United States
Environmental Protection
Agency
Environmental Monitoring Systems
Laboratory
Las Vegas NV 89114
Research and Development
EPA-600/S4-84-087 Jan. 1985
EPA Project Summary
Toxicity Persistence in
Prickly Pear Creek, Montana
John R. Baker and Barry P. Baldigo
I nstream toxicity tests using the larval
fathead minnow Pimephales promelas
and the cladoceran Ceriodaphnia reticu-
lata were conducted on Prickly Pear
Creek, Montana, waters to study toxic-
ity persistence in a stream. The toxicity
source was Spring Creek, a tributary of
Prickly Pear Creek. Gold mining tailing
and settling ponds in the Spring Creek
drainage release zinc, copper and cad-
mium to Prickly Pear Creek. Flow
regimes, water quality, and biotic con-
ditions were characterized in conjunc-
tion with toxicity testing. The study
objectives were to: (1) develop a data
base for validating a toxicity persistence
model; (2) assess the applicability of
data from the Prickly Pear Creek study
relative to model assumptions; and (3)
assess field techniques for acquiring
model input data.
Toxicity to the test organisms was
primarily due to zinc and copper in
Spring Creek waters. Changes in Prickly
Pear Creek toxicity downstream from
the Spring Creek confluence were pri-
marily due to dilution and complied
with model assumptions. However,
other unidentified toxicants were pre-
sent in other tributary waters, and
Spring Creek was not the sole source of
toxicity in Prickly Pear Creek waters. C.
reticulate was highly sensitive to toxic-
ity in Spring Creek waters and provided
model input data. P. promelas had a
higher tolerance, and bioassay data
from these organisms could not be used
for model input. In the field, test orga-
nism nutritional problems were encoun-
tered using procedures described in
bioassay protocols for both of these
organisms, suggesting that either a
quantitative food regime should be
developed or a non-feeding test be used
in the future.
This Project Summary was developed
by EPA's Environmental Monitoring
Systems Laboratory, Las Vegas, NV, to
announce key findings of the research
project that is fully documented in a
separate report of the same title (see
Project Report ordering information at
back).
Introduction
The U.S. Environmental Protection
Agency's (EPA) Office of Water Regula-
tions and Standards, Monitoring and Data
Support Division (MDSD), is examining
persistence and degradation rates of
industrial and municipal toxic wastes
discharged to streams. MDSD is seeking
to identify methods most suitable for
assessing instream persistence of whole
effluent toxicity in receiving waters.
Methods are required for site specific
assessment of effluent toxicities, both
acute and chronic, prior to discharge, at
the discharge point, and at downstream
locations where dilution, degradation,
and partitioning to other compartments
result in reduced toxicant concentrations.
Particular interest centers on validation
of toxicity models designed to predict
instream toxicity persistence and valida-
tion of methods for acquiring input data
for these models.
Instream toxicity testing has recently
been conducted at several sites by EPA's
Environmental Monitoring Systems
Laboratory-Las Vegas (EMSL-LV) and
EPA's Environmental Research Labora-
tory-Duluth. Model validation will be
based on results from these investiga-
tions. A stream dilution model is presently
being assessed. Model assumptions are:
(1) toxic chemicals and toxicity itself
follow conservative (not enhanced or
degraded) mixing behavior; (2) physical.
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chemical, and biological interactions do
not substantially alter toxicity at the point
of complete mixing; and (3) variations in
effluent toxicity are reflected in varying
toxicity of the receiving waters and can be
described by mass balance relationships.
In order to provide additional instream
toxicity persistence data to MDSD, the
EMSL-LV conducted a stream toxicity
study in fall 1983 at Prickly Pear Creek,
Montana. The objectives of this study
were to: (1) develop a data base to be used
for model validation; (2) assess the appli-
cability of data from Prickly Pear Creek
relative to model assumptions; and (3)
assess field techniques for acquiring
model input data. The study consisted of
short-term acute and chronic toxicity tests
using two test organisms, and stream
survey characterization of flow regimes,
water quality, and biotic conditions. Data
for model validation are given in Appen-
dices of the parent report.
