I. Cakmak Sabanci University Istanbul , Turkey cakmak@sabanciuniv

1. Detrimental Effects of Glyphosate on Nutritional Status of Plants with Micronutrients I. Cakmak Sabanci University Istanbul, Turkey cakmak@sabanciuniv.edu

2. Glyphosate: extensively applied herbicide • Increasing usage in minimum- and no-tillage agricultural practices • Extensive application on RR crops .

3. In regions with extensive use of glyphosate there are increasing reports on: • reductions in growth and yield, • increases in disease problems, • increased use of insecticides and fungicides, • inhibition of N fixing bacteria • increased use of foliar micronutrient fertilizers • micronutrient deficiencies

4. Particular micronutrient deficiencies induced by Glyphosate:Manganese and Iron Deficiencies

5. Repeated use ofglyphosateinduces Fe and Mn deficienciesin soybeans in USA (Photo: Prof. Don Huber) (areas which are not treated with glyphosate)

6. Glyphosate-induced Fe-deficiency chlorosis + gyphosate + seed Fetreatment - gyphosate +seed Fe treatment + gyphosate –seed Fe treatment Photo: N.C. Hansen, Fort Collins, USA (Jolley et al., Soil Sci Plant Nutr. 50, 793-981, 2004)

7. visual chlorosis scroregrain yieldTreatment (1=green to 5 =severe) (t/ha) – Fe + Fe* – Fe + Fe* Control 3.1 2.8 1.01 1.70(no herbicide) Glyphosate 3.7 3.3 0.27 0.61 Interaction of seed applied Fe and glyphosate application on Fe deficiency chlorosis in soybeans;Minnesota, USA * 50g Fe/ha as FeEDDHA applied to seeds (Jolley et al., Soil Sci Plant Nutr. 50, 793-981, 2004)

8. Foliar and Soluble Micronutrient* Sales in USA RoundUp Ready Corn, Soy (*represents approx 30% of US Sales) (Courtesy of Prof. P. Brown, UC Davis)

9. Glyphosate istoxic to plants whether from a drift or residues in soil A significant amount of glyphosate applied to target plants reaches the soil as a result of direct contact, wash off from leaves and exudation from roots of the treated weeds. Up to 10 % of the applied glyphosate move to non-target plants entweb.clemson.edu. http://www.uky.edu/Ag/Tobacco/Pages/RoundupFloat.html

10. no Glyphosate with Glyphosate

11. Application of glyphosate on target plants (weeds)/glyphosate resistant cultivars; very quick uptake by leaves. Rapid translocation of glyphosate from shoot to root REMEMBER Glyphosate is highly phloem mobile Release of glyphosate into the rhizosphere

12. Adaptive Root Responses of Strategy-I Plants to Fe Deficiency Plasma Rhizosphere/Soil Apoplasm Cytoplasm Membrane FeII TR Chelator FeIII R FeIII Chelate FeIII FeIII PUMP H+ H+ FeIII Reductants (Chelators) R Marschner and Römheld, 1995; Plant and Soil R: Ferric Reductase

13. Effect of iron deficiency in cucumber on pH decreasing and reducing capacity of the roots +Fe -Fe +Fe -Fe Romheld et al., 1981

14. ) 1 - 80 root dry wt 3 h LSD n capacity 60 0.05 1 40 - Fe reductio (µmol Fe(II) g 20 0 -Glyp. +Glyp. +Glyp. +Glyp. -Glyp. 6h 12h 24h +Fe Effect of Glyphosate on Ferric Reductase Time dependent inhibition of ferric reductase activity in Fe-deficient sunflower plants (+Glyp: 1.89 mM glyphosateor 6% of the recommended rate for weed control) Fe deficient Control (+Fe) Ozturk et al., 2007

15. ) 1 - 80 root dry wt 3 h 60 LSD 0.05 Fe reduction capacity 1 - 40 (µmol Fe(II) g 20 0 -Glyp. +Glyp. +Glyp. +Glyp. -Glyp. 0.32 mM 0.95 mM 1.89 mM +Fe Dose dependent inhibition of ferric reductase activity by glyphosate (+Glyp: 0.32, 0.95 and 1.89 mM Fe deficient Control (+Fe) Ozturk et al., 2007

16. Foliar applications of glyphosate (Glyp) at 0.32 mM, 0.95 mM and 1.89 mM for 24h. Staining of Ferric Reductase Activity GLYPHOSATE Ozturk et al., 2007 Fe Deficient Plants

17. Effect of foliar applied of glyphosate (1.89 mM) for 24h on ferric reductase activity. -Fe -Fe +Glyp (1.89 mM) Ozturk et al., 2007

