Monitoring of Bt-Maize pollen exposure in the vicinity of the nature reserve Ruhlsdorfer Bruch in northeast Germany 2007 to 2008

Background, aim, and scope Basically, technological innovations are associated with benefits and risks. This is also true for the introduction of genetically modified organisms (GMO) into agriculture. In Germany, precautionary regulations currently demand isolation distances (i.?e. buffer zones) for the cultivation of genetically modified maize (Bt-maize) in the vicinity of conventional (150?m) and organic maize fields (300?m). The Bt-toxin may harm non-target organisms (NTO) such as Lepidoptera. Despite this, corresponding regulations for the protection of nature reserves are lacking to date. Conventional and Bt-maize have been grown in the vicinity of the Flora-Fauna-Habitat (FFH) Ruhlsdorfer Bruch in Brandenburg, Germany. The aim of this study was to investigate whether exposure of maize pollen from surrounding Bt-maize fields to NTOs in the nature protection area could be excluded or not. Two types of exposure were investigated: Firstly, whether maize pollen was dispersed by wind into the nature reserve area exposing resident NTOs. Secondly, foraging NTOs from the nature reserve are exposed by roaming the surrounding fields and collecting maize pollen. In order to fulfil the precautionary principle defined by law, the study should help to determine appropriate isolation distances for the cultivation of Bt-maize with regard to sustainable protection of NTOs in the FFH Ruhlsdorfer Bruch. In 2007, the local authorities issued an isolation distance between Bt-maize fields and nature reserves of 100?m and in 2008, this became 250?m in the northern and 500?m in the westerly direction, respectively. Materials and methods Standardised methods for biological and technical pollen sampling issued by the Association of German Engineers (VDI 4330 Part 3, 2007 and VDI 4330 Part 4, 2006) were applied providing a quality controlled and methodologically harmonised database which does not only serve the needs to be fulfilled by the present case-specific monitoring study but can also be used as a reference database for further investigations. Maize pollen exposure was measured within the FFH Ruhlsdorfer Bruch and its immediate vicinity in July and August 2007 and 2008. In 2007, the sampling was performed at three sites using 12 technical samplers (Sigma-2/PMF) placed at five measuring points at distances from 5?m to 120?m from the maize field edges. Additionally, for biological pollen sampling six bee colonies were situated at these three sites (two colonies at each site). The technical sampler Sigma-2/PMF enables point sampling which is primarily influenced by wind and topography providing information on effective maize pollen input (flow and deposition) at the measuring sites. Honey-bees roaming in the surrounding area with typical foraging distances of several kilometres may act as planar collectors. They may serve as indicators for the exposure to pollen-collecting NTOs. Furthermore, biological sampling is more selective due to the organism’s preferences, whereas the technical sampling is neutral. Hence, both the technical and the biological sampling complement each other in their scope of application. The pollen samples of both matrices were analysed microscopically and the maize pollen loads were quantified. Pollen-DNA was analysed by means of the quantitative PCR-method (qPCR) identifying conventional and Bt-maize pollen by two independent laboratories. In 2008, the monitoring was repeated with additional sites. Eighteen technical samplers were exposed at five sites with eight measuring points at distances from 5?m to 250?m from the maize field edges. Two honey-bee colonies for biological sampling were placed at one site for control purposes. PCR-analyses were performed to measure the amounts of Bt-maize pollen in the samples. Results The results of pollen monitoring at the Ruhlsdorfer Bruch revealed maize pollen exposure for all monitoring sites in both surveys. In 2007, up to 1.75 million maize pollen/m² were deposited at sites closest to the maize field. At 120?m from field edge in the middle of the FFH-reserve, 99,000 maize pollen/m² were detected. In 2008, similar results were found, at distances up to 250?m from the field edges deposition of 164,000 maize pollen/m² was detected. Data on maize pollen deposition show a clear distance relationship and are in accordance with results of further comprehensive surveys based on the same methodology (Hofmann 2007). The results of the microscopic analysis of the pollen pellets demonstrated that bees collected maize pollen at all sites, in 2007 and 2008. Although maize pollen is not the main food source (2007:0.1–0.3?