- Volume 128, Issue 10, 1982
Volume 128, Issue 10, 1982
- Obituary
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- Biochemistry
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Use of Lectins to Characterize the Receptor Sites for Bacteriophage PL-1 of Lactobacillus casei
More LessThe polysaccharide (PS) located outside the peptidoglycan layer in Lactobacillus casei ATCC 27092 was found to inhibit the adsorption of PL-1 phage to cell wall preparations without inactivating free phage. Electron microscopic examination of adsorption mixtures showed that the phage were adsorbed to fragments of PS material in a tail-first orientation. Phage did not adsorb to isolated peptidoglycan. The PS was composed of l-rhamnose, d-glucose, d-glucosamine and d-galactosamine, with the hexosamines possibly in N-acetylated form. Prior treatment of L. casei with Streptomyces haemagglutinin, an anti-human blood group B agglutinin that binds specifically to l-rhamnose, resulted in a concentration-dependent inhibition of phage adsorption. Phage adsorption was inhibited partially by lectins specific for d-glucose and N-acetyl-d-glucosamine, but not by lectins specific for N-acetyl-d-glucosamine or N-acetyl-d-galactosamine. The inhibition of phage adsorption occurred immediately upon the addition of the effective lectins, and was reversed by the addition of the respective lectin inhibitors, α-phenyl galactoside or α-methyl glucoside. The results indicate that l-rhamnosyl residues are the main determinants of the PL-1 phage receptor sites, while d-glucosyl residues may be involved more indirectly.
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Characteristics of Fructose-1,6-bisphosphatase from the Methanol-utilizing Yeast Hansenula polymorpha
More LessFructose-1,6-bisphosphatase (EC 3.1.3.11) has been partially purified from methanol- and ethanol-grown Hansenula polymorpha. The enzyme was specific for fructose 1,6-bisphosphate as a substrate and did not react with sedoheptulose 1,7-bisphosphate. The enzyme purified from ethanol-grown cells resembled that from methanol-grown cells in all properties investigated, namely, temperature stability, substrate affinity, pH-dependent inhibition by AMP, and pH-dependent stimulation of activity by EDTA. It is concluded that, although the metabolic role of fructose-1,6-bisphosphatase during growth on methanol and ethanol is completely different, growth on these substrates probably involves the same enzyme.
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Enzymes of N-Acetylglucosamine Metabolism during Germ-tube Formation in Candida albicans
More LessThe enzymes of N-acetyl-d-glucosamine (GlcNAc) metabolism, GlcNAc-6-phosphate deacetylase and GlcN-6-phosphate deaminase were found to be inducible in Candida albicans. The pattern of induction for these enzymes was the same under conditions of germ-tube formation (37 °C) and where yeast cells metabolized GlcNAc with no change in morphology (28 °C); this indicates that these enzymes are not control points in the dimorphic development of C. albicans. During induction there was a 40- and 25-fold increase in specific activity for the deacetylase and the deaminase, respectively, and the maximum specific activity corresponded to the time when all the GlcNAc had been metabolized. The presence of lomofungin (an inhibitor of transcription) or trichodermin (an inhibitor of translation) in cell suspensions of C. albicans containing GlcNAc prevented the increase in specific activity of these enzymes. 2-Deoxyglucose inhibited germ-tube formation, partially inhibited the induction of the deacetylase (43%) and the deaminase (60%), but did not affect the growth of C. albicans on either Glc or GlcNAc. GlcN-6-phosphate was a competitive inhibitor of the deacetylase with a K i of 1·45 mm while the other product of the reaction, acetate, did not inhibit the enzyme. The K m value for GlcN-6-phosphate on GlcN-6-phosphate deaminase was 0·24 mm. Incubation of starved yeast cells with GlcNAc produced a four-fold increase in the specific activity of UDP-GlcNAc-pyrophosphorylase at either 28 °C or 37 °C.
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Properties of the β-Glucosidases of Talaromyces emersonii
More LessThe thermophilic fungus Talaromyces emersonii when grown on cellulose-containing medium produces three extracellular forms of β-glucosidase, I(M r 135000), II (M r, 100000), III (M r, 45700), and one intracellular form, β-glucosidase IV (M r, 57600). With the exception of β-glucosidase II which has yet to be isolated in quantities sufficient for characterization each has been shown to be a glycoprotein and to exist as a single polypeptide. Each of the purified enzyme preparations catalysed cellobiose hydrolysis but only β-glucosidase IV, at the concentrations of substrates used, also catalysed glucosyl transfer to cellobiose.
