bluuebunny/crossref_metadata_2025 huggingface dataset:
Inducible Synthesis of Bacterial Luciferase: Specificity and Kinetics of Induction
Authors: Coffey John J.
Published: January 2020

A nitrate-utilizing strain of marine bacteria was isolated in which luciferase was inducible by
l
-arginine. The induction was highly specific; structural analogues of arginine were ineffective, as were other natural amino acids. Several metabolites of arginine acted as weak inducers but did not affect the rate of induction in limiting concentrations of arginine. Hence, these compounds probably exerted a sparing effect on intracellular arginine. The kinetics of induction were followed by measurement of light output from intact cells, under conditions in which in vivo light output was determined only by the luciferase level. No enzyme appeared in the cells for 12 min after the addition of inducer, although the primary structure of the luciferase molecule was apparently synthesized within 2 to 4 min. It is proposed that during the remaining 8 to 10 min a precursor of luciferase was converted to active enzyme. The differential rate of synthesis rose during the first 5 min of induction, apparently as messenger ribonucleic acid accumulates in the cells; thereafter it remained constant for approximately 100 min.
Conjugation in
Escherichia coli:
Recombination Events in Terminal Regions of Transferred Deoxyribonucleic Acid
Authors: Pittard J. , Walker E. M.
Published: January 2020

An investigation of recombination events occurring in zygotes formed during conjugation has been carried out. The frequencies with which donor markers situated close to the origin are recovered indicates an obligatory interaction between the recipient chromosome and a region on the donor chromosome adjacent to the leading end. The partial exclusion from recombinants of some of the most proximal markers studied, however, indicates that this interaction does not occur at the free extremity of the transferred deoxyribonucleic acid. Alternate models to explain these facts are presented.
Induction of Penicillinase with Inorganic Phosphate
Authors: Imsande John , Gerber M. J.
Published: January 2020

Phosphate stimulates penicillinase formation in
Bacillus cereus
569. The rate of penicillinase synthesis in the presence of 0.3
m
phosphate,
p
H 7.0, is approximately 10-fold greater than that for uninduced cells, while the rate of synthesis in the presence of 0.3
m
phosphate and 1 unit/ml of penicillin is approximately fourfold greater than in the presence of penicillin alone. When phosphate-induced cells are transferred to low phosphate medium, the rate of penicillinase synthesis rapidly reverts to that of uninduced cells. Furthermore, the phosphate-induced synthesis of the enzyme is inhibited by either chloramphenicol or actinomycin D. These antibiotics are known to inhibit protein synthesis and deoxyribonucleic acid-dependent ribonucleic acid (RNA) synthesis, respectively. Thus, phosphate appears to induce the synthesis of a species of RNA that is required for the synthesis of penicillinase in
B. cereus
569. The penicillin-dependent induction lag for penicillinase was compared in high and low phosphate media. It was found that, at 37 C, the penicillin-dependent lag is approximately 3 min in the presence of 0.3
m
phosphate and approximately 6 min in low phosphate medium.
Polyamines and the Accumulation of Ribonucleic Acid in Some Polyauxotrophic Strains of
Escherichia coli
Authors: Raina Aarne , Jansen Miekie , Cohen Seymour S.
Published: January 2020

The cellular accumulations of polyamines and ribonucleic acid (RNA) were compared in the polyauxotrophic mutants of
Escherichia coli
strain 15 TAU and
E. coli
K-12 RC
re1
met
−
leu
−
. Putrescine, spermidine, and their monoacetyl derivatives were the main polyamines in both strains, when grown in glucose-mineral medium. No significant degradation of either
14
C-putrescine or
14
C-spermidine was found in growing cultures of strain 15 TAU, which requires thymine, arginine, and uracil for growth. Experiments with this organism showed that in a variety of different incubation conditions, which included normal growth, amino acid starvation, inhibition by chloramphenicol or streptomycin, or thymine deprivation, a close correlation was seen between the intracellular accumulation of unconjugated spermidine and RNA. In the presence of arginine, the antibiotics stimulated the production of putrescine and spermidine per unit of bacterial mass. Deprivation of arginine also resulted in an increase in the production of putrescine per unit of bacterial mass, most of which was excreted into the growth medium. However, in this system the antibiotics reduced the synthesis of putrescine. Furthermore, streptomycin caused a rapid loss of cellular putrescine into the medium. The latter effect was not seen in anaerobic conditions or in a streptomycin-resistant mutant of 15 TAU. Methionine added to the growth medium of growing TAU not only markedly increased the total production of spermidine, but also increased both the intracellular concentration of spermidine and the accumulation of RNA. Exogenous spermidine extensively relaxed RNA synthesis in amino acid-starved cultures of 15 TAU. Analysis in sucrose density gradients showed that the RNA accumulated in the presence of spermidine was ribosomal RNA.