%; 2008:2–3?%) the collection efficiency of the bee colonies resulted in high amounts of sampled maize pollen with 4 to 11 million per site in 2007 and up to 467 million in 2008. Molecular-biological analysis of maize pollen DNA by qPCR demonstrated that transgenic Bt-MON810 DNA was present in all technical and biological samples, corroborated by two independent laboratories. In 2007, the GMO-content in the samples ranged up to 44?% in the bioaerosols and 49?% in the pollen pellets. In 2008, GMO-proportions of up to 18?% were detected. Discussion The results of this study provide evidence that NTOs in the Ruhlsdorfer Bruch were exposed to Bt-maize pollen under the cultivation conditions in 2007 with a buffer zone of 100?m. The GMO-content reached up to 48?%. The results of the monitoring in 2008 confirmed these findings. Even though the exposure could be reduced by increasing the isolation distances to 250?m and 500?m respectively, the results still show percentages of up to 18?% Bt-MON810 in the pollen samples. The results on maize pollen deposition at the Ruhlsdorfer Bruch in 2007 and 2008 correspond to the results of an investigation which was conducted over several years applying the same standardised method, but covering a wider range of distances. The correlation between maize pollen deposition (n/m²) and distance to the source field (m) fitted best to a power function of the type y = 1.2086 · 10⁶ · x ^–0.548. Despite the same trend, the pollen deposition in the Ruhlsdorfer Bruch revealed above-average findings. Also the analysis of the pollen pellets collected by the bees showed an exposure in 2007 with values for the GMO-content of up to 49?%. For both methods, the exposure decreased in 2008 due to the greater buffer zones up to 500?m. Whereas the GMO-content for the biological sampling were reduced to values below 10?%, the values for the technical sampling were still higher indicating that greater buffer zones would be necessary for safety reasons under the precautionary principle. Conclusions The results of this investigation proved that maize pollen were dispersed by wind to distances farther than 250?m from field edge leading to maize pollen exposure in the centre of the nature reserve. The results also demonstrated that foraging NTOs living in the nature reserve were exposed to maize pollen from surrounding fields. Considering the cultivation of Bt-maize MON810, the assumption of the Environmental Risk Assessment (ERA) that there will be no relevant exposure beyond the Bt-maize fields, cannot be confirmed. Considering the results of this and related studies and with respect to the precautionary principle, one can state that buffer zones between Bt-maize fields and protected areas are an effective measure to minimise the exposure of Bt-maize pollen to NTOs and, thus, to prevent from adverse effects. Recommendations and perspectives Because of still insufficient ecotoxicological data for the risk assessment of Bt-maize MON810 considering butterflies and other protected NTOs, protection standards assuring the precautionary principle have to be implemented to avoid Bt-maize pollen exposure to NTOs. This applies for the case Ruhlsdorfer Bruch and for nature reserve areas in general. In order to exclude risks to protected NTOs occurring in nature reserves, sufficient buffer zones for Bt-maize cultivation should be considered. The statistical analysis revealed that distances of more than 500?m are necessary to decisively reduce exposure to foraging insects. In fact, distances of more than 1,000?m are necessary to prevent maize pollen deposition from values above 100,000 pollen/m² with a certainty of 90?%. An adequate risk assessment can only be attained if based on field measurements accounting for the high variation of pollen deposition due to local environmental site conditions and field management. The monitoring should be based on standardised methods. It should include locations with the highest expected deposition rates, the boundaries of the protected areas and sites of interest within those boundaries, e.?g., specific habitats of sensitive species.     

