Donana National Park Research Articles

A vegetation management experiment: Goats grazing shrublands in Doñana Natural Park

We have studied the effect of goat grazing on the shrub understory of a pine forest situated in a protected area (Donana Natural Park). Along three years we have studied the changes in phytovolume, flammability, species richness and diversity in a grazed shrubland and in a control area not grazed by the goats. We studied also food preferences of the goats and the ability of the goats to disperse the seeds of the plants they eat. Goat grazing significantly reduced shrub biomass and flammability, decreasing the risk of forest fire. Goats selected the species they eat, changing the pattern of selection throughout the year. Grazing reduced shrub diversity, but did not affected species richness. The goats can also potentially disperse the seeds of some of the species they eat, since the seeds were able to pass through the goat’s gut and germinated afterwards. Goat grazing on scrub vegetation can be used as an effective tool for the control of shrubs in protected forest areas, without losing biodiversity.

Use of metallomics and metabolomics to assess metal pollution in Doñana National Park (SW Spain).

Monitoring organism exposure to heavy metals has acquired increased importance in the last decades. The mouse Mus spretus has been used to assess the biological response to contaminants in the relevant ecological area of Doñana National Park (DNP) and surrounding areas (SW Spain), where many migrating birds land for breeding and feeding every year. A metallomics approach, based on the characterization of metal biomolecules using size exclusion chromatography coupled with inductively coupled plasma-mass spectrometry (SEC-ICP-MS) and a metabolomics approach based on direct infusion to a mass spectrometer (DI-ESI-QTOF-MS) followed by a partial linear square-discriminant analysis (PLS-DA), were used to compare the biological responses of M. spretus living in three areas of DNP (the reference) and surrounding areas (El Partido and El Matochal). The activities of key antioxidant enzymes, such as Cu/Zn-SOD, Mn-SOD, CAT, GR, and guaiacol peroxidase, were also determined in connection with environmental contamination issues. The results show differences caused by the presence of metals in the ecosystem that affected to the levels of metals and metalloproteins, such as MT, Cu/Zn-SOD, or Mn-CA, the breakdown of membrane phospholipids, perturbations in metabolic pathways, related to energy metabolism, and oxidative stress.

Redox proteomics as biomarker for assessing the biological effects of contaminants in crayfish from Doñana National Park

Despite its environmental relevance and sensitivity, Doñana National Park (DNP) is under high ecological pressure. In crayfish (Procambarus clarkii), the utility of redox proteomics as a novel biomarker was evaluated in the aquatic ecosystems of DNP and its surroundings, where agricultural activity is a serious concern. After fluorescence labeling of reversibly oxidized Cys and 2-DE separation, the total density of proteins with reversibly oxidized thiols was found to be much higher in animals from the Matochal (MAT) and Rocina (ROC) streams, while no difference was found in crayfish from Partido (PAR) stream compared to those from the DNP core at Lucio del Palacio (the negative control). The 2-DE analysis revealed 35 spots with significant differences in thiol oxidation, among which 19 proteins were identified via MALDI-TOF/TOF. While 3 spots, identified as ferritin, showed higher oxidation levels in ROC, other identified proteins were more intense at MAT than at ROC (superoxide dismutase, protein disulfide isomerase and actin) or were overoxidized only in MAT (nucleoside diphosphate kinase, fructose-biphosphate aldolase, fatty acid-binding protein, phosphopyruvate hydratase). For most of the identified proteins, spots corresponding to different Cys oxidized forms were detected, and the native forms, without oxidized thiol groups were also found in some of them. Evidence of reversible oxidation was found for specific Cys residues, including Cys13 in ferritin as well as Cys76 and Cys108 in nucleoside diphosphate kinase. The identified thiol-oxidized proteins provide information about the metabolic pathways and/or physiological processes affected by pollutant-elicited oxidative stress.

