MERCURY, SULFUR-METABOLIZING BACTERIA AND ORGANIC MATTER IN
THE SEDIMENTS OF SUBARCTIC KUSAWA LAKE, YUKON

by

Jocelyn Anne Joe-Strack
B.Sc. University of Victoria, 2006

THESIS SUBMITTED IN PARTIAL FULFILLMENT OF
THE REQUIREMENTS FOR THE DEGREE OF
MASTER OF SCIENCE
IN
NATURAL RESOURCES AND ENVIRONMENTAL STUDIES
(GEOGRAPHY)

UNIVERSITY OF NORTHERN BRITISH COLUMBIA
April 2015

© Jocelyn Anne Joe-Strack, 2015

UMI Number: 1526529

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Abstract

Abstract

Recent studies of Arctic and Subarctic environments have detected rising levels of
natural and anthropogenic mercury (Hg), putting northern residents at risk for Hg exposure.
W ithin lake sediments, Hg can be methylated by certain species of Sulfate-Reducing
Bacteria (SRB), a subset of Sulfur-Metabolizing Bacteria (SMB). This research assessed the
controls of Subarctic SRB Hg-methylation in proglacial Kusawa Lake, Yukon, Canada.
Kusawa was found to be oligotrophic, with very low primary productivity and an orthograde
oxygen profile, conditions that inhibit Hg-methylation. In addition, the SMB proportion of
total bacteria was small (1 .9 x l0 '3 %), no known SRB Hg-methylators w ere detected, the
total Hg sediment concentration was 0.022 ± 0.0009 p,gg-1 (±SE) and methylm ercury was
undetectable. The results support previous research that suggests the factors influencing
SRB Hg-methylation in Kusawa Lake are: (i) the rate of algal-derived Hg-scavenging, (ii) the
sediment concentration of total Hg and (iii) the diversity of sediment SRB.

Frontispiece: Kusawa Lake, Arc M ountain, near sample site, March 2010. (Photo credit: Pat
Roach)

ii

Contents
Abstract............................................................................................................................................ i
Contents.........................................................................................................................................iii
List of Tables................................................................................................................................ vii
List of Figures..............................................................................................................................viii
Acronym s..................................................................................................................................... xii
Acknow ledgem ents..................................................................................................................xiii
Chapter 1

Introduction.............................................................................................................1

1.1

Introduction........................................................................................................................1

1.2

Research Objectives.......................................................................................................... 4

Chapter 2
2.1

Study Area, Kusawa Lake, Yukon........................................................................6

Creation and Physical Characteristics.............................................................................. 6

2.1.1

Landscape and Sub-basins.................................................................................................. 6

2.1.2

Glacial H is to ry.......................................................................................................................6

2.1.3

Bathym etry.............................................................................................................................9

2.1.4

Core Sample Location.......................................................................................................... 9

2.2

Climate History................................................................................................................. 12

2.2.1

T e m p e ra tu re .......................................................................................................................12

2.2.2

Precipitation........................................................................................................................ 14

2.2.3

Takhini River Discharge.................................................................................................... 15

2.2.4

Regional Disturbances.......................................................................................................16

2.3

2.2.4.1

Torrent Systems..........................................................................................................16

2 .2.4.2

Forest Fires..................................................................................................................16

Vegetation....................................................................................................................... 17

Chapter 3

Limnology and Sedimentology of Proglacial Kusawa L a k e ...................... 18

3.1 Introduction......................................................................................................................18
3.1.1

Glacial Contributions.........................................................................................................18

3.1.2

Terrestrial Contributions..................................................................................................19

3.1.3

Fluvial Transport and S to rag e........................................................................................ 20

3.1.4

Lacustrine Processes......................................................................................................... 21

3.1.4.1

Stratification............................................................................................................... 21

3.1.4.2

River and stream inflow............................................................................................22

3.1.4.3

W ind and currents..................................................................................................... 23

3.1.5

Varves....................................................................................................................................24

iii

Contents
3.1.6

Lead-210 and Caesium-137 Dating and Sedimentation R ates.................................25

3.1.7

Trace M e ta ls......................................................................................................................... 26

3.1.7.1

3.2

Redox Conditions.........................................................................................................27

Methods...........................................................................................................................28

3.2.1

Core Sam pling...................................................................................................................... 28

3.2.2

Aerial Photos........................................................................................................................ 29

3.2.3

Limnology.............................................................................................................................. 30

3.2.4

Lead-210 and Caesium-137 Dating and Sedimentation R ate................................... 30

3.2.5

Particle S ize...........................................................................................................................31

3.2.6

Trace M e ta ls......................................................................................................................... 32

3.3

Results and Discussion....................................................................................................33

3.3.1

Aerial Photos........................................................................................................................ 33

3.3.2

Limnology...............................................................................................................................35

3.3.3

Lead-210 and Caesium-137 Dating and Sedimentation R ate................................... 38

3.3.3.1

Lead-210 and Caesium-137 D a tin g ........................................................................ 38

3.3.3.2

Sedimentation R ate.................................................................................................... 4 0

3.3.3.3

1911 Sedimentation A n o m aly................................................................................. 41

3.3.4

Particle S ize........................................................................................................................... 42

3.3.5

Trace M e ta ls......................................................................................................................... 43

3.3.5.1

Core A lignm ent............................................................................................................ 44

3 .3.5.2

Trace M e ta l Distribution............................................................................................ 44

3.3.5 .3

Redox conditions revealed by trace m etal reductions....................................... 46

Chapter 4
4.1

Rock-Eval Pyrolysis of Organic M a tte r in Kusawa Lake Sedim ents

48

Introduction.................................................................................................................... 48

4.1.1

Sediment-Associated Organic M a tte r ............................................................................ 48

4.1.2

Organic M a tte r Applications............................................................................................ 50

4.2

4.1.2.1

Rock-Evat Pyrolysis..................................................................................................... 51

4.1.2.2

Rock-Eval Pyrolysis and Algal-Derived Organic M a tte r ..................................... 53

Methods........................................................................................................................... 53

4.2.1

4.3

Rock-Eval Pyrolysis.............................................................................................................. 53

Results and Discussion................................................................................................... 55

4.3.1

TOC, RC and PC Fractions.................................................................................................. 55

4.3.1.1

S I and S 2 .......................................................................................................................55

4.3.2

HI versus OICO...................................................................................................................... 58

4.3.3

Organic M a tte r and Particle Size..................................................................................... 60

4.4

Summary of Rock-Eval Pyrolysis.................................................................................. 62

Chapter 5

Total Bacteria and Sulfur M etabolizing Bacteria in Kusawa Lake

Sediments

................................................................................................................................. 63

5.1

Introduction.................................................................................................................... 63
iv

Contents
5.1.1

Review of Bacterial M etabolism .....................................................................................63

5.1.2

Sulfur-Metabolizing Bacteria.......................................................................................... 65

5.2

5.1.2.1

Sulfur Origin...................................................................................................................65

5 .1.2.2

Sulfur Cycling................................................................................................................ 66

5 .1.2.3

Organic M a tte r M etabolism ......................................................................................68

5.1.2.4

Sulfur cycling in Oligo trophic Lakes.........................................................................71

5.1.2.5

M e ta l Ion M etab olism ................................................................................................ 72

5.1 .2.6

Phytogeny.......................................................................................................................72

Methods............................................................................................................................ 75

5.2.1

Nucleic Acid Extraction..................................................................................................... 75

5.2.2

Sequence Analysis of DsrAB............................................................................................76

5.2.3

Quantification of DsrA...................................................................................................... 79

5.2.4

Quantification of 16S rD N A ............................................................................................. 80

5.2.2

qPCR Calculations and Analysis...................................................................................... 81

5.3

5 .2 .2 .1

qPCR Calculations........................................................................................................ 81

5 .2.2.2

qPCR Analysis and Statistics......................................................................................83

Results and Discussion.................................................................................................... 85

5.3.1

Total Bacteria Q u an tification ......................................................................................... 85

5.3.1.1
5.3.2

Sulfur-Metabolizing Bacteria Quantification...............................................................87

5 .3.2.1
5.3.3

Total Bacteria and Particle Size................................................................................ 86
Sulfur Bacteria and Particle Size.............................................................................. 89

Sulfur-Metabolizing Bacteria Diversity......................................................................... 89

5 .3.3.1

M axim um Likelihood Tree......................................................................................... 90

5.3.4

Outcomes of Phylogeny................................................................................................... 93

Chapter 6

M ercury in Kusawa L a k e ...................................................................................95

6.1

Introduction..................................................................................................................... 95

6.1.1

Origins of Mercury in the Y u k o n ....................................................................................95

6.1.2

Long-Range Atmospheric Transport.............................................................................. 98

6.1.3

Terrestrial and Aquatic Transport and Fates.............................................................. 99

6.1.3.1

Algal-Derived Organic M a tte r Hg Scavenging....................................................100

6.1.4

M ercury M ethylation and Sulfate-Reducing Bacteria............................................ 101

6.1.5

Biochemical Controls of M ercury M e th y la tio n ........................................................ 103

6.1.6

Bioaccumulation, Biomagnification and T o xicity.................................................... 105

6.2

Methods.......................................................................................................................... 108

6.2.1

6.3

Total M ercury and M ethylm ercury Q uantification.................................................108

Results and Discussion...................................................................................................108

6.3.1

Total M ercu ry................................................................................................................... 108

6.3.2

M ethylm ercury................................................................................................................. 109

6.3.3

Total M ercury and Particle Size....................................................................................110

6.3.4

Total Mercury and Organic M a t t e r .............................................................................I l l

v

Contents
6.3.5

Total Mercury and Total Bacteria................................................................................ 113

6.3.6

Total M ercury and Sulfur-Metabolizing B acteria.................................................... 115

6.3.7

Summary of Sediment, Organic M a tte r and BacterialInteractions w ith M ercury.
...............................................................................................................................................117

6.4

Mercury in Kusawa Lake and the Implications of Climate Change.........................118

6.4.1

Increased Allochthonous Sediment and Organic M a t te r ...................................... 118

6.4.2

Increased Primary Productivity.................................................................................... 119

6.4.3

SRB Community Composition....................................................................................... 119

Chapter 7

Conclusion.......................................................................................................... 121

7.1

Limitations and Future W ork.......................................................................................125

References................................................................................................................................. 127
Appendix: Trace M etal Profiles............................................................................................140

List of Tables

List of Tables
Table 2.1: Overview of Kusawa Lake sub-basins. Glacial cover is as of 2004 (Gilbert &
Desloges, 2 0 0 5 )................................................................................................................................... 7
Table 2.2: Description of bathym etry of five regions in Kusawa Lake starting from head of
the lake at Kusawa River (region V) to outlet at Takhini River (region I) based on
acoustic profiles collected and characterized by Gilbert & Desloges (2005) and Chow
(20 09 )................................................................................................................................................... 11
Table 3.1: Comparison of calculated year vs. depth and sedimentation rates for the original
2010 210Pb profile th at was adjusted to the 2005 core taken from ~700 m west of the
2010 core............................................................................................................................................ 40
Table 3.2: Down core particle size and % average sediment size class distributions

42

Table 4.1: Overview of O M fractions released and measured from Rock-Eval Pyrolosis 6
(version 6 ) th at are measured as TOC per g of dry weight sediment. TOC (Total organic
carbon) = PC (pyrolysable carbon) + RC (residual carbon) and PC=S1+S2 +S3 C02 +S3 C0
(Carrie et al., 20 12 )........................................................................................................................... 52
Table 4.2: Comparison of PC components as percentages of TOC and PC..................................57
Table 5.1: Overview of redox potentials
reduction cycle, where

of reduction intermediates of the sulfate

are at pH 7 (Thauer et al., 2007).................................................. 66

Table 6.1: Speciation and the common measurements of Hg in environm ental samples.
Adapted from (Chetelat et al., 2012)........................................................................................... 95

vii

List of Figures

List of Figures
Figure 2.1: Overview of southwest Yukon drainage systems and topology surrounding
Kusawa Lake. Regions over 1000 m a.s.l. are shaded in grey and spot elevations are
identified for valley floors and lake surfaces (Gilbert & Desloges, 2 0 0 5 )............................. 7
Figure 2.2: Overview of the Takhini River drainage basin and Kusawa Lake sub-drainage
basins. The extent of glacial ice as of 2004 is shown in grey (Chow, 2 0 0 9 )........................ 8
Figure 2.3: Bathymetry of Kusawa Lake determ ined from the July 2004 acoustic survey
completed by Gilbert & Desloges (2005). Isobath interval is 20 m (solid lines) and 10 m
(broken lines) below the normal summer w ater level. Maxim um depths are indicated
and acoustic transects are shown as horizontal lines. The velocity of sound in w ater
was assumed to be 1430 m /s (corresponding to 5 .5 ’C).......................................................... 10
Figure 2.4: Acoustic section of region IV in Kusawa Lake. The perspective is looking down
lake, to the North.

Location of transect is shown in Figure 2.3 as a horizontal line

across region IV (Gilbert & Desloges, 2 0 0 5 )............................................................................... 11
Figure 2.5: Average annual air tem perature at W hitehorse Airport, located ~80km northeast
of Kusawa Lake, from 1905-2010.

The dash-dot regression line indicates a 2.14‘ C

tem perature increase from 1970-2010, r2=0.229, p<0.01.

(Environment Canada,

2 0 1 4 a )................................................................................................................................................. 13
Figure 2.6: Average monthly discharge of the Takhini River near W hitehorse from the W ater
Survey of Canada (09AC001) from 1948-2012.

Daily flows are shown for the tw o

highest discharge years in 1948 and 2000 and minimal year in 1970 (Environment
Canada, 2014b ).................................................................................................................................. 15
Figure 2.7: Particles of char associated with frequency of wildfire in the region determ ined
by a core taken in 2005 where frequent wildfires w ere observed from 1917-1954
(adapted from Stern et al., 2 0 0 9 )................................................................................................. 17
Figure 3.1: Overview of the 210Pb cycle and decay of 238U to supported (sediment) and
unsupported (atmospheric) 222Rn decay sources..................................................................... 25
Figure 3.2: Kusawa Lake core sampling field trip in March 2010 showing the KB gravity corer
at the sample site............................................................................................................................. 29
Figure 3.3: Aerial photos taken in, a: August 2007, b: July 1987, c: August 1972/1 979, and d:
1948 unknown tim e of year. The w hite circle identifies the sample site, located within
the sediment plume from the Upper-Takhini and Kusawa Rivers........................................34
Figure 3.4: Graph of under ice limnology measurements taken on March 2 9 /2 0 1 2 by P.
Roach (unpublished) at four of the five bathymetric regions of Kusawa Lake. The max

List of Figures
depth at regions I, III, IV and V are 21 m, 58 m, 126 m and 104 m, respectively (a:
tem perature; b: pH and c: dissolved oxygen)........................................................................... 36
Figure 3.5a: Alignment of 2005 and 2010 137Cs and 210Pb profiles to compare and interpolate
differences. The estimated dates w ere determ ined by linear alignment betw een peaks
at 1 9 9 9 /1 9 9 6 ,1 9 7 2 /1 9 6 4 and 1 9 2 5/1 911 for the 2 0 1 0 /2 0 0 5 cores, respectively

39

Figure 3.5b: Estimated 210Pb downcore activities anchored to the 137Cs peak at 1963 with
accumulated dry weight and depth of sediment. The annual linear sedimentation rate
is 0.092 cm /yr.................................................................................................................................... 39
Figure 3.6: Sedimentation rate determ ined by adjusted 210Pb dating profile and Equation 3.1.
Note the anomalous sedimentation event at 1911................................................................. 41
Figure 3.7: Downcore Percent Sediment Size Class (%) variations.

Note that the sample

consists predom inately of clays and silt......................................................................................43
Figure 3.8: PCA analysis of downcore trace metal distribution. Note shift betw een 5-7 cm,
likely associated w ith enrichment of redox sensitive metals, 9-10 cm, likely associated
with the 1911 sedimentation event and 26 cm, likely associated with the W hite River
Ash layer..............................................................................................................................................45
Figure 3.9: Determ ination of down core redox values based on M n and Fe reduction at 5.5
cm and 7 cm, respectively.

The hypothesized trend in oxygen and hydrogen sulfide

levels are shown as dotted sigmoidal lines as suggested by Schaller (1997) and
Manceau (1 9 92 )................................................................................................................................ 47
Figure 4.1: Overview of Rock Eval variables measured from Core B (sub-sectioned in 0.5 cm
increments from 0-9.5 cm and 1 cm increments from 10-30 cm).

Rock Eval fractions

are arranged as a matrix that is reflective of the relationship betw een them , where: a:
TOC is a measurem ent of Rock Eval at, b: maximum tem perature of pyrolysis;TOC = c:
RC + d: PC;PC= e: S I + f: 52+ S3C02 (not shown)+ g: S3CO; h: Hl= S2/TOC andi: OICO =
S3CO /TO C.......................................................................................................................................... 56
Figure 4.2: Biplot of S2 vs. max tem perature, where sediment subsamples th at released OM
below 400°C are designated S240o and those above 400"C are S2b..................................... 58
Figure 4.3. Pseudo Van Krevelen diagram of HI vs. OICO. The dotted line is the 1 HkOICO
ratio line and the dot-dash line is the 0.5 HkOICO ratio line.

The oval indicates a

negative downcore degradation trend that excludes samples from 7 cm and 26 c m ... 60
Figure 4.4: Relation betw een TOC and fine sediment particles (r2=0.194, p<0.01). The oval
displays tw o visually observed clusters that include sediments from below 12 cm,
excluding 15 cm.................................................................................................................................61
Figure 5.1: Schematic microbial processing of organic m atter in lakes sediments and the
associated reduction pathways and resultant products of various modes of metabolism.
From Jprgensen (20 00 )....................................................................................................................64

List of Figures
Figure 5.2: Generalized overview of the SMB enzyme pathway for sulfate reduction and
sulfide oxidation,
electron

a: Sulfur-Oxidizing Bacteria (SOB) that take up H2S or S2O 32 as an

donor to

produce

S 042' through

the

Dsr/Apr/SAT or Sox pathways,

respectively (adapted from Stewart et al., 2011; Gregerson et al., 2011); b: SulfateReducing Bacteria (SRB) take up extracellular S 0 42 and reduce it to H2S through the
SAT/Apr/Dsr pathway to produce S 042' that is excreted (adapted from Cao et al., 2014).
............................................................................................................................................................... 67
Figure 5.3: Schematic of the role of SMB in the carbon and sulfur cycle to provide an
overview of coupling mechanism of using sulfate as an electron acceptor to degrade
ferm ented organic m atter (i.e. simple sugars such as lactate) to produce hydrogen
sulfide and either C 0 2 or acetate. Figure from Zhou (2011)................................................. 69
Figure 5.4: General representation of phylogenetic tree that depicts the broad diversity
sulfur-metabolizing bacteria and archaea and their major phylogenetic lineages and
method of sulfur metabolism.

PS: Phototrophic Sulfur Bacteria; SOB: Sulfur-Oxidizing

Bacteria and Archaea; SB: Sulfur (So)-Reducing Bacteria and Archaea; SRB: SulfateReducing Bacteria and Archaea; OS: Organic Sulfur Utilizing Bacteria and Archaea.
Adapted from Sievert et al. (2007)............................................................................................... 73
Figure 5.5: Overview of the downcore distribution SMB (DsrAB) and Tbac (16S rDNA) by
gene copy#/g sediment w ith standard deviation error bars and the redox boundaries
proposed by M n and Fe reduction peaks (Chapter 3). Ponar grabs are representative of
top ~7 cm ............................................................................................................................................ 85
Figure 5.6: Boxplot of %SMB of Total Bacterial qPCR replicates (16 ratios for each sample).
The black line in each box represents the sample median and the overall median is
shows by the vertical line as 1 .9 x 1 0 3. The Ponar Grabs are separated to the top but
are presumed to be a homologous sampling of the top ~ 7 c m .............................................87
Figure 5.7: M axim um likelihood tree of truncated dsrA genes (177 amino acids) using the
W helan and Goldman model.

Numbers correspond to 500 bootstrap replicates and

percentage of replicate trees in which the associated taxa w ere clustered as shown.
Clusters are based on 87% amino acid identity and associated Classes are divided by
the dotted lines.

Operational taxonomic units (OTU) are based on 97% amino acid

identity and require at least tw o environmental clones........................................................ 92
Figure 6.1: Global anthropogenic emissions of mercury to air by continent from 1 9 9 0 ,1 9 9 5 ,

2000 and 2005 that depicts a decrease in developed countries and increase in
developing countries, particularly Asia. The inset shows a breakdown of the industrial
activity associated w ith Hg emissions by each continent, where again Asia is the
primary producer. Both figures from United Nations Environmental Programme (2013).
...............................................................................................................................................................96

x

List of Figures
Figure 6.2: General overview of the global Hg cycle from the atmosphere to terrestrial and
aquatic systems and the transfer to and bioaccumulation through the food web as
methylmercury. From GMOS (2 0 1 2 ).......................................................................................... 99
Figure 6.3: Overview of Hg cycling in the sediments of estuarine and coastal environments
as described by M e rritt et al. (2008).

W here Hg enters oxic soils from the w ater

column and is transported to anoxic sediments by diffusion via sediment association
w here it is m ethylated by SRB.

MeHg is then re-transported to the sediment w ater

interface (SWI) w here it may enter the food chain or be converted to another species.
Circles depict sediment particles, Hgi=inorganic divalent Hg or Hg(ll), RD=reduction,
OD=oxidation, MiR =bacterially independent reduction. (M e rritt & Amirbahman, 2009)
.............................................................................................................................................................103
Figure 6.4: Level of Hg detected in trout tissue from Kusawa Lake from 1993-2010.

From

Stern et al. (2011)........................................................................................................................... 107
Figure 6.5: Downcore profile of Hg concentration in Kusawa Lake sediments. There is a 1.5fold increase in THg concentration from 1915-2010 over the top 10 cm ........................ 109
Figure 6 .6 : Downcore profile of 1/Hg association with % Fine Silt.

There are tw o

associations of sediments associated with the top 7cm, 12-18cm and 28cm (solid line
oval) and the lower sediments (dotted line oval). Outliers are 1 cm, 11 cm and 24 cm.

110
Figure 6.7:

Biplot of Hg'1 and HI'1 with the regressions denoted for the strong relation

betw een Hg and the HI400 (solid line) labile O M measurements and the degraded Hlb
fractions (dotted line).................................................................................................................... 112
Figure 6 .8 : Biplot of 1/Hg association with Log(Total bacteria). The sediments from 1-5 cm
show a steep positive relation at the left of the graph.

The solid line oval includes

sediments where a negative relation between Hg and Tbac for samples from 6-18 cm
and 26-29 cm. The dotted oval contains samples from below 12 cm. Samples from 2022 cm are considered outliers from these trends.................................................................. 114
Figure 6.9: Biplot of log(%SMB) vs. 1/Hg where, a: depicts the separation of the oxic and
anoxic zones in the top 10 cm, and b: shows the lack of structure within the anoxic
sediments from below 11 cm...................................................................................................... 116
Figure 7.1: Summary of findings and fates of Hg delivered to bottom sediments of Kusawa
Lake.

The estimates of redox zones, bacterial metabolism and state of O M

degradation are also described............................................................................................... 134

Acronyms

OIRE 6 oxygen index

Acronyms
O M organic m atter
a.s.l above sea level
PC pyrolysable carbon
A D W accumulated dry weight
PCA Principle Components Analysis
AM DE atmospheric mercury depletion
events

PCR polymerase chain reaction

Apr APS reductase

PSB purple sulfur bacteria

APS adenosine 5'-phosphate

qPCR quantitative polymerase chain
reaction

BLAST Basic Local Alignment Search Tool
RC residual carbon
CRS constant rate of supply model
SAT sulfate adenylyltransferase
dH 20 distilled w ater
SE standard error
DO dissolved oxygen
SB sulfur-reducing bacteria
Dsr dissimilatory sulfite reductase
SMB sulfur-metabolizing bacteria
EMR Energy Mines and Resources, Yukon
SOB sulfur-oxidizing bacteria
GEM gaseous elem ental mercury
SRB sulfate-reducing bacteria
GSB green sulfur bacteria
SSC suspended sediment concentration
HC hydrocarbon
THg total mercury
HI hydrogen index
TOC total organic carbon
LOI loss on ignition
Tbac total bacteria
M eH g methylmercury
UNBC University of Northern British
OICO carbon monoxide index

Columbia

OICO 2 carbon dioxide index

W RA W hite River Ash

Acknowledgements

Acknowledgements
This is the last section I am writing and I can only think of the fact that completing
this project and writing this document has been extraordinarily challenging.

I am amazed

th at it's almost complete and there are many to thank.
W hen I began my Master's, this project was intended to be different and I thank my
supervisor, Dr. Ellen Petticrew for allowing me to follow my own direction and guiding me
through this process.

