Showing papers on "Growth medium published in 1989"

Dispersed growth of streptomyces in liquid culture

Abstract: The study of the physiology of the filamentous bacterium Streptomyces is inhibited by its formation of mycelial pellets in liquid cultures. It is demonstrated that dispersed growth may be achieved by the addition of polymers to the culture medium. Uncharged polymers, such as polyethylene glycol, are relatively ineffective but polyanions such as agar, Carbopol and Junlon produce dispersed cultures when included in a defined growth medium at low concentrations. Junlon-containing media enable optical density measurements to be used to follow batch growth of Streptomyces. Improvements in both biomass yield and product yield of the pigmented antibiotic actinorhodin were found to result from the incorporation of Junlon into minimal medium.

Ethanol production by Escherichia coli strains co-expressing Zymomonas PDC and ADH genes

Abstract: A novel operon and plasmids comprising genes which code for the alcohol dehydrogenase and pyruvate decarboxylase activities of Zymomonas mobilis are described. Also disclosed are methods for increasing the growth of microorganisms or eukaryotic cells and methods for reducing the accumulation of undesirable metabolic products in the growth medium of microorganisms or cells.

Aggregation in Azospirillum brasilense Cd: conditions and factors involved in cell-to-cell adhesion

Abstract: Aggregation of the root-inhabiting, asymbiotic N-fixingAzospirillum brasilense Cd (ATCC-29729), was studied Aggregation occurred towards the end of the exponential phase and during the stationary phase More aggregates were formed in media supplemented with organic acids than in those containing sugars as a sole carbon source Maximum growth with no aggregation was obtained in a medium containing both fructose and malate as carbon sources Aggregation was increased by poly-L-lysine and carbodiimide as well as by increasing the C/N ratio and decreasing combined nitrogen in the growth medium Aggregates were stable at pH levels of >8 and <6, but dispersed at pH 71 Treatment of Azospirillum with NaEDTA resulted in loss of both aggregative capacity and the ability of adsorb to wheat roots without losing cell viability When extracted bacteria were suspended in their dialysed NaEDTA extract, both their aggregative and adsorptive capacities were restored

Stimulation of Growth in Human and Murine Cells by Adriamycin

Abstract: Adriamycin causes a variety of biological actions and is an effective cytotoxic agent against proliferating cells. In this paper we show that the drug is not limited in its action solely to cytotoxicity, but can also stimulate cell growth under the appropriate conditions. Using the survival assay of cloning in soft agar, we present data showing that the conditions for Adriamycin-induced growth stimulation are that the drug be in a subtoxic concentration range of 10(-10)-10(-9) M (greater than 10(-8) M causes cytotoxicity) and that the growth medium be suboptimal. This latter condition is satisfied by either growing cells for an extended period in order to exhaust the growth supporting capacity of the medium, or by growing the cells at low (less than 10%) serum concentrations. Several active anthracycline congeners also have the ability to stimulate growth. The results indicate that the cytotoxic anticancer agent Adriamycin can stimulate the proliferation of some cells.

Transport and catabolism of proline betaine in salt-stressed Rhizobium meliloti

Abstract: Exogenous proline betaine (N,N-dimethylproline or stachydrine) highly stimulated the growth rate of Rhizobium meliloti, in media of inhibitory concentration of NaCl whereas proline was ineffective. High levels of proline betaine uptake occurred in cells grown in media of elevated osmotic strength; on the contrary, only low activity was found in cells grown in minimal medium. The apparent Km was 10 μM with a maximal transport rate of 25 nmol min-1 mg-1 of protein in 0.3 M NaCl-grown cells. The concentrative transport was totally abolished by KCN (2 mM), 2,4-dinitrophenol (2 mM), and carbonyl cyanide-m-chlorophenyl hydrazone (CCCP 10 μM) but was insensitive to arsenate (5 mM). Glycine betaine was a very potent inhibitor of proline betaine uptake while proline was not. Proline betaine transport was not reduced in osmotically shocked cells and no proline betaine binding activity was detected in the crude periplasmic shock fluid. In the absence of salt stress, Rhizobium meliloti actively catabolized proline betaine but this catabolism was blocked by increasing the osmotic strength of the medium. The osmolarity in the growth medium regulates the use of proline betaine either as a carbon and nitrogen source or as an osmoprotectant.

