Genomic and physiological diversity amongst strains of Thiobacillus ferrooxidans,and genomic comparison with Thiobacillus thiooxidans

Twenty-three strains of Thiobacillus ferrooxidans of known pedigree were examined. Thirteen strains survived 65° C for 5 min and 7 of these for 10 min, but sporulation was never observed. All strains grew between 25° C and 35° C and some strains grew at 5° and 40° C. They were genomically diverse, comprising 7 DNA homology groups, and the GC content varied from 55–65 mol %. Correlation between genomic group and growth temperature was noted. All strains grew on ferrous sulfate as energy source, but some failed to utilize elemental sulfur. Acidified thiosulfate supported growth of most of the strains examined but it was judged to be a poor substrate upon which to base taxonomic conclusions because of decomposition of thiosulfate in acid. Six strains of Thiobacillus thiooxidans showed negligible genomic affinity to T. ferrooxidans, and they comprised 2 DNA homology groups and their GC content varied from 52–62 mol%. Anomalies due to contaminants in cultures of T. ferrooxidans were resolved, and the contaminants were identified.     

Silver toxicity to ferrous iron and pyrite oxidation and its alleviation by yeast extract in cultures ofThiobacillus ferrooxidans

Summary Ferrous-ion oxidation byThiobacillus ferrooxidans was inhibited by 10^–6 M Ag⁺ while a slight inhibition of growth was apparent with 10^–7 M Ag⁺. The threshold toxic concentration was the seme for four different test strains. While prolonged lag phases resulted from culture exposure to Ag⁺, Fe²⁺ oxidation rates after the onset of growth showed little variation under these conditions. Yeast extract (0.02%) partially alleviated the toxicity of silver-ion by reducing the lag periods. Pyrite oxidation byT. ferrooxidans and mixed cultures of acidophiles was tested at 8.3×10^–7 to 8.3×10^–5 M Ag⁺. Strong inhibition was apparent at 8.3×10^–5 M Ag⁺ and little to no inhibition was observed at 8.3×10^–7 M Ag⁺.     

Transformation of pickling cucumber with chitinase-encoding genes using Agrobacterium tumefaciens

Summary Transformation of cucumber cv. Endeavor was attempted using three Agrobacterium tumefaciens strains (a supervirulent leucinopine type, an octopine type and a nopaline type), each harbouring one of three binary vectors which contained an acidic chitinase gene from petunia, and basic chitinase genes from tobacco and bean, respectively, driven by the CaMV 35S promoter. Petiole explants were inoculated with a bacterial suspension (10⁸ cells·ml^–1), cocultivated for 48–96 h and placed on Murashige and Skoog (MS) medium with 5.0 M each of 2,4-D and BA, 50 mg·l^–1 kanamycin and 500 mg·l^–1 carbenicillin. The frequency of embryogenic callus formation ranged from 0 to 12%, depending on strains/vectors used and length of cocultivation, with the highest being obtained using the leucinopine strain with petunia acidic chitinase gene. The kanamycin-resistant embryogenic calli were used to initiate suspension cultures (in liquid MS medium with 1.0/1.0 M 2,4-D/BA, 50 mg·l^–1 kanamycin) for multiplication of embryogenic cell aggregates. Upon plating of cell aggregates onto solid MS medium with 1.0/1.0 M NAA/BA and 50 mg·l^–1 kanamycin, calli continued to grow and later differentiated into plantlets. Transformation by the leucinopine strain and all three vectors was confirmed by PCR amplification of the NPT II gene in transgenic calli and plants, in addition to Southern analysis. Expression of the acidic chitinase gene (from petunia) and both basic chitinase genes (from tobacco and bean) in different transgenic cucumber lines was confirmed by Western analyses.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - BA 6-benzyl-aminopurine - CaMV cauliflower mosaic virus - NAA naphthaleneacetic acid - NPT II neomycin phosphotransferase II - PCR polymerase chain reaction     

Plant regeneration from protoplasts isolated from embryogenic suspension cultured cells of Cinnamomum camphora L.

