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There has been much effort to assess the efficacy of sewage treatment plants to remove chemicals with estrogenic activity, but other modes of toxicity have received less attention. This study assessed the efficacy of advanced tertiary technologies to remove estrogen receptor (ER), aryl hydrocarbon receptor (AhR), retinoic receptor (RAR) agonists and genotoxicity using recombinant gene bioassays. Untreated sewage contained ER, AhR and RAR agonists and was genotoxic. Activated sludge treatment removed over 69% of the ER, AhR and RAR aqueous phase activities. Ozonation and biologically activated carbon processes removed genotoxicity and most receptor agonists to below detection limits. Estrogenic activity was associated with the semi-polar fraction of the aqueous phase and non-polar phase of the influent (maximum 30%). AhR and RAR activities were found in both the aqueous and particulate phases of influent. Only RAR activity was frequently found in the polar fraction of aqueous and particulate phases of the process effluent. In the influent, the greatest RAR activity was found in the polar fraction of the aqueous phase (>65%). Genotoxic activity was detected in all influent fractions but rapidly decreased. The results showed that ER, AhR and RAR agonists were greatly reduced by the advanced tertiary treatment processes.

Thousands of organic micropollutants and their transformation products occur in water. Although often present at low concentrations, individual compounds contribute to mixture effects. Cell-based bioassays that target health-relevant biological endpoints may therefore complement chemical analysis for water quality assessment. The objective of this study was to evaluate cell-based bioassays for their suitability to benchmark water quality and to assess efficacy of water treatment processes. The selected bioassays cover relevant steps in the toxicity pathways including induction of xenobiotic metabolism, specific and reactive modes of toxic action, activation of adaptive stress response pathways and system responses. Twenty laboratories applied 103 unique in vitro bioassays to a common set of 10 water samples collected in Australia, including wastewater treatment plant effluent, two types of recycled water (reverse osmosis and ozonation/activated carbon filtration), stormwater, surface water, and drinking water. Sixty-five bioassays (63%) showed positive results in at least one sample, typically in wastewater treatment plant effluent, and only five (5%) were positive in the control (ultrapure water). Each water type had a characteristic bioanalytical profile with particular groups of toxicity pathways either consistently responsive or not responsive across test systems. The most responsive health-relevant endpoints were related to xenobiotic metabolism (pregnane X and aryl hydrocarbon receptors), hormone-mediated modes of action (mainly related to the estrogen, glucocorticoid, and antiandrogen activities), reactive modes of action (genotoxicity) and adaptive stress response pathway (oxidative stress response). This study has demonstrated that selected cell-based bioassays are suitable to benchmark water quality and it is recommended to use a purpose-tailored panel of bioassays for routine monitoring.

To understand mechanisms of tufa biofilm calcification, selected karstwater stream stromatolites in Germany have been investigated with regard to their hydrochemistry, biofilm community, exopolymers, physicochemical microgradients, calcification pattern and lamination. In stream waters, CO2 degassing drives the increase in calcite saturation to maximum values of approximately 10-fold, independent from the initial Ca2+/alkalinity ratio. For the cyanobacteria of tufa biofilms, a culture-independent molecular approach showed that microscopy of resin-embedded biofilm thin sections underestimated the actual diversity of cyanobacteria, i.e. the six cyanobacteria morphotypes were opposed to nine different lineages of the 16S rDNA phylogeny. The same morphotype may even represent two genetically distant cyanobacteria and the closest relatives of tufa biofilm cyanobacteria may be from quite different habitats. Diatom diversity was even higher in the biofilm at the studied exemplar site than that of the cyanobacteria, i.e. 13 diatom species opposed to 9 cyanobacterial lineages. The non-phototrophic prokaryotic biofilm community is clearly different from the soil-derived community of the stream waters, and largely composed of flavobacteria, firmicutes, proteobacteria and actinobacteria. The exopolymeric biofilm matrix can be divided into three structural domains by fluorescence lectin-binding analysis. Seasonal and spatial variability of these structural EPS domains is low in the investigated streams. As indicated by microsensor data, biofilm photosynthesis is the driving mechanism in tufa stromatolite formation. However, photosynthesis-induced biofilm calcification accounts for only 10-20% of the total Ca2+ loss in the streams, and occurs in parallel to inorganic precipitation driven by CO2-degassing within the water column and on biofilm-free surfaces. Annual stromatolite laminae reflect seasonal changes in temperature and light supply. The stable carbon isotope composition

Modified protocols of fluorescence in situ hybridization (FISH) and catalyze reporter deposition fluorescence in situ hybridization (CARD-FISH) were developed in order to detect bacteria in situ in calcified stromatolite biofilms. Smooth, well-preserved thin sections of calcified biofilms (~ 5 µm thin, vertical sectioning of ~ 1 cm deep) were obtained by cryo-sectioning using the adhesive tape-stabilization technique. A modified hybridization buffer was applied during hybridization to prevent calcite dissolution as well as false binding of oligonucleotide probes to the charged mineral surfaces. Particularly, bright and specific CARD-FISH signals allowed the detection of bacteria in intensively calcified biofilms even at low magnification, which is suitable for investigating millimeter- to centimeter-scale vertical distribution patterns of bacteria.