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- Ecology
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Maintenance of the Normal Flora of Human Skin Grafts Transplanted to Mice
More LessFull-thickness human cadaver skin was maintained on the dorso-lateral thoracic region of hairless mice whose immune rejection mechanism was suppressed using anti-mouse-thymocyte globulin. The bacterial profile of the pregrafted skin did not differ significantly from the normal human microflora. In contrast, the murine skin exhibited quantitative and qualitative differences from the human flora, in particular by the complete absence of Propionibacterium acnes, the dominant bacterium on sebum-rich areas of human skin. The normal microbial profile of the human grafts was maintained throughout the experimental period despite the novel environmental milieu. There was little contamination of the grafts from the normal murine flora. It was concluded that the grafted human skin would provide a realistic model for studying the ecology of human cutaneous micro-organisms.
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Nitrate Dissimilation by Vibrio spp. Isolated From Estuarine Sediments
More LessThe generic distribution of nitrate-respiring bacteria in tidal mudflats of six Scottish east-coast river estuaries has been determined. The predominant bacteria involved were identified as Aeromonas/Vibrio spp., together with lower numbers of enterobacteria. Pseudomonads accounted for only a small proportion of the nitrate-respiring populations. Samples of sediment from all the estuaries produced nitrite and ammonia after 7 d incubation at 15 °C with added nitrate. Studies with the isolate Vibrio V48 and strains of Vibrio parahaemolyticus (NCMB 1903 and NCMB 2047) demonstrated that when these bacteria were grown anaerobically under conditions of glucose limitation, nitrite was the primary product of nitrate respiration, whereas ammonia was excreted into the medium when nitrogen was limiting. Nitrate reduction was accompanied by the synthesis of a particulate nitrate reductase and under N-limitation a nitrite reductase was also induced.
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Pectinases in Leaf Degradation by Aquatic Hyphomycetes: the Enzymes and Leaf Maceration
More LessAll seven species of aquatic hyphomycetes tested produced both polygalacturonases and pectin lyases. The polygalacturonases were constitutive, whereas the pectin lyases were induced on pectic substrates at pH 6.5 and above. Some species could grow on pectic substrates at both pH 5 and pH 7; other species at pH 7 only. Tricladium splendens grew as well on a polypectate substrate as it did on glucose. This fungus elaborated three endo-polygalacturonase isoenzymes, as did Articulospora tetracladia. Tetrachaetum elegans secreted a presumed exo-pectin lyase and a pectin esterase, and Mycocentrospora angulata an endo-pectin lyase and a pectin esterase. The four species macerated alder-leaf strips totally within 9 to 12 d at stream pH 7, utilizing predominantly pectin lyases and pectin esterases. These enzymes are stimulated by Ca2+, and this may explain why plant decay proceeds most rapidly in calcium-rich streams of pH 6.5 and above. The frequency with which the four species of aquatic hyphomycetes were observed on experimental leaf packs placed in a stream does not appear to be related to their pectolytic capability.
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- Genetics And Molecular Biology
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Amplification and Product Identification of the fnr gene of Escherichia coki
More LessThe position of a gene (fnr) that is essential for growth of Escherichia coli with fumarate or nitrate as electron acceptor was located within an 11·5 kb HindIII fragment of bacterial DNA by deletion analysis with fnr transducing phages (λfnr) and by sub-cloning restriction fragments into multicopy plasmids. The functional gene was isolated in a 1·65 kb BamHI-HindIII fragment of a hybrid plasmid pGS24. The fnr gene product wasidentified as a protein of M r 31000, by post-infection labelling and by the ‘maxicell’ method. Organisms containing the multicopy plasmid (pGS24) overproduced fumarate reductase to the same extent as cultures containing a comparable fumarate reductase plasmid (pNU3 1) during anaerobic growth; the fnr plasmid also overcame the repression of fumarate reductase synthesis that is normally observed during aerobic growth. Similar effectson nitrate reductase synthesis were also observed. The results support the view that the fnr gene product functions as a positive regulator or a specific sigma factor for expression of anaerobic energy-generating systems.
