Interactive Effects of Two Environmental Stressors, PAH Contamination and Hypoxia, in the
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Transcript of Interactive Effects of Two Environmental Stressors, PAH Contamination and Hypoxia, in the
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Interactive Effects of Two Environmental Stressors, PAH Contamination and Hypoxia, in the
Brown Shrimp, Penaeus aztecus
Enmin Zou
Department of Biological SciencesNicholls State University
Thibodaux, LA 70310
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Dissolved OxygenContours (mgl)
Hypoxic Zone in NorthernGulf of Mexico
Algal Blooms
Depletion of Oxygen
Degradation of Organic Materials
Nutrients input
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Hypoxia is an environmental stress for aquatic organisms, such as penaeid shrimps, inhabiting thesewaters.
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Contamination of polycylcic aromatic hydrocarbons (PAHs) from gas and petroleum production on the northern continental slope of the Gulf of Mexico
Total PAH in Gulf of Mexico sediments:
< 0.005 – 36.7 ppm< 0.005 – 36.7 ppm (Wade et al., 1988)
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Aquatic animals in the northern Gulf of Mexico are subject to dual stresses of hypoxia and PAH contamination
Hypoxia PAH Contamination
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The brown shrimp, Penaeus aztecus, is animportant commercial shrimp, whose life history overlaps with the hypoxic zone in the northern Gulf of Mexico.
Penaeus aztecus is faced with two stressors:• Hypoxia• PAH contamination
Hypoxia
PAHs
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Objectives:
1. To investigate whether acute exposure to naphthalene, a representative petroleum PAH, can make shrimps more vulnerable to hypoxia.
2. To investigate whether hypoxia can promotenaphthalene bioaccumulation.
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Oxy
gen
Con
sum
ptio
n R
ate
(OC
R)
0O2 Concentration (C)
Oxyregulation
Oxy
conf
orm
atio
n
Cc
OCR = C/(a + bC)
Impacts of hypoxia on oxyregulation of the brown shrimp,Penaeus aztecus
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O2 Concentration (C)
Oxy
gen
Con
sum
ptio
n R
ate
(OC
R)
0Cc Cs
Cc = (aCs/b)1/2
Oxyregulating Capacity = a/b
OCR = b – a/C (Zou et al., 1993)
S = ∫Ca/ba/b(OCR-A)dC + (OCR-A)dC + ∫Cs
C (OCR-B)dC(OCR-B)dC = minimum
Oxy
conf
orm
atio
nA
Oxyregulation
B
Or when S = 0•
a/b
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a/b, an index for oxyregulation, reflects an animal’s susceptibility to hypoxia.
The greater a/b, the smaller the oxyregulatingcapacity, i.e. the sooner onset of hypoxic stress.
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Four groups of brown shrimps were respectively exposed to:
• Clean seawater
• Seawater with 0.05% v/v acetone
• Seawater with 0.5 mg/l naphthalene
• Seawater with 2.0 mg/l naphthalene
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0.00
1.00
2.00
3.00
4.00
5.00
6.00
0.00 2.00 4.00 6.00 8.00
Oxygen Concentration (mg/l)
OC
R (
gO2
• g
-1 •
min
-1)
OCR = 3.64 - 1.54/C ( p < 0.05)A
0.00
1.00
2.00
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6.00
0.00 2.00 4.00 6.00 8.00
Oxygen Concentration (mg/l)
OC
R (
gO2
• g
-1 •
min
-1)
OCR = 4.81 - 2.18/C (p < 0.001)
B
0.001.002.003.004.005.006.00
0.00 2.00 4.00 6.00 8.00
Oxygen Concentration (mg/l)
OC
R (
gO2
• g
-1 •
min
-1)
OCR = 4.99 - 4.32/C (p < 0.0005)C
0.001.002.003.004.005.006.00
0.00 2.00 4.00 6.00 8.00
Oxygen Concentration (mg/l)
OC
R (
gO2
• g
-1 •
min
-1)
OCR = 7.21 - 14.55/C (p < 0.0005)
D
Clean Seawater (wt = 9.00 g) 0.05% v/v Acetone (wt = 7.02 g)
0.5 mg/l Naphthalene (wt = 8.17 g) 2.0 mg/l naphthalene (wt = 7.53 g)
Cc = 2.53 mg/l
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0.00
0.50
1.00
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2.00
2.50
3.00
3.50a/
b
(6)(20)
(12)
(18)
Control A Control B 0.5 mg/lNaphthalene
2.0 mg/lNaphthalene
#***
Effects of acute exposure to naphthalene on oxyregulating capacity of Penaeus aztecus
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Changes in critical oxygen concentration (Cc) due to naphthalene
Clean Seawater 2.53 mgO2/l
Seawater with 0.05%v/v acetone
2.89 mg O2 /l
0.5 mg/l Naphthalene
3.76 mg O2 /l
2.0 mg/l Naphthalene
4.22 mgO2/l
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Conclusion:
Acute exposure to naphthalene reduces oxyregulating capacity of the brown shrimp, Penaeus aztecus, subjected to progressive hypoxia,thereby making shrimps more suceptible to hypoxia.
