Regulation and Spatiotemporal Patterns of Extracellular Enzyme Activities in a Coastal,Sandy Aquifer System (Doñana,SW Spain)

A seasonal study of extracellular enzyme activities (EEA) was conducted in the coastal, sandy aquifer system located in the greater fluvial–littoral ecosystem of Doñana (SW, Spain). Glucosidase, leucine aminopeptidase, alkaline phosphatase, and phenol oxidase activities were determined over a 2-year period in 30 piezometers spread in an area of approximately 100 km². Taking into account all enzymes, piezometers, and seasons, EEA ranged over several orders of magnitude, from 1.01?×?10^?5?±?2.92?×?10^?6 to 1.37?±?0.13 nmol (methylumbelliferyl, amido-4-methylcoumarin, or dihydroxyphenylalanine) mL^?1 h^?1. The quality, much more than the quantity, of organic matter and nutrients seemed to be the major variables that controlled the spatiotemporal patterns showed by EEA. EEA patterns obtained in this study agree with several functional models of microbial communities, such as optimal resource allocation and nutrient co-limitation. This study probably represents the first one in which these functional models have been tested in subsurface systems. Results obtained in this study seem to suggest that microbial communities inhabiting groundwaters in Doñana are not dead or compromised cells. By contrast, these communities play relevant roles in carbon and nutrient cycling, continue with the decomposition process that begins in the sediments of the shallow lakes located in the area, provide remineralized carbon and nutrients to producers of these surface aquatic systems, and close energy and matter cycles. This study proposes that groundwater systems should be considered dynamic systems, comparable in functional complexity to surface systems.     

Copepods and branchiopods of temporary ponds in the Doñana Natural Area (SW Spain): a four-decade record (1964–2007)

The Doñana Natural Area includes a large array of wetlands with the highest degree of environmental protection in Spain, and so it has long attracted many studies. We present a cumulative list of zooplankton taxa (Copepods and Branchiopods) based on a collection of 18 publications (1964–2007) and 4 unpublished studies. Seventy-eight taxa have been recorded in a set of 55 ponds, and 72 taxa at 38 sites spread over the Doñana marshland. In total, 96 taxa have been recorded, including 50% of all branchiopod species reported for the whole Iberian Peninsula. Taxa composition was significantly segregated between ponds and marshland during floods (ANOSIM test, R = 0.929, P < 0.01), but this segregation disappeared at a larger spatio-temporal scale when a non-metric MDS ordination produced a gradient from ponds to marshland (ANOSIM test, R = 0.272, P < 0.01). The lack of segregation between ponds and marshland sites, and among ponds with different hydroperiods, was not due to a large number of cosmopolitan species, but to a random distribution of a large number of low-occurrence species (67% of total taxa occurred with a frequency <15%). Long-hydroperiod ponds occupy a key position among the Doñana wetlands in terms of biodiversity as these ponds accumulated a high crustacean richness over time. They also supported a significantly higher cumulative number of cladoceran and harpacticoid taxa, while short-hydroperiod ponds accumulated the lowest number of diaptomid taxa. Our data indicate the need for recording biodiversity in the long term as richness on a short-temporal scale is not a good indicator of the number of crustacean species that would be encountered with a longer sampling period in Mediterranean temporary wetlands.     

First report of Eimeria myoxi in the garden dormouse (Eliomys quercinus Linnaeus, 1766) from Doñana Natural Area (Andalusia,SW Spain)

