- Volume 22, Issue 1, 1960

SUMMARY: A strain of Pseudomonas whose motility could be activated under anaerobic conditions by arginine was shown to decompose arginine to ornithine by enzymic reactions involving citrulline and adenosine phosphates. The enzymes were readily separable from the particulate cell fractions and were not associated with the external flagellar apparatus. Citrulline also activated anaerobic motility but only after a period of pre-aeration or subsequent to its aerobic intracellular synthesis from ornithine. Permeability experiments suggested strongly that the cells of the pseudomonad were more permeable to arginine and citrulline under aerobic than anaerobic conditions. The promotion of motility by citrulline and the dependence of arginine-activated motility on the activity of the arginine dihydrolase system indicate that arginine acts as a direct source of energy for motility.

SUMMARY: The normally slow rates of growth and of cellulose synthesis by certain strains of Acetobacter in static culture were accelerated greatly when these strains were cultured in shaken, indented flasks or in stirred aerated fermenters, whereby the time factor was reduced from 3–4 weeks to 2–4 days. Optimum yields of cellulose from Acetobacter acetigenum, strain EA-I and A.xylinum strain HCC B-155 were produced in shaken, indented flasks, in a medium of hydrolysed molasses which contained 4–5 % of sugar and about 0·05 % of nitrogen (as ammonia). Comparison of eleven strains of cellulose-forming Acetobacter sp. in static and shaken culture showed that all produced less cellulose and all except two showed less growth in shaken culture. Yields of cellulose in stirred, aerated fermenters decreased with increasing air flow rates, although the growth level remained constant. The results suggest that increased aeration decreases the yield of cellulose by causing decreased synthesis of cellulose per unit cell weight, but the involvement of celluloseless mutants cannot be excluded. Addition of neutralizing agents to the fermenter cultures increased the yields of cellulose by the order of 50 %.

SUMMARY: The kinetics of the early stages of infection by herpes virus were studied in the chorio-allantoic membrane (CAM). Adsorption followed approximately first-order kinetics and was temperature-dependent. Adsorbed virus was not removed from the CAM by repeated washing after various adsorption periods up to 60 min. Forty to 70 % of the adsorbed firmly bound virus was neutralized by anti-herpes serum at the end of the adsorption period, mid became gradually insusceptible to antiserum over the next three hours. Only about 0·1 % of this adsorbed virus was recovered by freezing and grinding infected CAM’s at the end of adsorption, although the recovery procedures themselves did not inactivate the virus. Because of the failure to demonstrate any other reason for the loss of over 99 % of adsorbed virus it is suggested that virus had lost its infectivity very rapidly upon adsorption at a time when a considerable part of it was susceptible to antiserum. Growth of new virus occurred from the non-recoverable virus.

SUMMARY: Evidence is presented which suggests that after a period of adsorption of 30 min. the herpes virus which could not be removed from HeLa cells by repeated washing existed in four fractions: Fraction A, about 63 % which became non-infective and insusceptible to antiserum over the next 2 hr.; fraction B, about 16 % which remained susceptible to antiserum and infective for up to 6 hr. after adsorption; fraction C, about 2 %, which remained infective and insusceptible to antiserum for at least 6 hr. after adsorption; fraction D, about 19 %, which became non-infective and insusceptible to antiserum during the adsorption period. Replication occurred from fractions A and D. Inactivation and the development of insusceptibility to antiserum occurred very close together in time, possibly simultaneously. Incubation of infected cells in the presence of antiserum lowered the yield of virus obtained after 48 hr. incubation; this could not be explained simply by prevention of secondary infection through the medium.

SUMMARY: Only one of sixteen strain of Tetrahymena, studied, was found to be completely dependent on an exogenous source of serine for growth. A second strain appears to lade threonine aldolase, as it can synthesize serine from added glycine but not from added threonine. The remaining fourteen strains will utilize either glycine or threonine (as a source of glycine) for serine synthesis provided the folic acid concentration in the medium is high.

SUMMARY: Metabolic products of fermentable carbohydrates, as well as certain fatty acids, ethanol and acetaldehyde can serve as sources for serine synthesis in Tetrahymena. Exogenous hydroxypyruvate and related compounds were inactive.

Sparing experiments indicate that serine contributes to the formation of glycine, cysteine (presumably via cystathionine), aminoethanol and thymine.

Summary: A suspension of isolated cells of Mycobacterium tuberculosis was mixed with melted, watery agar and used to prepare thin films of agar gel suspended in ⅜ in. diameter wire loops. These films were incubated in culture media and at intervals of time sample films were transferred to glass slides, fixed, stained and ‘the mean count per colony’ determined. Some batches of agar inhibited growth, but this .inhibition was reduced by extracting the powdered agar with methanol, and was abolished by adding serum, egg or charcoal to the medium. Strain H37Rv and 5 freshly isolated strains growing in a good medium all showed lag periods of 14–17 hr. and generation times of 13–18 hr. In unfavourable media the lag extended to 10–15 days.

