Mercury Assessment and Monitoring Protocol for the 

Bear Creek Watershed, Colusa County, California 

Scientific Investigations Report 2010-5018

Prepared for the Bureau of Land Management

U.S. Department of the Interior

U.S. Geological Survey
Cover: Historical photograph of the 10-pipe retort furnace, Manzanita Mine, Colusa County, which was used to process 

previously concentrated mercury ore, circa 1900. This photograph is from the 1903 California State Mining Bureau Bulletin 27. 

Credit to Ron Churchill, California Geological Survey. 
Mercury Assessment and Monitoring 

Protocol for the Bear Creek Watershed, 

Colusa County, California

By Thomas H. Suchanek, Roger L. Hothem, James J. Rytuba, and Julie L. Yee

Prepared for the Bureau of Land Management

Scientific Investigations Report 2010-5018

U.S. Department of the Interior

U.S. Geological Survey
U.S. Department of the Interior

KEN SALAZAR, Secretary

U.S. Geological Survey

Marcia K. McNutt, Director

U.S. Geological Survey, Reston, Virginia: 2010

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Suggested citation:

Suchanek, T.H., Hothem, R.L., Rytuba, J.J., and Yee, J.L., 2010, Mercury assessment and monitoring protocol for the 

Bear Creek Watershed, Colusa County, California: U.S. Geological Survey Scientific Investigations Report 2010-5018, 

34 p.
iii

Contents

Abstract...........................................................................................................................................................1

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

Objectives...............................................................................................................................................1

Study Area..............................................................................................................................................1

Geology..........................................................................................................................................4

Sources of Mercury ...........................................................................................................4

Geothermal Springs............................................................................................................4

Cold Carbonate Springs.....................................................................................................7

Bedrock ................................................................................................................................7

Mercury and Gold Mines ..................................................................................................7

Mercury Assessments in the Bear Creek Watershed..................................................................10

Physical/Chemical Assessments ............................................................................................10

Upper Bear Creek .............................................................................................................10

Sulphur Creek....................................................................................................................10

Middle Bear Creek Downstream of Sulphur Creek.....................................................11

Biological Assessments ...........................................................................................................11

Invertebrates .....................................................................................................................12

Fish ................................................................................................................................12

Amphibians and Birds......................................................................................................16

Overview of Historical Data ..............................................................................................................16

Mercury Monitoring Protocol....................................................................................................................17

Objective...............................................................................................................................................17

Sampling...............................................................................................................................................17

Seasonality..................................................................................................................................17

Physical/Chemical Matrices....................................................................................................17

Sample Collection and Processing................................................................................18

High-Flow Event Sampling ..............................................................................................18

Sediment Collection and Analysis .................................................................................19

Biological Matrices ...................................................................................................................19

Invertebrates .....................................................................................................................19

Fish ................................................................................................................................20

Preparation for Collections .............................................................................................21

Quality Assurance and Quality Control (QA/QC)..........................................................21

Sample Collection and Processing................................................................................22

Invertebrates ............................................................................................................22

Fish .......................................................................................................................23

Sample Shipment..............................................................................................................23

Statistics...............................................................................................................................................23

Power and Sample Size for Testing Percent Difference Between Pre-R and 

Post-R Periods ..............................................................................................................24

Analytical Costs...................................................................................................................................25
iv

Summary and Conclusions.........................................................................................................................27

Acknowledgments.......................................................................................................................................27

References Cited..........................................................................................................................................27

Appendix A. Fish Processing Form for Bear Creek Mercury Study ....................................................31

Appendix B. Invertebrate Laboratory Processing Form for Bear Creek Mercury Study ................32

Appendix C. Labels for Sample Containers .............................................................................................33

Appendix D. Example Chain of Custody Form .........................................................................................34

Contents—Continued

Figures

Figure 1. Map showing Bear Creek Watershed with locations of mines, springs, and

proposed sampling locations for mercury monitoring ................................. 2

Figure 2. Geological map and ore deposits and geothermal springs located in the 

Clear Lake volcanic field, including the Bear Creek watershed ..................... 3

Figure 3. Map showing geology and Hg and Au deposits located in the Sulphur Creek

tributary to Bear Creek ..................................................................... 6

Figure 4. Map showing location of the Rathburn and Petray Hg mines and tributaries 

1-4 that drain the mine area and release Hg-enriched sediment into Bear Creek... 8

Figure 5. Map showing Cache Creek Watershed, including its two major tributaries,

Bear Creek and North Fork Cache Creek; red dots indicate invertebrate

mercury bioindicator sampling sites used by Slotton and others during 1995–97 ... 13

Figure 6. Map showing mercury concentrations (ppm dw) in stream invertebrates from

the Cache Creek watershed in 1995 ...................................................... 14

Figure 7. Map showing mercury (ppm dw) in stream invertebrates in the Bear Creek

Watershed from D. Slotton collections in 1996 .......................................... 15

Tables

Table 1. Bear Creek watershed monitoring sites ................................................ 5

Table 2. Available and Target* invertebrate families, Bear Creek .............................. 20

Table 3. Available and Target* fish species, Bear Creek ....................................... 21

Table 4. Summary of invertebrate and fish sample collections for 

Bear Creek Watershed ..................................................................... 21

Table 5. Equipment checklist ........................................................................ 22

Table 6. Fish and invertebrate collection checklist................................................ 22

Table 7. Estimated costs of individual analyses ................................................... 25

Table 8. Summary of water and sediment chemical analyses per site for 

Bear Creek watershed ..................................................................... 25

Table 9. Summary of invertebrate and fish chemical analyses per site for 

Bear Creek watershed ..................................................................... 26

Table 10. Estimated budget (in dollars) for Federal staff to conduct Bear Creek 

sampling based on 2009 costs ............................................................ 26
v

Conversion Factors, Datum, and Acronyms

Conversion Factors

Multiply By To obtain

Length

inch (in.) 2.54 centimeter (cm)

inch (in.) 25.4 millimeter (mm)

foot (ft) 0.3048 meter (m)

mile (mi) 1.609 kilometer (km)

Area

acre 4,047 square meter (m2)

acre 0.4047 hectare (ha)

acre 0.004047 square kilometer (km2)

Volume

gallon (gal) 3.785 liter (L) 

gallon (gal) 0.003785 cubic meter (m3) 

cubic inch (in3) 16.39 cubic centimeter (cm3) 

cubic inch (in3) 0.01639 liter (L)

cubic foot (ft3) 0.02832 cubic meter (m3) 

Flow rate

cubic foot per second (ft3/s) 0.02832 cubic meter per second (m3/s)

gallon per minute (gal/min) 0.06309 liter per second (L/s)

Mass

gram (g) 0.03527 ounce, avoirdupois (oz)

kilogram (kg) 2.205 pound, avoirdupois (lb)

microgram (μg) 0.0000000353 ounce, avoirdupois (oz)

milligram (mg) 0.0000353 ounce, avoirdupois (oz)

nanogram (ng) 0.0000000000353 ounce, avoirdupois (oz)

ounce, avoirdupois (oz) 28.35 gram (g) 

pound, avoirdupois (lb) 0.4536 kilogram (kg) 

ton, short (2,000 lb) 0.9072 megagram (Mg) 

ton, long (2,240 lb) 1.016 megagram (Mg) 

Temperature in degrees Celsius (°C) may be converted to degrees Fahrenheit (°F) as follows:

°F = (1.8×°C)+32.

Temperature in degrees Fahrenheit (°F) may be converted to degrees Celsius (°C) as follows:

°C = (°F-32)/1.8.

Specific conductance is given in microsiemens per centimeter at 25 degrees Celsius (μS/cm at 

25 °C).

Concentrations of chemical constituents in water are given in milligrams per liter (mg/L, parts 

per million), micrograms per liter (μg/L, parts per billion), or nanograms per liter (ng/L, parts per 

trillion).

Concentrations of chemical constituents in tissues are given in micrograms per gram (μg/g, 

parts per million).

Concentrations of chemical constituents in sediments are given in micrograms per gram (μg/g, 

parts per million) or nanograms per gram (ng/g, parts per billion).
vi

Conversion Factors, Datum, and Acronyms—Continued

Datum

Horizontal coordinate information is referenced to the North American Datum of 1983 (NAD 83).

Acronyms

ANOVA Analysis of variance

ASTM American Society for Testing and Materials

Au Gold

B Boron

BLM Bureau of Land Management

CDFG California Department of Fish and Game

Cl- Chloride

CVRWQCB Central Valley Regional Water Quality Control Board

DOC Dissolved organic carbon

Fe Iron

FLPE Fluorinated polyethylene

HCl Hydrochloric acid

Hg Mercury

K Potassium

Li Lithium

MMeHg Monomethyl mercury

Na Sodium

NAWQA National Water-Quality Assessment

ORP Oxidative-reductive potential

Rb Rubidium

S Sulfur

Se Selenium

Sr Strontium

TMDL Total maximum daily load

TotHg Total mercury

USBM U.S. Bureau of Mines

USEPA U.S. Environmental Protection Agency

USGS U.S. Geological Survey

W Tungsten

YSI Yellow Springs Instruments
Mercury Assessment and Monitoring Protocol for the 

Bear Creek Watershed, Colusa County, California

By Thomas H. Suchanek, Roger L. Hothem, James J. Rytuba, and Julie L. Yee

Abstract

This report summarizes the known information on 

the occurrence and distribution of mercury (Hg) in physical/

chemical and biological matrices within the Bear Creek 

watershed. Based on these data, a matrix-specific monitoring 

protocol for the evaluation of the effectiveness of activities 

designed to remediate Hg contamination in the Bear Creek 

watershed is presented. The monitoring protocol documents 

procedures for collecting and processing water, sediment, 

and biota for estimation of total Hg (TotHg) and monomethyl 

mercury (MMeHg) in the Bear Creek watershed. The 

concurrent sampling of TotHg and MMeHg in biota as well 

as water and sediment from 10 monitoring sites is designed 

to assess the relative bioavailability of Hg released from 

Hg sources in the watershed and identify environments 

conducive to Hg methylation. These protocols are designed 

to assist landowners, land managers, water quality regulators, 

and scientists in determining whether specific restoration/

mitigation actions lead to significant progress toward 

achieving water quality goals to reduce Hg in Bear and 

Sulphur Creeks. 

Introduction

Objectives

The objectives of this report are (1) to summarize the 

known information on the occurrence and distribution of 

mercury (hereafter Hg) in physical/chemical and biological 

matrices within the Bear Creek watershed, and (2) using this 

information as a basis, develop a matrix-specific monitoring 

protocol for the evaluation of the effectiveness of activities 

designed to remediate Hg contamination in the Bear Creek 

watershed.

Study Area

Bear Creek, Cache Creek, and the North Fork of Cache 

Creek are the major tributaries of the Cache Creek watershed, 

encompassing 2,978 km2 (fig. 1). The Cache Creek watershed 

contains soils naturally enriched in mercury (Hg) as well as 

natural springs (both hot and cold) with varying levels of 

aqueous Hg. All three tributaries are known to be significant 

sources of anthropogenically derived Hg from historic mines, 

both Hg and gold (Au), and associated ore storage/processing 

sites and facilities (D.G. Slotton, unpub. data, 1995; Foe and 

Croyle, 1998; Schwarzbach and others, 2001; Tetra Tech EMI, 

2003; Domagalski and others, 2004; Slotton, 2004; Suchanek 

and others, 2004, 2008a, 2009; Gassel and others, 2005). 

The Bear Creek watershed is located in the northeastern 

part of the Clear Lake volcanic field that is composed of 

Pliocene to Holocene volcanic centers (fig. 2). It is the 

youngest of the volcanic fields that formed along the margin 

of the North American plate as the Mendocino triple junction 

migrated northward along the coast of California. The high 

heat flow associated with the volcanism resulted in the 

formation of Hg and gold-silver (Au–Ag) ore deposits. In 

addition, numerous active hot springs and gas vents are 

present throughout the volcanic field, including the Bear 

Creek watershed (figs. 1, 2, 3, 4). The ore deposits are the 

northernmost and youngest in the Coast Range Hg mineral 

belt. Mercury and Au–Ag deposits are present on the west 

side of the Bear Creek watershed and in the drainage area of 

Sulphur Creek, a major tributary of Bear Creek. Hot springs 

that are actively depositing Hg and Au are associated with 

some of these Hg–Au deposits. The geothermal springs reflect 

the waning stage of the hydrothermal systems that formed 

the Hg and Au deposits. Cold to weakly thermal carbonate 

springs also are present in the Bear Creek watershed. These 

springs are actively depositing travertine, and some of these 

fluids contain anomalously high concentrations of Hg. The 

hot springs and cold carbonate springs are characterized by 

high salinity waters that are derived from evolved connate 

water (water trapped in pores during deposition of a sediment) 

present in the sedimentary rocks of the Great Valley sequence. 

The Bear Creek watershed is unusual in that meteoric 

water (groundwater derived from precipitation) does not 

always dominate the composition of waters in Bear Creek 

and some of its tributaries. High salinity effluent from the 

numerous geothermal springs, cold carbonate springs, and 

high salinity groundwater is a significant source of water to 

Bear Creek and contributes to its relatively high conductivity. 
2 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Figure 1. Bear Creek Watershed with locations of mines, springs, and proposed sampling locations for 

mercury monitoring. Locations of figures 3 and 4 are indicated on this figure.

