1. “DEBtox”, a brief history and extension to mixtures and plants Tjalling Jager Dept. Theoretical Biology

2. Contents  Brief history of toxicants in DEB –the “DEBtox” family tree  Case study with toxicants in DEB3 –nonylphenol in Capitella teleta  How to extend DEBtox to mixtures –overview of progress made in NoMiracle framework  How to extend DEBtox to plants –what makes plants different from animals?

3. Background external concentration (in time) repro growth survival feeding hatching … Descriptive analysis you don’t learn anything, ignore that effects change in time ignore that endpoints are linked cannot extrapolate to other situations

4. The DEBtox concept external concentration (in time) toxico- kinetics toxico- kinetics internal concentration in time DEB parameters in time DEB model DEB model repro growth survival feeding hatching …

5. The DEBtox concept external concentration (in time) toxico- kinetics toxico- kinetics internal concentration in time DEB parameters in time DEB model DEB model Internal concentration are often not measured … repro growth survival feeding hatching … DEB parameter cannot be measured …

6. DEB theory  Organisms obey mass and energy conservation –find the simplest set of rules... –over the entire life cycle... –for all organisms (related species follow related rules) –most appropriate DEB model depends on species and question resources waste products growth maintenance maturation offspring Kooijman (2010)

7. Generalised life cycle feeding gametes

8. eggs mobilisation Standard DEB model for animal structure somatic maintenance  growth maturity maintenance 1-  reproduction maturity buffer maturation p foodfeces assimilation reserve b

9. Targets link to DEB parameters Assumption –toxicant affects target site linked to one (or more) DEB parameter(s) target sitetoxicant DEB parameter organism Assumptions –there is a threshold for effects (the NEC) –above the NEC, parameter changes linearly with internal concentration...

10. A brief history of ‘DEBtox’ Corresponds with origin of DEB in 1979 egg

11. A brief history of ‘DEBtox’ The 80’s …  Kooijman (1981) –toxicokinetics determines survival pattern  Kooijman & Metz (1984) –toxicants affect energy budgets and thereby population response egg

12. A brief history of ‘DEBtox’ egg The early 90’s …  Parallel to OECD trajectory –review test guidelines with respect to statistical analysis

13. A brief history of ‘DEBtox’ Birth in 1996 …  Windows software/booklet –Kooijman & Bedaux, 1996  Series of papers –Bedaux & Kooijman (1994) –Kooijman & Bedaux (1996) –Kooijman et al (1996)

14. A brief history of ‘DEBtox’ And 10 years later …  ISO/OECD (2006) –DEBtox next to methods for NOEC and EC50  ECB workshop (2007) –presenting DEBtox to EU regulators

15. eggs mobilisation Simplified DEB animal structure somatic maintenance  growth maturity maintenance 1-  reproduction maturitybuffer maturation p foodfeces assimilation reserve

16. A brief history of ‘DEBtox’ The 2000’s …  Péry et al (2002, 2003) –modifications for midges  Ducrot et al (2004, 2007) –midges and snails  Lopes et al (2005) –link to matrix models  Billoir et al (2007, 2008) –Bayes, new derivation  Muller et al (2010) –alternative formulation division

17. A brief history of ‘DEBtox’ In our group …  Jager et al (2004), Alda Álvarez et al (2005, 2006) –multiple endpoints and ageing –population (Euler-Lotka)  Baas et al (2007, 2009) –mixtures: lethal effects division

18. A brief history of ‘DEBtox’ Embryo division …  Klok & De Roos (1996), Klok et al (1997, 2007) –earthworm matrix model, Bayesian approach division

19. A brief history of ‘DEBtox’ division Applying ‘DEB3’ …  Jager et al (2010) –basis and mixtures  Jager & Klok (2010) –compare methods and population level  Jager & Selck (acc.) –worked-out case study Kooijman (2010)

20. A brief history of ‘DEBtox’ New offspring …  E.g., from current PhD projects … division

21. A brief history of ‘DEBtox’ Increase in... –flexibility –complexity –computation time –data needs –new questions division Kooijman-Metz classic DEBtox DEB3

22. A “DEB3” DEBtox case study Tjalling Jager Dept. Theoretical Biology

23. Toxicant case study  Marine polychaete Capitella (Hansen et al, 1999) –exposed to nonylphenol in sediment –body volume and egg production followed –no effect on mortality observed Jager and Selck (acc. J. Sea Res.)

