EFFECT OF HYPOXIA/ANOXIA ONN CYCLING PROCESSES IN THE WATER

COLUMN

Sasha Jenkins, Matt Hipsey, Perran Cook, Ian Waite, Alice

Gedaria, Tony O’Donnell

N cycling dynamics in estuaries

Conceptual model of current understanding of N cycle under anoxia conditions (Burgin and Hamilton, 2007)

Water column

Sediment

Prolonged hypoxic/anoxic conditionsHypoxia = dO2 ≤ 2 mg/L

Anoxia = dO2 ≤ 0.1 mg/L

Anaerobic processes may become significant in water column

Aerobic N

cycle

processes

(with O2)

Anaerobic

N cycle

processes

(w/o O2)

How frequent are anoxia/hypoxia events in the Swan Estuary?

Seasonal salinity & dissolved oxygen profiles

Winter Spring

Summer Autumn

↑rainfall

↑rainfall↑ algae

Salt wedge Salt wedge

sea upriver

What effect does this have on N cycle processes?

Possible N cycle microbial processes induced

by hypoxia/anoxia

NO2- NO N2O N2

N2O NO3-

NH4+ NH2OH NO2

- NO N2O N2

Nitrification

Nitrifier Denitrification

Denitrification

N2O?

NO2- NO N2O N2

N2O NO3-

NH4+ NH2OH NO2

- NO N2O N2

Nitrification

Nitrifier Denitrification

Denitrification

N2O?

NH4+ + NO2

- → N2 + 2 H2O

Anammox

Normally

aerobic

All pathways = N

removal

(net loss of N

from estuary)

Contribute to

total N budget

By-product nitrous oxide (N2O) =

potent Greenhouse gas (GHG)

How significant are N2O emissions from the

water column during hypoxia/anoxia events?

Anaerobic

Anaerobic

Objectives & study site

• To quantify (qPCR, process measures) N-cycle processes during anoxia/hypoxia events throughout the yr (4 seasons), 2 depths, 6 sites

• Characterise and identify key microorganism involved in these processes using molecular community analysis techniques (cloning, DGGE and T-RFLP)

• Provide better estimates of the process rates for validation of models

Abundance N cycle functional

groups (e.g. nitrifiers)– SIZE?Quantitative PCR

DGGE

Community diversity-

RESPONSE?

Id species involved- WHO?

Process measures

& Gas analysis

Sample collection

DGGE

Experimental Approach

Microbial community analysis

Cycles

Product

Microbial

Activity –

WHICH

PATHWAY?

Results Spatial/temporal variation N2O production

Temporal & spatial variation in N2O production

NAR & STJ sites

Mainly in Winter & Autumn

Absent in Summer

UpriverSea

Results Factors influencing N2O production

1. O2 concentration

N2O maxima :

O2 concentration

= 0.5-2 mg/l

Hypoxic

Dissolved O2 profile

2. Stratification

Extent of mixing

Hypoxia develops

under Salt wedge

in Winter/Spring

= N2O maxima

Salt wedge

BLA ARM NAR STJ RON SUC

N2O

N2O

3. Inorganic substrate availability

Autumn

Winter

a) NH4 (ammonium) -Nitrifiers b) NOx (nitrate/nitrite) -Denitrifiers

Winter

4. Organic substrate availability

a) DOC (carbon) –heterotrophic denitrifiers b) DON (organic N)- N source

Winter

Autumn

Upriver

sites

Results

Which N cycle microbial process is responsible for N2O production?

Possible N cycle microbial processes

responsible for N2O

NO2- NO N2O N2

N2O NO3-

NH4+ NH2OH NO2

- NO N2O N2

Nitrification

Nitrifier Denitrification

Denitrification

N2O?

NO2- NO N2O N2

N2O NO3-

NH4+ NH2OH NO2

- NO N2O N2

Nitrification

Nitrifier Denitrification

Denitrification

N2O?

Normally

aerobic

Anaerobic

Nitrifier & denitrifier abundance in winter

Denit Denit/

NitrifBoth highly

abundant

Nitrifiers

significantly

increased

compared to

other site

Nitrifiers are N2O

producers?

Nitrifier & denitrifier abundance in Autumn

Denitrifiers ?

Denit/

Nitrif ? Not,

conclusive

B A N ST R SU

Gammaproteobacteria

Community

Analysis

Experimental manipulations

• Microbial data not conclusive

• To assess the microbial response to environmental change (temperature,

salinity, dissolved oxygen) and resource inputs of C and N

• Aim: to unravel N cycle pathways

AnaerobicAnalysis

Gas & microbial analysis

at regular intervals over 0,

6, 12, 24, 48 hrs period

Parameters

Serum bottle

Air space = 1/3

Sample water (50ml)

Substrate (10 pmol)

Microbial response to environmental change & resource inputs

AnoxicHypoxicSuboxicOxic

Environmental variables: Salinity (36, 28, 22, 19 ppt), DO2 (6, 2,1, 0.5 mg/l) gradients

Substrates for N cycle processes: Control (no amendment)

NO3 (Dentification)

Treatments NO3+NH4 (Nitrification, Dentification, Anammox)

NH4 (Nitrification/Anammox)

NO3 + glucose (Heterotrophic Denitrification)

N2O production & Substrate availability

Denitrifiers

Nitrifiers

Nitrifiers/Denitrifiers

Heterotrophic Denitrifiers

C-limited

Winter/Spring N2O production

BLA ARM NAR STJ RON SUC

N2O

N2O

N2O

N2O

BLA ARM NAR STJ RON SUC

Rainfall

Salt wedge

downriver

High NH4,

NOx &

DOC

Salt wedge

migrates

upriver

Lower NH4,

NOx &

DOC

Winter

Spring

Nitrifiers ?

Nitrifiers ?

Denitrifiers ?

Summer/Autumn N2O production

BLA ARM NAR STJ RON SUC

N2O

N2O

Rainfall

Discharge

high NH4,

DOC

NH4, NOx

& DOC

virtually

depleted

Summer

AutumnDenitrifiers ?Nitrifiers ?

BLA ARM NAR STJ RON SUC

Summary

• N uptake and/or removal (via nitrification or

denitrification) are significant in water column

- contributes to N budget

• N2O maxima occurs under hypoxic conditions (O2 =

0.5- 2 mg/l)

– Nitrifiers are possibly responsible

• Increased N20 production associated with salt wedge

(winter/spring), rainfall, substrate availability (NH4,

NOx)

• Denitrification C limited - DOC & DON could promote

activity (Autumn & Winter)

Management implications

Need to develop best management practices (BMP) to mitigate N2O

• Oxygenate bottom water STJ-SUC ≥ 0.5-2 mg/l during Autumn

• Rehabilitation/stream restoration

– reduce DIN (NH4) inputs for nitrifiers during the high discharge

in winter

– reduce DON inputs for nitrifiers from agricultural & urban

catchments during summer/autumn

Acknowledgements

• Ian Waite (UWA) & Alice Gedaria for technical support

• Perran Cook (Monash University) for GCMS analysis & data analysis

of gas samples

• Jeff Cosgrove and Kerry Trawler (SRT) for comments & input

throughout project

• All crew on the lower Swan River sampling regime especially Naomi

Hellriegel from Dept of Water

• Swan River Trust for funding research