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Distribution of the Escherichia coli K12 Insertion Sequences IS1, IS2 and IS3 Among Other Bacterial Species
More LessA total of 15 bacterial species were screened for the presence of the Escherichia coli K12 insertion sequences IS1, IS2 and IS3 by Southern blotting and DNA hybridization. The only positive signals obtained were for IS1 in Serratia marcescens, where the sequence was present on a plasmid, and a weak hybridization to IS3 in Erwinia carotovora. This shows that the capacity for transposition alone is not sufficient to allow the spread of sequences through a wide range of bacterial species.
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Loss of Nitrofuran Resistance in Fusarium oxysporum is Correlated with Loss of a 46·7 kb Circular DNA Molecule
More LessStrain FC1, a wild-type isolate of the phytopathogenic fungus Fusarium oxysporum, is resistant to nifuroxime, a nitrofuran derivative. Mutants sensitive to this compound arise at high frequency by vegetative segregation. Resistance to nifuroxime is inherited as an extranuclear marker in crosses between a nifuroxime-sensitive variant and the wild-type. Nifuroxime-sensitive strains are also unable to express an exopolygalacturonase activity present in the wild-type. For one strain tested, it was shown that the ability to parasitize a susceptible tomato plant variety was also lost. Loss of nifuroxime resistance was concomitant with the loss of a 46·7 kb circular DNA molecule.
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Characterization of Saccharomycopsis lipolytica Mutants Producing Lowered Levels of Alkaline Extracellular Protease
More LessMutants affected in production of alkaline extracellular protease (xpr mutants) have been characterized. The alkaline protease produced by the xpr mutants, except for that of the structural gene mutant, was indistinguishable from the wild-type protease on the basis of isoelectric focusing in polyacrylamide gels, SDS-PAGE and thermal stability. Some revertants of xpr mutants were temperature sensitive for protease production, but the thermal stability of the revertant protease was not altered. The xpr mutants grew as rapidly as the wild-type with glutamic acid, leucine or urea as sole nitrogen source. The pleiotropic xpr mutants which produced lowered levels of extracellular RNAase also produced less extracellular acid proteases (4 to 40% of wild-type). These results suggest that the pleiotropic xpr mutants may affect secretion. However, production of phosphatase(s) (which was secreted but located primarily in the cell envelope) was much less affected in these mutants (65 to 100% of wild-type). A possible explanation for these results is that at least one step (component) of the secretion pathway for the extracellular proteases and RNAases is not shared by the phosphatase(s).
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The Plasmid-coded Metabolism of Naphthalene and 2-Methylnaphthalene in Pseudomonas Strains: Phenotypic Changes Correlated with Structural Modification of the Plasmid pWW60-1
More LessPseudomonas sp. NCIB 9816 contains two plasmids: pWW60, an IncP9 plasmid of 87 kb encoding genes for the catabolism of naphthalene, and pWW61, a cryptic plasmid of about 65 kb. The ability to degrade naphthalene was transferred at low frequency by conjugation from strain NCIB 9816 into a plasmid-free strain of Pseudomonas putida, PaW340. A transconjugant, PaW701, containing the naphthalene plasmid pWW60-1, metabolized naphthalene and salicylate via the ortho pathway. 2-Methylnaphthalene was not a growth substrate but was partly metabolized with accumulation of a brown compound in the medium (λ max = 440 nm). Spontaneous mutants of PaW701 with the ability to grow on 2-methylnaphthalene arose at a frequency of about 10−5. These fell into two groups. Group A mutants had no detectable salicylate hydroxylase activity and accumulated salicylate from naphthalene in culture supernatants: they appeared to grow on the pyruvate released from oxidation of the first ring of both substrates. Their plasmids all contained a 16·7 kb insert in different sites within a small, limited region of the plasmid. Group B mutants used a meta pathway for catabolism of naphthalene and 2-methylnaphthalene. Their plasmids had undergone a small deletion of from 1·2 to 1·6 kb in a region of the plasmid close to the sites of the insertions in the group A mutants.
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A Genetic System for Pachysolen tannophilus, a Pentose-fermenting Yeast
More LessThe genetics of Pachysolen tannophilus, a yeast able to ferment xylose to alcohol, was investigated. This yeast is a strongly homothallic organism in which the haplophase is predominant. Typically, diploidy is a consequence of fusion of two mitotic products within a specialized conjugant cell. The diploid nucleus proceeds directly to meiosis and the production of a four-spored ascus. A technique for isolating both homozygous and heterozygous diploids has been devised. The species is amenable to tetrad analysis, and is well-suited to the study of pentose fermentation in yeast.