24. Control growth  Volumetric body length in control –here, assume no contribution reserve to volume … 01020304050607080 0 0.5 1 1.5 2 2.5 3 time (days) volumetric body length (mm) 0

25. Control growth Assumption –effective food density depends on body size 01020304050607080 0 0.5 1 1.5 2 2.5 3 time (days) volumetric body length (mm) 0

26. Control growth 01020304050607080 0 0.5 1 1.5 2 2.5 3 time (days) volumetric body length (mm) 0 Assumption –initial starvation (swimming and metamorphosis)

27. Control reproduction  Compare to mean reproduction rate DEB –ignore reproduction buffer … 01020304050607080 0 500 1000 1500 2000 2500 3000 3500 time (days) cumulative offspring per female 0

28. NP effects  Compare the control to the first dose

29. “Hormesis”  Requires a mechanistic explanation … –conservation of mass and energy must hold! Potential assumptions –NP is a micro-nutrient –decreased investment elsewhere (e.g., immune system) –NP relieves a secondary stress (e.g., parasites or fungi) –NP increases the food availability/quality

30. NP effects Assumption –NP increases food density/quality

31. NP effects Assumption –NP affects costs for making structure –somatic = maturity maintenance coefficient

32. Standard DEB animal structure foodfeces maturity maintenancesomatic maintenance assimilation  1-  growth reproduction maturity buffer maturation reserve mobilisation eggs

33. NP effects Assumption –NP also affects costs for maturation and reproduction

34. Standard DEB animal structure foodfeces maturity maintenancesomatic maintenance assimilation  1-  growth reproduction maturity buffer maturation reserve mobilisation eggs

35. fit not satisfactory? fit Strategy for data analysis actual DEB model experimental data additional experiments literature educated guesses mechanistic hypothesis standard DEB model Accept the simplest explanation … for now

36. DEBtox extensions: mixtures of toxicants Tjalling Jager Dept. Theoretical Biology

37. The challenge …. –some 100,000 man-made chemicals … –large range of natural toxicants and other stressors … –for animals alone, >1 million species described … –complex exposure situations … –effects depend on exposure time, endpoint, …

38. The challenge …. –some 100,000 man-made chemicals … –large range of natural toxicants and other stressors … –for animals alone, >1 million species described … –complex exposure situations … –effects depend on exposure time, endpoint, … Descriptive approaches like CA and IA are of little use! external concentration A at time t one life-history trait at time t external concentration B at time t

39. external concentration A (in time) external concentration B (in time) toxico- kinetics toxico- kinetics toxico- kinetics toxico- kinetics internal concentration A in time internal concentration B in time DEB parameters in time DEB model DEB model life-history traits theory implies interactions … growth reproduction Mixtures in DEB

40. external concentration A (in time) toxico- kinetics toxico- kinetics external concentration B (in time) toxico- kinetics toxico- kinetics DEB model DEB model internal concentration A in time DEB parameters in time internal concentration B in time life-history traits Mixtures in DEB

41. Simple mixture rules assimilation maintenance … compound‘target’ DEB parameter toxicity parameters linked (compare CA)

42. Simple mixture rules assimilation maintenance … compound‘target’ DEB parameter

43. Simple mixture rules assimilation maintenance … toxicity parameters independent (compare IA) compound‘target’ DEB parameter

44. Visual representation Baas et al (2007)  For binary mixture, model represents surface over time … –example Cu and Cd in Folsomia candida

45. fluoranthene pyrene PAHs in Daphnia  Based on standard 21-day OECD test –10 animals per treatment –length, reproduction and survival every 2 days –no body residues (TK inferred from effects) Jager et al (2010)

46. costs reproduction (and costs growth) costs reproduction (and costs growth) same target

47. Iso-effect lines for body length <50% effect

48. Mixture conclusions  Multiple TK modules link to one DEB model –either through the same or different target  We have ‘proof of concept’ –but lack of suitable data … –more data need to be analysed  For more complex mixtures, other interactions are inevitable –which require mechanistic assumptions

49. DEBtox extensions: what makes plants different? Tjalling Jager Dept. Theoretical Biology

50. About reserve and structure reserve structure  Abstractions, in DEB theory: –reserves buffer metabolism from environment –structure and reserve have constant composition –composition can be different between both –their can be multiple reserves and structures... reserve structure reserve structure reserve structure reserve structure reserve structure

51. maintenance Animals that feed on animals reserve structure food  food has similar composition as body

52. Animals that feed on plants  food has different (but constant) composition as body maintenance reserve structure food excess carbon

53. maintenance Animals that feed on anything carbohydrate and lipids structure food  food has different (varying) composition as body protein maintenance excess

54. maintenance Plants that use light carbohydratenitrogen structure lightCO 2 NO 3 excess  energy and building blocks separately -

55. Roots and shoots... shoot root

56. How many reserves? Depends on organism and on question  Nutrients may need a reserve compartment –if they are (can be) limiting –if they vary in time  DEBtox for algal growth in standard test... –constant light, excess nutrients –thus: no reserves needed –(Kooijman et al, 1996) reserve

57. Looking for simplicity For things like algae and Lemna...  Maturation and reproduction may be ignored –not so clear how these concepts work in plants  No large distinction between individual and population –population can be modelled as one individual –with surface area proportional to volume

58. Wrapping up What is DEBtox? –a specific piece of software (not available anymore) –principle: toxicants affect parameters of any DEB-based model Analysis of mixtures –follows naturally from the DEB perspective –more work needed to see how far this idea can get us Extension to plants –differ from animals; resources not from one flux –DEB theory includes plants, so DEBtox approaches can be developed

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