I thank my com m ittee members, Dr. Phil Owens for sedimentology

advice and Dr. Keith Egger for use of his lab and helping me plan and troubleshoot my lab
work.

I especially thank Pat Roach for inviting me to participate and contribute to the

mercury research at Kusawa Lake and introducing me to Dr. Gary Stern.
I w ant to express my great appreciation to Dr. Stern and his collaborators, as I would
not have been able to complete this work w ithout their generous support.

Particularly, I

would like to thank them for collecting my samples and analyzing them for several of the
variables used in my study. As well, I would like to thank Dr. Stern and Dr. Jessie Carrie for
helping me interpret the Rock Eval results.
At UNBC, I w ant to thank Dr. Brent M urray for sharing his molecular biology
expertise and Dr. Dezene Huber for use of his lab. As well, I'd like to thank the members of
Dr. Egger's lab for their support and assistance in troubleshooting.

I am very thankful for

my fellow UNBC grad students for the opportunities to mutually commiserate in this
experience.

I'd especially like to thank Leah Vanden Busch for sharing the challenge of

undertaking a similar path of writing from the north.
I am indebted and grateful to Champagne and Aishihik First Nation for supporting
me throughout my post-secondary career and for donating resources for my lab work.
I'm also greatly appreciative to Darielle Talarico, for offering me the opportunity to
advance my career while providing the tim e and wonderful support needed to complete
this work.
So, in final reflection, I realize that when I think of this period of my life, I think of my
m other. I strive to rem em ber and invoke her lessons of how to live, how to work and how
to be kind and patient and I thank her. W hen I moved to Prince George to begin this she
missed me and as I finish, I miss her; but I can reflect in parallel the work this took, the path
I've followed and people who held me along the way.
I would like to thank my family from both my m other and father's side who have
been not only supportive of my educational goals but have also simply let me know they are
proud of me.

I have many wonderful friends and sisters that I thank for their love, for

listening and sharing.

I thank the Creator for my home the Yukon, W hitehorse and

Riverdale, places that w ithout I would have struggled to find the grounding needed to
complete this task.

Finally, I would like to thank sweet Scott Osborne, who has patiently

listened, endured my exasperation and given me back the com fort and security of family.
I raise my hands to you all. Shaw nithan, kwanaschis, thank you.

xiii

Chapter 1 Introduction

1.1

Introduction
The United Nations Environmental Programme has identified M ercury (Hg) as a global

threat to human and environmental health.

It is responsible for toxicity that affects the

gastrointestinal system and brain and is of particular concern to pregnant and nursing
w om en (United Nations Environment Programme, 2013). Arctic and Subarctic residents are
at risk for Hg exposure from naturally occurring sources of Hg and atmospherically
transferred Hg em itted from distant industrial sources (Schroeder et al., 1998; Dunford et
al.,

2010).

In

lakes,

algae and

algal-derived

organic

m atter

(O M )

anthropogenic and naturally occurring Hg from the w ater column.

can

scavenge

They then form

aggregates or floes with suspended sediment that settle to the lake bottom where Hg
species may be stored or re-suspended to the w ater column (Outridge et al., 2007; Stern et
al., 2009; Sanei et al., 2014). At the sediment oxic/anoxic transition zone, Sulfate-Reducing
Bacteria (SRB), a subset of Sulfur-Metabolizing Bacteria (SMB), can convert Hg to organic
m ethym ercury (M eHg) (Benoit et al., 1999; Gilmour et al., 2011).

M ethylm ercury is of

concern as it readily crosses cell walls to bioaccumulate and biomagnify up the food chain
(Fitzgerald & Lamborg, 2003; Chetelat et al., 2012).
In the Yukon Territory of northwestern Canada, many residents still harvest fish,
animals and vegetation from the lakes and forests. Therefore, the need to determ ine the

1

Introduction
risk of Hg exposure is im portant for those who regularly consume wild foods. This study
intends to contribute to the understanding of the factors that influence the ability of SRB to
m ethylate Hg in Subarctic lake sediments and inherently enable Hg uptake to the food web.
Much of this study augments previous work by Dr. Gary Stern and his team from the
University of M anitoba w here they have been examining the interaction betw een Hg and
certain carbon fractions of organic m atter (O M ) as detected by Rock Eval Pyrolysis (Outridge
et al., 2007; Stern et al., 2009; Carrie et al., 2012; Hare et al., 2014; Sanei et al., 2014). Their
hypothesis proposes th at climate change related increases in algal m atter have resulted in a
concurrent increase in algal scavenging of Hg from the w ater column to the lake sediments.
In 2005, Dr. Stern and his team collected duplicate sediment cores from the deepest
region of Subarctic, Kusawa Lake (60',19'55"N, 136°4'48"W ) in southwestern Yukon. From
these tw o cores they analyzed the components of Total Organic Carbon (TOC) using RockEval Pyrolysis and the Total Hg (THg) by cold vapour atomic absorption spectrometry. The
outcome was th at both cores exhibited a strong relation between THg and the Rock Eval
fraction, S2 th at consists of labile hydrocarbon-rich algal-derived O M . They also noted that
the 20th Century S2 levels w ere 26-29% higher than the pre-1900 levels (Stern et al., 2009).
This finding com plem ented previous works (Outridge et al., 2007) and was further
investigated in a recent study th at found that the ability of S2 algal-derived O M to scavenge
Hg from lakes is dependent on the lake's overall productivity and the level of available Hg
(Sanei et al., 2014).

The algal-Hg scavenging hypothesis also suggests that post-1950

increases observed in sediment Hg records are not directly related to atmospheric Hg

2

Introduction
deposits.

Instead, the available organic carbon in a basin has the ability to strongly

influence the rate of Hg sequestration in sediments (Outridge et al., 2007; Stern et al., 2009;
Sanei et al., 2014).
In M arch 2010, members of Dr. Stern's research team took another set of duplicate
sediment cores from ~700 m east of the previous 2005 sample site in Kusawa Lake, Yukon.
These w ere the cores used in this study to assess the environmental controls of Hgmethylation and dem ethylation by SRB in Kusawa Lake's bottom sediments. To achieve this,
the cores w ere analyzed to measure the concentration of total mercury, total bacteria and
SMB, Rock Eval Pyrolysis, trace metals, particle size and 210Pb and 137Cs dating. Aspects of
Kusawa's hydrology, climate conditions and other regional features were also considered.
The overall intent of this study is to contribute to the understanding of Hg cycling in
Subarctic aquatic systems so current and future generations of Yukoners can continue to
safely harvest food from the lakes and forests and enjoy the Yukon's spectacular beauty.
An example of the importance of the wilderness to Yukoners is the joint creation of
Kusawa Lake Park by an agreem ent between the Government of Yukon, the Champagne
and Aishihik First Nation, Carcross/Tagish First Nation and Kwanlin Dun First Nation. These
three First Nations have im memorially used Kusawa Lake when it was known as Nakhq M an,
"rafting-across lake" by the Southern Tutchone and Kusawau.a meaning, "long narrow lake"
by the inland and coastal Ttingit. There is a large delta on the lake created by the Primrose
River th at was a gathering place to discuss traditional governance, concerns regarding
animals and the forest and to celebrate in the abundance of fish and caribou th at migrated

3

Introduction
through the narrows. In planning the park, many of these traditional ideals are applied to
ensure the longevity and vitality of the lake (Kusawa Park, 2014).

Establishing a baseline

biogeochemical understanding of mercury cycling in Kusawa Lake and the potential
influence of climate change will aid current and future generations in monitoring and
recognizing the extent of mercury accumulation and its impact on our culture, health and
wellbeing.

1.2

Research Objectives
This project is a study of the interactions between O M , bacteria, and Hg in the

bottom sediments of Kusawa Lake, Yukon Territory. It also takes into account the influence
of the lake's limnology and sedimentation.

In essence, the objective of this study is to

evaluate and characterize the potential factors that control the interaction between SMB,
particularly SRB and Hg in the bottom sediments of Kusawa Lake. It will also contribute to
the body of research on Hg cycling and accumulation in Subarctic ecosystems. Overall, my
thesis results are intended for researchers, the First Nations of Kusawa Lake and the current
and future generations of Yukon people.
The primary objectives of this study are to:

1 . examine the origin, concentration and composition of organic m atter in the
lacustrine sediments of Kusawa Lake (Chapters 3 and 4);

4

Introduction

2 . characterize the phylogeny and concentrations of sulfur-metabolizing bacteria and
their interactions with the components of organic m atter and sediment processes
(Chapter 5);
3.

discuss the mechanisms involved in mercury transport to the sediments and
evaluate the

potential for mercury

methylation

by sulfate-reducing

bacteria

(Chapter 6 ), and;
4.

discuss the potential of climate change to influence the interactions between
organic m atter, sulfur-metabolizing bacteria and mercury (Chapter 6 ).

This thesis is structured into seven chapters where each chapter builds on the previous.
Chapter 1 outlines the overarching questions and research objectives. Chapter 2 introduces
the Kusawa Lake drainage basin, its historic climate conditions and other local features.
Chapter 3 describes the limnology, bathym etry and the features of the sediment core's
subsamples including the 210Pb and 137Cs dating, particle size and trace m etal profiles.
Chapter 4 is an overview of the Rock Eval Pyrolysis O M results and Chapter 5 depicts the
downcore concentrations of total bacteria and SMB as well as phylogeny of SMB. Finally,
Chapter 6 incorporates the findings from the previous four chapters to discuss the various
influences on SRB-mediated methylation of Hg in the sediments.

This chapter also

postulates the potential influences of climate change on the processes studied and the
implications for Yukon people. Finally, Chapter 7 summarizes the conclusions of the study,
significance of this work and suggests supplementary research.

5

Chapter 2 Study Area, Kusawa Lake, Yukon

2.1

Creation and Physical Characteristics
"When Crow was making the world he was flying over thinking hard about w h at to

do with the fish. He decided to go to Klukshu and held one wing towards the coast, the Alsek
drainage, and declared th a t it should be filled with Sockeye, Coho and Chinook salmon. He
held his other wing towards Kusawa, the Yukon drainage and said th at will be filled with
Chinook. That is why Klukshu has more salmon than the Yukon"
-

2.1.1

Adapted from Mrs. Annie Ned (Kusawa Park, 2014)

Landscape and Sub-basins
Kusawa Lake (60°19'55"N , 136°4'48"W ) is a proglacial fluvial system (Church &

Gilbert, 1975) w ith a lake surface area of 142 km 2 that drains a watershed of 4292 km 2 to
the Takhini River (Gilbert & Desloges, 2005), which enters the Yukon River just before Lake
Laberge outside of W hitehorse, Yukon (Fig 2.1). The lake is located ~80 km southwest of
W hitehorse and is accessible off the Alaska Highway betw een the Champagne and Aishihik
First Nation communities of Takhini and Champagne. It has five major sub-basins shown in
Figure 2.2 and described in Table 2.1.

2.1.2

Glacial History
Kusawa Lake lies at the southern extension of the Late Pleistocene glacially dammed

Glacial Lake Champagne, which reached a maximum elevation of 853 m above sea level
(a.s.l.) and spanned several valleys along the Shakwak Trench and Boundary Range
Mountains. Its boundaries have not been definitively defined, but its strandlines have been

6

Study Area, Kusawa Lake, Yukon
Yukon

V , YUKON
Takhini R.
Haines Jn.

Whitehorse

PLATEAU

Marsh
L.

Figure 2.1: Overview of southwest Yukon drainage systems and topology surrounding
Kusawa Lake.

Regions over 1000 m a.s.l. are shaded in grey and spot elevations are

identified for valley floors and lake surfaces (Gilbert & Desloges, 2005).

Table 2.1: Overview of Kusawa Lake sub-basins. Glacial cover is as of 2004 (Gilbert &
Desloges, 2005)________________________________________________________________________

Sub-Basin

Area (km2)

Glacial Cover %

Description

Upper-most
Takhini River

545

1 3 .9

Smallest sub-basin; greatest glacial cover;

Jo-Jo Creek

Takhini lake likely a sediment trap

Jo-Jo lake at 953 m a.s.l. in steep-walled
valley; acts as a sediment trap

7

Study Area, Kusawa Lake, Yukon
136*40trw

135*35'0"W

80*35TTN

N.0.SC.09

1

R

136‘4(K m

%

135*35*0*%V

Figure 2.2: Overview of the Takhini River drainage basin and Kusawa Lake sub-drainage
basins. The extent of glacial ice as of 2004 is shown in grey (Chow, 2009).

8

Study Area, Kusawa Lake, Yukon
identified along the Dezadeash valley, including offshoots from the Kathleen valley and
through the Frederick Lake valley to Kusawa Lake and the Takhini River valley (Fig 2.1)
(Hughes et al., 1969; Jackson et al., 1991; Gilbert & Desloges, 2005). In the Kusawa region,
Glacial Lake Champagne was dammed by a trunk glacier located at the southern end of the
Takhini valley that resulted in moraines and a large delta at the outlet of Kusawa Lake.
Failure of this ice dam lowered Lake Champagne and led to the separation and form ation of
Kusawa, which was further lowered to the modern day level of 671 m a.s.l. after a sediment
plug at the lake outlet eroded enough to allow drainage (Gilbert & Desloges, 2005).

2.1.3

Bathymetry
In July 2004, Gilbert & Desloges (2005) conducted an acoustic survey of Kusawa Lake

to determ ine the events involved in its form ation during the Holocene deglaciation of Lake
Champagne.

From this, they identified five distinct bathymetric regions based on

morphology, the acoustic character of the sediments and by inference of the sedimentary
history and environm ent (Fig 2.3, Table 2.2).

2.1.4

Core Sample Location
The core sediment samples were taken from the area below the transect line shown

in region IV (Fig 2.3).

Region IV is the deepest part of the lake with the second deepest

sediment fill. The acoustic profile of region IV taken by Gilbert & Desloges (2005) is shown

9

Study Area, Kusawa Lake, Yukon
Figure 2.3: Bathymetry of Kusawa Lake
determ ined from the July 2004 acoustic
survey

completed

Desloges (2005).

by

Gilbert

&

Isobath interval is 20

m (solid lines) and 10 m (broken lines)
below the normal summer w ater level.
Maxim um

depths are

acoustic

transects

indicated and

are

shown

as

horizontal lines. The velocity of sound
in w ater was assumed to be 1430 m/s
(corresponding to 5.5°C).
Primrose
River

Sandpiper
River

Acoustic transects

in Figure 2.4.

From this profile, they

84 m

channel

inferred that region IV consists of one
facies of well-layered sediment with
reflectors parallel to the present lake
bedrock.
Devtmoie
Creek

Further, they suggest that

this sedimentation regime is due to a
long period of deposition, largely from
low-density

turbidity

currents

that

distribute sediment along the length of
the

lake floor

(Gilbert

&

Desloges,

Region V

/

2005; Chow, 2009).

Kusawa River

10

Study Area, Kusawa Lake, Yukon
Table 2.2: Description of bathym etry of five regions in Kusawa Lake starting from head of
the lake at Kusawa River (region V) to outlet at Takhini River (region I) based on acoustic
profiles collected and characterized by Gilbert & Desloges (2005) and Chow (2009)._________

Region

Water
Depth (m)

Sediment
Depth (m)

IV

135

82

Description

Deepest part of lake, U-shaped valley. W ell-layered sediment
fill lies conformably on top of underlying basement (further
discussion in Section 2.1.4 and Figure 2.4)

II

40-84

~40-100

After elbow, several depressions, thickest glacial-lacustrine
sediment layer in lake. Before Primrose Delta in northern
portion, the narrowest part of the lake there is nearly flat
shallow sediment fill

Figure 2.4: Acoustic section of region IV in Kusawa Lake. The perspective is looking down
lake, to the North.

Location of transect is shown in Figure 2.3 as a horizontal line across

region IV (Gilbert & Desloges, 2005).

11

Study Area, Kusawa Lake, Yukon
2.2

Climate History
W ith the exception of the Primrose River narrows, Kusawa Lake is ice covered from

Novem ber/D ecem ber until late May. One Yukoner who owns a cabin near the campground
shared th at the ice typically goes out around M ay long weekend and can be described as
"thousands of smashing dishes, and it doesn't stop, its goes on and on for about 24 hours
and then the lake's quiet again" (personal communication, Janssens, M , 2014).
Kusawa is located within the Dezadeash region and is m oderated by warm air
masses from the Pacific Ocean. It lies just to the east of the Boundary Range Mountains and
can experience considerably lower tem peratures and precipitation than locations further
inland due to orographic effects cause by the Coastal and St. Elias mountain ranges (Lowey,

2002 ).
There are no long-term meteorological stations in the direct vicinity of Kusawa. The
precipitation trends are assumed from the Environment Canada's W hitehorse Airport
station located ~80 km northeast of the lake.

2.2.1 Temperature
Tem perature was first recorded in W hitehorse in 1905, when the small town was the
main rail and sternwheeler transportation hub for Stampeders on route to Dawson City
during the Klondike Gold Rush. It was also a respite stop for rafters after Miles Canyon and
the tow n's namesake, the treacherous W hitehorse Rapids. Activity then dropped off and
tem perature was not recorded from 1910 until 1941 when construction commenced for the
Alaska Highway from Dawson Creek, B.C. to Fairbanks, Alaska during W orld W ar II.

The

12

Study Area, Kusawa Lake, Yukon
average annual tem perature record at the W hitehorse Airport is shown in Figure 2.5. From
1970-2010 there is a 2 .14 ‘ C increase in annual tem perature with the warm est year recorded
in 2004. This is evidence of climate warming, which is also observed in the longest climate
proxy in the Yukon, the Yukon River break-up at Dawson City. The tim e and date of break­
up has been recorded every year since 1896, with a raffle awarded to the ticket with the
closest guess to when the ice goes out.

In the first 20 years of the record, break up

occurred around M ay 10th; in the past 20 years to date it has now advanced to M ay 4th
(Janowicz, 2010).

Another notable trend is that six of the seven April break-ups have

occurred within the last 10 years (Joe-Strack, 2012).

✓

>4;

Average Annual Temperature (C°)

Figure 2.5: Average annual air tem perature at W hitehorse Airport, located ~80km northeast
of Kusawa Lake, from

1905-2010.

The dash-dot regression line indicates a 2.14’ C

tem perature increase from 1970-2010, rz=0.229, p<0.01. (Environment Canada, 2014a)

13

Study Area, Kusawa Lake, Yukon
2.2.2

Precipitation
The average rainfall and snowfall at W hitehorse Airport from the Environment

Canada Canadian Climate Normals 1981-2010 was 160.9 mm and 141 cm, respectively
(Environment Canada, 2014a).

Ric Janowicz, the Yukon's hydrologist with Environment

Yukon's W ater Resources Branch noted inconsistent changes in annual precipitation
throughout the Yukon.
northern

and

southeast

He states th at w inter precipitation has generally increased in
regions

and

decreased

in the

southwest,

while

summer

precipitation has increased slightly throughout the Territory, with greater increases in the
southeast and central areas (Janowicz, 2010). The Kusawa region has experienced variable
snow pack loads, for example, during the 2007 Southern Lakes flood event, Kusawa Lake
had a 131-150% greater than normal snow pack, while in 2008 the snow-water equivalence
was between 71-90 and 91-100% of normal and in 2013 the nearby measured snow packs
w ere within the normal range (Environment Yukon, 2007, 2008 and 2010).

These

fluctuations may be due to the strong influence of atmospheric systems from the Pacific
Ocean mixing with drier, cooler inland air masses (Lowey, 2002).

Overall, the Kusawa

watershed may have experienced a small increase in summer rain and possible decrease in
w inter snow, though this is uncertain due to the recent variability in snow pack.

14

Study Area, Kusawa Lake, Yukon
2.2.3

Takhini River Discharge
The W ater Survey of Canada and Yukon W ater Resources Branch have operated

three hydrometric stations th at recorded discharge from the Takhini River watershed: the
Primrose River above Kusawa 29AC006, operational from 1990-1998; the Takhini River at
Kusawa outlet 09AC004, operational from 1952-1986 and 40 km downstream from there
near the Champagne and Aishihik First Nation community of Takhini, the Takhini River near
W hitehorse 09AC001, operational from 1948-current (Environment Canada, 2014b). The

300-

*%*

'(A

I
5

2000

1
<r

- -

c

-

Max (1949)
Min (1970)

£

<ft.
Month

Figure 2.6: Average m onthly discharge of the Takhini River near W hitehorse from the W ater
Survey of Canada (09AC001) from 1948-2012.

Daily flows are shown for the tw o highest

discharge years in 1948 and 2000 and minimal year in 1970 (Environment Canada, 2014b).

15

Study Area, Kusawa Lake, Yukon
average discharge for the Takhini River near W hitehorse station (Fig 2.6) is shown as it has
the longest record. The daily average hydrographs from 1949, 1970 and 2000 are shown
and represent the maximum, minimum and second maximum discharge years on record.
The peak flow occurs later in the summer as the watershed is glacially dom inated as
opposed to freshet, which typically occurs in early June in unglaciated systems.

2.2.4

2.2.4.1

Regional Disturbances

Torrent Systems
The Kusawa watershed is identified as a torrent system, which is common in

mountainous terrain and implies th at the system is prone to sporadic and sudden terrestrial
discharges due to a cycle of subdued w ater discharge and short pulses of dynamic debris
transport (Eisbacher & Clague, 1984). The most recent event was in 1982 where a flood
event at the Kusawa Lake territorial campground was caused by a combination of rainfall
flooding and slope failure and resulted in a catastrophic mudslide (Lowey, 2002); the
affected area is shown in Figure 2.2 as the Campground Alluvial Fan.

2 .2.4.2

Forest Fires
In 2005, Stern et al. (2009) analyzed a core from region IV for char deposited by

nearby forest fire activity and detected major fire activity between 1917-1948, which has
the potential to influence benthic activity and O M composition (Fig 2.7). M ore recent

16

Study Area, Kusawa Lake, Yukon

%
I

PvtRte* Ow

Figure 2.7: Particles of char associated with frequency of w ildfire in the region determ ined
by a core taken in 2005 w here frequent wildfires were observed from 1917-1954 (adapted
from Stern et al., 2009).
w ildfire events were also reported in the Upper-Takhini and Kusawa watershed in 2000 and
1987, respectively (Kusawa Park, 2014).

2.3

Vegetation
The northern part of the Kusawa Lake watershed lies within the Yukon Southern

Lakes ecoregion and the south within the Yukon-Stikine Highlands ecoregion. In the valleys,
W hite Spruce and to a limited extend, Trembling Aspen dom inate with a small population of
Lodgepole Pine at the north end of the lake. The conifer and mixed forest trees are present
to the tree line (~1200 m a.s.l.) where herbs, shrubs, willow, moss and lichen dom inate the
alpine tundra terrain (Smith et al., 2004).

17

Chapter 3 Limnology and Sedimentology of Proglacial Kusawa Lake

3.1

Introduction
Lake sedimentation in proglacial systems is dependent on several external factors

such as summ er and annual tem perature, precipitation, rate of snow m elt and rain storm
events (Hodder et al., 2007). These climate driven factors control localized systems such as
hydrology, lake stratification and terrestrial slope stability, all of which influence lake
sedimentation and interpretation of lacustrine sediment core profiles.

3.1.1

Glacial Contributions
Glaciers act as w ater storage bodies that supply m eltw ater and sediment to a

drainage basin on a diurnal, seasonal and tem poral scale (Jansson et al., 2003).

The

quantity of glacial m eltw ater is highly seasonal, with peaks in larger southern Yukon
drainage basins typically occurring in July or August (Environment Yukon, 2011).

Glacial

derived sediment transported with m eltw ater originates from ice shearing erosion of
bedrock and can contribute up to 75-83% of the fine sediment load (Desloges & Gilbert,
1994).

There is a strong relation betw een m eltw ater discharge and suspended sediment

concentration (SSC) that fluctuates throughout the season and with extrem e events such as
landslides or rainstorms (Gurnell et al., 1994).

18

Limnology and Sedimentology of Proglacial Kusawa Lake
The sediment load originating from glaciers is influenced by events such as glacial
surging, ongoing cycles of retreat (Gilbert et al., 2002) and jokulhlaups, which are
catastrophic outburst flood events th at release and transport large volumes of w ater and
sediment (Desloges & Church, 1992; Ng & Bjomsson, 2003). These extrem e glacial events
are predom inately climate driven and can strongly influence the sediment record.

3.1.2

Terrestrial Contributions
O ther sediment sources in proglacial systems include terrestrial inputs from

land/m ud slides and peri-glacial sediment slumped from lateral valley sides (Chow, 2009).
W ithin the Yukon's various permafrost zones, land and mud slide potential is increasing due
to climate warming and the concurrent increase of permafrost melt, leading to decreased
land stability (Environment Yukon, 2011). Kusawa Lake lies within the sporadic continuous
permafrost zone (10-50%

permafrost coverage) of the Yukon, therefore the

higher

elevations of the southern basin are possibly permafrost bearing (Yukon Permafrost
Network, 2011).