A new growth medium for the study of siderophore-mediated interactions

Abstract: A microbial growth medium, RSM, was developed to study the role of siderophores (microbial Fe-transport compounds) in the inhibition of the take-all pathogen, Gaeumannomyces graminis var. tritici, by Pseudomonas putida strain B10. The inorganic constituents of the medium were designed to mimic the rhizosphere while the organic composition was designed to promote rapid growth and siderophore production. The antibiosis experiments were highly reproducible and the antagonism appeared to be due to production of pseudobactin, the siderophore of B10. On plates amended with chrome azurol S, G. graminis did not produce siderophores while other fungi did. The growth of G. graminis on plates prepared with Fe chelate buffers was inhibited at a free ferric ion concentration of 10−24.6 M, although three other fungi were not inhibited, even at 10−25.5 M, presumably due to their greater production of siderophores. In liquid medium amended with Fe chelate buffers, both the doubling time and the lag phase of P. putida increased as the free ferric ion concentration was reduced. A wide variety of fungi and bacteria were found to grow on this medium. Because the inorganic composition of RSM is based on that of the rhizosphere, the development of this medium may be a first step towards the study of the chemistry and biology of the rhizosphere under well defined conditions.

Retroviruses expressing different levels of the normal epidermal growth factor receptor: biological properties and new bioassay.

Abstract: Two retroviral DNAs that encode the normal human epidermal growth factor (EGF) receptor hEGFR have been generated by inserting a hEGFR cDNA into two different retroviral vectors. One DNA (pCO11-EGFR-neo) also contained a linked selectable marker gene (neoR). The other (pCO12-EGFR) only expresses hEGFR. When introduced into NIH3T3 cells, the two DNAs and the viruses derived from them induced a fully transformed phenotype, including focal transformation and growth in agar or low serum, but transformation depended entirely upon EGF being present in the growth medium. Compared with pCO11-EGFR-neo, pCO12-EGFR induced EGF-dependent transformation 2-5 times more efficiently and expressed higher numbers of receptors (4 x 10(5) vs. 1 x 10(5) EGF receptors per cell). The results indicate that transforming potential is directly related to the number of EGF receptors. In defined, serum-free medium that contained only very low concentrations of insulin (0.6 microgram/ml) and transferrin (0.6 micrograms/ml), hEGFR-virus infected cells were able to grow with EGF as the only growth factor. Moreover, daily incubation of the cells with EGF for only 30 min was sufficient to induce growth. NR6 cells, which lack endogenous EGF receptors, were transformed as efficiently as NIH3T3 cells by the hEGFR virus. The dose-dependent growth response to EGF of infected NR6 cells grown in serum-free medium can be used as a highly sensitive bioassay for the quantitative assessment of EGF and transforming growth factor type alpha (TGF alpha). This bioassay is at least as sensitive as previously reported radioimmunoassays and can measure a much wider concentration range (10 pg-100 ng/ml). Uninfected NR6 cells or NR6 cells infected by helper virus alone can be used as controls for the EGF specificity of growth stimulation.

The Mechanism of Intracellular Acidification Induced by Glucose in Saccharomyces cerevisiae