Omics technologies and their applications to evaluate metal toxicity in mice M. spretus as a bioindicator

Metals are important components of living organisms since many biological functions critically depend on their interaction with some metal in the cell. However, human activities have increased toxic metal levels in the terrestrial and aquatic ecosystems affecting living organisms. The impact of metals on cellular metabolism and global homeostasis has been traditionally assessed in free-living organisms by using conventional biomarkers; however, to obtain a global vision of metal toxicity mechanisms and the responses that metals elicit in the organisms, new analytical methodologies are needed. We review the use of omics approaches to assess the response of living organisms under metal stress illustrating the possibilities of different methodologies on the basis of our previous results. Most of this research has been based on free-living mice Mus spretus, a conventional bioindicator used to monitor metal pollution in Doñana National Park (DNP) (SW Spain), which is an important European biological reserve for migrating birds affected by agricultural, mining and industrial activities. The benefits of using omic techniques such as heterologous microarrays, proteomics methodologies (2-DE, iTRAQ®), metallomics, ionomics or metabolomics has been remarked; however, the complexity of these areas requires the integration of omics to achieve a comprehensive assessment of their environmental status. This article is part of a Special Issue entitled: Environmental and structural proteomics. This work presents new contributions in the study of environmental metal pollution in terrestrial ecosystems using Mus spretus mice as bioindicator in Doñana National Park (SW Spain) and surroundings. In addition, it has been demonstrated that the integration of omics multi-analytical approaches provides a very suitable approach for the study of the biological response and metal interactions in exposed and free-living mice (Mus musculus and Mus spretus, respectively) under metal pollution.

Rainfall stochasticity controls the distribution of invasive crayfish and its impact on amphibian guilds in Mediterranean temporary waters

Invasive crayfish have severely impacted invaded aquatic ecosystems worldwide. We studied temporal and spatial variation in the range expansion of the red swamp crayfish at one of the first European localities to which it was introduced: Donana National Park (SW Spain). In contrast to the rapid range expansion witnessed in other areas, this invasive crayfish has not spread across the entire park. Instead, its distribution has expanded during wet periods, but contracted during drought periods. The red swamp crayfish has caused steep amphibian declines in other invaded areas. However, after approximately 35 years of crayfish presence in Donana National Park, we have yet to detect a reduction in the number or occurrence of amphibian species. Amphibians may thus be protected by the large abundance of temporary ponds in the area, which provides them with an effective refuge network. We show that natural fluctuations in annual rainfall and in the number of ponds filled can temporarily eliminate invasive crayfish from particular areas. This fact should be taken into account when attempting to reduce the impact of crayfish on aquatic communities, intensifying crayfish removal during particularly dry years, when it is most effective.

Inorganic mass spectrometry-based metallomics for environmental monitoring of terrestrial ecosystems affected by metal pollution using Mus spretus as bioindicator

A metallomic approach based on the use of size-exclusion chroma tography coupled to inductively coupled plasma-mass spectrometry (SECICP-MS) has been used to characterize the bio logical response in liver, brain, kidneys, lungs and plasma of the free-living mou se Mus spretus in polluted areas located in Donana National Park (southwest Spain) and the surroundings, mainly affected by agriculture, mining and industry activities, which are responsible for the presence of metallic contaminants. It is remarkable the high presence of Cu, Z n, Cd, As, Pb and Ni in the cytosolic extracts of different organs and plasma, especially in co ntaminated areas. In liver extracts, high intensity peaks traced by Cu, Zn, Pb and Cd at 7 kDa (matching with metallothionein I standard) are triggered by the presence of contaminants. In kidn ey, similar Cu and Cd-peaks at 7 kDa were observed but the equivalent Zn-peak was depleted by the competitiv e interactions of Cu-Cd-Zn for the active sites of these molecules. In addition, peaks traced by Cu and Zn at about 32 kDa in liver extract match with superoxide dismuta se standard (Cu,Zn-SOD), which increase in accordance to contamination. An analogous behavior was observed for a Zn,Cu-peak at about 67 kDa that can be related with the bovine serum albumin standard (Cu,Zn-BSA ) or other carrier protein such as transferrin (Cu-Tf) present in liver and plasma. Finally, low molecular mass arsenic metabolites were detec ted in mice captured in MAT site affected by mine waste.

