Fixation of [13N]N2 and transfer of fixed nitrogen in the Anthoceros-Nostoc symbiotic association

The initial product of fixation of [¹³N]N₂ by pure cultures of the reconstituted symbiotic association between Anthoceros punctatus L. and Nostoc sp. strain ac 7801 was ammonium; it accounted for 75% of the total radioactivity recovered in methanolic extracts after 0.5 min and 14% after 10 min of incubation. Glutamine and glutamate were the primary organic products synthesized from [¹³N]N₂ after incubation times of 0.5–10 min. The kinetics of labeling of these two amino acids were characteristic of a precursor (glutamine) and product (glutamate) relationship. Results of inhibition experiments with methionine sulfoximine (MSX) and diazo-oxonorleucine were also consistent with the assimilation of N₂-derived NH ₄ ⁺ by Anthoceros-Nostoc through the sequential activities of glutamine synthetase (EC 6.3.1.2) and glutamate synthase (EC 1.4.7.1), with little or no assimilation by glutamate dehydrogenase (EC 1.3.1.3). Isolated symbiotic Nostoc assimilated exogenous ¹³NH ₄ ⁺ into glutamine and glutamate and their formation was inhibited by MSX, indicating operation of the glutamine synthetase-glutamate synthase (GS-GOGAT) pathway: However, relative to free-living cultures, isolated symbiotic Nostoc assimilated 80% less exogenous ammonium into glutamine and glutamate, implying that symbiotic Nostoc could assimilate only a fraction of N₂-derived NH ₄ ⁺ . This implication was tested by using Anthoceros associations reconstituted with wild-type or MSX-resistant strains of Nostoc incubated with [¹³N]N₂ in the presence of MSX. The results of these experiments indicated that, in situ, symbiotic Nostoc assimilated about 10% of the N₂-derived NH ₄ ⁺ and that NH ₄ ⁺ was made available to Anthoceros tissue where it was apparently assimilated by the GS-GOGAT pathway. Since less than 1% of the fixed N₂ was lost to the suspension medium, it appears that transfer of NH ₄ ⁺ from symbiont to host tissue was very efficient in this extracellular symbiotic association.Abbreviations DON 6-diazo-5-oxo-l-norleucine - GDH glutamate dehydrogenase - GOGAT glutamate synthase - GS glutamine synthetase - MSX l-methionine-dl-sulfoximine     

Capacity of chromatiaceae for chemotrophic growth. Specific respiration rates of Thiocystis violacea and Chromatium vinosum

The capacity for chemoautotrophic, mixotrophic and organotrophic growth in the dark was tested with 45 strains of 17 species (11 genera) of the Chromatiaceae. The auxanographic deep agar shake culture method was used; the gas phase contained 5% O₂ and 1% CO₂ in N₂. All strains tested of Chromatium vinosum, C. minus, C. violascens, C. gracile, Thiocystis violacea, Amoebobacter roseus, Thiocapsa roseopersicina gave positive growth responses under chemoautotrophic and mixotrophic conditions (extra carbon source acetate); one strain of Thiocapsa roseopersicina grew also organotrophically on acetate alone. No growth was obtained with the remaining 17 strains of ten species. None of the five type species (three genera) of the Chlorobiaceae grew under chemotrophic conditions. With Thiocystis violacea 2311 a growth yield of 11.3g dry weight per mol thiosulfate consumed was obtained under chemoautotrophic conditions; under mixotrophic conditions with acetate the yield increased to 69g dry weight per mol thiosulfate consumed. With Thiocystis violacea 2311 maximal specific respiration rates were obtained with thiosulfate as electron donor irrespective of the presence or absence of sulfur globules in the cells; organic substrates served as carbon sources only and did not support respiration. With Chromatium vinosum D utilization of thiosulfate was not constitutive; maximal respiration rates on thiosulfate were obtained only with thiosulfate grown cells containing sulfur globules. Respiration rates were further increased by malate, fumarate or propionate; these substrates also served as sole electron donors for respiration. Acetate and pyruvate were used as carbon sources only. The ecological significance of the chemotrophic metabolism is discussed.     

