Acid-Base Regulation by Azolla spp. with N2 as Sole N Source and with Supplementation by NH+4 or NO−3

Diazotrophic cultures of three species of Azolla (Az. caroliniana, Az. microphylla, Az. pinnata) symbiotic with Anabaena azollae accumulated some 0.10–0.24 mol organic anion per mol N assimilated (0.010–0.018 mol organic anion per mol C assimilated), with a corresponding efflux of 0.05–0.11 mol H⁺ per mol N assimilated (0.006–0.009 mol H⁺ per mol C assimilated). These values are lower than those found for terrestrial diazotrophic vascular plants; this may be related to the decreased possibility of increasing Fe and P availability by rhizosphere acidification in a free-floating plant. Modification of the organic anion content, and of the quantity (and direction) of H⁺ exchange with the medium, with 5 mol m^?3 NH⁺₄ or NO^?₃ added to diazotrophic cultures, are consistent with substantial N acquisition from combined N as well as N₂ assimilation. This conclusion is consistent with previously published work with ¹⁵N.     

Carbon transfer associated with assimilation of organic nitrogen sources by silver birch (Betula pendula Roth.)

Summary Qualitative and quantitative aspects of heterotrophic carbon assimilation by mycorrhizal plants of birch (Betula pendula) were examined. Plants were grown aseptically from seed in the mycorrhizal condition with the fungus Hebeloma crustuliniforme and in the non-mycorrhizal condition, with protein as their sole exogenous nitrogen source. Yields and nitrogen contents were determined in some of the plants, while the roots of others were supplied with ¹⁴C-labelled protein and their shoots exposed for up to 72 h to different irradiance regimes. Only mycorrhizal plants utilised the organic nitrogen. Uptake of carbon associated with this utilisation and its translocation to the leaves was demonstrated directly by means of autoradiography. Amounts of activity transferred to shoots were greatest in low irradiance regimes. Calculation of net carbon gain from the heterotrophic source, based upon the assumption that breakdown products of protein are assimilated as amino-acids, indicates that over a 55-day growth period up to 9% of plant C may be derived from protein. The physiological and ecological significance of these findings are discussed.     

CARBON SKELETON SOURCES FOR AMMONIUM ASSIMILATION IN N-SUFFICIENT AND N-LIMITED CELLS OF CYANIDIUM CALDARIUM (RHODOPHYTA)1

The possible origin of carbon skeletons for ammonium assimilation in Cyanidium caldarium (Tilden) Geitler was investigated. N-sufficient cells assimilated ammonium at a rate of 182 ± 18 μmol·mL packed cell volume (pcv)^-1· h^-1. Removal of CO₂ or darkening almost immediately prevented ammonium assimilation. N-limited cells in light assimilated ammonium at a rate of 493 ± 45 μmol · mL pcv^-1· h^-1 in the presence of CO₂ and at a lower rate of 168 ± 17 μmol · mL pcv^-1· h^-1 in the absence of CO₂. In darkness they assimilated ammonium at a rate of 293 ± 29 μmol · mL pcv^-1 h^-1 in the presence of CO₂, only 60% of the assimilation rate in light. In the absence of CO₂, ammonium was assimilated at a similar rate of 325 ± 14 μmol · mL pcv^-1· h^-1. Under the latter conditions, however, assimilation was inhibited after 40 min and ceased after 70 min; it resumed upon resupply of CO₂. We suggest that N-sufficient cells of C. caldarium obtain carbon skeletons for ammonium assimilation exclusively by photosynthetic reactions. Upon N-limitation they develop the ability, apparently through derepression or activation of regulatory enzyme system(s), to obtain a consistent quantity of additional carbon skeletons and ATP from mobilization of carbon reserves. This enables the N-limited cell to assimilate ammonium not only in light but also in darkness, and at a higher rate than N-sufficient cells. The fact that ammonium assimilation in light occurs at a higher rate than in darkness suggests that ammonium assimilation in light is the sum of both light and dark ammonium assimilation, which implies separate metabolic reactions for the two processes. These results suggest the existence of two distinct and differently controlled pathways in N-limited cells, but not in N-sufficient cells, through which carbon skeletons for ammonium assimilation originate. An important role for dark CO₂ fixation in dark or light ammonium assimilation is also indicated.     

