Tuberculate spore formation by thirty-two strains of Histoplasma capsulatum

Summary 1. Thirty-two strains ofHistoplasma capsulatum were studied concerning their ability to form tuberculate spores and their conversion into the yeast phase.2. Nine strains did not produce tuberculate spores on Sabouraud's agar, on corn meal agar, on spent medium, on media with pH adjusted from 4.5 to 7.0 or on the first passage through hamsters.3. Tuberculate spore production did occur in these nine strains when Sabouraud's medium was enriched with phosphate, especially KH₂PO₄. In addition, all but two strains produced tuberculate spores after a second passage through hamsters.4. Growth on KH₂PO₄ enriched Sabouraud's agar led to a greater yield of yeast phase as compared to yeast phase resulting from colonies of the same strain grown on plain Sabouraud's agar. This may be due to the greater number of spores produced on the KH₂PO₄ enriched medium.5. A grinding technique of preparing inocula improved slightly the facility of obtaining the yeast phase over heavy inoculation with unground pieces of mold culture.     

Influence of the phosphorus and nitrogen source on nisin production in Lactococcus lactis subsp. lactis batch fermentations using a complex medium

The influence of different phosphorus and nitrogen sources on Lactococcus lactis subsp. lactis NIZO 22186 growth and nisin production was studied in batch fermentations using a complex medium. KH₂PO₄ was found to be the best phosphorus source for nisin production. Increasing initial phosphate concentrations from 0 to 5% KH₂PO₄ exerted a double effect, creating favourable pH conditions and particularly stimulating the nisin production levels, which were highest at 5% KH₂PO₄. Up to now, no such high initial phosphate concentrations have been reported for the production of other antibiotics or bacteriocins. Nisin, a lanthionine-containing peptide antibiotic with bacteriocin properties, clearly behaved as a primary metabolite, since its formation was linked with active growth and was not suppressed by phosphate concentrations up to 5%. A complex medium supplemented with cotton seed meal as nitrogen source also gave very high nisin yields. Correspondence to: L. De Vuyst     

Shoot specific fungal endophytes alter soil phosphorus (P) fractions and potential acid phosphatase activity but do not increase P uptake in tall fescue

Aims

An experiment was performed to test how different fungal endophyte strains influenced tall fescue’s ability to access P from four P sources varying in solubility.

Methods

Novel endophyte infected (AR542E+ or AR584E+), common toxic endophyte infected (CTE+), or endophyte-free (E-) tall fescues were grown for 90 days in acidic soils amended with 30 mg kg^?1 P of potassium dihydrogen phosphate (KH₂PO₄), iron phosphate (FePO₄), aluminum phosphate (AlPO₄), or tricalcium phosphate ((Ca₃(PO₄)₂), respectively.

Results

Phosphorus form strongly influenced plant biomass, P acquisition, agronomic P use efficiency, microbial communities, P fractions. P uptake and vegetative biomass were similar for plants grown with AlPO₄, Ca₃(PO₄)₂, and KH₂PO₄ but greater than in control and FePO₄ soils. Infection with AR542E+ resulted in significantly less shoot biomass than CTE+ and E- varieties; there was no influence of endophyte on root biomass. The biomarker for arbuscular mycorrhizal fungi (AM fungi, 16:1ω5c) was selected as an effective predictor of variations in P uptake and tall fescue biomass. Potential acid phosphatase activity was strongly influenced by endophyte x P form interaction.

Conclusions

Endophyte infection in tall fescue significantly affected the NaOH-extractable inorganic P fraction, but had little detectable influence on soil microbial community structure, root biomass, or P uptake.

