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Protocol no. 45
THE DUNALIELLA TERTIOLECTA TEST A MARINE ALGAL ASSAY PROCEDURE

This test examines the effect of contaminated sea-water on the growth of the marine alga, Dunaliella tertiolecta. Inhibition of growth provides an indication of likely toxicity.

CONTACT

Dr Mario Bucci and Dr Giancarlo Sbrilli
Unita' Sanitaria Locale N.25
Servizio Multizonale di Prevenzione S.S. No 398
Loc. Montegemoli I-57025 Piombino (LI) Italy
Tel: 0565 67807 or 67810 Fax: 0565 276472

RATIONALE

Ecotoxicology has been defined as the study of harmful effects of chemicals upon ecosystems. In practice, much ecotoxicological work is done to establish effects on individuals, although the ultimate concern of the ecotoxicologist is to establish the consequences of these effects at the levels of populations and whole ecosystems. Ecotoxicology may also differ from classical toxicology in a number of important aspects. In the first place, the entry of pollutants into ecosystems, and their subsequent movement, bioaccumulations, binding and degradation, involve processes not normally encountered in toxicology. Thus the movement of pollutants through ecosystems depends upon such factors as the feeding ecology, behaviour, migration, detoxifying systems, and sensitivities to toxic action of a wide variety of species. There is also the question of the movement and distribution of pollutants in air, aquatic systems and in soil. If the aquatic environment is considered, there is a considerable problem with saline industrial effluent affecting aquatic systems. Several different species might be considered as test organisms to monitor the effects of such effluent. In this test, the marine unicellular green alga Dunaliella tertiolecta was chosen for study for several reasons, such as ease of culture, availability, and use by the Italian National Institute for Water Research (I.R.S.A.-C.N.R.).

BASIC PROCEDURE

The marine alga, Dunaliella tertiolecta, is grown in samples of saline industrial effluents or in samples of sea-water taken from areas contaminated by industrial waste, environmental pollutants, etc.. At the end of the experimental period the number of alga is assessed using an automatic particles counter. Any inhibition of algal growth is used as an indication of toxicity.

CRITICAL ASSESSMENT

The total biological effect of saline wastewater may be evaluated. Algal growth may either be stimulated or inhibited by effluent. In the authors opinion, the low cost and simplicity of this assay makes its ideal for obtaining quantitative data on the biological effects of effluent. Wastewater samples from chemical, iron and steel industries, and from an oil-fired power plant have been submitted to Dunaliella tertiolecta, MicrotoxR and Poecilia reticulata toxicity test. Experimental data indicates that algal tests show greater sensibility than MicrotoxR and the fish test. It therefore appears to be useful in assessing acute toxicity of diluted saline wastewaters.

TEST STATUS

Undergoing in-house development. At present only a few samples have been passed through the test. The results from this assay are being compared with other in vitro tests such as the Luminescent Bacterial test, MicrotoxR, and the fish test, Poecilia reticulata.

CHEMICALS TESTED

The samples of sea-water tested have come from effluent obtained from industrial plants on the coast of the Mediterranean. They include effluent form the chemical, iron and steel industries, as well as an oil-fired power plant. Also, a phenol and cyanide solution made up in filter-sterilized natural sea-water.

REFERENCES

  1. APHA-AWWA-WPCF (1980) Standard methods for the examination of water and wastewater. 15th Edition. Washington, DC, American Public Health Association. Chiaudani, G. & Vighi. M. (1978) Metodologia standard di saggio algale per lo studio della contaminazione delle acque marine. Quaderni dell'Istituto di Recerca Sulle Acque, C.N.R., n. 39. Elnabarawy, M.T. & Welter, A.N. (1984) Utilization of algal cultures and assays by industry. In: Algae as ecological indicators. (ed. L. Elliot Schubert) Academic Press, pp. 317-328.
  2. EPA, Environmental Protection Agency (1974) Marine algal assay procedure bottle test. Corvallis, Oregon 97330, pp.43.
  3. EPA, Environmental Protection Agency (1978) The Selenastrum capricornutum Printz algal assay bottle test. Experimental design, application and data interpretation protocol. EPA-600/9-78-018. Corvallis Environmental Research Laboratory, Office of Research and Development, Corvallis, Oregon. EPA, Environmental Protection Agency (1982) Second US/USSR Symposium: Biological aspects of pollutant effects on marine organisms. EPA-600/3-82-034, pp. 112-122.
  4. Finney, D.J. (1971) Probit analysis. Cambridge University Press, Cambridge, pp. 333. Joubert, G. (1983) Detailed method for quantitative toxicity measurements using the green algae Selenastrum capricornutum. In: Aquatic Toxicology - advance in environmental science and technology. (ed. Nriagu, J. O.) John Wiley & Sons, Vol. 13, pp. 467-485.
  5. Puddu, A. (1989) Programma di calcolo per l'elaborazione dei risultati di un saggio di tossieita mediante analisi dei probits. Metodi analitici per le acque, Notiziario C.N.R.- Istituto di Ricerca sulle Acque, ISSN:0392-1425, 2:19-37.
  6. Sbrilli, G., Bucci, M. & Luti, R. (1990) Saggio algale di tossieita. Utilizzazione di Dunaliella tertiolecta nella valutazione degli effetti biologici di scarichi industriali a prevalente componente marina. Acqua-Aria. 6, 483-489.
  7. Trevors, J.T. (1986) A basic program for estimating LD50 values using the IBM-PC. Bull. Environ. Contam. Toxicol. 37, 18-26. Van Coillie, R., Couture, P. & Visser, S.A. (1983) Use of algae in aquatic ecotoxicology. In: Aquatic Toxicology - Advances in Environmental Science and Technology. (ed. Nriagu, J.O.) John Wiley and Sons, Vol. 13, pp. 487-502.
  8. Walsh, G.E. & Alexander, S.V. (1980) A marine algal bioassay method: results with pesticides and industrial wastes. Wat. Air, Soil Pollut., 13, 45-55. Walsh, G.E., Bahner, L.H., & Horning, W.B. (1980) Toxicity of textile mill effluent to freshwater and estuarine algae, crustaceans and fishes. Environ. Pollut. Ser. A, 21, 169-179.
  9. Walsh, G.E. & Merril, R.G. (1984) Algal bioassays of industrial and energy process effluent. In: Algae as ecological indicators. (ed: L. Elliot Schubert) Academic Press, pp. 329-360.

IP-45 © February 1992