Aquatic Toxicology 124125 (2012) 209216

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Aquatic Toxicology
journal homepage: www.elsevier.com/locate/aquatox

Acute effects of deltamethrin on swimming velocity and biomarkers of the

common prawn Palaemon serratus
Cristiana Oliveira a,b, , Joana Almeida a,c , Lcia Guilhermino a,c , Amadeu M.V.M. Soares b , Carlos Gravato a
a
b
c

CIIMAR Centro Interdisciplinar de Investigaco Marinha e Ambiental, Universidade do Porto, Porto, Portugal
CESAM Centro de Estudos do Ambiente e do Mar, Departamento de Biologia, Universidade de Aveiro, Aveiro, Portugal
ICBAS Instituto de Cincias Biomdicas Abel Salazar, Departamento de Estudos de Populaces, Universidade do Porto, Porto, Portugal

a r t i c l e

i n f o

Article history:
Received 19 February 2012
Received in revised form 9 August 2012
Accepted 14 August 2012
Keywords:
Swimming velocity
Cholinesterase
Palaemon serratus
Deltamethrin
Ecological relevant endpoint

a b s t r a c t
The main purpose of the present study was to investigate the effects of deltamethrin on biomarkers and
behavior of Palaemon serratus (common prawn), since this attempt to link different levels of biological
organization will allow determining which biomarkers might be ecologically relevant and will be useful
to complement the information about the effects of pesticides by using behavioral parameters. Therefore,
parameters of liver antioxidant status, energy metabolism and neurotransmission were determined in
different tissues of the common prawn and used to assess the effects at sub-individual level, whereas
swimming velocity was used to assess the effects at the individual level. It was also investigated if the
swimming velocity can be used as an endpoint in ecotoxicology bioassays and if it can be as sensitive as
biomarker endpoints.
Swimming velocity was signicantly reduced in prawns exposed to deltamethrin, showing a lowest
observed effect (LOEC) of 0.6 ng L1 . Eye acetylcholinesterase (AChE) activity was signicantly increased
in prawns exposed to 0.6, 1.2 and 2.4 ng L1 deltamethrin, whereas muscle cholinesterase (ChE) activity
was signicantly increased in prawns exposed to 19 and 39 ng L1 . On the other hand, lactate dehydrogenase (LDH) activity was signicantly increased in muscle of prawns exposed to 0.6, 1.2, 2.4,
4.9 ng L1 deltamethrin, showing that organisms were requiring additional energy, but probably using it
for detoxication processes rather than locomotion, since swimming velocity was inhibited. Glutathione
S-transferase (GST) activity was signicantly increased in the digestive gland of common prawn exposed
to 19 and 39 ng L1 deltamethrin. Catalase (CAT) activity was signicantly increased in digestive gland of
prawn exposed to 19 ng L1 deltamethrin. However, CAT activity decreased in digestive gland of prawn
exposed to 39 ng L1 , suggesting an antioxidant defense system failure concomitant with high levels of
lipid peroxidation. Thus, global results showed that decreased swimming velocity was not associated
with cholinesterase inhibition. In fact, the impairment of swimming velocity may be due to allocation
of energy for detoxication and antioxidant protection instead of swimming activity. The present study
showed that swimming velocity could be used as an ecologically relevant tool and a sensitive endpoint
to assess and complement the study of pesticide effects on marine organisms.
2012 Elsevier B.V. All rights reserved.

1. Introduction
Pyrethroids are a class of neurotoxic pesticides, the use of which
has been continuously increasing during the last two decades
(Amweg et al., 2005; Oros and Werner, 2005; Meacham et al., 2008;
Wolansky and Harrill, 2008). These compounds are derivatives and
synthetic analogues of natural pyrethrins (Soderlund et al., 2002;

Corresponding author at: CIIMAR Centro Interdisciplinar de Investigaco Marinha e Ambiental, Laboratrio de Ecotoxicologia e Ecologia, Universidade do Porto,
Rua dos Bragas, 289, 4050-123, Porto, Portugal. Tel.: +351 223401828.
E-mail addresses: colivei@ciimar.up.pt, cristianav.oliveira@gmail.com
(C. Oliveira).
0166-445X/$ see front matter 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.aquatox.2012.08.010