Application of metallomic and metabolomic approaches in exposure experiments on laboratory mice for environmental metal toxicity assessment

Metals have a central role in biological systems, regulating numerous cellular processes, and in other cases having toxic or deleterious effects on the metabolism. Hence, the study of metal-induced changes in cellular metabolic pathways is crucial to understanding the biological response associated with environmental issues. In this context, the finding of biomarkers has great interest, representing -omics techniques, such as metallomics and metabolomics, powerful tools for this purpose. The present work evaluates the exposure of mice Mus musculus to toxic metals (As, Cd and Hg), considering the changes induced in both the metallome and metabolome as a consequence of the high genetic homology between Mus musculus/Mus spretus mice, which allows the use of the database from M. musculus to identify the proteins and metabolites expressed by M. spretus. For this purpose a metallomic approach based on size exclusion chromatography (SEC) in combination with other complementary orthogonal separation techniques and heteroelement monitoring by ICP-ORS-qMS was performed, followed by identification of metallobiomolecules by organic mass spectrometry. In addition, simultaneous speciation of selenoproteins and selenometabolites in mouse plasma was accomplished by tandem (double) SEC-(dual) affinity chromatography (AF)-HPLC and online isotope dilution analysis (IDA)-ICP-ORS-qMS. Finally, the simultaneous changes in metabolic expression in mice caused by metal exposure (metabolome) were considered, using direct infusion mass spectrometry (DI-ESI-QqQ-TOF-MS) of extracts from mice plasma. Subsequently altered metabolites were identified using MS/MS experiments. The results obtained under controlled conditions were extrapolated to homologous free-living mice captured in Doñana National Park (DNP) and surroundings (southwest Spain) affected by As, Cd and Hg pollution. In summary, such studies are needed to understand the effect of heavy metal exposure and cope with heavy metal toxicity.

Distribution and Risk Assessment of Pharmaceutical Compounds in River Sediments from Doñana Park (Spain)

The occurrence of 16 pharmaceutical compounds in river sediments from Donana National Park was investigated, as well as the ecotoxicological risk to the ecosystems of this well-known protected zone. Eight of the 16 pharmaceuticals were present in sediments. The highest concentrations in river sediments concern to diclofenac, salicylic acid and caffeine (52.1, 27.2 and 25.4 μg/kg dm). Naproxen, carbamazepine, propranolol, 17β-estradiol and estriol were the other pharmaceutical compounds detected in sediment samples. The relation between the concentration of these pharmaceutical compounds in sediments and river water in contact with was also investigated. While some compounds were detected exclusively in sediment samples (diclofenac, 17β-estradiol and estriol), others were detected only in surface waters in contact with (ibuprofen, ketoprofen and gemfibrozil). The outcome analyses of the ecotoxicological risk assessment showed that the presence of all studied pharmaceutical compounds involved a high toxicological risk at short and long term to Donana ecosystem.

Influence of grazing on the decomposition of Pinus pinea L. needles in a silvopastoral system in Doñana, Spain

Aims The aim of this study was to determine whether goat grazing in the understory of a pine forest at Donana Natural Park could accelerate the decomposition of the pine needles accumulated on the soil surface and, if so, through which mechanisms. Specifically, the roles of trampling (mechanical fragmentation) and nutrient enrichment through defecation (fertilization) were evaluated in terms of their effect on pine needle decomposition rates.

