Response of nitrogenase to altered carbon supply in a Frankia-Alnus incana symbiosis

To study the effect of altered carbon supply on nitrogenase (EC 1.7.99.2), plants of Alnus incana (L.) Moench in symbiosis with the local source of Frankia were exposed to darkness for 2 days, and then returned to normal light/dark conditions. During the dark period nitrogenase activity in vivo (intact plants) and in vitro ( Frankia cells supplied with ATP and reductant), measured as acetylene reduction activity, was almost completely lost. Western blots for both the Fe-protein (dinitrogenase reductase) and the MoFe-protein (dinitrogenase) showed that, in particular, the amount of MoFe-protein was strongly reduced during darkness. Protein stained sodium dodecyl sulphate-polyacrylamide gels of Frankia protein showed that the nitrogenase proteins were the only abundant proteins that clearly decreased during darkness. During recovery, studied for 4 days, nitrogenase activity in vivo recovered to the level before dark treatment but was still only half of control activity, Nitrogenase activity in vitro and the amount of MoFe-protein, both expressed per Frankia protein, recovered and reached similar values in previously dark treated plants and in control plants. The rate of recovery was similar to the increase in activity of control plants, suggesting growth of Frankia in addition to synthesis of nitrogenase proteins during the recovery after carbon starvation.     

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.     

Comparative studies of ectomycorrhiza formation in Alnus glutinosa and Pinus resinosa with Paxillus involutus

 Mycorrhiza ontogeny and details of Hartig net and mantle structure were compared in ectomycorrhizas synthesized in growth pouches between the broad host range fungus Paxillus involutus and the tree species European black alder (Alnus glutinosa) and red pine (Pinus resinosa). In Alnus glutinosa, a paraepidermal Hartig net was restricted to the proximal (basal) portion of first-order laterals; the hypodermal layer appeared to be a barrier to fungal penetration. Phi-thickenings were present in some cortical cells but these were not related to lack of fungal ingress into the cortex. The mantle was often present close to the root apex but in many roots it was loosely organized and patchy. In several instances, the mantle formed around the root apex was only temporary; renewed root growth occurred without the formation of a mantle. In Pinus resinosa, the Hartig net developed between cortical cell layers of monopodial and dichotomously branched first–order laterals. Fungal hyphae in the Hartig net exhibited a complex labyrinthine mode of growth. The mantle had a pseudoparenchymatous structure and covered the root, including apices of dichotomously branched roots. The Paxillus–Pinus resinosa interaction had all the characteristics of a compatible ectomycorrhizal association. The Paxillus–Alnus glutinosa interaction, however, showed only aspects of superficial ectomycorrhizas, including the presence of a minimal (sometimes absent) and mostly proximal Hartig net and variable mantle development. Sclerotia were produced in the extraradical mycelium of Paxillus involutus when associated with either Alnus glutinosa or Pinus resinosa. Accepted: 22 October 1998     

The potential of some soil amendments to improve tree growth on metalliferous mine wastes

Trees are commonly planted on metalliferous mine wastes as a means of reclaiming these contaminated substrates. Other land-uses are often limited. However tree establishment is often poor owing to hostile substrate properties such as acidity, low macronutrient concentrations, high trace element concentrations, poor substrate structure and poor water retention. Soil amendments are a potentially cheap amelioration option. A factorial experiment was set-up to investigate the effects of seven amendments on the substrate chemistry and the growth and leaf chemistries of alder and sycamore. The amendments included ammonium nitrate, mineral phosphate and sulphate of potash fertilizers, calcareous sand, a superexpanding polyacrylamide gel polymer, cake sewage sludge and diatomite. The last two amendments are used here in a novel capacity. The most successful amendments with regard to their growth and leaf chemistry effects were found to be cake sludge and diatomite for alder and mineral phosphate, calcareous sand and cake sludge for sycamore. The least successful amendments were ammonium nitrate, sulphate of potash and calcareous sand for alder and ammonium nitrate for sycamore. The results differed according to the autoecological properties of the two species, with alder being easily the most productive. The implications of these results for tree-planting schemes are discussed.     