Hossain et al., 2005; Wolansky and Harrill, 2008), and can be categorized in two types, based on structure-activity and symptomology
(Soderlund et al., 2002; Nasuti et al., 2003; Hossain et al., 2005;
Meacham et al., 2008; Anadn et al., 2009). Nevertheless, both
types act mainly on the nervous system, especially on the voltagedependent sodium channels of excitable membranes, inducing a
prolongation of the sodium current during excitation caused by
membrane depolarization (Narahashi, 1992).
Wolansky and Harrill (2008) have reviewed a large number of
studies showing behavioral alterations after pyrethroid exposure.
Since most of them were performed with mammals, there is a need
to evaluate the effects of these pesticides in aquatic organisms, as
pyrethroids may be spread to aquatic environments by agricultural
and urban runoff (Wang et al., 2009). In fact, these compounds are

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C. Oliveira et al. / Aquatic Toxicology 124125 (2012) 209216

highly toxic to aquatic invertebrates, with LC50 values less than

1 ppb, and even at non-lethal concentrations they are able to induce
signicant behavioral changes in aquatic invertebrates affecting
their survival (Snchez-Fortn and Barahona, 2005). According to
Wolansky and Harrill (2008), all pyrethroid compounds, regardless of species, are able to impair motor function that is the most
extensively characterized neurobehavioral endpoint for pyrethroid
effects. Therefore, the study of locomotor performance, particularly
swimming behavior, after exposure to pyrethroids is of considerable interest. Swimming behavior is considered a main character
determining survival in many species of aquatic organisms and is
frequently used as a behavioral endpoint (Plaut, 2001; Roast et al.,
2000a,b, 2001; Garca-de la Parra et al., 2006; Zhang et al., 2006; De
Lange et al., 2009; Gravato and Guilhermino, 2009; Almeida et al.,
2010, 2012).
Other endpoints, such as biomarkers that provide useful information on the mechanism of pyrethroid toxicity, should be used
in conjunction with neurobehavioral effects generating relevant
information for estimating the effects of hazards (Wolansky and
Harrill, 2008). Biomarkers are based on perturbations of physiological and biochemical parameters, which persist after exposure,
and have been widely used to assess the exposure of organisms to a
range of chemicals in the environment (Badiou et al., 2008; van der
Oost et al., 2003). Among the most commonly used biomarkers,
those associated with oxidative stress are particularly important
(Dorts et al., 2009; Abele et al., 2011), since the mechanisms of toxicity for most pesticides, including pyrethroids, are the production
of free radicals, induction of lipid peroxidation (LPO), and disturbance of the total antioxidant capability of the body (Mohammad
et al., 2004). A suite of biochemical defense mechanisms, called the
antioxidant defense system, are found in all aerobic organisms to
prevent cellular damage caused by reactive oxygen species. Both
enzymatic and non-enzymatic antioxidants counteract the deleterious effects of reactive oxygen species, protecting cells from
oxidative damage (Livingstone, 2001).
Changes in energy metabolism are also used to assess the effects
of pollutants on aquatic organisms (Lee et al., 2002; Lima et al.,
2007; Rao, 2006; Sancho et al., 1997, 1998), since additional energy
for detoxication may be needed to maintain physiological or biochemical functions at a normal level (Choi et al., 2001; Mouneyrac
et al., 2011).
Acetylcholinesterase (AChE) is an enzyme responsible for the
rapid degradation of acetylcholine at the cholinergic synapses
allowing precise control and modulation of the neural transmission.
Its activity changes have been initially used to show the effects of
anticholinesterase chemicals, like organophosphate and carbamate
pesticides (Badiou et al., 2008). In addition, inhibition of swimming velocity may be associated with inhibition of AChE activity
(Blint et al., 1995). However, the few studies concerning the effects
of pyrethroids on AChE have shown contradictory results (Badiou
et al., 2008; Blint et al., 1995; Elhalwagy and Zaki, 2009; Hossain
et al., 2004, 2005; Velsek et al., 2007).
Deltamethrin is one of the most toxic type II pyrethroids (Dorts
et al., 2009; Snchez-Fortn and Barahona, 2005; Velsek et al.,
2007) and has been used as an alternative pesticide in animal
health, in vector control and in public health (Dorts et al., 2009).
Prawns are good indicators of aquatic contamination, because
their biochemical stress responses are quite similar to those
found in other crustaceans and invertebrates species (Vijayavel
and Balasubramanian, 2009). The common prawn, Palaemon serratus, was chosen in the present study to assess the effects of
deltamethrin.
The main purposes of this study were to investigate the effects
of deltamethrin on biomarkers and behavior of P. serratus, since
such information on marine organisms is still scarce, and to establish possible associations between different levels of biological

