Effects of Copper Ions on Non-target Species: A Case Study Using the Grazer Theodoxus fluviatilis (Gastropoda: Neritidae).

The various uses of copper or copper compounds in industrial, biocidal or pesticidal products lead to inputs of the metal into aquatic environments. To assess effects of copper ions on non-target organisms, the freshwater snail Theodoxus fluviatilis was used as test organism for a three-week laboratory experiment. Snails were exposed to four copper concentrations ranging from 4 to 39 µg Cu2+/L, and besides mortality, several sublethal parameters were evaluated. Concerning survival, an aqueous copper concentration of 6 µg/L was determined as NOEC, and 16 (± 0.1) µg/L as LC50. Negative sublethal effects on reproduction, activity and pathological modifications in the snails were detected in the treatment with an aqueous copper concentration of 15 µg/L. Our results using T. fluviatilis as a mere grazer exclusively feeding on biofilms contribute to findings that field relevant copper concentrations have a significant effect on non-target organisms in aquatic environments.

Compound-specific δ(13)C analyses reveal sterol metabolic constraints in an aquatic invertebrate.

RATIONALE: Dietary sterol deficiencies can have severe life history consequences for consumers. Compound-specific stable isotope analysis (CSIA) was applied to the exploration of the sterol metabolic constraints and bioconversion capacities of the amphipod Gammarus roeselii. Evaluating structural sterol requirements has great potential to improve our understanding of the ecological relevance of sterols as limiting nutrients. METHODS: Juvenile G. roeselii were reared on food mixtures consisting of different ratios of the two algae Scenedesmus obliquus (cultivated with (13)C-labeled NaHCO3) and Nannochloropsis limnetica (unlabeled), which have been shown previously to differ in food quality. We measured the sterol content and composition using a gas chromatograph equipped with a flame ionization detector and the δ(13)C values of sterols using compound-specific isotope ratio mass spectrometry to examine potential sterol-mediated nutritional constraints of G. roeselii. RESULTS: In the food mixtures, δ(13)C values of cholesterol, synthesized by N. limnetica, were -25‰ and those of the Δ(7)-phytosterols, chondrillasterol and fungisterol, synthesized by S. obliquus, were 7 and 18‰, respectively. Although the cholesterol concentrations in G. roeselii decreased with increasing proportion of dietary S. obliquus, the δ(13)C values remained constant at -25‰. Lathosterol, which appeared in G. roeselii at high dietary proportions of S. obliquus, had a δ(13)C value of 35‰. CONCLUSIONS: We provide evidence that the the Δ(7)-phytosterols present in S. obliquus cannot be metabolized to cholesterol in G. roeselii, resulting in the accumulation of lathosterol in the animals and potentially in sterol-limited growth. These findings emphasize the advantage of CSIA in revealing the physiological mechanisms associated with nutritional constraints.

Better define beta-optimizing MDD (minimum detectable difference) when interpreting treatment-related effects of pesticides in semi-field and field studies.

The minimum detectable difference (MDD) is a measure of the difference between the means of a treatment and the control that must exist to detect a statistically significant effect. It is a measure at a defined level of probability and a given variability of the data. It provides an indication for the robustness of statistically derived effect thresholds such as the lowest observed effect concentration (LOEC) and the no observed effect concentration (NOEC) when interpreting treatment-related effects on a population exposed to chemicals in semi-field studies (e.g., micro-/mesocosm studies) or field studies. MDD has been proposed in the guidance on tiered risk assessment for plant protection products in edge of field surface waters (EFSA Journal 11(7):3290, 2013), in order to better estimate the robustness of endpoints from such studies for taking regulatory decisions. However, the MDD calculation method as suggested in this framework does not clearly specify the power which is represented by the beta-value (i.e., the level of probability of type II error). This has implications for the interpretation of experimental results, i.e., the derivation of robust effect values and their use in risk assessment of PPPs. In this paper, different methods of MDD calculations are investigated, with an emphasis on their pre-defined levels of type II error-probability. Furthermore, a modification is suggested for an optimal use of the MDD, which ensures a high degree of certainty for decision-makers.

