Cellular biomarkers of exposure to the fungicide copper oxychloride, in the common garden snail Helix aspersa, in Western Cape vineyards
Thesis (PhD(Agric)(Botany and Zoology))--University of Stellenbosch, 2001.
Copper oxychloride (Cu2CI(OH)3) is a broad-spectrum fungicide, intensively sprayed in many South African vineyards and orchards. It is necessary to find accurate and effective methods of monitoring the effects of this fungicide on the biota of the agricultural environment. The use of biomarkers may be a possible method to employ for this purpose. This study investigated the effects of copper, as a result of copper oxychloride exposure, on the biology of the common garden snail Helix aspersa, as welI as a number of cellular responses to exposure to the fungicide. The possible use ofthese responses as biomarkers was also investigated. Two groups of snails were exposed to 80 and 240 tJg g-! copper oxychloride respectively, for six weeks. A third group served as control. On a weekly basis, body mass, number of eggs produced, neutral red retention (NNR) times of haemocytic lysosomes, and body copper concentrations were determined for each individual. At the end of the experiment, the digestive glands, ovotestes and hermaphrodite ducts of a number of snails were prepared for histological analysis. The following parameters were investigated: tubule area, epithelium height and area in the digestive gland, spermatozoan area in the vesicula seminalis and ovotestis, as wen as oocyte numbers in the ovotestis. To test the validity of the laboratory results, a field survey was conducted. Snails were colIected from an uncontaminated vineyard and on two occasions from a contaminated vineyard in the Western Cape. The same cellular responses were investigated as in the laboratory study. The results showed that growth, egg production and hatching success in Helix aspersa were affected by experimental exposure to copper oxychloride. In both the laboratory study and field survey, copper in the body of H. aspersa was shown to be compartmentalized and the digestive gland was the most important site of copper accumulation. NNR times of haemocytic Iysosomes were shown to be affected by copper oxychloride exposure, already during the first week of exposure. A time evolution of copper accumulation and lysosomal damage existed. Epithelium height and area of digestive gland tubules, and spermatozoan and oocyte densities in the ovotestis, were also affected by copper oxychloride exposure and the concomitant copper burdens in the respective organs. Through the field survey it was ascertained that these histopathological changes were largely dependent on exposure time. It was concluded that lysosomal response of H. aspersa haemocytes, as measured by the NNR time assay, could be considered a useful biomarker of copper oxychloride exposure, since it provides an early warning of stress induced by this fungicide. Changes in digestive gland epithelium cells, and gametes in the ovotestis, can also possibly serve as biomarkers of copper oxychloride exposure. However, these can not serve as an early warning. All of the cellular responses identified in the present study can be used in combination with other cellular and physiological parameters and toxicological endpoints in order to improve the reliability and accuracy of interpretations regarding cause and effect.