Environmental factors affectingin vitro nitrogenase activity of cyanobacteria isolated from rice-fields

The nitrogen-fixing capacity of four cyanobacterial strains was tested in relation to heterotrophic ability, tolerance to combined nitrogen and adaptive capacity to changes in light intensity and pH. Strains (Anabaena variabilis UAM 202;Calothrix marchica UAM 214;Nodularia spumigena UAM 204,Nostoc punctiforme UAM 205) were isolated from the rice-fields of Valencia (Spain).C. marchica, was the only strain able to grow and to fix dinitrogen under heterotrophic conditions, with fructose and glucose. Fructose was the best substrate supporting growth and dinitrogen fixation in mixotrophy (growth in the light under conditions where CO₂ and organic carbon are assimilated simultaneously), photoheterotrophy (growth in the light on an organic compound in the absence of net CO₂ fixation) and heterotrophy (growth on an organic compound in the dark). Ammonium repressed nitrogenase more than nitrate. Full repression was observed only at concentrations of combined nitrogen higher than those usually found in rice-fields. Carbohydrates had a protective effect on nitrogenase against ammonium inhibition inC. marchica. All four strains showed increased nitrogenase activity when the light intensity was increased during assays. Variations of pH normally occurring in rice fields led to no important changes in nitrogenase activity inC. marchica. This fact, together with its potential for heterotrophic growth and tolerance to combined nitrogen, make this the most suitable of the four strains for inoculation experiments in rice fields.     

Regulating the ratio of photoautotrophic to heterotrophic metabolic activities in photoheterotrophic culture of Euglena gracilis and its application to α-tocopherol production

Methods of regulating the ratio of photoautotrophic to heterotrophic growth rates in photoheterotrophic culture of Euglena gracilis were investigated. In normal photoheterotrophic culture (in the presence of excess organic carbon), the cells grew mainly by organic carbon assimilation (heterotrophic metabolism). The relative contribution of photoautotrophic metabolism increased with the increase in the light supply coefficient, the increase in the CO₂ concentration in the aeration gas and the decrease in the feed rate of organic carbon source. However, limiting the organic carbon supply was the most effective method of shifting the metabolic balance to the photoautotrophic side. In the presence of excess organic carbon source, the -tocopherol contents of the cells in photoheterotrophic culture were low even when the light supply coefficient and CO₂ concentration in the aeration gas were high. By limiting the organic carbon supply to the photoheterotrophic culture, the intracellular content of -tocopherol increased to the same level as those obtained in photoautotrophic cultures.     

Heterotrophic growth of Thiobacillus acidophilus in batch and chemostat cultures

Heterotrophic growth of the facultatively chemolithoautotrophic acidophile Thiobacillus acidophilus was studied in batch cultures and in carbon-limited chemostat cultures. The spectrum of carbon sources supporting heterotrophic growth in batch cultures was limited to a number of sugars and some other simple organic compounds. In addition to ammonium salts and urea, a number of amino acids could be used as nitrogen sources. Pyruvate served as a sole source of carbon and energy in chemostat cultures, but not in batch cultures. Apparently the low residual concentrations in the steady-state chemostat cultures prevented substrate inhibition that already was observed at 150 M pyruvate. Molar growth yields of T. acidophilus in heterotrophic chemostat cultures were low. The Y _max and maintenance coefficient of T. acidophilus grown under glucose limitation were 69 g biomass · mol^–1 and 0.10 mmol · g^–1 · h^–1, respectively. Neither the Y _max nor the maintenance coefficient of glucose-limited chemostat cultures changed when the culture pH was increased from 3.0 to 4.3. This indicates that in T. acidophilus the maintenance of a large pH gradient is not a major energy-requiring process. Significant activities of ribulose-1,5-bisphosphate carboxylase were retained during heterotrophic growth on a variety of carbon sources, even under conditions of substrate excess. Also thiosulphate- and tetrathionate-oxidising activities were expressed under heterotrophic growth conditions.     

