Integral assessment of estrogenic potentials in sediment-associated samples

Background, aim, and scope  The enzyme-linked receptor assay (ELRA) detects estrogenic and anti-estrogenic effects at the molecular level of receptor binding and is a useful tool for the integrative assessment of ecotoxicological potentials caused by hormonally active agents (HAA) and endocrine disrupting compounds (EDC). The main advantage of the ELRA is its high sample throughput and its robustness against cytotoxicity and microbial contamination. After a methodological adaptation to salinity of the ELRA, according to the first part of this study, which increased its salinity tolerance and sensitivity for 17-β-estradiol, the optimised ELRA was used to investigate 13 native sediments characterised by different levels of salinity and chemical contamination. The applicability of the ELRA for routine analysis in environmental assessment was evaluated. Salinity is often a critical factor for bioassays in ecotoxicological sediment assessment. Therefore, salinity of the samples was additionally adjusted to different levels to characterise its influence on elution and binding processes of receptor-binding substances. Materials and methods  The ELRA was carried out with the human estrogen receptor α (ER) in a 96-well microplate format using the experimental setup known from the competitive immunoassay based on ligand–protein interaction. It is an important improvement that a physiologically relevant receptor was used as a linking protein instead of an antibody. The microplates were coated with a 17-β-estradiol-BSA conjugate, and dilution series of estradiol and of native sediment samples were added and incubated with the ER. After a washing step, a biotinylated mouse anti-ER antibody was added to each well. Receptor binding to estradiol, agonistic and antagonistic receptor binding, were determined by a streptavidin-POD-biotin complex with subsequent measurement of the peroxidase activity at the wavelength of 450 nm using a commercial ELISA multiplate reader. The sediment elutriates and pore water samples of sediments were tested in a dilution series to evaluate at which dilution step the receptor-binding potential ends. In the elution process (see Section 2.1 to 2.2), a method was developed to adjust the salinity to the levels of the reference testings, which offers an appropriate option to adjust the salinity in both directions. Statistical evaluation was made with a combination of the Mann–Whitney U test and the pT-method. Results  This part of the study characterised the environmental factor ‘salinity’ for prospective applications of the ELRA. Using reference substances such as 17-β-estradiol, the ELRA showed sigmoid concentration-effect relations over a broad range from 0.05 μg/l to 100 μg/l under physiological conditions. After methodological optimisation, both sensitivity and tolerance of the assay against salinity could be significantly raised, and the ELRA became applicable under salinity conditions up to concentrations of 20.5‰. The mean relative inter-test error (n = 3) was around 11% with reference substances and below 5% for single sediments elutriates in three replicates each. For sediment testings, the pore water and different salinity-adjusted elutriates of 13 sediments were used. A clear differentiation of the receptor-binding potential could be reached by application of the pT-method. Thereby, pT-values from one to six could be assigned to the sediments, and the deviation caused by the different salinity conditions was one pT-value. The mean standard deviation in the salinity adaptation procedure of the elutriates was below 5%. Discussion  Although the ELRA has already been used for assessments of wastewater, sludge and soil, its applicability for samples to different salinity levels has not been investigated so far. Even if the ELRA is not as sensitive as the E-screen or the YES-assay, with regard to reference substances like 17-β-estradiol, it is a very useful tool for pre-screening, because it is able to integrate both estrogenic as well as anti-estrogenic receptor-binding effects. According to the results of sediment testing, and given the integrative power to detect different directions of effects, the ELRA shows sufficient sensitivity and salinity tolerance to discriminate receptor-binding potentials in environmental samples. Conclusions  The optimised ELRA assay is a fast, cost-effective, reliable and highly reproducible tool that can be used for high-throughput screening in a microplate format in detecting both estrogenic and anti-estrogenic effects. Additionally, the ELRA is robust against microbial contaminations, and is not susceptible towards cytotoxic interferences like the common cell-culture methods. The general applicability and sufficient sensitivity of the ELRA was shown in freshwater environments. Marine and brackish samples can be measured up to salinity levels of 20.5‰. Recommendations and perspectives  In view of the proven sensitivity, functionality and the fastness of the ELRA, it is recommendable to standardise the test method. At the moment, no adequate in vitro test procedure exists which is standardised to DIN or ISO levels. The E-screen and the yeast estrogen/androgen screens (YES/YAS) sometimes underlie strong cytotoxic effects, as reported in the first part of this study. Further development of an ELRA assay using human androgen receptors appears to be very promising to gain information about androgenic and anti-androgenic effects, too. This would offer a possibility to use the ELRA as a fast and reliable pre-screening tool for the detection of endocrine potentials, thus minimising time and cost-expensive animal experiments.     

