Global rates of atmospheric nitrogen deposition

1. ELEVATED ATMOSPHERIC NITRATE DEPOSITION IN NORTHERN HARDWOOD FORESTS: IMPACTS ON MICROBIAL MECHANISMS OF PLANT LITTER DECOMPOSITION Jared L. DeForest Earth, Ecological, & Environmental Sciences University of Toledo

2. Global rates of atmospheric nitrogen deposition 50.0 20.0 10.0 7.5 5.0 2.5 1.0 0.5 0.3 0.1 kg N ha-1 Galloway & Cowling, (2002)

3. Total Nitrogen Deposition (2002)

4. Human activities have doubled the amount of available nitrogen Global Nitrogen Cycle 150 Values in 1012 g; From Schlesinger (1997)

5. Human activities have doubled the amount of available nitrogen Global Nitrogen Cycle 150 Values in 1012 g; From Schlesinger (1997)

6. The deposition of nitrogen can be in two forms: Nitrate (NO3-) or Ammonium (NH4+) Nitrate represents the majority of total nitrogen deposition in the Midwest Nitrate is rapidly assimilated by the microbial community and through the process of cell death, that nitrogen is released as ammonium Ammonium can represent 75% of extractable total inorganic nitrogen in soil

7. Human Nitrate Deposition

8. 120 Gt C yr-1 60 Gt C yr-1 The doubling of available nitrogen can be a potent modifier of the carbon cycle Atmosphere 750 Gt C GPP Land Plants 560 Gt C 60 Gt C yr-1 Respiration Decomposition Soils 1500 Gt C Adapted from Schlesinger (1997)

9. Increases in nitrogen deposition can inhibit decomposition because high levels of soil nitrogen can suppress the activity of enzymes that degrade plant litter Lignin degrading enzymes are the most likely to be suppressed by increases in soil ammonium availability

10. Ligninolytic activity is often inhibited by ammonium (NH4+) Extracellular Ammonium Ligninolytic Activity Ammonium (mM) Ligninolytic Activity Culture Age (days) Adapted from Keyser et al., 1978

11. Basidiomycetes are the primary decomposers of lignin Degrading lignin is a specialized function giving lignin-degrading microorganism access to lignified carbohydrates. A relatively small population of soil bacteria, actinomycete, and fungi have the ability to depolymerize lignin by non-enzymatic and enzymatic means. White-rot fungi are considered the primary decomposers of lignin because they produce an array of enzymes that can fully degrade lignin.

12. White-rot fungi are a physiological, rather than a taxonomic, grouping of fungi. At least 21 genera are considered white-rot fungi. Evidence of White-Rot Decomposition

13. Lignin The decomposition of lignin is important because: Lignin is the second most abundant organic molecule Lignin protects plant tissue from decomposition

14. The Decomposition of Plant Litter Labile Compounds Remaining Mass Non Lignified Cellulose Lignified Cellulose Lignin Time  Adapted from Berg (1986)

15. Phase regulated by nutrient level and readily available carbon Phase regulated by lignin decomposition rate Labile Compounds Remaining Mass Non Lignified Cellulose Lignified Cellulose Lignin Time  Adapted from Berg (1986)

16. Phase regulated by lignin decomposition rate Phase regulated by nutrient level and readily available carbon Ambient Nitrogen Elevated Nitrogen Mass Loss Time Adapted from Fog (1988)

17. Microbial Nitrate Assimilation And Turnover Human Nitrate Deposition Less Lignolytic Enzyme Activity More Available NH4+ Less Litter Decomposition Reduced Carbon Flow Less Lignin Decay

18. Hypothesis Chronic nitrate additions can suppress the lignin-degrading activity of soil microbial communities

19. Predictions Nitrate amended soils will have: A microbial community composition with less fungi Lower activity of enzymes that degrade lignin and cellulose

