Showing papers in "Folia Microbiologica in 1974"

Isolation and screening of microorganisms dissolving low-grade rock phosphate.

Abstract: Several yeasts, fungi and bacteria isolated from the rhizosphere of leguminous crops and soils of rock phosphate deposit area were found to solubilize low-grade Mussorie rock phosphate. Of the several yeasts and fungi,Schawanniomyces occidentalis, Aspergillus awamori andPenicillium digitatum were better than others in rock phosphate solubilization. Among bacterial isolates from soils of rock phosphate deposits, Gram-negative motile rods were more effective than Gram-negative non-motile rods in dissolving rock phoshates. The most efficient bacteria were identified as strains ofPseudomonas striata. All the microorganisms acidified the liquid medium but there was no relationship between the rock phosphate dissolved and the decrease in pH of the culture broth.

Studies on methanol-oxidizing yeasts. I. Isolation and growth studies.

Abstract: The yeast strain 11Bh was studied from the aspect of qualitative and qunatitative composition of lipids formed in cells during growth on methanol, synthetic ethanol and glucose. The strain was found to form some 3% free fatty acids toward the end of the growth phase. More esterified fatty acids are formed on ethanol and glucose (2.75 and 2.86%, respectively) than on methanol (1.6%). The composition of lipids and representation of the various fatty acids in the lipids is similar on all three substrates. The cell lipids contain over 40 rel.% oleic and about 16 rel.% each of palmitoleic, palmitio and linolenic acid. Odd-numbered fatty acids are present after growth on any of the three substrates, amounting to at most 4 rel.%. Of the extracellular fatty acids formed toward the end of growth of cells on methanol, propionic and acetic acid occurred in largest amounts in the medium.

Production of nematode-attracting and nematicidal substances by predacious fungi

Abstract: Arthrobotrys conoides Drechsler,Arthrobotrys oligospora Fressenius andMonacrosporium rutgeriensis R. C. Cooke, Pramer belong to the peculiar group of predactious fungi which trap and kill nematodes. We have found that these cultures produce nematode-attracting and nematicidal substances the production of which is potentiated in the presence of nematodes. Our method of nematode attraction assay is also described.

Effect of growth rate on the glucose metabolism of yeast grown in continuous culture. Radiorespirometric studies.

Abstract: Specifically labelled14C-d-glucose was used to estimate the percentage participation of glycolysis and pentose phosphate cycle in the glucose catabolism ofCandida utilis andSaccharomyces cerevisiae. The two yeasts were cultivated at various growth rates (0.1 to 0.5 h−1) in a chemostat on synthetic medium limited with glucose under aerobic conditions. The results show a considerable increase in the percentage participation of pentose phosphate cycle in the glucose catabolized bySaccharomyces cerevisiae with the increase in specific growth rate. However, inCandida utilis, the specific growth rate does not influence significantly the part of glucose catabolized via pentose phosphate cycle, but its absolute values are relatively higher than inSaccharomyces cerevisiae. A rough quantitative estimate indicates that a maximum of 60 to 72% of the assimilated glucose is catabolized through the pentose phosphate cycle while inSaccharomyces cerevisiae the percentage participation of the pentose phosphate cycle varies from 24 to 60% (maximum) and 9 to 34% (minimum).

Immunochemical studies on mannans of the genusSaccharomyces

Abstract: Structural and immunochemical studies were carried out on mannans isolated from various species of the genusSaccharomyces. It has been found that mannans ofSaccharomyces cerevisiae, Saccharomyces bayanus, Saccharomyces chevalieri, Saccharomyces italicus, Saccharomyces logos, Saccharomyces oviformis, Saccharomyces uvarum andSaccharomyces willianus are structurally and antigenically closely related, they gave high cross-reactions with anti-Saccharomyces cerevisiae serum. Mannotetraosa was found to be the immunodominant group of these mannans. Mannans ofSaccharomyces rouxii andSaccharomyces rosei showed different structural features and immunochemical properties.

