The document summarizes a study on the effects of resmethrin and piperonyl butoxide (R-PBO), two common pesticides, on the foraging ability of juvenile blue crabs. Crabs were exposed to control water or 1:3 ppb or 10:30 ppb R-PBO for various time periods and then placed in tanks with shrimp to observe predation rates over time. Crabs exposed to 10:30 ppb R-PBO ate significantly fewer shrimp than other groups, likely due to impaired motor skills from pesticide exposure. However, crabs exposed to 1:3 ppb R-PBO ate more shrimp than controls, contradicting expectations. The results suggest pesticide exposure

1. Effects of Resmethrin and PBO Juvenile Blue Crab (Callinectes sapidus) Foraging
Erin Plachy
Texas A&M University Corpus Christi
6300 Ocean Dr. Corpus Christi, Texas 78412
erin.plachy@spartans.ut.edu
Keywords: Callinectes sapidus, pesticide, resmethrin, Piperonyl butoxide, foraging ability,
mesocosm, brown shrimp

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ABSTRACT
The effects of resmethrin and PBO (Piperonyl Butoxide), two common toxicants used to
control mosquito populations, on juvenile blue crab foraging behavior was studied. Resmethrin
and PBO are commonly sprayed together in a 1-to-3 ratio to control mosquito populations.
Juvenile male blue crabs (Callinectes sapidus) and brown shrimp were collected from Corpus
Christi, Texas and kept in tanks for a minimum of 48 hours in a salinity of 18-20 PSU. Before
being placed in mesocosms, crabs were exposed in to control water or 1:3 ppb R-PBO for 12
hours, or 10:30 ppb resmethrin-PBO (R-PBO) for 3 or 12 hours. Shrimp were similarly exposed
to control water. One crab and eight shrimp were placed in each mesocosm, and the number of
shrimp alive at 1, 2, 3, 6, 8, 12, and 24 hours was recorded. The 10:30 ppb R-PBO crabs ate less
shrimp than both the control (p=0.0796) and the 1:3 ppb R-PBO crabs (p=0.00035). The 1:3 ppb
R-PBO crabs ate significantly more than the control ethanol crabs (p=0.0079). Resmethrin
affects the nervous system which reduces a crab’s motor skills, making it difficult to catch prey.
A decrease in predation pressure by blue crabs on prey species could cause a shift in predator-
prey dynamics in estuarine ecosystems.
INTRODUCTION
The blue crab (Callinectes sapidus) is essential to many ecological communities and is a
key predator and prey species in areas along the eastern North America to South America
(Williams 1973). In addition to their ecological importance, blue crabs are also commercially
important; however, crab landings are at their lowest since the late 1960s, indicating a decline in
crab abundance (Sutton and Wagner 2007). The decline is primarily attributed to loss of habitat

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and overfishing (Guillory et al. 2001), but there is a growing concern for insecticide chemicals
entering blue crab habitats through runoff.
Pennington et al. (2001) found that especially after periods of significant rainfall, 93.4%
of all mid-Texas coast estuarine waters contained triazine herbicides from local spraying. Since
blue crabs spend much of their life in estuarine systems, they are likely exposed to runoff
pollution from surrounding human populations/cities. SCOURGE™ is an insecticide often used
for mosquito control and contains the chemicals resmethrin and piperonyl butoxide (Zulkosky et
al. 2005). Resmethrin interferes with the nervous system of insects (Zulkosky et al. 2005) and is
commonly used with Piperonyl butoxide (PBO), a synergist. PBO inhibits an organism from
metabolizing toxins, which amplifies the effect of the resmethrin (WHO 2001; Zulkosky et al.
2005). Several studies have been conducted on the effect of pesticides on blue crabs (Osterburg
et al. 2012; Horst and Walker 1999), but the sublethal effects of resmethrin with PBO (R-PBO)
on juvenile blue crabs are basically unknown.
The objective of this project was to study the sublethal effects of R-PBO as it pertains to
foraging ability of juvenile blue crabs. Since the pesticide’s target, insects, are part of the same
phylum as crabs, there are likely similar side effects from exposure. Because blue crabs are so
important in the estuarine ecosystems and to commercial fisheries in the U.S., it is necessary to
mitigate any additional pressures on the existing population.
MATERIALS AND METHODS
Juvenile male blue crabs and brown shrimp were caught in Corpus Christi Bay and the
Upper Laguna Madre (TX). Animals acclimated to lab conditions for a minimum of 48 hours and
were kept in tanks with untreated salt water (salinity 18-20). Crabs were caught via crabbing

