Transcript Reece.pps - the National Sea Grant Library

Genetic Considerations in Broodstock Selection for
Oyster Restoration, Aquaculture Development, and
Non-native Species Introductions
Kimberly S. Reece
Virginia Oyster Landings 1880 - 2005
Chesapeake Bay Market Oyster Landings 1931-2005
7
7
Morta lity from H. n elsoni (MSX)
Virginia
be gins
Maryland
5
6
Millions of Bushels
Millions of Bushels
6
5
Total
4
4
P. marin us (Derm o)
3
3
in tens ifie s
2
2
1
1
0
1930
0
1940
1950
1960
1970
1980
1990
2000
Year
What is the best approach to restoration, protection and
preservation of the oyster resource?
Preferred Approach May Depend on Motivations and
Perspectives
What is the goal of oyster restoration?
Industry
Restorationobjective to
become
profitable and
self-sustaining
Ecological
Restoration
To restore habitat
and
populations of
native oysters
To rebuild a
sustainable
harvest
fishery
Native oyster
Develop a
new oyster
industryaquaculture
Non-native oyster
Not necessary exclusive approaches, but emphasis and
measures of success may differ
Possible Solutions
1.
2.
3.
Oyster reef restoration- build/restore habitat (reefs) and establish sanctuaries.
1.
Reefs provide substrate for natural spatfall, sanctuaries protect from fishing
pressure.
2.
Stock reefs with oysters from hatcheries-goal self-sustaining
1.
wild broodstock
2.
selected / domesticated strains?
Aquaculture development through improved selective breeding practices:
1.
Enhanced disease tolerance
2.
Enhanced growth rate
Consideration of alternative Crassostrea species for Chesapeake Bay aquaculture
and maybe restoration of the fishery (or ecological restoration).
1.
Asian oysters are significantly more resistant (tolerant) to MSX and Dermo.
2.
Crassostrea ariakensis tested in Chesapeake Bay has shown:
1.
rapid growth
2.
taste that is acceptable to market
3.
disease tolerance in field trials
Genetic Considerations
Stocking reefs with hatchery oysters
Does it work?
Is it a good idea from the genetics point of view?
Which oysters to use? Wild or Selected?
What is the genetic impact on extant natural
Aquaculture Development
populations?
oyster stocks
to use? but
Diploids
Ultimate goal =Which
self-sustaining
populations,
of or triploids?
Special
genetic
lines might be selected for particular
what
genetic
make-up.
traits of interest.
Maintain genetically healthy lines.
Is there any genetic impact on extant natural
populations?
Introduction of a Non-native Oyster
Aquaculture or on bottom fishery?
Which species? Genetic identification needed.
Which stock? Broodstock source?
Oregon strain too genetically bottlenecked?
Genetic Considerations
(Restoration)
Stocking reefs with hatchery oysters
Does it work?
Is it a good idea from the genetics point of view?
Which oysters to use? Wild or Selected?
What is the genetic impact on extant natural
populations?
Ultimate goal = self-sustaining populations, but of
what genetic make-up?
Should Reefs be Stocked?
•Supportive breeding - adding hatchery broodstock to reefs to
supplement natural populations.
•If we do stock, what is the best broodstock?
• Hatchery oysters from wild broodstock too wimpy? ie. Subject to
high disease mortality?
•Any selected line?
• Different lines (or wild broodstock) be used for different
systems/environments?
wild
I like a pale
ale- 10 ppt
Make mine a
stout-30 ppt
VIMS
selected
Genetic variation
High
Natural spatfall- natural populations
Hatchery oysters from wild broodstock
Selected lines
Low
Highly inbred lines
The answer to the questions of whether to stock and with what,
depends on:
1. The genetic structure of the historical and the extant
populations.
2. The phenotypic relevance of any detected genetic variation. Is
there local adaptation?
3. The genetic impact of hatchery (planted) oysters on the wild
populations and overall genetic variance (Ne).
Do the disease-tolerant oysters, selected lines have a better chance
of survival in the face of disease challenges?
Maybe yes, in the short term, but what about longer term?
Risks of inbreeding?
Environmental change
New stress/challenge:
Selected stock may not be able to survive different challenges-may really be
“wimpy” under a different set of conditions.
Inbreeding may lead to increasing deleterious allele frequencies = line crash
Genetic diversity (higher effective population
size) can be important for survival of a species
Environmental change = new stress/challenge and can results in
elimination of some genetic types :
Others may survive:
Some “natural” populations are demonstrating disease tolerance.