Ramihyphins—Antifungal and morphogenic antibiotics from Fusarium sp. S-435

Abstract: Fusarium sp. S-435 produces a group of related antifungal antibiotics. Three components of a mixture were isolated in a pure state from the mycelium and cultivation medium of the productive strain: ramihyphins A, B and C. Another two components (D and E) were demonstrated by thin-layer chromatography. Ramihyphin A is the main component of the mixture. Its basic physical and chemical properties are presented. The antibiotic is not active against bacteria, yeast-like microorganisms, oomycetes and several protozoa. It inhibits the growth of higher filamentous fungi, stimulating ramification of hyphae when used at subfungistatic concentration. Ramihyphin A does not inhibit germination of conidia of sensitive fungi but stops their growth after their germination.Blastomyces dermatitidis, Coccidioides immitis andHistoplasma capsulatum belong to most sensitive pathogenic fungiin vitro. Ramihyphin A inhibits considerably the growth of the following phytopathogenic fungi:Stereum hirsutum, Neurospora sitophila, Botrytis cinerea, Monilia fructigena andAlternaria tenuis. The antibiotic exhibits antifungal effects alsoin vivo and is relatively little toxic for HeLa cells.

Inhibition of coagulase activity and growth of Staphylococcus aureus by garlic extracts.

Abstract: Garlic extract (1 mg dry weight/ml) produced an inhibition of the coagulase reaction and increased the time of coagulation by a factor of 1.5, whereas 4 mg dry weight/ml increased the coagulation time factor by 2.75. At this latter concentration of garlic, coagulation was only partially complete at 4 h. Garlic extract (1.4 mg dry weight/ml) reduced growth in nutrient broth whereas 5.6 mg dry weight/ml was completely inhibitory. These effects were not observed until 8 h after exposure of the organisms to the garlic extracts. There were 5.9% more survivors among garlic treated mice compared to non-treated animals, but this difference is not significant.

Requirements of Acholeplasma laidlawii A, strain LA 1, for nucleic acid precursors.

Abstract: The requirements for nucleic acid precursors byAcholeplasma laidlawii type A, strain LA 1, were studied. The absence of a requirement for ribose or deoxyribose and the interconvertibility of uridine and cytidine contrasts with previously reported results with strain pG 8. Growth of the LA 1 strain was only moderately inhibited by a ten-fold excess of RNA over DNA. The only required nucleic acid precursors were guanine, thymine and either cytosine or uracil.

D-amino acids in the cell pool of bacteria

Abstract: About 30 different bacterial species were tested for the possible presence of freed-amino acids in their cell pool. Gram-positive bacteria particularly the species of the genusBacillus have a fairly large pool of freely extractabled-amino acids. Varied quantities of freed-amino acids were detected inBacillus subtilis B3,Bacillus subtilis Marburg,Bacillus licheniformis, Bacillus brevis, Bacillus stearothermophilus, Lactobacillus fermenti, Lactobacillus delbrueckii, Staphylococcus aureus andClostridium acetobutylicum. The individual components ofd-amino acids were identified in 5Bacillus species referred to above,d-alanine is the major component; the otherd-amino acids identified are aspartic acid, glutamic acid, histidine, leucines, proline, serine and tyrosine. Thed-amino acid pool size inBacillus subtilis B3 varies with different culture conditions. The pool size is maximum when growth temperature is 30°C and it fluctuates with change in pH of the medium. The maximum quantity ofd-amino acids could be recovered when the culture was at mid log phase. O2 supply to the medium has little effect ond-amino acid pool size. The starvation of cells leads to depletion of thed-amino acid pool which is exhausted almost completely within 4 hours by incubation in nutrient-free medium.

The study of variability and strain selection in Streptomyces atroolivaceus

Abstract: A seven-step selection procedure (repeated UV irradiation, single-step application of nitrous acid, and natural selection, following each mutagenic treatment) made it possible to increase production of mithramycin byStreptomyces atroolivaceus from 40–50 μg/ml to 680–830 μg/ml,i.e. roughly by 15 to 19-fold. The UV radiation was more effective when applying lower doses, yielding about 5% survival, 2.3% survival was obtained after the treatment with nitrous acid.N-Methyl-N′-nitro-N-nitrosoguanidine applied in a buffer pH 9.0) at doses yielding more than 99% killing was less effective than the two former mutagens.