Abstract: Addition of glucose or fructose to cells of Saccharomyces cerevisiae adapted to grow in the absence of glucose induced an acidification of the intracellular medium. This acidification appeared to be due to the phosphorylation of the sugar since: (i) glucose analogues which are not efficiently phosphorylated did not induce internal acidification; (ii) glucose addition did not cause internal acidification in a mutant deficient in all the three sugar-phosphorylating enzymes; (iii) fructose did not affect the intracellular pH in a double mutant having only glucokinase activity; (iv) glucose was as effective as fructose in inducing the internal pH drop in a mutant deficient in phosphoglucose isomerase activity; and (v) in strains deficient in two of the three sugar-phosphorylating activities, there was a good correlation between the specific glucose- or fructose-phosphorylating activity of cell extracts and the sugar-induced internal acidification. In addition, in whole cells any of the three yeast sugar kinases were capable of mediating the internal acidification described. Glucose-induced internal acidification was observed even when yeast cells were suspended in growth medium and in cells suspended in buffer containing K+, which supports the possible signalling function of the glucose-induced internal acidification. Evaluation of internal pH by following fluorescence changes of fluorescein-loaded cells indicated that the change in intracellular pH occurred immediately after addition of sugar. The apparent Km for glucose in this process was 2 mM. Changes in both the internal and external pH were determined and it was found that the internal acidification induced by glucose was followed by a partial alkalinization coincident with the initiation of H+ efflux. This reversal of acidification could be due to the activity of the H+-ATPase, since it was inhibited by diethylstilboestrol. Coincidence between internal alkalinization and the H+ efflux was also observed after addition of ethanol.

Toxin production by a rhizobacterialPseudomonas sp. that inhibits wheat root growth

Abstract: We studied the production of a toxin inhibitory to both winter wheat (Triticum aestivum L.) root growth andEscherichia coli that was produced by a rhizobacterial pseudomonad. Of several carbon sources tested, the most rapid growth and highest toxin concentrations were obtained with glucose, glycerol, or trehalose. Toxin production was repressed with L-cysteine as the nitrogen source. Toxin was produced during the late exponential and early stationary phase of growth by the bacterium and, contrary to studies with other toxins, was unaffected by Fe and P concentrations in the growth medium. Toxin production by the bacterium was the same at growth temperatures of 25 and 15°C while it produced less at 5°C. If the bacterium was able to grow, it produced toxin. No compound tested induced an increase in toxin production indicating toxin production is constitutive.

Induction of D-amino-acid oxidase by D-alanine in Rhodotorula gracilis grown in defined medium

Abstract: SUMMARY: The obligate aerobe yeast Rhodotorula gracilis was grown in batch culture on a chemically defined, pH-controlled medium containing glucose or d-alanine as carbon sources, ammonium or d-alanine as nitrogen sources, and d-alanine as a sole carbon and nitrogen source. Under these conditions, d-alanine induced the synthesis of d-amino-acid oxidase (EC 1.4.3.3) to an extent depending on the nutrients, the highest specific activity of the enzyme [up to 0.6 U (mg protein)−1] being detected when both d-alanine and glucose were present in the growth medium. In contrast, enzyme activity was negligible when both ammonium and glucose were present in the growth medium, even in the presence of d-alanine. The racemic mixture dl-alanine was also utilized as a source of both carbon and nitrogen for the growth of R. gracilis, but the enzyme activity appeared only after the depletion of l-alanine from the medium. Data on transmembrane transport of d-alanine in the presence of different nutrients clearly indicated that the l-isomer prevented induction by d-alanine through inhibition of the transport of the d-amino acid into cells. However, such an effect was not exerted by ammonium, indicating that this compound probably acts at the level of enzyme synthesis.

Effect of date extract on growth and spore germination of Bacillus subtilis.

Abstract: Berhi date extract was studied for its effect on the growth of Bacillus subtilis, Staphylococcus aureus, Salmonella typhi and Pseudomonas aeruginosa. From 80 to 99% inhibition was observed in nutrient broth cultures containing 20% (w/v) date extract. Spore germination of B. subtilis was inhibited completely using various concentrations of date extract. Cell elongation and depression in the cell wall of B. subtilis were seen by scanning electron microscopy as a result of incubation in a growth medium containing dates.

