CATALOG DOCUMENTATION
EMAP-ESTUARIES PROVINCE LEVEL DATABASE
CAROLINIAN PROVINCE 1994-1997
SEDIMENT TOXICITY DATA

TABLE OF CONTENTS

1.	DATA SET IDENTIFICATION

2.	INVESTIGATOR INFORMATION

3.	DATA SET ABSTRACT

4.	OBJECTIVES AND INTRODUCTION

5.	DATA ACQUISITION AND PROCESSING METHODS

6.	DATA MANIPULATIONS

7.	DATA DESCRIPTION

8.	GEOGRAPHICAL AND SPATIAL INFORMATION

9.	QUALITY CONTROL/QUALITY ASSURANCE

10.	DATA ACCESS

11.	REFERENCES

12.	TABLE OF ACRONYMS

13.	PERSONNEL INFORMATION

1. DATA SET IDENTIFICATION

1.1	Title of Catalog Document

EMAP-Estuaries Province Level Database
Carolinian Province
Sediment Toxicity Data

1.2	Authors of the Catalog entry

Timothy R. Snoots,

Dr. Jeffrey L. Hyland

1.3	Catalog Revision Date

November 23, 1999

1.4	Data Set Name

CP TOX.DAT


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1.5 Task Group

Estuari es

1.6	Data set identification codes

7

1.7	Version

001

1.8	Requested Acknowledgment

If you plan to publish these data in any way, EPA requires
a standard statement for work it has supported:

"Although the data described in this article have been
funded wholly or in part by the U. S. Environmental
Protection Agency through its EMAP-Estuaries Program, it
has not been subjected to Agency review, and therefore does
not necessarily reflect the views of the Agency and no official
endorsement should be inferred."

2. INVESTIGATOR INFORMATION

2.1	Principal Investigator

J. Hyland (N0AA/N0S/NCC0S/CCMA) - Carolinian Province Manager
A. Ringwood (SCDNR) - Lead P.I. for SC/GA region team
C. Hackney (UNC-W) - Lead P.I. for NC region team
G. McRae, G. Nelson, J. McKenna, J. Landsberg (FLDEP) -
Lead P.I.s for FL region team (depending on year)

2.2	Investigation Participant - Sample Collection

Field Sample Collection

T.	Alphin, S. Bowen, C. Byrum, D. Dye, A. Gospodarek,

J.	Grace, J. Grimley, C. Hackney, C. Powell, C. Preziosi,

H.	Riley, S. Roberts, M. Smith, K. Stokesbury,

D.	Tremain, T. Wheeler (UNC-W); S. Ross (NCNERR);

M.	Armstrong-Taylor, J. Jones, M. Levinson, P. Powers,

A.	Ringwood, T. Snoots, G. Steele (SCDNR); L. Balthis,

T.	Herrlinger, C. Keppler, M. Wert (UC); D. Adams,

K.	Amendola, D. Cook, C. Harnden, B. Heagey, J. Mckenna,

G.	Nelson, C. Nowicki, R. Paperno, B. Rosenblatt,

M.	Wessel (FLDEP); J. Hyland, S. Kokkinakis
(N0AA/N0S/0RCA)

Field Training and Coordination

S. Kokkinakis (N0AA/N0S/0RCA); J. Macauley (EPA-GED);
T. Hei tmuller (USGS-GB); D. Keith (EPA-AED)


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2.3 Sampling Processing -

Principal Investigator

Program Management and Coordination

J. Hyland, A. Robertson (NOAA/NOS/NCCOS/CCMA); K. Summers (EPA);

F.	Holland, A. Ringwood (SCDNR); C. Hackney,

T. Wheeler (UNC-W); S. Ross (NCNERR);

J. Landsberg, J. McKenna, G. McRae, G. Nelson,

R. Paperno (FLDEP)

Toxicity Testing

M.	DeLorenzo (CU) ; J. Grimley (UNC-W); J. Jones,

C.	Keppler, P. Maier, A. Ringwood, R. Van Dolah (SCDNR);

P.	Ross (Citadel); C. Mueller, J. Scott,

G.	Thursby (SAIC)

QA/QC

T.	Heitmuller (USGS-GB), S. Kokkinakis (NOAA/NOS/ORCA)

Data Management and Statistical Support

T.	Snoots, F. Holland, R. VanDolah (SCDNR); L. Balthis,

T.	Herrlinger (UC); J. Rosen, L. Zimmerman (TPMC);

S.	Rathbun (UGA); M. Adams, L. Harwell (JCWS);

V.	Engle (EPA-GED); Z. Malaeb (USGS-GB);

S.	Hale (EPA-AED); K. Summers (EPA); T. Wilson (CU)

3. DATA SET ABSTRACT

3.1 Abstract of the Data set

The CP_T0X.DAT data set contains results of up to four different
toxicity assays (10-day Ampelisca abdita mortality assay, 10-day
Ampelisca verrilli mortality assay, 7-day Mercenaria mercenaria
growth assay, and Microtox solid-phase assay) performed on sediment
samples from each station sampled in the EMAP Carolinian Province
from 1994-1997.

The following reports are products of these and other data
collected during the 1994-1997 Sampling period in the Carolinian
Province. These reports may contain additional information and
summary statistics that are not contained in this data set catalog
or its respective data sets. We therefore recommend referring to
them when using these data.

Hyland, J.L., T.J. Herrlinger, T.R. Snoots, A.H. Ringwood, R.F.
Van Dolah, C.T. Hackney, G.A. Nelson, J.S. Rosen, and
S.A. Kokkinakis. 1996. Environmental quality of
estuaries of the Carolinian Province: 1994. Annual
statistical summary for the 1994 EMAP-Estuaries
Demonstration Project in the Carolinian Province.

NOAA Technical Memorandum NOS ORCA 97. NOAA/NOS,

Office of Ocean Resources Conservation and Assessment,

Silver Spring, MD. 102 p.