Progesterone transformations as a diagnostic feature in the generaAlternaria, Stemphylium andCladosporium

Abstract: Various species of the generaAlternaria, Stemphylium andCladosporium were shown to display a specific reaction characteristic for the given genus In theAlternaria genus this is a 1-2-dehydrogenation of the A ring of the steroid molecule, inStemphylium 14α-hydroxylation and inCladosporium 7β-hydroxylation This chemotaxonomic feature may supplement morphological and functional criteria in the taxonomy of filamentous fungi (Hyphomycetes)

Turnover of murein in cellular and filamentous populations of Bacillus megaterium.

Abstract: Bacillus megaterium grows in the form of filaments at temperatures above 45°C. The rate of turnover of the cell wall begins to decrease gradually under these conditions. At the same time sensitivity of the filamentous forms to lysozyme decreases. Filaments outgrown at 48°C retain the decreased rate of turnover of the cell wall for a certain time after transfer to 30°C, in spite of the fact that septa are formed and filaments are converted to cells. However, a population incubated longer than 2 h at 48°C often ceases to grow and the growth is not restored even after transfer to 30°C. Three clones of the asporogenic strainBacillus megaterium KM differing somewhat in their ability to form filaments at 35°C differ mutually also in the rate of turnover of the cell wall. However, the decreased rate of the turnover cannot be unambiguously correlated with the increased tendency to form filaments.

Microbial glucosidation of dihydroxyanthraquinones. General properties of the glucosidation system

Abstract: Corresponding mono-β-d-glucosides were obtained by fermentation ofStreptomyces aureofaciens B 96 with four isomeric dihydroxyanthraquinones (alizarin, quinizarin, chrysazin and anthraflavin). The effect of some factors (sugar source, concentration of substrates, pH) on biosynthesis of 1-hydroxy-2-(β-d-glucopyranosyloxy)anthraquinone was studied and optimum conditions for the microbial glucosidation were determined.

Mutagenesis by N-methyl-N-nitroso-N'-nitroguanidine in synchronized cultures of Mycobacterium phlei.

Abstract: Conditions of maximum induction of back mutations byN-methyl-N-nitroso-N′-nitroguanidine (“nitrosoguanidine”) were studied in auxotrophic mutants ofMycobacterium phlei. In asynchronous cultures the effects of pH, buffer molarity and concentration and exposure time to nitrosoguanidine were studied. It was shown that between 6 and 10, pH does not affect the induction of back mutations but that with increasing pH up to 9 the lethal effect of nitrosoguanidine on cells is increased. Protracted treatment with nitrosoguanidine or buffer molarity did not affect the induction of back mutations. It was found with several strains ofMycobacterium phlei that it is most efficient to treat a culture with 0.5 mg or 1 mg nitrosoguanidine/ml for 20 min at pH 6. On the basis of these findings a method of induction of back mutations by nitrosoguanidine was developed for populations with synchronous cell division.

Relationships between plant roots, proteolytic organisms and activity of protease

Abstract: Proteolytic bacteria represented 18–58% of the bacterial population isolated from the rhizoplane of different crops. The activity of protease was considerably higher on roots of wheat growing in the soil than in the rhizosphere or free soil. However, only a slightly positive rhizosphere effect in the relative occurrence of casein-hydrolyzing bacteria could be observed. An indirect relationship between numbers of bacteria hydrolyzing casein and the activity of the enzyme could be found. The activity of protease related to a unit of culturable proteolytic bacteria was considerably higher on the root than in the rhizosphere and in the soil. A relationship between characteristics of the production of the enzyme by proteolytic bacteria and the protease activity on the surface of roots was demonstrated. The resulting enzyme activity on the surface of roots depended apparently on growth conditions of the plant and nature of root exudates and was influenced both by inactivation and protection due to adsorption of the enzyme by roots.

Effect of glucose on vanillic acid oxidation inCellulomonas sp.