     

Characterization of phosphate absorption in maize root cortex segments

Uptake of phosphate ions by 1 mm segments of isolated maize root cortex layers was studied. Cortex segments (from roots of 8 days old maize plants) absorb phosphate ions from 1 mM KH₂PO₄ in 0.2 mM CaSCO₄ at the average rate of 34.3 ±3.2 μg P_i g^?1 (fr. m.) h^?1,i.e. 0.35± 0.02 μmol P_i g^?1 (fr. m.) h^?1. Phosphate uptake considerably increases after a certain period of “augmentation”,i.e. washing in aerated 0.2 mM CaSO₄. This increase is completely blocked by the presence of 10 μg ml^?1 cycloheximide. The relation of uptake rate to phosphate concentration in the medium was shown to have 3 phases in the concentration range of 0.02 - 40 mM. Transition points were found between 0.8–1 mM and 10–20 mM. Following K_m and V_max values were found: K_m[mM] : 0.37 - 3.82 - 27.67 V_max[μg P_i g^?1 (fr. m.) h^?1] : 3.33 - 39.40 - 66.67 We have found no sharp pH optimum for phosphate uptake. It proceeds at almost constant rate till pH 6.0 and then the uptake rate drops with increasing pH. At low phosphate concentrations (1 mM) the lowest uptake rate was found at 5 and 13 °C, while the uptake is higher at 5 °C than at 13 °C at phosphate concentrations higher than 1 mM. At these concentrations uptake rate at 35 °C is lower than at 25 °C. Phosphate uptake considerably decreased in anaerobic conditions. DNP and iodoacetate (0.1 mM) completely blocked phosphate uptake from 1 mM KH₂PO₄, while uptake from 5 and 10 mM KH₂PO₄ was left unaffected by these substances. The inhibitors of active - SH groups NEM and PCMB inhibited phosphate uptake: 10^?3 M NEM by 81.6%, 10⁴ M NEM by 42% and 10^?4 M PCMB by 42%.     

Effect of temperature and pH on absorption of carbon dioxide by a free level of mixed solutions of some buffers

The rate of absorption of carbon dioxide by solutions of NaHCO₃, KH₂PO₄, hydrogencarbonate, phosphate and borate buffers at 20, 30 and 40°C was determined manometrically. The absorption rate increases for all buffers tested with increasing pH. The CO₂ absorption rate by KH₂PO₄ and by the phosphate buffer at low pH is lower than that of water. For other buffers tested it is equal to or higher than that of water, especially at higher temperatures.     

Increasing phosphorus concentration in the extraradical hyphae of <Emphasis Type="Italic">Rhizophagus irregularis</Emphasis> DAOM 197198 leads to a concomitant increase in metal minerals

Plants associated with arbuscular mycorrhizal fungi (AMF) acquire phosphorus via roots and extraradical hyphae. How soil P level affects P accumulation within hyphae and how P in hyphae influences the accumulation of metal minerals remains little explored. A bi-compartmented in vitro cultivation system separating a root compartment (RC), containing a Ri T-DNA transformed carrot root associated to the AMF Rhizophagus irregularis DAOM 197198, from a hyphal compartment (HC), containing only the extraradical hyphae, was used. The HC contained a liquid growth medium (i.e., the modified Strullu-Romand medium containing P in the form of KH₂PO₄) without (0 μM) or adjusted to 35, 100, and 700 μM of KH₂PO₄. The accumulation of P and metal minerals (Ca, Mg, K, Na, Fe, Cu, Mn) within extraradical hyphae and AMF-colonized roots, and the expression of the phosphate transporter gene GintPT were assessed. The expression of GintPT in the extraradical hyphae did not differ in absence of KH₂PO₄ or in presence of 35 and 100 μM KH₂PO₄ in the HC but was markedly reduced in presence of 700 μM KH₂PO₄. Hyphal P concentration was significantly lowest in absence of KH₂PO₄, intermediate at 35 and 100 μM KH₂PO₄ and significantly highest in presence of 700 μM KH₂PO₄ in the HC. The concentrations of K, Mg, and Na were positively associated with the concentration of P in the extraradical hyphae developing in the HC. Similarly, P concentration in extraradical hyphae in the HC was related to P concentration in the growth medium and influenced the concentration of K, Mg, and Na. The accumulation of the metal mineral K, Mg, and Na in the extraradical hyphae developing in the HC was possibly related to their function in neutralizing the negative charges of PolyP accumulated in the hyphae.     