Another terrestrial sediment pathway is via torrent systems where

sediment is displaced and deposited to form alluvial fans or deltas along lakes. The system
can be described as either dormant, where the region is stable and w ater is discharged
w ithout m ajor landscape disruption or active, where short pulses of bulk debris transport
result in catastrophic events such as mud or landslides and floods (Eisbacher & Clague,
1984).

Torrent systems are also subject to climate change impacts such as changes in

19

Limnology and Sedimentology of Proglacial Kusawa Lake
vegetation, frequencies of storm events, permafrost melt and other factors th at may alter
slope stability (Stoffel & Huggel, 2012)

3.1.3

Fluvial Transport and Storage
River and stream dynamics play a key role in sediment transport and fate.

The

fluvial discharge (m 3/s) is directly related to seasonal tem perature and precipitation.
W inter river and stream discharges are mainly fed by groundwater and carry little sediment.
W inter low flows in the southern Yukon typically occur just before the snow begins to m elt
in late March (Janowicz, 2008).

The spring nival peak normally occurs in early June

(Environment Yukon, 2011). During summer, rainfall may also cause peak discharge events,
especially in the southwest Yukon where climate systems are strongly influenced by the
Pacific Ocean and many ocean-derived storms can deliver large volumes of precipitation
inland (Lowey, 2002).

In the glacially dom inated systems of the southwest Yukon, the

annual peak w ater levels from glacial m eltw ater typically occur in early to mid August
(Environment Yukon, 2011).
For coarse sediments, the available energy for a given discharge is related to a fluvial
system's capacity to mobilize a given SSC and grain size. In other words, lower discharges
are only capable of transporting a limited sediment load and maximum grain size, whereas
higher discharges, such as flood events, have more energy and can transport a more
substantial load and larger grain sizes (Knighton, 1998; M arren, 2005). The seasonal and
annual variations in discharge energy and the corresponding sediment load are reflected in

20

Limnology and Sedimentology of Proglacial Kusawa Lake
lake sediment records, for example larger particles are transported and deposited during
higher flow years and flood events.

Fine sediments fluxes are controlled by supply, from

sources such as land and channel bank erosion and typically are not limited by transport
capacity as coarse sediments are.
Before sediment is delivered to lakes, it can be stored in sediment traps located
higher in the basin, resulting in a lag in the lake sediment record if remobilization occurs
(Hodder et al., 2007). Types of sediment traps include other upstream lakes that cause a
decrease in flow and therefore deposit larger particle sizes, alluvial fans, deltas, sandbars
and flood plains (Parker et al., 1998; Dirszowsky & Desloges, 2004; Lipowsky, 2006).

3.1.4

Lacustrine Processes
The dispersal of suspended sediment in lakes is influenced by a num ber of factors.

One of interest is the Coriolis Effect, where the currents of all lakes in the northern
hemisphere are deflected to the right due to the acceleration generated by rotation of the
Earth (Hamblin

&

Carmack,

1978).

O ther significant factors

include

particle

size,

stratification, inflow, wind and turbidity currents.

3.1.4.1 Stratification
Characterization of lake types requires knowledge of stratification and mixing
regimes. Lakes stratify into layers based on tem perature and chemistry into the: epilimnion
- the surface waters th at are generally w arm er and less dense, metalimnion - the transition

21

Limnology and Sedimentology of Proglacial Kusawa Lake
zone betw een the surface and bottom w ater, thermocline - located within the metalimnion
and a region of rapid tem perature shift, and hypolimnion - the bottom waters that are
generally colder and more dense. Lakes are considered isothermal during mixing or when
there is continuous mixing and as a result no stratification is observed.
W ater column differences are also characterized by dissolved oxygen w here regions
may be: oxic - containing oxygen, anoxic - oxygen depleted or suboxic - region of low
oxygen levels betw een the oxic and anoxic zones.

Beyond this there are various

classifications of vertical oxygen distribution, tw o of note are: orthograde profiles - when
oxygen is present throughout the w ater column and is typical of low productivity lakes or
oligotrophic systems, and clinograde - often occurs as a result of bacterial decomposition of
organic m atter and high algal productivity at some depth within the w ater column resulting
in low oxygen availability in the hypolimnion, typical in more eutrophic systems (Dodson,
2005).
The frequency and mechanism of mixing these layers depends on various lake
conditions, such as hydrostatic pressure, hydrology and climate.

A previous study

monitored the tem perature profiles of several Yukon lakes in a summer and w inter month
and the majority of the lakes w ere found to be dimictic (Shortreed & Stockner, 1986),
indicating they destratify twice a year (Dodson, 2005).

3 .1.4.2

River and stream inflow
The differences in density between river and streams flowing into the main body of

w ater in a lake results in three possible flow patterns (w here density = p): overflow -

22

Limnology and Sedimentology of Proglacial Kusawa Lake
hypopycnal (piake>Pinfiow), interflow - homopycnal (piake=Pmfiow) and underflow - hyperpycnal
(piake<Pinfiow)- The patterns are further dependent on the lake's hydrostatic pressure and
tem perature stratification (M iddleton & Hampton, 1976; Edwards, 1992; Knighton, 1998).
Underflows are one of the principle mechanisms of sediment distribution in
proglacial lakes (Chow, 2009). There are tw o types of underflows that are typical of glacialfed lakes.

First, quasi-continuous flows, where high-density river w ater flows under the

hypolimnon as a bottom current resulting in long-term delivery of fines with interm ittent
pulses from upper basin activities (Smith & Ashley, 1985). The second is from catastrophic
events and mass movements resulting in short-lived surge-type currents th at are usually
more localized and isolated (Serink, 2004).

These tw o types of flows can be considered

turbidity currents or underflows that carry sediment along lake or ocean beds (Chow, 2009).
The ability of turbidity currents to transport the sediment is dependent on
tem perature, salinity, hydrostatic pressure, current velocity and sediment composition. Any
change in these variables can result in sedimentation of material to the lake bottom (Serink,
2004).

3.1.4.3

W ind and currents
W ind also plays a significant role in water-circulation and transporting sediment,

though it is difficult to model and understand due to variations in wind strength and
direction and the complexity of lake basins (Csanady, 1978).

W ind stress primarily

influences surface currents and is capable of re-suspending sediments at shallow locations

23

Limnology and Sedimentology of Proglacial Kusawa Lake
(Hiikanson, 1977).

Internal lake oscillations or lake basin-scale waves, called seiches, also

contribute to ongoing sediment distribution and lake mixing (W uest & Lorke, 2003).

3.1.5

Varves
The highly seasonal nature of sediment delivery in proglacial lakes often results in

the form ation of varves, seasonal layers of sediment deposited over the course of one year
in deep-w ater lakes. Under desirable conditions, annual sediment couplets are form ed with
one thick light coloured sand/silt (minerogenic) layer derived from the nival peak and
summer flows and the second, a thin darker layer of organics and silt/clay (organic-rich
layer) accumulated during the autumn and w inter (Desloges & Gilbert, 1994; Lamoureux &
Bradley, 1996; Slaymaker et al., 2003).

Due to their annual nature, varves are commonly

used in paleolimnology to gain insight into a lake's environmental history and contemporary
bed sediment processes, such as variations induced by climate change (de Geer, 1912;
Desloges et al., 2002).

However, the sedimentary record may not always reflect the true

chronology, as the sediment lamination may not be truly laminar in nature due to post­
settlem ent remobilization, discontinuous sedimentary processes or other interferences that
disrupt the varve record (Hodder et al., 2007). To compensate for these potential errors in
sediment chronology, other methods of dating, such as isotope dating can be used to verify
accuracy.

24

Limnology and Sedimentology of Proglacial Kusawa Lake
3.1.6

Lead-210 and Caesium-137 Dating and Sedimentation Rates
Radiometric dating is a common technique used to date various materials such as

carbon, oxygen and inorganics based on the comparison between the abundance of a
naturally or artificial occurring radioactive isotope to its decay products and their known
rates of decay (Olsson, 1986).
The annual laminar deposited sediment in lakes can be dated by measuring the
downcore "unsupported" or excess 210Pb isotope activity compared to the surface 210Pb
activity that corresponds to the sampling date (Fig 3.1). There are tw o natural origins of
210Pb, the atmospheric decay

of 222Rn (unsupported)

226Ra within the sediments (supported).

and 222Rn production from natural

The unsupported or excess 210Pb is calculated by

subtracting the supported component from the total 210Pb activity within each sediment
layer.

The

determ ined
surface

dates

by

activity

are

then

comparing
to

the

the

decayed

activity at depth (de Souza et al.,

2012 ).

Figure 3.1: Overview of the 210Pb
cycle

and

decay

of

238U

to

supported

(sediment)

and

unsupported

(atmospheric)

222Rn

decay sources.

25

Limnology and Sedimentology of Proglacial Kusawa Lake
The resultant dates can be further confirmed by observing the independent 137Cs peak,
which is observed as a result of fall out during therm onuclear weapons testing between
1954 and 1963, the latter corresponding to the tim e of maximum fallout (Foster et al., 2006;
Appleby, 2008).

Once the sediment dates are known the sedimentation rate can be

calculated as grams of sedim ent/m 2/y e a r to indicate changes in the rate of sediment
delivery to the sample site (de Souza et al., 2012).

3.1.7

Trace Metals
Trace metals profiles are commonly measured in lake sediments to discern an array

of sediment and O M processes and mechanisms. They can be used to track sediment from
their origin, assess contaminant transportation, heavy metal pollution and enrichment
(AMAP, 1998).

Once deposited in lakebed sediments, trace metals undergo various

transformations depending on the metals reactivity, concentrations of other trace metals,
redox conditions, and interactions with O M and sediment. In lakes, trace metals originate
from: autochthounous sources from within the lake basin, allochthonous sources from
within the local environm ent but outside the lake basin or anthropogenic sources from
environm ental pollution due to human activity (such as Hg, V, Cr, Ni, Zn, Cu, Cd and Ag)
(Outridge et al., 2005; Heimburger et al., 2012).

26

Limnology and Sedimentology of Proglacial Kusawa Lake
3.1.7.1

Redox Conditions
Trace metals can also be used to approximate redox conditions in downcore

sediment profiles w ithout oxygen or pH measurements. Under oxidizing conditions, redox
sensitive metals are more soluble and under reducing conditions they are less soluble,
which

causes enrichm ent of redox sensitive trace

metals during oxygen depletion

(Tribovillard et al., 2006). Trace metals known to be redox sensitive include: As, Cu, Fe, M n,
M o, Ni, Pb, Cd, Co, Cr, V, U and Zn (Boyle, 2001; Audry et al., 2006; Tribovillard et al., 2006;
Ye et al., 2013).
One w ell-docum ented redox effect on trace metals is the reduction of soluble Fe and
M n oxyhydroxides (M n or Fe-OH), which scavenge elements such as As, Cd, Cu, M o, Pb and
Zn to M n /F e - 0 + adsorption sites (Boyle, 2001). Reduction of insoluble Fe2+ to soluble Fe3+
and resultant loss of the hydroxide group at approximately -100 mV - 100 mV, results in a
sharp precipitation peak observable in the Fe trace m etal profile along with subsequent
peaks of previously bound metals (Audry et al., 2006). A similar sequence occurs with M n at
around approximately 100 - 300 mV redox potential. By noting the reduction peaks of Fe
and M n and associated precipitation of scavenged metals, the redox boundaries of the
sediments can be hypothesized.

27

Limnology and Sedimentology of Proglacial Kusawa Lake
3.2

Methods

3.2.1 Core Sampling
Pat

Roach,

Contaminants

Scientist

with

Aboriginal

Affairs

and

Northern

Developm ent Canada and a team from Dr. Gary Stern's research group at the University of
M anitoba collected duplicate 30 cm sediment cores from under ice in March 2010.
samples w ere collected from

bathymetric region

The

IV of Kusawa Lake at 60°15"46',

136‘ 11"39', using a 10 cm diam eter KB gravity corer (m anufactured by Departm ent of
Fisheries and Oceans) (Fig 3.2). On site the first core, Core A, was sub-sectioned into 1 cm
intervals from 0-30 cm; the second core, Core B, was sub-sectioned into 0.5 cm intervals
from 0 -10 cm and 1 cm intervals from 10-30 cm. All sub-sections w ere stored in W hirlpak
bags at am bient tem perature (0-10°C). Core B subsamples w ere shipped to the University
of M anitoba in W innipeg while Core A was stored at -20°C and transferred to UNBC.
In Septem ber 2010, Pat Roach collected three sediment grabs via boat using an
Ekman grab within the vicinity of the main duplicate cores to increase statistical certainty.
Each grab was assumed to reach approximately 7 cm in depth.

The three surface grabs

w ere shipped on ice to UNBC and subject to the same tests as Core A.

28

Limnology and Sedimentology of Proglacial Kusawa Lake

Figure 3.2: Kusawa Lake core sampling field trip in March 2010 showing the KB gravity corer
at the sample site

3.2.2

Aerial Photos
The Governm ent of Yukon's Energy Mines and Resources (EMR) SkyLine Air Photo

Locator website (EMR, 2013) was used to determ ine the location and coverage of aerial
photos available for Kusawa Lake, region IV.

Photo originals taken in 2007, 1987, 1979,

1972 and 1948 w ere retrieved from the EMR Library and scanned into TIFF files. To stich
the photos into panoramas, AutoPano Giga 3.0 software, developed by Kolor (Kolor SARL,
2013) was used.

Final photos w ere optimized and cropped to highlight any tem poral

variations in topography, sediment plumes and overall conditions of the lake section. The
sample site was identified in each photo by measuring relative distances from select
topographic features, such as shoreline and creek outlets.

29

Limnology and Sedimentology of Proglacial Kusawa Lake
3.2.3

Limnology
Under ice w ater measurements for Kusawa bathymetric regions I and III and tw o

locations w ithin region IV near the sample site (IVa) and south of the sample site (IVb), were
taken using the Sonde YSI 6000 (YSI Inc., Yellow Springs, OH, USA), which was lowered to
depth using an electric winch through an ice auger hole on March 28, 2012 by Pat Roach.
Measurem ents w ere taken to determine: dissolved oxygen (DO), tem perature and pH.

3.2.4

Lead-210 and Caesium-137 Dating and Sedimentation Rate
Isotope analyses of 210Pb and 137Cs were conducted on Core B at the University of

M anitoba's D epartm ent of Soil Science. Sediments w ere tem porally characterized by depth
and sedimentation rates w ere calculated using the linear and the constant rate of supply
model (CRS) (Robbins, 1978). The CRS model assumes that the absolute 210Pb flux rate of
the bottom sediments remains constant regardless of background sedimentation, which is a
more suitable assumption as compared to other sedimentation models.

Caesium-137

activity is measured to verify 210Pb dates in correspondence with nuclear weapons testing
from 1954-1963 (Robbins, 1978; Stern et al., 2009). Final dates and trends were compared
to the 2005 core taken by Stern et al. (2009) from approximately 700 m west of the current
sample location.

30

Limnology and Sedimentology of Proglacial Kusawa Lake
3.2.5

Particle Size
Particle size distributions of each Core A sediment increment w ere measured using

the M alvern Mastersizer 3000 (M alvern Instruments Ltd., M alvern, UK) at UNBC.

The

Mastersizer 3000 determ ines particle sizes from 0.01-3500 pm by measuring the intensity
of light scattered as a laser beam passes through a dispersed particulate sample. Samples
w ere not subject to H 2O 2 digestion or other methods to remove organics. Approximately 1
g of sediment from each subsection was resuspended in 5 mL of distilled w ater (dH20) and
left to soak overnight. A few drops of the sample solution w ere added to 120 mL of reverse
osmosis dH20 in the Mastersizer reservoir until ~11% light obscurity was measured.

The

samples w ere subject to pulse sonication for 60 s to break-up and prevent floe formations
and false measurements of larger particle sizes. Particles sizes w ere represented using D50,
the median diam eter grain size, Dgoand D i0, to reflect the size of the 90th and 10th percentile
of the grain size distribution respectively. The D i0, D50 and D90and percent sediment size
class were calculated from Udden (1914) and W entw orth (1922) using a sem i-autom ated
macro on MS Excel (Microsoft Inc.) called GRADISTAT v.4.0. (Blott, 2000). Changes in down
core sediment size w ere used to identify historic sediment deposition events and to
compare with other measured parameters

31

Limnology and Sedimentology of Proglacial Kusawa Lake
3.2.6

Trace Metals
Subsamples of Core B w ere sent to AcmeLabs in Vancouver, B.C. w here 0.25 g of

sediment sample was subject to four acid digestions in preparation for Inductively Coupled
Plasma-Mass Spectrometry (ICP-MS) to determ ine the concentrations of 60 trace elements.
ICP-MS detects sediment metals concentrations by counting the atoms for each elem ent
present in the solution (Acme Labs, 2013). The subsamples for Core B w ere divided into 0.5
cm increments from 0-10 cm and 1 cm increments for the rem ainder of the core to 30 cm.
To allow direct comparison to Core A, which was sliced in 1 cm increments throughout the
core, each x.5 and x.O m easurem ent was averaged to provide one measurem ent per
centim eter. For example, 0-0.5 cm + 0.5-1 cm was averaged to yield the overall 0-1 cm or 0
cm m easurem ent used in analysis.

The downcore trace metal distribution trend was

analyzed using Principle Components Analysis (PCA) in a variance-covariance cross-products
m atrix on three axes to discern tem poral trends, major historic events and other notable
downcore variations. The downcore trend of trace metals with known redox sensitivities
w ere also used to estimate sediment redox boundaries and hypothesize the boundaries of
the sediment oxic, suboxic and anoxic zones.

32

Limnology and Sedimentology of Proglacial Kusawa Lake
3.3

Results and Discussion

3.3.1 Aerial Photos
It is assumed that all the aerial photos were taken in the late summer (Fig 3.3).
Though no specific date was recorded for the 1948 session, informal communication with
EMR staff responsible for the Air Photo library confirmed that very few aerial flights have
occurred outside of July or August in the Yukon.

As well, visual observation of the 1948

photo (Fig 3.3d) indicates similar a sediment plume trend to other years. Photos from both
1972 and 1979 w ere stitched together to represent the 1970s.
In Chow's study (2009), she suggests that the Kusawa River carries a high sediment
load from its upper tributaries of high valley walls and alluvial fans. The river then braids
and decreases m om entum as it enters Kusawa Lake, resulting in mass settling and
sedimentation in bathymetric region V (Figure 2.3). Upper Takhini Lake has been reported
to act as an efficient sediment trap (Chow, 2009), though the Upper-most Takhini River
continuously delivers sediment to Kusawa Lake. The Upper-most Takhini sub-basin also has
the greatest extent of glacial cover and is frequently subject to mass wasting events (Ng &
Bjdrnsson, 2003; Gilbert & Desloges, 2005; Chow, 2009).

Both of these sediment sources

are observed in Figure 3.3, w here sediment plumes from both the Kusawa River and Upper
Takhini River are observed in all years, varying in their dispersal and turbidity depending on
the year and exposure of the photographs.
Observations of the sediment plume and flow patterns in Figure 3.3 show th at this
narrow, deep valley lake exhibits river-like characteristics. One illustration of this is th at the

33

Limnology and Sedimentology of Proglacial Kusawa Lake

\
I

4

/ \
h

'
•

v

-

^ *

Figure 3.3: Aerial photos taken in, a: August 2007, b: July 1987, c: August 1972/1 979, and d:
1948 unknown tim e of year. The w hite circle identifies the sample site, located within the
sediment plume from the Upper-Takhini and Kusawa Rivers.

34

Limnology and Sedimentology of Proglacial Kusawa Lake
highest sediment concentration occurs around the outside edge of the lake bends, which
could be considered river-like meanders. This is most evident in the westward turn around
Arc M ountain where the sediment is concentrated to the outside eastern shore.

The

Coriolis Effect may also influence sediment distribution by moving w ater currents to the
right or east in the Northern Hemisphere (Hamblin & Carmack, 1978).
The w hite dot on each panorama indicates the sample site, which is located within
the dispersal path of the sediment plume in each photo.

Therefore, the cores collected

w ere subject to ongoing sedimentation from mass wasting, flooding and other events that
impact the volume of sediment delivered from the Kusawa and Upper Takhini Rivers. As a
result, when interpreting the core sedimentation rates and downcore relations, it cannot be
assumed th at sediment delivery is constant, although the sample was taken from the
deepest region of the lake. A more ideal sample site would have been on the western side
of the m eander around Arc M ountain, where Stern's et al. (2009) first core in 2005 was
collected.

3.3.2

Limnology
The w ater column tem perature, pH and

DO w ere

measured

under ice for

bathym etric regions I, III, IVa (near sample site) and IVb (south of sample site) on March 28,
2012 (Fig 3.4). For regions I, III and IVa there was a thermocline observed at approximately
2.5 m below the w ater surface (Fig 3.4a). Below 2.5 m the tem perature increased gradually

35

Limnology and Sedimentology of Proglacial Kusawa Lake
b.

Temperature (°C)

pH
5.0

I

I________ 1

I________ '

I

5.5

6.0

I

6.5

10

DO (mg/L)
11

12

13

1___________I___________i__________ I

O

>

E

f
&
c

E
3
8
4>
I
Region I
Region III
Region IVa
Region IVb

Figure 3.4: Graph of under ice limnology measurements taken on March 29 /2 0 1 2 by P.
Roach (unpublished) at four of the five bathymetric regions of Kusawa Lake. The max depth
at regions I, III, IV and V are 21 m, 58 m, 126 m and 104 m, respectively (a: tem perature; b:
pH and c: dissolved oxygen)

to the bottom at each location. The most distinct therm al stratification was observed at
region I, the shallowest section of the lake.
The pH profiles were more varied between regions with region III being the most
acidic and region V the least (Fig 3.4b).

At each site the pH increased gradually with

increasing depth but did not exceed 6.5. The most notable trend was a sharp increase in pH
at region IVa around depths of 100 m and 112 m.

These tw o distinct shifts may be

36

Limnology and Sedimentology of Proglacial Kusawa Lake
indicative of high-density underflow currents that originated from the glacially dominated
Upper-Takhini basin. Another possibility is that the bathymetric hole located in region IVa
may have a distinct current from the southern overflow, resulting in a shift in pH.
One interesting observation from the limnology measurements is the presence of
oxygen throughout the w ater column in all regions.

These orthograde oxygen profiles

indicate abundant oxygen availability even in late winter, which suggests oligotrophic
conditions. A small decrease in DO is also observed in region IVa around the pH shift at 100
m, further suggesting a separate underflow current and independent cycling in the deep
bathym etric hole.
W hile minimal stratification is observed at near the lake outlet, little to no layering is
displayed near the lake head. Chow's (2009) took a similar set of limnology measurements
in July 2004, the warm est summer recorded in Yukon history (Environment Canada, 2010).
W hen compared to the 2012 w inter measurements used in this study, an inverse trend
betw een sum m er and w inter is observed for tem perature. This supports classification of
Kusawa Lake as dimictic, indicating that it destratifies twice a year, which is typical of Yukon
lakes (Shortreed & Stockner, 1986).
Together, analysis of the sediment plume from the aerial photos along with the
summer and w inter limnological record suggests that the flow pattern at the head of the
lake, and the core sample site, is more fluvial.

W hen the lake current slows before the

Takhini River outlet in region I more lake-like characteristics such as distinct therm al
stratification are observed.

37

Limnology and Sedimentology of Proglacial Kusawa Lake
3.3.3

Lead-210 and Caesium-137 Dating and Sedimentation Rate

3.3.3.1

Lead-210 and Caesium-137 Dating
W hen initially reviewing the original 210Pb and 137Cs down core profiles (Fig 3.5a) tw o

anomalies w ere noted: first, if the dating is based on the 210Pb measurements, then the
137Cs peak aligned to 1972, instead of the expected peak in 137Cs fallout in 1963 and second,
there was a large dilution event observed at 1925.

Aside from the 1925 event, the

downcore pattern of excess 210Pb exhibited a consistent linear decrease with accumulated
dry weight.
To provide some insight into these tw o anomalies, the measurements were
compared to the core taken in 2005 by Stern et al. (2009) (Fig 3.5a) that was sampled from
~700 m west of the 2010 core site within region IV.

The tw o cores w ere not directly

comparable, as the 2005 core was taken from outside of the major lake current described in
section 3.3.2. The 2005 core has a lower linear sedimentation rate of 323 g /m 2/y r (Stern et
al., 2009) compared to 518 g /m 2/y r for the 2010 core. As it could not be confirmed w hether
the 210Pb measurements or 137Cs measurements for the 2010 core were incorrect, a method
of relative interpolation was used to provide a reasonable estimate of the sediment dates.
Two similar peaks w ere identified in each profile at the 210Pb dates of 1 9 9 9 /1 9 9 6 and
1 9 2 5 /1 9 11 for the 2 0 1 0/2 00 5 cores, respectively.