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Constitutive and Gamma Ray Modified Uptake of Labelled Precursors into the DNA of Dictyostelium discoideum during Development
More LessThe uptake of the labelled precursors, thymidine, deoxyadenosine and adenine, into the nuclear and mitochondrial DNA of the cellular slime mould, Dictyostelium discoideum, at various stages of development was studied. Labelling of the different species of DNA, nuclear main-band, mitochondrial, and satellite, was analysed by CsC1 gradients using the AT-specific drug netropsin to enhance the density resolution. Constitutive and gamma ray modified uptake patterns were obtained. During early development, through late aggregation, constitutive uptake of thymidine and deoxyadenosine was exclusively into DNA at the density of mitochondrial and nuclear satellite I DNA, believed to be primarily due to mitochondrial DNA labelling. Labelled adenine was not incorporated into any DNA before early culmination. During the period of early culmination uptake of all three of these precursors into the main-band nuclear DNA increased dramatically, to considerably exceed uptake into the mitochondrial DNA. The molecular basis for these constitutive uptake patterns during development is not understood, but they appear to involve developmentally associated periods of DNA replication in some or all of the cells, possibly accompanied by changes in precursor transport and/or pool sizes that are different for the mitochondrial and nuclear DNA metabolic pathways. Early in development gamma rays had little effect on the constitutive uptake of precursor into the mitochondrial DNA but induced new dose-dependent uptake of thymidine and deoxyadenosine into the nuclear DNA. This unscheduled nuclear DNA synthesis may be a manifestation of gamma ray induced repair replication. At the early culmination stage gamma rays severely depressed the constitutive uptake of thymidine, deoxyadenosine and adenine into the nuclear DNA, to a level of about 10% at 20 krad. Higher doses induced a slight increase over this level. These changes may be due to the inhibition of the constitutive semiconservative replication by low gamma ray doses with the superposition of some induced repair replication.
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Phages C-2 and J: IncC and IncJ Plasmid-Dependent Phages, Respectively
More LessPhages C-2 and J were isolated from sewage. Phage C-2 was filamentous and formed plaques on Salmonella typhimurium strains carrying various C plasmids. It also plated on Proteus mirabilis and Serratia marcescens strains carrying particular C plasmids, but failed to form plaques on lines of Escherichia coli K12 strains harbouring most of these plasmids, although in all cases, phage multiplication on the strains was demonstrated. No phage increase occurred in any strain which lacked a C plasmid or contained plasmids of other incompatibility groups. The phage was sensitive to chloroform and, unlike other filamentous bacterial viruses, adsorbed to shafts of conjugative pili. It had a disc-like structure at the end which attached to the pilus. Phage C-2 had a buoyant density of 1·30 g cm-3 and a single-stranded circular DNA genome of 3·0 MDal. Phage J had an hexagonal head with an inter-apical distance of 40 nm and a short non-contractile tail. It was resistant to chloroform and diethyl ether. The phage formed plaques or propagated on E. coli strains harbouring some IncC plasmids and all IncJ and IncD plasmids tested. The phage did not form plaques but propagated on P. mirabilis and Ser. marcescens strains carrying these plasmids. It did not plate or propagate on S. typhimurium strains harbouring the plasmids. The plaques were very hazy and variable in size. The phage attached sparsely, at a site which appeared to be located at the base of the tail, to sides of conjugative pili.
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- Pathogenicity And Medical Microbiology
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Effect of Glucose and Amino Acids on Expression of K99 Antigen in Escherichia coli
More LessK99 antigen production by enterotoxigenic Escherichia coli strains of bovine origin was investigated by slide agglutination and in vitro attachment to intestinal villi. Work with two strains (B41 and B44) showed that on minimal medium M2, K99 antigen was not repressed by a high concentration of glucose (2%, w/v). Growth on synthetic or complex medium did not affect K99 antigen detection, which was independent of capsular antigens, and its synthesis was not repressed by Casamino acids or glucose. A survey of 12 strains revealed two groups: in one group K99 antigen production was constitutive on basal medium without glucose, and in the second group K99 antigen was produced only in the presence of glucose. Immunoelectrophoresis patterns, and the results of slide agglutination and attachment tests, were dependent upon K99 type, whereas haemagglutination patterns were not.