Enhanced curdlan production in <Emphasis Type="Italic">Agrobacterium</Emphasis> sp. ATCC 31749 by addition of low-polyphosphates

A large amount of adenosine triphosphate with high energy phosphate bonds is required for uridine triphosphate regeneration during curdlan biosynthesis by Agrobacterium sp. ATCC 31749. To supply high energy for curdlan synthesis, three low-polyphosphates (Na₄P₂O₇, Na₅P₃O₁₀, and (NaPO₃)₆) with higher energy phosphate bonds were employed to substitute for KH₂PO₄-K₂HPO₄ in fermentation medium. Two genes encoding the polyphosphate metabolizing enzymes, polyphosphate kinase and exopolyphosphatase, were amplified and showed 95% homology to those in Agrobacterium sp. C58 by sequence analysis. The curdlan yields were enhanced by 23 and 134% when phosphate concentrations 0.024 mol/L of Na₅P₃O₁₀ and 0.048 mol/L of (NaPO₃)₆ respectively, were added in the medium. The maximum curdlan yield of 30 ± 1.02 g/L was obtained with the addition of 0.048 mol/L of (NaPO₃)₆ with 5 g/L CaCO₃ in the medium. When CaCO₃ was removed from the culture and the three lowpolyphosphates were added, the pH and biomass yield dropped remarkably and little or no curdlan was produced. The culture containing 0.048 mol/L of (NaPO₃)₆ was mixed with KH₂PO₄-K₂HPO₄ and CaCO₃ in the medium, but showed no effect on curdlan production. However, curdlan yield was improved by 49 ∼ 60% when CaCO₃ was removed from the medium and KH₂PO₄-K₂HPO₄ acted as a buffer. It appears that the positive effect of (NaPO₃)₆ on curdlan production required the buffering capacity of CaCO₃ and the absence of KH₂PO₄-K₂HPO₄ competing as a phosphate supplier.     

Uncoupling of Methanogenesis from Growth of Methanosarcina barkeri by Phosphate Limitation

Production of methane by Methanosarcina barkeri from H₂-CO₂ was studied in fed-batch culture under phosphate-limiting conditions. A transition in the kinetics of methanogenesis from an exponentially increasing rate to a constant rate was due to depletion of phosphate from the medium. The period of exponentially increasing rate of methanogenesis was extended by increasing the initial concentration of phosphate in the medium. Addition of phosphate during the constant period changed the kinetics to an exponentially increasing rate of methanogenesis, indicating the reversibility of phosphate depletion. The relation between methanogenesis and growth of M. barkeri was investigated by measuring the incorporation of phosphorus, supplied as KH₂³²PO₄, in the medium. At a low (1 μM) initial concentration of phosphate in the medium and during the constant period of methanogenesis, there was no net cell growth. At a higher (10 μM) initial concentration of phosphate, cell growth proceeded linearly with time after phosphate had been removed from the medium by uptake into cells.     

Study on the Relationship between Soil Selenium and Plant Selenium Uptake

Various extraction methods have been used to determine selenium (Se) concentrations in soils and plants in the second seleniferous regions of China. Our results show tea Se contents in the study area range from 1.009 to 2.6 mg/kg, which reveal that the tea areas in Ziyang County are in seleniferous regions. The four extraction methods evaluated in this study provide different information concerning soil and plant Se levels. The quality control/quality assurance program for this project indicated there is excellent agreement between total soil Se and extractable Se. For example, phosphate extractable Se results from the field investigation and greenhouse study were found to be highly correlated (R ² > 0.91) by linear regression analyses. Results from rye seedling experiments further show phosphate extractable Se has significant correlations with plant Se uptake and that a 0.1 M solution of KH₂PO₄ can be used as the extractant of soil available Se. In the acid soil, the Brassica campestris yield could be significantly reduced when the content of Se⁶⁺–Se ≥ 0.5 mg/kg, and the influence on the yield was not as obvious when the content of Se⁶⁺–Se reached up to 2.0 mg/kg. The uptake by Brassica campestris of Se⁶⁺–Se is higher than that of Se⁴⁺–Se. The main factors influencing the biological availability of soil Se, in order of their importance are CaCO₃, the presence of silt grains, organic matter and the presence of clay grains. pH could affect KH₂PO₄ extractable Se through CaCO₃.     