Identification and physiological characterization of the nitrogen fixing bacterium Corynebacterium autotrophicum GZ 29

The coryneform hydrogen bacterium strain GZ 29, assigned to Corynebacterium autotrophicum fixed molecular nitrogen under autotrophic (H₂, CO₂) as well as under heterotrophic (sucrose) conditions. Physiological parameters of nitrogen fixation were measured under heterotrophic conditions. The optimal dissolved oxygen concentration for cells grown in a fermenter with N₂ was rather low (0.14 mg O₂/l) compared with cells grown in the presence of NH ₄ ⁺ (4.45 mg O₂/l). C. autotrophicum GZ 29 had a doubling time of 3.7 h at 30°C with N₂ as N-source and sucrose as carbon source and at optimal pO₂. Acetylene reduction reached values of 12 nmoles of ethylene produced/minxmg protein. Although the oxygen concentration in the growing culture was kept constant, the optimal dissolved oxygen tension for the acetylene reduction assay shifted to higher pO₂-values. The overall efficiency of nitrogen fixation amounted to 22 mg N fixed/g sucrose consumed; it reached a maximal value of 65 mg N fixed/g sucrose consumed at the beginning of the exponential growth phase. Intact cells reduced acetylene even under anaerobic test conditions; further anaerobic metabolic activity could not be ascertained so far.     

不同营养方式对小球藻FACHB 484生长的影响及其非自养生长机理研究

系统研究了小球藻FACHB 484在含有葡萄糖的不同营养方式下的生长情况,并通过抑制试验探讨葡萄糖在小球藻FACHB 484光异养和兼养生长条件下所起的作用以及小球藻FACHB 484是否存在氧化呼吸系统的关键酶类。结果表明:小球藻FACHB 484可利用葡萄糖进行化能异养、光激活异养、光异养及兼养生长,其生长速率大小为:兼养光异养光激活异养化能异养光合自养。兼养培养的最大生物量和比生长速率分别是自养培养的8.6和3.4倍,其比生长速率接近于光合自养和光异养培养下的比生长速率之和。葡萄糖主要作为小球藻FACHB 484兼养和光异养培养的碳源,而能量主要源自光。小球藻FACHB 484存在氧化呼吸链代谢途径,其细胞中有琥珀酸脱氢酶和细胞色素氧化酶。       

Hydrogen oxidation and efficiency of nitrogen fixation in succinate-limited chemostat cultures ofRhizobium ORS 571

Rhizobium ORS 571, isolated from stem nodules of the tropical legumeSesbania rostrata is able to grow in the chemostat with molecular nitrogen as sole nitrogen source at a specific growth rate of 0.1 h^-1. Samples from nitrogenfixing cultures showed high acetylene reduction activities: 1,500 nmol ethylene formed per milligram dry weight per hour. Under nitrogen-fixing conditions an uptake hydrogenase is induced. Ammonia-assimilating cultures, without additional hydrogen, did not induce hydrogenase. The addition of hydrogen to succinate-limited nitrogen-fixing cultures resulted in an increase in the molar growth yield on succinate (Y _succinate) from 27 to 35 and a slight decrease in the molar growth yield on oxygen ( ), showing that hydrogen oxidation is less energy-yielding than the oxidation of endogenous substrates. Respiration-driven proton translocation measured with starved cells indicated the functioning of site 1 and 2 of oxidative phosphorylation. Cytochrome spectra showed that cytochromea ₆₀₀, present at high dissolved oxygen tension (d.o.t.) almost completely disappeared at low d.o.t. In flash-photolysis spectra only thea-type cytochrome could be detected as an oxidase in cells both grown at high and low d.o.t. Growth yields in ammonia-assimilating cultures were higher than those measured in nitrogen-fixing cultures. Assuming two sites of oxidative phosphorylation, a molar growth yield on ATP (Y _ATP) of about 3 and 6 was calculated for respecticely nitrogen-fixing and ammonia-assimilating cultures. TheY _ATP under nitrogen-fixing conditions is dependent on the amount of H₂ formed per mol N₂ fixed (H₂/N₂ ratio). A method has been described to calculate the total amount of ATP use by nitrogenase during the fixation of 1 mol N₂ (ATP/N₂ ratio) and H₂/N₂ ratios in aerobic nitrogen fixing organisms. This calculation yielded that nitrogen fixation inRhizobium ORS 571 is a high ATP-consuming process. The calculated ATP/N₂ and H₂/N₂ ratios were respectively 42 and 7.5.Abbreviations d.o.t. dissolved oxygen tension A preliminary account of this work was presented at the 5th International Symposium on Nitrogen Fixation, September 1983, Noordwijkerhout, The Netherlands     

