- Volume 23, Issue 2, 1960
Volume 23, Issue 2, 1960
- Article
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Further Mutational Studies on the PAS Resistance of Mycobacterium tuberculosis var. hominis
More LessSUMMARY: Mutational studies on the PAS (sodium p-aminosalicylate) resistance of Mycobacterium tuberculosis var. hominis, strain H37RV have been carried out. It has been assumed that one type of population structure of clones with relation to drug resistance corresponds to one genotype. Clones derived from single colonies obtained on different PAS concentrations produced three types of population structure; the first type is similar to the population structure of the parent strain which has not been previously exposed to the drug and its degree of resistance is up to 0·2 μg. PAS/ml. The second and the third types are different from the parent strain and their degrees of resistance are up to 3 and 1000 μg. PAS/ml., respectively. Neither intermediate population structures nor intermediate degrees of resistance were clearly observed. From these results, it may be concluded that there are only two genotypes responsible for PAS resistance, one determining a lower, and the other a higher degree of resistance.
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Morphological Variants of Proteus hauseri
More LessSUMMARY: Cultures of Proteus hauseri may consist of one or more of five colonial variants. Population pressure experiments started with one variant eventually yielded all other variants. Y variants form raised non-swarming colonies on a MacConkey-type agar at 37°, but swarm in concentric step-like rings at room temperature on this medium; they swarm in step-like concentric rings on nutrient agar at room temperature and at 37°. This variant is stable under conditions of logarithmic growth and is regarded as the wild type. Variant X is a typical Rough form and does not swarm under any circumstances. In continuous culture it mutates to the Y type at a high rate. The W variant forms colonies very similar to those of the Y variant, but like the X variant does not swarm. It is stable under conditions of continuous culture, but mutates on MacConkey agar to X and Y variants at low rates. On MacConkey agar at 37° the Z type appears as a flat amorphous colony surrounded by a continuous thin film of swarm. It also swarms at room temperature. This variant is replaced by a mixture of Y and W colonial forms in continuous culture experiments. Drop-like colonies, the ‘Tröpfchenform’ previously described by Weil & Felix (1917) were also encountered. Drop-like colonies are unpredictable in appearance and were not studied in detail.
Hanging-drop preparations of 4 hr. old broth cultures at 37° of either W or X colonial variants (which never swarm) reveal many motile forms. Young broth cultures of the other three variants consist of forms which are all actively motile. Lytic phage for a particular strain is active on all its variants except the X or Rough type which is also the only variant to form unstable suspension in saline and in broth. The Y, W and Z variants described here correspond reasonably well to the A, B and C phases of Belyavin (1951). The H, O terminology of Weil & Felix (1917) is not suitable and the nomenclature of Belyavin (1951) may be more satisfactory.
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Nutrition of Flavobacterium aquatile Strain Taylor and a Microbiological Assay for Thiamine
More LessSUMMARY Flavobacterium aquaiile strain Taylor grew readily in a medium containing (%, w/v); trypticase, 0·5; glucose, 0·5; K2HPO4, 0·05; KH2PO4, 0·05; MgSO4.7H2O, 0·02; supplemental thiamine. Growth was proportional to the thiamine-HCl content in the range 1·80 mμg./10 ml. medium. Larger quantities of thiamine did not increase growth. Maximum growth responses at all concentrations of thiamine were obtained after 48 hr. incubation at 30° when the cultures were shaken during incubation. The response of F. aquatile to thiamine gives a basis for a microbiological assay. Of the compounds tested only thiamine pyrophosphate and acetylthiamine replaced thiamine. None of the other vitamins, or various purines, pyrimidines, nucleosides, or miscellaneous compounds tested except certain carbohydrates, altered the response to thiamine. Maltose or ribose (0·:5 %, w/v) stimulated F. aqua-tile in the range 0–5 mμg. thiamine/10 ml. At greater concentrations of thiamine, and in the presence of glucose, slight growth inhibitions were caused by maltose and ribose. Sodium chloride at concentrations greater than 0·:05 m retarded growth generally and therefore the response to thiamine was modified. None of the interfering compounds appeared to invalidate the use of the bacterium as an assay organism for thiamine.
