DOC
EPA
United States
Department of
Commerce
National Oceanic and
Atmospheric Administration
Boulder CO 80303
United States
Environmental Protection
Agency
Office of Environmental
Engineering and Technology
Washington DC 20460
EPA-600/7-81-093
July 1981
Research and Development
The Effects of
Petroleum
Hydrocarbons on
Chemoreception and
Behavior in the
Dungeness Crab
Cancer magister
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EPA-600/7-81-093
July 1981
THE EFFECTS OF PETROLEUM HYDROCARBONS ON CHEMORECEPTION
AND BEHAVIOR IN THE DUNGENESS CRAB
CANCER MAGISTER
by
B.L. Olla
NOAA, NMFS
Northeast Fisheries Center
Sandy Hook Laboratory
Highlands, New Jersey 07732
W.H. Pearson, P.C. Sugarman, D.L. Woodruff, and J.W. Blaylock
Battelle Pacific Northwest Laboratories
Marine Research Laboratory
Sequin, Washington 98382
NOAA Project Officer: Douglas A. Wolfe (NOAA/Boulder, CO)
This study was conducted as part of the Federal Interagency
Energy/Environment Research and Development Program
Prepared for the Office of Research and Development
U.S. Environmental Protection Agency
Washington, D.C.
T
5, Librauy (5PL-16)
B^om Stroet. Soos le?
. IL 60604
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DISCLAIMER
This work is the result of research sponsored by the Environmental
Protection Agency and administered by the Environmental Research Labora-
tories of the National Oceanic and Atmospheric Administration.
The Environmental Research Laboratories do not approve, recommend or
endorse any proprietary product or proprietary material mentioned in this
publication. No reference shall be made to the Environmental Research
Laboratories or to this publication furnished by the Environmental Research
Laboratories in any advertising or sales promotion which would indicate or
imply that the Environmental Research Laboratories approve, recommend, or
endorse any proprietary product or proprietary material mentioned herein,
or which has as its purpose an intent to cause directly or indirectly the
advertised product to be used or purchased because of this Environmental
Research Laboratories publication.
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FOREWORD
The accelerated development of petroleum resources on the continental shelves
of the United States, along with continued importation of petroleum from
foreign sources, is expected to increase the transfer and refinement of
petroleum in coastal areas. In order to properly evaluate the potential
consequences of this increased petroleum flow, NOAA is conducting studies in
the Fate and Effects of Petroleum Hydrocarbons in Selected Marine Organisms
and Ecosystems. The overall objectives of this project are to study experi-
mentally those specific processes controlling the distribution, transport,
and effects (physiological, behavioral, and ecological) of petroleum hydro-
carbons in coastal marine systems. These studies are expected to facilitate
the assessment of environmental impacts of petroleum releases and to serve as
the basis for developing regulatory measures for suitable protection of the
marine environment. The study reported here addresses the question of
whether chronic low-level petroleum inputs might interfere with critical,
life-sustaining behavioral processes in a nearshore, commercially-important
marine species. This report presents the results of three years of coopera-
tive effort between B. L. Olla, Chief of Behavioral Investigations at NOAA,
NMFS Sandy Hook Laboratory, Highlands, New Jersey, and W. H. Pearson of
Battelle's Marine Research Laboratory, Sequim, Washington. Each section of
the report represents work published, in press, or near final preparation for
journal submission.
Douglas A. Wolfe
NOAA/Office of Marine Pollution Assessment
TM
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ABSTRACT
The behavior of Dungeness crabs, Cancer magister, was observed to
determine not only whether oil exposure produced behavioral effects, but
also whether crabs could change their behavior to mitigate any exposure
effects. Dungeness crabs clearly detected the presence of petroleum hydro-
carbons but did not avoid oil under all circumstances. Changes in the
antennular behavior of crabs showed that they detected naphthalene at 10 2
mg/L and the water-soluble fraction of Prudhoe Bay crude oil at 4 x 10 4
mg/L. Thus, the crabs detected dissolved petroleum hydrocarbons at concen-
trations below those typical of oil spills. Behavioral observations of
crabs given a choice between clean and oiled sand indicated that crabs
avoided highly oiled sand (1000 - 2000 ppm) to some extent but spent more
time in moderately oiled (<200 ppm) than clean sand. Because the extent of
avoidance varied highly and was apparently related to factors intrinsic to
the animal and its environment, we must assume that avoidance of oiled sand
is not assured.
The effects of oil exposure on chemoreception and feeding behavior in
Dungeness crabs were determined after measuring the high sensitivity of the
crabs to chemical food cues. Abrupt changes in antennular orientation and
sharp increases in antennular flicking rate indicated that crabs detected
an extract of littleneck clam, Protothaca staminea, at 10 7 mg/L. At 10
mg/L of the extract crabs probed the substrate with the chelae or showed
other feeding behavior. After 24 h of continuous exposure to 0.3 mg/L of
oil-contaminated water and with oil still present, the proportion of crabs
showing the changes in antennular behavior indicating detection of chemical
food cues was significantly reduced. In contrast, the proportion showing
chelae probing was not. Within one hour after return to clean water the
antennular response recovered. Such rapid recovery suggests that the
impairment was due to light anesthesia of chemosensory cells or, more
likely, masking of food cue odor by oil. Petroleum hydrocarbons impaired
the distance chemoreception seated in the antennules of Dungeness crabs
and, thereby, could cause crabs some difficulty in finding food. Field and
laboratory experiments then examined how oiled sediment influenced predation
on littleneck clams by Dungeness crabs. In field enclosures, crabs consumed
more clams from oiled than clean sand. A laboratory experiment indicated
that the observed increase in predation derived in large part from increased
prey accessibility due to an oil-induced change in clam burrowing behavior.
The potential difficulty in finding food due to chemosensory disruption by
petroleum hydrocarbons was apparently offset by an oil-induced change in
prey behavior. To the extent that oiled sediment renders prey species more
vulnerable to crab predation and crabs switch prey, harvesting of vulnerable
prey by crabs would reduce their representation in the benthic fauna and
produce ecological effects far different than those predicted from a series
of conventional toxicity tests.
IV
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ANNOTATED TABLE OF CONTENTS
Page
FOREWARD iii
ABSTRACT iv
FIGURES vii
TABLES ..... viii
INTRODUCTION 1
CONCLUSIONS AND RECOMMENDATIONS .... 2
I. BEHAVIORAL MITIGATION OF PETROLEUM EFFECTS ... 4
A. Detection of petroleum hydrocarbons by the
Dungeness crab, Cancer magister 7
Materials and Methods . 7
Results 0
Discussion 12
B. Avoidance of oiled sediment by the Dungeness crab,
Cancer magister 13
Materials and Methods 14
Results 16
Discussion 25
II. BEHAVIORAL EFFECTS OF PETROLEUM EXPOSURE 26
A. Thresholds for detection and feeding behavior
in the Dungeness crab, Cancer magister 26
Materials and Methods 27
Results 31
Discussion 37
B. Impairment of chemosensory food detection in
the Dungeness crab, Cancer magister, by
petroleum hydrocarbons 39
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Materials and Methods 39
Results 41
Discussion 47
C. Effects of oiled sediment on predation of the
littleneck clam, Protothaca staminea, by
the Dungeness crab, Cancer magister 48
Materials and Methods 49
Results 51
Discussion 61
REFERENCES 64
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FIGURES
Figure 1. A flow-chart indicating possible behavioral
responses to an environmental perturbation and
their consequences
Page
Figure 2. The percentage of Dungeness crabs detecting naphthalene
and the water soluble fraction (WSF) of crude oil as
a function of the logarithm of concentration
Figure 3. A schematic diagram of the chemosensory testing
apparatus
11
29
Figure 4. The percentage of Dungeness crabs detecting the
freeze-dried clam extract (FDCE) or beginning feeding
behavior as a function of the logarithm of the FDCE
concentration within the testing chamber
34
vn
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TABLES
Page
Table 1. The composition of the WSF of Prudhoe Bay crude oil 10
Table 2. Experimental conditions in experiments concerning
avoidance of oiled sand 18
Table 3. Time spent by Dungeness crabs in clean and oiled sand
in the unpartitioned tanks 19
Table 4. Oil-in-sand concentrations in partitioned tanks determined by
IR spectrophotometry 20
Table 5. Total concentrations of monoaromatic hydrocarbons in
partitioned tanks determined by helium equilibration
gas chromatography 21
Table 6. Average percentages of the total crab-hours observed
in partitioned tanks that Dungeness crab spent in
oiled sand 22
Table 7. Average percentages of the crab-hours observed in
partitioned tanks buried and resting that Dungeness crabs
spent buried and resting in oiled sand 23
Table 8. Average percentages of total crab-hours observed in
partitioned tanks that Dungeness crabs were active 24
Table 9. The occurrences of specific components of feeding
behavior at various FDCE concentrations 35
Table 10. The antennular flicking rate ratios at various FDCE
concentrations 36
Table 11. The concentrations of monoaromatic hydrocarbons in the
chemosensory testing chambers 43
Table 12. Percentage of Dungeness.crabs detecting the clam extract
[FDCE] after exposure to continuously-flowing sea water
contaminated with Prudhoe Bay crude oil 44
Table 13. Percentage of Dungeness crabs probing with the chelae
upon presentation of a clam extract [FDCE] after
exposure to continuously-flowing sea water contaminated
with Prudhoe Bay crude oil 45
vm
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Page
Table 14. Percentage of Dungeness crabs responding to a clam
extract [FDCE] presented in clean water 1 h after a
24-h exposure to oil-contaminated sea water 46
Table 15. The total concentration of oil in sand inside the
field enclosures ............ 54
Table 16. Hydrocarbon concentrations in sand during 29-day field
experiment 55
Table 17. Percentage of total number of clams and estimated
shucked wet weight that were consumed by crabs 56
Table 18. Average number of intact unburied clams observed in
the field and laboratory experiments . 57
Table 19. Depth distribution of clams recovered at the end of
the field and laboratory experiments 58
Table 20. Clams consumed by crabs as percentage of total number
initially added in each size class 59
Table 21. Clams consumed in each size class as percentage of total
number of clams consumed 60
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INTRODUCTION
Changes in behavior have been shown to be quite sensitive indicators
of environmental stress in marine animals (Olla, 1974; Olla et al. , 1980a &
b). Generally, behavioral studies of pollutant effects have simply sub-
stituted behavioral disruption for the normal end-point of death used in
conventional toxicity assays. Rarely have these kinds of studies either
choosen ecologically pertinent behavior for investigation or interpreted
any observed behavioral changes on the basis of a thorough understanding of
the organism's behavioral ecology (Oila et al., 1980a & b). Still, more
rarely have such studies empirically followed the observed behavioral
effects to their ecological consequence (see Ward & Busch (1976), Ward et
al. (1976), and Krebs & Burns (1977) for examples of these rarer studies).
In the work presented here we have addressed broader questions than just
whether oil disrupts behavior and done so within a developing theoretical
framework (Olla et al., 1980a & b) that points out the ecologically meaningful
questions concerning behavioral ecological processes and the consequences
of their disruption. Our aim in this work was to examine the effects of
petroleum hydrocarbons on the behavioral ecology of adult Dungeness crabs,
Cancer magister.
Our research took two pathways. One pathway examined whether Dungeness
crabs could, within the scope of their genetically based norm of reaction,
change their behavior to mitigate any effects of petroleum and, thereby,
enhance the probability of survival. The other pathway investigated whether
exposure to petroleum would produce untoward changes in some important
behavior where the crab was unable to mitigate exposure behaviorally. The
crab's failure to mitigate petroleum effects could derive from an inherent
inability to change its behavior in the presence of petroleum or from an
externally imposed inability to change behavior adaptively under particular
environmental conditions. Research concerning behavioral mitigation of
petroleum effects centered on laboratory experiments that measured the
crab's ability to detect petroleum and then behave appropriately, i.e.,
avoid contact with oil-contaminated sand. Research concerning adverse
petroleum effects centered on laboratory experiments that determined how
petroleum modified chemosensory detection of food cues and field experi-
ments that examined crab predation on a natural prey buried in oiled sand.
The two sections of the report reflect our two pathways of research.
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CONCLUSIONS AND RECOMMENDATIONS
Our results demonstrate that the Dungeness crab, Cancer magister, can
detect dissolved petroleum hydrocarbons at concentrations well below those
typical of oil spill situations. In so doing, the crab can meet the first
prerequisite for any subsequent behavioral response that would mitigate any
effects of oil exposure.
Whereas detection of petroleum hydrocarbons clearly occurred, whether
the crab could behave appropriately after detection was not as clear.
Avoidance of oiled sediment did occur but not under all circumstances. The
extent of avoidance was not dramatic and seemed to vary as factors intrinsic
to the crab, the oil, and the environment interacted. Petroleum hydrocarbons
are one complex chemical stimulus among a host of other stimuli, both
chemical and of other modalities. How the animal responds depends on where
this complex stimulus is in a ranking that constantly changes as the animal's
environmental needs change. The variation in activity level and pattern
seen among the various experiments were related to variations in the phase
relationships between tidal cycles and photoperiod and were one example of
how the characteristics of the crab and the environment interacted.
Changes in the extent of avoidance with the oil concentration of the
sand were noted. Crabs showed no difference between control and oiled sand
in low concentrations (^20 ppm), seemed attracted to moderately oiled sand
(~200 ppm), and avoided the highly oiled sand (-^2000 ppm). These results
suggest to us that the crab is capable of avoiding oiled sediment at high
levels but that the occurrence and extent of avoidance depends on the
environmental conditions and such factors as hunger level, reproductive
condition, and migratory state, that are intrinsic to the crab. We must
conclude, therefore, that it is presently best to be conservative and
assume that avoidance of oiled sand is not assured.
In our laboratory studies of avoidance behavior in the Dungeness crab
(Pearson et al., in prep.) and elsewhere with hake (Pearson et al., in
prep.), and the blue crab (Pearson et al., in prep.), it is becoming
increasingly clear that factors intrinsic to the animal and the environment
apparently can override or at least lessen the extent of avoidance. Knowledge
of these factors is crucial both to realistic experimental design and to
meaningful extrapolation of laboratory results to the natural environment.
Even our brief field observations coupled with the laboratory studies make
us confident that more extensive study of crab behavior under natural
conditions would enable us to develop a model that would indicate the
circumstances under which crabs do and do not avoid oiled sand.
After establishing the acute sensitivity of Dungeness crabs to chemical
food cues, we exposed crabs under continuous-flow for 24 h to petroleum
hydrocarbons at concentrations typical of oil spill situations. Under the
24-h exposure, crabs showed impaired ability to detect chemical food cues.
Recovery from impairment was observed one hour after ending exposure to
oil-contaminated water. This rapid recovery suggested that either the
hydrocarbons anesthetized the chemosensory cells or, more likely, masked
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the food odor. Whichever the mechanism, the impairment would probably
continue as long as petroleum hydrocarbons were present, and the continuing
presence of petroleum hydrocarbons would likely make food finding difficult
for the Dungeness crab. Because the crab detects dissolved petroleum
hydrocarbons at 10 4 mg/L, we suspect that masking by even lower hydrocarbon
levels than we tested could disrupt the sensing of chemical food cues,
especially if the cues are quite dilute. Because even our brief exposure
at one level produced a clear impairment in a chemosensory ability that
supports a crucial process in the behavior and ecology of the crab, we feel
that the effects of chronic low level exposure on chemosensory processes
need study.
Whereas the observed impairment of distance chemoreception suggests
that crabs would have more difficulty sensing prey, the field and laboratory
experiments on crab predation unexpectedly showed crabs consuming more
clams from oiled than clean sand. The potential difficulty in detecting
prey by chemoreception in an oiled environment was apparently offset by a
change in the burrowing behavior of the clam that increased prey accessi-
bility. The implication of this change in prey accessibility is that
whereas oiling of the sediment may not directly lead to the death of buried
clams, oiled conditions could very well lead to serious inroads on the clam
population by crab predation. It is still possible that under other oiled
conditions, oiled sediment may act as a chemosensory barrier to the detection
of buried prey. Our attempt to trace empirically an observed behavioral
effect to its ecological consequences gave unexpected results that revealed
the complexity of the situation. Our experience here only reinforces our
view that the fruitfulness of examining pollutant effects rests on a com-
prehensive understanding of the behavioral ecology of all the organisms
involved.
In the continual search for sublethal effects of petroleum hydro-
carbons, there are clearly some sublethal effects on the Dungeness crab.
Our studies have only revealed a small portion of these effects. Even so,
we can clearly see how these observed effects influence the animal's relation-
ship to its environment. Any attempt to predictively model the fate of Dungeness
crabs under oil exposure must consider these demonstrated sublethal effects.
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I. BEHAVIORAL MITIGATION OF PETROLEUM EFFECTS
A change In behavior is the initial response of an animal to an environ-
mental perturbation (Slobodkin, 1968). If the behavioral response removes
or lessens the effect of the perturbation, the probability of death or the
cost incurred by other adaptive responses to maintain homeostasis may be
lowered or eliminated. The response may be movement away from the area of
the perturbation or other changes in behavior that effectively reduce
exposure. Generally, it is a fair statement that behavioral mitigation of
potentially stressful environmental changes is evident at almost all phylo-
genetic levels (for examples related to petroleum effects see Johnson,
1977;' 01 la et al. , 1980a).
For a successful behavioral response to an environmental perturbation
to occur, the animal must be capable of: 1) sensing it; 2) recognizing it
as aversive; and 3) responding appropriately (Pearson & 01 la, 1979; 01 la et
al., 1980a & b). In Figure 1, we have diagrammed possible response pathways
that may occur when an animal is subjected to an environmental perturbation.
Obviously no behavioral act can be elicited if the perturbation is not
sensed or is sensed at a level that will not permit the animal to respond
behaviorally before becoming debilitated. The inability to sense a stress-
inducing change in the environment is most likely to occur when the change
is novel, i.e., one which bears little similarity to past events (Slobodkin
& Rapoport, 1974).
Either behavioral or neurophysiological techniques may be used to
establish sensitivity, depending upon the specific question to be answered.
In this work we have used behavioral criteria exclusively because they have
proven to be quite effective for determining sensitivity to various sub-
stances. For example, utilizing changes in the rate of antennule flicking
and gill bailing, Pearson and 01 la (1977) showed the threshold concentration
at_which blue crabs, Callinectes sapidus, detected a food extract to be
10 12 mg/L. Blue crabs were also able to detect naphthalene (Pearson &
Olla, 1979) at a threshold concentration of 10~7 mg/L (Pearson & 01 la,
1980) and the water-soluble fraction (WSF) of crude oil at 10 6 mg/L (Pearson
et al., 1980a).
While detection may occur at very low concentrations, the elicitation
of more complex behaviors appears to require much higher concentrations.
In the blue crab, food searching did not occur until the concentration of a
food extract reached 10 mg/L, 13 orders of magnitude higher than the detection
level (Pearson & Olla, 1977). When presented with naphthalene, the blue
crab showed active locomotory behavior at a concentration of 2 mg/L, about
7 orders of magnitude higher than the detection level (Pearson & Olla,
1980).