These values w ere interpolated to

calculate new dates, which w ere anchored at 1963 to reflect the 137Cs peak (Fig 3.5b).

In

other words, the excess 210Pb profile for the 2010 core was "forced" by the 1963 137Cs peak.
This approach was employed, as the 137Cs peak is considered a more reliable chronological
38

Limnology and Sedimentology of Proglacial Kusawa Lake
excess 210Pb (Bq g '1)

2010 1990 -

1970 -

1950 -

1910 -

1870 -

1850 0.00

0.05

0.15

0.10

0.20

0.25

'Cs (Bq g’ ’)

Figure 3.5a: Alignment of 2005 and 2010 137Cs and 210Pb profiles to compare and interpolate
differences. The estimated dates were determ ined by linear alignment betw een peaks at
1 9 9 9 /1 9 9 6 ,1 9 7 2 /1 9 6 4 and 19 25 /1 911 for the 2 0 1 0/2 005 cores, respectively.

excess z10Pb (Bqg"1)
0.001

0.002

0.005

0.010

0.020

0.050

0.100

0.200

0.500

- 2.5

t f ' CM -

- 10

- 12

0.00

0.05

0.15

0.20

Figure 3.5b: Estimated 210Pb downcore activities anchored to the 137Cs peak at 1963 with
accumulated dry weight and depth of sediment.

The annual linear sedim entation rate is

0.092 cm /yr.

39

Limnology and Sedimentology of Proglacial Kusawa Lake
measure than the excess 210Pb profile (Benoit & Rozan, 2001), especially when dealing with
tw o cores from different sedimentation environments. This new data set does not provide
reliable dates, however, given the similarity of the shapes of the tw o data sets, the
downcore trends are considered valid.

Correction for focusing and the 210Pb flux for the

2010 210Pb profile was not calculated. Table 3.1 describes the differences and results for the
original and adjusted 210Pb profile along with sedimentation rates, which are discussed in
the next section.

Table 3.1: Comparison of calculated year vs. depth and sedimentation rates for the original
2010 210Pb profile that was adjusted to the 2005 core taken from ~700 m west of the 2010
core.__________________________________________________________________________________

2010 210Pb
Version

137Cs
Peak

10 cm dilution
event

# Years
recorded

Adjusted________ 1963_________ 1911___________ 162

3 .3.3.2

Avg Linear
Sed Rate

Annual Linear
Sed Rate

548 g /m 2/y r

0.092 cm /yr

Sedimentation Rate
The sedimentation rate was calculated by:
ADW(g) /
/ Volume Sediment (m 3)
# years
slice (m)

,
'

= Sedim entation Rate ( g /m 2/ y r )

(Equation 3.1)

W here: ADW is Accumulated Dry W eight and volume of sediment is determ ined using n r 2,
(r=radius of corer or d iam eter/2 = 0.05 m) by thickness of slice and #years are based on the
num ber of years calculated by the 210Pb and 137Cs dating profile per slice thickness. These
results are shown in Figure 3.6 plotted against both depth and year.

The average linear

sedimentation rate was calculated to be 548 ± 47 g /m 2/y r (± SE). The annual linear

40

Limnology and Sedimentology of Proglacial Kusawa Lake
Sedimentation Rate (g/m /yr)
250

500

i_

_i_

750

1000

1250

1500

1750
2010
2007
2002
1992
1982
1969
1959
1950
1940

£
I

1928
1915
1911

1
w

1903
1885
1863

in

1847

CM

Figure 3.6: Sedimentation rate determ ined by adjusted 210Pb dating profile and Equation 3.1.
Note the anomalous sedimentation event at 1911.

sedimentation rate was determ ined by the slope of the ADW versus excess 210Pb curve to be
0.092 cm /yr, r2=0.87 (Fig 3.5b).

3 .3.3.3

1911 Sedimentation Anomaly
There is an anomalous sediment delivery event of 1837 g /m 2/y r that occurred in

1911 at 11 cm (Fig 3.6) in the core that is also reflected as a dilution in 210Pb activity (Fig
3.5b). Unfortunately, no historical climate data was recorded at that tim e period. It can be
speculated th at some large event such as a flood, jokulhlaup or release of previously
damned sediment from moraines, ice or some other mechanical barrier occurred. Prior to

41

Limnology and Sedimentology of Proglacial Kusawa Lake
the event the average sedimentation rate was 350 g /m 2/y r and after it was 500 g /m 2/y r.
This suggests that a previously unconnected sediment source was activated and increased
the annual volume of sediment delivered to region IV. The source is either from the UpperTakhini River or Kusawa River, as both are glacially m oderated, it is difficult to determ ine
which sub-basin the large sediment load originated from.

3.3.4

Particle Size
Overall there is little downcore particle size variability in the core subsamples from

0-30 cm (Table 3.2, Fig 3.7). The particle size is very small with a D5o of 3.9 ± 0.087 pm (±
SE).

These and other grain size measurements are shown in Table 3.2 and depicted in

Figure 3.7.

The 19 1 1/1 1 cm sediment event is reflected as a sharp increase in the

proportion of clays and medium silts and then a decrease in these fractions at 10 cm. The
average D i0 is 0 .7 0 pm prior to the event and 0.77 pm after. This supports the hypothesis
presented in section 3.3.2 that a previously inactive sediment source released a higher
concentration of fine sediment to region IV.

Table 3.2: Down core particle size and % average sediment size class distributions.

Particle Size

Average
Standard Error
42

Limnology and Sedimentology of Proglacial Kusawa Lake

Clay
Fine Silt
Medium Silt
Sand

Percent Sediment Size Class (%)

Figure 3.7: Downcore Percent Sediment Size Class (%) variations.

Note that the sample

consists predom inately of clays and silt.

3.3.5

Trace Metals
Of the 60 trace metals analyzed, Au and Re were below the minimum detection limit

and the following trace metals displayed little to no down core variation and w ere not
included in any analysis: Be, Lu, Tm, Ho, Tb, S, Se, Eu, Er, Yb and Te.

Sulfur (S) was only

detected in the top centim eter at 0.045% and was below the limit of detection for the
rem ainder of the core. Full sediment profiles of each detected trace metal are presented in
the Appendix.

43

Limnology and Sedimentology of Proglacial Kusawa Lake
3.3.5.1

Core Alignm ent
The trace metal and sediment size profiles w ere used to align the tw o 30 cm

duplicate cores for cross-core correlation. Core A was subsampled in 1 cm increments and
Core B in 0.5 cm increments to 10 cm and 1 cm slices to 30 cm. As the tw o cores were
taken from the same ice hole in March 2010 in succession, they are assumed to have
comparable depths and sedimentation rates, though they are not identical.

The strong

association betw een sediment size and trace metal concentrations is well documented
(Horowitz, 1991; Sutherland, 2000; Outridge et al., 2005; Ye et al., 2013). W hen sediment
size and trace elem ent trends w ere compared directly, no significant correlations were
observed. However, when the surface of Core A was aligned to 1 cm on Core B (therefore,
losing the surface measurem ent for Core B), 41.5% of the trace metal profiles had a
significant regression relation with Dio (p<0.05). This alignment was further confirmed by
visual inspection and alignment of similar peaks found on both the trace m etal and
sediment size profiles. Therefore, all cross-core correlations and regressions used in this
study w ere constructed by aligning Core A at 0 cm to Core B at 1 cm.

3.3.5.2

Trace M e ta l Distribution
A 3D-PCA was prepared to analyze the overall trace metal distribution and variation

by sediment depth (Fig 3.8). W hile the majority of the depths group together, there are a
few notable exceptions. The surface sample at 0 cm divergence is likely due to sedimentw ater interactions occurring at the bed sediment surface. The samples from 9 cm and 10
cm also separate and are potentially affiliated with the 1911 sedimentation event.

44

Limnology and Sedimentology of Proglacial Kusawa Lake

Sediment Depth
■

0-4cm

g

■

5-9cm

o
5

T D
■
__ ■

10-14cm
1 5 -19cm
20-24cm

°

■

25-29cm

8

6

0.03

0.02
0.01

..

0.00

8

b
- 0.01

8
- 0.02

S
-0 .0 3

-0 .0 4
-0 .0 4

- 0.02

0.00

0.02

0.04

0.06

Axis 1 (25.48%)

Figure 3.8: PCA analysis of downcore trace metal distribution. Note shift betw een 5-7 cm,
likely associated w ith enrichment of redox sensitive metals, 9-10 cm, likely associated with
the 1911 sedimentation event and 26 cm, likely associated with the W hite River Ash layer.

One of the more interesting deviations occurs at 26 cm. A small increase in % clay is
noted here along w ith a concurrent decrease in Dio at 27 cm. This drastic change in trace
m etal composition is hypothesized to be the W hite River Ash (WRA) layer. The WRA is a
product of tw o volcanic eruptions that are estimated to have occurred at 1150 calendar yr
BP from M ount Churchill located in the southeast portion of the Wrangell Mountains in
Alaska.

The

resultant ash covered over 340,000

southwestern Yukon (Clague et al., 1995).

km2 of eastern Alaska and the

The ash layer is visible along clay cliffs and

rem em bered in local First Nations legends as it resulted in a major displacement of many
inhabitants (W orkm an, 1979, personal communication with local elders).

Chakraborty

(2010) identified the ash layer at 20 cm in a core taken from Kusawa Lake in region IV. The
annual linear sedimentation rate of that core was 0.022 cm /yr, compared w ith the current

45

Limnology and Sedimentology of Proglacial Kusawa Lake
core of 0.092 cm /yr (Chakraborty et al., 2010). The higher rate of sediment delivery to the
2010 core would account for observing the ash layer at a deeper level.
Depths 5-7 cm diverge to the far right of the PCA distribution due to large peaks in
M n, M o and As at 5, 6 and 7 cm, respectively. W hen these elements are excluded from the
trace m etal composition, the samples group with the m ajority of samples.

The peak

succession is likely due to changes in redox potential and is discussed in the next section.

3.3.5.3

Redox conditions revealed by trace m etal reductions
As no oxygen or sulfur species measurements were taken, trace metal reductions

w ere used to estim ate redox potentials and oxygen availability within the sediments. One
well-understood oxidation-reduction processes is the reduction of M n followed by Fe oxides
th at scavenge metals to the pore w ater across the oxic-suboxic boundary (Schaller et al.,
1997; Audry et al., 2006; Ye et al., 2013). The redox zones w ere estimated using the M n and
Fe reduction peaks, which occurred at 5.5 cm and 7 cm, respectively.

The resultant

proposed oxic, suboxic and anoxic boundaries are depicted in Figure 3.9, w here the
oxic/anoxic boundary is estimated to occur around 9 cm depth.

The assumed sigmoidal

depletion of oxygen across the suboxic zone and the onset of sulfate reduction in the anoxic
zone are also presented for reference. The subsequent M n and Fe oxyhydroxide scavenging
metals released w ere observed as major enrichment of As, M o, Cr, Zn and V (Appendix)
across the suboxic zone (Boyle, 2001).

46

Limnology and Sedimentology of Proglacial Kusawa Lake
Iron Fe (%)
7.0

7.5

8.0

8.5

9.0

9.5
-

2010

-

2002

0 2 Depletion

1982

E
o
a

Mn Reduction 1Q0-300mV

©

o

- 1959

c

o

Fe Reduction -100-100mV

E

a>
co
SUBOXIC -1 0 0 -3 0 0 m V

- 1940

- 1915
Sultate Reduction

- 1903

2000

3000

4000

5000

6000

Manganese Mn (ppm)

Figure 3.9: Determ ination of down core redox values based on M n and Fe reduction at 5.5
cm and 7 cm, respectively. The hypothesized trend in oxygen and hydrogen sulfide levels
are shown as dotted sigmoidal lines as suggested by Schaller (1997) and Manceau (1992).

47

Chapter 4 Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake
Sediments

4.1

Introduction
The carbon cycle is one of the most im portant on the planet as it drives every

process necessary for life along with oxygen and hydrogen. Carbon is responsible for joining
with other elements to form organic compounds such as lipids, carbohydrates and proteins
and accounts for half the dry weight of each cell (Nelson & Cox, 2000).

To achieve this

organic carbon undergoes several transformations depending on its state and association
w ith other elements (UNH, 2011). Carbon may also be present in inorganic forms, such as
charcoal, graphite or coal or as inorganic m atter compounds like calcite (CaC03) and
dolom ite [CaM g(C03)2]

(Schumacher, 2001).

Organic M a tte r (O M ) is a measure of the

compounds that make up dead cells and includes elements carbon, oxygen, hydrogen,
nitrogen, phosphorus and other organic and inorganic elements and molecules (Hedges &
Keil, 1995).

4.1.1

Sediment-Associated Organic Matter
W ithin aquatic systems, OM can associate with fine sediments to form floes an d /o r

aggregates th at may become suspended or dissolved and transported downstream with

48

Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake Sediments
lake and fluvial currents. This sediment-associated O M then settles out through the lake
w ater column and is deposited at the sedim ent-water interface of the lake bottom (von
W achenfeldt & Tranvik, 2008).

The deposited O M typically originates from

allochthonous, or autochthonous sources (Dodson, 2005).

either

Allochthonous O M includes

terrestrial sediments and soils th at are introduced to aquatic systems via shoreline erosion,
river discharge or more extrem e events such as land mass movements. This type of OM
consists of forest m aterial, degraded animal m atter, humic substances and soil-associated
prokaryotes and other primary consumers such as fungi. In aquatic systems, allochthonous
O M and sediment also originates from outer basin sources such as incoming rivers and
upper basin stream that feed into the lake.

Autochthonous O M is derived from primary

producers and autotrophs such as algal and bacterial m atter that reside in the lake basin
and benthic sediment (Brady & W eil, 1996).
All O M found within the sediments can be altered or degraded via microbes,
photochemical reactions, biotic alterations to the w ater column or benthic zone, chemical
redox reactions, diagenesis and other reactions (Hedges & Keil, 1995; Carrie et al., 2012).
The rate of bacterial degradation is influenced by several factors such as tem perature, redox
potential, nutrient availability and carbon sources (Staehr et al., 2012).

The influence of

climate change on primary productivity and bacterial metabolism has been the focus of
several studies. M any have observed an increase in algal m atter in recent sediment records
that is attributed to an increase in primary productivity caused by w arm er annual

49

Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake Sediments
tem peratures (Macdonald et al., 2005; McBean, 2005; Walsh, 2005; Arctic Council, 2007;
Outridge et al., 2007; Stern et al., 2009; Van Oostdam et al., 2009; Jiang et al., 2011).
Another product of O M

degradation is petroleum hydrocarbons.

W hen OM

contained in sedimentary rock is subject to subsurface extrem e therm al degradation at
constant tem peratures betw een 50"C and 175’C they can produce petroleum molecules.
These molecules may then migrate to a reservoir and undergo further alteration to yield
economical and retrievable hydrocarbons (Nunez-Betelu & Baceta, 1994). Several methods
and applications have been developed to determ ine the viability, state, presence and
quality of hydrocarbons associated with sedimentary rock formations; these procedures are
also relevant in environmental studies that evaluate more recent and less degraded
hydrocarbons and carbon compounds.

4.1.2

Organic M atter Applications
There has been great interest in developing methods to determ ine the amount,

source and make up of total organic carbon (TOC) in the environment. Total organic carbon
is a measure of the carbon components of O M in w ater, soils and sediments and other
environm ental samples (Dodson, 2005). Examples of some of these methods include: loss
on ignition (LOI), hydrogen peroxide digestion, isotope markers (6 13C) and chemical or
therm al production of C 0 2 (Schumacher, 2001; Carrie et al., 2012).

50

Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake Sediments
4.1.2.1

Rock-Eval Pyrolysis
Rock-Eval Pyrolysis is an analysis of TOC that was traditionally used to assess

economic hydrocarbon potential in sedimentary rock. The technique is based on therm al
release of carbon compounds in a two-step process: first, pyrolysis in an inert atmosphere
(nitrogen), followed by combustion in an oxic atmosphere (air) (Nunez-Betelu & Baceta,
1994; Lafargue et al., 1998). This characterizes TOC (w t % of sediment) as either Residual
Carbon (RC) or Pyrolysable Carbon (PC). Residual carbon consists of degraded material and
humic and detritus (dead organic m aterial) substances that are not readily available for
further metabolic processing.

Pyrolysable carbon consists of labile m aterial and smaller

molecules such as amino acids, fatty acids, vitamins, nucleotides and steroids (Carrie, 2012)
and is partitioned into hydrocarbon (HC) containing fractions, S I and S2, and oxygen
containing fractions, S3CC>2 and S3CO. In some studies, the S2 fraction is further defined by
separation into S240o (detected from 300-400°C during pyrolysis) and S2b (400-650°C).
Carrie et al. (2012) observed that the S2400 fraction is likely better suited for assessing the
interaction betw een elements bound to algal-derived organic sulfur compounds and labile
O M in sediments and soils (Lafargue et al., 1998; Disnar et al., 2003; Sanei et al., 2005;
Carrie et al., 2012). A summary of each of these fractions is outlined in Table 4.1.
Rock-Eval Pyrolysis was originally used to determ ine the HC ratio of kerogen, the OM
portion of sedimentary rock that is used as a measure of petroleum

potential in

hydrocarbon exploration. Kerogen can be broadly classified using a Van Krevelen Diagram,

51

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments
Table 4.1: Overview of O M fractions released and measured from Rock-Eval Pyrolosis 6
(version 6) th at are measured as TOC per g of dry weight sediment.

TOC (Total organic

carbon) = PC (pyrolysable carbon) + RC (residual carbon) and PC=S1+S2+S3C02+S3C0 (Carrie
et al., 2012).________________________________________________________________________

TOC

Fraction

PC
si

Measure

Temperature

mg HC g 1

0-300'C

Ul
* V

Description
Small (<500 Daltons) volatile and
easily degraded molecules such as
small sugars and lipids

S2b

mg HC g

40 0 -650‘C

M ore refractory larger kerogen-like
biomoecules

S3C02

mg C 0 2 g

100-650"C

C 0 2 containing molecules such as
carbohydrates, lignins

RC

wt %

650-850'C

Strongly refractory and refractory

__________________________ compounds___________

a calculated plot that segments the ratio of hydrogen to carbon (H/C) using Rock-Eval
measurements (Nunez-Betelu & Baceta, 1994). This m ethod has been reform ed to study
more recent sediments and assess the tem poral variations in O M as well as its content and
origin. M ore recent O M sources typically have lower H/C ratios, as they are associated with
less degraded material (Stern et al., 2009; Sanei et al., 2005 & 2014; Carrie et al., 2012). A
study by Carrie (2012) worked towards standardizing the Rock-Eval technique by comparing
known amino acid, sugar and lipid compounds, with source m atter such as terrestrial and
aquatic material and measuring their contribution to each O M fraction.

52

Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake Sediments
4.1.2.2

Rock-Eval Pyrolysis and Algal-Derived Organic M a tte r
Another recent application of the Rock Eval technique is to study the role of algal-

scavenging of Hg from the w ater column to lake sediments in Arctic and Subarctic lakes
(Outridge et al., 2007 & 2011; Stern et al., 2009; Sanei, 2005 & 2014). A key finding from
these studies is that in many of the lakes the S2 fraction of PC, related to the am ount of
algal-derived O M found in the sediments, is strongly correlated with the concentration of
total Hg (THg). These studies and others (Dirszowsky & Desloges, 2004; Carrie et al., 2009;
Hare et al., 2014) have generally observed a recent tem poral increase in S2 with correlating
Hg taken from northern lake sediments. The hypothesis is that the increase in S2 and THg is
due to climate change related increases primary productivity and a corresponding increase
in algal Hg-scavenging from the w ater column, which will be discussed further in Chapter 6.

4.2

Methods

4.2.1

Rock-Eval Pyrolysis
Rock-Eval 6 (Vinci Technologies, France) was used to assess the O M content of a 30

mg sediment subsample of each slice of Core B by Dr. Hamed Sanei and Dr. Jessie Carrie at
the University of M anitoba.
To determ ine the quantity and type of O M present, tw o heating stages progressing
at a rate 25°C/m in are required. The first stage takes place in a pyrolysis oven under inert,

53

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments
02-free conditions (nitrogen) to measure S I and S2 hydrocarbon pyrolysates (300-650'C ) by
flam e ionization detection.

At the same tim e, online infrared detectors continuously

measure the tw o S3 components (100-650*C), S3C0 and S3C02, which are released due to
therm al cracking of oxygen bearing compounds.

For the second step, the sample is

transferred to an oxidation oven and heated to 850"C to burn o ff the remaining O M and
release the RC fraction. TOC can then be determ ined by the sum of the total quantity of
carbon detected during pyrolysis, PC and oxidation, RC (Nunez-Betelu & Baceta, 1994;
Lafargue et al., 1998).

In this study, only the S I, S2 and S3CO fractions of PC and their

derivatives, HI and OICO, are considered.
From these components, the Hydrogen Index (HI) and carbon monoxide index
(OICO) can be calculated using the following equations (Carrie et al., 2012):
HI = 100 x S2/TOC (mg HC g 1 T O C 1)

(Equation 4.1)

OICO = 100 x S3CO/TOC (mg CO g 1 T O C 1)

(Equation 4.2)

For comparison w ith each other the S I, S2 and S3CO measurements w ere converted
to % TOC and % PC. To achieve this, S I and S2 were multiplied by 0.083, as 83% of the
signal is related to C by mass and the conversion of mg/g to wt%, i.e. 0.01. Using the same
logic, S3CO was multiplied by 1 2 /2 8 0 to determ ine the am ount of carbon in the CO signal
(Carrie et al., 2012).
To allow direct comparison to Core A, (sliced in 1 cm intervals throughout) the first
10 cm of Core B (sliced in 0. 5cm intervals) w ere averaged and aligned as described in
Section 3.3.5.1.

54

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments
4.3

Results and Discussion

4.3.1

TOC, RC and PC Fractions
Due to laboratory error, no measurem ent was recorded for RC at 4.5 cm; therefore

the 4.0-4.5 cm measurem ent was used to represent the 4-5 cm or 4 cm value for TOC, RC, HI
and 0 IC 0 2, OICO and OIRE6. This is not expected to drastically influence the overall results.
A downcore view of the major OM components determ ined by Rock-Eval Pyrolysis is shown
in Figure 4.1.
The am ount of TOC (Fig 4.1a) in the sediments was very low at an average of 0.70 ±
0.005 wt% (± SE), indicating the sediments were highly inorganic, as expected in a glaciolacustrine oligotrophic system. The greatest amount of TOC was observed in the top 5 cm
of sediment along w ith one notable peak at 15 cm. The %RC (Fig 4.1c) was quite high with
an average of 77.9% of TOC and made up 0.54 ± 0.003 wt% of the sediments, indicating that
majority of the carbon compounds are strongly resistant and refractory (Carrie et al., 2012).
Accordingly, the average labile O M or PC (Fig 4 .Id ) fraction had an average of 22.1% TOC
and contributed 0.16 ± 0.001 wt%.

4.3.1.1

S I and S2
The breakdown of PC components is shown in Figure 4.1 along with their

percentages of both TOC and PC in Table 4.2. S I (Fig 4.1e), being the most labile portion of
organic m atter, contributes the lowest am ount of carbon of all the PC components with a
mean of 5.37 ± 0.29 % PC. This is expected as S I consists of small compounds such as lipids,
55

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments
Max Temp <UC)

Total Organic Carbon TOC (wt %)

0.6

0.8

1.0

1.2

400

420

440

460

i_________I_________ I_________ I

O ~|

Figure 4.1: Overview of Rock
Eval variables measured from
Core B (sub-sectioned in 0.5

^L O

-

cm increments from 0-9.5 cm
and 1 cm increments from 1030 cm). Rock Eval fractions are
arranged as a matrix that is
reflective of the relationship
betw een them , where:
a: TOC is a m easurem ent of

d.

Residual Carbon RC (wt %)

C
04

0.5

U-

I__

0.6

I

0.7

1.0

1

|

Rock

Pyrolysable Carbon PC (wt %)
015

0 20

0.25

0 30

Eval

at,

b:

maximum

tem perature of pyrolysis;
TOC = c: RC + d: PC;

in

PC= e: S I + f: S2+ S3C02 (not

—o .

shown)+ g: S3CO;
h: Hl= S2/TOC and
i: OICO = S3CO/TOC

O -i

. C t-

f.

S1 (mg HC g~1)

e

g.

S2 (mg HC g~1)

S3CO (mg CO g“1)

0.1

02

0.3

1.4

0.6

l_

l

1

1

I________ I________ I________ I________ I

0.7

0.8

0.9

1.0

_

CM

H.