Maybe these are a better source for supportive breeding broodstock
Shell Bar Reef, Great Wicomico River
June-September 2006: biweekly analysis of P. marinus in samples (each n = 25)
of deployed DEBYs and naturally recruited C. virginica
P. marinus Weighted Prevalence
3
2.5
Natural
DEBY
2
1.5
1
0.5
0
6/8/06
6/22/06
Carnegie and Burreson
7/6/06
7/20/06
8/3/06
8/17/06
8/31/06
9/14/06
York River-Disease Data
•Cumulative mortality higher in Ross Rocks -- approaching 100% by
September -- than in DEBYs (63% in October)
•Cumulative mortality in Aberdeen Rocks (58% by October) similar to
DEBYs; Wreck Shoals slightly higher (72%; MSX disease?)
Mortality, York River, 2006
100
Cumulative Mortality (%)
90
80
DEBYs, Resistant
Ross Rock, Susceptible
Aberdeen Natives
Wreck Natives
70
60
50
40
30
20
10
0
Carnegie and
Burreson
1
June
2
July
3
Aug
4
5
Sept
Oct
Motivations for, and the risks of, supportive breedingusing selected/hatchery stocks for restoration efforts.
Motivations
•Increase the chances of survival/reproduction in the face of disease.
•Genetic rehabilitation-introgression of “disease resistance” alleles into
natural populations.
•Ability to genetically track the success and dispersal patterns at restored
sites-experiments to help design/improve restoration strategies.
However, (the risks)
•Calculations and analyses indicate population bottlenecking possible by
deploying highly inbred selected lines (Hare and Rose)
•Little evidence to date that the selected lines are doing well and
reproducing. Are we wasting $? (Carlsson et al.--stay tuned)
•Evidence of resistance (tolerance) in natural populations (Carnegie and
Burreson), which are genetically more diverse and therefore risk can be
reduced by using wild broodstock.
Need Basic Genetic Data
Chesapeake Bay
What is the Crassostrea virginica population genetic
structure?
Ongoing studies-published and preliminary results
What is the effective population size in CB and how would
selectively bred stock impact this?
Matt Hare’s presentation on Thursday:high risk with current selected highly
inbred lines with low Ne.
What are the larval dispersal patterns around restored
reefs?
Ongoing studies-published and preliminary results
What is the genetic structure of the extant native oyster populations?
What historically was the genetic structure of the native oyster populations?
The BAYLOR SURVEY of OYSTER
GROUNDS
1892 survey of most productive oyster
grounds in Virginia (8 million bushels/year)
Chesapeake Bay Oysters
One panmictic population
OR
Isolated, genetically distinct
populations?
One population, which over time declined to an extent that there are
now individual populations that have become genetically isolated?
Retentive/trap-like estuaries with low gene flow among systems?
Microsatellites
+High power of discrimination for populations genetics and restoration
monitoring
+Highly variable
+High throughput
+Nuclear marker-biparentally inherited
Microsatellite- simple sequence repeats often varying lengths
among copies (alleles)
ATCTATATATATATATATATATATCGTGG
Chromosome (allele) from
TCGATATATATATATATATATATAGCACC
♀ (TA)10
ATCTATATATATATATATATCGTGG
Chromosome (allele) from
(TA)8
TCGATATATATATATATATAGCACC
♂
Evidence of Genetic Structure in the Bay using Microsatellite Markers
But Weak Structure
Buroker et al. 1983. Evidence of differentiation using allozyme markers
Rose, Paynter and Hare. 2006. J Hered. 97:158-170
Populations may be
genetically different.
There is evidence that
more distant populations
are more distinct.
Pairwise Comparisons of 10 Chesapeake Bay Populations
FST
2
3
4
5
6
7
8
9
10
1
0.00023
0.00010
0.00102
0.00129
0.00051
0.00070
0.00101
0.00018
-0.00055
2
3
4
5
6
7
8
9
10
1
0.4268
0.5576
0.1162
0.0762
0.3262
0.2168
0.0488
0.5557
0.9102
P
Carlsson et al.
2
3
4
5
6
7
8
9
0.00033
0.00167
0.00092
-0.00049
0.00050
0.00111
0.00005
-0.00057
0.00176
0.00033
-0.00010
0.00025
0.00121
0.00034
0.00027
0.00121
0.00124
0.00160
0.00062
0.00097
-0.00039
0.00223
0.00119
0.00171
0.00072
0.00089
-0.00012
0.00094
0.00052
-0.00050
-0.00061
0.00065
0.00075
0.00102
0.00121
0.00030
2
3
4
5
6
7
8
9
0.3916
0.0186
0.1846
0.8965
0.2793
0.0557
0.6826
0.8916
0.0586
0.4902
0.6260
0.4277
0.1162
0.5352
0.4482
0.2158
0.1621
0.0713
0.3574
0.3057
0.7666
0.0332
0.2022
0.0781
0.4453
0.2763
0.6641
0.1787
0.4854
0.8271
0.9326
0.3848
0.2382
0.2285
0.1445
0.5479
Is structure relevant?
