Foliar nitrogen uptake in broadleaf evergreen Mediterranean forests: Fertilisation experiment with labelled nitrogen.

Atmospheric nitrogen (N) deposition in Mediterranean sclerophyllous forests of Holm oak (Quercus rotundifolia, Q. ilex) in Spain often exceeds empirical critical loads established for ecosystem conservation. There are still uncertainties on the capacity of canopy retention and uptake of the atmospheric N deposited of these forests. Studying and analysing all the forest nitrogen-cycle processes is essential to understand the potential effect of N deposition in these ecosystems. This study conducted a year-long short-term fertilisation experiment with labelled ammonium (15N-NH4) and nitrate (15N-NO3) to estimate foliar N absorption rates and assess the influence of leaf phenology and meteorological seasonal variations. Fertilising solutions were prepared to simulate low and high wet N deposition concentration, based on data reported from previous studies. Additionally, ecophysiological and meteorological measurements were collected to explore potential relationships between absorption rates, plant activity, and weather conditions. The results showed that Holm oak leaves were able to absorb both oxidised and reduced N compounds, with higher rates of NH4+ absorption. N recovery of both NH4+ and NO3- was higher in the low concentration treatments, suggesting reduced effectiveness of absorption as concentration increases. Foliar absorption rates were leaf-age dependent, with the highest values observed in young developing leaves. Foliar uptake showed seasonal changes with a clear reduction during the summer, linked to drought and dry weather conditions, and showing also smaller leaf net assimilation and stomatal conductance. During the rest of the year, foliar N absorption was not clearly associated to plant physiological activity but with environmental conditions. Our findings suggest that Holm oak canopies could absorb an important part of the incoming N deposition, but this process is compound, season and leaf phenology dependent. Further research is therefore needed to better understand and model this part of the N cycle.

Ozone and Temperature May Hinder Adaptive Capacity of Mediterranean Perennial Grasses to Future Global Change Scenarios.

Climate warming is recognized as a factor that threatens plant species in Mediterranean mountains. Tropospheric ozone (O3) should also be considered as another relevant stress factor for these ecosystems since current levels chronically exceed thresholds for plant protection in these areas. The main aim of the present study was to study the sensitivity of four Mediterranean perennial grasses to O3 and temperature based on plant growth, gas exchange parameters (photosynthesis-A, stomatal conductance-gs, and water use efficiency-WUE), and foliar macro- (N, K, Ca, Mg, P, and S) and micronutrients (B, Cu, Fe, Mn, Mo, and Zn) content. The selected species were grasses inhabiting different Mediterranean habitats from mountain-top to semi-arid grasslands. Plants were exposed to four O3 treatments in Open-Top chambers, ranging from preindustrial to above ambient levels, representing predicted future levels. Chamber-less plots were considered to study the effect of temperature increase. Despite the general tolerance of the grasses to O3 and temperature in terms of biomass growth, WUE and foliar nutrient composition were the most affected parameters. The grass species studied showed some degree of similarity in their response to temperature, more related with phylogeny than to their tolerance to drought. In some species, O3 or temperature stress resulted in low A or WUE, which can potentially hinder plant tolerance to climate change. The relationship between O3 and temperature effects on foliar nutrient composition and plant responses in terms of vegetative growth, A, gs, and WUE constitute a complex web of interactions that merits further study. In conclusion, both O3 and temperature might be modifying the adaptation capacity of Mediterranean perennial grass species to the global change. Air pollution should be considered among the driving favors of biodiversity changes in Mediterranean grassland habitats.

The Effects of Ozone on Visual Attraction Traits of Erodium paularense (Geraniaceae) Flowers: Modelled Perception by Insect Pollinators.

Ozone (O3) effects on the visual attraction traits (color, perception and area) of petals are described for Erodium paularense, an endangered plant species. Plants were exposed to three O3 treatments: charcoal-filtered air (CFA), ambient (NFA) and ambient + 40 nL L-1 O3 (FU+) in open-top chambers. Changes in color were measured by spectral reflectance, from which the anthocyanin reflectance index (ARI) was calculated. Petal spectral reflectance was mapped onto color spaces of bees, flies and butterflies for studying color changes as perceived by different pollinator guilds. Ozone-induced increases in petal reflectance and a rise in ARI under NFA were observed. Ambient O3 levels also induced a partial change in the color perception of flies, with the number of petals seen as blue increasing to 53% compared to only 24% in CFA. Butterflies also showed the ability to partially perceive petal color changes, differentiating some CFA petals from NFA and FU+ petals through changes in the excitation of the UV photoreceptor. Importantly, O3 reduced petal area by 19.8 and 25% in NFA and FU+ relative to CFA, respectively. In sensitive species O3 may affect visual attraction traits important for pollination, and spectral reflectance is proposed as a novel method for studying O3 effects on flower color.

Ozone uptake at night is more damaging to plants than equivalent day-time flux.

