U.S. Department of the Interior

U.S. Geological Survey

Scientific Investigations Report 2008–5198

Toxic Substances Hydrology Program

Fisheries: Aquatic and Endangered Resources Program

Contaminant Biology Program 

Prepared in cooperation with the

U.S. Environmental Protection Agency and the

Massachusetts Division of Fisheries and Wildlife 

Prevalence of Tumors in Brown Bullhead from 

Three Lakes in Southeastern Massachusetts, 2002
Cover. Photographs (clockwise from top left) showing (A) deployment of a fyke net in Ashumet Pond, (B) oral lesions in brown 

bullhead, (C) side view of brown bullhead as its length is being measured, and (D) front view of brown bullhead.
Prevalence of Tumors in Brown Bullhead 

from Three Lakes in Southeastern 

Massachusetts, 2002

By Paul C. Baumann1, Denis R. LeBlanc1, Vicki S. Blazer1, John R. Meier3, 

Stephen T. Hurley2, and Yasu Kiryu1

1

 U.S. Geological Survey

2

 Massachusetts Division of Fisheries and Wildlife, Buzzards Bay, Massachusetts

3

 U.S. Environmental Protection Agency, Cincinnati, Ohio

Toxic Substances Hydrology Program

Fisheries: Aquatic and Endangered Resources Program

Contaminant Biology Program

Prepared in cooperation with the

U.S. Environmental Protection Agency and the

Massachusetts Division of Fisheries and Wildlife

Scientific Investigations Report 2008–5198

U.S. Department of the Interior

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

DIRK KEMPTHORNE, Secretary

U.S. Geological Survey

Mark D. Myers, Director

U.S. Geological Survey, Reston, Virginia: 2008

For more information on the USGS—the Federal source for science about the Earth, its natural and living resources, 

natural hazards, and the environment:

World Wide Web: http://www.usgs.gov

Telephone: 1-888-ASK-USGS

Any use of trade, product, or firm names is for descriptive purposes only and does not imply endorsement by the 

U.S. Government.

Although this report is in the public domain, permission must be secured from the individual copyright owners to 

reproduce any copyrighted materials contained within this report.

Suggested citation:

Baumann, P.C., LeBlanc, D.R., Blazer, V.S., Meier, J.R., Hurley, S.T., and Kiryu, Yasu, 2008, Prevalence of tumors in 

brown bullhead from three lakes in southeastern Massachusetts, 2002: U.S. Geological Survey Scientific Investigations Report 2008–5198, 43 p., available online at http://pubs.usgs.gov/sir/2008/5198.
iii

Contents

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

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

Biology of the Brown Bullhead...........................................................................................................3

Geographic and Hydrologic Setting ..................................................................................................4

Previous Studies of Tumor Prevalence.............................................................................................4

Lake Selection and Field and Laboratory Methods .................................................................................4

Lake Selection and Characteristics...................................................................................................5

Field Collection Methods.....................................................................................................................7

External Examination, Blood Sampling, and Necropsy ................................................................10

Histopathology.....................................................................................................................................10

Age Estimation.....................................................................................................................................10

Blood Analysis by Comet Assays.....................................................................................................12

Statistical Methods.............................................................................................................................12

Prevalence of Tumors in Brown Bullhead...............................................................................................13

Size and Age Distributions ................................................................................................................13

Prevalence of External Raised Lesions...........................................................................................13

Prevalence of Barbel Deformities....................................................................................................14

Histopathology of External Lesions .................................................................................................14

Histopathology of the Liver, Spleen, and Kidney ...........................................................................16

Hepatosomatic Index .........................................................................................................................16

DNA Damage in Red Blood Cells .....................................................................................................16

Discussion.....................................................................................................................................................17

Summary........................................................................................................................................................20

Acknowledgments.......................................................................................................................................20

References Cited..........................................................................................................................................20

Glossary.........................................................................................................................................................25

Appendix 1. Selected Characteristics of Brown Bullhead (Ameiurus nebulosus) Collected 

from Ashumet, Santuit, and Great Herring Ponds, Southeastern Massachusetts, and 

Examined Externally in the Field, May–July 2002 .....................................................................28

Appendix 2. Selected Characteristics of Brown Bullhead (Ameiurus nebulosus) Collected 

from Ashumet, Santuit, and Great Herring Ponds, Southeastern Massachusetts, and 

Necropsied for Internal Examination and Tissue Sampling, May–June 2002......................35

Figures

1. Map showing locations of Ashumet, Johns, Santuit, and Great Herring Ponds, 

the Massachusetts Military Reservation (MMR), and the plumes of ground-water 

contamination from the MMR in 2007, southeastern Massachusetts.................................2

2. Photograph showing brown bullhead (Ameiurus nebulosus) from Ashumet Pond, 

Cape Cod, Massachusetts, with normal barbels and no apparent external lesions, 

May 2002 ........................................................................................................................................3
iv

3–5. Maps showing—

3. Topographic setting, water-table contours, plumes of ground-water 

contamination in 2007, and paths of drainage swales from the Massachusetts 

Military Reservation near Ashumet Pond, Falmouth and Mashpee, Cape Cod, 

Massachusetts.....................................................................................................................6

4. Topographic setting, water-table contours, and plumes of ground-water 

contamination from the Massachusetts Military Reservation in 2007 near 

Santuit Pond, Barnstable, Cape Cod, Massachusetts...................................................8

5. Topographic setting, water-table contours, and the sea-level Cape Cod Canal 

near Great Herring Pond, Plymouth, southeastern Massachusetts ...........................9

6. Photographs showing two examples of brown bullhead (Ameiurus nebulosus) from 

Ashumet Pond, Cape Cod, Massachusetts, with body-surface and oral lesions and 

shortened barbels, May 2002....................................................................................................11

7. Images of Comet assays showing (A) assay of blood sample from a Cape Cod, 

Massachusetts, brown bullhead (Ameiurus nebulosus) and (B) interpretation of a 

typical assay................................................................................................................................12

8–9. Graphs showing—

8. Prevalences of large (greater than 6 millimeters) raised oral and body-surface 

lesions in brown bullhead (Ameiurus nebulosus) from Great Herring, Santuit, 

and Ashumet Ponds, Cape Cod, Massachusetts, May–July 2002.............................14

9. Prevalences of multiple external lesions and melanistic patches in brown 

bullhead (Ameiurus nebulosus) from Great Herring, Santuit, and Ashumet 

Ponds, Cape Cod, Massachusetts, May–July 2002 .....................................................14

10. Images showing selected microscopic findings in tissues of brown bullhead 

(Ameiurus nebulosus) from Ashumet Pond, Cape Cod, Massachusetts, May 2002........15

11. Graph showing prevalence of raised external lesions in brown bullhead 

(Ameiurus nebulosus) from tributaries of the Great Lakes, rivers in the eastern 

United States and Canada, and Ashumet, Santuit, and Great Herring Ponds on 

Cape Cod, Massachusetts ........................................................................................................18

Tables

1. Chi-square statistical comparison of combined raised-lesion prevalence in brown 

bullhead (Ameiurus nebulosus) in the 1990s from Ashumet and Johns Ponds to the 

combined prevalence in three reference lakes, Cape Cod, Massachusetts .....................5

2. Hydrologic characteristics of Ashumet, Santuit, and Great Herring Ponds, 

southeastern Massachusetts.....................................................................................................5

3. Numbers of brown bullhead (Ameiurus nebulosus) sampled by location and date 

from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, 

May–July 2002...............................................................................................................................7

4. Percentage by location of brown bullhead (Ameiurus nebulosus) having external 

raised lesions, Ashumet, Santuit, and Great Herring Ponds, southeastern 

Massachusetts, May–July 2002...............................................................................................13

5. Percentage by location of brown bullhead (Ameiurus nebulosus) having barbel 

abnormalities, including missing, shortened, and knobbed barbels, Ashumet, 

Santuit, and Great Herring Ponds, southeastern Massachusetts, May–July 2002.........14
v

6. Percentage of necropsied brown bullhead (Ameiurus nebulosus) having selected 

microscopic lesions observed in liver tissue, Ashumet, Santuit, and Great Herring 

Ponds, southeastern Massachusetts, May–July 2002.........................................................16

7. Hepatosomatic index (liver weight divided by body weight) of brown bullhead 

(Ameiurus nebulosus) from Ashumet, Santuit, and Great Herring Ponds, 

southeastern Massachusetts, May–July 2002 ......................................................................16

8. Red blood cell nucleic DNA damage as determined by the Comet assay for 

brown bullhead (Ameiurus nebulosus) from Ashumet and Great Herring Ponds, 

southeastern Massachusetts, May–July 2002 ......................................................................17

Conversion Factors, Datum, and Abbreviations

Multiply By To obtain

Length

inch (in.) 2.54 centimeter (cm)

inch (in.) 25.4 millimeter (mm)

inch (in.) 25,400 micrometer (μm)

foot (ft) 0.3048 meter (m)

mile (mi) 1.609 kilometer (km)

Area

acre 0.4047 hectare (ha)

Volume

ounce, fluid (fl. oz) 29.57 milliliter (mL) 

ounce, fluid (fl. oz) 29,570 microliter (μL) 

gallon (gal) 3.785 liter (L) 

Mass

ounce, avoirdupois (oz) 28.35 gram (g)

ounce, avoirdupois (oz) 28,350 milligram (mg) 

pound, avoirdupois (lb) 0.4536 kilogram (kg) 

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

 °F=(1.8×°C)+32

Vertical coordinate information is referenced to the National Geodetic Vertical Datum of 1929 

(NGVD 29).

Altitude, as used in this report, refers to distance above the vertical datum.

Chemical concentration in water is given in units of milligrams per liter (mg/L) or micrograms 

per liter (μg/L). Chemical concentration in sediment is given in units of micrograms per kilogram 

(μg/kg). Sediment concentrations are sometimes expressed in parts per billion (ppb), which is 

equivalent to μg/kg.
vi

ABBREVIATIONS USED IN REPORT

AFCEE Air Force Center for Engineering and the Environment

AFCEE Air Force Center for Environmental Excellence (prior to 2007)

DNA deoxyribonucleic acid

EMS ethyl methane sulfonate

EROD 7-ethoxyresorufin-O-deethalyase

GLM general linear model

HAZWRAP Hazardous Waste Remedial Actions Program

HSI hepatosomatic index

IARC International Agency for Research on Cancer

IRP Installation Restoration Program

LMA low-melting agarose

mA milliampere

MADFW Massachusetts Division of Fisheries and Wildlife

MANG Massachusetts National Guard

MassGIS Massachusetts geographic information system

MMR Massachusetts Military Reservation

MS-222 tricaine methane sulfonate

PAH polycyclic aromatic hydrocarbon

PCB polychlorinated biphenyl

PCE tetrachloroethene

TCE trichloroethene

TRET Technical Review and Evaluation Team

USAEC U.S. Army Environmental Command

USEPA U.S. Environmental Protection Agency

USGS U.S. Geological Survey

V volt
Abstract

The Massachusetts Military Reservation (MMR) has 

been a military base on western Cape Cod since the early 

1900s. Contaminated surface water and ground water from 

the MMR have discharged into several kettle lakes on or 

near the base. To discover whether the prevalences of tumors 

and other lesions in brown bullhead (Ameiurus nebulosus) 

in these lakes, particularly Ashumet Pond, were elevated 

above normal, the U.S. Geological Survey (USGS), assisted 

by the U.S. Environmental Protection Agency (USEPA) 

and the Massachusetts Division of Fisheries and Wildlife 

(MADFW), conducted a study in 2002 of brown bullhead 

in Ashumet Pond and in two reference lakes, Santuit Pond 

(on Cape Cod) and Great Herring Pond (on the mainland of 

Massachusetts). Brown bullhead from Great Herring Pond 

had few external raised lesions (2.8 percent), a low prevalence 

of liver neoplasms (5 percent), and little genetic damage to 

their red blood cell nuclei. Brown bullhead from Ashumet 

Pond had a high prevalence of raised lesions (62.1 percent), 

which included histopathologically verified papillomas and 

squamous carcinoma; an elevated incidence of liver neoplasms 

(16.7 percent); and an elevated level of genetic damage to 

their red blood cell nuclei. Because red blood cells in fish 

have a lifespan of about 100 days, these results indicate an 

ongoing exposure to genotoxins in Ashumet Pond. Brown 

bullhead from Santuit Pond also had elevated prevalences of 

raised lesions (48.3 percent) and liver neoplasms (15 percent), 

although the prevalences of large and multiple lesions were 

significantly lower than those in fish from Ashumet Pond. 

