Project: PS4CTX - A novel approach for monitoring, toxicity evaluation and risk assessment of cyanobacterial toxins – a use of passive samplers
Person in Charge: doc. Ing. Blahoslav Maršálek, CSc.
Host institution: Institute of Botany AS CR
Country of Origin: Czech Republic
Country of scientific activity: USA
Project duration: 24 months
Scientific panel: Environment and Geosciences
The project aims to combine the great expertise and unique position of the host institution in the field of cyanotoxins with an excellent experience of the reintegrating researcher in the same research area, and with the extensive knowledge in tumor promotion and epigenetic toxicity acquired by the reintegrating researcher during his post-doctoral stay at Michigan State University, in order to address the issue of cyanotoxin contamination in South Moravian region (SMR) using innovative interdisciplinary approaches combining environmental chemistry, risk analysis, in vitro toxicology and most advanced and the-state-of-the-art methods. Production of tumor promoting microcystins (MCs) by cyanobacterial water blooms has been recognized as an important health problem in the Czech Republic, since MCs can be found in about 70% water bodies, including drinking reservoirs in SMR. Little is known about MC contamination in drinking waters, hence the project aims to use a novel and world-unique approach based on passive sampling for monitoring of MCs in drinking water and treatment plants; and use the collected data for analysis of risks associated with MC-contaminated drinking water. The next aim is to develop and construct a quite new and original passive sampler for cylindrospermopsin, an emerging cyanotoxin whose detection frequency is increasing worldwide, including the Czech Republic. Cyanobacteria produce a variety of bioactive chemicals other than well-known cyanotoxins, and some of them might induce tumor promotion activity. Thus, evaluation of cyanobacterial samples from drinking water reservoirs using relevant in vitro liver tumor promotion models will provide an important data for identification of new hazards represented by toxic cyanobacteria. The funding will facilitate reintegration of the researcher and support the transfer of advanced knowledge to the host, which will contribute to both scientific excellence at SMR and professional growth of the researcher.
The ongoing project summary
Project focuses on the issue of mass proliferations of toxic cyanobacteria, contamination of drinking water reservoirs with cyanobacterial toxins (cyanotoxins) and associated human health risks.
Project aims were following:
1) provide information on contamination of drinking water reservoirs and drinking waters with cyanotoxins microcystins and on cyanotoxin human health risks with the use of traditional sampling and monitoring approaches as well as with the use of innovative methods based on passive sampling;
2) develop and optimize passive sampler for sequestration of another hazardous cyanotoxin cylindrospermopsin;
3) use in vitro methods for studies of tumor promotional and carcinogenic effects of cyanobacteria and cyanotoxins.
Within the first specific aim, field measurements, sample collection (water, phytoplankton) and exposures of passive samplers were performed at selected drinking water reservoirs and adjacent treatment plants during June-November 2011, upon numerous laboratory optimizations of sampler design and sampling procedures. After the end of field sampling period, the collected samples and data are being processed and analyzed.
For the needs of the second project aim, cyanobacterial biomass for cylindrospermopsin isolation was prepared by laboratory cultivation, cylindrospermopsin extraction was optimized, as well as HPLC method for cylindrospermopsin analysis was optimized and HPLC calibrated. In addition to originally planned activities, methods potentially applicable for cyanotoxin removal in drinking water production were also investigated. In relation to the third specific aim, cell culture methods and techniques were introduced in the laboratories at the host institution, cell lines WB-F344, HL1-hT1 and HaCaT were obtained, methods for evaluation of gap junctional intercellular communication (GJIC), cell proliferation and cytotoxicity were also introduced. These cell lines and methods were used not only for the planned research of tumor promotional effects of cyanobacteria, but also for investigation of dermatotoxicity of cyanobacteria.
Preliminary data from traditional sampling at reservoirs and drinking water treatment plants under the study in the year 2011 show that microcystins were commonly detected in surface layers of drinking water reservoirs , often at concentrations exceeding limit 1 ug/L defined by National Drinking Water Decree No. 252/2004 Coll. However, samples of intake water at drinking water treatment plants which originated from deeper layers of reservoirs with lower concentrations of cyanobacteria contained microcystins less often and concentrations above 1 ug/L were detected only exceptionally. Concentrations of microcystins in final treated water were below 1 ug/L throughout the sampling period and thus did not pose human health risks for consumers. Amounts of microcystins accumulated in exposed passive samplers well-corresponded to seasonal changes of microcystin concentrations obtained from traditional sampling and also to the observed trends in microcystin concentrations during drinking water treatment process. These results will be published at conference SETAC 2012 (Berlin, 20-24.5.2012) and 64th meeting of Association of Czech and Slovak chemical societies (Olomouc, 25-27.6.2012).
Several grams (dry weight) of biomass of Aphanizomenon flos-aquae PCC7905 containing about 5-10 mg of cylindrospermopsin were produced by laboratory cultivation and will be used in the next phase of project. For realization of experiments with cylindrospermopsin passive sampling, isolation of cylindrospermopsin from cyanobacterial biomass was optimized and analytical method was optimized. Based on literature review, sorbents for further development of cylindrospermopsin passive sampler were selected. The introduced in vitro cell culture methods were used for research of mixture toxicity of compounds whom human population might be exposed to. It was found that cyanobacterial extracts significantly potentiated tumor promotional effects of common anthropogenic environmental contaminants and the results were accepted for publication in Chemosphere journal. The next study focused on elimination of toxicity of cyanobacterial extracts and cyanotoxins by processes relevant for drinking water treatment and demonstrated that ozonation rapidly and effectively removes not only microcystins but also tumor promotional and cytotoxic effects of cyanobacterial extratcs, whereas chlorination was much less effective.
The results were presented at conference on Toxicity and biodegradability of chemicals (2011), in Bulletin of VURH and are being prepared for publication in peer-reviewed journal.The next study realized with the support of the project investigated dermatotoxic effects of cyanobacteria with the use of in vitro models. The first results showed that the most toxic effects were elicited by hydrophobic fraction extracted from cyanobacterial biomass, whereas toxicity of commonly studied hydrophilic extracts was lower, which indicates an existence of novel not-yet-identified cyanobacterial dermatotoxins.
The main expected outcomes and deliverables of the project will be not only the development and introduction of methods valuable for future research but in the first place publications of experimental results and data at national and international scientific conferences and in scientific journals. These publications will present new information on contamination of drinking water reservoirs and drinking waters with cyanobacterial toxins, case studies of assessment of human health risks associated with the occurrence of microcystins , introduction of novel approaches and tools applicable for scientific and regulatory monitoring of cyanobacterial toxins. The findings on toxicity of cyanobacteria and further health hazard characterization and information useful for minimizing health risks will be also presented. Project will thus significantly contribute to the effort for protection of human health from adverse effects of cyanotoxins.