MAIN CONCLUSION: Plants exposed to equivalent ozone fluxes administered during day-time versus night-time exhibited greater losses in biomass at night and this finding is attributed to night-time depletion of cell wall-localised ascorbate. The present study employed Lactuca sativa and its closest wild relative, L. serriola, to explore the relative sensitivity of plants to ozone-induced oxidative stress during day-time versus night-time. By controlling atmospheric ozone concentration and measuring stomatal conductance, equivalent ozone uptake into leaves was engineered during day and night, and consequences on productivity and net CO2 assimilation rate were determined. Biomass losses attributable to ozone were significantly greater when an equivalent dose of ozone was taken-up by foliage at night compared to the day. Linkages between ozone impacts and ascorbic acid (AA) content, redox status and cellular compartmentation were probed in both species. Leaf AA pools were depleted by exposure of plants to darkness, and then AA levels in the apoplast and symplast were monitored on subsequent transfer of plants to the light. Apoplast AA appeared to be more affected by light-dark transition than the symplast pool. Moreover, equivalent ozone fluxes administered to leaves with contrasting AA levels resulted in contrasting effects on the light-saturated rate of CO2 assimilation (Asat) in both species. Once apoplast AA content recovered to pre-treatment levels, the same ozone flux resulted in no impacts on Asat. The results of the present investigation reveal that plants are significantly more sensitive to equivalent ozone fluxes taken-up at night compared with those during the day and were consistent with diel shifts in apoplast AA content and/or redox status. Furthermore, findings suggest that some thought should be given to weighing regional models of ozone impacts for extraordinary night-time ozone impacts.

Increased tropospheric ozone levels enhance pathogen infection levels of amphibians.

As a result of anthropogenic activities, changes to the chemistry of Earth's atmosphere pose a threat to ecosystem health and biodiversity. One such change is the increase in tropospheric ozone (O3), which is particularly severe in the Mediterranean basin area, where the levels of this pollutant are chronically high during spring and summer time. Within this region, Mediterranean mountain ecosystems are hot spots for biodiversity which may be especially vulnerable to changes in O3 levels. Declines in montane amphibian populations have been recorded worldwide, including the Mediterranean basin. A significant driver of these declines is the emerging infection disease, chytridiomycosis, caused by the aquatic fungus Batrachochytrium dendrobatidis (Bd). Chytridiomycosis has negatively affected populations of several amphibian species in the Spanish Central Range, including in the Sierra Guadarrama, and interactions with other biotic and abiotic factors are an important part of these declines. However, there is little evidence or knowledge of whether tropospheric O3 levels may be another factor in the outbreaks of this disease. To test the hypothesis that O3 levels are another interactive driver of Bd infection dynamics, two different approaches were followed: 1) an experimental study in open top chambers was used to quantify the aspects of how Bd infection progressed throughout the metamorphic process under four different O3 levels; and 2) a field epidemiological study was used to analyse the relationship between the Bd infection load in the Sierra de Guadarrama and tropospheric O3 levels during a 9 year period. Our results suggest that high O3 levels significantly delayed the rate of development of tadpoles and increased Bd infection, providing empirical evidence of two new separate ways that may explain population declines of montane amphibians.

Joining empirical and modelling approaches to estimate dry deposition of nitrogen in Mediterranean forests.

In Mediterranean areas, dry deposition is a major component of the total atmospheric N input to natural habitats, particularly to forest ecosystems. An innovative approach, combining the empirical inferential method (EIM) for surface deposition of NO3- and NH4+ with stomatal uptake of NH3, HNO3 and NO2 derived from the DO3SE (Deposition of Ozone and Stomatal Exchange) model, was used to estimate total dry deposition of inorganic N air pollutants in four holm oak forests under Mediterranean conditions in Spain. The estimated total deposition varied among the sites and matched the geographical patterns previously found in model estimates: higher deposition was determined at the northern site (28.9 kg N ha-1 year-1) and at the northeastern sites (17.8 and 12.5 kg N ha-1 year-1) than at the central-Spain site (9.4 kg N ha-1 year-1). On average, the estimated dry deposition of atmospheric N represented 77% ±â€¯2% of the total deposition of N, of which surface deposition of gaseous and particulate atmospheric N averaged 10.0 ±â€¯2.9 kg N ha-1 year-1 for the four sites (58% of the total deposition), and stomatal deposition of N gases averaged 3.3 ±â€¯0.8 kg N ha-1 year-1 (19% of the total deposition). Deposition of atmospheric inorganic N was dominated by the surface deposition of oxidized N in all the forests (means of 54% and 42% of the dry and total deposition, respectively). The relative contribution of NO2 to dry deposition averaged from 19% in the peri-urban forests to 11% in the most natural site. During the monitoring period, the empirical critical loads provisionally proposed for ecosystem protection (10-20 kg N ha-1 year-1) was exceeded in three of the four studied forests.

Validation of ozone response functions for annual Mediterranean pasture species using close-to-field-conditions experiments.