This work reports for the first time the presence and molecular characterization of Eimeria myoxi in the garden dormouse (Eliomys quercinus) from the Doñana Natural Area (Andalusia, SW Spain). Fresh faecal samples were collected from a total of 28 garden dormice, which were caught following current guidelines for the ethical use of animals in research, and processing by a standard flotation technique with saturated saline solution. Then, wet drops were examined microscopically, and the number of oocysts was semi-quantified. Eimeria oocysts were observed in 16 of the 28 (57.1%) faecal samples, showing most of them a very low number of oocysts (≤1 oocyst per microscopic field × 400). The unsporulated oocysts visualized in 16 faecal samples were subspherical and of length 19.2 ± 1.2 μm and width 17.4 ± 1.1 μm, being morphologically compatible with E. myoxi. This finding was supported by molecular analysis of the small subunit ribosomal RNA (SSU-rRNA) gene, identifying the same species in 22 of the 28 (78.6%) dormice, including 15 samples in which oocyst size was compatible with E. myoxi. Moreover, the subsequent analyses of the apicoplast open reading frame 470 (ORF470) and the mitochondrial cytochrome c oxidase subunit I (COI) genes confirmed the molecular identification of the isolates as E. myoxi. The phylogeny analyses were consistent with previous phylogenetic studies and support the existence of three lineages of rodent-infecting Eimeria species.     

Recovery of zooplankton diversity in a restored Mediterranean temporary marsh in Doñana National Park (SW Spain)

Over 30 years after drainage for agriculture, a 2700 ha temporary marshland was recently restored in Doñana National Park. We describe the recovery of zooplankton communities (copepods, cladocerans and rotifers) in 47 new temporary ponds excavated as part of the restoration project during the first two hydroperiods (April 2006 and 2007), and compare them to those of eight reference sites in the surrounding marshland. Major changes in the species composition and abundance occurred in new ponds between years. While rotifers and cyclopoid copepods dominated in terms of number of individuals in 2006, calanoid copepods and cladocerans were the most abundant groups in 2007. Rotifer species richness was significantly lower in 2007, but there was an increase in Simpson and β-diversity in 2007 owing largely to a dramatic decline in the abundance of Hexarthra cf. fennica (rare in reference sites) from 93% of all rotifer individuals in new ponds in 2006 to only 32% in 2007. In contrast, species richness of copepods and cladocerans was significantly higher in new ponds in 2007, but there were no changes in Simpson diversity. β-Diversity of cladocerans was also significantly higher in 2007. In 2006, the species richness of cladocera and copepods was significantly lower in new ponds than in reference sites, but by 2007 there were no differences in richness or Simpson diversity. Overall, 7 copepod, 13 cladoceran and 26 rotifer taxa were recorded in new ponds, including 80% of taxa recorded in reference sites. These results indicate that zooplankton communities can be rapidly restored in Mediterranean temporary wetlands, at least when large source populations in the surrounding area reduce dispersal limitation. They also illustrate the importance of comparing different metrics of richness and diversity in studies of zooplankton restoration.     

Variability of the sediment phosphate composition of a temporary pond (Doñana National Park,SW Spain)

This study reports on the spatial and temporal variability of the phosphate composition in the sediment of a temporary pond over a period of 3 years using the EDTA-method for P-fractionation. Sediment samples were collected at three different sites (open-water, littoral and flood plain) to compare the effect of the length of the wet/dry phase on the sediment phosphate composition, with special emphasis on the potential bioavailability of the P-fractions.Fine sediments (<0.1 mm) were rich in organic matter (9–25%) and contained high mean concentrations of Tot-P (182–655 mg kg^–1 d.w.), especially in the flood plain sediment. The sediment P composition was dominated by P-organic fractions at all sites (64–94%). The average C/N ratios were 8.8, 6.0 and 5.9 for sediments of the flood plain, littoral and open-water sites, respectively. The flood plain sediment was significantly poorer in iron-bound P (FeOOHP), but richer in the P-organic fractions extracted by EDTA than the sediment of the open-water site (P<0.01). The percentage of organic matter increased significantly in the sediment of the open-water site at the end of each dry season (P<0.05), while it decreased in the sediment of the flood plain site (P<0.01). In all sediments, the fraction of Fe(OOH)P decreased at the end of each dry season and some of these changes were significant (P<0.05). The decrease in the fraction of Fe(OOH)P was not related to changes in the sediment redox potential. Although the flood plain site was dry longer than the open-water site during the study period, the differences between the sediment composition of both sites were probably due to the effect of plant growth on the dry sediments of the flood plain site rather than to a direct effect of desiccation.     