Are populations locally adapted?
4 microsatellite markers
What happens to the oysters deployed on reefs?
Molecular markers to track deployed oysters.
•Do they reproduce?
•Genotype (genetically fingerprint) the spatfall.
•Are progeny purebred deployed or wild oysters?
AND/OR
•Hybrids?
•Do the deployed oysters survive? How long?
•Yearly genetic assessments of oysters at experimental
deployment sites.
•What impact do they have on surrounding populations?
•Screening populations-follow through time.
Wild stocks
Planted hatchery
stocks
Spat population:
Progeny from wild,
hatchery or hybrids? Are
they more or less fit than
wild?
Molecular markers can help us discriminate among stocks/lines
and allow us to learn more about the reef recruitment shadow
and the results of the inter-breeding of wild and hatchery
stocks.
Genetic analyses tracking the success of reef stocking
Objective: Monitoring the breeding success, and longer-term
relative survivability, of oysters planted on reefs
7
Experiment designed
for the Great
Wicomico River
system using the
genetically unique,
disease tolerant
aquaculture strains
(DEBYs).
6
5
4
3
2
1
Spat collected at sample sites every 2 weeks from June -October for
genetic typing in the years 2002-2006.
GWR has been seeded multiple times over the years with
several different stocks
YearYear Number
Number Stock
Stock
19961996 750000
Sound
750000Tangier
Tangier Sound
19971997 150000
Sound
150000Tangier
Tangier Sound
19981998 150000
150000Deep
Deep Rock
Rock
19981998
2500
Sound
2500Tangier
Tangier Sound
19991999
5000
Sound
5000Tangier
Tangier Sound
20002000
24750
24750CROSBreed
CROSBreed
20002000 210000
210000Deep
Deep Creek
Creek
20012001
10000
10000CROSBreed
CROSBreed
20012001 300000
300000Deep
Deep Creek
Creek
20012001 200000
Creek
200000Lynnhaven/Plantation
Lynnhaven/Plantation Creek
20022002
13500
13500CROSBreed
CROSBreed
20022002 795700
795700DEBY
DEBY
20032003 292060
292060DEBY
DEBY
20042004
18000
18000CROSBreed
CROSBreed
20042004 1400000
1400000DEBY
DEBY
20052005-06
15000000
15000000DEBY
DEBY
Since 2002 primarily DEBY deployments as part of the
experimental design to track success of planted oysters.
Why did we choose DEBYs for the GWR experiment?
DEBYs are genetically unique. Maternal signal-mtDNA.
DEBYs Show High Frequency of Mitochondrial Haplotypes (DNA
fingerprint patterns) that are Rare in Natural Chesapeake Bay Populations
B
A
Hinf I digest of mt coIII in DEBY strain
A
Hinf I Digest of mt coIII in a Natural Population
Frequency of the B alleles is relatively low in natural populations: <2%.
Frequency of the B alleles is much higher in the DEBY stock, generally
ranging from 25-50% depending on the spawn.
Microsatellite markers allow clear discrimination between
hatchery lines and natural populations
Rappahannock
wild – yellow
Deployed spaton-shell - blue
Example Rappahannock River, Drumming Ground
Have the deployed DEBYs contributed significantly to spat
production in GWR?
Mt DNA Analyses
PRIOR TO
DEPLOYMENT
DEPLOYED
DEBY
PRODUCED
SPAT
AA
BB
Rare
Carlsson et al. Great Wicomico 2002-2006
Mt DNA and microsatellite analyses
•1579 spat collected in the summer of 2002
•1 individual confidently assigned to DEBY
•~10% DEBY/WILD hybrids
Hare et al. 2006- form Great Wicomico River 2002
Overall, data to date suggest that the DEBY contribution has been low: predation, poor
survival and reproduction? Recently there have been much larger deployments with efforts
and protecting plants and genetic signal needs to be followed over several years.
Genetic Considerations
(Aquaculture)
Aquaculture Development
Which oyster stocks to use? Diploids or triploids?
Special genetic lines might be selected for particular
traits of interest.
Maintain genetically healthy lines.