An efficient and reproducible protocol is described for the regeneration of Cinnamomum camphora protoplasts isolated from cultured embryogenic suspension cells. Maximum protoplast yield (13.1±2.1×10⁶/g FW) and viability (91.8±3.8%) were achieved using a mixture of 3% (w/v) cellulase Onozuka R10 and 3% (w/v) macerozyme Onozuka R10 in 12.7% (w/v) mannitol solution containing 0.12% (w/v) MES, 0.36% (w/v) CaCl₂·2H₂O, and 0.011% (w/v) NaH₂PO₄·2H₂O. First divisions occurred 7–10 days following culture initiation. The highest division frequency (24.6±2.9%) and plating efficiency (6.88±0.8%) were obtained in liquid medium (MS) supplemented with 30 g l^–1 sucrose, 0.7M glucose, 0.1 mg l^–1 NAA, 1.0 mg l^–1 BA, and 1.0 mg l^–1 GA₃. After somatic embryo induction and then shoot induction, the protoplast-derived embryos produced plantlets at an efficiency of 17.5%. Somatic embryos developed into well-rooted plants on MS medium supplemented with 1.0 mg l^–1 3-indole butyric acid (IBA). Regenerated plants that transferred to soil have normal morphology.     

Dynamics of production oftrans-zeatin andtrans-zeatin riboside by immobilized cytokinin-autonomous and cytokinin-dependent tobacco cells

Two strains of cultured tobacco cells (Nicotiana tabacum L. cv. Wisconsin 38) differing in their requirement for exogenous cytokinins (cytokinin-dependent and cytokinin-autonomous) were immobilized on polyphenylenoxide (Sorfix) activated with glutaraldehyde. Columns packed with immobilized cells were continually eluted with diluted Murashige and Skoog's medium lacking or supplemented with synthetic cytokinin (6-benzylaminopurine; BA). Purified samples of column eluates were fractionated by HPLC, andtrans-zeatin (t-Z) andtrans-zeatin riboside (t-ZR) content was estimated by enzyme immunoassay. Both cytokinin-autonomous and cytokinin-dependent tobacco cells produced and excretedt-Z and its riboside, and there were significant quantitative differences between the strains. The steady-state excretion rate oft-Z was 19.8 ng · g^–1 dw · h^–1 and 4 ng · g^–1 dw · h^–1, respectively, and that oft-ZR 4 ng · g^–1 dw · h^–1 and 1 ng · g^–1 dw · h^–1, respectively. Exposure of cytokinin-dependent cells to BA after 72 h of starving for this synthetic cytokinin caused temporary increase in excretion of both zeatin and its riboside. After the application of 5 M BA for 24 h, the excretion rate oft-ZR reached 5 ng · g^–1 dw · h^–1 (5-fold increase), and that oft-Z achieved 12 ng · g^–1 dw · h^–1 (3-fold increase). The elevation oft-Z excretion was delayed about 13 h compared witht-ZR excretion, which started increasing almost immediately after BA application. A pulse of BA in lower concentration (1.5 M for 30 h) provoked lower response.     

Marine bacterioplankton at the Weddell Sea ice edge,distribution of psychrophilic and psychrotrophic populations

Summary In the eastern Weddell Sea on several transects from ice-covered, through ice melt, to open-ocean stations, total and heterotrophic bacteria were estimated to document an enhanced bacteriological biomass expected near the ice edge. The highest numbers of bacteria were found in melted ice cores, with 4.2·10³ CFUml^–1 and 1.1·10⁷ Cells ml^–1. Although brine from pore water samples average more than one order of magnitude less cells per ml, the highest bacterial production, 2.2·10⁷ cells l^–1 day^–1, was recorded in brine samples. All quantitatively studied bacterial parameters were lower under the ice than in the ice samples but there were no clear vertical gradients in the water column. In the studied spring situation, sea ice occurrence seems to play only a minor role in the general distribution of the seawater bacterioplankton. The bacterial community structure was investigated by carrying out 29 morphological and biochemical tests on 118 isolated strains. The bacterial communities inhabiting Antarctic pack ice differ from those found in underlying seawater. Although non fermentative Gram-negative rods were always dominant in seawater, Vibrio sp. represented more than 25% of the strains isolated from some ice samples. The results clearly indicated that a large majority of the bacteria isolated from seawater must be considered psychrotrophic but that truly psychrophilic strains occurred in melted ice and brine samples.Data presented here were collected during the European Polarstern Study (EPOS) sponsored by the European Science Foundation     

Alcoholic fermentation of glucose and xylose by Pichia stipitis,Candida shehatae,Saccharomyces cerevisiae and Zymomonas mobilis: oxygen requirement as a key factor