Formation and growth of a heronry in a managed wetland in Doñana,southwest Spain

Capsule A newly created wetland has been beneficial for the breeding of five heron species and for the settlement and expansion of Glossy Ibises in the region.

Aim To determine whether created wetland would lead to the establishment of a heron and ibis breeding colony, and if so, to determine its effects on the distribution of these species in the wider region.

Methods Number of pairs breeding at the new wetland and in the region were estimated annually from 1996 to 2008. We investigated whether the growth of the new colony was explained by redistribution of breeding pairs within the region.

Results Purple Herons, Squacco Herons and Glossy Ibises colonised the new wetland in 1996. Black‐crowned Night Herons settled in 1998, while Cattle and Little Egrets were the last species to establish in the mixed colony in 2001. The population increase was particularly marked for Glossy Ibises (which had bred in Doñana only sporadically through the 20th century). Our findings suggest that simple redistribution of pairs does not explain the formation and growth of the new colony. Since the first year of establishment numbers of Glossy Ibises and Purple Herons have shown a significant growth trend at both the new colony and in Doñana.

Conclusion The creation and management of a new wetland has been successful for the conservation of heron and egret species and promoted the expansion of the Glossy Ibis population across the western Mediterranean region.     

Acidobacteria in freshwater ponds at Doñana National Park, Spain

The Acidobacteria show a widespread distribution in natural ecosystems. In this study, we analyzed the presence of Acidobacteria in freshwater ponds at Do?ana National Park (southwestern Spain). Nucleic acid sequence analysis, quantitative, real-time RT-PCR, and fluorescence in situ hybridization (FISH) were carried out. Acidobacteria in these aquatic environments were investigated using their 16S and 23S rDNA sequences and acidobacterial specific primer pairs through phylogenetic approaches. The presence of up to five subdivisions of Acidobacteria was detected during this study. The analyzed ponds exhibited distinctive patterns of acidobacterial clades. In order to detect their role in ecosystem functions, metabolically active Acidobacteria were detected based upon rRNA analyses. Quantitative, real-time RT-PCR showed a low percentage of metabolically active Acidobacteria at suboxic zones within the water column covered by surface Fe-rich films. Oxygen-saturated areas showed around 4% of total bacterial RNA belonging to Acidobacteria both in the water column and the sediment surface. The morphology of the most abundant Acidobacteria was revealed by FISH as cocci generally in pairs or chains. Enrichment cultures were also obtained and indicated a putative metabolism based on aerobic and heterotrophic characteristics likely taking advantage of the abundant organic matter present at the investigated sites. These results represent a significant contribution toward understanding the distribution and ecological role of the phylum Acidobacteria in natural ecosystems, specifically at Do?ana National Park freshwater ponds.     

Long-term vegetation changes on the stabilized dunes of Doñana National Park (SW Spain)

Changes in the vegetation of stabilized sand dunes of Doñana National Park in the last four centuries have been analysed, combining ecological and historical sources of information.Quantitative vegetation data have been obtained from 5 inventories of a 15 km long transect, consisting of 34 points where nearby woody plants have been recorded. Inventories were taken in 1636, 1647, 1652, 1682 and 1859. The analysis of these data, combined with climatic and management information on the area, reveals that until 1636 a mediterranean forest dominated byQuercus suber, existed; it was used as a game preserve. From 1636 to 1682 vegetation underwent profound changes with a marked drop in trees and mature scrub. The vegetation was replaced by a pioneer scrub species. This rapid change was brought about by the introduction of cattle and pigs, timber and cork exploitation, and the use of prescribed fire. The climatic crisis that arose circa 1700 led to erosion of the sandy soils, which made the vegetation change irreversibly. Livestock pressure hampered natural forest regeneration until 1737 whenPinus pinea plantations started. From then onwards little change in vegetation is known to have occurred.It is suggested that the main steps of change in Doñana vegetation history may have occurred in many other areas of the Mediterranean Basin.     