Cells of
E. coli
K-12 RC
rel
met
−
leu
−
, grown in a complete medium, had approximately the same ratio of free spermidine to RNA as did strain 15 TAU. However, the relaxed strain showed a much lower ratio of putrescine to spermidine than the stringent 15 TAU. Omission of methionine stopped spermidine synthesis and markedly increased both the intracellular accumulation and the total production of putrescine. It seems that a high intracellular level of spermidine acts as a feedback inhibitor in the biosynthesis of putrescine in this strain. The hypothesis that the intracellular concentration of polyamines may participate in the control of the synthesis of ribosomal RNA in bacteria is discussed.
Macromolecule Synthesis in Yeast Spheroplasts
Authors: Hutchison H. Terry , Hartwell Leland H.
Published: January 2020

Conditions have been established for the preparation of spheroplasts of
Saccharomyces cerevisiae
which are able to increase their net content of protein, ribonucleic acid (RNA), and deoxyribonucleic acid (DNA), several-fold upon incubation in a medium stabilized with 1
m
sorbitol. The rate of RNA and protein synthesis in the spheroplasts is nearly the same as that occurring in whole cells incubated under the same conditions; DNA synthesis occurs at about half the whole cell rate. The spheroplasts synthesize transfer RNA and ribosomal RNA. The newly synthesized ribosomal RNA is incorporated into ribosomes and polysomes. The polysomes are the site of protein synthesis in these spheroplasts. Greater than 90% of the total RNA can be solubilized by treatment of the spheroplasts with sodium dodecyl sulfate or sodium deoxycholate. These spheroplast preparations appear to be a useful subject for the study of RNA metabolism in yeast.
Identification of
Mycoplasma
and Other Microorganisms by Polyacrylamide-Gel Electrophoresis of Cell Proteins
Authors: Razin Shmuel , Rottem Shlomo
Published: January 2020

The proteins of
Mycoplasma
cells of various species produce highly reproducible and species-specific electrophoretic patterns in polyacrylamide gels containing 5
m
urea and 35% acetic acid. These electrophoretic patterns can be used for the rapid identification and classification of
Mycoplasma
. Preliminary results indicate that this method may also be used for the identification and classification of other microorganisms.
Localization of Indigenous Yeast in the Murine Stomach
Authors: Savage Dwayne C. , Dubos Rene J.
Published: January 2020

Certain strains of yeast were cultured frequently from the feces of adult CFW mice and Long-Evans and Sprague-Dawley rats, but not from infants of those murine strains, or from adults or infants of NCS or NCS-D mice. When the yeasts could be cultured from the feces, they could also be grown from all areas of the digestive tracts of the animals, but especially from the stomachs, where they formed layers on the epithelium of the glandular mucosa. Three of the yeast isolates, one each from the three murine colonies, were provisionally classified in the genus
Torulopsis
of the asporogenous yeasts. These yeast strains failed to colonize the digestive tubes of suckling infant mice of either the CFW or NCS-D colonies. In contrast, they colonized the guts of adult NCS-D mice and formed layers in their stomachs; tests with the yeast from CFW mice revealed that this strain colonized the guts and formed layers in the stomachs of germ-free CFW mice. When established in NCS-D mice, the yeast strains did not affect qualitatively or quantitatively the growth of the animals or the composition of the bacterial flora in the gastrointestinal tracts. Moreover, they did not elicit an unusual inflammatory response in the digestive tracts; nor were they pathogenic for NCS mice when injected by the intraperitoneal or intravenous routes. The yeasts thus appear to be harmless saprophytes that are able to flourish in the environment of the surface of the secreting epithelium of the murine stomach. The findings conform with the view that some types of microorganisms of the gastrointestinal tract are not just mixed randomly but rather occupy microenvironments in almost pure culture. This concept is important to the understanding of the ecology of the gut microflora.
Endotoxic Glycolipid from a Heptoseless Mutant of
Salmonella minnesota
Authors: Kasai N. , Nowotny A.
Published: January 2020

The endotoxin of a heptoseless mutant of
Salmonella minnesota
R595 was extracted with phenol-water. Most of this material was found distributed in the insoluble fraction of the extract. The results showed that the R595 endotoxin behaved as a lipid rather than as a lipopolysaccharide (LPS). The preparation, although it does not contain O-specific polysaccharides, does contain 2-keto-3-deoxyoctonic acid (KDO), hexosamine, and several other unidentified compounds. Therefore, the term “glycolipid” is used in this paper instead of lipopolysaccharide. The crude glycolipid fraction, which was soluble in a mixture of chloroform-methanol (8:2), was purified by a procedure including fractionation with organic solvents and by different-column chromatographic methods. Although a chromatographic fraction of the glycolipid showed homogeneity in most systems investigated, the presence of contaminants could not be excluded. Chemical analysis of the glycolipids showed the absence of hexoses and heptoses. Constituents which were found were hexosamine, KDO, fatty acids, and phosphorus, which showed a relatively simple chemical composition. Partial acidic hydrolysis of the glycolipid yielded hexosamine-phosphates, as described in “Lipid A” fractions of smooth LPS preparations. Thin-layer chromatography of the partially hydrolyzed glycolipid showed a pattern similar to “Lipid A” fractions of other strains. The biological activity of the glycolipid was at the same level as that of other gram-negative endotoxins. Pyrogenicity, Shwartzman reactivity, and chick embryo
ld
50
values were as high or higher than those of purified
Serratia marcescens
endotoxin preparations, but mouse
ld
50
measurements gave significantly lower results.
Organization of the Tryptophan Pathway: a Phylogenetic Study of the Fungi
Authors: Hütter R. , DeMoss J. A.
Published: January 2020

The enzymes involved in tryptophan biosynthesis have been analyzed in a variety of fungal strains and a few other microorganisms. The same five biosynthetic reactions occur in all organisms tested, but differences have been found in the stability requirements for the enzymes, in their differential precipitation with ammonium sulfate, and in their sedimentation pattern after zone centrifugation. Based on the sedimentation behavior of anthranilate synthetase, phosphoribosyl-transferase,
N
-(5′-phosphoribosyl)-anthranilate isomerase, and indole-3-glycerophosphate synthetase, five different patterns of enzyme association could be recognized. The distribution of these patterns was used to evaluate several specific features of proposed phylogenetic relationships in the fungi. A closer relationship between
Chytridiales
and
Aspergillales
is postulated, eliminating the
Zygomycetes
and the
Endomycetales
as probable intermediates; the latter groups are considered to be sidelines. The data support the idea of a polyphyletic origin of the phycomycetes and suggest that anascosporogenous yeasts tested are related to the heterobasidiomycetes rather than to the
Endomycetales
. A possible sequence of changes leading to the various patterns of organization of the tryptophan pathway during the course of evolution is also proposed.
Release of Surface Enzymes in
Enterobacteriaceae
by Osmotic Shock
Authors: Neu Harold C. , Chou James
Published: January 2020