These differences may indicate differing causes of pathology 

in the two lakes. The high prevalence of melanistic lesions 

on brown bullhead from Ashumet Pond, combined with the 

tumor pathology and genetic damage, implicates chemical 

carcinogens as one of the causal factors in that lake.

Introduction

Many kettle lakes in the glacial sand and gravel deposits 

of southeastern Massachusetts support populations of brown 

bullhead (Ameiurus nebulosus). These fish, a species of catfish 

also known locally as hornpout, feed on the organisms in the 

bottom sediments of the lakes. Bullhead use barbels, which are 

whisker-like sensory organs in contact with the sediments, to 

find their food (Scott and Crossman, 1973). Because bullhead 

spend much of their time in contact with the bottom sediments, the development of liver tumors1

, including hepatic 

(liver cell) and biliary (bile duct cell) tumors, in these fish has 

been linked to contaminants, particularly polycyclic aromatic 

hydrocarbons (PAHs), in the sediments (Baumann and others, 

1996; Baumann and Okihiro, 2000). Although skin tumors, 

including papillomas and squamous carcinoma, have been 

initiated in brown bullhead in the laboratory by using PAHs 

extracted from sediment (Black, 1983), there are probably 

multiple causal factors for such external tumors (Baumann and 

others, 1996). These tumors have been (and continue to be) 

used as an indicator of environmental health (Baumann, 1992).

The Massachusetts Military Reservation (MMR), which 

has been a multiuse military facility on western Cape Cod 

(fig. 1) since the early 1900s (Rolbein, 1995; Massachusetts 

National Guard (MANG), 2008), is adjacent to several large 

kettle lakes. Plumes of contaminated ground water from the 

MMR (fig. 1) discharge to several of these lakes (Savoie and 

others, 2000; McCobb and others, 2003; Air Force Center 

for Environmental Excellence (AFCEE), 2005, 2007), and 

in the past, storm drainage from the runways was directed to 

adjacent lakes (Hazardous Waste Remedial Actions Program 

(HAZWRAP), 1986).

Several studies by the Installation Restoration Program 

(IRP) at the MMR in the 1990s reported that the prevalence 

(measured as percentage) of tumors on brown bullhead in 

two lakes (Ashumet and Johns Ponds, fig. 1) near the MMR 

was elevated relative to the prevalence in several reference

lakes. There were concerns that these tumors were related to 

sources of contamination at the MMR, although cause and 

effect could not be established (AFCEE, 1997). In a review 

of these earlier studies, the Technical Review and Evaluation 

Team (TRET), a technical advisory group to the IRP, agreed 

with the finding of an elevated prevalence but noted that the 

numbers of fish sampled in the studies were small (AFCEE, 

2008a). The TRET recommended that any future examination 

1

Terms listed in the glossary at the back of this report are in bold type 

where first used in the text.

Prevalence of Tumors in Brown Bullhead from 

Three Lakes in Southeastern Massachusetts, 2002

By Paul C. Baumann, Denis R. LeBlanc, Vicki S. Blazer, John R. Meier, Stephen T. Hurley, and Yasu Kiryu
2 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Figure 1. Locations of Ashumet, Johns, Santuit, and Great Herring Ponds, the Massachusetts Military Reservation (MMR), and 

the plumes of ground-water contamination from the MMR in 2007, southeastern Massachusetts.

CAPE COD

BAY

BUZZARDS

BAY

NANTUCKET SOUND

VINEYARD SOUND

Falmouth

Sandwich

Bourne

Plymouth

Mashpee

Wareham

Barnstable

see figure 3

see figure 4

see figure 5

70°40' 70°30'

41°50'

41°40'

41°35'

41°45'

GREAT 

HERRING

POND

SANTUIT

POND

JOHNS

POND

Cape Cod Canal

ASHUMET

POND

Base from Office of Geographic and Environmental Information (MassGIS), 25:000, 2005

Massachusetts State Plane Coordinate System projection, North American Datum of 1983

Plumes mapped by Air Force Center for

Engineering and the Environment (2007)

and U.S. Army Environmental Command (2007a)

EXPLANATION

Areal extent of plumes of 

ground-water contamination, 2007

Massachusetts Military 

Reservation

Ocean or lake

Cranberry bog

Town boundary

0 4 MILES

0 4 KILOMETERS
Introduction 3

of the problem include a statistically based prevalence study 

that sampled more fish from the lakes of concern and similar 

reference lakes.

As a result of the TRET’s recommendations, the U.S. 

Geological Survey (USGS) conducted a study in 2002 of the 

prevalence of tumors in brown bullhead in Ashumet Pond and 

two reference lakes, Santuit and Great Herring Ponds (fig. 1). 

The study was done in cooperation with the U.S. Environmental Protection Agency (USEPA) and the Massachusetts Division of Fisheries and Wildlife (MADFW). The objective of 

the study was to collect a sufficient number of brown bullhead 

from each lake to allow a test of the statistical significance of 

differences in tumor prevalence among the lakes. This study 

was not designed to address the causes of any elevated tumor 

frequency (see Discussion section).

This report describes the sampling effort, presents the 

data on the prevalence and characteristics of the tumors, and 

discusses the statistical significance of differences in tumor 

prevalence between Ashumet Pond and the two reference 

lakes. The report also presents results on the pathology of the 

tumors, as well as evidence from blood analysis of genetic 

damage in the brown bullhead. Finally, the results are compared with findings from the earlier studies at the MMR and 

from investigations of brown bullhead tumors in other lakes 

and rivers in North America.

Biology of the Brown Bullhead

The brown bullhead (fig. 2) is the most widely distributed 

of the native bullhead catfish, found from the Atlantic Ocean 

to the Gulf Coast watersheds and from eastern Canada to 

Alabama (Scott and Crossman, 1973). It is also native to the 

Great Lakes system, Hudson Bay, and the Mississippi River 

watershed. The specific epithet “nebulosus” means “clouded,” 

referring to the fish’s mottled sides. Brown bullhead can be 

distinguished from other bullhead by the whitish color at 

the base of the barbels and the square-tipped or only slightly 

rounded caudal fin. This species is common to lakes and slowmoving areas of rivers and is tolerant of warm water temperatures, low oxygen levels, and contamination. Like other 

catfish, brown bullhead are active primarily at night, when 

their sensory barbels help them locate food in the darkness. 

They are omnivorous bottom feeders that eat a wide variety of 

plant and animal material, including aquatic insects and other 

benthic invertebrates, small fish, and even algae. This feeding 

regime can result in a high exposure to contaminants in sediment, bottom waters, or invertebrates living in the sediment.

Figure 2. Brown bullhead (Ameiurus nebulosus) from Ashumet Pond, Cape Cod, Massachusetts, with normal 

barbels and no apparent external lesions, May 2002.
4 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Geographic and Hydrologic Setting

The study area in southeastern Massachusetts is 

characterized by extensive sand and gravel deposits from the 

last ice age (LeBlanc and others, 1986; Oldale and Barlow, 

1986; Masterson and others, 1997). These glacial deposits are 

pitted with kettle lakes. Direct surface runoff to the lakes is 

limited because the soils are sandy and precipitation infiltrates 

the ground rapidly. The lakes are in direct hydraulic contact 

with the surrounding aquifer, and typically their water budgets 

are dominated by ground-water inflow and outflow seepage 

rather than by surface-water inflow and outflow (Masterson 

and others, 1998; Walter and others, 2002; Walter and 

Whealan, 2005).

Ashumet and Santuit Ponds are on Cape Cod and are separated hydraulically from the mainland by the sea-level Cape 

Cod Canal (fig. 1). Great Herring Pond is on the mainland in 

the Plymouth-Carver aquifer (Hansen and Lapham, 1990). 

The ground-water watersheds of the three lakes have similar 

land use and vegetation, except that the area north of Ashumet 

Pond includes the MMR (fig. 1).

Past military activities since the early 1900s at the MMR 

resulted in contamination of the Cape Cod aquifer (AFCEE, 

2007; U.S. Army Environmental Command (USAEC), 2007a). 

More than a dozen plumes of contaminated ground water 

emanate from the MMR and extend toward the coast (fig. 1). 

The plumes contain industrial solvents, fuels and fuel additives, landfill leachate, treated-wastewater effluent, and, in 

the northern part of the base, explosives and propellants from 

military ordnance. These plumes are being remediated by the 

U.S. Department of Defense (AFCEE, 2007; USAEC, 2007b). 

Several plumes discharge to lakes and streams adjacent to the 

reservation (Savoie and others, 2000; AFCEE, 2005, 2007), 

although there is no information about these discharges prior 

1986, the year when the IRP’s plume investigations began.

Previous Studies of Tumor Prevalence

The earliest known observations of external lesions on 

brown bullhead on western Cape Cod were made by MADFW 

field crews in Johns Pond and its outflow, the Quashnet 

River, in October 1991 and May 1992 (Stephen T. Hurley, 

MADFW, Southeast Region, unpublished files, December 

2007). Concerns that fish populations in Ashumet and Johns 

Ponds may have been exposed to contaminants from the MMR 

prompted a series of studies in the early 1990s. The first of 

these projects, conducted in 1992, noted a high prevalence of 

external tumors in brown bullhead from both Ashumet Pond 

(42 percent) and Johns Pond (67 percent) (HAZWRAP, 1995). 

In 1993, fish were collected again from Ashumet and Johns 

Ponds, as well as from Long, Coonamessett, and Grews Ponds 

in Falmouth (Stegeman, 1997). Finally, brown bullhead were 

examined from Ashumet and Johns Ponds, as well as from 

Long Pond in Brewster and Harwich, in spring 1994 and from 

these three lakes plus Upper Mill Pond in Brewster in fall 

1994 (HAZWRAP, 1996). The numbers of fish collected for 

each site and time were relatively small, ranging from 2 to 30, 

but were sufficient to indicate a potential problem.

Studies in the early 1990s (Stegeman, 1997) reported 

moderately but significantly elevated 7-ethoxyresorufinO-deethylase (EROD) and P4501A activity in brown bullhead 

collected from Ashumet and Johns Ponds compared to brown 

bullhead collected from Long and Coonamessett Ponds. The 

EROD and P450 assays are designed to measure the activity 

of these liver enzymes, which metabolize a variety of organic 

contaminants. Subsequent examination by other investigators 

of these enzymes in Cape Cod fish was inconclusive, however, 

because the measured enzyme activity varied significantly 

within and among the lakes sampled (AFCEE, 1997). 

Prior to the initiation of the study described in this report, 

the tumor-prevalence data from the earlier studies near the 

MMR were combined from all sampling times for each location in order to do statistical testing (AFCEE, 2008b, c). Numbers of fish with and without external raised lesions from each 

location were entered in a contingency table (table 1), and 

tumor prevalence was examined for independence with a chisquare test. When all the sites were compared separately with 

each other, tumor prevalence was found to be significantly 

different among sites (p < 0.001). When the data from 

Ashumet and Johns Ponds were combined, and data from 

all of the other lakes were similarly combined as a reference 

prevalence, Ashumet and Johns Ponds were found to have a 

significantly higher incidence of external tumors (p < 0.001) 

(table 1). In combining the data to obtain sufficient sample 

numbers, seasonal and yearly fluctuations in tumor frequency 

were not considered. Also, data on fish age were not available. 

Despite these limitations, the large difference in tumor prevalence (37 percent for Ashumet and Johns Ponds compared to 

7 percent for the reference lakes), along with the strong statistical significance, indicated that tumor prevalence in brown 

bullhead from Ashumet and Johns Ponds was higher than that 

in the reference lakes. This study was designed to provide data 

to determine tumor prevalence based on a larger sample size.