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with raw chicken, and shrimp were caught using a push net or a seine net. Crab size ranged from
35-55 mm, and shrimp size ranged from 35-50 mm. Crabs were required to have both chelipeds
and swimming legs as well as most of their walking legs.
Fourteen-liter plastic tubs equipped with filters, oyster shells, and untreated saltwater
were set up. Twelve hours before the experiment, shrimp and crabs were measured and assigned
to individual bowls or jars with either R-PBO water or control water. Control water included
ethanol because ethanol was present in the pesticide stock solution. Crabs and shrimp were
exposed to 1:3 ppb R-PBO water, 10:30 ppb R-PBO water, or water with ethanol. Since more
than half of the 10:30 ppb R-PBO crabs died overnight, so the effects after 3 hours of exposure
to 10:30 were also studied (Table 1). Shrimp were placed into mesocosm tubs first, and allowed
to hide for 5-10 minutes before one crab was added. The number of shrimp alive in each
mesocosm was recorded at hour 1, 2, 3, 6, 8, 12, 24, 36, 48, 60, and 72 hours, or until the crab
had eaten all the shrimp. Other endpoints recorded included the number of shrimp that were not
hiding, if/where the crab was hiding, and whether or not the crab had clumped the oyster shells.
Table 1. Mesocosm crab and shrimp treatment with corresponding Resmethrin-PBO
concentrations and exposure time
Mesocosm Crab Exposure Shrimp Exposure Time Exposed
Control Ethanol 0 0 12 hours
Crab Low Exposure 1:3 ppb R-PBO* 0 12 hours
Shrimp Low Exposure 0 1:3 ppb R-PBO 12 hours
Both Low Exposure 1:3 ppb R-PBO 1:3 ppb R-PBO 12 hours
Crab High Exposure 3-hr 10:30 ppb R-PBO 0 3 hours
Crab High Exposure 12-hr 10:30 ppb R-PBO 0 12 hours
*Resmethrin to PBO ratio is labeled as R-PBO. “0” exposure indicates control ethanol water.

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RESULTS
Statistical tests were complete in JMP and Microsoft Excel (2010). The 10:30 ppb
exposure 12-hour treatment was not used due to lack of replicates. Data was analyzed using a
repeated measures ANOVA (JMP), with pesticide treatment as a fixed factor. Treatment was
nearly significant (p=0.0630), indicating that treatment had an effect on the number of shrimp
alive. Because of the small number of replicates (i.e., n<6 in some cases) in the treatments, this
result should likely be considered significant in order to avoid making a Type II error. Time was
a significant factor (p< 0.0001), and the time-treatment interaction was not significant (p=0.280).
In Excel, a students t-test (assuming equal variances) was completed to compare
treatment levels to each other and to the controls, with a Bonferroni correction. The number that
survived at each time point differed between treatment groups (Figure 1). Crabs exposed to
10:30 ppb for 3 hours tended to eat significantly less shrimp than the control group (p=0.0796)
whereas crabs exposed to 1:3 ppb for 12 hours ate significantly more than the control group
(p=0.0079). In comparison to each other, crabs exposed to 10:30 ppb (3-hour) ate significantly
less than the crabs exposed to 1:3 ppb (12-hours).

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Fig 1. Average Shrimp Alive Over Time. Lines show average number of shrimp alive for crabs
exposed to control (green, dashed line), 10:30 ppb resmethrin:PBO (red), or 1:3 ppm
resmethrin:PBO. Error bars are ±SE.
DISCUSSION
Results indicate that that crabs exposed to higher concentrations (10:30 ppb R-PBO)
have suppressed foraging abilities whereas crabs exposed to lower concentrations (1:3 ppb R-
PBO) seem to have slightly enhanced foraging abilities in comparison to the control group.
Though there are virtually no published studies done on crabs with R-PBO, some studies have
investigated the effects of mercury and other heavy metals on other aquatic organisms that
produce similar effects. Heavy metal contamination can reduce feeding and foraging ability in
aquatic organisms, including species such as killifish (Reichmuth et al. 2009; Weis et al. 1999,
2000, 2001, 2011). Weis et al. (2001) concluded that contaminants (e.g., heavy metals) can alter
an organism’s motivation to feed and reduce foraging search effectiveness, and their ability to
0
1
2
3
4
5
6
7
8
9
0 1 2 3 6 8 12 24
NumberofShrimpAlive
Hour
Average Shrimp Alive: Res. + PBO
Control Ethanol 10:30 ppb 3-hr 1:3 ppb 12-hr

