Chapter 14 - Biogeochemical Cycling

Objectives• Be able to give an explanation of why biogeochemical cycles are important• Be able to explain what the GAIA hypothesis is• Be able to list three major biogeochemical changes between early and

modern earth • Be able to define the term reservoir and give an example of a small easily

perturbed reservoir and a large stable reservoir• Be able to list the three major plant polymers• Be familiar with all parts of the carbon, nitrogen, and sulfur cycles • Be able to draw each cycle and describe the microbial activities associated

with each leg of the cycles • Be able to give an example of a microbe associated with each leg of the

cycle

Elemental Breakdown

% dry mass of an E. coli cell

Major elements Carbon Oxygen Hydrogen Nitrogen Sulfur Phosphorus

Minor elements Potassium Calcium Magnesium Chlorine Iron Trace elements Manganese Molybdenum Cobalt Copper Zinc

50208

1413

20.050.050.050.2

All trace elements combined comprise 0.3% of dry weight of cell

Chemical composition of an E. coli cell

Gaia Hypothesis: earth acts like a self-

regulating superorganism

How has earth maintained conditions favorable for life? Compare atmospheres and temperatures on Earth, Venus, and Mars.

0.03%

79%

21%

1%

1.7ppm

13

98%

1.9%

0

0.1%

0

290 50

95%

2.7%

0.13%

1.6%

0

-53

96.5%

3.5%

Trace

70 ppm

0

459

Carbon dioxide

Nitrogen

Oxygen

Argon

Methane

Surface temperature 0C

Earth

with life

Earth

no lifeMarsVenusGas

Atmosphere and Temperatures found on Venus, Mars, and Earth

Biogeochemical activities are:

unidirectional on a geologic time scale

cyclical on a contemporary scale

To understand cycling of elements, the size and cycling activity level of the reservoirs of the element must be defined. atmospheric CO2 is a relatively small reservoir of carbon that is actively cycled. Such small, actively cycled reservoirs are most subject to perturbation.

H2O O2

CO2

Turnover rates3 x 10 yr2 x 10 yr2 x 10 yr

2

3

6

atmosphere

lithospherehydrosphere

Relative reservoir sizes: H2O > O2 >> CO2

The concept of a reservoir

Physical transformationsdissolutionprecipitationvolatilizationfixation

Chemical transformationsbiosynthesisbiodegradationoxidoreductive-biotransformations

What reactions drive biogeochemical cycling?

Driving force for biogeochemical cycles is sunlightEnergy Flow

Primary producers

Grazers

Predators

Predators

Dec

om

pos

ers

CO

and

min

era

ls2

100%

15%

2%

0.3%

CO 2

<0.1%The ability to photosynthesize allows sunlight energy to be trapped and stored. This is not an efficient process although some environments are more productive than others. Only 10-15% of the energy trapped in each trophic level is passed on to the next level.

Description of ecosystem

Net primary productivity

(g dry organic matter/m2/yr)

Tundra

Desert

Temperate grassland

Temperate forest

Tropical rainforest

Cattail Swamp

Freshwater pond

Open ocean

Coastal seawater

Upwelling area

Coral reef

Corn field

Rice paddy

Sugarcane field

400

200

Up to 1,500

1,200 – 1,600

Up to 2,800

2,500

950 – 1,500

100

200

600

4,900

1,000 – 6,000

340 – 1,200

up to 9,400

Net primary productivity of some natural and managed ecosystems

The Carbon CycleThe development of photosynthesis allowed microbes to tap into sunlight energy and provided a mechanism for the first carbon cycle. At the same time the carbon cycle evolved, the nitrogen cycle emerged because nitrogen was limiting for microbial growth. Although N2 was present, it was not in a usable form for microbes.

Aerobic Anaerobic

Fossil fuels

FermentationPhotosynthesis

Respiration

Methanogenesis

CO + H O2 2 O + CH O2 2 Alcohols, acids,H + CO2 2

CH 4

CH O2

Carbon Reservoir Metric tons carbon

Actively cycled

Atmosphere

CO2

Ocean

Biomass

Carbonates

Dissolved and particulate organics

Land

Biota

Humus

Fossil fuel

Earth’s crust

6.7 x 1011

4.0 x 109

3.8 x 1013

2.1 x 1012

5.0 x 1011

1.2 x 1012

1.0 x 1013

1.2 x 1017

Yes

Yes

No

Yes

Yes

Yes

Yes

No

Global Carbon Reservoirs

The carbon cycle is a good example of one that is undergoing a major perturbation due to human activity.

