GROWTH AND NITROGEN FIXATION OF THE DIAZOTROPHIC FILAMENTOUS NONHETEROCYSTOUS CYANOBACTERIUM TRICHODESMIUM SP. IMS 101 IN DEFINED MEDIA: EVIDENCE FOR A CIRCADIAN RHYTHM1

Trichodesmium sp. IMS 101, originally isolated from coastal western Atlantic waters by Prufert-Bebout and colleagues and maintained in seawater-based media, was successfully cultivated in two artificial media. Its characteristics of growth, nitrogen fixation, and regulation of nitrogen fixation were compared to those of natural populations and Trichodesmium sp. NIBB 1067. Results indicate that the culture grown in artificial media had nitrogen fixation characteristics similar to those when the culture is grown in seawater-based medium and to those of Trichodesmium sp. in the natural habitat. The study provides practical artificial media to facilitate the physiological studies of these important diazotrophic cyanobacteria, as well as the cultivation of other Trichodesmium species in future studies. Manipulations of the light/dark cycle were performed to determine whether or not the daily cycle of nitrogen fixation is a circadian rhythm. Cultures grown under continuous light maintained the cycle for up to 6 days. We demonstrated that the daily cycle of nitrogen fixation in Trichodesmium sp. IMS 101 was at least partially under the control of a circardian rhythm.     

OXYGEN-INDUCED CHANGES IN MORPHOLOGY OF AGGREGATES OF APHANIZOMENON FLOS-AQUAE (CYANOPHYCEAE): IMPLICATIONS FOR NITROGEN FIXATION POTENTIALS1

The filamentous nitrogen-fixing cyanobacterium Aphanizomenon flos-aquae (L.) Ralfs forms bundle or fake shaped aggregates which can provide buoyancy control, protection against intense illumination, enhancement of phycosphere nutrient regeneration, and which may result from size-selective herbivory by zooplankton. The dimensions of aggregates can change quickly. In this study, after a period of darkness, illumination caused aggregates to elongate approximately five-fold over a 10-15 min period. The metamorphosis was reversible upon cessation of illumination and through successive light-dark cycles. Manipulations of environmental oxygen concentration and photosystem Ü activity (via DCMU amendment), together with measurements made inside flakes with O₂-sensitive microelectrocles, showed that the metamorphosis was a response to oxygen concentration and operated to enhance diffusive efflux of photosynthetically produced oxygen during illumination. During darkness oxygen concentration within contracted aggregates became severely depleted relative to the environment. We propose that metamorphic minimization of local oxygen concentration is an adaptation that enhances the ability of Aphanizomenon flos-aquae to fix atmospheric nitrogen via the oxygen-labile nitrogenase enzyme system.     

Heterotrophic metabolism and regulation of uptake hydrogenase activity in symbiotic cyanobacteria

The H₂ uptake activity of three cyanobionts isolated fromCycas revoluta, C. circinalis andazolla filiculoides was shown to be related primarily to the growth rate and independent of the main mode of carbon nutrition. Significant H₂ uptake was found in the coralloid roots ofCycas revoluta andZamia furfuracea (3 and 22 times higher than the respective C₂H₂ reduction activities). The results attained allow us to conclude that in cyanobacteria, in contrast to most nitrogen-fixing heterotrophs, uptake hydrogenase activity is not repressed by carbon substrates and that cyanobacteria in association seem to be endowed with sufficient H₂ uptake capacity to recover all of the H₂ released during the process of N₂-fixation.     

First data on epilithic heterocystous cyanobacteria from Roman hypogea

Abstract

Epilithic species of heterocystous cyanobacteria were isolated from Roman hypogea, grown at low photon flux densities and morphologically characterized. Determination of photosynthesis versus irradiance curves and nitrogen fixation activities on three of these strains, showed that their photosynthetic efficiency was low but sufficient to sustain nitrogen fixation.     

Respiration accumulates Calvin cycle intermediates for the rapid start of photosynthesis in Synechocystis sp. PCC 6803

We tested the hypothesis that inducing photosynthesis in cyanobacteria requires respiration. A mutant deficient in glycogen phosphorylase (?GlgP) was prepared in Synechocystis sp. PCC 6803 to suppress respiration. The accumulated glycogen in ΔGlgP was 250–450% of that accumulated in wild type (WT). The rate of dark respiration in ΔGlgP was 25% of that in WT. In the dark, P700⁺ reduction was suppressed in ΔGlgP, and the rate corresponded to that in (2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone)-treated WT, supporting a lower respiration rate in ?GlgP. Photosynthetic O₂-evolution rate reached a steady-state value much slower in ?GlgP than in WT. This retardation was solved by addition of d-glucose. Furthermore, we found that the contents of Calvin cycle intermediates in ?GlgP were lower than those in WT under dark conditions. These observations indicated that respiration provided the carbon source for regeneration of ribulose 1,5-bisphosphate in order to drive the rapid start of photosynthesis.     