Crustacean and rotifer composition of temporary ponds in the Doñana National Park (SW Spain) during floods

The zooplankton of 18 temporary ponds in the Doñana National Park (SW Spain) was studied during floods in February and May 1997. A total of 37 rotifer taxa and 34 crustaceans species were identified (17 cladocerans, 2 ostracans, 4 diaptomids, 7 cyclopodids, 1 harpacticoid, 1 anostracan, 1 notostracan and 1 conchostracan). Zooplankton samples were collected separately from the littoral and the open-water of 12 different ponds. Commonly distributed zooplankton species (frequency of appearance 50%) were not segregated to either the littoral or the open-water according to a chi-square test (P>0.005). The study ponds were divided into seasonal, intermediate and ephemeral ponds according to the length of their hydroperiod. The total numbers of both crustacean and rotifer taxa were highest in the intermediate-hydroperiod ponds (26 and 32 taxa, respectively). The total number of zooplankton taxa collected in both February and May was not significantly correlated to the hydroperiod of the temporary ponds of Doñana during the study period (r=0.165, P=0.526). 21 rotifer taxa and 20 crustacean species were found in the ephemeral ponds; the number of restricted species was also relatively high (3 rotifers and 4 crustaceans). Therefore, the ephemeral ponds held a relatively rich community during floods compared to other temporary ponds of Doñana despite their small size and short wet phase.     

A quantitative hydrogeomorphic approach to the classification of temporary wetlands in the Doñana National Park (SW Spain)

A quantitative hydrogeomorphic approach was applied to an extensive survey of temporary wetlands in the Doñana National Park (SW Spain) in search for quantitative thresholds for wetland classification. Twenty freshwater ponds on the aeolian sand mantle and 46 sites on silty-clay substrate, located in the southern marshland, were surveyed during the heaviest rainy period so far recorded (October 1995–September 1997). On average, temporary ponds showed higher water depth, longer flooding period, lower conductivity (<0.5 mS cm^?1), lower pH (6.7), lower phosphate concentration (0.4 μM) and a more balanced proportion of Ca²⁺/Na⁺ than temporary marshes. During floods, marshland sites exhibited higher water transparency, pH (9.5), alkalinity (3.5 meq l^?1), conductivity (8.2 mS cm^?1), phosphate concentration (0.7 μM), Na⁺ and Ca²⁺ concentrations (97.2 and 3.5 meq l^?1, respectively) than ponds. Study sites were significantly segregated (ANOSIM test: R = 0.88, P < 0.01, n = 92) in relation to water depth and conductivity. A conductivity of 1.6 mS cm^?1 is proposed as a threshold between marshland sites and ponds during floods. Marshland sites were further segregated into two groups (ANOSIM test: R = 0.777, P < 0.01, n = 23) according to the Na⁺/Ca²⁺ ratio (in meq l^?1) at a threshold value of 25. An ordination by PCA showed that five variables grouped 81.4% of the total variance in two axes. The first PCA axis (60.7% of variance) separated temporary wetlands into ponds and marshland sites according to variables related to substrate and hydrology (Na⁺/Ca²⁺ ratio, conductivity, water depth and flooding period). Other variables (e.g., water transparency, alkalinity, pH, submersed macrophyte biomass, phosphate, nitrate and planktonic chlorophyll concentrations) did not produce a significant segregation between marshland and pond sites during floods. Further discrimination within each wetland type was thus not achieved.     

Bovine tuberculosis in a free ranging red fox (Vulpes vulpes) from Doñana National Park (Spain)

During 1997 and 1998, a survey of Iberian carnivores was conducted to study the epidemiology of bovine tuberculosis in the Do?ana National Park and surrounding areas in southwestern Spain. Post-mortem examinations were done on seven red foxes (Vulpes vulpes), two Egyptian mongoose (Herpestes ichneumon), one weasel (Mustela nivalis), two genets (Genetta genetta), one Iberian lynx (Lynx pardinus), one Eurasian badger (Meles meles), and two polecats (Mustela putorius). Lesions suggestive of bovine tuberculosis were not detected but, in culture, Mycobacterium bovis was isolated from the retropharyngeal lymph nodes of one adult male red fox. This is the first report of M. bovis infection in red fox in Spain.     