The process of osmotic shock, which has been used to release degradative enzymes from
Escherichia coli
, can be applied successfully to other members of the
Enterobacteriaceae
. Cyclic phosphodiesterase (3′-nucleotidase), 5′-nucleotidase (diphosphate sugar hydrolase), acid hexose phosphatase, and acid phenyl phosphatase are released from
Shigella, Enterobacter, Citrobacter
, and
Serratia
strains. Some strains of
Salmonella
also release these enzymes. Members of
Proteus
and
Providencia
groups fail to release enzymes when subjected to osmotic shock and do not show a lag in regrowth, although they do release their acid-soluble nucleotide pools. In contrast to
E. coli
, release of enzymes from other members of the
Enterobacteriaceae
studied is affected by growth conditions and strain of organism. None of the organisms was as stable to osmotic shock in exponential phase of growth as was
E. coli
. Exponential-phase cells of
Shigella, Enterobacter
, and
Citrobacter
could be shocked only with 0.5 m
m
MgCl
2
to prevent irreparable damage to the cells. These observations suggest that this group of degradative enzymes is probably loosely bound to the cytoplasmic membrane through the mediation of divalent cations.
Recovery of Division Ability in Ultraviolet-irradiated
Escherichia coli
Induced by Photoreactivation, Photoprotection, and Liquid Holding Treatment
Authors: Kantor George J. , Deering R. A.
Published: January 2020

Small doses of ultraviolet light (UV, 265 mμ) cause
Escherichia coli
B to grow into long, multinucleate, nonseptate, filamentous cells. This UV-induced filament formation can be prevented by irradiating with photoprotecting light (335 mμ) prior to UV irradiation, and by irradiating with photoreactivating light (406 mμ), or by liquid holding treatment, after UV irradiation. It is concluded that UV-induced division inhibition in
E. coli
B is initially induced by repairable lesions in the deoxyribonucleic acid, probably pyrimidine dimers.
Analysis of Sporulation Mutants I. Response of Uracil Incorporation to Carbon Sources, and Other Mutant Properties
Authors: Freese Ernst , Fortnagel Peter
Published: January 2020

Mutants deficient in sporulation were isolated and characterized with respect to antibiotic and protease activity, transformability, growth, and sporulation. All but two mutants could grow on minimal medium containing glucose. The inability of most mutants to incorporate uracil into trichloroacetic acid-precipitable material (ribonucleic acid) during the developmental period, and their response to a number of carbon sources, were used to characterize their biochemical blocks. Reproducible measurements of these responses were possible when the
p
H of the culture, which changed during growth and greatly influenced the rate of uracil uptake, was adjusted to 6.5. By their response to ribose and glutamate, the sporulation mutants could then be divided into four groups. All mutants of the first three groups produced antibiotic activity against
Staphylococcus aureus
, whereas all mutants, except one, of the fourth group produced none or very little of this activity. Mutants which did not respond to glutamate belonged to the first three groups; they also grew slowly or not at all on glutamate as sole carbon source. One of these mutants lacked succinic dehydrogenase activity. The results indicate that most of our sporulation mutants are unable to produce or utilize a natural carbon precursor, which is normally used as a slowly available carbon and energy source via the Krebs cycle when other carbon sources are used up. It enters the Krebs cycle as a precursor of α-ketoglutarate, probably via acetylcoenzyme A. All mutants of group four are blocked in this pathway before α-ketoglutarate.
Development and Structure of Bacterial Leaf Nodules in
Psychotria bacteriophila
Val. (Rubiaceae)
Authors: Lersten Nels R. , Horner Harry T.
Published: January 2020

The development and mature structure of bacterial leaf nodules in
Psychotria bacteriophila
were studied by using light and electron microscopy. Bacteria in mucilage surrounding the shoot apex pass through certain stomates in leaf primordia into the substomatal chamber. These chambers enlarge and become nodules as the yound leaves grow out of the apical region. Surrounding mesophyll cells grow into each nodule and form a cellular reticulum whose interstices are occupied by bacteria. Each intrusive mesophyll cell wall is unusually thick and continually supplemented by vesicles originating from dictyosomes. The gram-negative bacteria are often surrounded by capsules. Nodule bacteria contain several crystal-like dense bodies. A population of normal, dividing, and degenerating bacteria is found in each nodule. Extensive membranes occur between the bacteria. A hypothesis is proposed to explain certain aspects of this obligate symbiotic relationship.
Cell Wall Composition in Relation to the Taxonomy of Some
Actinoplanaceae
Authors: Szaniszlo Paul J. , Gooder Harry
Published: January 2020

Hydrolytic residues of the cell walls of 48 strains of
Actinoplanaceae
, previously assigned to 10 species and the four genera,
Actinoplanes, Ampullariella, Amorphosporangium
, and
Pilimelia
, were examined by paper chromatography and column chromatography. Comparisons were made for taxonomic purposes between the groupings obtained, by use of chemical characters and the groupings currently recognized morphologically. Most of the species investigated had qualitatively distinct cell wall compositions. Often, however, the cell wall compositions of species in different genera were more similar, in some respects, than were those of species in the same genus. Quantification of the cell wall amino acids and amino sugars substantiated that cross-generic similarities existed. Based on these results and the morphological conclusions reached by other investigators, a single-genus concept is suggested for the
Actinoplanaceae
examined.
Giant Cells of
Escherichia coli
Authors: Adler Howard I. , Terry Claude E. , Hardigree Alice A.
Published: January 2020