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Hyland, J.L., L. Balthis, C.T. Hackney, G. McRae, A.H. Ringwood,
T.R. Snoots, R.F. Van Dolah, and T.L. Wade. 1998.
Environmental quality of estuaries of the Carolinian
Province: 1995. Annual statistical summary for the 1995
EMAP-Estuaries Demonstration Project in the Carolinian
Province. NOAA Technical Memorandum NOS ORCA 123
NOAA/NOS, Office of Ocean Resources Conservation and
Assessment, Silver Spring, MD. 143 p.

See also: Grimley and Hackney (1996), Ringwood et al. (1997b),
Ringwood and Keppler (In Press), Ringwood et al.(1998),
Ringwood et al. (1996), Ringwoodet al. (1995),

Ringwood et al. (1997a), Science Applications
International Corp. (1998a), Science Applications
International Corp. (1998b).

3.2 Keywords for the Data Set

Sediment toxicity, amphipod, Ampelisca abdita,

Ampelisca verrilli, Microtox, Mercenaria mercenaria,

EMAP Carolinian Province

4. OBJECTIVES AND INTRODUCTION

4.1	Program Objective

EMAP has three primary objectives:

1.	To estimate the current status, extent, changes, and trends

in indicators of the Nation's ecological resources on a
regional basis;

2.	To monitor indicators of pollutant exposure and habitat

condition, and to seek correlative relationships between
human-induced stresses and ecological condition that
identify possible causes of adverse effects; and

3.	To provide periodic statistical summaries and interpretive

reports on ecological status and trends to the EPA
Administrator and to the public.

4.2	Data Set Objective

The CP_T0X.DAT data set contains results of up to four different
toxicity assays (10-day Ampelisca abdita mortality assay, 10-day
Ampelisca verrilli mortality assay, 7-day Mercenaria mercenaria
growth assay, and Microtox solid-phase assay) performed on sediment
samples from each station sampled in the EMAP Carolinian Province
from 1994-1997.

Where applicable, both assay end results (i.e., % survival,
% growth, or EC50) , and statistical test results (P-value for
test of control vs. sample results) are reported. This allows
a duel criteria to be used in the determination of a sample's
toxicity (i.e., test results should be both statistically and
biologically significant to declare the overall toxicity result


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significant). These duel criteria have been applied for each
test and results are reported in a coded fashion
(1 = significant toxicity, 0 = not toxic).

4.3 Data Set Background Information

The standard 10-day sediment bioassay with the marine amphipod
Ampelisca abdita (ASTM 1993) has been used to assess sediment
toxicity in other EMAP provinces. This assay was also used in the
Carolinian Province in 1994, 1995 and 1997 to provide a basis
for comparisons among EMAP provinces and between years within
the Carolinian Province. However, because Ampelisca abdita
proved to be relatively insensitive to sediment contaminants
in prior surveys conducted in both the Carolinian and Louisianian
Provinces (Hyland et al. 1996, Macauley et al. 1994), an
additional amphipod assay with the congeneric species Ampelisca
verrilli was included in the 1995 effort. Preliminary testing
with A. verrilli and a subset of the 1994 sediment samples
indicated that this species was more sensitive to sediment
contamination than A. abdita (Ringwood et al. 1995).

Furthermore, A. verrilli is a more common member of the infaunal
benthos of southeastern estuaries.

A third bioassay used to measure potential sediment toxicity
at all base sites and selected supplemental sites was the
Microtox solid-phase test with the photoluminescent bacterium
Vibrio fischeri (formerly Photobacterium phosphoreum).

This assay provides a sublethal measure of toxicity based on
attenuation of light production by the bacterial cells due
to exposure to the sediment sample (Bulich 1979, Ross et al.
1991, Microbics 1992a and b) . Microtox has not been used in
other EMAP-E provinces, but its recent application in other
coastal assessment programs suggested that it might be a
useful tool to consider for the Carolinian Province. Small
sample sizes (a 100-mL subsample of the composited surface
sediment from each station) and a short processing time
(20-min exposures) provide clear logistical advantages.

Results of the Carolinian Province 1993 pilot study
(Ringwood et al. 1996) and 1994 monitoring demonstration
(Hyland et al. 1996) also suggested that this test is more
powerful in its ability to discriminate between degraded and
reference sites than the amphipod toxicity test.

A fourth sediment bioassay used in the 1995-1997 surveys was
a 7-d sublethal test of the effects of sediment exposure on
growth of juvenile Mercenaria mercenaria (referred to
hereafter as "seed clams"). The seed-clam bioassay was
developed during the Carolinian Pilot Study (Ringwood et al.
1996, Ringwood and Kepler In Press). Field-validation testing
on a subset of the 1994 sediment samples indicated that this
bioassay was a more sensitive indicator of sediment
contamination than the A. abdita bioassay (Ringwood et al.
1995). There are other practical advantages. For example,
newly metamorphosed clams exhibit very rapid growth, thus
effects on growth can be detected within a short time frame.
Second, because seed clams can be obtained from cultured
populations (available ~ 3 months after fertilization),


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experiments can be conducted with animals of similar size, age,
and pre-exposure histories. Third, a relatively small sample
volume (500 mL) is required, thus minimizing sampling time and
storage needs. Lastly, Mercenaria feed at the sediment-water
interface, where maximum contaminant exposure would be
expected. Thus the bioassay is representative of a realistic
exposure scenario.

4.4	Summary of Data Set Parameters

See 4.2 (Data Set Objective)

4.5	Year-Specific Information about Data

Toxicity testing in the Carolinian Province from 1994-1997
was performed by the following laboratories:





Test

Type





Ampeli sea

Ampeli sea

Mercenari a



Year

abdi ta

verri Hi

mercenari a

Mi crotox

1994

SCDNR,SAIC

-

-

SCDNR

1995

SAIC

SCDNR

SCDNR

SCDNR

1996

-

-

SCDNR

UNC-W

1997

SCDNR,SAIC

-

SCDNR

SCDNR,SAIC

Hyphen (-) indicates test was not performed in that year.