Comparative adaptations of Aphanizomenon and Anabaena for nitrogen fixation under weak irradiance

1. In situ measurements of nitrogen fixation rates for Aphanizomenon in fertile Colorado lakes with low inorganic nitrogen concentrations demonstrated high efficiency of nitrogen fixation at low irradiance. 2. For study populations, rates of N₂ fixation in darkness and with alternating exposure to light and darkness were a higher percentage of light‐saturated rates for Aphanizomenon than for Anabaena, suggesting storage of reduced metabolites at high irradiance that are used subsequently by Aphanizomenon when cells are forced by mixing into zones of low irradiance. Also, saturation of N₂ fixation occurred over a lower range of irradiance for Aphanizomenon than for Anabaena. 3. High efficiency of N₂ fixation in Aphanizomenon at low or fluctuating irradiance is complementary to its previously demonstrated high efficiency of photosynthesis at low irradiance. Nitrogen fixation rate was also strongly related to DIN concentration; fixation was highest at low DIN (maximum < 5 μg L^?1) but was also most vulnerable to photoinhibition under such conditions. 4. The fixation capabilities of Aphanizomenon under weak or varying irradiance could explain its commonly observed domination over Anabaena when transparency is low and available nitrogen is scarce.     

Nitrate reductase activity, nitrogenase activity and photosynthesis of black alder exposed to chilling temperatures

Actinorhizal ( Frankia -nodulated) black alder [ Alnus glutinosa (L.) Gaertn.] seedlings fertilized with 0.36 m M nitrate (low nitrate fertilizer treatment) or 7.14 m M nitrate (high nitrate fertilizer treatment) and acclimated in a growth chamber for 2 weeks were exposed to 2.5 h of night-time chilling temperatures of −1 to 4°C. Cold treatment decreased nitrogenase activity (acetylene reduction activity) 33% for low nitrate fertilized plants and 41% for high nitrate fertilized plants. Recovery of nitrogenase activity occurred within 7 days after chilling treatment. In contrast, in vivo nitrate reductase (NR) activities of leaves and fine roots increased immediately after chilling then decreased as nitrogenase activities recovered. Fine roots of alder seedlings exhibited NR activities proportional to the amounts of nitrate in the rooting medium. In contrast, the NR activities of leaves were independent of substrate and tissue nitrate levels and corresponded to nitrogenase activity in the root nodules. In a separate experiment, net photosynthesis (PS) of similarly treated black alder seedlings was measured before and after chilling treatments. Net PS declined in response to chilling by 17% for plants receiving low nitrate fertilizer and 19% for plants receiving high nitrate fertilizer. After chilling, stomatal conductance (g_s) decreased by 39% and internal CO₂ concentration (c_i) decreased by 5% in plants receiving the high nitrate fertilizer, whereas plants receiving the low nitrate fertilizer showed no change in g_s and a 13% increase in c_i. Results indicate that chilling stimulates stomatal closure only at the high nitrate level and that interference with biochemical functions is probably the major impact of chilling on PS.     

‘Respiratory protection’ of the nitrogenase in dinitrogen-fixing cyanobacteria

Several filamentous and unicellular cyanobacteria were grown photoautotrophically with nitrate or dinitrogen as N-sources, and some respiratory properties of the cells or isolated plasma (CM) and thylakoid (ICM) membranes were compared. Specific cytochrome c oxidase activities in membranes from dinitrogen-fixing cells were between 10- and 50-times higher than those in membranes from nitrate-grown cells, ICM of heterocysts but CM of unicells being mainly responsible for the stimulation. Whole cell respiration (oxygen uptake) of diazotrophic unicells paralleled increased cytochrome oxidase activities of the isolated membranes. Mass spectrometric measurements of the uptake of isotopically labeled oxygen revealed that (low) light inhibited respiration of diazotrophic unicells to a much lesser degree than that of nitrate-grown cells which indicates the prevailing (respiratory) role of CM in the former. Normalized growth yields of diazotrophic unicells grown in continuous light were significantly higher than those of cells grown in a 12/12 hrs light/dark cycle. Mass spectrometry showed that overall nitrogen uptake by the former was higher than by the latter; in particular, and in marked contrast to the time course of nitrogenase activity (acetylene reduction) there was no appreciable nitrogen uptake or protein synthesis during dark periods; likewise, there was no 14-CO₂ fixation, nor chloropholl synthesis, nor cell division in the dark. By contrast, growth in continuous light gave sustained rates of nitrogen and carbon dioxide incorporation over the whole time range. Our results will be discussed in terms of respiratory protection as an essential strategy of keeping apart nitrogenase and oxygen, either atmospheric or photosynthetically produced within the same cell.     

Advantages of the division of labour for the long-term population dynamics of cyanobacteria at different latitudes

A fundamental advancement in the evolution of complexity is division of labour. This implies a partition of tasks among cells, either spatially through cellular differentiation, or temporally via a circadian rhythm. Cyanobacteria often employ either spatial differentiation or a circadian rhythm in order to separate the chemically incompatible processes of nitrogen fixation and photosynthesis. We present a theoretical framework to assess the advantages in terms of biomass production and population size for three species types: terminally differentiated (heterocystous), circadian, and an idealized species in which nitrogen and carbon fixation occur without biochemical constraints. On the basis of real solar irradiance data at different latitudes, we simulate population dynamics in isolation and in competition for light over a period of 40 years. Our results show that in isolation and regardless of latitude, the biomass of heterocystous cyanobacteria that optimally invest resources is comparable to that of the idealized unconstrained species. Hence, spatial division of labour overcomes biochemical constraints and enhances biomass production. In the circadian case, the strict temporal task separation modelled here hinders high biomass production in comparison with the heterocystous species. However, circadian species are found to be successful in competition for light whenever their resource investment prevents a waste of fixed nitrogen more effectively than do heterocystous species. In addition, we show the existence of a trade-off between population size and biomass accumulation, whereby each species can optimally invest resources to be proficient in biomass production or population growth, but not necessarily both. Finally, the model produces chaotic dynamics for population size, which is relevant to the study of cyanobacterial blooms.