Hydrogen Index (mg HC g“1TOC~1)
70

80

i

I

90

*•

OICO (mg CO g"1TOC~1)

100

110

120

90

100

J

I__ __ I

I

I__

— __

110

120

I___

Q.

© _

Q lO

O_
CO

56

Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake Sediments
Table 4.2: Comparison of PC components as percentages of TOC and PC.

Fraction

smaller proteins and sugars that are quickly degraded through various mechanisms (i.e.
bacterial degradation) (Stern et al., 2009; Carrie et al., 2012).
The S2 (Fig 4 .If ) made up a higher percentage of the HC portion of PC than S I with a
mean of 27.84 ± 0.72% PC.

S2 is more stable than S I and is known to consist of algal-

derived lipid cell wall material and some minor inputs from plant material (Carrie et al.,
2012). As it is more stable and less prone to bacterial degradation, S2 is considered a better
representative of algal-derived OM in the sediments than S I.

Similar to S I and TOC, the

largest concentration of S2 is near the sedim ent-water interface with a 5.5-fold and 3.5-fold
post-1950 increase in S I and S2, respectively. Stern et al.'s (2009) 2005 core observed a 4fold, post-1950 increase in S I and S2 HCs that was attributed to an increase in primary
productivity related to climate change, specifically the annual tem perature increase of
2.14°C since 1970.
Separation of the S2400 and S2b fractions can be visually inferred by comparing S2 to
the maximum tem perature of pyrolysis (i.e. the tem perature at which the S2 signal was
detected) (Fig 4.2). There is a distinct separation between the surface layers to 8 cm along
with 14 cm that are detected below 400°C and the rem ainder of the core samples, detected
57

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments

S2400

Sediment Depth (cm)

0
1

♦

0-4cm, OXIC

▲

5-9cm SUBOXIC

■

10-29cm, ANOXIC

AW” 2 7 *'T B »

380

Max Temperature (°C)

Figure 4.2: Biplot of S2 vs. max tem perature, where sediment subsamples that released OM
below 400°C are designated S24oo and those above 400'C are S2b.

above 400°C. This indicates th at 0-8 cm and 14 cm samples are composed of more labile
O M and from 9 cm and below (excluding 14 cm) consist of more degraded and processed
O M (Disnar et al., 2003; Carrie et al., 2012).

4.3 .2

HI versus OICO
A Pseudo Van Krevelen diagram of HI vs. OICO (Fig 4.3) was used to infer between

allochthonous and autochthonous sources of O M .

Carrie et al. (2012) concluded that

58

Rock-Eval Pyrolysis of Organic M atter in Kusawa Lake Sediments
terrigenous and aquatic OM have significantly different HhOICO ratios for recent sediments.
However, they did caution that interpretations of the S3CO measurements are influenced
by mineral carbon and therefore decarbonation, which was not completed for this study, is
recom m ended.

They also noted that this interpretation is intended for recent, less

degraded O M , as the ratio is influenced by the state of degradation. Therefore, it will be
assumed th at decreasing HhOICO ratios are related to a higher degree of degradation. In
Figure 4.3, there is a downcore degradation trend (indicated by the oval) w here all samples
have an HhOICO <1.5, implying that all samples are from

a similar source, likely

autochthonous aquatic O M . Two exceptions to this are from the 7 cm and 26 cm depths,
which are visually considered outliers from

the general trend.

Note that in this

arrangem ent the 14 cm sample groups as expected, indicating that its divergence is not due
to a shift in OICO or HI and it is likely of the same or similar O M origin. At this stage, it is
assumed th at the 7 cm sample diverges due to the increase in proposed redox activity and
the 26 cm outlier can be attributed to the WRA layer. Both occurrences w ere identified in in
Section 3.3.5.2 in the trace metal PCA diagram (Fig. 3.6). Overall, with the exception of 26
cm, it appears th at the OM source is consistent and is inherently assumed to be
autochthonous aquatic O M with little to no major terrestrial or upper basin input. This is
also supported by the relatively stable particle size profile, excluding the 1911 large
sedimentation event, which will be addressed in the next section.

59

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments
120

slope ratios 1

100-

19

j*

S*dlm «nt Depth {cm)

39
49

CTJ

o
X

9

0-4cm , OXIC

▲

5-9cm , SUBOXIC

■

10-29cm , ANOXIC

X

169
21 B

24 B
15B

27!

6026 B

100

110

120

130

OICO (mg CO g^TOC"1)
'TOC'■
’)

Figure 4.3. Pseudo Van Krevelen diagram of HI vs. OICO. The dotted line is the 1 HhOICO
ratio line and the dot-dash line is the 0.5 HhOICO ratio line. The oval indicates a negative
downcore degradation trend that excludes samples from 7 cm and 26 cm.

4.3.3

Organic Matter and Particle Size
Autochthonous O M in the sediments is presumed to originate in the lake w ater

column and

by association w ith suspended mineral particles from

aggregates that

ultim ately settle to the lake bottom (Dodson, 2005). To further assess O M transport and
source, TOC was compared to fine particle content (% particles <3.9 pm ) in a bi-plot (Fig 4.4).

60

Rock-Eval Pyrolysis of Organic Matter in Kusawa Lake Sediments

t#

0 .9 -

15«

3»
Sediment Depth (cm)

2#

5
£

I

•

0-4cm , QXIC

▲

5 - 9cm, SUSOXIC

■

1 0 -29cm, ANOXIC

10B
0.7
2 6 ft

29 ■
28 ■

25 ■

166
17*
27 ■

0. 6 14*

18*
13*
1 9 ft

24

2 1 *»

28

32

36

Fine Sediment (% particles <3.9//m)

Figure 4 .4: Relation betw een TOC and fine sediment particles (r2=0.194, p<0.01). The oval
displays tw o visually observed clusters that include sediments from below 12 cm, excluding
15 cm.

The surface sediments to 4 cm and the sample from 15 cm have the highest TOC while the
m ajority remains clustered together as indicated by the oval. The outliers at 10 cm and 11
cm are linked to the 1911 major sedimentation event where the higher proportion of small
sized particles at 11 cm are associated with a lower TOC. Overall, the figure suggests that
the O M association w ith small particles is relatively low and constant being between 0.7 and
0.5%, indicating th at variable inputs of O M from the watershed are not apparent here but
rather the m ajority of the OM is autochthonous from in-lake sources. This also implies that

61

Rock-Evai Pyrolysis of Organic Matter in Kusawa Lake Sediments
settling of this m aterial did not involve O M -m ediated aggregation or flocculation and that
sedim entation from the watershed does not deliver large amounts of O M to the lake
bottom . These findings correspond with the assumption from Figure 4.3 that majority of
the O M originates from autochthonous sources and is likely associated w ith primary
productivity and bacterial m atter.
One notable discrepancy is the high TOC %wt at 15 cm core depth.

W ithin the

assessed OM profiles a 15 cm sample peak is observed in all measurements except S I, the
most labile and easily degraded O M fraction.

Figure 4.4 also indicates that there are no

obvious allochthonous inputs to the core with the exception of the WRA layer at 26 cm
depth, making it likely that the sediments at 14-15 cm contain some unknown labile
authochthonous O M accumulation.

4.4

Summary o f Rock-Eval Pyrolysis
Evaluation of the sediment core using Rock-Eval Pyrolysis 6 suggests that the O M is

predom inately from autochthonous limnetic sources. The surface sediments from 0-4 cm
are associated with recently deposited labile OM and sediment associated O M found below
9 cm in the anoxic zone is largely refractory with the exception of some unknown possibly
labile O M accumulation at 14-15 cm depth. Finally, the WRA layer was identified as an
allochthonous O M source observed at 26 cm depth.

62

Chapter 5 Total Bacteria and Sulfur Metabolizing Bacteria in
Kusawa Lake Sediments

5.1

Introduction

5.1.1

Review of Bacterial Metabolism
Bacteria are capable of using several different compounds and molecules for their

metabolism.

How each microorganism generates energy is dependent on oxygen

availability and w hat molecule it uses as its source of electrons.

In aerobic respiration,

oxygen is used as the term inal electron acceptor, while in anaerobic respiration a
compound other than oxygen such as nitrate (NO 3 ) or sulfate (S0 42 ) is used. Heterotrophic
bacteria are capable of using a variety of inorganic and organic electron acceptors under
either oxic or anoxic conditions for their metabolism. W hile, obligate or facultative aerobes
require high or limited levels of oxygen and conversely obligate or facultative anaerobes
require strictly anoxic or tolerate suboxic conditions, respectively.
W ithin lake sediments the mode of bacterial respiration is primarily driven by
sediment depth, oxygen availability and redox potential (Fig 5.1). This relates to downcore
microbial m ediated OM degradation and the corresponding available electron acceptors
and donors.

First, large labile OM macromolecules are deposited at the sedim ent-w ater

interface and subject to microbial aerobic respiration and hydrolysis within the oxic and

63

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
suboxic zones.

These marcromolecules are broken down into O M monomers such as

nucleic acids, sugars and amino acids. As oxygen is depleted through the suboxic zone, the
molecules are ferm ented to produce carboxyl groups, such as lactate and propionate and
used as substrates by microbes in the suboxic and anoxic sediments.

The final OM

breakdown of carbon takes place in the strictly anoxic lower sediments w here CO2 is
reduced to either m ethane (methanogenesis) or acetate (acetogenesis) and carbon
mineralization may occur (Jprgensen, 2000; Madigan et al., 2003).

PRIMARY
PRODUCTION
CO,.
NH,\
HPO/,
_ etc.

WATER

METAZOAN &
MICROBIAL 0,
RESPIRATION

f

MACRO \
MOLECULAR
ORGANIC
V MATTER 1

OXIC
ZONE
NO,
REDUCTION
Mn(IV)
REDUCTION

EXOENZYMATIC
HYDROLYSIS
/

SUBOXIC
ZONE
Fb(III)
REDUCTION

SUGARS \

' AMINO ACIDS '
LONG CHAIN
FATTY ACIDS
.NUCLEIC ACIDS,
BURIED
ORGANIC
MATERIAL,

so ■
f

(

REDUCTION

<£butyrate}

(

CH,
-C p ro p io n a te
ACETATE

FERMENTATION

CH,
FORMATION

SULFATE
REDUCTION
ZONE
METHANOQENIC
ZONE

Figure 5.1: Schematic microbial processing of organic m atter in lakes sediments and the
associated reduction pathways and resultant products of various modes of metabolism.
From Jprgensen (2000).

64

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.1.2

Sulfur-Metabolizing Bacteria
There has been extensive study on sulfur-metabolizing prokaryotes from both

Bacteria and Archaea.

Sulfur-Metabolizing Bacteria (SMB) are studied for their metabolic

diversity, environm ental ubiquity and usefulness in biotechnology applications such as soil
rem ediation, m etal precipitation and w astew ater treatm ent (M uyzer & Stams, 2008). They
are one of the key groups of organisms that drive the global sulfur cycle and also play a
significant role in the cycling of carbon, nitrogen and various metals (Barton 8t Hamilton,
2007).

5.1.2.1

Sulfur Origin
In freshwater environments, elem ental sulfur (So) originates from autochthonous

bacterial storage of sulfur globules, allochthonous w eathered rocks, terrestrial O M , wildfire
and volcanic ash and anthropogenic sources such as fossil fuels, w astew ater discharge, long
range atmospheric transfer and acid mine drainage (Holm er 8i Storkholm, 2001).

In

oligotrophic lake sediments, the overall level of sulfate, the most oxidized form of sulfur, is
typically <300m M , while it can occur at 700-800 m M in some eutrophic lakes (Pester et al.,
2012).

W ithin

low

productivity

lake

sediments,

autochthonous

sulfur-species

and

compounds generally originate from organic sulfur compounds, such as the proteins of
aquatic organisms (Holm er & Storkholm, 2001).

65

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5 .1.2.2

Sulfur Cycling
Sulfur-metabolizing bacteria (SMB) are capable of either the reduction or oxidation

of various inorganic chemicals such as iron, nitrogen, hydrogen and phosphorus, however
they are most recognized for their role in sulfur cycling. Sulfur is typically involved in redox
processes (Norici et al., 2005) with an overview of the redox couples involved in the sulfurreduction pathway are show in Table 5.1.

_

_

_

_

_

_

_

_

_

_

p

_

_

_

_

Table 5.1: Overview of redox potentials E° of
S20 32‘ +2e' -+ S 0 3 + H2S

-402

reduction intermediates of the sulfate reduction
cycle, where £° are at pH 7 (Thauer et al., 2007).

S 0 42 + 8 e — H2S

-217

To utilize sulfur, SMB employ a suite of enzymes th at catalyze either the oxidation
from hydrogen sulfide to sulfate or the reduction in reverse. The different species of SMB
and their mode of respiration are discussed in the next section.
protein clusters to drive sulfur metabolism (Fig 5.2).

SMB use four primary

The first three are located in the

cytoplasm and utilize sulfite as an interm ediate as follows: (i) the first reaction in the
reduction pathway is catalyzed by sulfate adenylyltransferase (SAT) for S 042‘<-»adenosine 5'phosphosulfate (APS); (ii) APS reductase (Apr complex) catalyzes APS*-»S03 ; (iii) and the
final reaction is facilitated by dissimilatory sulfite reducatase (Dsr complex) for S 0 3 <-»H2S.
The fourth protein is located in the periplasm, where sulfur and thiosulfate are oxidized to
sulfite using the SOX protein complex that converts So«-»S20 32 <-»S042' (Madigan et al., 2003;
M uyzer & Stams, 2008; Hokenbrink et al., 2011; Stewart et al., 2011).

66

a-

b.

Sulfur-Oxidizing Bacteria
S20 32-

\

\

so42
^

so42

H,S

t

Outer Membrane
HYD
sax comp/ex

2e- +H

5^

C Y T eS

RSn„H

flavocytochrome c
sulfide dehydrogenase

SO''' +H
orH-S

APS reductase

Pwlplasm
C ytoplasm

sulfide:quinone reducta •

NADP’
+

acceptor

NADPH + H

H ,0

6e + 6H*

H ,S —

—

PR

SO .

sulfate adenylyltransferase
(ATP sulfurylase)

dissimilatory
sulfite reductase

Figure 5.2: Generalized overview of the SMB enzyme pathw ay for sulfate reduction and sulfide oxidation,

a: Sulfur-

Oxidizing Bacteria (SOB) th at take up H2S or S20 32 as an electron donor to produce S 0 42 through the D sr/A pr/SA T or Sox
pathways, respectively (adapted from Stewart et al., 2011; Gregerson et al., 2011); b: Sulfate-Reducing Bacteria (SRB)
take up extracellular S 042' and reduce it to H2S through the SAT/Apr/Dsr pathw ay to produce S 0 42' th a t is excreted
(adapted from Cao e t al.. 2014).
<31

'si

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments

h 2s

Sulfate-Reducing Bacteria

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.1.2.3

Organic M a tte r Metabolism
In highly active environments, such as shallow marine systems, SMB can contribute

to more than 50% of organic carbon mineralization (Jprgensen, 2000). They can be broadly
categorized into five groups based on their carbon metabolism and preferred electron
acceptor or donor: (i) phototrophic sulfur-oxidizing bacteria; (ii) sulfur-oxidizing bacteria
(SOB); (iii) sulfur(S0)-reducing bacteria (SB); (iv) sulfate-reducing bacteria (SRB) and (v)
organic sulfur utilizing bacteria (Giovannoni & Stingl, 2005; Sievert et al., 2007). A general
overview of sulfur-based reactions coupled with O M breakdown is depicted in Figure 5.3
(Zhou et al., 2011; Pester et al., 2012).
Anoxygenic photosynthetic Purple Sulfur Bacteria (PSB) and Green Sulfur Bacteria
(GSB) contain highly efficient bacteriochlorophylls that drive photosynthetic growth and
require very little light. Manske et al. (2005) successfully cultured GSB from a depth of 100
m in the Black Sea where only 0.00075 pmol quanta m '2 s'1 of light was measured, the
lowest reported values for photosynthetic growth at that tim e. Cold and low light adapted
photosynthetic proteins have also been described for a strain of GSB isolated from an
Antarctic Lake (Ng et al., 2010). Phototrophic SMB use H2S as an electron donor for C 0 2
fixation and store globules of elem ental sulfur that are eventually oxidized to produce SO42'.
They can also use S2C>32 or Fe2+ as an electron donor instead of H 2S. One notable difference
betw een PSB and GSB is th at the elem ental S0 globules produced by GSB are stored outside
of the cell, whereas PSB store them intracellularly (Madigan et al., 2003; Dahl et al., 2005;
Hokenbrink et al., 2011).

68

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments

ffwtwyntheMand
chemobthotrophy j

Atmtnpfwm deposition

5* reduction «md m lphur

I Chemkal oxidation

1

• d te p ro p o rtK w u M to n

Sulphur <fcproportioft*tion
Chem oiithotrophtc oxidation :
ChemofrtHotrophic oxidation

* [ * £ £ * ] -------* Aerobic end aneerohlc recpicedon ^

Figure 5.3: Schematic of the role of SMB in the carbon and sulfur cycle to provide an
overview of coupling mechanism of using sulfate as an electron acceptor to degrade
ferm ented organic m atter (i.e. simple sugars such as lactate) to produce hydrogen sulfide
and either CO2 or acetate. Figure from Zhou (2011).

Chemolithotrophic SOB oxidize a broad array of reduced inorganic sulfur-compounds
such as S0 and H2S via the Dsr and Sox enzymes or the reverse sulfate-reduction pathways to
produce SO42' and H+.

W hile the majority of these species are aerobic there are also

facultative and obligate anaerobes that oxidize sulfur and are commonly found in
hydrotherm al vents or hot springs (Ruby, 1981; Madigan et al., 2003). SOB can use an array
of carbon substrates, though it appears most are autotrophic and therefore, only consume
C 0 2 (Cypionka, 2000).
Sulfate-reducing bacteria (SRB) are facultative and obligate anaerobes that reduce
S 042' to H2S and can be divided into tw o groups based on their completeness of carbon
breakdown.

The first group degrades carbon compounds to acetate and the second

69

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
completely oxidizes to carbon dioxide.

Common SRB substrates include H2, acetate,

ferm entation products (i.e. succinate, malate, lactate, pyruvate and form ate), aromatics (i.e.
toluene and benzene) and alcohols, particularly ethanol and propanol. Several species th at
completely oxidize carbon compounds to C 0 2 use fatty acids as electron donors, and are
also capable of autotrophic growth using H2 as their electron source. Under both regimes,
produced H2S is either assimilated and converted to amino acids or dissimilated and
excreted from the cell (Madigan et al., 2003).
M any SRB are found in oxic environments and are presumed to still be utilizing the
sulfate-reduction pathway, though this mechanism is poorly understood (Sass & Cypionka,
2007). Some SRB are capable of using oxygen as an electron acceptor though it does not
sustain growth and can lead to the production of oxygen radicals along with increased pH,
which in turn is toxic. This suggests that it is not the oxygen itself the SRB are sensitive to
but the products of its metabolism (Cypionka, 2000).

Several mechanisms have been

proposed to describe how SRB cope with oxygen stress in sediments: (i) they utilize various
enzymes that detoxify oxygen radicals (Cypionka, 2000); (ii) they form cell aggregates with
anoxic interiors (Sass et al., 1998); (iii) they migrate diurnally to avoid high oxygen
concentrations produced by photosynthetic organisms during the day (Krekeler et al., 1998)
or (iv) they employ a mechanism called aerotaxis, where several cells migrate to some
oxygen threshold with an adequate substrate supply and remove oxygen to restore anoxic
conditions (Cypionka, 2000).

70

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
Sulfur (So)-reducing bacteria (SB) are unable to reduce sulfate to sulfide (S 042 - * H 2S)
but they can reduce elem ental sulfur to sulfide and therefore live in syntrophy (shared use
of metabolic products between various bacterial species) with bacteria that oxidize H2S-»So
(Madigan et al., 2003; Stahl et al., 2007).
Under

extrem e

methanogenesis

and

anoxic

conditions

acetogenesis

found

occurs.

much

Here,

deeper

in

the

sediments,

autotrophic

SRB

com pete

with

methanogens and acetogens for C 0 2, as majority of the bacteria present are autotrophs
th at use H2 as an electron donor and C 0 2 as their electron acceptor (Madigan et al., 2003;
Thauer et al., 2007).

5 .1.2.4

Sulfur cycling in Oligotrophic Lakes
Under low sulfate conditions, such as in oligotrophic lakes, sulfate levels are typically

below 1 m M and SMB sustains growth by continuously cycling H2S to S 042 across
oxic/anoxic interfaces within the sediment or w ater column. As a result, SMB communities
support each other w here SOB and phototrophic bacteria provide S 042' for SRB and SB,
which in turn provide S0 and H2S for oxidation. For this reason, many SRB are found near
the oxic/anoxic interface. As a consequence, in low sulfate environments, growth of SMB
populations is restricted by the concentration of metabolically available sulfur compounds.
(Holm er & Storkholm, 2001; Thauer et al., 2007).

71

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.1.2.5

M e ta l Ion Metabolism
In aquatic systems, SRB interact with various metals to influence their speciation,

solubility and mobility. This is driven by a unique metal interaction with sulfides to produce
bioprecipitated metal-sulfides, a phenomenon that is applied in biotechnical methods to
remove metals from w astewater, such as acid mine-drainage runoff. Common metals that
bind with sulfide include Cd, Co, Cu, Fe, M n, Hg, Ni and Zn (Hockin & Gadd, 2007).
SRB are also capable of using Fe(lll) and M n(IV) as term inal electron acceptors
instead of sulfate under anaerobic conditions and play an im portant role in the redox
cycling of Fe2+/F e 3+ and M n 2+/M n 4+ across the oxic/anoxic interface. One hypothesis is that
reduced sulfide can be re-oxidized to sulfate by coupling and stabilizing with M n and Feoxides in the suboxic transition zone (Thamdrup et al., 1994; Thauer et al., 2007).

5.1.2 .6

Phytogeny
An overview of the diversity and metabolic preferences of SMB and their distribution

within 11 of the 21 phyla of the Domain Bacteria is shown in Figure 5.4. Traditionally the
16S rDNA gene was used to detect unculturable SMB until degenerate primers were
designed to target AprA, the alpha subunit of APS reductase and DsrAB, the alpha and beta
subunits of the dissimilatory sulfite reductase enzyme complex (W ager et al., 1998; Klein et
al., 2001; Leloup et al., 2009). Both Apr and Dsr gene targets recover representatives from
almost all SMB lineages (W agner et al., 1998; Klein et al., 2001; Loy et al., 2004; Stahl et al.,
2007), although some SMB may not contain AprA and/or DsrAB if sulfur cycling occurs via

72

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
Euryarchaeota
SRB: Archaeoglobales

ARCHAEA

Crenarchaeota

SB: Thermococcales,

SB: Thermoproteales

Thermoplasmatales

SB: Desulfurococcales
SOB: Sulfolobales

OS: some methanogens
M ethanosarcina

BACTERIA
Nitrospiracea
SRB: Thermodesulfovibrio

a-Proteobacteria
OS: Roseobacter-cladeSAR-11
SOB: Paracoccus
PS: Various genera

Aquificae
SOB: Aquiflex,
Persephonella

P-Proteobacteria

SB: Desulfobacterium,

SOB: Thiobacillus„Thiomonas

Thermovibrio

OS: Alcaligenes
PS: Rubrivivax, Rhodoferax

SRB: Thermodesulfobacteriaceae

Y-Proteobacteria

SRB: Therm odesulfatator

SOB: Various genera

Thermodesulfobacterium

PS: Chloroflexaceae

OS: M ethlophaga, Pseudomonas
PS: Chromatiaceae,

PS: Chloroflexus

Ecothiorhodospiraceae

Firmicutes
SRB: Desulfotomaculum,
Desultosporosinus

6-Proteobacteria

SB: Am moniflex

SRB: Desulfovibrio, Desulfotalea,
Desulfonema, Desulfobacter
SB: Desulfuromonas, Desulfurella,

PS: Chlorobiaceae

Geobacter

e-Proteobacteria

OS: some sulfate reducers,

SOB: Arcobacter, Thiovulum, Sulfurimonas

Desulfovibrio

SBSulfurosprillum, Nautilia, Caminibacter

Figure 5.4: General representation of phylogenetic tree that depicts the broad diversity
sulfur-metabolizing bacteria and archaea and their major phylogenetic lineages and method
of sulfur metabolism.

PS: Phototrophic Sulfur Bacteria; SOB: Sulfur-Oxidizing Bacteria and

Archaea; SB: Sulfur (So)-Reducing Bacteria and Archaea; SRB: Sulfate-Reducing Bacteria and
Archaea; OS: Organic Sulfur Utilizing Bacteria and Archaea.

Adapted from Sievert et al.

(2007).

