Time-related alterations and other confounding factors in direct sediment contact tests

Sediments are an important sink for and source of environmental pollutants. Because of these two qualities, sediments have been acknowledged in environmental risk assessment and corresponding regulations. In case of the European Water Framework Directive (EWFD), for instance, sediments were addressed by means of an amendment published in 2008. In order to adequately response to sediment contamination within the context of this legal obligation, however, there is still a need for research in several key fields. One open question is the correlation between the effects of pollutants in sediments and factors which can potentially confound such effects. The present thesis aimed to investigate potential confounding impacts in order to identify the importance of the given influences on biotests results and eventually the assessment of sediment contamination. Impact of contact-time on results obtained in biotests appeared to be of particular priority. Hence, time-dependence of data acquired in direct sediment contact tests (SCTs) was the major focus of the given studies. Further investigated were the impacts of test vessel material, extraction methodology, and difference between single- and multi-metal exposures. A literature survey and two experimental studies were carried out in order to address the time-dependence of results obtained in SCTs. The main experimental investigation compared the impact of time on the effects of 3,4-dichloroaniline (DCA), fluoranthene (FA) and pentachlorophenol (PCP) in the zebrafish (Danio rerio) embryo SCT, the nematode SCT with Caenorhabditis elegans, and the bacterial SCT with Arthrobacter globiformis. The second experimental study assessed the impact of short-term (two week) contact-time between sediments and pollutants on bioaccumulation of FA and PCP in the earthworm species Lumbriculus variegatus. All three studies concurrently revealed that time can severely impact results of SCTs. Literature indicated a complete loss of initially observable effects within weeks and statistically significant changes in effects in different SCT which mostly utilised invertebrates. These changes in effects included both increases and decreases, depending on the respective combination of species, substances, sediments, and additional accompanying parameters. In the two experimental studies, the impact of time was sufficiently severe that distinct changes of results were observed. In the case of the fish embryo SCT, a complete loss of originally observed toxicity of all three substances was recorded within two to sixteen weeks. In the nematode contact test, low number of juveniles in comparison to controls indicated toxicity in all four tested combinations of sediments and substances. Reproduction of worms exposed to FA-spiked natural sediment increased in the test conducted after 4 weeks of contact between FA and the sediment, and reached reproduction in the controls after six weeks. In the other three combinations of sediment and substance that were tested in the nematode SCT, no changes were observed until the end of experiments after six weeks. Toxicity of PCP remained unchanged in the bacterial SCT over the course of 44 weeks in both sediments. Accumulation of PCP in earthworms significantly decreased in FA-spiked natural sediment within 13 days of ageing and exhibited no change in PCP-spiked artificial sediment and both FA-spiked sediments. In an additional study, four different sediment extraction methodologies and one corresponding SCT were intercompared with regard to biological effectiveness on zebrafish embryos: Soxhlet extraction (SOX), membrane dialysis extraction (MDE), hydroxypropyl-beta-cyclodextrin extraction (HPCD) and Tenax TA® extraction (TNX). TNX was identified as a putatively suitable method for the prediction of toxicological bioavailability in the SCT with zebrafish embryos. SOX, MDE and HPCD overestimated bioaccessibility, as was expected for SOX and MDE. HPCD is a methodology that was reported to be predictive of bioaccessibility, but induced effects at least as great as effects induced by the two vigorous extraction methodologies Soxhlet and MDE. This result may be explained by short sediment-pollutant contact-time, and indicated that results obtained from different extraction methodologies can be severely altered by time. In an investigation comparing the impact of glass or plastic test vessel material on results in the aqueous zebrafish embryo test, glass vessels were shown to be better suited for testing of DCA. In the case of plastic, 16 hours of DCA-vessel contact prior to embryo exposure resulted in an almost complete loss of effects in the tested concentrations, whereas in glass vessels, an LC50 could still be obtained. These results confirm that currently discussed measures, such as semi-static tests, passive dosing and flow-through renewal, are necessary to minimise the influence of vessel material. Furthermore, experimental timing, i.e. the exact time test compounds are introduced into test vessels, can be crucial for the outcomes of experiments. The final study assessed uptake, accumulation, and corresponding changes in transcript abundance of selected genes in zebrafish embryos exposed to Cd-, Cu-, Ni- and Zn-spiked sediment. In terms of potentially confounding impacts, multi- vs. single-metal exposures were compared. Results emphasized the great relevance of Cu as toxicant on fish. Furthermore, it was shown that multi-metal exposure can impact uptake and bioaccumulation, most likely by means of inhibition and competition. This correlation was established for the first time concerning the zebrafish embryo sediment contact test. Literature indicated that this impact can be further influenced by additional confounding factors like pH or time. Transcript abundances were not sensitive to these changes. This finding could indicate that molecular biomarkers are more robust in their response to confounding factors than other endpoints, but needs to be further investigated. In most studies in the present thesis, a recurring comparison was conducted on the impact of sediment type, either artificial or natural sediment, on results in spiking-based investigations of sediment contamination. Results indicated no preference for either type, and depended, again, on the specific combination of species and substance. It is concluded that both artificial and natural sediments are very useful tools with advantages as well as inherent limitations. A parallel application should be considered in spiking-based assessments. The observed significant alterations of results induced by time concurrently indicated a need to developed research strategies to sustainably address time-dependence as potential confounding impact. Difference between exposures to single substances or mixtures was also identified as a relevant impact. Both identified influences might result in severe over- or underestimations of risks associated with contamination of sediments. However, acknowledgement of factors impacting results in environmental assessment creates a novel challenge. Assessing confounding impacts apparently calls for more tests, whereas economical and animal welfare considerations prioritise a minimisation of test demands. Hence, a three step, long-term strategy is proposed, which can eventually reduce necessity for extensive testing and address confounding impacts at the same time: a) Pilot-studies on the targeted confounding impacts (such as given in the present thesis), b) Integrative studies to assess the interlinking of the addressed impacts between different organisational levels and c) Inclusion of the confounding impacts into holistic, predictive models. In addition, the novel idea of adverse outcome pathways was identified as promising conceptual strategy to characterise and understand the mechanisms of confounding impacts. To realise such long-term research strategies on the organisational level, concentrated cooperational efforts in research frameworks are necessary. Current EU-founded projects such as MODELKEY provide examples how frameworks can be conceived that are developed to address the challenge posed by confounding impacts. The impact of time, in particular, was identified to be sufficiently severe to justify its inclusion into such frameworks.

Grantor dc:publisher

Publikationsserver der RWTH Aachen University

Year dc:date

2011

Author dc:creator

• Zielke, Hanno

Contributors dc:contributor

• Hollert, Henner

• info:eu-repo/classification/ddc/570
• Aquatisches Sediment
• Biotest
• Zebrabärbling
• Umweltgift
• Alterung
• Extraktion
• Biowissenschaften, Biologie
• Optimierung
• Testsystem
• direkter Sedimentkontakttest
• test optimisation
• direct sediment contact test
• zebrafish
• ageing

Statement dc:rights

• info:eu-repo/semantics/openAccess

Language dc:language

eng

Identifier

https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-115054%22

OAI record oai:identifier

https://publications.rwth-aachen.de/oai2d?verb=GetRecord&metadataPrefix=oai_dc&identifier=oai%3Apublications.rwth-aachen.de%3A52861