Prickly Pear Creek forms its headwaters
in the Elkhorn Mountains approximately
32 km southeast of Helena, Montana. The
stream flows north for 64 km before
entering Lake Helena and the Missouri
River. Gold mining in the Corbin and
Spring Creek drainage basin (draining
into Prickly Pear Creek) began in the early
1860s. Tailing and settling ponds remain
as prominant features within these
drainages and release high concentra-
tions of zinc, copper, and cadmium, which
are carried into Prickly Pear Creek via
Spring Creek. Areas along Prickly Pear
Creek were also subjected to extensive
mining operations in the early 1900s.
Over 75 percent of Prickly Pear Creek was
subjected to stream bed modifications
and dredging during the mining process.
Methods
Spring Creek toxicity and instream
toxicity persistence in Prickly Pear Creek
were determined using static renewal
bioassays designed to measure both acute
and chronic toxicity. Test organisms were
the cladoceran Ceriodaphnia reticulata
andthe larvalfathead mmnowPimephales
promelas. Toxicity tests were conducted
with water collected from September 30
through October 9, 1983. Prickly Pear
Creek stations were 011, a control station
located 1.1 km upstream from the Spring
Creek confluence; 013 and 014 were
biological impact stations located 300 m
and 3.8 km, respectively, downstream
from the Spring Creek confluence; and
018, a biological recovery station, was
located 12 km downstream from the
Spring Creek confluence. Water quality
and hydrological parameters were also
measured as part of the study. For toxicity
tests, control test waters were collected
from the upstream Prickly Pear Creek
station 011. These waters were diluted
with Spring Creek water (the toxicity
source) to obtain dilution test volumes
with varying metals concentrations for
comparison to ambient Prickly Pear Creek
water toxicity.
Results and Discussion
Metal Concentrations
Spring Creek metal contributions
caused significant increases in concen-
trations of metal in Prickly Pear Creek
water. There was a consistent decline in
downstream Prickly Pear Creek metal
concentrations, with approximately a
two-fold decrease between stations 013
and 018, due primarily to other tributary
inflow dilution. Dissolved metal concen-
trations were low in all of these tributary
streams.
Total recoverable cadmium, zinc, and
copper concentrations in Spring Creek
and Prickly Pear Creek samples consis-
tently exceeded EPA-recommended acute
criteria for aquatic life during the toxicity
testing period. Concentrations of arsenic
and lead were below the aquatic life
criteria at all stations. Silver exceeded the
acute criteria on October 6 at station 013
but was well below the acute criteria for
all other dates and stations. Although
cadmium exceeded the acute criteria,
concentrations were below reported toxic
levels for C. reticulata and larval fathead
minnows. Toxicity in test organisms was
attributed to zinc and copper, but C.
reticulata bioassays indicated that another
unidentified toxicant was present. Zinc
and copper concentrations in Spring
Creek were variable over the 10-day
testing period with peaktotal recoverable
concentrations on test days 1 and 5 (test
numbers 0 through 9 refer to dates,
September 30 - October 9). A small
rainstorm occurred on September 30 and
resulted in the October 1 (test 1) concen-
tration peak. The cause of the October 5
peak in total recoverable concentrations
was not determined.
Toxicity Tests
Ceriodaphnia reticulata
Acute, and Chronic Toxicity in Dilution
Tests—Dilution tests with Spring Creek
water produced acute effects (LC-50s) to
C. reticulata at dilution volumes varying
from approximately 5 to 20 percent. There
were no significant differences in Spring
Creek acute toxicity in tests 2 through 5,
8, and 9, but toxicity was significantly
higher in tests 1, 6, and 7. Higher toxicity
in test 1 corresponded to high total
recoverable and dissolved concentrations
of zinc and copper in Spring Creek on
October 1. An increase in total recover-
able concentrations of these metals on
October 5 had no apparent effect on
toxicity. Metal concentrations generally
declined on October 6 and 7 and the
increase in toxicity in tests 6 and 7 was
not due to an increase in any of the metals
analyzed in this investigation. There was
no mortality in the control organisms for
tests 6 and 7, indicating that mortality in
the Spring Creek dilution tests was due to
toxicity. Chemical analyses for other
parameters were not possible and the
toxicant was not identified. - -
Chronic toxicity, resulting in reduced
neonate production, was only evident in
tests 5 through 8 and occurred at dilution
volumes of 5 to 10 percent Spring Creek
water. Reduced neonate production in
tests 1 through 4 and in test 9 was in part
or totally due to mortality, and chronic
effects were not evident. Spring Creek
toxicity, resulting in chronic effects, was
greatest in tests 5 through 7 with signifi-
cantly lower neonate production at dilu-
tion volumes of 5 percent Spring Creek
water. Greater chronic toxicity in tests 6
and 7 was associated with greater acute
toxicity, as previously stated, and was due
to the unidentified toxicant. The increased
toxicity resulting in chronic effects in test
5 was due to either the initial occurrence
of the unidentified toxicant or to the
increase in total recoverable zinc and
copper on that day. Overall, the relation-
ship between toxicity and metal concen-
trations was poor. This was primarily due
to the occurrence of the unidentified
toxicant.