Laboratory Protocol for Genetic Gut Content Analyses of Aquatic Macroinvertebrates Using Group-specific rDNA Primers.

Analyzing food webs is essential for a better understanding of ecosystems. For example, food web interactions can undergo severe changes caused by the invasion of non-indigenous species. However, an exact identification of field predator-prey interactions is difficult in many cases. These analyses are often based on a visual evaluation of gut content or the analysis of stable isotope ratios (δ15N and δ13C). Such methods require comprehensive knowledge about, respectively, morphologic diversity or isotopic signature from individual prey organisms, leading to obstacles in the exact identification of prey organisms. Visual gut content analyses especially underestimate soft bodied prey organisms, because maceration, ingestion and digestion of prey organisms make identification of specific species difficult. Hence, polymerase chain reaction (PCR) based strategies, for example the use of group-specific primer sets, provide a powerful tool for the investigation of food web interactions. Here, we describe detailed protocols to investigate the gut contents of macroinvertebrate consumers from the field using group-specific primer sets for nuclear ribosomal deoxyribonucleic acid (rDNA). DNA can be extracted either from whole specimens (in the case of small taxa) or out of gut contents of specimens collected in the field. Presence and functional efficiency of the DNA templates need to be confirmed directly from the tested individual using universal primer sets targeting the respective subunit of DNA. We also demonstrate that consumed prey can be determined further down to species level via PCR with unmodified group-specific primers combined with subsequent single strand conformation polymorphism (SSCP) analyses using polyacrylamide gels. Furthermore, we show that the use of different fluorescent dyes as labels enables parallel screening for DNA fragments of different prey groups from multiple gut content samples via automated fragment analysis.

Using stable isotope analysis in stream mesocosms to study potential effects of environmental chemicals on aquatic-terrestrial subsidies.

While recent research has provided evidence that the emergence of merolimnic insects (species with an aquatic larval stage) provides a considerable energy subsidy to riparian food webs, it has also shown that merolimnic insects may serve as a vector for contaminants. Therefore, riparian food webs may be at risk from either an aquatic-terrestrial transfer of contaminants or from the contaminant-driven reductions of emerging merolimnic insects. The objective of the present study was to develop an integrated stream mesocosms test design capable of identifying these inter-ecosystem boundary effects and to provide a comprehensive approach as a basis for ecotoxicological testing. We chose the widely distributed web-building spider Tetragnatha extensa as a representative species for riparian predators. Trophic aspects of riparian food webs were investigated by stable isotope analysis of carbon (δ(13)C) and nitrogen (δ(15)N). Utilization of stable isotope ratios provided detailed information on the riparian food web structure and the dietary composition of T. extensa. Merolimnic invertebrates (mainly Cloeon spp. and Chironomidae) were found to contribute up to 71 % of T. extensa's diet, demonstrating their importance in riparian food webs in ecotoxicological mesocosm testing. This study provides a conceptual and methodological basis for assessing aquatic insect emergence-related pollutant transfer or effect translation from aquatic to adjacent terrestrial systems.

Review on environmental alterations propagating from aquatic to terrestrial ecosystems.

