Early Logarithmic Phase Research Articles

Establishment of cell lines in vitro from a mammary ascites tumor of mouse and biological properties of the established lines in a serum containing medium.

1) Two cell lines were established in vitro from a mouse ascites mammary carcinoma FM3A and they were similar to each other both in their in vitro growth rate and their morphological identity with in vivo tumor cells. 2) Both A and B lines had three characteristic chromosomes and high transplantability, and the transplanted subcutaneous tumors showed the morphology of undifferentiated adenocarcinoma. 3) The cloning of the present cell lines were made with the use of agar. 4) The study on the nutritional requirement of the present lines showed that Bactopepton, pyruvate and serine were growth enhancing respectively and the combination of these additives were more effective than single addition. 5) The glucose utilization of the lines in the course of the growth was enhanced at the growth stage of the lag phase and early logarithmic phase but decreased after the stage of late logarithmic phase. 6) The capacity to grow in vitro without lag time could be maintained for a pretty long time after transplantion to the host animal, but it fell gradually.

Studies on the Proteolytic Enzymes of Thermophilic Streptomyces Part II

Taxonomical characteristics of a thermophilic streptomycete which had been reported to produce a relatively thermostable proteinase were examined, and it was found to belong to a variety of Streptomyces rectus for which the name Streptomyces rectus var. proteolyticus was suggested. Some cultural conditions for proteinase formation by this strain were investigated. The formation of proteinase was greatly stimulated by the addition of Mn and affected by the composition of medium. The proteinase formation occurred during the stationary phase of growth at 50°C, while intracellular peptidase was formed in early logarithmic phase. No presence of precursor of the proteinase was suggested.

DIFFERENTIATION BETWEEN RADIATION SENSITIVE AND -RESISTANT MUTANTS OF ESCHERICHIA COLI, STRAIN B.

Morphological heterogeneity in cell sizes of Escherichia coli, strains B, B/r, and BSprevents estimation of the viable cell count by the indirect turbidimetric method. The average dry weights of E. coli B, B/r, and BS, when grown in acetate synthetic medium, are 0.16, 0.10, and 0.13 picograms per cell respectively. Increased radiation sensitivity was correlated with those bacterial populations showing decreased intracellular glycogen content, decreased lipopolysaccharide–lipoprotein content, R-colony forms, and cellular elongation. The radiation-sensitive strains contained more DNA, RNA, and protein per cell than the radiation-resistant mutant; however, no significant differences were evident in the DNA, RNA, and protein contents on a dry weight basis. No qualitative differences in the monosaccharides or their derivatives were found. Intracellular glucose synthesis and accumulation preceded DNA, RNA, and protein synthesis throughout the lag and early logarithmic growth phases for all three strains. Multiple forms of intracellular glycogen complexes were present in all three strains. In addition to unbound glycogen and protein-bound glycogen, a bound form of glycogen was consistently associated with crude fractions which contained the DNA, RNA, and lipopolysaccharide constituents. Increased lability of the cold TCA-insoluble bound glycogen to hot TCA and alpha-amylase hydrolysis was correlated with increased radiosensitivities of the organisms.

NUCLEASES OF MYCOPLASMA.

SUMMARY: Nuclease activity was observed in several saprophytic and parasitic Mycoplasma organisms; the nucleases of Mycoplasma laidlawii were studied in detail. Nuclease activity of this organism was highest at the early logarithmic phase of growth, and was found mainly the soluble fraction of the organisms. Anion-exchange chromatography of the proteins of the soluble fraction separated ribonuclease from deoxyribonuclease activity. Each enzyme had an alkaline pH optimum, required magnesium ions for activity, and degraded native and heat-denatured nucleic acids. M. laidlawii RNase degraded RNA-core. RNA inhibited the degradation of DNA by M. laidlawii deoxyribonuclease. The implications of these findings with respect to the effects of RNA and DNA on growth of M. laidlawii are discussed.

The mode of action of certain 3- and 5-nitropyridines and pyrimidines. III. Biochemical lesions in Trichomonas vaginalis

Some 5-nitropyridines and pyrimidines inhibit the growth of Trichomonas vaginalis in vitro and in vivo. They are detoxified by reduction to their corresponding amino analogs. The chemical 2-amino-5-nitropyrimidine has been used as a model compound to study the inhibitory pathway. During growth with inhibitory concentrations of 2-amino-5-nitropyrimidine the synthesis of DNA, RNA and protein paralleled multiplication. Polysaccharide accumulated in cells grown with inhibitor when maltose was used as the carbon source, but not when glucose was used. The organism normally stores carbohydrate as glycogen. The polysaccharide that accumulated in inhibited cells was shown by quantitative iodine staining to be less highly branched than glycogen. Addition of inhibitor to resting cells or cell-free extracts did not affect metabolism. Consequently, cells were grown for varying periods of time with inhibitor, and the metabolism of intact cells or dialyzed cell-free extracts compared to similar preparations of control cells grown simultaneously. Up to 2 hours growth with inhibitor did not affect gas exchange as measured with a Warburg apparatus, or dehydrogenation reactions as measured by following the reduction of triphenyl-tetrazolium chloride. Cells up to 2 hours of age were less active metabolically than older cells. After 3 hours growth with inhibitor, aerobic respiration and endogenous anaerobic fermentation remained intact, but anaerobic utilization of carbon sources was inhibited as were dehydrogenation reactions. It was found that the internal environment of inhibited cells was more aerobic than control cells. As compared to preparations from control cells, enzyme extracts of cells grown for 4 hours with inhibitor contained more glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase; the same amounts of aldolase and “malic enzyme”; and less malic dehydrogenase. The data have been interpreted as indicating that there is some sort of developmental cycle during the lag and early logarithmic phases of growth of T. vaginalis and that the inhibitor, by acting on an energy yielding mechanism, kills the cells by holding them in this early pattern. Growth with inhibitor did not affect the purine and pyrimidine base composition of RNA and DNA. For DNA, using cytosine as 1, these were cytosine:guanine:adenine: thymine as 1:.95:2.1:2.0. For RNA, they were cytosine:guanine:adenine:uracil as 1:1.2:2.6:2.2. The total cellular RNA of this organism is unique since its base composition closely mimics that of its DNA.