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Interrelationships Between Physical State, Phenotypsic Stability and Transferability of β-Lactamase Genes in Haemophilus influenzae
More LessWe have evaluated 66 β-lactamase-producing (β-Lac+) clinical isolates of Haemophilus influenzae for the presence of plasmid DNA, ability to conjugate with an H. influenzae type b recipient, and stability of the β-Lac+ phenotype. Of the 66 strains, 23 (35%) contained a plasmid demonstrable by either of two methods of cleared cell lysate preparation. The molecular weight of 17 of the plasmids was approximately 30 × 106. Four strains carried a plasmid of mol.wt 3 × 106 and single strains carried plasmids of mol. wt 36 × 106 or 50 × 106. Of 23 plasmids that were unstable in vitro, 10 (43%) had molecular weights of 30 × 106 and coded only for β-lactamase. Stability of the β-Lac+ phenotype was observed in all 43 strains in which plasmid DNA was not demonstrated by physical methods. When β-Lac+,‘plasmid-free’ strains were conjugated with strain Eagan (H. influenzae type b), ampicillin-resistant transconjugants were isolated in 21 out of 43 matings (the lower limit of detection was 10−7 per donor cell per 30 min mating). With few exceptions, homologous (type b × type b) crosses with β-Lac+, plasmid-containing donors were more efficient (frequency 10−2–10-3) than heterologous (untypable × type b) matings (frequency 10−6–10−7). β-Lac+,‘plasmid-free’ strains were inefficient donors (frequency 10−6–10−7) irrespective of serotype, but all transconjugants from these crosses were efficient donors (frequency 10−1–10−4) and contained a plasmid of mol. wt 30 × 106 that was unstable in 9 cases out of 26 (35%). The increase in transferability and phenotypic instability of plasmid-bearing β-Lac+ strains reinforces the concept of a chromosomal locus for β-lactamase genes in‘plasmid-free’ β-Lac+ strains. Carriage of β-lactamase genes on 30 × 106 mol. wt R plasmids aids transferability at the expense of genetic stability.
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Enhancement of Non-specific Resistance to Pseudomonas Pneumonia by a Synthetic Derivative of Muramoyl Dipeptide in Immunosuppressed Guinea Pigs
More LessA synthetic derivative of muramoyl dipeptide, 6-O-stearoyl-N-acetylmuramoyl-l-alanyl-d-iso-glutamine [L18-MDP(A)], showed a protective effect against bacteraemic and non-bacteraemic pneumonia caused by Pseudomonas aeruginosa in immunosuppressed guinea pigs. In about half of the animals treated with the compound before infection, death from bacteraemic pneumonia produced by intratracheal inoculation of P. aeruginosa was delayed for 7 d, although all of the animals infected without prior treatment with the compound died within 4 d of infection. Multiplication of the organisms in the lung was also suppressed for at least 10 d by treatment with the compound when the animals inhaled an aerosol of P. aeruginosa. In contrast, in untreated animals the numbers of bacteria in the lung gradually increased from 106 to 109 c.f.u. g−1, and a few animals in which the organism increased to 109 c.f.u. g−1 had died by 6 and 10 d after infection. In both healthy and immunosuppressed animals, the accumulation of polymorphonuclear leukocytes (PMNs) in a subcutaneous air-pouch injected with heat-killed organisms was augmented by subcutaneous treatment with L18-MDP(A) 1 d before bacterial injection. The phagocytic activity of peritoneal PMNs was also increased by treatment with this compound. The augmentation of protective mechanisms against pseudomonas pneumonia by L18-MDP(A) may be attributed at least partly to the increased chemotactic and phagocytic activity of PMNs.
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Antibacterial Effect of the Scandium and Indium Complexes of Enterochelin on Escherichia coli
More LessEnterochelin, the iron chelator produced by a number of pathogenic enterobacteria, appears to be an essential metabolite for multiplication within the host, where it transports iron from the host iron-binding proteins to the bacteria. Previous work showed that complexes of enterochelin containing either scandium (Sc3+) or indium (In3+) exerted a bacteriostatic effect on Klebsiella pneumoniae in serum, whilst the Sc3+ complex exerted a significant therapeutic effect on mice infected with K. pneumoniae. These observations have now been extended to a number of pathogenic serotypes of Escherichia coli including those carrying either the K1 antigen or the ColV plasmid. The Sc3+ and In3+ complexes each exert a bacteriostatic effect on these organisms growing in either whole serum or media containing an iron-binding protein. Evidence is presented that the Sc3+ complex may act as a competitive inhibitor of the Fe3+ complex. In contrast to their effects on K. pneumoniae, sideramines other than enterochelin fail to reverse the bacteriostatic effect of the Sc3+ complex of enterochelin in E. coli, suggesting that the complex produces a more profound derangement of metabolism in this organism. The Sc3+ complex exerts a significant therapeutic effect on E. coli infections in mice although the In3+ complex is less active.