Clearlake

Brushy

Sky High

932 m

Round

Mountain

651 m

Schoolteacher

Hill

504 m

Sky High

829 m

Split

Rock

937 m

Stony Top

394 m

Sugarloaf

496 m

Three

Sisters

486 m

Whiskey

Hill

346 m

Round

Mountain

545 m

Anderson

Mountain

558 m

Bald Hill

351 m

Bald

Mountain

631 m

Baldy

Mountain

638 m

Baldy

Mountain

1020 m

Bally

Peak

657 m

Bear Valley

Buttes

596 m

Billys

Hill

492 m

Quackenbush

Mountain

635 m

Buck

Mountain

815 m

Chalk

Mountain

472 m

Chocolate

Peak

1003 m

Cold Spring

Mountain

1059 m

Cooks

Mountain

644 m

Coyote

Peak

485 m

Gibson

Hill

697 m

Glascock

Mountain

703 m

Gravelly

Buttes

524 m

Helmer

Hill

544 m

Jackson

Mountain

800 m

La Coma

Tallac

725 m

Lambert

Hill

520 m

Langs

Peak

780 m

Lodoga

Peak

694 m

Lytle

Mountain

635 m

Pacific

ce

ntain

m

Red Rocks

484 m

Reister

Knoll

774 m

Reister

Rock

947 m

Clear

Lake

Indian

Valley

Reservoir

Burns Valley

Antelope Creek

Freshwater Creek

East Fork Sulphur Creek

Lurline Creek

Trout Creek

West Fork Spa

Calvins Creek

Sulphur Creek

Stanton Creek

Long Valley Creek

Little Stony Creek

Kilpepper Creek

Glenn Valley Slough

Wolf Creek

Herndon Creek

Little Indian Creek

Trout Creek

Rocky Creek

Mill Creek

North Fork Cache Creek

North Fork Cache Creek

Bear Creek

Sweet Hollow Creek

Perkins Creek

Dry Creek

Phipps Creek

Hog Hollow Creek

Chandans Creek

Middle Creek

Salt Creek

Molesworth Creek

Cache Creek

Indian Creek

Copsey Creek

Brushy Creek

Trout Creek

Grizzly Creek

Indian Creek

Manzanita Creek

Salt Branch

Mill Creek

Soda Creek Spanish Creek

South Fork Mill Creek

errington Creek

WaltersCreek

Manor Slough

SpringCreek

East Fork Spanish Creek

Davis Creek

Empire Mine

Rathburn Mine

Turkey Run

Wide Awake Mine

Petray North Pit

Petray South Pit

1

2

3

4

5

7

8

9

10

6

53 16

29

20

Colusa Co Yolo Co

Colusa Co

Lake Co

Yolo Co Lake Co

122°30'W

122°30'W

39°15'N

39°15'N

39°N

39°N

2 0 2 4

Kilometers

Bear Creek Watershed Overview

1

Project

Location

Sampling Stations

Named Mines

Unnamed Mines

Bear Creek Watershed

Watershed Boundaries

Land Status

Bureau of Land Management

US Forest Service

State

Private/Other

Clyde Mine

Elgin Mine

Manzanita Mine

Abbott Mine

Area in

Figure 3

Area in

Figure 4
Introduction

3

Figure 2. Geological map and ore deposits and geothermal springs located in the Clear Lake volcanic field, including the Bear Creek watershed. Wilbur 

Springs and adjacent Hg and Au deposits and geothermal springs are significant sources of Hg to Bear Creek.

Lake

Berryessa

Clear

Lake

McLaughlin

mine

10 KILOMETERS

38o3945'

o

39o15'

122o45' 122o15'

The Geysers

Steam Field

EXPLANATION

QUATERNARY DEPOSITSCLEAR LAKE VOLCANIC ROCKSSONOMA VOLCANIC ROCKSGREAT VALLEY SEDIMENTARY ROCKSOPHIOLITE ROCKSFRANCISCAN FORMATIONHOT SPRINGSMERCURY DEPOSITSGOLD DEPOSITS

FAULT

Cherry Hill

Wilbur

Springs

Clyde

Abbott

Petray 

Rathburn 

Wide Awake

Empire 
4 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

watersheds. The geothermal springs are located within the 

Sulphur Creek watershed and include Wilbur Springs, and, 

farther upstream, Jones Fountain of Life, Elbow, Blank, 

and several hot springs within the Elgin Hg deposit. The 

Jones Fountain of Life and two smaller hot springs (Blank 

and Elbow) occur on the eastern margin of the Cherry Hill 

Au deposit and adjacent to the Manzanita Hg deposit. Cold 

carbonate springs are present throughout the Bear Creek 

watershed and are localized along faults that bound the east 

and west side of Bear Valley and the east side of the lower 

reach of Bear Creek. A large number of cold saline springs 

are localized along the Bear Fault (located on the west side 

of Bear Valley and to the east of the Rathburn and Petray 

Hg mines). Effluent from these springs, along with saline 

groundwater, significantly increases the conductivity of Bear 

Creek water (Slowey and Rytuba, 2007). Downstream of the 

input from the cold springs, the chloride (Cl-) concentration 

of Bear Creek is 94 milligrams per liter (mg/L, ppm = parts 

per million) (Site 3, table 1) as compared to 30 ppm upstream 

of the input of the springs (Site 2, table 1), an increase of 

300 percent. Because of the addition of saline groundwater, 

the Cl- concentration of Bear Creek increases to 150 ppm 

just upstream of the confluence with Sulphur Creek (Site 4, 

table 1). However, sulfate concentration in the 6.5-km reach of 

Bear Creek from Site 2 to Site 4 remains about the same, at a 

relatively high concentration of 17–19 ppm. The conductivity 

of Bear Creek increases substantially downstream of its 

confluence with Sulphur Creek because of the effluent from 

high salinity hot springs in the Sulphur Creek watershed. 

During the dry season, Sulphur Creek waters are composed 

primarily of saline, hot spring effluent that also contains 

elevated concentrations of sulfate, Hg, W, B, and iron (Fe).

Geothermal Springs

In the lower part of the Sulphur Creek watershed, 

geothermal springs are localized along the Resort fault zone. 

At Wilbur Springs, several hot spring vents form a coalescing 

travertine terrace along the northern bank of Sulphur Creek 

(figs. 2 and 3). The temperature of the Wilbur Hot Springs 

geothermal waters ranges as high as 56oC, and the waters 

have very high concentrations of Cl-

 (10,900 ppm) and B 

(280 ppm). The concentration of total Hg (TotHg) in the 

Wilbur springs ranges from 6.4 to 6.7 μg/L (ppb = parts 

per billion) (Janik and others, 1994). Suchanek and others 

(2004) measured the geothermal waters where they feed 

the hot baths at the Wilbur Springs Resort and reported 

TotHg concentrations ranging from 3.5 to 7.3 ppb with 26 

to 69 percent of the TotHg present in the dissolved fraction. 

Black sediment that precipitates from the hot spring water 

contains 27 ppm TotHg and 4.3 ppm Au (Peters, 1991). This 

Hg-enriched sediment enters Sulphur Creek from the hot 

spring vent area as the hot spring effluent cools and flows over 

the travertine terrace. 

Conductivity under low-flow conditions is 800 μS/cm in the 

upper part of the watershed and increases to 1,200 μS/cm just 

upstream of the confluence with Sulphur Creek. During the 

dry season, the effluent from saline springs and groundwater 

predominate, and Bear Creek water reaches its maximum 

salinity. During the wet season, runoff reduces the salinity of 

Bear Creek, and the waters of Bear Creek and its tributaries 

are predominantly meteoric. The effluent from the high salinity 

springs also contains elevated concentrations of Hg, sulfate, 

boron (B), and tungsten (W) that significantly affect water 

quality, especially under low-flow conditions during the dry 

season, and during low rainfall years. Important components 

that contribute to the methylation of Hg include sulfate, 

salinity, dissolved organic carbon (DOC), and Hg. Thus, 

seasonal changes in the water chemistry of Bear Creek and 

some of its tributaries, especially Sulphur Creek, can affect the 

physical and chemical variables that control Hg methylation. 

It is, therefore, important that the monitoring protocol specify 

sampling under similar seasonal and flow conditions so that 

results may be compared under similar flow conditions.

Geology

The north-south trend of Bear Creek reflects a major 

geologic boundary in California that is termed the Coast 

Range fault. The fault separates Cretaceous sedimentary 

rocks of the Great Valley sequence and the Coast Range 

ophiolite to the east from rocks of the Franciscan Complex 

to the west (fig. 2). Initial movement along the Coast Range 

fault thrust Great Valley sequence rocks above the similar 

age Franciscan Complex. This initial period of compressional 

faulting was followed by downward displacement along lowangle detachment faults. The hills that comprise the east side 

of Bear Valley consist of Great Valley sequence sandstones, 

siltstones, and shales that have been tilted upward and dip to 

the east at angles up to 45 degrees. These sedimentary rocks 

comprise the floor of Bear Valley as well as the first low hills 

on the west side of the valley. The uplifted west side of Bear 

Valley is bounded by a series of faults that separates Great 

Valley sequence rocks to the east from rocks of the Franciscan 

Complex and Coast Range ophiolite to the west where these 

rocks form the high hills of Walker Ridge. The Rathburn and 

Petray Hg deposits are hosted in hydrothermally altered Coast 

Range ophiolite, however, the Hg deposits in the Sulphur 

Creek watershed are hosted in sedimentary rocks of the Great 

Valley sequence.

Sources of Mercury

Both natural and anthropogenic sources contribute 

Hg to Bear Creek and its tributaries. The natural sources 

consist of geothermal springs, cold carbonate springs, and 

hydrothermally altered rock associated with Hg and Au 

deposits, and unaltered rock. The anthropogenic sources 

include abandoned Hg mines, Au mines, and ore-processing 

facilities present in the Bear Creek and Sulphur Creek 
Introduction 5

Elbow spring, located on the southern bank of Sulphur 

Creek (fig. 3), has the highest water Cl- concentration 

(13,390 ppm) and TotHg concentration (61.0 ppb) of the 

geothermal springs in the Sulphur Creek watershed. The black 

precipitate at the vent contains 179 ppm TotHg and 12.1 ppm 

Au (Janik and others, 1994). Several unnamed hot spring vents 

occur in Sulphur Creek downstream of the Elbow spring and 

precipitate similar black sediment. Under low-flow conditions, 

the fine-grained precipitate accumulates in this section of 

Sulphur Creek until the first high-flow event of the wet 

season transports the Hg-enriched sediment downstream into 

Bear Creek. Upstream of the Elbow spring is Jones Fountain 

of Life, a spring that erupts about every 30 minutes with a 

maximum flow of 1.5 L/sec. The water has a Cl-

 concentration 

of 11,260 ppm, a TotHg concentration of 22.0 ppb, and an 

estimated TotHg flux from this spring of 1.1 kg/yr (Janik 

and others, 1994). Suchanek and others (2004) reported that 

the TotHg concentration in this spring ranges from 24.3 to 

39.7 ppb, with 14 to 35 percent of the TotHg in the dissolved 

fraction. As the geothermal water cools and flows from the 

vent, it precipitates black, fine-grained sediment that contains 

157 ppm TotHg and 4 ppm Au. The black precipitate consists 

of an amorphous aluminum silicate that contains Fe, sulfur 

(S), and potassium (K). Crystals of cinnabar (up to 1 mm in 

size) and barite (up to 6 mm) occur within the amorphous 

black precipitate. Blank hot spring is located south of Sulphur 

Creek, and effluent and precipitate from the hot spring do 

not enter the creek except under very high-flow conditions. 

The hot spring water has a Cl-

 concentration of 8,700 ppm 

and TotHg concentration of 6.9 ppb. However, because of its 

relatively low flow (0.25 L/sec), the estimated TotHg flux from 

this spring is only about 0.055 kg/yr (Janik and others, 1994).

In the upper part of the Sulphur Creek watershed, several 

hot springs and gas vents are present in and adjacent to the 

Elgin Hg mine. The Elgin hot spring, located within the open 

cut of the Elgin Hg mine, has the highest temperature, 67oC, 

of all geothermal springs in the Sulphur Creek watershed. The 

Cl- concentration of the water is 11,400 ppm, and the TotHg 

concentration is 11.0 ppb. The flow from the hot spring vent is 

1.07 L/sec, and the estimated TotHg flux is 0.15 kg/yr (Janik 

and others, 1994). Waters from other hot springs adjacent to 

the Elgin Hg mine have lower Hg concentrations, 0.7 ppb. 

Table 1. Bear Creek watershed monitoring sites (NAD 83).

Site No. Monitoring site Latitude Longitude

Distance from 

Cache Creek 

(km)

1 Milk Creek at Brim Road; upper watershed 

characterization above known mercury sources

39° 09’ 46.90” N 122° 26’ 50.20” W 34.80

2 Bear Creek at bridge above tributaries from 

Rathburn Petray; above all known mine input 

and to be used for comparison with site 3

39° 05’ 51.40” N 122° 24’ 48.90” W 25.78

3 Bear Creek downstream of Site 2; assess input 

from tributaries from Rathburn-Petray mines 

and cold saline springs along Bear Fault

39° 4’ 50.27” N 122° 24’ 48.1” W 23.65

4 Bear Creek upstream of Sulphur Creek 

(Hamilton); assess attenuation of input from 

Rathburn-Petray mines

39° 03’ 24.00” N 122° 24’ 41.00” W 20.74

5 Sulphur Creek above Wilbur Hot Springs and 

most mines; assess input from upper watershed 

and Elgin mine and geothermal springs

39° 02’ 0.40” N 122° 25’ 49.80” W 21.41

6 Sulphur Creek at USGS Gauge; assess input 

from geothermal springs and mines

39° 02’ 19.0” N 122° 25’ 8.0” W 19.92

7 Bear Creek downstream of Sulphur Creek;

assess impact of input from Sulphur Creek

39° 02’ 20.23” N 122° 24’ 25.93” W 18.46

8 Bear Creek at Highway 20 Bridge; assess Hg 

methylation and transport from watershed 

impacted by mines and geothermal springs

39° 0’ 41.78” N 122° 21’ 40.34” W 12.33

9 Bear Creek at Thompson Canyon Bridge;

assess Hg methylation in area of low gradient 

and cattle grazing

38° 58’ 18.60” N 122° 20’ 26.60” W 6.73

10 Bear Creek just upstream of the confluence 

with Cache Creek; assess Hg and MMeHg 

released from entire Bear Creek watershed

38° 55’ 37.00” N 122° 20’ 0.00” W 0.08
6 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Sulphur Creek watershed are significant sources of particulate 

TotHg to Sulphur Creek, with average estimates ranging from 

0.6 to 10.7 kg/yr (Suchanek and others, 2004). The relative 

importance of the Hg–Au mine and geothermal Hg sources 

in Sulphur Creek has not been sufficiently documented, 

but under low-flow conditions and low rainfall years, the 

geothermal Hg source predominates. 

All geothermal springs in the Sulphur Creek watershed are 

elevated in sulfate and sulfide concentration, resulting in 

7–16 × 103 kg of sulfate being added to Sulphur Creek from 

these sources per year (Churchill and Clinkenbeard, 2002).

Sulphur Creek water downstream of all geothermal 

spring inputs has very high concentrations of TotHg, 1.3 ppb, 

88 percent of which is in the particulate form (Suchanek and 

others, 2004). The geothermal springs and mine sources in the 

Figure 3. Geology and Hg and Au deposits located in the Sulphur Creek tributary to Bear Creek. Geothermal springs adjacent 

to the ore deposits are actively depositing Hg and Au and are sources of Hg-enriched sediment to Bear Creek.