Evolución paleoambiental del sector meridional del Parque Nacional de Doñana desde el Plioceno Inferior a la actualidad

The analysis of the geological units and the ostracod assemblages of a long core collected in the southern Donana National Park (SW Spain) permits to deduce an evolution from shallow marine palaeoenvironments (Lower Pliocene) to a brackish lagoon (Upper Pleistocene-Holocene) and the deposit of aeolian sediments (<1900 yr BP), with an intermediate alluvial stage during the Pleistocene. In the Late Holocene, a tsunamigenic event was detected, with the erosion of aeolian sediments and a subsequent deposit on subtidal environments.

The impact of the common rabbit on cork oak regeneration in SW Spain

Seed predation has traditionally been considered as simply a progressive loss of seeds. However, partial consumption of tree seeds may actually assist regeneration. We quantified partial consumption of acorns by the common rabbit (Oryctolagus cuniculus) in a cork oak forest in southwest Spain (Donana Natural Park). To examine the effects of partial consumption on germination and seedling emergence, we simulated two different levels of acorn predation (30 and 50 % cotyledon loss, representing light and heavy partial consumption, respectively). One-third of the sampled acorns presented evidence of consumption but most of the damage caused by partial consumption did not affect the acorn embryo (damage was mainly to non-vital parts of the propagule). Partial consumption caused more rapid germination and emergence than that was observed in intact acorns, but germination and emergence rate was unaffected. Heavy consumption of the acorn caused differences in root and shoot biomass as a consequence of cotyledon loss. In this greenhouse study, partial consumption was non-lethal to the acorns, and cotyledon biomass loss did not decrease the probability of successful establishment. Partially consumed seeds can therefore contribute significantly to the natural regeneration of Quercus coccifera, although it would be beneficial to conduct long-term studies focusing on seed traits and animal behaviour under field conditions.

Antibodies to West Nile virus and related flaviviruses in wild boar, red foxes and other mesomammals from Spain

Red foxes (Vulpes vulpes), wild boar (Sus scrofa) and Iberian pigs (Sus scrofa domestica) that are raised extensively outdoors, as well as other wild mesomammals from south central Spain and wild boar from Doñana National Park (DNP), were tested for antibodies against related flaviviruses by ELISA and for antibodies against WNV by VNT. Mean flavivirus seroprevalence according to ELISA was 20.4±7.8% (21 out of 103) in red foxes, 12.6±2.8% (69 out of 545) in wild boars, and 3.3±2.7% (6 out of 177) in Iberian pigs. A stone marten (Martes foina) also tested positive. Flavivirus seroprevalence in wild boar was significantly higher in DNP, and increased with age. Haemolysis of the serum samples limited interpretation of VNT to 28 samples, confirming WNV seroprevalence in one red fox, four Iberian pigs and nine wild boars. ELISA positive, microVNT negative samples suggest presence of non-neutralizing antibodies against WNV or antibodies to other antigenically related flaviviruses. Despite the importance of wetlands for flavivirus maintenance and amplification, WNV/flavivirus seroprevalence in wild boar and red foxes was not associated to wetland habitats. This is the first report of exposure of red foxes to WNV. With view to use of the tested species as sentinels for flavivirus activity, limited exposure of Iberian pigs that would be available for regular sampling, low numbers of foxes collected and concentration of wild boar harvest in the winter season are major drawbacks.

Batrachochytrium dendrobatidis infection of amphibians in the Doñana National Park, Spain

Amphibian chytridiomycosis, caused by infection with the non-hyphal, zoosporic chytrid fungus Batrachochytrium dendrobatidis (Bd), is an emerging infectious disease recognised as a cause of recent amphibian population declines and extinctions worldwide. The Doñana National Park (DNP) is located in southwestern Spain, a country with widespread Bd infection. This protected area has a great diversity of aquatic habitats that constitute important breeding habitats for 11 native amphibian species. We sampled 625 amphibians in December 2007 and February to March 2008, months that correspond to the early and intermediate breeding seasons for amphibians, respectively. We found 7 of 9 sampled species to be infected with Bd and found differences in prevalence between sampling periods. Although some amphibians tested had higher intensities of infection than others, all animals sampled were apparently healthy and, so far, there has been no evidence of either unusually high rates of mortality or amphibian population declines in the DNP.

