Effect of ATP on the Calcium Efflux in Dialyzed Squid Giant Axons

Dialysis perfusion technique makes it possible to control the internal composition of squid giant axons. Calcium efflux has been studied in the presence and in the virtual absence (<5 µM) of ATP. The mean calcium efflux from axons dialyzed with 0.3 µM ionized calcium, [ATP]_i > 1,000 µM, and bathed in artificial seawater (ASW) was 0.24 ± 0.02 pmol·cm^-2·s^-1 (P/CS) (n = 8) at 22°C. With [ATP]_i < 5 µM the mean efflux was 0.11 ± 0.01 P/CS (n = 15). The curve relating calcium efflux to [ATP]_i shows a constant residual calcium efflux in the range of 1–100 µM [ATP]_i. An increase of the calcium efflux is observed when [ATP]_i is >100 µM and saturates at [ATP]_i > 1,000 µM. The magnitude of the ATP-dependent fraction of the calcium efflux varies with external concentrations of Na⁺, Ca⁺⁺, and Mg⁺⁺. These results suggest that internal ATP changes the affinity of the calcium transport system for external cations.     

A study of lipid bilayer membrane stability using precise measurements of specific capacitance

A method is described for measuring the specific capacitance (C_m) of lipid bilayer membranes with an estimated experimental error of only 1%. The gross capacitance was measured with an AC Wheatstone bridge and a photographic technique was used to determine the area of thin membrane. The results of measurements on oxidized cholesterol-decane membranes formed in 1 × 10^-2 M KCl show that C_m depends upon temperature, voltage, time, and the age of the bulk membrane solutions. For a freshly thinned membrane (from 5 week old solution), C_m increases exponentially from an initial value of 0.432 ±0.021 (SD) μF/cm² with a time constant of ~15 min. A 100 mv potential applied across the membrane for 10-20 min prior to making measurements eliminated this time dependence and produced final-state membranes. C_m of final-state membranes depends upon applied voltage (V_a) and obeys the equation C_m = C₀ + βV_a² where V_a V_DC + V^rms_AC. C₀ and β depend upon temperature; C₀ decreases linearly with temperature while β increases linearly. At 20°C, C₀ = 0.559 ±0.01 (SD) μF/cm² and β = 0.0123 ±0.0036 (SD) (μF/cm²)/(mv²) and at 34°C, C₀ = 0.472 ±0.01 and β = 0.0382 ±0.0039. These variations in C_m are interpreted as resulting from thickness changes. The possibility that they result from diffuse layer and/or membrane dielectric phenomena is discussed and found to be unlikely. The results are discussed in terms of membrane stability by constructing hypothetical potential energy vs. thickness curves.     

Photosynthetic Response of Soybean to Microclimate in 26-Year-Old Tree-Based Intercropping Systems in Southern Ontario,Canada

In order to study the effect of light competition and microclimatic modifications on the net assimilation (NA), growth and yield of soybean (Glycine max L.) as an understory crop, three 26-year-old soybean-tree (Acer saccharinum Marsh., Populus deltoides X nigra, Juglans nigra L.) intercropping systems were examined. Tree competition reduced photosynthetically active radiation (PAR) incident on soybeans and reduced net assimilation, growth and yield of soybean. Soil moisture of 20 cm depth close (< 3 m) to the tree rows was also reduced. Correlation analysis showed that NA and soil water content were highly correlated with growth and yield of soybean. When compared with the monoculture soybean system, the relative humidity (RH) of the poplar-soybean, silver maple-soybean, and black walnut-soybean intercropped systems was increased by 7.1%, 8.0% and 5.9%, soil water content was reduced by 37.8%, 26.3% and 30.9%, ambient temperature was reduced by 1.3°C, 1.4°C and 1.0°C, PAR was reduced by 53.6%, 57.9% and 39.9%, and air CO₂ concentration was reduced by 3.7μmol·mol^-1, 4.2μmol·mol^-1 and 2.8μmol·mol^-1, respectively. Compared to the monoculture, the average NA of soybean in poplar, maple and walnut treatments was also reduced by 53.1%, 67.5% and 46.5%, respectively. Multivariate stepwise regression analysis showed that PAR, ambient temperature and CO₂ concentration were the dominant factors influencing net photosynthetic rate.     

