Early Logarithmic Phase Research Articles

Ecological Application of Antibiotics as Respiratory Inhibitors of Bacterial Populations1

Two terregenous and four marine bacterial isolates were treated with six antibiotics and antibiotic combinations. Comparisons made between responses of cells in early and late logarithmic and stationary growth phases indicated variable sensitivity to the agents. Bacteria in stationary and late log-phase cultures exhibited the greatest resistance, whereas the early log-phase cells exhibited greatest antibiotic susceptibility. We conclude that the tested antibiotics cannot be used for ecological purposes to delineate bacterial respiration in mixed microbial communities.

Role of zinc in the production of Clostridium perfringens alpha toxin.

Clostridium perfringens was found to produce alpha toxin in a synthetic medium containing zinc; in medium containing no zinc, a little toxin was detected in the early logarithmic phase of growth and it disappeared rapidly. No intracellular accumulation of alpha toxin protein occurred whether or not zinc was present in the medium. In zinc-deficient medium, the organisms produced and released into the surrounding medium the protein specifically precipitable with alpha antitoxin in an amount comparable to that of alpha toxin produced in the zinc-containing medium. The protein combined rapidly in some unknown way with zinc to form the active and stable alpha toxin.

The release of alkaline phosphatase and of lipopolysaccharide during the growth of rough and smooth strains of Salmonella typhimurium.

A rough strain of Salmonella typhimurium, which has a defect in the "inner core" region of its lipopolysaccharide, was shown to release more alkaline phosphatase into the medium during growth than did an isogenic smooth strain of the same organism. The pattern of enzyme release varied during growth in that early logarithmic phase cells retained alkaline phosphatase in their periplasmic space and at their cell surface while cells in later growth phases shed as much as 6% of their content of this enzyme into the medium. The reaction product localization technique used in this study enables us to assess enzyme production and localization in individual cells of a culture. The release of the enzyme was paralleled by a release of lipopolysaccharide from the cells. The released alkaline phosphatase shows a strong tendency to associate with purified phenol-extracted lipopolysaccharide.

Biotin deficiency in Arthrobactger globiformis: comparative cell ultrastructure and nonreplacement of the vitamin by structurally unrelated compounds.

The development of aberrant cell forms of Arthrobacter globiformis 425 due to biotin deficiency was followed by electron microscopy of ultrathin sections. Upon comparison with normal cell growth, aberrant cells developed in the early logarithmic growth phase. Several membrane-bound bodies were embedded in a thick matrix, showing that cell division was impaired. Mesosomes and cytoplasmic membranes were still present in the abnormal cell although the normal cell wall was absent. This condition persisted throughout the growth cycle. This pattern of morphological development was correlated with changes in macromolecular composition of the cells. Various structurally unrelated compounds were tested for their ability to replace biotin. These included aspartic acid, oxalacetic acid, coenzyme A, oleic acid, linoleic acid, linolenic acid, and Tween 80. Only Tween 80 was able to spare biotin to a limited degree. However, this sparing action was eliminated in the presence of avidin.

Characterization and Synthesis of Anacystis Ribonucleic Acids

Photoautotroph cultures of the blue-green alga Anacystis nidulans show a pronounced maximum of 10 −7 µ g total RNA/cell in the early logarithmic growth phase and a subsequent 50% decrease during the stationary phase. The alteration of the total RNA content influences all RNA fractions in the same way. Two high-molecular weight ribosomal RNAs of 1.1 and 0.56 × 10 6 daltons, respectively, are obtained if Mg 2+ (5–10 mM) is present during the RNA extraction. Absence of Mg 2+ during the RNA extraction at 0° or heating the isolated total RNA for 1 min at 60°C in the presence of Mg 2+ results in an irreversible conversion of the 1.1 × 10 6 rRNA to 0.9 plus 0.2 × 10 6 rRNA. The 0.56 × 10 6 rRNA is found to be a very stable molecule After less than 1 min pulse-labelling of A. nidulans cells with Na 2 H 32 PO 4 and separation of the RNA fractions by MAK chromatography the radioactivity is eluted with the first part of the DNA fraction and in front of the small ribosomal RNA. Extension of the pulse-labelling to a few minutes yields radioactivity in all MAK fractions. In pulse-chase experiments the small rRNA is shown to have the most rapid turnover compared to the other RNA fractions. DNA-RNA hybridization studies demonstrate that all 3 rRNA fractions contain rapidly-labelled RNA of the non-ribosomal type, however, as competition experiments indicate, in different proportions to the amount of ribosomal precursor RNAs. Thus, the 0.9 × 10 6 fraction does not contain rapidly labelled ribosomal RNA.