Abstract: Respirometric and radioisotopic methods were used to study the stimulating effect of glucose on the breakdown of14COOH-vanillic acid by aCellulomonas species. Glucose was found to reduce the lag phase in vanillic acid oxidation apparently by enabling vanillic acid to cross the cell membrane and thus to reach the appropriate metabolic enzymes. Glucose also eliminated the inhibitory effect of higher concentrations of vanillic acid on the breakdown process and stimulated incorporation of the carboxyl carbon from vanillic acid into cell structures. A similar effect was observed with succinate and acetate.

Antimicrobial agents. XX. Ergosterol content of Candida albicans cells during adaptation to antimycotics.

Abstract: Adaptation is a fundamental property by means of which microbial cells react to adverse environmental conditions. In transiently resistant strains of derm~tophytes adapted to antimycotics, we demonstrated a decrease in the ergosterol content of the cells, as well as loss of sensitivity (~apek and Simek, 1972). We also studied these two factors in strains of Candida albicans obtained from the Clinic of Skin and Venereal Diseases of the Faculty of Medical Hygiene, Charles University, Prague, which were adapted by the gradient plate method (Szybalski and Bryson, 1952) to the antimycotic VUFB-9244, i.e. 2-methylthio-5-(3-idopropargyloxy) pyrimidine:

Effect of proteases, phospholipases and polysaccharide-splitting enzymes on plasma membrane particles and on the synthesis of the fibrillar cell wall component in yeast protoplasts.

Abstract: Plasma membrane particles demonstrable by the freeze-etching technique, play, according to some authors, a role in the cell wall synthesis. On a model of yeast protoplast capable of regenerating the cell wall we studied the morphology of plasma membrane particles and the synthesis of the fibrillar cell wall component following a treatment with various enzymes and with lysolecithin. The enzymes used included proteases (trypsin, papain, pronase), polysaccharide-splitting enzymes (snail enzyme complex, mannosidase), phospholipases (A, C, D) and lipase. Upon treating living protoplasts with these substances in no case did we observe any morphologically demonstrable change in the particle structure or in their distribution in the plasma membrane. The fibrillar cell wall component was synthetized even in the presence of proteases and phospholipases. If the plasma membrane particles are assumed to represent enzyme systems synthesizing the cell wall component then in living protoplasts they are not located on the outer plasma membrane face or else are protected by some mechanism against the action of the corresponding enzymes.

Effect of derivatives of 3-quinolinecarboxylic acid on DNA synthesis, growth and division in Escherichia coli 15 TAU.

Abstract: The inhibitory effect of derivatives of 3-quinolinecarboxylic acid on replication of DNA, growth, division and colony-forming ability ofEscherichia coli 15 TAU was compared with the effect of nalidixic acid. Oxolinic acid was found to be most effective. It brings about an immediate inhibition of DNA replication even at a concentration 10-times lower than that of nalidixic acid. The importance of 6,7-methyleneoxy-, 1-ethyl and 3-carboxyl groups on the quinoline ring for maximum effectivity of the preparation was verified. The question of the primary importance of the inhibition of replication for antibacterial effects is discussed.

The effect of microorganisms and sorption on the protease activity of plant roots.

Abstract: The protease activity of sterile roots of wheat was zero or very low, so that the determined values did not exceed limits of the experimental error. Roots colonized by microorganisms had a significant protease activity. The activity of protease on seeds and roots of the plants growing in a medium inoculated with the soil microflora was higher than in cases when only the epiphytic microflora of seeds served as a source of microorganisms. Sterile roots inoculated with three different strains of bacteria isolated from the rhizosphere and producing protease exhibited a considerable protease activity. The protease activity of non-sterile roots of plants growing in the dark was higher than that of plants growing under normal illumination. Crystalline proteinase was adsorbed on sterile roots and the activity of the enzyme was decreased in this adsorbed state. The adsorption of the enzyme was only slightly higher in the presence of calcium ions. Treatment of roots with a sodum chloride solution, with dextran and ethanol increased the adsorption of the proteinase by roots.

Mutarotase in galactose-induced baker's yeast.