For the Dungeness crab we used techniques similar to those we had with
the blue crab to ask whether and at what concentration the Dungeness crab
detected petroleum hydrocarbons. Detection of petroleum hydrocarbons is
the first step to behavioral mitigation of their effects. The following
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section, I.A., presents work soon to be published by Pearson, Sugarman,
Woodruff, Blaylock, and 01 la (1980b) and describes how Dungeness crabs
detect dissolved petroleum hydrocarbons at concentrations below those
typical of oil spills.
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ENVIRONMENTAL
PERTURBATION
ANIMAL
NO DETECTION
NO AVOIDANCE OR
OTHER NON-ADAPTIVE
BEHAVIORAL RESPONSE
DETECTION
EXPOSURE
LETHAL EFFECTS
AVOIDANCE OR
OTHER ADAPTIVE
BEHAVIORAL RESPONSE
REDUCED
EXPOSURE
Li
NO EXPOSURE
NO LETHAL EFFECT^
SUB-LETHAL
EFFECTS
INITIAL SURVIVAL
REDUCED LONGEVITY
DUE TO PHYSIOLOGICAL
OR BEHAVIORAL
DISRUPTION
REDUCED LONGEVITY
DUE TO '^OLOGICAL
STRESS
LONG-TERM
SURVIVAL
Figure 1. A flow-chart indicating possible behavioral responses to an
environmental perturbation and their consequences.
From 01 la et al.. 1980a.
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A. Detection of Petroleum Hydrocarbons by the Dungeness Crab, Cancer magister.
For decapod crustaceans the antennules have been considered the site
of distance chemoreception (Hazlett, 1971a), and their flicking may be
analogous to sniffing in vertebrates (Fuzessery, 1978). Previous work
has shown that in the blue crab, Callinectes sapidus, the antennular behavior
indicating detection of food substances (Pearson & Olla, 1977) also indicated
detection of the petroleum hydrocarbon naphthalene (Pearson & Olla, 1979,
1980) and the water soluble fraction of crude oil (Pearson et al., 1980a).
In the Dungeness crab, similar antennular behavior, i.e., a change in
orientation and increased flicking rate, also indicated detection of food
substances (Pearson et al., 1979). Here we used these changes in antennular
behavior to determine chemosensory detection thresholds in the Dungeness
crab for naphthalene and the water soluble fraction (WSF) of Prudhoe Bay
crude oil.
Materials and Methods --
Dungeness crabs trapped in the Strait of Juan de Fuca, Washington,
were held outdoors in 1200-liter tanks under the conditions described by
Pearson et al. (1979). The seawater temperatures during the naphthalene
and WSF experiments were 12.7 (± 0.6 SD)°C and 10.6 (± 0.3)°C; the salinities,
31.6 (± 0.9) °/00 and 32.0 (± 0.0 )°/00; the dissolved oxygen, 6.9 (±0.7)
mg/L and 7.3 (± 0.5) mg/L; and the pH, 8.12 (± 0.17) and 8.02 (±0.16),
respectively.
Experimental Solutions -- Saturated solutions of naphthalene were
prepared by adding naphthalene crystals to seawater filtered through a 0.4
(jm Nucleopore membrane. These stock solutions were stirred continuously at
room temperature on a magnetic stirrer and were used after at least 18 h of
stirring and no more than five days from first use. On each day of testing,
a portion of the stock solution was siphoned off and passed through a 100
ml glass syringe fitted with a Millipore prefilter (Type A025) to remove
any naphthalene crystals. Less than one hour before testing, serial dilutions
of this filtered stock naphthalene solution were made with seawater freshly
filtered through a 0.4pm membrane. An aliquot of the filtered seawater
used for dilution served as the control solution. Experimental and control
solutions were kept in a water bath at ambient sea-water temperature during
testing.
On each day of testing, samples of the stock solution and 10 1 dilution
were analyzed for naphthalene content. Ten milliliters of hexane were
vigorously shaken with 50 ml of sample solution for one minute. This
hexane was removed and analyzed for napthalene content by capillary GC
methods (Bean et al., 1978). The stock naphthalene solution was 22.9
(± 2.1) mg/L, and the 10'1 dilution was 2.2 (± 0.2) mg/L.
The water soluble fraction (WSF) of Prudhoe Bay crude oil was prepared
freshly each day by methods similar to Anderson et al. (1974). In a 19-liter
glass bottle, one part oil was gently poured over nine parts membrane-filtered
seawater. Before the oil was added, a glass siphon tube inserted through
a stopper covered with aluminum foil was placed in the filtered seawater.
With the bottle stoppered, the seawater was slowly stirred on a magnetic
stirrer for 20 h at room temperature. The stirring speed was adjusted so
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that the vortex did not extend more than 25% of the distance to the bottom
of the bottle. After mixing, the oil and water phases were allowed to
separate for one hour. The water phase was then siphoned from below the
oil phase and filtered through a prefilter under very low pressure to
remove any remaining oil droplets. Serial dilutions of the resulting WSF
were then immediately made with freshly membrane-filtered seawater and
kept in a water bath at ambient seawater temperature during use. The
membrane-filtered seawater used for dilution was the control solution.
The stock WSF was analyzed by capillary gas chromatography for di- and
tri-aromatic hydrocarbons (Bean et al. , 1978), and by gas partitioning
analysis modified from McAuliffe (1971) for monoaromatics.
Chemosensory Threshold Determination -- The apparatus and procedures
of Pearson et al. (1979) were used here. In brief, glass testing chambers
were arranged on four trays, 10 chambers to a tray, and the trays were
surrounded by blinds. The experimental solutions were introduced into each
testing chamber through an inlet manifold connected to a glass funnel. Sea-
water from dripper arms entered each funnel at a rate of 1.0 L/min. A
teflon delivery tube carried the experimental solutions to the funnel from
a buret calibrated to deliver 20 ml in 15 s.
To obtain a dilution factor for estimating the effective concentration
of experimental solutions within a testing chamber, seawater solutions of
14Onaphthalene (sp. act. 3.6 mCi/mmole, Amersham-Searle Corporation) were
introduced and samples taken at timed intervals from the midpoint of the
chamber and counted for radioactivity by liquid scintillation spectrometry.
The chamber contained a crab model displacing 701 ml, a volume typical of
the crabs tested. The maximum concentration in the chamber occurred 45 s
after 14C-naphthalene was added and was 0.0188 (± 0.0058 SD) times the
concentration of the introduced solution. This dilution factor did not
differ significantly from that found by Pearson et al. (1979) using a
visible dye.
Approximately 24 h before testing, crabs were transferred to the
testing chambers from the holding tanks where they had been fed an ad
libitum diet of blue mussels, Myti1 us edulis. Because, in preliminary
experiments, tidal phase was found to influence chemosensory responses
(Pearson et al. , 1979), testing was synchronized to begin and end within
either a rising or falling tide. The seawater for the test dilutions and
control was drawn and filtered one hour after a tidal change. Testing then
began as soon as possible and stopped before the next tidal change.
Each day a maximum of 40 crabs were presented individually with 20 ml
of either one of nine dilutions of naphthalene stock solution, one of eight
dilutions of WSF, or a control of filtered seawater. Molting and mating
crabs were not tested. The order in which individual crabs were watched
and the choice of experimental solution were randomized except that active
crabs and ones with retracted antennules were passed over. The observer
did not know the identity of any test solution. Individual crabs were
observed for 1.0 min prior to introduction of the experimental solution,
and their antennular flicking rate and other behavior recorded. The flicking
rate of one antennule was measured using a hand-held counter. The solution
was then introduced, and the observations continued for 1.0 min after the
beginning of solution addition. The behavior was scored with the criteria
used by Pearson et al. (1979\
8
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To be scored as detecting an experimental solution, a crab had to
exhibit an abrupt change in the orientation of the antennules within 30 s
after solution introduction, and the ratio of the antennular flicking rate
for 1.0 min after solution introduction to that for 1.0 min before had to
be 1.50 or above. This value was determined previously by Pearson et al.
(1979) from observations of crabs in the testing apparatus without any
solutions present. Because 1.50 was the 95th percentile of these anten-
nular flicking rate ratios, the a priori probability that a flicking rate
ratio greater than 1.50 represented a spontaneous increase rather than a
reaction to the experimental solution was less than 5%.
Results --
Composition of the WSF -- The monoaromatic hydrocarbons by far dominated
the WSF (Table 1.) and comprised 99.1% of the total hydrocarbons measured.
The remaining aromatic hydrocarbons, mostly the naphthalenes, were present
at concentrations 100 times less than that of the monoaromatics. The
hydrocarbons partitioned into the WSF from the crude oil in proportion to
their solubility in seawater (Clark & MacLeod, 1977; Bean et al., 1978).
Detection Thresholds — Dungeness crabs detected both naphthalene and
the water soluble fraction (WSF) of Prudhoe Bay crude oil, with the complex
mixture (WSF) being more readily and consistently detected. Because the
percentage of crabs detecting naphthalene varied widely over the range of
concentrations presented, the regression equation relating percentage
detection and the logarithm of concentration was not significant (F = 1.3,
P = 0.30) (Fig. 2). The curve for naphthalene detection was sawtooth-shaped
with the percentage of detection being high at 10_8 mg/L and approaching
control valves at other concentrations. Above 10 6 mg/L, however, the
percentage of crabs detecting naphthalene rose linearly with concentration
and the regression equation was significant (F = 29.1; P = 0.01) and of low
variability (R2 = 94%). The threshold concentration at which 50% of the
crabs detected naphthalene, calculated from this latter regression equation,
was 3 X 10 2 mg/L. In contrast to naphthalene, the percentage of crabs
detecting the WSF decreased in a consistent way with the WSF concentration
(Fig. 2). The regression equation was significant (F = 60.4, P « 0.01),
and the variability was low (R2 = 91.0%). The 50% detection threshold was
4 x 10 4 mg/L, about 100 times lower than that for naphthalene.
When a crab detected naphthalene or WSF, the response was usually
distinct. For crabs meeting the detection criteria, the median ratios of
the antennular flicking rates did not vary with concentration (Median
Tests, x2 = 2.38, P = 0.12 for naphthalene; x2 = 9-07, P = 0.75 for WSF),
so that what varied with concentration was the percentage of crabs responding
and not the magnitude of the response. Also, the magnitudes of the increase
in antennular flicking were the same for both naphthalene and WSF. For
naphthalene, the grand median of the antennular flicking rate ratios was
2.04; for the WSF, the grand median was 1.96.
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Table 1. The composition of the WSF of Prudhoe Bay crude oil. Sample
size was 3 for the di- and triaromatics and 6 for the mono-
aromatics.
mg/L
TOTAL ALKANES < 0.001
NAPHTHALENE 0.0851 ± 0.0088
TOTAL METHYLNAPHTHALENES 0.0766 ± 0.0080
TOTAL DIMETHYLNAPHTHALENES 0.0269 ± 0.0015
PHENANTHRENE 0.0006 ± 0.0004
METHYLPHENANTHRENE < 0.0001
DIMETHYLPHENANTHRENE < 0.0001
TOTAL POLYNUCLEAR AROMATICS 0.1892 ± 0.0175
BENZENE
TOLUENE
ETHYLBENZENE
m + p XYLENE
o-XYLENE
TOTAL TRIMETHYL BENZENES
TOTAL MONOAROMATICS
TOTAL HC MEASURED
10.00
6.74
0.30
1.12
1.12
0.46
19.75
19.94
± 0.29
± 0.42
± 0.02
± 0.06
± 0.08
± 0.12
± 0.86
From Pearson et al., 1980b.
10
-------
Q
OQ
O
t/i
70
60
50
40
30
A NAPHTHALENE
O-— WSF
Y- 54.1 +1.22 X
R2 = 91%
LOG 1QCONCENTRATION (MG/L)
Figure 2. The percentage of Dungeness crabs detecting naphthalene
and the water soluble fraction (WSF) of crude oil as a
function of the logarithm of concentration (mg/L). The
percentage of crabs detecting a control of membrane-
filtered seawater was 28.8% (n = 66) for naphthalene
and 26.8% (n = 41) for WSF. The number beside each point
is the number of trials at the concentration.
From Pearson et al., 1980b.
11
-------
Discussion --
When presented with naphthalene or WSF of crude oil, Dungeness crabs,
Cancer magister, changed antennular orientation and flicking rate in the
same manner as when presented with a clam extract. The blue crab, Cal1inectes
sapidus, also gives the same detection behaviors for hydrocarbons as for
food (Pearson & Olla 1977, 1979, 1980; Pearson et al., 1980a), and the
similar findings in both species indicate that chemoreception by these
crustaceans is not restricted to chemical cues for food and, thus, agree
with Ache's (1975) suggestion that the chemical spectrum sensed by decapod
crustaceans is really quite broad.
While the manner of antennular response to naphthalene and WSF was the
same as that to a clam extract, the magnitudes of the flicking increase
were slightly less and the chemosensory thresholds were 105 and 103 times
higher than those found for the clam extract (Pearson et al., 1979). The
grand median ratios of flicking rates for naphthalene and WSF, 2.04 and
1.96, respectively, were lower than that for the clam extract, 2.67. Also,
the ranges of flicking ratios for the hydrocarbons were less than 30% of
that for the clam extract. The slightly less intense response and much
higher thresholds suggest that the petroleum hydrocarbons rank as much less
potent chemical cues than sapid chemicals from a natural food.
Previously, Pearson and Olla (1980) had hypothesized that the chemical
and chemosensory processes producing a higher detection threshold for a
single petroleum hydrocarbon, naphthalene, than for a complex mixture of
hydrocarbons, the WSF of crude oil, are analogous to the processes producing
a similar relationship of thresholds for single amino acids and complex
mixtures. Usually, food extracts and complex mixtures of amino acids and
other chemicals have a lower detection threshold than that of a single
amino acid (Mackie, 1973; McLeese, 1974). Indeed, with the Dungeness crab
the detection threshold for WSF was two orders of magnitude lower than that
for naphthalene. Also, the variability in detection was much less for WSF
than for naphthalene. This apparent greater difficulty in detecting the
single hydrocarbon than the more complex WSF is presumptive evidence for
the hypothesized analogy. With naphthalene constituting only 0.4% of the
total hydrocarbons in the WSF, the crabs were probably responding primarily
to other compounds or, perhaps, to some sort of odor medley.
One possible explanation for the extreme variability in naphthalene
detection is that detection at high naphthalene concentrations was inhibited
by some toxic, narcotic, or anesthetic action not present or much reduced
at low concentrations. The blocking of chemosensory feeding and mating
responses in the crab, Pachygrapsus crassipes, after 24~h exposure to
naphthalene at 10 3 mg/L (Takahashi & Kittredge, 1973) supports the pos-
sibility of such inhibition. If the threshold concentration for chemosensory
inhibition was within the range of concentrations we presented, then a
sharp increase in the percentage of crabs detecting naphthalene would be
expected below the inhibition threshold and would produce the sawtooth-shaped
curve seen for naphthalene in Figure 2. A sawtooth-shaped curve would also
result if the sensitive antennular chemoreceptors were more impaired than
the less sensitive body chemoreceptors on the dactyls, chelae, and mouthparts.
If so, detection would occur mainly through the body chemoreceptors at high
naphthalene levels and would switch to the antennules at low levels below
12
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the relatively high detection threshold of the body chemoreceptors and
where any inhibitory effects of the naphthalene on the antennules were
lessened.
For both food extract and petroleum hydrocarbons, the blue crab has
exhibited more acute chemoreception than the Dungeness crab (Pearson &
Olla, 1977, 1979, 1980; Pearson et al., 1980a). Pearson et al. (1979)
hypothesized that the lower detection threshold for clam extract seen in
the blue crab was a consequence of the blue crab's greater ability to
sample the chemical environment with its higher flicking rate and larger
antennules. This hypothesis would apply equally to the differences between
the two crabs in the hydrocarbon detection thresholds.
An important practical question is how the ability of the Dungeness
crab to detect petroleum hydrocarbons compares with the range of hydro-
carbon concentrations likely to be encountered by the crab. In the water
column during an oil spill, McAuliffe et al. (1975) found concentrations of
dissolved hydrocarbons ranging from 2 x 10 3 to 2 x 10 1 mg/L. Of these
dissolved hydrocarbons about one half were the monoaromatics dominating the
WSF used here. During a spill from a North Sea platform, Grahl-Nielson
(1978) found petroleum hydrocarbon concentrations ranging up to 4 x 10 1
mg/L. In the open sea between Nova Scotia and Bermuda, Gordon_et al.
(1974) found petroleum hydrocarbon concentrations of 2.04 x 10 2, 8 x 10 4,
and 4 x 10 4 mg/L at the surface, 1 m and 5 m, respectively. These con-
centrations roughly agree with those given for relatively uncontaminated
oceanic areas by Clark and MacLeod (1977), who also stated that chronically
contaminated areas have hydrocarbon concentrations about two orders of
magnitude higher than those of the open sea. Unfortunately, analytical
difficulties in distinguishing petrogenic from biogenic hydrocarbons at low
environmental concentrations make estimates of oil levels in chronically
contaminated areas uncertain. For the North Sea, Grahl-Nielsen et al.
(1979) found that despite considerable oil production there was no apparent
standing crop of petroleum hydrocarbons, but rather petroleum contamination
occurred as localized, transient patches. Thus, the petroleum hydrocarbon
concentrations in uncontaminated (10 4 to 10 3 mg/L), chronically contami-
nated (10~4 to 10"2 mg/L), and oil spill (10~3 to ID'1 mg/L) situations
are all at or above the WSF detection threshold (10 4 mg/L) so that Dungeness
crabs can detect hydrocarbons readily at the concentrations found in oil
spill situations, probably in chronically contaminated situations, and
marginally in uncontaminated situations. In being able to detect the
petroleum hydrocarbons at concentrations at and below those found in oil
spill situations, Dungeness crabs can achieve the first step to any sub-
sequent behavioral response to petroleum.
B. Avoidance of Oiled Sediment by the Dungeness Crab, Cancer magister.
Our next series of experiments examined whether the crab could change
its behavior to eliminate or reduce petroleum exposure, and this section
presents work in preparation for submission for publication by Pearson,
Sugarman, Woodruff, Blaylock and Olla.
Whether crabs could avoid contaminated sand was considered the eco-
logically relevant question because sandy bottoms are an important environ-
mental resource for the Dungeness crab. Their importance is indicated by
13
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the considerable extent to which crabs bury in the sand during their daily
activity cycle (Pearson et al. , 1979) and by the high degree to which the
crab's diet consists of animals, e.g., clams, that arc buried in sandy
substrates (Butler, 1954; Gotshall, 1977). In addition, some sandy bottoms
may be more suitable than others for various life habits. For example, our
field observations indicated that crabs were burying in distinct patches of
sand where the shear force of sediment took a very narrow value compared to
the range of penetrability found in the general area.
Because of the importance of the sandy bottoms to Dungeness crabs, the
long-term effects of an oil spill may derive not from oil-contaminated
water, but from contact with oiled sediment. Krebs and Burns (1977) presented
evidence that at least seven years after the West Falmouth Oil Spill contact
with oiled sediment was still producing behavioral abnormalities and reduced
population densities in the fiddler crab, Uca pugnax. Because the Dungeness
crab may be similarly vulnerable, the experiments reported here examined
the ability of the crab to avoid oiled sediment. Ecological consequences
from either avoidance or failure to avoid can be expected. Avoidance
probably denies the crab a patch of important resources whereas failure to
avoid could lead to adverse effects deriving from oil exposure.