Control water toxicity was also evident
in tests 1 through 3 with significantly
lower neonate production in the control
tests relative to the culture water tests.
Bioassays conducted on water collected
from the tributary streams on October 16
revealed a potential source of control
water toxicity due to Copper Creek inflow,
located 100 m upstream from control
station 011. Copper Creek water was
chronically toxic, resulting in low neonate
production, and therefore may have been
a source of control water toxicity. A
significant difference in neonate produc-
tion was also found in test5, but this was
probably due to nutritional differences in
the culture water and control treatments,
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and not to control water toxicity. The
culture water supported high concentra-
tions of algae (Closterium) and bacteria,
and provided additional food for C.
reticulata in the culture water treatments.
This resulted in higher neonate produc-
tion in the culture water treatments for
almost all tests.
Downstream Station and Dilution Test
Comparisons—Prickly Pear Creek was
toxic to C. reticulata at the downstream
stations. Toxicity in the Spring Creek
dilution tests and in the downstream
Prickly Pear Creek tests was compared to
determine if downstream changes in
toxicity were due strictly to inflow of
Spring Creek water. Validity of test com-
parisons was based on dilution volumes
of Spring Creek water in the dilution and
downstream tests. Dilution volumes of
Spring Creek water at the downstream
stations 013, 014, and 018 were 17.3,
7.2, and 2.4 percent, respectively, and
were similar to dilution volumes of Spring
Creek water used in the C. reticulata
dilution tests (20, 10, and 2.5 percent).
Mortality in dilution and downstream
tests having comparable Spring Creek
dilution volumes showed a high degree of
similarity. However, higher mortality in
some downstream tests indicated that
there was an additional downstream
source of toxicity and that the toxicant
may have been similar in nature to the
unidentified toxicant in Spring Creek.
Neonate production in dilution and
downstream test comparisons also
showed no significant difference in a
majority of the tests. However, there was
a trend for lower neonate production in
the downstream tests in most tests. This
trend in highertoxicity in the downstream
tests resulted from either additional
downstream sources of toxicity or
downstream enhancement of Spring
Creek toxicity.
Although one or both of the above
processes may have occurred,
differences in test comparisons were
minimal and did not refute that variations
in Prickly Pear Creek toxicity were
primarily due to downstream dilution of
the Spring Creek inflow. Downstream
toxicity persistence, therefore, did appear
to follow a conservative behavior.
Pimephales promelas
Larval fathead minnows were more
tolerant to Spring Creek toxicity than
were C. reticulata. Estimated LC-50s for
fathead minnows were at control dilution
volumes greater than 25 percent Spring
Creek water, which were greater than
dilution volumes found for downstream
Prickly Pear Creek stations. This was also
reflected in the downstream station tests
which showed little or no mortality.
Fathead minnow LC-50s indicated that
Spring Creek toxicity was highly variable
for this species. Minimal mortality occur-
red in tests 2, 8, and 9, and acute effects
were not evident for those days. There
was a significant decline in toxicity in
tests 6 and 7 indicating thatthe unidenti-
fied toxicant resulting in toxicity to C.
reticulata was not at toxic concentrations
for fathead minnows. Higher toxicity in
tests 0,1, and 5 did correspond to higher
total recoverable concentrations of zinc
and copper; however, a strong relation-
ship for these metal concentrations and
toxicity was not clearly evident.
Part of the variability found in the
fathead minnow test was probably not
inherently related to Spring Creek toxicity.