5. DATA ACQUISITION AND PROCESSING METHODS
5.1 Data Acqui si ti on

5.1.1	Sampling Objective

See section 4.3 (Data Set Background Information)

5.1.2	Sample Collection Method Summary

A 1/25 m2, Kynar-coated stainless steel, Young Grab sampler
was used to collect sediments. This grab sampled an area of
440 cm2 and a maximum depth of penetration in the sediment
of 10 cm. Stainless steel utensils were used to remove the
top 2-3 cm of sediment from a grab. The sediment was removed
to a stainless steel bowl and placed in a cooler of ice to
remain cold, but unfrozen. The grab sampler was rinsed and
re-deployed. This procedure was repeated until the volume
of sediment required for all contaminant, toxicity, and
sediment characteristics analyses had been collected.
The sediment was mixed by hand until thoroughly homogenized,
and aliquots were placed into pre-cleaned containers
(Polyethylene for A. abdita and A. verrilli, and Polypropylene
for Microtox and M. mercenaria). In addition, Microtox
samples were wrapped in foil immediately to exclude light.
All toxicity samples were immediately stored on ice following
collection in the field, and then refrigerated at 4 C once
back in the lab.


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5.1.3 Beginning Sampling Dates

30	June 1994
05	July 1995
09	July 1996
07 July 1997

5.1.4	Ending Sampling Dates

31	August 1994

14 September 1995
19 September 1996
25 August 1997

5.1.5	Platform

Samples were collected from various gasoline or diesel
powered boats equipped with at least the following
equipment: "A" frame boom or davit, winch, LORAN-C or
GPS for location, and a depth finder.

5.1.6	Sampling Equipment

A 1/25 m2, Kynar-coated stainless steel, Young Grab sampler.
This grab sampled an area of 440 cm2 and a maximum depth
of penetration in the sediment of 10 cm.

5.1.7	Manufacturer of Sampling Equipment

Ted Young
Falmouth, MA

5.1.8	Key Variables

5.1.9	Sample Collection Method Calibration

The sampling gear does not require any calibration.
It required inspection for deformities incurred due to
mishandling or impact on rocky substrates.

5.1.10	Sample Collection Quality Control

Field technicians were trained to follow Standard Operating
Procedures to insure the collection of representative,
uncontaminated and high quality samples. QA/QC measures
were taken in the field to avoid or reduce contamination
and insure the collection of representative samples. These
included: use of stainless steel instruments, thorough
cleaning of the sampler between grabs, use of pre-cleaned
containers for sediment storage and ensuring that engines
were shut down when a sample was exposed to the air.
A successful grab had relatively level, intact sediment over
the entire area of the grab and a sediment depth of 7-10
centimeters. Unacceptable grabs included those: containing
no sediments, which were partially filled or had shelly
substrates or grossly slumped surfaces. Grabs completely


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filled to the top, where the sediment was oozing out of the
hinged top, were also unacceptable.

See: Kokkinakis et al. (1994a)

5.1.11	Sample Collection Method References

See: Hyland et al. (1996),

Hyland et al. (1998),

Kokkinakis et al. (1994b)

5.1.12	Sample Collection Method Deviations
None

5.2 Data Preparation and Sample Processing

5.2.1	Sample Processing Objective

Determine toxicity of sediment samples using up to four
toxicity assays (10-day Ampelisca abdita mortality assay,
10-day Ampelisca verrilli mortality assay, 7-day Mercenaria
mercenaria growth assay, and Microtox solid-phase assay).

5.2.2	Sample Processing Methods Summary

5.2.2.1	Field Summary
NA

5.2.2.2	Laboratory Summary

A. abdita and A. verrilli Testing

Procedures followed the general guidelines provided in
ASTM Protocol E1367-92 (ASTM 1993) and the EMAP-E
Laboratory Methods Manual (U.S. EPA 1994a, 1994b).This is
an acute toxicity test which measures the effect of
sediment exposure on amphipod survival under static
conditions. Approximately 3-3.5 L of surface sediments
(composite of upper 2 cm from multiple grabs) were
collected for each type of assay from each station and
stored in 3.7-L polyethylene jars at 4 C in the dark until
testing. Tests were conducted with subsamples of the same
sediment on which analyses of contaminants and other
sediment characteristics were performed. Wherever
possible, sediment samples were tested within 30 days
of collection as recommended in the EMAP-E protocol.

Amphipods were collected from unpolluted tidal flats.

Prior to testing, the animals were acclimated at 20 C
for 2-9 days in the case of A. abdita, or for 2-4 days
in the case of A. verrilli. During the acclimation
period, the amphipods were fed the diatom Phaeodactylum
tricornutum. Wherever possible, juvenile amphipods of
approximately the same size (usually 3-5 mm in length
for A. abdita, and 3-10 mm in length for A. verrilli)


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were used to initiate the tests.

The general health of each batch of amphipods was
evaluated by a reference toxicity test (i.e., "positive
control"). These tests were run in a dilution series
with seawater (no sediment phase) and the reference
toxicant sodium dodecyl sulfate (SDS). Tests for both
species were run under static conditions in dark and
followed the basic methods described by ASTM (1993).
The exposure period was 96 h for A. abdita and 24 h for
A. verrilli. The shorter exposure period was used for
A. verrilli to match previous reference toxicant tests
conducted with this species by MRRI. LC50 values were
computed for each batch of test animals for comparison
against background toxicity data on these same species
and reference toxicant. Animals were not used in
definitive tests with field samples unless acceptable
reference toxicant results were obtained. A test was
considered acceptable if the LC50 value was within
+/- 2 SD of the mean LC50 based on the preceding 20
(A. abdita) to 22 (A. verrilli) reference toxicant
tests.