Summary To investigate simultaneous alcoholic fermentation of glucose and xylose derived from lignocellulosic material by separate or co-culture processes, the effect of oxygen transfer rate (OTR) on the fermentation of 50 g/l xylose by Pichia stipitis NRRL Y 7124 and Candida shehatae ATCC 22984, and the fermentation of 50 g/l glucose by Saccharomyces cerevisiae CBS 1200 and Zymomonas mobilis ATCC 10988 was carried out in batch cultures. The kinetic parameters of the xylose-fermenting yeasts were greatly dependent on the OTR. The optimum OTR values were found to be 3.9 and 1.75 mmol·1^–1·h^–1 for C. shehatae and P. stipitis, respectively. By contrast the fermentative parameters of S. cerevisiae were poorly affected by the OTR range tested (0.0–3.5 mmol·l^–1·h^–1) Under these conditions the ethanol yields ranged from 0.41 g·g^–1 to 0.45 g·g^–1 and the specific ethanol productivity was around 0.70 g·g^–1·h^–1. Z. mobilis gave the highest fermentative performance under strictly anaerobic conditions (medium continually flushed with nitrogen): under these conditions, the ethanol yield was 0.43 g·g^–1 and the average specific ethanol productivity was 2.3 g·g^–1·h^–1. Process considerations in relation to the effect of OTR on the fermentative performance of the tested strains are discussed. Offprint requests to: J. P. Delgenes     

Lateral hydraulic conductivity of early metaxylem vessels in Zea mays L. roots

The hydraulic conductivity of the lateral walls of early metaxylem vessels (Lp_x in m · s^–1 · MPa^–1) was measured in young, excised roots of maize using a root pressure probe. Values for this parameter were determined by comparing the root hydraulic conductivities before and after steam-ringing a short zone on each root. Killing of living tissue virtually canceled its hydraulic resistance. There were no suberin lamellae present in the endodermis of the roots used. The value of Lp_x ranged between 3 · 10^–7 and 35 · 10^–7 m · s^–1 · MPa^–1 and was larger than the hydraulic conductivity of the untreated root (Lp_r = 0.7 · 10^–7 to 4.0 · 10^–7 m · s^–1 · MPa^–1) by factor of 3 to 13. Assuming that all flow through the vessel walls was through the pit membranes, which occupied 14% of the total wall area, an upper limit of the hydraulic conductivity of this structure could be given(Lp_pm=21 · 10^–7 to 250 · 10^–7 m · s^–1 · MPa^–1). The specific hydraulic conductivity (Lp_cw) of the wall material of the pit membranes (again an upper limit) ranged from 0.3 · 10^–12 to 3.8 · 10^–12 m² · s^–1 · MPa^–1 and was lower than estimates given in the literature for plant cell walls. From the data, we conclude that the majority of the radial resistance to water movement in the root is contributed by living tissue. However, although the lateral walls of the vessels do not limit the rate of water flow in the intact system, they constitute 8–31% of the total resistance, a value which should not be ignored in a detailed analysis of water flow through roots.Abbreviatations and Symbols k_wr (T ₁ ^2/W ) rate constant (half-time) of water exchange across root (s^–1 or s, respectively) - Lp_cw specific hydraulic conductivity of wall material (m² · s^–1 · MPa^–1) - Lp_pm hydraulic conductivity of pit membranes (m · s ^–1 · MPa^–1) - Lp_r hydraulic conductivity of root (m · s^–1 · MPa^–1) - Lp_x lateralhydraulic conductivity of walls of root xylem (m · s ^–1 · MPa^–1) This research was supported by a grant from the Bilateral Exchange Program funded jointly by the Natural Sciences and Engineering Research Council of Canada and the Deutsche Forschungsgemeinschaft to C.A.P., and by a grant from the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 137, to E.S. The expert technical help of Mr. Burkhard Stumpf and the work of Ms. Martina Murrmann and Ms. Hilde Zimmermann in digitizing chart-recorder strips is gratefully acknowledged.     