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Preferential Compatibility in Streptomyces violaceoruber
More LessSUMMARY A compatibility system, or system of relative mating potency, has been found among mutants of Streptomyces violaceoruber (S. coelicolor). Certain combinations of growth factor-dependent mutants failed to form nutritionally independent (prototrophic) recombinants. Incompatibility was not the result of inability of identical mutant genes to interact, failure of hyphae to fuse, or inhibition of proto-troph development by incompatible mixtures. No infective or diffusible factors were found which induced or suppressed fertility. The yield of recombinants was affected by the relative growth rates of the two parental types, the time elapsed between the planting of each parent and the relative and absolute numbers of the parents. Sporogenesis was not a prerequisite for recombination.
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Structure of the Mate-Killer (mu) particles in Paramecium aurelia, stock 540
G. H. Beale and A. JurandSUMMARY The mate-killer (mu) particles in the cytoplasm of Paramecium aurelia, stock 540, variety (syngen) 1, were studied by: Feulgen-staining; phase-contrast microscopy; fluorescence under ultraviolet irradiation after staining with acridine orange; electron microscopy (including the ‘silver-Feulgen’ technique). The appearance of the particles following treatment with DNAase and RNAase was observed. It was found that the mu particles were capsulated rod-shaped structures, 2–10 μ long and about 0·3 μ in diameter (excluding the capsule); they appeared to reproduce by transverse fission without formation of cross-walls. There was an external double membrane and the internal material consisted largely of DNA, which was not limited to particular zones but spread throughout the interior of the particle. RNA was also present. The relationships of the particles to other micro-organisms is discussed, and it is concluded that they do not differ from bacteria in any important respect.
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Interference with Multiplication of Pc Phage by a Factor Produced by Exposure of Pseudomonas aeruginosa to Inactivated Pc Phage
More LessSUMMARY Pseudomonas aeruginosa strain C10 when exposed to phage Pc which had been inactivated by heat or acid produced a factor which decreased the yield of Pc phage in a single cycle of growth on P. aeruginosa C10. Some of the factor appeared in the culture fluid, but most could be obtained only by disrupting the bacteria. The factor did not inactivate free Pc phage. It decreased phage yield most when it was mixed with P. aeruginosa C10 1 hr. before infection with Pc phage and was then kept present to the end of the experiment. When the factor was diluted out immediately after adsorption of Pc phage on P. aeruginosa C10, or when it was present only from the start of adsorption, it had less effect on phage yield.
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Replica Plating and Rapid Ascus Collection of Neurospora
More LessTwo colonial strains of Neurospora crmsa have been found suitable for testing nutritional requirements by replica plating. One strain (crisp) has been replicated with a set of needles and the other (crisp, ragged) with filter paper. When ripe perithecia from crisp × crisp crosses are submerged in water, complete asci can be collected rapidly with a capillary pipette.
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A Technique for Obtaining Uniform Inocula of Actinomyces israelii
More LessSUMMARY: The anaerobic filamentous organism Actinomyces israelii grows as discrete colonies which sediment in static liquid media and are too large and variable in size to be suitable for quantitative inoculations of experimental media. A stirred- culture technique yielded uniformly small viable colonies of Actinomyces israelii. The microcolonies obtained still sedimented rapidly but could be kept in even suspension by bubbling nitrogen through the suspension. Such suspensions gave uniform inocula from a graduated Pasteur pipette as judged by total-nitrogen determinations and showed the growth characteristics of A. israelii grown in static liquid media.