Summary: By selection with concentrations of streptomycin from 25 to 250 μg./ml., auxotrophic mutants were obtained with high frequency from Salmonella typhimurium strain LT-2. All of them were found to require both thiamine and nicotinic acid for their growth. The requirement for thiamine can be satisfied by either 4-methyl-5-hydroxyethyl-thiazole, cystine or cysteine but not by methionine. Nicotinic acid can be replaced by nicotinamide. No other amino add, vitamin, purine or pyrimidine can replace these vitamins.

In addition to the nutritional requirement, all of these mutants are streptomycin resistant slow growers. These four characters, i.e. thiamine and nicotinic acid- requirement, slow growth and streptomycin resistance are converted to wild-type in a single step by transduction with phage PLT-22, grown on wild-type bacteria as well as on prototrophic one-step intermediate streptomycin resistant mutants and streptomycin indifferent mutants. Spontaneous mutants with wild phenotype were also obtained from some unstable mutants of this group, apparently in a single mutational event. The transductional findings suggest that these auxotrophic mutants are probably due not to some chromosomal aberration but to mutation within a single locus. The data from reciprocal transductions between these mutants suggest that multiple sites of the same locus are responsible. The mutations at different sites seem to have different levels of mutability and some of them have different levels of streptomycin resistance.

SUMMARY: Faster growth of Escherichia coli strain B cells is observed in media which Contain factors previously found to increase the numbers killed by radiation. Incubation in the presence of metabolic inhibitors after irradiation allows many more irradiated cells to originate colonies. Holding the cells on complete media at low temperatures after irradiation causes more irradiated cells to die, an exception to the principle that greater survival of strain B is associated with slower growth. The difference in the radiosensitivity of strain B and two resistant mutants, strains B/r and Bpr5, depends on differences in their utilization of organic nutrients, since: (i) all three strains have about the same sensitivity if they are plated on a minimal medium; (ii) if strains B and B/r are treated with chloramphenicol immediately after ultraviolet, the survival of strain B increases, and that of strain B/r decreases, both reaching the same level as if they had been plated immediately on minimal medium; (iii) if irradiated cells of strains B and B/r are incubated on nutrient medium for a short period before chloramphenicol treatment, the survival of strain B roughly equals that of the resistant mutants, while that of B/r is unaffected. When logarithmic phase broth-grown cells of all three strains are exposed to visible light after ultraviolet, only strain B is ‘photoreactivated’, and then only when viable counts on nutrient medium are compared, the counts on synthetic medium being unaffected by the visible light.

Summary: The conditions for the formation, the isolation and the purification of a new class of disaccharide derivatives, namely 3-ketoglycosides, are described. These compounds are formed by the action of certain bacteria, provisionally called Alcaligenes faecalis, on lactose, maltose, lacto- or maltobionate. From lactose about 76 % of the total amount was converted into the corresponding 3-ketoglyco-side. Neither the corresponding monoses, gluconate nor equimolar mixtures of these substances were oxidized this way. The new substances were ultimately purified by paper chromatography. No crystalline preparations have so far been obtained. These bacteria contain strong hydrolases for either maltose or lactose, and weak ones for the bionates. The yield of 3-ketoglycosides is inversely proportional to the hydrolase activity.

Summary: Bacteria, provisionally called Alcaligenes faecalis, oxidize lactose, maltose and the corresponding bionates aerobically to 3-ketoglycosides. Several properties of the pure compounds are described, as well as the arguments, leading to the proposed chemical formula.

SUMMARY: The sonic disruption of spores of Bacillus cereus gives multi-hit kinetics. The first hit destroys the exosporium, which protects the spore body from destruction; the second hit destroys the spore body. Spores which have been stripped of their exosporia are still viable and heat resistant. From the rate of release during sonic treatment it appears that alanine racemase, adenosine deaminase and hexos- amine are located in the exosporium while ribosidase is in the spore body. The rate of release of dipicolinic acid does not identify its location.

SUMMARY: The structure conferring rigidity and shape on the complex cell wall of Escherichia coli strain B has been isolated in a state virtually free from other wall material. It shows a characteristic surface pattern which, together with observations on its mode of disintegration by phage enzyme or lysozyme, indicates the fairly simple principles of its construction.

SUMMARY: The wild-type strain of Rhodopseudomonas spheroides 2.4.1 does not utilize exogenously supplied gluconate and grows poorly with glucose, fructose or mannose as substrates, accumulating aldonic acids and 2-keto-3-deoxy gluconic acid in media containing these sugars. Mutants which are capable of growing well with glucose without the accumulation of acids acquire the enzyme ‘phosphoglu-conic acid dehydrase’ and oxidize hexoses mainly via the pathway described previously in Pseudomonas saccharophila. The Embden-Meyerhof pathway is constitutive in the wild-type and mutant strains of R. spheroides but appears to have a limited function as a result of very low aldolase activity. Phosphogluconic acid dehydrogenase is either absent or too low in activity to permit the utilization of the pentose-phosphate oxidative pathway.

