David X. Soto

Contaminant accumulation and multi-biomarker responses in field collected zebra mussels (Dreissena polymorpha) and crayfish (Procambarus clarkii), to evaluate toxicological effects of industrial hazardous dumps in the Ebro river (NE Spain).

Large amounts of industrial waste containing high concentrations of mercury, cadmium and organochlorine residues were dumped in a reservoir adjacent to a chlorine-alkali plant in the village of Flix(Catalonia, Spain), situated at the shore of the lower Ebro river. Effects of these contaminants to aquatic river invertebrates were assessed by integrating analyses of metals and organochlorine residues in field collected zebra mussels and crayfish with a wide range of biomarkers. Biological responses included levels of metallothioneins, activities of ethoxyresorufin-O-deethylase, oxidative stress biomarkers (glutathione content, enzymatic activities of superoxide dismutase, catalase, glutathione s-transferase, glutathione peroxidise and glutathione reductase), levels of lipid peroxidation and of DNA strand breaks. The results obtained evidenced similar response patterns in mussels and crayfish with increasing toxic stress levels from upper parts of the river towards the meander located immediately downstream from the most polluted site, close to the waste dumps. The aforementioned stress levels could be related with concentrations of mercury, cadmium, hexachlorobenzene, polychlorobiphenyls and dichlorodiphenyltrichloroethanes from 4- to 195-fold greater than local background levels. The response of biomarkers to these pollutant concentrations differences was reflected in high activities and levels of antioxidant enzymes, metallothioneins, lipid peroxidation and DNA strand breaks and decreased levels of glutathione.

Assessment of mercury and methylmercury pollution with zebra mussel (Dreissena polymorpha) in the Ebro River (NE Spain) impacted by industrial hazardous dumps

Large amounts of industrial waste containing high concentrations of mercury (up to 436 microg/g) are dumped in a reservoir adjacent to a chlor-alkali plant in Flix (Catalonia, Spain), on the lower Ebro River. In order to assess the spatial redistribution of mercury from the point source and its bioavailability to the aquatic food web, zebra mussels (Dreissena polymorpha) were collected at several sites. The highest total Hg (THg) and methylmercury (MeHg) concentrations ever reported for zebra mussels were found (THg: 0.02 to 0.81 microg/g ww; MeHg: 0.22 to 0.60 microg/g ww). At the most polluted site, close to the waste dump, the mean values were 20 times greater than the local background level. Concentrations decreased with increasing mussel size at all sites. The MeHg/THg ratio was ca. 60% (range: 50-80%). A comparison of similar size classes clearly indicated the hot spots of Hg bioavailability to the aquatic food web and downstream transport.

Stable hydrogen and oxygen isotopes in aquatic food webs are tracers of diet and provenance

Summary 1. The stable hydrogen and oxygen isotope (d 2 H and d 18 O) composition of animal tissues are well-established tracers for terrestrial migration ecology and wildlife forensics. However, the behaviour of these isotopes in aquatic ecosystems and their potential as tracers of diet and provenance are complicated because of inputs from ambient H2O and diet. 2. We conducted controlled aquaria-based experiments to quantify the mechanisms that drive the H and O isotopic flow within and among aquatic species. The isotopic composition of water and diet of two aquatic species (Chironomus tentans and Poecilia reticulata), representing two trophic levels, was varied in six isothermal treatments. Both species were raised from juvenile to adult to ensure that tissues were in isotopic equilibrium with their dietary and environmental conditions (ambient water, food, dissolved oxygen). We measured water, dissolved O2, diet, tissue protein and lipids for d 2 Ho rd 18 O. 3. The flows of H and O isotopes for tissue formation in aquatic organisms were parameterized using a steady-state multi-pool mass-balance model. The ambient H2O contribution to tissue protein H and O isotopes in both species was significant (30–50% for 2 H and >80% for 18 O). An apparent trophic effect for d 2 H and isotopic discrimination between water and protein for d 18 O( c .1 5&) were identified. 4. Our isotopic data and model revealed potential applications and cautions in using d 2 H and d 18 O measurements for ecological studies in aquatic food webs. Tissue d 2 H values may be a complementary trophic tracer in aquatic food webs, but only when the main controlling mechanisms are properly accounted for (i.e., H isotopic exchange with water during protein synthesis and influence of metabolic water). Measurements of d 18 O, on the other hand, reflect that of water and so can be used for predicting isotopic assignment to origin of aquatic organisms as there is no complicating trophic effect, but more d 18 O field data and improved analytical precision may be required to better establish the strengths to ecological applications.