Lake Selection and Field and 

Laboratory Methods

The study included collection and examination of brown 

bullhead from three lakes to determine the prevalences of 

tumors and other lesions; histopathology of selected tissue 

samples to diagnose lesion types, which can provide evidence 

of lesion causation; and analysis of red blood cell nucleic 

damage, which can provide evidence of ongoing exposure to 

genotoxins. A detailed description of these methods is beyond 

the scope of this report. The reader is referred to Blazer and 

others (2007), Rafferty and Grazio (2006), and the other references at the end of the report for additional information.
Lake Selection and Field and Laboratory Methods 5

Lake Selection and Characteristics

Three lakes were included in this study—Ashumet Pond 

adjacent to the MMR and two reference lakes, Santuit and 

Great Herring Ponds (fig. 1). Selected characteristics of the 

three lakes are given in table 2 and described in MADFW 

(2008) and AFCEE (1997). Although it was included in the 

original design of the study, Johns Pond (fig. 1) was dropped 

because too few brown bullhead were collected from this lake.

The reference lakes were selected because they are similar to Ashumet and Johns Ponds in several ways. The groundwater contributing areas to the lakes are comparable in size 

(Masterson and Walter, 2000; John P. Masterson, U.S. Geological Survey, written commun., January 2008), and all the 

lakes receive substantial inflow from ground-water seepage, 

which generally is focused near the lake shore. The land use 

near the lakes, with the exception of the unique land use on the 

MMR near Ashumet and Johns Ponds, is similar among the 

lakes and includes residential areas with onsite septic systems, 

small cranberry bogs, and undeveloped, forested land. All the 

lakes are used for recreational motorized boating and fishing, 

particularly during the summer months. Finally, both Santuit 

and Great Herring Ponds were known to have large enough 

populations of brown bullhead to provide an adequate sample 

size in 2–3 days of collecting, and these lakes were within 

10 mi of the MMR to facilitate the field effort.

Ashumet Pond (fig. 3) is a classic ground-water flowthrough kettle lake (McCobb and others, 2003). The lake 

has no surface outlet and only a minor surface inlet from an 

abandoned cranberry bog at its northern end. Two drainage 

swales from the MMR airfield reach the lake (fig. 3). The 

swales channeled surface runoff to the cranberry bog and 

lake during particularly intense storms until 2002, when 

infiltration basins were installed on the MMR to capture the 

runoff (Robert Burt, 102nd Fighter Wing, Otis Air National 

Guard Base, written commun., December 2007). The runoff 

during the early 1950s to the late 1960s reportedly included 

dissolved and fluid-phase hydrocarbons (HAZWRAP, 1986; 

K-V Associates, Inc., and IEP, Inc., 1991; Stephen T. Hurley, 

MADFW, Southeast Region, unpublished files, December 

2007); in the late 1960s, the Air Force began efforts, such 

as the installation of oil-water separators and berms, to limit 

hydrocarbon-contaminated runoff from reaching the lake.

PAHs, including known carcinogens, were found in 

sediment collected from Ashumet Pond (AFCEE, 1997). The 

results of sediment chemical analyses (Jon Davis, AFCEE, 

data retrieved from the IRP database on November 15, 2006) 

indicated that levels of PAHs from one sample location were 

somewhat elevated (850 μg/kg for phenanthrene, fluoranthene, 

and pyrene combined) above values for these same compounds at five Great Lakes reference sites (304–527 μg/kg). 

Table 1. Chi-square statistical comparison of combined raised-lesion prevalence in brown bullhead 

(Ameiurus nebulosus) in the 1990s from Ashumet and Johns Ponds to the combined prevalence in three 

reference lakes, Cape Cod, Massachusetts. 

[Expected counts are included below observed counts. Source: Air Force Center for Engineering and the Environment 

(AFCEE) (2008b)]

Characteristic Category

Number of bullhead

Total number of 

observed bullhead Ashumet and 

Johns Ponds Reference lakes

With external lesions Observed 40 8 48

Expected 23.35 24.65

Without external lesions Observed 67 105 172

Expected 83.65 88.35

Total number of observed 

bullhead at all locations 107 113 220

Chi-square statistic = 11.881 + 11.250 + 3.316 + 3.140 = 29.587

Degrees of freedom = 1, p < 0.001

Table 2. Hydrologic characteristics of Ashumet, Santuit, and 

Great Herring Ponds, southeastern Massachusetts. 

[Source: Massachusetts Division of Fisheries and Wildlife, 2008]

Characteristic Ashumet 

Pond

Santuit 

Pond

Great Herring 

Pond

Area in acres 202 172 376

Average depth in feet 23 5 20

Maximum depth in feet 65 9 42
6 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Figure 3. Topographic setting, water-table contours, plumes of ground-water contamination in 2007, and paths of drainage swales 

from the Massachusetts Military Reservation near Ashumet Pond, Falmouth and Mashpee, Cape Cod, Massachusetts.

40

45

50

55

60

65

35

30

25

20

70°35' 70°32'30" 70°30'

41°40'

41°37'30"

0 1 MILE

0 1 KILOMETER

Base from Office of Geographic and Environmental Information (MassGIS), DRG, 24:000, 2001

Massachusetts State Plane Coordinate System projection, North American Datum of 1983

Plumes mapped by AFCEE (2007) and USAEC (2007a)

Water table mapped by Walter and Whealan (2005)

50

EXPLANATION

Areal extent of plumes of ground-water contamination, 2007

Water-table contour—altitude of water table. Contour interval is 5 feet. Datum is NGVD 29

Drainage swale

Boundary of Massachusetts Military Reservation
Lake Selection and Field and Laboratory Methods 7

These compounds are in PAH combustion mixtures and serve 

as good markers for the longer chain carcinogens. The detection limit used for these analyses was fairly high (220 μg/kg) 

and precluded the detection of most of the carcinogenic PAHs, 

such as benzo(a)pyrene. These compounds commonly occur 

at lower concentrations than this limit, although benzo(b)fluoranthene, a known carcinogen, was detected at 270 μg/kg. The 

PAH levels measured in the sediments from Ashumet Pond, 

however, were substantially lower than the levels measured at 

USEPA-designated “Area of Concern” sites where livertumor epizootics have been identified (such as the Black and 

Cuyahoga Rivers in Ohio); at these sites, concentrations 

of these three compounds were measured at levels of 

3,500–109,000 μg/kg (Paul C. Baumann, U.S. Geological 

Survey, unpublished files, 2007).

Several ground-water contaminant plumes from the 

MMR were known to discharge to Ashumet Pond at the time 

of this study (2002). Part of the Chemical Spill 10 plume, 

which contained trichloroethene (TCE) at concentrations 

near the lake as high as 1,962 μg/L in 1999, was believed to 

be discharging to the northwestern part of the lake (AFCEE, 

2000). Similarly, parts of the Ashumet Valley plume, which 

contained tetrachloroethene (PCE) concentrations as high as 

14.6 μg/L in 2002 (AFCEE, 2003) and various contaminants 

associated with treated municipal wastewater (Walter and 

others, 1996; McCobb and others, 2003), also was discharging 

to the lake. Both TCE and PCE have recently been designated 

as animal carcinogens and probable human carcinogens by 

the International Agency for Research on Cancer (IARC, 

1997). Concentrations of TCE and PCE are diluted rapidly as 

ground water mixes with the lake water, and these compounds 

generally are not detectable in the lake water at the discharge 

locations (AFCEE, 1999). Both plumes continue to discharge 

to the lake, but at increasingly lower concentrations because of 

ongoing ground-water remediation. No information is available on contaminants that may have discharged to the lake 

prior to about 1986, when efforts to identify and characterize 

the plumes began.

Santuit Pond (fig. 4) also is a ground-water flow-through 

kettle lake. In addition to ground-water inflow and outflow, 

Santuit Pond has a surface inlet from an active cranberry 

bog and surface outlets at its southern end to the Santuit and 

Little Rivers and a small, unnamed bog. Flow to the streams 

and bogs is controlled for cranberry production. There is no 

evidence that contamination from the MMR reaches the lake. 

Santuit Pond is shallower and smaller than the other two lakes 

in the study (table 2).

Great Herring Pond (fig. 5) has a surface outlet to the 

Herring River and a surface inlet at its northern end from a 

small stream that drains several cranberry bogs and Little 

Herring Pond. Recreational motorized boating, including the 

use of personal watercraft vehicles (for example, the Jet Ski), 

particularly during the summer, is heavier on Great Herring 

Pond than on the other two lakes in the study (Stephen T. 

Hurley, MADFW, Southeast Region, unpublished files, 

December 2007). Contaminants from the MMR cannot reach 

Great Herring Pond, which is on the mainland side of the 

Cape Cod Canal.

Field Collection Methods

Brown bullhead were collected with fyke nets and by 

electrofishing. The fyke nets had D-shaped first hoops with 

openings of 0.6–0.76 m, mesh with 12- or 25-mm openings 

when stretched, and leads 9–15 m long. Each afternoon, 

6–16 nets were placed near shore with leads attached to 

posts driven at locations suggested by the MADFW crew 

on the basis of their knowledge of the lake or in places with 

habitat known to be favored by bullhead. All nets were 

fished overnight and sampled by afternoon of the next day. 

Electrofishing was done by the MADFW with a pulsed 

direct-current electrofishing boat (Smith-Root, Inc., Model 

GPP Electrofisher). The sampling dates and numbers of fish 

caught and examined for each lake are summarized in table 3. 

Brown bullhead with a minimum length of 250 mm (selected 

to maximize the number of fish of age 3 and older) were kept 

for examination. The first 20–24 fish captured were taken as 

a necropsy subsample and transported from the collection 

site to a field laboratory in water-filled coolers. The longest 

transportation time was 30 minutes. Brown bullhead continued 

to be caught and processed daily until about 60 fish had been 

collected from each lake. If insufficient time was available to 

examine all fish that had been caught in a day, the fish were 

held in cages in the lake where they had been captured until 

the examination could be finished.

Table 3. Numbers of brown bullhead (Ameiurus nebulosus) sampled by location and date from Ashumet, Santuit, 

and Great Herring Ponds, southeastern Massachusetts, May–July 2002.

Location Date sampled (2002) Number of fish,

necropsy and field exam

Number of fish,

field exam only

Total number of 

fish sampled

Ashumet Pond May 21–22, June 7 24 34 58

Santuit Pond June 25–28, July 1 20 39 59

Great Herring Pond June 4–7, June 24–26 20 50 70
8 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Figure 4. Topographic setting, water-table contours, and plumes of ground-water contamination from the Massachusetts Military 

Reservation in 2007 near Santuit Pond, Barnstable, Cape Cod, Massachusetts.

40

45

50

55

60

65

35

30

25

20

70°30' 70°27'30"

41°42'30"

41°40'

Base from Office of Geographic and Environmental Information (MassGIS), DRG, 24:000, 2001

Massachusetts State Plane Coordinate System projection, North American Datum of 1983

Plumes mapped by AFCEE (2007) and USAEC (2007a)

Water table mapped by Walter and Whealan (2005)

0 1 MILE

0 1 KILOMETER

50

EXPLANATION

Areal extent of plumes of ground-water contamination, 2007

Water-table contour—altitude of water table. Contour interval is 5 feet. Datum is NGVD 29

Boundary of Massachusetts Military Reservation
Lake Selection and Field and Laboratory Methods 9

Figure 5. Topographic setting, water-table contours, and the sea-level Cape Cod Canal near Great Herring Pond, Plymouth, southeastern Massachusetts.

50

55

40

30

20

10

60

65

45

35

25

15

5

40

30

20

10

45

35

25

15

5

5

Base from Office of Geographic and Environmental Information (MassGIS), DRG, 24:000, 2001

Massachusetts State Plane Coordinate System projection, North American Datum of 1983

Water table adapted from Hansen and Lapham (1990)

0 1 MILE

0 1 KILOMETER

50

EXPLANATION

Water-table contour—altitude of water table. Contour interval is 5 feet. Datum is NGVD 29

70°35' 70°32'30"

41°50'

41°47'30"
10 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

External Examination, Blood Sampling, and 

Necropsy

Fish were processed on site at the lake in several steps. 

The lengths of the fish were measured, raised lesions on the 

body surface and in the oral cavity (fig. 6) were counted and 

measured, and the positions of the lesions were recorded. 

Areas of nonraised melanistic spotting were also recorded. 

Abnormal barbels were classified into one of three categories: 

missing, shortened, or knobbed. Open lesions, melanistic 

patches, and eye abnormalities also were described. Gender 

was not recorded for fish processed at the lake. All fish not to 

be subsampled for necropsy were finclipped for identification 

and released at their collection site.