Cherry

 Hill

West End

Blank Spring

In BetweenManzanita

Jones Fountain

 of Life

Wide Awake

Elbow Spring

Central Mine

Empire EXPLANATION

ALLUVIUM

TRAVERTINE

SERPENTINITE

CONGLOMERATE

SANDSTONE,

SILTSTONE

ALTERATION

SILICIFICATION

ARGILLIC

FAULT

HOT SPRING

Sulphur

Creek

MERCURY MINE

X GOLD MINE

X

X

X

X

X

X

X X

X

0 125 250 M

Retort

N

X

X

X

X

X
Introduction 7

Cold Carbonate Springs

Cold carbonate springs are present along several 

faults in the Bear Creek watershed. A large number of these 

springs are localized along the Bear Fault, which is located 

to the west of Bear Creek and east of the Rathburn-Petray 

mines (fig. 4). These springs consist of variable mixtures of 

meteoric water and saline groundwater derived from connate 

fluids in sedimentary rocks in the Great Valley sequence. 

The cold spring waters have a highly variable proportion of 

saline groundwater, ranging from 13 to 100 percent. Springs 

along the central part of the Bear Fault have the highest 

component of saline groundwater and have exceptionally 

high conductivities (9,400 and 19,000 μS/cm). The most 

saline cold spring waters, 6,065 mg/L Cl-, are comparable to 

saline geothermal hot spring waters located in the Sulphur 

Creek watershed. The cold springs are characterized by high 

concentration of the cations sodium (Na), K, rubidium (Rb), 

lithium (Li), selenium (Se), strontium (Sr), and W, as well 

as nitrate, sulfate, and DOC. Plots of these cations against 

Cl-

 for the various cold springs in the Bear Valley watershed 

define a two-component linear mixing line, indicating that the 

spring waters are mixtures of meteoric and saline groundwater. 

Alkalinity is relatively high in cold carbonate springs and 

surface waters that drain the Rathburn-Petray mine area. This 

results from release of carbonate from saline groundwater, and 

leaching of carbonate from serpentinite and mafic bedrock 

of the Coast Range ophiolite and the Franciscan Formation. 

The presence of alkaline pH (> 8.0) indicates that the cold 

carbonate springs and some surface waters, with a significant 

component of saline groundwater, are in aqueous equilibrium 

with calcium carbonate. The alkalinity of spring waters ranges 

from 2.8 meq/L as carbonate (less groundwater-dominated) to 

29 meq/L in saline cold spring water that is actively depositing 

travertine. Bicarbonate is the dominant buffering agent, 

although organic acids also may buffer these waters. 

High concentrations of TotHg, up to 0.7 ppb, occur in 

some cold carbonate springs and saline surface waters that 

drain the Rathburn-Petray mine area. In cold carbonate spring 

water, Hg is present primarily in the filtered fraction, typically 

comprising from 78 to 100 percent of the TotHg present. In 

saline surface waters that are dominantly meteoric with a 

small component of high salinity groundwater, the amount 

of TotHg in the filtered fraction is highly variable, ranging 

from 100 percent to less than 4 percent. The concentrations of 

TotHg in both filtered and unfiltered waters are not correlated 

with Cl- concentration or any other major or minor element. 

Processes that control TotHg concentration in saline surface 

and spring waters are not conservative and may include 

precipitation and dissolution of carbonate, dissolution of HgS 

by DOC, and seasonal changes in surface-water flow. The 

highest TotHg concentration (0.9 ppb measured under dry 

season low-flow conditions) occurs in surface water from a 

tributary that drains the north pit of the Petray mine (North 

fork of Tributary 1 in Slowey and Rytuba, 2008). The water 

in this tributary is dominantly meteoric (< 1.5 percent saline 

groundwater), and the creek sediment has a high concentration 

of TotHg (350 ppm), which is present as cinnabar and 

metacinnabar. 

Bedrock

The Hg concentrations in unaltered bedrock exposed 

in the Bear Creek watershed are typically less than 154 ng/g 

(parts per billion = ppb), with higher Hg concentrations 

occurring in sedimentary rocks of the Great Valley sequence 

being higher in the range of Hg values (Smith and others, 

2008). The Hg concentration in unaltered shales, siltstones, 

and sandstones in the Great Valley sequence ranges from 31 

to 154 ppb. Much lower Hg concentrations, less than 24 ppb, 

occur in mafic rocks of the Coast Range ophiolite, serpentinite 

derived from the ophiolite, and mafic rocks in the Franciscan 

Formation. The Hg concentration in greywacke, blueschist, 

and cherty mudstone of the Franciscan Formation ranges from 

35 to 88 ppb. Volcanic rocks in the Clear Lake volcanic field 

typically have low Hg concentrations (<92 ppb). However, 

volcanic rocks are not present in the Bear Creek watershed, 

and only minor intrusive basaltic rock has been mapped in the 

Sulphur Creek watershed. Erosion of hill slopes composed of 

Great Valley sequence sedimentary rocks would be expected 

to produce sediment with a Hg concentration in the range of 

30 to 150 ppb. Sediment derived from hill slopes composed 

of Franciscan and Coast Range ophiolite would produce 

sediment with lower Hg concentrations, ranging from 20 to 

90 ppb. 

An estimate of the annual TotHg flux of 0.45 to 9.8 kg 

from background soils in Sulphur Creek has been calculated 

by assuming erosion rates between 0.5 and 10 (tons/acre)/yr 

applied over the entire watershed (Churchill and Clinkenbeard, 

2002), but the actual amount of regional background TotHg 

entering the watershed is unknown. No estimate has been 

made for the Bear Creek watershed.

Mercury and Gold Mines 

Mines in the Bear Creek watershed include several 

Hg and Au mines in Sulphur Creek, and the Rathburn and 

Petray Hg mines located in the hills (Walker Ridge) on the 

western side of Bear Valley. All of these Hg mines have 

had relatively small production, but the surface disturbance 

and mine wastes at the mines are highly variable. Surface 

exposures of cinnabar mineralization in the Petray mine open 

pits have high TotHg concentrations (up to 2,490 ppm) and are 

a continuing source of Hg-enriched sediment that is released 

from the mine site resulting in high concentrations of Hg 
8 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Figure 4. Location of the Rathburn and Petray Hg mines and tributaries 1- 4 that drain the mine area and release Hg-enriched sediment into Bear Creek. 

Some saline cold springs along the Bear Fault release Hg into Bear Creek. Cold carbonate springs east of the Petray Mine have relatively low concentrations of Hg as does the saline Cain Spring located east of Bear Creek. Bear Creek sample sites discussed in text and listed in table 1 are shown by red dots.
Introduction 9

in tributaries that drain the mine area (Slowey and Rytuba, 

2008). In Sulphur Creek, erosion of waste rock and tailings 

from the mine sites is a source of Hg-enriched sediment but 

Hg-enriched precipitates from geothermal springs located in 

and adjacent to the mines are likely more significant. Of the 

10 major tributaries that enter Bear Creek, Sulphur Creek has 

the highest sediment Hg concentrations with 91.8 ppm in the 

grain size fraction < 63 μm (Bosworth and Morris, 2009). 

In comparison, the other tributaries to Bear Creek have Hg 

concentrations in sediment that range from 0.04 to 0.53 ppm 

in the grain-size fraction < 63 μm (Bosworth and Morris, 

2009). The average background Hg concentration in sediment 

within the Cache Creek watershed is 0.06 ppm in the grain 

size fraction < 63 μm (Foe and Bosworth, 2008). Mercury 

mine tailings (calcines) with TotHg concentrations up to 

1,020 ppm are present at only three of the mines. Waste rock 

containing variable concentrations of TotHg ranging from 10 

to 1,020 ppm, and Hg-enriched soils ranging in concentration 

from 10 to 300 ppm are present at all of these mines and 

are a volumetrically more significant potential source of 

Hg-enriched sediment to the Bear Creek watershed (Churchill 

and Clinkenbeard, 2002).

The Manzanita Hg-Au mine and the nearby West End 

Hg-Au mine and In Between Au prospect (fig. 3) were 

developed on the periphery of the larger Cherry Hill hot spring 

Au-Hg system that was discovered by Homestake Mining 

Company in 1977 (Nelson and others, 1993). The mines occur 

on either side of Sulphur Creek (fig. 3). The Manzanita Au–Hg 

mine was discovered in 1863 and operated until 1942. Total 

production was 2,500 flasks of Hg (U.S. Bureau of Mines, 

1965). About 3,000 oz (about 85 kg) of Au was produced 

from the Cherry Hill and Manzanita mines from 1865 to 1891. 

The Au–Hg ores are hosted in silicified shales, sandstones, 

and conglomerates in the basal section of the Great Valley 

sequence. Cinnabar is the main Hg-bearing phase, but minor 

metacinnabar also was present. Where native Au was present, 

Hg also occurred as an amalgam (Churchill and Clinkenbeard, 

2002). Other sulfides present in the ore included pyrite, 

marcasite, and stibnite. Bitumen and petroleum occur in some 

of the gold-quartz veins, especially at the In Between mine. 

The Manzanita mine workings include several adits and shafts 

and, in the upper part of the deposit, ores were mined from 

a glory hole and open cut. Various processing techniques 

were used at the Manzanita mine. These included stamp mills 

that introduced Hg to recover the Au by amalgamation, and 

mechanical concentration of cinnabar to create a cinnabar 

concentrate from which Hg was recovered by a retort 

(Churchill and Clinkenbeard, 2002). In the 1940s, the Hg 

concentrates from the mine were processed at the Wide Awake 

mine, located about 1 km to the south. There are presently no 

tailings at the Manzanita, West End, In Between, and Cherry 

Hill mines. There is a limited amount of waste rock at each 

of the mine sites, but exposures of altered rock are potential 

sources of Hg-enriched sediment. 

The Central and Empire Hg mines have produced a small 

amount of Hg estimated to be about 170 flasks from ores that 

were processed primarily at the Wide Awake mine. There are 

small amounts of tailings and waste rock at the Central mine, 

and waste rock is present around and within a small retort 

at the Empire mine. The mine site is vegetated and does not 

appear to be a source of Hg-enriched sediment to Sulphur 

Creek (Tetra Tech, 2003).

The Wide Awake mine was discovered in the 1870s 

and produced 1,800 flasks of Hg until the mine was closed 

in the 1940s. Some of the production came from ores that 

were mined at the Manzanita and Empire mines. The Hg ore 

occurs in the basal section of the Great Valley sequence at 

the contact between shale and sediments composed of detrital 

serpentinite. Cinnabar is the main ore mineral and occurs in 

opalized serpentinite that locally contains abundant petroleum. 

Mine workings include several surface cuts and underground 

workings that were accessed by a 152-meter shaft (U.S. 

Bureau of Mines, 1965). Mercury ores were processed in a 

24-ton Scott furnace and several smaller retorts. The pile of 

tailings that includes some waste rock is estimated to contain 

11,000 tons of material (Churchill and Clinkenbeard, 2002) 

and is a source of Hg-enriched sediment during runoff in the 

wet season.

The Elgin Hg mine is located in the headwaters of 

Sulphur Creek, and several hot springs and gas vents are 

present in and adjacent to the mine workings. The mine was 

discovered in the early 1870s, and only a small amount of 

production, about 50 flasks, has been reported for the mine. 

The Hg ores occur along the contact of serpentinite and shale 

of the Great Valley sequence and are hosted in silicified 

serpentinite and in acid-leached serpentinite in the upper part 

of the ore body. Cinnabar is the primary ore mineral, and 

native sulfur occurs in the acid-leached part of the ore body. 

Pyrite is present in the lower part of the ore body. The mine 

workings consist of surface cuts, adits, and underground 

workings that were limited in development because the hot 

springs and associated sulfur and methane gases precluded 

extensive underground mining (U.S. Bureau of Mines, 1965). 

The small amount of ore was processed in a retort, and the 

amount of tailings on the mine site is minor, but waste rock in 

the open cuts is a potential source of Hg-enriched sediment. 

The Elgin hot spring vents in the central part of the main open 

cut, and effluent from the spring is collected in a small pond. 

A black precipitate that is enriched in Hg forms in the hot 

spring vent and collects in the pond but is only released to the 

watershed under high flows.

The Clyde Au mine (fig. 1) ore body was discovered 

in the early 1860s and has produced a small amount of Au. 

Native Au and pyrite occur in a silicified body of serpentine. 

The Au was recovered in a mill that apparently did not use 

Hg; more recent reprocessing of the tailings used gravity 

separation rather than Hg to recover the Au. This mine has 

only a small potential to release Hg-enriched sediment into 

Sulphur Creek.
10 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

in Bear Creek water downstream of the Rathburn and Petray 

mine input under low-flow conditions is very low (2.2 ng/L; 

pptr = parts per trillion), it is still higher than in water 

upstream of the mine input, which has a TotHg concentration 

of 0.45 pptr.

Mercury Assessments in the Bear Creek 

Watershed

Physical/Chemical Assessments

Water samples were collected during 2000–2001 

from a number of sites within the Cache Creek watershed 

and analyzed for TotHg and monomethyl Hg (MMeHg) 

(Domagalski and others, 2004; Slotton and others, 2004; 

Suchanek and others, 2004). A subset of those sites was within 

the Bear Creek Watershed and their locations are identified 

below.

Upper Bear Creek

This site is located at the Bear Valley Road bridge 

crossing (N: 39o5.83’, W: 122o24.71’). This site was believed 

to be upstream of all known mine loading of Hg, and it is 

located upstream of the Hg inputs into Bear Creek identified 

by Slowey and Rytuba (2008). Results from Slotton and others 

(2004) collections are presented below. For both unfiltered and 

filtered aqueous TotHg and MMeHg, the Upper Bear Creek 

site exhibited concentrations that were among the lowest in the 

entire Cache Creek and Bear Creek watersheds. At the Upper 

Bear Creek site, unfiltered aqueous TotHg ranged from ca. 0.5 

to 4.0 pptr and filtered aqueous TotHg ranged from ca. 0.4 to 

1.0 pptr, respectively. These data suggest that the TotHg at this 

site is mostly associated with particles. MMeHg in unfiltered 

aqueous samples ranged from ca. 0.05 to 0.11 pptr and filtered 

aqueous samples ranged from ca. 0.02 to 0.10 pptr, suggesting 

that a large proportion of MMeHg is found in the dissolved 

form. Because the Upper Bear Creek site is upstream of all 

known mining sites, these results could indicate a geothermal 

or saline spring (as opposed to a mining source) as the primary 

source of both TotHg and MMeHg in this region of Bear 

Creek. 