Effect of Tetrathionate on the Growth of a Marine Thiobacillus

SUMMARY: Thiobacillus sp. strain IV-85, an isolate from the marine environment, oxidized thiosulphate in two sequential logarithmic phases. Tetrathionate accumulated in the partially spent broth during the first logarithmic phase, and reached a maximum concentration at the time of transition to the second logarithmic phase. Tetrathionate added during the early first logarithmic phase caused a decrease in the rate of thiosulphate utilization similar to that observed after the transition from the first to second logarithmic phase. Bacteria harvested from the first logarithmic phase resisted 0.1 mM-cyanide which inhibited oxygen uptake by bacteria harvested from the second logarithmic phase. Rhodanese (thiosulphate: cyanide sulphur transferase, EC. 2.8.1.1) was detected in extracts of bacteria from both phases. Tetrathionate accumulation in the growth medium inhibited rhodanese activity, resulting in the transition of thiosulphate utilization from the first to second logarithmic phase.

Correlation of M protein production with those factors found to influence growth and substrate utilization of Streptococcus pyogenes.

In the absence of proteinase formation, factors reported to influence the growth or fermentation by streptococci have been evaluated to determine their quantitative effect upon the production of M protein during the growth of Streptococcus pyogenes. Buffers, amino acids, peptides, gross organic additions, and carbohydrate substrates were tested under a variety of cultural conditions. The M protein content was remarkably constant throughout the late logarithmic period of growth, i.e., when the cell population doubled, the M protein doubled. However, several factors affected the M protein content per milligram of cells (dry weight). When types 1, 12, and 22 were grown aerobically in a semidefined medium, the M protein content of the cell population essentially doubled; in Todd-Hewitt broth, this aerobic effect on M protein synthesis was not observed. When cells grown on Todd-Hewitt broth were transferred to medium containing 0.1% starch and no added glucose, the M protein content per milligram of cells (dry weight) increased as much as fourfold. When growth was initiated in glucose, the rate of M protein formation was at a maximum in the early logarithmic phase of growth and was comparatively greater than the rate of cellular multiplication. When the amount of substrate fermented was greater than 0.2%, increased M protein was not observed. An evaluation of the effects of medium or conditions of growth showed the units of M per milligram of cells (dry weight) were not influenced by a shift in the stoichiometry of either the anaerobic or aerobic fermentation, substrate used, or adenosine triphosphate utilized for growth. These results show that M protein synthesis is subject to limited glucose repression or substrate catabolite repression.

ハマチ類結節症の原因細菌の分類学的研究

Morphological and biochemical characteristics were examined on four strains of the bacteria causative of “pseudotuberculosis” among yellowtail (Seriola quinqueradiata), and on a strain of Pasteurella piscicida, Janssen et Surgalla. The observation that the pseudotuberculosis organisms were identical with P. piscicida was confirmed. These organisms were all Gram-intermediate, non-motile rods. A peculiar morphological feature of the organisms is the transformation from rod-shaped cells in the early logarithmic growth phase to small coccoids or cocci in the stationary phase. The organisms gave positive Kovacs' oxidase reaction; fermented glucose in Hugh-Leifson's medium, and produced acid but no gas from a few other carbohydrates such as fructose, galactose, and mannose. They were sentistive to most of the antibiotics and antimicrobial drugs, including penicillin and polymyxin B; were halophilic; and did not grow at pH lower than 6.1. Although the cells of these organisms did not show any typical coryneform appearance, nevertheless, the above characters suggest that they have a close relationship with coryneform bacteria and genus Arthrobacter seemed to be the most suitable genus to which they belong.

