centroptilum triangulifer) for ecotoxicology and ... · selenium 101 • selenium is an essential...
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Development of a mayfly model (Centroptilum triangulifer) for
ecotoxicology and bioaccumulation studies: Selenium trials
Justin Conley1, David Funk2 and David Buchwalter1
1Department of Environmental and Molecular Toxicology, NCSU2Stroud Water Research Center, Avondale, PA
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Aquatic insects are under- represented in ecotoxicology
• 70-90% of the invertebrate species pool in most lotic systems
• Ecologically important• Extensively used as ecological indicators
• Typically difficult to culture• Complex life cycles, dietary requirements• Flow
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What about Chironomous?
• Too tolerant
• Sediment associated
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Introducing: C. triangulifer• Native to the Eastern
US and Canada• Only lives in rivers,
but prefers marginal habitats
• Parthenogenic• Imago lays eggs
(~1000) upon wetting• Eggs can be stored at
4°C
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Life cycle
Temperature °C Days to adulthood
25 27
20 45
10 179
Source: Sweeney and Vannote, 1984 Freshwater Biology
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Diet: diatoms
Since the biggest “jump” in bioaccumulation of most metals/metalloids in food websoccurs at the incorporation stage (primary producers), trophic transfer from the base of food webs is central to understanding exposure and effects
A culturable mayfly that grazes on periphyton and diatoms is a more environmentallyrelevant test species for streams and rivers than Daphnids or other common “lab rats”
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Selenium movement through food webs: Insights from marine studies
Log
scal
e
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Selenium 101
• Selenium is an essential element with a narrow window between essentiality and toxicity
• Kesterson, Belews lake and other areas of intense Se contamination show population crashes and teratogenicity (birth defects) in fish and birds
• Bioaccumulation of Se in aquatic systems is primarily thought to be via diet.
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Selenium
• Surprisingly little work done in stream systems
• Conventional wisdom is that insects are simply conduits of Se to higher trophic level organisms (fish and birds)
• Limited evidence that invertebrates are affected by Se (but see Debruyn and Chapman, 2007 ES&T)
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Diet
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Experimental design
1.8 L water (ASTM artificial soft water)
Controls
5 µg/L
10 µg/L
20 µg/L
75Se as selenite added as a tracer
This allowed us to quantify Se in waterand in periphyton scrapings over time
Algal plates exposed for 1 week prior totransfer to clean water
20 Centroptilum larvae (7 days old) added to each bottle
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Concentration of Se by algae
5 10 200.0
2.5
5.0
7.5
10.0
12.5
dissolved Se g/L
g
Se /
g al
gae
(dry
wei
ght)
~450 fold enrichment
~490 foldenrichment
~530 foldenrichment
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1.96 2.20 4.44 8.72 11.310
10
20
30
food [Se]
may
fly
g Se
/g(d
ry w
eigh
t)
Trophic transfer of Se from periphyton to mayflyadults (after laying eggs)
2.91 1.89
2.19
1.86
2.43
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450-530 fold(up to 1275)
1.9-2.9 fold1.1-1.3 fold
Log
scal
e
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Effects of dietary Se on mayflies?
Note that these experiments were run with different batches of algal plates. Food quality/algal species differences are important.
Dietary Se in this range does notappear to affect growth
contro
l 1co
ntrol 2 1.9
6 2.2 4.44
8.72
11.31
0100200300400500600700800900
body
wei
ght
g (d
ry)
µg Se / g algae (dry wt)
Cntrl 21.5
0
200
400
600
800
Bod
y m
ass
( g)
*p=0.023
At higher dietary Se, growth is affected
µg Se / g algae (dry wt)
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Cntrl Se
0
500
1000
1500
2000
Eggs
(#/c
lutc
h)
p<0.0001
*
~40 µg Se/gin maternaltissues (postEgg laying)
At higher maternal selenium concentrations, fecundity is affected
~20 µg Se/g in diet
TTF = ~2
n=10
n=10
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0 500 1000 15000
500
1000
1500
2000
2500
Se, r2=0.049Cntrl, r2=0.683
Adult mass
Clu
tch
Size
At high maternal Se concentration, growth is stunted, and fecundity is affected
but reduction of fecundity is not directly tied to reduced growth
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Maternal transfer of Se to eggs is consistent across all maternal Se body burdens/dietary treatments
0 20 40 60 800
10
20
30
402.22.04.48.711.325.517.5
[Se] Mother - Egg complex (ng)
[Se]
Egg
s (n
g)
r = 0.9120p < 0.0001
Diet treatments
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What does it all mean?• The general paradigm of Se movement through
food webs that has been established for marine systems seems to hold for freshwater environments
• Contrary to conventional wisdom, insects can be affected by high selenium content in their diets
• Given the importance of insects in the diets of fish and birds, it is important to understand their bioaccumulation of Se
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Uncertainties and data gaps• One major difference between fresh and marine systems
could relate to inorganic Se forms: selenate uptake into marine algae is quite slow (sulfate competition), whereas selenite uptake is important
• How do primary producers (algae, diatoms) in freshwater vary in their Se bioaccumulation?
• The assumption that dissolved Se uptake is unimportant is not universally true for insects
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Future directions
• Development of Centroptilum as a model test species– TDS work, other elements (Cd, nanoparticles)– Need to normalize diet
• Larval Se work with different mayfly grazers– Bioaccumulation and antioxidant responses
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We recently sequenced a cDNA library from animals treated with:Cu, Cd, Hg mixturehydrogen peroxide20-OH ecdysonephenobarbitolchlorpyrifos/atrazine mixturecontrols
A model for toxicogenomic studies?
7,000 contigs
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Acknowledgements
Dr. Lingtian Xie
NCSU College of Agriculture and Life Sciences
EPA: Greg Pond and MaggiePassmore