1. BIOTIC REGULATION OF SEDIMENT ORGANIC PHOSPHORUS MINERALIZATION IN SUBTROPICAL LAKES Isabela Claret Torres

2. Introduction –Eutrophication of freshwater ecosystems Strongly related to human activities High load of nutrient input:  wastewater of anthropogenic activities: » agriculture and urban activities Paths of Cultural Eutrophication (anthropogenic eutrophication)

3. Dominance by cyanobacteria, diatom, and green algae  input nutrients (N & P)  light availability  phytoplankton biomass  biological diversity occurrence of algae ‘bloom’  oxygen concentration change in heterotrophs composition death of fish ecosystem change, loss of species diversity and decrease in water quality

4. –Oligotrophic/ Eutrophic/Hypereutrophic: ≠ Characteristics –Nutrients will accumulate in the sediment: Altering Properties of Sediments –Internal load: Chemical, Physical and Biological –Benthic Sediments  Nutrient Cycling: Sources or Sinks  Nutrients

5. –Reduction and control of external load Internal load :  Trophic state / time lag for recovery –Phosphorus (P) Eutrophication – Control of P availability: Controlling and reversing eutrophication

6. Microbial Community Organic P Labile Slowly Available (DRP) Refractory Organic P Sediment Sediment P release DRP Water Column P Input (external) Living Particles Non Living Particles Dissolve P Particulate P

7. –Earlier studies: Abiotic regulation of P solubility in sediments Disregarding the microbial communities –Microbial communities: First to respond to an environmental change Affected or regulated by environmental factors First to change due to eutrophication

8. Torsvik et al. (2002)  complexity food web  diversity Few dominant species  diversity and distribution  [nutrient]  Levels of Eutrophication  [nutrient]  Trophic levels

9. –Microbial community can influence the organic P dynamics in sediments Directly:  Hydrolyzed by enzymes Indirectly:  Metabolism can alter pH, redox potential, and other chemical characteristics of the sediment

10. –Microorganisms: sense + respond  environment Basic understanding of their functional role in relation to nutrient cycles –Concerns about internal loading of nutrients: Better understanding the biotic factors that regulate nutrients cycling and ultimately impact water quality –Few studies have been conducted on the role of microbial communities and their activities in regulating P solubility in sediments

11. I propose to study the microbial community biomass, diversity and activity as related to P solubility in sediments of subtropical lakes with different trophic conditions

12. OBJECTIVES AND HYPOTHESIS To develop a linkage between the biogeochemical characteristics of benthic sediments and the microbial community in relation to their activities Hypothesis ≠ Trophic Conditions Sediments ≠ Biogeochemical Characteristics Microbial Community Organic P Mineralization

13. 1 – OBJECTIVE –To determine the biogeochemical properties of benthic sediments in lakes with different trophic conditions –Hypothesis: Nutrient accumulation rates in sediments are directly related to trophic conditions Sediments of lakes:  ≠ trophic state  ≠ biogeochemical properties

14. 2 – OBJECTIVE –Characterize the microbial community biomass and diversity –Hypothesis:  [nutrient]  microbial biomass  diversity  nutrient availability will be a selection pressure that will favor organisms that are well adapted to this condition, thus supporting high biomass but decreasing diversity

15. 3 – OBJECTIVE –Characterize distribution of heterotrophic microbial activities: Enzyme activities:  Phosphatase and 5’nucleotidase Heterotrophic microbial respiration:  Mineralization of organic P –Hypothesis: Sediments of lakes with different biogeochemical characteristics due to trophic conditions will hold distinct microbial communities that will be reflected on their activities and ultimately will influence organic P mineralization

16. EXPERIMENTAL APPROACH 3 Lakes Selected

17. Lake Annie (Highlands County) Located in south central Florida Small egg shaped lake (36.6 ha) Sediments vary from organic to sand Thermal stratification from March  October Maximum depth of 20.7 m (mean 9.09 m) Oligotrophic Lake Low nutrients concentration: 1) TP mean: 5 µg l -1 2) TN mean: 373 µg l -1 3) Chlorophyll-a mean: 3.6 µg l -1

