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Use of re-exposures to examine the effect of watercress derived isothiocyanates on Gammarus pulex

Use of re-exposures to examine the effect of watercress derived isothiocyanates on Gammarus pulex. Melanie Dixon. Why use re-exposures?. watercress farm & wash/pack house factory chalk stream headwater variable factory discharge ~8h per day weight/type products being washed.

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Use of re-exposures to examine the effect of watercress derived isothiocyanates on Gammarus pulex

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  1. Use of re-exposures to examine the effect of watercress derived isothiocyanates on Gammarus pulex Melanie Dixon

  2. Why use re-exposures? • watercress farm & wash/pack house factory • chalk stream headwater • variable factory discharge ~8h per day • weight/type products being washed Watercress farm outfall (Spring 08)

  3. Isothiocyanates • glucosinolates occur naturally; watercress and other crucifers • enzyme released when plant wounded • isothiocyanates produced as secondary metabolites • important in defence of plant against herbivorous invertebrates Parabolic screen above outfall

  4. Hypothesis • Gammarus pulex shows an increased sublethal response following repeated exposure to pure and watercress derived phenethyl isothiocyanate (PEITC). Gammarus pulex (L.)

  5. Method Development • selection of test organism; depletion of G. pulex • acute juvenile test • sublethal endpoint, precopular separation • time constraints, degradation rate G. pulex precopular pairs

  6. Method • prepare wash water or PEITC solution • expose pairs for 2h • return to clean water • count pairs re-formed • re-expose for 2h in fresh wash water

  7. Results • response during 2h exposure • comparison of pairs separated: exposure & re-exposure • pairs reforming after 24h in clean water • ET50 values

  8. PEITC (1µl/l) Wash water Control Re-exposures Pairs separated % Response during 2h exposure 100 80 60 40 20 0 0 15 30 45 60 75 90 105 120 Time (mins)

  9. Comparison of pairs separatedafter 2h (exposure/re-exposure) 100 80 % 60 40 Control Upper 95% CI 20 Control Lower 95% CI 0 Wash Wash Wash PEITC 3 PEITC 5 water 2 water 4 Water 5

  10. Pairs re-forming after 24h in clean water

  11. 120 100 80 60 40 20 0 ET50 (95% CI) values Mean 81±9 mins Washwater Minutes 140 120 Mean 71±14 mins 100 80 60 40 PEITC 20 0 Individual Tests

  12. Mean ET50 (± SE) 100 80 60 Wash water Minutes WW re-exp. PEITC (1ug/l) 40 PEITC re-exp. 20 0

  13. Environmentally relevant concentration used. • Baseline sensitivity /variability in mate guarding phase. • Potential pairs ‘lost’ during exposure; 3rd exposure possible? Watercress (Nasturtium officianale (R.Br.))

  14. Conclusions • re-exposures could be successfully carried out • re-exposures resulted in slight (not statistically significant) increase of impact on most occasions • transient impact? Watercress in flower

  15. We gratefully acknowledge the support of the Vitacress Conservation Trust and use of facilities at the National Oceanographic Centre and Environment Agency Biological Effects Laboratory. Vitacress Salads Ltd, Hampshire, UK

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