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Volume 9,
Issue 2,
1953
Volume 9, Issue 2, 1953
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The Influence of Cultural Conditions on Polysaccharide Production by Aerobacter aerogenes
More LessSUMMARY: The polysaccharide content of Aerobacter aerogenes surface cultures grown on defined media was estimated by the anthrone method and related to the amount of growth as measured by the non-dialysable nitrogen content. Observations after different periods of incubation at 35° showed that on a medium containing excess sugar, growth was complete after 24 hr. and polysaccharide production was complete after 4 days. While most of the polysaccharide was produced after the cessation of growth, the rate of production per cell was greatest in the logarithmic phase. The polysaccharide: nitrogen ratio was low in 4-day cultures on media wherein cessation of growth was due to exhaustion of sugar (the sole carbon and energy source) and also was low in growths limited by anaerobic conditions or by the development of acidity. It was increased 10–20-fold in cultures wherein cessation of growth was due to exhaustion of limited nitrogen, phosphorus or sulphur sources, so that an excess of sugar remained available for extra polysaccharide synthesis. This great increase in the polysaccharide: nitrogen ratio of the culture was paralleled by similar increases in the degree of mucoidness as measured by the wet weight: nitrogen ratio, in the intracellular polysaccharide content as measured by periodate-Schiff staining, in capsular polysaccharide as measured by capsule diameter, and in loose slime polysaccharide as measured chemically in the supernatant after centrifugation of the culture. After 4 days at 35° the loose slime accounted for about half of the total polysaccharide in the highly mucoid cultures and about a fifth of that in the non-mucoid cultures.
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A Strain of Lactobacillus bifldus which requires Strepogenin
More LessSUMMARY: A strain of Lactobacillus bifidus was isolated from the faeces of a breastfedinfant and subcultivated in the minimal medium of Hassinen, Durbin, Tomarelli & Bernhart (1951) . The cysteine requirement for maximal growth of the strain was c. 100 μg. cysteine/ml. Human and cow milk were found to contain another factor, besides cysteine, which was essential for optimal growth. The active material in milk could be replaced by enzymic hydrolysates of some commercial proteins; egg albumin, and blood albumin were the most potent sources. The active material was heat stable, but was completely inactivated by treatment with strong acid. All the properties of this active material studied showed a close resemblance to those of strepogenin.
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The Study of Cell Rupture in Staphylococcus aureus
More LessSUMMARY: In order to prepare staphylococcal cell walls as free as possible from intact organisms yet with minimum chemical and mechanical degradation, a rapid turbidimetric method was developed to measure the proportion of staphylococci ruptured by shaking with glass beads. The assay took less than a minute, and was used to find the bacterial concentration and speed and time of shaking which gave the biggest yields of cell walls in the shortest time.
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A Method for Direct Viable Counts of the Facultatively Anaerobic Microflora in the Rumen of a Sheep maintained on a Hay Diet
More LessSUMMARY: By direct viable counts in deep cultures in a basal nutrient agar, consisting of buffered and very highly clarified rumen liquor, supplemented only with vitamin-free acid hydrolysate of casein, tryptophan, a fermentable carbohydrate and agar, the crude wet rumen contents of a hay-fed sheep has been shown to contain a population of c. 108 viable and facultatively anaerobic bacteria per g., mostly Gram-positive cocci capable of fermenting a wide range of soluble carbohydrates including starch and inulin but not mannitol. The mannitol-fermenting population is of the order of 106 organisms per g., consisting mostly of Gram-negative rods, including coliform bacteria. Addition of reducing substances such as sodium sulphide or mercaptoacetate (thioglycollate) to the basal agar had little or no effect in increasing the counts. An extract of hay made with cleared and sterile rumen liquor did not yield the typical rumen streptococcal flora when counted by this method in raffinose + rumen liquor nutrient agar.
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The Effect of Fatty Acids on the Growth, Metabolism and Morphology of the Anaerobic Cocci
More LessSUMMARY: The sodium salts of a number of fatty acids had little, if any, effect on the metabolism or morphology of representative strains of anaerobic cocci of groups I, II, V and VI ( Hare, Wildy, Billett & Twort, 1952 ). However, strains of groups III and IV in the presence of an adequate concentration of fatty acid showed much greater activity than in media without substances of this nature; growth was very much better, gas was formed in larger amounts and organic acids (e.g. pyruvate) or carbohydrates (e.g. fructose) were attacked. The morphology of these group III and IV organisms was also affected in that the cocci were small (0·5–0·8 μ.) and well formed in the presence of adequate concentrations of fatty acid, whereas in control preparations they were larger (1·2–1·6 μ.) and sometimes had transverse septa.