organization. This approach will allow determining which

biomarkers might be ecologically relevant (if they are linked to
alterations in behavior) and to demonstrate if behavioral parameters can be useful to complement the information obtained with
biomarkers about the effects of pesticides. In this context, it was
also investigated if behavior can be used as endpoint for ecotoxicological bioassays and if this behavioral endpoint can be as sensitive
as biomarker endpoints. Swimming velocity was used as behavioral endpoint. Eye AChE and muscle cholinesterase (ChE) activities,
muscle isocitrate dehydrogenase (IDH) and lactate dehydrogenase
(LDH), and digestive gland glutathione S-transferase (GST), glutathione peroxidase (GPx) and catalase (CAT) activities were used
to assess the effects of deltamethrin on key enzymes. LPO was
assessed as a marker of oxidative damage.
2. Material and methods
2.1. Chemicals
(S)--Cyano-3-phenoxybenzyl (1R)-cis-3-(2,2-dibromovinyl)2,2-dimethylcyclopropane carboxylate (deltamethrin, minimum
98%) of analytical standard grade was purchased from
SigmaAldrich Chemical Corporation. All other chemicals were of
analytical grade and were obtained from SigmaAldrich (USA),
Boehringer (Germany), Merch (Germany) and Bio-Rad (Germany).
2.2. Prawn capture and maintenance
P. serratus (common prawn) specimens were captured at Praia
Norte of Viana do Castelo (Northwest of Portugal) using a handoperated net during low tide. Selected organisms were transported
to laboratory and allowed to recover for 15 days in 200 L tanks with
aerated saltwater ltered by activated carbon (chemically) and a
bacteria lter (biologically) (salinity 32 g L1 ). Prawns were kept at
18 C with a 14 h:10 h (light:dark) photoperiod and fed (2 g/100
prawns) three times per week with commercial food (Complete
feedingstuff for sh, 4.5 mm, mashed; composition: 40.3% crude
protein, 16.3% crude fat, 9.2% crude ber and 10.3% ash). Water
renewal and determination of abiotic parameters (salinity, temperature, pH and dissolved oxygen) was made once per week.
2.3. Acute bioassay
After recovery, prawns were individually exposed to
deltamethrin (0.6, 1.2, 2.4, 4.9, 9.8, 19.5, 39, 78, 156 and 313 ng L1 )
for 96 h in 700 mL of each test solution (salinity 32 g L1 ). Test
concentrations of deltamethrin were prepared by serial dilutions
of a stock solution (10 mg L1 ) prepared in dimethyl sulfoxide
(DMSO). Two more conditions were used: a control and a control
with the solvent (by adding the same volume of DMSO used on
each exposure aquarium, i.e. 28 L control + DMSO). Prawns
with 2.78 0.04 cm length, weighing 0.65 0.03 g, were used in
bioassays. Prawns were not fed 2 days before assays and during
the exposure periods. Ten prawns were used per treatment. The
bioassays were carried in a room with controlled photoperiod
(14 h:10 h light:dark) and temperature (18 C). Abiotic parameters
(salinity, temperature, pH and dissolved oxygen) were monitored
daily during the exposure period.
2.4. Swimming velocity
After 96 h of exposure, each prawn was individually transferred
to a 4 m long track race with 12 cm diameter and allowed to swim
against a water ow of 7 L min1 . The swimming velocity (cm s1 )
was calculated as the quotient between the distance (cm) covered
and the time (s) of swimming (Gravato and Guilhermino, 2009).