Human activity has had a large impact on the atmospheric CO2 reservoir beginning with industrialization. As a result, the level of CO2 in the atmosphere has increased 28% in the past 150 years.

Carbon source metric tons carbon/yrRelease by fossil-fuel combustion 7 x 109

Land clearing 3 x 109

Forest harvest and decay 6 x 109

Forest regrowth -4 x 109

Net uptake by oceans -3 x 109

Annual flux 9 x 109

Natural sources of CO2

• respiration• ocean degassing• terrestrial degassing• wildfires

Anthropogenic sources of CO2

• fossil fuel combustion• cement production• land use changes

Natural sinks for CO2

• terrestrialuptake by plantsuptake by soils

• oceanic partitioningbiomass production

Anthropogenic sinks for CO2

• chemical production• biological materials

Natural and anthropogenic CO2 sources and sinks

CO2

(ppm)

CH4

(ppm)

N2O

(ppm)

SF6

(ppt)

PFC

(ppt)

Preindustrial

1992

278

356

0.700

1.714

0.275

0.311

0

32

0

70

Atmospheric Lifetime

(years)

50-200 12 120 3,200 50,000

Global Atmospheric Concentrations of Selected Greenhouse Gases

CO2 is not the only problem!

CH4 is 22 times stronger as a greenhouse gas than CO2

The term reservoir can be used on a global scale or on a smaller scale such as a habitat.

How does carbon cycle within a habitat?

Macro vs. microorganisms

simple vs. simple to complex substrates

aerobic vs. aerobic/anaerobic redox conditions

What are the major carbon inputs into the environment?

plant materials (through photosynthesis)cellulose 15 – 60% hemicellulose 10-30%lignin 5- 30%protein/nucleic acids 2-15%

fungal cell walls/arthropodschitin

Carbon cycling on the habitat scale

nGlucosesubunit

G lucose subunits 1 - 4 linked

Molecular weightup to 1.8 x 10 6

Cellulose

1-4 exoglucanase 1-4 endoglucanase

1-4 glucosidase (cellobiase)

+

(shorter pieces)

Cellobiose(can be transported into cell)

Transport across membrane

AerobicTCA cycle

Anaerobic

Fermentation

Cellu lose

Glucose

Cellulose degradation begins outside the cell with a set of three exoenzymes:

β-1,4- endoglucanseβ-1,4- exoglucanaseβ-1,4- glucosidase

n

Molecular weight~ 40,000

Galacturonicacid

Methylatedgalacturonic acid

Hemicellulose

Amino linkage Acetylgroup

Chitin

For the more complex polymers such as lignin a variety of oxidizing enzymes are used. A specific example is the combination of lignin peroxidase and oxidase which produce H2O2 to aid in degradation of lignin.

Lignin due to its complexity is generally degraded much more slowly than cellulose or hemicellulose.

0 100 200Days

Per

cent

age

rem

ain

ing

L ignin

Wheat straw

Cellu lose

Hemicellulose

Succinicacid Aceticacid - oxyadipic acid-carboxy-cis, cis-muconic acidProtocatechuicacidVanillin Vanillic acidConiferylalcoholConyferylaldehyde

Ferulicacid Caffeic acidAdjacent hydroxylgroups allow ringcleavage+ TCA CO + H O2 2

Lignin polymerExtracellular enzymesLignin monomers (transported into the cell)Other phenols andvarious portionsof lignin molecules

Succinicacid

Aceticacid

- oxyadipic acid

-carboxy-cis, cis-muconic acid

Protocatechuicacid

Vanillin

Vanillic acid

Coniferyla lcohol

Conyferyla ldehyde

Ferulicacid

Caffeic acid

Adjacent hydroxylgroups allow ring

cleavage

+

TCA CO + H O2 2

Lignin polymer

Extracellular enzymes

Lignin monomers (transported into the cell)

Other phenols andvarious portions

of lignin molecules

The most complex organic polymer found in the environment is humus. Formation of humus is a two-stage process that involves the formation of reactive monomers during the degradation of organic matter, followed by the spontaneous polymerization of some of these monomers into the humus molecule.

