MicroMundo: experimental project fostering contribution to knowledge on antimicrobial resistance in secondary school.

Antimicrobial resistance (AMR) has become a serious global health threat. Education could play a decisive role, so the scope of innovative educational projects, such as MicroMundo, should be analysed. MicroMundo is a service-learning project for the exploration of microbial biodiversity in soils in search of new antibiotics. In order to evaluate the contribution of MicroMundo to the improvement of knowledge about AMR as well as the optimal age of the participants, an open questionnaire before and after the educational intervention (for secondary school students) and a satisfaction survey (for all participants) were carried out. Programme instruction began in university and continued through secondary school; 14 teachers (phase 1) and 137 students from three educational levels (phase 2) participated. MicroMundo was successfully implemented, showing a statistically significant improvement in knowledge about antibiotics, resistance, health and environmental consequences, and possible measures to reverse the problem, at all educational levels. Furthermore, the satisfaction surveys revealed a growing interest in research and science. Therefore, students of general branches who have not yet opted for a scientific line would be the most suitable to conduct the programme. In conclusion, MicroMundo is recommended as a training method to raise awareness about AMR and promote scientific vocations.

Agricultural impacts on streams near Nitrate Vulnerable Zones: A case study in the Ebro basin, Northern Spain.

Agricultural intensification during the last century has caused river degradation across Europe. From the wide range of stressors derived from agricultural activities that impact rivers, diffuse agricultural pollution has received most of the attention from managers and scientists. The aim of this study was to determine the main stressors exerted by intensive agriculture on streams around Nitrate Vulnerable Zones (NVZs), which are areas of land that drain into waters polluted by nitrates according to the European Nitrate Directive (91/676/EEC). The study area was located in the NW of La Rioja (Northern Spain), which has some of the highest nitrate concentrations within the Ebro basin. The relationships between 40 environmental variables and the taxonomic and functional characteristics of the macroinvertebrate assemblages (which are useful indicators of water quality) were analyzed in 11 stream reaches differentially affected by upstream agricultural activity. The streams affected by a greater percentage of agricultural land cover in the surrounding catchment had significantly higher nitrate concentrations than the remaining sites. However, hydromorphological alteration (i.e. channel simplification, riparian forest and habitat degradation), which is closely linked to agricultural practices, was the main factor affecting macroinvertebrate assemblages. We suggest that "good agricultural practices" should be implemented in streams affected by NVZs to reverse stream degradation, in concordance with the European Water Framework Directive (WFD).

Effects of Didymosphenia geminata massive growth on stream communities: Smaller organisms and simplified food web structure.

This study aims to contribute to the understanding of the impact of Didymosphenia geminata massive growths upon river ecosystem communities' composition and functioning. This is the first study to jointly consider the taxonomic composition and functional structure of diatom and macroinvertebrate assemblages in order to determine changes in community structure, and the food web alterations associated with this invasive alga. This study was carried out in the Lumbreras River (Ebro Basin, La Rioja, Northern Spain), which has been affected by a considerable massive growth of D. geminata since 2011. The study shows a profound alteration in both the river community composition and in the food web structure at the sites affected by the massive growth, which is primarily due to the alteration of the environmental conditions, thus demonstrating that D. geminata has an important role as an ecosystem engineer in the river. Thick filamentous mats impede the movement of large invertebrates-especially those that move and feed up on it-and favor small, opportunistic, herbivorous organisms, mainly chironomids, that are capable of moving between filaments and are aided by the absence of large trophic competitors and predators -prey release effect-. Only small predators, such as hydra, are capable of surviving in the new environment, as they are favored by the increase in chironomids, a source of food, and by the reduction in both their own predators and other midge predators -mesopredator release-. This change in the top-down control affects the diatom community, since chironomids may feed on large diatoms, increasing the proportion of small diatoms in the substrate. The survival of small and fast-growing pioneer diatoms is also favored by the mesh of filaments, which offers them a new habitat for colonization. Simultaneously, D. geminata causes a significant reduction in the number of diatoms with similar ecological requirements (those attached to the substrate). Overall, D. geminata creates a community dominated by small organisms that is clearly different from the existing communities in the same stream where there is an absence of massive growths.

Carbon metabolism and bacteroid functioning are involved in the regulation of nitrogen fixation in Medicago truncatula under drought and recovery.

Regulation of symbiotic nitrogen fixation (SNF) during drought stress is complex and not yet fully understood. In the present work, the involvement of nodule C and N metabolism in the regulation of SNF in Medicago truncatula under drought and a subsequent rewatering treatment was analyzed using a combination of metabolomic and proteomic approaches. Drought induced a reduction of SNF rates and major changes in the metabolic profile of nodules, mostly an accumulation of amino acids (Pro, His, and Trp) and carbohydrates (sucrose, galactinol, raffinose, and trehalose). This accumulation was coincidental with a decline in the levels of bacteroid proteins involved in SNF and C metabolism, along with a partial reduction of the levels of plant sucrose synthase 1 (SuSy1). In contrast, the variations in enzymes related to N assimilation were found not to correlate with the reduction in SNF, suggesting that these enzymes do not have a role in the regulation of SNF. Unlike the situation in other legumes such as pea and soybean, the drought-induced inhibition of SNF in M. truncatula appears to be caused by impairment of bacteroid metabolism and N(2)-fixing capacity rather than a limitation of respiratory substrate.

Reduced carbon availability to bacteroids and elevated ureides in nodules, but not in shoots, are involved in the nitrogen fixation response to early drought in soybean.