Pesticide concentrations in agricultural storm drainage inlets of a small Swiss catchment

Agricultural pesticides transported to surface waters pose a major risk for aquatic ecosystems. Modelling studies indicate that the inlets of agricultural storm drainage systems can considerably increase the connectivity of surface runoff and pesticides to surface waters. These model results have however not yet been validated with field measurements. In this study, we measured discharge and concentrations of 51 pesticides in four out of 158 storm drainage inlets of a small Swiss agricultural catchment (2.8 km²) and in the receiving stream. For this, we performed an event-triggered sampling during 19 rain events and collected plot-specific pesticide application data. Our results show that agricultural storm drainage inlets strongly influence surface runoff and pesticide transport in the study catchment. The concentrations of single pesticides in inlets amounted up to 62 µg/L. During some rain events, transport through single inlets caused more than 10% of the stream load of certain pesticides. An extrapolation to the entire catchment suggests that during selected events on average 30 to 70% of the load in the stream was transported through inlets. Pesticide applications on fields with surface runoff or spray drift potential to inlets led to increased concentrations in the corresponding inlets. Overall, this study corroborates the relevance of such inlets for pesticide transport by establishing a connectivity between fields and surface waters, and by their potential to deliver substantial pesticide loads to surface waters.

     

Wax perception in honeybees: contact is not necessary

In social insects, much progress has been made in identifying variations in the cuticular signatures of sexes, castes, kin and reproductive status. In contrast to this, we still do not know how the receivers perceive these recognition cues. This study was designed to investigate whether honeybees use contact-chemosensory or olfactory sensilla to perceive wax components. To answer this question in a behavioral assay, we combined classical conditioning of the proboscis extension reaction and a recently established method using zinc sulfate to selectively block antennal contact-chemosensory sensilla. Comparison of the responses to sucrose, wax and geraniol before and after antennal zinc sulfate treatment revealed that the sucrose response is lost after treatment but the responses to wax and geraniol are maintained. As sucrose is perceived by the contact-chemosensory sensilla, the retention of the wax response indicates that contact-chemosensory sensilla are not necessary for wax perception.     

Learning from local knowledge: modeling the pastoral-nomadic range management of the Himba, Namibia.

It is widely accepted that successful grazing management strategies in semiarid ecosystems need to be adapted to the highly temporal and spatially heterogeneous forage production. Nevertheless, a full understanding of the key factors and processes for sustainable adaptive management has yet to be reached. The investigation of existing, successful range management systems by simulation models may help to derive general understanding and basic principles. The semi-nomadic Himba in northern Namibia applied a sophisticated management system until the mid-1990s which combined season-dependent pasture use (resulting in rainy-season pastures and dry-season pastures), preservation of reserves for drought and sanctions for rule breaking. A stochastic ecological simulation model has been developed here which represents the main aspects of this management system. With this model we analyze (1) which components of the traditional Himba strategy are essential for sustainability and (2) what happens to the state of the rangeland system under socioeconomic changes. This study shows that temporally and spatially heterogeneous pasture use yields higher productivity and quality of a pasture area than the pressure of homogeneous permanent grazing. Two aspects are of importance: (1) intra-annual heterogeneous use (resting of the dry-season pastures during the rainy season) and (2) interannual heterogeneous use (spatial extension of grazing in years of drought). This management system leads to an effective build-up and use of a buffer in the system: the reserve biomass (the non-photosynthetic reserve organs of the plants), an indicator for grazing and management history. Analyzing purchase as one form of socioeconomic change, we demonstrate that easier market access to purchase livestock may lead to a decline in vegetation quality. However, cattle production increases as long as rest periods on parts of the pasture during the rainy season are granted. Methodologically, we emphasize that simulation models offer an excellent framework for analyzing and depicting basic principles in sustainable range management derived from local knowledge. They provide the opportunity of testing whether these basic principles are also valid under different ecological and socioeconomic settings.     

Volatile nanoparticle formation and growth within a diluting diesel car exhaust

A major source of particle number emissions is road traffic. However, scientific knowledge concerning secondary particle formation and growth of ultrafine particles within vehicle exhaust plumes is still very limited. Volatile nanoparticle formation and subsequent growth conditions were analyzed here to gain a better understanding of "real-world" dilution conditions. Coupled computational fluid dynamics and aerosol microphysics models together with measured size distributions within the exhaust plume of a diesel car were used. The impact of soot particles on nucleation, acting as a condensational sink, and the possible role of low-volatile organic components in growth were assessed. A prescribed reduction of soot particle emissions by 2 orders of magnitude (to capture the effect of a diesel particle filter) resulted in concentrations of nucleation-mode particles within the exhaust plume that were approximately 1 order of magnitude larger. Simulations for simplified sulfuric acid-water vapor gas-oil containing nucleation-mode particles show that the largest particle growth is located in a recirculation zone in the wake of the car. Growth of particles within the vehicle exhaust plume up to detectable size depends crucially on the relationship between the mass rate of gaseous precursor emissions and rapid dilution. Chassis dynamometer measurements indicate that emissions of possible hydrocarbon precursors are significantly enhanced under high engine load conditions and high engine speed. On the basis of results obtained for a diesel passenger car, the contributions from light diesel vehicles to the observed abundance of measured nucleation-mode particles near busy roads might be attributable to the impact of two different time scales: (1) a short one within the plume, marked by sufficient precursor emissions and rapid dilution; and (2) a second and comparatively long time scale resulting from the mix of different precursor sources and the impact of atmospheric chemistry.     