Sulphur Creek

This site, located in Sulphur Creek upstream of the 

confluence with Bear Creek (N: 39o 2.21’, W: 122o 24.56’), 

represents Hg inputs from Hg mines, Au mines, and 

geothermal springs along Sulphur Creek. Data from Slotton 

and others (2004) on aqueous TotHg and MMeHg in Sulphur 

Creek demonstrate a different result in comparison with 

the Upper Bear Creek site. Unfiltered and filtered TotHg 

ranged from ca. 0.3 to 1.1 ppb and from ca. 0.09 to 0.3 ppb, 

respectively, suggesting that at this site, the largest component 

of TotHg is found in the particulate form, not in the dissolved 

Estimates of yearly TotHg flux from mine wastes into 

Sulphur Creek have been based on estimates of erosion rates 

from mine wastes with known Hg concentrations. The Hg flux 

into Sulphur Creek is estimated to range from 4.4 to 18.6 kg, 

while the range of Hg flux into Bear Creek is from 0.7 to 

23.5 kg (Churchill and Clinkenbeard, 2002). However, there is 

high uncertainty in these estimates and only very limited data 

are available to quantify the Hg flux from the mine sites. Only 

two samples of water from the intermittent creek that drains 

the Wide Awake mine area have been collected, and the data 

indicate significant release of Hg from the mine site during 

the wet season. The concentration of TotHg in water from 

the Wide Awake mine drainage measured in February 2000 

and 2001 was 2.5 and 4.3 ppb respectively, and from 52 to 

58 percent of the TotHg was in the filtered fraction (Suchanek 

and others, 2004). 

The Rathburn Hg mine ore body was discovered and 

initially mined in the early 1890s. The Rathburn and the more 

recently developed Petray open pit mines are localized along 

fault zones in serpentinite that has been altered and cut by 

quartz and chalcedony veins. Cinnabar is the primary ore 

mineral, and metacinnabar has been identified in sediment 

derived from the Petray mine. The Hg ores formed in a 

hot spring system in the steam-heated environment present 

above boiling groundwater table. However, no active hot 

springs are present in the deposits. The Rathburn Hg mine is 

relatively small, having produced 100 flasks of Hg. Mining 

in the late 1960s and early 1970s recovered about 400 flasks 

of Hg from the Petray open pit mine (U.S. Bureau of Mines, 

1965). At the Rathburn Mine, Hg ores were processed in a 

brick retort, and small amounts of calcines are present that 

contain up to 1,020 ppm Hg (Churchill and Clinkenbeard, 

2002; Slowey and Rytuba, 2008). Waste rock derived from 

open cuts (in altered serpentinite) contains less than 39 ppm 

TotHg. Ores from the Petray Mine were processed offsite in 

a rotary furnace at the Abbott Mine, and, as a result, there 

are no mine tailings at the Petray mine. However, waste rock 

and ore in altered serpentinite are sources of Hg-enriched 

sediment that are released from the mine during storm events. 

The Hg concentration in sediments of the two tributaries 

that drain the Petray mine is highly elevated for the sampled 

section extending 2 km from the mine site. Both cinnabar 

and metacinnabar are present in sufficient quantity to be 

panned easily from the sediment (Slowey and Rytuba, 2008). 

The relative importance of Hg-enriched sediment released 

from the Rathburn and Petray mines as compared with Hg 

released from cold carbonate springs along the Bear Fault 

has not been determined, but both sources contribute Hg to 

Bear Creek (Slowey and Rytuba, 2008). Sediment in Bear 

Creek just downstream of the input from tributaries that drain 

the Rathburn and Petray mines (Monitoring Site 3, table 1) 

has a highly elevated TotHg concentration, 9,290 ppb. The 

TotHg concentration in Bear Creek sediment is considerably 

lower, 80 ppb, upstream of the Rathburn and Petray input 

(Monitoring Site 2, table 1), which indicates a significant Hg 

release from the mine sites. Although the TotHg concentration 
Introduction 11

form. MMeHg ranged from ca. 0.2 to 20 pptr in unfiltered 

samples and from ca. 0.09 to 1.5 pptr in filtered aqueous 

samples, again suggesting a significant component derived 

from particulate MMeHg. Although there are several large 

geothermal springs in and along Sulphur Creek, numerous 

adjacent Hg and Au mines erode into Sulphur Creek. Thus, 

although the dissolved fraction of both TotHg and MMeHg 

derived from the geothermal springs likely contribute 

significantly to Hg loading into Sulphur Creek, the particulate 

Hg found in the eroding soils from the surrounding mines 

likely overwhelms the contribution from the springs. 

Within Sulphur Creek, Suchanek and others (2004) 

also analyzed unfiltered water collected during February 

2000 and 2001. These data indicated that the highest TotHg 

concentrations were extremes observed from geothermal 

spring sites as follows: (1) The Jones Fountain of Life spring: 

24.3 ppb in 2000 and 39.7 ppb in 2001, and (2) geothermal 

springs from which the Wilbur Hot Springs Resort draws hot 

water for their baths at 4.0–7.3 ppb in 2001. However, TotHg 

concentrations immediately downstream of the Resort at the 

USGS gaging station (Sulphur Creek Index Station) were 

considerably reduced to 1.0 ppb (90 percent of which was in 

particulate form) in 2000 and 1.3 ppb (88 percent of which 

was in particulate form) in 2001. 

Middle Bear Creek Downstream of Sulphur Creek

This site, located about 10 km downstream of the 

confluence of Sulphur Creek (at approximately Monitoring 

Site 9 on fig. 1) (N: 38o58.88’, W: 122o20.94’), represents a 

reach of Bear Creek that has approximately 10-fold dilution of 

Sulphur Creek inputs. Bear Creek exhibits TotHg and MMeHg 

concentrations that are intermediate between the Upper Bear 

Creek site and the Sulphur Creek site. TotHg in unfiltered 

and filtered aqueous samples exhibited ranges from ca. 11 to 

150 pptr and from ca. 8 to 40 pptr, respectively. MMeHg in 

unfiltered and filtered aqueous samples exhibited ranges from 

ca. 0.2 to 1.5 pptr and from ca. 0.09 to 0.5 pptr, respectively. 

The distribution of these ranges of TotHg and MMeHg with 

respect to the proportions represented by filtered and unfiltered 

(that is, dissolved vs. particulate) components suggest a 

mixture of contributions between dissolved and particulate 

sources. This interpretation would be consistent with the 

multiple sources influencing the composition of the water 

mass at a site 10 km downstream of the confluence of Sulphur 

Creek with Bear Creek. Data on TotHg and MMeHg from the 

Upper Bear Creek site suggested contributions from a source 

that is primarily in a dissolved form. This source water would 

then be mixed with a Hg source that was primarily particulate 

from Sulphur Creek. As particulate material from the Sulphur 

Creek source is deposited downstream along the Bear Creek 

streambed, the influence of particulate Hg diminishes, 

consistent with the greater overlap in the ranges of Hg in 

filtered and unfiltered samples for both TotHg and MMeHg. 

A visual analysis of time-series data for TotHg and 

MMeHg at the Middle Bear Creek site from January 2000 to 

September 2001 revealed that TotHg peaks occurred during 

the winter rainy season (February) when maximum erosion 

typically would send Hg-laden particles downstream. MMeHg 

peaks at this site occurred during the summer (July/August) 

both years. A further analysis of the MMeHg:TotHg ratios1

for water samples at these three sites reveals that this ratio 

is highest for the Upper Bear Creek site (0.1 for unfiltered 

water; 0.05 for filtered water), lowest for the Sulphur Creek 

site (0.001 for unfiltered water; 0.002 for filtered water), 

and intermediate for the Middle Bear Creek site (0.01 for 

unfiltered water; 0.01 for filtered water). 

Biological Assessments

Several types of bacteria can convert naturally occurring 

inorganic Hg to its more toxic form, MMeHg, which is 

a neurotoxin and mutagen. MMeHg may affect several 

physiological functions, including vision, response to stimuli, 

growth, and reproduction in both vertebrates and invertebrates. 

In some cases, MMeHg can cause mortality. The effects of Hg 

toxicosis are exhibited most prominently in the egg or fetal 

stage, but may affect juvenile and adult stages as well. Thus, 

it is important to understand what concentrations of Hg are 

present, and evaluate whether corrective measures are feasible 

to lessen the impacts of Hg to wildlife and humans in this 

region.

Wildlife resources of significance in the Bear Creek 

watershed include wintering bald eagles (Haliaetus 

leucocephalus), prairie falcons (Falco mexicanus), greater 

roadrunner (Geococcyx mexicanus), tule elk (Cervus elaphus 

nannodes), river otters (Lontra canadensis), and foothill 

yellow-legged frogs (Rana boylii) (Schwarzbach and others, 

2001; Bureau of Land Management, 2004). An estimated 

154 bird species have been documented in the watershed 

(Bureau of Land Management, 2004). BLM recognizes 

several sensitive species found in the watershed, including: 

Townsend’s western big-eared bat (Plecotus townsendii), 

Pallid bat (Antrozous pallidus), burrowing owl (Athene 

cunicularia), St. Helena mountain king snake (Lampropeltis 

zonata zonata), foothill yellow-legged frog, and western pond 

turtle (Actinemys marmorata), and 17 sensitive plant species. 

Two of these species, foothill yellow-legged frog and the 

western pond turtle, are California Species of Special Concern 

(Jennings and Hayes 1994; California Department of Fish and 

Game, 2009).

1

MMeHg:TotHg ratios typically are regarded as a proxy for MMeHg 

bioavailability.
12 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

proportion of MMeHg in these samples was lowest (17–33 

percent) at the Sulphur Creek site, intermediate at the Middle 

Bear Creek site (66–84 percent), and highest at Upper Bear 

Creek (81–97 percent). A visual analysis of time-series data 

for MMeHg concentrations in these invertebrates at the 

Middle Bear Creek site exhibited trends similar to water, with 

MMeHg peaks occurring during the summer (July/August) 

both years. That same analysis at the Upper Bear Creek site 

was not as conclusive, but exhibited a maximum in MMeHg 

during the summer of 2001 only.

Schwarzbach and others (2001) also analyzed aquatic 

insects (e.g., Trichoptera, Zygoptera, Anisoptera, and 

Megaloptera) in Bear Creek as well as other locations within 

the Cache Creek watershed. Their data revealed a pattern 

of lower TotHg concentrations (< 0.5 ppm dw) in the North 

Fork of Cache Creek, Mill Creek, and Bear Creek upstream 

of Sulphur Creek, but much higher concentrations at sites in 

Bear Creek downstream of Sulphur Creek (0.5–5.2 ppm dw) 

and in Sulphur Creek proper (5.0–8.7 ppm dw). In general, 

their results agreed with those of Slotton and others (1997) for 

comparable taxa and sites.

Fish

Slotton and others (2004) focused their fish Hg analyses 

from their wider Cache Creek watershed study primarily on 

California roach (Hesperoleucus symmetricus), speckled dace 

(Rhinichthys osculus), and red shiner (Notropis lutrensis), 

all three of which exhibited average MMeHg concentrations 

as a percentage of TotHg of 90 percent or greater. However, 

only California roach were abundant enough within the Bear 

Creek watershed to obtain sufficient samples to analyze trends. 

No fish were found in Sulphur Creek. However, an analysis 

of the MMeHg:TotHg ratio was conducted on California 

roach from Upper Bear Creek and Middle Bear Creek. Upper 

Bear Creek roach (N = 29) exhibited a MMeHg percentage 

of 97.66 percent, whereas the roach at the downstream site, 

Middle Bear Creek (N = 31), yielded a MMeHg percentage of 

86.77 percent. These results are similar to those from analyses 

on water and invertebrates from those same sites. Time-series 

data for California roach at the Upper Bear Creek site and 

the Middle Bear Creek site demonstrated MMeHg peaks in 

summer June/July/August time periods for both 2000 and 

2001.

Schwarzbach and others (2001) also collected California 

roach from Bear Creek sites upstream and downstream of 

the confluence with Sulphur Creek in April and August 1997. 

Results were consistent with Slotton and others (2004), with 

TotHg concentrations in roach that were significantly higher 

downstream than upstream of the confluence with Sulphur 

Creek: 3 times greater in April (upstream = 0.1 ppm, ww; 

downstream = 0.3 ppm, ww) and 4 times greater in August 

(upstream = 0.4 ppm, ww; downstream = 1.7 ppm, ww). 

Sacramento pikeminnows (Ptychocheilus grandis) also were 

collected in Bear Creek in 1998 and exhibited the highest 

TotHg concentrations among all pikeminnows from a total of 

five sites throughout the entire Cache Creek watershed. 

Invertebrates

Beginning in April 1995, Slotton and others (1997) 

conducted a survey of TotHg in benthic stream invertebrates 

at 38 sites along Cache Creek and the Bear Creek 

watershed (figs. 5, 6, 7). This survey involved collections 

of several trophic levels of benthic invertebrates including: 

Ephemerellidae (mayflies – herbivores), Siphlonuridae 

(swimming mayflies – herbivores), Hydropsychidae (netspinning caddisflies – drift feeders), Pteronarcidae (giant 

stoneflies – leaf shredders), Perlodidae (yellow sally 

stoneflies – small predators), Calopterygidae (damselflies 

– small predators), Sialidae (alderflies – small predators), 

Naucoridae (creeping water bugs – large predators), Aeshnidae 

(dragonflies – large predators), Libellulidae (dragonflies – 

large predators), Tipulidae (craneflies – large predators), and 

Corydalidae (hellgrammites – large predators). 

Slotton and others (1997) found significant variability 

in the concentration of TotHg in invertebrate tissues among 

different sites within the Cache Creek watershed with a range 

of about 0.05–22.74 μg/g (parts per million = ppm) dry weight 

(dw). The highest TotHg concentrations were obtained from 

Harley Gulch at Highway 20 (up to 22.74 ppm, dw) and 

within Sulphur Creek (up to 2.69 ppm, dw) (note asterisks 

in fig. 6). Upstream of the confluence with Sulphur Creek, 

Bear Creek invertebrate whole body TotHg typically ranged 

from about 0.1 to 0.2 ppm (dw) but as soon as Sulphur Creek 

waters enter Bear Creek, invertebrate TotHg concentrations 

increased to about 1.2–1.5 ppm (dw). However, one site about 

0.4 km upstream of the confluence of Sulphur Creek with 

Bear Creek had elevated invertebrate TotHg concentrations to 

about 0.3 ppm (dw). Although this increase appears to be real, 

it is from a single site at a single sampling point in time and 

its significance should not be over-interpreted. It is possible 

that there are additional Hg sources entering Bear Creek 

upstream of the confluence of Sulphur Creek, possibly from 

the Rathburn-Petray mine area described above. In addition, 

roadbed materials and/or grazing effects in Bear Valley 

could affect Hg concentrations upstream of Sulphur Creek. 

