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)

Phosphorus Supply

Nutrient Limitation

• Liebig's law of the minimum.• Cellular elemental balance as a index.

• Environmental elemental balance.• Enzyme expression as an index.

Alkaline Phosphatase (AP) Activity of Aquatic Bacteria Indicates P-Bioavailability

de novo synthesis

Nucleic Acids

= energetically costlyDissolved Organic Phosphorus (DOP)

Phospholipids

PO43-

PO43-

High PO43- supply:

AP activity is repressed or

inhibited

PO43-PO4

3-

Low to No PO43-

supply:

AP activity is expressed at high

levels.

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.

Control of role in N-cycling

Carbohydrates

Amino Acids

NH4+

N-repletebacterium

Regenerate

Low C:N ratio of organic substrates

Carbohydrates

Amino Acids

NH4+

N-depletebacterium

Uptake

High C:N ratio of organic substrates

NO3-

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+, PO4

3- & 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.

Did the community change? Denaturing Gradient Gel Electrophoresis (DGGE)

% chemical denaturant

25 %

55 %

(-)

(+)

(-)

(+)

(-)

(+)

(-)

(+)

Lake Williams 16SrDNA DGGE

Minor richness increase after 24 h in +P and +P+G treatments.

+G+P

+P

+P

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.

(Susquehanna River Basin Commission, 2001)

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)

ParameterSummer 2006

Lake Williams(n = 6)

Lake Pinchot(n = 6)

bacteria (106 ml-1) 2.2 ± 0.52 4.3 ± 0.81

chlorophyll a (μg l-1) 32.7 ± 10.4 70.0 ± 21.4

TN:TP ratio (atom) 290 ± 36 17 ± 6.2

total N (μM) 220 ± 21 63 ± 18

total P (μM) 0.78 ± 0.14 3.7 ± 0.47

bacterial AP(nmol h-1 μg protein-1)

5.4 ± 1.2 0.73 ± 0.38

bacterial GDHT:GS 560 ± 110 72 ± 23

Lakes of Contrasting TN:TP Ratio

Watershed Summary

• Low GDHT:GS due to greater supply of labile organic-C and PO4

3-; 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.