THE LYSOSOME PERIPHERY: BIOCHEMICAL AND ELECTROKINETIC PROPERTIES OF THE TRITOSOME SURFACE

Normal rat liver lysosomes were isolated by the technique of loading with Triton WR-1339. Purity of the preparation was monitored with marker enzymes; a high enrichment in acid hydrolases was obtained in the tritosome fraction. In 0.0145 M NaCl, 4.5% sorbitol, 0.6 mM NaHCO₃, pH 7.2 at 25°C the tritosomes had an electrophoretic mobility of -1.77 ± 0.02 µm/s/V/cm, a zeta potential of 23.2 mV, a surface charge of 1970 esu/cm², and 33,000 electrons per particle surface assuming a tritosome diameter of 5 x 10^-7 m. Treatment of the tritosomes with 50 µg neuraminidase/mg tritosome protein lowered the electrophoretic mobility of the tritosome to -1.23 ± 0.02 µm/s/V/cm under the same conditions and caused the release of 2.01 µg sialic acid/mg tritosome protein. Treatment of the tritosomes with hyaluronidase did not affect their electrophoretic mobility, while trypsin treatment elevated the net negative electrophoretic mobility of the tritosomes. Tritosome electrophoretic mobilities indicated a homogeneous tritosome population and varied greatly with ionic strength of the suspending media. pH vs. electrophoretic mobility curves indicated the tritosome periphery to contain an acid-dissociable group which likely represents the carboxyl group of N-acetylneuraminic acid; this was not conclusively proven, however, since the tritosomes lysed below a pH of 4 in the present system. Total tritosome carbohydrate (anthrone-positive material as glucose equivalents) was 0.19 mg/mg tritosome protein while total sialic acid was 3.8 µg (11.4 nmol)/mg tritosome protein. A tritosome "membrane" fraction was prepared by osmotic shock, homogenization, and sedimentation. Approximately 25% of the total tritosome protein was present in this fraction. Analysis by gas-liquid chromatography and amino acid analyzer showed the following carbohydrate composition of the tritosome membrane fraction (in microgram per milligram tritosome membrane protein): N-acetylneuraminic acid, 14.8 ± 3; glucosamine, 24 ± 3; galactosamine, 10 ± 2; glucose, 21 ± 2; galactose, 26 ± 2; mannose, 31 ± 5; fucose, 7 ± 1; xylose, 0; and arabinose, 0. The results indicate that the tritosome periphery is characterized by external terminal sialic acid residues and an extensive complement of glycoconjugates. Essentially all the tritosome N-acetylneuraminic acid is located in the membrane and about 53% of it is neuraminidase susceptible.     

Elastic-Mathematical Theory of Cells and Mitochondria in Swelling Process: The Membranous Stresses and Modulus of Elasticity of the Egg Cell of Sea Urchin, Strongylocentrotus purpuratus

To the revolution-ellipsoidal and spherical membranous shell (cell mitochondrion) are introduced the equations for the calculation of both the modulus of elasticity (Young's modulus) and the stresses, which exist at the membrane. The existing pressure difference between the inner and outer surface of the membrane is calculated in the dilution of seawater media in the osmotic steady state. The experimental results are obtained by using egg cells of the sea urchin, Strongylocentrotus purpuratus. Up to the specific volume of the egg cell (V_E ≈ 35·10^-8 cm³) Boyle-van't Hoff's law is valid (defined as the subelastic range) beyond that the elastic stresses exist (elastic range). For the maximum value of the stresses existing at the cell wall one obtains σ ≈ 5.5·10⁶ dyne/cm² and for the modulus of elasticity E = 1.0·10⁷ dyne/cm², which is constant when the value of relative strain ε_ν > 15%. The breaking limit by an approximate calculation is σ_U ≈ 11·10⁶ dyne/cm². The membrane is assumed to be convoluted and its hypothetical degree of folding was calculated [unk]_a = 34%. The results are compared with the values existing in the literature and other types of cells are found to have values of elasticity in the same range as values of the membrane of S. purpuratus. Both compression and cell elastometer methods are criticized and in certain cases results of these methods are considered to belong to the subelastic domain.     