Impact of aeration on the metabolic end‐products formed from glucose and galactose by Streptococcus lactis

Abstract: Compared with cultures grown aerobically in batch culture with glucose, aerated cultures of lactic streptococci had a less homolactic type of metabolism when galactose was the carbohydrate source in batch cultures, or when glucose was limiting in chemostat cultures. Differences in end-products of sugar metabolism between aerated and unaerated cultures were observed. In addition to lactate, formate, acetate and ethanol were produced in anaerobic cultures, whereas acetate and acetoin were formed in aerated cultures. Acetate production in aerated cultures depended on lipoic acid, an essential cofactor of the pyruvate dehydrogenase complex. In a chemically defined medium with glucose as the energy substrate, lipoic acid (or acetate) was an essential growth factor. Formation of acetoin was inversely related to lipoic acid concentration in the growth medium. Although not observed in unaerated cultures, acetoin (and 2,3-butanediol) was produced in unaerated buffered suspensions metabolizing pyruvate. Aeration caused a modest increase in the activities of aP-acetolactate synthetase and phosphate acetyl trans-ferase, but it is unlikely that the increases were sufficient to account for the changes in end-products of sugar metabolism observed.

Changes in internal and external pH accompanying growth of Candida albicans: studies of non-dimorphic variants.

Abstract: Non-dimorphic variants of Candida albicans, which were unable to form germ tubes or mature hyphae in media containing amino acids, glucose and salts or N-acetylglucosamine or serum, were prepared from two hyphal positive laboratory strains using a physical separation method. The hyphal-minus phenotype was stable and may be due to mutations or phenotypic variation. The variant strains maintained their internal pH within narrower bounds as compared to their parental wild-types. When exposed to conditions that normally supported the induction of germ tubes the cytoplasmic pH of the wild type strains increased from 6.8 to over pH 8.0 within 5 min while in the variants the rise in internal pH was only about 0.3 pH units. The wild type strains acidified the growth medium more rapidly than the variants. The results suggest that the control of internal pH is directly or indirectly associated with the regulation of dimorphism. The variants had unaltered cell volumes and specific growth rates. The hyphal-minus phenotype was however fully reversible since revertants occurred spontaneously on serum containing agar.

The Influence of C:N Ratio in the Growth Medium on the Cellular Composition and Regulation of Enzyme Activity in Hyphomicrobium X

Abstract: SUMMARY: The regulation of the enzymes associated with one-carbon metabolism and the assimilation of nitrogen, together with the cellular composition of Hyphomicrobium X, were investigated. The effect of changing the methanol-carbon concentration with the NH+ 4-nitrogen concentration remaining constant (C: N ratio) in the medium during chemostat growth at a constant dilution rate was studied. As the medium changed from a C-limitation to a dual C- and N- and finally a N-limitation, the culture gradually passed through three definite growth phases. In response to these environmental conditions the cellular composition and the specific enzyme activity patterns changed. The C-content of the cells changed very little. The N- and protein-content was constant over C-limiting conditions, but under dual C- and N-limiting and N-limiting conditions an accumulation of poly-β-hydroxybutyrate (PHB) occurred and as a consequence the N-content and protein-content of the cells decreased. The enzyme associated with N-assimilation during C-limitation was an NADP+-dependent glutamate dehydrogenase which was replaced by the high affinity glutamine synthetase and glutamate synthase pathway immediately the NH+4-N concentration in the medium became limiting. Similarly the specific activity of methanol dehydrogenase, which was high during C-limiting conditions, dropped to a low level as the NH+ 4-N concentration decreased. Finally carbon balances were constructed throughout the experiment which showed that irrespective of the C:N ratio in the medium during C-limitation, the methanol-carbon was fluxed into biomass and CO2 only; during dual limitation the carbon was channelled into biomass, CO2 and PHB; and finally when the growth was in the presence of excess carbon no methanol-carbon was directed into over-metabolite production but, instead, the excess carbon was oxidized through the dissimilatory pathway.

