Nutrient Dynamics • Nutrient Uptake and Growth Models • Nitrogen Assimilation & Preference • Phosphorus • Nutrient Limitation Assays • Nutrient Regeneration • How are rates of uptake and regeneration often measured (plankton versus benthic)?
Nutrient Uptake and Growth Models • Uptake rate vs substrate concentration in environment (Michaelis-Menten model). • Growth rate vs substrate concentration intracellular (Droop model). • Growth rate vs substrate concentration in environment (Monod model)
Nutrient Competition • Large algal cells may perform luxury uptake and storage (e.g., diatoms) • Small algal cells out-compete at lower concentrations. • Bacteria can do both for phosphate; they compete with phytoplankton.
Nitrogen “Preference” • Phytoplankton: NH4+ > NO3- ≈ (urea) • Bacteria: aa> NH4+ > NO3- ≈ (urea) N2-fixation last (most E)
Nutrient Limitation • Liebig's law of the minimum. • Cellular elemental balance as a index. • Environmental elemental balance. • Enzyme expression as an index.
PO43- PO43- PO43- Alkaline Phosphatase (AP) Activity of Aquatic Bacteria Indicates P-Bioavailability Low to No PO43- supply: AP activity is expressed at high levels. High PO43- supply: AP activity is repressed or inhibited de novosynthesis Nucleic Acids PO43- Phospholipids Dissolved Organic Phosphorus (DOP) = energetically costly
Nutrient Regeneration • Microbial food web dominates regeneration. • Bacteria important when organic matter consumed is C:N < 10 or C:P < 60. • Often U = R. • R > U; concentration increases. • R < U; concentration decreases.
How is uptake and regeneration measured in the field? • Net effects (difference in U and R). • Incubation with 15N labeled compounds: • 0.3663% of 15N +14N as 15N (add < 10%) • Uptake is what accumulates in particles. • Regeneration is by “isotope dilution” of DIN. • Whole system budgets: • Upstream addition of a conservative tracer. • Again use 15N added directly to the ecosystem.
High C:N ratio of organic substrates Uptake Carbohydrates NH4+ NO3- N-replete bacterium N-deplete bacterium Amino Acids Control of role in N-cycling Low C:N ratio of organic substrates Regenerate Carbohydrates NH4+ Amino Acids
Response to C & N supply: • GDH Regulation: • Expression and activation at low C:NLDOM ratio • Repression and inactivation at high C:NLDOM ratio • GS Regulation: • Reverse of GDH. • GDH:GS activity ratio (Hoch et al., 2006).
Assess the bioavailability of N and P in freshwater bacterioplankton. • Does GS & GDH activity respond to amendments of C and N in lake bacterioplankton cultures? • Does P supply (assessed by AP activity) affect N-metabolism? • Are results influenced by community composition?
Amendment Experiments • < 0.8 μm filtrate is inoculum and media. • Amend replicates with NH4+, PO43- & glucose. • Monitor parameters initially and after 24 h.
Lake BacteriaAmendmentExperiments • Expected +N response. • Unexpected glucose response (need P). • +P repressed AP. • Increasing GS activity requires +P; DIN uptake increased.
% chemical denaturant 25 % 55 % (-) (-) (-) (-) (+) (+) (+) (+) Did the community change? Denaturing Gradient Gel Electrophoresis (DGGE)
+P +P +G+P Lake Williams 16SrDNA DGGE Minor richness increase after 24 h in +P and +P+G treatments.
Relationships among N-metabolism and that of P and C. • More N-replete bacteria are more P-limited. • More N-replete bacteria have less efficient growth. • More P-replete bacteria have more efficient growth.
Lake Sites:Contrasting TN:TP ratio • Both lakes: • Lower Susquehanna River Basin • Piedmont region • Eutrophic • Lake Williams: • East Branch Codorus Watershed • ≈ 80% agriculture land use • TN:TP = 286 • Lake Pinchot: • Conawego Watershed • ≈ 40% agriculture landuse • TN:TP = 17.1 (sewage-P) (Susquehanna River Basin Commission, 2001)
Watershed Summary • Low GDHT:GS due to greater supply of labile organic-C and PO43-; DIN uptake. • Low GDHT:GS suggest N-replete bacteria that regenerate NH4+. • Bacterial community composition does not appear to greatly influence enzyme activity. • TN:TP ratio of lake ecosystems influences bacterial nutrient dynamics (sewage effect). • Similar results with periphyton (“rock slime”) communities in streams.