C. Oliveira et al. / Aquatic Toxicology 124125 (2012) 209216

211

After swimming trials, each prawn was allowed to recover during

2 h in the respective exposure aquarium.
2.5. Assessment of biomarkers
After 2 h recovery, total body weight and the total length
(rostrumtelson) of each prawn were determined before they were
sacriced by decapitation and dissected. Eyes, digestive gland and
the abdominal muscle were rapidly isolated on ice, frozen, and
maintained at 80 C until further analysis. Eyes and a portion
of muscle were homogenized in 1 mL of ice-cold k-phosphate
buffer (pH 7.2, 0.1 M) and centrifuged at 3300 g for 3 min at
4 C. The resulting supernatants from eye and muscle samples
were used to measure ChE activity by Ellmans technique (Ellman
et al., 1961) adapted to microplate by Guilhermino et al. (1996).
No distinction was made between different forms of ChE and
acetylthiocholine was used as substrate in all the assays. Eye of
P. serratus contains mainly AChE activity (Frasco et al., 2006) and
thus, the enzyme activity measured in the eye was considered to
represent AChE activity, while in the muscle the activity was designated as ChE activity. The supernatant from muscle samples was
also used to determine IDH activity by measuring the increase of
NADPH at 340 nm according to Ellis and Goldberg (1971) adapted
to microplate (Lima et al., 2007).
Another portion of muscle was used to analyze LDH activity. For
LDH determination, samples were submitted to a cycle of 3 freezing
and thawing cycles and homogenized in 1 mL of ice-cold Tris/NaCl
buffer (pH 7.2, Tris 81.3 mM; NaCl 203.3 mM) and centrifuged at
3300 g for 3 min at 4 C. The resulting supernatant was collected
and LDH activity determined by measuring the amount of pyruvate
consumed due to NADH oxidation at 340 nm according to Vassault
(1983) adapted to microplate (Diamantino et al., 2001).
Digestive gland was homogenized in k-phosphate buffer (pH
7.4, 0.1 M). Part of the homogenate was used to determine the
LPO by measuring the thiobarbituric acid reactive species (TBARS),
according to Ohkawa (1979) and Bird and Draper (1984), with the
adaptations of Torres et al. (2002). LPO was expressed in nmol
TBARS/g. The remaining homogenate was centrifuged at 10,000 g
for 20 min at 4 C to isolate the post-mitochondrial supernatant
(PMS). The PMS was used for determinations of GST, GPx and
CAT activities. GST activity was quantied by the conjugation
of reduced glutathione (GSH) with 1-chloro-2,4-dinitrobenzene
(CDNB) at 340 nm (Habig et al., 1974) adapted to microplate (Frasco
and Guilhermino, 2002). GPx activity was determined by measuring the decrease of NADPH at 340 nm using H2 O2 as substrate (Floh
and Gunzler, 1984). CAT activity was determined by measuring the
H2 O2 consumption at 240 nm according to Clairborne (1985).
All enzyme activities are expressed as nmol min1
mg protein1 , except for catalase that was expressed as
mol min1 mg protein1 . Protein of all samples was quantied according to Bradford (1976), adapted to microplate as
indicated in Guilhermino et al. (1996), using bovine -globulin as
protein standard and a wavelength of 600 nm. All measurements
were done at a constant temperature of 25 C.
2.6. Statistical analysis
LC50 and 95% condence limits were calculated with Probit
Analysis using SPSS 16.0 software. All data were rst analysed by
KolmogorovSmirnov and Bartletts tests to check normality and
homoscedasticity, respectively. Appropriate data transformations
were made whenever necessary. Then, one-way analysis of variance (ANOVA) was used to compare different treatments, followed
by Dunnetts test to determine the differences when compared
to control + DMSO, and also the lowest observed effect concentration (LOEC). For all data, outlier values (more than 3.5 standard

Fig. 1. Swimming velocity of Palaemon serratus exposed to 0.639 ng L1

deltamethrin for 96 h. Values are the mean of 10 prawns per treatment with the
corresponding standard error bars. 0 and 0 represents control only with saltwater and control with solvent (control + DMSO), respectively. * indicates statistically
signicant differences (p < 0.05, Dunnett test) relative to control (control + DMSO).