At the field laboratory, fish from the necropsy subsample 

were anesthetized with tricaine methane sulfonate (MS-222), 

measured (total length), and weighed. Mixed arteriovenous 

blood was drawn from the caudal artery and vein by using 

the lateral approach described by Schmitt and others (1999). 

A heparinized vacutainer tube was used to collect a small 

amount of blood (less than 0.5 mL). This blood was stored 

on ice and shipped overnight in ice-containing coolers to 

the USEPA Ecological Exposure Research Division of the 

National Exposure Research Laboratory in Cincinnati, Ohio. 

The blood was then processed within 24 hours through the 

slide-storage step until later analysis was done by using the 

Comet assay (Tice, 1995). Two blood smears were also made 

immediately on site.

After the blood was sampled, the fish were euthanized 

by cervical dislocation. Each individual was inspected for 

grossly observable lesions involving the skin, eyes, and tissues 

within the oral cavity by following the same procedure used in 

the field. Visible abnormalities, including the number of each 

type of barbel abnormality (nasal, maxillary, and chin), were 

recorded. Raised lesions were counted, and the prevalence was 

calculated. Fish were then necropsied. An incision was made 

in the abdomen from the anus to the isthmus, and the viscera 

were examined for lesions. Any abnormalities or unusual 

features that were grossly visible were noted, and a subsample 

was preserved for histopathology. The sex of the fish was 

determined and recorded. The liver and spleen were excised, 

examined, and weighed. Any visible lesions were recorded, 

and the organs were immediately preserved in 10-percent 

neutral-buffered formalin for histopathology. Pieces about 

2 cm in size of the trunk kidney were also removed and placed 

in formalin for histopathology, as were any gross lesions noted 

on any other tissues. For brown bullhead from Ashumet Pond, 

a subsample of barbels and gills was preserved. One pectoral 

spine was removed whole for later determination of fish age in 

the laboratory.

Histopathology

Preparation of tissues for histopathology followed 

standard procedures currently on file at the Fish Health Branch 

of the USGS Leetown Science Center, Kearneysville, W.V. 

Tissue sections were processed and stained with hematoxylin 

and eosin. In addition, all liver, spleen, and kidney sections 

were stained with Perl’s Prussian blue stain, and selected 

slides were stained with von Kossa and Giemsa stains (Luna, 

1992). Light-microscope observations of bile duct hyperplasia 

(increased number of bile ducts), altered foci (areas that have 

altered staining characteristics and are generally accepted as 

preneoplastic), neoplasia, parasites, granulomas (a specific 

type of chronic inflammation), multinucleate giant cells 

(a characteristic of some chronic inflammatory reactions), 

and thickening of the serosa (lining of the liver) were scored 

as present or absent. Macrophage aggregate scoring used a 

severity scale of 0 (none), 1 (mildly scattered), 2 (moderate), 

and 3 (severe). Tissue slides were archived at the Fish 

Health Branch.

Age Estimation

Spines were sectioned near the base, and ages were 

estimated by two experienced technicians. If their independent 

estimates did not agree, both technicians reexamined the cross 

sections in question to reach an age estimate by consensus. 

The overall percent agreement between technicians was 

80 percent, which is high for age estimates from spine cross 

sections. Age estimates that seemed low for the size of the fish 

were redetermined by using a variable-setting fiber-optic light 

source, which allowed the use of higher intensity light to look 

for annuli near the spine edge. Of 12 fish from two sites that 

were reexamined, the second age estimate agreed with the first 

for 8 fish. Of the other 4 fish, all from Ashumet Pond, the age 

was increased by 1 year (2 fish), 2 years (1 fish), and 3 years 

(1 fish).
Lake Selection and Field and Laboratory Methods 11

Figure 6. Two examples of brown bullhead (Ameiurus nebulosus) from Ashumet Pond, Cape Cod, Massachusetts, 

with body-surface and oral lesions and shortened barbels, May 2002.
12 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Blood Analysis by Comet Assays

The single-cell gel electrophoresis, also called the 

“Comet” assay, is a method that measures deoxyribonucleic 

acid (DNA) damage at the individual cell level by analysis of 

DNA migration in gel-embedded cells following electrophoresis (Singh and others, 1988). The name “Comet” refers to 

the formation of a long comet-like tail of DNA fragments as a 

result of DNA damage (fig. 7). This assay has been used as an 

indicator of DNA damage in wild fish from sites contaminated 

with a variety of compounds, including PAHs, polychlorinated 

biphenyls (PCBs), and heavy metals (Chang and others, 2005; 

Devaux and others, 1998; Winter and others, 2004; Yang and 

others, 2006). Because of funding constraints, blood samples 

from fish caught only in Ashumet and Great Herring Ponds 

were analyzed with this method.

The Comet assay was performed as described by Singh 

and others (1988) and modified by Tice (1995). To explain 

briefly, 3 μL of fish blood were diluted with 1 mL of cold 

mincing solution. Then 10 μL (in duplicate) of the diluted 

blood were mixed with 75 μL of 0.5-percent low-melting 

agarose (LMA) at 37°C and pipetted onto a microscope slide 

precoated with a layer of 1.2-percent normal-melting agarose. 

The slides were cooled on ice until the agarose layer hardened; 

then another layer of LMA was added, and the slides were 

cooled on ice again. Next the slides were placed overnight 

in cold (about 4°C) lysing solution. This step was followed 

by placement in alkaline electrophoresis buffer (pH greater 

than 13) for 15 minutes to allow for unwinding of the DNA. 

Electrophoresis was then run for 10 minutes (25 V, 265 mA) 

at room temperature. After electrophoresis, the slides were 

neutralized, fixed by immersion in cold methanol, air-dried, 

and stored at room temperature.

The slides were stained with ethidium bromide and were 

scored on a fluorescent microscope at 400X magnification 

by using Comet image-analysis software (Komet 4.0, Kinetic 

Imaging, Liverpool, United Kingdom). The Comet parameters 

that were analyzed for each cell included the length of DNA 

migration (tail length, in micrometers), the percentage of 

migrated DNA (percentage of DNA in the tail relative to total 

DNA in the image), and the tail-extent moment (calculated 

as the tail length multiplied by the percentage of DNA in the 

tail). Fifty cells per slide and a total of 100 cells per fish were 

analyzed to evaluate the extent of DNA damage for each fish.

Statistical Methods

Tukey’s studentized range test was used to compare fish 

length and hepatosomatic index (HSI) among fish from the 

different lakes. The chi-square test, or Fisher’s exact test when 

sample-size conditions of the chi-square test were not met, 

was used to compare the prevalences of lesions and barbel 

abnormalities among fish caught in different locations. For 

the Comet assay, statistical analyses were performed by using 

SAS version 8.1 software (SAS Institute, Inc., Cary, N.C.). 

The general linear model (GLM) included the variables sampling site (contaminated versus reference), fish gender (female 

versus male), and length (as a measure of age). This method of 

analysis was used to detect whether the extent of DNA damage 

measured by the Comet assay (tail length, percent tail DNA, 

tail-extent moment) was affected by these variables. Logistic 

regression models were developed to examine the relationship 

between the probability of lesion or deformity occurrence and 

DNA damage measured by the Comet assay.

Figure 7. Images of Comet assays showing (A) assay of blood 

sample from a Cape Cod, Massachusetts, brown bullhead 

(Ameiurus nebulosus) and (B) interpretation of a typical assay.

Background

Head extent Tail extent

B

A
Prevalence of Tumors in Brown Bullhead 13

Prevalence of Tumors in Brown 

Bullhead

The prevalence of tumors was determined from the 

examination of 187 brown bullhead collected in spring 

2002 from Ashumet, Santuit, and Great Herring Ponds 

(table 3). Selected data from the field examination of the fish 

are reported in appendix 1. The combination of fyke nets and 

electrofishing resulted in complete or nearly complete samples 

from the three lakes. The histopathology of the skin, body 

tumors, and liver tissue and the genetic damage to red blood 

cells were determined from subsamples collected from the 64 

necropsied brown bullhead from the lakes. Selected data from 

the laboratory examination of the necropsied fish are reported 

in appendix 2.

Size and Age Distributions

The distributions of brown bullhead lengths differed 

among the three lakes. Brown bullhead from Ashumet Pond 

were largest in average size (379 mm), and those from 

Santuit Pond were significantly (α = 0.05, Tukey’s studentized 

range test) shorter (286 mm) than fish from either Great 

Herring Pond (336 mm) or Ashumet Pond (appendix 1). 

The size difference between the brown bullhead specimens 

from Santuit and Ashumet Ponds was great enough that the 

range overlapped for only 3 fish (1 from Santuit Pond and 

2 from Ashumet Pond) of the 117 fish sampled. Fish from 

Great Herring Pond were closer in size to fish from Ashumet 

Pond but were still significantly smaller (α = 0.05, Tukey’s 

studentized range test).

Ages were obtained only for the fish that were necropsied 

(appendix 2). Brown bullhead from Ashumet Pond were 

older on average than those from either of the other lakes. 

One-third of the fish from Ashumet Pond were age 7 or older, 

whereas only one-fifth or fewer of brown bullhead from the 

other lakes reached that age. The ages of insufficient numbers 

of fish were obtained to permit determination of age-specific 

lesion frequencies. Also, only spines were used for aging, 

and these have recently been shown to underestimate the true 

age of fish, particularly at year 5 and older (Maceina and 

Sammons, 2006).

Prevalence of External Raised Lesions

The prevalences of body-surface, oral, and total external 

raised lesions, presumed by field observation to be tumors, 

were dependent on location (chi-square test, p < 0.0001) 

(table 4). Fish from Ashumet Pond had the highest prevalence, 

and fish from Great Herring Pond had the lowest prevalence. 

Fish from Great Herring Pond had a prevalence of total 

external raised lesions that was less than 3 percent 

(table 4). Prevalences of oral and body-surface lesions in 

brown bullhead from Santuit Pond were each about one-half 

of that in brown bullhead from Ashumet Pond, although the 

prevalences of total external lesions in the two lakes were 

similar. The prevalence of large, raised lesions also differed 

between Ashumet and Santuit Pond fish (fig. 8). A greater 

percentage of fish had large (greater than 6 mm) oral and 

body-surface lesions in Ashumet Pond. If medium and large 

oral lesions were grouped (all lesions 3 mm or larger), they 

were observed on 33.8 percent of brown bullhead from Santuit 

Pond but on 56.9 percent of brown bullhead from Ashumet 

Pond. Similarly, medium and large body-surface lesions 

(3 mm or larger) were observed on 22.1 percent of Santuit 

Pond fish but on 70.6 percent of Ashumet Pond fish.

The percentage of fish having single and multiple lesions 

also differed by location. The prevalence of single external 

lesions was significantly different on fish from the three 

lakes (chi-square test, p = 0.0443), although the prevalence of 

single lesions was similar for fish from Santuit and Ashumet 

Ponds; fish from Great Herring Pond had a significantly 

lower prevalence of single lesions than did the fish from the 

other two lakes. The occurrence of multiple lesions also was 

significantly different on fish from the three lakes (chi-square 

test, p < 0.0001) (fig. 9). Multiple lesions were observed 

almost twice as often on fish from Ashumet Pond as on fish 

from Santuit Pond and were not found on any fish from Great 

Herring Pond. Similarly, melanistic patches—abnormally 

darkened, but not raised, areas of skin—were absent on fish 

from Santuit Pond but were observed on almost 70 percent 

of the fish from Ashumet Pond (fig. 9). Melanistic patches 

on brown bullhead from Great Herring Pond had less than 

10 percent prevalence, significantly lower than on fish in 

Ashumet Pond (chi-square test, p < 0.0001).

Table 4. Percentage by location of brown bullhead (Ameiurus 

nebulosus) having external raised lesions, Ashumet, Santuit, 

and Great Herring Ponds, southeastern Massachusetts, May–

July 2002.

Location Oral lesions Body-surface 

lesions

Total external 

lesions

Ashumet Pond 46.6 48.3 62.1

Santuit Pond 28.8 20.3 48.3

Great Herring Pond 0 2.8 2.8
14 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Prevalence of Barbel Deformities

Fish from Santuit Pond had the highest prevalence of 

missing barbels, whereas fish from Ashumet Pond had the 

lowest prevalence (table 5). This difference was significant at 

the p < 0.05 level (chi-square test, p = 0.0378). The frequency 

of shortened barbels was similar among fish from the three 

lakes. Whereas fish from Ashumet Pond had at least twice the 

prevalence of knobbed barbels as fish from the other lakes, the 

prevalences were not significantly different (chi-square test, 

p = 0.0496). A comparison of total barbel lesions on fish from 

the three lakes did not indicate any significant differences 

(chi-square test, p = 0.1843).