Stability of the new alluvial fan of el partido stream, located in the eastern side of el rocío village in the doñana national park (spain), after the actions carried out to restore its hydrological regime

We present, first of fall, a recall of the formation of a new alluvial fan at El Partido stream between 1982 and 2006, located in the eastern side of El Rocio village, over the Donana National Park Marshes, as a result of the channelization of the mentioned stream with crop purposes in 1981. The settlement of the new alluvial fan, over an area of special protection, and the concerns linked to its expansion and growth, hindering the drainage of the urbanized area of El Rocio village after flood events, caused a strong impact. The analysis of the situation drove to the conclusion that the most suitable measure to stabilize the new alluvial fan was to restore the functionality of the former alluvial fan located upstream the study area. The restoration works were executed in 2006 and from the year 2007 the monitoring of its behavior in several flood events that occurred in the stream has been carried out. This paper summarizes the followed restoration scheme of the former alluvial fan and describes the main monitoring tasks, especially during the last 2 years focused specifically in the neighborhood of El Rocio village. To conclude, a description of the recent flood behavior along the final reach of the stream, as a result of the restoring works and its impact on the current stability of the new alluvial fan has been provided. Keywords flood defense, hydrological correction, stabilization of an alluvial fan, torrential event Language: en

Germination of hyphal bodies of Pythium spiculum isolated from declining cork oaks at Doñana National Park (Spain)

Pythium spiculum , a recently described new taxon, has been isolated from declining cork oaks ( Quercus suber L.) at Donana National Park (south-western Spain). The microorganism can infect and cause root disease in Quercus , but currently it is unknown whether its hyphal bodies can germinate and infect host trees. These hyphal bodies, regardless of shape, have been shown to be able to germinate, producing long germ tubes, sometimes ramified. Zoospore production was not recorded, but hyphal bodies are potential host infective structures in dry soil conditions.

Heavy flooding effects on home range and habitat selection of free-ranging Iberian hares (Lepus granatensis) in Doñana National Park (SW Spain)

The Iberian hare Lepus granatensis is a common and abundant species throughout the Iberian Peninsula; however, studies documenting its ecology are scarce. Between August 1996 and September 1997, after a long drought, we studied the effects of a widespread flood on the Iberian hare ecology in ecotone pastures of Donana National Park. Hares were captured using vertical nets through part of 1996 and 1997. Fourteen adult hares (nine females and five males) were tagged with radiocollars and located by triangulation on a daily basis at least once a week. Habitat use was estimated using a geographical information system. The average home range size was 28 ha for males and 24 ha for females. No significant differences were observed between sexes. Significant differences were observed in the use of scrub areas and in dry pastures. During the dry and wet seasons, males and females were most often located in the pastures of the ecotone, rush stands and scrub. The dense bracken (Pteridium aquilinum) and seaside bulrush (Scirpus maritimus) communities were avoided. These results are the first on the spatial ecology of this species in its native habitat.

Integrating detection probabilities in species distribution models of amphibians breeding in Mediterranean temporary ponds