Ozone (O3) critical levels have been established under the Long-Range Transboundary Air Pollution Convention to assess the risk of O3 effects in European vegetation. A recent review study has led to the development of O3 critical levels for annual Mediterranean pasture species using plants growing in well-watered pots at a coastal site and under low levels of competition. However, uncertainties remain in the extrapolation of the O3 sensitivity of these species under natural conditions. The response of two O3-sensitive annual Mediterranean pasture Trifolium species at the coastal site was compared with the response of the same species growing at a continental site, in natural soil and subject to water-stress and inter-specific competition, representing more closely their natural habitat. The slopes of exposure- and dose-response relationships derived for the two sites showed differences in the response to O3 between sites attributed to differences in environmental growing conditions, growing medium and the level of inter-specific competition, but the effect of the individual factors could not be assessed separately. Dose-based O3 indices partially explained differences due to environmental growing conditions between sites. The slopes showed that plants were more sensitive to O3 at the continental site, but homogeneity of slopes tests revealed that results from both experimental sites may be combined. Although more experimental data considering complex inter-specific competition situations and the effect of important interactive factors such as nitrogen would be needed, these results confirm the validity of applying the current flux-based O3 critical level under close to natural growing conditions. The AOT40-based O3 critical level derived at the coastal site was also considered a suitable risk indicator in close to natural growing conditions in the absence of soil moisture limitations on plant growth.

Atmospheric deposition of inorganic nitrogen in Spanish forests of Quercus ilex measured with ion-exchange resins and conventional collectors.

Atmospheric nitrogen deposition is one of the main threats for biodiversity and ecosystem functioning. Measurement techniques like ion-exchange resin collectors (IECs), which are less expensive and time-consuming than conventional methods, are gaining relevance in the study of atmospheric deposition and are recommended to expand monitoring networks. In the present work, bulk and throughfall deposition of inorganic nitrogen were monitored in three different holm oak forests in Spain during two years. The results obtained with IECs were contrasted with a conventional technique using bottle collectors and with a literature review of similar studies. The performance of IECs in comparison with the conventional method was good for measuring bulk deposition of nitrate and acceptable for ammonium and total dissolved inorganic nitrogen. Mean annual bulk deposition of inorganic nitrogen ranged 3.09-5.43 kg N ha(-1) according to IEC methodology, and 2.42-6.83 kg N ha(-1) y(-1) using the conventional method. Intra-annual variability of the net throughfall deposition of nitrogen measured with the conventional method revealed the existence of input pulses of nitrogen into the forest soil after dry periods, presumably originated from the washing of dry deposition accumulated in the canopy. Important methodological recommendations on the IEC method and discussed, compiled and summarized.

Atmospheric pollutants in peri-urban forests of Quercus ilex: evidence of pollution abatement and threats for vegetation.

Peri-urban vegetation is generally accepted as a significant remover of atmospheric pollutants, but it could also be threatened by these compounds, with origin in both urban and non-urban areas. To characterize the seasonal and geographical variation of pollutant concentrations and to improve the empirical understanding of the influence of Mediterranean broadleaf evergreen forests on air quality, four forests of Quercus ilex (three peri-urban and one remote) were monitored in different areas in Spain. Concentrations of nitrogen dioxide (NO2), ammonia (NH3), nitric acid (HNO3) and ozone (O3) were measured during 2 years in open areas and inside the forests and aerosols (PM10) were monitored in open areas during 1 year. Ozone was the only air pollutant expected to have direct phytotoxic effects on vegetation according to current thresholds for the protection of vegetation. The concentrations of N compounds were not high enough to directly affect vegetation but could be contributing through atmospheric N deposition to the eutrophization of these ecosystems. Peri-urban forests of Q. ilex showed a significant below-canopy reduction of gaseous concentrations (particularly NH3, with a mean reduction of 29-38%), which indicated the feasibility of these forests to provide an ecosystem service of air quality improvement. Well-designed monitoring programs are needed to further investigate air quality improvement by peri-urban ecosystems while assessing the threat that air pollution can pose to vegetation.

Modelling the influence of peri-urban trees in the air quality of Madrid region (Spain).

Tropospheric ozone (O(3)) is considered one of the most important air pollutants affecting human health. The role of peri-urban vegetation in modifying O(3) concentrations has been analyzed in the Madrid region (Spain) using the V200603par-rc1 version of the CHIMERE air quality model. The 3.7 version of the MM5 meteorological model was used to provide meteorological input data to the CHIMERE. The emissions were derived from the EMEP database for 2003. Land use data and the stomatal conductance model included in CHIMERE were modified according to the latest information available for the study area. Two cases were considered for the period April-September 2003: (1) actual land use and (2) a fictitious scenario where El Pardo peri-urban forest was converted to bare-soil. The results show that El Pardo forest constitutes a sink of O(3) since removing this green area increased O(3) levels over the modified area and over down-wind surrounding areas.