Pre-validation of choriogenin H transgenic medaka eleutheroembryos as a quantitative estrogenic activity test method.

The choriogenin H - EGFP transgenic medaka (Oryzias melastigma) has been used to test estrogenic substances and quantify estrogenic activity into 17ß-estradiol (E2) equivalency (EEQ). The method uses 8 eleutheroembryos in 2 ml solution per well and 3 wells per treatment in 24-well plates at 26 ± 1 °C for 24 ± 2 h, with subsequent measurements of induced GFP signal intensity. EEQ measurements are calculated using a E2 probit regression model with a coefficient of determination (R2) > 0.90. The selectivity was confirmed evaluating 27 known estrogenic and 5 known non-estrogenic compounds. Limit of quantitation (LOQ), recovery rate and bias were calculated to be 1 ng/ml EEQ, 104% and 4% respectively. Robustness analysis revealed exposure temperature is a sensitive parameter that should be kept at 26 ± 1 °C. The repeatability of intra- and inter-laboratories achieved CV < 30% for most tested food and cosmetics samples. The lot-lot stability was confirmed by the stable EEQ qualitative control (QC, 1 ng/mL E2) and calibration curve results. The stability of standard reagents, samples and sample extracts was also investigated. These data demonstrated this method to be an accurate indicator of estrogenic activity for both chemicals and extracts.

The potential of foodwaste leachate as a phycoremediation substrate for microalgal CO2 fixation and biodiesel production.

Foodwaste leachate (FWL) is often generated during foodwaste treatment processes. Owing to its high nutrient content, FWL has high potential for phycoremediation, a microalgal technology application for water treatment while acting as CO2 fixation tank. Additionally, the end product of microalgal from phycoremediation can be potentially used for biodiesel production. Therefore, the phycoremediation has drawn a lot of attention in recent decades. This study evaluates the performance of microalgal foodwaste leachate treatment and the potential of utilizing FWL as medium for microalgal biodiesel production. Two microalgal species, Dunaliella tertiolecta and Cyanobacterium aponinum, were selected. For each species, two experimental levels of diluted FWL were used: 5 and 10% FWL. The partial inhibition growth model indicates that some inhibit factors such as ammonia; total suspended solids and oil and grease (O&G) content suppress the microalgal growth. Most of the nutrient such as nitrogen and phosphorus (> 80%) can be removed in the last day of phycoremediation by D. tertiolecta. C. aponinum also show considerable removal rate on total nitrogen ammonia and nitrate (> 60%). Biomass (0.4-0.5 g/L/day) of D. tertiolecta and C. aponinum can be produced though cultivated in diluted FWL. The bio-CO2 fixation rates of the two species were 610.7 and 578.3 mg/L/day of D. tertiolecta and C. aponinum. The strains contain high content of saturated fatty acid such as C16 and C18 making them having potential for producing good quality biodiesel.

Prioritising local action for water quality improvement using citizen science; a study across three major metropolitan areas of China.

Streams in urban areas are prone to degradation. While urbanization-induced poor water quality is a widely observed and well documented phenomenon, the mechanism to pinpoint local drivers of urban stream degradation, and their relative influence on water quality, is still lacking. Utilizing data from the citizen science project FreshWater Watch, we use a machine learning approach to identify key indicators, potential drivers, and potential controls to water quality across the metropolitan areas of Shanghai, Guangzhou and Hong Kong. Partial dependencies were examined to establish the direction of relationships between predictors and water quality. A random forest classification model indicated that predictors of stream water colour (drivers related to artificial land coverage and agricultural land use coverage) and potential controls related to the presence of bankside vegetation were found to be important in identifying basins with degraded water quality conditions, based on individual measurements of turbidity and nutrient (N-NO3 and P-PO4) concentrations.

The Mechanism of Diarrhetic Shellfish Poisoning Toxin Production in Prorocentrum spp.: Physiological and Molecular Perspectives.