Application of Enzyme Production Properties in Subtyping of Group A Streptococci According to T Type

The production of extracellular nicotinamide adenine dinucleotide glycohydrolase (NADG) and the cell-bound lipoproteinase (serum opacity reaction, SOR) by strains of different serological types of group A streptococci, in relation to the T typing, was studied. The production of both NADG and SOR, or only one of them, was found to be characteristic of serotypes, as determined by M and T antigen. No difference in the production of these enzymes was found in relation to M-positive and M-negative variants. Investigation into NADG and SOR production as related to the T type enabled the division of a single agglutination pattern into four main groups, each of which corresponds to one specific M type or more. Of the 370 strains belonging to 12 different T-agglutination patterns, 21% produced both enzymes and 42.5% failed to produce any of them, whereas the remaining 36.5% produced only one out of the two enzymes. Five streptococcal types which did not produce NADG and SOR also failed to synthesize streptolysin S at the early logarithmic phase of growth, indicating that streptolysin S production by young cultures may be also related to serotype. No correlation was found between the production of NADG-SOR as related to serotype and the production of streptolysin O, acid phosphotase, esterase, N -acetylglucosaminidase, hyaluronidase, streptokinase, and the cell-sensitizing factor. The practical and potential usefulness of NADG and SOR production in epidemiological studies is discussed.

Application of Enzyme Production Properties in Subtyping of Group A Streptococci According to T Type

The production of extracellular nicotinamide adenine dinucleotide glycohydrolase (NADG) and the cell-bound lipoproteinase (serum opacity reaction, SOR) by strains of different serological types of group A streptococci, in relation to the T typing, was studied. The production of both NADG and SOR, or only one of them, was found to be characteristic of serotypes, as determined by M and T antigen. No difference in the production of these enzymes was found in relation to M-positive and M-negative variants. Investigation into NADG and SOR production as related to the T type enabled the division of a single agglutination pattern into four main groups, each of which corresponds to one specific M type or more. Of the 370 strains belonging to 12 different T-agglutination patterns, 21% produced both enzymes and 42.5% failed to produce any of them, whereas the remaining 36.5% produced only one out of the two enzymes. Five streptococcal types which did not produce NADG and SOR also failed to synthesize streptolysin S at the early logarithmic phase of growth, indicating that streptolysin S production by young cultures may be also related to serotype. No correlation was found between the production of NADG-SOR as related to serotype and the production of streptolysin O, acid phosphotase, esterase, N-acetylglucosaminidase, hyaluronidase, streptokinase, and the cell-sensitizing factor. The practical and potential usefulness of NADG and SOR production in epidemiological studies is discussed.

Identification of the cytokinin-active ribonucleosides in pure Escherichia coli tRNA species.

The distribution of the two cytokinin-active ribonucleosides 6-(3-methyl-2-butenylamino)-9-beta-D-ribofuranosylpurine (1) and 6-(3-methyl-2-butenylamino)-2-methylthio-9-beta-D-ribofuranosylpurine (2) in tRNA(Phe), tRNA(UUG) (Leu), tRNA(UCR) (Ser), tRNA(Tyr), tRNA(Cys), and tRNA(2) (Trp) from stationary phase Escherichia coli has been studied. Compound 2 predominated in all these species except tRNA(Phe), in which compound 1 was greatly in excess. Compound 1 was present in relatively small amounts in tRNA(UUG) (Leu), tRNA(Cys), and tRNA(2) (Trp), and was not detected in tRNA(UCR) (Ser) and tRNA(Tyr). Cells from the early logarithmic phase had the highest cytokinin content in their tRNA. Stationary phase tRNA had a slight increase in the amount of 1, which may be explained by the appearance of the new tRNA(Phe). No differences were observed between the chromatographic patterns of tRNAs from early-, mid-, and latelogarithmic phases.

Electron transport and oxidative phosphorylation in the blue-green alga Anabaena variabilis.

It is shown that secretion of bacteriolytic complex enzymes in a bacterium of the family Pseudomonadaceae begins in the latent and early logarithmic culture growth phases. A maximum of specific bacteriolytic activity falls in the same time interval with a maximum of specific activity of lytic proteinase L2 and muramidase. The activity of concomitant enzymes--non-specific proteinase and neutral phosphatase--peaks in the middle of the logarithmic growth phase. Secretion of exopolysaccharide into the external medium begins in the second half of the logarithmic growth phase and, thus, is not coordinated with secretion of bacteriolytic enzymes. It is shown that relative content of enzymes and polysaccharides depends upon the composition of growth medium for the producer.