Ultimately, these large polymers are degraded and produce new cell mass, CO2 (which returns to the atmosphere), and contribute to the formation of a stable organic matter fraction, humus. Humus turns over slowly, at a rate of 3 to 5% per year.

In addition to mineralization to CO2, a number of small carbon molecules are formed largely as a result of anaerobic activities and in some instances as a result of anthropogenic activity. These include:

Methane generationThe methanogens are a group of obligately anaerobic Archaea that can reduce CO2 to methane (use CO2 as a terminal electron acceptor) both chemoautotrophically or heterotrophically using small MW molecules such as methanol or acetate.

4H2 + CO2 CH4 + 2H2O G0 = -130.7 kJ

Although much methane is microbially produced, there are other sources as well. What happens to the methane? This is of concern because methane is a greenhouse gas 22 times more effective than CO2 in trapping heat.

Estimates of methane released into the atmosphere

S o urc e M e tha n e e m iss io n(1 06 m e tric to n s /ye a r)

B io ge n ic R u m ina n ts T e rm ite s Pa d d y f ie ld s N a tura l w e t la nd s L a nd fil ls O ce a n s a n d la ke s T u nd ra

A b io ge n ic C o a l m in ing N a tura l g a s fla rin g a nd v e nt in g Ind us t ria l a nd p ip e lin e losse s B io m a ss b u rn ing M e th a ne h yd ra te s V o lca n o es A uto m o b ile s

T o ta l T o ta l b io ge n ic T o ta l a b io ge n ic

80 - 1 0025 - 1 5070 - 1 20

1 20 - 20 05 - 7 01 - 2 01 - 5

10 - 3510 - 3515 - 4510 - 40

2 - 40.50.5

3 49 - 82 03 02 - 66 548 - 1 55

8 1 - 8 6 % o f to ta l1 3 - 1 9 % o f to ta l

Anthropogenic 190 – 405 54 - 49% of total

Methane utilization

In most environments, the methane produced is utilized by methanotrophic microbes as a source of carbon and energy. The first enzyme in the biodegradation pathway of methane is methane monooxygenase (MMO). This enzyme is of interest because it can aid in the degradation of highly chlorinated materials such as TCE (trichloroethylene). The oxidation of TCE does not provide energy for the microbe, it is simply a result of nonspecific catalysis by the MMO enzyme. This is also called cometabolism.

CH4 + O2 CH3OH HCHO HCOOH CO2 + H2O

methanol formaldehyde formic acid

MMO

Carbon monoxide- a highly toxic molecule that is produced largely as a result of fossil fuel burning and photochemical oxidation of methane in the atmosphere. Despite the fact that this is a highly toxic molecule, some microbes can utilize is as a source of energy.

In summary, there is huge variety in the types of carbon-containing molecules found in the environment. Similarly microbes have developed an equal variety in their metabolic approaches to deriving carbon and energy from these compounds.

CO CO2

CO CO2

CO CO2

The Nitrogen CycleN is cycled between: NH4

+ (-3 oxidation state) and NO3- (+5 oxidation state)

Nitrogen Reservoir Metric tons nitrogen Actively cycled

Atmosphere

N2

Ocean

Biomass

Soluble salts (NO3, NO2-, NH4

+)

Dissolved and particulate

organics

Dissolved N2

Land

Biota

Organic matter

Earth’s crust

3.9 x 1015

5.2 x 108

6.9 x 1011

3.0 x 1011

2.0 x 1013

2.5 x 1010

1.1 x 1011

7.7 x 1014

No

Yes

YesYes

No

Yes

Slow

No

Global Nitrogen Reservoirs

Biological inputs of nitrogen from N2 fixation

land - 135 million metric tons/yr (microbial)

marine - 40 million metric tons/yr (microbial)

fertilizers - 30 million metric tons/yr (anthropogenic)

Nitrogen must be fixed before it can be incorporated into biomass. This process is called nitrogen fixation.

The enzyme that catalyzes nitrogen fixation is nitrogenase.

