Abstract

1. Locke, A., and N.G. MacNair Gulf Fisheries Centre, Fisheries and Oceans Canada, P.O. Box 5030, Moncton, New Brunswick PEI Department of Fisheries and Aquaculture, Charlottetown, Prince Edward Island Abstract Since about the year 2000, invasive tunicates (Styela clava, Botrylloides violaceus, Ciona intestinalis and to a lesser extent Botryllus schlosseri) have become major pests on blue mussel (Mytilus edulis) aquaculture sites in PEI estuaries. Large biomasses of tunicates create problems for handling and harvesting of the mussels, so growers in heavily infested estuaries sometimes use dilute (5%) acetic acid (household vinegar) or suspension solutions of calcium hydroxide (hydrated lime) to kill tunicates. The extent to which these solutions may affect other biota in the estuaries has recently been questioned. We report here our preliminary investigation. Footprint and Effect of Treatment Agents on Non-target Marine Organismsassociated with Tunicate-Infested Mussel Socks in PEI Estuaries Acetic Acid Background Acetic acid is a weak organic acid, which, diluted to 4-5%, is ordinary household vinegar. Acetic acid is registered as a pesticide in Canada with the Pesticide Management Regulatory Agency. It has previously been used as an organic herbicide in terrestrial applications. It was first tested as a treatment against tunicates in Nova Scotia (Carver et al. 2003). Immersion in acetic acid, or application of acetic acid inside wrapped structures, are considered to be promising methods for eradication of invasive tunicates in New Zealand (Coutts and Forrest 2005; Forrest et al. 2007). How it is used in aquaculture in PEI In laboratory studies (e.g., MacNair unpub. data; LeBlanc et al. 2007), immersion in 5% acetic acid for <1 minute kills tunicates very effectively. For use in aquaculture, immersion is considered impractical because of the rapid dilution of the acid by water from the immersed mussel socks. Sprayers have been developed for experimental applications of acetic acid to mussel socks in PEI (Fig. 1). So far, no spray application methods have been as effective against tunicates as the immersion trials. With both immersion and spray, there is some incidental mortality of mussels. If the mussels are shaken so they close their valves before exposure to acetic acid, mortality/loss of attachment is in the range of 10-20% (LeBlanc et al. 2007). Toxic action The most obvious effect of acetic acid is that it may lower the pH of the surrounding water, i.e., make it more acid, however we predict that only a very small area adjacent to the sock is being affected. The pH of 5% acetic acid is 2.6. Chemical reactions that occur when acetic acid is added to salt water tend to buffer the pH, i.e., maintain it closer to the natural pH of sea water which is also the preferred pH of marine organisms, around 8.2. This reaction can take place rapidly when acetic acid is diluted in salt water. The efficacy of acetic acid as a biocide is apparently due to factors other than just pH. Previous work has shown it to be more effective than other acids adjusted to the same pH (e.g., Forrest et al. 2007). We also found that acetic acid was more effective than citric acid when used as an immersion treatment adjusted to similar pH (Fig. 2). One breakdown product of acetic acid in sea water is sodium acetate. This chemical, at pH<6.5, acts to uncouple cellular communication by disrupting the gap junction channels between cells (Germain and Anctil 1996). Any toxic action of sodium acetate would be limited to a small footprint and short time-frame around the treatment site, given the high buffering capacity of sea water. Lime Background Heating limestone (calcium carbonate, CaCO3) to drive off the carbon dioxide produces quicklime (CaO); adding water to quicklime in turn produces hydrated lime (calcium hydroxide, Ca(OH)2). Quicklime has a long history of use in bivalve culture in Long Island Sound as a treatment to clear oyster beds of starfish (MacKenzie 1977) and in California to eliminate sea urchins from kelp beds (Bernstein and Welsford 1982). How it is used in aquaculture in PEI Hydrated lime has been used as an immersion treatment against starfish on mussel socks for years in PEI. The method has been adapted for tunicate control (Fig. 3). A typical mortality of mussels following treatment is 10-15% (MacNair, unpub. data). Toxic action Most studies have addressed the toxic action of quicklime rather than hydrated lime. Quicklime, a very alkaline substance, burns the surface of echinoderms, producing lesions which over time enlarge and allow bacterial invasion which eventually kills the animal (Loosanoff and Engle 1942). Presumably the method by which hydrated lime kills tunicates is similar. The pH of a supersaturated solution of hydrated lime is 12-12.5. However, lime exposed to air or water rapidly combines with carbon dioxide to form calcium carbonate. This reaction may be completed within 24 hr (some sources indicate up to 5 days for completion of the reaction, but this probably depends on the rate of dilution and mixing of the material with water). The precipitated calcium carbonate sinks to the bottom. Initial observations have shown the pH of the water adjacent to treated mussel socks to rapidly return to normal. As well, underwater observations showed marine organisms adjacent to a treatment location to be unaffected by the precipitating lime as it sinks in the water column. Applications of quicklime to oyster beds for predator control (at rates up to 6.75 tonnes/ha) have apparently resulted in no detrimental effects on oyster spat, other bivalves (mussels, clams, scallops), or mature lobsters, crabs or fish in the area (McKenzie 1977; Shumway et al. 1988). We are not aware of any previous effects studies with hydrated lime. Fig. 3. A bag of hydrated lime is added to a trough of sea water, then the mussel sock is dragged through the trough (Fig. 3) Fig. 1. This sprayer applies 5% acetic acid to all sides of the mussel sock. Literature cited Bernstein and Welsford. 1982. Can. Tech. Rep. Fish. Aquat. Sci. 968. Carver et al. 2003. J. Shellfish Res. 22: 621-631. Coutts and Forrest. 2005. Cawthron Rep. 1110. Forrest et al. 2007. Aquaculture 262: 319-332. Germain and Anctil 1996. Biol. Bull. 191: 353-366. LeBlanc et al. 2007. Aquaculture 264: 205-213. Loosanoff and Engle. 1942. Use of lime in controlling starfish. Res. Rep. 2, US Fish and Wildlife Serv., p. 1-29. McKenzie 1977. Aquaculture 10: 45-51. Shumway et al. 1988. Bull. Environ. Contam. Toxicol. 40: 503-509. Fig. 2. The results of an experiment comparing mortality of C. intestinalis exposed for 5 and 10 sec intervals to acetic (A5 and A10) and citric (C5 and C10) acids at pH 2.6-2.8, and control (ctrl). • Next steps • Calculation of present and potential rates of application in selected estuaries • Determination of mass balance calculations – to evaluate the rate of neutralization of hydrogen ion imbalances • Determine the “pH footprint” around treatment sites • Bioassays of effects on bacteria (Microtox test), sand shrimp (Crangon septemspinosa), three-spined stickleback (Gasterosteus aculeatus) to be carried out by Dr. K. Doe, Environment Canada, Moncton. Acknowledgements We thank P. Yeats for explaining the chemistry, W. Fairchild for discussions of toxicity, T. Gallant for contributions to literature review, and R. Bernier, A. Morrison, E. Reese and K. Swan for their assistance with experiments. This work in progress has been supported by DFO’s AIS Research Funds and by PEI-DFA.