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The Oxidation of Tricarboxylic Acid Cycle Intermediates by a Strain of Corynebacterium erythrogenes
More LessSUMMARY: Washed suspensions of Corynebacterium erythrogenes strain 142, grown on a yeast extract medium, were unable to oxidize some members of the tricarboxylic acid cycle immediately unless these compounds had been added to the growth medium. At 25° the organism eventually acquired the ability to oxidize these compounds; with succinate rapid oxidation was not obtained even after 6 hr. Rapid oxidation of succinate did occur when washed suspensions were incubated first with l- or d-malate. The lag periods were shortened by the addition of ammonium sulphate, but adaptation was prevented by chloramphenicol, 2:4-dinitrophenol, azide or an increase in temperature to 37°. Neither chloramphenicol nor the increase in temperature affected oxidation by adapted organisms; 2:4-dinitrophenol and azide were inhibitory, especially when added with the substrate. Extracts of unadapted organisms oxidized immediately all those compounds which intact organisms did not. Chloramphenicol and the respiratory uncoupling agents did not inhibit oxidations by the cell extracts. Extracts from adapted or unadapted bacteria reacted with succinate, malate and fumarate in the presence of hydroxylamine to form substances which gave coloured compounds with ferric chloride. The compounds with fumarate and malate were not identified. These compounds and succinhydroxa- mate were formed equally well by extracts of organisms grown in the absence or presence of the dicarboxylic acids. There was no evidence that the enzymes responsible for these reactions are directly concerned with the penetration of the acids into C. erythrogenes..
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Differentiation of Acetomonas and Pseudomonas
More LessSUMMARY: A comparative study of 50 strains of Acetomonas and 165 strains of Pseudomonas showed that the two genera could be easily and rapidly differentiated by the use of four criteria; these were: (a) production of acid on ethanol + CaCO3 agar plates, (b) oxidation of calcium lactate to carbonate, (c) production and accumulation of dihydroxyacetone in glycerol media, (d) growth at (initial) pH 4·5. Acetomonas was positive with (a), (c) and (d), but negative with (b); Pseudomonas was positive only with (b). Liquefaction of gelatin and production of a greenish yellow fluorescent pigment, being variable with Pseudomonas and absent from Acetomonas, were useful only when positive.
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The Dissociation of Morphogenesis from Cell Division in the Cellular Slime Mould, Dictyostelium discoideum
More LessSUMMARY: Dictyostelium discoideum, strain Fty-1, myxamoebae were harvested from the stationary phase and dispensed in small population samples on washed agar. Total cell counts were made at intervals between deposition and fruit construction. Within an uncertainty level of 2·5 %, the data indicate that the populations remained constant and the terminal fruits contained the same number of cells as had formed the aggregates. Subsidiary evidence based on viability determinations and the known morphology of moribund cells suggested that new cells did not arise nor old cells die.
When myxamoebae were harvested from the log phase while still feeding on a high density of bacteria, a substantial degree of cell division did-occur during morphogenesis. But the increase in cell number of myxamoebae which had not aggregated corresponded approximately, with that of myxamoebae which had aggregated and fruited. Consequently such increases appear to be coincidental with and not causally related to the morphogenetic sequence.
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Yeasts from Antarctica
More LessSUMMARY: The yeast flora of moss from Granite Harbour and of soils from Wright Dry Valley in the McMurdo Sound region of Antarctica was examined. There were 75,000 yeasts/g. on the moss; species recovered were Cryptococcus laurentii, C. albidus and Rhodotorula minuta. Yeasts were isolated from 8 of the 12 soil samples in numbers up to 5000/g. Strains of Candida scottii, whose maximum growth temperature was c. 15°, were the predominant soil yeasts.
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The Keto Acid Metabolism of Corynebacterium diphtheriae growing in Submerged Culture
More LessSUMMARY: The keto acid production of Corynebacterium diphtherias (strain CN2000) growing in submerged culture in complex medium was investigated. Pyruvic acid production followed a pattern in which there was a very rapid production of acid which reached a peak about 12 hr. after inoculation. The acid concentration decreased rapidly after 12 hr. and the final pyruvic acid concentration appeared to be very variable. The source of the pyruvic acid is believed to be lactic acid, the concentration of which decreased rapidly as pyruvic acid increased. The presence in the medium of sufficient iron to abolish toxin production completely did not affect the pattern of pyruvic acid production. α-Ketoglutaric acid production in the cultures appeared to vary in a random manner. C. diphtheriae strain G12/6 produced a maximum yield of pyruvic acid after only 6 hr. growth but was otherwise similar to strain CN2000. C. diphtherias strain SM1 gave a maximum production of pyruvic acid after 24 hr. but in the presence of 3 μg. Fe/ml. production was more rapid and was like that found for strain CN2000. No correlation was found between toxin production and the production of pyruvic acid or α-keto glutaric acid.