A mutant strain of
Escherichia coli
K-12 produced amorphous cells when grown in a variety of media. The
lon
−
allele, known to increase the radiation sensitivity of the cytokinesis mechanism, was introduced into the mutant by means of conjugation. Cells of this recombinant strain grew, after exposure to radiation, into giant amorphous cells, approximately 500 to 1,000 times the volume of a normal
E. coli
cell. These giant cells are analogous to the filaments formed after the irradiation of
lon
−
rod-shaped cells.
Synthesis of M Protein by Group A Hemolytic Streptococci in Completely Synthetic Media During Steady-State Growth
Authors: Davies Helen C. , Karush Fred , Rudd Joanne H.
Published: January 2020

Strains of type 6 (S 43) and type 14 group A streptococci were grown with M-protein production in the presence of chemically defined synthetic media slightly modified from that previously employed for the growth of a nonproducer of M protein (type 4). The M protein, which is associated with virulence in group A streptococcus, was previously produced in growing cultures only with complex media. The bacterial growth with the biosynthesis of M protein in synthetic medium was obtained by successive adaptation in steady-state culture with decreasing amounts of Todd-Hewitt broth. The synthesis continued for at least 480 generations at
p
H 7.3 and with a generation time of 84 min. Glucose was the limiting nutrilite and the concentration of reducing agents in the medium was critical. The M protein was identified by gel diffusion against type-specific antisera from the Communicable Disease Center and from R. Lancefield. The yield of M protein obtained from organisms grown in the continuous-culture device was comparable to that from standard broth stationary cultures.
Diacetyl Biosynthesis inStreptococcus diacetilactisandLeuconostoc citrovorum
Authors: Speckman R. A. , Collins E. B.
Published: January 2020

Pyruvate was shown to be the precursor of diacetyl and acetoin inStreptococcus diacetilactis, but dialyzed cell-free extracts ofS. diacetilactisandLeuconostoc citrovorumthat had been treated with anion-exchange resin to remove coenzyme A (CoA) formed only acetoin from pyruvate in the presence of thiamine pyrophosphate (TPP) and Mg++or Mn++ions. The ability to produce diacetyl was restored by the addition of acetyl-CoA. Acetyl-phosphate did not replace the acetyl-CoA. Neither diacetyl nor acetoin was formed when the otherwise complete reaction system was modified by using boiled extract or by omitting the extract, pyruvate, TPP, or the metal ions. Free acetaldehyde was not involved in the biosynthesis of diacetyl or acetoin from pyruvate, dialyzed cell-free extracts of the bacteria produced only acetoin (besides CO2) from α-acetolactate, and acetoin was not involved in the biosynthesis of diacetyl. Only one of the optical isomers present in racemic α-acetolactate was attacked by the extracts, and there was no appreciable spontaneous decarboxylation of the α-acetolactate at thepH (4.5) used in experiments.
Use of Nystatin to Eliminate Spontaneous Revertants in Yeast
Authors: Strömnaes Öistein , Mortimer Robert K.
Published: January 2020

Studies of induction of suppressor mutants may be obscured by revertants already present in the treated population. Such revertants, which appear as a minority of prototrophs among auxotrophs, can be eliminated by pretreatment with nystatin before application of the mutagen. Treatment with nystatin, under the proper conditions, decreased the frequency of prototrophs about 30-fold. This is sufficient for studying induced mutation at frequencies close to the spontaneous frequency.
Formation of Filaments and Synthesis of Macromolecules at Temperatures Below the Minimum for Growth of
Escherichia coli
Authors: Shaw Maxwell K.
Published: January 2020

When
Escherichia coli
ML30 was transferred during exponential growth at a temperature near the minimum for growth to temperatures just below the minimum for growth, optical density increased for a considerable period of time and considerable synthesis of ribonucleic acid, deoxyribonucleic acid, protein and mucopeptide also occurred. Synthesis of deoxyribonucleic acid was inhibited slightly before the cessation of synthesis of other macromolecules. At 6 C, filaments up to 300 μ in length were formed. Cross walls were not formed, but on transfer to 30 C the filaments rapidly fragmented into short, single cells. The filaments had abundant nuclear material distributed along their length, in contrast to filaments formed by
E. coli
15T
−
in the absence of thymine. There was evidence for false division points and incomplete septum formation.
Salt Specificity of a Reduced Nicotinamide Adenine Dinucleotide Oxidase Prepared from a Halophilic Bacterium
Authors: Hochstein L. I. , Dalton B. P.
Published: January 2020