Our field observations of the behavioral response of crabs to plumes
of freshwater during high runoff produced a preliminary hypothesis concerning
one possible behavioral response to oiled sediment that might occur. When
freshwater plumes were present Dungeness crabs left the eelgrass beds and
moved deeper with the downstream current to bury in the sand. Some preliminary
experiments suggested that a downstream movement, followed by burying, may
also occur with oiled sediment. A portion of the experiments reported here
addressed this question.
We performed a series of replicate experiments. The initial experi-
ments compared crab behavior in large partitioned and unpartitioned tanks
to see whether the tendency of crabs to bury downstream influenced the
avoidance of oiled sand. The next experiments examined how the oil-in-sand
concentration influenced avoidance. The last experiments concerned how
weathering of the oil might change avoidance behavior.
Materials and Methods —
The general approach in determining the occurrence and extent of oiled
sediment avoidance was to place Dungeness crabs in large tanks with oiled
and clean sediment and then observe the time spent in various clean and
oiled areas. Both partitioned and unpartitioned tanks were used to examine
whether avoidance behavior would differ if the crab's downstream movement
did not permit avoidance of oiled sediment. Table 2 gives some of the
parameters of the experiments reported here.
Animal Collection and Maintenance -- Dungeness crabs, Cancer magister,
were trapped in the Strait of Juan de Fuca, Washington, and held outdoors
in eight 1200-liter tanks. Molting and mating crabs were isolated from
those to be tested. Seawater drawn from the entrance of Sequim Bay entered
each holding tank through a manifold under a 15-cm layer of gravel and
sand. The crabs readily buried in the sand. Carapace size of crabs used
in the various experiments is given in Table 2. Individual crabs were
marked with variously shaped pieces of flat, white teflon secured with
stainless steel wire wrapped about the carapace.
14
-------
Experimental Tanks — The experimental tanks were 2.4 x 1.2 x 0.6 m
and constructed of 1.2-cm plywood coated with finishing type fiberglass
resin. A continuous flow of filtered sea water entered each tank at 5 L/min
at the midpoint behind a fiberglass baffle that extended across the
entire tank width. Holes were drilled on a 2.5 cm square grid across the
entire baffle. Hole sizes increased from the midpoint of baffle to each
side and were adjusted to promote an even cross-sectional flow from one end
of the tank to the other. An identical baffle was positioned at the down-
stream end of the tank 5 cm from a solid fiberglass end plate. After
passing through the downstream baffle, seawater spilled over this 47-cm
high end plate into a drain. Sediment depth was 7 cm and oiled and unoiled
sediment were separated by a barrier extending from bottom of the tank to
the top of the sediment.
Both unpartitioned and partitioned tanks had oiled sediment covering
the same surface area but in different positions relative to water flow.
Unpartitioned tanks had the downstream one-half covered with either clean
or oiled sand. The upstream end always had clean sand. A crab could walk
in a straight line from any position in the tank to any other. Partitioned
tanks had a solid fiberglass sheet running down the middle for three quarters
the length of the tank. The whole length of the tank was covered with
clean or oiled sediment on one side and with clean sand on the other. A
crab could move into or out of the oiled sediment only on the downstream
one quarter.
Preparation of Oiled Sediment — Sand was mixed with Prudhoe Bay crude
oil for addition to the experimental tanks. The sand was washed over a
0.32 mm mesh Nitex screen to remove the fine silts. The resulting geometric
mean size was 1.2 mm.
To produce oiled sand with nominal concentrations of 10000 ppm,
1000 ppm, and 100 ppm, 135 L of coarse washed sand was mixed to 1350 ml,
135 ml, and 13.5 ml of Prudhoe Bay crude oil , respectively, in a cement mixer
for 30 min. Equal amounts of oil and seawater were stirred in a high speed
blender for 30 sec before being mixed into the sand. The minimum of addi-
tional seawater necessary to yield a smoothly flowing mixture was added to
the cement mixer. During the addition of the oiled sand to the tanks and
introduction of seawater, the oil concentrations were expected to fall to
between 10 and 20% of the nominal value. Such reduction would have produced
oil concentrations typical of polluted sediment (Clark & MacLeod, 1977).
The actual oil concentrations in the sediment were determined from
composite core samples taken at the beginning of the first and end of each
subsequent experiment. Each composite consisted of 5 sediment cores (7 cm
x 2.5 cm diameter). Three composite samples were taken from each control
tank and three were drawn from each half of the oiled tanks. Total oil
levels were measured by infrared spectrophotometry (Simard et al., 1951;
Anderson et al., 1979).
Oil concentrations in the water column were determined from 50 ml
water samples drawn from the midpoint of the downstream end of the parti-
tioned oiled tanks in about 15 cm above the oiled sand. Samples were
drawn twice daily 12 hours apart. To test the oil distribution within a
tank, samples were drawn from the ends and center of each side. The hydro-
carbon concentration and components were analyzed by helium equilibration
gas chromatography (Bean & Blaylock, 1977).
15
-------
Procedure -- Forty-eight hours after the oiled sand had stood in
running seawater, 6 crabs were added to each tank in the middle of the
upstream end. Observations began one hour later and were taken hourly for
the duration of the experiments. The activity and position, by quadrant,
of the crabs were observed and recorded on a data sheet for each tank, the
activities assigned relative values and recorded by shorthand codes were:
ACTIVE - walking (2), climbing (2), standing (1); INACTIVE - resting (0),
and buried (-1). Behavior scores were tallied hourly for each tank and the
behaviors by quadrant were summed daily.
Ambient tidal levels were monitored hourly with a Metercraft tide
chart recorder (Style No. 80-H).
For the weathering experiment all crabs were removed from all tanks
after the first 5-d observation period. After the tanks had stood for
another 2 d, a new group of crabs were introduced and observed for 3 d
after they were in turn removed. After 8 weeks, another group of crabs
were introduced and observed for 3 d. Clean seawater entered the tanks
continuously whether crabs were present or not.
Results --
In the unpartitioned tanks Dungeness crabs generally avoided oiled
sand. In the control tanks crabs spent more time downstream than upstream,
but in the tanks with oiled sand downstream significantly decreased
time was spent downstream (Table 3).
The measured concentration of oil in the sand fell during the experi-
ments (Table 4). During the first five days that crabs were present, oil
concentrations fell about 20%. As the crabs buried themselves in the oiled
sand, we observed droplets of oil floating up through the water column.
After two months with a total of 11 days during which crabs were present,
oil concentrations were about half of the initial concentration. The loss
of oil then was probably heavily influenced by the activity level of the
crabs. Indeed the monoaromatic content of the water column often reached
its highest levels (up to 297 ppb) during periods of high activity. Great
amounts of monoaromatic hydrocarbons were found only over the highly oiled
sand (Table 5).
In the partitioned tanks crabs avoided oiled sand but not under all
circumstances. In control tanks crabs did not spend more time on one side
than the other, but in tanks with oiled sand on one side, crabs spent less
time on highly oiled sand and more time on oiled sand of moderate and low
concentration than on clean sand (Table 6). For only the time spent inactive,
i.e., buried in or resting on sand, the same pattern emerges (Table 7). In
this case the crabs spent 70% of the inactive time either buried in or
resting on the moderately oiled sand.
Analysis of variance indicated a significant variation with the oil
concentration but no significant weathering or interaction effects. None-
theless, after two months crabs on the highly oiled sand were apparently no
longer spending less time on oiled sand while crabs on the lowly and moderately
oiled sand continued to spend more time on oiled than clean sand.
16
-------
We could not detect any changes in the overall activity level of the
crabs due to oiled sand (Table 8). Activity levels did vary from experiment
to experiment, but this variation was apparently related to the tidal cycle
prevailing at the time of the experiment. The crabs were generally active
at night but inactive, i.e., buried and resting, during the day. During an
extreme low tide during the night or in extreme high tide during the day,
crabs tended to decrease activity while during a large change in tidal
height crabs increased activity. How the tidal cycle interacted with the
day-night cycle apparently determined the level and pattern of activity.
17
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Table 2. Experimental conditions in experiments concerning avoidance of oiled sand.
oo
TEST
I
II
III
IV
VA
VIA
VBC
VIBC
VICd
DATES
12/17 -
5/6 -
9/30 -
1/28 -
10/21 -
12/9 -
10/30 -
12/18 -
2/4 -
12/22/78
5/11/79
10/5/79
2/2/79
10/26/79
12/14/79
11/2/79
12/21/79
2/7/80
CRAB SIZE
X ± SD cm
(n = 48)a
N.D.
15.2 ±
15.1 ±
15.5 ±
15.9 ±
15.7 ±
16.1 ±
15.5 ±
16.4 ±
b
1.2
.7
.7
1.4
.7
.8
.6
1.1
TEMPERATURE
x ± SD OC
(n n 3)
7.0 ±
11.2 ±
10.1 ±
6.3 ±
10.1 ±
9.5 ±
10.1 ±
8.7 ±
6.9 ±
.2
.4
.2
.2
.2
.1
. 1
.1
.1
SALINITY _ D.O.
x ±°/00 x ± SO mg/L
(n=3) (n=3)
32.
32.
32.
32.
32.
32.
32.
32.
31.
0 8.6 ± .2
0 7.4 ± .4
0 7.8 ± .3
0 8.4 ± .2
0 6.9 ± .7
0 N.D.
0 7.9 ± .3
0 N.D.
0 8.1 ± .2
NO. RtPLICATES/TREATMENT
Control Oil
"V
.j * >*•
£• • C) C, C3 Cl (^
S. t'' "^ S; § § ^
1 1
2 2
2 2
1 1
2 2
2 2
2 2
2 2
2 2
1
2
2
2
2
2
1 ].
2 2
2 2
1
2
2
2
2
2
" Except for I and II where n = 24
N.D. = net determined
Oil weathered in experimental apparatus 1 week
Oil weathered in experimental apparatus 3 w
-------
Table 3. Time spent by Dungeness crabs in clean and oiled sand in
the unpartitioned tanks. Initially the oil-in-sand concen-
tration was 2882 (±854) ppm; at the end, 2315 (±496) ppm.
Results from 5 replicate tanks are pooled for each treatment.
NUMBER OF CRAB HOURS OBSERVED
UPSTREAM DOWNSTREAM
CONTROL
OILED
Buried & Resting 849
Total 1470
Buried & Resting 1108
Total 1935
1494 (63.8%)'
2130 (59.2%)*
948 (46.1%)'
1665 (46.2%)1
Oiled differs significantly from control, X2 = 138.2; p > 0.999
Oiled differs significantly from control, X2 = 120.5; p > 0.999
19
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Table 4. Oil-in-sand concentrations in partitioned tanks determined by
IR spectrophotometry.
OIL CONCENTRATION (ppm) IN SAND
TIME CONTROL LOW MODERATE HIGH
First Week
Beginning 5.1 ± 1.8 16.2 ± 4.4 192 ± 71 2508 ± 978
End 4.1 ± 0.4 17.8 ± 5.5 146 ± 39 2021 ± 686
Second Week
Beginning 4.4 ± 0.7 16.8 ± 5.7 160 ± 27 1788 ± 406
End 4.5 ± 1.0 10.0 ± 2.8 141 ± 65 1281 ± 223
At Two Months
Beginning 4.0 ± 0 8.0 ± 0 85 ± 10 1222 ± 4
End 6.0 ± 2.8 7.0 ± 4.2 72 ± 4 1381 ± 16
20
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Table 5. Total concentrations of monoaromatic hydrocarbons in
partitioned tanks determined by helium equilibration gas
chromatography. (The number of samples is enclosed in
parentheses.)
TOTAL MONOAROMATIC CONCENTRATION (ppb)
LOW MODERATE HIGH
FIRST WEEK (20) 0.2 ±0.9 (20) 0.4 ±1.3 (40) 45.8 ±62.3
SECOND WEEK ( 4) 2.1 ± 3.4 ( 4) 0.7 ± 0.5 ( 8) 9.2 + 8.7
THIRD WEEK N.D. N.D. ( 8) 4.3 ± 3.6
21
-------
Table 6. Average percentages of the total crab-hours observed in partitioned
tanks that Dungeness crab spent in oiled sand. The left half of the
tanks was used for control. (The number in parentheses is the number
of replicate tanks.)
PERCENTAGE OF TIME SPENT ON OILED SAND
Control Low Moderate High
FIRST WEEK
SECOND WEEK
AFTER TWO MONTHS
OVERALL
52
56
41
50
(10)
(4)
(2)
51
58
57
55
(5)
(4)
(2)
58
58
66
61
(5)
(4)
(2)
42
40
55
46
(10)
(4)
(2)
22
-------
Table 7. Average percentages of the crab-hours observed in partitioned tanks
buried and resting that Dungeness crabs spent buried and resting in
oiled sand. For controls the left side of the tank was used to
calculate the percentage. (The number of replicate tanks is enclosed
in parentheses.)
PERCENTAGE OF INACTIVE TIME SPENT INACTIVE IN OIL
Control Low Moderate High
FIRST WEEK
SECOND WEEK
AFTER TWO MONTHS
OVERALL
51
58
38
49
(10)
(4)
(2)
55
60
58
58
(5)
(4)
(2)
67
67
76
70
(5)
(4)
(2)
41
31
54
42
(10)
(4)
(2)
23
-------
Table 8. Average percentages of total crab-hours observed in partitioned tanks
that Dungeness crabs were active.
Control
x S.D.
PERCENTAGE OF TIME ACTIVE
_Low Moderate
x S.D. x S.D.
High
x S.D.
FIRST WEEK
SECOND WEEK
THIRD WEEK
46 ± 22
52 ± 4
46 ± 20
52 ± 16
59 ± 6
48 ± 1
57 ± 15
65 ± 10
43 ± 0
43 ± 9
56 ± 4
50 ± 3
24
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Discussion --
Before detailed discussion is possible, more analysis of the data only
briefly presented here is necessary. Nonetheless, several trends are
emerging.
Dungeness crabs avoid oiled sand, but not under all circumstances.
There was not a threshold oil concentration above which the crabs avoided
oiled sand and below which they were indifferent to it. Oiled sand with
1000 to 2000 ppm was avoided but sand with less than 200 ppm attracted
crabs.
Although no significant weathering effects were found, the number of
replicates at two months was low. Quite possibly a longer experiment with
more replication would detect a weathering effect.
While any effects of oiled sand on the level or pattern of activity
was not obvious, the variation in the activity levels and patterns indicates
a need to consider seasonal changes in behavior in designing and interpreting
experiments such as these. Seasonal variation in activity could well have
contributed to the observed high variability in the extent of avoidance.
Factors characteristic of the animal, the petroleum, and the environ-
ment all interact in determining the occurrence and extent of avoidance of
oiled sand. Taken as a whole, these experiments indicate that avoidance of
oiled sediment is more than a possibility but not a certainty, and, therefore.
the need exists to understand both the circumstances that favor or preclude
avoidance and the effects of exposure to oiled sand when avoidance is less
than complete.
25
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II. BEHAVIORAL EFFECTS OF PETROLEUM EXPOSURE
Because successful behavioral mitigation of exposure effects was not
assured under all circumstances, we examined the effects of petroleum
exposure on the chemosensory-directed feeding behavior of the Dungeness
crab. Chemoreception was chosen because chemoreception is the primary
sense in many life habits of marine organisms (Grant & Mackie, 1974) and
because ecologically important behavorial effects from petroleum would most
likely derive from disruption of chemoreception and the associated behaviors
(Blumer, 1969; Takahashi & Kittredge, 1973; Olla & Samet, 1974). We took
three steps on this pathway, establishing a baseline for sensitivity to
chemical food cues, examining whether such sensitivity changed under petroleum
exposure and studying how well crabs could find prey buried in oiled sand.
These steps are described in the following sections.
Section II.A. is work already published by Pearson, Sugarman, Woodruff,
and Oil a (1979) in the Journal of Experimental Marine Biology and Ecology
and describes the high sensitivity of Dungeness crabs to chemical food
cues. Section II.B., a manuscript submitted for publication, shows how a
24-h exposure to water contaminated with crude oil at concentrations typical
of an oil spill impaired the distance chemoreception seated in the crab's
antennules. Section II. C. , another manuscript ready for submission,
describes how we examined the ability of Dungeness crabs to find and take
clams buried in oiled sand. Due to a change in the clam's burrowing behavior
that increased its accessibility, crabs consumed more clams from oiled than
clean sand. Thus, a behavioral change tending to mitigate exposure led the
clam into vulnerability to ecological stressors. We indicated this pos-
sibility in our theoretical schematic (Figure 1).
A. Thresholds for Detection and Feeding Behavior in the Dungeness Crab,
Cancer magister.
Chemoreception is a sensory modality important to many life habits in
marine organisms (Grant & Mackie, 1974). A necessary step towards under-
standing the relation between chemoreception and ecology is to determine
the chemosensory acuity or threshold of an organism for particular chemical
signals (Wilson, 1970). Commonly, feeding responses have been used as
behavioral criteria for determining chemosensory abilities in crustaceans
(McLeese, 1970, 1974; Mackie & Shelton, 1972; Mackie, 1973; Fuzessery &
Childress, 1975), but Pearson and Oil a (1977) have suggested that overt
feeding responses may not be the most sensitive behavioral indicators of
chemical detection by crustaceans. By observing changes in antennular
flicking and gill bailing, they found that the threshold concentration at
which the blue crab, Callinectes sapidus, detected a food extract was
10 15 g/L which was many orders of magnitude below the concentration at which
chelae probing began. Here we report chemosensory thresholds for the
Dungeness crab, Cancer magister (Dana), also determined through measurement
of changes in antennular movement.
Despite the demonstrated importance of the antennules to chemoreception,
antennular movements have been sparsely studied and hardly ever used to
26
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measure chemosensory thresholds. The antennules of decapod crustaceans
have been shown to be the distance chemoreceptors involved in feeding
behavior (Hazlett, 1971a), orientation to odors (McLeese, 1973), sex recog-
nition (Ameyaw-Akumfi & Hazlett, 1975), host location (Ache, 1975) and
sex pheromone reception (Christofferson, 1970, 1972). Although crustacean
antennules have been the subject of considerable electrophysiological
investigation (Levandowsky & Hodgson, 1965; Van Weel & Christofferson,
1966; Ache & Case, 1969; Ache, 1972; Shepheard, 1974; Fuzessery & Childress,
1975; Ache et al., 1976; Price & Ache, 1977; Fuzessery, 1978; Fuzessery et
a!., 1978), antennular behavior in whole animals has been rarely studied.
Snow (1973) described antennular activities in the hermit crab, Pagurus
alaskensis, and discussed the factors influencing flicking rate. In an
attempt to develop artificial bait for the Dungeness crab, Allen et al.
(1975) have used changes in antennule orientation as one measure among
several to score the intensity of feeding responses to various natural and
artificial substances. Antennular flicking may facilitate chemoreception,
and increased flicking rates in the presence of sapid chemicals have been
reported for Homarus gammarus (Mackie & Shelton, 1972), Pleuroncodes planipes,
Cancer antennarius, Spirontocaris taylori, Pagurus hirsutiusculus (Fuzessery
& Childress, 1975) and Panulirus argus (Price & Ache, 1977). Only Pearson
and Olla (1977) have explicitly used antennular flicking rates as
the basis for measuring chemosensory acuity.