The Glyoxylate Cycle in an Arbuscular Mycorrhizal Fungus. Carbon Flux and Gene Expression

The arbuscular mycorrhizal (AM) symbiosis is responsible for huge fluxes of photosynthetically fixed carbon from plants to the soil. Lipid, which is the dominant form of stored carbon in the fungal partner and which fuels spore germination, is made by the fungus within the root and is exported to the extraradical mycelium. We tested the hypothesis that the glyoxylate cycle is central to the flow of carbon in the AM symbiosis. The results of (13)C labeling of germinating spores and extraradical mycelium with (13)C(2)-acetate and (13)C(2)-glycerol and analysis by nuclear magnetic resonance spectroscopy indicate that there are very substantial fluxes through the glyoxylate cycle in the fungal partner. Full-length sequences obtained by polymerase chain reaction from a cDNA library from germinating spores of the AM fungus Glomus intraradices showed strong homology to gene sequences for isocitrate lyase and malate synthase from plants and other fungal species. Quantitative real-time polymerase chain reaction measurements show that these genes are expressed at significant levels during the symbiosis. Glyoxysome-like bodies were observed by electron microscopy in fungal structures where the glyoxylate cycle is expected to be active, which is consistent with the presence in both enzyme sequences of motifs associated with glyoxysomal targeting. We also identified among several hundred expressed sequence tags several enzymes of primary metabolism whose expression during spore germination is consistent with previous labeling studies and with fluxes into and out of the glyoxylate cycle.     

Activity of the Enzymes of Carbon Metabolism in Sulfobacillus sibiricus under Various Conditions of Cultivation

The thermoacidophilic iron-oxidizing chemolithotroph Sulfobacillus sibiricus N1^T is characterized by steady growth and amplified cell yield when grown in vigorously aerated medium containing Fe²⁺, glucose, and yeast extract as energy sources. In this case, carbon dioxide, glucose, and yeast extract are used as carbon sources. Glucose is assimilated through the fructose-bisphosphate pathway and the pentose-phosphate pathway. The glyoxylate bypass does not function in S. sibiricus, and the tricarboxylic acid cycle is disrupted at the level of 2-oxoglutarate dehydrogenase. The presence of ribulose-bisphosphate carboxylase indicates that carbon dioxide fixation proceeds through the Calvin cycle. The activity of ribulose-bisphosphate carboxylase is highest in autotrophically grown cells. The cells also contain pyruvate carboxylase, phosphoenolpyruvate carboxylase, phosphoenolpyruvate carboxykinase, and phosphoenolpyruvate carboxytransphosphorylase.     

Differential utilization of nutrients during development by the xylophagous leafhopper,Homalodisca coagulata

Utilization of nutrients at different stages of development was examined for the xylophage,Homalodisca coagulata (Say). Survivorship and consumption rates of second-instar, fourth-instar and adult leafhoppers were measured daily on the hostsLagerstroemia indica L. andEuonymus japonica Thumb. Rates of consumption, assimilation efficiencies and daily assimilation of nitrogen, carbon, and individual organic compounds were calculated based on chemical analyses of xylem fluid and insect excreta. Gross growth efficiencies of diet utilization were also estimated by comparing biomass of young adults to estimates of nutrient utilization of the two host species. Different instars survived and utilized nutrients at varying rates on the two hosts. Second-instar leafhoppers survived at higher rates and utilized nitrogen more efficiently onE. japonica than onL. indica. However, assimilated nitrogen was much less as a result of lower consumption rates. In contrast, adults onL. indica had increased longevity, utilized carbon more efficiently, and assimilated higher quantities of both carbon and nitrogen than those onE. japonica. Efficiencies of nutrient utilization were high forH. coagulata compared to folivores or phloem feeders, particularly in the conversion of ingested nutrients to assimilated compounds. Variations in diet utilization during development are discussed in terms of polyphagy.     