Optimization of Growth Medium and Poly-$\beta$-hydroxybutyric Acid Production from Methanol in Methylobacterium organophilum

Abstract: Methylobacterium organophilum, a facultative methylotroph was cultivated on a methanol as a sole carbon and energy source. The cell growth was affected by the various components of minimal synthetic medium and the medium composition was optimized with 0.5% (v/v) methanol at pH 6.8 and at 3. The maximum specific growth rate of M. organophilum was achieved to 0.26 hr in the optimized medium which has following composition: Methanol, 0.5% (v/v):(NH)SO, 1.0g/l:KHPO, 2.13g/l:KHPO, 1.305g/ι:MgSO.7. 45g/l and trace elements (CaCl.2, 3.3mg:FeSO.7, 1.3mg:MnSO.4, 130:ZnSO.5, 40:NaMoO.2, 40:CoCl.6, 40:HBO, 30 per liter). By the limitation of nitrogen and deficiency of Mn or Fe, the cell growth was significantly repressed. Methanol greatly repressed the cell growth and the complete inhibition was observed at concentration above 4% (v/v). In order to overcome the methanol inhibition and to prevent the methanol limitation, intermittent feeding of methanol was conducted by a D.O.-stat technique. PHB production by M. organophilum was stimulated by deficiency of nutrients such as NH, SO, , , or PO in the medium. The maximum PHB content was obtained as 58% of dry cell weight under deficiency of potassium ion in the optimized synthetic medium.

Estrogen Synthetase (Aromatase) Activity in Primary Culture of Human Term Placental Cells: Effects of Cell Preparation, Growth Medium, and Serum on Adenosine 3′,5′-Monophosphate Response*

Abstract: The establishment of human term trophoblast cells in culture is dependent on the method of cell preparation, growth medium used, and presence of serum. Using freshly isolated term placental cells, we investigated 1) the effects of two different methods of removal of nontrophoblast cells and two culture media on trophoblast aromatase specific activity, cellular morphology, and hCG secretion over 72 h; and 2) the sensitivity of these hormonal parameters to cAMP added 24 h after the initiation of culture. Under conditions in which aromatase is responsive to cAMP, we studied the effect of removing serum after 24 h in culture with serum. After trypsin digestion of placental villi, isolated trophoblast cells were either treated with ammonium chloride (A) or purified on Percoll density gradients (P) and then grown in monolayer culture with medium 199 plus 10% fetal bovine serum (M) or Dulbecco’s Modified Eagle’s Medium (DMEM) plus 20% fetal bovine serum (D). Regardless of the method of treatment or growth medium...

Multiple forms of endo‐1,4‐β‐D‐glucanase in the extracellular cellulase of Penicillium pinophilum

Abstract: The influence of the composition of the growth medium on the production of endo-1,4-beta-D-glucanase (CM-cellulase) activity by P. pinophilum was studied in shake flask cultures using Avicel PH101 as the carbon source. It was observed that the culture conditions had a profound effect on the level of endoglucanase (CM-cellulase) produced by P. pinophilum. However, isoelectric focusing of the endoglucanase activity obtained from shake flask and fermenter cultures using the same growth medium revealed that the enzyme system found in both cultures was identical qualitatively, and contained seven or eight different endoglucanase components. All the endoglucanase components appeared simultaneously in the early stages of culture and prolonged incubation resulted only in an increase in the concentration of these enzymes. Protease levels were found to be low in both types of culture but were particularly so in the growth medium which contained corn steep liquor. The proteases were unable to release low molecular weight peptides when P. pinophilum cellulase protein was used as a substrate. The results were interpreted to indicate that the multiplicity of endoglucanase components found in cultures of P. pinophilum is most likely the result of expression of a number of specific genes rather than by post-secretional modification of one or more endoglucanase(s) synthesized by the fungus.

Clonal derivation of a rat muscle cell strain that forms contraction-competent myotubes