After observation of the behavior of Dungeness crabs in laboratory and
field, the threshold concentrations at which crabs detected a food extract
and exhibited feeding behavior were measured. To facilitate interspecific
comparison we used experimental apparatus and procedures similar to those
previously used for the blue crab (Pearson & Olla, 1977).
Materials and Methods --
Animal Collection and Maintenance -- Dungeness crabs, Cancer magi star
were trapped in Sequim Bay and the Strait of Juan de Fuca, Washington, and
held outdoors in eight 1200-L tanks. Molting and mating crabs were isolated
from those to be tested. Seawater, drawn from the entrance to Sequim Bay,
entered each holding tank through a manifold under a 15-cm layer of gravel
and sand. The crabs readily buried themselves in the sand. Large diameter
PVC pipe provided additional shelter. The temperature and salinity of the
seawater were 10.4 (± 0.8 S.D.)°C and 30.0 (± 0.1)°/005 respectively; the
dissolved oxygen, 6.5 (± 1.6) mg/L and pH, 8.04 (± 0.12). Clumps of blue
mussels, Mytilus edulis (L.), provided an ad libitum diet.
Observations of Feeding Behavior In the Field -- Underwater observations
using SCUBA diving were made in Possession Sound, Everett, Washington. The
site had a shallow, sandy shelf extending from shore and breaking at a
depth of 4 m into a steep slope that continued to depths of over 140 m.
Observations with SCUBA were confined to depths above 20 m and were con-
centrated on the shelf and upper portion of the slope. Here were eelgrass
beds up to 500 m2 in area and to the south the sand graded into pebbly
sand, pebbles, and then cobbles. Starting from a landmark we repeatedly
followed a transect west across the shelf, then at the upper portion of
the slope we went either north or south against the prevailing current.
A return transect was made over the shelf about 3 to 6 m from the beginning
of the slope and paralleling the previous transect. The number of crabs
27
-------
encountered, their activity, and other behavioral data were recorded in
situ on waterproof paper. A stopwatch in an underwater case was used~to
measure the time (sec) for 10 flicks of one antennule in buried crabs.
Twelve dives averaging 44 min each were made during daylight at different
points in the tidal and seasonal cycles.
Observations of Feeding Behavior In the Laboratory -- Feeding behavior
was observed in the outdoor holding tanks described above and in 265-L
fiberglass aquaria with glass fronts and sandy substrates. Reactions of
individual crabs to clam juice and chopped clam were observed from behind a
blind positioned before the glass front.
Determination of Chemosensory Thresholds -- In the apparatus (Fig. 3)
modified from Pearson and Olla (1977) crabs were presented with experimental
solutions and their subsequent behavior observed. From the main seawater
supply, seawater at 10°C passed through wound cellulose filters and a
polypropylene filter bag of 100 ym pore size into head tanks that in turn
delivered the seawater to four PVC manifolds (6.35 cm diam) each with 10
glass dripper arms. The dripper arms were adjusted to deliver 1.0 L/min into
each 5.68-L testing chamber. Seawater passed through a glass funnel into an
inlet manifold of plexiglass tubing (12 mm diam) that was connected to the -
white translucent plexiglass cover clamped to each chamber. Seawater entered
each chamber through three slits in the inlet manifold; the bottom one
(1 x 26 mm) was horizontal, and the upper two (1 x 30 mm) were at 30° to the
horizontal. The three slits were positioned 2, 3, and 4.5 cm from the
chamber bottom.
The glass testing chambers were arranged in a single line on four
trays; 10 chambers to a tray. Fluorescent lights with a daylight spectrum
provided 508 (± 80 S.D.) lux of illumination with a photoperiod synchronized
to civil sunrise and sunset. Illumination at night was 0.40 (± 0.18 S.D.)
lux. Blinds surrounded the trays so that the crabs could be observed and
the experimental solutions added from either side.
The experimental solutions were introduced into each testing chamber
via the glass funnel receiving the seawater from the dripper arms. A
teflon delivery tube (7 mm diam) carried the solution to the funnel from a
buret calibrated to deliver 20 ml in 15 sec. Initially, the delivery tubes
were inserted through PVC tubes fixed in position, but later this stationary
PVC tubing was removed.
To obtain a dilution factor for estimating the effective concentration
within a chamber, seawater solutions of azorubin red were introduced, and
water samples from six positions within the chamber compared with standard
dilutions of the dye solution in a spectrophotometer at 520 nm. During the
dye studies the chamber contained a crab model displacing 530 mL, a volume
typical of the crabs tested. The peak concentration was in the lower
midpoint of the chamber 10 sec after the dye solution was added and was
0.011 (± 0.003) times the concentration of the introduced solution.
Therefore, the effective concentration of experimental solutions reported
here were calculated by multiplying the concentrations of the introduced
solutions by 0.011.
28
-------
SEAWATER
FROM HEAD TANK
CALIBRATED
BURETTE
TEFLON DELIVERY TUBE
SEAWATER MANIFOLD
ffl\lllllllllllllll
1
1
c
VflM
^~
INLET
MANIFOLD
-»• _ •
llllllll\ s-
~^t
. JJ
Figure 3. A schematic diagram of the chemosensory testing apparatus.
From Pearson et al., 1979.
29
-------
Because clams have been found to be a major portion of the diet of
Dungeness crabs (Butler, 1954; Gotshall, 1977), seawater solutions of a
freeze-dried extract of the littleneck clam, Protothaca staminea (Conrad),
were presented to the crabs. Clams were held in the laboratory for several
days to purge themselves of sediment. The shucked meats and clam liquor
were then freeze-dried and stored at -60°C. The resultant freeze-dried
clam extract (FDCE) was first powdered in a blender with a stainless steel
cup chilled to -60°C, and then stored at -60°C. FDCE stock solutions were
prepared by mixing a weighed portion of the powdered FDCE with seawater
filtered through a 0.4 [jm membrane. After 2 h on a magnetic stirrer, stock
solutions were filtered through tared Whatman #4 and GF/C filter paper.
FDCE concentrations reported here have been corrected for the loss of the
FDCE retained on the filters, which was 48.6 (± 1.5)% of the initial FDCE
weight. The stock solutions, which averaged 2.05 (± 0.06)g FDCE/L, were
refrigerated and used for no more than five days. Serial dilutions of the
stock FDCE solution were made less than 1 h before testing with seawater
freshly filtered through a 0.4 pm membrane. An aliquot of the filtered sea-
water used for dilution constituted the control solution. The experimental
and control solutions were kept in a water bath at the ambient seawater
temperature and shaken immediately before use.
Approximately 24 h before testing, crabs were transferred to the
testing chambers from the holding tanks where they had an aid 1 ibitum diet.
In preliminary experiments tidal phase was found to influence chemosensory
responses. Therefore, while all testing was done during daylight hours, it
was synchronized to begin and end within either a rising or falling tide.
The seawater for the FDCE dilutions and control was drawn and filtered 1 h
after a tidal change. Testing then began as soon as possible and stopped
before the next tidal change.
Each day a maximum of 40 crabs were each presented with 20 ml of
either one of eight dilutions of FDCE stock solution or a control of filtered
seawater. Molting and mating crabs were not tested. The order in which
individual crabs were watched and the choice of experimental solution were
randomized except that active crabs and ones with retracted antennules were
passed over. The identity of the experimental solution was not known to
the observer. Individual crabs were observed for 1.0 min prior to intro-
duction of the experimental solution, and their antennular flicking rate
and other behavior recorded. The flicking rate of one antennule was measured
using a counter activated by a hand-held switch. The experimental solution
was then introduced and the observations continued for 1.0 min after the
beginning of solution addition. The behavior was scored with criteria
selected after observations of feeding in laboratory and field and numerous
trials in the apparatus.
A crab was considered to have detected an experimental solution when
there was an abrupt change in the orientation of the antennules within 30 sec
after solution introduction and if the ratio of the antennular flicking
rate for 1.0 min after solution introduction to that for 1.0 min before was
1.50 or higher. The antennular flicking rate for a resting Dungeness crab
in the apparatus ranged from 5 to 47 flicks/min with a median of 30 flicks/min
(n = 30). In contrast, the flicking rate observed in the field was higher.
At 9-ll°C observations with SCUBA showed the antennular flicking rate of
buried crabs to range from 36 to 103 flicks/min with a median of 70 flicks/min
30
-------
(n = 27). When crabs were observed for two consecutive minutes in the
chemosensory testing apparatus, without any addition of experimental or
control solution, the ratio of the antennular flicking rate of the second
minute to that of the first ranged from 0.49 to 2.00 and had a median of
0.94 (n = 30). The criterion value of 1.50 was determined a priori from
observations of crabs in the testing apparatus following the experimental
protocol except no solution was added. After ranking the resultant ratios
by magnitude, the ratio at the 95 percentile (n = 30) proved to be 1.50.
Thus, the a priori probability that an antennular flicking ratio greater
than 1.50 represents a spontaneous increase rather than a reaction to the
FDCE was less than 5%.
Feeding behavior was taken to begin when a Dungeness crab probed the
substrate with its chelae and/or exhibited a rapid and coordinated movement
in which the dactyls and chelae moved to bring an object, under the crab,
forward and up to its mouth. These coordinated movements were essentially
the same as those described as capture responses by Fuzessery and Chi 1 dress
(1975) for the crab Cancer antennarius. Because the sequence and character
of behavior of the Dungeness crab in the testing chambers correspond to
that in the laboratory aquaria, the behavioral criteria should be viewed as
indicating the onset of feeding behavior that would have led to ingestion,
given the presence of food.
At each FDCE concentration the percentage of crabs meeting the assay
criteria was recorded. Then the threshold concentration for detection was
calculated following the regression analysis of Pearson and Olla (1977).
The threshold concentration was that concentration at which 50% of the
crabs exhibited the behavior of interest.
Results --
Observations of Feeding Behavior In the Field — SCUBA observations of
Dungeness crabs showed that their activities differed with habitat and
tidal phase. During high tides crabs were walking and feeding on the sandy
shelf. At low tides crabs were buried on the upper portion of the slope
and the number on the shelf, even in the eelgrass, was low. In 6 out
of the 12 dives made, feeding behavior was observed. Three Dungeness crabs
were seen with clams. The first crab was ingesting the foot, gills, and
viscera of a clam. The second was carrying a littleneck clam, Protothaca
staminea, held tightly against its body by both chelae. The third crab was
carrying and feeding on a butter clam, Saxidomus giganteus (Deshayes).
Both valves of the butter clam had been broken away near the siphon, and
the crab was inserting one chela and tearing away tissue for ingestion.
Four Dungeness crabs were observed feeding on barnacles by using both
chelae to hold barnacle covered rocks (1 to 4 cm in diameter) up to the
maxillipeds and mandibles, which were crushing the barnacles for ingestion.
Red rock crabs, Cancer productus (Randall), were also observed carrying
Protothaca staminea and feeding on barnacles.
Observations of Feeding Behavior In the Laboratory -- In aquaria,
resting crabs typically had their ventrum touching the sand substrate and
chelae drawn up close to or touching the body. The eyestalks were extended
but occasionally bobbed into and out of their sockets for a few seconds.
The two antennules were often flicking in different directions and at
31
-------
different rates. When clam juice or a piece of clam was introduced into an
aquarium, crabs exhibited an abrupt change in the orientation of the antennules,
closely followed by a sharp increase in the antennular flicking rate. The
antennules became parallel as they oriented in the same direction, usually
upstream. The rate of antennular flicking rose from a base of 20-40 flicks/min
to a peak of as much as 120 flicks/min. Rhythmic beating of the maxillipedal
flagellae, which is presumptive of gill bailing (Burrows & Willows,
1969), began with, or immediately followed, the changes in antennular
behavior. Crabs then rose from the resting position and began walking with
the chelae extended forward and held above the substrate surface. The
ambulatory dactyls probed the sand with an inward motion. The chelae
probed the substrate with inwardly directed, arcing motions across or
slightly beneath the sand surface. When a dactyl contacted a clam piece,
the clam was quickly swept inward and then forward by the dactyls and
upward to the maxillipeds by the chelae, which were simultaneously brought
inward and upward. The chelae held the clam while the maxillipeds and
mandibles tore off pieces for ingestion. When feeding on live blue mussels,
crabs probed the mussel clump with the chelae and then either pulled one
mussel away or crushed it in place. Separated or attached mussels were
crushed between the fingers of the chelae, which then held pieces of broken
mussel shell while the maxillipeds and mandibles cleaned off the adhering
tissue. In contrast, the blue crab has been observed by Pearson and Olla
(1977) to separate one mussel completely from the clump by cutting the
byssum threads and then to pry the valves apart with both chelae as one
would open a book.
Determination of Chemosensory Thresholds -- In the chemosensory testing
apparatus, specific components of feeding behavior occurred more frequently
and intensely with increasing FDCE concentration (Table 9). Thus, when low
FDCE concentrations (10 16 to 10 8 g/L) were introduced, crabs typically
showed little or no activity beyond the changes in antennular movement
indicative of detection. At high levels (10 6 to 10 3 g FDCE/L) crabs made
probing motions with the dactyls and chelae, especially while walking. The
dactyl and chelae probing was sometimes accompanied by movement of the
chelae to the mouth. At the highest level (10 3 g FDCE/L) crabs rapidly
moved the dactyls inward and forward and the chelae inward and upward in
the same manner in which, in the laboratory aquaria, they had seized and
brought to the maxillipeds clam pieces encountered by the dactyls. Also
occurring at the highest extract level was the approach to and manipulation
of the inlet manifold, the source of the FDCE-laden seawater.
The detection of the clam extract was distinct and readily discernible.
When a crab changed antennular orientation and increased antennular flicking,
the flicking ratios were usually considerably above the criterion value of
1.50 (overall median ratio = 2.67, Table 10). Further, the magnitude of
the detection response itself did not increase with FDCE level. Although
the flicking ratios for all crabs observed differed with FDCE concentration
£P = 0.999, median test, Conover, 1971), the median ratios for responding
individuals did not _(P = 0.379, median test). Even though the overall
median flicking ratio for crabs showing feeding activity (4.61) was higher
than that for crabs detecting but not feeding (2.57), the difference was
not quite significant £P = 0.938, median test). The number of animals
detecting, not the magnitude of the detection response, increased with FDCE
level, and the percentage of crabs detecting the FDCE was used to determine
the threshold concentration.
32
-------
Crabs readily detected solutions of the clam extract. The percentage
of crabs detecting the FDCE was nearly 100% at the highest levels tested
and decreased with the FDCE concentration (Fig. 4). The threshold concen-
tration at which 50% of the crabs detected the FDCE was calculated to be
4.8 x 10~10 g/L (Fig. 4). Back calculation of the 95% confidence limits
about the Y estimate of 50% showed that the detection threshold could have
been as low as 10 14 g/L. The percentage of crabs (n = 30) that detected
the control seawater was 33.3%.
Because feeding responses only occurred above 10 6 g FDCE/L (Fig. 4),
there were not enough data for an adequate regression equation. Therefore,
a median line was drawn in Fig. 4, and the threshold concentration at which
50% of the crabs_began feeding behavior estimated graphically. The feeding
threshold was 10 2 g FDCE/L, some 8 orders of magnitude higher than the
detection threshold.
33
-------
CO
-pi
DETECTION
?= 82.407 + 3.478 X
LOG1Q OF
FDCE CONCENTRATION Ig/I I
Figure 4.
The percentage of Dungeness crabs detecting the freeze-dried clam extract
(FDCE) (circles) or beginning feeding behavior (triangles) as a function
of the logarithm of the FDCE concentration (g/L) within the testing
chamber. The number of crabs at each concentration is indicated. The
control seawater was detected by 33.3% of the crabs (n = 30).
From Pearson et al., 1979.
-------
CO
en
Table y. The occurrences of specific components of feeding behavior at various FDCE concentrations. The
tabulated values are percentages of the total number cf crabs observed at eai,h concentration.
The control values werj not included in the Cox and Stuart, tests for trend (Conover, 1971).
BEHAVIOR
Change in antennu'lar
orientation &
increased flicking
Beating of niaxil li-
pedal flagellae
Gaping and labiating
of maxi 1 1 ipeds
Antennal whipping
Grooming
Body raising
Walking
Dactyl searching
Chelae probing
Total No. of Crabs
-2.6
93.9
90.9
36.4
42.4
9.1
36.4
24.2
27.3
42.4
33
-4.
75.
69.
6.
12.
6.
12.
0
3.
6.
32
6
0
8
2
5
2
5
1
2
'-eg 10
-5.5
41.7
45.8
4.2
16.7
4.2
16.7
0
16.7
4.2
24
of FDCE Concentration (g/L
-7.t
45.8
33.3
0
4.2
8.4
4.2
0
0
0
24
-9.6
2S.O
40.0
4.0
24.0
0
8.0
0
4.0
0
25
-31 6
50. 0
57.7
4.0
7.7
7.7
11.5
0
0
0
25
N
-13.6
44.0
3b.O
4.0
12.0
12.0
11.5
0
4.0
0
25
-15.6
33.3
37.5
4.0
4.2
0
4.2
0
4.2
0
24
CONTROL
33.3
20.0
0
6.7
3.3
10.0
0
0
0
30
Test for
Trend
P
. 9375
.9375
.9375
.875
.6875
1.000
—
.9375
1.000
From Pearson et al., 1979.
-------
Table 10. The antennular flicking rate ratios at various FDCE concentrations. The ratios were calculated
by dividing the flicking rate for 1.0 min after FDCE introduction by that 1.0 min before.
All Crabs
Log10[FDCE] N
-2.6
-4.6
-5-6
en
-7.6
-9.6
-11.6
-]3.&
-15.6
Control
Total
33
32
24
24
25
26
25
24
30
243
0
0
0
0
0
0
0
0
0
0
Range
of Ratios
.88 -
.26 -
.50 -
.58 -
.50 -
.55 -
.33 -
.64 -
. 01 -
.01 -
30.7
13.8
13.0
38.0
7.60
5.40
18.0
12.5
9.60
38.0
Median
Ratio
3.50
2.43
1.74
1,75
1.44
2.35
1.70
1,43
1.39
1.78
Crabs fulfil
detection cri
N
31
24
10
11
7
13
11
8
10
125
1 i rKj
tsria
Median
Ratio
3.
2.
2.
2.
2.
3.
L. .
7
2.
2 .
82
68
32
40
85
17
20
19
60
67
Crabs detecting
but net feeding
N
17
22
9
11
7
13
11
8
10
108
Median
Ratio
2.
2.
0
c. •
2.
2.
3.
2.
3.
2.
2.
81
54
04
40
85
17
20
IS
60
57
Crabs feeding
N Median
Ratio
14 4.53
2 S.30
1 2.54
0
0
0
r>
•J
n
0
17 4.61
From Pearson et al., 1979.
-------
Discussion --
The antennules of many decapods have been shown to function as distance
chemoreceptors (Hazlett, 1971a), and now for at least three decapods changes
in antennular behavior have been found to indicate chemical detection. For
the blue crab, Callinectes sapidus, abrupt increases in the antennular
flicking rate accompanied by the onset of vigorous and continuous rhythmic
beating of the maxillipedal flagellae indicated detection not only of a
clam extract but also of the petroleum hydrocarbon naphthalene (Pearson &
01 la, 1977, 1979). The antennular flicking rate increased upon presentation
of glycine or glutamic acid to the spiny lobster, Panulirus argus, (Price &
Ache, 1977). With the Dungeness crab an increase in flicking rate along
with a change in orientation also indicated detection of a clam extract.