Abstract: Cell-free extracts of baker's yeast possess mutarotase activity only after induction of cells in the presence of galactose. The mutarotase activity appears 1 h after transfer to a galactose-containing medium and rises in synchrony with the utilization of galactose. Cycloheximide blocks the induction completely at a concentration of 100 μg/ml. InSaccharomyces fragilis the mutarotase is constitutive but its activity is strikingly increased after growth on galactose. The yeast mutarotase resembles in some respects analogous enzymes from other cells (pH dependence, substrate specificity, heat lability). Its affinity ford-galactose is substantially greater than ford-glucose. There may exist a coupling between mutarotase activity and the anomer-specific galactokinase.

Microbial decomposition of coumarin in soil.

Abstract: Microbial decomposition of coumarin was studied in samples of chernozem soil by manometric measurement of oxygen consumption, paper chromatography of aromatic metabolic intermediates in soil extract and measurement of their UV spectra, and by the technique of simultaneous adaptation. Coumarin is decomposed in soil viao-coumaric and melilotic acids and at least one other compound of aromatic character. The metabolic pathway including salicylic acid and catechol was not proved. A total of 39 strains of coumarin-decomposing bacteria were isolated from the soil, out of which 25 belong to the genusPseudomonas, 7 to the genusCellulomonas and 7 to the genusAchromobacter. A comparison of the counts of bacteria utilizing coumarin as a sole carbon source in garden soil, in two chernozem soil samples and in acidic brown soil showed that their occurrence bears no relation to the so-called total number of bacteria (grown on agar medium with yeast and soil extracts and with tryptone) or to the content of carbon and nitrogen in the soil, or to its acidity.

Characterization of degradation products of the cell wall released during growth and sporulation of Bacillus megaterium.

Abstract: Asporogenic and sporogenic strains ofBacillus megaterium KM release during growth heterogeneous fragments of the cell wall into the medium the non-dialyzable fraction representing 50–90% by the total. During lysis of sporangia the non-dialyzable fraction represents only 30% of the soluble fraction of autolyzed walls. Gel filtration on Sephadex permits to separate the non-dialyzable fragments of the cell wall released during growth into two fractions contaning simultaneously peptidoglycan and phosphorus. The two fractions contain peptidoglycan components in the same ratio as in the cell wall. Only one peptidoglycan macromolecular fraction, smaller than the fractions released during growth, was detected by gel filtration in the material released during lysis of sporangia.

DNA synthesis in pretreated and ultraviolet-irradiated cells ofEscherichia coli B/rHcr+ andHcr−

Abstract: DNA synthesis after the ultraviolet irradiation was followed in the excision proficient strainEscherichia coli B/rHcr +, in which the ability to excise thymin dimers was suppressed by a preirradiation inhibition of DNA and protein syntheses and in the excision deficient strainEscherichia coli B/rHcr −. Synthesis of pulse-labeled DNA, its stability and semiconservative DNA synthesis were compared in both strains. It was found that cells of theHcr + strain restore semiconservative DNA synthesis and the pulselabeled DNA appears stable, in spite of the fact that dimers are not excised under these conditions. On the other hand, cells of theHcr − strain are unable to restore semiconservative DNA synthesis and the pulselabeled DNA is degraded. As the repair by the excision of dimers under the used experimental conditions may be excluded in both strains, it is possible to assume that activity of enzymes included in theHcr + marker is prerequisite for restoring the DNA synthesizing system in theHcr + strain.

Carbon sources forAspergillus niger growth under different shaking programmes

Abstract: In a 36-h shaken submerged culture ofAspergillus niger M1, maltose supported growth substantially more than either sucrose, glucose or fructose. If the culture was kept static for 6–36 h, the differences gradually disappeared. Lactose does not serve as growth substrate.

Changes of microflora of wheat roots after foliar application of urea.