20. Study Sites 7 9 12 12 (kg N ha-1 y-1)

21. PLOTS Ambient N Deposition Plus 30 kg N-NO3- ha-1 y-1 Ambient Nitrogen Deposition A B C D

22. Cell membranes can be used to determine microbial community composition Lipid bilayer Cell membrane Microbial cell Phospholipid

23. Phospholipid Fatty Acids Unique to fungi Fatty Acids Tails Common to many soil microorganisms The length of fatty acid tails and position of double bonds on the tails can be unique to broad taxonomic groups

25. Extracellular Enzymes Plant Litter Compound Cellobiohydrolase Cellulose Cellulose b-glucosidase Peroxidase Lignin Lignin Phenol oxidase Enzyme Analysis

26. Nitrate additions had no noticeable effect on microbial community composition % mol fraction

27. Nitrate additions decreased microbial biomass Total PLFA (nmol PLFA mg-1 C)

28. -glucosidase b * Peroxidase * Cellobiohydrolase Phenol Oxidase * p < 0.05 -40% -30% -20% -10% 0% 10% Change in Enzyme Activity Nitrate addition suppressed activity of soil lignin & cellulose degrading enzymes

29. * p < 0.05 -glucosidase b Peroxidase Cellobiohydrolase * Phenol Oxidase -40% -30% -20% -10% 0% 10% Change in Enzyme Activity Nitrate addition suppressed activity of lignin degrading enzymes in litter

30. Nitrate Additions Microbial Community Composition Lignolytic Activity Total PLFA (Microbial Biomass) No Apparent Change Decrease Decrease

31. Decreases in b-glucosidase activity can help explain lower microbial biomass in nitrate amended soils. Reductions in b-glucosidase activity can diminish the physiological capacity of the microbial community to metabolize cellulose. This reduction could reduce the energy enzymatically derived from cellulose degradation.

32. Conclusions Anthropogenic nitrate deposition may diminish the physiological capacity of soil microbial communities to degrade plant litter.

33. Does a suppression of lignin & cellulose degrading enzymes indicate a reduction in the flow of carbon from these compounds?

34. Hypothesis Nitrate additions will inhibit the ability of soil microorganisms to metabolize and assimilate the products of lignin and cellulose degradation

36. CHO OCH3 H OH 13C Vanillin Lignin Microbial Assimilation

37. Cellulose Microbial Assimilation 13C Cellobiose

38. 13C Sequential Extractions: Soil was incubated for 48 hours and 13C was traced into respiration, dissolved organic carbon (DOC), microbial carbon, and soil carbon.

39. 13C PLFA Analysis: Traced the flow of labeled 13C vanillin and cellobiose into cell membranes.

40. CHO OCH3 H OH Microbial Membrane Extraction & Separation 13C 13C PLFA Analysis 13C 13C 13C 13C 13C 13C 13C 13C 13C Analysis

41. N additions increased the incorporation of vanillin into PLFAs * Vanillin * * Cellobiose

42. N additions did not alter the flow of 13C vanillin into carbon pools

43. N additions did not alter the flow of 13C cellobiose into carbon pools

44. N additions increased soil organic carbon Soil Organic Carbon (mg C g-1)

45. Chronic nitrate additions Excess nitrogen likely inhibits lignocellulose degradation more than vanillin or cellobiose degradation UNCHANGED Vanillin or Cellobiose into Carbon Pools INCREASED Soil Organic Carbon

46. Conclusions Nitrate additions have apparently stemmed the flow of carbon through the soil food web evident by increasing soil organic matter formation through a reduction in lignolytic activity.

47. Northern Hardwood Forests Implications Atmospheric CO2 Pools Slower Decomposition

48. Human Nitrogen Deposition

49. Global Implication The same mechanism that decreases lignin decomposition could be used to understand the impact nitrogen deposition may have on broad global patterns of decomposition

50. Environmental Conditions Litter Biochemistry Global Controls of Decomposition Plant litter decay