High control mortality occurred after the
third or fourth day and at test termination,
mortality was greater than 30 percent in
six of the 10 tests (0, 1, 2, 4, 7, and 8).
High control mortalities are usually indic-
ative of procedural problems; however,
mortality declined in the lower dilution
treatments with little or no mortality at
either 12.5 or 25 percent in all tests. The
consistent decline in lower dilution treat-
ment mortality relative to high control
mortality strongly suggested that Spring
Creek water was ameliorating conditions
in the control water. This mortality decline
may have been due either to dilution of
control water toxicity or to the addition of
some factor enhancing survival. Control
water toxicity was evident in C. reticulata
bioassays; however, reconstituted water
controls included in fathead minnowtests
6 and 7 suggested that control mortality
was not due to toxicity.
The inherent growth variability in fat-
head minnows precluded demonstrating
chronic effects. Growth was significantly
increased with increased feeding in a
separate feeding experiment, indicating
test fish were probably underfed and that
a quantitative food regime should be
developed for future tests. Nevertheless,
fathead minnows raised at the Las Vegas
Laboratory from identical egg batches
showed variations in length approaching
400 percent after 30 days. This kind of
growth variability would highly influence
test results and a nonfeeding lethality
test may be more appropriate for field
testing.
Stream Survey
Water Quality
Nonmetal water quality parameters
measured in the stream survey did not
reveal any other sources of toxicity or
toxicants. Total organic carbon concen-
trations were low, ranging from 2 to 3
pg/l, and ammonia concentrations were
below detection limits, indicatng little or
no contributions from either septic tanks
or domestic animals within the study
area. Cyanide and chlorine were also
below detection limits. Spring Creek ion
concentrations were moderate, with a
conductivity of 421 /jmhos/cm,2.7times
greater than at control station 011.
Conductivity at station 013 was 226
(jmhos/cm, but increased to 269
/umhos/cm at station 018 as a result of
additional secondary inflow sources high
in ion concentrations downstream from
Spring Creek. This downstream increase
was also reflected in alkalinity and hard-
ness, which showed similar downstream
trends. Turbidity in Spring Creek was
higher than in Prickly Pea/ Creek; how-
ever, water clarity or suspended solids
were not a water quality problem during
this investigation. Temperature, dissolved
oxygen and pH levels were typical of fall
conditions for temperate streams and
were indicative of good water quality.
Hydrology
Stream flow at the U.S. Geological
Survey gaging station, located 2 km
downstream from station 018, ranged
from 35 to 42 cubic feet per second (cfs)
during the toxicity testing period and was
typical of seasonal low flows over the last
3 years. Peak flow on October 2 was due
to a small storm event that occurred on
September 30 and to snow melt from a
storm that occurred on September 18.
Stream flow in Prickly Pear Creek
increased from 11 cfs at station 011 to 37
cfs at station 018. Measured tributary
inflows accounted for 62 percent of the
increase in flow. Estimated unmeasured
inflows between stations 013, 014, and
018 were approximately 5 and 8 cfs,
respectively. There were no other major
surface inflows and the majority of the
unmeasured increase in flow was due to
groundwater inputs.
The percent of Spring Creek water
volume to the total water volume at
downstream Prickly Pear Creek stations
013,014, and 018 was 17.3, 7.2, and 2.4
percent, respectively, based on concen-
trations of Rhodamine WT injected into
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Spring Creek on September 23. Dye
retention time from the Spring Creek
confluence to station 018 was just over
11 hours.
Biota
Salmonid fishes were abundant at all
Prickly Pear Creek stations. However,
there was a downstream shift in species
abundance. Brook trout (Salvelinus fon-
tinalis) was the only salmonid found at
station 011. Both brooktrout and rainbow
trout (Salmo gairdneri) were abundant at
stations 013 and 014, and the brown
trout (Salmo trutta) also occurred with the
other salmonid species at station 018.
The species shift in salmonids was prob-
ably not due to metal toxicity from Spring
Creek, but rather to the increased fre-
quency of pool habitats downstream.
Previous investigations have shown
major reductions in both macroinverte-
brate and periphyton numbers and diver-
sity in the Prickly Pear Creek impact zone
(station 013) and a gradual downstream
reoccurrence of these species between
stations 014 and 018, the recovery zone.