The pathway of carbon dioxide assimilation in rhodospirillum rubrum grown in turbidostat continuous-flow culture

Summary A comparison of light and dark short-term incorporation of [¹⁴C]-carbon dioxide by Rhodospirillum rubrum grown in turbidostat continuous-flow culture at two different steady states on medium containing malate has shown that the labelling of phosphate esters was the main light-dependent process. Thus, the reductive pentose phosphate cycle appears to be the major pathway of carbon dioxide assimilation in the light under these growth conditions.The labelling of glutamate was also light-dependent and was most marked in the most rapidly growing steady state culture.The assimilated [¹⁴C]carbon was transferred to metabolites of the tricarboxylic acid cycle, particularly C₄-dicarboxylic acids, and the transfer involved additional carboxylations which were not light-dependent. The activity of these reactions accounted for initial high rates of carbon dioxide assimilation in the dark.In the dark assimilated [¹⁴C]carbon accumulated in succinate.     

Métabolisme de l’Acide 2-Aminoéthylphosphonique (Ciliatine) chez le Poulet

For Podospora anserina, several studies of cellulolytic enzymes have been established, but characteristics of amylolytic enzymes are not well understood. When P. anserina grew in starch as carbon source, it accumulated glucose, nigerose, and maltose in the culture supernatant. At the same time, the fungus secreted α-glucosidase (PAG). PAG was purified from the culture supernatant, and was found to convert soluble starch to nigerose and maltose. The recombinant enzyme with C-terminal His-tag (rPAG) was produced with Pichia pastoris. Most rPAG produced under standard conditions lost its affinity for nickel-chelating resin, but the affinity was improved by the use of a buffered medium (pH 8.0) supplemented with casamino acid and a reduction of the cultivation time. rPAG suffered limited proteolysis at the same site as the original PAG. A site-directed mutagenesis study indicated that proteolysis had no effect on enzyme characteristics. A kinetic study indicated that the PAG possessed significant transglycosylation activity.     

Anaplerotic metabolism of Aspergillus nidulans and its effect on biomass synthesis in carbon limited chemostats

Anaplerotic fixation of carbon dioxide by the fungus Aspergillus nidulans when grown under carbon-limited conditions was mediated by pyruvate carboxylase and a phosphoenol pyruvate (PEP)-metabolising enzyme which has been tentatively designated as PEP carboxylase. The activities of both enzymes were growth rate dependent and measurements of H¹⁴CO₃ incorporation by growing mycelium indicated that they were responsible for almost all the assimilated carbon dioxide. In carbon-limited chemostats, the maximum rate of bicarbonate assimilation occurred at a dilution rate of 0.11 h^–1, equivalent to 1/2 _max. The affinity of the pyruvate carboxylase for bicarbonate was twice that of the PEP carboxylase under the conditons of growth used. The effect of changing the bicarbonate concentration in carbon-limited chemostats was substantial: increasing the HCO ₃ ^– concentration over the range 0.7–2.8 mM enhanced biomass synthesis by 22%. Over-shoots in bicarbonate assimilation and carboxylase activity occurred when steady state chemostat cultures were subjected to a step down in dilution rate.     

Both Forward and Reverse TCA Cycles Operate in Green Sulfur Bacteria

The anoxygenic green sulfur bacteria (GSBs) assimilate CO₂ autotrophically through the reductive (reverse) tricarboxylic acid (RTCA) cycle. Some organic carbon sources, such as acetate and pyruvate, can be assimilated during the phototrophic growth of the GSBs, in the presence of CO₂ or HCO₃⁻. It has not been established why the inorganic carbonis required for incorporating organic carbon for growth and how the organic carbons are assimilated. In this report, we probed carbon flux during autotrophic and mixotrophic growth of the GSB Chlorobaculum tepidum. Our data indicate the following: (a) the RTCA cycle is active during autotrophic and mixotrophic growth; (b) the flux from pyruvate to acetyl-CoA is very low and acetyl-CoA is synthesized through the RTCA cycle and acetate assimilation; (c) pyruvate is largely assimilated through the RTCA cycle; and (d) acetate can be assimilated via both of the RTCA as well as the oxidative (forward) TCA (OTCA) cycle. The OTCA cycle revealed herein may explain better cell growth during mixotrophic growth with acetate, as energy is generated through the OTCA cycle. Furthermore, the genes specific for the OTCA cycle are either absent or down-regulated during phototrophic growth, implying that the OTCA cycle is not complete, and CO₂ is required for the RTCA cycle to produce metabolites in the TCA cycle. Moreover, CO₂ is essential for assimilating acetate and pyruvate through the CO₂-anaplerotic pathway and pyruvate synthesis from acetyl-CoA.     