Schwarzbach and others (2001) also sampled invertebrates in 

Bear Creek near Hamilton Canyon, about 1.2 km upstream of 

the Slotton site, and found TotHg concentrations ranging from 

0.02 to 0.04 ppm, ww (about 0.10–0.20 ppm, dw).

Further studies conducted from 1999 to 2003 by 

Slotton and others (2004) provided additional Hg data 

from water, invertebrates, and fish (where present) at three 

sites in the Bear Creek watershed (in addition to other sites 

along the entire Cache Creek watershed). Their findings 

mirror the relative ranges of TotHg and MMeHg found in 

aqueous samples presented in section, “Physical/Chemical 

Assessments.” Average TotHg in aquatic insects ranged from 

ca. 19–41 ppb, wet weight (ww) at the Upper Bear Creek site 

to ca. 168–465 ppb (ww) at the Middle Bear Creek site, and 

ca. 416–1,987 ppb (ww) at the Sulphur Creek site. MMeHg 

ranged from ca. 18–33 ppb (ww) at the Upper Bear Creek site, 

to ca. 138–359 ppb (ww) at the Middle Bear Creek site, and 

ca. 139–290 ppb (ww) at Sulphur Creek. Interestingly, the 
Introduction

13

Figure 5. Cache Creek Watershed, including its two major tributaries, Bear Creek and North Fork Cache Creek; red dots indicate invertebrate mercury bioindicator sampling sites used by Slotton and others during 1995–97 (Slotton and others, 1997).
14 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Figure 6.

Mercury concentrations (ppm dw) in stream invertebrates from the Cache Creek watershed in 1995. (Slotton and others, 1997).
Introduction 15

Figure 7. Mercury (ppm dw) in stream invertebrates in the Bear Creek Watershed from D. Slotton collections in 

1996 (Slotton and others, 1997).
16 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

maximum concentrations at the Sulphur Creek site (no 

fish data available for Sulphur Creek), and intermediate 

concentrations downstream of the confluence of Sulphur 

Creek with Bear Creek at the Middle Bear Creek site. In 

addition, the ratio of MMeHg:TotHg (or the percentage of 

TotHg as MMeHg) exhibited similar trends for all matrices; 

i.e., the highest ratios were found at the Upper Bear Creek 

site, the lowest ratios at Sulphur Creek, and intermediate ratios 

at the Middle Bear Creek site (Slotton and others, 2004). As 

discussed above, because no known mining sites are located 

near or upstream of the Upper Bear Creek site, the Hg source 

influencing this site is most likely a geothermal spring, 

which could easily be undetected if it lies within the stream 

bed itself. In addition, evidence from the Sulphur Creek site 

suggests that the Hg source for that site is dominated by Hg 

mines. 

Several recent publications provide evidence that 

cinnabar and/or metacinnabar, the primary forms of Hg at 

Hg mines, are more refractory and less bioavailable than 

other forms of Hg (Kim and others, 2000; Bloom and others, 

2003; Suchanek and others, 2008b). Thus, the Bear Creek 

watershed data also are consistent with an interpretation that 

Hg derived from springs is more bioavailable and thus could 

be bioaccumulated more efficiently than Hg derived from 

mining sources. That is, despite the absolute concentrations of 

Hg, if the primary source of Hg at the Upper Bear Creek site 

is primarily from springs (see discussions above for water), 

and the primary source of Hg at Sulphur Creek was mining 

(see discussion above for water), this would result in a higher 

proportion of MMeHg to TotHg in water, invertebrates and 

fish from the Upper Bear Creek site (which may derive its 

Hg from springs), a lower proportion of MMeHg:TotHg at 

the Sulphur Creek site (which is dominated by mines), and 

an intermediate proportion at the Middle Bear Creek site 

(which has a mixture of the two types of sources). Therefore, 

sites having geothermal or saline spring influence may play 

a relatively more significant role in producing MMeHg than 

those sites dominated by mines. For example, even though 

the absolute amount of TotHg at the Upper Bear Creek site is 

quite low, the amount of MMeHg produced per unit of TotHg 

is greatest at this site (likely dominated by Hg sources from 

springs) and lowest at Sulphur Creek (likely dominated by Hg 

sources from mining). 

Domagalski and others (2004) estimated annual TotHg 

loading from Bear Creek as well as Sulphur Creek in relation 

to their contributions to the water discharges of the Cache 

Creek watershed and the Yolo Bypass for water years 2000 

and 2001. Interestingly, Sulphur Creek contributed ca. 1.8–2.8 

× 106 m3 annually (representing ca. 1–2 percent of the total 

flow of downstream Cache Creek) and Bear Creek contributed 

ca. 18.3–33.3 × 106 m3 annually (representing ca. 12–19 

percent of the total flow of downstream Cache Creek). 

Amphibians and Birds

Schwarzbach and others (2001) collected six foothill 

yellow-legged frogs (Rana boylii) from Bear Creek, three 

from upstream, three from downstream of the Sulphur Creek 

confluence, and a sample from Bear Creek at Highway 20. The 

range of TotHg concentrations was 0.075–0.538 ppm (ww) 

with the highest value obtained at the Highway 20 site. The 

mean TotHg was 0.11 ppm (ww) in upstream samples and 

0.31 ppm (ww) in downstream samples. 

At 22 sites in 1997 and 19 sites in 1998, Hothem and 

others (in press) collected and analyzed TotHg in three 

anuran species from the Cache Creek watershed, including 

sites within the Bear Creek watershed: American bullfrogs 

(Lithobates catesbeianus), Foothill yellow-legged frogs 

(Rana boylii), and Pacific chorus frogs (Pseudacris sierra). 

Results were comparable between 1997 and 1998. Pooling 

data for both years, the geometric means for TotHg in Bear 

Creek bullfrogs in 1997 were 0.032–0.390 ppm (ww) at sites 

upstream of Sulphur Creek and 0.423–0.561 ppm (ww) for 

sites downstream of Sulphur Creek. For yellow-legged frogs, 

the pattern was similar; using pooled data, TotHg ranged from 

0.082 to 0.159 ppm (ww) upstream of the confluence with 

Sulphur Creek, and 0.328 to 0.846 ppm (ww) downstream of 

Sulphur Creek. Similarly, Pacific chorus frog TotHg at a single 

site was 0.166 ppm (ww) upstream of Sulphur Creek and 

0.258 ppm (ww) downstream of Sulphur Creek.

In 1997, Schwarzbach and others (2001) collected and 

analyzed three killdeer (Charadrius vociferus) eggs from 

separate nests in the Bear Creek region. One from a nest near 

the Jones Fountain of Life yielded a TotHg concentration 

of 0.26 ppm (ww) and eggs from two nests on Bear Creek 

ca. 0.4 km downstream of the Sulphur Creek confluence 

contained TotHg concentrations of 0.10 and 0.90 ppm (ww). 

Hothem and others (2008) reported TotHg concentrations 

from cliff swallow (Petrochelidon pyrrhonota) eggs and 

nestling carcasses collected in 1997 and 1998 from nests 

within the Cache Creek watershed, including the Bear Creek 

watershed. Geometric mean values for egg TotHg ranged 

from 0.02 to 0.04 ppm (ww) at a reference site upstream of 

the confluence of Sulphur Creek, 0.097–0.208 ppm (ww) 

at Sulphur Creek sites and 0.070–0.118 ppm (ww) for Bear 

Creek sites downstream of the Sulphur Creek confluence. 

TotHg geometric means in nestling carcasses ranged from 

0.047 ppm (ww) at a reference site upstream of the Sulphur 

Creek confluence to 0.116 ppm (ww) downstream of the 

confluence. 

Overview of Historical Data

Cumulative data for water, invertebrates, and fish 

matrices show similar patterns of minimum TotHg and 

MMeHg concentrations at the Upper Bear Creek site, 
Mercury Monitoring Protocol 17

Mercury Monitoring Protocol

Objective

The objective of this monitoring protocol is to document 

procedures for collecting and processing water, sediment, and 

biota for estimation of TotHg and MMeHg in the Bear Creek 

watershed. Concurrent sampling of TotHg and MMeHg in 

biota as well as water and sediment will provide information 

regarding the relative bioavailability of Hg released from Hg 

sources in the watershed and identify environments of Hg 

methylation. 

These protocols will assist landowners, land managers, 

water-quality regulators, and scientists in detecting whether 

restoration/mitigation actions lead to significant progress 

toward achieving water-quality goals to reduce Hg in Bear 

Creek and Sulphur Creek to meet total maximum daily load 

(TMDL) standards established by the Central Valley Regional 

Water Quality Control Board (CVRWQCB).

Sampling

Water, sediment, and biota should be sampled for a 

minimum of 2 years before any remediation activity (Pre-R). 

Because of potentially significant inter-annual variability in 

precipitation events that drive erosion of Hg-laden sediments 

from the landscape into streams within the Bear Creek 

watershed, as well as inter-annual variability in the production 

of Hg-charged fluids from cold or hot springs, at least 2 

years is needed to provide adequate baseline data prior to the 

commencement of remediation. However, if time and funding 

allow, it would be advantageous to collect Pre-R data for 

up to 5 years to confirm trends from past studies. Following 

remediation, water, sediment, and biota should be sampled 

for several years (Post-R). For a number of reasons, including 

the physical disturbance effects of remediation, reduction in 

loading, the inter-annual variability factors discussed above, 

and changes in bioaccumulation rates, it is unknown how long 

it will be before potential changes in Hg concentrations occur 

in abiotic and biotic matrices after remediation. Therefore, 

the length of time needed to detect changes will most likely 

be longer than the time needed to establish a reliable baseline. 

Up to 10 years of Post-R sampling would be desirable to 

account for those factors identified above. At a minimum, 

Post-R sampling should be conducted for 5 years. If additional 

remediation projects are begun or completed during the initial 

Post-R phase, then the sampling plan should revert to the 

Pre-R protocol, which would start a new Post-R phase after 

that subsequent remediation was completed. 

Seasonality

Summer (June-August) appears to be the season when 

MMeHg production reaches a maximum at all sites within 

the Bear Creek watershed as evidenced by data from water, 

sediments, invertebrates, and fish. During Pre-R monitoring, 

water, sediment, and biota should be sampled in the spring 

and late summer/early fall and analyzed for TotHg and 

MMeHg. This would serve two purposes. First, it would 

provide information to assess what concentrations of TotHg 

and MMeHg are present in spring during typical breeding 

periods for many wildlife species. Second, it would confirm 

that late summer/early fall is a period of higher MMeHg 

concentrations (based on data from previous studies). If these 

results are consistent with data from previous studies, water, 

sediment, and biota could be collected only in the late summer 

in subsequent Pre-R years. During Post-R years for any of the 

mine sites in the watershed, water, sediment, and biota should 

be collected in the spring and late summer and analyzed for 

both TotHg and MMeHg. If these results also are consistent 

with previous data, water, sediment, and biota could be 

collected only in the late summer. 

Physical/Chemical Matrices

Based on existing data, and the need to identify potential 

changes in Hg loading and bioaccumulation in relation to 

remediation actions, we recommend 10 monitoring sites for 

physical and biological matrices (table 1). Sampling sites 

for the Bear Creek watershed were selected based on spatial 

relationships to known natural and anthropogenic sources of 

Hg within the watershed (fig. 1), and the justification for each 

sample site is provided in table 1. The frequency and timing 

of water and sediment sampling should be coordinated with 

biota sampling. All samples at a site should be collected on the 

same day during similar flow and weather conditions in the 

watershed. It is important that water and sediment samples be 

collected prior to biota sampling to avoid any changes in water 

quality that arise from disturbance of the stream during biota 

collection. Water should be collected for analysis of TotHg and 

MMeHg in unfiltered and filtered samples. Sediment should 

be collected for analysis of TotHg and MMeHg. The unfiltered 

water sample provides information on TotHg and MMeHg 

associated with particulate phases in the water. The filtered 

water sample provides information on Hg that is dissolved 

as well as TotHg and MMeHg present in colloids that pass 

through a 0.45-μm filter (Babiarz and others, 2001). Based 

on results from historical studies (see above), late summer to 

early fall appears to be the season during which maximum 

MMeHg concentrations were observed in water, sediments, 

and biota. Because this is the most critical toxic form of Hg 
18 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

For collection of filtered water for TotHg and MMeHg 

analysis, water should be filtered in the field during collection 

through an in-line 0.45-mm filter pack assembly using a 

peristaltic pump with Teflon® tubing. Water samples should 

be acidified with ultra-pure 6N HCl in the field immediately 

after collection (Olson and DeWild, 1999). Samples should be 

kept on ice until shipped. Water samples should be shipped on 

wet ice to arrive the next morning at the analytical facilities at 

temperatures ranging from 1 to 4oC; the temperature specified 

by EPA Method 1631E to minimize biologically induced 

phase changes and MMeHg degradation.

Water-quality parameters, including pH, conductivity, 

dissolved oxygen, oxidation-reduction potential (ORP), and 

temperature should be measured in the field with a multi-probe 

metering device (for example, HydrolabTM or YSITM), either 

by placing the probe directly into the flowing stream water, 

or by filling a container and placing it over the probe. If a 

multi-probe metering device is not available, the following 

meters are recommended. Values of pH should be determined 

using an Orion 250A or 290A meter with a gel-filled Triode 

electrode, or comparable pH meter. The pH meter should be 

calibrated using commercially available pH 10 and 7 buffers. 

Water temperatures should be measured using a thermometer 

that was checked against a thermometer certified by the 

American Society for Testing and Materials (ASTM). Specific 

conductance should be determined using a Cole-Parmer 

meter and probe; a one-point calibration should be done with 

a standard in the approximate concentration range of the 

unknown water. Dissolved oxygen should be determined using 

a YSI meter and probe, calibrated using standard procedures.

High-Flow Event Sampling

The high flow sampling should be carried out using 

the same protocol used for the seasonal sampling of water 

under low flow conditions. The high flow sampling should 

be carried out by starting at the upper part of the watershed 

and continuing to the lowest sample site in the watershed. 

It is important to collect the sample in the centroid of the 

stream, if possible. In order to assess Hg release from the 

Rathburn and Petray mines, Tributary 1 (fig. 4) should be 

sampled just above its intersection with the Bear Fault. Water 

samples should be collected for analysis of Hg and MMeHg 

on unfiltered and filtered fraction. It is not necessary to collect 

a sediment sample. Automated samplers should be considered 

for use because staff may not always be available for a highflow event. The automated ISCO samplers with Teflon®

tubing are suggested for use to collect stream water samples. 