In addition, Mackie and Shelton (1972) and Fuzessery and Childress (1975)
reported increased antennular flicking in several crustaceans as the first
indication of "awareness" of sapid solutions but did not incorporate anten-
nular flicking into their measurements of chemosensory ability.
Antennular flicking is widespread among crustaceans, and several
explanations for the function of flicking in chemoreception have been
given. Snow (1973) suggested that flicking in a hermit crab facilitated
chemoreception by splaying out the aesthetasc hairs and thereby increasing
the passage of water over the chemoreceptors. The neurophysiological
observation that flicking produced renewed nervous response after fading of
the initial response led Price and Ache (1977) to propose that an increased
rate of flicking served to extend the time period during which the chemo-
receptors of the lobster were sensitive. After suggesting that antennular
flicking in the lobster was analogous to sniffing in vertebrates, Fuzessery
(1978) discussed how flicking may be involved in orientation to odor. Our
observations that antennular flicking increases in the presence of sapid
chemicals in Callinectes sapidus and Cancer magister are more presumptive
evidence that flicking enhances chemoreception in some way but do not allow
us to choose any one explanation from among those proposed.
Interspecific comparisons of chemosensory ability are difficult because
experimental techniques and, especially, criteria vary greatly, and of the
other crustaceans studied only three appear comparable on the basis of
close phylogeny or similar experimental treatment. The best comparison is
between blue and Dungeness crabs because the behavioral criteria and the
testing methods were essentially the same. The detection threshold for the
blue crab was 10 15 g/L (Pearson & Olla, 1977), which_is 5 orders of magnitude
lower than that found here for the Dungeness crab (10 10 g/L). The lobster,
Panulirus argus, presented with the amino acids glycine and glutamic acid,
showed increased antennular flicking at 10 10 M (about 10 8 g/L), the
lowest concentration tested (Price & Ache, 1977). Our regression of the
data of Fuzessery and Childress (1975) showed the threshold concentration
for the release of feeding motions by an equimolar mixture of three amino
acids to be 10 9 g/L for the crab Cancer antennarius. Amino acids alone
and in combination have been found to be less attractive than food extracts
(Shelton & Mackie, 1971; Mackie, 1973) so that we would predict that testing
with a food extract would yield lower thresholds in £. argus and C. antennarius.
The feeding threshold for the Dungeness crab was not distinct from
that found for the blue crab. The feeding thresholds for the blue crab
37
-------
varied from IC)"1 to 10 3 g FDCE/L (Pearson & 01 la, 1977). In comparison
the Dungeness crab had a feeding threshold of 10 2 g FDCE/L. For both
crabs the feeding thresholds were many orders of magnitude above those of
detection.
There are several explanations for the difference in the detection
thresholds seen for Callinectes sapidus and Cancer magister. Ultimately
the threshold differences derive from ecological and evolutionary differences.
Proximately the threshold differences may derive from differences in the
morphology and movement of the antennules. Although a larger crab than the
blue crab, the Dungeness crab has smaller antennules. When flicked through
equal arcs the smaller antennules of Cancer magister would sweep out a
smaller water volume than those of Callinectes sapidus. In addition, the
flicking rate of a resting Dungeness crab (30 flicks/min) is less than that
of a resting blue crab (85 flicks/min). Given equal numbers of chemo-
receptors per unit surface area, a resting Dungeness crab could be sampling
a smaller volume of water per unit of time than a similar blue crab and,
consequently, would exhibit a higher threshold. Perhaps detailed morpho-
metric analysis could relate increasingly chemoreceptive area to decreasing
thresholds in a series of crustaceans.
Blue and Dungeness crabs live under two different natural regimes of
temperature. The two species were tested at temperatures 10°C apart, and
this temperature difference may partially account for the difference in
detection thresholds. At 10°C Dungeness crabs feed, mate, and are generally
active while at the same temperature blue crabs feed little, if at all,
(Leffler, 1972) and are lethargic and intermittently dormant (Olla & Pearson,
unpubl.). It is, therefore, doubtful that testing both species at the same
temperature would produce a meaningful comparison.
Field observations reported here and elsewhere (Butler, 1954; Gotshall,
1977) have shown that Dungness crabs find, capture, and consume clams and
small crustaceans buried in the sand. In our laboratory and in the field
(Butler, 1954) crabs search for buried prey by probing the substrate with
their dactyls and chelae. During such searching the crab uses both chemical
and tactile cues, and Fuzessery and Childress (1975) suggest that because
the capture responses seen both here and in their study occur with chemical
cues alone, chemoreception predominantly controls the location and capture
of prey.
Before searching intensely with dactyls and chelae, however, the crab
has found, presumably through some chemical cue, a site potentially productive
of food, and it is in finding such a site that the efficiency with which
the antennules function in distance chemoreception influences foraging
success. Generally as the chemosensory thresholds lowers, the active space
widens (Wilson, 1970). With the two threshold values given here and field
measurements of current velocities one could construct two active spaces.
The first would predict the area within which a crab can detect the presence
of an aggregation of clams, and the second, the area within which the crab
begins active search for buried prey. This type of analysis has obvious
application to ecological questions, especially those concerning predator-prey
relationships.
38
-------
B. Impairment of Chemosensory Food Detection in the Dungeness Crab,
Cancer magister, by Petroleum Hydrocarbons
For marine organisms, disruption of chemoreception by oil is viewed as
both likely and of important ecological consequence (Blumer, 1969; Olla &
Samet, 1974; Olla et al., 1980a). Chemosensory disruption by various petroleum
hydrocarbons and oil fractions has been reported in snails (Jacobsen &
Boylan, 1973; Hyland & Miller, 1979), lobsters (Atema & Stein, 1974), and
in shore crabs (Takahashi & Kittredge, 1973). In some of these early
studies the exposure regime was not well defined and did not always compare
well with the length and level of exposure likely to be encountered in an
actual oil spill. Here we report on the ability of the Dungness crab,
Cancer magister, to detect and respond to a food extract after 24-h exposure
to seawater contaminated with Prudhoe Bay crude oil. To improve the
correspondence of our petroleum exposure to that of actual situations, we
exposed and tested crabs in a continuously-flowing seawater system at an
oil level reasonably expected to occur during an oil spill.
Behavioral criteria can be used to measure the acuity of chemoreception
in various organisms (Olla et al., 1980a). Changes in_antennular behavior
indicate that detection of a clam extract occurs at 10 15g/L for the blue
crab, Callinectus sapidus, (Pearson & Olla, 1977) and at 10~10g/L for the
Dungeness crab (Pearson et al. , 1979). Such high sensitivity implies high
dependence on chemoreception in finding food.
To determine whether exposure to petroleum hydrocarbons impaired this
acute detection ability, we exposed Dungeness crabs to oil-contaminated sea-
water for 24 h, presented them with a clam extract in the presence of the
oil-contaminated seawater, and recorded the percentages of crabs showing
the changes in antennular behavior indicative of detection and of those
showing the chelae probing indicative of food searching. At 24 h and 48 h
after stopping the flow of oil-contaminated seawater, we retested the
crabs to determine the time period necessary for recovery of detection
ability. Because this first experiment indicated rapid recovery, we per-
formed a similar second experiment in which we presented the clam extract
to crabs 1 h after stopping the flow of contaminated seawater.
Materials and Methods --
Animal Collection and Maintenance -- Dungeness crabs, Cancer magister
(Dana), trapped in the Strait of Juan de Fuca, Washington, were held under
the conditions described by Pearson et al. (1979). Seawater temperatures
during the two experiments were 8.9 (± 2.7 S.D.)°C (n = 16) and 9.2 (±0.5)°C
(n = 16); salinities, 31.8 (±0.4)%o (n = 5) and 32.0 (±0.0)°/00 (n = 4);
and dissolved oxygen 7.6 (±0.7) mg/1 (n = 16) and 8.0 (±0.3) mg/1 (n = 9).
Clumps of blue mussels, Mytilus edulis, and live clams, Protothaca staminea,
provided an ad 1ibitum diet.
Experimental Apparatus -- We coupled the oil delivery system developed
by Vanderhorst et al. (1977), and used extensively by Anderson et al.
(1979, 1980a & b), to the Chemosensory testing apparatus of Pearson et al.
(1979). Oil-contaminated seawater was delivered to 20 of the 40 Chemo-
sensory testing chambers from dripper arms situated along manifolds connected
to the oil delivery system. Contaminated water entered each exposure
39
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chamber at 0.1 L/min while clean water entered at 0.9 L/min. Control
chambers received clean water at 1.0 L/min. Seawater entered each chamber
through a glass funnel connected to a slotted inlet tube within the chamber.
Teflon tubes carried seawater solutions of the clam extract to the funnels
from burets calibrated to deliver 20 ml within 15 sec. Previous dye studies
of Pearson et al. (1979) showed that the maximum concentration of an introduced
solution within a chamber occurs 10 sec after introduction and is 0.011
(±0.003) times the concentration of the introduced solution.
The delivery system produced oil-contaminated seawater that was
largely a water-soluble fraction with some finely dispersed droplets. The
chemical composition of this oil-contaminated seawater has been well
characterized by Bean et al. (1978) and reported in detail by Anderson et
al. (1979; 1980a). Here we sampled seawater in the testing chambers by
the resin column absorption technique of Bean et al. (1978) and analyzed
the samples by infrared (IR) spectrophotometry. The data of Bean et al. (1978)
and Anderson et al. (1979, 1980a) show the correlations between the values
determined by IR and the concentration of specific hydrocarbons determined
by other methods for the same system. To determine how fast hydrocarbon
concentrations dropped after stopping the flow of oil-contaminated water in
the second experiment, we supplemented IR analyses with analyses for mono-
aromatic hydrocarbons by a helium gas partitioning technique modified from
McAuliffe (1971).
Experimental Solutions -- The experimental solutions were seawater
solutions of freeze-dried clam extract (FDCE) of littleneck clam, Protothaca
staminea. The solutions were prepared following Pearson et al. (1979).
Stock solutions averaging 1.89 (±0.12) g FDCE/L (n = 6) for the first
experiment and 2.06 (±0.22) g FDCE/L (n = 5) for the second were refrigerated
and used within five days. A 10 6 dilution of the stock FDCE solution was
made 1 h before testing with seawater freshly filtered through a 0.4 pm
membrane. An aliquot of the filtered seawater used for dilution was used
as the control solution. All solutions were held in a water bath at ambient
seawater temperature.
Procedures — After the oil delivery system had been operating for
several days and the hydrocarbon concentrations examined, a single Dungeness
crab was added to each of the 20 exposure and 20 control chambers. Chemo-
sensory testing was synchronized to begin and end within either a rising or
falling tide and after 24 h exposure to oil-contaminated seawater. In the
first experiment, the FDCE solutions were presented with oil-contaminated
seawater still flowing through the chambers. Each crab was presented with
either one of two dilutions of FDCE or a control of filtered seawater.
After correction_for dilution within a chamber, these FDCE concentrations
were 10~2 and 10"8 g/L. The choice of dilution and the order of presentation
were randomized except that active crabs and those with retracted antennules
were passed over. The observer did not know the identity of any solution.
An individual crab was observed for 60 sec prior to introduction of experi-
mental solution, and the antennular flicking rate and other behavior recorded.
The observer depressed a switch of an event counter for each flick of one
antennule. The solution (20 mL) was then introduced and observation continued
for another 60 sec from onset of introduction.
40
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The criteria of Pearson et al. (1979) were used to score the behavior.
Detection was indicated when a crab abruptly changed antennular orientation
and increased antennular flicking rate so that the ratio of the rate after
solution introduction to that before was 1.50 or higher. Previous observations
indicate that the a priori probability that such an increase in antennular
flicking is spontaneous, rather than in response to the solution, is less
than 5% (Pearson et al., 1979). The onset of food searching was indicated
when a crab probed the substrate with its chelae or exhibited the capture
response described by Pearson et al. (1979).
To examine recovery of detection ability, we stopped the flow of
oil-contaminated water after the first presentation of FDCE. Clean sea-
water then entered the chambers at 0.9 L/min. After 24 h and 48 h both
exposed and control crabs were again presented with experimental solutions
and their behavior observed and scored.
Because the first experiment indicated rapid recovery, we wished to
see if such recovery could occur within one hour and, therefore, repeated
the exposure phase of the first experiment. Instead of presenting FDCE
with oil-contaminated water still present, we turned off the contaminated
water and presented the FDCE 1 h later. The start and finish of exposure
for individual crabs was staggered to achieve this one-hour clearance of
oil from the chambers.
Statistical Analysis — The experiments were run until about 30 crabs
had been tested under each experimental condition. The numbers of crabs
detecting and not detecting the various experimental solutions were totaled
for exposed and control conditions. Although data is presented as the
percentage of crabs detecting the FDCE, chi-square analysis was done on 2 x
2 contingency tables of the number of crabs detecting or not detecting
under control or exposed conditions. Data for crabs showing chelae probing
was treated similarly.
Results --
Hydrocarbon Concentrations -- During the first experiment, where the
clam extract was presented in the presence of oil-contaminated seawater,
the total hydrocarbon concentrations by IR analyses were 0.27 (±0.04) ppm
(n = 22) during the 24-h exposure and 0.013 (±0.004) ppm (n = 6) 24 h after
the oil-contaminated water was stopped. During the second experiment,
where the clam extract was presented 1 h after stopping the oil-contaminated
seawater, the total hydrocarbon concentration by IR averaged 0.34 (±0.07)
ppm (n = 10). Also, in the second experiment after 1 h the concentration
of monoaromatic hydrocarbons (Table 11) fell to 0.008 times the exposure
level.
Impairment and Recovery of Chemosensory Detection -- After 24-h exposure
to and still in the presence of oil-contaminated seawater, the percentages
of exposed crabs detecting the clam extract was about half those of control
crabs (Table 12). In contrast, the percentage of crabs probing with chelae
did not differ significantly between control and_exposed conditions (Table
13). Some exposed individuals presented with 10 2 g FDCE/L probed the substrate
with their chelae without the normally proceeding changes in antennular
behavior.
41
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Recovery of detection ability occurred rapidly. In the first experiment
the percentage of crabs detecting FDCE at both levels did not differ between
control and exposed conditions for both 24 h and 48 h (Table 12). In the
second experiment, where the FDCE was presented 1 h after the flow of
oil-contaminated seawater was stopped, again the percentage of crabs
detecting did not differ significantly between control and exposed conditions
(Table 14).
42
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Table 11. The concentrations of monoaromatic hydrocarbons in the
chemosensory testing chambers. Determined by helium gas
partitioning, n = 4.
Hydrocrabon
Total Trimethyl
Benzenes
HYDROCARBON CONCENTRATIONS (PPB)
During 24 h of
Continuous Flow
x S.D.
40.6 ± 11.9
1 h after flow
Stopped
x S.D.
Benzene
Toluene
Ethyl Benzene
m + p Xylene
o Xylene
50.1 ±
85.0 ±
13.8 ±
38.0 ±
19.5 ±
10.8
12.7
2.8
5.2
3.4
0.13
0.16
0.74
0.94
0.90
± 0.24
± 0.31
± 0.95
± 1.36
± 1.22
<0.01
TOTAL
247.0 ± 34.7
1.98 ± 3.76
43
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Table 12. Percentage of Our.geness crabs detecting the clam extract [FDCE] after exposure to continuously-
flowing sea water contaminated with Prudnoe Bay crude oil.
[FDCE]
g/L
io~2
io~s
Control
Treatment
Control
Exposed
Control
Exposed
Control
Exposed
AFTER 24-H
No. of Crabs
Tested %
32
30
34
31
18
20
EXPOSURE
'• Detecting x2
97
16.0
53
32
3.46
13
17
0.40
25
AFTER 24-H
No. of Crabs
p Tested %
37
0.999
31
38
0.937
33
17
0.473
22
IN CLEAN WATER
Detecting x2
95
1.18
87
42
0.05
39
35
0.13
41
AFTER 48-H
No. of Crabs
p Tested %
31
0.723
31
31
0.177
31
16
0.282
17
IN CLEAN WATER
Detecting x2
97
1.96
87
36
1.06
48
40
1.97
18
P
0.838
0.697
0.840
-------
Table 13. Percentage of Oungeness crabs probing with the chelae upon presentation of a clam extract [f-DCE]
after exposure to continuously-flowing sea water contaminated with prudhce Bay crude oil.
[FDCE]
g/L
10"2
i(f *
Control
Treatment
Control
Exposed
Control
Exposed
Gnat re 1
Exposed
AFTER
24-H EXPOSURE
No. of crabs
Tested % Detecting x2
32
30
34
31
18
20
37
2.18
87
6
0.329
10
0
0.924
5
AFTER 24-
No. of CT
p Tested
37
0.860
31
38
0.434
33
17
C.S64
22
H I,N CLEAN WATER
abs
% Detecting x2
84
0.114
Si
5
0.3S5
9
0
0./93
4
AFTER
Nc, of cr?.
p Tested
31
0.265
?:
33
G.470
3i
16
0.627
17
48-H IN CLEAN WATER
•bs
% Detecting x2
84
2.20
68
0
1.02
3
6
1.10
0
P
0.862
0.687
0.705
-------
Table 14. Percentage of Dungeness crabs responding to a clam extract [FDCE] presented in clean water 1 h
after a 24-h exposure to oil-contaminated sea water.
[FDCE]
g/L Treatment
10~2
Control
Exposed
10~8
Control
Exposed
Control
Control
Exposed
No. of Crabs
Tested
28
30
33
30
19
13
CRABS DETECTING
% x2 p
96
0.002 0.040
97
42
0.551 0.542
33
37
0.681 0.591
23
CRABS CHELAE
% X2
89
0.952
80
0
1.118
3
0
0
PROBING
P
0.671
0.710
-------
Discussion —
The hydrocarbon concentration to which we exposed crabs were typical
of those found during actual oil spills. McAuliffe et al. (1975) estimated
that concentrations of hydrocarbons dissolved in the water column reached
0.2 ppm during a spill from an oil platform. About one alf of these
dissolved hydrocarbons were the monoaromatics present in our system at 0.2
ppm (Table 11). During another platform spill hydrocarbon concentrations
between 0.1 and 0.4 ppm were found in the water column by Grahl-Nielsen
(1978). Because alkanes were detected, these concentrations represent
emulsified oil as well as dissolved hydrocarbons. During the AMOCO CADIZ
spill initial measurements of the subsurface water of the L'Aber Wrach
estuary showed hydrocarbon concentrations between 0.026 to 0.330 ppm (Wolfe,
1978). Measurements with a towed fluorometer showed that oil-in-water
concentrations exceeded 0.5 ppm throughout the estuary and rose above 1.0
ppm in shoal areas (Calder & Boehm, in press). These high levels resulted
from turbulent mixing of surface mousse into the water column, and here,
too, the detection of alkanes indicated an oil-in-water emulsion. Because
the contaminated water produced by our oil delivery system contains pre-
dominantly dissolved monoaromatics and very little alkanes (Bean et al.,
1978; Anderson et al., 1980a), our exposure regime best represents the
platform spill characterized by McAuliffe et al. (1975) and our results
indicate more the effects of dissolved hydrocarbons than of emulsified oil.