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Genetic Analysis of Valine Inhibition in the K12 Strain of Bacterium coli
More LessSUMMARY: Development of resistance to valine by the K12 strain of Bacterium coli (Escherichia coli) has been found to occur by a one-step process leading to full resistance in every case. The genetic factors controlling this valine resistance have been studied by recombinant analyses. The results indicate a very close linkage between valine resistance and leucine independence. These substances have previously been shown to bear special relationship to each other with respect to the K12 strain.
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The Classification of Lactobacilli by means of Physiological Tests
More LessSUMMARY: Four hundred and fifty-two strains of lactobacilli, including representatives of all recognized species and 197 freshly isolated cultures, were studied. It was possible to classify 390 (86·3%) of these into eight groups on the basis of the results of six physiological tests. The tests used were the production of gas from glucose and citrate, and of ammonia from arginine, growth at 15, 45 and 48°, heat survival at 60 and 65 and tolerance of 4, 6 and 8% (w/v) sodium chloride.
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The Precipitating Antigens of Friedländer’s Bacillus
More LessSUMMARY: The biochemical characteristics of approximately eighty strains of Friedläder’s bacillus are described. Certain aspects of their antigenic patterns were studied by testing the precipitating antigens of phase variants of originally mucoid strains, as well as the parent strains themselves. Evidence is presented that an as yet undescribed antigenic layer exists between those characteristic of the mucoid and smooth phases. This layer is most pronounced in the Sm phase organisms. The Sm antigen was shown to be present in nearly all Friedländer’s strains and absent in the Bacterium coli strains studied; hence an Sm antiserum should be valuable for diagnostic purposes. The biochemical and serological results are discussed in regard to the taxonomy of this group with particular reference to the consolidation, already proposed by other workers, of the genera Klebsiclla and Aerobacter.
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Paths of Phosphate Transfer in Micrococcus pyogenes: Phosphate Turnover in Nucleic Acids and other Fractions
More LessSUMMARY: The paths of transfer of phosphate groups during uptake of inorganic orthophosphate by Micrococcus pyogenes var. aureus (strain Duncan) have been studied by following the incorporation of 32P-labelled phosphate groups into the phospholipid (LP), acid-soluble inorganic (AI) and organic (AO), deoxyribonucleic acid (DNA), ribonucleic acid (RNA) and glycerophospho-protein complex (PGP) fractions of the cells.
In resting cells a strictly reciprocal exchange of inorganic phosphate across the osmotic barrier is half complete in 70 min. During respiration or growth, phosphate moves inward through the osmotic barrier at the same rate as during rest, but the outward movement is abolished. Phosphate accumulates mainly in the AI, AO and LP during respiration, but in all the fractions during growth. Most of the phosphate of the organic fractions is drawn through the AI. The RNA is formed from the nucleotides of the AO, while the DNA phosphate is derived either directly from the AI or from intermediates in the AO. Little or no turnover of RNA or DNA phosphate occurs during growth, and phosphate is not transferred directly or indirectly from RNA to DNA or vice versa. The PGP, although the major constituent of the mechanically rigid cell envelope, does not participate as a whole in phosphate transfer reactions during rest or respiration; but during growth the amount of PGP increases in proportion to the expansion of cell envelope area. The LP phosphate exhibits rapid turnover during respiration and growth.
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Transport of Phosphate across the Surface of Micrococcus pyogenes: Nature of the Cell ‘Inorganic Phosphate’
More LessSUMMARY: An osmotic barrier, impermeable to inorganic phosphate ions, is present near the external surface of Micrococcus pyogenes var. aureus (strain Duncan). The barrier encloses a volume of 1·66 ± 0·07 ml./g. dry-wt. cells, within which the inorganic phosphate of the cell is confined at a concentration of c. 0·1 m. The inorganic phosphate of the internal and external media exhibit a rapid reciprocal exchange, sensitive to traces of heavy metals. The existence of the osmotic barrier and the characteristics of the phosphate exchange are not in accord with an exchange-adsorption hypothesis; they are in accord with an exchange-diffusion hypothesis, according to which phosphate is shuttled across the osmotic barrier by carrier molecules. There is no experimental justification for supposing the acid-soluble inorganic phosphate of M. pyogenes to exist as an acid-labile compound.
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Electron Microscopy of a Staphylococcal Bacteriophage
More LessSUMMARY: The staphylococcal bacteriophage 3A consists of a flat, oblong head, c. 600 A. by c. 1000 A., and a long tail, c. 120 A. by c. 2900 A. Fixation with formalin destroys the integrity of the head. Cells infected with the phage release it at the end of the latent period by a process resembling the bursting of a bag. After the phage is released, the empty cell membranes persist for a short time before being dissolved in the medium. Correlation of these findings with turbidity readings and Gram staining of the infected cultures indicate that the turbidity falls only when the shells of the cocci are dissolving.