Oxidation and Assimilation of Atmospheric Methane by Soil Methane Oxidizers

The metabolism of atmospheric methane in a forest soil was studied by radiotracer techniques. Maximum (sup14)CH(inf4) oxidation (163.5 pmol of C cm(sup-3) h(sup-1)) and (sup14)C assimilation (50.3 pmol of C cm(sup-3) h(sup-1)) occurred at the A(inf2) horizon located 15 to 18 cm below the soil surface. At this depth, 31 to 43% of the atmospheric methane oxidized was assimilated into microbial biomass; the remaining methane was recovered as (sup14)CO(inf2). Methane-derived carbon was incorporated into all major cell macromolecules by the soil microorganisms (50% as proteins, 19% as nucleic acids and polysaccharides, and 5% as lipids). The percentage of methane assimilated (carbon conversion efficiency) remained constant at temperatures between 5 and 20(deg)C, followed by a decrease at 30(deg)C. The carbon conversion efficiency did not increase at methane concentrations between 1.7 and 1,000 ppm. In contrast, the overall methane oxidation activity increased at elevated methane concentrations, with an apparent K(infm) of 21 ppm (31 nM CH(inf4)) and a V(infmax) of 188 pmol of CH(inf4) cm(sup-3) h(sup-1). Methane oxidizers from soil depths with maximum methanotrophic activity respired approximately 1 to 3% of the assimilated methane-derived carbon per day. This apparent endogenous respiration did not change significantly in the absence of methane. Similarly, the potential for oxidation of atmospheric methane was relatively insensitive to methane starvation. Soil samples from depths above and below the zone with maximum atmospheric methane oxidation activity showed a dramatic increase in the turnover of the methane assimilated (>20 times increase). Physical disturbance such as sieving or mixing of soil samples decreased methane oxidation and assimilation by 50 to 58% but did not alter the carbon conversion efficiency. Ammonia addition (0.1 or 1.0 (mu)mol g [fresh weight](sup-1)) decreased both methane oxidation and carbon conversion efficiency. This resulted in a dramatic decrease in methane assimilation (85 to 99%). In addition, ammonia-treated soil showed up to 10 times greater turnover of the assimilated methane-derived carbon (relative to untreated soil). The results suggest a potential for microbial growth on atmospheric methane. However, growth was regulated strongly by soil parameters other than the methane concentration. The pattern observed for metabolism of atmospheric methane in soils was not consistent with the physiology of known methanotrophic bacteria.     

Genetic Variation in Wild and Hatchery Stocks of Suminoe Oyster (Crassostrea ariakensis) Assessed by PCR-RFLP and Microsatellite Markers

Genetic variation in wild Asian populations and U.S. hatchery stocks of Crassostrea ariakensis was examined using polymerase chain reactions with restriction fragment length polymorphism (PCR-RFLP) analysis of both the mitochondrial COI gene and the nuclear internal transcribed spacer (ITS) 1 region and using 3 microsatellite markers. Hierarchical analysis of molecular variance and pairwise comparisons revealed significant differentiation (P < 0.05) between samples from the northern region, represented by collections from China and Japan, and 2 of 3 samples from southern China. PCR-RFLP patterns were identified that were diagnostic for the northern (N-type) and southern (S-type) groups. Microsatellite marker profiles were used to assign each oyster to one of the two northern or two southern populations. Results for more than 97% of the oysters were consistent with the PCR-RFLP patterns observed for each individual in that oysters with N-type patterns were assigned to one of the northern populations and those with S-type patterns to one of the southern populations. At one site of the Beihai (B) region in southern China a mix of individuals with either the N-type or S-type PCR-RFLP genotypes was found. No heterozygotes at the nuclear ITS-1 locus were found in the sample, possibly indicating reproductive isolation in sympatry. Microsatellite assignment test results of the B individuals were also consistent with identifications as either the N-type or S-type based on PCR-RFLP patterns. The parental population for one hatchery stock was this B sample, which initially was composed of almost equal numbers of northern and southern genetic types. After hatchery spawns, however, more than 97% of the progeny fell into the northern genetic group by PCR-RFLP and microsatellite assignment test analyses, indicating that the individuals with the southern genotype contributed little to the spawn, owing to gametic incompatibility, differential larval survival, or a difference in timing of sexual maturity. Overall, results suggested that oysters collected as C. ariakensis in this study, and likely in other studies as well, include two different sympatric species with some degree of reproductive isolation.     