The vertical structure of mediterranean scrub in Doñana national park (SW spain)

The vertical structure of pioneer and stabilized mediterranean sand vegetation at Doñana National Park has been compared using a non destructive method based on organ frequency analysis. Vertical distribution of organ frequencies and light intensity through the canopy, leaf chlorophyll content, leaf area index, organ biomass, are also discussed forArtemisia campestris subsp.maritima, Scrophularia frutescens, Halimium halimifolium, andCistus salvifolius.     

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 Do?ana 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.     

Antibodies to Mycobacterium bovis in wild carnivores from Doñana National Park (Spain)

We conducted a retrospective serologic survey for antibodies against the MPB70 protein of Mycobacterium bovis in wild carnivores from Do?ana National Park (southwestern Spain). Serum samples from 118 red foxes (Vulpes vulpes), 39 Iberian lynx (Lynx pardinus), 31 Eurasian badgers (Meles meles), five Egyptian mongoose (Herpestes ichneumon), four European genet (Genetta genetta), and one Eurasian otter (Lutra lutra) were analyzed using an indirect competitive enzyme-linked immunoassay. Antibodies against the MPB70 protein of M. bovis were detected in seven badgers, five foxes, and one lynx. The frequency of positive animals was significantly higher in badger (23%) than in lynx (3%) and fox (4%). Antibodies were not detected in other species. Annual antibody frequency peaked at 38% in badgers and 11% for red fox. These species may contribute to persistence of bovine tuberculosis in Do?ana.     

Fire Management in a Natural Protected Area: What Do Key Local Actors Say?

Public policies on fire in forest ecosystems are changing from fire-fighting and suppression to an integrated management approach that incorporates ecological and social considerations. However, policy implementation is usually directed by central governments without considering local actors. We identified key local actors in the Monarch Butterfly Biosphere Reserve, Mexico. This is an important natural protected area of high socio-environmental complexity and the overwintering sites of monarch butterfly that migrate from Canada and United States every year. We applied network and grounded theories to analyze qualitative information derived from semi-structured interviews with 28 key local actors including government employees, local inhabitants, non-governmental organization and academic personnel. We identified actors who play essential roles in local fire management. Aside from fire-fighting, local actors engage in fire prevention, habitat restoration, research, training, planning, coordination, and communication activities that are specific to a spatial, temporal, institutional, and environmental context of fire management. The incorporation of the concepts that local actors associate with fire knowledge, behavior, and regimes would result in better planning for fire management in the short, medium, and long-term.     

Flowering and fruiting phenology in the coastal shrublands of Doñana,south Spain

Flowering and fruiting phenological patterns at the individual-, population-, and community level were studied in a southern Spanish scrub community composed of 30 shrub species. Few individuals of any species produced a high number of flowers. Intrapopulation deviation in the peak time of flowering showed a strong and positive skewness. Relative flowering duration, however, displayed a virtually normal distribution. Generally, species flowering in spring have a short flowering time, while species flowering earlier or later in the year show significantly longer flowering periods. Species were in bloom throughout the year, but there was a major peak during spring and two lesser ones in autumn and early summer. Shallow rooting taxa in typically mediterranean genera displayed a strategy of spring flowering and summer fruiting. Summer and autumn flowering occurred among heath-like shrubs of relatively wet places, and forest-associated, vertebrate-dispersed species which commonly have underground storage organs. Species with ripe fruits presented two peaks, the major one during the summer including the majority of taxa with seeds dispersed by non-vertebrate agents. There was a minor fruiting peak in autumn dominated by taxa that rely on vertebrates for dispersal. The complex seasonal patterns observed are interpreted in relation to environmental conditions and physiological constraints on species living in a highly seasonal climate.     