Effects of Light, Carbon Dioxide, and Temperature on Photosynthesis, Oxygen Inhibition of Photosynthesis, and Transpiration in Solanum tuberosum

Individual leaves of potato (Solanum tuberosum L. W729R), a C₃ plant, were subjected to various irradiances (400-700 nm), CO₂ levels, and temperatures in a controlled-environment chamber. As irradiance increased, stomatal and mesophyll resistance exerted a strong and some-what paralleled regulation of photosynthesis as both showed a similar decrease reaching a minimum at about 85 neinsteins·cm⁻²·sec⁻¹ (about ½ of full sunlight). Also, there was a proportional hyperbolic increase in transpiration and photosynthesis with increasing irradiance up to 85 neinsteins·cm⁻²·sec⁻¹. These results contrast with many C₃ plants that have a near full opening of stomata at much less light than is required for saturation of photosynthesis.     

Construction and Characterization of Salmonella typhimurium Strains That Accumulate and Excrete α- and β- Isopropylmalate

Two Salmonella typhimurium strains, which could be used as sources for the leucine biosynthetic intermediates α- and β-isopropylmalate were constructed by a series of P22-mediated transductions. One strain, JK527 [flr-19 leuA2010 Δ(leuD-ara)798 fol-162], accumulated and excreted α-isopropylmalate, whereas the second strain, JK553 (flr-19 leuA2010 leuB698), accumulated and excreted α- and β-isopropylmalate. The yield of α-isopropylmalate isolated from the culture medium of JK527 was more than five times the amount obtained from a comparable volume of medium in which Neurospora crassa strain FLR₉₂-1-216 (normally used as the source for α- and β-isopropylmalate) was grown. Not only was the yield greater, but S. typhimurium strains are much easier to handle and grow to saturation much faster than N. crassa strains. The combination of the two regulatory mutations flr-19, which results in constitutive expression of the leucine operon, and leuA2010, which renders the first leucine-specific biosynthetic enzyme insensitive to feedback inhibition by leucine, generated limitations in the production of valine and pantothenic acid. The efficient, irreversible, and unregulated conversion of α-ketoisovaleric acid into α-isopropylmalate (α-isopropylmalate synthetase K_m for α-ketoisovaleric acid, 6 × 10⁻⁵ M) severely restricted the amount of α-ketoisovaleric acid available for conversion into valine and pantothenic acid (ketopantoate hydroxymethyltransferase K_m for α-ketoisovaleric acid, 1.1 × 10⁻³ M; transaminase B K_m for α-ketoisovaleric acid, 2 × 10⁻³ M).     

Cloning,purification, kinetic and anion inhibition studies of a recombinant β-carbonic anhydrase from the Atlantic salmon parasite platyhelminth Gyrodactylus salaris

A β-class carbonic anhydrase (CA, EC 4.2.1.1) was cloned from the genome of the Monogenean platyhelminth Gyrodactylus salaris, a parasite of Atlantic salmon. The new enzyme, GsaCAβ has a significant catalytic activity for the physiological reaction, CO₂ + H₂O ⇋ HCO₃⁻ + H⁺ with a k_cat of 1.1 × 10⁵ s⁻¹ and a k_cat/K_m of 7.58 × 10⁶ M⁻¹ × s⁻¹. This activity was inhibited by acetazolamide (K_I of 0.46 µM), a sulphonamide in clinical use, as well as by selected inorganic anions and small molecules. Most tested anions inhibited GsaCAβ at millimolar concentrations, but sulfamide (K_I of 81 µM), N,N-diethyldithiocarbamate (K_I of 67 µM) and sulphamic acid (K_I of 6.2 µM) showed a rather efficient inhibitory action. There are currently very few non-toxic agents effective in combating this parasite. GsaCAβ is subsequently proposed as a new drug target for which effective inhibitors can be designed.     