Ultrastructural Studies of the Intercellular Hypha aud Haustorium of Puccinia recondita f.sp. tritici

The ultrastructure of intercellular hyphae and D-hati-storia of P. recondita f.sp. tritici, and the host response to haustorial invasion, was investigated. The intercellular hyphae share common characteristics with those of other uredial stage rust fungi. Anastomosis was observed between intercellular hyphae. Two nucleoli were frequently observed in a single nucleus in the haustorium, indicating possible nuclear fusion between the two nuclei in D-haustoria of this fungus. The close association of host organelles, such as the nucleus, Golgi bodies, endo-plasmic reticulum, vesicles and mitochondria, with the developing haustorium, is described.     

Fusion of yeast protoplasts and isolated nuclei of Fusarium moniliforme

Protoplasts of a xylose-fermenting yeast strain (a fusion product of Pachysolen tannophilus and Saccharomyces cerevisiae) were fused with isolated nuclei of the xylan degrading filamentous fungus Fusarium moniliforme. Polyethyleneglycol 4000 was used as the fusogenic agent. Fourteen stable hybrids showing xylanase activity were obtained. It can be assumed that this ability was acquired from the nuclear genome of the fungus, since the parental yeast strain did not show any xylanase activity. The enzymatic activity was determined quantitatively. The parental strain of the fungus reached its maximum xylanase activity of 796 nkat/ml at 96 h of growth. Four of the hybrids had a xylanase activity of between 211 and 297 nkat/l at 24 h of growth. Zymograms of these hybrids showed the presence of xylanases when grown on xylan as the sole carbon source. Using pulse field electrophoresis gels, no difference between the chromosome pattern of the fusion products and the parental yeast strain was observed.     

Polysaccharide production by immobilized Aureobasidium pullulans cells in batch bioreactors

Cells of the fungus Aureobasidium pullulans ATCC 201253 were entrapped within 4% agar cubes or 5% calcium alginate beads and were examined for their production of the polysaccharide pullulan in batch bioreactors. The batch bioreactors were utilized twice for 168 hours of polysaccharide production in medium containing corn syrup as a carbon source. The agar-entrapped cells produced nearly equivalent pullulan concentrations during both production cycles. The alginate-entrapped cells produced higher polysaccharide levels during the second cycle compared to the levels observed during the initial cycle. The agar-entrapped cells elaborated a polysaccharide with a higher pullulan content than did the alginate-entrapped cells during both production cycles.     

Agricultural encroachment: implications for carbon sequestration in tropical African wetlands

Tropical wetlands have been shown to exhibit high rates of net primary productivity and may therefore play an important role in global climate change mitigation through carbon assimilation and sequestration. Many permanently flooded areas of tropical East Africa are dominated by the highly productive C₄ emergent macrophyte sedge, Cyperus papyrus L. (papyrus). However, increasing population densities around wetland margins in East Africa are reducing the extent of papyrus coverage due to the planting of subsistence crops such as Colocasia esculenta (cocoyam). In this paper, we assess the impact of this land use change on the carbon cycle and in particular the impacts of land conversion on net ecosystem carbon dioxide exchange. Eddy covariance techniques were used, on a campaign basis, to measure fluxes of carbon dioxide over both papyrus and cocoyam dominated wetlands located on the Ugandan shore of Lake Victoria. Peak rates of net photosynthetic CO₂ assimilation, derived from monthly diurnal averages of net ecosystem exchange, of 28–35 μmol CO₂ m^?2 s^?1 and 15–20 μmol CO₂ m^?2 s^?1 were recorded in the papyrus and cocoyam wetlands, respectively, whereas night‐time respiratory losses ranged between 10 and 15 μmol CO₂ m^?2 s^?1 at the papyrus wetland and 5–10 μmol CO₂ m^?2 s^?1 at the cocoyam site. The integration of the flux data suggests that papyrus wetlands have the potential to act as a sink for significant amounts of carbon, in the region of 10 t C ha^?1 yr^?1. The cocoyam vegetation assimilated ~7 t C ha^?1 yr^?1 but when carbon exports from crop biomass removal were accounted for these wetlands represent a significant net loss of carbon of similar magnitude. The development of sustainable wetland management strategies are therefore required to promote the dual wetland function of crop production and the mitigation of greenhouse gas emissions especially under future climate change scenarios.     