These studies were conducted in the
summer. Quantitative analyses of peri-
phyton and macroinvertebrate samples
were not part of this investigation. A
superficial examination of the macroin-
vertebrate samples at the time of collec-
tion indicated no evident reduction in
either species types or species number in
the impact zone. This lack of reduction
may have been a physiological response
to temperature and needs to be validated
quantitatively. Water temperatures dur-
ing this investigation were approximately
7°C compared to summer temperatures
of 16 to 20°C.
Metals in Sediment and Tissue
Sediment metal concentrations at
station 013 were approximately an order
of magnitude higher than those found at
the upstream stationO! 1. Sediment metal
concentrations at Spring Creek and sta-
tion 013 were similar, indicating high
sediment deposition from Spring Creek in
the area of station 013. Sediment con-
centrations decreased downstream from
station 013 and were four to five times
lower at station 018. However, concen-
trations at station 018 were substantially
higher than concentrations found at the
control station 011, further demonstra-
ting the extent of downstream impacts
from Spring Creek. Sediment metals were
a potential source of downstream toxicity.
However, hydrological conditions during
the testing period were very stable, and
increased downstream water metal con-
centrations resulting from sediment
resuspension probably did not occur or
were very minimal. Sediment-water in-
teractions were not determined in this
study and should be investigated to
determine the extent sediments act as a
source of or sink for metals under various
hydrological conditions in Prickly Pear
Creek.
Tissue metal concentrations were
highest in periphyton followed by macro-
invertebrates and fish. Periphyton and
macroinvertebrate tissue concentrations
were substantially higher at station 013
and decreased downstream relative to
ambient water and sediment concentra-
tions. Metal uptake by periphyton and
macroinvertebrates represented a poten-
tial metal sink; however, these organisms
are also a source of metals when ingested
by other organisms. Fish tissue metal
concentrations were not exceptionally
high and there was no substantial dif-
ference in tissue concentrations between
stations. Previous investigations have
found significantly higher tissue metal
concentrations in most organs (kidneys,
gills, brains, heart, and gonads) from fish
collected in the impact areas of Prickly
Pear Creek. However, muscle tissue did
not have elevated metal concentrations.
In this investigation, whole fish were
used for tissue analyses and the inclusion
of muscle tissue probably masked metal
concentrations in the organs.
General Discussion
The results of the Prickly Pear Creek
instream toxicity measurements (after
mixing of the effluent), indicate such
methods along with assessments of
stream biota communities can provide a
great deal of information about the nature
and extent of effluent impacts on resident
biota. These analyses aid in identifying
needs for limiting effluent toxicity.
An important and often overlooked
aspect of effluent and instream toxicity is
the persistence of toxicity within a re-
ceiving system and the potential spatial
extent and severity of impact to the biota.
Obviously, pollutants that are rapidly
degraded to nontoxic forms, lost to the
atmosphere, or rendered unavailable
through other processes pose far less
threat to biota than the more persistent
forms.
Conditions in Prickly Pear Creek were
such that conservative behavior in toxicity
was favored. Hydrological retention time
from the Spring Creek confluence to the
downstream Prickly Pear Creek stations
was relatively short (11 hours) and would
limit oxidation and reduction processes,
especially at stream temperatures (7°C)
found during this investigation. Sus-
pended solids and organic compounds
were also very low and toxicity was not
highly influenced by particle adsorption
or by complexing with organic com-
pounds. Prickly Pear Creek was not
acutely toxic to larval fathead minnows,
and bioassay results were not applicable
to test ing the fate of Spring Creek toxicity.
C. reticulata was more sensitive than
fathead minnows and bioassay results
did demonstrate conservative behavior in
toxicity.
Toxicity found in both test organisms
clearly paralleled past changes in native
fish and macroinvertebrate communities
attributed to toxicity. Distribution and
abundance of fish and macroinverte-
brates in Prickly Pear Creek have been
well documented for summer conditions
in previous investigations. These studies
have shown that toxicity from Spring
Creek has little or no effect on native fish,
but a definite impact zone and recovery
zone were found in Prickly Pear Creek
relative to macroinvertebrate community
diversity and species abundance. If fat-
head minnows and C. reticulata bioassays
from this study were used to predict
downstream biotic conditions in Prickly
Pear Creek, identical impacts and/or
zones would be designated. Therefore, it
does appear that these bioassays reflect
summer levels of toxicity affecting native
fish and macroinvertebrate communities
in this system.