18. Lake Okeechobee Cultural eutrophication 50 yrs Becoming hypereutrophic Littoral Mud Peat Sand Rock Located in south Florida Largest lake in Florida (Surface area of 1890 Km 2 ) Shallow lake (mean depth of 2.7 m) Eutrophic Lake High Nutrient Concentration: 1) TP: 80 µg l -1 2) TN: 1,700 µg l -1 3) Chlorophyll-a: 25 µg l -1 Reddy et al. 1991

19. Lake Apopka Clear water-macrophyte  turbid algal dominated Nutrient input : 1) discharge of sewage (since 1922) 2) agriculture wastewater (since 1942) Even though these inputs have been controlled and regulated, the eutrophication process continued Large surface area (125 Km 2 ) Shallow lake (mean depth 2 m) Located in central FL Sediments divided in two main layers (Reddy & Graetz 1991) 1) surficial (0-35 cm) unconsolidated material  recent algal deposits and allochthonous material 2) subsurface layer (35-78 cm) consolidated material  partially decomposed algal cells and particulate OM Hypereutrophic Lake Very High Nutrient Concentration: 1) TP mean = 200 µg l -1 2) TN mean = 5,140 µg l -1 3) Chlorophyll-a mean = 92 µg l -1

20. EXPERIMENTAL APPROACH 1) Sediment Types Surface sediments (0 to 10 cm) 4 sites - Lake Annie 4 sites - Lake Apopka 9 sites - Lake Okeechobee 2) Depth study 1 site - Lake Annie 1 site - Lake Apopka 3 sites - Lake Okeechobee – All sites will be sampled in 3 replicates

21. Task 1: Biogeochemical Properties Methods 1) Water Content - Bulk Density (Blake & Hartge 1986) 2) Total P - LOI: Ignition Method ( Anderson 1976) 3) Total C - N: Carlo Erba NA 1500 CNS ( Nelson & Sommers 1996) 4) MBC - TOC: Fumigation-extraction (Vance et al. 1987) 5) MBN - TKN: Fumigation-extraction (Howarth & Paul 1994) 6) MBP: Fumigation-extraction (Hedley & Stewart 1982) 7) Phosphorus Forms: Fractionation method (Ivanoff et al. 1998) 8) Polyphosphate: 31 P NMR (Hupfer et al. 1995)

22. Task 1: Anticipated Results Deeper sediment Total organic P Refractory Labile Fig. D [P] [C] [N] Annie Okeechobee Apopka Fig. A Labile Organic P Surface Deeper Sediment Surface sediment Total organic P Fig. C Labile Refractory

23. Task 2 Microbial community as influenced by redox zones –Characterize the diversity and biomass of the microbial community –PLFA (phospholipids ester linked fatty acids) Phospholipids: constituents of bacterial cell Good estimative of viable biomass Not at a taxonomic level Functional groups (sulfate reducers, methanogens) Information: shifts in community composition

24. Task 2 Microbial community as influenced by redox zones – Characterize the within group diversity – DNA analysis After PLFA analysis groups will be selected Verify if: Trophic state Nutrient availability within group diversity

25. Task 2: Anticipated Results Microbial Biomass [P] Fig. E Microbial Biomass Depth Fig. F Microbial Diversity C:N:P ratio Fig. H Microbial Diversity [P] Fig. G

26. Task 3: Microbial activities as influenced by redox zones in sediments 1) Enzymes –Alkaline and acid phosphatase activity: Rapid fluorometric assay Fluorescence model substrate MUF-P (4-methylumbelliferyl-phosphate) Enzyme activity will be determined from the difference between the amount of fluorescent substrate liberated over incubation time –5‘-nucleotidase activity: ATP measurements following the procedure described in Siuda & Gude (1994)