Effect of nitrogen starvation on ammonium-inhibition of nitrogenase activity in Azotobacter chroococcum

When Azotobacter chroococcum cells grown in batch culture under N₂-fixing conditions were transferred to a medium lacking a nitrogen source, the cellular C/N ratio, the amount of alginic acid released into the external medium and the rate of endogenous respiration increased appreciably after 6 h to the exclusion of dinitrogen, whereas nitrogenase activity did not undergo any significant change. Nitrogen deficiency caused a decrease in the ammonium inhibition of nitrogenase activity from 95% inhibition at zero time to 14% after 6 h incubation under dinitrogen starvation, with no difference in the rate of ammonium utilization by N₂-fixing and N₂-starved cells being observed. This suggests that a balance of nitrogen and carbon assimilation is necessary for the ammonium inhibition of nitrogenase activity in A. chroococcum to take place.     

Using reverse micelles as microreactor for hydrogen production by coupled systems of Nostoc / R. palustris and Anabaena / R. palustris

Uptake hydrogenase negative mutants of bloom forming cyanobacteria (Nostoc and Anabaena) and the fermentative bacteria Rhodopseudomonas palustris P₄ were used together for producing hydrogen within the reverse micelles fabricated by N-ethyl hexyl sodium sulfosuccinate (AOT) in isooctane and cetyl trimethyl ammonium bromide (CTAB) in benzene. The rate of H₂ production in AOT/isooctane reverse micellar system was found to be more promising in comparison to the CTAB/Benzene reverse micellar entrapment. After mutagenesis in 2.0% (v/v) ethyl methane sulphonate (EMS) mutants of Nostoc and Anabaena were selected on BG-11 plates (containing 2% agar) and then used for analysis of produced hydrogen. In comparison to the unmutated Nostoc with R. palustris (within AOT/isooctane) the coupled system of mutated Nostoc and R. palustris produced H₂ by 3.9-fold higher rate, which is 8.6 mmol H₂/h/mg protein. Whereas, mutated Anabaena coupled with R. palustris produced 4.8 times higher hydrogen production within (AOT)/isooctane reverse micelles in comparison to the unmutated Anabaena with R. palustris. Effect of nitrogen to carbon ratio (N/C) on hydrogen production was studied and Anabaena/R. palustris and Nostoc/R. palustris systems were, respectively, found to generate 11.2 and 9.8 mmol H₂/h/mg protein continuously for 3 days. Effects of temperature and light intensity were also investigated and we found that 32°C temperature and 1,000 Lux light intensity are the optimum values in these systems. Addition of sodium dithionite also resulted in further enhancement of the rate and duration of hydrogen production in both (mutated Nostoc/R. palustris and mutated Anabaena/R.␣palustris) systems.     

Isolation and characterization of a marine Anabaena sp. capable of rapid growth on molecular nitrogen

A marine filamentous cyanobacterium capable of rapid growth under N₂-fixing conditions has been isolated from the Texas Gulf Coast. This organism appears to be an Anabaena sp. and has been given the strain designation CA. Cultures grown on mineral salts medium bubbled with 1% CO₂-enriched air at 42°C show a growth rate of 5.6±0.1 generations per day with molecular nitrogen as the sole nitrogen source. This growth rate is higher than any other reported in the literature to date for heterocystous cyanobacteria growing on N₂. Under similar growth conditions, 7.5 mM NH₄Cl yields a growth rate of 6.6±0.1 generations per day while 7.5 mM KNO₃ allows for a growth rate of 5.8±0.4 generations-day. Nitrogen-fixation rates, as measured by acetylene reduction, show maximum activity values in the range of 50–100 nmoles ethylene produced/minxmg protein. These values compare favorably with those obtained from heterotrophic bacteria and are much higher than values reported for other cyanobacteria. Growth experiments indicate that the organism requires relatively high levels of sodium and grows maximally at 42°C. Because of its high growth rate on N₂, this newly isolated organism appears ideal for studying nitrogen metabolism and heterocyst development among the cyanobacteria.     