Abstract: A muscle cell strain capable of forming contracting myotubes was isolated from an established rat embryo cell line. The myogenic cells, termed rat myoblast omega or RMo cells, have a diploid complement of chromosomes (n=42). In the presence of mitogen-containing growth medium, RMo cells proliferated with a cell generation time of about 12 hours. In mitogen-depleted medium, RMo cells withdrew from the cell cycle and formed myotubes that spontaneously contracted. Differentiated RMo cells produced creatine kinase isozymes in a ratio characteristic of skeletal muscle cells. RMo cells were easy to cultivate. Cells proliferated and differentiated equally well on gelatin-coated or noncoated culture dishes, at clonal or mass culture densities, and in all basal media tested. In most experiments, growth medium consisted of horse serum-containing medium supplemented with either chicken embryo extract or FGF activity; cells proliferated equally well in medium containing unsupplemented calf serum. RMo cells differentiated if growth medium was not replenished regularly. Alternatively, differentiation was induceable by incubation in mitogen-depleted medium consisting of basal medium supplemented either with 10−6 M insulin, 0.5% serum, or 50% conditioned growth medium. RMo cells were competently transformed with cloned exogenous genes. Because it forms functional myofibrils, the RMo cell line constitutes a useful model system for studying the cell biology and biochemistry of proteins involved in contractile apparatus assembly and muscle disease.

The Regulation of Exopolysaccharide Production and of Enzymes Involved in C1 Assimilation in Methylophilus methylotrophus

Abstract: Summary: Methylophilus methylotrophus produced viscous and non-viscous exopolysaccharides (EPS) when grown in batch culture. Both types contained glucose, galactose, mannose and an unidentified 6-desoxyhexose, and were substituted with pyruvate and acetate residues. When the organism was grown in continuous culture only the non-viscous EPS was synthesized; the rate of production was 18·5 mg h−1 (g biomass)−1 in methanol-limited cultures and increased by approximately 3- and 4-fold when growth was limited by oxygen or nitrogen respectively. The specific activity of methanol dehydrogenase in cell extracts was relatively low when bacteria were grown under conditions of methanol excess and increased 2-fold in carbon-limited cells, reflecting the need to ‘scavenge’ the small amounts of available methanol. In contrast, the specific activities of several key enzymes of the ribulose monophosphate (RuMP) pathway were greater in cells grown under conditions of nitrogen or oxygen limitation than when growth was Limited by the availability of carbon, indicating the potential for increased carbon flux round the cycle when excess methanol was present in the growth medium. When methylotrophs are grown under conditions of methanol excess it is important that there is a mechanism to prevent the overproduction of formaldehyde, and we suggest that these changes in EPS production and in the specific activities of the key enzymes of the RuMP cycle are necessary for the efficient removal of this toxic metabolite of methanol.

A medium designed for monitoring pitching yeast contamination in beer using a conductimetric technique

Abstract: Nitrogen assimilation is the most readily utilized source of conductance changes when pitching yeast is grown in a glucose-based medium. A simple growth medium comprising yeast nitrogen base, in which nitrogen is supplied as ammonium sulphate, and glucose gave good growth but little change in conductance. Inclusion of a succinate buffer in the medium to remove protons liberated as a result of nitrogen uptake produced a large decrease in conductance and detection times that correlated well with enumeration of yeast by plate counting. The medium will allow more rapid and automated detection of pitching yeast survival in pasteurized beer although individual calibration for each beer type will be necessary.

Inoculation of cowpea and wheat with strains of Bradyrhizobium sp. that differ in their production of indole acetic acid

Abstract: Sufficient indole acetic acid (IAA) was produced in yeast extract-mannitol (YM) medium by each of two isolates of indigenous Bradyrhizobium sp., to inhibit primary root growth of cowpea. Similar inoculation of cowpea with YM-cultivated strains from a culture collection or with washed cell suspensions of the two IAA-producing isolates had no apparent inhibiting effect on primary root growth. YM-cultivated cultures of each of the two high IAA-producing strains did not prevent nodulation of secondary roots and did not affect plant growth. In tests on agar plates a Bradyrhizobium sp. strain, CB756, frequently used for commercial inoculant production, stimulated root growth of cowpea and wheat seedlings, but when used to inoculate wheat in sand culture, neither strain CB756 nor the IAA-producing isolates measurably affected wheat growth. Production of IAA by the isolates and strains was strongly influenced by growth medium; all required tryptophan to excrete at least 0,5 μg IAA cm−3. Nine 5-methyl-tryptophan-r...