Calcium Efflux from Internally Dialyzed Squid Giant Axons

Calcium efflux has been studied in squid giant axons under conditions in which the internal composition was controlled by means of a dialysis perfusion technique. The mean calcium efflux from axons dialyzed with 0.3 µM calcium and 5 mM ATP was 0.26 pmol/cm²·s at 22°C. The curve relating the Ca efflux with the internal Ca concentration had a slope of about one for [Ca]_i lower than 0.3µM and a slope smaller than one for higher concentrations. Under the above conditions replacement of [Na]_o and [Ca]_o by Tris and Mg causes an 80% fall in the calcium efflux. When the axons were dialyzed with a medium free of ATP and containing 2 mM cyanide plus 5µg/ml oligomycin, analysis of the perfusion effluent gave values of 1–4 µM ATP. Under this low ATP condition, replacement of external sodium and calcium causes the same drop in the calcium efflux. The same effect was observed at higher [Ca]_i, (80 µM). These results suggest that the Na-Ca exchange component of the calcium efflux is apparently not dependent on the amounts of ATP in the axoplasm. Axons previously depleted of ATP show a significant transient drop in the calcium efflux when ATP is added to the dialysis medium. This effect probably represents the sequestering of calcium by the mitochondrial system. The consumption of calcium by the mitochondria of the axoplasm in dialyzed axons was determined to be of the order of 6.0 x 10^-7 mol Ca⁺⁺/mg of protein with an initial rate of 2.6 x 10^-8 mol Ca⁺⁺/min·mg of protein. Axons dialyzed with 2 mM cyanide after 8–10-min delays show a rise in the calcium efflux in the presence of "normal" amounts of exogenous ATP. This effect seems to indicate that cyanide, per se, can release calcium ions from internal sources.     

Activation of Haem-Oxidized Soluble Guanylyl Cyclase with BAY 60-2770 in Human Platelets Lead to Overstimulation of the Cyclic GMP Signaling Pathway

Background and Aims

Nitric oxide-independent soluble guanylyl cyclase (sGC) activators reactivate the haem-oxidized enzyme in vascular diseases. This study was undertaken to investigate the anti-platelet mechanisms of the haem-independent sGC activator BAY 60-2770 in human washed platelets. The hypothesis that sGC oxidation potentiates the anti-platelet activities of BAY 60-2770 has been tested.

Methods

Human washed platelet aggregation and adhesion assays, as well as flow cytometry for α_IIbβ₃ integrin activation and Western blot for α1 and β1 sGC subunits were performed. Intracellular calcium levels were monitored in platelets loaded with a fluorogenic calcium-binding dye (FluoForte).

Results

BAY 60-2770 (0.001–10 µM) produced significant inhibition of collagen (2 µg/ml)- and thrombin (0.1 U/ml)-induced platelet aggregation that was markedly potentiated by the sGC inhibitor ODQ (10 µM). In fibrinogen-coated plates, BAY 60-2770 significantly inhibited platelet adhesion, an effect potentiated by ODQ. BAY 60-2770 increased the cGMP levels and reduced the intracellular Ca²⁺ levels, both of which were potentiated by ODQ. The cell-permeable cGMP analogue 8-Br-cGMP (100 µM) inhibited platelet aggregation and Ca²⁺ levels in an ODQ-insensitive manner. The cAMP levels remained unchanged by BAY 60-2770. Collagen- and thrombin-induced α_IIbβ₃ activation was markedly inhibited by BAY 60-2770 that was further inhibited by ODQ. The effects of sodium nitroprusside (3 µM) were all prevented by ODQ. Incubation with ODQ (10 µM) significantly reduced the protein levels of α1 and β1 sGC subunits, which were prevented by BAY 60-2770.

Conclusion

The inhibitory effects of BAY 60-2770 on aggregation, adhesion, intracellular Ca²⁺ levels and α_IIbβ₃ activation are all potentiated in haem-oxidizing conditions. BAY 60-2770 prevents ODQ-induced decrease in sGC protein levels. BAY 60-2770 could be of therapeutic interest in cardiovascular diseases associated with thrombotic complications.     

Physicochemical properties of tipula iridescent virus

The molecular weight of Tipula iridescent virus, based on sedimentation and diffusion coefficients, was 5.51 × 10⁸, with hydration of 0.57 g of water per g of virus. Deoxyribonucleic acid content, based on total inorganic phosphorus liberated, was 19 ± 0.2%. At 260 mμ, the virus gave an uncorrected absorbance of 18.2 cm²/mg of virus and a light-scattering corrected absorbance of 9.8 cm²/mg of virus. Amino acid analyses of the virus protein revealed a remarkable similarity to Sericesthis iridescent virus. The possibility is discussed that the four iridescent insect viruses reported to date bear a strain relationship.     