Histopathology of External Lesions

Of the 24 necropsied fish collected from Ashumet Pond, 

10 fish were reported by gross observation to have raised 

body-surface lesions. Samples of lesions from six of the fish 

were submitted for histopathology. One lesion was a nonneoplastic (hyperplastic) lesion of epidermal thickening 

(fig. 10A), two were squamous cell carcinomas, and three 

were papillomas (fig. 10B). Thirteen of the necropsied fish 

were reported by gross observation to have raised lesions in 

the oral cavity. Pieces of eight of these lesions were taken for 

histopathology. Three of the eight lesions were papillomas, 

and five were squamous cell carcinomas (fig. 10C). Thus, of 

the 14 lesions collected from fish from Ashumet Pond (diagnosed by gross observation as tumors) and fixed for histopathology, 13 lesions (or 93 percent) were verified as neoplasia.

At Santuit Pond, 8 of the 20 necropsied fish were 

reported by gross observation to have raised body-surface 

or oral tumors; two samples were submitted for histopathology. Both were diagnosed as papillomas. At Great Herring 

Pond, two fish were reported to have raised lesions; however, 

samples were not taken from either fish.

Figure 8. Prevalences of large (greater than 6 millimeters) 

raised oral and body-surface lesions in brown bullhead (Ameiurus 

nebulosus) from Great Herring, Santuit, and Ashumet Ponds, 

Cape Cod, Massachusetts, May–July 2002.

Figure 9. Prevalences of multiple external lesions and melanistic 

patches in brown bullhead (Ameiurus nebulosus) from Great 

Herring, Santuit, and Ashumet Ponds, Cape Cod, Massachusetts, 

May–July 2002.

Table 5. Percentage by location of brown bullhead (Ameiurus 

nebulosus) having barbel abnormalities, including missing, 

shortened, and knobbed barbels, Ashumet, Santuit, and Great 

Herring Ponds, southeastern Massachusetts, May–July 2002.

Location Missing 

barbels

Shortened 

barbels

Knobbed 

barbels

Ashumet Pond 6.9 22.4 22.4

Santuit Pond 23.7 28.8 10.2

Great Herring Pond 14.3 20.0 8.6

0

10

20

30

40

50

P

Great Herring Pond Santuit Pond Ashumet Pond REV

ALENCE OF LARGE RAISED LESIONS, IN PERCENT

Oral lesions

Body-surface lesions

STUDY-AREA LAKES

0

10

20

30

40

50

60

70

80

Great Herring 

Pond

Santuit

Pond 

Ashumet

Pond 

PREV

ALENCE, IN PERCENT

Multiple external lesions

Melanistic patches

STUDY-AREA LAKES
Prevalence of Tumors in Brown Bullhead 15

200 μm

200 μm

200 μm 100 μm

200 μm

200 μm

A

a

B

a

C

a

D

a

a

a

b

E

a

a

F

a

Figure 10. Selected microscopic findings in tissues of brown bullhead (Ameiurus nebulosus) from Ashumet 

Pond, Cape Cod, Massachusetts, May 2002. (A) Hyperplastic skin lesion with a proliferation of epidermal 

cells, primarily alarm substance cells (a). (B) Papilloma of the body surface. The proliferating cells (a) do 

not invade beyond the basement membrane (arrow). (C) Squamous cell carcinoma from the oral cavity. 

Proliferating cells (a) do invade through the basement membrane, and nests of neoplastic cells can be 

seen within connective tissue (arrow). (D) Section of a barbel with trematodes (a) within the nerve tissue 

underlying a papillomatous skin lesion (b). (E) Cholangioma (a) within the liver. Misshapen bile ducts with 

proliferating epithelium (arrows) are observed but are not invading into the surrounding liver. (F) Area of 

chronic inflammation (a) and multinucleate giant cells (arrows) within hepatic tissue in response to helminth 

parasites. Hematoxylin and eosin stain.
16 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Ashumet Pond but was not observed in fish from the other two 

lakes. Macrophage aggregates and parasites were observed in 

fish from all the lakes. The number of macrophage aggregates 

in the liver, spleen, and kidney did not vary consistently 

among sites. Heavy infections of immature cestodes were 

found in almost all fish. Often these parasites were associated 

with an intense inflammatory response and multinucleate giant 

cells (fig. 10F). Bile duct myxozoans were found in some fish 

at every site (37.5 percent at Ashumet Pond, 25 percent at 

Santuit Pond, and 20 percent at Great Herring Pond) and may 

be related to bile duct proliferation.

Hepatosomatic Index

A variety of investigators have noted a positive 

correlation between exposure to PAHs and the ratio of the 

liver weight to body weight (hepatosomatic index, or HSI) 

in fish (Fabacher and Baumann, 1985; Pinkney and others, 

2001). The relationship has been most striking at very high 

levels of PAH contamination, however, and is less obvious 

for intermediate levels of contamination (such as in Ashumet 

Pond), where other factors affecting liver size (such as 

nutrition, disease, and seasonal vitellogenin synthesis) may 

play a more dominant role (Yang and Baumann, 2006). The 

HSI was significantly greater for fish from Ashumet Pond 

than for fish from Great Herring and Santuit Ponds (α = 0.05, 

Tukey’s studentized range test); the indexes for fish from the 

latter two lakes had similar HSI values (table 7). The range of 

values from these sites also indicates that the brown bullhead 

population in Ashumet Pond had a higher proportionate liver 

mass (table 7).

DNA Damage in Red Blood Cells

The Comet assay parameters—tail length, percentage of 

tail DNA, and tail-extent moment—did not differ significantly 

with fish length (GLM, p > 0.05). Although female and male 

fish from Ashumet and Great Herring Ponds differed in the 

percentage of tail DNA (GLM, p < 0.05), they did not differ in 

tail length and tail-extent moment (GLM, p > 0.10). Therefore, 

the age (with length as a surrogate) and gender of fish did not 

seem to exert a major effect on the Comet assay results. All 

three of the measurements of DNA damage were significantly 

greater in fish from Ashumet Pond than in fish from Great 

Herring Pond (GLM, p < 0.05) (table 8). For fish from the two 

ponds, logistic regression analysis demonstrated that the tail 

length measured in the Comet assay was positively related to 

the odds ratio of fish having external raised lesions 

(ln[P(x)/(1–P(x))] = -10.75 + 0.2322 × tail length, Wald’s 

test, p < 0.05).

Table 7. Hepatosomatic index (liver weight divided by body 

weight) of brown bullhead (Ameiurus nebulosus) from Ashumet, 

Santuit, and Great Herring Ponds, southeastern Massachusetts, 

May–July 2002.

Statistic Ashumet Pond Santuit Pond Great Herring Pond

Mean 0.0242 0.0156 0.0152

Standard error .00117 .00066 .00078

Maximum .042 .022 .024

Minimum .016 .011 .011

Barbels from fish taken from Ashumet Pond contained 

metacercariae of digenetic trematodes along and within the 

nerve bundles (fig. 10D). These organisms were associated 

with hemorrhage and inflammation within the dermis and 

along and within the nerve fibers. Papillomas, squamous cell 

carcinomas, and dysplasia of the pseudocartilage were also 

observed in the barbels.

Histopathology of the Liver, Spleen, and Kidney

Regeneration of liver cells, altered cell foci, and 

neoplasia were all found more often in brown bullhead from 

Ashumet Pond than from the other two lakes (table 6). Most 

of the neoplasms were of bile duct origin—cholangioma 

(benign) or cholangiocarcinoma (malignant) (fig. 10E). One 

fish collected from Ashumet Pond had a malignant hepatic 

cell carcinoma, and one fish collected from Santuit Pond had 

a benign hepatic cell adenoma. Vacuolation of the intrahepatic 

exocrine pancreas was observed in 25 percent of the fish from 

Table 6. Percentage of necropsied brown bullhead (Ameiurus 

nebulosus) having selected microscopic lesions observed in liver 

tissue, Ashumet, Santuit, and Great Herring Ponds, southeastern 

Massachusetts, May–July 2002.

Lesion type Ashumet 

Pond

Santuit 

Pond

Great Herring 

Pond

Liver cell regeneration 45.8 0.0 15.0

Exocrine pancreas 

vacuolation

25.0 .0 .0

Bile duct hyperplasia 50.0 60.0 60.0

Altered cell foci 37.5 25.0 15.0

Neoplasia 16.7 15.0 5.0
Discussion 17

Discussion

External raised lesions were significantly more common in brown bullhead from Ashumet and Santuit Ponds 

than in brown bullhead from Great Herring Pond (table 4). 

Ashumet Pond was the primary target site because previous 

fish surveys had found an unusually high prevalence of raised 

lesions on brown bullhead captured there (HAZWRAP, 1995, 

1996; Stegeman, 1997). Great Herring and Santuit Ponds were 

sampled as reference sites; therefore, the elevated prevalence 

of raised lesions at Santuit Pond was unexpected. Comparison 

with the prevalence of raised lesions recorded in fish from 

Great Lakes locations (fig. 11) indicates that fish from both 

Ashumet and Santuit Ponds are at the high end of the range 

of raised-lesion prevalence seen in fish from contaminated 

locations, with prevalences close to the levels of occurrence 

documented in fish collected in Hamilton Harbour, Ontario, 

Canada (Smith and others, 1989) in the mid-1980s and in 

Presque Isle Bay, Penn. (Obert, 1994) in the early 1990s. 

Similarly, brown bullhead from Great Herring Pond mimic the 

raised-lesion prevalence between 2 and 4 percent seen in fish 

from Great Lakes reference locations (fig. 11). Of the 16 external raised lesions submitted for histopathology from Ashumet 

and Santuit Ponds fish, 15 (or 94 percent) were diagnosed as 

neoplasms. This result provides a level of confidence for the 

comparison in figure 11.

Fish age is correlated with the prevalences of both internal tumors (Baumann and others, 1990; Pinkney and others, 

2004) and external raised lesions (Maccubbin and Ersing, 

1991; Pinkney and others, 2004). Thus, tumor prevalences are 

best compared among fish of the same or similar age groups. 

The estimated ages of some fish from both Ashumet Pond and 

Great Herring Pond were only 3 or 4 years, despite the fact 

that the fish were very large. These data may indicate that the 

age was underestimated, as compression and loss of annuli are 

a well-documented problem in determining the ages of older 

fish by examination of spines. Future studies should collect 

otoliths as well as spines for aging. Nevertheless, the order-ofmagnitude difference in the prevalence of raised lesions and 

the levels of significance achieved because of the numbers of 

Table 8. Red blood cell nucleic DNA damage as determined by the Comet assay for brown bullhead (Ameiurus nebulosus) from 

Ashumet and Great Herring Ponds, southeastern Massachusetts, May–July 2002. 

[Values are means with standard deviation in parentheses. DNA, deoxyribonucleic acid; μm, micrometer]

Location Number of bullhead 

sampled

Tail length of migrated DNA

(μm)

DNA in tail relative to total 

DNA in image

(percent)

Tail-extent moment

(μm)

Ashumet Pond 10 46.95 (3.11) 19.22 (1.90) 10.27 (1.34)

Great Herring Pond 11 40.89 (2.90) 15.28 (1.64) 7.06 (1.03)

fish examined in this study leave no doubt that external raised 

lesions are much more common in brown bullhead from both 

Ashumet and Santuit Ponds than would be expected.

The prevalences of shortened and knobbed barbels were 

not significantly different in fish collected from the three 

lakes. Fish from Santuit Pond had the highest occurrence of 

missing barbels; of the three barbel anomalies, however, this 

abnormality is the least connected with chemical exposure and 

often results from physical stress. Therefore, barbel deformities were not useful in characterizing potential contaminant 

exposure among fish in these lakes.