Extracts prepared from a halophilic bacterium contained a reduced nicotinamide adenine dinucleotide (NADH
2
) oxidase active at high solute concentrations. The cation requirement was nonspecific, since KCl, RbCl, and CsCl replaced NaCl with little or no loss of activity, and NH
4
Cl was only partially effective. Only LiCl failed to replace NaCl. No specific chloride requirement was observed although not all anions replaced chloride. Bromide, nitrate, and iodide were essentially ineffective, whereas acetate, formate, citrate, and sulfate proved suitable. The presence of sulfate affected the ability of a cation to satisfy the solute requirement. Sulfate enhanced the rate of NADH
2
oxidation when compared with the rate observed in the presence of chloride. Cations which were inactive as chlorides (LiCl and MgCl
2
at high concentrations) satisfied the cation requirement when added as sulfate salts. Although magnesium satisfied the cation requirement, a concentration effect, as well as an anion effect, was observed. In the presence of MgCl
2
, little NADH
2
oxidation was observed at concentrations greater than 1
m
. At lower concentrations, the rate of oxidation increased, reaching a maximal value at 0.1
m
and remaining constant up to a concentration of 0.05
m
MgCl
2
. Magnesium acetate and MgSO
4
also replaced NaCl, and the maximal rate of oxidation occurred at 0.05
m
with respect to magnesium. There was no change in the rate of oxidation at high magnesium acetate concentrations, whereas the rate of NADH
2
oxidation increased at higher concentrations of MgSO
4
.
Effects of the Rare Earth Cerium on
Escherichia coli
Authors: Sobek Joseph M. , Talburt Dwight E.
Published: January 2020

The rare earth cerium was found to bind rapidly to
Escherichia coli
. Cerium inhibited oxygen uptake in the presence of glucose as well as the endogenous respiration of glucose-grown cells. For a cell concentration of 4 mg per ml, maximal inhibition was obtained at 120 μg per ml. Greater concentrations did not increase the inhibitory effect. Cerium inhibited
14
CO
2
evolution and
14
C uptake from uniformly labeled glucose. Marked changes in the distribution of
14
C incorporated into different chemical fractions of the cell were noted. The most striking changes occurred in the alcohol- and alcohol ether-soluble fractions, in which the
14
C activity was increased 5- to 20-fold in the presence of cerium.
Lactate Dehydrogenase Activity in Certain Strains of
Staphylococcus aureus
Authors: Stockland A. E. , Clemente C. L. San
Published: January 2020

Lactate dehydrogenase (LDH) was studied in phage-propagating strains 29, 3A, 6, 81, and 42D of
Staphylococcus aureus
selected from the five groups in the International-Blair series. Cells were cultivated in Brain Heart Infusion (Difco) under nearly anaerobic conditions and were harvested near the end of the log phase. LDH activity was maximal at the end of the exponential growth period and was measured spectrophotometrically by reduction of
p
-nitro-blue tetrazolium, with phenazine methosulfate as a coupling agent. Crude enzyme extracts were prepared both by an acetone extraction technique and by sonic treatment. LDH activity for these enzyme preparations was determined by the colorimetric method mentioned and also by measuring the rate of nicotinamide adenine dinucleotide reduction at 340 mμ. The order of activity observed, by use of both assay methods, was 29 > 81 > 6 > 3A > 42D. LDH forms (possibly isoenzymes) for each of 15 strains, which represent the five phage-propagating groups of the International-Blair series, were separated by acrylamide gel electrophoresis. Five forms were distinguished and arbitrarily numbered on the basis of their rate of migration, no. 5 being the slowest component. No one strain had more than four, nor fewer than two, LDH forms. Form 3 appeared in 13 of the 15 strains and was followed in frequency by no. 2, 1, 4, and 5.
Ribonucleic Acid, Deoxyribonucleic Acid, and Protein Content of Cells of Different Ages ofMycobacterium tuberculosisand the Relationship to Immunogenicity
Authors: Youmans Anne S. , Youmans Guy P.
Published: January 2020

The amount of ribonucleic acid (RNA), protein, and deoxyribonucleic acid (DNA) was determined in pellicle cultures of different ages of the H37Ra strain ofMycobacterium tuberculosis, grown on a synthetic medium. We found that the highest content of RNA and protein was present in 2-week-old cultures, indicating that these cells were in the logarithmic phase of growth. DNA content was highest at 1 and 2 weeks. The amount of all three compounds then decreased about 50% during the following 6 weeks. Two-week-old cells should therefore be used for preparation of the immunogenic ribosomal fraction. The optimal concentration of zinc chloride increased RNA and protein synthesis, and also improved the appearance of the pellicle growth. Two-week-old cells, which contained the largest amount of RNA and protein, immunized mice significantly better than older cells. Since protein and DNA are not involved in the production of immunity, a correlation could be made between amount of RNA and the capacity of viable H37Ra cells to immunize mice. The immunizing capacity of these cells was not affected by ribonuclease, probably because the ribonuclease did not penetrate into the whole cells.
Spirochete from the Rabbit Tick,Haemaphysalis leporispalustris(Packard) I. Isolation and Preliminary Characterization
Authors: Pickens Edgar G. , Gerloff Robert K. , Burgdorfer Willy
Published: January 2020

An unidentified spirochete, referred to as the 277F agent, was isolated fromHaemaphysalis leporispalustristicks from two cottontail rabbits by inoculation of the tick suspension into embryonated chicken eggs. Because of its minute width (0.1 μ), the organism was difficult to see when stained by the Giemsa method, but was readily demonstrated by silver impregnation or fluorescent-antibody procedures. In dark-field microscopy, the spirochetes appeared uniformly and rather tightly coiled, and exhibited typical corkscrewlike motility. After yolk-sac inoculation, the agent was highly lethal for chick embryos and was recovered in large quantity from several embryonic tissues and fluids. It could also be maintained in nonfertile eggs and in an enriched liquid medium. This previously undescribed spirochete could pass through Berkefeld N but not Seitz EK filters. It was relatively resistant to heat and to penicillin or sulfadiazine, but was markedly inhibited by streptomycin, broad-spectrum antibiotics, and homologous neutralizing antiserum. Of several species of animals tested for susceptibility to this spirochete, only the snowshoe hare gave evidence of infection.
Isolation of
Listeria monocytogenes
from Vegetation
Authors: Welshimer H. J.
Published: January 2020