Treatments for the definitive tests with field samples
consisted of a single concentration of each sediment
sample (100% sediment) and a negative control
[i.e., reference sediment], A negative control was
run with each batch of field samples. The tests were
conducted under static conditions at a temperature of
20 +/- 1 C and salinity range of 26-33ppt for A. abdita
and 26-35ppt for A. verrilli. Twenty amphipods were
randomly distributed to each of five replicates per
each treatment including the control. Amphipods were
not fed during the tests.

The negative controls provided a basis of comparison
for determining statistical differences in survival in
the field sediments. In addition, control survival
provided a measure of the acceptability of final test
results. Test results were considered valid if mean
control survival (among the 5 replicates) was >= 85%
and survival in any single control chamber was >= 80%.

One-liter glass containers with covers were used as
test chambers. Each chamber was filled with 200 mL of
sediment and 600-800 mL of filtered seawater.

The sediment was press-sieved through a 2.0-mm screen
to remove ambient fauna prior to placing it in a chamber.
All containers were illuminated constantly throughout the
10-day test to inhibit amphipod emergence from the
sediment, thus maximizing exposure to the test sediment.
Air was supplied using oil-free aerators and glass
pipettes inserted into the test chambers. Water tables
with recirculating chiller pumps were used to maintain
constant temperatures (20 +/- 1 C). Daily recordings
were made of temperature and the number of dead vs.
living animals. On days two and eight, two of the five


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replicate chambers for each treatment were selected
randomly and measured for salinity, dissolved oxygen,
pH, and total ammonia in the overlying water.

At the conclusion of a test, the sediment from each
chamber was sieved through a 0.5-mm screen to remove
amphipods. The number of animals dead, alive, or
missing was recorded. Sediments with missing A. abdita
were preserved in formalin containing Rose Bengal stain
and re-examined under a dissecting microscope to ensure
that no living specimens had been missed. Animals still
unaccounted for were considered to have died and
decomposed in the sediment. Because of their larger
size, A. verrilli were much easier to locate with the
unaided eye. Thus, if any of these animals were missing
after initial examination of the sieved sediment, then
they were assumed to have died and decomposed.

Differences between survival of Ampelisca abdita in
field versus control samples were evaluated by an
unpaired heteroscedastic t-test run on untransformed
percentage data, under the assumptions of normality
and unequal variances. For A. verrilli, differences
between field samples and controls were evaluated by
either: (i) an unpaired homoscedastic t-test in cases
of normal data with equal variances, or (ii) a
Mann-Whitney U-test in cases of non-normal data or
unequal variances. The A. verrilli comparisons also
were performed on untransformed percentage data.

For both bioassays, field samples were considered to
be significantly toxic if mean survival in comparison
to the corresponding negative control was < 80% and
statistically different at alpha = 0.05.

A variety of quality control procedures were
incorporated to assure acceptability of amphipod
test results and comparability of the data with
other studies. As described above, these provisions
included the use of standard ASTM and EMAP protocols,
positive controls run with a reference toxicant,
negative "performance" controls run with reference
sediment, and routine monitoring of water quality
variables to identify any departures from optimum
tolerance ranges. In addition, during the first year
of the program, an inter-laboratory comparison of
results using the A. abdita assay was performed by
the two participating testing facilities (SAIC and
SCDNR/MRRI). Samples from two of the base sites
collected in 1994 were tested by each facility.

Results were highly comparable: mean survival in
field samples relative to controls was 96% for both
samples by one lab, and 98 to 100% by the other lab.


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Microtox Testing

Tests were conducted in duplicate following the
"large-sample-size" protocol of Microbics Corporation
(1992b). Wherever possible, sediment samples were
tested within the recommended 10-d holding period.
A 7-g aliquot of each sediment sample was used to make a
dilution series ranging from 0.01 to 10% sediment in a
2% saline diluent. A reagent solution containing the
bacteria was then added to each sediment suspension.
After a 20-min incubation period, a column filter was
used to separate the liquid phase and bacterial cells
from the sediment. Post-exposure light output in each
of the filtrates was measured on a Microtox Model 500
Analyzer. A log-linear regression model was used to
determine an EC50 - the sediment concentration that
reduced light production by 50% relative to a control
(nontoxic reagent blank). EC50 values were corrected
for percent water content and reported as dry-weight
concentrati ons.

Assays were run with the reference toxicant phenol with
each new batch of bacteria. These tests provided
measures of the general quality of the bacterial
populations, as well as the ability of the laboratory
to produce results consistent with the expected phenol
toxicity range (i.e., Microtox EC50 values typically
between 13-26 mg/L). Use of the standard Microtox
equipment and protocol helped to assure data
comparability with results of other Microtox studies.

Mercenaria mercenaria Testing

Seed clams (~ 1 mm in length) were obtained from
Atlantic Clam Farms, Folly Beach, S.C. Replicate
subsets were dried and weighed to provide initial
weight estimates. On the day before initiation of a
test, sediment samples were sieved through a
500-micrometer screen (to remove ambient fauna) and
distributed to the test chambers. Approximately
50 mL of sieved sediment were added to each of four
replicate 250-mL beakers for each sediment sample.
A negative control (same Folly River sediments used
as controls in the Ampelisca verrilli assays) was run
with each batch of field samples. Filtered seawater
(1-micrometer filter), adjusted to 25 ppt with deionized
water, was added to each beaker to bring the total
volume up to 200 mL. The sediment suspension was
allowed to settle overnight and clams (30-50 per
replicate) were added the next day (which initiated
the test).

Tests were conducted for 7 d. All tests were conducted
at room temperature (23-25 C) under gentle aeration.
Animals were fed three times throughout the test with a
phytoplankton mixture consisting of equal volumes of
Isochrysis galbana and Chaetocerus gracilis.