Decapod crustaceans have two chemoreceptor systems, one for distance
chemoreception (seated in the antennules) and another for contact chemo-
reception (seated in the dactyls, chelae, and mouthparts) (Luther, 1930;
Case & Gwilliam, 1961; Levandowsky & Hodgson, 1965; Hazlett, 1968; 1971 a &
b). The observation that exposed crabs showed chelae probing but no
increases in antennular flicking when presented with FDCE suggests that
24-h exposure to oil-contaminated seawater depressed the functioning of
the distance chemoreceptor system in Dungeness crabs while, at least as far
as we can determine, not significantly affecting the contact chemoreceptor
system. Perhaps longer exposure would have affected the contact chemo-
receptor system.
The rapid recovery of detection ability suggests that the disruptive
effects of oil exposure may be due to masking of the extract odor or anesthesia
of the chemoreceptors, rather than direct cellular damage. Cellular damage
would have required a recovery period of days while masking or anesthesia
would have been readily reversible upon return to clean seawater (Johnson,
1977; Olla et al., 1980a). In addition to causing a readily reversible
effect, the oil-contaminated seawater contained several monoaromatic hydro-
carbons known to reduce anesthesia or reversible narcosis in barnacle
larvae (Crisp et al., 1967). Whereas direct cellular damage is easily
eliminated as the mechanism behind the chemosensory impairment, a recovery
time of one hour does not eliminate either masking or anesthesia as alter-
natives. Another mechanism, not so likely in our system but quite possible
in an actual oil spill, is disruption of antennular functioning through
physical effects on the sensory hairs, e.g. coating of surfaces or matting
hairs together.
Practically, the disruption of the distance chemoreceptor system
indicates that under oil spill situations the presence of dissolved hydro-
47
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carbons could cause the Dungeness crab some difficulty in finding food.
The apparent intactness of the contact chemoreceptor system implies that
the crab may have to touch food to detect its presence. Because chemical
stimulation of the antennules apparently is necessary to sustain food
searching when contact with food is not direct and immediate (Hazlett,
1971a & b), difficulty in food gathering even after contacting food is
quite possible under exposure to oil-contaminated seawater.
Because the radius of the active space within which an organism detects
a chemical cue is inversely proportional to the chemosensory detection
threshold (Bossert & Wilson, 1963), we can roughly calculate how oil exposure
decreases the active space for Dungeness crabs. By lowering the FDCE
detection threshold about 104 times oil exposure decreases the radius of
the active space by 104 times. To maintain the same rate of prey capture
as the presence of petroleum hydrocarbons shrinks the active space, the
Dungeness crab may be required to change its foraging behavior to rely on
other sensory modalities, perhaps touch. Fortunately, the recovery data
suggests that the necessity of such changes may be relatively transient if
the duration of exposure is transient.
C. Effects of Oiled Sediment on Predation of the Littleneck Clam,
Protothaca staminea, by the Dungeness crab, Cancer magister.
Oiling of the sediment is increasingly viewed as an important deter-
minant of long-term ecological effects of oil pollution (Wolfe et a!.,
1977; Cabioch et al., 1978). A good example is the 1969 West Falmouth Oil
Spill. After four years the benthic fauna was still undergoing rapid
changes in densities and species composition (Sanders, 1978). After seven
years high concentrations of oil still persisted, (Burns & Teal, 1979) and
exposure to oiled sediment was still producing lowered population densities
in the fiddler crab, Uca pugnax, through impaired escape behavior and
abnormal burrowing (Krebs & Burns, 1977). Because oil disrupts detection
of chemical food cues (Jacobsen & Boylan, 1973; Takahashi & Kittredge,
1973; Hyland & Miller, 1979; Pearson et al., in prep.), oiled sediment may
produce another more subtle behavioral effect, disruption of chemosensory-
directed predation on buried prey. Oiled sediment would, thereby, affect
not only the process of predation itself, but also ecosystem structure and
function. One predator-prey system perhaps vulnerable to the effect is
crab predation, which substantially influences species composition of
soft-bottom communities (Virnstein, 1977; Woodin, 1978; Reise, 1979) and
may affect bivalve distribution (Williams, 1978). Here we report field and
laboratory experiments that examined how oiled sediment influenced predation
on the littleneck clam, Protothaca staminea, by the Dungeness crab, Cancer
magister.
Clams are a major portion of the diet of Dungeness crabs (Butler,
1954; Gotshall, 1977) and Pearson et al. (1979) observed Dungeness crabs in
the field feeding on littleneck clams. Because the Dungeness crab readily
detects extracts of the littleneck clam (Pearson et al., 1979) and chemo-
sensory detection of this extract is reduced by exposure to Prudhoe Bay
crude oil in a continuous flow system (Pearson et al., in prep.), we hypo-
thesized that crabs would consume less clams buried in oiled sediment
because of impaired chemosensory ability. However, in two experiments with
field enclosures crabs consumed more clams on oiled than clean sediment. A
48
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subsequent laboratory experiment then indicated that the observed increase
in predation was in part due to increased prey accessibility from oil-induced
changes in clam burrowing behavior.
Materials and Methods --
Field Experiments — In two experiments lasting 13 and 29 d, we used
field enclosures to determine the rates of crab predation on clams buried
in oiled sediment. Cages similar to those of Virnstein (1977) were constructed
by bending and welding steel rod normally used for reinforcing concrete
(0.95 cm dia) into a frame 100 x 100 x 30 cm high. Polypropylene/polyethylene
screening of 3.2 x 5.5 cm mesh covered the cage sides and top. A square
frame of reinforcing rod formed a top that could be completely opened.
Rods extending from each corner 60 cm into the bottom anchored each cage.
Within each cage a wooden box (95 x 95 x 15 cm deep) with a screened bottom
was buried flush with the existing bottom and received the experimental
sediment and clams.
On the southern side of Travis Spit, Sequim Bay, Washington, eight
cages in two equal groups were set parallel to shore at a tidal height of
-0.76 m (-2.5 ft). Five meters separated each cage within a group, and
eighteen meters separated the two groups. In both experiments four cages
randomly received 135 L of clean sand and four cages, oiled sand. In the
29-d experiment an additional cage set between the two groups also received
oiled sand and was used to monitor oil concentration during the experiment.
During both experiments water depth varied tidally from 0.1 to 3.4 m.
Water temperatures were 14.5 (±1.4 S.D.)°C (n = 49) and 14.6 (±1.6)°C (n =
220), for the first and second experiments, respectively. Salinity was
32.0 (±0.1)°/00 (n = 11). Measurements of current velocity with parachute
drogues ranged from 2.0 to 16.1 cm/sec with a median of 5.0 cm/sec (n = 72)
and indicated current velocity to be a function of wind and tide. Current
velocity measured with a direct reading current meter surrounded by the
same mesh covering the cages averaged 2.2 (±0.6) cm/sec over four 6-h
periods during which the tide rose 3.2 (±0.1) m.
To produce oiled sand with a nominal concentration of 10000 ppm, 135 L
of coarse washed sand (Geometric mean size = 1.2 mm) was mixed with 1350 mL
of Prudhoe Bay crude oil in a cement mixer for 30 min. Equal amounts of
oil and seajwater were stirred in a high speed blender for 30 sec before i
being mixed into the sand. The minimum of additional seawater necessary
to yield a smoothly flowing mixture was added to the cement mixer. We
expected that during the addition of oiled sand to the cages the oil con-
centration would fall to between 10 and 20% of the nominal value. Such
reduction would have produced an oil concentration typical of polluted
sediment (Clark & MacLeod, 1977).
To find the actual oil concentration of sediment in the cages, two
types of composite core samples for hydrocarbon analysis were taken from
every cage at the beginning and end of both experiments. Because we expected
more rapid loss of oil from the surface, we took short (6 cm) and long (15
cm) sediment cores of 2.5 cm diameter. From control and oiled cages three
composite samples, each composed of five long cores, were taken. From
oiled cages six composite samples of eight short cores were taken. Total
oil levels were measured in three long core and three short core composite
49
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samples from each cage by infrared spectrophotometry (Simard et al., 1951;
Anderson et al. , 1979). Some of the remaining short-core composite samples
were analyzed for any changes in the relative proportion of hydrocarbons by
glass capillary gas chromatography (Anderson et al. , 1978). During the
29-d experiment one composite sample of 6 cm cores was taken every two to
three days from the additional cage and analyzed for total oil concen-
tration.
Littleneck clams, Protothaca staminea, collected from Sequim Bay were
divided by shell length into four size classes (26-35, 36-45, 46-55, and
56-65 mm). For each size class we used a differently colored permanent ink
to mark the umbo region. In the 13~d experiment each cage received 10
clams in each size class; in the 29-d experiment, 12 in each class. These
densities, 40 and 48 clams/m2, were typical for Sequim Bay (Vanderhorst &
Wilkinson, 1979). One day after oiled sand had been added to the cages,
clams were placed randomly on the sand surface of every cage and allowed to
bury. Clams remaining on the surface the following day were buried.
One male Dungeness crab (carapace length 14.1 (±0.3)cm and 15.2 (±0.7)
cm for 13~d and 29-d experiments) was placed into each cage two days after
addition of the clams. Observations were made by SCUBA or snorkel ing every
one to two days. Divers recorded the number of clams in each size class
that were unburied but intact as well as those eaten. The number of visible
siphons of buried clams, and the position and behavior of each crab within
a cage and of other animals around the cage were recorded. Every two to
four days divers carefully opened each cage and removed and saved all
visible shells and shell fragments. If needed, the mesh of the cages was
brushed clean. We counted the number of colored shells recovered to estimate
the number of clams in each size class consumed by the crabs.
To confirm the consumption of clams estimated by counting recovered
shells, we removed the uneaten clams remaining at the end of each experi-
ment. To see whether clams were buried shallower in the oiled sand, sand
and clams from all cages were carefully removed layer by layer and the
clams found at several depths were sorted by size and counted. In the 13-d
experiment, 2.2% (n = 320) of the clams added were not recovered either as
shells during the experiment or intact at the end. In the 29-d experiment
we could not account for 2.6% (n = 384) of the clams added. Data analysis
was based only on the clams recovered. In the 29-d experiment data from
one oiled cage where the crab died was not used.
Laboratory Experiment -- To test whether the increased predation rate
on clams in oiled sediment observed in both field experiments was due to
shallower burial, a 19-d laboratory experiment examined the predation rate
on clams buried in two different depths of sand. Eight 1100-liter tanks
(2.2 x 1.1 x 0.6 m) constructed of 1.2 cm plywood coated with finishing
type fiberglass resin were divided in half lengthwise by opaque fiberglass
dividers to yield chambers of the same surface area as the field enclosures.
Filtered seawater (13.1 (±1.0)°C, 32.0 (±0.1)°/0o, 6.9 (±0.2) mg DO/I)
entered each tank at 5 L/min through a 2.5 cm diameter PVC pipe positioned
in the middle of one end behind a fiberglass baffle that extended the width
of the whole tank. Holes were drilled on a 2.5 cm square grid across the
entire baffle and hole sizes adjusted to promote an even cross-sectional
flow through the tank. An identical baffle was positioned downstream 5 cm
before a solid fiberglass plate. After passing through the downstream
baffle, seawater spilled over the 47-cm high end plate into a drain.
50
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Daylight illumination (238 ± 115 lux) was provided by daylight spectrum
flourescent lighting on a photoperiod synchronized to civil sunrise and
sunset. Night illumination from incandescent bulbs was adjusted to less
than one lux to simulate moonlight.
The 16 chambers provided four replicates of the following treatments:
shallow-oiled (5 cm depth of oiled sand), shallow-clean (5 cm of clean
sand), deep-oiled (10 cm of oiled sand), and deep-clean (10 cm of clean
sand). Preparation of oiled sand was identical to that for the field
experiments except that all sand for the laboratory experiment was washed
over a 0.32 mm mesh screen to remove fine silts. Composite samples of five
cores of 1.9 cm diameter were taken from each tank at the end of the experi-
ment, and total oil concentration was determined by the IR analysis used in
the field experiments. In other experiments (Pearson et al., in prep.) the
total oil concentration after the sand had stood 24-h in the tanks with
flowing seawater was 2690 (±957) ppm (n = 42).
Each chamber received 12 marked littleneck clams in each of the four
size classes used in the field. One day after the oiled sand had stood in
running seawater, the clams were randomly placed on the sand and allowed
to bury. Those remaining on the surface the following day were partially
buried, siphon end uppermost, and were found completely buried the next
day. At this time one Dungeness crab (carapace width 15.5 ± 0.56 cm) was
placed into each chamber.
Initially crabs were observed once an hour to determine when feeding
activity occurred. Although most feeding occurred at night, some occurred
in late afternoon so that detailed observations on feeding behavior were
taken from 1600 to civil sunset. Shells from eaten clams were removed at
least once each day and the number in each size class recorded. Also, the
number of intact unburied clams was recorded at least once a day. At the
end of the experiment we measured the shear strength of the sand in the
four treatments with a hand-held shear vane and then recovered all remaining
uneaten clams. Of the clams added, 2.3% (n = 768) were not recovered as
shells or intact clams. Data from one oiled chamber in which the crab died
were not used.
To estimate the weight of clams consumed by the crabs, we used length-
weight data on littleneck clams collected from Sequim Bay. The shell
lengths of 30 clams in each of the four size classes were measured individually,
the clams shucked, and all tissue within the shell blotted dry and weighed.
From the data the regression equation, W = 2.089 x 10 5 L3'32, related the
shucked wet weight (W) in g to the shell length (L) in mm. The weight of
clam consumed by a crab during an experiment was estimated by summing over
all size classes the total number of clams in a size class times the shucked
wet weight for the median shell length in that size class determined from
the regression equation.
Results --
Oil Concentration in Sand -- The concentration of oil in the surface
sand dropped slightly but not significantly in the 13-d field experiment
(Table 15). In contrast, the oil concentration dropped to 12% of its
initial value in the 29-d field experiment. The difference in initial oil
51
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levels between the two field experiments was derived from a greater water
depth and slight current occurring while oiled sand was being added to the
cages in the 13-d experiment. Along with the overall decline in total oil
concentration in the 29-d experiment, the aromatic hydrocarbons showed a
slightly higher decline than the saturates (Table 16). Within these two
classes, relative proportions of individual hydrocarbons changed little or
not at all. Oil droplets were observed rising from the sand when crabs
burrowed or otherwise disturbed the sand so that the rate at which oil was
released from the sand was undoubtedly related to the activity of the
crabs. Oil concentration within the cage, monitored every two or three
days, did not show a steady decline probably because patches of undisturbed
sand of high oil concentration persisted amidst disturbed sand of much
reduced oil concentration.
In the 19-d laboratory experiment the oil concentration dropped to
1044 (±329) ppm (n = 16) and 1185 (±409) ppm (n = 16) in the shallow-oiled
and deep-oiled sand. These concentrations were slightly less than half the
initial oil level.
Crab Predation On Clams -- We observed Dungeness crabs unearthing
littleneck clams with their chelae and dactyls. Crabs unearthed more than
they consumed and opened large (>45 mm) clams differently than small (<45
mm) ones. Small clams were usually crushed, and we recovered not whole
valves but small fragments. Large clams were pried open after being held
for several minutes. After picking up a large clam, a crab tumbled it
until the clam was held against the body with the inside of one chela. The
umbo faced the elbow of the cheliped and the commissure between the valves
was horizontal. The crab spread the fingers of its other chela and placed
their tips into the commissure between the valves. After holding this
position for several minutes, many crabs dropped the clam and moved away.
Where a crab continued holding a clam, we could not discern whether the
crab then forced the valves apart by wedging in its fingertips or simply
waited until the clam relaxed slightly and inserted the fingertips in the
gap. Once the fingertips were inside, the crab would quickly pry the
valves apart. Shells from large clams were usually recovered intact or
with chipping or partial breakage along the valve edge.
Effects of Oiled Sediment — In both field and laboratory experiments
crabs ate significantly greater numbers and weights of clams from oiled
than clean sand (Table 17). During the field experiments divers observed
more intact unburied clams, especially large ones, on the surface of the
oiled sand (Table 18). At the end of the field experiments the clams were
significantly shallower in the oiled sand (Table 19). The shallow burial
suggested that the higher consumption of clams on oiled sediment was due to
their higher accessibility.
The laboratory experiment indicated that shallow burial could have
accounted for much but not all of the higher consumption of clams buried in
oiled sand. If shallow burial allowed increased consumption of clams by
increasing their accessibility, then we would expect and did indeed find
greater consumption in shallow-clean than deep-clean sand. Crabs on shallow-
clean sand ate about twice the number of clams as those on deep-clean sand
52
-------
(Table 17). If shallow burial were the only mechanism producing an increased
consumption rate, we would expect but did not find equal consumption rates
for shallow-oiled and shallow-clean sand. Crabs on shallow-oiled sand ate
clams at about 1.5 times the rate of those on shallow-clean sand. If
oiling the sand increased the ease with which sand could be moved and
thereby the ability of crabs to unearth clams, shear strength among the
four treatments would have but did not parallel consumption rates. Listed
in order of increased consumption of clams shear strengths were as follows:
Deep-clean, 1.98 (±0.67) KPa (n = 6); Shallow-clean, 0.97 (±0.45) KPa;
Deep-oiled, 2.83 (±0.72) KPa; Shallow-oiled, 0.23 (±0.08) KPa.
Supplemental observations suggest that other aspects in burrowing
behavior may have changed and, thereby, increased clam accessibility. We
observed that clams unearthed by crabs reburrowed with individual clams
remaining on the surface from several hours to several days. We could not
discern whether individual clams on oiled sand reburrowed more slowly. If
reburrowing was slower in oiled sand, we would expect and did find more
clams on the surface of oiled sand (Tables 18 and 19). Over all the experi-
ments the number of intact unburied clams was positively correlated with
the consumption rate (Spearman's Rho - 0.83; compare Table 17 and the last
column in Table 18).
From the oiled sand of all experiments and the clean-shallow sand of
the laboratory experiment, crabs ate significantly greater proportions of
the available clams in the two smaller size classes than in the larger ones
(Table 20). In addition, few small clams (<45 mm) were observed intact at
the sand surface (Table 18). When comparing only the clams actually eaten,
the size class distribution of consumption did not differ between oiled and
clean sand (Table 21).
53
-------
Table 15. The total concentration of oil in sand inside the field
enclosures. From oiled cages three composite samples of eight
6~cm cores or five 15-cm cores were analyzed by infrared
spectrophotometry. From control cages two composite samples
of five 15-cm cores were analyzed.
TOTAL OIL CONCENTRATION
x ± S.D. (n) ppm
Oiled Oiled Control
6-cm Core 15-cm Core 15-cm Core
13-Day Field Experiment
Beginning 811 ± 511 (12) 709 ± 350 (12) 4.4 ± 3.4 (8)
End 657 ± 368 (11) 711 ± 291 (12) 6.8 ± 5.0 (8)
29-Day Field Experiment
Beginning 1345 ± 297 (15) 1500 ± 465 (15) 7.8 ±4.9 (8)
End 151 ± 128 (15) 154 ± 106 (14) 6.6 ± 1.1 (8)
54
-------
Table 16. Hydrocarbon concentrations in sand during 29-day field experiment.