27. Task 3: Microbial activities as influenced by redox zones in sediments 2) Heterotrophic microbial respiration: Represented by CO 2 and CH 4 production rates Aerobic and Anaerobic conditions Mineralization OP under aerobic and anaerobic  Methods described by Wright and Reddy (2001) (SRP + OrgP) experiment(SRP + OrgP) Control -

28. Alkaline /Acid Phosphatase Activity [Labile Inorganic P] Fig. L Microbial Biomass Enzyme Activity Fig. I Fig. J Enzyme Activity Depth 5'-Nucleotidase [P] Fig. M Task 3: Anticipated Results

29. Organic P Mineralization Fig. N Anaerobic Aerobic

30. 1: Biogeochemical properties of sediments Preliminaries results from sampling in Lake Okeechobee

31. Objective To characterize the biogeochemical properties of sediments at several locations and different sediment types of Lake Okeechobee

32. O11 M17 KR FC J7 J5

33. Material & Methods Biogeochemical Parameters Water Content/ Bulk Density/pH Total P Total C and Total N Microbial Biomass C/N/P Organic P Forms Anaerobic [carbon dioxide (CO 2 )]

34. Results

35. SitespH Bulk Density (g cm -3 ) Moisture Content (%) LOI (%) M17 (Peat)7.30.1081.571.2 O117.50.0984.940.2 M9 (Mud)7.60.1377.828.5 K87.50.0885.636.5 J78.00.7428.72.1 TC (Sand)7.20.6330.62.4 KR7.30.2364.923.4 J5 (Littoral)7.50.6730.11.6 FC7.10.7025.61.1 Table I: pH, Bulk Density (gcm -3 ), Moisture Content (%), and Organic Matter Content (LOI %)

36. Table II: Extractable Total Organic Carbon (mg kg -1 ) and Labile Total Kjeldahl Nitrogen (mg kg -1 ) SitesTOC (mg kg -1 )TKN-NF (mg kg -1 ) M17 (Peat)1,325156 O11420170 M9 (Mud)31694 K8436135 J76719 TC (Sand)7724 KR17168 J5 (Littoral)8921 FC2910

37. MBC x Labile Po (NaHCO 3 ): p = 0.003, r 2 = 0.74 MBC x Moderate Labile Po (NaOH): p = 0.006, r 2 = 0.69 MBC x Labile Pi (NaHCO 3 ): p = 0.003, r 2 = 0.74 MBC x TKN: p = 0.003, r 2 = 0.74

38. Total Phosphorus Concentration

39. 813 mg kg -1 Moderately Labile Po (NaOH) Readily Available Pi (NaHCO 3 Pi) Inorganic P (HCl Pi) Highly Resistant Po (Residue P) Readily Available Po (NaHCO 3 Po) 1,166 mg kg -1 Peat Mud Littoral Sand 369 mg kg -1 30 mg kg -1

40. CO 2 Production Rates

41. CO 2 Production Rates Correlations: TP: p = 0.000, r 2 = 0.85 TC: p = 0.000, r 2 = 0.89 TN: p = 0.009, r 2 = 0.65 Labile Po (NaHCO 3 ): p = 0.000, r 2 = 0.91 Moderate Labile Po (NaOH): p = 0.000, r 2 = 0.94 Labile Pi (NaHCO 3 ): p = 0.002, r 2 = 0.77 TKN: p = 0.000, r 2 = 0.92 TOC: p = 0.002, r 2 = 0.75 MBC: p = 0.002, r 2 = 0.79 Indication that nutrient availability is influencing microbial activities in Lake Okeechobee sediments

42. Acknowledgements Dr. K. R. Reddy (Advisor) Dr. A. Ogram (Co-advisor) Dr. M. Brenner (Geological Sciences) Dr. E. Phlips (Fisheries and Aquatic Science) Dr. K. Portier (Statistics) Dr. K. Havens ( South FL Water Management District ) Committee Members

43. Acknowledgements Dr. Hari Pant Dr. Kanika Sharma Dr. Ashvini Chauhan Matt Fisher Noel Cawley Mrs. Yu Wang Thank You !