Components of fermentation medium regulate bacteriocin synthesis by the recombinant strain Lactococcus lactis subsp. lactis F-116

The regulation of the synthesis of bacteriocin produced by the recombinant strain Lactococcus lactis subsp. lactis F-116 has been studied. The synthesis is regulated by the components of the fermentation medium, the content of inorganic phosphate (KH₂PO₄), yeast autolysate (source of amine nitrogen), and changes in carbohydrates and amino acids. The strain was obtained by fusion of protoplasts derived from two related L. lactis subsp. lactis strains, both exhibiting a weak ability to synthesize the bacteriocin nisin. Decreasing the content of KH₂PO₄ from 2.0 to 1.0 or 0.5% caused bacteriocin production to go down from 4100 to 2800 or 1150 IU/ml, respectively; the base fermentation medium contained 1.0% glucose, 0.2% NaCl, 0.02% MgSO₄, and yeast autolysate (an amount corresponding to 35 mg % ammonium nitrogen). The substitution of sucrose for glucose (as the source of carbon) increased the antibiotic activity by 26%, and the addition of isoleucine, by 28.5%. Elevation of the concentration of yeast autolysate in the low-phosphate fermentation medium stimulated both the growth of the lactococci and the synthesis of bacteriocin. Introduction of 1% KH₂PO₄, yeast autolysate (an amount corresponding to 70 mg % ammonium nitrogen), 2.0% sucrose, and 0.1% isoleucine increased the bacteriocin-producing activity of the strain by 2.4 times.     

Mineral Uptake in Fusarium oxysporum f. sp. vasinfectum

A method for growing Fusarium oxysporum, a mycelial fungus, and a technique for its use in mineral uptake studies have been described. Some general characteristics of the uptake process were determined. The fungus, grown for 54 hours, was found to take up as much K as 15 to 20 meq/100 g dry weight in 2 to 4 hours from a solution of 5 meq/l KCl. Approximately 3 to 5 meq of this uptake was readily removed by a CaCl₂ rinse. The uptake was only slightly sensitive to pH over the range of 4 to 9. Below pH 4 uptake dropped rapidly. The age of the culture appeared to be the dominant factor in determining the rate of uptake. In contrast to other fungi, the presence of glucose during uptake was detrimental to K uptake. Conditions unfavorable for metabolic activity as low temperature, anaerobiosis, or the presence of DNP markedly reduced the uptake rate. Although the fungus took up Na from single salt solutions nearly as well as K, the latter ion was much preferred in mixtures of the two ions. The organism showed no significant metabolic uptake of Ca or Cl. During uptake from KCl solutions, the organic acid content increased. The increase, chiefly in succinic acid and to a lesser extent in acetic and citric acids, amounted to about half the K uptake. The remainder of the K taken up was correlated with a roughly equivalent efflux of cellular Mg.     

Proton/Phosphate Stoichiometry in Uptake of Inorganic Phosphate by Cultured Cells of Catharanthus roseus (L.) G. Don

Upon absorption of phosphate, cultured cells of Catharanthus roseus (L.) G. Don caused a rapid alkalinization of the medium in which they were suspended. The alkalinization continued until the added phosphate was completely exhausted from the medium, at which time the pH of the medium started to drop sharply toward the original pH value. Phosphate exposure caused the pH of the medium to increase from pH 3.5 to values as high as 5.8, while the rate of phosphate uptake was constant throughout (10-17 micromoles per hour per gram fresh weight). This indicates that no apparent pH optimum exists for the phosphate uptake by the cultured cells. The amount of protons cotransported with phosphate was calculated from the observed pH change up to the maximum alkalinization and the titration curve of the cell suspension. Proton/phosphate transport stoichiometry ranged from less than unity to 4 according to the amount of phosphate applied. At low phosphate doses, the stoichiometries were close to 4, while at high phosphate doses, smaller stoichiometries were observed. This suggests that, at high phosphate doses, activation of the proton pump is induced by the longer lasting proton influx acidifying the cytoplasm. The increased H⁺ efflux due to the proton pump could partially compensate protons taken up via the proton-phosphate cotransport system. Thus, the H⁺/H₂PO₄⁻ stoichiometry of the cotransport is most likely to be 4.     