Means are based on three composite samples of eight 6-cm cores
analyzed Ly glass-capillary gas chromatography.
CONCENTRATION
HYDROCARBON
C12
13
14
IS
16
17
Pristane
18
Phytane
19
20
21
22
23
24
25
26
27
28
TOTAL
SATURATES
Naphthalene
2 Methyl Naph.
1 Methy! Naph.
1& 2 Ethyl Nanh
Dimethyl Naph.
2,6 & 2,7
1,3 & 1,6
1,7
1,4 & 2,3
and 1,5
1,2
Pnenanthrene
Methyl Phen.
Dimethyl Phen.
TOTAL
AROMATICS
BEGINNING
x z S
4.579
5.735 1.
5.775
6.936 1.
5.437
5.875
3.803
4.C77
2.272
5.064
4.: J86
3.648
3.622
3.111
3.096
2.602
2.264
1.638
1.215
75.332 8.
1.214
2.904 1.
2.539 . 3.
. .843
1.624
1.792
2. 136
1.708
1.082
.352
.109
.144
16.451 6.
. D. ppm
681
572
868
872
332
315
296
S42
299
554
375
285
159
237
466
330
125
095
053
195
b!6
101
092
319
689
7.15
844
681
535
146
026
040
301
X
.410
.565
.638
.786
707
.718
.472
.570
.237
.580
.502
.465
.457
.422
.408
.356
.342
.216
.108
9.010
.023
.115
,098
.036
.146
.115
.140
.086
.063
.021
.003
.011
.856
END
± S. 0. ppm
.279
.336
.£02
i ~"~
.4/u
.386
.387
.233
.299
.147
. 353
.292
.263
.254
.232
.226
.206
.176
.182
.159
5.2-46
.C31
.096
082
027
.083
.060
.1.16
.072
.053
.013
. 001
.010
.614
% REMAINING
9.0
9.8
11.0
Jj.3
13.0
12.2
12.2
12.2
12.6
11.5
12.0
12.7
1? 6
13.6
13.2
1"J 7
i5.1
13.2
8.9
12.1 -h 1,5
1.9
4.0
3.9
'4.3
9.0
6.4
6.6
5.0
5.8
6.0
2.8
7.6
b.O 1 2.5
55
-------
Table 17. Percentage of total number of clams and estimated shucked wet weight that v,ere consumed by crabs.
Data from 4 field enclosures or 4 laboratory chambers were summed to provide 2x2 contingency tables
for X2 analysis of the numbers of clams consumed during an experiment or intact at, the end versus
control or oiled sand. Footnotes indicate other comparisons. See text for procedures for
estimating weight consumed.
en
01
PERCENTAGE OF TOTAL NUMBER OF CLAMS CONSUMED BY CRABS
CONTROL OILED
EXPERIMENT %(n) %(n) X2 p
13-d field
29-d field
Laboratory
Shal low sand
Deep sand
17 (156)
14 (183)
32 (186)a
17 (190)a
36 (157) 13.5 0.999
46 (192) 46.1 >0.999
48 (145)b 9.4 0.998
41 (185)b 26.9 >0.999
MEDIAN WEIGHT OF TOTAL CLAMS CONSUMED
CONTROL OILED
g g
48 80C
50 157C
121 125
44 101C
Clean-shallow versus clean-deep sand; X2 - 11.3; p = 0.999
Oiled-s-hallow versus oiled-deep sand; X2 = 1.7; p = 0.802
Oiled treatments greater than control, one-tailed Mann Whitney test; p > 0.05
-------
Table 18. Average number of intact unburied clams observed in the field and laboratory experiments. One
observation per day was made. Maximum numbers of clams in a size class were 40 for the 13-d
field experiment and 48 for all other experiments.
n
13-d Experiment
Control 11
Oiled 11
29-d Experiment
Control 29
Oiled 29
Laboratory
Shal low
Control 11
Oiled 11
Deep
Control 11
Oiled 11
NUMBER OF INTACT
26-35 mm 36-45 mm
0 0.09 (±0.30)
0 O.C9 (±0.30)
0.03 (±0.18) 0
0.03 (±0.18) 0.4 (±0.6)
0.4 (±0.7) 4.4 (±2.2)
0.7 (10.8) 8.4 (±3.2)a
C 0.4 (±0.5)
0 4.4 (±2.0)b
UNBURIED CLAMS
46-55 mm
0.7 (±0.5)
2.3 (±l.I)a
0.4 (±0.6)
3.1 (±1.4)b
11.4 (±3.4)
17.0 (±6.7)
1.5(±0.8)
4.8 (±1.8)b
IN SIZE CLASS
56-65 mm
1.3 (±0.7)
2.8 (±1.2)a
0.2 (±0.4)
5.4 (±4.5)b
13.9 (±5.8)
22.1 (±7.0)
2.3 (±2.0)
8.3 (±3.5)b
All sizes
2.1 (±1.0)
5.2 (±1.9)a
0.6 (±1.0)
8.1 (±2.6)b
30.1 (±10.9)
48.2 (±15.8)a
4.2 (±2.b)
17.2 (±4.0)b
f. Differs from control p < 0.05
Differs from control p < 0.01
-------
Table 19. Depth distribution of clams recovered at the end of the field and laboratory experiments.
Footnotes indicate comparisons of Distributions.
on
00
13-d experiment
Control
Oiled
29-d experiment
Control
Oiled
Laboratory
Shal low
Control
Oiled
Deep
Control
Oiled
, Control vs. oiled;
Control vs. oiled;
PERCENTAGE OF TOTAL CLAMS RECOVERED
Unburied 0-4 cm 4-8 cm 8-15 cm
2 2 32 64a
3 17 15 65a
Unburied 0-8 cm 8-15 cm
1 14 85b
6 49 45b
Unburied 0-5 cm
17 83 c e
69 31 C f
Unburied 0-10 cm
2 98 d e
15 85 d f
X2 = 21.0; p > 0.999
X2 = AS. 4; p > 0.999
Total Number
Recovered
129
101
158
102
127
75
158
109
^ Control-shallow vs. oiled shallow; X2 -• 55.0; p > 0.999
Control-deep vs oiled-deep; X2 - 15,9; p > 0.999
&f Control-shallow vs. control-deep; X2 = 20.9; p > 0.999
Oiled-shallow vs. Oiled-deep; X2 -- 57.0; p > 0.99'j
-------
Table 20. Clams consumed by crabs as percentage of tot.i1 number initially added in each size class.
X2 analyses were performed on contingency tables of the number of clams consumed in each
class vs number of clams initially in that size class.
Initial
Number CLAMS CONSUMED AS PERCENTAGE OF
In Each
EXPERIMENT Size Class 26-35 mm 36-45 mm 46-55 mm
13-d Field
Control 40 22 18 10
Oiled 40 7S 30 12
29-d Field
Control 48 21 12 8
Oiled 48 62 62 27
Laboratory
Shal low
Control 48 56 27 17
Oiled 36 97 47 22
Deep
Control 48 ?j 19 14
Oiled 48 56 54 33
TOTAL NUMBER IN EACH SIZE CLASS
56-65 mm X2 p
20 1.8 0.385
28 17.0 0.939
10 2.8 0.576
31 8.2 0.957
23 10.1 0.982"
28 15.6 0.999
10 2.2 0.468
1-1 10. 8 0.987
-------
[able 21. Clams consumed in each size class as percentage of total number of clans consumed. X2 analyses
were performed c-> contingency tables of trio numbers of clans consumed in each size class in
clean &and vs those in oiled sand.
EXPERIMENT
13-d Field
Control
Oiled
2S-d Field
CT> Control
o
Oi led
Laboratory
Shallow
Control
Oiled
Deep
Control
Oiled
Total Number
Clams Consumed
27
56
25
88
59
70
32
76
CLAMS CO
26-35 mm
30
50
40
34
46
50
34
36
NSUMFO IN SIZE
36-45 mm
26
21
24
34
22
24
28
34
CLASS AS PERCENTAGE
46-55 .Tim
15
9
16
15
14
11
22
21
OF TOTAL NUMBER CONSUMED
55-65 mm X2
30
3.28 0.
20
20
0.93 0.
17
19
0.68 0.
14
16
1.11 0.
9
P
650
182
122
225
-------
Discussion —
Predator-prey relationships are a dynamic balance between ability of
the predator to detect, locate, pursue, capture, and consume prey and the
ability of the prey to escape such events. Because exposure to oil-contaminated
water impairs detection of the clam extract by Dungeness crabs (Pearson et
al., in prep.)s we expected oil to place the predator at a disadvantage by
making chemosensory detection of prey more difficult. Our results indicate
that this potential loss was offset by a behavioral change in the prey that
increased its accessibility. Despite this, our original hypothesis may
still be true for prey unaffected by oil. Macoma balthica behaved differently
depending on how the oil is applied to the sand (Taylor & Karinen, 1977).
We would suggest that a layer of oil or oiled sediment laid over clean
sediment would act as a chemosensory barrier to detection and decrease
predation on clams by the Dungeness crab. Other experimental designs are
needed to understand not only how crabs act on a bed of clams but how they
find the bed. Crabs may use distance chemoreception more to find a bed of
clams than to find an individual clam on a bed. Carriker's (1959) observation
that blue crabs readily find and devour clam spat before they bury encourages
our suspicion that distance chemoreception is important in crab predation
on clams. The implication here is that whereas oil might impair the location
of clam beds, upon entering a bed with oiled sand crabs could make sizable
inroads on the clam population.
The results of our field and laboratory experiments show that the
Dungeness crab is an active predator quite capable of finding, unearthing,
and opening buried clams. Like other crabs (Walne & Dean, 1972), the
Dungeness crab consumes more small than large clams, and the differential
consumption appears related to the effectiveness of the methods that the
crab uses to open clams. The two approaches to opening bivalves, crushing
and prying, are used by a variety of crabs besides the Dungeness crab, and
appear important in determining the critical size above which shelled prey
is considerably less vulnerable to predation (Landers, 1954, Ebling et al.,
1964; Walne & Dean, 1972; MacKenzie, 1977; Williams, 1978; Vermeij, 1978;
Zipser & Vermeij, 1978). The crushing of small clams probably accounts for
our inability to account for all the clams added. Like Landers (1954), we
recovered only small fragments of the crushed clams from 26-45 mm and
determined the consumption by counting hinges. Of the unaccounted for
clams in the field and laboratory experiments 80% and 64% were 26-45 mm.
If these were, in fact, eaten but their small hinges overlooked, then we
underestimated consumption. Such underestimation would be 2-3% at most and
is too slight to influence our conclusions.
Here we had concluded that the increased predation of littleneck clams
in oiled sand by Dungeness crabs was derived mainly from shallow burial
that increased clam accessibility. Similarly, seasonal variation in burial
depth of the clam, Macoma balthica, changes its accessibility to predatory
wading birds (Reading & McGrorty, 1978) and the summertime upward movement
of the quahog, Mercenaria mercenaria, allows serious predation by the blue
crab, Callinectes sapidus. (Carriker, 1951). Increases in vulnerability to
predation have also been seen in fish and crustacean prey exposed to pesti-
cides and heavy metals and usually derive from impaired escape behavior
(Hatfield & Anderson, 1972; Kania & O'Hara, 1974; Tagatz, 1976; Ward &
Busch, 1976; Ward et al., 1976; Farr, 1977; Sullivan et al., 1978).
61
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Burial depth was only one aspect of clam burrowing behavior and accessi-
bility to crab predation, and shallow burial depth alone could not account
for all the greater consumption of clams seen on oiled sand. Distribution
of clams with depth, including resting on the surface, resulted from several
competing processes involving both clam and crab. Active emergence by
clams and their exhumation by crabs determined the rate at which clams
reached the surface and shallow depths. Reburrowing determined the rate at
which clams left the surface and moved down through the sand. Consumption
by crabs determined the rate at which clams were lost from the system.
Various combinations of these processes could have produced our observed
results. For example, high rates of exhumation or active emergence or a
low rate of reburrowing would have led to more intact clams on the surface
and at shallow depth in the oiled sand. Exposure to oiled sand could have
changed these processes in various ways to contribute to the increased
consumption of clams.
Many burrowing bivalves use active emergence from the sediment to move
to more favorable locations (Ansell & Trevallion, 1969; Ansell et al. ,
1972), and some clams show active emergence and slower reburrowing rates in
response to contaminated sediment. Macoma balthica comes to the sand
surface when stressed by starvation (de Wilde, 1975; and decreases burrowing
and increases emergence in proportion to the degree to which oil and heavy
metal contaminates the sediment (Taylor & Karinen, 1977; McGreer, 1979).
Emergence from contaminated sediment also occurs in Tellina tenuis (Stirling,
1975) and Macoma inquinata (Roesijadi & Anderson, 1979). These studies
indicate the possibility of active emergence, but because crabs were also
unearthing and eating clams, we could not discern the extent to which
active emergence and slower reburrowing actually occurred here.
If active emergence occurs as an avoidance reaction to oiled sand,
then slow reburrowing may be another type of avoidance reaction. Alter-
nately, however, slow reburrowing could result from debility. Exposure to
a water-soluble fraction of crude oil decreases burrowing rate into uncon-
taminated sediment in Macoma balthica (Linden, 1977) so that this decrease
presumably results from debility rather than avoidance. Besides slow
reburrowing, debility could also lead to poorer escape from digging crabs
and less resistance to opening. By decreasing the clam's ability to escape
predation, oil-induced debility then could have contributed to the observed
higher consumption rate. However, we could not directly discern whether
crabs unearthed or opened clams more readily on oiled sand.
Besides changes in the clam's behavior or ability to escape predation,
exposure to oiled sand may increase the attack rate on clams by increasing
the appetite of the crabs. Increasing appetite could derive from the
increased metabolic demands due to the stress of oil exposure. Enhanced
feeding rates under sublethal stress due to exposure to oil-in-water dis-
persions of crude oil have been seen in the shrimp, Pandalus danae, and the
English sole, Parophrys vetulus (Anderson et al., 1980b). Whereas increased
clam consumption on oiled sand through active emergence, slow reburrowing,
debility, or increased appetite all appear possible, without more specific
observations we cannot state whether any of these possibilities actually
contributed to the increased clam consumption observed here.
62
-------
We also observed a preponderance of large clams (>45 mm) among the
intact unburied clams on both clean and oiled sand. Variation among the
size classes in rates of exhumation, emergence, or reburrowing could have
led to this prevalence of large clams. Alternatively, if emergence, ex-
humation, and reburrowing rates were the same for all size classes over the
whole of the experiments, then simply a lower rate of attack or success in
opening large clams would lead to their prevalence. Walne and Dean (1972)
found that the shore crab, Carcinus maenas, selected the smallest
quahogs and mussels offered and that ease of attack rather than availability
seemed to influence consumption. Except for consumption we could not
determine the relative magnitudes of the other processes involved. Despite
this, we believe the decrease in burrowing rate seen in other clams exposed
to oil and the greater work required to open clams seen here suggest a
greater likelihood that slower reburrowing by the clam and less success in
opening large clams contribute to the observed results.
Predation is an important determinant of species composition and
diversity (Paine, 1966; Glasser, 1979) and recent work (Virnstein, 1977;
Woodin, 1978; Reise, 1979) has confirmed the importance of crab predation
in soft-bottom communities. An important long-term implication of our
experimental results is the ecological consequence of a shift in crab
predation under oiled conditions. To the extent that oiled sediment renders
prey species more vulnerable to crabs, and crabs switch to more vulnerable
prey, we would expect their harvesting by the crab to reduce the prey's
representation in the benthic fauna. Such an ecological effect might be
far different than that predicted from a series of standard toxicity tests.
63
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REFERENCES
Ache, B.W. 1972. Amino acid receptors in the antennules of Homarus
americanus. Comp. Biochem. Physiol., 42A:807-811.
Ache, B.W. 1975. Antennular mediated host location by symbiotic
crustaceans. Mar. Behav. Physiol. , 3:125-130.
Ache, B.W. and J. Case. 1969. An analysis of antennular chemoreception
in two commensal shrimps of the genus Betaeus. Physiol. Zool., 42:361-371.
Ache, B.W., A.M. Fuzessery, andW.E.S. Carr. 1976. Antennular
chemosensitivity in the spiny lobster, Panulirus argus: Comparative
tests of high and low molecular weight stimulants. Biol. Bull.,
151:273-282.
Allen, W.V., E.G. Frederick, and R. Wong. 1975. Experiments on the
development of an artificial bait for the Dungeness crab, Cancer
magister (Dana). Sea Grant Publ. No. H.S.U. - S.G.-7, Humboldt State
Univ., 25 pp.
Ameyaw-Akumfi, C. and B.A. Hazlett. 1975. Sex recognition in the
crayfish Procambarus clarkii. Science, 190:1225-1226.
Anderson, J.W., S.L. Kiesser, and J.W. Blaylock. 1979. Comparative
uptake of napthalenes from water and oiled sediment by benthic amphi-
pods. In: Proceedings of the 1979 Oil Spill Conference. American
Petroleum Institute, Los Angeles, CA-, P- 579-584.
Anderson, J.W., S.L. Kiesser, and J.W. Blaylock. 1980a. The cumulative
effect of petroleum hydrocarbons on marine crustaceans during constant
exposure. Rapp. P. -v. Reun. Cons, int. Explor. Mer., 179:62-70.
Anderson, J.W., J.M. Neff, B.A. Cox, H.E. Tatum, and G.M. Hightower.
1974. Characteristics of dispersions and water-soluble extracts of
crude and refined oils and their toxicity to estuarine crustaceans and
fish. Mar. Biol. , 27:75-88.
Anderson, J.W., R.G. Riley, and R.M. Bean. 1978. Recruitment
of benthic animals as a function of petroleum hydrocarbons concen-
trations in the sediment. J. Fish. Res. Bd. Canada, 35:776-790.
Anderson, J.W., R.G. Riley, and R.M. Bean. 1980b. Long-term effects
of petroleum hydrocarbons from water on marine organisms. In: Pacific
Northwest Annual Report for 1979, PNL-3300, UC-11, Part 2. Pacific
Northwest Laboratories, Richland, WA., p. 103-104.
Ansell, A.D., P. Sivadas, B. Narayanan, and A. Trevallion. 1972. The
ecology of two sandy beaches in southwest India. III. Observations on
the population of Donas incarnatus and D. spiculum. Mar. Biol. ,
17:318-332.
Ansel!, A.D. and A. Trevallion. 1969. Behavioral adaptations of intertidal
molluscs from a tropical sandy beach. J. Exp. Mar. Biol. Ecol. , 4:9-35.
64
-------
Atema, J. and L.S. Stein. 1974. Effects of crude oil on the feeding
behavior of the lobster Homarus americanus. Environ. Pollut., 6:77-86.
Bean, R.M. and J.W. Blaylock. 1977. Characterization of volatile
hydrocarbons in flowing seawater suspensions of Number 2 fuel oil.
In: Fate and Effects of Petroleum in Marine Ecosystems and Organisms.
D. Wolfe, (ed.), Proceedings of NOAA Symposium, Seattle, WA- Pergamon
Press, Oxford, p. 397-403.