Morphological and cultural comparison of microorganisms in surface soil and subsurface sediments at a pristine study site in Oklahoma

Surface-soil and subsurface microfloras at the site of a shallow aquifer in Oklahoma were examined and compared with respect to (1) total and viable cell numbers, (2) colony and cell types that grew on various plating media, (3) cell morphologies seen in flotation films stripped from sample particles, and (4) cellular ultrastructure. Appreciable numbers of microbial cells were present in the subsurface (total counts: 10⁶–10⁷ cellsg^–1; viable counts up to 10⁶ cells · g^–1), but the subsurface microflora was considerably less populous than that of the surface soil (total counts: 10⁹ cells·g^–1; viable counts: 10⁷–10⁸ cells · g^–1). The subsurface microflora (especially that of the saturated zone) also appeared to be much less diverse, containing fewer microbial types that would grow on enumeration plates (on nutrient-rich media, 3–4 colony types versus 19–22 for the surface soil) and fewer cell types that could be distinguished by direct microscopy (3–4 types versus 17 for the surface soil). The specific types of microorganisms that were numerically predominant in the aquifer sediments were entirely different from those that were predominant in the surface soil. Moreover, the predominant types varied from one depth to another within the saturated zone. The potential metabolic capability of the subsurface microflora, as indicated by its readiness to grow rapidly on nutrient-rich media, also varied with depth.     

Alpha and beta adrenergic agonists stimulate water absorption in the rat proximal tubule

Summary Simultaneous capillary and luminal microperfusion studies were performed in the rat proximal tubule to determine the effects of the beta agonist isoproterenol and the alpha agonist phenylephrine on water absorption. Capillary and luminal perfusion solutions were composed such that organic solutes were not present, no bicarbonate was present in the lumen, and no chloride gradient was imposed. Under such conditions, water absorption (Jv) averaged 0.36±0.11 nl·min^–1·mm^–1. The addition of isoproterenol to the capillary solution in concentrations of 10^–6 and 10^–4 m resulted in significantly higherJv's of 0.68±0.10 and 0.71±0.11 nl·min^–1·mm^–1, respectively. The enhancing effect of isoproterenol was inhibited by the beta blocker propranolol (10^–4 m), but not by the alpha blocker phentolamine (10^–7 m). The addition of phenylephrine (10^–6 m) to the capillary perfusion solution also resulted in a significantly higherJv of 0.84±0.14 nl·min^–1·mm^–1, an effect inhibited by phentolamine (10^–7 m), but not by propranolol (10^–4 m). Neither phentolamine nor propranolol alone in the concentrations indicated had an effect on water absorption. These experiments indicate that both alpha and beta agonists stimulate water absorption in the superficial proximal tubule of the rat. This effect appears to be relatively specific for each class of agonist, as demonstrated by the effects of the specific antagonists.     

Isolation and study of two strains ofLeptospirillum-like bacteria from a natural mixed population cultured on a cobaltiferous pyrite substrate

Two strains ofLeptospirillum-like bacteria, L6 and L8, have been isolated from a mixed inoculum, also containingThiobacillus ferrooxidans andT. thiooxidans, cultured for one year with a colbaltiferous pyrite as energy substrate in a 100 I continuous bioleaching laboratory unit. Several physiological properties of the strains are described. The vibrio-shaped microorganisms grew at pH values lower than 1.3. Their growth rate was maximum between 2.5 and 8.0 g l¹ ferrous iron. The optimal growth temperature was 37.5° C. Ferric iron had a stimulative effect on bacterial development up to 8 g l^–1, and growth was as rapid at 14 g l^–1 ferric iron as at 8 g l^–1. The negative influence of cobalt on the final cell concentration was observed at 0.5 g l^–1, but the growth rate was not affected up to 2 g l^–1. The G + C content of strains L8 is 55.6 mol%.     

Role of non-quantum acetylcholine secretion in neural control of the membrane potential in skeletal muscle fibers of rats