The histopathology of the liver, although done by using 

a small number of fish from each location (N = 20 to 24), 

showed a prevalence of neoplasms (mainly of bile duct origin) 

15 percent or higher for both Ashumet and Santuit Ponds. This 

prevalence is comparable to prevalences measured in fish from 

a number of Great Lakes Areas of Concern with contaminantrelated tumor epizootics and is greater than the 5-percent 

prevalence level thought to indicate a reference condition 

(Baumann and others, 1996). Because the only known cause 

of liver tumors in fish is chemical carcinogens, these data, 

combined with the external-tumor data, indicate a role for 

carcinogenic contaminants in the tumor pathology of brown 

bullhead in Ashumet and Santuit Ponds.

The Comet assay was used to quantify genetic damage 

and provide an index of exposure to tumor-inducing chemicals 

in the brown bullhead population. This method can be used 

with any age class of fish and provides a measure of genetic 

damage at the time of animal capture. Logistic regression indicated that higher DNA damage measured by the Comet assay 

was associated with higher prevalence of external lesions. The 

results also indicated that DNA damage was greater in brown 

bullhead from Ashumet Pond than in bullhead from Great 

Herring Pond (table 8). The data from the present study, along 

with a previous analysis of brown bullhead collected at four 

other locations (tributaries to Lake Erie), indicate that DNA 

damage does not differ significantly with fish length (Yang 

and others, 2006). This observation indicates that the greater 

DNA damage observed in brown bullhead from Ashumet Pond 

than in bullhead from Great Herring Pond is not confounded 

by differences in the ages of fish collected at the two locations. 
18 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

0

20

40

60

80

100

A

S

H

BLA

B

UF

C

U

Y

H

A

M

PIB

B

U

C

M

U

N

O

W

C84

O

W

C88

A

N

A

T

U

C

A

S

HP

S

A

NP

G

HP

PREV

ALEN

CE OF EX

TER

N

AL LESIO

N

S, IN PERCENT

Eastern 

sites 

Cape Cod 

sites 

Reference sites

Contaminated sites

Cape Cod sites

Great Lakes

sites 

Figure 11. Prevalence of raised external lesions in brown bullhead (Ameiurus nebulosus) from tributaries of the Great 

Lakes, rivers in the eastern United States and Canada, and Ashumet, Santuit, and Great Herring Ponds on Cape Cod, 

Massachusetts. Sample sizes ranged from 48 to 179 fish, with a mean sample size of more than 100 fish per location. 

ASH, Ashtabula River, Ohio (Mueller and Mac, 1994); BLA, Black River, Ohio (Baumann and others, 1996); BUF, Buffalo 

River, New York (Baumann and others, 1996); CUY, Cuyahoga River, Ohio (P.C. Baumann, unpublished data); HAM, Hamilton 

Harbour, Ontario, Canada (Smith and others, 1989); PIB, Presque Isle Bay, Pennsylvania (Obert, 1994); BUC, Buckeye Lake, 

Ohio (P.C. Baumann, unpublished data); MUN, Munuscong Bay, Michigan (Baumann and others, 1991); OWC84, Old Woman 

Creek, Ohio, 1984–85 (Baumann and others, 1996); OWC88, Old Woman Creek, Ohio, 1988 (P.C. Baumann, unpublished data); 

ANA, Anacostia River, Washington, D.C. (Pinkney and others, 2004); TUC, Tuckahoe River, Maryland (Pinkney and others, 

2004); ASHP, Ashumet Pond; SANP, Santuit Pond; GHP, Great Herring Pond.
Discussion 19

These results imply that fish from Ashumet Pond may have 

been exposed to higher levels of genotoxins than have fish 

from Great Herring Pond.

Several studies employing the Comet assay have examined the relation between the extent of induced DNA damage 

and the time elapsed after the treatment of fish with genotoxic 

chemicals. Pandrangi and others (1995) reported increased 

damage to erythrocytes (red blood cells) from brown bullhead 

within 6 hours following a single intraperitoneal injection 

of cyclophosphamide; the response peaked at 48–72 hours. 

Belpaeme and others (1998) observed increased DNA damage 

to erythrocytes in marine flatfish (turbot) (Psetta maximus) 

exposed by static renewal to ethyl methane sulfonate (EMS) in 

water; the response in several tissues was higher after 7 days 

of exposure than after 3 days of exposure. Belpaeme and others (1998) also reported an increasing response with increasing 

exposure for brown trout (Salmo trutta fario) for up to 14 days 

after static renewal to EMS in water. These studies show that 

the Comet assay provides a relatively rapid indicator of exposure. Although there are no known long-term studies of the 

persistence of DNA damage in fish erythrocytes, these results 

and the fact that the lifespan of erythrocytes is probably only 

about 100 days (based on erythrocyte kinetics in other fish 

species) suggest that there is ongoing exposure of the brown 

bullhead in Ashumet Pond to genotoxic substances.

Although causation was not investigated during this 

study, some discussion is warranted, particularly because 

causation remains elusive without further research. External 

tumors are known to have a viral etiology in several species 

of fish, including white sucker (Catostomus Commersonii) 

(Baumann and Okihiro, 2000), but experimentation to date 

has not demonstrated a viral etiology in brown bullhead. 

Painting of brown bullhead with sediment extract containing 

PAHs from the Buffalo River, N.Y., produced skin tumors at a 

38-percent prevalence after 2 years (Black and others, 1985). 

Similarly, epizootics of neoplasia in wild fish have been 

associated with contaminants (primarily PAHs) in freshwater, 

marine, and estuarine habitats in more than 20 locations in 

North America alone (Baumann, 1998). Even tumors having 

a partial viral etiology, such as papillomas on white suckers, 

have been more prevalent in populations from contaminated 

areas (Smith and others, 1989; Premdas and others, 1995). 

This observation indicates that tumor viruses may be activated 

at times of immune system dysfunction or inhibition caused 

by carcinogenic or immune-system suppressing compounds 

(Anders and Yoshimizu, 1994). Etiologies of such tumors 

would therefore be multifactoral (Premdas and Metcalfe, 

1996). Similarly, parasite infections could cause tissue damage 

and thereby stimulate cell replication and increase the chances 

of tumor progression.

For Ashumet Pond, both the known PAH concentrations 

in sediment and PCE and TCE levels in water are far below 

concentrations known to induce tumors or tumor epizootics. It 

must be noted, however, that few studies using PCE and TCE 

have been conducted on fish. One study, involving ground 

water contaminated primarily with TCE (Gardner and others, 

1998), found the ground water to be a tumor promoter but not 

a tumor initiator, although reagent-grade TCE did not have any 

tumor promoter effects. Gardner and others (1998) concluded 

that, although chemical analysis of the ground water found 

TCE to be the only reportable contaminant, other compounds 

below reportable limits may have had a synergistic effect on 

tumor promotion. Different mixes of chemical compounds can 

cause different suites of pathology (Yang and others, 2002).

The population pathology profiles differed for brown 

bullhead from Ashumet and Santuit Ponds. Large oral and 

medium body-surface lesions were more than twice as 

common, and large raised lesions on the body were more than 

four times as common, in Ashumet Pond fish than in Santuit 

Pond fish. These differences could be caused by the apparent 

younger age distribution of the Santuit Pond fish, or they also 

could be caused by differences in exposure to contaminant 

mixtures. The HSI for Ashumet Pond fish was greater than 

that for Santuit Pond fish. Similarly, dark skin patches were 

observed on almost two-thirds of the fish from Ashumet 

Pond, while none were seen on fish from Santuit Pond. Such 

melanistic patches have been associated with fish exposed to 

PAHs (Black, 1983; Black and others, 1985; Maccubbin and 

Ersing, 1991).

The high prevalence of melanistic lesions on Ashumet 

Pond brown bullhead, combined with the tumor pathology 

and genetic damage, implicates chemical carcinogens as one 

of the causal factors in that lake. Because many of the brown 

bullhead were very large and ages may have been underestimated, chemical exposure contributing to the pathology may 

have occurred as long ago as the early 1990s. Although there 

is no direct evidence of a link to contaminants from the MMR, 

remedial actions begun in the early 1990s have reduced and 

will continue to reduce the discharge of contaminants from 

the drainage system and ground-water plumes to Ashumet and 

Johns Ponds. The current (2008) brown bullhead population 

probably consists mostly of new individuals born since the 

2002 sampling and the beginning of remediation efforts.

For these reasons, future studies could repeat the prevalence work at Ashumet and Santuit Ponds and at least one 

other reference lake to determine whether the pathology 

patterns have changed. The work could include, for at least 

50 fish per lake, (1) measurement of length and weight, 

(2) recording of external lesions and preservation of samples 

(including all skin, oral, and barbel abnormalities) for histopathology, (3) determination of sex, (4) determination of 

age from analysis of otoliths and scales, and (5) preservation 

of liver samples for histopathology. In addition, for at least 

20 fish per lake, the analysis could include (1) collection of 

bile for determination of PAH metabolites, (2) collection of 

samples from the kidneys and spleen for determination of 

macrophage aggregates, (3) preservation of liver samples for 

DNA adduct analysis, and (4) collection of blood samples 

from a subset of younger (1–2 years) and older (greater than 

3 years) fish for Comet assays.

Additional studies could be conducted on the factors that 

may influence the elevated prevalence of neoplasms in brown 
20 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

bullhead from Ashumet and Santuit Ponds on the basis of the 

assumption that the causal factors may differ between the two 

lakes. These studies could include (1) environmental sampling 

of contaminants of concern in the lake water and sediments, 

(2) cage studies in Ashumet Pond to test the pathological 

response of brown bullhead collected from a reference lake to 

exposure for 6–8 weeks to Ashumet Pond water and sediment, 

and (3) laboratory studies to examine possible synergistic 

effects of the combined exposure of brown bullhead to PAHs 

and volatile organic compounds.

Summary

Brown bullhead were sampled and examined for pathology from three lakes in southeastern Massachusetts—Ashumet, Santuit, and Great Herring Ponds. Fish from Great 

Herring Pond had low levels of raised lesions and low levels 

of genetic damage to red blood cell nuclei. Brown bullhead 

from Ashumet Pond, which has been subjected to contamination from the Massachusetts Military Reservation, had a high 

prevalence of raised lesions, which included histopathologically verified papillomas and squamous cell carcinoma, an 

elevated incidence of liver neoplasms, and an elevated level 

of genetic damage to red blood cell nuclei. Because red blood 

cells in fish have a lifespan of about 100 days, these results 

indicate an ongoing exposure to genotoxins in Ashumet Pond. 

Brown bullhead from Santuit Pond also had elevated prevalences of raised lesions and liver neoplasms, although the 

prevalences of large and multiple lesions were significantly 

lower than that in Ashumet Pond fish. These differences, along 

with additional differences in internal pathology, may point to 

differing causes of the raised lesions in the two lakes. The high 

prevalence of melanistic lesions on Ashumet Pond brown bullhead, combined with the tumor pathology and genetic damage, 

implicates chemical carcinogens as one of the causal factors 

in that lake. Because many of the brown bullhead were large 

and ages may have been underestimated, chemical exposure 

contributing to the pathology may have occurred as long ago 

as the early 1990s. An additional prevalence survey would 

help to clarify whether the causal factors are still active.

Acknowledgments

The authors would like to acknowledge the assistance of 

Daniel Peterson and Michael Rowan of Ohio State University; Andrew Massey, Joshua Koch, and Caroline Olson of 

the U.S. Geological Survey; and Ed Kraus, Jeff Breton, and 

Salvatore Paterno of the Massachusetts Division of Fisheries 

and Wildlife for their assistance in the field. We also thank the 

members of the Technical Review and Evaluation Team and 

Spence Smith of the Air Force Center for Engineering and the 

Environment for their advice and support during the design of 

the study, and Kathy Spring of the U.S. Geological Survey for 

preparation of the histopathology slides.

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Schmitt, C.J., Blazer, V.S., Dethloff, G.M., Tillitt, D.E., Gross, 

T.S., Bryant, W.L., DeWeese, L.R., Smith, S.B., Goede, 

R.W., Bartish, T.M., and Kubiak, T.J., 1999, Biomonitoring 

of environmental status and trends (BEST) program—field 

procedures for assessing the exposure of fish to environmental contaminants: U.S. Geological Survey Information 

and Technology Report USGS/BRD-1999-0007, 68 p.

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Canada: Ottawa, Fisheries Research Board of Canada, 

Canada Bulletin 184, 966 p.

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1988, A simple technique for quantitation of low levels 

of DNA damage in individual cells: Experimental Cell 

Research, v. 175, p. 184–191.