A rural area in Virginia, in which clinical listeriosis of man and animals has been rare, was selected for this study. Vegetatino, which had died and remained in the fields over the winter, was collected and examined for
Listeria monocytogenes
by Gray's cold-holding procedure. Twelve fields were sampled and cultured, and eight strains of
L. monocytogenes
were isolated. Only two of the strains were pathogenic for the mouse. Compared with strains of
L. monocytogenes
isolated from listeric humans and animals in Virginia and the United States as a whole, these represented serotypes of low frequency.
Acridine Binding by
Escherichia coli
:
p
H Dependency and Strain Differences
Authors: Silver Simon , Levine Elaine , Spielman Philip M.
Published: January 2020

Acridine dye binding by cells of
Escherichia coli
has been characterized in terms of a number of parameters. There is a temperature-dependent, readily
reversible
binding of acriflavine which occurs to a greater extent with acridine-sensitive mutants of
E. coli
K-12 than with wild-type
E. coli
B or K-12. There is an essentially
irreversible
internal binding of acriflavine which occurs when the cellular permeability barriers are destroyed or altered by heat-treatment, elevated
p
H, treatment with toluene or phenethyl alcohol, or infection with bacteriophage T2 or T4. Both the reversible and the irreversible binding of acridines occurs more effectively with the acridine dye acriflavine than with the related dye proflavine, and still less effectively with 9-aminoacridine and quinacrine. These properties of acridine binding can be correlated with various inhibitory effects of the dyes on the cells.
Biosynthesis of Ergothioneine from Endogenous Hercynine in
Mycobacterium smegmatis
Authors: Genghof Dorothy S. , Van Damme Olga
Published: January 2020

Ergothioneine was synthesized and accumulated in growing cultures of
Mycobacterium smegmatis
when the medium was adequately supplied with sulfur. In a low sulfur medium, the accumulation was sharply limited although growth of the organism was apparently normal. Synthesis of hercynine, the precursor of ergothioneine, was unaffected by low sulfur levels and was markedly increased by addition of
l
-histidine, the precursor of hercynine. Resting-cell pellicle experiments, performed with cells grown on the low sulfur high histidine medium, showed that ergothioniene was synthesized from endogenous hercynine, when cysteine or compounds readily converted to cysteine (such as cystine, lanthionine, cystathionine, and thiazolidine carboxylic acid) were added. Homocysteine and djenkolic acid allowed for minimal synthesis of betaine, whereas methionine,
S
-methylcysteine, sodium sulfate, and sodium thiosulfate were unable to donate sulfur for ergothioniene synthesis under the experimental conditions employed. Addition of cysteine to a resting pellicle preparation caused the formation of 100 to 200 μg of ergothioneine per g of dry cells in 2.5 to 3 hr. A modified procedure for isolating ergothioneine and hercynine, employing a 75% ethyl alcohol extraction of wet organisms, followed by a single alumina column separation of the compounds, is described.
Stereospecificity of Citrate Synthetase in Relation to Glutamate Biosynthesis by Extracts of
Chloropseudomonas ethylicum
Authors: O'Brien R. W. , Stern Joseph R.
Published: January 2020

The synthesis of citric and glutamic acids by extracts of
Chloropseudomonas ethylicum
was studied with labeled precursors. When acetyl-coenzyme A-
1
-
14
C
was used as substrate, only 0.1% of the total radioactivity was found in the C-5 position of citric acid; whereas, with oxalacetate-
4
-
14
C
as substrate, 100% of the total radioactivity was found in C-5. These results demonstrated that the
Chloropseudomonas
citrate synthetase had an absolute stereospecificity, identical to that of the pig heart synthetase. The distribution of radioactivity in the glutamic acid synthesized from acetyl-coenzyme A-
1
-
14
C
was 0% in C-1 and 94.0% in C-5; whereas the glutamic acid formed from oxalacetate-
4
-
14
C
contained 89.6% in C-1 and 0.5% in C-5. This distribution is entirely consistent with the biosynthesis of glutamic acid from citric acid via aconitase,
d
s
-isocitrate, and
l
-glutamate dehydrogenases. The presence of
l
-glutamate dehydrogenase in extracts was demonstrated. The stereospecificity of the citrate synthetase and the pattern of glutamate labeling further establish that the aconitase of
Chloropseudomonas
is completely stereospecific.
Glutamate Catabolism ofRickettsia rickettsiand Factors Affecting Retention of Metabolic Activity
Authors: Rees Horace B. , Weiss Emilio
Published: January 2020

Glutamate catabolism and the factors contributing to metabolic stability of purified suspensions ofRickettsia rickettsiwere investigated. By use of14C-glutamate, it was shown that CO2was produced from all carbons of glutamate and that14CO2production was reduced by the addition of most of the unlabeled intermediates of the citric acid cycle and pyruvate. Oxalacetate, added in various concentrations, did not stimulate glutamate utilization. When the cells were suspended in bovine plasma albumin (BPA), CO2production from glutamate proceeded at a nearly uniform rate for 8 hr at 32 C and for 24 hr at 15 C. When BPA was used, the cells retained their metabolic activity at 0 or 30 C regardless of cell concentration, and were not influenced by the addition of varoius metabolites. Without BPA, metabolic stability was directly related to concentration. Of the stabilizers tested on low concentrations of rickettsiae, reduced glutathione was the most effective, provided that the gas phase contained predominantly N2. Under these conditions of low partial pressure of O2, glutamate further stabilized metabolic activity and was actively metabolized. The cells were also stabilized by oxidized glutathione in a gas phase of air, but under these conditions glutamate was utilized at a more moderate rate and it impaired metabolic stability.
Interactions of the Complement System with Native and Chemically Modified Endotoxins
Authors: Gewurz H. , Mergenhagen S. E. , Nowotny A. , Phillips J. K.
Published: January 2020