Experiments on muscle fibers of the rat diaphragm (in vitro denervation) showed that their three-hour incubation in the cultural medium results in an 8-mV drop in the resting membrane potential (RMP). Addition of 5·10^–8 M carbacholine to the cultural medium, mimicing the effect of non-quantum acetylcholine, delayed depolarization of the denervated muscle. The effect of carbacholine could not be eliminated byd-tubocurarine (5·10^–6 M), a postsynaptic acetylcholine receptor blocker, and by ouabain (1·10^–4 M), and inhibitor of Na⁺, K⁺-ATPase of the membrane. At the same time, the effect could be completely eliminated by Mg²⁺ ions (5·10^–3 M), which blocked Ca²⁺ channels of the membrane, by N-nitroarginine (1·10^–4 M), which inhibited the enzyme NO-synthase, and by hemoglobin (2·10^–5 M), which inactivated the extracellular NO molecules. It is concluded that the released non-quantum acetylcholine can contribute to neural control of RMP of cross-striated muscle fibers via the Ca²⁺-dependent activation of NO synthesis in the sarcoplasm. The NO molecules can play the role of a retrograde signal indicative of the normal functioning of the neuromuscular synapse. The impairment of this link caused by a denervation-induced cessation of the non-quantum secretion can serve as a signal triggering the early changes in the muscle membrane following nerve transection.Neirofiziologiya/Neurophysiology, Vol. 27, No. 1, pp. 67–71, January–February, 1995.     

Ribonuclease III reduces the efficiency of bacteriophage gy1 propagation inE. coli

TheE. coli rnc gene encodes the double-stranded, RNA-specific ribonuclease III (RNaseIII). A novel bacteriophage, gy1, was isolated, and its propagation inE. coli was shown to depend on the expression of RNaseIII in the cell. (a) gyl has a low efficiency of plating on rnc⁺ strains and a high efficiency of plating on a rnc^– E. coli strain harboring the rnc 105 point mutation that renders its RNaseIII product inactive. (b) gy1 has a high efficiency of plating on rnc^– strains in which thernc gene is disrupted by a Tn10 insertion. (c) Plasmids harboring a rnc⁺ gene that were introduced into the rnc^– strains described above reduced the efficiency of plating of gy1.     

Regulation of phospho(enol)-pyruvate-and oxaloacetate-converting enzymes in Corynebacterium glutamicum

The presence and properties of the enzymes involved in the synthesis and conversion of phospho(enol)pyruvate (PEP) and oxaloacetate (OAA), the precursors for aspartate-derived amino acids, were investigated in three different Corynebacterium strains. This study revealed the presence of both PEP carboxykinase 0.29 mol·min^–1·mg^–1 of protein [units (U)·mg^–1] and PEP synthetase (0.13 U·mg^–1) in C. 2 glutamicum as well as pyruvate kinase (1.4 U·mg^–1) and PEP carboxylase (0.16 U·mg^–1). With the exception of PEP carboxykinase these activities were also present in glucose-grown C. flavum and C. lactofermentum. Pyruvate carboxylase activity was not detected in all three species cultivated on glucose or lactate. At least five enzyme activities that utilize OAA as a substrate were detected in crude extracts of C. glutamicum: citrate synthase (2 U·mg^–1), malate dehydrogenase (2.5 U·mg^–1), glutamate: OAA transaminase (1 U·mg^–1), OAA-decarboxylating activity (0.89 U·mg^–1) and the previously mentioned PEP carboxykinase (0.29 U·mg^–1). The partially purified OAA-decarboxylase activity of C. glutamicum was completely dependent on the presence of inosine diphosphate and Mn²⁺, had a Michaelis constant (K _m) of 2.0mm for OAA and was inhibited by ADP and coenzyme A (CoA). Examination of the kinetic properties showed that adenine nucleotides and CoA derivatives have reciprocal but reinforcing effects on the enzymes catalyzing the interconversion of pyruvate, PEP and OAA in C. glutamicum. A model for the regulation of the carbon flow based on these findings is presented.Correspondence to: M. S. M. Jetten     

Bacterial productivity in the water column and sediments of the Georgia (USA) coastal zone: Estimates via direct counting and parallel measurement of thymidine incorporation

Three methods of estimating bacterial productivity were compared using parallel samples of Atlantic Ocean water (within 0.25–15 km of the Georgia coast). The frequency-of-dividing cells (FDC) method and the [³H]thymidine incorporation method gave results which were strongly correlated (r=0.97), but the FDC estimates were always higher (X2 to X7) than the [³H]thymidine estimates. Estimates of bacterial productivity ranged from 2–4×10⁸ cells·l^–1·h^–1 at 0.25 km from shore to 1–9×10⁷cells·l^–1·h^–1 at 15 km. A method involving incubation of 3-m filtrates and direct counting gave results that could not be easily translated into estimates of bacterial productivity. Application of the FDC method to sediment samples gave high productivity estimates, which could be not reconciled with productivity estimates based on sediment oxygen uptake.     