Control studies using ISCO samplers have documented only 

minimal contamination of sample bottles from dry deposition 

of airborne Hg, and reliable results are obtained when Hg 

concentrations are greater than 5 pptr (Riscassi and others, 

2008).

in this system, it is advisable that samples be collected during 

periods when MMeHg is highest in order to relate results to 

impacts on biota. However, it is possible that different weather 

or precipitation conditions from those that were present during 

the previous studies could alter the seasonality of maximum 

MMeHg. Therefore, some sampling at other seasons (for 

example, spring) should be incorporated into this monitoring 

plan. In order to relate physical/chemical data to biological 

data, it is recommended that for the first 2 years, physical/

chemical data be collected in both spring and late summer/

early fall.

Episodic transport of TotHg and MMeHg caused by high 

flows associated with storm events has been documented for 

several watersheds (Babiarz and others, 1998; Hurley and 

others, 1998; Scherbatskoy and others, 1998; Balogh and 

others, 2005). Because most Hg transported in a watershed 

moves during a few high-flow events, it also is necessary to 

sample Bear Creek during at least one high-flow event per 

year in order to monitor Hg dynamics in stream water and 

assess Hg transport from both natural and mine impacted 

areas. 

Sample Collection and Processing

Water and sediment should be collected using standard 

ultra-clean sampling and handling protocols (Gill and Bruland, 

1990; Bloom, 1995) to avoid introduction of extraneous Hg 

into the samples. Sampling personnel must wear clean, nonpowdered gloves made of polyethylene, latex, or PVC at all 

times when handling sampling equipment and containers, 

and gloves must be changed between sample collection sites. 

At a sampling site, one person is designated as “dirty hands” 

and the other person who is sampling is designated as “clean 

hands.” The dirty hands person is responsible for all activities 

that do not involve direct handling of the sample and its 

container, such as handling all other potentially contaminated 

equipment, clothing, etc. The clean hands sampler is involved 

in direct handling of the sample container and transfer of the 

sample from the collection device to the sample container. 

Water samples should be collected in ultra clean 250-mL 

FLPE bottles. A Hg laboratory with experience in low level 

Hg analysis, such as the USGS Mercury Research Laboratory 

or Brooks Rand LLC, should provide sample bottles. Grab 

samples should normally be taken near the centroid of flow, 

but during high flow, grab samples may occasionally be taken 

from the streambank. Care should be taken not to disturb bed 

sediments before sample collection. For TotHg, the sample 

container should be filled partially and emptied 3 times prior 

to its final filling. The sample container should then be rapidly 

submersed, filled, and capped while submersed so that there 

is no headspace of air. Water samples for Hg and MMeHg 

analysis should be preserved with 2 mL of Ultrex 6N HCl 

(Olson and DeWild, 1999).
Mercury Monitoring Protocol 19

Sediment Collection and Analysis

Sediment samples collected using polycarbonate or 

Teflon® spoons should be transferred to Teflon® or acidwashed glass containers, following the protocols described 

by Shelton and Capel (1994). Representative sediment should 

be collected from in-stream sites, avoiding large pebbles, 

twigs, and roots. Excess water should be decanted from the 

container. Sediment samples collected for analysis of TotHg 

and MMeHg are frozen with dry ice immediately after 

collection, kept frozen, and shipped on dry ice so that they are 

received frozen at the analytical facility, and kept frozen there 

until analysis is initiated. Sediment should be analyzed for 

TotHg and MMeHg, and percent solids should be determined 

for reporting results on a dry weight basis. Analysis of TotHg 

should be carried out according to USEPA method 1631 and 

for MMeHg according to USEPA method 1630.

Biological Matrices

This protocol documents procedures for collecting and 

processing aquatic invertebrates and fish for estimation of 

Hg bioaccumulation in biota of the Bear Creek watershed. 

Concurrent sampling for Hg and MMeHg in water and 

sediments will provide information regarding sources and 

relative bioavailability of Hg at each site (see above). 

Clean techniques are essential to minimize potential 

contamination, including contact with personnel and 

equipment. The field methods described here are based on 

guidelines developed in conjunction with the National WaterQuality Assessment (NAWQA) Program, the Toxic Substances 

Hydrology Programs, and researchers from other disciplines in 

the USGS as described by Scudder and others (2008).

This protocol will focus on smaller individuals of upper 

trophic level predatory fish whenever possible. Smaller fish 

provide a more short-term representation of changes in Hg 

concentrations of prey species, which in turn will reflect 

short-term changes in Hg in water and sediments. Fish of 

lower trophic levels also will be collected, especially where 

predominantly piscivorous species are not available. TotHg 

and MMeHg will be analyzed in individual samples of fish 

during the first year of the study to confirm the percentage of 

TotHg as MMeHg. If, as predicted, the percentage of TotHg 

that is MMeHg is high (> 95 percent) in the sampled fish, 

then it will be sufficient in subsequent years to analyze the 

fish samples only for TotHg. Once the percentage of TotHg 

as MMeHg is established for those species, only TotHg need 

to be analyzed. Previous studies have shown that about 95 

percent of TotHg in fish muscle tissue is MMeHg (Huckabee 

and others, 1979; Bloom, 1992; Wiener and Spry, 1996). 

Skin-off fillets of top-level piscivores will be analyzed as well 

as individual whole bodies (less gut contents) of the lower 

trophic level fish. In addition, because the cost for MMeHg 

analyses greatly exceeds that for TotHg, the elimination of 

the redundant MMeHg analyses will provide significant 

cost savings yet will not compromise the validity of the 

data. Whole-body aquatic invertebrates, however, should be 

analyzed for both TotHg and MMeHg every year, because 

the ratios of MMeHg to TotHg tend to be inconsistent among 

invertebrate taxa, among sites and among years (Mason and 

others, 2000; Haines and others, 2003; Wiener and others, 

2007). 

Biological samples, as well as water and sediment 

samples, need to be collected concurrently or within a short 

time period (2 weeks for invertebrates and forage fish; 4 weeks 

for top predator fish) to minimize changes that might affect 

relative Hg concentrations in the various media collected. 

In addition, sample collection of each media type must be 

coordinated to minimize risk of site disturbance and sample 

contamination. For example, collection of biota at a given site 

before water and sediment will likely stir up bottom sediment 

producing conditions unsuitable for water and sediment 

sampling. At each site, a typical order for sample collection 

would be water, followed by sediment, then invertebrates, 

and finally fish. Protocols for sample collection, processing, 

labeling, submission, and for data management should be 

reviewed by all field personnel prior to sample collection. See 

Scudder and others (2008) for examples. 

Sampling sites for the Bear Creek watershed (table 1, 

fig. 1) were selected based on spatial relationships (both above 

and below) to known natural and anthropogenic sources of 

Hg within the watershed and potential for bioaccumulation 

by biota. In addition, data from previous biological studies 

(Schwarzbach and others, 2001; Slotton and others, 2004; 

Hothem and others, in press) were used to assess the most 

useful sites for a comprehensive, but cost-effective, evaluation 

of Hg bioaccumulation before and after restoration.

Invertebrates

The target macroinvertebrates for this study (Merritt and 

Cummins, 1995) should be predatory or filter-feeding insects, 

depending on their abundance and availability at sampling 

sites. Aquatic invertebrates can serve as excellent bioindicators 

of metals contamination (Cain and others, 1992). At least three 

key invertebrate taxa should be targeted for sampling (table 2). 

As feasible, these taxa should represent different functional 

feeding categories (for example, scrapers, shredders, grazers, 

collectors/gatherers) and should be taxa and sizes that are 

considered important prey items for target fish. For the 

lotic habitats (streams and flowing channels) of the Bear 

Creek watershed, preferred macroinvertebrates should be 

larval caddisflies (Trichoptera, family Hydropsychidae), 

larval dragonflies (Odonata, Anisoptera, family Gomphidae, 

Aeshnidae, and Libellulidae), and adult water striders 

(Hemiptera, family Gerridae). Invertebrates should be sorted 

to the lowest practical taxon, normally family, and processed 

as single-taxon composites. For example, one or more species 

of net-spinning caddisfly larvae in the family Hydropsychidae 

could form a composite sample. If available, water striders 

might represent a good sentinel species because they have 

been found to quantify accurately the food chain entry of Hg 
20 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

(Jardine and others, 2005). However, because of the high 

variability of Hg in water striders from aquatic systems, 

especially among different seasons, care should be taken to 

understand the potential short-term variations in Hg sources to 

the streams being sampled (see Jardine and others 2009). For 

all sites, alternate species of invertebrates should be identified 

and collected the first year to serve as functionally equivalent 

substitutes in case adequate numbers of primary target species 

are not available. Some invertebrates may be unsuitable 

because of their small size (for example, Chironomidae), 

because expending the time needed to collect sufficient mass 

for chemical analyses may not be feasible. 

Fish

Black basses (Centrarchidae: Micropterus spp.) are 

excellent target species (May and others, 2000), but previous 

studies have not been successful at collecting black bass 

from Bear Creek sites. Predatory fish can serve as good 

bioindicators of metals contamination in the long term 

(Wiener and Spry, 1996), but considering the ephemeral nature 

of the streams in parts of the Bear Creek watershed, other 

fish should be collected wherever possible. One or two other 

species will need to be identified that can be collected across 

all study sites. Species that have been collected in previous 

studies (Slotton and others, 1997, 2004; Schwarzbach and 

others, 2001) include Sacramento pikeminnow, Sacramento 

sucker, and California roach. The species of fish that is most 

likely to be collected is the California roach. The collection 

of the same species at each site allows for comparison among 

sites across geographical areas. The use of target taxa allows 

for comparison among sites across geographical areas. 

Alternate species will be identified for each site. Two species 

of forage fish and one species of top predator fish (table 3) will 

be collected, as available, at each site; these will be identified 

in the field to the lowest possible taxonomic category and 

processed individually. Alternate species of acceptable forage 

fish and top predator fish also will be identified for all sites 

to serve as functionally equivalent substitutes in the case that 

adequate numbers of primary target species are not collected. 

Preference should be given to fish that are resident 

(for example, speckled dace), avoiding whenever possible 

sampling areas where fish are likely to move in and out of 

other major water bodies. Although speckled dace represent 

a good potential target species because they have relatively 

limited movements, short lifespan, and are benthically 

dependent foragers (for example, on insects), they have not 

been documented during previous collections and may not 

be available. Collections during the first year will determine 

the size ranges available for various fish species and provide 

guidance for future collections. Depending on the fish species, 

all individuals used in composite samples should be as similar 

in size as possible. If large variation in fish size affects the 

variation in Hg concentrations, then size should be used as a 

covariate during statistical analysis to control for this effect 

while comparing differences in Hg concentrations among 

sites. 

Table 2. Available and Target* invertebrate families, Bear Creek.

Order Common name Family name Trophic level Previously sampled

Hemiptera Water strider adult* Gerridae Large predator No

Creeping water bug adult Naucoridae Large predator Slotton and others 1997, 2004

Odonata Dragonfly larvae* Gomphidae Large predator Schwarzbach and others 2001; Slotton and 

others 2004

Dragonfly larvae* Aeshnidae Large predator No

Dragonfly larvae* Libellulidae Large predator Slotton and others 1997, 2004; Schwarzbach 

and others 2001

Damselfly larvae Calopterygidae Small predator Slotton and others 1997; Schwarzbach and 

others 2001

Damselfly larvae* Coenagrionidae Small predator Slotton and others 2004

Trichoptera Net-spinning caddisfly larvae* Hydropsychidae Drift feeder Slotton and others 1997, 2004; Schwarzbach 

and others 2001

Ephemeroptera Mayfly larvae Ephemerellidae Herbivore Slotton and others 1997, 2004

Mayfly larvae Siphlonuridae Herbivore Slotton and others 1997

Diptera Cranefly larvae Tipulidae Large predator Slotton and others 1997, 2004

Soldier fly larvae Stratiomyidae Small predator Slotton and others 2004

Plecoptera Golden stonefly larvae Perlidae Small predator Slotton and others 1997

Megaloptera Alderfly larvae Sialidae Small predator Slotton and others 1997

Dobsonfly larvae Corydalidae Large predator Slotton and others 1997, 2004

Coleoptera Riffle beetle adult Elmidae Large predator Slotton and others 2004
Mercury Monitoring Protocol 21

Table 3. Available and Target* fish species, Bear Creek.

Family name Common name Scientific name Previously sampled

Centrarchidae Smallmouth bass Micropterus dolomieui No

Bluegill Lepomis macrochirus No

Green sunfish Lepomis cyanellus Slotton and others 2004

Cyprinidae Sacramento pikeminnow* Ptychocheilus grandis Slotton and others 2004; Schwarzbach and others 2001

California roach* Hesperoleucus symmetricus Slotton and others 2004; Schwarzbach and others 2001

Speckled dace* Rhinichthys osculus No

Catastomidae Sacramento sucker* Catostomus occidentalis Slotton and others 2004; Schwarzbach and others 2001

Table 4. Summary of invertebrate and fish sample collections for Bear Creek Watershed.

Collection Season

Pre Cleanup Post Cleanup

Year 1 Years 2-5 Year 1 Years 2-5

Fish Inverts Fish Inverts Fish Inverts Fish Inverts

Spring (April –May) 3 species 3 taxa 2 species 2 taxa

Fall (Sept. – Oct.) 3 species 3 taxa 3 species 3 taxa 2 species 2 taxa 3 species 3 taxa

Preparation for Collections

Suggested equipment and supplies for use in these 

biological studies are listed in tables 5 and 6. Equipment 

and supplies need to be properly prepared to minimize the 

potential for sample contamination. New, sealed supplies such 

as zip-seal plastic bags and plastic vials with plastic caps do 

not need pre-cleaning. However, reusable equipment will need 

to be cleaned properly prior to field collections (see Scudder 

and others, 2008, for techniques). After tools are cleaned, 

supplies and smaller equipment will be double-bagged in 

new plastic bags and stored in sealed containers to minimize 

contamination; supplies should be cleaned and packed 

separately for each site to minimize the need for field cleaning 

(Brumbaugh and others, 2001). In the field, all equipment 

should be cleaned between sites. 

Field forms (appendixes A and B), sample labels 

(appendix C), and laboratory submission forms (appendix D) 

should be prepared prior to collecting samples and should be 

printed on durable water-proof paper (for example, Rite-InThe-RainTM paper). Field forms and labels should be preprinted 

with station name, analyte, medium code, and contact 

information (name and telephone number). For other examples 

of such forms, see Scudder and others (2008).