Endotoxic lipopolysaccharides (LPS) isolated fromSerratia marcescens, Veillonella alcalescens, andSalmonella typhosawere potent in their ability to induce fixation of complement (C′) in normal guinea pig, rabbit, mouse, and human serum. The C′-fixing ability of LPS was pronounced even when assays were performed in undiluted serum, and was lost after each of four chemical modifications which resulted in loss of biological toxicities. The detoxification procedures had in common the cleavage of ester-bound, long-chain carboxylic acids. The ability of biologically active LPS to fix C′ innormalguinea pig serum was reflected chiefly in dramatic uptake of classical C′3 (C′3t); fixation of C′1, C′4, and C′2 was virtually undetectable. Hence, it was the capacity for fixation of C′3t which was lost most overtly during detoxification. Addition of immune serum to the assay mixtures resulted in detectable fixation of C′1 and C′4. Biologically active LPS also fixed more of these components than did detoxified LPS. Immune serum restored the ability of detoxified LPS to fix C′3t, but whether this is by the original pathway is not yet clear. We concluded that the loss of certain biological activities and the loss of ability to fix C′3t in normal serum after LPS detoxification involved loss or rearrangement of substrates on LPS which either initiated or supported, or both, its interaction with the complement system. It was apparent that the ability to fix C′ can serve as a valuable in vitro indicator of the integrity of the toxic conformation of biologically active LPS membrane fragments. These experiments supported the hypothesis that certain of the biological activities induced by endotoxins are mediated via the complement system.
2-Deoxyribose Gene-Enzyme Complex in
Salmonella typhimurium
I. Isolation and Enzymatic Characterization of 2-Deoxyribose-Negative Mutants
Authors: Hoffee Patricia A.
Published: January 2020

Salmonella typhimurium
was found to utilize 2-deoxyribose as a sole carbon and energy source. Cells grown in the presence of deoxyribose contained increased levels of deoxyribose kinase, thymidine phosphorylase, and two forms of deoxyribose-5-phosphate aldolase (DR5P aldolase). One form of DR5P aldolase was induced by deoxyribose and coordinately regulated with deoxyribose kinase. The second form of DR5P aldolase was induced by deoxyribose-5-phosphate and coordinately regulated with thymidine phosphorylase. Mutants unable to ferment deoxyribose have been isolated and shown to be lacking either deoxyribose kinase or deoxyribose permease, but none has been found from which DR5P aldolase is missing. Thymine-requiring mutants which are able to grow on low levels of thymine have been isolated and shown, in some cases, to be lacking one or both DR5P aldolases.
Identification of
d
-Mannosamine and Quinovosamine in
Salmonella
and Related Bacteria
Authors: Lüderitz O. , Gmeiner J. , Kickhöfen B. , Mayer H. , Westphal O. , Wheat R. W.
Published: January 2020

Lipopolysaccharides of several strains of
Salmonella, Arizona
, and
Proteus vulgaris
, have been found to contain two unusual amino sugars not previously reported. These amino sugars were identified as
d
-mannosamine and quinovosamine (2-amino-2,6-dideoxy glucose).
Inhibition of Replication of an F′lacEpisome in Hfr Cells ofEscherichia coli
Authors: Dubnau Eugenie , Maas Werner K.
Published: January 2020

Hfr strains ofEscherichia coliK-12 were found capable of accepting a F′lacepisome during mating, with a frequency approximating that of F−strains. However, the F′lacepisome was unable to replicate in the Hfr cells, and was diluted out during the growth of the culture. Thelac+gene of the episome can be “rescued” by recombination into the host chromosome, as shown by the appearance of variegated recombinant colonies on a lactose-fermentation indicator medium. InrecAHfr strains, however, nolac+offspring were obtained in crosses with F′lacdonors. The induced synthesis of β-galactosidase in F′lac+× Hfr zygotes was studied. Rates of enzyme synthesis were approximately constant with respect to time as expected from unilinear inheritance of the F′lacepisome. However, the rate of synthesis eventually increased, presumably due to integration of thelac+gene in some of the zygotes. In F′lac+×recAHfr zygotes the rate of β-galactosidase synthesis remained constant with respect to time, as expected.
Chemical and Serological Relationships Between the Heteropolysaccharides of
Mycobacterium tuberculosis
and
Mycobacterium kansasii
Authors: Birnbaum Susan E. , Affronti Lewis F.
Published: January 2020

The identity of a heteropolysaccharide from cell walls of
Mycobacterium tuberculosis
H37Ra with Seibert's tuberculopolysaccharide I was demonstrated by thin-layer chromatography, chemical analysis, and antigenic tests. The polysaccharide of
M. kansasii
was shown to be identical with that of
M. tuberculosis
. Defatted cells were disintegrated by ultrasonic treatment in the presence of glass beads; cell walls were obtained by differential ultracentrifugation. Ethyl alcohol-precipitated carbohydrate extracts were analyzed for protein and nucleic acid; these impurities were removed. Tuberculopolysaccharide I from the mycobacterial culture filtrate is probably derived from a lipopolysaccharide of the cell wall, which is partially removed by chloroform in the intact state. Alkaline extraction releases additional polysaccharide, in varying degrees of association with cell wall murein.
N
-Acetylglucosamine Assimilation in
Escherichia coli
and Its Relation to Catabolite Repression
Authors: Dobrogosz Walter J.
Published: January 2020