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At the end of the 7-d exposure period, clams were
sieved from the sediments, placed in clean seawater,
and allowed to depurate for ~ 1 h. Clams were
re-captured on a sieve and rinsed briefly with distilled
water to remove excess salt. Dead clams were removed
and not included in subsequent growth estimates
(mortality rates generally were < 10%). The remaining
live clams were dried overnight (60-70 C), counted, and
weighed on a micro-balance. The pre- and post-exposure
measurements were then used to determine growth rates,
expressed as ug/clam/d. Effects of sediment exposure
on growth rates were evaluated using either a t-test
or Mann-Whitney U-test (when assumptions of the
parametric test were violated). Samples were
considered to be significantly toxic if mean growth
rate in comparison to the control was < 80% and
statistically different at alpha = 0.05.

Each new batch of seed clams was evaluated for
suitability and relative sensitivity with a reference
toxicant test ("positive control"). These tests were
run under static conditions, at room temperature, in a
dilution series with 25ppt seawater (no sediment phase)
and the reference toxicant cadmium. Treatments within
each test consisted of a seawater control and four
cadmium concentrations (25, 50, 100, 200 ug/L as CdCl2) .
Each treatment was represented by 3-4 replicates. The
effective Cd concentration that reduced growth by 50%
(EC50) relative to the seawater control was estimated by
regression analysis.

5.2.3	Sample Processing Method Calibration

See Section 5.2.2.2 (Sample Processing Methods Summary/
Laboratory Summary)

5.2.4	Sample Processing Quality Control

See Section 5.2.2.2 (Sample Processing Methods Summary/
Laboratory Summary)

5.2.5	Sample Processing Method Reference

See Section 5.2.2.2 (Sample Processing Methods Summary/
Laboratory Summary)

5.2.6	Sample Processing Method Deviations

None


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6. DATA ANALYSIS AND MANIPULATIONS

6.1 Name of New or Modified Value

P_VALUE

RESULT

SIG

P-value for statistical test
Test result (relative to control)
Toxicity Test Result (1=Hit)

6.2 Data Manipulation Description

P VALUE

For A. abdita and A. verrilli, P_VALUE is the probability
value associated with a statistical test of mean survival
of Amphipods in field versus control samples. The test
performed was either: (1) an unpaired homoscedastic t-test
in cases of normal data with equal variances, or (2) a
Mann-Whitney U-test in cases of non-normal data or unequal
variances. The A. verrilli comparisons were performed on
untransformed percentage data.

For M. mercenaria, P_VALUE is the probability value associated
with a statistical test of mean growth of M. mercenaria in
field versus control samples. Effects of sediment exposure
on growth rates were evaluated using either a t-test or
Mann-Whitney U-test (when assumptions of the parametric test
were violated).

For A. abdita and A. verrilli, RESULT is mean survival in
field sediment as a percent of mean survival in control
sediment (e.g., RESULT = field/control * 100)

For Microtox, values reported in RESULT are EC50 values
(sediment concentration that reduced light production by 50%
relative to a control). These EC50 values have been
corrected for percent water content and are reported as
percent (dry-weight) sediment concentrations.

For M. mercenaria, RESULT is mean clam growth in field
sediment as a percent of mean growth in control sediment
(e.g., RESULT = field/control * 100).

The variable SIG reports a final, coded toxicity result based
on a duel criteria of statistical significance (P_VALUE)
and biological significance (RESULT). If both the P_VALUE
and RESULT for an assay meet the criteria for significance,
then the response is considered to be significantly toxic and
SIG = 1. If either P_VALUE or RESULT for an assay do not meet
the appropriate criteria for significance, then the response
is considered not toxic and SIG = 0. Where available data are
not adequate to evaluate using this duel criteria approach
(e.g., P_VALUE or RESULT data are missing), SIG = (missing).
The criteria used to evaluate toxicity responses in each of the

RESULT

SIG


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four assays are discussed below.

For both A. abdita and A.verrilli assays, field samples were
considered to be significantly toxic if mean survival of field
samples as a percent of the mean survival for the corresponding
negative control (RESULT) was < 80% and statistically different at
alpha =0.05 (P_VALUE).

For Microtox assays, evaluation criteria were established for
two separate silt-clay classes because of the strong inverse
relationship between Microtox EC50 values and percent
silt-clay content. Sediments with >= 20% silt-clays
were classified as being toxic if EC50 values were <= 0.2%;
sediments with < 20% silt-clays were classified as being
toxic if EC50 values were <= 0.5% (sensu Ringwood et al. 1995).
Lower EC50 values in muddier sediments are believed to be
caused by physical adsorption of the bacteria to the
sediment particles. Ringwood et al. (1995, 1997)
demonstrated this effect by conducting Microtox assays
in artificial sediment mixtures of pure sand and kaolin
clay and evaluating the EC50 values as a function of the
finer-particle content.

For M. mercenaria assays, samples were considered to be
significantly toxic if mean growth rate in field samples as
a percent of mean growth rate in the controls (RESULT) was
< 80% and statistically different at alpha = 0.05 (P_VALUE).

6.3 Data Manipulation Examples

7. DATA DESCRIPTION

7.1 Description of Parameters

Variable Type Format Label

DATE

P_VALUE

QC_C0DE

RESULT

SIG

STA_NAME

TEST

UNIT

Num
Num

YYMMDD6.
6.4

Char 30.

Num
Num
Char
Char 22.
Char 10.

8.3
1.
7.

Sample collection date (YYMMDD)

P-value for statistical test
QC Code

Test result (relative to control)
Toxicity Test Result (1=Hit)

Carolinian Province Office Station Name
Test Species
Test result unit code

Note the conventions used in the Format column above:

For character (Char) variables, the number given is the
maximum width (number of characters) for that variable.

For numeric (Num) variables, the format is given in W.D
format, where W = maximum width (number of characters)
for the number (including all digits and the decimal
point), and D = number of digits to the right of the
decimal point.