Inorganic carbon uptake kinetics of the stream macrophyte Callitriche cophocarpa Sendt.

Photosynthetic carbon uptake of Callitriche cophocarpa Sendt. was examined in plants collected from six Danish streams and in plants grown under variable inorganic carbon conditions in the laboratory. Both field and laboratory plants showed a low affinity for inorganic carbon (CO₂ compensation points ranging from 0.7 to 22 μM, and K_0.5(CO₂) from 51 to 121 μM), consistent with C-3 photosynthesis and use of CO₂ alone. Variation in inorganic carbon uptake characteristics was low in both groups of plants. Only in laboratory-grown plants was a coupling found between carbon uptake and the inorganic carbon regime of the medium. The carbon extraction capacity, expressed as a percentage of the initial amount of dissolved inorganic carbon (DIC) assimilated in PH-drift experiments, increased from −1.4 to 11.8% with declining external carbon availability, and the initial slope of the CO₂ response curve increased from 6.4 to 15.3 g⁻¹ h⁻¹ dm³. The plasticity of the inorganic carbon uptake system of C. cophocarpa was very low compared to the plasticity observed for submerged macrophytes with accessory carbon uptake systems (i.e. HCO₃⁻ use or C-4 photosynthesis), suggesting that the plasticity of the C-3 photosynthetic apparatus as such is restricted. The low carbon affinity of C. cophocarpa indicates that this species depends on CO₂ oversaturation for a sufficient supply of CO₂ for photosynthesis and growth.     

Changes in size,biomass and production of Euchlanis dilatata lucksiana Hauer during its lifespan

Changes in body length, carbon content and somatic and reproductive partitioning of assimilated carbon during the lifespan of Euchlanis dilatata lucksiana Hauer are described. The greatest increase in animal size was observed within the first two days of its lifespan, the biomass (expressed in carbon units) of females increased sharply between 1 and 22 h after hatching due to high somatic production, and between 53 and 65 h after hatching due to high reproduction rate. In her lifetime the Euchlanis female used only about 1% of the assimilated carbon for somatic production and as much as 16% of this carbon for reproduction. The possible reasons for, and adaptive advantages of, this phenomenon are discussed.     

Carbon assimilation and extracellular antigens of some yeast-like fungi

Many yeast-like fungi assimilated n-hexadecane, butylamine and putrescine as sole carbon sources. Methanol was not assimilated. This points to a physiological similarity to endomycetous, hydrocarbon-utilizing yeasts. Stephanoascus ciferrii assimilated uric acid, adenine and allantoin as sole source of carbon and nitrogen. All strains of Geotrichum candidum and many other yeast-like fungi assimilated acetoin and butan-2,3-diol. Assimilation tests for adenine, uric acid, allantoin, acetoin and butan-2,3-diol were found to be suitable for taxonomic purposes.Extracellular antigens immunologically related to those produced by Geotrichum candidum were detected in the cell-free culture liquids of several yeast-like fungi. The extracellular antigen excreted by Stephanoascus ciferrii was species-specific.     

Detailed description of developmental growth stages of Diplocarpon rosae in Rosa: a core building block for efficient disease management

Black spot disease of roses caused by the ascomycetous fungus Diplocarpon rosae teleomorph (Marssonina rosae anamorph) is a widespread and devastating disease. Despite considerable progress in the management of black spot disease in the recent years, it is still unclear by which mechanisms this fungus colonises and invades the host system, and without a good knowledge of such infection machineries, it not possible to fully overcome the challenges of D. rosae infection. By exploring research contributions up to date, we highlight in this review the ultrastructure of D. rosae infection cycle in the host cell by emphasising on several aspects related to its in vitro and in vivo germinations, the infection mechanisms of the fungus, the different fungal structures formed in the host cells and the optimum storage conditions for D. rosae to retain its viability and pathogenicity over time. Here, attention is particularly focused on the asexual life cycle of D. rosae, with the sexual cycle being briefly discussed. In addition, a new dimension of research approaches to effectively control black spot disease of roses, that is, how to accurately use the advanced biotechnology tools to speed up the current state of the disease management, is proposed here.