European geography of Alnus incana leaf variation

The intra- and inter-population and geographic differentiation of Alnus incana were examined biometrically on the basis of morphological character of the leaves. The study results only partly confirmed the assumption that Carpathian and Alpine populations of the species differ significantly. This may indicate that gene exchange continued during glacial periods as these regions were not sufficiently isolated from each other. The geographic variation of grey alder along the Balkans–Scandinavia axis is more eminent. It is debatable whether A. incana has migrated northwards from montane refugia, along Central European rivers, mostly the Vistula and Oder. It can be, however, noted that the Masurian, Sudetan and East Carpathian populations are similar to one another.     

Variations in stomatal density and index: implications for palaeoclimatic reconstructions

The variation in stomatal characters in leaves from one Alnus glutinosa (L.) Gaertn. tree is analysed. Measurements were taken from over 70 sites on the abaxial surfaces of representative ‘sun’ and ‘shade’ leaves having the same insertion point. The mean values of stomatal density and index in the shade leaf were significantly lower (71 and 93%, respectively) than those for the sun leaf. Within leaves, up to 2.5-fold differences in stomatal density values were observed. Contour maps derived from the data reveal non-random trends over the leaf surface. Correlations between stomatal density, epidermal cell density and stomatal index indicate that the variation in stomatal density within leaves arose primarily from local differences in stomatal differentiation, rather than from local differences in leaf expansion. This research demonstrates that a high level of variation in stomatal characters occurs both within and between leaves. We conclude that a well-defined sampling strategy should be used when estimating stomatal characters for (tree) leaves. Furthermore, the leaf's insertion point and situation within the tree crown should be taken into account. We discuss the implications of these findings for palaeoclimatic interpretations and emphasize the need for great caution when drawing conclusions based solely on stomatal characters.     

Effects of elevated oxygen tensions on acetylene reduction in Alnus incana-Frankia symbioses

An open flow-through gas system was used to determine the effect of C₂H₂ and elevated O₂ on acetylene reduction activity (ARA) and respiration of the intact, potted root system of Alnus incana (L.) Moench in symbiosis with Frankia Avcll or with a local source of Frankia . Both symbiotic systems responded to C₂H₂ by an immediate plateau range in ARA. The Plateau in ARA was in some cases followed by a decline of less extent than reported for many legumes. A concurrent decline in net respiration of the root system was on average 8% of the CO₂ efflux prior to C₂H₂ introduction.
Respiration of the root systems in both symbioses responded to elevated oxygen levels in the 10 kPa C₂H₂ atmosphere by an increase of up to 17% of the net respiration prior to C₂H₂ introduction in 21 kPa O₂. In contrast, the elevated oxygen levels resulted in an immediate drop in ARA followed by a minor increase to a stable level lower than that at the preceding, lower oxygen tension. The symbiosis with the local Frankia had lost all ARA when the partial pressure of O₂ exceeded 50 kPa, whereas the symbiosis with Avcll still had some activity at 80 kPa O₂. This difference in tolerance of elevated O₂ clearly shows that the oxygen exclusion mechanisms may be controlled by the microsymbiont in Alnus-Frankia symbioses. The symbiotic systems recovered ARA to a similar extent when returned from elevated O₂ levels to 21 kPa O₂.     

Effect of changes in nitrogen nutrition on the polyamine content of Alnus glutinosa

Abstract. The principal polyamines in Alnus glutinosa roots, nodules and root pressure sap, putrescine, spermidine and spermine, were quantified by reversed-phase, high-performance liquid chromatography with fluorescence detection following precolumn derivatization with 9-fluorenylmethyl chloroformate and 1-ada-mantanamine. Putrescine was the major component of all tissues and sap. It comprised 70% or more of the polyamine pool except in roots of KNO₃-fed plants, in which similar amounts of putrescine and spermidine were present at levels five-fold lower than plants fed (NH₄)₂SO₄. Polyamine levels in nodules were 50% greater than in roots. The polyamine content of roots and nodules was not altered significantly when the nitrogen nutrition was changed from sole reliance on nitrogen fixation to partial or complete utilization of (NH₄)₂SO₄. However, the polyamine content of root pressure sap from nodulated plants increased almost four-fold when they were fed with increasing concentrations of NH₄NO₃, although the total polyamine content remained low (5mmol m⁻³ sap). The polyamine content of the Alnus root system changed with plant age. In particular, the spermidine content of both roots and nodules was higher in 10- as compared to 16-week-old plants.