Background: Microinjection was tested as a potentially powerful tool to introduce natural and anthropogenic pollutants directly into fish eggs to determine their toxicological impact on fish. With this technique, parental transfer of lipophilic contaminants may be mimicked. Here, we investigated the applicability of pollutant injection into the yolk of early zebrafish (Danio rerio) eggs with special regard on survival after vehicle injection. Tested vehicles were autoclaved tap water, dimethyl sulfoxide (DMSO), methanol, and triolein. Results: Highest mortality occurred after the injection of DMSO and methanol. The lethality rates were up to 40% higher than under control conditions. Best survival rates were obtained after triolein and water injections. However, the triolein droplet was not assimilated by the embryo within 96 h post fertilization suggesting an incomplete uptake of triolein-solubilized chemicals. Technical aspects concerning microinjection in zebrafish eggs are discussed with special emphasis on quantitative injection. Conclusions: Microinjection into the yolk cell of zebrafish eggs is feasible, but the application of exact volumes appears problematic. However, microinjection is a powerful tool for studies without the demand for high volume accuracy. Adopting microinjection for pollutant research requires further investigation.

Maternal transfer of emerging brominated and chlorinated flame retardants in European eels AbstractThe European eel (Anguilla anguilla) is regarded as a critically endangered species. Scientists are in agreement that the "quality of spawners" is a vital factor for the survival of the species. This quality can be impaired by parasites, disease and pollution. Especially endocrine disrupting organic chemicals pose a potential threat to reproduction and development of offspring.To our knowledge, the findings in this publication for the first time describe maternal transfer of contaminants in eels. We analysed the concentrations of in total 53 polybrominated diphenyl ethers (PBDEs) and their halogenated substitutes in muscle, gonads and eggs of artificially matured European eels and in muscle and gonads of untreated European eels that were used for comparison. We found evidence that persistent organic pollutants such as PBDEs, as well as their brominated and chlorinated substitutes are redistributed from muscle tissue to gonads and eggs. Concentrations ranged from 0.001 ng g -1 ww for sum Dechlorane metabolites (DPMA, aCL 10 DP, aCl 11 DP) to 2.1 ng g -1 ww for TBA in eggs, 0.001 ng g -1 ww for Dechlorane metabolites to 9.4 ng g -1 ww for TBA in gonads and 0.002 ng g -1 ww for Dechlorane metabolites to 54 ng g -1 ww for TBA in muscle tissue.Average egg muscle ratios (EMRs) for compounds detectable in artificially matured eels from both Schlei Fjord and Ems River ranged from 0.01 for Dechlorane 602 (DDC-DBF) to 10.4 for PBEB. Strong correlations were found between flame retardant concentrations and lipid content in the analysed tissue types, as well as transfer rates and octanol-water partitioning

Persistent organic pollutants (POPs) are monitored regularly in water, sediment, and biota in the Baltic Sea. Lipophilic substances are measured in remarkable concentrations especially in the fatty parts of fish, such as herring (Clupea harengus). However, less lipophilic POPs, e.g. perfluorinated compounds (PFCs), can also be detected. For the first time to our knowledge, this study provides a broad range of contaminant concentrations simultaneously measured in filet, liver, and gonads of both sexes of Baltic herring. We analysed organochlorines, polybrominated diphenyl ethers (PBDEs), and PFCs in mature autumn-spawning individuals and found distinct organ pollutant pattern for all POPs in both sexes. POP concentrations found in the gonads of both sexes indicate that not only females but also males tend to reduce contaminants via reproduction. However, sex-dependent differences could be identified for hexachlorobenzene, PBDEs, and were most remarkable for PFCs. This transfer of contaminants to the gonads in both male and female herring is being underestimated, as it may directly affect the general reproduction success as well as the healthy development of the next generation. Hence, the accumulation of contaminants in the gonads should be considered one possible threat to a healthy wildlife as its achievement is stated by the Baltic Sea Action Plan. Inclusion of a periodic monitoring of POP concentrations in gonads of fish may be an important bioeffect measure to assess the environmental status of biota in the Baltic Sea.

Background: The fish embryo toxicity test (FET) is an established method in toxicology research for quantifying the risk potential of environmental contaminations and other substances. The typical results of the method are the half maximal effective concentration (EC 50 ) or the no observed effect concentration (NOEC). However, from an environmental perspective, it is most important to safely identify the concentration of the substance effect which lies above the effect under control condition (spontaneous effect). The common FET is not optimal to detect ECs for small target effects. This paper shows how to optimize the efficiency and consequently the benefit of the FET for small effects using an adequate experimental design. The approach presented here can be carried over to all test systems generating binary (yes/no) outcomes. Results: The experimental design has three components in this context: determination of spontaneous response, sample size calculation, and dose allocation. A strategy for all three components is proposed from which a design is given including precision requirements and makes the most effective use of the experimental effort. This strategy amounts to expanding the usual FET guidelines of Organisation for Economic Co-operation and Development, German Institute for Standardization, or American Society for Testing and Materials by adding a planning step that adapts the test to the specific user's need. Conclusions: For the practical calculation of an adapted design, a newly developed software is presented as R package toxtestD. It provides a user-friendly way of developing an optimal experimental design for the FET without in-depth statistical knowledge. The programme is suited for all experimental problems involving a binary outcome and a continuous concentration.