Production of the Fimbrial Adhesin 987P by Enterotoxigenic Escherichia coli during Growth under Controlled Conditions in a Chemostat

Abstract: Summary: The effects of growth conditions on the production of 987P fimbriae by the enterotoxigenic Escherichia coli strain 1592 were examined in steady state chemostat experiments at different specific growth rates. The amount of fimbriae produced by fimbriate cells (P+) was dependent on the specific growth rate (μ). Under aerobic growth conditions fimbriae production increased with higher μ values till μ = 0·40 h−1and decreased again at μ values close to μmax (0·48 h−1). Under anaerobic growth conditions the maximal production was comparable to that under aerobic growth conditions, and was also maximal close to μmax (0·16 h−1). Phase variation, measured as the percentage of fimbriate cells in a particular population, was independent of μ. The composition of the growth medium influenced both phase variation and overall production of fimbriae. A shift from minimal to a complex medium induced a rapid reduction in the amount of fimbriae per P+cell and a slower reduction in the percentage of P+cells. A shift from complex to minimal medium resulted in an increase in the percentage of P+cells and a constant amount of fimbriae per P+cell. The frequency of the phase switch was calculated for different growth conditions. The frequency of the P+→ P−switch between two steady states was 2·7 x 10−2. In batch culture the frequency of the P−→ P+switch was minimally 2·9 x 10−2. The results indicate that phase variation and the production of 987P fimbriae by fimbriate cells are under independent physiological control.

Structural variations and growth potential of Yersinia enterocolitica under different culture conditions.

Abstract: The growth potential and the polypeptide composition of Yersinia enterocolitica serotype 0:3 isolated from patients with uncomplicated diarrhoea, reactive arthritis or septicemia were evaluated under different culture conditions. The expression of polypeptides varied with presence of the virulence-associated 40-48 Mdal plasmid, growth medium, growth temperature and gas composition of the culture (air, carbon dioxide, oxygen). Also the initial growth medium at 26 degrees C, before temperature shift to 37 degrees C, influenced the subsequent growth potential and expression of polypeptides. The plasmid encoded at least 7 polypeptides. This plasmid also inhibited the multiplication of bacteria under defined culture conditions. The dominating plasmid-encoded polypeptides were optimally expressed in air or oxygen-supplemented growth medium. The majority of the chromosomally encoded polypeptides were expressed independently of presence of the plasmid, whereas the expression of at least 8 were repressed by the plasmid. Five chromosomally encoded polypeptides were expressed only in carbon dioxide and five only in oxygen environment. These results indicate that Y. enterocolitica may express different molecules in different environments in vivo. This may be of importance for host-parasite relationship and immune response.

Biotransformation of l-tyrosine and l-phenylalanine to 2,5-dihydroxyphenylacetic acid

Abstract: Disclosed is a method for the production of homogentisic acid which involves growing a fungus (typically a yeast of the species Yarrowia lipolytica) which has been mutated to provide a strain which is unable to grow on L-tyrosine and/or L-phenylalanine as the sole carbon source in a suitable growth medium containing L-tyrosine and/or L-phenylalanine together with a sub-optimal concentration of carbohydrate assimilable by the yeast. The yeast secretes recoverable quantities of homogentisic acid which can be rapidly and completely polymerized by raising the pH of the medium to above 10 thereby forming a melanin pigment.

Cell cycle progression delay in conditioned medium does not play a role in the repair of X-ray damage in Chinese hamster V79 cells.