deviations from the group mean) were removed. Signicance level

was 0.05 and SPSS 16.0 package was used for statistical analysis.
3. Results
The estimated 96 h LC50 value for common prawn exposed
to deltamethrin was 48.4 ng L1 with a CI95% between 34.1 and
67.4 ng L1 .
The swimming velocity of common prawn was signicantly
decreased after 96 h exposure to deltamethrin compared to control + DMSO (F(8;83) = 15.607; p = 0.000), showing a LOEC value
of 0.6 ng L1 (58% inhibition) and reaching 87% inhibition at
39 ng L1 (Fig. 1). Eye AChE activity was signicantly increased
(F(8;77) = 3.613; p = 0.001) in prawns exposed to 0.6, 1.2 and
2.4 ng L1 deltamethrin when compared to control + DMSO, with
increases of 78, 66 and 100%, respectively (Fig. 2A). Muscle ChE
activity was signicantly increased (F(8;80) = 2.662; p = 0.013) in
prawns exposed to 19.5 and 39 ng L1 deltamethrin compared
to control + DMSO, with inductions of 73 and 81%, respectively
(Fig. 2B).
GST activity was signicantly increased (F(8,82) = 4.210;
p = 0.000) in digestive gland of prawns exposed to 19.5 and
39 ng L1 deltamethrin compared to control + DMSO, with inductions of 221 and 255%, respectively (Fig. 3A). CAT activity was
signicantly increased (F(8,80) = 2.389; p = 0.024) in digestive
gland of prawns exposed to 19.5 ng L1 deltamethrin, showing an increase of 118% when compared to control + DMSO
(Fig. 3B). No signicant differences (F(8,74) = 0.732; p = 0.663) were
found for GPx activity in digestive gland of prawns exposed to
deltamethrin (Fig. 3C). Level of LPO was signicantly increased
(F(8,73) = 2.546; p = 0.018) in digestive gland of prawns exposed to
39 ng L1 deltamethrin, reaching almost 90% increase compared to
control + DMSO (Fig. 3D).
LDH activity was signicantly increased (F(8,80) = 27.326;
p = 0.000) in muscle of prawns exposed to 0.6 (283% increase),
1.2 (300% increase), 2.4 (374% increase) and to 4.9 ng L1 (340%
increase) deltamethrin (Fig. 4A) compared to control + DMSO.
Signicant inductions were observed in muscle IDH activity
(F(8,81) = 2.139; p = 0.043) of prawns exposed to 19.5 and 39 ng L1
deltamethrin (Fig. 4B).
4. Discussion
The estimated 96 h LC50 value for P. serratus exposed to
deltamethrin was 48.4 ng L1 (CI95% 34.167.4 ng L1 ). This value
for prawns is in a similar range (ng L1 ) as the value previously

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C. Oliveira et al. / Aquatic Toxicology 124125 (2012) 209216

60

50

Eye AChE activity

(nmol mg-1 protein-1)

Muscle ChE activity

(nmol mg-1 protein-1 )

70

40
30
20
10

19.5

39

6
5
4
3
2
1
0

0
0

0.6

0'

1.2

2.4

4.9

9.8

19.5

39

0.6

0'

1.2

2.4

4.9

9.8

Deltamethrin concentration (ng L -1)

Deltamethrin concentration (ng L-1)

Fig. 2. Eye AChE (A) and muscle ChE (B) activities of P. serratus exposed to 0.639 ng L1 deltamethrin for 96 h. Values are the mean of 10 prawns per treatment with the
corresponding standard error bars. 0 and 0 represents control only with saltwater and control with solvent (control + DMSO), respectively. * indicates statistically signicant
differences (p < 0.05, Dunnett test) relative to control (control + DMSO).

reported for Paratya australiensis exposed to deltamethrin in articial saltwater at different salinities (about 35 ng L1 ) (Thomas
et al., 2008). Moreover, Viran et al. (2003) observed that the 48 h
LC50 value for guppies (Poecilia reticulata) exposed to deltamethrin
was 5130 ng L1 and Velsek et al. (2007) observed that the
96 h LC50 value for rainbow trout (Oncorhynchus mykiss) larvae and fry exposed to deltamethrin was 1170 and 1700 ng L1 ,
respectively. Thus, the current results show that P. serratus is

more sensitive to deltamethrin than sh species (Fig. 5). In fact,

Thomas et al. (2008) also observed that crustacean species, namely
P. australiensis and Ceriodaphnia cf. dubia, were more sensitive
to deltamethrin than larvae of the rainbow sh (Melanotaenia
duboulayi). Therefore, the present result is in good agreement with
previous studies, since deltamethrin is an insecticide, and phylogenetically crustaceans are more closely related to insects than
sh.