Alterations in growth and crassulacean acid metabolism (CAM) activity of in vitro cultured cactus

Unlike C-3 plants, cacti possess a crassulacean acid metabolism (CAM) physiology that can alter the pattern of carbon uptake and affect plant growth under artificial environmental conditions, especially in tissue culture. In vitro-derived plantlets of Coryphantha minima grew 7-fold larger than plants cultured under similar ex vitro conditions. Growth regulators incorporated into the culture media during shoot proliferation stage of micropropagation had a strong influence on this increased growth. Other important factors that contributed to increased growth under in vitro conditions were high relative humidity and sugar in the culture medium. An analysis of gas exchange and daily fluctuations of malic acid levels revealed an increase in net photosynthetic rate, in terms of carbon assimilation, by in vitro plants compared with that of ex vitro plants. This stimulated photosynthesis in the presence of an external carbon source was unexpected but apparently true for cacti exhibiting CAM physiology. Unlike CAM plants grown in ex vitro conditions, net CO₂ uptake by in vitro-cultured cacti occurred continuously in the light as well as the dark. Once regenerated, cacti were transferred to ex vitro conditions where the normal CAM pathway resumed with a concomitant reduction in growth and CO₂ uptake. These results showed that growth of cacti can be considerably accelerated by in vitro culture. This revised version was published online in June 2006 with corrections to the Cover Date.     

Nitrate respiration,denitrification, and utilization of nitrogen sources by aerobic carbon monoxide-oxidizing bacteria

We describe the ability of carboxydotrophic bacteria for nitrate respiration or denitrification. Four out of fourteen strains examined could denitrify heterotrophically forming N₂ (Pseudomonas carboxydoflava) or N₂O (Pseudomonas carboxydohydrogena, Pseudomonas compransoris, and Pseudomonas gazotropha). Three carried out a heterotrophic nitrate respiration (Arthrobacter 11/x, Azomonas B1, and Azomonas C2). P. carboxydohydrogena could use H₂ as electron donor for nitrate respiration under chemolithoautotrophic growth conditions. CO did not support denitrification or nitrate respiration of carboxydotrophic bacteria, although the free energy changes of the reactions would be sufficiently negative to allow growth. CO at 50 kPa was a weak inhibitor of N₂O-reduction in carboxydotrophic and non-carboxydotrophic bacteria and decelerated denitrifying growth. Carboxydotrophic bacteria could utilize a wide range of N-sources. Results obtained with a plasmid-cured mutant of Pseudomonas carboxydovorans OM5 showed, that genes involved in nitrogen assimilation entirely reside on the chromosome. In the presence of an suitable electron donor, most carboxydotrophic bacteria could carry out a reduction of nitrate to nitrite that did not support growth and did not lead to the formation of ammonia.This article is dedicated to Professor Hans G. Schlegel on the occasion of his 65th birthday and in admiration for his élan and eternal idealism     

Lichens toGunnera — with emphasis onAzolla

Summary N₂-fixing cyanobacteria occur in symbiotic associations with fungi (ascomycetes) as lichens and with a few green plants. The associated cyanobacterium is always a species ofNostoc orAnabaena. Only a small number of plant genera are involved but there is a remarkable range of host diversity. Associations occur with several bryophytes (e.g.Anthoceros, Blasia, Cavicularia), a pteridophyte (Azolla), cycads (nine genera includingMacrozamia andEncephalartos) and an angiosperm (Gunnera). Except forGunnera, where the cyanobacterium penetrates the plant cells, the cyanobacteria are extracellular with specialized morphological modifications and/or structures of the host plant organs providing an environment which facilitates interaction with the prokaryote.Salient aspects of current knowledge pertaining to the establishment, perpetuation, and functioning of the individual symbioses are summarized. Where possible this includes information concerning recognition and specificity, mode(s) of infection, morphological modifications/adaptations of the host plant and a synopsis of morphological, physiological and biochemical changes common to the symbiotic cyanobacteria. The latter encompasses heterocyst frequencies, enzymes involved in ammonia assimilation, photosynthetic capability and metabolic interaction with the host.TheAzolla-Anabaena symbioses, which have potential agronomic significance as an alternative nitrogen source and maintain continuity with the endophyte through the sexual cycle, are emphasized.     