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Impacts of hypoxia on naphthalene bioaccumulationin the brown shrimp, Penaeus aztecus
Working Hypothesis
Hypoxia
Increased Ventilation
Increased Water-flow over Gills
Enhanced Bioaccumulationof Naphthalene
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N2 Tank
Degassing Tank
Exposure Tank
Oxygenation Tank
Pump
Schematic diagram of hypoxia experimental system
Shrimps
Exposure Tank O2 Concentration: 1.32 – 2.61 mg/l
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Hypoxia system 1 Exposure tank
2 Oxygenation tank
3 Nitrogen saturated tank
3
2
1
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• Clean Seawater• 0.005% v/v Acetone + Normoxia• 0.005% v/v Acetone + Hypoxia• 10 g/l Naphthalene + Normoxia• 10 g/l Naphthalene + Hypoxia
•Clean Seawater• 0.01% v/v Acetone + Normoxia• 0.01% v/v Acetone + Hypoxia• 250 g/l Naphthalene + Normoxia• 250 g/l Naphthalene + Hypoxia
Ten groups of 25 shrimps each were respectively exposed to:
At days 3 and 7 shrimps were sacrificed for branchial andHepatopancreatic tissues.
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0
5
10
15
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25
Normoxia Hypoxia Normoxia Hypoxia
A
Hepatopancreas Gill
0
5
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15
20
25
Normoxia Hypoxia Normoxia Hypoxia
B
Hepatopancreas Gill
0
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15
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Normoxia Hypoxia Normoxia Hypoxia
C
Hepatopancreas Gill
01020304050607080
Normoxia Hypoxia Normoxia Hypoxia
D
Hepotapancreas Gill
10 g/L, day3 10 g/L, day 7
250 g/L, day 3 250 g/L, day 7
7.62 (8) 9.59 (8)
0.51 (8)1.37 (8)
3.35 (6)
9.8 (8)
0.43 (8) 0.67 (8)
8.16 (7)
4.06 (7)
1.21 (7) 1.00(7)
18.98 (7)
26.33 (6)
0.66 (7)0.80 (6)
Naphthalene bioaccumulation in Penaeus aztecusunder hypoxia
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Conclusions:
• Naphthalene was found to mainly accumulate in the hepatopancreas---Naphthalene burden in the hepatopancreas up to 60 times that in the gills.
• Hypoxia did not significantly alter naphthalene bioaccumuluation in either organs.
NaphthaleneBioaccumulation
Uptake
Elimination
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Naphthalene
Naphthalene epoxide
Glutathione conjugates
Naphthalene Metabolism
CYP 1A (Ethoxyresorufin O-deethylase, or EROD)
Glutathione S-transferase(GST)
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EROD activities in the hepatopancreas of Penaeus aztecus subjected to various treatments (pM/min/mg protein)
Ace, acetone; Nor, normoxia; Hyp, hypoxia; Nap, naphthalene; (A) * p < 0.05, versus Nap + Nor; (B) * p < 0.05, versus Nap + Nor; (C) * p< 0.05, ** p< 0.01, versus Clean + Nor; (D) ** p < 0.01, versus Ace + Nor ;
0
20
40
60
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120
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
A
01020304050607080
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
C
*
0
20
40
60
80
100
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
D
01020304050607080
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
B10 g/L, day 3 10 g/L, day 7
250 g/L, day 3 250 g/L, day 7
*
** **
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Summary of results for EROD activities in the hepatopancreas
• Naphthalene significantly induced hepatopancreatic EROD activity under normoxia.
