis rate increased as the cells progressed through S phase during exponential growth. In contrast, the DNA synthesis rate pattern had a relative peak in the middle of S phase during growth restriction, which is otherwise characterized by a low mean DNA synthesis rate.     

Fungal melanin inhibitor and related compounds from Penicillium decumbens

In Silene vulgaris (M.) G. cell culture three growth phases were distinguished, namely, a lag phase, an exponential phase and a stationary phase. Pectin termed silenan and an acidic arabinogalactan were isolated as cell wall polysaccharides of S. vulgaris callus at the different growth phases during culture. Production of silenan as the galacturonan (or rhamnogalacturonan) core was observed at the beginning of the exponential phase and at the stationary phase of the callus growth. Arabinogalactan, containing the galacturonic acid residues, is formed at the exponential phase followed by attachment to the core of silenan in the middle of the exponential phase. The arabinogalactan constituent of silenan appeared to be destroyed gradually at the stationary growth phase. The monosaccharide compositions of silenan and arabinogalactan were determined at various phases of the callus growth. Silenan was found to be formed in maximum amounts at the exponential phase of the cell growth. Insignificant alterations of the yields of acidic arabinogalactan were found during culture while total productivity per litre of medium and rate of production per day of arabinogalactan were found to be maximal at the exponential phase of growth.     

5-Oxo-prolinase activity in tobacco suspension cultures: Regulation by sulfate nutrition

In heterotrophic and photoheterotrophic tobacco ( Nicotiana tabacum L., var. Samsun) suspensions cultured with growth-limiting amounts of sulfate, 5-oxo-prolinase activity declines at the same time as the growth rate of the cells decreases. However, 5-oxo-prolinase activity is reduced to a greater extent than growth. As a result, the specific activity of 5-oxo-prolinase also declines when sulfur is scarce. The decrease in both growth and 5-oxo-prolinase activity can be prevented by adding sulfate to the suspensions during exponential growth. Addition of sulfate after the exponential growth phase restored neither growth nor 5-oxo-prolinase activity. These observations show that 5-oxo-prolinase activity in tobacco cells is regulated by the sulfate supply in the medium. Such a regulation is an essential prerequisite, but not a proof, for a role of 5-oxo-prolinase as the rate-limiting factor in glutathione degradation.
During exponential growth the average specific activity of 5-oxo-prolinase in heterotrophic tobacco cells is twice as high as in photoheterotrophic cells. This difference is consistent with the idea that green cells are equipped for glutathione synthesis and export, and chloroplast-free cells for uptake and degradation of this peptide.     

Cytotoxin production by a merineLyngbya strain (cyanobacterium) in a large-scale laboratory bioreactor

A cytotoxic compound was produced by the marine cyanobacteriumLyngbya sp. Pearl strain in large laboratory-scale batch cultures. Adsorption and fractionation of methanol extracts with reverse phase (C-18) cartridges provided a rapid method for removal of bioassay interference from salts, biopolymers and pigments and concentration of the cytotoxic principles. Cytotoxicity to the murine leukemia cell line P-388 was produced in two cycles coinciding with the initiation of exponential growth and again during the late exponential growth phase. Antiviral activity against influenza virus PR8 was found in extracts prepared from early exponential growth phase cells but antiviral activity was not detected in extracts of mid-log or late-log growth phase cells. These differences in bioactivity suggests that the cytotoxic principles produced during early and late exponential growth may be different compounds. Cytotoxicity assays using murine P-388 leukemia indicates that the semi-pure compound has an IC₅₀ of < 0.25 μg ml⁻¹ to this cell line. P-388 cytotoxicity in cell extracts increased during the late exponential growth phase and the specific yield was estimated at approximately 0.14 mg g⁻¹ (dry cells).     

Inference of Super-exponential Human Population Growth via Efficient Computation of the Site Frequency Spectrum for Generalized Models

The site frequency spectrum (SFS) and other genetic summary statistics are at the heart of many population genetic studies. Previous studies have shown that human populations have undergone a recent epoch of fast growth in effective population size. These studies assumed that growth is exponential, and the ensuing models leave an excess amount of extremely rare variants. This suggests that human populations might have experienced a recent growth with speed faster than exponential. Recent studies have introduced a generalized growth model where the growth speed can be faster or slower than exponential. However, only simulation approaches were available for obtaining summary statistics under such generalized models. In this study, we provide expressions to accurately and efficiently evaluate the SFS and other summary statistics under generalized models, which we further implement in a publicly available software. Investigating the power to infer deviation of growth from being exponential, we observed that adequate sample sizes facilitate accurate inference; e.g., a sample of 3000 individuals with the amount of data expected from exome sequencing allows observing and accurately estimating growth with speed deviating by ≥10% from that of exponential. Applying our inference framework to data from the NHLBI Exome Sequencing Project, we found that a model with a generalized growth epoch fits the observed SFS significantly better than the equivalent model with exponential growth (P-value = 3.85 × 10^?6). The estimated growth speed significantly deviates from exponential (P-value  ? 10^?12), with the best-fit estimate being of growth speed 12% faster than exponential.     