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7.1.6 Precision to which values are reported

Variable RESULT is reported to 0.001 units, however, values
are only valid to: 0.1 units for A. abdita and A. verrilli
results, 0.01 units for M. mercenaria results, and 0.001
units for Microtox results. Variable P_VALUE is reported
to 0.0001 units, however, values are only valid to: 0.001
units for A. abdita and A. verrilli results, and 0.0001
units for M. mercenaria results.

7.1.7 Minimum Value in Data Set

Variable Minimum

P_VALUE 0.0000
RESULT -148.690
SIG	0

7.1.8 Maximum Value in Data Set

Variable Maximum

P_VALUE 1.0000
RESULT 219.420
SIG	1

7.2 Data Record Example

7.2.1	Column Names for Example Records
STA_NAME;DATE;TEST;RESULT;UNIT;P_VALUE;SIG;QC_C0DE

7.2.2	Example Data Records

CP95121
CP95121
CP95121
CP95121
CP95122
CP95122
CP95122
CP95122
CP95123
CP95123
CP95123
CP95123

950720
950720
950720
950720
950720
950720
950720
950720
950726
950726
950726
950726

Ampeli sea abdi ta;104.200;Survi val_%;0.0790;0;AST-E
Ampeli sea verrilli;103.000;Survi val_%;0.2000;0;
Mercenaria mercenaria;135.320;Growth_%;.;0;CST-E, CST-
Microtox (V. fischeri);0.550;EC50_%;.;0;

Ampeli sea abdi ta;100.000;Survi val_%;0.5000;0;AST-E
Ampelisca verrilli;97.000;Survival_%;0.3700;0;
Mercenaria mercenaria;78.710;Growth_%;0.0286;1;CST-F
Microtox (V. fischeri);0.640;EC50_%;.;0;

Ampeli sea abdi ta;92.500;Survi val_%;0.0080;0;

Ampeli sea verri11i;104.000;Survi val_%;0.3700;0;
Mercenaria mercenaria;52.540;Growth_%;0.0024;1;CST-F
Microtox (V. fischeri);9.520;EC50_%;.;0;

GEOGRAPHIC AND SPATIAL INFORMATION

8.1 Minimum Longitude

-81 Degrees, 43.83 Minutes West Longitude


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8.2	Maximum Longitude

-75 Degrees, 33.82 Minutes West Longitude

8.3	Minimum Latitude

27 Degrees, 12.07 Minutes North Latitude

8.4	Maximum Latitude

36 Degrees, 43.43 Minutes North Latitude

8.5	Name of area or region

Coastal distribution of sampling is along the southeastern US
from Cape Henry, VA, through St. Lucie Inlet, FL. States
represented: Virginia, North Carolina, South Carolina, Georgia,
and Florida.

9. QUALITY CONTROL/QUALITY ASSURANCE

9.1	Measurement Quality Objectives

See Section 5.2.2.2 (Sample Processing Methods Summary/

Laboratory Summary)

9.2	Quality Assurance/Control Methods

See Section 5.2.2.2 (Sample Processing Methods Summary/

Laboratory Summary)

9.3	Quality Assessment Results

Unless flagged with one of the following QC codes, all data reported
in the CP_T0X.DAT data set met the QA/QC guidelines given above and
are acceptable without further qualification.

Where necessary, the following QC codes are reported under the
variable QC_C0DE and should be considered when interpreting
test results.

QC Code Description

AST-C Fewer than 5 replicates tested

AST-E Sample held for longer than 30 days

AST-M More than 20 animals inoculated into replicate

CST-A Fewer than 4 replicates were tested

CST-B Sediment too coarse to sieve through 0.5 mm mesh

therefore making it difficult to recover clams.

CST-C Sediment held longer than 30 days prior to testing
CST-D Folly River control sediment not used. Note that this
occurred only once. EXPTNUM 950928, sediment from
Breach Inlet used.

CST-E Statistical analysis not run because the mean growth
rate was >100% of the mean control growth rate


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CST-G Very high to complete mortality of clams in sample

(i.e., sample i s toxi c)

CST-H Fewer than 3 replicates were tested

(cadmium exposures only)

MST-A Samples were processed within 14 days of sampling
MST-E Sample held for > 10 days prior to testing
MST-F Unable to calculate an EC50 value for this sample due
to an insignificant decrease in luminescence or
an increase in luminescence
(i.e., little or no toxic effects)

MST-X Calculated EC50 result was greater than the highest test
concentration of 10%. Because the accuracy of an EC50
value above 10% is unknown, EC50 values greater that 10%
have been reported as 10.000%.

MST-Y Hit/Miss result could not be determined due to missing
silt-clay data

DATA ACCESS

10.1	Data Access Procedures

Data can be downloaded from the WWW site.

10.2	Data Access Restrictions

Data can only be accessed from the WWW site.

10.3	Data Access Contact Persons

For programmatic/policy matters, contact:

Dr. Jeffrey L. Hyland

N0AA/N0S National Centers for Coastal Ocean Science

Center for Coastal Monitoring and Assessment - Charleston Lab

217 Fort Johnson Rd. (P.O. Box 12559)

Charleston, SC 29422-2559

(843)762-5415 (Tel.)

(843)762-5110 (FAX)

jeff.hyland0noaa.gov (e-mail)

For data-related questions, contact:

Dr. W. Leonard Balthis

N0AA/N0S National Centers for Coastal Ocean Science

Center for Coastal Monitoring and Assessment - Charleston Lab

217 Fort Johnson Rd. (P.O. Box 12559)

Charleston, SC 29422-2559

(843)762-5652 (Tel.)

(843)762-5110 (FAX)

len.balthis0noaa.gov (e-mail)


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Data Librarian, EMAP-Estuaries
Melissa M. Hughes
OAO Corporation
U.S. EPA NHEERL-AED
27 Tarzwell Drive
Narragansett, RI 02882-1197
(401) 782-3184 (Tel.)