N2 fixing system

Nitrogen fixation

(kg N/hectare/yr)

Rhizobium-legume

Anabaena-Azolla

Cyanobacteria-moss

Rhizosphere assoc.

Free-living

200-300

100-120

30-40

2-25

1-2

Rates of Nitrogen Fixation

1-2 kg N/hec/yr 2- 25 kg/N/hec/yr

Free-living bacteria must also protect nitrogenase from O2

complex is membrane associated

slime production

high levels of respiration

conformation change in nitrogenase when O2 is present

Azotobacter - aerobic

Beijerinckia - aerobic, likes acidic soils

Azospirillum - facultative

Clostridia - anaerobic

Examples of free-living bacteria:

Summary for nitrogen fixation:

energy intensive

inhibited by ammonia

occurs in aerobic and anaerobic environments

end-product is ammonia

nitrogenase is O2 sensitive

Fate of ammonia (NH3) produced during nitrogen fixation

plant uptake

microbial uptake

adsorption to colloids (adds to CEC)

fixation within clay minerals

incorporation into humus

volatilization

nitrification

} assimilation and mineralization

NH3 is assimilated by cells into:

proteins

cell wall constituents

nucleic acids

Ammonia assimilation and ammonification

Release of assimilated NH3 is called ammonification. This process can occur intracellularly or extracellularly

proteases

chitinases

nucleases

ureases

- -

- -

+

+

H O2

glutamatedehydrogenase

NAD NADH

= O

glutamate - ketoglutarate

+ NH 3

NH 3

A

At high N concentrations

-

-

-

-

-

-

+

+

+

= O

glutamate

glutamate

glutam ine - ketoglutarate

NH 3

NH 3

NH 3

NH 2

ATP

ADP + Pi

glutam ine synthetase

Ferredoxin

2H+

2e -

glutamate-synthase

(GOGAT)

Transamination

B

At low N concentrations

Summary for ammonia assimilation and ammonification

Assimilation and ammonification cycles ammonia between its organic and inorganic forms

Ammonification predominates at C:N ratios < 20

Assimilation predominates at C:N ratios > 20

Fate of ammonia (NH3) produced during nitrogen fixation

plant uptake

microbial uptake

adsorption to colloids (adds to CEC)

fixation within clay minerals

incorporation into humus

volatilization

nitrification

Nitrification - Chemoautotrophic aerobic process

Nitrosomonas NitrobacterNH4

+ NO2- NO3

-

Nitrosomonas: 34 moles NH4

+ to fix 1 mole CO2

Nitrobacter: 100 moles NH4

+ to fix 1 mole CO2

Summary for nitrification

Nitrification is an chemoautotrophic, aerobic process

Nitrification in managed systems can result in nitrate leaching and groundwater contamination

Nitrification is sensitive to a variety of chemical inhibitors and is inhibited at low pH. (There are a variety of nitrification inhibitors on the market)

Nitrification is important in areas that are high in ammonia (septic tanks, landfills, feedlots, dairy operations, overfertilization of crops). The nitrate formed is highly mobile (does not sorb to soil). As a result, nitrate contamination of groundwater is common. Nitrate contamination can result in methemoglobenemia (blue baby syndrome) and it has been suggested (not proven) that high nitrate consumption may be linked to stomach cancer.

What is the fate of NO3- following nitrification?

accumulation (disturbed vs. managed)

fixation within clay minerals

leaching (groundwater contamination)

dissimilatory nitrate reduction• nitrate ammonification• denitrification

plant uptake

microbial uptakebiological uptake (assimilatory nitrate reduction)}

Assimilatory nitrate reductionmany plants prefer nitrate which is reduced in the plant prior to use however, nitrogen in fertilizer is added as ammonia or urea.