CADMIUM ACCUMULATION AND TOXICITY IN SUBMERGED PLANT HYDRILLA VERTICILLATA (l.f.) ROYLE

Cadmium accumulation and its toxicity in relation tochlorophyll, protein, cysteine contents and in vivo nitrate reductaseactivity were studied under controlled conditions in Hydrillaverticillata, a submerged commonly occurring macrophyte. Plants weresubjected to six different concentrations of Cd ranging from 1.0 to 25.0 µM for 24, 48, 72 and 168 h. Tissue Cd concentration was maximum (13.71 µmoles/g dw) at 25 µM background concentration. At this concentration, a decrease of approximately 79 and 72% was found in chlorophyll and protein content. In vivo nitrate reductase activity was stimulated at 1.0 µM; however, the activity gradually declined beyond this concentration. Exposure to various cadmium concentrations resulted in an increase in cysteine content of the plant.     

Bryophyte responses to microclimatic edge effects across riparian buffers.

Although riparian buffers are an important aspect of forest management in the boreal forest of Canada, little is known about the habitat conditions within buffers, due in part to complex edge effects in response to both the upland clearcut and the stream. We investigated microclimatic conditions and bryophyte growth and vitality in seven locations between the stream edge and 60 m into the upland undisturbed conifer forests and at the clearcut sites with riparian buffer 30 km northwest of Thunder Bay, Ontario, Canada. We hypothesized that the growth and vitality of a pleurocarpous moss, Hylocomium splendens, and an acrocarpous moss, Polytrichum commune, would be directly related to the microclimatic gradients detected. We further hypothesized that sensitivity of the bryophytes to environmental factors will vary depending on their life form type, i.e., pleurocarpous moss will respond differently than the acrocarpous moss. Both bryophyte species were transplanted in pots and placed at 10-m intervals along 60-m transects perpendicular to the stream across the buffer and undisturbed sites. Bryophyte growth, cover, and vitality, as well as microclimatic parameters and plant cover, were measured over the summer in 2003. The riparian buffers were simultaneously affected by microclimatic gradients extending from both the clearcut edge and the riparian-upland ecotonal edge. Both bryophyte species responded to changes in the microclimatic conditions. However, vapor pressure deficit (VPD) was the most important factor influencing the growth of H. splendens, whereas for P. commune growth soil moisture was most important. Our study confirms earlier findings that interior forest bryophytes such as H. splendens can be used as indicators to monitor edge effects and biodiversity recovery following forest harvesting. We demonstrate that growth and vitality of these bryophytes reflect the prevailing near-ground microclimatic conditions at the forest edges. Abundance estimates of such bryophytes can be used to determine the depth of edge effects across both ecotonal edges (e.g., riparian-upland forest edge) and anthropogenically created edges (e.g., clearcut edge). Forest management practices must consider depth of edge in determining the appropriate width of riparian buffers that would be necessary to sustain biodiversity and associated values at the land/water interface.     

Modeling the heat transfer in acetylene cylinders during and after the exposure to fire

In this paper two different approaches for predicting the heating-up of an acetylene cylinder involved in a fire and the afterward cooling with water are presented. In the simulations polynomial functions were used to describe the temperature dependency of the thermal properties of the cylinder interior, which is a complex system composed by a solid porous material, a solvent (typically acetone) and acetylene dissolved in it. Model equations covered heat conduction in the cylinder interior and at its walls. In the first approach the cylinder surroundings were simulated as a further domain constituted by CO₂, during the fire exposure, and for water, during the cooling. In this domain the coupled heat transfer (convection and conduction) and the momentum equation (Navier–Stokes) were solved. In the second approach no further domain was considered but boundary conditions were set directly on the cylinder walls. Results of the calculations performed with both approaches are presented. This work could prove helpful in predicting to which extent the interior of an acetylene cylinder exposed to fire reaches temperatures capable of initiating the decomposition of acetylene and to determine how long a water cooling should be applied, so that the system is brought again under non-critical conditions.     

Screening and identification of early warning algal species for metal contamination in fresh water bodies polluted from point and non-point sources

The water bodies of Lucknow, Unnao and Kanpur (U.P.), India polluted through various point and non point sources were found to be either eutrophic or oligotrophic in nature. These water bodies supported a great number of algal diversity, which varied seasonally depending upon the physico-chemical properties of water. Further, the water bodies polluted through non point sources supports diverse algal species, while the water bodies polluted through point sources supports growth of tolerant blue green algae. High biomass producing algal species growing in these water bodies have accumulated significant amount of metals in their tissues. Maximum amount of Fe was found accumulated by species of Oedogonium sp. II (20,523.00 mug g(-1) dw) and Spirogyra sp. I (4,520.00 mug g(-1) dw), while maximum Chromium (Cr) was found accumulated in Phormedium bohneri (2,109.00 mug g(-1) dw) followed by Oscillatoria nigra (1,957.88 mug g(-1) dw) and Oedogonium sp. I (156.00 mug g(-1) dw) and Ni in Ulothrix sp. (495.00 mug g(-1) dw). Results showed that some of these forms growing in polluted environment and accumulating high amounts of toxic metals may be used as bioindicator species, however, their performance in metal contaminated water under different ecological niche is to be ascertained.     

Eine Weight-of-Evidence-Studie zur Bewertung der Sedimentbelastung und des Fischrückgangs in der Oberen Donau

Background and aim Despite intensive and continuous stocking and improvement of water quality since the 1970s, fish populations, especially those of the grayling (Thymallus thymallus), have declined over the last two decades in the upper Danube River (Germany). In order to assess 1) possible links between molecular/biochemical responses and ecologically relevant effects, and 2) if ecotoxicological effects might be related to the decline in fish catches in the upper Danube river, sediment samples and fish were collected at different locations and analyzed using a weight-of-evidence (WOE) approach with several lines of evidence. The objective of the presentation is to introduce the conceptual framework and to review results of the ongoing study. As previously addressed by Chapman and Hollert (2006) a variety of lines of evidence can be used in WOE studies. Briefly, 1) a comprehensive battery of acute and mechanism-specific bioassays was used to characterize the ecotoxicological hazard potential. 2) Histopathological investigations and the micronucleus assay with erythrocytes were applied, analyzing in situ parameters. 3) Diversity and abundance of benthic macroinvertebrates and fish as well as 4) persistent organic pollutants, endocrine disrupting substances, limnochemical parameters and the concentration of heavy metals were recorded. To identify organic contaminants a spotential causes of sediment toxicity assays, 5) effect directed analysis was applied.