Direct assessment of mineral phosphorus availability to tropical crops using 32P labelled compounds

The availability to plants of phosphorus (P) derived from sparingly soluble iron and aluminium phosphates was directly assessed with ³²P labelled compounds in two glasshouse trials.In the first experiment, the comparative availability of all mineral phosphate (P) compounds to maize increased with time (14 to 42 days post emergence) and plant total P uptake, but P source did not affect the growth or total plant uptake of P. The comparative availability of the amorphous AlPO₄ (Al-P), crystalline AlPO₄ (variscite), amorphous FePO₄ (Fe-P), and crystalline FePO₄ (strengite) compared to KH₂PO₄ (=100) was 53.1, 3.4, 38.9, and 1.9%, respectively. In the second experiment, the availability of Fe-P, strengite, and KH₂PO₄ to several topical crop species was examined. There was no difference between maize, sorghum, mungbean, cowpea or soybean in their ability to utilise Fe-P or KH₂PO₄, although maize utilised strengite more than the other species. The major difference between these species in their ability to acquire P appears to be a difference in ability to locate soluble soil P rather than differences in their ability to access different pools of soil P.The advantages of using neutron irradiation to directly measure P absorption from mineral P compounds over traditional methodologies is discussed.     

Modeling some mineral nutrient requirements for micropropagated wild apricot shoot cultures

A response surface methodology (RSM) experimental design was applied for improving micropropagation of a wild apricot, Prunus armeniaca Lam., from the mountains of Kazakhstan. In an initial study, woody plant medium (WPM) mineral nutrients [calcium nitrate, ammonium nitrate, mesos (calcium chloride, potassium phosphate and magnesium sulfate) potassium sulfate and minor nutrients] were tested in a response surface methodology (RSM) experiment. Shoot quality was the best when nitrogen and mesos (CaCl₂, MgSO₄, K₂SO₄, KH₂PO₄) compounds were altered. In this study an expanded mesos optimization experiment was run. Data taken included a subjective quality rating, shoot length, shoot number, leaf color and size, callus and physiological disorders. Data were analyzed by Classification and Regression Tree Analysis (CART), a data mining technique that provides specific cutoff values for data and easy to interpret data trees. The CART analysis indicated that the best quality would be with ≤2.4× WPM levels of KH₂PO₄ and ≤0.75× MgSO₄. Shoot length was affected by K₂SO₄, but most shoots were of good size at any concentration. Shoot multiplication was affected by KH₂PO₄, but there were >5 shoots at any concentration. Leaf color was best with ≤2.41× KH₂PO₄ and ≤1.22× K₂SO₄. Based on the CART analysis, a recommendation for improved mesos compounds was developed. Each of the individual trees was analyzed and the cutoff points determined so that all the growth characteristics could be considered in the final concentrations chosen. Using the combined results from the CART analysis, the suggested medium would include WPM with CaCl₂ 2.7×, MgSO₄ 2.7×, K₂SO₄ 0.8×, KH₂PO₄ 0.75×.     

Improvement of glycerol production by Candida krusei in batch and continuous cultures using corn steep liquor

No fermentation parameter was affected at phosphate concentration above 0.4 g l^–1 when KH₂PO₄ was used as phosphate source and the glucose consumption rate was difficult to control when corn steep liquor (CSL) was adopted as the phosphate source. However, if CSL was supplemented as a source of growth factors instead of as the phosphate source, not only glucose uptake and glycerol was improved, but also fermentation became easy to control and a steady state of continuous culture was easily obtained.     

Effect of nutritional factors on lipase biosynthesis by Aspergillus niger

Summary Lipase biosynthesis occured in medium without lipids, but for improved production an inducer was needed. The source and concentration of an inducer had no signifficant effect. Starch as an additional carbon source stimulated lipase biosynthesis when used in small amounts. Addition of NH₄NO₃ as a nitrogen source, KH₂PO₄ as a phosphate source as well as Mg ions to the medium with inital pH 5.0 gave the best yield.     

Solubility of phosphorus added to four Manitoba soils with different calcium and magnesium contents

Summary The solubility of phosphorus was found to approximate that of dicalcium phosphate dihydrate and/or dimagnesium phosphate trihydrate when KH₂-PO₄, H₃PO₄ and K₂HPO₄ were added to four Manitoba soils. Eighty to one hundred, seventy to ninety and sixty to eighty per cent of the phosphorus added remained in solution when H₃PO₄, KH₂PO₄ and K₂HPO₄ were added, respectively. The solubility of the added phosphorus was high in all samples and relatively soluble compounds, dicalcium phosphate dihydrate and dimagnesium phosphate trihydrate, were most likely formed in the samples indicating that phosphorus added to these soils would be readily available to plants. Associate Professor and Professor respectively.     