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Dimethylation of Adenine and the Resistance of Streptomyces erythraeus to Erythromycin
More LessThe erythromycin-producing organism, Streptomyces erythraeus, is highly resistant to erythromycin and also to other members of the macrolide, lincosamide and streptogramin type B (MLS) group of antibiotics. Resistance results from the presence of a single residue of N 6,N 6-dimethyl-adenine in S. erythraeus 23S rRNA. The enzyme which produces this residue (‘the erythromycin-resistance methylase’) has been isolated and characterized in vitro.
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- Physiology And Growth
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Correlation between Catabolite Repression of Arginine Transport and Repression of Anabolic Ornithine Carbamoyltransferase in Pseudomonas putida
More LessSynthesis of anabolic carbamoyltransferase (EC 2.1.3.3.) of Pseudomonas putida A90 was repressed two- to five-fold when arginine was added to cultures growing at the expense of a readily utilizable substrate, such as asparagine; synthesis was repressed 16- to 17-fold when arginine was the sole carbon or carbon and nitrogen source. The most likely explanation is that asparagine exerted some form of catabolite repression on arginine transport. Bacteria grown in the presence of asparagine plus arginine transported arginine at only 13 % of the rate of bacteria grown at the expense of arginine alone.
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The Requirement of Oxygen for the Active Transport of Sugars into Yeasts
More LessThe inducible systems of uptake of β-d-galactopyranosides into Kluyveromyces fragilis, Kluyveromyces marxianus and Debaryomyces polymorphus were studied under aerobic and anaerobic conditions, using both lactose and the non-metabolized analogue, methyl 1-thio-β-d-galactopyranoside (TMG). A common carrier served both substrates, and aerobic entry was by active transport, involving proton symport. The rate of uptake and the final equilibrium concentration were decreased on adding N,N′-dicyclohexylcarbodiimide, diethylstilbestrol or antimycin A, and active transport was completely abolished by carbonyl cyanide m-chlorophenylhydrazone. Uptake under anaerobic conditions differed markedly from that occurring aerobically: TMG was not concentrated anaerobically, even by strongly fermenting yeasts. Instead, it was transported into the cells by facilitated diffusion, which could sustain a rate of entry of over half the maximum rate observed aerobically. That this difference between aerobic and anaerobic transport into yeasts might apply to glycosides in general, was suggested by the finding that a strain of Saccharomyces cerevisiae also took up maltose by active transport under aerobic conditions, but by facilitated diffusion anaerobically. By contrast, the amino acid, 2-aminoisobutyric acid, was concentrated by Kluyveromyces fragilis, even under anaerobic conditions. The significance of these findings in relation to the Kluyver effect is discussed.
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Interaction of Polygalactosamine with Conidia of Neurospora crassa
More LessNeurospora crassa produces a phase-specific cationic mucopolysaccharide composed primarily of galactosamine (galactosaminoglycan mucopolysaccharide; GAG-MP), which becomes part of the cell wall and is later secreted into the medium. Its appearance coincides with the onset of the restricted phase of growth, and causes efflux of small molecular weight metabolites when incubated with conidial cells. This activity may be a product of electrostatic interactions of the GAG-MP with the conidial plasma membrane. The activity is blocked by acetylation of the primary amines on the molecule, digestion by enzymes that hydrolyse carbohydrate linkages and the inclusion of NaCl (1 m) in the GAG-MP/conidia reaction mixture. The physiological activity of the GAG-MP mimics that of known depolarizing agents. A model is proposed in which GAG-MP depolarizes the plasma membrane at the onset of the restricted phase of growth, stimulating a cell surface enzyme to produce endogenous cyclic AMP which in turn switches on the enzymic and genetic machinery necessary for that phase of growth.
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The Effect of Organic Compounds on the Encystment, Viability and Germination of Zoospores of Phytophthora cinnamomi
More LessA number of organic compounds were tested to determine their ability to induce encystment or germination of zoospores of the fungus, Phytophthora cinnamomi. The basic amino acids, lysine and arginine, induced encystment at concentrations in the millimolar range. Pectin (500 μg ml−1) also induced encystment and in addition stimulated the encysted cells to germinate. A wide range of carbohydrates and the amino acids, glutamate and aspartate, stimulated cysts to germinate although they did not themselves induce encystment.