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The Influence of Trace Elements on the Metabolism of Aromatic Compounds by Soil Fungi
More LessSUMMARY: The effect of copper, iron, manganese and zinc on the metabolism of catechol, o-, m- and p-hydroxybenzoic, o-, m- and p-methoxybenzoic, 3:4-dimethoxybenzoic and vanillic acids was investigated. Aspergillus niger, Hormodendrum sp. and Penicillium sp. mycelia which were deficient in the various trace elements were the organisms used. Only iron had any marked effect on the rates of metabolism and on the accumulation of intermediate products of metabolism in the substrate solutions. The intermediate products identified were: protocatechuic acid from m- and p-hydroxybenzoic and p-methoxybenzoic acids; catechol from o -hydroxy- benzoic acid; vanillic acid from 3:4-dimethoxybenzoic acid; o-, m- and p-hydroxy- benzoic acids from the corresponding mono-methoxybenzoic acids.
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Isolation of a Corynebacterium and its Transitional Forms from a Case of Subacute Bacterial Endocarditis treated with Antibiotics
More LessSUMMARY: A bacteriological study was made of blood specimens taken repeatedly during a 2-year period from a child with subacute bacterial endocarditis who received intensive treatment with antibiotics. The conventional bacillary form of Corynebacterium sp. was present in the blood and bone marrow of the patient before the beginning of antibiotic therapy and on occasions when the administration of antibiotics was suspended. These were the periods when the patient showed overt symptoms of clinical illness. When antibiotic therapy was adequate to produce clinical remission of symptoms, the infecting organism was not eradicated, but persisted in the blood in a small granule-like form that could be demonstrated and cultured only by highly specialized techniques. The cultural procedures required to bring about reversion of the granule-like form to the conventional bacillary form and the morphology of the various transitional forms that the organism assumed during the reversion process are described and illustrated.
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The Location of the Group Antigen of Group D Streptococcus
More LessSUMMARY: Gel-diffusion precipitin tests showed that the Streptococcus group D antigen, as usually demonstrated by ring-precipitin tests, was serologically identical in all four species (Streptococcus faecalis, S. faecium, S. durans, S. bovis). Cell walls of representative cultures of all group D species had a similar chemical composition (determined by chromatograpy) as found by other workers. The one strain of S. bovis examined lacked galactosamine which was present in the cell walls of the other group D species. The group D specific antigen was found in the residue (cell contents) after the cell-wall fraction had been removed, unlike the group A specific antigen which forms an integral part of the cell wall.
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Studies on the Mode of Action of Antifungal Heptaene Antibiotics
More LessSUMMARY: The effect of heptamycin (an antifungal heptaene antibiotic closely related to the ascosin-candicidin-trichomycin group) on the respiration of Candida albicans was studied. Heptamycin inhibited the oxidation of pyruvate, lactate and trehalose to a considerable degree. The oxidation of glucose, acetate, acetaldehyde and ethanol was only slightly affected by heptamycin. Anaerobic glycolysis was significantly inhibited. No inhibition of cell-free preparations of trehalase (obtained from C. albicans) or pyruvic carboxylase (from C. albicans and Saccharomyces carlsber-gensis) was observed; CO2 output by cell-free preparations was not affected either. The uptake of pyruvate and trehalose by C. albicans cells is apparently due to an active and inducible mechanism; heptamycin inhibits oxidation of these substances by blocking their penetration into the yeast cell. The oxidation of trehalose in induced organisms was not inhibited by heptamycin whereas that of pyruvate was. Heptamycin, candicidin B, ascosin, trichomycin and amphotericin B inhibited phosphate uptake by non-proliferating suspensions of C. albicans. The inhibition was observed at fungistatic concentrations (0·5–5μg./ml.) and without any preincubation with the antibiotic. Phosphate uptake by Debaryomyces nicotianae, Pichia farinosa, Cryptococcus albidus and Saccharomyces cerevisiae was similarly affected by heptamycin. Esterification of inorganic phosphate by cell-free extracts of C. albicans was not affected by heptamycin. It is suggested that the fungistatic effect of heptaene antibiotics upon sensitive yeasts is mainly due to the prevention of phosphate uptake.