Buffering capacity of the chief components of nutritive media for algae

The buffering capacity of solutions of KH₂PO₄ and NaHCO₃ increases with their concentration, the behaviour being describable by mathematical expressions. Solutions of KH₂PO₄ prepared from tap water exhibit a buffering capacity higher by an order of magnitude than those prepared from distilled water. However, there is no difference between the buffering capacities of solutions of NaHCO₃ prepared from tap and distilled water. Urea and NH₄NO₃ have almost no effect on the buffering capacities of sol utions.     

Characteristics of Aluminum-Phosphate-Adapted Carrot Cells: Uptake and Utilization of the Phosphate

To elucidate the mechanism responsible for the superior growthof a selected line of carrot cells (Daucus carota L. cv MS Yonsun)in medium that contained AIPO₄, kinetic studies of the uptakeof phosphate and the efficiency of utilization of phosphatewere performed with the selected cells and the wild-type cells.When the two cell lines were grown in a medium with adequatesoluble phosphate (2 mM), there was no difference between theirgrowth rates. Rates of increase in fresh weight as a functionof increasing concentration of phosphate in the medium werealso identical between the cell lines, indicating that the efficiencyof utilization of phosphate by the selected cell line was similarto that by the wild-type cells. However, rate of uptake of phosphateby the selected cells under phosphate limited conditions (20µMNaH₂PO₄ at pH 5.6) was about 5-fold higher than that by thewild-type cells. Apparent K_m values for the uptake of phosphatewere calculated to be 13.6 and 9.1 µM for the selectedand the wild-type cells, respectively. The V_max valuewas estimatedto be 88.8 nmol per g fresh weight per min for the selectedcells and 28.2 for thewild-type cells. Thus, the selected cellshas an enhanced system for uptake of phosphate wherebytherewas an increase in the rate of the uptake without any dramaticchange in the affinity for phosphateions. (Received September 21, 1991; Accepted December 25, 1991)     

Energetic performance is improved by specific activation of K fluxes through KCa channels in heart mitochondria

Mitochondrial volume regulation depends on K⁺ movement across the inner membrane and a mitochondrial Ca²⁺-dependent K⁺ channel (mitoK_Ca) reportedly contributes to mitochondrial K⁺ uniporter activity. Here we utilize a novel K_Ca channel activator, NS11021, to examine the role of mitoK_Ca in regulating mitochondrial function by measuring K⁺ flux, membrane potential (ΔΨ_m), light scattering, and respiration in guinea pig heart mitochondria. K⁺ uptake and the influence of anions were assessed in mitochondria loaded with the K⁺ sensor PBFI by adding either the chloride (KCl), acetate (KAc), or phosphate (KH₂PO₄) salts of K⁺ to energized mitochondria in a sucrose-based medium. K⁺ fluxes saturated at ∼ 10 mM for each salt, attaining maximal rates of 172 ± 17, 54 ± 2.4, and 33 ± 3.8 nmol K⁺/min/mg in KCl, KAc, or KH₂PO₄, respectively. NS11021 (50 nM) increased the maximal K⁺ uptake rate by 2.5-fold in the presence of KH₂PO₄ or KAc and increased mitochondrial volume, with little effect on ΔΨ_m. In KCl, NS11021 increased K⁺ uptake by only 30% and did not increase volume. The effects of NS11021 on K⁺ uptake were inhibited by the K_Ca toxins charybdotoxin (200 nM) or paxilline (1 μM). Fifty nanomolar of NS11021 increased the mitochondrial respiratory control ratio (RCR) in KH₂PO₄, but not in KCl; however, above 1 μM, NS11021 decreased RCR and depolarized ΔΨ_m. A control compound lacking K_Ca activator properties did not increase K⁺ uptake or volume but had similar nonspecific (toxin-insensitive) effects at high concentrations. The results indicate that activating K⁺ flux through mitoK_Ca mediates a beneficial effect on energetics that depends on mitochondrial swelling with maintained ΔΨ_m.