Modulation of steroidogenesis by coastal waters and sewage effluents of Hong Kong, China, using the H295R assay

BACKGROUND, AIM, AND SCOPE: The presence of a variety of pollutants in the aquatic environment that can potentially interfere with the production of sex steroid hormones in wildlife and humans has been of increasing concern. The aim of the present study was to investigate the effects of extracts from Hong Kong marine waters, and influents and effluents from wastewater treatment plants on steroidogenesis using the H295R cell bioassay. After exposing H295R cells to extracts of water, the expression of four steroidogenic genes and the production of three steroid hormones were measured. MATERIALS AND METHODS: Water samples were collected during the summer of 2005 from 24 coastal marine areas and from the influents and effluents of two major waste water treatment plants (WWTPs) in Hong Kong, China. Samples were extracted by solid phase extraction (SPE). H295R cells were exposed for 48 h to dilutions of these extracts. Modulations of the expression of the steroidogenic genes CYP19, CYP17, 3betaHSD2, and CYP11beta2 were determined by measuring mRNA concentrations by real-time polymerase chain reaction (Q-RT-PCR). Production of the hormones progesterone (P), estradiol (E2), and testosterone (T) was quantified using enzyme linked immunosorbent assays (ELISA). RESULTS: Extracts from samples collected in two fish culture areas inhibited growth and proliferation of H295R cells at concentrations greater or equal to 10(5) L equivalents. The cells were exposed to the equivalent concentration of active substances in 10,000 L of water. Thus, to observe the same level of effect as observed in vitro on aquatic organisms would require a bioaccumulation factor of this same magnitude. None of the other 22 marine samples affected growth of the cells at any dilution tested. Twelve of the marine water samples completely inhibited the expression of CYP19 without affecting E2 production; inhibition of CYP17 expression was observed only in one of the samples while expression of CYP11beta2 was induced as much as five- and ninefold after exposure of cells to extracts from two locations. The expression of the progesterone gene 3betaHSD2 was not affected by any of the samples; only one sample induced approximately fourfold the production of E2. Although more than twofold inductions were observed for P and T production, none of these values were statistically significant to conclude effects on the production of these two hormones. While influents from WWTPs did not affect gene expression, an approximately 30% inhibition in the production of E2 and a 40% increase in P occurred for the exposure with influents from the Sha Tin and Stonecutters WWTPs, respectively. Effluents from WWTPs did not affect the production of any of the studied hormones, but a decrement in the expression of the aldosterone gene CYP11beta2 was observed for the Sha Tin WWTP exposure. No direct correlation could be established between gene expression and hormone production. DISCUSSION: Observed cytotoxicity in the two samples from fish culture areas suggest the presence of toxic compounds; chemical analysis is required for their full identification. Although effluents from WWTPs did not affect hormone production, other types of endocrine activity such as receptor-mediated effects cannot be ruled out. Interactions due to the complexity of the samples and alternative steroidogenic pathways might explain the lack of correlation between gene expression and hormone production results. CONCLUSIONS: Changes observed in gene expression and hormone production suggest the presence in Hong Kong coastal waters of pollutants with endocrine disruption potential and others of significant toxic effects. The aromatase and aldosterone genes seem to be the most affected by the exposures, while E2 and P are the hormones with more significant changes observed. Results also suggest effectiveness in the removing of compounds with endocrine activity by the WWTPs studied, as effluent samples did not significantly affect hormone production. The H295R cell showed to be a valuable toll in the battery required for the analysis of endocrine disrupting activities of complex environmental samples. RECOMMENDATIONS AND PERSPECTIVES: Due to the intrinsic complexity of environmental samples, a combination of analytical tools is required to realistically assess environmental conditions, especially in aquatic systems. In the evaluation of endocrine disrupting activities, the H295R cell bioassay should be used in combination with other genomic, biological, chemical, and hydrological tests to establish viable modes for endocrine disruption and identify compounds responsible for the observed effects.     

