Toxicity testing in Japan and the use of Toxkit microbiotests

Management of toxic chemicals in the aquatic environment is performed in Japan by setting environmental standards by law. Forty-nine chemicals are regulated and measured once a month at specific sites in rivers and lakes. Many of these chemicals are detected at various places in Japan, but in concentrations below the standard values. Dioxins, which are not regulated, have been detected in leachates from a landfill area of industrial wastes and also in human mother milk. In Japan, bioassays are not used to date as a monitoring tool. However, a manual of ecotoxicological test methods for chemicals is presently under examination by the Ministry of international Trade Industry, the Ministry of Agriculture, Forestry and Fisheries and the Environmental Agency. In most cases, the methods proposed are in accordance with OECD Guidelines for the testing of chemicals; they comprise algal, Daphnia and fish tests. Many toxicity test methods have already been described in Japanese scientific literature, based on various types of test organisms from different phylogenetic groups such as e.g. bacteria, yeasts, protozoans, micro- and macro-algae, crustaceans, molluscs, insects, amphibians, fish and birds. Taking into account the difficulties associated with the culturing and/or maintenance of live stocks, the "culture free" Toxkit microbiotest technology appeared appealing to the authors and is now used for environmental monitoring purposes in their laboratory. A new toxicity testing method using mobility analysis of Daphnias obtained from ephippia has recently been developed and will be reported. The test criterion is the change in mobility of Daphnia neonates, which is measured by computer analysis of videocamera-pictures (0.013 sec) during 5 seconds. The mobility per time unit is calculated subsequently. Toxicity is evaluated by comparing the change in mobility of Daphnias exposed to chemicals, versus that in the controls. The technology allows to detect the increase in mobility by stimulus at low concentration and the decrease at higher concentrations of toxicants. The method has a high sensitivity and allows for toxicity detection in less than 3 hours. Results with the new mobility analysis method with Daphnia neonates will be compared with those obtained with the conventional Daphnia method prescribed by the OECD.