Diarrhetic shellfish poisoning (DSP) is a gastrointestinal disorder caused by the consumption of seafood contaminated with okadaic acid (OA) and dinophysistoxins (DTXs). OA and DTXs are potent inhibitors of protein phosphatases 2A, 1B, and 2B, which may promote cancer in the human digestive system. Their expression in dinoflagellates is strongly affected by nutritional and environmental factors. Studies have indicated that the level of these biotoxins is inversely associated with the growth of dinoflagellates at low concentrations of nitrogen or phosphorus, or at extreme temperature. However, the presence of leucine or glycerophosphate enhances both growth and cellular toxin level. Moreover, the presence of ammonia and incubation in continuous darkness do not favor the toxin production. Currently, studies on the mechanism of this biotoxin production are scant. Full genome sequencing of dinoflagellates is challenging because of the massive genomic size; however, current advanced molecular and omics technologies may provide valuable insight into the biotoxin production mechanism and novel research perspectives on microalgae. This review presents a comprehensive analysis on the effects of various nutritional and physical factors on the OA and DTX production in the DSP toxin-producing Prorocentrum spp. Moreover, the applications of the current molecular technologies in the study on the mechanism of DSP toxin production are discussed.

Quantification of phytoplankton bloom dynamics by citizen scientists in urban and peri-urban environments.

Freshwater ecosystems are severely threatened by urban development and agricultural intensification. Increased occurrence of algal blooms is a main issue, and the identification of local dynamics and drivers is hampered by a lack of field data. In this study, data from 13 cities (250 water bodies) were used to examine the capacity of trained community members to assess elevated phytoplankton densities in urban and peri-urban freshwater ecosystems. Coincident nutrient concentrations and land use observations were used to examine possible drivers of algal blooms. Measurements made by participants showed a good relationship to standard laboratory measurements of phytoplankton density, in particular in pond and lake ecosystems. Links between high phytoplankton density and nutrients (mainly phosphate) were observed. Microscale observations of pollution sources and catchment scale estimates of land cover both influenced the occurrence of algal blooms. The acquisition of environmental data by committed and trained community members represents a major opportunity to support agency monitoring programmes and to complement field campaigns in the study of catchment dynamics.

Toxicity and estrogenic endocrine disrupting activity of phthalates and their mixtures.

Phthalates, widely used in flexible plastics and consumer products, have become ubiquitous contaminants worldwide. This study evaluated the acute toxicity and estrogenic endocrine disrupting activity of butyl benzyl phthalate (BBP), di(n-butyl) phthalate (DBP), bis(2-ethylhexyl) phthalate (DEHP), diisodecyl phthalate (DIDP), diisononyl phthalate (DINP), di-n-octyl phthalate (DNOP) and their mixtures. Using a 72 h zebrafish embryo toxicity test, the LC50 values of BBP, DBP and a mixture of the six phthalates were found to be 0.72, 0.63 and 0.50 ppm, respectively. The other four phthalates did not cause more than 50% exposed embryo mortality even at their highest soluble concentrations. The typical toxicity symptoms caused by phthalates were death, tail curvature, necrosis, cardio edema and no touch response. Using an estrogen-responsive ChgH-EGFP transgenic medaka (Oryzias melastigma) eleutheroembryos based 24 h test, BBP demonstrated estrogenic activity, DBP, DEHP, DINP and the mixture of the six phthalates exhibited enhanced-estrogenic activity and DIDP and DNOP showed no enhanced- or anti-estrogenic activity. These findings highlighted the developmental toxicity of BBP and DBP, and the estrogenic endocrine disrupting activity of BBP, DBP, DEHP and DINP on intact organisms, indicating that the widespread use of these phthalates may cause potential health risks to human beings.

Effective growth of dinoflagellate Prorocentrum minimum by cultivating the cells using municipal wastewater as nutrient source.

Several studies have been conducted worldwide in order to develop a more economical method for mass algal cultivation so that more cost-effective biomass production can be accessed. One of the directions is to reduce production costs by using wastewater as a nutrient source in algal cell cultivation. The growth ability of a red-tide causative dinoflagellate species, Prorocentrum minimum, in various concentrations of local urban wastewater was examined in this study. The highest exponential growth rate and maximum cell density (MCD) were achieved when autoclaved 10% wastewater was used for cell cultivation, although the cells could survive in 0.01-100% wastewater. Both growth rate and MCD of the cells in wastewater were found to be substantially higher than that in optimized L1 culture medium. This research highlights the potential of using wastewater as a cost-effective approach for mass cultivation of dinoflagellate cells with consequent production of valuable microalgal biomass.