The polypeptide, poly-l-lysine and the basic proteins, histone 1, histone 4 and lysozyme together with the lectin, concanavalin A, immobilized swimming zoospores at concentrations between 0.3 and 10 μg ml−1. At concentrations below 1 μg ml−1 cysts were formed but at higher concentrations cells lysed and the viability of the population was reduced. These compounds did not induce germination of the encysted zoospores. Ethanol, isovaleraldehyde and hexanal also induced encystment but only at concentrations in the millimolar range.
These data, together with previous work on cations show that the triggering of motile cells to encyst does not in this species invariably result in the cells being committed to germination.
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Characterization of Cytochromes in Lithotrophically and Organotrophically Grown Cells of Thiobacillus A2
More LessThe dominant cytochrome in thiosulphate-grown Thiobacillus A2 was found to be of the c-type with a reduced α-band at 548 nm (c 548). This c 548 component did not constitute an integral part of the membrane carrier system. It did, however, appear to be part of a large complex not tightly bound to membranes. Reconstitution experiments showed that cytochromes of the membrane ‘abc’ system could be reduced by the c 548 component and vice versa. The reduction of membrane cytochromes of either lithotrophic or organotrophic origin by thiosulphate electrons was achieved, but it required the presence of a soluble fraction containing cytochrome c 548. Evidence tending to rule out a reductive cleavage as the first step of the thiosulphate oxidation pathway in Thiobacillus A2 was obtained by following the reduction of partially purified c 548 particles under various conditions.
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Toxic Effect of Manganese on Growth and Sporulation of Bacillus stearothermophilus
More LessThe growth rate of Bacillus stearothermophilus cells in a chemically defined medium was inversely proportional to the concentration of Mn2+ between 15 and 300 mu;m. In the presence of >100 mu;m-Mn2+, cells grew with doubling times of >60 min. Cessation of exponential growth due to a high concentration of Mn2+ was most pronounced in cultures low in sulphur or carbon. This was due to the toxic effect of high Mn2+ concentrations since dilution of cultures to a final Mn2+ concentration of 15 mu;m or less restored the growth rate to maximum. Sporulation depended upon the nature of the growth-limiting nutrient. The manganese effect on sporulation of sulphur-limited cells depended upon the concentration of glucose and the aeration rate in a qualitatively similar manner. The highest spore yield at optimal aeration rates was obtained when the initial Mn2+ concentration in the medium was 10–30 μm. Although the growth rate of bacteria in low-sulphate media was determined by the manganese concentration, its effect on sporulation frequency was independent of the growth rate.
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Dependence of Bacillus stearothermophilus Spore Germination on Nutrient Depletion and Manganese
More LessSpores of Bacillus stearothermophilus NCTC 10003 germinated without delay in glucose/glutamate chemically defined medium at 60 °C. The nature of the nutrient limitation inducing sporulation affected the ability of spores to germinate under the conditions studied. Spores produced after glucose depletion of the medium (carbon-depleted spores) germinated faster and to a greater extent than did spores produced after sulphate depletion of the medium (sulphur-depleted spores). Sub-lethal heat treatment (80 °C, 10 min) prior to germination enhanced dormancy. l-Alanine (0·2 mg ml−1) did not reverse this effect, nor did it enhance the germination of sulphur-depleted spores. The rate and extent of germination for both kinds of spores was proportional to the intrasporal manganese concentration. In contrast, extrasporal manganese exerted an inhibitory effect and it was not required for germination. We conclude that the nature of the nutrient depletion and the level of intrasporal manganese distinctly affect the extent and rate of spore germination.
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Glucose Repression of Carbon Source Uptake and Metabolism in Streptomyces coelicolor A3(2) and its Perturbation in Mutants Resistant to 2-Deoxyglucose
More LessA newly devised method to obtain diffuse growth of Streptomyces coelicolor A3(2) in liquid minimal medium was used to study glucose repression. Although diauxic growth was not obtained, glucose repression of uptake of 14C-labelled carbon sources was demonstrated. Active, arabinose-induced, arabinose transport was repressed at the level of transcription by glucose. Of two glycerol-inducible glycerol transport systems, one was glucose-inhibited but not repressed (and operated by facilitated diffusion), whilst the other (an active transport system) was glucose-repressed. Active transport systems for galactose and fructose which did not require induction by their respective sugars were both inhibited by glucose. Galactose- and fructose-metabolizing enzymes were inducible by the respective sugars, but only in the absence of glucose. This was because glucose both inhibited galactose and fructose transport and repressed the metabolic enzymes concerned. Constitutive active glucose uptake was also demonstrated in arabinose-grown cells. Mutants that grew on arabinose or glycerol in the presence of 2-deoxy-glucose were glucose-derepressed for both soluble carbon source utilization and extracellular agarose. Three glucose-derepressed mutants were studied in detail. One of these could not utilize glucose (and probably lacks glucose kinase), whilst the other two could utilize glucose to differing degrees.