Cation transport in Escherichia coli. I. Intracellular Na and K concentrations and net cation movement.

Methods have been developed to study the intracellular Na and K concentrations in E. coli, strain K-12. These intracellular cation concentrations have been shown to be functions of the extracellular cation concentrations and the age of the bacterial culture. During the early logarithmic phase of growth, the intracellular K concentration greatly exceeds that of the external medium, whereas the intracellular Na concentration is lower than that of the growth medium. As the age of the culture increases, the intracellular K concentration falls and the intracellular Na concentration rises, changes which are related to the fall in the pH of the medium and to the accumulation of the products of bacterial metabolism. When stationary phase cells, which are rich in Na and poor in K, are resuspended in fresh growth medium, there is a rapid reaccumulation of K and extrusion of Na. These processes represent oppositely directed net ion movements against concentration gradients, and have been shown to be dependent upon the presence of an intact metabolic energy supply.

The Division Index and Multiplication in a Mass Culture of Tetrahymena Following Inoculation*

SUMMARY. Test tube cultures of Tetrahymena pyriformis GL in the early maximum stationary phase of growth were used as inocula. After the elapse of a short lag phase a rapid transition to the logarithmic phase of growth was observed in cultures grown on peptone media(4,6). During the early phase of exponential multiplication more cells (as expressed on a percentual basis of the population number) were in the visible stage of cytoplasmic fission than after the elapse of 5 to 6 generations. Analysis of comprehensive data suggests that the higher division index in the early logarithmic phase of growth is not the expression of a synchronization of cells in metabolic respects but rather indicates a prolongation of the stage of cell fission during this phase.

Studies on the nature of hemophilus influenzae cells susceptible to heritable changes by desoxyribonucleic acids.

In E. influenzae the highly specific desoxyribonucleic acids (DNA's) which play the role of heredity determinants of type specificity and SM resistance, have induced these traits in only a small proportion of the population exposed to their action. The evidence suggests that this small proportion, "the susceptible cells" possess a property or substance needed by the DNA in order to induce an heritable change. The size of the small proportion of susceptible cells can be influenced significantly by a number of factors; when all the factors now to be listed are operating the frequency has not exceeded 1:1000. The Type of Origin of Recipient Cells.-Type a exhibits the lowest frequency, about 1:10,000,000, and type d the highest, approximately 1 per 1000 cells exposed. This type-specific property which controls the frequency of susceptible cells is an inherited trait; repeated change to a heterologous type shows no influence on the incidence of these cells. Concentration of DNA.-Within certain limits increase in the concentration of DNA controlling streptomycin resistance can increase the size of the proportion of cells in which streptomycin resistance can be induced. However, increases in concentrations greater than 10(-1) microg. per ml. have not induced streptomycin resistance in a higher proportion of cells. Phase of Growth Cycle.-Predictable fluctuations in frequency of induced heritable changes have been demonstrated in both Rb and Rd populations during growth. There is no evidence that susceptible cells reproduce their kind; they emerge in all experiments when the population reaches the end of the logarithmic period and a density of 2 to 4 x 10(8) cells per ml. In the early logarithmic phase it is difficult to demonstrate the presence of susceptible cells. The peak frequency of susceptible cells occurs in the early stationary phase of the growth cycle. Thereafter, the decline in frequency is a gradual one. The data suggest that in a given population the same cells are susceptible to a number of different type-specific DNA's and the DNA controlling SM resistance. Comparison of Frequency of Cells Susceptible to Different DNA's.-In a given population the frequency of cells susceptible to different type-specific DNA's and the DNA controlling streptomycin resistance is not significantly different. Competition between Type-Specific DNA's-The data suggest that DNA's of types a, b, and c compete for the same cells in Rd populations. When Rd populations are exposed simultaneously to 2 of these 3 DNA's in different concentrations the proportion which each type contributes to the total type-specific cells induced is closely correlated with the concentration of the corresponding DNA. Exclusion of DNA's.-Induction of one type specificity or streptomycin resistance can be completely prevented in a population containing susceptible cells by previous exposure for 15 minutes to a 1000-fold higher concentration of another type-specific DNA.

Some recent advances in the bacteriology of pasteurized milk

AbstractThe keeping quality of pasteurized milk is influenced to a considerable degree by the atmospheric temperature on the day of collection of the raw milk from the farms, a marked deterioration being recorded at collection temperatures of over 21° C. (70° F.). This deterioration is apparent even when the raw milk collected at different atmospheric temperatures has apparently the same bacterial activity, as measured by dye reduction, on delivery to the dairy. It is possible that an increase in heat resistance of certain acid‐producing organisms during the early logarithmic phase of growth may account for this phenomenon.Raw milk produced under satisfactory hygienic conditions has a satisfactory keeping quality after pasteurization, but the addition of 1% of an unsatisfactory raw supply to the bulk before pasteurization will cause marked deterioration.In mild weather the infection of pasteurized milk with B. Coli at any point in its travel results in a marked loss of keeping quality, but in hot weather this loss is masked by the lower natural keeping quality of the bulk milk as revealed by laboratory pasteurization.