Effects of different nitrate and phosphate concentrations on the growth and toxin production of an Alexandrium tamarense strain collected from Drake Passage.

Nitrate (N) and phosphate (P) are believed to be two of the most important nutrients for the growth and toxin production for Alexandrium species. The study of the growth and toxicity characteristics of the Alexandrium spp. under the change of N and P can help us to understand the dynamics of algal bloom and toxification events in natural environments. A strain of Alexandrium tamarense (designated as Kci) was successfully isolated from the Drake Passage in 2001 and the clonal culture has been kept in our laboratory (Ho et al., 2003, 2012). In order to extend our understanding on the growth physiology and toxicity of this A. tamarense strain, growth and cellular toxin content were examined in unialgal batch cultures under different concentrations of N and P. The effects of variable N, P concentrations on growth, cellular toxicity (fg STXeq. cell(-1)), and toxin composition (% molar) were determined in both exponential and stationary growth phases. The toxin profile, determined by high-performance liquid chromatography with fluorescence detection (HPLC-FD), was found to be remained relatively stable and was consistently dominated by the N-sulfocarbamoyl C-toxins (>90%) under different conditions and growth phases. There were also trace amounts of other carbamate gonyautoxins consistently expressed. The cellular toxicity varied under different N and P concentrations, as well as different growth stages. A positive correlation was observed between cellular toxicity and N concentrations, but the toxicity was enhanced when P was depleted. Both cell densities and growth rate of the cells were severely suppressed under N- or P-depletion. However, the biovolume of the cells tended to be larger at N- or P-depleted cultures. Results from the present study provide valuable insight for the ecophysiology of Alexandrium species in the coastal ecosystem of Drake Passage.

Development of a relative risk model for evaluating ecological risk of water environment in the Haihe River Basin estuary area.

Ecological risk assessment for water environment is significant to water resource management of basin. Effective environmental management and systems restoration such as the Haihe River Basin require holistic understanding of the relative importance of various stressor-related impacts throughout the basin. As an effective technical tool for evaluating the ecological risk, relative risk model (RRM) was applied in regional scale successfully. In this study, the risk transfer from upstream of basin was considered and the RRM was developed through introducing the source-stressor-habitat exposure filter (SSH), the endpoint-habitat exposure filter (EH) and the stressor-endpoint effect filter (SE) to reflect the meaning of exposure and effect more explicit. Water environment which includes water quality, water quantity and aquatic ecosystems was selected as the assessment endpoints. We created a conceptual model which depicting potential and effect pathways from source to stressor to habitat to endpoint. The Haihe River Basin estuary (HRBE) was selected as the model case. The results showed that there were two low risk regions, one medium risk region and two high risk regions in the HRBE. The results also indicated that urbanization was the biggest source, the second was shipping and the third was industry, their risk scores are 5.65, 4.71 and 3.68 respectively. Furthermore, habitat destruction was the largest stressor with the risk scores (2.66), the second was oxygen consuming organic pollutants (1.75) and the third was pathogens (1.75). So these three stressors were the main influencing factors of the ecological pressure in the study area. For habitats, open waters (9.59) and intertidal mudflat were enduring the bigger pressure and should be taken considerable attention. Ecological service values damaged (30.54) and biodiversity decreased were facing the biggest risk pressure.

Isolation and identification of a new tetrodotoxin-producing bacterial species, Raoultella terrigena, from Hong Kong marine puffer fish Takifugu niphobles.

Puffer fish, Takifugu niphobles, collected from the Hong Kong coastal waters were screened for tetrodotoxin-producing bacteria. A Gram-negative, non-acid-fast, non-sporing and rod shaped bacterial strain (designated as gutB01) was isolated from the intestine of the puffer fish and was shown to produce tetrodotoxin (TTX). Based on the Microbial Identification (MIDI) and 16S-23S rDNA internal transcribed spacer (ITS) phylogenetic analysis, the strain was identified as Raoultella terrigena. The TTX production ability of the strain was confirmed by mouse bioassay, ELISA and mass spectrometry (MALDI-TOF). Our results reiterate that the TTX found in puffer fish was likely produced by the associated bacteria and TTX are widely produced amongst a diversity of bacterial species.