California scientific collection permits, obtained from the 

California Department of Fish and Game, will be required for 

collections of invertebrates and fish. Landowner permissions 

will be required for access to sites located on private property 

or when site access requires crossing private property. 

A consultation with local U.S. Fish and Wildlife Service 

personnel with regard to Section 7 of the Endangered Species 

Act is recommended and may be required in some locations 

if a threatened or endangered species is known or thought 

to be present (http://www.fws.gov/endangered/factsheets/

consultations.pdf). 

Quality Assurance and Quality Control (QA/QC)

Quality-assurance samples are collected to investigate 

field and laboratory variability. Triplicate invertebrate 

composite samples (three individuals per species) and multiple 

individual fish samples in this protocol serve as qualityassurance replicates for field variability. Quality assurance for 

laboratory analyses will include duplicate runs, blanks, spikes, 

and use of certified or standard reference sample materials. 

Voucher specimens for taxonomic confirmation should be 

preserved in 70-percent ethanol (invertebrates). Techniques 

used for biota must ensure that tissues are not contaminated 

during collection or during sample processing. Processing on 

site or at a nearby outdoor area is acceptable if a stable, clean 

work area is available. However, processing in an enclosed 

facility, such as a field laboratory, is preferred. Disposable, 

powder-free latex gloves must be worn during all sample 

processing and must be changed frequently, particularly after 

touching any unclean surface.
22 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Table 5. Equipment checklist (modified from Scudder and others 

2008).

Carboys, 5-gallon (2, tap; 2, DI water; 1, 5 percent HCl)

Camera, digital

Chairs, folding

Clipboard(s)

Coolers (4, wet ice; 2, dry ice)

Collection permits/licenses

Detergent, phosphate-free

Field data sheets (pre-printed, water-resistant paper)

First aid kit

Flashlights and headlamps

Garbage bags, large, plastic

Global Positioning System (GPS) unit

Ice, dry (about 15 lb per day of trip)

Ice, “wet”

Insect repellent

Keys, identification, for invertebrates and fishes

Labels, (waterproof, pre-printed)

Maps 

Markers, fine-tip, alcohol/solvent resistant

Markers, wide-tip, alcohol/solvent resistant

Personal Flotation Device (1 per person)

Phone, cell

Phone numbers, emergency

Protocol (on water-resistant paper)

Sunscreen

Table, folding

Tape, flagging, biodegradable

Tape, clear, 2", for shipping

Tray or tub, large (for dishwashing/storage)

Trays, plastic, large (for cleaning equipment)

Trays, plastic, shallow (for weighing fish)

Towels, paper rolls

Trash bags, heavy duty

Waders/hip boots/wading boots

Wader repair kit

Wash Bottles (500 mL, Teflon®; 1, tap water; 1, DI water; 1, dilute 

HCl; 1,dilute detergent)

Water, high-purity DI

Water, tap

Table 6. Fish and invertebrate collection checklist (modified from 

Scudder and others, 2008).

Dipnets, D-frame 

Forceps, plastic

Counters, hand-held

Gloves, powder-free (nitrile, vinyl, other non-latex)

Vials with caps, plastic, 20 mL

Magnifying glass/hand lens/binocular microscope

Scale, top-loading, accuracy to 0.01 g

Trays, plastic, shallow (for picking invertebrates)

Batteries or fuel for electrofishing unit

Buckets, 10 gallon

Electrofishing unit (backpack)

Gloves, rubber, safety (insulated)

Polarizing sunglasses

Nets, large dipnet

Nets, small

Anesthetic CO2 tablets (e.g., AlkaSeltzer®)

Bags, plastic, heavy-weight, zipper-seal, 1 L

Bags, plastic, heavy-weight, zipper-seal, 4–8 L

Cutting sheet/mat/board, plastic

Knives, filleting

Measuring board (non-metallic)

Pliers, needle-nose

Rulers, 6-inch plastic

Scale, top-loading, accuracy to 0.01 g

Scale, top-loading, accuracy to 0.1 g

Scale, hanging or hook, for large fish

Scalpels, high-grade stainless

Scissors, dissecting, high-grade stainless steel

Vials with caps, plastic, 20 mL

Sample Collection and Processing

A summary of invertebrate and fish sample-collection 

plans is provided in table 4. Additional taxa may be collected 

as available. 

Invertebrates

Invertebrate sampling should be conducted in spring 

(April–May) and late summer/fall (August–September) in 

the first year of the study (Pre-R) and the first year after 

restoration (Post-R) to evaluate inter-seasonal differences in 

Hg bioaccumulation. Invertebrates may be collected using dip 

nets, kick nets, or by hand and placed into labeled plastic bags 

or containers. At least 1 g wet weight of each taxon will be 

required to ensure sufficient biomass for analyses (minimum 

of 0.1 g dry weight) (Hall and others, 1998). Depending on the 

size of the invertebrates collected, the number of individuals 

needed to obtain 1 g wet weight will vary but, in any case, will 

likely not be less than three individuals. Small invertebrates, 

such as caddisfly larvae, may require up to 150 individuals. 

Collectors should attempt to be consistent with selection of 

species and size classes within a species and should collect 

each taxon from as broad a range of locations within a reach as 

possible. Because the number of individuals comprising each 

sample will drive Hg concentration variability, that number 

should be held constant for each species being sampled. This 

will be determined most easily after the first Pre-Remediation 

sampling and attempts should be made to maintain this 

number of individuals throughout the study period.

Invertebrates should be processed as three replicate 

composite samples for each taxon with the same number of 

organisms, of similar size, in each composite sample. Holding 

(depuration) times should typically range from a few to 24 

hours per site, depending on the time of day collected. Voucher 

specimens of each uncertain taxon should be collected for 

taxonomic confirmation. Within 24 hours, individuals should 

be sorted by family and placed in disposable dishes using 

Teflon®-coated forceps or gloved-hand. Organisms are to be 
Mercury Monitoring Protocol 23

rinsed clean with DI water and patted dry with a clean paper 

towel. Composite samples, consisting of 3–150 individuals of 

the same family (1–5 g, wet weight), should then be weighed 

and placed into chemically cleaned glass jars with Teflon®-

lined lids. Processed samples should then be preserved 

immediately on dry ice for transport to a freezer where they 

should be kept frozen for no more than 6 months until they 

can be shipped to an approved analytical laboratory. Each 

composite sample of invertebrates should be analyzed for 

TotHg and MMeHg. If sample mass is insufficient for both 

analyses, the priority analysis is MMeHg. 

Fish

A variety of fish-collection procedures may be 

appropriate, depending on site conditions and target species. 

General fish-collection procedures are described elsewhere 

(Meador and others, 1993; Moulton and others, 2002), and 

rely primarily on backpack electrofishing. Other methods in 

combination with or in lieu of electrofishing, such as seining 

or rod-and-reel (with artificial lures), or passive gear, such as 

traps or nets, may be more effective at some sites and would 

be acceptable. After capture, fish should be placed in a large 

plastic bucket in native water until they can be processed. 

Guidelines for live specimen handling and care are provided 

in Walsh and Meador (1998). Fish can be anesthetized using 

carbon dioxide from carbon dioxide-producing tablets, such 

as Alka-Seltzer® (2–4 tablets per gallon of water in bucket or 

other container). After anesthetization, fish can be euthanized 

by additional carbon dioxide (recommended by Walsh and 

Meador, 1998). Fish that are to be processed in a location 

other than the collection site should be placed in clean zip-seal 

bags on wet ice and processed within 24 hours. If they cannot 

be processed within 24 hours, they should be placed in a 

cooler on dry ice and kept frozen until they can be processed.

Fish should be collected both in spring and fall in the 

first Pre-R year of the study and in the first Post-R year of 

the study. Fish should be collected from the same sites as the 

water, invertebrates, and sediments and at each site, a typical 

order for sample collection would be water, followed by 

sediment, then invertebrates, and finally fish. Three species of 

fish, depending on availability, should be collected from each 

sample site. Initially, at each site, the goal is to collect five 

individuals of each species of similar size/length. Fish should 

be analyzed for MMeHg and TotHg during year 1 of Pre-R 

sampling and TotHg only during subsequent years. Detailed 

procedures for processing fish are provided by Scudder and 

others (2008). Upper trophic level fish are longer lived with 

slower tissue turnover than forage fish, so they do not have to 

be collected at exactly the same time as the rest of the biota. 

At each site, five similar-sized mature individuals of the target 

top predator fish should be collected. This size should be 

maintained throughout the entire study. A second species of 

piscivorous fish may be sampled if availability of the primary 

target species is limited. 

As indicated earlier, additional species of fish should 

be collected wherever possible. The preferred fish that may 

be collected is the California roach. Captured fish should be 

held in native water in buckets until they are weighed and 

measured for standard and total length. Each individual fish 

should then be labeled, placed in a polyethylene zip-lock bag 

and placed on dry ice in a cooler. Samples should be stored 

frozen until they can be processed further. Each fish should 

be measured for standard and total length (± 0.5 mm) and 

the total mass determined (± 0.5 g) on an electronic balance. 

In the laboratory, fillet tissue from top-level predatory fish 

should be dissected following procedures described by May 

and others (2000). The whole bodies of small fish should be 

analyzed, after removing the contents of the gastrointestinal 

tract. Tissues to be analyzed should be placed in chemically 

clean glass sample jars with Teflon®-lined lids. Processed 

samples should be preserved immediately on dry ice for 

transport to a freezer or analytical laboratory and analyzed as 

soon as possible, preferably within 6 months. 

Sample Shipment

Before they are shipped to the contract laboratory for 

analysis, all sample data must be entered on the appropriate 

sample chain-of-custody forms (included in the packaging), 

and the analytical laboratory must be notified to ensure that 

they can receive the samples. Ship frozen samples for next-day 

delivery (for example, FedEx®); ensuring that sufficient dry 

ice is included in the packaging so that a 1-day delay will not 

adversely affect the samples. Samples should be shipped on 

Mondays or Tuesdays to minimize the likelihood of a shipping 

error causing samples to sit over a weekend.

Statistics

TotHg and MMeHg concentrations in all matrices 

(water, sediments, invertebrates, fish) should be compared 

statistically among sites and sampling periods. However, 

limited availability of certain invertebrate and fish taxa at 

some sites and limited replication may restrict the statistical 

power (probability) for detecting certain effects, in which case 

only qualitative inter-site comparisons may be useful. Using 

the scientific literature, potential impacts of the observed Hg 

concentrations on the reproduction, growth, metamorphosis, 

and survival of each taxon should also be evaluated for 

the species sampled as well as other species present in the 

watershed. 

To avoid biasing the results of the data analysis towards 

spurious patterns in the data, any hypotheses intended for 

statistical evaluation should be developed a priori. For 
24 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

example, if a simple assessment of change is desired, the 

following hypotheses can be developed a priori:

Ho = Hg concentrations for [matrix] during Post-R year(s) are

 statistically indistinguishable from Hg concentrations

 during Pre-R year(s).

H1 = Hg concentrations for [matrix] during Post-R year(s) are

 statistically different from Hg concentrations during

 Pre-R year(s).

If desired, these hypotheses can be generalized to include 

trends characterized by relatively sustained rates of continuous 

change. For example, if an analysis of change in Post-R years 

is desired, then a linear regression model could be applied and 

the following hypotheses developed:

Ho = The slope of Hg concentrations for [matrix] over time

 during Post-R year(s) is statistically indistinguishable

 from zero (that is, no change). 

H1 = The slope of Hg concentrations for [matrix] over time

 during Post-R year(s) is statistically different than zero

 (that is, indicating an increase or decrease in Hg

 concentration).

These hypotheses can be evaluated with a wide variety 

of models ranging from simple (that is, two-sample t-test 

and simple linear regression) to complex models depending 

on additional sampling factors that could influence and/or 

confound with the variation in Hg concentrations (such as site, 

year, flow events, and fish size). However, complex models 

require larger sample sizes in order to reliably identify and 

account for the effects associated with these additional factors. 

Therefore, even if additional factors were to confound with the 

difference between Pre-R and Post-R periods, small sample 

sizes could reduce the ability to account for them and this 

would diminish the interpretability of the comparisons. 

Because the extent to which variation in the data exists 

cannot be determined a priori, then the specific appropriate 

statistical model might best be chosen after patterns in the 

initial data are evaluated. At a minimum, a one-factor ANOVA 

model comparing Pre-R and Post-R, essentially equivalent to 

a two-sample t-test, can evaluate the first hypothesis. Ideally, 

a Repeated Measures ANOVA model (for example, based on 

a linear mixed effects model structure) should be considered, 

using data from multiple sites as the repeated measures 

sampling units at each time interval. Covariates such as site, 

timing, water flow, and fish size should be added to this type 

of model as appropriate. With sufficiently abundant data, 

multiple comparisons tests (for example, Student-NewmanKeuls or Tukey’s Multiple Comparisons Tests) can be used to 

compare Pre-R data from 1 or 2 years versus multiple years of 

Post-R data. This would allow statistical comparisons between 

and among individual year pairs as well as groupings of years. 

However, if data are limited, this approach would have less 

statistical power than using simpler and more direct statistical 

approaches. 

Because inter-annual variability, especially with respect 

to the frequency and intensity of rainfall events (and possibly 

the flow rates of Hg-laden cold or hot springs), may result in 

significant variations in the Hg concentrations observed in 

water, sediments, and biota, it is important to assess multiple 

years for both the Pre-R Phase and Post-R Phase. In order 

to account for inter-annual variability in rainfall and natural 

spring activity, it is recommended that 5 years of Pre-R and 

5–10 years of Post-R sampling should be conducted, with 

preference for more years when possible. 

Power and Sample Size for Testing Percent 

Difference Between Pre-R and Post-R Periods

Because the statistical power to detect changes in 

Hg concentrations for all matrices would be enhanced by 

increasing sample sizes, to offset the uncertainty associated 

with sampling variation, a formula for determining adequate 

sample size to detect specific levels of change is provided 

below. This formula only applies to situations where the twosample t-test will be used. A comparable formula for repeated 

measures ANOVA would require more variables and is 

difficult to express without yet having a detailed specification 

for that model. Therefore, this formula is intended only to 

provide a starting point for determining the adequacy of the 

protocol sample size after initial data has been gathered. 