Photocatalytic and combined anaerobic-photocatalytic treatment of textile dyes

A photocatalytic process based on immobilized titanium dioxide was used to treat crude solutions of azo, anthraquinone and phthalocyanine textile dyes. In addition, the process was applied to the treat autoxidized chemically reduced azo dyes, i.e. representatives of recalcitrant dye residues after biological sequential anaerobic-aerobic treatment. Photocatalysis was able to remove more than 90% color from crude as well as autoxidized chemically reduced dye solutions. UV-absorbance and COD were also removed but to a lower extent (50% in average). The end products of photocatalytic treatment were not toxic toward methanogenic bacteria. The results demonstrate that photocatalysis can be used as a pre- or post-treatment method to biological anaerobic treatment of dye-containing textile wastewater.     

Functionally relevant microorganisms to enhanced biological phosphorus removal performance at full-scale wastewater treatment plants in the United States

The abundance and relevance ofAccumulibacter phosphatis (presumed to be polyphosphate-accumulating organisms [PAOs]), Competibacter phosphatis (presumed to be glycogen-accumulating organisms [GAOs]), and tetrad-forming organisms (TFOs) to phosphorus removal performance at six full-scale enhanced biological phosphorus removal (EBPR) wastewater treatment plants were investigated. Coexistence of various levels of candidate PAOs and GAOs were found at these facilities. Accumulibacter were found to be 5 to 20% of the total bacterial population, and Competibacter were 0 to 20% of the total bacteria population. The TFO abundance varied from nondetectable to dominant. Anaerobic phosphorus (P) release to acetate uptake ratios (P(rel)/HAc(up)) obtained from bench tests were correlated positively with the abundance ratio of Accumulibacter/(Competibacter +TFOs) and negatively with the abundance of (Competibacter +TFOs) for all plants except one, suggesting the relevance of these candidate organisms to EBPR processes. However, effluent phosphorus concentration, amount of phosphorus removed, and process stability in an EBPR system were not directly related to high PAO abundance or mutually exclusive with a high GAO fraction. The plant that had the lowest average effluent phosphorus and highest stability rating had the lowest P(rel)/HAc(up) and the most TFOs. Evaluation of full-scale EBPR performance data indicated that low effluent phosphorus concentration and high process stability are positively correlated with the influent readily biodegradable chemical oxygen demand-to-phosphorus ratio. A system-level carbon-distribution-based conceptual model is proposed for capturing the dynamic competition between PAOs and GAOs and their effect on an EBPR process, and the results from this study seem to support the model hypothesis.     

Simplified contaminant source depletion models as analogs of multiphase simulators

Four simplified dense non-aqueous phase liquid (DNAPL) source depletion models recently introduced in the literature are evaluated for the prediction of long-term effects of source depletion under natural gradient flow. These models are simple in form (a power function equation is an example) but are shown here to serve as mathematical analogs to complex multiphase flow and transport simulators. The spill and subsequent dissolution of DNAPLs was simulated in domains having different hydrologic characteristics (variance of the log conductivity field=0.2, 1 and 3) using the multiphase flow and transport simulator UTCHEM. The dissolution profiles were fitted using four analytical models: the equilibrium streamtube model (ESM), the advection dispersion model (ADM), the power law model (PLM) and the Damkohler number model (DaM). All four models, though very different in their conceptualization, include two basic parameters that describe the mean DNAPL mass and the joint variability in the velocity and DNAPL distributions. The variability parameter was observed to be strongly correlated with the variance of the log conductivity field in the ESM and ADM but weakly correlated in the PLM and DaM. The DaM also includes a third parameter that describes the effect of rate-limited dissolution, but here this parameter was held constant as the numerical simulations were found to be insensitive to local-scale mass transfer. All four models were able to emulate the characteristics of the dissolution profiles generated from the complex numerical simulator, but the one-parameter PLM fits were the poorest, especially for the low heterogeneity case.     