Abstract: We tested our hypothesis that the lower survival of X-irradiated cells in growth medium (GM) relative to that in conditioned medium (CM) is due to differences in nutrient concentration levels rather than to differential effects on cell progression and growth. Chinese hamster V79 cells in log and unfed plateau phase, grown in Eagle's minimal essential medium (MEM) with 15% serum (100% GM), were irradiated. Before plating, cells were incubated in situ in various concentrations of MEM with serum (GM, normal cell progression) or MEM without serum or in CM (no cell progression). Cell survival was the lowest in 100% MEM with or without serum and increased with the decrease in MEM and serum concentrations, reaching a plateau in 40% MEM or 40% growth medium (40% MEM with 6% serum), similar to that in conditioned medium. Growth kinetics was the same in 40 and 100% growth medium, but the D0 of cells in 40% growth medium was higher than that of cells in 100% GM. Similarly, the D0 of cells in 40% MEM was higher than that of cells in 100% MEM, although cell progression was absent in both media. The radiation sensitivity of cells was the same in 40% GM with progression and in 40% MEM and CM with no progression. Cells in low-nutrient media were flatter than those in 100% MEM or GM. There was a correlation between the nutrient concentration in the medium postirradiation and the D0. This correlation was independent of the presence or absence of serum and thus independent of cell cycle progression. The cell morphology which is dependent on the nutrient concentration appears to influence the ability of a fraction of cells to repair their radiation damage.

Degradation of some sugars and sugar acids by the nonphosphorylated D-gluconate pathway in Aspergillus ustus

Abstract: The degradation of fifteen sugars, sugar acids and related substrates were examined using cellfree extracts of Aspergillus ustus growing on D-glucose, D-mannose, D-galactose or D-gluconate as the only carbon source. D-gluconate was superior for the induction of the enzymes capable for the degradation of some of these substrates. The addition of 0.5% malt extract with D-gluconate to the growth medium or the presence of shaking conditions resulted to an increase in the degradation of those substrates, whileas the incorporation of 0.5% malt extract alone to the medium has no effect. Extracts of D-gluconate-grown mycelia of A. ustus degraded D-gluconate > D-galactonate > 1 : 5 gluconolactone and > L-arabonate nonphosphorolytically more effectively. Optimum pH and temperature for the degradation of D-gluconate were found to be 8.0 and 40°C, respectively. Thermal stability studies on the behaviour of D-gluconate dehydratase showed that this enzyme was stable at 50°C and 60°C for 30 and 5 minutes, respectively. Specific activity of this enzyme was increased three times when cell-free extracts were incubated at 60°C for 5 minutes. MgCl2 and CoSO4 were good activators, while CaCl2 p-mercurychlorobenzoate (PMCB), sodium arsenite, ZnSO4, CuSO4, iodoacetic acid, MnCl2 and FeSO4 were potent inhibitors for D-gluconate dehydratase activity. Km was calculated for D-gluconate and found to be 2.5 × 10−2 M.

Selection and characterization of an L-thiazolidine-4-carboxylic acid resistant callus culture of Vigna radiata (L.) Wilczek var. radiata

Abstract: Callus cultures were established from seedling root tips of mungbean (Vigna radiata (L.) Wilczek var. radiata) cv. K 851. The growing calli were exposed to increasing concentrations of thioproline — an analog of proline, in the medium. A concentration of 3.0 mM thioproline completely inhibited the growth of the cells. However, after 25 days incubation 5 cell clones were obtained which could grow on this concentration of thioproline. Out of them one vigorously growing cell clone was further characterized. This selected clone contained higher endogenous levels of free proline (5 fold) and K+ (1.5 fold) and exhibited elevated tolerance, not only to thioproline but also to exogenously applied NaCl in the growth medium, as compared to the normal sensitive callus cells. Higher endogenous levels of free proline and K+ appear to impart dual resistance to thioproline and NaCl to the selected cell strain.

Peculiarities of amino acid transport in Schizosaccharomyces pombe: effects of growth medium.

Abstract: Transport systems for amino acids in the wild-type strain ofSchizosaccharomyces pombe are not constitutive. During growth on different media no transport of acidic, neutral and basic amino acids is detectable. To acquire the ability to transport amino acids, cells must be preincubated with a metabolic source of energy, such as glucose. The appearance of transport activity is associated with protein synthesis (suppression by cycloheximide) at all phases of culture growth. After such preincubation the initial rate of amino acid uptake depends on the phase of growth of the culture and on the amount of glucose in the growth medium but not on the nitrogen source used.l-Proline and 2-aminoisobutyric acid are practically not transported under any of the conditions tested.