Nutritional and hormonal requirements of Ginkgo biloba embryo-derived callus and suspension cell culture

Calli were obtained from Ginkgo biloba embryos grown on Murashige and Skoog (MS) medium. The G. biloba cells could grow on either MS or Gamborg B5 mineral salt medium supplemented with sucrose (3% and 2%, respectively) and naphthaleneacetic acid (NAA) and kinetin (K) in concentrations ranging from 0.1 to 2.0 mg·L^–1. Best growth and maintenance of callus cultures were achieved using MS medium supplemented with 2 mg·L^–1 NAA and 1 mg·L^–1 K (N₂K₁MS). Light was required to maintain healthy growth of the callus tissue.In both MS and B5 based media, sucrose was hydrolyzed extracellularly before being taken up by Ginkgo cell suspension cultures. Specific growth rates of 0.13 d^–1 and 0.08 d^–1 were obtained in MS medium supplemented with 1 mg·L^–1 NAA, 0.1 mg·L^–1 K and 30 g·L^–1 sucrose (N₁K_0.1MS) and B5 medium supplemented with the same growth regulator regime and 20 g·L^–1 sucrose (N₁K_0.1B5) respectively. Complete phosphate and ammonium uptake was observed in 11 days when cultured in MS medium and 10 days and 4 days respectively when cultured in B5 medium. During the culture, G. biloba cells consumed only 64% and 29% of the nitrate content of N₁K_0.1MS and N₁K_0.1B5 media respectively. Maximum dry biomass concentrations were 13.4 g·L^–1 and 7.9 g·L^–1, and yields on carbohydrate were 0.39 and 0.45 in N₁K_0.1MS and N₁K_0.1B5 media respectively. The better performance of MS cultures came from the higher sucrose and nitrogen salts concentrations of this medium.Abbreviations B5 Gamborg mineral salt medium - d.w. Dry weight - K Kinetin - MS Murashige and Skoog mineral salt medium - N or NAA Naphthaleneacetic acid - N_iK_jMS i and j are the respective concentrations (mg·L^–1) of NAA and K - n Number of experimental points - r Linear regression correlation coefficient     

Influence of water activity on Streptococcus diacetylactis metabolism

During the cheese-making process, water activity (a_w) is one of the essential environmental parameters acting on bacterial growth and metabolic pathways. The influence of a_w on Streptococcus diacetylactis growth and lactic acid production was studied. The specific growth rate was linearly related to water availability in the milk medium. The cell behaviour was quite different above and below a_w=0.95, which can be considered a limiting value. Below this value, the lactic acid production reached 1.4–6.1 mg·g^–1, whereas the specific productivity was 2.0–2.6 mg·10^–10 cells·h^–1. Changes in the consumption of lactose and amino acids during the different growth phases was completely modified by decreasing the water availability in the medium. Correspondence to: N. Cochet     

Medium optimization for a methanol utilizing bacterium based on chemostat theory

Summary In the productions of biomass and vitamin B₁₂ using methanol as the sole carbon source, it is necessary to use a medium in which methanol is the growth limiting substrate. Other inorganic salts should be in slight excess so that the yield of cells and the intracellular content of vitamin B₁₂ do not vary. From basic principles of chemostat culture, a medium was optimized for Pseudomonas AM-1 a methanol utilizing bacterium, for the concentrations of various inorganic salts. This was done in a series of chemostat cultures at a dilution rate of 0.1 h^–1. Optimum amounts of NH₄ ⁺, PO₄ ^3- and Mg²⁺ were estimated from the minimum concentration of the salt at which methanol became growth limiting. The optimum concentrations of Ca²⁺, Fe²⁺, Mn²⁺, and Zn²⁺ as a group were determined in the same way. Cu²⁺, Mo⁶⁺, Co²⁺ and B³⁺ are required at concentrations of g/l and they were not studied as these very low level can be introduced as contaminants from other salts. The optimum medium composition (in g/l) was as follows: (NH₄)₂SO₄, 1.0; H₃PO₄, 75×10^–3; MgSO₄ · 7H₂O, 30×10^–3; CaCl₂ · 2H₂O, 3.3×10^–3; FeSO₄ · 7H₂O, 1.3×10^–3, MnSO₄ · 4H₂O, 0.13×10^–3; ZnSO₄ · 7H₂O, 0.13×10^–3; CuSO₄ · 5H₂O, 40×10^–6; Na₂MoO₄, 40×10^–6; CoCl₂ · 6H₂O, 40×10^–6; H₃BO₃, 30×10^–6 and methanol 4.     