3.5
40

*
*

30
25
20
15
10

B
3
CAT activity
(nmol mg-1 protein-1 )

GST activity
(nmol mg-1 protein-1 )

35

2.5
2
1.5
1
0.5

0
0

0'

0.6

1.2

2.4

4.9

9.8

19.5

39

0'

800

2.4

4.9

9.8

19.5

39

700
LPO
nmol TBARS g wt-1

5
GPx activity
(nmol mg-1 protein-1 )

1.2

Deltamethrin concentration (ng L-1 )

Deltamethrin concentration (ng L-1 )

6

0.6

4
3
2

600
500
400
300
200

1
100

0
0

0'

0.6

1.2

2.4

4.9

9.8

19.5

Deltamethrin concentration (ng L-1 )

39

0
0

0'

0.6

1.2

2.4

4.9

9.8

19.5

39

Deltamethrin concentration (ng L-1)

Fig. 3. Digestive gland GST (A), CAT (B), and GPx (C) activities and LPO levels (D) of P. serratus exposed to 0.639 ng L1 deltamethrin for 96 h. Values are the mean of 10
prawns per treatment with the corresponding standard error bars. 0 and 0 represents control only with saltwater and control with solvent (control + DMSO), respectively. *
indicates statistically signicant differences (p < 0.05, Dunnett test) relative to control (control + DMSO).

C. Oliveira et al. / Aquatic Toxicology 124125 (2012) 209216

Muscle LDH activity

(nmol mg-1 protein-1)

35

14

*
*

30

12
Muscle IDH activity
(nmol mg-1 protein-1)

40

213

25
20
15
10

10
8
6
4
2

5
0

0
0'

0.6

1.2

2.4

4.9

9.8

Deltamethrin concentration (ng

19.5

39

0'

L-1 )

0.6

1.2

2.4

4.9

9.8

19.5

39

Deltamethrin concentration (ng L-1)

Fig. 4. Muscle LDH (A) and IDH (B) activities of P. serratus exposed to 0.639 ng L1 deltamethrin for 96 h. Values are the mean of 10 prawns per treatment with the
corresponding standard error bars. 0 and 0 represents control only with saltwater and control with solvent (control + DMSO), respectively. * indicates statistically signicant
differences (p < 0.05, Dunnett test) relative to control (control + DMSO).

5130 ng L-1 LC50 (48 h) in Poecilia reticulata (Viran et al., 2003)

4000 ng L-1 Lost of general activity and equilibrium of Poecilia reticulata (Viran et al., 2003)
2000 ng L-1 Concentration-dependent effects on swimming behavior of Oreochromis niloticus larvae
and fry (Karasu Benli et al., 2009)
1170 and 1700 ng L-1 LC50 (48 h) in Oreochromis niloticus larvae and fry, respectively (Karasu Benli
et al., 2009)
1500 ng L-1 LC50 (96 h) in Channa punctatus (Sayeed et al., 2003)
1000 ng L-1 LC50 (96 h) in Oncorhynchus mykiss juveniles (Velek et al., 2007)

Deltamethrin concentration (g L-1)

1000 ng L-1 Lost of movement coordination of Oncorhynchus mykiss juveniles (Velek et al., 2007)
1000 ng L-1 Significantly lower values of plasma glucose, alanine aminotransferase, cholinesterase
and significantly higher values of erythrocyte count, haemoglobin content, haematocrit and plasma
total protein, albumins, ammonia, aspartate aminotransferase, creatinekinase and calcium in
Oncorhynchus mykiss juveniles (Velek et al., 2007)
750 ng L-1 Significant increase of LPO levels and GSH content in liver, kidneys and gills; significant
induction of GST and GPx activities in liver and kidneys, and inhibition in gills; and significant
inhibition of CAT activity in liver, kidneys and gills in Channa punctatus (Sayeed et al., 2003)
135 to 212 ng L-1 LC50 (96 h) in Melanotaenia duboulayi exposed in water at different salinities
(Thomas et al., 2008)
100 ng L-1 Increase of tGSH content and CAT activity and inhibition of hepatopancreas GPx and
muscle AChE activities in Penaeus monodon. No significant effects on LPO levels neither on GST
activity (Tu et al., 2012)
100 ng L-1 Significantly increase od LPO levels in gills, however no significant effects on CAT, GPX
and GST activities (Dorts et al., 2009)
48 ng L-1 LC50 (96 h) in Palaemon serratus (in the present study)
39 ng L-1 Statistically significant increase of MDA content and GSH levels in Tinca tinca brain during
the recovery time, after 60 days of exposure (Hernndez-Moreno et al., 2010)
39 ng L-1 Significant increase of LPO levels in Palaemon serratus (in the present study)
32 to 37 ng L-1 LC50 (96h) in Paratya australiensis exposed in water at different salinities (Thomas et
al., 2008)
20 to 29 ng L-1 LC50 (96h) in Ceriodaphnia cf. dubia exposed in water at different salinities (Thomas
et al., 2008)
19.5 ng L-1 Significantly increase of muscle ChE activity and digestive gland GST and CAT
activities in Palaemon serratus (in the present study)
6 ng L-1 Significantly inhibition of Palaemon serratus swimming velocity and significantly
increase of eye AChE and muscle LDH activities (in the present study)
Fig. 5. Sensitivity of biomarkers and behavioral responses to deltamethrin exposure compared to lethal effects.