Nitrogen fixation (NF) in soybean (Glycine max L. Merr.) is highly sensitive to soil drying. This sensitivity has been related to an accumulation of nitrogen compounds, either in shoots or in nodules, and a nodular carbon flux shortage under drought. To assess the relative importance of carbon and nitrogen status on NF regulation, the responses to the early stages of drought were monitored with two soybean cultivars with known contrasting tolerance to drought. In the sensitive cultivar ('Biloxi'), NF inhibition occurred earlier and was more dramatic than in the tolerant cultivar ('Jackson'). The carbon flux to bacteroids was also more affected in 'Biloxi' than in 'Jackson', due to an earlier inhibition of sucrose synthase activity and a larger decrease of malate concentration in the former. Drought provoked ureide accumulation in nodules of both cultivars, but this accumulation was higher and occurred earlier in 'Biloxi'. However, at this early stage of drought, there was no accumulation of ureides in the leaves of either cultivar. These results indicate that a combination of both reduced carbon flux and nitrogen accumulation in nodules, but not in shoots, is involved in the inhibition of NF in soybean under early drought.

Medicago truncatula root nodule proteome analysis reveals differential plant and bacteroid responses to drought stress.

Drought is one of the environmental factors most affecting crop production. Under drought, symbiotic nitrogen fixation is one of the physiological processes to first show stress responses in nodulated legumes. This inhibition process involves a number of factors whose interactions are not yet understood. This work aims to further understand changes occurring in nodules under drought stress from a proteomic perspective. Drought was imposed on Medicago truncatula 'Jemalong A17' plants grown in symbiosis with Sinorhizobium meliloti strain 2011. Changes at the protein level were analyzed using a nongel approach based on liquid chromatography coupled to tandem mass spectrometry. Due to the complexity of nodule tissue, the separation of plant and bacteroid fractions in M. truncatula root nodules was first checked with the aim of minimizing cross contamination between the fractions. Second, the protein plant fraction of M. truncatula nodules was profiled, leading to the identification of 377 plant proteins, the largest description of the plant nodule proteome so far. Third, both symbiotic partners were independently analyzed for quantitative differences at the protein level during drought stress. Multivariate data mining allowed for the classification of proteins sets that were involved in drought stress responses. The isolation of the nodule plant and bacteroid protein fractions enabled the independent analysis of the response of both counterparts, gaining further understanding of how each symbiotic member is distinctly affected at the protein level under a water-deficit situation.

The response of carbon metabolism and antioxidant defenses of alfalfa nodules to drought stress and to the subsequent recovery of plants.

Alfalfa (Medicago sativa) plants were exposed to drought to examine the involvement of carbon metabolism and oxidative stress in the decline of nitrogenase (N(2)ase) activity. Exposure of plants to a moderate drought (leaf water potential of -1.3 MPa) had no effect on sucrose (Suc) synthase (SS) activity, but caused inhibition of N(2)ase activity (-43%), accumulation of succinate (+36%) and Suc (+58%), and up-regulation of genes encoding cytosolic CuZn-superoxide dismutase (SOD), plastid FeSOD, cytosolic glutathione reductase, and bacterial MnSOD and catalases B and C. Intensification of stress (-2.1 MPa) decreased N(2)ase (-82%) and SS (-30%) activities and increased malate (+40%), succinate (+68%), and Suc (+435%). There was also up-regulation (mRNA) of cytosolic ascorbate peroxidase and down-regulation (mRNA) of SS, homoglutathione synthetase, and bacterial catalase A. Drought stress did not affect nifH mRNA level or leghemoglobin expression, but decreased MoFe- and Fe-proteins. Rewatering of plants led to a partial recovery of the activity (75%) and proteins (>64%) of N(2)ase, a complete recovery of Suc, and a decrease of malate (-48%) relative to control. The increase in O(2) diffusion resistance, the decrease in N(2)ase-linked respiration and N(2)ase proteins, the accumulation of respiratory substrates and oxidized lipids and proteins, and the up-regulation of antioxidant genes reveal that bacteroids have their respiratory activity impaired and that oxidative stress occurs in nodules under drought conditions prior to any detectable effect on SS or leghemoglobin. We conclude that a limitation in metabolic capacity of bacteroids and oxidative damage of cellular components are contributing factors to the inhibition of N(2)ase activity in alfalfa nodules.

Nitrogen fixation control under drought stress. Localized or systemic?

Legume-Rhizobium nitrogen fixation is dramatically affected under drought and other environmental constraints. However, it has yet to be established as to whether such regulation of nitrogen fixation is only exerted at the whole-plant level (e.g. by a systemic nitrogen feedback mechanism) or can also occur at a local nodule level. To address this question, nodulated pea (Pisum sativum) plants were grown in a split-root system, which allowed for half of the root system to be irrigated at field capacity, while the other half was water deprived, thus provoking changes in the nodule water potential. Nitrogen fixation only declined in the water-deprived, half-root system and this result was correlated with modifications in the activities of key nodule's enzymes such as sucrose synthase and isocitrate dehydrogenase and in nodular malate content. Furthermore, the decline in nodule water potential resulted in a cell redox imbalance. The results also indicate that systemic nitrogen feedback signaling was not operating in these water-stressed plants, since nitrogen fixation activity was maintained at control values in the watered half of the split-root plants. Thus, the use of a partially droughted split-root system provides evidence that nitrogen fixation activity under drought stress is mainly controlled at the local level rather than by a systemic nitrogen signal.