Seasonal variations of pollution indicators in a wildfowl reserve (Do?ana National Park, Spain)

Pollution indicators in Do?ana National Park show a cyclic seasonal pattern with maximum level in the autumn and minimum in the spring. While faecal streptococci proved to be the best indicator micro-organisms in this type of salty, alkaline environment, total coliforms gave an overestimation of pollution because of the presence of hydroteluric coliforms.     

Utility of proteomics to assess pollutant response of clams from the Doñana bank of Guadalquivir Estuary (SW Spain)

The utility of proteomics to assess pollutant response of Scrobicularia plana clams from three sites of Guadalquivir Estuary at the southern end of the National Park of Do?ana (SW Spain) has been studied. Protein expression profiles were analyzed by 2-DE in soluble fractions of S. plana gills. Nearly 2000 well-resolved spots were detected in silver-stained gels, with focused areas in the 4-6.5 pH range. Different protein expression signatures were found at each site, with the highest number of more intense spots in animals with the highest metal content. Nineteen more intense protein spots were analyzed out by nanospray-ion trap tandem mass spectrometry, de novo sequencing and a bioinformatics search for their possible identification. While sequence tags of 16 more intense protein spots were obtained, including several proteins induced by pollutant exposure of model organisms, only 2 proteins were unambiguously identified: hypoxanthine-guanine phosphoribosyltransferase (HPRT) and glyceraldehyde-3-phosphate dehydrogenase (G3PDH). Both enzymes were significantly higher in animals with the highest metal contents. Thus, we propose these two proteins, HPRT and G3PDH, as novel pollution biomarkers.     

Primary moult of continental Black-tailed Godwits Limosa limosa limosa in the Doñana wetlands,Spain

Capsule: Most Continental Black-tailed Godwits Limosa limosa limosa using the Doñana wetlands during post-breeding migration appear to begin moult before they arrive and suspend moult before they migrate onwards to West Africa.

Aims: We aim to describe the primary moult strategies and patterns in the Continental Black-tailed Godwits using the Doñana wetlands, a major passage and wintering area for waterbirds in southern Spain.

Methods: Individual godwits were captured, marked and their primary moult was scored in Doñana during the non-breeding season (June–March) in 2011 and 2012. Data from resightings of colour-marked godwits and birds equipped with satellite transmitters were used to estimate stopover duration during post-breeding migration (June–September) to determine if godwits move to West Africa before completing their primary moult.

Results: Average primary moult duration was estimated to be 84 days?±?9 se, during 29 June–21 September and did not differ between sexes. Only 2% of individuals were observed with suspended moult. We estimated stopover duration in Doñana to be 13 days?±?2 se before migrating to West Africa.

Conclusions: Most godwits stage for about two weeks in the Doñana wetlands during southward migration, moult their primaries and appear to suspend moult before crossing the Sahara. Others may complete their primary moult in Doñana, or elsewhere in Europe and overwinter in Doñana where increasing numbers of godwits have been detected in recent years. A few individuals may finish the moult in Doñana and migrate to West Africa late in the post-breeding season.     

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 Doñana 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.

Methods

An experiment was conducted featuring the following 4 treatments: 1) intact needles (control), 2) trampled needles, 3) intact needles fertilized with liquid manure, and 4) trampled needles fertilized with liquid manure. Litter decomposition was determined as a function of mass loss over time, using the litter-bag method. Bags were recovered 4, 8, 16, 24 and 36 months after burial in soil, dried and weighed. Needle length, leaf mass per area and C and N concentration were also measured in the buried litter-bags.

Results

Four months after burial, mass loss was greater in the trampled (23–27 %) than non-trampled (14–16 %) treatments. However, from 8 months onwards, decomposition rates in the fertilized treatments were significantly higher than those in the non-fertilized treatments (between 5 % and 15 % less mass loss). Meanwhile, fertilized treatments presented higher N content (2.1 %) than the non-fertilized ones (1.2 %), with a significantly lower C:N ratio also found in the in the fertilized treatment.

Conclusions

Trampling and fertilization during grazing accelerates litter decomposition and thus promotes the incorporation of N into the system. Acceleration of decomposition reduces the accumulation of pine needles on the soil surface, reducing the risk of fire.