The work reported here, addressing
toxicity persistence in a receiving stream,
represents an initial step toward field
validation of procedures for establishing
water quality based effluent limits using
biological data. Although the primary
source of metals to Prickly Pear Creek
was a tributary stream rather than an
effluent pipe. Spring Creek was treated as
an effluent for which load limits and
required reductions could be established
and a permit issued. Data from this and
other projects will provide information on
the conservative (or nonconservative)
nature of various types of pollutants in a
range of receiving systems. Such case-
history information will enable EPA's
Office of Water Regulations and Stand-
ards to assess the validity of the mass
balance modeling approach to predicting
instream toxicity persistence. An eventual
goal is to include in this testing all steps
leading to, and including, the issuance of
permits using biological data. Analyses to
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be included in these tests would consist
of:
1. identification of water quality lim-
ited systems;
2. water body survey and assessment;
3. review and, if necessary, revision of
designated uses;
4. establishment of appropriate criter-
ia;
5. performance of waste load alloca-
tion; and
6. identification of control technology
requirements.
These analyses, when completed, would
result in assurance of a water quality
based permit that would allow the water
quality standard to be met. Issuance of
the permit would be followed by monitor-
ing to ensure water quality improvements
are being achieved.
Conclusions
Metal concentrations in Prickly Pear
Creek were significantly increased down-
stream from its tributary Spring Creek,
which produced elevated levels due to
gold mining tailing and settling ponds in
the drainage basin. Concentrations of
cadmium, zinc, and copper measured over
a 10-day period exceeded EPA acute
criteria for aquatic life at one or more of
the downstream sampling stations in
Prickly Pear Creek. Sediments were a
potential downstream source of metals,
but probably did not contribute to ambient
water metal concentrations due to stable
hydrological conditions. Elevated metal
concentrations were the only water qual-
ity problems observed in Prickly Pear
Creek during this investigation.
Spring Creek toxicity to test organisms
(C. reticulata and P. promelas) was pri-
marily due to zinc and copper. Other
unidentified toxicants were present and
Spring Creek was ot the only tributary
serving as a source of toxicity for Prickly
Pear Creek waters. Although there were
additional sources, changes in toxicity
(persistence) in Prickly Pear Creek were
primarily due to downstream dilution of
Spring Creek water. Therefore, Spring
Creek toxicity did exhibit a conservative
behavior in its downstream distribution in
Prickly Pear Creek and complied with
toxicity model assumptions.
Sensitivity of the two test organisms to
toxicity in Spring Creek and Prickly Pear
Creek was very different. C. reticulata
was highly sensitive, and bioassay results
were applicable in assessing toxicity
persistence in Prickly Pear Creek. P.
promelas had a higher tolerance and
could not be used in assessing toxicity
persistence. Although sensitivity of larval
fathead minnows and the cladoceran was
different, both appeared to be highly
representative of toxic effects in Prickly
Pear Creek native fish and macroinverte-
brate communities found in previous
studies.
Problems were encountered in the field
bioassay procedures used for both orga-
nisms. These problems were related to
the food regime used in each of the
bioassays. Cerophyl proved to be a better
food source than yeast in C. reticulata
tests. Chronic toxicity was not measured
in P. promelas,, apparently because of
underfeeding, and either a quantitative
food regime should be developed for this
test or a nonfeeding test should be used
in future field testing.
JohnR. Baker and Barry P. Baldigoare with Lockheed Engineering & Management
Services Co., Inc., Las Vegas, NV 89114.
Wesley L. Kinney is the EPA Project Officer (see below).
The complete report, entitled "Toxicity Persistence in Prickly Pear Creek,
Montana," (Order No. PB 85-137 149; Cost: $11.50, subject to change) will be
available only from:
National Technical Information Service
5285 Port Royal Road
Springfield, VA 22161
Telephone: 703-487-4650
The EPA Project Officer can be contacted at:
Environmental Monitoring Systems Laboratory
U.S. Environmental Protection Agency
P.O. Box15027
Las Vegas, NV 89114
. S. GOVERNMENT PRINTING OFFICE:1985/559-l 11/10768
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