Differential accumulation of mercury and other trace metals in the food web components of a reservoir impacted by a chlor-alkali plant (Flix, Ebro River, Spain): Implications for biomonitoring

Comparative studies of biomonitors of trace metal contamination are relatively scarce. We took advantage of a point source pollution in a reservoir (Flix, Spain) to compare trace metal (Hg, Pb, Cd, Se, As, Zn, Cu, Cr) bioaccumulation patterns among 16 food web components. Our results indicate that most organisms are suitable for Hg biomonitoring, whereas other metals are better monitored by only some of them. Biofilms and zebra mussel were the organisms with larger and more diverse biomonitoring capacity. However, we show that using groups of biomonitors increase the scope and strengths of the conclusions and specific goals can be better addressed. We conclude providing an overview of the strengths and weaknesses of the main organisms considered for biomonitoring trace metals in rivers and reservoirs.

Isotopic Evidence That Dragonflies (Pantala flavescens) Migrating through the Maldives Come from the Northern Indian Subcontinent

Large numbers of the Globe Skimmer dragonfly (Pantala flavescens) appear in the Maldives every October–December. Since they cannot breed on these largely waterless islands, it has recently been suggested that they are “falling out” during a trans-oceanic flight from India to East Africa. In addition, it has been suggested that this trans-oceanic crossing is just one leg of a multi-generational migratory circuit covering about 14,000–18,000 km. The dragonflies are presumed to accomplish this remarkable feat by riding high-altitude winds associated with the Inter-tropical Convergence Zone (ITCZ). While there is considerable evidence for this migratory circuit, much of that evidence is circumstantial. Recent developments in the application of stable isotope analyses to track migratory dragonflies include the establishment of direct associations between dragonfly wing chitin δ 2H values with those derived from long-term δ 2H precipitation isoscapes. We applied this approach by measuring wing chitin δ 2H values in 49 individual Pantala flavescens from the November–December migration through the Maldives. Using a previously established spatial calibration algorithm for dragonflies, the mean wing δ 2H value of −117±16 ‰ corresponded to a predicted mean natal ambient water source of −81 ‰, which resulted in a probabilistic origin of northern India, and possibly further north and east. This strongly suggests that the migratory circuit of this species in this region is longer than previously suspected, and could possibly involve a remarkable trans-Himalayan high-altitude traverse.

Expanding the Isotopic Toolbox: Applications of Hydrogen and Oxygen Stable Isotope Ratios to Food Web Studies

The measurement of stable carbon (δ13C) and nitrogen (δ15N) isotopes in tissues of organisms has formed the foundation of isotopic food web reconstructions, as these values directly reflect assimilated diet. In contrast, stable hydrogen (δ2H) and oxygen (δ18O) isotope measurements have typically been reserved for studies of migratory origin and paleoclimate reconstruction based on systematic relationships between organismal tissue and local environmental water. Recently, innovative applications using δ2H and, to a lesser extent, δ18O values have demonstrated potential for these elements to provide novel insights in modern food web studies. We explore the advantages and challenges associated with three applications of δ2H and δ18O values in food web studies. First, large δ2H differences between aquatic and terrestrial ecosystem end members can permit the quantification of energy inputs and nutrient fluxes between these two sources, with potential applications for determining allochthonous vs. autochthonous nutrient sources in freshwater systems and relative aquatic habitat utilization by terrestrial organisms. Next, some studies have identified a relationship between δ2H values and trophic position, which suggests that this marker may serve as a trophic indicator, in addition to the more commonly used δ15N values. Finally, coupled measurements of δ2H and δ18O values are increasing as a result of reduced analytical challenges to measure both simultaneously and may provide additional ecological information over single element measurements. In some organisms, the isotopic ratios of these two elements are tightly coupled, whereas the isotopic disequilibrium in other organisms may offer insight into the diet and physiology of individuals. Although a coherent framework for interpreting δ2H and δ18O data in the context of food web studies is emerging, many fundamental uncertainties remain. We highlight directions for targeted research that will increase our understanding of how these markers move through food webs and reflect ecological processes.