Smith, I.R., Ferguson, H.W., and Hayes, M.A., 1989, Histopathology and prevalence of epidermal papilloma epidemic in 

brown bullhead, Ictalurus nebulosus (Lesueur), and white 

sucker, Catostomus sommersoni (Lacepede) populations 

from Ontario, Canada: Journal of Fish Diseases, v. 12, 

p. 373–388.

Stegeman, J.J., 1997, Biomarkers in fish from Cape Ponds, in

Air Force Center for Environmental Excellence (AFCEE), 

Fish health in the upper Cape Cod ponds near the Massachusetts Military Reservation, Plume Response Program, resource document: Otis Air National Guard Base, 

prepared for AFCEE by the Jacobs Engineering Group, 

Inc., AFC-J23-35K78412-M18-0001, July 1997, variously 

paged.

Tice, R.R., 1995, The single cell gel/Comet assay—a microgel 

electrophoretic technique for the detection of DNA damage 

and repair in individual cells, in Phillips, D.H., and Venitt, 

S., eds., Environmental mutagenesis: Oxford, United 

Kingdom, Bios Scientific Publishers, Ltd., p. 315–339.

U.S. Army Environmental Command (USAEC), 2007a, 

Impact Area Groundwater Study Program, overview and 

update—transitioning to cleanup: Camp Edwards, Massachusetts Military Reservation, U.S. Army Environmental 

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January 18, 2008, at http://groundwaterprogram.army.mil/

community/facts/overview-2007.pdf

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Impact Area Groundwater Study Program, investigation and 

cleanup update: Camp Edwards, Massachusetts Military 

Reservation, U.S. Army Environmental Command, fact 

sheet, summer 2007, 32 p., accessed January 18, 2008, at 

http://groundwaterprogram.army.mil/community/facts/

MMR_Plume_Book_2007_vFINAL_HiRes.pdf

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Jennifer, 1996, Geochemical and hydrologic controls on 

phosphorus transport in a sewage-contaminated sand and 

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sources and effects of pumping on surface and ground 

water, Sagamore and Monomoy flow lenses, Cape Cod, 

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p. 163–175.
24 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

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P.C., 2006, DNA damage and external lesions in brown 

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v. 29, no. 1, p. 116–123.
Glossary

C

carcinoma Malignant neoplastic lesions 

with invasion into the surrounding tissue.

E

epizootic A disease that is prevalent among 

a group of animals.

etiology The cause of a disease or disorder.

H

histopathology Microscopic examination of 

tissue pathology.

L

lesion Any grossly visible or microscopic 

abnormality, including parasites, inflammation, reddened areas, eroded areas, and 

raised growths.

M

melanistic Abnormally darkened areas 

of skin.

N

neoplasm New, abnormal, unregulated, or 

uncoordinated growth of specific cell types.

O

otolith Bone that is part of the ear of 

the fish.

P

papilloma Benign neoplastic lesions 

composed of proliferating, but not invasive, 

epidermal cells.

pathology The study and diagnosis of 

disease through the examination of organs, 

tissues, bodily fluids, and whole bodies.

R

reference lake A lake believed to be 

“unpolluted” by the contaminants of concern 

that serves as an “uncontaminated” background site for comparison to the lake 

of interest.

T

tumor By strict definition, a raised growth, 

but often used interchangeably with cancer 

or neoplasm.
THIS PAGE INTENTIONALLY LEFT BLANK
Appendixes 1 and 2
28 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002 Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued [Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data] Fish No. Date examined Necropsy sample Length (mm) Melanistic patches No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels <3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Ashumet Pond

1 05/21/02

Y 390

N 0 5 3 0 8 15 0 0 0 0 0 0 0 0 0

2 05/21/02

Y 380

N 0 0 3 0 0 0 0 0 0 0 0 0 0 0 0

3 05/21/02

Y 365

Y 0 0 0 0 0 0 0 0 1 0 0 0 0 0 0

4 05/21/02

Y 375

Y 0 2 0 0 0 1 0 0 0 0 0 0 0 0 0

5 05/21/02

Y 390

N 0 0 0 0 0 0 0 0 0 0 0 0 1 0 1

6 05/21/02

Y 420

Y 0 0 1 0 0 0 0 0 0 0 0 0 1 0 0

7 05/21/02

Y 355

Y 0 1 8 0 0 14 0 0 0 0 0 0 0 0 0

8 05/21/02

Y 380

Y 9 2 1 0 1 0 0 0 0 0 0 0 0 0 0

9 05/21/02

Y 380

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

10 05/21/02

Y 400

Y 3 1 5 0 0 0 0 1 0 2 0 1 1 0 4

11 05/22/02

Y 330

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

12 05/22/02

Y 410

N 0 1 3 0 2 13 0 0 0 1 0 1 0 0 0

13 05/22/02

Y 360

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

14 05/22/02

Y 380

N 2 0 0 0 0 0 0 0 0 0 0 0 0 0 0

15 05/22/02

Y 380

Y 0 1 1 0 0 0 0 0 0 0 0 0 0 0 0

16 05/22/02

Y 390

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

17 05/22/02

Y 420

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

18 05/22/02

Y 365

N 0 1 5 0 1 0 0 0 0 0 0 0 0 0 0

19 05/22/02

Y 375

Y 0 3 10 1 0 1 0 0 0 0 0 0 0 0 0

20 05/22/02

Y 400

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

21 05/22/02

Y 385

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

22 05/22/02

Y 360

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

23 05/22/02

Y 410

N 0 1 1 0 0 0 1 0 0 0 0 0 0 0 0

24 05/22/02

Y 390

Y 3 3 0 0 2 7 0 0 0 0 0 0 0 0 0

25 05/23/02

N 405

Y 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0

26 05/23/02

N 370

Y 0 1 0 0 0 6 0 0 1 0 0 0 0 0 0

27 05/23/02

N 350

Y 0 0 0 1 0 1 0 0 0 0 0 0 0 0 0

28 05/23/02

N 345

Y 0 0 2 0 0 0 1 0 2 0 0 0 0 0 0

29 05/23/02

N 350

Y 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0

30 05/23/02

N 400

N 0 0 1 0 0 0 0 0 0 0 0 0 0 0 0
Appendix 1 29

Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued[Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data]

Fish 

No.

Date 

examined

Necropsy 

sample

Length 

(mm)

Melanistic 

patches

No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels

<3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed

Ashumet Pond—Continued

31 05/23/02 N 390 Y 0 0 3 2 0 0 0 0 0 0 0 0 0 0 032 05/23/02 N 385 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 033 05/23/02 N 410 Y 0 1 1 1 0 5 0 0 0 0 0 0 0 0 034 05/23/02 N 370 N 0 0 0 0 0 1 0 0 0 0 0 1 0 0 035 05/23/02 N 395 Y 0 0 2 0 1 1 0 0 0 0 0 0 0 0 036 05/23/02 N 395 Y 0 0 0 2 1 0 0 0 0 1 0 1 0 0 137 05/23/02 N 400 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 038 05/23/02 N 335 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 039 05/23/02 N 350 Y 0 0 0 0 0 0 0 0 0 0 0 0 1 0 140 05/23/02 N 400 Y 0 0 0 0 1 0 0 0 0 0 0 0 0 0 041 05/23/02 N 340 N 0 1 6 1 2 11 0 0 0 2 0 0 0 0 042 05/23/02 N 360 Y 0 0 0 0 0 1 0 0 0 0 0 0 0 0 043 05/23/02 N 345 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 044 05/23/02 N 385 Y 1 1 4 1 0 6 0 0 0 0 0 0 0 0 045 05/23/02 N 395 N 2 6 8 3 2 10 0 0 0 0 0 0 1 0 146 05/23/02 N 330 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 047 05/23/02 N 375 Y 0 0 0 2 1 0 0 0 0 0 0 0 0 0 048 05/23/02 N 380 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 1 049 05/23/02 N 345 Y 1 0 1 0 0 1 1 0 1 0 0 0 0 0 050 05/23/02 N 390 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 051 05/23/02 N 385 Y 0 0 0 0 1 4 0 0 0 0 0 0 0 0 052 05/23/02 N 400 N 2 0 1 0 0 1 0 0 0 0 0 0 1 0 053 06/07/02 N 340 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 054 06/07/02 N 400 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 055 06/07/02 N 400 Y 0 0 0 1 0 0 0 0 0 0 0 1 2 0 256 06/07/02 N 375 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 057 06/07/02 N 400 N 0 1 1 0 1 2 0 0 0 0 0 0 0 0 058 06/07/02 N 385 Y 1 1 1 1 0 2 0 0 0 0 0 1 0 2 0
30 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002 Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued [Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data] Fish No. Date examined Necropsy sample Length (mm) Melanistic patches No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels <3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Santuit Pond

1 06/25/02

Y 298

N 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0

2 06/25/02

Y 301

N 0 1 0 0 0 0 1 0 0 0 0 0 0 0 0

3 06/25/02

Y 297

N 1 2 0 0 0 0 0 0 0 0 0 0 1 0 0

4 06/25/02

Y 300

N 0 0 0 2 0 1 0 0 0 0 0 0 0 0 0

5 06/26/02

Y 271

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

6 06/26/02

Y 302

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

7 06/26/02

Y 290

N 0 0 0 0 0 0 0 0 0 0 0 0 1 2 0

8 06/26/02

Y 272

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

9 06/27/02

Y 326

N 0 0 0 0 1 1 0 0 0 0 0 0 1 0 0

10 06/27/02

Y 309

N 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0

11 06/27/02

Y 328

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

12 06/27/02

Y 295

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

13 06/27/02

Y 281

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

14 06/27/02

Y 284

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

15 06/27/02

Y 290

N 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0

16 06/27/02

Y 303

N 3 3 3 3 2 0 1 0 0 0 0 0 0 1 0

17 06/27/02

Y 272

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

18 06/27/02

Y 280

N 0 0 4 0 0 0 0 0 0 0 0 0 0 0 0

19 06/27/02

Y 284

N 0 0 0 3 3 2 0 0 0 0 0 0 0 0 0

20 06/27/02

Y 285

N 1 3 0 0 0 0 0 0 0 0 0 0 0 0 0

21 06/27/02

N 255

N 0 0 0 0 0 1 0 0 0 0 0 0 0 0 0

22 06/27/02

N 288

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

23 06/27/02

N 275

N 0 0 0 0 0 0 1 0 1 0 0 0 0 0 0

24 06/27/02

N 275

N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0

25 06/27/02

N 280

N 0 0 0 0 2 3 0 1 0 0 0 0 0 0 0

26 06/27/02

N 285

N 0 0 0 0 0 1 0 0 0 0 0 0 0 1 0

27 06/27/02

N 282

N 0 0 0 1 0 0 0 0 0 1 0 0 0 0 0

28 06/27/02

N 271

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

29 06/28/02

N 287

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1

30 06/28/02

N 291

N 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0
Appendix 1 31

Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued[Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data]

Fish 

No.