Drimanes from Drimys brasiliensis with leishmanicidal and antimalarial activity

This paper evaluates CHCl₃ and CH₃OH extracts of the stem bark, branches and leaves of Drimys brasiliensis and drimane sesquiterpenes isolated from the stem bark against strains of Leishmania amazonensis and Leishmania braziliensis promastigotes and Plasmodium falciparum trophozoites. All of the extracts and compounds were tested in cell lines in comparison with reference standards and cell viability was determined by the XTT method. The CHCl₃ and CH₃OH extracts from the stem bark and branches yielded promising results against two strains of Leishmania, with 50% inhibitory concentrations (IC₅₀ ) values ranging from 39-100 µg/mL. The CHCl₃ extract of the stem bark returned IC₅₀ values of 39 and 40.6 µg/mL for L. amazonensis and L. braziliensis, respectively. The drimanes were relatively effective: 1-β-(p-coumaroyloxy)-polygodial produced IC₅₀ values of 5.55 and 2.52 µM for L. amazonensis and L. braziliensis, respectively, compared with 1-β-(p-methoxycinnamoyl)-polygodial, which produced respective IC₅₀ values of 15.85 and 17.80 µM. The CHCl₃ extract demonstrated activity (IC₅₀ of 3.0 µg/mL) against P. falciparum. The IC₅₀ values of 1-β-(p-cumaroyloxyl)-polygodial and 1-β-(p-methoxycinnamoyl)-polygodial were 1.01 and 4.87 µM, respectively, for the trophozoite strain. Therefore, the results suggest that D. brasiliensis is a promising plant from which to obtain new and effective antiparasitic agents.     

Rhizoremediation of Trichloroethylene by a Recombinant, Root-Colonizing Pseudomonas fluorescens Strain Expressing Toluene ortho-Monooxygenase Constitutively

Trichloroethylene (TCE) was removed from soils by using a wheat rhizosphere established by coating seeds with a recombinant, TCE-degrading Pseudomonas fluorescens strain that expresses the tomA⁺ (toluene o-monooxygenase) genes from Burkholderia cepacia PR1₂₃(TOM_23C). A transposon integration vector was used to insert tomA⁺ into the chromosome of P. fluorescens 2-79, producing a stable strain that expressed constitutively the monooxygenase at a level of 1.1 nmol/min · mg of protein (initial TCE concentration, 10 μM, assuming that all of the TCE was in the liquid) for more than 280 cell generations (36 days). We also constructed a salicylate-inducible P. fluorescens strain that degraded TCE at an initial rate of 2.6 nmol/min · mg of protein in the presence of 10 μM TCE [cf. B. cepacia G4 PR1₂₃(TOM_23C), which degraded TCE at an initial rate of 2.5 nmol/min · mg of protein]. A constitutive strain, P. fluorescens 2-79TOM, grew (maximum specific growth rate, 0.78 h⁻¹) and colonized wheat (3 × 10⁶ CFU/cm of root) as well as wild-type P. fluorescens 2-79 (maximum specific growth rate, 0.77 h⁻¹; level of colonization, 4 × 10⁶ CFU/cm of root). Rhizoremediation of TCE was demonstrated by using microcosms containing the constitutive monooxygenase-expressing microorganism, soil, and wheat. These closed microcosms degraded an average of 63% of the initial TCE in 4 days (20.6 nmol of TCE/day · plant), compared to the 9% of the initial TCE removed by negative controls consisting of microcosms containing wild-type P. fluorescens 2-79-inoculated wheat, uninoculated wheat, or sterile soil.     

Flavobacterium plurextorum sp. nov. Isolated from Farmed Rainbow Trout (Oncorhynchus mykiss)

Five strains (1126-1H-08^T, 51B-09, 986-08, 1084B-08 and 424-08) were isolated from diseased rainbow trout. Cells were Gram-negative rods, 0.7 µm wide and 3 µm long, non-endospore-forming, catalase and oxidase positive. Colonies were circular, yellow-pigmented, smooth and entire on TGE agar after 72 hours incubation at 25°C. They grew in a temperature range between 15°C to 30°C, but they did not grow at 37°Cor 42°C. Based on 16S rRNA gene sequence analysis, the isolates belonged to the genus Flavobacterium. Strain 1126-1H-08^T exhibited the highest levels of similarity with Flavobacterium oncorhynchi CECT 7678^T and Flavobacterium pectinovorum DSM 6368^T (98.5% and 97.9% sequence similarity, respectively). DNA–DNA hybridization values were 87 to 99% among the five isolates and ranged from 21 to 48% between strain 1126-1H-08^T, selected as a representative isolate, and the type strains of Flavobacterium oncorhynchi CECT 7678^T and other phylogenetic related Flavobacterium species. The DNA G+C content of strain 1126-1H-08^T was 33.2 mol%. The predominant respiratory quinone was MK-6 and the major fatty acids were iso-C_15∶0 and C_15∶0. These data were similar to those reported for Flavobacterium species. Several physiological and biochemical tests differentiated the novel bacterial strains from related Flavobacterium species. Phylogenetic, genetic and phenotypic data indicate that these strains represent a new species of the genus Flavobacterium, for which the name Flavobacterium plurextorum sp. nov. was proposed. The type strain is 1126-1H-08^T ( = CECT 7844^T = CCUG 60112^T).     

