Transcript Slide 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
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)
Total Nitrogen Deposition
(2002)
Human activities have doubled the amount of
available nitrogen
Global Nitrogen Cycle
150
Values in 1012 g;
From Schlesinger (1997)
Human activities have doubled the amount of
available nitrogen
Global Nitrogen Cycle
150
Values in 1012 g;
From Schlesinger (1997)
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
Human
Nitrate
Deposition
The doubling of available nitrogen can be a
potent modifier of the carbon cycle
GPP
Land Plants
560 Gt C
Respiration
120 Gt C yr-1
60 Gt C yr-1
Decomposition
Soils
1500 Gt C
Adapted from Schlesinger (1997)
Atmosphere
750 Gt C
60 Gt C yr-1
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
Ligninolytic activity is often inhibited by
ammonium (NH4+)
2.5
100
Extracellular
Total
Extracellular
Ammonium
75
Nitrogen
1.5
50
Ligninolytic
Activity
1
25
0.5
Ligninolytic Activity
0
0
0
1
2
3
Culture Age (days)
Adapted from Keyser et al., 1978
4
5
6
Ligninolytic Activity
Ammonium (mM)
2
Basidiomycetes are the primary
decomposers of lignin
Degrading lignin is a specialized function giving lignindegrading microorganism access to lignified carbohydrates.
A relatively small population of soil bacteria, actinomycete,
and fungi have the ability to depolymerize lignin by nonenzymatic 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.
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
The decomposition
of lignin is important
because:
Lignin is the second
most abundant organic
molecule
Lignin protects
plant tissue from
decomposition
Lignin
Remaining Mass
The Decomposition of Plant Litter
Labile Compounds
Non Lignified Cellulose
Lignified Cellulose
Lignin
Adapted from Berg (1986)
Time 
Remaining Mass
Phase regulated
by nutrient
level and readily
available carbon
Phase regulated by lignin
decomposition rate
Labile Compounds
Non Lignified Cellulose
Lignified Cellulose
Lignin
Adapted from Berg (1986)
Time 
Phase regulated
by lignin
decomposition rate
Mass Loss
Phase regulated
by nutrient
level and readily
available carbon
Adapted from Fog (1988)
Ambient
Nitrogen
Elevated
Nitrogen
Time
Human Nitrate
Deposition
Less
Lignolytic
Enzyme
Activity
Less Lignin
Decay
More
Available
NH4+
Less
Litter
Decomposition
Microbial
Nitrate Assimilation
And Turnover
Reduced
Carbon
Flow
Hypothesis
Chronic nitrate additions can suppress
the lignin-degrading activity of soil
microbial communities
Predictions
Nitrate amended soils will have:
A microbial community composition with
less fungi
Lower activity of enzymes that degrade
lignin and cellulose
7
9
12
12
(kg N ha-1 y-1)
Study
Sites
Ambient
Nitrogen
Deposition
A
Ambient N
Deposition Plus
30 kg N-NO3ha-1 y-1
B
C
D
PLOTS
Cell membranes can be used to determine
microbial community composition
Cell membrane
Lipid bilayer
Microbial
cell
Phospholipid
Phospholipid Fatty Acids
Fatty
Acids
Tails
Unique to
fungi
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
Enzyme Analysis
Extracellular Enzymes
Plant Litter Compound
Cellobiohydrolase
Cellulose
b-glucosidase
Cellulose
Peroxidase
Lignin
Phenol oxidase
Lignin
0%
Bacteria
Actinomycete
20:4 w6
18:1 w9 c
18:2 w6
20%
10me18:0
25%
18:0
17:0
a17:0
i17:0
16:1 w5 c
16:1 w9 c
15:0
a15:0
14:0
cy19:0a
18:1 w7t
18:1 w7c
cy17:0
16:1 w7c
10me16:0
i16:0
i15:0
% mol fraction
Nitrate additions had no noticeable effect
on microbial community composition
30%
N Ambient
N Amended
15%
10%
5%
Fungi
Protozoan
Total PLFA (nmol PLFA mg-1 C)
Nitrate additions decreased microbial
biomass
10
8
6
p = 0.012
4
2
0
Control
N Amended
Nitrate addition suppressed activity of soil
lignin & cellulose degrading enzymes
b-glucosidase
*
Peroxidase
*
Cellobiohydrolase
Phenol Oxidase
* p < 0.05
-40%
-30%
-20%
-10%
Change in Enzyme Activity
0%
10%
Nitrate addition suppressed activity of lignin
degrading enzymes in litter
* p < 0.05
b-glucosidase
Peroxidase
Cellobiohydrolase
*
-40%
Phenol Oxidase
-30%
-20%
-10%
Change in Enzyme Activity
0%
10%
Nitrate
Additions
Microbial
Community
Composition
No
Apparent
Change
Total PLFA
(Microbial Biomass)
Lignolytic
Activity
Decrease
Decrease
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.