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C. Oliveira et al. / Aquatic Toxicology 124125 (2012) 209216

Traditionally, mortality has been taken as an endpoint to

assess the risk of pesticides for organisms. However, exposure to
pesticides below lethal concentrations may already affect the performance of organisms. In fact, in the present study, it was observed
that deltamethrin decreased the swimming velocity of prawn at a
concentration 80 times lower than its LC50 . The decrease of swimming velocity by deltamethrin is in good agreement with recent
studies, which showed that deltamethrin affected the behavior of
sh species (Fig. 5): loss of general activity and equilibrium of P.
reticulata (Viran et al., 2003); concentration-dependent effects on
swimming behavior of Oreochromis niloticus larvae and fry (Karasu
Benli et al., 2009); and loss of movement coordination of O. mykiss
juveniles (Velsek et al., 2007). However, studies including behavioral effects of deltamethrin in crustaceans are scarce. Nevertheless
the current study is in good agreement with previous studies concerning the adverse effects of other pyrethroids in crustaceans:
Christensen et al. (2005) observed that cypermethrin decreased the
ability of Daphnia magna to swim after 48 h exposure to concentrations equal or higher than 100 ng L1 ; Nrum et al. (2010) observed
that 100 ng L1 lambda-cyhalothrin caused the immobilization of
Leuctra nigra, Gammarus pulex and Heptagenia sulphurea.
The impairment of swimming may have serious consequences
for macroinvertebrates, since the ability of individuals to acquire
food, search for mates or to avoid predators may be reduced (Nrum
et al., 2010). Thus, swimming velocity seems to be an ecologically
relevant parameter that must be taken into consideration when one
aims to assess the ecologically relevant effects of contaminants on
aquatic organisms. In addition, high sensitivity, simplicity of testing procedures, and reproducibility across studies are some of the
advantages that were previously mentioned when using measurements of motor activity as an endpoint to assess the neurotoxicity
induced by pyrethroids (Wolansky and Harrill, 2008). Pyrethroids
are known to act mainly on the voltage-dependent Na+ channels
of the nerve cell membrane, although some studies have demonstrated that they may have secondary effects on the cholinergic
system, particularly on AChE activity (Badiou et al., 2008; Hossain
et al., 2004, 2005; Velsek et al., 2007). Behavioral disruption and
neurological dysfunction have been investigated in many studies,
and relationships between behavioral impairments and AChE inhibition are well reviewed in a recent publication (Amiard-Triquet,
2009). However, in the present study, behavioral impairment was
not followed by an inhibition of AChE or ChE: eye AChE activity was signicantly increased in prawns exposed to 0.6, 1.2 and
2.4 ng L1 deltamethrin. Nevertheless, current results are in good
agreement with a previous study performed with bees exposed to
deltamethrin (Badiou et al., 2008) that showed increased activity of
AChE in the head of surviving insects. However, other studies show
contradictory results, since mostly inhibition of AChE activity after
exposure to pyrethroids has been observed in studies using sh
and rats (Badiou et al., 2008; Badiou and Belzunces, 2008; Blint
et al., 1995; Elhalwagy and Zaki, 2009; Hossain et al., 2004, 2005;
Velsek et al., 2007). The induction of ChE activity observed in the
present study may be due to an increased release of ACh in the
hippocampus (Hossain et al., 2004), which may lead to an overcompensation, namely the increase of AChE activity in the cholinergic
system (Badiou et al., 2008). Therefore, the present study showed
that behavioral impairments are not always related to AChE inhibition. Thus, other metabolic or physiological disturbances could be
the cause of the observed impairment of swimming velocity.
It is well documented that deltamethrin may induce oxidative
stress (Fig. 5, Sayeed et al., 2003; Hernndez-Moreno et al., 2010; Tu
et al., 2012). To mitigate and repair the oxidative damage caused by
free radicals, organisms have evolved complex enzymatic antioxidant systems, such as the enzymatic activities of GST, GPx and
CAT (van der Oost et al., 2003; Abele et al., 2011; Regoli et al.,
2011). In the current study, increased levels of LPO were observed