Time course of vasopressin-induced formation of microvilli in granular cells of toad urinary bladder

Summary Vasopressin-induced transformation of ridges to microvilli on the surface of granular cells of toad urinary bladder occurs in conjunction with induced alterations in the water permeability of the luminal membrane. This study was designed to establish the relationship between the time course for induction of microvilli and the time course for induction of increased water permeability after vasopressin stimulation. Hemibladders were examined at 2.5, 5, 10, 20 and 30 min following exposure to 20 mU/ml of vasopressin and at 5, 10, 20, 30, 40, 50 and 60 min after washout of vasopressin. Within 2.5 min, vasopressin initiated complete transformation of ridges to microvilli on approximately 13% of the granular cells, while osmotic water flow (Jv) was 0.31±0.10 l·min^–1·cm^–2. Five minutes following vasopressin stimulation, microvilli were present on approximately 30% of granular cells andJv was 2.27±0.13 l·min^–1·cm^–2. At 10 minJv was maximum at 4.03±0.15 l·min^–1·cm^–2 and 50% of the granular cells were covered with microvilli. This percentage increased to 70% at 20 min and was maintained at 30 min, althoughJv decreased to 3.9±0.35 l·min^–1·cm^–2 at 30 min. Five minutes following vasopressin washout, ridges interspersed with microvilli reappeared asJv fell to 1.10±0.30 l·min^–1·cm^–2. At 10 min after vasopressin washout,Jv approached basal levels, but the reversal of microvilli to ridges remained incomplete. At 60 min after vasopressin washout, the granular cells had regained their original ridgelike surface structures. Thus, these studies establish a temporal relationship between the induction and reversibility of vasopressin-induced microvillous formation and alterations in the osmotic water permeability of the apical plasmalemma.     

The effect of carbon dioxide on the growth kinetics of fructose-limited chemostat cultures of Acetobacterium woodii DSM 1030

The growth of the anaerobic acetogenic bacterium Acetobacterium woodii DSM 1030 was investigated in fructose-limited chemostat cultures. A defined medium was developed which contained fructose, mineral salts, cysteine · HCl and Ca pantothenate (1 mg · 1^–1) supplied in a vitamin supplement. Growth at high dilution rates was dependent on the presence of CO₂ in the gas phase. The ^max was found to be 0.16 h^–1 and the fructose maintenance requirement was 0.1 to 0.13 mmol fructose · (g dry wt)^–1 · h^–1. A growth yield of 61 g dry wt · (mol fructose)^–1, corrected for the cell maintenance requirement and for incorporation of fructose carbon into cell biomass, was determined from the fructose consumption. A corresponding growth yield of 69 g dry wt · (mol fructose)^–1 was calculated from the acetate production assuming that fructose fermentation was homoacetogenic. A Y_ATP of 12.2 to 13.8 g dry wt · (mol ATP)^–1 was calculated from these growth yields using a value of 5 mol ATP · (mol fructose)^–1 as an estimate of the amount of ATP synthesised from fructose fermentation. The addition of yeast extract (0.5 g · 1^–1) to the medium did not influence the ^max or cell yield. After prolonged growth under fructose-limited conditions the requirement of the culture for CO₂ in the gas phase was reduced.Abbreviations YE yeast extract - IC inorganic carbon - D fermenter dilution rate : h^–1 - M_X maintenance requirement for X: mmol X · (g dry wt)^–1 · h^–1 - X may be fructose (Fruct), fructose consumed in energy metabolism (Fruct [E]), acetate (Ac) - ATP CO₂, NH _inf4 ^sup+ or Pi - qX specific rate of utilisation or consumption of X: mmol X · (g dry wt)^–1 · h^–1 - V fermenter volume: litre - rC · Cell, fermenter cell carbon production: mmol C · h^–1 - Y_X yield of cells on X: g dry wt · (mol X)^–1 - Y _infx ^supmax the yield corrected for cell maintenance: g dry wt · (mol X)^–1 - S_ATP stoichiometry of ATP synthesis from fructose: mol ATP · (mol frucose)^–1 - x cell concentration: g dry wt · 1^–1 - specific growth rate : h^–1 - ^max maximum specific growth rate: h^–1