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The Differentiation Responses of Dictyostelium discoideum Amoebae at Various Times During Synchronous Growth
More LessSynchronous cultures of the cellular slime mould, Dictyostelium discoideum, were prepared by a selection method which involved low-speed centrifugation of exponential cultures. The method gave a relatively high degree of synchronous growth and cell doubling and a discrete phase of DNA synthesis was discernible. Amoebae which were induced to sporulation at different times during synchronous growth differed in their differentiation responses. Those which were about to divide produced more fruiting bodies and had higher levels of α-mannosidase and N-acetylglucosaminidase than cells removed to sporulation conditions at other times during synchronous growth.
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Acetylene as an Inhibitor of Methanogenic Bacteria
More LessGrowth of six pure cultures of methanogens was inhibited by low concentrations of dissolved acetylene (C2H2); other archaebacteria (three Halobacterium species) and several eubacteria were not similarly affected. The minimum concentration of dissolved C2H2 required to inhibit growth of Methanospirillum hungatei completely was about 8 mu;m; dissolved ethylene at 20 μm had little effect on growth. Dissolved acetylene (33 mu;m) did not alter the E h of the medium, or result in a loss in viability of M. hungatei after 16 h exposures. In anaerobic cell extracts of M. hungatei, activities of hydrogenase, NADP reductase, methyl-coenzyme M reductase and ATP hydrolase were not inhibited by C2H2 concentrations several times higher than those required for growth inhibition. The intracellular ATP content of all of the methanogens dropped dramatically on exposure to C2H2. Moreover, cells of M. hungatei and Methanobacterium bryantii on exposure to C2H2 lost their ability to maintain a transmembrane pH gradient. We suggest that exposure to C2H2 results in a decline in methanogen functions which require a H+-flux, including ATP synthesis, Ni2+ uptake and methanogenesis.
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- Short Communication
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Invasion by Toxoplasma gondii of ATP-depleted and ATP-restored Chick Embryo Erythrocytes
More LessChick embryo erythrocytes (CEE) were treated with NaF and KCN to deplete their ATP levels, and their susceptibility toward Toxoplasma gondii invasion was examined. A marked reduction in the invasion rate was observed in ATP-depleted CEE but the rate was recovered in ATP-restored CEE. These results indicate that T. gondii invasion is dependent on the ATP level of the CEE. Consequently, energy-dependent host cell activity is considered to be prerequisite for the invasion of T. gondii.
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- Taxonomy
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Determination of the Gram Type Using the Reaction Between Polymyxin B and Lipopolysaccharides of the Outer Cell Wall of Whole Bacteria
More LessThe polymyxin B-dependent formation of protrusions (blebs) of lipopolysaccharides on the cell wall of whole bacteria is suggested as a specific reaction for the determination of the Gram type of newly isolated bacteria. A total of 35 different bacterial species from 27 genera were studied as well as the influence of various test parameters. The reaction was specific: all 13 species of the positive Gram type gave a negative reaction, but all bacteria of the negative Gram type gave a distinct positive reaction.
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Distribution of Cytochromes in Selected Species of Corynebacteria Pathogenic to Animals
More LessEvidence is presented for the first time for the presence of a b-type cytochrome in extracts of Corynebacterium renale, C. bovis, C. kutscheri and C. pseudotuberculosis, and b- and c-type cytochromes in extracts of Rhodococcus equi (syn. C. equi). Previous observations on the presence of a b-type cytochrome in C. pyogenes were extended.
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Differentiation between the Genera Rhodococcus and Nocardia and between Species of the Genus Mycobacterium by Susceptibility to Bleomycin
More LessSusceptibility to bleomycin proved to be a useful criterion for differentiating Rhodococcus from Nocardia, the former being susceptible and the latter resistant to this agent. The test was also useful for discriminating between various species of the genus Mycobacterium.
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