assimilatory nitrate reduction is inhibited by ammonium

nitrate is more mobile than ammonium leading to leaching loss

microorganisms prefer ammonia since uptake of nitrate requires a reduction step

Dissimilatory nitrate reduction

Dissimilatory reduction of nitrate to ammonia (DNRA)

use of nitrate as a TEA (anaerobic process) – less energy produced

inhibited by oxygen

not inhibited by ammonium

found in a limited number of carbon rich environments

stagnant watersewage plantssome sediments

Denitrification

use of nitrate as a TEA (anaerobic process) – more energy produced

many heterotrophic bacteria are denitrifiers

produces a mix of N2 and N2O

inhibited by oxygen

not inhibited by ammonium

100 80 60 40 20 0

0 20 40 60 80 100

RumenDigested sludge

Estuarine sediments

Lake sediments

Soil + C

Soil

% NH (Dissimilatory reduction)4+

C/e

- ac

cep t

or(r

ela

tive

scal

e )

% N (Denitrification)2

NO 3

-

NO 3

-

NO 2

-

NO 2

- NO N O2 N 2

N 2N O2NONO 2

-

Cytoplasm

Periplasm

Innermembrane

Outermembrane

Outside cell

nitratereductase

nitrite reductase

nitric oxidereductase

nitrous oxidereductase

Denitrification requires a set of 4 enzymes:

nitrate reductase

nitrite reductase

High [NO3-] favors N2 production

Low [NO3-] favors N2O production

nitric oxide reductase

nitrous oxide reductase

Denitrification

NO, N2O deplete the ozone layer

Reaction of N2O with ozone

O2 + UV light O + O

O + O2 O3 (ozone generation)

N2O + UV light N2 + O*

N2O + O* 2NO (nitric oxide)NO + O3 NO2 + O2 (ozone depletion)NO2 + O* NO + O2

returns fixed N to atmosphere:

get formation of NO, N2O

NO3 NO N2O N2

Summary for nitrate reduction

Nitrate assimilated must be reduced to ammonia for use.

Oxygen does not inhibit this process

Nitrate assimilation is inhibited by ammonia

1. Assimilatory nitrate reduction

2. Dissimilatory nitrate reduction to ammonia (DNRA)

Anaerobic respiration using nitrate as TEA

Inhibited by oxygen

Limited to a small number of carbon-rich, TEA poor environments

Fermentative bacteria predominate

3. Dissimilatory nitrate reduction (denitrification)

Anaerobic respiration using nitrate as TEA

Inhibited by oxygen

Produces a mix of N2 and N2O

Many heterotrophs denitrify

Sulfur Cycle

10th most abundant element

average concentration = 520 ppm

oxidation states range from +6 (sulfate) to -2 (sulfide)

Sulfur Reservoir Metric tons sulfur

Actively cycled

Atmosphere SO2/H2S

Ocean Biomass Soluble inorganic ions (primarily SO4

2- )

Land Biota Organic matter Earth’s crust

1.4 x 106

1.5 x 108

1.2 x 1015

8.5 x 109

1.6 x 1010

1.8 x 1016

Yes

Yes

Slow

Yes

Yes

No

Global Sulfur Reservoirs

1. Assimilatory sulfate reduction

The form of sulfur utilized by microbes is reduced sulfur. However, sulfide (S2-) is toxic to cells. Therefore sulfur is taken up as sulfate (SO4

2-), and in a complex series of reactions the sulfate is reduced to sulfide which is then immediately incorporated into the amino acid serine to form cysteine.

Sulfur makes up approx. 1% of the dry weight of a cell. It is important for synthesis of proteins (cysteine and methionine) and co-enzymes.

Assimilatory sulfate reduction (requires a reduction of SO42- to S2-)

SO42- + ATP APS + Ppi

adenosine phosphosulfate

APS + ATP PAPS + ADP 3’ – phosphoadenosine – 5-phosphosulfate

PAPS + 2e- SO32- + PAP

SO32- + 6H+ + 6e- S2-

S2- + serine cysteine + H2O

Sulfur Mineralization

SH – CH2- CH - COOH + H2O

NH2

- OH – CH2- CH – COOH + H2S

NH2

-

terrestrial environments

cysteine serine

marine environments

algae dimethylsulfoniopropionate Dimethylsulfide (DMS)

At a C:S ratio < 200:1, sulfur mineralization is favored

At a C:S ratio > 400:1, sulfur assimilation is favored

Sulfide oxidation (nonbiological)

H2S and DMS are photooxidized to SO42- in the atmosphere

Normal biological production = 1 kg SO4/ha/yr

Rural production = 10 kg SO4/ha/yr

Urban production = 100 kg SO4/ha/yr

acid rain – pH < 5.6

Both the H2S and the DMS generated during sulfur mineralization are volatile and therefore significant amounts are released to the atmosphere. Here they are photooxidized to sulfate.

