Manuel E. Ortiz-Santaliestra

Influence of developmental stage on sensitivity to ammonium nitrate of aquatic stages of amphibians

In static renewal experiments, we studied how developmental stage influences the effect of ammonium nitrate on embryonic and larval stages of anuran amphibians. The observed lethal effects caused by ammonium nitrate increased with both concentration and duration of exposure. Significant differences were observed in sensitivity to ammonium nitrate as a function of developmental stage in Discoglossus galganoi, Pelobates cultripes, and Bufo calamita. In D. galganoi and P. cultripes, younger individuals displayed greater acute effects from the chemical fertilizer compared with older individuals. For example, 100% of P. cultripes hatchlings died after 4 d of exposure to a nominal concentration of 225.8 mg N-NO3NH4/L, whereas less than 40% of individuals from older larval stages died when exposed to this concentration. A delay of 4 d in the beginning of the exposure to the chemical was enough to cause significant differences in sensitivity. Bufo calamita showed a higher sensitivity in later larval stages after 12 d of exposure. Hyla meridionalis and B. calamita were less sensitive than the other two species. Peak ammonium nitrate concentrations usually occur when amphibians are breeding and, thus, when the most sensitive aquatic stage is in the water. The developmental stage of the test animals should be considered when evaluating the risk of ammonium nitrate to amphibians.

Imidacloprid-treated seed ingestion has lethal effect on adult partridges and reduces both breeding investment and offspring immunity☆

The ingestion of imidacloprid treated seeds by farmland birds may result in exposure to toxic amounts of this insecticide. Here we report on the effects that the exposure to the recommended application rate and to 20% of that rate may produce on birds feeding on treated seeds. Experimental exposure to imidacloprid treated seeds was performed on red-legged partridges (Alectoris rufa) (n=15 pairs per treatment group: control, 20% or 100% of the recommended application rate) during two periods that corresponded to the autumn (duration of exposure: 25 days) and late winter (10 days) cereal sowing times in Spanish farmlands. We studied effects on the survival, body condition, oxidative stress biomarkers, plasma biochemistry, carotenoid-based coloration, T-cell mediated immune response and reproduction of exposed adult partridges, and on the survival and T-cell immune response of their chicks. The high dose (recommended application rate) killed all partridges, with mortality occurring faster in females than in males. The low dose (20% the recommended application rate) had no effect on mortality, but reduced levels of plasma biochemistry parameters (glucose, magnesium and lactate dehydrogenase), increased blood superoxide dismutase activity, produced changes in carotenoid-based integument coloration, reduced the clutch size, delayed the first egg lay date, increased egg yolk vitamins and carotenoids and depressed T-cell immune response of chicks. Moreover, the analysis of the livers of dead partridges revealed an accumulation of imidacloprid during exposure time. Despite the moratorium on the use of neonicotinoids in the European Union, birds may still be at high risk of poisoning by these pesticides through direct sources of exposure to coated seeds in autumn and winter.

The pond network: can structural connectivity reflect on (amphibian) biodiversity patterns?

Landscape connectivity is a very recurrent theme in landscape ecology as it is considered pivotal for the long term conservation of any organism’s populations. Nevertheless, this complex concept is still surrounded by uncertainty and confusion, largely due to the separation between structural and functional connectivity. Amphibians are the most threatened vertebrates around the globe, in Europe mostly due to habitat alteration, and to their particular life cycle. Pond breeding amphibians are considered to be organised in metapopulations, enhancing the importance of landscape connectivity in this group of animals. We sampled the amphibian species present in two pond groups in Central Western Spain. We applied the graph theory framework to these two pond networks in order to determine the importance of each pond for the entire network connectivity. We related the pond importance for connectivity with the species richness present in each pond. We tested if connectivity (partially) determined the presence of the amphibian species sampled using logistic regression. The results show that the structural connectivity of the pond network impacts on the amphibian species richness pattern and that the importance of the pond for the connectivity of the network is an important factor for the presence of some species. Our results, hence, attest the importance of (structural) landscape connectivity determining the pattern of amphibian (functional) colonization in discrete ponds.