Ethylene Removal at Low Temperatures under Biofilter and Batch Conditions

Removal of the plant hormone ethylene (C₂H₄) is often required by horticultural storage facilities, which are operated at temperatures below 10°C. The aim of this study was to demonstrate an efficient, biological C₂H₄ removal under such low-temperature conditions. Peat-soil, acclimated to degradation of C₂H₄, was packed in a biofilter (687 cm³) and subjected to an airflow (~73 ml min⁻¹) with 2 ppm (μl liter⁻¹) C₂H₄. The C₂H₄ removal efficiencies achieved at 20, 10, and 5°C, respectively, were 99.0, 98.8, and 98.4%. This corresponded to C₂H₄ levels of 0.022 to 0.032 ppm in the biofilter outlet air. At 2°C, the average C₂H₄ removal efficiency dropped to 83%. The detailed temperature response of C₂H₄ removal was tested under batch conditions by incubation of 1-g soil samples in a temperature gradient ranging from 0 to 29°C with increments of 1°C. The C₂H₄ removal rate was highest at 26°C (0.85 μg of C₂H₄ g [dry weight]⁻¹ h⁻¹), but remained at levels of 0.14 to 0.28 μg of C₂H₄ g (dry weight)⁻¹ h⁻¹ at 0 to 10°C. At 35 to 40°C, the C₂H₄ removal rate was negligible (0.02 to 0.06 μg of C₂H₄ g [dry weight]⁻¹ h⁻¹). The Q₁₀ (i.e., the ratio of rates 10°C apart) for C₂H₄ removal was 1.9 for the interval 0 to 10°C. In conclusion, the present results demonstrated microbial C₂H₄ removal, which proceeded at 0 to 2°C and produced a moderately psychrophilic temperature response.     

Isolation and Characterization of Carbendazim-degrading Rhodococcus erythropolis djl-11

Carbendazim (methyl 1H-benzimidazol-2-yl carbamate) is one of the most widely used fungicides in agriculture worldwide, but has been reported to have adverse effects on animal health and ecosystem function. A highly efficient carbendazim-degrading bacterium (strain dj1-11) was isolated from carbendazim-contaminated soil samples via enrichment culture. Strain dj1-11 was identified as Rhodococcus erythropolis based on morphological, physiological and biochemical characters, including sequence analysis of the 16S rRNA gene. In vitro degradation of carbendazim (1000 mg·L⁻¹) by dj1-11 in minimal salts medium (MSM) was highly efficient, and with an average degradation rate of 333.33 mg·L⁻¹·d⁻¹ at 28°C. The optimal temperature range for carbendazim degradation by dj1-11 in MSM was 25–30°C. Whilst strain dj1-11 was capable of metabolizing cabendazim as the sole source of carbon and nitrogen, degradation was significantly (P<0.05) increased by addition of 12.5 mM NH₄NO₃. Changes in MSM pH (4–9), substitution of NH₄NO₃ with organic substrates as N and C sources or replacing Mg²⁺ with Mn²⁺, Zn²⁺ or Fe²⁺ did not significantly affect carbendazim degradation by dj1-11. During the degradation process, liquid chromatography-mass spectrometry (LC-MS) detected the metabolites 2-aminobenzimidazole and 2-hydroxybenzimidazole. A putative carbendazim-hydrolyzing esterase gene was cloned from chromosomal DNA of djl-11 and showed 99% sequence homology to the mheI carbendazim-hydrolyzing esterase gene from Nocardioides sp. SG-4G.     