73

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
another enzyme pathway (Stahl et al., 2007). In Figure 5.4, the DsrAB gene is not confirmed
in all of the species shown; it has been detected in phyla that contain phototrophic SOB,
SOB or SRB and is unconfirmed in phylum e-proteobacteria, Aquificae and Crenarchaeota
(W agner et al., 1998; Stahl et al., 2007; Zhou et al., 2011).
The evolution of the dsrAB gene is dominated by vertical transmission along each
lineage with a few lateral transfer events from 6-Proteobacteria species to Archaeoglobus,
Thermodesulfobacterium, phototropic SMB and members of the Gram-positive Firmicutes
(Klein et al., 2001). The exact mechanism of this transfer is unknown with theories that it
occurred through xenologous (horizontal gene flow) displacement of a single gene in situ
(Omelchenko et al., 2003) or a large 'metabolic island' containing sulfate-reduction genes
was transferred by some mobile elem ent in one single event (Mussmann et al., 2005). As a
result, when constructing dsrAB dendrograms, tw o distinct lineages of Firmicutes species
are commonly observed (Castro et al., 2002; Dhillon et al., 2003; Kaneko et al., 2007;
Schmalenberger et al., 2007; W u et al., 2009).

74

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.2
5.2.1

M ethods
Nucleic Acid Extraction
Nucleic acids were isolated from each 1 cm slice of Core A from 1-30 cm in duplicate

using the M O BIO Laboratories Inc. Ultra Clean Soil DNA Isolation Kit (M O BIO Laboratories
Inc., 2010).

Approximately 0.25-1 g of dry sediment was weighed on a precision scale

before extraction.

DNA concentration was determ ined using the Quanti-iT dsDNA HS kit

w ith the Qubit fluorom eter (Invitrogen, 2010b) and the Nanodrop N D -1000 spectrometer;
2 pL of tem plate was used for both apparatuses. DNA concentration decreased with sample
depth until no DNA could be detected past 22 cm.

The low extraction efficiency was

presumed to be from a combination of increased DNA degradation at lower levels and
possible changes in sediment composition that impeded DNA extraction (Frostegard, 1999).
To increase extraction efficiency the M O BIO Laboratories Inc. Power Soil DNA Isolation Kit
(M O BIO Laboratories Inc., 2009) was used for samples from 11-30 cm. W hile extraction
concentrations w ere still low, they were detectable on the Qubit fluorom eter.
All extractions w ere cleaned and concentrated using the Favorgen Genomic DNA
Clean-up kit (Favorgen Biotech, 2009). The 50 pL of cleaned tem plate was eluted from the
full 50 pL and 100 pL original tem plate obtained from both the Ultra Clean and Power Soil
kits, respectively.

75

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.2.2

Sequence Analysis of DsrAB
The dsrAB gene was PCR-amplified using primers Dsr-lF-GC (5'-ACS CAY TGG AAR

CAC G-3') and Dsr-4Rdeg (5'-GTG TAR CAG TTD CCR CA-3') to yield a 1.7 kb fragm ent as
described by Klein (2001) and W agner (1998).

The optim al MgCI2 concentration was

determ ined to be 2.5 m M , w here higher concentrations increase annealing efficiency but
decreased specificity.

A touch down gradient for the annealing tem perature from 65°C-

59°C was also used to increase annealing efficiency as well as decrease specificity. The final
PCR reaction conditions using the Platinum Taq PCR Kit (Invitrogen, 2010c) w ere as follows:
1 pL of 10 pm ol/pL of each primer, 5 pL of DNA tem plate, 2.5 pL of 10X PCR buffer, 1.25 pL
of MgCI2 (2.5 m M ), 0.675 p i of BSA (3 mg/mL), 0.2 pL of deoxynucleoside triphosphate mix
(dNTPs) (2 m M [each] dATP, dCTP, dGTP and dTTP), 1 Unit of Taq Platinum polymerase and
nuclease free w ater to the final reaction volume of 25 pL. Reactions w ere distributed to 0.2
mL PCR tubes and amplified under the following conditions: initial denaturation of 90 s at
94°C followed by 30 cycles of 15 s at 94°C, touch down gradient from 65°C-59°C for 20 s, 54
s at 72°C and a final extension of 1 min at 72°C.
Clone libraries w ere constructed using TOPO® XL PCR cloning kit (Invitrogen, 2010a).
The initial attem p t of gel purification using the Crystal Violet provided w ith the TOPO XL
cloning kit was not successful due to low PCR concentrations and inability to visualize the
band. Alternatively, the Qiagen QIAQuick Gel Extraction kit (Qiagen ®, 2008) was used as
per the manufacturer instructions to isolate the 1.7 kb fragm ent. Successful PCR reactions

76

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
w ere combined for ligation as follows: the three Ponar Grabs and core slices from 1-3 cm, 45 cm, 6-7 cm, 8-9 cm, 10-13 cm, 14-17 cm, 18-21 cm, 22-25 cm and 26-30 cm.

Cloning

attem pts using the purified combined PCR products with the TOPIO E. coli strain provided
w ith the TOPO XL kit w ere unsuccessful due to the long-term storage and improper handling
of the cells th at rendered them no longer viable. Instead, E. coli JM 109 cells (provided by B.
M urray, UNBC) w ere used to transfect the ligated TOPO XL plasmid and PCR products. Cells
w ere grown overnight at 37°C on LB-Kanamycin (50 pg/mL) - X-Gal (80 pg/mL) - IPTG (0.05
mM)

plates and

blue-white screening was used to determ ine

Approximately 1-12 successful bacterial colonies were

successful reactions.

picked from

each plate and

inoculated into 5 mL of LB Broth with 50 pg/m L Kanamycin and incubated overnight at 37°C
with shaking at 225 rpm. Plasmids w ere isolated using the QIAgen MiniPrep Kit as per the
manufacturer's instructions (Qiagen ®, 2006). The plasmid concentration was determ ined
on the Nanodrop. Insert size was verified using universal M 13 forward and reverse primers
under the following conditions: 0.5 pL of 10 pm ol/pL of each primer, 1 pL of 1:10 plasmid
dilution, 2.5 pL of 10X PCR buffer, 0.75 pL of MgCI2 (2.5 m M ), 0.25 pL of deoxynucleoside
triphosphate mix (dNTPs) (2 m M [each] dATP, dCTP, dGTP and dTTP), 1 Unit of Taq Platinum
polymerase and nuclease free w ater to the final reaction volume of 25 pL. Reactions w ere
distributed to 0.2 mL PCR tubes and amplified under the following conditions: initial
denaturation of 93°C for 5 min followed by 35 cycles of 94°C for 30 s, 56°C for 30 s, 72°C for
90 s and a final extension of 72°C for 5 min (Invitrogen, 2010a).

77

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
Plasmids with the correct fragm ent length w ere sequenced using the universal
primers M13F-Forward and M13R-Reverse to sequence the dsrA and dsrB subunits found at
either end of the 1.7 kb fragm ent. Plasmids w ere prepared for sequencing in 96-w ell plates
w ith ~450 ng of plasmid, 10 pmol of prim er to a final volume of 15 pL and sent to Fragment
Analysis and DNA Sequencing Services at the University of British Columbia, Okanagan in
Kelowna, BC for sequencing using the ABI 3130x1 Genetic Analyzer (UBC Okanagan, 2012).
Obtained

sequences w ere

manually

reviewed

using CodonCode Aligner 4.04

(CodonCode Corporation, 2013) to optimize and verify the nucleotide signal.

Sequences

w ere then compared to the online Basic Local Alignment Search Tool (BLAST) to verify the
presence of the dsrA or dsrB gene and select the nearest related sequences from the
GenBank database for alignment and comparison. All nucleotide sequences obtained in this
study and from GenBank w ere aligned using MUSCLE (a multiple sequence comparison by
log-expectation com puter program) in MEGA 5.2.2 (Tamura et al., 2011). Sequences were
then translated to amino acids and the best-fit MEGA 5.2.2 model analyzer was run to
determ ine the optimal model to construct a maximum-likelihood tree using 500 replicate
bootstrap analysis.

The same tree building process was done for the un-translated

nucleotide sequences to visually assess any ambiguities. The tree was grouped into clusters
based on 87% amino acid sequence homology and into operational taxonomic units based
on 97% amino acid sequence homology. Environmental conditions such as climate, sulfate
and oxygen levels of the sequences obtained from the BLAST database w ere also recorded
(W agner et al., 1998; Klein et al., 2001; Dhillon et al., 2003; Leloup et al., 2009).

78

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.2.3

Quantification of DsrA
Sulfur-metabolizing bacteria w ere quantified using the SyBr® Green qPCR method by

targeting the dsrA gene (Applied Biosystems, 2006).

Previously prepared Plasmid 13167

from the cloning experim ent was used as the standard and the plasmid concentration was
confirmed using the Qubit. The plasmid was linearized using restriction enzyme Hindlll to
prevent supercoiling and enable qPCR amplification as follows: 5 pL enzyme buffer, 1 pL
Hindlll, 5 pL plasmid and fill to 50 pL with nuclease free H20. The reaction was incubated at
37°C using a heat block for 1 hr (Promega, 2011).
A 6-tim es 10-fold serial dilution of the standard was run on every plate in duplicate
to determ ine the standard curve used for absolute quantification. A duplicate 'no-tem plate'
negative control was also run for each plate to account for contamination and prim er
dimers.

A melting curve for each plate was run from 65°C-95°C to assess non-specific

amplification and the presence of primer dimers.
The dsrA genes present is each DNA sample was amplified and detected in duplicate
using the Bio-Rad MiniOpticon™ Real-Time PCR Detection System (Bio-Rad, 2014).

The

primers used are described by Kondo et al. (2004) and Leloup et al. (2007) and are as
follows: forw ard DSR-F+ (5'-ACS CAC TGG AAG CAC GGC GG-3') and reverse DSR-R (5'-GTG
GMR CCG TGC AKR TTG G-3') to yield a 221 bp fragment.

Primers w ere optimized by

running variations of concentration for each forward and reverse prim er at 50 nM , 300 nM
and 900 nM . The lowest positive detection concentration was determ ined to be 50 nM for
both the forward and reverse primers.

79

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
Annealing tem perature was optimized to 63°C and the following conditions w ere
used for all unknown samples: in 48-w ell plates, 2 pL DNA tem plate or plasmid standard,
12.5 pL SyBr® Green PCR M aster Mix, 50 nM DSR-F+ and DSR-R primers and filled to 25 pL
with nuclease free H2O and run on the RT-PCR machine at 50°C for 2 min, 94°C for 15 min
and 35 cycles of 94°C for 15 s, 63°C for 60 s and 72°C for 45 s (W agner et al., 1998; Kondo et
al., 2004; Applied Biosystems, 2006; Leloup et al., 2007).

5.2.4

Quantification of 16S rDNA
Total Bacteria (Tbac) w ere quantified using the SyBr® Green qPCR method by

targeting the 16S rDNA gene. The standard was isolated from a stock of E. coli that was
grown overnight at 37°C on LB plates and one colony was picked to inoculate 5 mL LB Broth,
which was grown overnight at 225 rpm at 37°C. Total genomic DNA of the inoculation was
isolated using the M O BIO UltraClean® Microbial DNA Isolation Kit (M O BIO Laboratories
Inc., 2010).

DNA was quantified using the Qubit and used as the standard for Tbac

quantification.
A 6-times 10-fold serial dilution of the standard was run on every plate in duplicate
to determ ine the standard curve used for absolute quantification. A duplicate no-tem plate
negative control was also run for each plate to account for contamination and prim er
dimers. A melting curve for each plate was run from 65°C-95°C to determ ine non-specific
amplification and the presence of prim er dimers.

qPCR products w ere amplified and

80

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
detected in duplicate using the Bio-Rad MiniOpticon™ Real-Time PCR Detection System.
Two universal primers w ere used as described by Lopez-Gutierrez et al. (2004) to detect
mem bers of Eubacteria to amplify a 174 bp region and are as follows: 341f (5V-CCT ACG
GGA GGC AGC AG-3V) and 515r (5V-ATT CCG CGG CTG GCA-3V). Primers w ere optimized by
running variations of concentration for each forward and reverse prim er at 50 nM , 300 nM
and 900 nM . The lowest positive detection concentration was found to be 300 nM for the
forw ard and reverse primer.
Annealing tem perature was also optimized to 56°C. The following conditions were
used for all unknown samples: in 48-well plates, 5 pL DNA tem plate or plasmid standard,
12.5 pL SyBr® Green PCR M aster Mix, 300nM 341f and 515r primers and filled to 25 pL with
nuclease free H20 and run on the RT-PCR machine at 50°C for 2 min, 95°C for 15 min and 40
cycles of 95°C for 15 s, 56°C for 30 s and 72°C for 30 s (Lopez-Gutierrez et al., 2004).

5.2.2
5.2.2.1

qPCR Calculations and Analysis
qPCR Calculations
The following equations and variables were used to calculate the final DNA

concentration per gram of sediment at each sediment depth:
•

Cf, the num ber of cycles required to reach the certain threshold florescence measured
and reported by the RT-PCR detection system;

81

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
•

E, amplification efficiency was determ ined using the following equation:

E = 1 0 (-1 /a )

(Equation

5.1)

w here, a is the slope of the curve calculated by the known concentration of the
serial dilution of the sample vs. the threshold florescence.
•

Environmental inhibitors of amplification efficiency (versus the cleaned and concentrated
plasmid tem plates) w ere corrected for using the One-Point-Calibration m ethod described
by Brankatschk (2012) where,
£
N o s a m p le =

N 0 s ta n d a rd x

r
g

t sta n d ard
t sample
sam ple

(Equation 5.2)

w here, N0 is the concentration of the standard and tem plate in ng/pL of DNA based
on the known tem plate concentration. Therefore the ng/pL DNA of each sample can be
calculated to incorporate the differences in efficiency due to environmental impurities.
1 The num ber of copies of genes detected is then calculated by:

C o p y# = ctarget x

- £f£^ d-

(Equation 5.3)

*F r a g m e n tx “ bp

w here, Copy # is the num ber of gene copies detected per pL, ctorgef is the estimated
copy num ber DNA strand (for 16S rDNA estimate 3.6 copies/cell and dsrAB one copy per
cell), N a is Avogadro constant (6 .0 2 2 x l0 23 bp/m ol); I'fragment is the length of the amplified
DNA fragments, in this study 174 bp for 16S rDNA and 221 bp for dsrA and M bp is the
average w eight of a double-stranded base pair (6.6xlO n ng/mol).

82

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
•

From this the num ber of detected gene copy num ber per gram of sediment was
calculated by determining the dilution factors imposed by the DNA extraction, cleanup
and PCR steps and dividing by the grams of sediment analyzed (Leloup et al., 2009;
Applied Biosystems, 2010; Brankatschk et al., 2012).

5 .2.2.2

qPCR Analysis and Statistics
The SMB and Tbac ratio was calculated by dividing each of the four SMB qPCR

replicates with each of the four Tbac qPCR replicates to yield 16 ratios that were multiplied
by 100 to reflect percent and were arranged into a boxplot.

The median of each 16

replicate set was used as the final percentage to assess its downcore relation with other
variables.
All variables used in correlation or regression analysis were tested for normality
using the Sharpiro-Wilk normality test (p>0.05 indicates normal distribution).

All particle

size datasets and trace metals of interest with the exception of M n, As and M o were
normally distributed. The three bacteria variables and several of the Rock-Eval variables,
with the exception of TOC, RC, S3CO and OICO, were all found to be non-normal. Tbac, SRB
and Ratio w ere normalized by log-transformation and Hg and S2 were normalized by
reciprocal (1/x) transformation.

For visual ease, all the figures displaying bacterial

distributions use a log-scale axis.
The statistical test selected to measure the significance of a relation between
variables was dependent on the variables tested and their alignment with the requirements

83

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
to satisfy the assumptions for correlation or regression. The Spearman's Rank-Ordered
Correlation assumes th at the variables are continuously random intervals or ratios and they
do not need to be normally distributed. It gives a measure of the strength of a non-linear
monotonic relationship (as the value of one variable increases or decreases the other
variable also increases or decreases) betw een the variable, which was verified graphically if
required (Laerd Statistics, 2013). The Pearson's Correlation includes the assumption of a
normal distribution, though it has been recognized as a powerful and useful test for
continuous non-normal data (Chok, 2010). This correlation test was used in certain cases
w here a non-linear monotonic relation was not observed graphically or statistically but a
non-normal linear one was. If both variables were normally distributed then simple linear
regression was used.

84

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.3

Results and Discussion

5.3.1

Total Bacteria Quantification
The

downcore

distribution

of Tbac

is shown

in Figure

5.5

along with

the

corresponding SMB distributions, which will be discussed in the next section. The minimum
Tbac concentration was 1 .8 1 x l0 1016S rDNA/g sediment at the surface sample, 1 cm of; the

Copy #/g sediment
105

106

107

10 ®

109

10 1°

1011

1012

i___________i___________i___________i___________ i________ _ j ___________ i___________ i

oxic

r

2010

-

2002

1982

in -

- 1959

SUBOXIC

- 1940 $
-

1915

- 1903
Q.

0)
O
c
0)
E

L 1863
ANOXIC

(O

*

SMB
SMB Ponar
Total Bacteria

*

Total Bacteria Ponar
Suboxic Zone

O _
CO

Figure 5.5: Overview of the downcore distribution SMB (DsrAB) and Tbac (16S rDNA) by
gene copy#/g sediment with standard deviation error bars and the redox boundaries
proposed by M n and Fe reduction peaks (Chapter 3). Ponar grabs are representative of top
~7 cm.

85

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
maximum of 3 .2 1 x l0 1216S rDNA/g sediment occurred at 6 cm and the overall average was
5 .4 0 x l0 n ± 3 .2 2 x l0 9 16S rDNA/g (± SE) of sediment. The average of the three Ponar Grabs
(representative of a mixture of the top 7 cm from the surrounding region) is shown as
asterisks * in Figure 5.5 and was 2.28xlO n 16S rDNA/g of sediment, which is comparable to
the average num ber of bacteria in the sediment core. The number of bacteria increases at
the onset of the suboxic zone and maintains a high concentration until 6 cm w here it rapidly
decreases and remains somewhat constant to the core bottom.

5.3.1.1

Total Bacteria and Particle Size
No significant correlation was found between Tbac and any of the particle size

measurements indicating th at there is no obvious relationship between particle size and the
num ber of bacteria present in the bottom sediments. However, within the w ater column,
bacteria are dependent on the large surface area of suspended clay that adsorbs dissolved
organic m atter to provide a concentrated nutrient and energy source (Donohue & Molinos,
2009). The lack of correlation observed in the sediment core may be due to several factors.
For example, once the clay-OM-bacteria aggregates are deposited to the sediment, bacteria
may migrate to the optimal redox zone for their metabolism (von W achenfeldt & Tranvik,
2008; Gudasz et al,, 2012). As well, the capacity of clay particles to form an aggregate is not
necessarily dependent on particle size as it also involves the strength of ion adsorption,
physical structure of the clay, extent of surface area and its affinity for O M (Donohue &
Molinos, 2009).

86

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
5.3.2

Sulfur-Metabolizing Bacteria Quantification
The downcore distribution of SMB in the sediments is shown with Tbac in Figure 5.5.

The minimum, 2.94x10s dsrAB/g sediment occurs at 30 cm, the maximum, 7 .8 6 x l0 6 dsrAB/g
sediment occurs at 9 cm and the overall average was 2 .0 1 x l0 6 ± 5.22x10s dsrAB/g (±SE)
sediment. The three Ponar Grabs, shown as the three boxes located about the 0 cm line
represent the mixture of the top 7 cm of the region surrounding the core site and have an

Ponar Grabs
2010

Core Samples

-

2002

- 1982
- 1959
- 1940

E

o.

- 1915

10 -

£
D .
01
Q
§

- 1903
- 1863
15 -

E

1

to
20 -

25 -

30 -

2-3
Percent SMB of Total Bacteria [%)
Figure 5.6: Boxplot of %SMB of Total Bacterial qPCR replicates (16 ratios for each sample).
The black line in each box represents the sample median and the overall median is shows by
the vertical line as 1 .9 x 1 0 3. The Ponar Grabs are separated to the top but are presumed to
be a homologous sampling of the top ~7cm.

87

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
average of 6 .5 4 x l0 6 dsrAB/g sediment.
Visually, the SMB and Tbac downcore profiles (Fig 5.5) appear to follow a similar
trend, especially within the top 10 cm, however the Spearman Correlation of Log(Tbac) to
Log(SMB) is very weak (p=0.30, p<0.1). A better representation of the abundance of SMB is
shown as the percent SMB of total bacteria (%SMB) as a box plot in Figure 5.6. This is a
depiction of the percent SMB of Tbac calculated from the four qPCR replicates, to give a
total of 16 ratios used to calculate the median SMB% of each sediment sample. The vertical
line indicates the overall %SMB median,

1 .9 x l0 3 %SMB, while the

maximum was

0.013 %SMB and the minimum 1 .6 x 1 0 5 %SMB. The three Ponar grabs are shown at the top
w ith a median of 0.01%.
The downcore trend of %SMB indicates the overall abundance is very low.

Other

studies have reported %SMB anywhere from 1 0 5 to 3% with the higher concentrations
typically found in marine environments (Holmer, 2001; Barton et al, 2007; Leloup, 2009;
Pester 2012).

The low proportion is likely related to the Subarctic oligotrophic lake

conditions of Kusawa, such as cooler tem peratures, higher oxygen and low er sulfate levels.
The greatest proportion of SMB occurs at the surface and then decreases to 5 cm. A
second peak follows from 5 cm to 11 cm with the apex at 9 cm. A sharp peak with the
second highest abundance occurs at 12 cm, which is followed by several irregular
fluctuations to 19 cm.

A third peak is then observed from 19-24 cm and the remaining

samples are all below the overall median. Several publications (e.g. Mussmann et al., 2005;
Stahl et al., 2007; M uyzer & Stams, 2008) have reported that SMB typically exhibit tw o

88

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
population peaks within the oxic and anoxic zone, here it is proposed that these are located
at 5-9cm and 19-24cm respectively.

5.3.2.1

Sulfur Bacteria and Particle Size
No significant relation was found between SMB or %SMB and any of the particle size

measurements indicating that there is no direct relation betw een particle size and the
num ber of SMB present in the sediments. These relationships were evaluated here as SMB
are presumed to be a part of the community developed in the suspended clay-organicbacterial aggregate formations discussed in Section 5.3.1.1.

5.3.3

Sulfur-Metabolizing Bacteria Diversity
The dsrAB gene was sequenced to assess the diversity of the SMB present in Kusawa

Lake sediments.

PCR amplifications that contained the expected 1.7 kb fragm ent were

isolated and combined as follows: Ponar Grabs, 1-3 cm, 4-5 cm, 6-7 cm, 8-9 cm, 10-13 cm,
14-17 cm, 18-21 cm, 22-25 cm and 26-30 cm.

Up to tw elve clones were sequenced from

each group, except for the following groups w here only a few clones w ere grown: 6-7 cm (7
colonies); 8-9 cm (6 colonies); 10-13 cm (1 colony); 14-17 cm (0 colonies) and 26-30 cm (2
colonies).

The unsuccessful cloning was possibly due to laboratory error or inadequate

com petent cells fo r transformation.

Sequences obtained from the 76 successful clones

w ere compared to the GenBank database and 39 dsrAB sequences w ere confirmed. The

89

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
unsuccessful sequences w ere either not the dsrAB gene, the sequencing reaction was
contam inated or it was unsuccessful.

5 .3.3.1

M axim um Likelihood Tree
The nearest BLAST sequence for each clone was added to a dsrAB library along with

the Kusawa clones. Using MEGA 5.2.2 the sequences w ere aligned and a 177 amino acid
maximum likelihood tree was constructed using the W helan and Goldman model based on
500 bootstraps (Fig 5.7).

Eight clusters based on 87% amino acid identity and 16

Operational Taxonomic Units (OTUs) based on 97% amino acid identity w ere identified. The
probable bacterial order for each cluster was determ ined

by comparison with the

associated clones obtained from the BLAST database. The environmental condition of each
BLAST clone isolate was also considered to make inferences about the sediment conditions
at Kusawa.
As w ith many environmental studies of dsrAB (Holm er & Storkholm, 2001; Loy et al.,
2004; Leloup et al., 2009; Pester et al., 2012), uncultured SMB w ere detected in clusters I
and II and comprised of five OTUs. The associated BLAST sequences w ere all uncultured
SMB from freshw ater wetlands and paddy soils from a range of sulfate and oxygen
concentrations. Cluster I includes all the Ponar Grabs except one and isolates from 6-7 cm,
14-17 cm, 22-25 cm or 26-30 cm. As no BLAST search results w ere linked to OTU1 and OTU2,
it is proposed th at these comprise a novel lineage of subarctic, psychrophilic (cold tolerant)
SMB th at function under limited sulfate availability (Madigan et al., 2003).