Ambient ultraviolet B radiation and prevalence of infection by Batrachochytrium dendrobatidis in two amphibian species

Chytridiomycosis, the emerging disease caused by the chytrid fungus Batrachochytrium dendrobatidis (Bd) is responsible for declines and extirpations of amphibian populations worldwide. Environmental covariates modify the host-Bd interaction and thus affect the ongoing spread of the pathogen. One such covariate may be the intensity of ultraviolet B (UV-B) radiation. In a field experiment conducted in Laguna Grande de Peñalara (central Spain), a mountainous region where the presence of Bd has been documented since 1997, we analyzed the potential effect of environmental UV-B (daily maximum 2.5-3.9 W/m(2) ) on the susceptibility of larvae of the common toad (Bufo bufo) to Bd. The proportion of infected individuals increased as tadpoles developed. The prevalence of Bd was significantly lower in tadpoles exposed to environmental UV-B intensities (2.94%) than in tadpoles not exposed to the radiation (9.72%). This finding mirrors that seen for a second amphibian species, the European midwife toad (Alytes obstetricans), for which conditional prevalence (i.e., prevalence of infection conditioned on the probability of a site being infected) across the Iberian Peninsula was inversely correlated with the intensity of UV-B.

Reducing Pb poisoning in birds and Pb exposure in game meat consumers: The dual benefit of effective Pb shot regulation

The use of lead (Pb) ammunition in the form of shot pellets has been identified as a Pb exposure risk in wildlife and their human consumers. We explore the hypothesis that Pb shot ban enforcement reduces the risk of avian Pb poisoning as well as Pb exposure in game meat consumers. We assessed compliance with a partial ban on Pb shot commencing in 2003 by examination of 937 waterbirds harvested by hunters between 2007 and 2012 in the Ebro delta (Spain). Prevalence of Pb shot ingestion was determined, as were Pb concentrations in liver and muscle tissue to evaluate the potential for Pb exposure in game meat consumers. Hunted birds with only embedded Pb shot (no steel) declined from 26.9% in 2007-08 to <2% over the following three hunting seasons after ban reinforcement. Pb shot ingestion in mallards decreased from a pre-ban value of 30.2% to 15.5% in the post-ban period. Liver Pb levels were predominantly defined by the presence of ingested shot, whereas muscle levels were defined by the presence of both ingested and embedded shot. Only 2.5% of mallard muscle tissue had Pb levels above European Union regulations for meat (0.1μg/g wet weight) in the 2008-09 season, when Pb shot ingestion prevalence was also at a minimum (5.1%). Effective restrictions in Pb ammunition use have a dual benefit since this reduces Pb exposure for game meat consumers due to embedded ammunition as well as reducing Pb poisoning in waterbirds.

Adaptation to osmotic stress provides protection against ammonium nitrate in Pelophylax perezi embryos.

The negative effects of pollution on amphibians are especially high when animals are additionally stressed by other environmental factors such as water salinity. However, the stress provoked by salinity may vary among populations because of adaptation processes. We tested the combined effect of a common fertilizer, ammonium nitrate (0-90.3 mg N-NO3NH4/L), and water salinity (0-2 per thousand) on embryos of two Pelophylax perezi populations from ponds with different salinity concentrations. Embryos exposed to the fertilizer were up to 17% smaller than controls. Survival rates of embryos exposed to a single stressor were always below 10%. The exposure to both stressors concurrently increased mortality rate (>95%) of embryos from freshwater. Since the fertilizer was lethal only when individuals were stressed by the salinity, it did not cause lethal effects on embryos naturally adapted to saline environments. Our results underscore the importance of testing multiple stressors when analyzing amphibian sensitivity to environmental pollution.

Altered immune response in mallard ducklings exposed to lead through maternal transfer in the wild.

Lead (Pb) poisoning has caused significant mortality in waterfowl populations worldwide. In spite of having been banned since 2003, prevalence of Pb shot ingestion in mallards (Anas platyrhynchos) from the Ebro delta was still 15.5% in 2011-12. We collected mallard eggs from this area to study the effects of maternally transferred Pb on eggshell properties and on immune response and oxidative balance of ducklings. Eggshell Pb levels were positively correlated with Pb levels in the blood of ducklings. Ducklings with blood Pb levels above 180 ng mL(-1) showed reduced body mass and died during the first week post hatching. Blood Pb levels positively correlated with humoral immune response, endogenous antioxidants and oxidative stress biomarkers, and negatively correlated with cellular immune response. Pb shot ingestion in birds can result in maternal transfer to the offspring that can affect their developing immune system and reduce their survival in early life stages.

Levels of organochlorine pesticides and polychlorinated biphenyls in the critically endangered Iberian lynx and other sympatric carnivores in Spain.