SO42- + water H2SO4 (sulfuric acid)

fossil fuel burning releases SO2 H2SO3 (sulfurous acid)

Aerobic sulfur oxidation

H2S + 1/2O2 S0 + H20 G = -50.1 kcal/mol

Chemolithotrophic bacteria

Beggiatoa

Thioplaca

Thiothrix

Thermothrix

Thiobacillus

H2S not released to the atmosphere acts as substrate for sulfur-oxidizers.

Under aerobic conditions:

What unusual community is based on the chemoautrophic sulfur oxiders?

What is the conundrum for these organisms?

Most of these microbes deposit S0 as granules inside the cell. They can further oxidize S0 but this is not preferred. However, there are some sulfur oxidizers most notably Thiobacillus thiooxidans that are acidophilic and prefer to oxidize S0 to SO4

2-.

0 0.32 0.64 0.96 1.28 1.6

2.0

2.4

2.8

3.2

O 2

H S2

Dep

th (

mm

)

D issolved O (mg/l)2

Air

Beggiatoa

Mineral mediumwith 0.2% agar

M ineral mediumwith 1.5% agar1 - 8 mM Na S2

Beggiatoa

Acidothiobacillus - obligate aerobesacid intolerant spp.

Acidophilic sulfur-oxidizers:

H2S + 1/2O2 S0 + H2O

acid tolerant spp.

S0 + 3/2O2 + H2O H2SO4

G = -149.8 kcal/mol

All sulfur oxidizers are aerobic with the exception of:

Acidothiobacillus denitrificans - uses nitrate as TEA

4NO3- + 3S0 3SO42- + 2N2

Phototrophic oxidation

anaerobic photoautotrophic process:

Chromatium

Ectorhodospirillum

Chlorobium

Under anaerobic conditions, H2S is utilized by photosynthetic bacteria:

CO2 + H2S C(H2O) + S0

Anaerobic photosynthesis

CO2 + H2O C(H2O) + O2

Aerobic photosynthesis

Green and purple sulfur bacteria

Summary - Consequences of Sulfur Oxidation

• Solubilization and leaching of minerals, e.g., (phosphorus) due to decreased pH

• Acid mine drainage

• Acid rain

Dissimilatory sulfate reduction and sulfur respiration

Heterotrophic reduction of sulfur 1. respiratory S0 reduction

2. dissimilatory SO42- reductionanaerobic

heterotrophic

limited number of electron donors (substrates) lactic acid pyruvic acid H2

small MW alcohols

Desulfuromonas acetoxidans CH3COOH + 2H2O + 4S0 2CO2 + 2H2S

Desulfovibrio

DesulfotomaculumH2 + SO4

2-H2S + 2H2O- + 2OH-

Example of a heterotrophic sulfate reducer:

Examples of autotrophic sulfate reducers:

Summary - Sulfate Reduction:

• inhibited by oxygen

• can result in gaseous losses to atmosphere

• produces H2S which can result in anaerobic corrosion of steel and iron set in sulfate-containing soils

Winogradsky column – great illustration of sulfur cycling

Set up:

Soil is mixed with 1 g CaCO3, 1 g CaSO4, and shredded paper (cellulose). Soil is added to a column and saturated with water. A soil-water slurry is poured on top of this layer to the desired thickness.

Column is incubated under lights or in a window.

Initial conditions – aerobic, but O2 is used up quickly – aerobic

chemoheterotrophs

Second population – anaerobic, chemoheterotrophs ferment cellulose to low molecular weight fatty acids and alcohols

Third population – anaerobic, chemoheterotrophs respire the low molecular weight fatty acids and alcohols using SO4 as the TEA.

SO4 H2S (black) + CO2 Sulfate reducers

Fourth population – anaerobic, photoautotrophs photosynthesize using H2S and CO2.

CO2 + H2S S0 + C(H2O) Green and purple sulfur

bacteria

Population development

9/26/03

9/19/039/12/03

9/5/0310/2/03

10/17/03