The wide range of plant responses to ammonium nutrition can be used to study the way ammonium interferes with plant metabolism and to assess some characteristics related with ammonium tolerance by plants. In this work we investigated the hypothesis of plant tolerance to ammonium being related with the plants' capacity to maintain high levels of inorganic nitrogen assimilation in the roots. Plants of several species (Spinacia oleracea L., Lycopersicon esculentum L., Lactuca sativa L., Pisum sativum L. and Lupinus albus L.) were grown in the presence of distinct concentrations (0.5, 1.5, 3 and 6 mM) of nitrate and ammonium. The relative contributions of the activity of the key enzymes glutamine synthetase (GS; under light and dark conditions) and glutamate dehydrogenase (GDH) were determined. The main plant organs of nitrogen assimilation (root or shoot) to plant tolerance to ammonium were assessed. The results show that only plants that are able to maintain high levels of GS activity in the dark (either in leaves or in roots) and high root GDH activities accumulate equal amounts of biomass independently of the nitrogen source available to the root medium and thus are ammonium tolerant. Plant species with high GS activities in the dark coincide with those displaying a high capacity for nitrogen metabolism in the roots. Therefore, the main location of nitrogen metabolism (shoots or roots) and the levels of GS activity in the dark are an important strategy for plant ammonium tolerance. The relative contribution of each of these parameters to species tolerance to ammonium is assessed. The efficient sequestration of ammonium in roots, presumably in the vacuoles, is considered as an additional mechanism contributing to plant tolerance to ammonium nutrition.

Key enzymes of the urea cycle and 15 N-labeling patterns of arginine (Arg) were measured to elucidate the involvement of Arg in nitrogen translocation by arbuscular mycorrhizal (AM) fungi. Mycorrhiza was established between transformed carrot (Daucus carota) roots and Glomus intraradices in two-compartment petri dishes and three ammonium levels were supplied to the compartment containing the extraradical mycelium (ERM), but no roots. Time courses of specific enzyme activity were obtained for glutamine synthetase, argininosuccinate synthetase, arginase, and urease in the ERM and AM roots.15 NH 4 1 was used to follow the dynamics of nitrogen incorporation into and turnover of Arg. Both the absence of external nitrogen and the presence of L-norvaline, an inhibitor of Arg synthesis, prevented the synthesis of Arg in the ERM and resulted in decreased activity of arginase and urease in the AM root. The catabolic activity of the urea cycle in the roots therefore depends on Arg translocation from the ERM.15 N labeling of Arg in the ERM was very fast and analysis of its time course and isotopomer pattern allowed estimation of the translocation rate of Arg along the mycelium as 0.13 mg Arg mg 21 fresh weight h 21 . The results highlight the synchronization of the spatially separated reactions involved in the anabolic and catabolic arms of the urea cycle. This synchronization is a prerequisite for Arg to be a key component in nitrogen translocation in the AM mycelium.

Summary1. Atmospheric ammonia (NH 3 ) is one of the main drivers for ecosystem changes world-wide, including biodiversity loss. Modelling its deposition to evaluate its impact on ecosystems has been the focus of many studies. For that, universal indicators are needed to determine and compare the early effects of NH 3 across ecosystems. 2. We evaluate the effects of atmospheric NH 3 in ecosystems using lichens, which are one of the most sensitive communities at the ecosystem level. Rather than measuring total diversity, we use a functional diversity approach because this is potentially a more universal tool. 3. We evaluated the spatial and temporal patterns of atmospheric NH 3 concentrations ([NH 3 ] atm ) emitted from a point-source over a 1-year period in a cork oak Mediterranean woodland. We observed a temporal pattern of [NH 3 ] atm , with maximum concentrations during autumn. 4. The distribution of lichen species was c. 90% explained by [NH 3 ] atm . The tolerance of lichen species to atmospheric NH 3 , based on expert knowledge from literature, was tested for the first time against direct measurements of atmospheric NH 3 . Most species were well classified, with the exception of Lecanora albella and Chrysothrix candelaris, which were more tolerant than expected. Our updated lichen classification can be used to establish lichen functional groups that respond to atmospheric NH 3 , and these can be used in other Mediterranean countries. 5. Increasing [NH 3 ] atm led to a complete replacement of oligotrophic by nitrophytic species within 65 m of the NH 3 source. The geostatistical analysis of functional diversity variables yielded a spatial model with low non-spatial variance, indicating that these variables can cope robustly with high spatial variation in NH 3 . 6. Synthesis and applications. Our results support the use of functional diversity variables, such as a lichen diversity value, as accurate and robust indicators of the effects of atmospheric NH 3 on ecosystems. The spatial modelling of these indicators can provide information with high spatial resolution about the effects of atmospheric NH 3 around point-and diffuse sources. As this methodology is based on functional groups, it can be applied to monitor both the impact of atmospheric NH 3 and the success of mitigation strategies.

Ammonium nutrition is of interest as an alternative to that of using nitrate. However, the former has been reported as stressful to many plant species especially to some important crops, as most abiotic stresses may trigger oxidative imbalances in plants. In this work, we investigate the response of oxidative metabolism of two plant species, spinach (Spinacia oleracea L. cv. Gigante de invierno) and pea (Pisum sativum L. cv. Rondo), which have distinct tolerance to ammonium. Plants were grown in the presence of 1.5 and 3.0 mM N as ammonium and compared with equivalent nitrate nutrition. The antioxidant enzymes and metabolites as well as oxidative damage to proteins were determined. Protein and amino acid contents in both types of plants were also analysed. Ammonium nutrition in sensitive spinach or in the tolerant pea plants does not alter the redox status of ascorbate and glutathione or the phenolic contents, while no clear effect is seen in the antioxidant enzymes. The results showed that the stress originated from applying ammonium as the only N source is not an oxidative stress, independent of the ammonium tolerance of the plant species studied. Moreover, ammonium stress diminishes oxidative damage to proteins in the spinach plants. The data of the protein oxidation together with those from N metabolism highlight the relation between the stress induced by ammonium and an increased protein turnover.