Changes in superoxide dismutase activity and photosynthetic pigment content during growth of marine phytoplankters in batch-cultures

The ability of phytoplankton to cope with oxidative stress is one of the main factors that influence its survival in the marine environment, when senescence conditions prevail. In a first attempt to investigate the antioxidant strategies of different phytoplanktonic groups face to oxidative stress, the superoxide dismutase (SOD; EC 1.15.1.1) activity and photosynthetic pigment content along the growth curves of the dinoflagellate Lingulodinium polyedrum (Stein) Dodge, the prasinophycean Tetraselmis gracilis (Kylin) Butcher and the diatom Minutocellus polymorphus (Hargraves and Guillard) Hasle, von Stosch and Syvertsen were evaluated in batch-cultures. Total SOD activity was determined by an indirect method involving the inhibition of cytochrome c reduction. The contents of photosynthetic pigments were analysed by HPLC using a reverse phase column (RP-18), based on a ternary gradient. A peak of total SOD activity was detected at the beginning of the T. gracilis and M. polymorphus exponential growth. In L. polyedrum and M. polymorphus, SOD activity increased approximately three times by day 17 of growth, compared to the values obtained on day 3 (exponential phase) of the growth curve. All three species of microalgae had reduced SOD activity at the end of their growth. The levels of peridinin in L. polyedrum increased about 60% by day 17 of growth compared to the values obtained at exponential phase. Tetraselmis gracilis exhibited a remarkable increase (approximately 85%) in beta-carotene concentration after 10-14 days of growth whereas the beta-carotene levels in M. polymorphus decreased about 85% along its growth curve. These findings suggest that the antioxidant response during senescence in batch-cultures differ according to the species. Induction of SOD activity may occur either in the early exponential or stationary growth phases, which is important to prevent oxidative stress triggered by a number of factors that affects growth, such as nutrient and light availability.     

Regulation of catalase synthesis in Salmonella typhimurium.

The specific activity of catalase in Salmonella typhimurium and other enteric bacteria decreased during the logarithmic phase of growth and increased at the onset and during the stationary phase. The increase in catalase synthesis at the end of the exponential phase in S. typhimurium cells coincided with the lowest pH value reached by the culture. Maintenance of the pH at a constant neutral value did not alter the typical pattern of synthesis in contradiction of the results previously reported (McCarthy and Hinshelwood. 1959). A sudden decrease in the pH value of an S. typhimurium culture during exponential growth by addition of HC1 did not cause an alteration in the catalase synthesis pattern. Addition of hydrogen peroxide to S. typhimurium cultures within the range 1 muM TO 2MM during the exponential growth phase stimulated catalase synthesis. The extent of catalase synthesis depended on the concentration of hydrogen peroxide; the maximum stimulation was observed at 80 muM. Increased catalase synthesis was not detected for 10 to 15 min after hydrogen peroxide addition. Hydrogen peroxide was produced by S. typhimurium cultures during the exponential and stationary growth phases. However, no direct relationship between hydrogen peroxide accumulation and synthesis of catalase was observed.     

A nonlinear mathematical model of cell turnover, differentiation and tumorigenesis in the intestinal crypt

We present a development of a model [Tomlinson, I.P.M., Bodmer, W.F., 1995. Failure of programmed cell death and differentiation as causes of tumors: Some simple mathematical models. Proc. Natl. Acad. Sci. USA 92, 11130-11134.] of the relationship between cells in three compartments of the intestinal crypt: stem cells, semi-differentiated cells and fully differentiated cells. Stem and semi-differentiated cells may divide to self-renew, undergo programmed death or progress to semi-differentiated and fully differentiated cells, respectively. The probabilities of each of these events provide the most important parameters of the model. Fully differentiated cells do not divide, but a proportion undergoes programmed death in each generation. Our previous models showed that failure of programmed death--for example, in tumorigenesis--could lead either to exponential growth in cell numbers or to growth to some plateau. Our new models incorporate plausible fluctuation in the parameters of the model and introduce nonlinearity by assuming that the parameters depend on the numbers of cells in each state of differentiation. We present detailed analysis of the equilibrium conditions for various forms of these models and, where appropriate, simulate the changes in cell numbers. We find that the model is characterized by bifurcation between increase in cell numbers to stable equilibrium or explosive exponential growth; in a restricted number of cases, there may be multiple stable equilibria. Fluctuation in cell numbers undergoing programmed death, for example caused by tissue damage, generally makes exponential growth more likely, as long as the size of the fluctuation exceeds a certain critical value for a sufficiently long period of time. In most cases, once exponential growth has started, this process is irreversible. In some circumstances, exponential growth is preceded by a long plateau phase, of variable duration, mimicking equilibrium: thus apparently self-limiting lesions may not be so in practice and the duration of growth of a tumor may be impossible to predict on the basis of its size.