Intracytoplasmic membrane structures in Vibrio marinus.

An electron microscope study of Vibrio marinus strains MP-1, an obligate psychrophile, and PS-207, a moderate psychrophile, revealed numerous intracellular membranous structures. The structures were found to occur more frequently in V. marinus strain MP-1 than in strain PS-207. The frequency of occurrence and complexity of structure were related to age of the culture. In early logarithmic phase, cells revealed invaginations of the plasma membrane. More complex membrane forms, found in late logarithmic and stationary phase, were either myelin-like sheaths, for which the term "myelemma" is proposed, or membranes randomly arranged throughout the cells. The complex membrane forms were not observed to be directly connected with the plasma membrane. However, they were often found in approximation to the plasma membrane or associated with vacuoles and circular membrane profiles. Individual membranes were of a tripartite structure and of dimensions similar to the cell wall and plasma membrane.

Release of protoplasts in the yeast phase of Histoplasma capsulatum without added enzyme.

Cells of the yeast phase of the dimorphic systemic fungus pathogen Histoplasma capsulatum readily released large numbers of intact protoplasts without degradation of their cell walls by snail or microbial enzymes, previously regarded as a requirement for all yeast and mycelial fungal forms. Over 90 percent of "B" type yeast cells in the early logarithmic phase of growth released living protoplasts when incubated at 37 degrees C with 2 molar magnesium sulfate, whereas "A" type yeast cells required prior exposure for 24 hours to 2-deoxy-D-glucose before incubation in the 2 molar magnesium sulfate.

Aminoacetone formation and utilization by pseudomonads grown on DL-1-aminopropan-2-ol.

SUMMARY: Pseudomonas sp. 8/IX isolated from soil and Pseudomonas sp. NCIB 8858 are capable of growth on DL-I-aminopropan-2-ol or aminoacetone as sole sources of carbon, nitrogen and energy. During growth on the amino alcohol, small amounts of aminoacetone initially accumulate in the medium but disappear during the early logarithmic phase. Aminoacetone accumulation is favoured by high pH and high growth-substrate concentrations. Washed suspensions of the pseudomonads grown on DL-I-aminopropan-2-ol accumulate aminoacetone when incubated with the amino alcohol in the presence of metabolic inhibitors, and rapidly utilize aminoacetone in their absence. Aminoacetone formation is optimal at about pH 10 in the presence or absence of the most effective inhibitor iodoacetate. Aminoacetone utilization occurs optimally at pH 7·7, the apparent K m for the amino ketone being about 0·1 mM. At an initial concn. of 1·0 mM, the rate of utilization is proportional to cell density up to at least 0·5 mg. dry wt organisms/ml. and is constant with time over almost the entire course of the reaction. The maximum rate of utilization is approximately 150 mμmoles/mg. dry wt organisms/min. at 30˚. The phenyl analogue 2'-aminoacetophenone is utilized at a comparable rate but 5-aminolaevulate is virtually unaffected. Anaerobic conditions and metabolic inhibitors prevent amino ketone utilization. Growth conditions markedly affect the ability of suspensions both to accumulate and utilize aminoacetone. Of the growth substrates tested, only L-threonine enabled Pseudomonas sp. 8/IX to utilize aminoacetone, the rate being approx. 60 % of that found after growthon DL-I-aminopropan-2-ol The patterns of oxidation of possible intermediates of DL-I-aminopropan-2-ol catabolism are similar using washed organisms grown on either the amino alcohol or aminoacetone. Organisms grown on acetate plus glycine oxidize such compounds at relatively low rates. Methylglyoxal is not a substrate for either growth or oxidation. Comparison of the rates of oxygen uptake and aminoacetone utilization, and measurement of the total oxygen consumed, indicate about 70% complete oxidation of the amino ketone by organisms grown on the amino alcohol. Similar maximum crop-sizes are obtained in media in which ammonium sulphate or different relative amounts of D- and L-I-aminopropan-2-ol serve as the sole sources of growth-limiting nitrogen.