18. Dose Dependent Shikimate Accumulation Shoot Shikimate accumulation (µmol g -1 FW) Ozturk et al., 2007

19. Dose Dependent Shikimate Accumulation Root Shikimate accumulation (µmol g -1 FW) Ozturk et al., 2007

20. Time Dependent Shikimate Accumulation Shoot Root Shikimate accumulation (µmol g -1 FW) Ozturk et al., 2007

21. Short-Term Experiments: Glyphosate inhibits root uptake and root-to-shoot transport of micronutrients Root Uptake Translocation 59Fe Effect of 1.89 mM glyphosate (equivalent to 6 % of the recommended dosage for weed control in the field) on root uptake and shoot translocation of 59Fe, 54Mn and 65Zn in sunflower plants. 54Mn 65Zn Eker et al., 2006: J. Agric. Food Chem. 26, 10019 -10025

22. Plasma Membrane Adaptive Root Responses to Iron and Zinc Deficiencies in Cereals Rhizosphere/Soil Apoplasm Cytoplasm PHYTOSIDEROPHORES FeIII ZnII FeIII FeIII-PS FeIII-PS TR ZnII ZnII-PS ZnII-PS TR: Transporter Protein Marschner and Römheld, 1995; Plant and Soil

23. Biosynthetic Pathway of Mugineic Acid Family Phytosiderophores Mugineic Acid Family Mori et al., 1990

24. DMA and NA Fe OH Fe NH2 Fe3+-Deoxymugineic acid Fe3+-Nicotianamine Similar Stereochemical Structure between Fe3+-DMA and Fe3+-NA Mori, et al, 1990

25. Ozturk et al., 2008

26. Phytosiderophore Release from Wheat Roots Glyphosate Applicaton Ozturk et al., 2008

27. Ozturk et al., 2008

28. Ozturk et al., 2008

29. Application of glyphosate on target plants (weeds)/glyphosate resistant cultivars; very quick uptake by leaves. Rapid translocation of glyphosate from shoot to root REMEMBER Glyphosate is highly phloem mobile Release of glyphosate into the rhizosphere

30. Glyphosate applied to target plants is released into the rhizosphere Induction of Fe deficiency chlorosis in non-target plants (sunflower) Sunflower Indicator Soybean Target Sunflower Indicator Soybean Target Soybean Target Soybean Target (Neumann et al., 2005, J. Plant Dis. Prot.) Development of Fe deficiency symptoms in non-target plants … Glyphosate application to target plants (soybean)

31. Glyphosate released in therhizosphere reduces Mn uptake 54Mn in shoots Inhibited Mn acquisition by non-target plants (sunflower) grown in nutrient solution together with glyphosate treated target plants (soybean) for 2, 4 and 6 days. Neumann et al., 2005, J. Plant Dis. Prot.

32. Significantly lower Mn uptake/accumulation in RR-soybeans KS 4202 KS 4202 RR (B.Gordon, 2006; Kansas State Univ.)

33. Inhibition of MnO2 Reduction in Soil-filled Rhizoboxes after Glyphosate Treatment to Target Plants (soybean) Control + Glyphosate MnO2 Reduction in Soil-filled Rhizobox Culture of sunflower (non-target) and RR-Soybean with and without foliar Glyphosate spray (Neumann et al., 2005)

34. Glyphosate Reduces Grain Mn, Mg and Ca 00.30.6 0.9 Glyphosate Application, %

36. Yazici et al., 2008

37. Yazici et al., 2008

38. Seed Mn and Ca very important for seed viability and seedling vigour

39. Yazici et al., 2008

40. Hande et al., 2008

41. Plants with low Mg are highly sensitive to glyphosate Hande et al., 2008

42. Plants with low Mg are highly sensitive to glyphosate Hande et al., 2008

43. What is the effect of Glyphosate??

44. Effectiveness of Glyphosate Effect of Glyphosate with and without calcium in the tank Glyphosate + Calcium Glyphosate Control Glyphosate binds with the cations to form a strong complex which is not bio-available. Only unbound glyphosate act as a herbicide. www.loveland.co.uk/ Gifs/X-Change-du-pont.gif

45. Glyphosatebinds with the cations to form a strong complex which is not bio-available. Only unbound glyphosate act as a herbicide.

46. Conclusions • Routine glyphosate use in agricultural systems results in considerable side effects on plant growth and mineral nutrition of plants • Glyphosate is antagonistic to the uptake, transport and accumulation (tissue concentration) of Fe,Mn, Ca and Mg possibly due to the formation of poorly soluble glyphosate-metal complexes (??)

47. Conclusions-cont • Glyphosate impairs genetic adaptation mechanisms of plants to Fe deficiency • Plants grown under low Mg are very sensitive to glyphosate • A new risk assessment for glyphosate is urgently needed,

49. Root Tips: Co-localization of FerricReductase and Glyphosate Accumulation

50. Leading 10 Health Risk Factors in Developing Countries, % Cause of Disease Burden WHO, 2002