(401) 782-3030 (FAX)
hughes.melissa0epa.gov (e-mail)

10.4	Data file Format

Delimited ASCII Text

10.5	Information Concerning Anonymous FTP

Not accessible

10.6	Information Concerning WWW

Data can be downloaded from the WWW.

10.7	EMAP CD-ROM Containing the Data file

Data not available on CD-ROM.

11. REFERENCES

American Society for Testing and Materials (ASTM). 1993. Guide for
conducting 10-day static sediment toxicity tests with marine
and estuarine amphipods. ASTM Standard Method E-1367-92,

ASTM, Philadelphia, PA. 24p.

Bulich, A.A. 1979. Use of luminescent bacteria for determining
toxicity in aquatic environments. In: L. L. Marking and
R. A. Kimerle (eds.), Aquatic Toxicology, pp. 98-106. ASTM
STP 667. American Society for Testing and Materials,
Philadelphia, PA

Grimley, J. and C.T. Hackney. 1996 Year end summary. EMAP

Carolinian Province: North Carolina Estuaries Component.

Final year-three report under N0AA Cooperative Agreement
No. NA470A0148. University of North Carolina at Wilmington,
Center for Marine Research, Wilmington, NC.

Hyland, J.L., T.J. Herrlinger, T.R. Snoots, A.H. Ringwood, R.F.
Van Dolah, C.T. Hackney, G.A. Nelson, J.S. Rosen, and S.A.
Kokkinakis. 1996. Environmental quality of estuaries of the
Carolinian Province: 1994. Annual statistical summary for the
1994 EMAP-Estuaries Demonstration Project in the Carolinian
Province. N0AA Technical Memorandum N0S 0RCA 97. NOAA/NOS,
Office of Ocean Resources Conservation and Assessment,

Silver Spring, MD. 102 p.


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Hyland, J.L., L. Balthis, C.T. Hackney, G. McRae, A.H. Ringwood,
T.R. Snoots, R.F. Van Dolah, and T.L. Wade. 1998.

Environmental quality of estuaries of the Carolinian Province:
1995. Annual statistical summary for the 1995 EMAP-Estuaries
Demonstration Project in the Carolinian Province. NOAA Technical
Memorandum NOS ORCA 123 NOAA/NOS, Office of Ocean Resources
Conservation and Assessment, Silver Spring, MD.

Kokkinakis, S.A., C. Mageau, and A. Robertson. 1994a. Carolinian
Demonstration Project - 1994 Quality Assurance Project Plan.

Joint National Status and Trends/Environmental Monitoring and
Assessment Program. NOAA/NOS/ORCA, Silver Spring, MD.

Kokkinakis, S.A., J.L. Hyland, and A. Robertson. 1994b. Carolinian
Demonstration Project - 1994 Field Operations Manual. Joint
National Status and Trends/Environmental Monitoring and
Assessment Program. NOAA/NOS/ORCA, Silver Spring, MD.

Macauley, J.M., J.K. Summers, P.T. Heitmuller, V.D. Engle,

G.T. Brooks, M. Babikow, and A.M. Adams. 1994. Annual
Statistical Summary: EMAP - Estuaries Louisiana Province -
1992. U.S. EPA Office of Research and Development,

Environmental Research Laboratory, Gulf Breeze, FL.
EPA/620/R-94/002.

Microbics Corporation. 1992a. Microtox Manual (5 volume set).

Carlsbad, CA.

Microbics Corporation. 1992b. Microtox Update Manual, 128 p.

Carlsbad, CA.

Ringwood, A.H., M.E. DeLorenzo, P.E. Ross, and A.F. Holland.

1997b. Interpretation of Microtox« solid-phase toxicity
tests: the effects of sediment composition. Environ.

Toxicol. Chem. 16(6):1135-1140.

Ringwood, A.H. and C. Keppler. In Press. Seed clam growth:

A sediment bioassay developed in the EMAP Carolinian
Province, Environ. Monitor. & Assess, Submitted May 1997.

Ringwood, A.H., C. Keppler, R.F. Van Dolah, P. Maier,

J. Jones, M.E. DeLorenzo. 1998. EMAP Activities in the
Carolinian Province by Marine Resources Research Institute
in 1997: Results and Summaries. Final Annual Report under
NOAA Cooperative Agreement No. NA770A0214. South Carolina
Department of Natural Resources, Marine Resources Research
Institute, Charleston, SC.

Ringwood, A.H., A.F. Holland, R. Kneib, and P. Ross. 1996.

EMAP/NS&T pilot studies in the Carolinian Province:

Indicator testing and evaluation in southeastern estuaries.

Final Report under Grant NA90AA-D-SG790 through S.C. Sea
Grant College Program. S.C. Dept. of Natural Resources,

Marine Resources Research Institute, Charleston, S.C. NOAA
Technical Memorandum NOS ORCA 102. 113 p.


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Ringwood, A.H., R. Van Dolah, A.F. Holland, and M.G. Delorenzo.
1995. Year one demonstration project studies conducted in
the Carolinian Province by Marine Resources Research
Institute: Results and summaries. Final Annual Report
under NOAA Cooperative Agreement No. NA470A0177. South
Carolina Department of Natural Resources, Marine Resources
Research Institute, Charleston, S.C

Ringwood, A.H., R. Van Dolah, A.F. Holland, M.G. Delorenzo,

C. Keppler, P. Maier, J. Jones, and M. Armstrong-Taylor.
1997a. Year two demonstration project studies conducted in
the Carolinian Province by Marine Resources Research
Institute: Results and summaries. Final Annual Report under
NOAA Cooperative Agreement No. NA470A0177. South Carolina
Department of Natural Resources, Marine Resources Research
Institute, Charleston, SC.

Science Applications International Corp. 1998a. Sediment toxicity
testing (MAIA 1997), Amphipod test results, final report
(21 Jan 1998). Science Applications International Corporation,
Narragansett, RI.