Accumulation of organochlorine compounds is well studied in aquatic food chains whereas little information is available from terrestrial food chains. This study presents data of organochlorine levels in tissue and plasma samples of 15 critically endangered Iberian lynx (Lynx pardinus) and other 55 wild carnivores belonging to five species from three natural areas of Spain (Doñana National Park, Sierra Morena and Lozoya River) and explores their relationship with species diet. The Iberian lynx, with a diet based on the consumption of rabbit, had lower PCB levels (geometric means, plasma: <0.01 ng mL(-1), liver: 0.4ngg(-1) wet weight, fat: 87 ng g(-1)lipid weight) than other carnivores with more anthropic and opportunistic foraging behavior, such as the red fox (Vulpes vulpes; plasma: 1.11 ng mL(-1), liver: 459 ng g(-1), fat: 1984 ng g(-1)), or with diets including reptiles at higher proportion, such as the Egyptian mongoose (Herpestes ichneumon; plasma: 7.15 ng mL(-1), liver: 216 ng g(-1), fat: 540 ng g(-1)), or the common genet (Genetta genetta; liver: 466 ng g(-1), fat: 3854 ng g(-1)). Chlorinated pesticides showed interspecific variations similar to PCBs. Organochlorine levels have declined since the 80s in carnivores from Doñana National Park, but PCB levels are still of concern in Eurasian otters (Lutra lutra; liver: 3873-5426 ng g(-1)) from the industrialized region of Madrid.

Plasma levels of pollutants are much higher in loggerhead turtle populations from the Adriatic Sea than in those from open waters (Eastern Atlantic Ocean)

In this paper we determined the levels of 63 environmental contaminants, including organic (PCBs, organochlorine pesticides, and PAHs) and inorganic (As, Cd, Cu, Pb, Hg and Zn) compounds in the blood of loggerhead turtles (Caretta caretta) from two comparable populations that inhabit distinct geographic areas: the Adriatic Sea (Mediterranean basin) and the Canary Islands (Eastern Atlantic Ocean). All animals were sampled at the end of a period of rehabilitation in centers of wildlife recovery, before being released back into the wild, so they can be considered to be in good health condition. The dual purpose of this paper is to provide reliable data on the current levels of contamination of this species in these geographic areas, and secondly to compare the results of both populations, as it has been reported that marine biota inhabiting the Mediterranean basin is exposed to much higher pollution levels than that which inhabit in other areas of the planet. According to our results it is found that current levels of contamination by organic compounds are considerably higher in Adriatic turtles than in the Atlantic ones (∑PCBs, 28.45 vs. 1.12ng/ml; ∑OCPs, 1.63 vs. 0.19ng/ml; ∑PAHs, 13.39 vs. 4.91ng/ml; p<0.001 in all cases). This is the first time that levels of PAHs are reported in the Adriatic loggerheads. With respect to inorganic contaminants, although the differences were not as great, the Adriatic turtles appear to have higher levels of some of the most toxic elements such as mercury (5.74 vs. 7.59μg/ml, p<0.01). The results of this study confirm that the concentrations are larger in turtles from the Mediterranean, probably related to the high degree of anthropogenic pressure in this basin, and thus they are more likely to suffer adverse effects related to contaminants.

Sublethal Pb Exposure Produces Season-Dependent Effects on Immune Response, Oxidative Balance and Investment in Carotenoid-based Coloration in Red-Legged Partridges

Ingestion of lead (Pb) shot pellets constitutes the main cause of Pb poisoning in avifauna. We studied the effects of sublethal Pb exposure on immunity, carotenoid-based coloration, oxidative stress and trade-offs among these types of responses during spring and autumn in red-legged partridges (Alectoris rufa). We evaluated constitutive immunity testing lysozyme and natural antibody levels, and blood bactericidal and phagocytic activities. We studied induced immunity by testing PHA and humoral responses. We analyzed fecal parasite and bacterial abundance and oxidative stress biomarkers. Pb exposure in spring reduced natural antibody levels, whereas in autumn, it reduced lysozyme levels and increased phagocytic activity. Pb exposure increased PHA response in both seasons, and decreased T-independent humoral response in autumn. Pb exposure also increased noncoliform and decreased coliform Gram-negative gut bacteria. In spring, Pb exposure decreased antioxidant levels and increased coloration in males, whereas in autumn, it increased retinol levels but reduced coloration in both genders. Our results suggest that in spring, Pb-exposed females used antioxidants to cope with oxidative stress at the expense of coloration, whereas Pb-exposed males increased coloration, which may reflect an increased breeding investment. In autumn, both genders prioritized oxidative balance maintenance at the expense of coloration.