Diffusion of PAH and PCB from contaminated sediments with and without mineral capping; measurement and modelling

A novel laboratory microcosm test was developed to measure the diffusion of native PAHs and PCBs from sediments in the presence and absence of a capping layer. Diffusive flux of 15 PAHs and 7 PCBs from uncapped sediment from Oslo harbour was 3.8+/-0.9 microg m(-2)d(-1) and 0.010+/-0.003 microg m(-2)d(-1), respectively. The flux from sediments capped with 1cm mineral cap (crushed limestone or crushed gneiss (0-2mm)), observed during the first 410 d, was 3.5-7.3% of the flux from uncapped sediments. By measuring freely dissolved pore water concentrations of 10 PAHs the flux in the microcosm was modelled with steady state and transient diffusion models. The measured flux from uncapped sediment was 27-290% of modelled steady state flux. Good agreement was also found between the measured flux of pyrene from capped sediment and the flux modelled with the transient model when fitting only with the distribution coefficients for pyrene between the cap material and water (Kd_pyr). Fitted Kd_pyr, (210 and 23 l kg(-1) for limestone and gneiss, respectively) was in the same order of magnitude as K(d) calculated from organic carbon content in the cap materials (68 and 14 l kg(-1) respectively). Calculation of the efficiency of a hypothetical cap with 10 cm diffusion path shows that the increased diffusion path length alone can yield a flux reduction >99% through a strong increase in the stagnant diffusive boundary layer from <1 to 100mm.     

Historical marine ecology: examining the role of fisheries in changes in North Sea benthos

The organisms living on and in the sea floor, the benthos, represent an important ecological group. Although some (shellfish) have an economic value, most do not, and so little long-term data are available. We have identified three sources of historic benthic data for the North Sea, a regional sea that has been subjected to multiple human impacts for at least several hundred years. Each dataset has its limitations, but by their use together some issues emerge. Wider community shifts were observed in the shorter term and a number of extirpations at the scale of the North Sea were seen over longer time scales. The extirpated taxa share a number of characteristics consistent with an effect of fisheries such as fragile morphology. We must concentrate now on furthering our understanding of the ecological significance of shifts in dominance of particular functional units and protecting those habitats and species most vulnerable to fisheries-driven extirpation.     

Influence of tidal regime on the distribution of trace metals in a contaminated tidal freshwater marsh soil colonized with common reed (Phragmites australis)

A historical input of trace metals into tidal marshes fringing the river Scheldt may be a cause for concern. Nevertheless, the specific physicochemical form, rather than the total concentration, determines the ecotoxicological risk of metals in the soil. In this study the effect of tidal regime on the distribution of trace metals in different compartments of the soil was investigated. As, Cd, Cu and Zn concentrations in sediment, pore water and in roots were determined along a depth profile. Total sediment metal concentrations were similar at different sites, reflecting pollution history. Pore water metal concentrations were generally higher under less flooded conditions (mean is (2.32 ± 0.08) × 10⁻³ mg Cd L⁻¹ and (1.53 ± 0.03) × 10⁻³ mg Cd L⁻¹). Metal concentrations associated with roots (mean is 202.47 ± 2.83 mg Cd kg⁻¹ and 69.39 ± 0.99 mg Cd kg⁻¹) were up to 10 times higher than sediment (mean is 20.48 ± 0.19 mg Cd kg⁻¹ and 20.42 ± 0.21 mg Cd kg⁻¹) metal concentrations and higher under dryer conditions. Despite high metal concentrations associated with roots, the major part of the metals in the marsh soil is still associated with the sediment as the overall biomass of roots is small compared to the sediment.     