in prawns exposed to 39 ng L1 even when the activity of GST was

increased, suggesting a detoxication failure. In addition, CAT activity was not signicantly increased in prawns exposed to 39 ng L1
deltamethrin, suggesting also a failure in antioxidant protection,
which may have led to the increased oxidative damage observed.
Oxidative stress has been suggested as one of the molecular mechanisms generally involved in pesticide-induced toxicity, particularly
that caused by pyrethroids (Elhalwagy and Zaki, 2009; Mohammad
et al., 2004; Vijayavel and Balasubramanian, 2009). In previous
works performed with prawns (Dorts et al., 2009; Vijayavel and
Balasubramanian, 2009) an increase in LPO levels in Penaeus monodon exposed to deltamethrin and fenvalerate, respectively, was
also observed. Increased levels of total glutathione and CAT activity
were also found in P. monodon exposed to deltamethrin, despite the
absence of increased levels of LPO (Tu et al., 2012). Concerning sh,
exposure to deltamethrin also signicantly increased the levels of
LPO, GSH, and the activities of GST, GPx and CAT in Channa punctatus
(Sayeed et al., 2003). Moreover, it was observed that high doses of
type I (permethrin) and type II (cypermethrin) pyrethroids induced
a signicant increase of the oxidation index of plasma membrane of
erythrocytes of rats (Nasuti et al., 2003). Furthermore, deltamethrin
increased the levels of LPO in liver and blood plasma of rats (Yousef
et al., 2006; Tuzmen et al., 2008).
The ability to resist a toxicant may be expensive in terms of
energy, compromising the amount of energy available for other
biological processes (Amiard-Triquet, 2009; Choi et al., 2001).
Therefore, the impairment of prawns swimming velocity was probably due to insufcient energy. The energy cost to increase the
activity of detoxication enzymes constitutes a signicant component of the energy budget of an organism (Mouneyrac et al.,
2011). Thus, the energy seems to be allocated for detoxication processes and repair. Concerning energy metabolism, LDH
activity was signicantly increased in prawns exposed to low concentrations of deltamethrin (0.6 up to 4.9 ng L1 ) demonstrating
metabolic changes induced by the pesticide. LDH activity is particularly important when a considerable amount of additional
energy is rapidly required (Diamantino et al., 2001). Thus, current results suggest that prawns exposed to low concentrations
of deltamethrin (0.64.9 ng L1 ) were able to get additional energy
for detoxication and antioxidant protection, since no oxidative
damage was observed. However, LDH activity decreased in prawns
exposed to the highest concentrations tested (>4.9 ng L1 ), whereas
IDH activity was signicantly induced in prawns exposed to 19.5
and 39 ng L1 that suggests a switch from anaerobic to aerobic
metabolism to produce the required energy to face the exposure
to the pesticide. The decrease of LDH activity may be due to hepatocellular necrosis leading to the loss of the enzyme and/or its
inhibition (Wang and Zhai, 1998). The observations in the present
study suggest that the activation of the detoxication system to face
exposure to deltamethrin has an energetic cost that compromises
the swimming velocity of prawn, despite the additional energy
produced.

5. Conclusion
The swimming velocity was decreased in prawns exposed to
deltamethrin. The impairment of swimming velocity was not concomitant with the inhibition of AChE and ChE activities, showing
that behavioral impairment is not always explained by neurological disturbances. In addition, swimming velocity was still inhibited
despite the production of additional energy, suggesting an allocation of energy for detoxication and antioxidant protection.
Swimming velocity is essential for the performance and survival of
prawns and should be considered as an ecologically relevant endpoint, since it can be linked with tness-related parameters, such

C. Oliveira et al. / Aquatic Toxicology 124125 (2012) 209216

as search for food and avoidance of predators. Furthermore, the

present work complemented the knowledge about the effects of
deltamethrin on biomarkers and behavior of a marine species.

Acknowledgements
This study was supported by the Portuguese Fundaco para a
Cincia e a Tecnologia (FCT), FEDER and European social funds of
MCTES (POPH-QREN-Tipology 4.2).

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