90

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
0T U 3 and 0T U 4 are more closely related to previously identified uncultured clones
and likely have a similar function associated with low sulfate and oxic-anoxic environments.
Cluster II only contains one isolate from 4-5cm and its closest relation is from an oxic, low
sulfate, freshw ater wetland in Australia (Rees et al., 2010).
Cluster III contains the first branch of Gram-positive Fimicutes, Desulfotomaculum
spp., associated w ith the 14-17 cm zone.

The BLAST clones are from high sulfate

environments, suggesting that either SRB or SB are likely present (Zhou et al., 2011).
Cluster IV groups with BLAST isolates from high sulfate, anoxic environments that
are most closely related to Syntrophobacteraceae from 6-Proteobacteria.

These bacteria

are most notable for their ability to oxidize propionate and for syntrophic growth with
acetogens when using acetate under low sulfate conditions (Loy et al., 2004; Stahl et al.,
2007).

All of the clones th at form OTU7 are located between 8-9 cm, the proposed

oxic/anoxic interface.

These bacteria are likely taking advantage of the abundance of

electron acceptors present and may be growing in syntrophy and using the byproducts from
other species.

They are also likely to be somewhat oxygen tolerant and may be the

organisms responsible for the transfer of sulfur-compounds across the interface to continue
the cross-boundary sulfur cycle.
OTU8 constitutes an unknown singleton, as it has no association with any of the
other isolated clones or the sequences from the GenBank database.
Cluster V is the second branch of Firmicutes, Desulfotomaculum. The separation of
Firmicutes branches is due to the lateral Dsr gene transfer described by Klein (2004). Four
91

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
Figure

5.7:

M axim um

likelihood

tree

of

truncated dsrA genes (177 amino acids) using
the W helan and Goldman model.
0TU1

Numbers

correspond to 500 bootstrap replicates and
percentage of replicate trees
associated

OTU2

taxa

were

in which the

clustered

as

shown.

Clusters are based on 87% amino acid identity
and

associated

dotted

lines.

Classes are

divided

by the

Operational

taxonomic

units

(OTU) are based on 97% amino acid identity

AY167474:done darSW-64

and require at least tw o environmental clones

AM179478:done_21_40

Unidentified and Uncultured SRB Clusters
Cluster I: oxic-anoxic, low -h ig h S 0 4 -

OTU4

OTU5

Cluster II: oxic, lo w S 0 4 I OTU6

Firmicutes, Desulfotomaculum (G ram Positive)
Cluster III: u n k n o w n oxygen, high S04-

OTU7

Deltaproteobacteria, Syn trophobacteraceae
Cluster IV: anoxic, high S 0 4 -

Unknown singleton^
Firmicutes, Desulfotomaculum, (G ram Positive)
FJ748840:donc_MttDar75

Cluster V: oxic-anoxic, low -high S 0 4 OTU9
I OTUlu

Deltaproteobacteria

AB263705:done:O16

Sytrophaceae
I OTU12
_e| OTU13

Cluster V I: oxic-anoxic, lo w 5 0 4 -

U n kn o w n Fam ily

Cluster V II: oxic, lo w S 0 4 OTU14

Sytrophaceae
Cluster V III: oxic-anoxic, lo w S 0 4 I OTU15
D Q655253:dom _W 10

H0C90095:dooe_Rlte2008_0e

| OTU16

92

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa lake Sediments
clones located betw een 6-25 cm are grouped into 0T U 9 and are most closely related to a
clone from an acidic fen.

The other environmental clones in the cluster are from both

oxic/anoxic and low/high sulfate conditions.
The final three clusters are all found in the 6-Proteobacteria with the associated
environm ental clones from low sulfate zones and mostly oxic with a few growing in anoxic
conditions. All of the isolates are from above 7 cm except one clone from 10-13 cm.
Both Clusters VI and VIII are associated with environm ental clones related to the
Syntrophacae

family,

Syntrophobacteraceae.

which

is

from

the

same

order

Syntrophobacteraeles

as

These bacteria are known to operate in syntrophy with other

bacteria. Between the tw o clusters there are four OTUs that are all associated w ith bacteria
from a low sulfate and a mix of oxic and anoxic environments.
These tw o Clusters are separated by Cluster VII, which is not identified to any family.
It is possible that these are also Syntrophacae, however they may be another bacterium and
separated due to lateral gene transfers or some other mechanism (Klein et al., 2001). This is
the only cluster that is associated with strictly oxic bacteria along with low sulfate.

5.3.4

Outcomes of Phylogeny
An interesting outcome of the phylogeny is the dom inant presence of syntrophic

bacteria. This suggests th at under oligotrophic, presumably low sulfur conditions, SMB are
dependent on other organisms to provide substrates and electron donors other than their

93

Total Bacteria and Sulfur Metabolizing Bacteria in Kusawa Lake Sediments
preferred sulfur-compounds to enable growth.

Another observation is the absence of 6-

Proteobacteria species that are typically associated with high sulfate anoxic environments
such as orders Desulfobacterales, Desulfovibrionales, and Desulfurellales. It is possible they
may be present in the deeper sediments or they w ere not successfully amplified, however it
does signify that they are likely not abundant in the Subarctic, low sulfate and low to high
oxygen environm ent of Kusawa Lake.

94

Chapter 6 Mercury in Kusawa Lake

6.1

Introduction

6.1.1

Origins of Mercury in the Yukon
M ercury (Hg) is a ubiquitous heavy metal that has been widely studied for its toxicity,

industrial applications and its volatile biogeochemical cycle around the globe.

M ercury is

most recognized as a silver liquid in therm om eters and has many other liquid, solid or gas
forms (Table 6.1).

Through these states, Hg is capable of transport within geological,

atmospheric, aquatic and biological systems (Fitzgerald & Lamborg, 2005).

Table 6.1: Speciation and the common measurements of Hg in environmental samples.
Adapted from (Chetelat et al., 2012)._____________________________________________________

Species

Atmosphere

Aquatic

Organic/Tissue______
W

Hg(ll)

Gas and

Inorganic reactive particulate

Particle-

and dissolved in w ater column

Associated

and sediments

f:

Inorganic

As a solid, Hg is commonly found mineralized with sulfur to form cinnabar, HgS,
which is found throughout the Yukon, especially in conjunction with antim ony deposits
(Panteleyev, 2005). Solid Hg is naturally released to the environm ent by rock weathering or

95

Mercury in Kusawa Lake
discharge of stores from the cryosphere, driven by permafrost and glacial melt (Poissant et
al., 2008).

Cinnabar has been mined for over 3000 years, starting in the Peruvian Andes

(Cooke et al., 2009) and has been used in many industrial products and applications
throughout history such as thermostats, batteries, fluorescent lights, dentistry, chemical
catalysis and pharmaceuticals (Fitzgerald & Lamborg, 2005).
M ercury

is anthropogenically

em itted

to

the

atmosphere

through

industrial

combustion processes such as coal and fossil fuel burning, waste incineration, cement

Emissions to air, t
1400

Emissions,tonnes
1400 r
Dental amaigam (cremation}

1200

incJfttnKtoa waste end other
Chkx-alfceli industry
Cement production
Artisanal and sme*-*cale gdd production

1000

UrgMcale gold production
M rnl production (ferrous and non-ferrous!
Fossiftie i combustion for power end heating

800
600
Asia

400

North
America

Europe

Russia

South
America

Africa

Oceania

200
0

Africa

Asia

Europe

Russia

North Oceania South
America
America

Figure 6.1: Global anthropogenic emissions of mercury to air by continent from 1 9 9 0 ,1 9 9 5 ,
2000 and 2005 that depicts a decrease in developed countries and increase in developing
countries, particularly Asia.

The inset shows a breakdown of the industrial activity

associated with Hg emissions by each continent, w here again Asia is the primary producer.
Both figures from United Nations Environmental Programme (2013).

96

Mercury in Kusawa Lake
production and smelting (Fitzgerald & Lamborg, 2005). This has led to a global increase in
Hg emissions th at parallels increasing C 0 2 emissions beginning at the onset of the industrial
revolution (Lamborg et al., 2002). Global Hg emissions produced by the developed world
peaked and began to decrease in the 1990s, however, developing countries, particularly
those in Asia, have rapidly increased Hg emissions from an array of industrial activities (Fig

6 . 1).
Traditionally, the primary source of anthropogenic Hg was from coal burning, more
recently this has been greatly augmented by artisanal and small scale gold mining. The inset
o f Figure 6.1 outlines the industrial-associated global anthropogenic Hg contributions by
continent in 2005. Pacyna et al. (2010) predicts that by 2020 Hg emissions should decrease
in all continents with the exception of Asia, where the most conservative model estimates
Asian emissions at 450 tonnes/year and the most pessimistic predicts almost no change
from 2005 (Pacyna et al., 2010).
M ercury is also naturally em itted to the atmosphere by volcanic eruptions, wildfire
and photo- or biological reduction of Hg(il) in w ater and soils.
compounded

by additions from

human

activity to the

This natural release is

point w here

anthropogenic

contributions have raised the Hg level several-fold above the pre-industrial conditions
(United Nations Environment Programme, 2013).

97

Mercury in Kusawa Lake
6.1.2

Long-Range Atmospheric Transport
Once em itted to the atmosphere, Hg is converted to Gaseous Elemental Mercury

(GEM) or Hg(0), which has a very long residence tim e from 6 months to 2 years in the
troposphere (Lamborg et al., 2002).

Regional and seasonal-associated shifts in the

atmospheric redox potential results in the oxidation of GEM and production of reactive
gaseous Hg and particle-associated Hg, both of which have an atmospheric residency of just
days. These are deposited to the Earth's surface through precipitation scavenging (i.e. with
snow and rain) (Chetelat et al., 2012). In the high Arctic, atmospheric Hg-scavenging is most
notable in the spring through a phenomenon called Atmospheric M ercury Depletion Events
(ADME) w here a substantial am ount of atmospheric Hg is suddenly deposited with snowfall
due to seasonal changes in ocean chemistry (Schroeder et al., 1998; Steffen et al., 2007;
United Nations Environment Programme, 2013). Dunford et al. (2010) catalogued the origin
of GEM in Arctic, Subarctic and m id-latitude regions, including the Little Fox Lake Air Quality
M onitoring Station, located ~125 km northeast of Kusawa Lake.
another

Subarctic

station,

ADMEs

were

not

observed,

At Little Fox Lake and

instead,

atmospheric

GEM

concentrations w ere somewhat constant throughout the year with the maximum occurring
in early summer and minimum from late summer into autumn. Inputs from Asian countries
w ere more prom inent in regions closer to the Pacific at both the Subarctic and m id-latitude
stations and inputs from Russia and North America w ere also detected at Little Fox Lake but
at substantially lower levels than the contributions from Asia (Durnford et al., 2010).

98

Mercury in Kusawa Lake
6.1.3

Terrestrial and Aquatic Transport and Fates
Local anthropogenic, natural and atmospherically deposited Hg can be cycled

betw een terrestrial, freshwater, atmospheric and if in proximity, marine environments.
Some

Hg transportation

mechanisms

include

rapid

reduction

and

evasion

to

the

atmosphere, transportation in fluvial systems and deposition throughout watersheds
including the ocean and bioaccumulation through the food w eb (Fig 6.2) (Schroeder et al.,
2005; Chetelat et al., 2012). M ercury enters aquatic fluvial systems via precipitation, runoff,
erosion and release from the cryosphere.

It is capable of flux back to the atmosphere,

depending on Hg concentration, aquatic pH, wind and available substrates such as

Hg cycle

Figure 6.2: General overview of the global Hg cycle from the atmosphere to terrestrial and
aquatic systems and the transfer to and bioaccumulation through the food web as
methylmercury. From GMOS (2012)

99

Mercury in Kusawa Lake
dissolved organic carbon and particulate m atter (Schroeder et al., 2005).

In Subarctic

systems, there are many long-term stores of Hg in soils, sediments and the cryosphere
(AMAP, 2011).

In lakes, a major storage site is the bottom sediment, where Hg has been

sequestered and deposited from the w ater column to the lower sediments w here it remains
(Evans et al., 2005).

6.1.3.1

Algal-Derived Organic M a tte r Hg Scavenging
There is a well-recognized interaction between Hg and O M in both terrestrial and

aquatic systems (Fitzgerald & Lamborg, 2 0 0 3 ) .
favourable bond between

This is due to the thermodynamically

Hg (II) ions and the thio (-SH) group of various organic

compounds (Hesterberg et al., 2 0 0 1 ) .

One well-studied affiliation is the ability of labile

algal-derived O M to scavenge Hg from the w ater column and transfer it to the bottom
sediments during settling of suspended particulate-associated O M .

This is measured by

assessing the correlation between the S 2 4oo fraction detected by Rock Eval Pyrolysis and
Total Hg (THg) content of lake sediment records (Stern et al., 2 0 0 9 ; Sanei et al., 2 0 1 4 ) . The
efficiency of this interaction is based on the concentration balance between Hg and labile
O M . For example, if Hg is in excess of available O M it will remain in the w ater column and
be flushed from the aquatic system or taken up by the food web, or if there is an abundance
of O M then available Hg can be readily transported to the sediments (Sanei et al. 2 0 1 4 ) .
Kusawa has been identified as a high Hg-binding capacity lake, meaning that although the
concentrations of both O M and THg are exceeding low, the sediment O M is saturated with

100

Mercury in Kusawa Lake
Hg. This also means that any small increase in THg input may quickly overwhelm the OM
scavenging capacity and lead to

increasing amounts of Hg available to enter and

bioccumulate through the food chain (Sanei et al., 2014).
Understanding algal-Hg scavenging is im portant in northern systems to ensure an
accurate estim ate of the contribution to historic and recent atmospheric inputs.

Several

studies have attributed the rising concentration of Hg in lake sediment records to industryrelated anthropogenic inputs, however this does not account for climate change related
inputs such as erosion from bedrock and release from the cryosphere.

It also does not

consider the increasing rates of lake primary productivity that corresponds to increased
rates of algal-derived O M Hg scavenging. As a result, current estimates of sinks in the global
Hg budget may be erroneous regarding the level of atmospheric Hg deposition and fates in
northern ecosystems (Outridge et al., 2007; Stern et al., 2009; Sanei et al., 2014).

6.1.4

Mercury Methylation and Sulfate-Reducing Bacteria
One of the main reasons for the extensive study of Hg is its ability to bioaccumulate

through the food chain as toxic organic methylmercury (MeHg).

M icrobial-m ediated

methylation of Hg(ll) takes place at oxic/anoxic interfaces found within the environment,
such as w ithin the w ater column of eutrophic systems or within soils and lake sediments (Fig
6.3). The most widely recognized and studied species of Hg-methylating bacteria are SRB,
lron(lll)-Reducing Bacteria and methanogens. Mercury m ethylation is an anaerobic process

101

Mercury in Kusawa Lake
that is inherently linked to both the sulfur and iron cycles and most readily occurs at the
zones of sulfate and ferric iron reduction (Benoit et al., 1999; Mason & Lawrence, 1999;
Hammerschmidt et al., 2006).
The ability of SRB to m ethylate Hg is dependent on community composition, as only
certain

strains

are

confirmed

Hg-methylators,

all

of

which

are

found

under

Deltaproteobacteria (Gilmour et al., 2011). The most common strain used to study SRB Hgmethylation is Desulfovibrio desulfricans, from the order Desulfovibrionales, which are
obligate anaerobic SRB. D. desulfuricans both produces and degrades MeHg, where in most
cases the rate of dem ethylation exceeds that of methylation.

Therefore, as observed in

many environments, only a small percentage of THg is present as M eHg (Hollweg et al.,
2009; Gilmour et al., 2011).
The exact mechanism of SRB Hg-methylation is still poorly understood. It has been
proposed th at dissolved HgS(aq) readily diffuses across SRB cell membranes, is methylated
and MeHg is then excreted. Some SRB may employ the acetyl-CoA pathway, which is used
to com pletely oxidize carbon to C 0 2, while incomplete oxidizers that are capable of growth
outside the acetyl-CoA pathway may m ethylate mercury through some other process
(Ekstrom et al., 2003).

Further study of 0. desulfuricans has identified tw o proteins,

punative corrinoid protein (HgcA) and a 2[4Fe-4S] ferredoxin protein (HcgB) that can carry a
methyl group from the reductive acetyl-CoA pathway and transfer it to Hg to produce MeHg
(Gilmour et al., 2011).

102

Mercury in Kusawa Lake

Evasion

Nat
DamathytaSon

Hfl°

Figure 6.3: Overview of Hg cycling in the sediments of estuarine and coastal environments
as described by M erritt et al. (2008). W here Hg enters oxic soils from the w ater column and
is transported to anoxic sediments by diffusion via sediment association where it is
m ethylated by SRB.

MeHg is then re-transported to the sediment w ater interface (SWI)

w here it may enter the food chain or be converted to another species. Circles depict
sediment particles, Hgi=inorganic divalent Hg or Hg(ll), RD=reduction, OD=oxidation, MiR
=bacterially independent reduction. (M e rritt & Amirbahman, 2009)

6.1.5

Biochemical Controls of Mercury Methylation
Methylm ercury

production

is dependent

on

several

factors.

First,

is the

concentration of THg where in many studies MeHg is positively correlated to THg
(Hammerschm idt et al., 2006; Goulet et al., 2007; Hollweg et al., 2009; Jiang et al., 2011).
However, in systems w ith very low microbial activity a threshold can be reached and a log
relation betw een increasing THg and stable or decreasing levels of M eHg is observed th at is
likely related to saturation of methylating enzymes (Gilmour et al., 2011).

103

Mercury in Kusawa Lake
Organic m atter also influences the bioavailability of Hg, w here S 2 40o has a strong
correlation with THg in Subarctic and Arctic lake sediment records and as discussed can
scavenge Hg from the w ater column and transport it to the bottom sediments (Stern et al.,
2 0 0 9 ; Sanei et al., 2 0 1 4 ) . .
Sulfate (S 042 ) also strongly regulates the rate of methylation, where increasing
levels of S 0 42' correspond to an increase in M eHg productivity in low sulfate environments,
such as Kusawa Lake. Under high sulfate conditions, such as marine systems, SRB enzymes
can be saturated with preferred electron acceptors, thereby inhibiting Hg-methylation
(Gilmour et al., 1 9 9 2 ; Fitzgerald & Lamborg, 2 0 0 3 ; Norici et al., 2 0 0 5 ; Sievert et al., 2 0 0 7 ;
Ouddane et al., 2 0 0 8 ) . Sulfide (H2S) also controls the rate of methylation as it promotes the
form ation of dissolved HgS(aq) to enable diffusion across cell walls.

However, under high

sulfide conditions other sulfur-Hg species are formed, such as Hg(SH)2, HgS2 and polysulfides
that cannot be m ethylated and consequently inhibit methylation. Sulfur-oxidizing bacteria
also play a role in this balance by continuously oxidizing H2S to sulfate to regulate H2S or
S 0 42' inhibited m ethylation. This is a reason why methylation typically occurs around the
oxic/anoxic zone as neither sulfate or sulfides are in excess (Gilmour et al., 1 9 9 2 ; Benoit et
al., 1 9 9 9 ; Ouddane et al., 2 0 0 8 ) .
In sediments, MeHg is produced almost completely within the pore w ater, where
diffused HgS(aq) is transported across cell walls, m ethylated by bacteria, excreted and finally
diffused to the w ater column. Studies w here MeHg was not diffused found it was promptly
dem ethylated in the presence of Fe-oxides within the suboxic zone. Once MeHg is in the
104

Mercury in Kusawa Lake
w ater column it can be readily taken up by the food chain or may be lost by photodem ethylation and photo-reduction and em itted to the atmosphere. (Hammerschm idt et al.,
2006; Goulet et al., 2007).

6.1.6

Bioaccumulation, Biomagnification and Toxicity
Bioaccumulation is defined as the ability of a toxic substance to reside within an

organism for longer than it takes to degrade. Biomagnification in when the concentration of
the toxic substance is compounded as it moves up the food chain. For example, the MeHg
concentrations of fish continuously increase as it ingests M eHg contam inated invertebrates
(United Nations Environment Programme, 2013).
Mercury is designated as a global threat to human and environmental health
because of the varying degrees of toxicity experienced by those exposed to

high

concentrations (United Nations Environment Programme, 2013). In elem ental form , Hg can
cause respiratory tract failure when inhaled as a highly concentrated vapour, inorganic
Hg(ll) can lead to kidney failure and gastrointestinal damage and finally MeHg, the most
toxic species, can be absorbed through the gastrointestinal tract and distributed throughout
the body.

These risks are of particular concern to pregnant and breastfeeding wom en

(Health Canada, 2009). Mercury is also known to reside for long periods in the brain and
historically the psychological symptoms associated with extrem e mercury poisoning from
felt hat manufacturing coined the term 'M ad as a Hatter'.

105

Mercury in Kusawa Lake
The W orld Health Organization guidelines set the tolerable daily intake of Hg in
adults to 0.71 M-g/kg body weight. Health Canada has also set the commercial standard that
fish and seafood Hg levels must be below 0.50 pg/g of tissue (Health Canada, 2009). In the
north, exposure to mercury is of large concern due to the sustained practice of harvesting
food such as plants and berries, small and large game and fish.

The Inuit Health Survey

found th at Inuit who regularly consume country foods have a higher blood mercury
concentration than other Canadians though they w ere generally below the Health Canada
blood toxicity thresholds, with a few exceptions.

The report also emphasized that the

nutrients and wellbeing obtained by consuming country foods outweighs the contaminant
risks (Chan, 2011).
In the Yukon, Hg concentrations in fish are at very low levels and therefore there is
no limit to the am ount of fish that may be consumed.

However, pregnant and nursing

w om en are advised to limit their intake, especially of predatory fish such as trout, burbot
and northern pike (Environment Yukon, 2010). The annual Hg concentrations of trout in
Kusawa Lake are shown in Figure 6.4 where the average level over a 17-year period was
0.38 pg/g.

The record trout Hg level was 0.54 pg/g in 1993 and the average level that year

was 0.45 pg/g.

This high decreased to 0.23 pg/g in 2009, with the most recent

measurem ent in 2010 at 0.31 pg/g. These levels are nearing the Health Canada standard
limit of 0.50 pg/g, indicating that Hg is present, bioaccumulating and biomagnifying in the
Subarctic environm ent (Stern et al., 2011). The decrease in average tro u t Hg levels from

106

Mercury in Kusawa Lake
1993 is presumably associated with the decrease in atmospherically generated Hg from
developed countries (i.e. North America and Europe), however the drop betw een 19992001 and 2007-2008 corresponds to the second and third highest recorded discharge years
in 2000 and 2007, respectively for the Takhini river (W ater Survey of Canada, 2012),
indicating that the drop in trout Hg concentrations may be attributed to flushing of the
drainage basin, photo-reduction and dilution of bioavailable Hg in the watershed.

0.60

Kusawa
O) 0.50

O) 0.40

0.00
1993 1999 2001

2002 2003 2004

2005 2006 2007 2008 2009 2010

Year
Figure 6.4: Level of Hg detected in trout tissue from Kusawa Lake from 1993-2010.

From

Stern et al. (2011).

107

Mercury in Kusawa Lake
6.2

Methods

6.2.1

Total Mercury and Methylmercury Quantification
Core A subsamples w ere used to quantify Total mercury and m ethylmercury at the

University of M anitoba by Cold Vapour Atomic Absorption Spectrometry under the QA/QC
set by the Northern Contaminants Program (Stern et al., 2009). The normalized downcore
Hg measurements (1/H g) w ere correlated with other variables including particle size, Rock
Eval Pyrolysis, Tbac and SMB.

6.3

Results and Discussion

6.3.1

Total Mercury
The downcore profile of THg is depicted in Figure 6.5. The maximum THg sediment

concentration of 0.034 p g g 1 occurs at 4 cm, the minimum, 0.016 pgg'1 at 19 cm and 22 cm
and the overall average was 0.022 ± 0.0009 pgg'1 (±SE). The THg concentrations below 15
cm remain som ewhat constant and fluctuate between 0.016 pgg'1 to 0.021 p g g 1, while the
top 5 cm has the highest concentration that continuously decreases down the core. A 1.5fold increase in the THg concentration is observed from 1915-2010 that corresponds with
increasing global tem perature and industrial activity.

108

Mercury in Kusawa Lake
Total Hg (/jgg-1 sed)
0.015

0.020

0.025

0.030

0.035

o

r 2010

oxic

-

2002

- 1982
- 1959

SUBOXIC

- 1940 $
- 1915
- 1903
L 1863

ANOXIC

■o
CM

Figure 6.5: Downcore profile of Hg concentration in Kusawa Lake sediments. There is a 1.5fold increase in THg concentration from 1915-2010 over the top 10 cm.