Long-distance autumn migration across the Sahara by painted lady butterflies: exploiting resource pulses in the tropical savannah

The painted lady, Vanessa cardui, is a migratory butterfly that performs an annual multi-generational migration between Europe and North Africa. Its seasonal appearance south of the Sahara in autumn is well known and has led to the suggestion that it results from extremely long migratory flights by European butterflies to seasonally exploit the Sahel and the tropical savannah. However, this possibility has remained unproven. Here, we analyse the isotopic composition of butterflies from seven European and seven African countries to provide new support for this hypothesis. Each individual was assigned a geographical natal origin, based on its wing stable hydrogen isotope (δ2Hw) value and a predicted δ2Hw basemap for Europe and northern Africa. Natal assignments of autumn migrants collected south of the Sahara confirmed long-distance movements (of 4000 km or more) starting in Europe. Samples from Maghreb revealed a mixed origin of migrants, with most individuals with a European origin, but others having originated in the Sahel. Therefore, autumn movements are not only directed to northwestern Africa, but also include southward and northward flights across the Sahara. Through this remarkable behaviour, the productive but highly seasonal region south of the Sahara is incorporated into the migratory circuit of V. cardui.

Small Tails Tell Tall Tales – Intra-Individual Variation in the Stable Isotope Values of Fish Fin

Background Fish fin is a widely used, non-lethal sample material in studies using stable isotopes to assess the ecology of fishes. However, fish fin is composed of two distinct tissues (ray and membrane) which may have different stable isotope values and are not homogeneously distributed within a fin. As such, estimates of the stable isotope values of a fish may vary according to the section of fin sampled. Methods To assess the magnitude of this variation, we analysed carbon (δ 13C), nitrogen (δ 15N), hydrogen (δ 2H) and oxygen (δ 18O) stable isotopes of caudal fin from juvenile, riverine stages of Atlantic salmon (Salmo salar) and brown trout (Salmo trutta). Individual fins were sub-sectioned into tip, mid and base, of which a further subset were divided into ray and membrane. Findings Isotope variation between fin sections, evident in all four elements, was primarily related to differences between ray and membrane. Base sections were13C depleted relative to tip (~ 1 ‰) with equivalent variation evident between ray and membrane. A similar trend was evident in δ 2H, though the degree of variation was far greater (~ 10 ‰). Base and ray sections were 18O enriched (~ 2 ‰) relative to tip and membrane, respectively. Ray and membrane sections displayed longitudinal variation in 15N mirroring that of composite fin (~ 1 ‰), indicating that variation in15N values was likely related to ontogenetic variation. Conclusions To account for the effects of intra-fin variability in stable isotope analyses we suggest that researchers sampling fish fin, in increasing priority, 1) also analyse muscle (or liver) tissue from a subsample of fish to calibrate their data, or 2) standardize sampling by selecting tissue only from the extreme tip of a fin, or 3) homogenize fins prior to analysis.

The influence of metabolic effects on stable hydrogen isotopes in tissues of aquatic organisms

A steady-state mass-balance model describing controls on the stable hydrogen isotopic ratios (δ2H) of tissues in fish was previously developed but physiological effects related to fish size and growth had not been tested. Here, we assessed the influence of size (or growth rate) on tissue δ2H composition of a fish species (Poecilia reticulata) and the incorporation of metabolic products derived from dietary lipids (water, NADH). Sampled tissues were obtained from individuals that grew at different rates while raised on an isotopically homogeneous commercial diet (lipid-free fraction, δ2H=−95±2 ‰; and dietary lipids,−198±11 ‰) under different controlled water hydrogen isotopic composition (δ2H=−128±3 ‰;+17±5 ‰; and+202±5 ‰). Our findings suggested that fish growth rate was correlated positively with the degree of incorporation of metabolic products from dietary lipids that, in turn, influenced both fish tissue protein and lipid δ2H values. We conclude that δ2H measurements of lipids (and, subsequently, of body water) in fish could become a physiological tool that provides insights into fish growth rates.

Combining Denitrifying Bacteria and Laser Spectroscopy for Isotopic Analyses (δ 15 N, δ 18 O) of Dissolved Nitrate

We present a novel approach for nitrogen (δ(15)N) and oxygen (δ(18)O) isotopic analysis of nitrate in water based on the isotopic analysis of N2O produced from the conversion of NO3(-) by cultured denitrifying bacteria and off-axis integrated cavity output spectroscopy (OA-ICOS). The headspace N2O was manually injected into an OA-ICOS isotopic N2O laser analyzer through a syringe septum port. Sample analysis time was ∼300 s. The use of OA-ICOS technology yields accurate and precise δ(15)N and δ(18)O results for dissolved nitrate samples when nonlinearity issues are considered. This new isotope analytical technique thus improves the isotopic analysis of nitrates by (i) providing accurate measurements of δ(15)N and δ(18)O without preconcentration, (ii) eliminating interferences by other gas substances (i.e., H2O and CO2), and (iii) reducing extensive maintenance and costs of isotope ratio mass spectrometers (IRMS). This approach will greatly streamline the identification and quantification of nitrate sources in aquatic systems.