Abstract: Upper parts of wheat grown in containers with brown soil were four-times repeatedly treated with a solution of urea. The foliar application of urea resulted in a considerable increase of the number of bacteria in the roots and a decrease of the number of fungi. Seventeen and 13 genera of fungi could be detected in the rhizosphere of the treated plants, 3 and 5 weeks, respectively, after the application, whereas 23 and 16 genera could be identified in the rhizoplane of the treated plants. Fungi of the generaFusarium andPenicillium predominated in the root region. The foliar application of urea resulted in an increased relative occurrence of the genusFusarium and decreased relative representation of the genusPenicillium. Changes in the relative occurrence of other 18 genera of fungi present in small quantities on wheat roots could also be observed. The foliar application of urea affected not only numbers of fungi determined by the dilution plate method but also the occurrence of active hyphae.

Regulation of biosynthesis of excessive metabolites. XVI. Origin of the terminal group of tetracyclines.

Abstract: Experiments with radioactive precursors showed that the terminal group of tetracyclines can originate by various pathways. Both malonylCoA originating by carboxylation of acetylCoA, and malonylCoA synthesized by alternative biosynthetic pathways, can be utilized. Radioactive malonate semiamide or its derivative was hydrolyzed to malonie acid during the fermentation and the acid was incorporated into tetracycline in the normal way. Of the other assumed precursors, U-laC-uraci). 1-14C- propionate, U-14C-alanine, 5AaC-glutamate were unspecifieally incorporated into the terminal group of tetracycline. The incorporation of 4-14C-ethyl suceinamate and 5-14C-ethyl glutaramate was negligible. The present knowledge of biosynthesis of tetracyclines is based on the discovery of condensation of malonate units giving origin to a hypothetic oligoketide chain (Gatenbeck, 1961; Mitscher, 1968). It is assumed that malonate semiamide is a terminal group of tetracycline. The malonate semiamide condensates with 8 mol- ecules of malonylCoA and the tetracycline skeleton is thus formed. As it was poss- ible to prepare 2-acetyldecarbam ido-oxytetracycl ine by adding acetoaeetate to mu- tants of streptomycetes (Hochstein et al., 1960), it may also be assumed that the enzyme system involved in the biosynthesis of tetracyclines is not specific with respect to the terminal group. In the present work we performed experiments with incorporation of labelled malonate semiamide into tetracycline in vivo and used also other substrates which could be considered as possible sources of the terminal group of tetracyclines.

Batch kinetics and oxygen consumption of Candida lipolytica 4-1 on n-alkanes.

Abstract: Detailed batch kinetics ofCandida lipolytica 4-1 onn-hexadecane for varying dispersed phase volume from 0.5 to 5% v/v is presented. All batch growth curves exhibited a linear growth region, indicating a substrate uptake limit. System productivities derived from the linear region were correlated to the dispersed phase volume. The correlation coefficient was identical with that obtained on gas oil. This implies that a correlation coefficient of interfacial area to the dispersed phase volume is identical for both systems. Dissolved oxygen profile and uptake of oxygen from gas phase were also measured. The oxygen uptake rate, volumetric oxygen transfer rate and oxygen demand (requirement) were calculated by means of the balance method. Under limiting dissolved oxygen concentration the maximal oxygen transfer of the fermenter was assessed.

Dimorphic and yeast-like mutants of the genus Cephalosporium Cda.

Abstract: A series of mutants, in which the mycelial type of growth gradually changes to the dimorphic and permanent yeast-like forms, were isolated from cultures ofCephalosporium sp. subjected to UV radiation. The intermediate stage between the mycelial and dimorphic strains (mutants 2/29 and 2/R) is characterized by the absence of aerial hyphae, ability to form conidiophores inside agar and by polymorphism of conidia. The Y-M transformation of two dimorphic mutants obtained from the 2/R mutant depends on temperature. Another mutant isolated from the 2/29 strain was found to form the mycelial phase only when osmolarity of the medium increased. At 22°C the transformation of all three dimorphic strains was influenced by the carbon source: the Y phase predominated in glucose-containing media, the M phase predominated in media with amino acids or citrate serving as carbon sources. Another mutant (2/7R) was found to grow permanently in the Y phase and was not influenced by temperature, osmolarity of the medium and by the carbon source. It is assumed that the dimorphism of the mutants is caused by a conformational mutation inhibiting the apical growth. This mutation can be phenotypically reversed by some factors of the environment.