A new type of particulate ‘aldose dehydrogenase system’, constitutive in all strains is involved in the oxidation of glucose and mannose to the corresponding aldonic acid lactones. Gluconic and mannonic acids are both converted to 2-keto- 3-deoxy acid through the action of two distinct dehydrating enzymes. A mutation leading to the utilization of gluconic acid is associated with the acquisition of a high level of ‘2-keto-3-deoxy gluconic acid kinase’, but not of phosphogluconic acid kinase. A permeability factor may also be involved in the adaptation of gluconate.

SUMMARY: A glutamic dehydrogenase from Neurospora requiring either DPNH or TPNH for activity has a pH optimum of 8·6 and is inhibited by a number of chelating agents. The specific activity of the enzyme was high during early growth, but it fell sharply from the 2nd to the 5th day after inoculation. A deficiency of Zn decreased enzyme activity and addition of the metal in vivo to Zn-deficient mycelial felts restored the enzyme to normal values after a few days of incubation. The enzyme was unchanged when pyridoxine or riboflavin was limiting in the growth of mutants which require these vitamins. With a purified enzyme the Km values in mole/l. were as follows: α-ketoglutarate 0·75 × 107#x2212;2; TPNH, 2 × l0−4; and DPNH, 2·3 × 10−4.

SUMMARY: Hydrogenasesfrom Azotobacter vinelandii and Clostridium pasteurianum reduced methylene blue, ferricyanide, benzyl- and methyl-viologens when hydrogen was the donor. Methylene blue was the most effective acceptor. Hughes press preparations of either organism in tris (2-amino-2-hydroxymethyl propane-1:3-diol) buffer (pH 8·0) resulted in the best extraction of the enzyme. Hydrogenase from C. pasteurianum was readily inactivated by traces of oxygen but this could be prevented by sodium dithionite. Pyridine nucleotides and cytochrome c. are reduced by hydrogenase, but in extracts of Azotobacter, however, the addition of iron was required to couple DPNH to cytochrome c. The mechanism appears to involve the reduction of Fe3+ to Fe2+ enzymically and the Fe2+ is oxidized by cytochrome c non-enzymically. The effect of pH value and nature of buffer on this system was examined. Mo5+ did not reduce cytochrome c, with or without hydrogenase.

Metal-deficiency experiments, inhibitor studies, activation of dialysed preparations, and the results of radioactive tracer assays of purified protein fractions, showed that iron is the main metal constituent of hydrogenase. Molybdenum, however, is required for the fixation of nitrogen.

Summary: Highly labile factors (I and II) of anthrax toxin were purified from the toxic plasma of guinea-pigs just dead from anthrax by fractionation methods which involved the minimum of inactivation. The final preparations of factors I and II, which still contained constituents of guinea-pig plasma, were toxic when injected together but innocuous when injected separately.

SUMMARY: A serological comparison on gel-diffusion plates of purified preparations of factors I and II of the anthrax toxin obtained in vitro and in vivo indicated that all but one (factor I from in vitro sources) of these preparations contained at least two major serological components. At least three serological components were involved in the total system: A (associated with factor I activity); B (associated with factor II activity); C (not associated with toxic activity). These results indicated that no preparation of factor II of the anthrax toxin so far examined was immunologically homogeneous and that the serological connexion between factor I of the anthrax toxin (produced in vivo) and the protective factor produced in vitro (Smith, 1958) was due to a common component not associated with factor I activity. There was some evidence that the method of measuring immunogenicity in certain culture filtrates of Bacillus anthracis by serological precipitation (Thorne & Belton, 1957) is not generally applicable to all products of this organism.

SUMMARY: The ability of washed suspensions of Escherichia coli Y44 (which requires histidine and p-aminobenzoic acid for growth) to synthesize methionine from homocysteine was decreased almost to zero when the organism was harvested from a growth medium containing methionine itself. There was a similar effect, though in no case quite so severe, with eleven other strains of the organism. In further experiments with strain Y 44 the effect was found to be specific to methionine; the presence during growth of no other amino acid tested decreased subsequent synthesis by more than 20 %. Ability to synthesize methionine was lost and regained on single successive cultivations in the presence and absence of the amino acid; there was also no evidence from viable counts on agar media ± methionine for a genetic Change. ‘Inactive’ organisms (methionine-grown) were not activated by changing the nature of the precursor of the methyl group nor by adding heated extracts of yeast or of ‘active’ organisms. There was no evidence that ‘inactive’ organisms produced an inhibitor of methionine synthesis. It is concluded that growth with methionine leads to the non-production of an enzyme(s) concerned directly in the addition of the one-carbon unit to homocysteine.