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A Specific Polysaccharide from Mycoplasma mycoides
More LessSUMMARY: Washed suspensions of Mycoplasma mycoides var. mycoides were found to contain c. 10 % by weight of carbohydrate. The only sugars detected in hydrolysates were galactose and ribose. A galactan was obtained by extraction with warm aqueous phenol. The galactan was separated from ribonucleic acid by ultracentrifugation or by treatment with an ion-exchange resin. The product was rapidly hydrolysed by acid, releasing galactose and c. 4 % of CHCl3- soluble lipid. It formed precipitates with specific antiserum and could be used to sensitize sheep erythrocytes to agglutination by the antiserum. It was not pyrogenic for rabbits and had little in common with the lipopolysaccharides of typical Gram-negative bacteria.
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The Influence of Proflavine on the Synthesis of Foot-and-Mouth Disease Virus
More LessSUMMARY: Infective ribonucleic acid (RNA) can be extracted with phenol from virus- infected pig kidney tissue-culture cells before the appearance of new complete virus. In the early stages of multiplication, however, infective RNA cannot be obtained from the cells if they are incubated with ribonuclease before treatment with phenol. This suggests that the infective RNA is in a ‘free’ enzyme-sensitive form at this time. At the end of the lag phase, as the RNA is incorporated into complete virus, there is a decrease in the proportion of infective RNA in the cells which is sensitive to the enzyme. Nine hours after infection all the material in the cells which yields infective RNA on treatment with phenol is insensitive to ribonuclease. Incorporation of low concentrations of proflavine into the incubation medium decreases considerably the yield of infective virus, although the amounts of infective RNA and specific complement-fixing antigen are not proportionately reduced. In the presence of proflavine the RNA in the cells remains sensitive to ribonuclease throughout the growth cycle indicating that the compound prevents the incorporation of RNA into complete virus. The complement-fixing antigen produced in the proflavine inhibited system is the 7 mµ non-infective component of the virus.
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An Immunological Study of Bacillus subtilis Penicillinases
More LessSUMMARY: Antisera to the penicillinases of Bacillus subtilis strains 749 and 6346 were prepared, and the neutralization of enzymic activity by homologous and heterologous antisera was studied. Antisera to 749 penicillinase had, as expected, greater neutralizing power against 749 penicillinase than against 6346 penicillinase. A 6346 antiserum, however, neutralized the heterologous enzyme more effectively than the homologous enzyme, and was also more effective in precipitating heterologous than homologous enzymic activity. The penicillinases of B. subtilis, unlike those of B. cereus, could not be adsorbed on glass, but could be partially purified by calcium phosphate gel-adsorption. There was no cross-reaction, as measured by enzyme neutralization, between the penicillinases of B. subtilis and that of B. cereus 569. Materials other than penicillinase in the enzyme preparations used did not interfere with the neutralization reactions between B. subtilis penicillinases and their antisera. The basal penicillinase produced by B. subtilis 749 had the same neutralization curve, with a homologous antiserum, as did the induced enzyme.
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Inter-relationships of Alleles at the am Locus in Neurospora crassa
More LessSUMMARY: It has been shown that the alleles 5, 6 and 7 at the am locus in Neurospora crassa backmutate; the alleles 4, 8, 9 and 11 do not backmutate at detectable frequencies. Wild-types were recovered in low frequencies from crosses between different am alleles. It has been shown that each of the twelve am alleles, so far investigated, is unique with respect to at least one of the following criteria: mutation, recombination, complementation, type of glutamic dehydrogenase produced. It is argued that the minimum number of possible mutations at the am locus, which can affect glutamic dehydrogenase production, is eighteen. The probability that this minimum estimate is too large is less than 1 %.
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