The occurrence of fimbriae on a N 2 2 -fixing cyanobacterium which occurs in lichen symbiosis

The Nostoc cyanobiont of the lichen Peltigera canina when grown on N₂ possesses, in the motile stage, discrete unbranched non-flagellar appendages (fimbriae or pili). These arise from the host cell surface in a peritrichous manner, have an axial hole, are 7.0 ±0.3 nm in diameter and are up to 3 m long. They do not haemagglutinate guinea pig red blood corpuscles and differ from the major fimbrial types reported for Gram-negative heterotrophic bacteria and from sex pili. They may be involved in motility and specificity in symbiotic cyanobacteria.     

Integrated nitrogen,carbon, and water relations of a xylem-tapping mistletoe following nitrogen fertilization of the host

Xylem-tapping mistletoes transpire large volumes of water (E) while conducting photosynthesis (A) at low rates, thus maintaining low instantaneous wateruse efficiency (A/E). These gas-exchange characteristics have been interpreted as a means of facilitating assimilation of nitrogen dissolved at low concentration in host xylem water; however, low A/E also results in substantial heterotrophic carbon gain. In this study, host trees (Juniperus osteosperma) were fertilized and gas exchange of mistletoe (Phoradendron juniperinum) and host were monitored to determine whether mistletoe A/E would approach that of the host if mistletoes were supplied with abundant nitrogen. Fertilization significantly increased foliar N concentrations (N), net assimilation rates, and A/E in both mistletoe and host. However, at any given N concentration, mistletoes maintained lower A and lower A/E than their hosts. On the other hand, when instantaneous water-use efficiency and A/N were calculated to include heterotrophic assimilation of carbon dissolved in the xylem sap of the host, both water-use efficiency and A/N converged on host values. A simple model of Phoradendron carbon and nitrogen budgets was constructed to analyze the relative benefits of nitrogen- and carbonparasitism. The model assumes constant E and includes feedbacks of tissue nitrogen concentration on photosyn-thesis. These results, combined with our earlier observation that net assimilation rates of mistletoes and their hosts are approximately matched (Marshall et al. 1994), support part of the nitrogen-parasitism hypothesis: that high rates of transpiration benefit the mistletoe primarily through nitrogen gain. However, the low ratio of A/E is interpreted not as a means of acquiring nitrogen, but as an inevitable consequence of an imbalance in C and N assimilation.This research was supported by the National Science Foundation (grants BSR-8706772 and 8847942).     

Effect of <Emphasis Type="Italic">Nostoc</Emphasis> sp. on soil characteristics,plant growth and nutrient uptake

The arid desert area is expanding by 6 million ha year⁻¹. Nostoc, a terrestrial cyanobacterium, is known as a pioneer organism which can photosynthesize, fix atmospheric nitrogen, and secrete polysaccharides. In order to explore the potential of Nostoc as a countermeasure to soil desertification, we have investigated the effects of Nostoc on the chemical and physical properties of soil and on plant growth in outdoor and laboratory experiments. The effect of Nostoc on soil properties, when applied on the surface of soil in plastic containers, was determined after cultivation for 90 days outdoors. Its effect on plant growth and nutrient uptake was determined by growing Nostoc with Komatsuna (Brassica rapa var. peruviridis) on a soil-less culture in a plastic petri dish incubated inside a growth chamber for 14 days. Two colonies of Nostoc with different shapes were also examined for their tolerance to dryness and salinity. Nostoc application resulted in an increase in the organic carbon and nitrogen content of the surface soil and enhanced plant growth and plant iron uptake, but soil temperature and moisture were maintained. Spherical-shaped Nostoc showed a higher tolerance to dryness and salinity than irregular-shaped Nostoc, and the former was able to rectify problematic soil better than the latter. The results indicate that the application of Nostoc to soil has a potential for increasing soil organic matter and reclaiming degraded soil ecosystems. Presented at the 6th Meeting of the Asian Pacific Society of Applied Phycology, Manila, Philippines.     