Dynamics of Bacterial Abundance,Biomass, Activity,and Community Composition in the Oligotrophic Traunsee and the Traun River (Austria)

The holomictic Traunsee is the deepest and second largest lakein Austria. The special characteristic of this ecosystem isthe fact that local salt and soda industries presumably alterthe lake by the discharge of waste materials. Since thebeginning of the 20th century salt and soda works areannually releasing up to 50,000 tons of solid wastes and up to150,000 tons of chloride into Traunsee. To assess potentialeffects of these anthropogenic impacts on the bacterioplanktonthree sampling sites, influenced as well as not influenced bythe industrial discharge, were chosen for comparison andsampled monthly from November 1997 to October 1998. Bacterialabundance ranged between 0.4 to 3.0 × 10⁶ cells ml^-1 with decreasing numbers along the depth profile. Theproportion of actively respiring bacteria, i.e. INT [2-(4-iodophenyl)-3-(4-nitrophenyl)-5-phenyltetrazolium chloride]reducing cells, never exceeded 10% of DAPI (4',6'-diamidino-2-phenylindole) stained cells. Fluorescence in situ hybridization (FISH) was used to examine the seasonal and spatial distribution of dominant phylogenetic groups of thebacterioplankton. Up to 84% of bacteria detected with DAPIcould be detected via FISH applying the universal bacterialprobe EUB338. Percentages of alpha- and beta-Proteobacteriaand members of the Cytophaga-Flavobacterium cluster did notexceed 60% of DAPI-stained cells.Beta-Proteobacteriaappeared to be the most abundant group, not only in Traunsee butalso in two reference lakes, Attersee and Hallstättersee. No significant differences in any of the bacterial parameters couldbe detected between the three sampling sites and all measurementswere found in the range reported for oligotrophic lakes. The highdischarge of the Traun River, resulting in a lake water renewaltime of only one year, may diminish possible effects of industrial waste discharge in the pelagic zone.     

Growth dynamics of Saccharina latissima (Laminariales,Phaeophyceae) in Aarhus Bay,Denmark, and along the species’ distribution range

Growth dynamics of Saccharina latissima in a Danish embayment (56°03.793N, 10°16.148E) were investigated through an annual cycle (March 1999–March 2000) and related to patterns found in previous studies covering the distribution range of the species. The kelps exhibited meristematic growth as well as distal tissue loss all year around. Elongation rate peaked in spring (March–May, 0.75 cm day^?1), whereas maximum biomass growth occurred in May–July at higher insolation. S. latissima accumulated nitrogen (N) in November–March, when ambient N levels were high and the fast growth in March–July was followed by a depletion of these reserves. Light regimes and seasonal fluctuations of nutrients were, thus, major factors explaining the seasonal growth pattern of S. latissima in this temperate bay. Differences in light and exposure along depth gradients affected the growth, loss and storage product dynamics. High losses of N and carbon (C) through summer abscission of distal tissue question the efficiency of translocation, especially at shallower depths, where losses are accelerated by, e.g., high temperature. A large-scale comparison further highlighted that warming advanced the timing, increased the level of peak growth and also seemed to expand the growth period. Growth rates of the studied Danish population fell in the low end of the range for similar latitudes and temperatures, probably due to sub-optimal salinities (avg. 23.3 psu) in combination with periodically high summer temperatures (max. 21.1 °C).     

塔里木河下游阿拉干断面胡杨树高的分布和密度

通过小尺度生态监测与遥感数据分析获取塔里木河下游胡杨(Populus euphratica Oliv.)林现状信息,将胡杨个体对输水的响应实测数据和QuickBird影像信息进行对比和分析,"面"上宏观调查与"点"上典型研究相结合,集中对塔里木河下游阿拉干断面设立的100 hm2长期监测样方内的4 500棵胡杨树进行了详细的调查.研究表明:胡杨密度随着离河岸距离增加而呈现降低的规律.不同树高的胡杨也呈一定规律分布.树高小于3 m的幼树主要分布在距河道50 m范围内;树高39 m之间的胡杨在研究区中占绝对优势;树高大于9 m的胡杨在距河道200 m内分布比较均匀,200 m之外分布稀少.