Growth and proteinase production inPseudomonasspp. cultivated under various conditions of temperature and nutrition

SummaryA study was made of the formation of the extracellular proteolytic enzymes during the growth cycle of several species ofPseudomonascultivated under different conditions of temperature and nutrition. Proteolytic activity was not proportional to growth. Expressed per unit of cell dry weight, the proteolytic activity showed a peak in the early logarithmic phase which was greater in cultures grown at 3 than at 28°C. Proteolytic enzyme was not formed in the absence of organic nitrogen. Of 16 organisms studied,Pseudomonas aeruginosaATCC 10145 was the most prolific producer of proteolytic enzyme.

ACID RESISTANCE AND INFECTIVITY OF FOOD-POISONING CLOSTRIDIUM PERFRINGENS

Cells of food-poisoning Clostridium perfringens were subjected to treatment with synthetic gastric fluid. The log10survival of the cells followed a descending convex curve, both with duration of treatment and with decreasing pH. Resistance was at a minimum during the early logarithmic phase and increased to a maximum at the end of the growth phase by a factor of 10,000. Cells at stages of relatively high resistance consistently and effectively produced the food-poisoning syndrome in human volunteers while cells at stages of low resistance produced only mild abdominal discomfort.The transient changes in resistance to simulated gastric fluid in the presence and absence of pepsin were basically the same, but pepsin and other proteins had a protective effect on the cells during acid treatment.

Lytic Action of β(1-3)-Glucanase on Yeast Cells

Candida utilis, Saccharomyces cerevisiae, S. fragilis, Pichia polymorpha, and Hansenula anomala yeast cells, harvested in the early logarithmic phase, were attacked with purified beta(1-3)-glucanase from Micromonospora chalcea, which resulted in the liberation of protoplasts. The treated cells were observed under the electron microscope before the protoplasts were liberated. Differences in the cell walls of the enzyme-treated and untreated cells were observed. The action of the glucanase was also tested against isolated walls of C. utilis. The enzyme attacked the S. cerevisiae cell wall in a uniform manner. The attack on S. fragilis was located in certain zones of the cell wall, where breakage occurred and through which the protoplast emerged. On the other three yeasts, an intermediate attack was observed, not as definitely located as in S. fragilis, yet less uniformly than in S. cerevisiae.

Biosynthesis of biotin in microorganisms: IV. Repression and derepression of (+)-biotin synthesis from (±)-desthiobiotin

The enzyme system which catalyzes the conversion of (±)-desthiobiotin to (+)-biotin in Escherichia coli is formed rapidly in a basal medium during the early logarithmic phase and is repressed during the later growth phases. The biotin synthesizing enzyme system is repressed when the medium is supplemented with (+)-biotin; complete repression occurs only when the biotin content of the medium is raised above the level found intracellularly in cultures of the organism upon cessation of growth. An excess amount of desthiobiotin or oxybiotin also causes repression to some extent. Repression by the exogenous biotin vitamers is relieved rapidly when avidin is added to the cultures. Homobiotin is even more effective in relieving the repression. The kinetics of derepression follow the pattern characteristic of biosynthetic enzymes that are repressible by their end products. These findings indicated that the level of the enzyme system varied according to the demand for (+)-biotin by the organism, and supported the role of this enzymatic reaction as the key step in the physiological control of biotin synthesis.

Growth phases of Mycoplasma in liquid media observed with phase-contrast microscope.

Razin, Shmuel (University of Connecticut, Storrs), and Benjamin J. Cosenza. Growth phases of Mycoplasma in liquid media observed with phase-contrast microscope. J. Bacteriol. 91:858-869. 1966-Growth of 11 Mycoplasma strains in liquid media was followed by phase-contrast microscopy. A similar pattern of development was common to all strains. Branching filaments, 0.3 to 0.4 mu thick, characterized the early logarithmic phase of growth. The length of the filaments varied according to the strain tested and the growth medium. Addition of oleic acid to the medium induced the formation of very long filaments by M. laidlawii strain B. Upon aging, the filaments were found to break up into chains of coccoid elements. These chains further fragmented to yield shorter chains and single coccoid elements, which characterized the stationary and decline phases of growth. The size of the coccoid elements increased from 0.3 to 0.4 mu, when formed in the filaments, to 0.6 to 0.8 mu after being released from the chains. Further increase in the size of the cells took place at the decline phase of growth, leading to the formation of very large cells reaching a diameter of 10 to 20 mu. However, these large cells had the appearance of empty vesicles and were apparently nonviable as indicated by viable-count experiments.