The ability of
N
-acetylglucosamine to enhance catabolite repression by glucose was studied by using cultures grown on a combination of these substrates. Under these conditions, it was shown that two-thirds of the
N
-acetylglucosamine utilized was routed into dissimilatory pathways, whereas the remaining one-third was channeled into biosynthesis. It was established that over 50% of the
N
-acetylglucosamine assimilated was incorporated directly into amino sugar polymers. It was also shown that this exogenous supply of
N
-acetylglucosamine was in fact used preferentially over glucose as the precursor for amino sugar polymer biosynthesis. These findings provided support for the prediction that catabolite repression in
Escherichia coli
may be interrelated with certain reactions involved in amino sugar biosynthesis.
Relationship Between Sugar Structure and Competition for the Sugar Transport System in Bakers' Yeast
Authors: Cirillo Vincent P.
Published: January 2020

Twenty-five sugars have been compared as inhibitors of
l
-sorbose or
d
-xylose transport by the constitutive, monosaccharide transport system in bakers' yeast.
d
-Glucose showed the highest activity (i.e., apparent
K
i
= 5 m
m
). Since all sugars except 2-deoxyglucose showed a decrease in activity relative to glucose (i.e., apparent
K
i
= 25 − >2,000 m
m
), an attempt was made to relate the activity of each sugar with the way its structure differs from that of
d
-glucose. Assuming that the inhibition was the result of sugar-carrier complex formation, the analysis showed that the transport system has a rather broad specificity for pyranoses. Single changes at each of the five carbons of
d
-glucose (except for the 2-deoxy derivative) result in variable decreases in activity depending upon the carbon number and the alteration. The largest decrease in activity effected by a single change is the methylation or glucosylation of the anomeric hydroxyl. The combination of two or more changes leads to a decrease which is greater than the decrease in activity resulting from the individual changes occurring alone.
Purification and Genetic Determination of Bacteriocin Production inEnterobacter cloacae
Authors: de Graaf F. K. , Tieze G. A. , Bonga Sj. Wendelaar , Stouthamer A. H.
Published: January 2020

Enterobacter cloacae(strain DF13) was found to produce a bacteriocin which could be induced by mitomycin C. In the supernatant fluid of the induced culture phagelike particles were found. The bacteriocin was partially purified from induced cultures by ammonium sulfate precipitation and gel-filtration on Sephadex G-150. Ultraviolet-absorbing material was eluted from the Sephadex column in three fractions. The biological activity was mainly present in the second fraction and is associated with a protein with a molecular weight of about 61,000. The phagelike particles were found in the first fraction and show no biological activity. Upon conjugation ofE. cloacaestrain DF13 with another strain of the same species and withEscherichia coliK-12S, the ability to produce bacteriocin was transferred. The new bacteriocinogenic strain produced bacteriocin, which could not be distinguished from that produced byE. cloacaestrain DF13. Although transfer of the bacteriocinogenic factor often occurred together with transfer of the ability to produce phagelike particles, it was shown that these two factors are two separate genetic entities. In addition to a bacteriocinogenic factor,E. cloacaestrain DF13 was found to carry two other transferable plasmids: one determining resistance against streptomycin and sulfanilamide and another determining resistance against penicillin.
Effect of High Oxygen Tensions on the Growth of Selected, Aerobic, Gram-negative, Pathogenic Bacteria
Authors: Gottlieb Sheldon F. , Pakman Leonard M.
Published: January 2020

The in vitro effects of high O
2
tensions (P

O
2

) on aerobic, enteric pathogens were examined at pressures of up to 3 atm absolute. Organisms from the genera
Salmonella, Shigella
, and
Vibrio
were usually subjected to 24-hr exposures. Tensions of 0.87, 1.87, and 2.87 atm absolute of O
2
(plus traces of CO
2
and N
2
) became progressively inhibitory for
Salmonella
and
Shigella
growth, but were bactericidal only for
V. comma
strains at tensions greater than 0.87 atm absolute of O
2
. Growth inhibition of enteric organisms resulted from increased P

O
2

, rather than pressure per se, and could be mitigated nutritionally; an appropriate carbohydrate source is at least partially involved. Further studies with vibrios indicated that such mitigation was independent of medium
p
H. In addition, a synergistic relationship existed between O
2
and sulfisoxazole when tensions from 0.87 to 2.87 atm absolute of O
2
were maintained for 3 to 24 hr. Synergism occurred even under nutritional conditions which negated growth inhibition by O
2
alone. Bactericidal concentrations of sulfisoxazole, in the presence of increased P

O
2

, were reducible up to 4,000-fold. The combined procedure employed in this investigation, by use of an antimicrobial drug of known action, which also synergizes with O
2
, plus nutritional studies, suggests a means for establishing a site of O
2
toxicity. These data support the concept that O
2
inhibition of growth represents a metabolic disturbance and that metabolic pathways involving
p
-aminobenzoic acid may be O
2
-labile. Such an approach could also guide development of antimicrobial agents as O
2
substitutes for promoting synergism.
Screening of
Escherichia coli
Temperature-sensitive Mutants by Pretreatment with Glucose Starvation
Authors: Boyd D. , Nixon R. , Gillespie S. , Gillespie D.
Published: January 2020

A system for screening
Escherichia coli
temperature-sensitive mutants is described. The system involves glucose starvation and minimizes ambiguities introduced by the interdependencies of macromolecular synthesis during balanced growth. The system permits the quick recognition of protein synthesis mutants and their classification into two general catagories. Complete protein synthesis mutants are unable to make any polypeptide material, whereas partial protein synthesis mutants are able to produce inactive proteins. The phenotypes of several mutants are described.