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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