Terrestrial inputs into freshwater ecosystems are a classical field of environmental science. Resource fluxes (subsidy) from aquatic to terrestrial systems have been less studied, although they are of high ecological relevance particularly for the receiving ecosystem. These fluxes may, however, be impacted by anthropogenically driven alterations modifying structure and functioning of aquatic ecosystems. In this context, we reviewed the peer-reviewed literature for studies addressing the subsidy of terrestrial by aquatic ecosystems with special emphasis on the role that anthropogenic alterations play in this water-land coupling. Our analysis revealed a continuously increasing interest in the coupling of aquatic to terrestrial ecosystems between 1990 and 2014 (total: 661 studies), while the research domains focusing on abiotic (502 studies) and biotic (159 studies) processes are strongly separated. Approximately 35% (abiotic) and 25% (biotic) of the studies focused on the propagation of anthropogenic alterations from the aquatic to the terrestrial system. Among these studies, hydromorphological and hydrological alterations were predominantly assessed, whereas water pollution and invasive species were less frequently investigated. Less than 5% of these studies considered indirect effects in the terrestrial system e.g. via food web responses, as a result of anthropogenic alterations in aquatic ecosystems. Nonetheless, these very few publications indicate far-reaching consequences in the receiving terrestrial ecosystem. For example, bottom-up mediated responses via soil quality can cascade over plant communities up to the level of herbivorous arthropods, while top-down mediated responses via predatory spiders can cascade down to herbivorous arthropods and even plants. Overall, the current state of knowledge calls for an integrated assessment on how these interactions within terrestrial ecosystems are affected by propagation of aquatic ecosystem alterations. To fill these gaps, we propose a scientific framework, which considers abiotic and biotic aspects based on an interdisciplinary approach.

Do differences in sensitivity between native and invasive amphipods explain their coexistence in Lake Constance? A case study with lambda-cyhalothrin.

Invasive species are considered as one of the major threats for biodiversity worldwide. The Ponto-Caspian species Dikerogammarus villosus, for instance, spread throughout continental Europe and was recorded for the first time also within Lake Constance in 2003. Although D. villosus is a highly competitive species it was not capable of replacing the native Gammarus roeselii completely in this ecosystem, especially in the riparian zones of the highly agriculturally used island "Reichenau". As differences in pesticide sensitivity between both amphipod species may explain their distribution, the present study assessed the implication of the highly toxic pyrethroid lambda-cyhalothrin, which is authorized for application in the Lake Constance region, assuming the invasive species being more sensitive than the native one. However, both the feeding activity bioassays, which measured the leaf consumption over 7d (n=20), as well as the predation bioassay, which measured the predation rate upon Baetis nymphs in concert with the feeding activity on leaf material over 96 h (n=13), revealed an up to 5-fold higher tolerance of D. villosus towards lambda-cyhalothrin. These results suggest the investigated insecticide not being the trigger for the observed distribution pattern of both amphipod species. Hence, other factors like the diversity of habitat structures or the levels of ammonia may have facilitated the coexistence. Nevertheless, the present study uncovered a high leaf-shredding efficacy of the invasive species D. villosus suggesting that its role in the leaf decomposition process may have been underestimated in the past.

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 August 2009-30 September 2009.

This article documents the addition of 238 microsatellite marker loci and 72 pairs of Single Nucleotide Polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Adelges tsugae, Artemisia tridentata, Astroides calycularis, Azorella selago, Botryllus schlosseri, Botrylloides violaceus, Cardiocrinum cordatum var. glehnii, Campylopterus curvipennis, Colocasia esculenta, Cynomys ludovicianus, Cynomys leucurus, Cynomys gunnisoni, Epinephelus coioides, Eunicella singularis, Gammarus pulex, Homoeosoma nebulella, Hyla squirella, Lateolabrax japonicus, Mastomys erythroleucus, Pararge aegeria, Pardosa sierra, Phoenicopterus ruber ruber and Silene latifolia. These loci were cross-tested on the following species: Adelges abietis, Adelges cooleyi, Adelges piceae, Pineus pini, Pineus strobi, Tubastrea micrantha, three other Tubastrea species, Botrylloides fuscus, Botrylloides simodensis, Campylopterus hemileucurus, Campylopterus rufus, Campylopterus largipennis, Campylopterus villaviscensio, Phaethornis longuemareus, Florisuga mellivora, Lampornis amethystinus, Amazilia cyanocephala, Archilochus colubris, Epinephelus lanceolatus, Epinephelus fuscoguttatus, Symbiodinium temperate-A clade, Gammarus fossarum, Gammarus roeselii, Dikerogammarus villosus and Limnomysis benedeni. This article also documents the addition of 72 sequencing primer pairs and 52 allele specific primers for Neophocaena phocaenoides.