Autotrophic growth of four Sulfolobus strains on tetrathionate and the effect of organic nutrients

Autotrophic growth yields of four strains of Sulfolobus using tetrathionate as sole energy substrate fell in the range 6.2–7.8 g dry weight (mol tetrathionate oxidized)^-1. Autotrophic organisms lacked ribulose 1,5-bis-phosphate carboxylase, but contained pyruvate and phosphoenolpyruvate carboxylases. S. brierleyi and strains B6-2 and LM exhibited mixotrophic growth, with tetrathionate oxidation, CO₂-fixation and organic substrate assimilation occurring concurrently, using media containing glucose or acetate. Yeast extract or succinate supported heterotrophic growth and showed strain-dependent repression of one or both of tetrathionate oxidation and CO₂-fixation resulting in biphasic growth. All four carbon atoms of succinate were assimilated to cell-carbon during growth. Acetate was the major source of cell-carbon during mixotrophic growth. These observations are not inconsistent with the possibility of a reductive carboxylic acid cycle in these organisms. Radiorespirometric analysis of glucose oxidation indicated CO₂ release to occur by means of an Entner-Doudoroff pathway (followed by pyruvate decarboxylation) and oxidative pentose phosphate pathway reactions. There was little evidence from the glucose radiorespirometry of the large-scale use of an oxidative tricarboxylic acid cycle for terminal oxidation of acetate derived from pyruvate. These results demonstrate the considerable metabolic versatility of Sulfolobus strains and show that there is significant variation among them.Abbreviations PIPES Piperazine-N,N-bis (2-ethane sulphonic acid)     

Photolithotrophy,photoheterotrophy, and chemoheterotrophy: Patterns of resource utilization on an annual and a diurnal basis within a pelagic microbial community

An annual investigation of rates of photolithotrophy, photoheterotrophy, and chemoheterotrophy utilizing glucose and bicarbonate was made within the pelagic zone of a small, hardwater, southwestern Michigan lake. Sampling proceeded on a monthly, diurnal, and depth-wise basis. Annual mean photoheterotrophic uptake was estimated at 2.6g C m^–3h^–1. Two periods of relatively high activity were observed: one during spring overturn and the second during the late summer period. In general, greatest contributions to overall carbon cycling occurred during morning to midday incubation periods and at intermediate depths within the water column. Rates of chemoheterotrophy averaged 6.9g C m^–3h^–1 and were relatively uniform throughout the annual period. Greatest overall chemoheterotrophic activity was associated with periods of overturn. In general, this activity increased throughout the day and with increasing depth within the water column. The annual mean for photolithotrophic fixation was 1.33 mg C m^–3h^–1. Greatest contributions to rates of photosynthesis were associated with epilimnetic waters during early morning and midday incubations. Relatively minor contributions to inorganic fixation were made by waters below the 6-meter contour. Spring overturn and late summer represented periods of particularly great photolithotrophic activity. Quantitative comparisons among carbon pathways indicate that rates of pelagic heterotrophy, both photo- and chemoheterotrophy combined, contribute small quantities of carbon to overall carbon metabolism in this oligotrophic system. Qualitative comparisons among pathways indicate strong spatial and temporal separation. The late summer period showed greatest seasonal separation of the three pathways. Spring values represented a period of relatively high activity for all three pathways. On a depth-wise basis, photolithotrophic activity was greatest near the surface and chemolithotrophic activity greatest near the bottom. Photoheterotrophy took an intermediate position between the two. Diurnally, photoheterotrophy and photolithotrophy showed greatest activity during midday and early morning periods, whereas chemoheterotrophy increased throughout the daylight period and reached maximal values in sunset incubations.     

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.