Effect of Microvilli on Lateral Diffusion Measurements Made by the Fluorescence Photobleaching Recovery Technique

To consider the effect of surface microvilli on measurements of lateral diffusion by fluorescence photobleaching recovery, we have measured the diffusion of the lipid probe 3,3′-dihexadecyl indocarbocyanine iodide on the villated main body and unvillated budding polar body of unfertilized mouse eggs. On the main body we found D = (6.41 ± 0.62) × 10^-9 cm²/s with (77.0 ± 2.1)% recovery, and on the budding polar body we found D = (7.05 ± 0.75) × 10^-9 cm²/s with (84.7 ± 1.3)% recovery. We thus find only slight differences in diffusion in the two regions.     

Thermal denaturation in acidic solutions of double-helical ribonucleic acid from virus-like particles found in Penicillium chrysogenum. A spectrophotometric study

1. Two species of double-helical RNA isolated from mycelium of Penicillium chrysogenum were titrated with acid at 25°C and 95°C (solvent 0.1m-sodium phosphate buffer). At 25°C denaturation occurred at about pH3. At 95°C in the denatured form cytosine residues titrated as a simple monobasic acid of pK3.9 compared with pK2.5 for the native form at 25°C. 2. On thermal denaturation in neutral and acidic solutions one species of RNA (38% rG·rC) `melted' in three distinct stages, equivalent to a mixture of three species, namely one of about 25% rG·rC, another of about 33% rG·rC and a third of about 46% rG·rC: the relative proportions were 0.25:0.35:0.40. 3. On thermal denaturation in acidic solutions the increase in the fraction of ionized cytosine residues concomitant with the `melting' of rG·rC base pair also affects the spectrum especially at 280nm and serves to enhance the contribution of rG·rC base pairs at this wavelength. The increment in ε_(P) at 280nm on `melting' an rG·rC base pair approaches 53501·mol<sup>−1</sup>·cm<sup>−1</sup> depending on pH, compared with 33501·mol<sup>−1</sup>·cm<sup>−1</sup> at pH7. In contrast ε<sub>(P)</sub> at 280nm is scarcely affected by `melting' rA·rU base pairs or by the protonization of adenine residues. 4. Changes in the spectrum of Escherichia coli rRNA on denaturation in acidic solutions were studied to yield the mole fractions of rA·rU and rG·rC base pairs `melting' at particular pH values.     

Chlorine Dioxide for Reduction of Postharvest Pathogen Inoculum during Handling of Tree Fruits

Alternatives to hypochlorous acid and fungicides are needed for treatment of fruit and fruit-handling facilities. Chlorine dioxide was evaluated and found effective against common postharvest decay fungi and against filamentous fungi occurring on fruit packinghouse surfaces. In vitro tests with conidial or sporangiospore suspensions of Botrytis cinerea, Penicillium expansum, Mucor piriformis, and Cryptosporiopsis perennans demonstrated >99% spore mortality within 1 min when the fungi were exposed to aqueous chlorine dioxide at 3 or 5 μg · ml^-1. Longer exposure times were necessary to achieve similar spore mortalities with 1 μg · ml^-1. Of the fungi tested, B. cinerea and P. expansum were the least sensitive to ClO₂. In comparison with the number recovered from untreated control areas, the number of filamentous fungi recovered was significantly lower in swipe tests from hard surfaces such as belts and pads in a commercial apple and pear packinghouse after treatment of surfaces with a 14.0- to 18.0-μg · ml^-1 ClO₂ foam formulation. Chlorine dioxide has desirable properties as a sanitizing agent for postharvest decay management when residues of postharvest fungicides are not desired or allowed.     

Denitrification in Low pH Spodosols and Peats Determined with the Acetylene Inhibition Method

Potential denitrification rates were determined for predominantly acid (pH ≥ 3.6) horizons of forestal, miry, and agricultural soils from 22 locations in southern Finland. The acetylene inhibition method was used with nitrate-amended water-logged soils incubated in an N₂ atmosphere containing 2.5 or 5% C₂H₂. Complete inhibition of the reduction of N₂O to N₂ was observed in 99.3% of the samples. The denitrification rates varied from 0.12 to 53.8 μg of N·cm^-3·day^-1. Correlation between denitrification rate and soil pH was highly significant: r = 0.84 on a volume basis, and r = 0.44 on a weight basis. Vegetation type and amount of soil organic matter had a minor or no effect, respectively. In spodosolized soils the rates were significantly higher for B horizons than for A horizons. These results show that denitrification can occur in acid soils.