• Hypoxia had no significant effects on hepatopancreatic EROD activity in the presence of naphthalene.
• Acetone significantly inhibited EROD activity in the hepatopancreas.
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0
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
B
EROD activities in the gills of Penaeus aztecus subjected to various treatments (pM/min/mg protein)
Ace, acetone; Nor, normoxia; Hyp, hypoxia; Nap, naphthalene; (A) ** p < 0.01, versus Nap + Nor; (C) ** p < 0.01, versus Clean + Nor; (D) * p < 0.05, versus Clean + Nor, ** p < 0.01, versus Ace + Nor and Nap + Nor, # p < 0.05, versus Ace + Hyp;
0
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
A
**
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
C**
0
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
D*
#
10 g/L, day 3 10 g/L, day 7
250 g/L, day 3250 g/L, day 7
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Summary of results for EROD activities in the gills
• Hypoxia significantly inhibited branchial EROD activities when naphthalene was present.
• Acetone significantly suppressed branchial EROD activities under normoxia
• Under hypoxia, naphthalene at 250 g/l inhibited EROD activities in the gills.
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GST activities in the hepatopancreas of Penaeus aztecus subjected to various treatments (nM/min/mg protein)
Ace, acetone; Nor, normoxia; Hyp, hypoxia; Nap, naphthalene; (B) * p <0.05, ** p < 0.01, versus Nap + Nor; (C) * p < 0.05, versus Clean + Nor, ** p < 0.01, versus Nap + Nor; (D) ** p < 0.01, versus Nap + Hyp;
0
5
10
15
20
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor0
5
10
15
20
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
B*
0
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
C
0
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
D
10 g/L, day 3 10 g/L, day 7
250 g/L, day 3 250 g/L, day 7
A
** ****
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Summary of results for hepatopancreatic GST activities
• Naphthalene significantly induced GST activities in the hepatopancreas under both normoxia and hypoxia.
• Hypoxia significantly elevated GST activities in the hepatopancreas when naphthalene was present.
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0
5
10
15
20
25
30
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
D
GST activities in the gills of Penaeus aztecus subjected to various treatments (nM/min/mg protein)
Ace, acetone; Nor, normoxia; Hyp, hypoxia; Nap, naphthalene; (A) * p < 0.05, ** p < 0.01, versus Nap + Nor; (B) ** p < 0.01, versus Nap + Nor; (C) * p < 0.05, ** p < 0.01, versus Ace + Nor;
B
0
5
10
15
20
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Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
A
02468
10121416
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
02468
10121416
Ace+Nor Ace+Hyp Nap+Nor Nap+Hyp Clean+Nor
C
10 g/L, day 3 10 g/L, day 7
250 g/L, day 3 250 g/L, day 7
*
**
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Summary of results for branchial GST activities
• Hypoxia significantly decreased branchial GST activities in the presence of naphthalene.
• Under normoxia, naphthalene at 250 g/l significantly suppressed branchial GST activities.
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EROD GST
Hepatopancreas No effect +
Gill – –
Effects of hypoxia on activities of EROD and GSTin the presence of naphthalene
NaphthaleneBioaccumulation
Uptake
Elimination
• The significant increase in GST activity may explain the absence of increased naphthalene bioaccumulation in the hepatopancreas when Penaeus aztecus is subjected to hypoxia.
• Alterations in activities of EROD and GST have no bearings on naphthalene bioaccumulation in the gills because gills are not an ideal organ for bioaccumulation of chemicals.
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Take-home messages:
• Acute exposure to PAH compound naphthalene reduces oxyregulating capacity of the brown shrimp.
• Hypoxia does not promote bioaccumulation of the PAH compound naphthalene in the brown shrimp. The absence of such a significant effect is attributed to increased naphthalene metabolism in the hepatopancreas when the shrimp is subjected to hypoxia
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Appreciation to the funding agencies, NOAA and USGS, and my students Rongzhong Ye and Ben Stueben.
Acknowledgements
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Questions?