Conclusions
Anthropogenic nitrate deposition
may diminish the physiological
capacity of soil microbial
communities to degrade plant
litter.
Does a suppression of lignin &
cellulose degrading enzymes
indicate a reduction in the flow of
carbon from these compounds?
Hypothesis
Nitrate additions will inhibit
the ability of soil microorganisms
to metabolize and assimilate the
products of lignin and cellulose
degradation
13C
Lignin
H
Vanillin
CHO
OH
OCH3
Microbial
Assimilation
Cellulose
13C
Cellobiose
Microbial
Assimilation
13C
Sequential Extractions:
Soil was incubated for 48 hours and 13C was
traced into respiration, dissolved organic
carbon (DOC), microbial carbon, and soil
carbon.
13C
PLFA Analysis:
Traced the flow of labeled 13C vanillin and
cellobiose into cell membranes.
13C
PLFA Analysis
Microbial
Membrane
CHO
13C
H13C
13C
13C
13C
13C
13C
OH
Extraction &
Separation
13C
OCH3
13C
Analysis
13C
250
13 C
150
excess) PLFA (nmol
100
13 C
200
6
PLFA (nmol
excess)
N additions increased the
incorporation of vanillin into PLFAs
4
Control
N Amended
*
50
0
8
*
Vanillin
*
Cellobiose
2
0
Bacteria Bacteria & Fungi
Fungi
Total
N additions did not alter the flow of
into carbon pools
% Recovery
13
C
80
60
13C
vanillin
Control
N Amended
40
20
0
Microbial
Respiration
DOC
Microbial
Soil Organic
Biomass
Matter
N additions did not alter the flow of
into carbon pools
% Recovery
13
C
80
60
13C
cellobiose
Control
N Amended
40
20
0
Microbial
Respiration
DOC
Microbial
Soil Organic
Biomass
Matter
Soil Organic Carbon (mg C g-1)
N additions increased soil organic
carbon
60
50
40
30
p < 0.001
20
10
0
Control
N Amended
Excess nitrogen likely inhibits lignocellulose degradation
more than vanillin or cellobiose degradation
Chronic
nitrate additions
UNCHANGED
Vanillin or
Cellobiose into
Carbon Pools
INCREASED
Soil Organic
Carbon
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.
Implications
Atmospheric
CO2
Pools
Northern Hardwood Forests
Slower
Decomposition
Human
Nitrogen
Deposition
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
Global Controls of Decomposition
Environmental
Conditions
Litter
Biochemistry
Plant litter decay
Environmental
Conditions
Actual
Evapotranspiration
(AET)
Slower Decomposition
Faster Decomposition
High > 1000 mm
350-550 mm
Low < 300 mm
Environmental
Conditions
Years Required to Decompose
95% of Leaf Litter
~0.5 years
Wet Tropical
~4 years
Temperate
Deciduous
~14 years
Boreal
Litter
Biochemistry
Lignin and
Nitrogen
Concentrations
Faster Decomposition
Slower Decomposition
Lower Lignin
Higher Nitrogen
Higher Lignin
Lower Nitrogen
Human
Nitrogen
Deposition
Increase Litter
Nitrogen Concentrations
Decrease the
Decomposition of Lignin
Increase Decomposition
Rates
Decrease Decomposition
Rates
Phase regulated
by lignin
decomposition rate
Mass Loss
Phase regulated
by nutrient
level and readily
available carbon
Adapted from Fog (1988)
Ambient
Nitrogen
Elevated
Nitrogen
Time
Nitrogen deposition impact on decomposition may
depend on lignin concentrations
Increased Decay
Decreased Decay
“Low”
Lignin
“High”
Lignin
Annual Decomposition Rate (%)
Lignin Control of Decay is Greater at Higher AET
As slope decreases, higher
lignin concentrations require
more energy and moisture to
cause decay.
AET
100
1000 (mm)
Slope = -1.50
75
50
500 (mm)
25
250 (mm)
Slope = -0.75
Slope = -0.40
0
0
5
Adapted from Meentemeyer (1978)
10
15
20
Lignin Concentration (%)
25
30
Anthropogenic nitrogen deposition may
have a larger impact on decomposition in
wet-tropical environments
Less
Impact
More
Impact
Summary
Nitrogen deposition has the potential to diminish the
physiological capacity of lignin-degrading
microorganisms to depolymerize lignin.
Reductions in lignocellulose-degrading enzymes and
microbial biomass suggests a reduction in energy
available for microbial metabolism
Nitrogen deposition may have a greater impact on
decomposition in wet tropical regions than arid or
cold regions