Performance of a naturally growing Parmelioid lichen Remototrachyna awasthii against organic and inorganic pollutants

The present study aims to assess the level and possible sources of organic and inorganic pollutants in Mahableshwar city in Western part of India and their effect on naturally growing foliose lichen Remototrachyana awasthii (Hale & Patw.) Divakar & A. Crespo. This lichen species growing abundantly in the area was collected from eight different sites on the basis of anthropogenic activities detected in the area. The concentration of inorganic heavy metals (Al, As, Cd, Cr, Fe, Pb, Mn, and Zn) and polycyclic aromatic hydrocarbons (PAHs) were analyzed and correlated with photosynthetic pigments (Chl a, Chl b, total chlorophyll, and carotenoid) together with chlorophyll degradation and protein contents. The concentration of most of the metals at different sites was significantly greater than at the control site (P?<?0.001). The highest metal content was found at Bus Stand and Panchgani, a tourist place that experiences heavy traffic activities. The concentration of PAHs, particularly of two-and three-ringed PAHs, was also found to be the highest in samples collected at Bus Stand area. The chlorophyll degradation and protein content were found to be the most sensitive parameters to assess the vitality of lichen thallus against wide range of air pollutants. The effectiveness of R. awasthii as a biomonitor will be investigated in the near future by comparing this species with other biomonitors.     

Compositional changes of crude oil SARA fractions due to biodegradation and adsorption on colloidal support such as clays using Iatroscan

The compositional changes of saturates, aromatics, resins and asphaltenes (SARA) fractions in aqueous clay/oil microcosm experiments with a hydrocarbon-degrading microorganism community were analysed using Iatroscan. The clay mineral samples used in this study were organomontmorillonite, acid-activated montmorillonite and K, Ca, Zn and Cr montmorillonites produced by modifying the original montmorillonite sample. The evaluation and quantification of biodegradation and adsorption were carried out using a combination of the Iatroscan and gravimetric analysis. The SARA compositions in the presence of organomontmorillonite and acid-activated montmorillonite after incubation follow the same pattern in which the aromatic fraction is higher than the other fractions unlike in the presence of unmodified, K, Ca and Zn montmorillonites, where the saturates fraction is higher than the other fractions. Changes in SARA fractions due to biodegradation seemed to occur most in the presence of unmodified and calcium montmorillonites; hence, the removal of SARA fractions due to biodegradation was significant and enhanced in the presence of these two clay samples. However, biodegradation in the presence of organomontmorillonite and acid-activated and Cr montmorillonites was hindered. The study indicated that Cr montmorillonite adsorbed resins most, whereas Zn and K montmorillonites adsorbed aromatics most after incubation.     

Partitioning, sources and variability of regional and local oxidant (OX = O3 + NO2) in a coastal rural area in the southwest of Iberian Peninsula

The purpose of this work is to investigate the behaviour and variability of oxidant levels (OX?=?NO₂?+?O₃), for the first time, in a rural coastal area in the southwest of the Iberian Peninsula, affected by several air masses types. Detailed database (built-up over the years 2008 to 2011, and containing around 500,000 data) from the Atmospheric Sounding Station “El Arenosillo” was used. The observed daily cycles of NO _x and OX were influenced by air masses coming from industrial and urban area. It can be seen that the concentration of OX is made up of a NO _x -independent ‘regional’ contribution (i.e. the O₃ background), and a linearly NO _x -dependent ‘local’ contribution from primary emissions, such as traffic. The local emission is very low in this area. Also, the regional contribution is similar to unpolluted sites and presents seasonal variation, being higher in May. However, our measurements showed that the proportion of OX in the form of NO₂ increases with the increase in NO _x concentration during the day. The higher proportion of NO₂ observed at night must be due to the conversion of NO to NO₂ by the NO?+?O₃ reaction. With regards to the source of the local NO _x -dependent contribution, it may be attributed to industrial emission, or the termolecular reaction 2NO?+?O₂?=?2NO₂, at high-NO _x levels and stagnant air during several days. Finally, we estimated the photolysis rate of NO₂, J _NO2, an important key atmospheric reaction coupled with ozone. We also present surface plots of annual variation of the daily mean NO _x and OX levels, which indicate that oxidants come from transport processes instead of local emissions associated as local photochemistry.