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capture prey. These findings are consistent with the 10:30 ppb 3-hr exposure treatment, as crabs
ate fewer shrimp at a slower pace compared to control crabs. Several crabs exposed to R-PBO
were observed with a shrimp in their claw for extended periods of time, indicating that they
might be spending a longer amount of time consuming a single shrimp. Additionally, treatment
crabs were often uncoordinated and where observed swiping/swinging their claws in the
direction of shrimp but were unsuccessful in actually catching shrimp.
For blue crabs specifically, Reichmuth et al. (2009) found that crabs exposed to mercury
were less able to capture active prey such as shrimp, which caused crabs to consume less active
prey like mussels. This indicates that blue crabs, both in this work and ours, may be more
affected by reduced coordination rather than lack of motivation or hunger. Since resmethrin
affects the nervous system, motor skills are greatly reduced, making it more difficult for the
crabs to catch active prey. In the 10:30 ppb R-PBO 12-hr treatment, the crabs that did survive
were observed waving their chelipeds erratically and flipping upside-down as if they had lost all
control of their muscular system; the dead crabs were found the next morning upside-down, with
several autotomized limbs. Though the 10:30 ppb R-PBO 3-hr treatment crabs did not react as
severely as the 10:30 ppb 12-hr treatment crabs, it is likely that they endured similar, but
reduced, side effects.
The implications of pesticide runoff entering estuaries could potentially change the
community structure. Weis et al. (2011) studied several estuarine organisms in a contaminated
estuary and their foraging behaviors. All the species, including blue crabs and killifish, were
negatively affected from exposure, in varying degrees, except for grass shrimp (Palaemonetes
pugio) which had unchanged predator avoidance and was found to have partitioned its energy,
favoring growth and reproduction. The grass shrimp were able to increase their reproduction,

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grow larger, and live longer because of the combination of decreased predation pressure and the
shrimps’ ability to partition resources (Weis et al. 1999, 2011). Exposed to contaminants like
heavy metals and pesticides, blue crabs would not be able to forage as well, having to select less
optimal prey. Their very wide diet variety would be reduced, affecting their life span and
numbers similar to the findings in a study by Weis et al. (1999) on mummichongs exposed to
contaminants. In addition, because blue crabs are such an important predator in estuaries,
predation pressure would be relieved for many prey species who would then be allowed to
increase in numbers. Community structure based on predator-prey interactions would shift if
prey species became more abundant.
The 1:3 ppb R-PBO 12-hr unexpectantly ate significantly more than the control
(p=0.0079), as we expected 1:3 ppb crabs to eat less than the control ethanol group, but more
than the 10:30 ppb 3-hr crabs. This result contradicts those found by the previously discussed
studies with contaminants, though work with this specific toxicant has not been studied. Further
experimentation is needed to better understand why and how exposure might cause in increase in
foraging success and needs. In the future I would repeat the experiment with incremental
increases in concentrations of R-PBO in order to investigate the trend in the number of shrimp
alive over time for each concentration. Also, I would add 10:30 ppb 6-hrs and 8-hrs to see if
there are any changes in results between the 3-hr and 12-hr exposure. If lower exposures of R-
PBO occur in estuaries, the effects in natural systems would be dramatically different than those
predicted in laboratory experiments at higher concentrations. These results indicated that
exposure may cause crabs to increase foraging, which inturn may increase predation pressure on
prey species if these results are indeed consistent.With the increased use of pesticides used to
control mosquitoes in the past few decades, aquatic organisms such as blue crabs will

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increasingly encounter these harmful chemicals; further research is necessary to understand how
pesticides affects these ecologically and commercially important species.
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