Science Applications International Corp. 1998b. Sediment toxicity
testing (MAIA 1997), Microtox test results, final report
(10 Feb 1998). Science Applications International Corporation,
Narragansett, RI.

Ross, P.E., L.C. Burnett, C. Kermode, and M. Timme. 1991.
Miniaturizing a toxicity test battery for screening
contaminated sediments. Can. Tec. Rep. Fish. Aquat. Sci.,
1774: 331-335.

Strobel, C.J., S.J. Benyi, D.J. Keith, H.W. Buffum, and E.A.

Petrocelli. 1994. Statistical summary: EMAP - Estuaries
Virginian Province - 1992. U.S. EPA Office of Research
and Development, Environmental Research Laboratory, Narragansett,
RI. EPA/620/R-94/019.

Summers, J.K., J.M. Macauley, P.T. Heitmuller, V.D. Engle, A.M. Adams,
and G.T. Brooks. 1993. Annual statistical summary: EMAP -
Estuaries Louisianian Province - 1991. U.S. EPA Office of
Research and Development, Environmental Research Laboratory,

Gulf Breeze, FL. EPA/600/R-93/001.

U.S. EPA. 1994a. Environmental Monitoring and Assessment Program
(EMAP): Laboratory methods manual - estuaries, Volume 1:
Biological and physical analyses. Office of Research and
Development, Environmental Monitoring and Systems Laboratory,
Cincinnati, OH. EPA/620/R-94/xxx.

U.S. EPA. 1994b. Methods for assessing the toxicity of sediment-

associated contaminants with estuarine and marine amphipods.
EPA 600/R-94/025. U.S. EPA Office of Research and Development,
Narragansett, R.I.


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U.S. EPA. 1995. Environmental Monitoring and Assessment Program

(EMAP): Laboratory Methods Manual-Estuaries, Volume 1: Biologi
and Physical Analyses. U.S. Environmental Protection Agency,
Office of Research and Development, Narragansett, RI.
EPA/620/R-95/008.

TABLE OF ACRONYMS

<=	Less than or equal to

>=	Greater than or equal to

C	Degrees Celsius

cm2	Square centimeters

CMBAD	Coastal Monitoring and Bioeffects Assessment Division

CU	Clemson University

EMAP	Environmental Monitoring and Assessment Program

EPA	U.S. Environmental Protection Agency

EPA-AED	EPA-Atlantic Ecology Division

EPA-GED	EPA-Gulf Ecology Division

EPA-RTP	EPA-Research Triangle Park, NC

FLDEP	Florida Dept. of Environmental Protection

FMRI	Florida Marine Research Institute

FTP	File Transfer Protocol

GIS	Geographical Information System

JCWS	Johnson Controls Word Services

km2	Square kilometers

m2	Square meters

mg/L	Milligrams per liter

mS/cm	MilliSiemens per centimeter (equiv. to milliohms/cm)

MRRI	Marine Resources Research Institute

NCNERR	North Carolina National Estuarine Research Reserve

NCSU	North Carolina State University, NC

NA	Not Applicable

ng/g	Nanograms per gram

NOAA	National Oceanic and Atmospheric Administration

NOS	National Ocean Service

ORCA	Office of Ocean Resources Conservation and Assessment

QA/QC	Quality Assurance/Quality Control

ppb	Parts per billion (equiv. to ng/g)

ppm	Parts per million (equiv. to ug/g)

ppt	Parts per thousand

SAIC	Science Applications International Corporation

SCDNR	South Carolina Dept. of Natural Resources

TOC	Total Organic Carbon

TAMU/GERG Texas A&M University, Geochemical and Environmental
Research Group

TPMC	Technology Planning and Management Corporation

ug/g	Micrograms per gram

um	Micrometers

UC	University of Charleston, SC

UGA	University of Georgia, GA

UNC-W	University of North Carolina - Wilmington, NC

USGS-GB	US Geological Survey - Gulf Breeze, FL

wt.	Weight

WWW	World Wide Web -Internet


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13.

PERSONNEL INFORMATION

Dr. Courtney T. Hackney

University of North Carolina - Wilmington,

Wilmington, NC

Melissa M. Hughes
Data Librarian, EMAP-Estuaries
0A0 Corporation
U.S. EPA NHEERL-AED
27 Tarzwell Drive
Narragansett, RI 02882-1197
(401) 782-3184 (Tel.)

(401) 782-3030 (FAX)
hughes.melissa0epa.gov (e-mail)

For programmatic/policy matters, contact:

Dr. Jeffrey L. Hyland

NOAA/NOS National Centers for Coastal Ocean Science

Center for Coastal Monitoring and Assessment - Charleston Lab

219 Fort Johnson Rd.

Charleston, SC 29412-9110

(843)762-8652 (Tel.)

(843)762-8700 (FAX)

jeff.hyland0noaa.gov (e-mail)

For data-related questions, contact:

Dr. W. Leonard Balthis

NOAA/NOS National Centers for Coastal Ocean Science

Center for Coastal Monitoring and Assessment - Charleston Lab

219 Fort Johnson Rd.

Charleston, SC 29412-9110

(843)762-8654 (Tel.)

(843)762-8700 (FAX)

len.balthis0noaa.gov (e-mail)

Jan Landsberg

Florida Dept. of Environmental Protection
Florida Marine Research Institute
St. Petersburg, FL

James McKenna

Florida Dept. of Environmental Protection
Florida Marine Research Institute
St. Petersburg, FL

Dr. Gil McRae

Florida Dept. of Environmental Protection
Florida Marine Research Institute
St. Petersburg, FL

Gary A. Nelson

Florida Dept. of Environmental Protection
Florida Marine Research Institute
St. Petersburg, FL


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Dr. Amy H. Ringwood

South Carolina Dept. of Natural Resources,
Marine Resources Research Institute
Charleston, SC


-------