Cylindrospermopsin in whole cell extracts and live cultures of Cylindrospermopsis raciborskii: Ecotoxicity, bioaccumulation and management

Cylindrospermopsin (CYN) is an alkaloid toxin produced by at least six bluegreen algal genera. In Australia, the most common producer organism is Cylindrospermopsis raciborskii. This research determined the toxic effects and potential for toxin bioaccumulation associated with toxin-producing C. raciborskii, using environmentally relevant exposure scenarios. Short-term semi-static renewal ecotoxicity tests were conducted on four freshwater test organisms, using freeze-thawed C. raciborskii whole cell extracts or live cultures of C. raciborskii containing CYN. Duckweed (Spirodela oligorrhiza) demonstrated variable responses to whole cellextract exposures containing 0 – 500 μg L-1 of CYN. Growth stimulation and growth inhibition were recorded from the duckweed, depending on toxin exposure concentrations and length of the incubation period. Chlorophyll content of Spirodela was also affected by toxin exposure, with small peaks in chlorophyll a recorded in conjunction with exposure to 8 μg L-1 CYN. Changes to frond morphology (chlorosis, necrosis) and reduced root lengths were also noted, but these effects were not consistent throughout all trials. Bioconcentration of free CYN was not detected in the tissues of Spirodela; small quantities of toxin recovered from the tissues probably represented toxin sorption to the cell walls. Growth of water thyme (Hydrilla verticillata) was stimulated by exposure to whole cell extracts of C. raciborskii containing a maximum of 400 μg L-1 CYN. Exposure to the test solutions appeared to promote the redistribution of plant resources in H. verticillata, and the possible benefits of this, particularly with respect to increased root production, were considered. Effects on the chlorophyll content of Hydrilla were variable and included decreases in total chlorophyll content and changes to the chlorophyll a:b ratio. Bioconcentration of free toxin was not detected in the tissues of Hydrilla. In trials with Melanoides tuberculata, exposure to whole cell extracts or live algal cultures did not significantly affect the behaviour or relative growth rates of adult snails. However, changes in the number of hatchlings released from parent snails were recorded: exposure to whole cell extracts corresponded with increased numbers of hatchlings compared with controls, whereas decreases in hatchling numbers compared with controls were recorded from treatments containing live C. raciborskii. Both bioconcentration and bioaccumulation occurred in the soft tissues of snails, although exposure to whole cell extracts resulted in only minor tissue contamination compared with that from live C. raciborskii exposures. Bioaccumulation of the analog deoxy-CYN was recorded in the soft tissues. M. tuberculata did not bioconcentrate CYN in the shell. Tadpoles of the cane toad (Bufo marinus) featured the most dramatic responses to the test solutions. Exposure to live C. raciborskii cultures resulted in up to 66% mortality of B. marinus, whereas all tadpoles survived exposure to whole cell extracts of comparable CYN concentrations. Decreases in the time spent swimming and relative growth rates were recorded from surviving tadpoles during both types of exposure regimes. Histological examination of Bufo tissues revealed tissue injuries to multiple organs, with particular severity noted in the liver, intestine, nephric ducts and gill epithelia. The extent of cellular damage was similar in whole cell extract (containing a maximum of 400 μg L-1 CYN) and the live culture trials (containing a maximum of 232 μg L-1), despite the unequal toxin concentrations. Bioconcentration of CYN was not evident during the whole cell extract trial, whereas exposure to live cultures resulted in maximum average tissue concentrations of 895 μg toxin kg-1 fresh weight. A secondary aim of the thesis was to identify and address gaps in management approaches for toxin bioaccumulation and possible environmental effects associated with toxin-producing C. raciborskii blooms. A predictive management strategy was developed to determine the likelihood of tissue contamination in aquatic organisms inhabiting water bodies affected by blooms. The ten-step framework for predicting bioaccumulation risk was based upon characteristics of cyanotoxin bioavailability, exposure and uptake routes during the progression of a toxic bloom. Key concepts included monitoring changes in toxin availability throughout the progression of a toxic bloom, and the prediction of bioaccumulation risks based on threshold toxin values for selected aquatic organisms. These threshold values were re-examined following the completion of the laboratory trials. Current approaches for the management of blooms with respect to environmental risks were discussed, and procedures for the proper evaluation of environmental risks associated with Cylindrospermopsis blooms were assessed. These included adequate detection and monitoring systems, setting of environmental guideline values, and options for the control and remediation of toxic blooms.