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Micromanipulation on an Agar Surface for the Isolation and Cultivation of Single Organisms
More LessSUMMARY: Organisms may readily be isolated and cultured separately by micro-manipulation on the upper surface of an agar block. A micromanipulator is not essential, its place being taken by a second microscope with simple attachments. Illumination methods by dark-ground and phase-contrast techniques are described. A modification using a simple micromanipulator enables isolations to be made very rapidly. A device is described for making the necessary micro-instruments in a standard microscope.
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Heterotrophy in Diatoms
More LessSUMMARY: Forty-two bacteria-free cultures of pennate diatoms from fresh water or soil were tested for ability to grow heterotrophically with glucose as the sole carbon source. Thirteen of these isolates proved capable of growth under these conditions. They comprised seven isolates of Navicula pelliculosa, five of other different species of Navicula, and one of Nitzschia (?) fonticola.
One isolate of Navicula pelliculosa was tested for its ability to grow heterotrophically on any one of sixty organic compounds. Only glucose, glycerol, and fructose were found to be effective in supporting growth. Glucose was metabolized both in the dark, and in the light in the absence of CO2. Glycerol and fructose were not utilized in the dark, but supported growth in the light in the absence of CO2.
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Antibiotic Production by the Microfungi of Acid Heath Soils
More LessSummary: A study has been made, using a plating technique, of the fungi occurring in acid sandy podsol soils and of their distribution at different levels of the soil profile. The most widespread species, characteristic of this type of soil, included: Absidia orchidis, Mortierella alpina, Mucor ramannianus, Penicillium frequentans, P. nigricans, P. roqueforti, P. terlikoiwskii and Trichoderma viride. A number of other species were locally abundant, including Absidia glauca, Conio-thyrium sp., Gliocladium sp. (roscum series), Penicillium citrinum and Zygorhyncus moelleri.
The plate counts of viable fungal units decreased sharply from upper to lower levels, though there was occasionally a secondary maximum in the upper part of the B horizon. Most species were characteristically found most abundantly in upper layers of the A horizon, but Mucor ramannianus was notable for its abundance at much lower levels; in several of the profiles examined it was isolated in almost pure culture from the B horizon.
Of the sixty-five species isolated, about half produced antibiotics. Most of these were toxic to fungi and bacteria; only a few cases of specific antifungal or antibacterial activity were encountered. The Phycomycetes differed from other groups in that none produced antifungal substances. The capacity to produce antifungal antibiotics appears to be correlated with the distribution of species; of the widespread and locally abundant fungi, 45% produced such antibiotics as compared with 15% of the rare fungi. Nevertheless, certain widespread species (e.g. Absidia orchidis, Mortierclla alpina, Mucor ramannianus, Penicillium roqueforti) did not produce antifungal antibiotics, so it cannot be considered to be a necessary character of widespread soil fungi. There is some evidence that such widespread non-producers of antibiotics are relatively resistant to the antibiotics produced by other species.
An attempt has been made to group the fungi isolated into Burges’ ecological categories of ‘sugar-fungi’ and ‘humus-fungi’ on the basis of their capacity to attack and utilize carboxymethylcellulose (CMC). Only 26% of the fungi were unable to utilize CMC, Phycomycetes being prominent among these. Whereas 35% of the rare fungi and 54% of locally abundant fungi were ‘sugar-fungi’, only 17% (all Phycomycetes) of the widespread fungi were ‘sugar-fungi’.
These results are believed to lend support to the view that antibiotic production is of ecological significance to soil fungi. They also provide information which can form the basis of an experimental search for more direct evidence.
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The Induction of Lysogenesis in Salmonella typhimurium
More LessSummary: After exposure to phage A1 an organism of an indicator strain of Salmonella typhimurium may burst with phage liberation, may become lysogenic, or may develop into a mixed progeny of lysogenic and phage-sensitive bacteria. By the methods we have used, the proportion of such mixed progenies is about 20% of the number of lysogenic colonies found. Evidence is given to show that members of the clone of phage A1 vary in virulence and that members of the indicator strain 1404 vary also in resistance to lysis. The establishment of lysogenesis depends on the interplay of these two factors when host and parasite meet.
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Growth of Streptococci in a Glucose Phenolphthalein Broth
More LessSummary: A liquid medium buffered at pH 9·6 is described which is modified from that of Shattock & Hirsch (1947) by the addition of phenolphthalein and in the method of preparation. Its use in distinguishing streptococcal species was investigated.
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