Date 

examined

Necropsy 

sample

Length 

(mm)

Melanistic 

patches

No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels

<3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed

Santuit Pond—Continued

31 07/01/02 N 287 N 0 0 0 0 0 0 0 0 0 0 0 0 1 1 132 07/01/02 N 299 N 0 0 0 1 0 0 0 0 0 1 0 0 0 0 033 07/01/02 N 310 N 0 0 0 0 0 0 0 0 0 1 0 0 1 0 034 07/01/02 N 284 N 0 0 0 0 0 0 0 1 0 0 0 0 0 0 035 07/01/02 N 305 N 0 0 0 0 1 0 0 0 0 0 0 0 0 0 036 07/01/02 N 277 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 037 07/01/02 N 266 N 1 0 0 0 0 0 0 0 0 0 0 0 0 0 038 07/01/02 N 263 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 039 07/01/02 N 292 N 0 0 0 1 1 0 0 0 0 0 0 0 0 0 040 07/01/02 N 273 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 041 07/01/02 N 270 N 0 0 0 0 0 0 1 0 0 1 0 0 0 0 042 07/01/02 N 276 N 1 3 1 0 0 0 0 0 0 0 0 0 0 3 043 07/01/02 N 284 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 144 07/01/02 N 279 N 0 0 0 3 0 1 0 0 0 0 0 0 0 0 045 07/01/02 N 284 N 0 0 0 9 2 0 0 0 0 0 0 0 0 1 046 07/01/02 N 268 N 0 1 1 15 1 3 0 0 0 1 0 0 0 1 047 07/01/02 N 300 N 0 1 0 2 1 0 0 0 0 0 0 0 0 0 048 07/01/02 N 276 N 0 0 0 0 0 0 0 1 0 1 0 0 0 0 049 07/01/02 N 286 N 0 2 1 2 2 1 0 0 0 0 0 0 1 0 050 07/01/02 N 289 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 051 07/01/02 N 296 N 0 0 0 0 0 0 0 0 0 0 1 0 0 2 052 07/01/02 N 291 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 053 07/01/02 N 299 N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 054 07/01/02 N 276 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 055 07/01/02 N 313 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 056 07/01/02 N 284 N 3 0 0 0 0 0 0 0 0 1 0 0 0 0 057 07/01/02 N 274 N 0 0 0 0 0 0 0 0 0 0 0 1 0 0 058 07/01/02 N 265 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 059 07/01/02 N 273 N 0 0 0 0 0 0 0 0 0 0 0 1 0 1 0
32 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002 Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued [Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data] Fish No. Date examined Necropsy sample Length (mm) Melanistic patches No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels <3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Great Herring Pond

1 06/24/02

Y 340

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

2 06/24/02

Y 375

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

3 06/24/02

Y 280

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

4 06/24/02

Y 365

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

5 06/24/02

Y 351

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

6 06/24/02

Y 340

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

7 06/25/02

Y 370

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

8 06/25/02

Y 365

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

9 06/25/02

Y 370

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

10 06/25/02

Y 345

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

11 06/25/02

Y 380

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

12 06/25/02

Y 387

N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0

13 06/25/02

Y 341

N 0 0 1 0 0 0 0 0 0 0 0 0 0 1 0

14 06/25/02

Y 329

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

15 06/26/02

Y 349

Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

16 06/26/02

Y 365

N 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0

17 06/26/02

Y 365

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

18 06/26/02

Y 372

N 0 0 1 0 0 0 0 0 0 0 0 0 1 1 0

19 06/26/02

Y 279

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

20 06/26/02

Y 363

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

21 06/07/02

N 357

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

22 06/07/02

N 334

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

23 06/07/02

N 357

N 0 0 0 0 0 0 0 1 0 0 0 0 0 0 0

24 06/07/02

N 399

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

25 06/07/02

N --

N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0

26 06/07/02

N 275

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

27 06/07/02

N 403

N 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0

28 06/07/02

N 340

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

29 06/07/02

N 405

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

30 06/07/02

N 325

N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0
Appendix 1 33

Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued[Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data]

Fish 

No.

Date 

examined

Necropsy 

sample

Length 

(mm)

Melanistic 

patches

No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels

<3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed

Great Herring Pond—Continued

31 06/07/02 N 366 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 032 06/07/02 N 384 N 0 0 0 0 0 0 0 0 0 0 1 0 0 0 033 06/07/02 N 374 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 034 06/07/02 N 351 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 035 06/07/02 N 367 N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 136 06/07/02 N 328 N 0 0 0 0 0 0 1 0 0 0 0 0 0 0 037 06/07/02 N 302 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 238 06/07/02 N 293 N 0 0 0 0 0 0 0 0 0 1 0 0 0 0 039 06/07/02 N 306 N 0 0 0 0 0 0 0 0 0 0 0 0 1 0 040 06/07/02 N 371 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 041 06/07/02 N 357 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 042 06/07/02 N 350 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 043 06/07/02 N 335 N 0 0 0 0 0 0 1 0 0 0 0 0 0 0 044 06/07/02 N 340 N 0 0 0 0 0 0 0 0 0 0 0 0 2 0 045 06/07/02 N 317 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 046 06/07/02 N 356 N 0 0 0 0 0 0 0 0 0 1 0 0 0 0 047 06/07/02 N 358 N 0 0 0 0 0 0 0 1 0 0 0 0 0 1 148 06/07/02 N 270 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 049 06/07/02 N 347 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 150 06/07/02 N 374 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 051 06/07/02 N 391 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 052 06/07/02 N 390 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 053 06/07/02 N 306 N 0 0 0 0 0 0 0 0 0 0 0 0 1 0 054 06/07/02 N 375 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 055 06/07/02 N 345 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 056 06/07/02 N 386 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 157 06/07/02 N 353 N 0 0 0 0 0 0 0 0 0 1 0 0 0 0 058 06/07/02 N 330 N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 059 06/07/02 N 366 Y 0 0 0 0 0 0 0 0 0 0 0 0 0 0 160 06/07/02 N 347 N 0 0 0 0 0 0 1 0 0 0 0 0 0 0 0
34 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002 Appendix 1. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring Ponds, southeastern Massachusetts, and examined externally in the field, May–July 2002.—Continued [Locations of lakes are shown in figure 1. Data from necropsy sample are shown in appendix 2. No., number; mm, millimeters; <, less than value shown; >, greater than value shown; N, no; Y, yes; --, no data] Fish No. Date examined Necropsy sample Length (mm) Melanistic patches No. of body-surface lesions No. of oral lesions Nasal barbels Maxillary barbels Chin barbels <3 mm 3–6 mm >6 mm <3 mm 3–6 mm >6 mm Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Short- ened Miss- ing Knob- bed Great Herring Pond—Continued 61 06/07/02

N 332

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

62 06/07/02

N 357

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

63 06/07/02

N 334

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

64 06/07/02

N 360

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

65 06/07/02

N 298

N 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0

66 06/07/02

N 332

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

67 06/07/02

N 386

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

68 06/07/02

N 297

N 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0

69 06/07/02

N 380

N 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

70 06/07/02

N 341

N 0 0 0 0 0 0 0 0 0 0 0 0 0 1 0
Appendix 2 35

Appendix 2. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring 

Ponds, southeastern Massachusetts, and necropsied for internal examination and tissue sampling, May–June 2002.—Continued

[Locations of lakes are shown in figure 1. Data from external examination are shown in appendix 1. No., number; mm, millimeters; g, grams; yr, year; 

HSI, hepatosomatic index; M, male; F, female; --, no data]

Fish No. Date 

necropsied

Length 

(mm)

Weight 

(g) Sex Estimated age (yr)

Liver weight 

(g) HSI Gonads weight (g)

Spleen weight 

(g)

Ashumet Pond

1 05/21/02 390 865 M 6 15.25 0.018 2.63 1.33

2 05/21/02 380 905 M 4 23.38 .026 2.37 1.48

3 05/21/02 365 820 F 3 22.75 .028 72.30 1.06

4 05/21/02 375 963 M 6 27.75 .029 2.94 1.80

5 05/21/02 390 947 M 4 22.98 .024 3.41 1.29

6 05/21/02 420 1,095 M 7 23.43 .021 2.99 1.57

7 05/21/02 355 694 F 5 28.90 .042 14.93 1.49

8 05/21/02 380 917 F 5 24.56 .027 76.33 1.67

9 05/21/02 380 932 F 6 24.29 .026 75.22 1.32

10 05/21/02 400 1,132 M 11 18.58 .016 3.69 1.75

11 05/22/02 330 668 F 7 13.53 .020 68.59 0.67

12 05/22/02 410 1,045 M 7 18.60 .018 4.27 1.19

13 05/22/02 360 780 M 5 17.10 .022 2.34 1.47

14 05/22/02 380 1,000 F 4 32.79 .033 79.32 1.24

15 05/22/02 380 975 M 4 23.40 .024 2.75 1.12

16 05/22/02 390 1,023 M 7 22.84 .022 3.43 2.13

17 05/22/02 420 1,120 M 8 24.29 .022 3.58 1.75

18 05/22/02 365 851 F 6 26.50 .031 69.92 2.87

19 05/22/02 375 1,002 F 7 26.12 .026 108.65 2.18

20 05/22/02 400 931 M 3 22.24 .024 3.61 1.56

21 05/22/02 385 905 M 3 17.80 .020 2.02 1.46

22 05/22/02 360 768 M 3 14.20 .018 2.09 .81

23 05/22/02 410 1,112 M 5 25.89 .023 4.18 2.21

24 05/22/02 390 871 M 9 17.71 .020 1.87 2.36

Santuit Pond

1 06/25/02 298 376 M 4 4.97 .016 1.38 .63

2 06/25/02 301 380 M 6 7.50 .020 0.86 .54

3 06/25/02 297 351 M 4 4.17 .012 1.13 .33

4 06/25/02 300 342 M 3 4.61 .013 .42 .40

5 06/26/02 271 306 F 5 5.10 .017 36.71 .35

6 06/26/02 302 357 M 4 4.70 .013 1.33 .62

7 06/26/02 290 300 F 3 4.46 .015 32.13 .31

8 06/26/02 272 262 F 3 5.19 .020 -- .33

9 06/27/02 326 481 M 7 9.08 .019 1.16 .73

10 06/27/02 309 409 M 3 6.22 .015 .89 .68
36 Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts, 2002

Appendix 2. Selected characteristics of brown bullhead (Ameiurus nebulosus) collected from Ashumet, Santuit, and Great Herring 

Ponds, southeastern Massachusetts, and necropsied for internal examination and tissue sampling, May–June 2002.—Continued

[Locations of lakes are shown in figure 1. Data from external examination are shown in appendix 1. No., number; mm, millimeters; g, grams; yr, year; 

HSI, hepatosomatic index; M, male; F, female; --, no data]

Fish No. Date 

necropsied

Length 

(mm)

Weight 

(g) Sex Estimated age (yr)

Liver weight 

(g) HSI Gonads weight (g)

Spleen weight 

(g)

Santuit Pond—Continued

11 06/27/02 328 508 M 7 11.43 0.022 1.51 0.89

12 06/27/02 295 353 F 4 5.87 .017 33.10 .45

13 06/27/02 281 285 F 4 4.22 .015 3.03 .38

14 06/27/02 284 315 F 4 5.95 .019 3.05 .50

15 06/27/02 290 348 M 6 5.18 .015 0.73 .52

16 06/27/02 303 378 M 3 4.07 .011 .88 .42

17 06/27/02 272 288 F 6 4.56 .016 3.69 .44

18 06/27/02 280 357 M 7 5.20 .015 .67 .53

19 06/27/02 284 363 M 6 4.78 .013 .77 .59

20 06/27/02 285 300 F 3 4.07 .014 5.62 .37

Great Herring Pond

1 06/24/02 340 551 M 3 8.77 .016 1.86 .51

2 06/24/02 375 820 M 7 10.23 .012 3.74 1.95

3 06/24/02 280 320 M 2 6.27 .020 .19 .59

4 06/24/02 365 737 M 3 8.55 .012 2.36 1.29

5 06/24/02 351 590 M 3 9.25 .016 2.48 1.53

6 06/24/02 340 558 F 3 9.10 .016 50.77 2.45

7 06/25/02 370 709 F 6 8.12 .011 77.14 .73

8 06/25/02 365 704 M 3 9.54 .014 1.20 1.31

9 06/25/02 370 706 M 3 9.59 .014 1.93 .83

10 06/25/02 345 534 M 3 7.82 .015 1.10 .58

11 06/25/02 380 769 M 8 18.33 .024 2.53 1.72

12 06/25/02 387 900 M 7 13.67 .015 2.28 1.83

13 06/25/02 341 538 M 3 6.63 .012 1.06 1.20

14 06/25/02 329 508 F 3 11.21 .022 22.65 .48

15 06/26/02 349 675 F 6 8.98 .013 77.56 .79

16 06/26/02 365 675 M 4 10.74 .016 2.68 1.23

17 06/26/02 365 663 M 3 8.21 .012 1.90 1.24

18 06/26/02 372 665 F 7 9.00 .014 46.93 .51

19 06/26/02 279 327 F 2 6.03 .018 42.65 .41

20 06/26/02 363 663 M 3 8.47 .013 2.22 .98
Prepared by the Pembroke Publishing Service Center.

For more information concerning this report, contact:

Director

U.S. Geological Survey

Massachusetts-Rhode Island Water Science Center

10 Bearfoot Road

Northborough, MA 01532

dc_ma@usgs.gov

or visit our Web site at:

http://ma.water.usgs.gov
Printedon recycled paper

Baumann and others—Prevalence of Tumors in Brown Bullhead from Three Lakes in Southeastern Massachusetts—Scientific Investigations Report 2008–5198