AbstractAim The imperfect detection of species may lead to erroneous conclusions about species–environment relationships. Accuracy in species detection usually requires temporal replication at sampling sites, a time‐consuming and costly monitoring scheme. Here, we applied a lower‐cost alternative based on a double‐sampling approach to incorporate the reliability of species detection into regression‐based species distribution modelling.Location Doñana National Park (south‐western Spain).Methods Using species‐specific monthly detection probabilities, we estimated the detection reliability as the probability of having detected the species given the species‐specific survey time. Such reliability estimates were used to account explicitly for data uncertainty by weighting each absence. We illustrated how this novel framework can be used to evaluate four competing hypotheses as to what constitutes primary environmental control of amphibian distribution: breeding habitat, aestivating habitat, spatial distribution of surrounding habitats and/or major ecosystems zonation. The study was conducted on six pond‐breeding amphibian species during a 4‐year period.Results Non‐detections should not be considered equivalent to real absences, as their reliability varied considerably. The occurrence of Hyla meridionalis and Triturus pygmaeus was related to a particular major ecosystem of the study area, where suitable habitat for these species seemed to be widely available. Characteristics of the breeding habitat (area and hydroperiod) were of high importance for the occurrence of Pelobates cultripes and Pleurodeles waltl. Terrestrial characteristics were the most important predictors of the occurrence of Discoglossus galganoi and Lissotriton boscai, along with spatial distribution of breeding habitats for the last species.Main conclusions We did not find a single best supported hypothesis valid for all species, which stresses the importance of multiscale and multifactor approaches. More importantly, this study shows that estimating the reliability of non‐detection records, an exercise that had been previously seen as a naïve goal in species distribution modelling, is feasible and could be promoted in future studies, at least in comparable systems.

A comparison of robust polynomial fitting, global geopotential model and spectral analysis for regional–residual gravity field separation in the Doñana National Park (Spain)

Abstract Donana National Park is a protected area of approximately 500 km 2 located on the SW coast of Spain with singular and interesting ecological and geological features. A gravimetric survey is presented where L&R gravity metres were used in the gravimetric observations with GPS and high-precision levelling positioning. Bouguer gravity anomalies were computed and least squares prediction was used for gross-error detection. Robust polynomial fitting, the recent EGM2008 global geopotential model (complete to degree and order 2159), and spectral analysis were tested for regional–residual gravity field separation. A detailed description of the gravimetric characteristics of the Donana National Park is presented and the values of residual gravity anomalies were correlated with geological features, where the use of the EGM2008 global geopotential model has revealed an interesting tool for regional–residual gravity field separation. Finally, the interpretation of the results is justified by the well-known geological aspects of the park, but some modifications in the boundaries of the different geological features are needed in order to fit the modelled gravity with the residual gravity anomalies in the two cross-sections analysed.

Nonbiological factors affecting track censuses: implications for sampling design and reliability

Track census is a widely used method for rapid faunal assessments, which assumes that differences in track count numbers mainly reflect differences in species abundance due to some biological factors. However, some methodological and climatic variables might affect results of track censuses. Here, we tested the effect of climatic variables, such as maximum temperature, humidity, wind speed or days since last rain, and methodological factors such as censusing day period, distance from transect to vegetation edge, substrate condition or observer, on the number of tracks of mammal carnivores and some of their potential prey detected in sandy substrates. We sampled 2 × 2 km2 located within the scrubland area of Donana National Park (southwestern Spain) for carnivore and several potential prey tracks. Our results showed differences in the number of tracks detected between observers and a significant interaction between observers and the day period when censuses were carried out. Moreover, the variables increasing the quality of the substrate (higher environmental humidity, lower wind speed and days since last rain) not only led to a greater detection of carnivore tracks but, depending on the size of the species sampled other variables such as distance from transects to the vegetation border, also affected results. We recommend restricting sampling to certain fixed weather conditions when planning to monitor relative animal abundance from track censuses. When not possible, climatic or methodological variables should be included as covariates in analyses that try to test for the biological factors affecting wildlife abundance, taking into account that these variables, which affect the number of tracks detected could vary between years.

Characterization of 13 microsatellite loci for the Pygmy Marbled Newt Triturus pygmaeus (Salamandridae)

We have developed thirteen polymorphic microsatellite loci for the newt Triturus pygmaeus using an enriched-library approach. We have detected 171 alleles in 60 individuals genotyped (mean number of alleles per locus was 13.15) in two different populations in the Donana National Park, Southwestern Spain. Expected heterozygosities ranged from 0.16 to 0.956 in one population (Mata de los Dominguez) and from 0.1 to 0.959 in the other population (Navazo de la Higuera). The levels of polymorphism of the developed markers render them readily applicable for population genetic studies of diversity, structure, and migration.