The minimum sample size for detecting an effect is 

determined by sampling variation (to be determined from 

initial data), the anticipated size of the effect, and the 

desired power for detecting that effect. A percent change in 

Hg between two periods can be expressed as an absolute 

difference in log-transformed Hg. For example, a hypothetical 

change of D percent (for example, D = –15 would be a 

15 percent decrease) in Hg concentrations from Pre-R to 

Post-R treatment periods is equivalent to having the ratio of 

Hg from Post-R to Pre-R treatment periods = 1+(D/100) (for 

example, 0.85). Equivalently, this ratio can be expressed in 

terms of the difference δ = ln(Post) – ln(Pre) = ln(Post/Pre) 

= ln(1+(D/100)), where ln(Post) and ln(Pre) correspond to 

average natural log transformed Hg levels from Post-R and 

Pre-R periods respectively. The minimum sample size (n) 

required for a minimum prescribed power (1-β) of detecting 

a D percent change sample when testing at the a significance 

level should satisfy the following inequality (Zar, 1999, 

section 8.4): 

n

s ≥ + (t t ) 2 2

2 1 1

2

δ α ν β ν ( ), ( ), ,

where s2 = within-period variance among samples (withinPre-R and within-Post-R sampling variance are assumed to 

be equal), v = 2(n-1), and t

α(1),v and tβ(1),v are 1-sided critical 

values of the t-distribution with v degrees of freedom and α 

and β tail probabilities respectively (Zar, 1999, table B.3 in 

appendix B). 
Mercury Monitoring Protocol 25

This formula can be rearranged algebraically to 

determine, for a given sample size, the power of detecting an 

effect, 1-β , where β satisfies the following inequality:

t

s n

t β ν α ν

δ

( ), 1 ( ), 2 1

2

≤ − .

Power, significance level, and sample size are interrelated 

such that by enhancing any one of the three, then one or both 

of the other two will be compromised. For example, an effect 

detected at the α=0.10 significance level would provide a 

reasonable confidence that a change occurred (as opposed to 

being a spurious statistical pattern). Comparatively, an effect 

detected at the α=0.05 significance level would provide even 

greater confidence that a change had occurred, but there will 

be either a reduced power of making that detection, or an 

increased sample size will be required to maintain the same 

power as for detecting effects at the α=0.10 significance level.

Analytical Costs

The estimated prices for chemical analyses of water, 

sediment, and biota are listed in table 7. The prices are based 

on a minimal sampling protocol, with 1 year of Pre-R and 5 

years of Post-R sampling. These prices do not include any 

potential volume discounts or other discounts that individual 

laboratories might offer. The cost of sampling at each site is 

based on analysis of TotHg and MMeHg on an unfiltered and 

filtered water sample and a sediment sample. The percent 

solids also will be determined for the sediment sample so 

that results can be reported on a dry weight basis. The highflow sampling event includes analysis of TotHg and MMeHg 

analysis on an unfiltered and filtered water sample at each site 

but no collection of a sediment sample. A summary of costs 

for water and sediment per site is presented in table 8.

The estimated costs for chemical analyses of biotic 

samples are shown in table 7. These prices do not include any 

potential volume or other discounts that individual laboratories 

might offer. The proposed numbers of samples of fish and 

invertebrates by year for 10 sites and the estimated costs 

per year are shown in table 9. The total numbers of samples 

collected in the first year are not all proposed for analysis 

(table 2). Instead, those collections are intended to determine 

the taxa available at each site. The taxa that are most common 

at the most sites are those that should be proposed for analyses 

in the first and subsequent years. Seasonal availability will be 

a factor to consider the first year as well. As discussed above, 

sampling during subsequent years should be performed during 

the season that is determined to be optimal for evaluation of 

Hg bioaccumulation in the Bear Creek watershed, likely the 

season with the highest production of MMeHg.

The total costs, with the assumptions listed in the 

footnotes, are presented in table 10. No cost of living or 

inflationary factors were included in the cost estimates. In 

addition, costs will vary depending on the availability of 

“in-kind” services, available equipment and supplies, and 

travel costs. 

Table 7. Estimated costs of individual analyses.1

Unit Unit Price

Water samples for total Hg analyses EA $80

Water samples for MMeHg analyses EA $160

Sediment samples for total Hg analyses EA $110

Sediment samples for MMeHg analyses EA $210

Sediment percent solids for result on dry 

weight basis

EA $10

Biological samples for total Hg analyses EA $110

Biological samples for MMeHg analyses EA $160

Determination of tissue percent solids EA $10

Sample homogenization EA $20

 Total cost for total Hg only $140

 Total cost for total and MMeHg $300

1 Based on November 2008 cost estimate from Brooks Rand Laboratories, 

Seattle, WA.

Table 8. Summary of water and sediment chemical analyses per site for Bear Creek watershed.

Collection Season

Pre Cleanup Post Cleanup

Year 1 Years 2-5 Year 1 Years 2-5

Water1 Sediment2 Water1 Sediment2 Water1 Sediment2 Water1 Sediment2

Spring (April –May) $530 $330 $530 $330

Fall (Sept. – Oct.) $530 $330 $530 $330 $530 $330 $530 $330

Cost per site per year $1,060 $660 $530 $330 $1,060 $660 $530 $330

Cost per 10 sites per year $10,600 $6,600 $5,300 $3,300 $10,600 $6,600 $5,300 $3,300

High flow 10 Sites per year $5,300 $5,300 $5,300 $5,300

Total cost per year $22,500 $13,900 $22,500 $13,900

Cost per period $22,500 $55,600 $22,500 $55,600

10-year total $156,200

1

 Based on one water sample per site per collection season for filtered and unfiltered Hg, and filtered and unfiltered MMeHg.

2

 Based on one sediment sample per site for Hg, MMeHg, and determination of percent solids for reporting concentration on a dry weight basis.
26 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Table 9. Summary of invertebrate and fish chemical analyses per site for Bear Creek watershed.

Collection Season

Pre Cleanup Post Cleanup

Year 1 Years 2-5 Year 1 Years 2-5

Fish1 Inverts2 Fish3 Inverts4 Fish3 Inverts4 Fish3 Inverts4

Spring (April –May) 15 9 10 6

Fall (Sept. – Oct.) 15 9 10 6 10 6 10 6

Cost per site per year $9,000 $5,400 $1,400 $1,800 $2,800 $3,600 $1,400 $1,800

Cost per 10 sites per year $90,000 $54,000 $14,000 $18,000 $28,000 $36,000 $14,000 $18,000

Total cost per year $144,000 $32,000 $64,000 $32,000

Cost per period $144,000 $128,000 $64,000 $128,000

10-year total $464,000

1

 Based on three species and five individual fish samples per species (whole bodies for forage fish; fillets for larger piscivorous species) per site; analyze for 

TotHg and MMeHg in years 1 and TotHg in subsequent years.

2

 Based on three composite samples of each analyzed taxon per site; all samples analyzed for both TotHg and MMeHg.

3

 Based on two species and five individual fish samples per species (whole bodies for forage fish; fillets for larger piscivorous species) per site; analyze for 

TotHg only. 

4

 Based on two composite samples of each analyzed taxon per site; all samples analyzed for both TotHg and MMeHg.

Table 10. Estimated budget (in dollars) for Federal staff to conduct Bear Creek sampling based on 2009 costs.

Year 1 

Pre

Year 2-5 Pre 

(per yr)

Year 1 

Post

Year 2-5 Post 

(per yr) Total

Personnel:

Biologist1 6,140 3,070 6,140 3,070

Geologist1 6,140 3,070 6,140 3,070

2 technicians2 5,590 2,795 5,590 2,795

Travel 1,800 900 1,800 900

Equipment3 11,000 1,000 1,000 1,000

Supplies4 2,000 1,000 1,000 1,000

Shipping Costs5 400 200 400 200

Chemical Analyses: biota 144,000 32,000 64,000 32,000

Chemical Analyses: water & sediments 22,500 13,900 22,500 13,900

Per year totals 199,570 57,935 108,570 57,935

10-year totals 199,570 231,740 108,570 231,740 $771,620

1

 Based on research scientist (GS-12/1) for 4 weeks/year in first years and 2 weeks/year in 2-5 years (not adjusted for COLA).

2

 Based on technician (GS-5/1) for 4 weeks/year in first years and 2 weeks/year in 2-5 years (not adjusted for COLA).

3

 Includes one-time purchase of backpack electrofisher: $10,000. Delete if one can be borrowed.

4

 Includes chemically cleaned jars, dissecting equipment, nets, waders, etc.

5

 Based on one 40-lb cooler per season FedEx priority overnight to contract laboratory.
References Cited 27

Summary and Conclusions

This report summarizes the known information on 

the occurrence and distribution of Hg in physical/chemical 

and biological matrices within the Bear Creek watershed. 

Multiple sources, both natural and anthropogenic, contribute 

Hg to Bear Creek and its tributaries. Previous studies of 

water, sediment, and biota from the Bear Creek watershed 

have revealed elevated concentrations of Hg and MMeHg 

in certain areas, notably downstream of sources of Hg that 

include both abandoned Hg and Au mines, ore-processing 

facilities or natural hot and cold springs. Cumulative data for 

water, invertebrates, and fish matrices show similar patterns of 

minimum TotHg and MMeHg concentrations at Upper Bear 

Creek sites, maximum concentrations in the Sulphur Creek 

area, and intermediate concentrations downstream of the 

confluence of Sulphur Creek with Bear Creek. 

Based on these data, a matrix-specific monitoring 

protocol was developed to be used for the evaluation of the 

effectiveness of Hg remediation activities in the Bear Creek 

watershed. The monitoring protocol documents procedures 

for collecting and processing samples of water, sediment, and 

biota for estimation of TotHg and MMeHg in the Bear Creek 

watershed. It also details timing of sampling before and after 

site remediation and techniques for evaluation of the results 

of chemical analyses and estimated costs for sampling and 

analyses.

The concurrent sampling of TotHg and MMeHg in 

biota as well as water and sediment from 10 monitoring sites 

will provide information on the relative bioavailability of 

Hg released from Hg sources in the watershed and identify 

environments conducive to Hg methylation. The sampling 

sites were selected based on spatial relationships (both above 

and below) to known natural and anthropogenic sources of 

Hg within the watershed and potential for bioaccumulation by 

biota. In addition, data from previous biological studies were 

used to assess the most useful sites for a comprehensive, but 

cost-effective, evaluation of Hg bioaccumulation before and 

after restoration.

This protocol is designed to assist landowners, land 

managers, water-quality regulators, and scientists in 

determining whether specific restoration/mitigation actions 

lead to significant progress toward achieving water quality 

goals to reduce Hg in the Bear Creek watershed. 

Acknowledgments

We thank Drs. Roger Ashley and Collin Eagles-Smith 

for peer review and significant improvements in this report. 

Special thanks also go to Francis Evanisko at the BLM 

California State Office who provided mapping services. Funds 

for research and preparation for this report have been provided 

by a grant from the CALFED Watershed Program and 

administered by the California Department of Water Resources 

to the Bureau of Land Management, grant agreement number 

4600004720. Additional support has come from the Bureau of 

Land Management, the Colusa County Resource Conservation 

District, and the U.S. Geological Survey’s Western Ecological 

Research Center. 

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Appendix A. Fish Processing Form for Bear Creek Mercury Study. 31

Appendix A. Fish Processing Form for Bear Creek Mercury Study.

49

Appendix 

A. Fish Processing form for Bear Creek Mercury Study 

Bear Creek Watershed Study Fish Processing Data SheetSite: GPS Coordinates

Latitude: 

Collection Date/Time: Longitude: Processing Date/Time: Accuracy (+/-):

Notes: 

Sample ID Species

Total Length 

(TL) (mm)

Standard Length 

(SL) (mm)

Total 

Wt (g)

Sample wt 

(g) NOTES
32 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Appendix B. Invertebrate Laboratory Processing Form for Bear Creek Mercury 

Study. 

50 

Appendix B. Invertebrate laboratory processing form for Bear Creek Mercury Study. 

Bear Creek Mercury Study 

Invertebrate Processing Data Sheet 

 

Site:___________________________________________________________________

Collection Date/Time:____________________________________________________ 

Processing Date/Time:___________________________________________________

GPS Coordinates 

Latitude:____________________________________________________ 

Longitude:__________________________________________________ 

Accuracy (+/-):_______________________________________________

Notes:______________________________________________________

 

 

Jar/ 

Sample ID 

Order Family Number Wt 

(g) 

Notes

NOTES:

 
Appendix C. Labels for Sample Containers. 33

Appendix C. Labels for Sample Containers. 

51 

Appendix C. Labels for sample containers. 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 

“Unique Sample ID Number”

Family:_____________ 

#/Wt:_____/__________g. 

“Site Name” 
34 Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California

Appendix D. Example Chain of Custody Form. 

52 

Appendix D. Example Chain of Custody Form.

 Chain of Custody Record Page_____of_____ 

White: LAB COPY 

Yellow: CUSTOMER COPY

Client: 

Contact 

COC receipt confirmation? Y / N If so, by: email / fax (circle one) 

Client project ID: 

Address: 

Email: 

PO #: Phone #: Fax #: 

Collection Miscellaneous Field 

Preservation 

Analyses required Comments Requested TAT in 

business days: 

20 (standard) 

 15  10 

5 Other ____ 

Surcharges apply for expedited turn around times.

Sample ID 

Date 

Time 

Sampler (initials) 

Matrix type 

# of containers 

Field filtered? (Y/N) 

Unpreserved / ice only 

 (circle one) 3 HCl / HNO

Other (specify) 

Total Hg, EPA 1631 

Methyl Hg, EPA 1630 

ICP-MS Metals (specify) 

As / Se species (specify) 

% Solids 

Filtration 

Other (specify) 

Other (specify) 

1 

2 

3 

4 

5 

6 

7 

8 

9 

10 

Relinquished by: Date: Time: Relinquished by: Date: Time: 

Received by: Date: Time: Received at BRL by: Date: Time: 

Shipping carrier: # of coolers: BRL work order ID: BRL project ID: 

3958 6th Avenue NW 

Seattle, WA 98107 Phone: 206-632-6206 

Fax: 206-632-6017 

samples@brooksrand.com 

www.brooksrand.com
Publishing support provided by the U.S. Geological Survey

Publishing Network, Sacramento and Tacoma Publishing Service 

Centers

For more information concerning the research in this report, contact the

 Director, Western Ecological Research Center

3020 State University Drive East

Modoc Hall, Room 3006

Sacramento, CA 95819

http://www.werc.usgs.gov/ 
Suchanek and others— Mercury Assessment and Monitoring Protocol for the Bear Creek Watershed, Colusa County, California—Scientific Investigations Report 2010-5018