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Worldwide transfer and introduction of non-indigenous species in ballast water causes significant environmental and economic impact. One way to address this problem is to remove or inactivate organisms that are found in ballast water. In this study, 3 experiments were conducted in Puget Sound, Washington, USA, using a prototype ozone treatment system installed on a commercial oil tanker, the S/T Tonsina. Treatment consisted of ozone gas diffused into a ballast tank for 5 and 10 h. Treatment and control tanks were sampled during the ozonation period for chemistry, cultur- able bacteria, phytoplankton and zooplankton. Selected fish and invertebrates were placed in cages deployed in the treatment and control tanks. Ozone introduced into seawater rapidly converts bro- mide (Br - ) to bromines (HOBr/OBr - ), compounds that are disinfectants. These were measured as total residual oxidant (TRO). Ozone treatment inactivated large portions of culturable bacteria, phyto- plankton and zooplankton. The highest reductions observed were 99.99% for the culturable bacteria, >99% for dinoflagellates and 96% for zooplankton. Caged animal results varied among taxa and locations in the ballast tank. Sheepshead minnows and mysid shrimp were most susceptible, shore crabs and amphipods the least. Distribution of ozone in the treatment tank was not homogenous during experiments, as suggested by the observed TRO concentrations and lower efficacies for inactivating the different taxa in selected ballast tank locations. Low concentrations of bromoform, a disinfection byproduct, were found in treated ballast water.