Bean, R.M., J.W. Blaylock, and R.G. Riley. 1978. Application of trace
analytical techniques to the study of hydrocarbon composition upon
dispersion of petroleum in a flowing seawater system. In: Proceedings
of a Symposium on Analytic Chemistry of Petroleum Hydrocarbons in the
Marine/Aquatic Environment. Division of Petroleum Chemistry, American
Chemical Society, p. 902-908.
Blumer, M. 1969. Oil pollution of the ocean. In: Oil on the Sea,
D.P. Hoult (ed.), Plenum Press, New York, p. 10-11.
Bossert, W.H. and E.O. Wilson. 1963. The analysis of olfactory
communication among animals. J. Theoret. Biol., 5:443-469.
Burns, K.A., and J.M. Teal. 1979. The West Falmouth oil spill: Hydro-
carbons in the salt marsh ecosystem. Estuar. Coast. Mar. Sci.,
8:349-360.
Burrows, M.A. and O.D. Willows. 1969. Neuronal co-ordination of
rhythmic maxilliped beating in brachyuran and anomuran Crustacea.
Comp. Biochem. Physiol., 31:121-135.
Butler, T.H. 1954. Food of the commercial crab in the Queen Charlotte
Islands Region. Progr. Rept., Pacific Coast Stat., Fish. Res. Bd. Can..
99:3-5.
Cabioch, L., J. Dauvin, and F. Gentil. 1978. Preliminary observations
on pollution of the sea bed and disturbance of sub-littoral communities
in Northern Brittany by oil from the Amoco Cadiz. Mar. Pollut. Bui 1.,
9:303-307.
Calder, J.A. and P.O. Boehm. (In press;. The chemistry of AMOCO CADIZ
oil in the Aber Wrach. In: Publications du Centre National Pour
L1 Exploitation Des Oceans.
Carriker, M.R. 1951. Observations on the penetration of tightly
closing bivalves by Busycon and other predators. Ecology, 32:73-83.
Carriker, M.R. 1959. The role of physical and biological factors in
the culture of Crassostrea and Mercenaria in a saltwater pond.
Ecol. Mongr., 29:219-266.
Case, J. and G.F. Gwilliam. 1961. Amino acid sensitivity of the dactyl
chemoreceptors of Carcinides maenas. Biol. Bull. , 121:449-455.
65
-------
Christofferson, J.P. 1970. An electrophysiological and chemical
investigation of the female sex pheromone of the crab Portunus
sanguinolentus (Herbst). Ph.D. dissertation, University of Hawaii,
95 pp.
Christofferson, J.P. 1972. The site of chemoreceptors sensitive to
the sex pheromone of the female crab, Portunus sanguinolentus (Herbst).
Am. Zool. , 12:690.
Clark, R.C., Jr. andW.D. MacLeod, Jr. 1977. Inputs, transport
mechanisms, and observed concentrations of petroleum in the marine
environment. In: Effects of Petroleum on Arctic and Subarctic Marine
Environments and Organisms. Vol. I. Nature and Fate of Petroleum.
D.C. Malins (ed.), Academic Press, New York, p. 91-223.
Conover, W.J. 1971. Practical Nonparametic Statistics. J. Wiley
& Sons, New York, 462 pp.
Crisp, D.J., A.O. Christie, and A.F.A. Ghobashy. 1967. Narcotic and
toxic action of organic compounds on barnacle larvae. Comp. Biochem.
Physiol., 22:629-649.
Ebling, F.J., J.A. Itching, L. Muntz, and C.M. Taylor. 1964. The
ecology of Lough Ine XIII: Experimental observations of the destruc-
tion of Mytilus edulis and Nucella lapillus by crabs. J. Am'm. Ecol. ,
33:73-82.
Farr, J.A. 1977. Impairment of antipredator behavior in Palaemonetes
pugio by exposure to sublethal doses of parathion. Trans. Am. Fish.
Soc., 106:287-290.
Fuzessery, Z.M. 1978. Quantitative stimulation of antennular chemo-
receptors of the spiny lobster, Panulirus argus. Comp. Biochem.
Physiol., 60A:303-308.
Fuzessery, Z.M., W.E.S. Carr, and B.W. Ache. 1978. Antennular
chemosensitivity in the spiny lobster, Panulirus argus. Studies of
taurine sensitive receptors. Biol. Bull., 154:226-240.
Fuzessery, Z.M. and J.J. Childress. 1975. Comparative chemosenstivity
to ami no acids and their role in the feeding activity of bathypelagic
and littoral crustaceans. Biol. Bull. , 149:522-538.
Glasser, J.W. 1979. The role of predation in shaping and maintaining
the structure of communities. Am. Nat., 113:631-641.
Gordon, D.C., Jr., P.O. Keizer, and J. Dale. 1974. Estimates using
fluorescence spectroscopy of the present state of petroleum hydro-
carbon contamination in the water column of the northwest Atlantic
Ocean. Mar. Chem., 2:251-261.
Gotshall, D.W. 1977. Stomach contents of northern California Dungeness
crabs, Cancer magister. Calif. Fish Game, 63:43-51.
66
-------
Grahl-Nielsen, 0. 1978. The Ekofisk-Bravo blowout: petroleum hydrocarbons
in the sea. In: Proceedings of the Conference on Assessment of Eco-
logical Impacts of Oil Spills. American Institute of Biological
Sciences, Arlington, VA, p. 476-487.
Grahl-Nielsen, 0., K. Westrheim, and S. Wilhelmsen. 1979. Petroleum
hydrocarbons in the North Sea. In: Proceedings of the 1979 Oil Spill
Conference. American Petroleum Institute, Washington, D.C., p. 629-632.
Grant, P.T. and A.M. Mackie. 1974. Chemoreception in Marine Organisms.
Academic Press, London, 295 pp.
Hatfield, C.T., and J.M. Anderson. 1972. Effects of two insecticides
on the vulnerability of Atlantic salmon (Salmo salar) parr to brook
trout (Salvelinus fontinalis) predation. J. Fish. Res. Bd. Can.
29:27-29.
Hazlett, B.A. 1968. Stimuli involved in the feeding behavior of the
hermit crab, C1ibanarius vittatus. Crustaceana, 15:305-311.
Hazlett, B.A. 1971a. Antennule chemosensitivity in marine decapod
Crustacea. J. Am'm. Morph. Physio!., 18:1-10.
Hazlett, B.A. 1971b. Chemical and chemotactic stimulation of feeding
behavior in the hermit crab, Petrochirus diogenes. Comp. Biochem.
Physiol., 39A:665-670.
Hyland, J.L. and D.C. Miller. 1979. Effects of No. 2 fuel oil on
chemically evoked feeding behavior of the mud snail, Ilyanassa obsolete.
In: Proceedings of the 1979 Oil Spill Conference. American Petroleum
Institute, Washington, D.C., p. 603-607.
Jacobsen, S.M. and D.B. Boylan. 1973. Effect of seawater soluble fraction
of kerosene on chemotaxis in a marine snail, Nassarius obsoletus.
Nature, 241:213-215.
Johnson, f.G. 1977. Sublethal biological effects of petroleum hydro-
carbon exposures: bacteria, algae, and invertebrates. In: Effects
of Petroleum on Artie and Subartic Marine Environments and Organisms,
Vol II. D.C. Mai ins (ed.), Academic Press, New York, p. 271-335.
Kania, H.J. and J. O'Hara. 1974. Behavioral alterations in a simple
predator-prey system due to sublethal exposure to mercury. Trans.
Am. Fish. Soc. , 1:134-136.
Krebs, C.T. and K.A. Burns. 1977. Long-term effects of an oil spill
on populations of the salt marsh crab, Pea pugnax. Science, 197:484-487.
Landers. W.S. 1954. Notes en the predation of the hard clam, Venus
mercenaria, by the mud crab, Neopanope texana. Ecology, 35:422.
Leffler, C.W. 1972. Some effects of temperature on the growth
and metabolic rate of juvenile blue crabs, Callinectes sapidus, in the
laboratory. Mar. Biol. , 14:104-110.
67
-------
Levandowsky, M., and E.S. Hodgson. 1965. Amino acid and amine
receptors of lobsters. Comp. Biochem. Physiol., 16:159-161.
Linden, 0. 1977. Sublethal effect-s of oil on mollusc species from the
the Baltic Sea. Water, Air, Soil Pollut., 8:305-313.
Luther, W. 1930. Versuche uber die chemorezeption der brachyuran.
Z.f. vergl. Physiologic, 12:177-205.
MacKenzie, C.L., Jr. 1977. Predation on hard clam (Mercenaria
mercenaria) populations. Trans. Am. Fish. Soc. , 106:530-537.
Mackie, A.M. 1973. The chemical basis of food detection in the
lobster, Homarus gammarus. Mar. Biol. , 21:103-108.
Mackie, A.M. and R.G.J. Shelton. 1972. A whole-animal bioassay for
the determination of the food attractants of the lobster Homarus
gammarus. Mar. Biol., 14:217-221.
McAuliffe, C.D. 1971. GC determination of solvents by multiple
phase equilibration. Chem. Techno!., 1:46-51.
McAuliffe, C.D., A.E. Smalley, R.D. Groover, W.M. Welsh, W.S. Pickle,
and G.E. Jones. 1975. Chevron Main Pass Block 42 oil spill: chemical
and biological investigations, In: Proceedings of the 1975 Joint
Conference on Prevention and Control of Oil Spills, American Petroleum
Institute, Washington, D.C., p. 555-566.
McGreer, E.R. 1979. Sublethal effects of heavy metal contaminated
sediments on the bivalve Macoma balthica (L.). Mar. Pollut. Bull. ,
10:259-262.
McLeese, D.W. 1970. Detection of dissolved substances by the American
lobster (Homarus americanus) and olfactory attraction between lobsters.
J. Fish. Res. Bd. Can., 27:1371-1378.
McLeese, D.W. 1973. Orientation of lobsters (Homarus americanus)
to odor. J. Fish. Res. Bd. Can., 30:838-840.
McLeese, D.W. 1974. Olfactory responses of lobsters (Homarus
americanus) to solutions from prey species and to seawater extracts
and chemical fractions of fish muscle and effects of antennule ablation.
Mar. Behav. Physiol., 2:237-249.
Olla, B.L. (ed.). 1974. Behavioral measures of environmental stress.
In; Proceedings of a Workshop on Marine Bioassays, Chairman G. V.
Cox. Marine Technological Society, Washington, DC, p. 1-31.
Olla, B.L. and C. Samet. 1974. Behavior of marine organisms as a
measure of petroleum contamination. In: Proceedings of Estuarine
Research Federation Outer Continental Shelf Conference and Workshop
on Marine Environmental Implications of Offshore Oil and Gas Development
in the Baltimore Canyon Region of the Mid-Atlantic Coast. College
Park, MD, p. 437-450.
68
-------
Olla, B.L., W.H. Pearson, and A. L. Studholme. 1980a. Applicability
of behavioral measures in environmental stress assessment. In:
Biological effects of marine pollution and the problems of monitoring.
A.D. Mclntyre and J.B. Pearce (eds.), Rapp. P.-v. Reun. Cons.Int.
Explor. Her., 179 (In Press).
Olla, B.L., A.L. Studholme, and W.H. Pearson. 1980b. Behavior as a measure
of adaptation. In: Proceedings of USSR-USA Symposium on Physiology
and Biochemistry of Aquatic Animals. Belle W. Baruch Inst. Mar.
Biol. and Coastal Res., Columbia, SC (In Press).
Paine, R.T. 1966. Food web complexity and species diversity. Am. Nat.,
100:65-75.
Pearson, W.H., S.E. Miller, J.W. Blaylock, and B.L. Olla. 1980a. Detection
of the water soluable fraction of crude oil by the blue crab, Cal1inectes
sapidus. Mar. Environ. Res., (In Press)-
Pearson, W.H. and B.L. Olla. 1977. Chemoreception in the blue crab,
Callinectes sapidus. Biol. Bull. , 153:346-354.
Pearson, W.H. and B.L. Olla. 1979. Detection of naphthalene by
the blue crab, Call inectes sapidus. Estuaries, 2:64-65.
Pearson, W.H. and B.L. Olla. 1980. Threshold for detection of naphthalene
and other behavioral responses by the blue crab, Callinectes sapidus.
Estuaries, 3 (In Press).
Pearson, W.H., P.C., Sugarman, D.L. Woodruff, J.W. Blaylock, and B.L. Olla.
1980b. Detection of petroleum hydrocarbons by the Dungeness crab, Cancer
magister, Fish. Bull. , U.S. , (In Press).
Pearson, W.H., P.C. Sugarman, D.L. Woodruff, and B.L. Olla. 1979.
Thresholds for detection and feeding behavior in the Dungeness crab,
Cancer magister (Dana). J. Exper. Mar. Biol. Ecol., 39: 65-78.
Price, R.B. and B.W. Ache. 1977. Peripheral modification of chemo-
sensory information in the spiny lobster. Comp. Biochem. Physiol.,
57A:249-253.
Reading, C.J. and S. McGrorty. 1978. Seasonal variations in the
burying depth of Macoma balthica (L.) and its accessibility to wading
birds. Estuar.Coast. Mar. Sci. , 6:135-144.
Reise, K. 1979. Moderate predation on meiofauna by the macrobenthos
of the Wadden Sea. Helgolander wiss. Meeresunters, 32:453-465.
Roesijadi, G. and J.W. Anderson. 1979. Condition index and free amino
acid content of Macoma inquinata exposed to oil-contaminated marine
sediments. In: Marine Pollution: Functional Responses, W.B. Vernberg,
A. Calabrese, P.P. Thurberg and F.J. Vernberg, (eds.), Academic Press,
New York, p. 69-84.
69
-------
Sanders, H.L. 1978. Florida oil spill impact on the Buzzards Bay
benthic fauna: West Falmouth. J. Fish. Res. Bd. Can. , 35:717-730.
Shelton, R.G.J. and A.M. Mackie. 1971. Studies on the chemical
preferences of the shore crab, Carcinus maenas (L.). J. Exp. Mar. Biol.
Ecol., 7:41-49.
Shepheard, P. 1974. Chemoreception in the antennule of the lobster,
Homarus americanus. Mar. Behav. Physiol., 2:261-273.
Simard, A.G., I. Hasegawa, W. Bandaruk, and C. Headington. 1951. Infrared
spectrophotometric determination of oils and phenols in water. Anal.
Chem., 23:1384-1387.
Slobodkin, L.B. 1968. Toward a predictive theory of evolution. _In:
Population Biology and Evolution. R.C. Lewontin, (ed.), Syracuse Univ.
Press, Syracuse, N.Y., p. 187-205.
Slobodkin, L.B. and A. Rapoport. 1974. An optimal strategy of evolution.
Q. Rev. Biol. , 49:181-200.
Snow, P.J. 1973. The antennular activities of the hermit crab,
Pagurus alaskensis (Benedict). J. Exp. Biol. , 58:745-765.
Stirling, E.A. 1975. Some effects of pollutants on the behavior of the
bivalve Tellina tenuis. Mar. Pollut. Bull. 6:122-124.
Sullivan, J.F., G.J. Atchison, D.J. Kolar, and A.W. Mclntosh. 1978.
Changes in the predator-prey behavior of fathead minnows (Pimephales
piomelas) and largemouth bass (Micropterus salmoides) caused by cadmium.
J. Fish. Res. Bd. Can. , 35:446-451.
Tagatz, M.E. 1976. Effect of Mirex on predator-prey interaction in an
experimental estuarine ecosystem. Trans. Am. Fish. Soc. 105:546-549.
Takahashi, F.T. and J.S. Kittredge. 1973. Sublethal effects of the
water soluble component of oil: Chemical communication in the marine
environment. In: The microbial degradation of oil pollutants. D.G.
Ahearn and S.P. Meyers (eds.J, Louisana State Univ. Press Publ. No.
LSU-SG-73-01, p. 259-264.
Taylor, T.L. and J.F. Karinen. 1977. Response of the clam, Macoma
balthica (Linnaeus), exposed to Prudhoe Bay crude oil as unmixed oil,
water soluble fraction, and oil contaminated sediment in the laboratory,
In: Fate and Effects of Petroleum Hydrocarbons in Marine Organisms
and Ecosystems, Wolfe, D.A., (ed.), Pergamon Press, Oxford, p. 119-237.
Vanderhorst, J.R., C.I. Gibson, L.J. Moore, and P. Wilkinson. 1977.
Continuous flow apparatus for use in petroleum bioassay. Bull.Environ.
Cont. Toxic., 17:577-584.
Vanderhorst, J.R. and Wilkinson, P. 1979. The littleneck clam, Protothaca
staminea, as a tool for potential oil pollution assessment: Part 1 -
Density of stock. Mar. Envir. Res., 2:223-237.
70
-------
Van Weel, P.B. andJ.P. Christofferson. 1966. Electrophysiological
studies on perception in the antennulae of certain crabs. Physiol. Zool.,
39:317-325.
Vermeij, G.J. 1978. Biogeography and Adaptation: Patterns of Marine Life.
Harvard Univ. Press, Cambridge, MA, 332 pp.
Virnstein, R.W. 1977. The importance of predation by crabs and fishes
on benthic infauna in Chesapeake Bay. Ecology, 58:1199-1217.
Walne, P.R. and G.J. Dean. 1972. Experiments on predation by the
shore crab, Carcinus maenas L on Mytil us and Merceneria. J. Cons, int.
Explor. Her., 34:190-199.
Ward, D.V. and D.A. Busch. 1976. Effects of Temefos, an organophosphorous
insecticide, on survival and escape behavior of the marsh fiddler
crab, Uca pugnax. Oikos, 27:331-335.
Ward, D.V., G.L. Howes, and D.F. Ludwig. 1976. Interactive effects of
predation pressure and insecticide (Temefos) toxicity on populations
of the marsh fiddler crab Uca pugnax. Mar. Biol. , 35:119-126.
Wilde, P.A.W.J., de. 1975. Influence of temperature on behavior, energy,
metabolism, and growth of Macoma balthica (L.). In: Proceedings of the
9th European Mar. Biol. Symposium, p. 239-256.
Williams, M.J. 1978. Opening of bivalve shells by the mud crab
Scylla serrata Forskal. Aust. J. Mar. Freshwater Res., 29:699-702.
Wilson, E.G. 1970. Chemical communication within animal species,
_In: Chemical Ecology, E. Sondheimer and J.B. Simeone (eds.), Academic
Press, New York. p. 133-155.
Wolfe, D.A. 1978. The AMOCO CADIZ oil spill. A summary of observations
made by U.S. Scientists 23 March - 10 May 1978. Mar. Poll. Bull.,
9:288-292.
Wolfe, D.A., J. Anderson, O.K. Button, D.C. Malins, E.W. Mertens, and G.
O'Neil. 1977. Panel discussion on research needs. In: Fate and Effects
of Petroleum Hydrocarbons in Marine Organisms and Ecosystems. D.A.
Wolfe (ed.), Pergamon Press, Oxford, p. 454-464.
Woodin, S.A. 1978. Refuges, disturbance, and community structure:
A marine soft-bottom example. Ecology, 59:274-284.
Zipser, E. and G.J. Vermeij. 1978. Crushing behavior of tropical
and temperate crabs. J. Exp. Mar. Biol. Ecol. , 31:155-172.
71
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