6.3.2

Methylmercury
The level of methylmercury (M eHg) throughout the core was below the lim it of

detection. This is not surprising considering the low THg concentration, low levels of O M ,
SRB and presumably sulfate concentrations.

As well, the majority of M eHg is likely

dissolved and associated w ith the pore w ater instead of the sediments.

The inability to

detect M eHg does not indicate that MeHg production is not occurring, only that the level is
so low th at it cannot be detected by the method used.

109

Mercury in Kusawa Lake
6.3.3

Total Mercury and Particle Size
The strongest association of Hg with the various particles size measurements was

with fine silt (8-16 pm) that makes up 41-49% of the sediment.
measurements resulted in correlations that had slopes close to zero.

The other size

O ther studies have

shown that Hg does not strongly associate w ith clay or very small particles.

Jiang et al.,

(2011) proposed that aluminosilicates and other clay associated metals such as tin and
potassium could be diluting the Hg concentrations in the sediments, rather than directly
binding Hg. A biplot relation of % fine silt w ith the 1/Hg profile is depicted in Figure 6.6

3*
2*

11*

7A

(cm)
22 ■

10* X *

•

0-4cm. OXIC

A

5-9cm, SUBOXIC

181
16*

29*

30

40

50

60

Figure 6.6: Downcore profile of 1/Hg association with % Fine Silt.

There are tw o

associations of sediments associated with the top 7cm, 12-18cm and 28cm (solid line oval)
and the lower sediments (dotted line oval). Outliers are 1 cm, 11 cm and 24 cm.

110

Mercury in Kusawa Lake
w here tw o notable trends are observed. The first is grouped by the solid line oval w here a
negative relation includes sediment from 2-7 cm, 12-18 cm and 28 cm indicating increasing
THg concentrations w ith increasing amounts of fine sediment.

The second association is

shown by the dotted line oval that contains sediments from 8-11 cm and <19 cm. There is
also a major overlap between the tw o ovals that encloses samples from 12-18 cm and 28
cm.

The outliers from these tw o groupings includes the surface at 1 cm, influenced by

reactions at the sedim ent-w ater interface, 11 cm, associated w ith the 1911 sedimentation
event and 24 cm, which cannot be accounted for with the available data. As others have
indicated th at clay minerals do not directly bind Hg, the process that may be reflected in the
pattern seen here could be O M -m ediated aggregation of fine inorganic sediment.

6.3.4

Total Mercury and Organic Matter
The association between organic m atter and Hg is well studied. There is a strong

regression (rz=0.51, p<0.001) between the normalized reciprocals of THg and the Hydrogen
Index (HI) (mg HC g'1 T O C 1), where HI is a measure of S2/TOC determ ined by Rock Eval
Pyrolysis (Fig 4.11). Hg is known to have a strong association with the labile HI40o fraction
and a w eak relation with Hlb (Sanei et al., 2014). In this study, sediment from the surface to
8 cm and 14 cm w ere all detected below 400"C, making the rem ainder a part of Hlb (Fig 4.2).
A full profile biplot of the reciprocal of both HI and Hg (Fig 6.7) shows a clear
separation of the HI40o and Hlbfractions. The affinity of Hg with the more labile HI40o

111

Mercury in Kusawa Lake

0.016-1
29 V

20V
28 V
28 V
15V

13V.
„ - - - TZ*

Sedim ent Depth (cm)

21V

10V

16V

0.014 H

O
X

19V

Hl„ r2 - -0.02

•

0-4cm, OXIC

▲ 5-9cm, SUBOXIC
V

10-29cm, ANOXIC

11V

0 .012 -

m

Figure 6.7: Biplot of Hg'1 and HI'1 with the regressions denoted for the strong relation
betw een Hg and the HUoo (solid line) labile O M measurements and the degraded Hlb
fractions (dotted line).

fraction is evident with its strong regression (r2=0.93, pcO.0001, slope=2.9xl0 '1), denoted by
the solid line. The relation with the remaining sediments below 9 cm and excluding 14 cm
are shown by the dotted line w here no significant relation was found (r2= -0.02, p>0.4).
These findings are expected and in line with Sanei et al.'s (2014) study that classifies
Kusawa as a high-capacity binding lake due to its constantly strong correlation between
HI4oo and THg.

Lakes w ith Hg-HI slopes >1 are considered low Hg binding capacity lakes

w here the input of Hg exceeds the available binding capacity of labile O M . This indicates

112

Mercury in Kusawa Lake
th at although there are very low levels of OM and Hg at Kusawa, there is still sufficient OM
available to scavenge the low Hg influxes from the w ater column and deliver it to the
bottom sediments.

This finding also suggests that any small increase in Hg may rapidly

overwhelm the system resulting in increased amounts of available Hg in the w ater column
(Sanei et al., 2014).

6.3.5

Total Mercury and Total Bacteria
To investigate patterns which could provide further insight to the various redox-

driven reactions occurring down the core, Figure 6.8 is presented to depict the interaction
betw een 1/Hg and Log(Tbac), where a three possible associations are observed. First, the
surface sediments to 5 cm have a weak and steep positive trend. The second interaction,
depicted by the solid line oval includes sediments associated with suboxic and anoxic zones
from 6-15 cm, 26 cm and 28-29 cm. Here the concentration of bacteria decreases, as does
the concentration of Hg. It is assumed that the decreases are related to downcore microbial
redox interactions and loss of M eHg to the pore w ater although this was not directly
measured. The third interaction occurs in the sediments below 11 cm and excluding 20-22
cm w here a tight association between increasing Tbac and decreasing Hg is observed by the
dotted line oval.
One interesting observation is that the relation between Hg and Tbac depicts a
similar pattern observed in Figure 6.6, the biplot between Hg and % fine silt. It is proposed

113

Mercury in Kusawa Lake

5*.

S «dlm *nt Depth (cm )
> 1011

•

<
z

e
8

0~4cm, OXIC

A

5-9cm, SUBOXIC

■

10-29cm, ANOXIC

10 1 1*

I

21 ■

10'° H

1*
30

40

50

60

Hg"1(ngg"1)

Figure 6.8: Biplot of 1/Hg association with Log(Total bacteria). The sediments from 1-5 cm
show a steep positive relation at the left of the graph.

The solid line oval includes

sediments w here a negative relation between Hg and Tbac for samples from 6-18 cm and
26-29 cm. The dotted oval contains samples from below 12 cm. Samples from 20-22 cm are
considered outliers from these trends.

th at these interactions are influenced by three different redox reaction regimes: (1) the
steep positive relation in the near surface oxic sediments corresponds to recently deposited
sedim ent-OM -bacterial-Hg aggregates form ed

in the w ater column, (2) a downcore

dependent negative trend through the suboxic zone and oxic/anoxic transition boundary
associated w ith microbial redox reactions and (3) the lower anoxic sediments w here Hg has
a w eak relation with both bacteria and sediment that is independent of core depth. The
114

Mercury in Kusawa Lake
samples from 20-22 cm do not fit into any of these redox regimes and cannot be accounted
for with the available data. These trends are suggestions based on correlation and cannot
be verified with the available data.

6.3.6

Total Mercury and Sulfur-Metabolizing Bacteria
The final explanatory variable to describe the presence and cycling of Hg in the

bottom sediment of Kusawa Lake is its interaction with SMB. The log(Ratio) was compared
to 1/Hg in tw o separate biplots in Figure 6.9. The top panel (Fig 6.9a) includes sediments
from the oxic (denoted by the solid oval) and suboxic (denoted by the dotted oval) zones.
The lower sediments observed in Figure 6.9b are scattered and exhibit no evident relation.
There is a distinct redox-dependent separation of the top 10 cm that complements earlier
suggestions that the near surface sediments consist of recently deposited sedim ent-OM -Hgbacterial aggregates followed by redox driven associations through the suboxic zone. Again
though, these inferences are based on visual pattern and correlation and further study is
required to verify these proposed trends. Recall from Figure 5.7 that sediments from 8-9
cm house Syntrophobacteraceae sp. that require high sulfate, anoxic conditions, which in
turn are favourable for Hg-methylation.

Therefore, the most probable depth for SRB-

m ediated Hg-methylation and demethylation is at the oxic/anoxic boundary from 8-9 cm.
Though, no species from known SRB Hg-methylating orders w ere detected.

Overall, the

results suggest th at conditions are not favourable for Hg-methylation by SRB as there are no
obvious Hg-SMB interactions and no known Hg-methylators w ere detected.

115

Mercury in Kusawa Lake

0-4cm, OXIC

r 25-9cm. SUBOXIC

3*

10|i

r 3-

30

40

50

60

b.
10"

128
231

228

If!
®

1

?§■

268

IB

£
o

1 6B

m 10-3
«

25 ■
191
27 ■

s
10"

118
248
291
— i—

— I—

— I—

— I—

30

40

50

60

Hg'1 (ngg-1)

Figure 6.9: Biplot of log(%SMB) vs. 1/Hg where, a: depicts the separation of the oxic and
anoxic zones in the top 10 cm, and b: shows the lack of structure within the anoxic
sediments from below 11 cm.

116

Mercury in Kusawa Lake
6.3.7

Summary of Sediment, Organic Matter and Bacterial Interactions with Mercury

There are several factors th at appear to control mercury in Kusawa Lake sediments:
•

The concentration of THg is very low in the sediment record.

•

THg is strongly correlated with S 2 40o characterizing Kusawa as a high-capacity binding
lake,
o

therefore, any increase in THg, w ithout a concomitant increase in algal O M could
overwhelm the OM-binding capacity and lead to increased Hg availability in the
w ater column for bioaccumulation.

•

Both Tbac and fine silt have a similar redox driven correlation to THg where three
proposed trends are noted:
1.

a positive association with recently deposited near surface sediments, suggesting
th at sediment-bacteria-Hg interactions are established in the w ater column,
presumably as floe formations;

2.

a negative trend through the suboxic zone w here it is proposed that bacteriam ediated breakdown of sedim ent-OM floes result in the release of Hg;

3.

a weak relation in sediments below 12 cm, indicating loss of interaction due to
breakdown of sedim ent-OM floes.

•

There is a very low %SMB of total bacteria and no known Hg-methylating SRB w ere
detected indicating th at Hg-methylation by SRB in the sediments is likely minimal.
o

Subarctic, oligotrophic and orthograde lake conditions are unfavourable for SMB
and SRB-mediated Hg-methylation.

Mercury in Kusawa Lake
6.4

Mercury in Kusawa Lake and the Implications of Climate Change
There are several measurable changes and potential risks associated with climate

change in the Subarctic. In regards to Kusawa Lake, the major impacts will be as a result of
annual w arm er tem peratures and varying levels of snow and rainfall (Environment Yukon,
2014).

6.4.1

Increased Allochthonous Sediment and Organic M atter
Increases in tem perature and precipitation will inherently cause an increase in

sediment transport from a range of origins. Glacial melt is expected to increase and release
large concentrations of fine sediment as well as anthropogenic contaminants such as Hg,
nutrients and other heavy metals. Similarly, the increased rates of permafrost m elt will also
release stored heavy metals and greenhouse gases such as m ethane and carbon dioxide.
Furtherm ore, the loss of permafrost will cause land instability leading to erosion, land and
mudslides and increased instability of torrent systems (Walsh, 2005; AMAP, 2011). All of
this along with the expected higher rates of discharge and w ater levels will result in the
increased distribution and diversity of both aquatic and terrestrial O M throughout Kusawa
Lake.

118

Mercury in Kusawa Lake
6.4.2

Increased Primary Productivity
Longer ice-free seasons and w arm er w inter and summer tem peratures will enable

higher rates of aquatic primary productivity.

Since algal scavenging of Hg to the lake

sediments has been demonstrated as the principle control of Hg delivery (Sanei et al., 2014),
w arm er tem peratures will likely result in an increase in stored Hg and in M eHg available for
bioaccumulation in the w ater column.

If more O M w ere available in the lake system it

would decrease the control O M has on Hg by becoming a low Hg-binding capacity binding
lake (Sanei et al., 2014).

Another outcome is that increased primary productivity could

prom ote environments more closely related to

meso-eutrophic systems w here the

sediments may be fully anoxic with an oxygen clinograde (oxygen depleted in the lower
w ater column), allowing Hg to be m ethylated within the w ater column and the sediments.

6.4.3

SRB Community Composition
A study by Robador et al. (2009) assessed the in situ adaptation of psychrophilic SRB

from the Arctic Ocean and mesophilic tem perate SRB to long-term tem perature changes.
The result was that the Arctic SRB communities incubated for one year at 20'C maintained
the capacity to reduce sulfate at 0 ‘ C and also gained sulfate reduction ability at w arm er
tem peratures. Concurrently, tem perate SRB were subject to the same conditions and never
acquired the ability to efficiently reduce sulfate at 0 ”C.

These findings indicate th at

psychrophilic bacteria are more readily adaptable to changing climates than tem perate

119

Mercury in Kusawa Lake
bacteria.

This is likely due to the extrem e shifts in seasonal tem peratures in cooler

compared to more tem perate climates.

Robador et al. (2009) also found that there was

more diversity in the com m unity composition of Arctic SRB compared to tem perate and
during incubation certain populations from the Arctic samples declined while others
increased, however no populations w ere completely lost. They concluded that Arctic SRB
select for organisms th at are more readily adapted to the current tem perature.
From this study, it can be assumed that in Kusawa Lake SRB communities are
adapted to both w inter and summer tem peratures and in the event of ongoing warming the
most efficient species of SRB will be selected for. If the lake becomes more eutrophic the
level of available sulfate increases it could prom ote population increases of Hg-methylating
SRB and as a result, MeHg production will increase.

120

Chapter 7 Conclusion

This thesis has provided background and insight into the biogeochemical sediment
process of Hg cycling by SMB in Subarctic oligotrophic Kusawa Lake, Yukon.

It also

contributes to the limited literature regarding SMB at Subarctic latitudes and establishes a
baseline for use in future studies.

Each chapter of this work expands on the previous to

develop a sediment-based concept of O M and SMB-mediated Hg-cyding from its delivery to
the bottom sediments to either its storage as inorganic Hg or release to the w ater column
as bioaccumulating MeHg.
Chapter 2 introduced Kusawa Lake, its history, climate and hydrology.

Chapter 3

provided an overview of the lake's limnology and sedimentology of the core sites to
elucidate the historic and contemporary sediment processes.

One major sedimentation

event was observed at 11 cm sediment depth, which was dated to 1911. As well, estimates
of the redox zones w ere established using trace metal profiles, which provided guidance for
interpretation of O M and bacterial processes. Chapter 4 presented O M composition and
source as determ ined from Rock Eval Pyrolysis. S24oo OM consisting of recently deposited
algal cell wall material was detected from the surface to 8 cm and 14 cm depth.

All

remaining samples grouped as S2b and consisted of more refractory compounds. All of the
samples w ere from autochthonous sources with the exception of 7 cm (unknown) and 26
cm, the proposed W hite River Ash layer. One major O M accumulation event was detected
from 14-15 cm and lack of direct evidence restricted identification of its source.

121

Conclusion
Chapter 5 introduced the downcore distribution of Tbac and SMB along with the
SMB diversity.

There was an extrem ely low ratio of SMB to Tbac likely associated with

oligotrophic Subarctic lake conditions. Four distinct layers of SMB w ere identified: (i) 1 — 5
cm: SOB (Deltaproteobacteria, unknown; (ii) 1 - 7 cm: SOB and SRB (Deltaproteobacteria,
Syntrophaceae); (ii) 8 - 9 cm: SRB (Deltaproteobacteria, Syntrophobacteraceae) and; (iii)
below 7 cm: SRB (uncultured clones and Firmicutes, Desulfotomaculum).

No known Hg-

methylators w ere detected, however, species that operate in syntrophy were identified.
This suggests that because the SMB are metabolizing under low sulfate conditions,
syntrophic species are selected for to enable ongoing cycling of S 042 and H2S across the
oxic/anoxic boundary.
These lines of evidence w ere used to draw conclusions about Hg transport and fate
in the lake sediments, which are summarized in Figure 7.1.

Overall, Kusawa Lake is

oligotrophic, has a low rate of primary productivity and O M , which strongly influences the
controls of Hg-methylation. The cycle of Hg in Kusawa Lake begins in the w ater column,
where it binds with labile O M associated with fine sediment and bacterial aggregates. This
is a strongly influential control as Kusawa is considered a high Hg-binding capacity lake, due
to the low levels of both Hg and O M . These aggregates or floes settle to the lake bottom
sediments. There is a very low percent of SRB present and no known Hg-methylating SRB
w ere detected, therefore, opportunities for SRB Hg-methylation are assumed to be minimal.
M ethylation is occurring at some location within the lake ecosystem as elevated levels w ere

122

Sedim en t-associated

^ H g -O M -B a c te ria l
ag g re g a te s

SEDIMENT-WATER INTERFACE

\

MeHg
Bioaccumulation

2010

o
------------------------------------------------------------------------------------------------------------- f

c

"

SOB

Labile OM - Hg

2002

Sediment Depth (cm)

OXIC >300mV

so42'
19 8 2

SUBOXIC -100-300mV

Bacterial-mediated
OM degradation

19 5 9
... 4
^ i * ~ ~ _MeHgSRS
M eHg---------------------------J
♦
W
,'* » - - » M e H g - s r a

1911 Sedimentation Event

ANOXIC <-100mV

& SOB

Fermentation

so42

-If
H2S

Minimal Hg-SRB interaction assumed
due to unfavourable lake conditions for
SRB and Hg-methylation

15 H,S

Figure 7.1: Summary of findings and possible fates of
Hg delivered to bottom sediments of Kusawa Lake.
The estimates of redox zones, bacterial metabolism

30
NJ
U>

and state of O M degradation are also described.

Methanogens &
Acetogens

1915
19 0 3
18 6 3

SRB

Long-term storage
in sediments

19 4 0

Conclusion
detected in the trout. The most likely location for methylation in the bottom sediments
would be at the oxic/anoxic transition boundary, located at 8-9 cm depth.

There,

particulate Hg can be dissolved in association with sulfide and cross SRB (or some other
methylating bacteria) cell walls, be methylated, excreted and diffuse through the pore
w ater to the w ater column where it readily enters the food chain.

Overall, the major factors regulating the potential for Hg-methylation by SRB in Subarctic
Kusawa Lake are:
1.

the rate of primary productivity and w ater column concentration of sedimentassociated algal-derived O M that can scavenge Hg and transport it to the sediments,

2.

the concentration of mercury in the w ater column for scavenging by algal-derived
O M and in the sediments for availability to SRB at the oxic/anoxic boundary and,

3.

the diversity and concentration of SRB, where more eutrophic lake conditions are
likely needed to promote SMB populations and select for SRB Hg-methylators.

124

Conclusion
7.1

Limitations and Future W ork
As this study was completed in conjunction with a larger Hg research project at

Kusawa, many of the limitations are related to the core sampling procedures that could
have been modified to allow for analysis of additional variables and sampling sites.
The following is a list of limitations to this study and suggestions for future work:
•

The sample site location appears to have been influenced by less regular sediment
delivery than initially assumed, therefore future cores should be taken closer to the
Stern et al. (2009) 2005 core,

•

Only a single core was collected for the bacterial component of the study, another
core from higher in the basin within a more eutrophic ecosystem, such as a wetland,
could provide better detail regarding the mechanism of Hg-methylation,

•

No sulfate reduction rates or sulfate or sulfide m easurem ent w ere taken, which
would have helped provide further insight to SMB metabolism,

•

Sediment-associated m ethymercury levels w ere below the limit of detection and no
pore w ater measurements were taken, which would

have provide a better

understanding of sediment-pore w ater exchanges, and
•

Only SMB distribution and diversity was assessed.

Detection of other known Hg-

methlyating microbes such as iron-reducing bacteria would have been beneficial.

125

Conclusion
One suggestion for a future study would be to collect a core from a more eutrophic
system in the upper reaches of the Takhini River basin and another core from the current
sample site for comparison. Along with some of the measurements used in this study, in
situ comparisons of sulfate reduction rates, mercury methylation rates, Hg-OM binding and
interactions of stored inorganic Hg would help contribute to the understanding of the fate
of Hg in Subarctic ecosystems.
Overall, this work contributes to the understanding of the controls of Hg transport
and potential for SRB-mediated m ethylation in the sediments.

M ore specifically it

elucidates the microbial distribution and diversity th at play a role in Hg processing, which
are factors that have not been adequately studied at Subarctic latitudes. As well, identifying
factors presumed to control Hg processes in Kusawa Lake will help infer and identify climate
change implications and aid in assessing the impacts of Hg inputs from distant industrial
sources, particularly Asia. Finally, this work will contribute to ensuring the environmental
safety for Yukon people and future Yukon generations.

126

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139

Sediment Depth (cm)

Antimony Sb (ppm)

Barium Ba (ppm)

Arsenic As (ppm)

Bismuth Bi (ppm)

o

2009

10

1975
1945 CO

o

1911
1847

CM
to

CO

Sediment Depth (cm)

Cadmium Cd (ppm)
0.4

0.S

0.6

0.7

0.6

I

1

I

l

l

Caesium Cs (ppm)

Calcium Ca (%)

17

1.e

1.9

2.0

Chromium Cr (ppm)

Cerium Ce (ppm)

21

45

50

55

I_____I____ I

2009
1975

in

1945 (0

€

o

1911 ^

o

CM

o

CO

Cobalt Co (ppm)

Sediment Depth (cm)

o
to

20

22

L-

-- 1 -—

1

Copper Cu (ppm)

24

I

Dysporosium Dy (ppm)

Qadolimium Gd (ppm)
46

i________i________L_

40

45

5.0

5.5

Gallium Ga (ppm)
6.0

J

I

I

I

I

I
2009
1975
1 9 4 5(0

o

£

1 9 1 1 *"
1847

o

CM

to

CM

o

CO

O

18

Appendix: Trace Metal Profiles

Aluminum AJ (%)

Hafnium Hf (ppm)
05

Indium in (ppm)

Lanthanum La (ppm)

Iron Fe (%)

0.6

0.7

08

09

10

0.06

0.08

0.10

0 12

0.14

0 16

I____L

I

I

I

I

I

1.

J

1

1

1

7.0

7.5

8.0

8.5

9.0

24

26

28

30

32

Lead Pb (ppm)
34

26

27

28

29

30

31

Sediment Depth (cm)

r

2009
1975

in

~ 1945 CO

o

- 1911

in

L 1847

cm

CM
CO

Sediment Depth (cm)

O

Magnesium Mg (%)

54

56

58

60

62

64

20

21

1

1

I

1

1

1

I

I

l

22
l

2.3
l

2.4
l

26

l

2000

3000

4000

5000

6000

I______ I______ 1______ I______ I

I

Neodymium Nd (ppm)

Moiybedeum Mo (ppm)

M anganese Mn (ppm)

25

2
-J_______ I_______ I_______ I

L_

24

26

28

30

I

-L

1

I
r

2009
- 1975

in

- 1945 «

o

- 1911

in

L 1847

o
c
o
Nickel Ni (ppm)
18

Sediment Depth (cm)

o

i

20

1

22

I

24

Phosphorous P (%)

Niobium Nb (ppm)
26

I___\

24

0 085

O

in

in

o

o
in .

o

CM

8
in

CM

8

o
co _

0.095

0.105

Praseodymium Pr (ppm)

Potassium K (% )
0 115
i

i

i

-

i

i

6.5

7.0

75

8.0

8.5

l

l - l

I

»

r 2009
1975

L

1847

Appendix: Trace Metal Profiles

Lithium Li (ppm)
52

Rubidium Rb (ppm)

Samarium Sm (ppm)

150

160

170

180

190

I

I

I

I

I

54

56

58

Silver Ag (ppm)

Scandium Sc (ppm)

60

62

15

16

17

18

300

350

400

450

500

550

Sodium Na (%)
600

1.50

1.55

1.60

1.65

1 70

1 75
2009
1975

E
o

1945

O

1911

<D
O
c
0)
E
s
to in

1847

I

330

1

I

350

I

I

I

370

1-35

1.40

1.45

I

I

I

1

1.50

1

Thorium Th (ppm)

1.55

1

1.60

!

12

13

14

15

1

1

1

1

1

Titanium Ti (ppm)

Tin Sn (ppm)

16

17

1

3.4

3.6

3.8

4.0

0.75 0.80 0.85 0.90 0.95

1.00

1____ I____ I____ I
2009

E
o

1975

in

4=
&
c

1847

10) 8
COtn
CO

Uranium U (ppm)

Vanadium V (ppm)

Yittrium Y (ppm)

Zinc Zn (ppm)

Zirconium Zr (ppm)
15

o

m

20

25

I____ I____ I
2009
1975
1945 <9

o

1911 ^
1847

c rvj
o

NJ

Appendix: Trace Metal Profiles

Thallium Tl (ppm)

Strontium Sr (ppm)
310