Is there any genetic impact on extant natural
populations?
Genetic impact of aquaculture lines on natural
populations is a concern in many aquatic systems. Eg.
Salmonids
But
Is this a concern for aquaculture development in oysters?
Little evidence of genetic impact to date
Analysis of oysters collected near two farms growing DEBYs
Site 1
4 microsatellites
2 mtDNA genes
Over 85% significantly not assigned to DEBY
1 individual assigned to DEBY
Site 2
4 microsatellites
2 mtDNA genes
Over 90% significantly not assigned to DEBY
No individuals assigned to DEBY
1 DEBY (natural collection)
There is evidence of reduced genetic variation in
hatchery lines of C. virginica
Allelic diversity of microsatellites reduced in DEBYS
compared to natural populations
Natural population
DEBY strain
Genetic Considerations
(Introduction)
Introduction of a Non-native Oyster
Aquaculture or on bottom fishery?
Which species? Genetic identification needed.
Which stock? Broodstock source?
Oregon strain too genetically bottlenecked?
1995 Virginia House of Delegates Resolution no. 450
“Requesting the Virginia Institute of Marine Science to develop a
strategic plan for molluscan shellfish research and begin the
process of seeking the necessary approvals for in water testing of
non-native oyster species.”
ICES Protocols
The International Council for the Exploration of the Seas (ICES) Code of
Practice on Introductions and Transfers of Marine Organisms (ICES,
1994):
“…prior to any introduction a detailed analysis should be conducted on
the ecological, genetic and disease relationships of the species in its
natural range and environment.”
EIS Currently Being Drafted
Genetic Analyses of Crassostrea ariakensis
Objectives:
• Inventory of germplasm resources in the species, Crassostrea ariakensis- Correct
identification of the species became a large concern.
• To examine genetic variation and differentiation (population structure), among
natural populations of the C. ariakensis from China, Korea and Japan
• To examine genetic variability.
• In US hatchery stocks (Oregon Strain)
• Compared to wild source populations
Jan Cordes and Jie Xiao
Ximing Guo’s group-Rutgers
There is Genetic Variation among Wild C. ariakensis Populations
linkage disequilibrium, and significant
deviations from Hardy-Weinberg
Equilibrium (HWE)
Sample
LD
HW E
IR
0 of 6
none
KR
1 of 6
none
YR
3 of 6
none
DR
0 of 6
none
IR
KR
YR
DR
IR
-
0.022
0.014
0.026
KR
<0.001
-
0.01*
0.014
YR
<0.001
0.007*
-
0.025
DR
<0.001
<0.001
<0.001
-
Population pairwise Fst (above) and P-values
(below). * indicates Non-significant values.
Genetic Variation among Wild Populations
Factorial Correspondence Analysis
(FCA) by Individuals
KR
YR
DR
IR
US Hatchery Stocks
“Oregon Strain”
Japan
F1
WC
Pacific
Northwest,
USA
WCA
P1
TUI
Yellow
F1
NCA
River
China
Beihai
F1
SCA99
+
SCA00
Genetic Variation in Hatchery Stocks vs. Wild Populations
Factorial Correspondence Analysis
(FCA) by Population
KR
IR
YR
DR
TUI
WCA
SCA
NCA
•Hatchery Stocks show reduction in genetic diversity compared
to wild populations
•Oregon strain is relatively highly inbred
Wild Populations
Allelic richness for four hatchery strains
and four wild populations of C. ariakensis.
25
Hatchery Strains
Wild Stocks
CarG110
CarG4-60
Car119-6a
Car11-70
20
Sample
LD
HW E
IR
0 of 6
none
KR
1 of 6
none
YR
3 of 6
none
DR
0 of 6
none
Hatchery Stocks
15
Sample
LD
HW E
TUI
1 of 6
4
WCA
3 of 6
2
NCA
5 of 6
1
SCA
0 of 6
1
10
5
0
TUI
WCA
NCA
SCA
IR
KR
YR
DR
Acknowledgements
Elizabeth Francis
Stan Allen
Francis O’Beirn
Georgeta Constantin
Roger Mann
Tommy Leggett
Jie Xiao
Missy Southworth
Ryan Carnegie
Qian Zhang
Juli Harding
Mark Luckenbach
Gail Scott
Aimim Wang
Ken Paynter
Cheryl Morrison
Dr. Wu
Matt Hare
Pat Gaffney
Dr. Ahn
Don Merritt
Sharon Furriness
Junya Higano
Wendi Ribeiro
US National Sea Grant-ODRP
NOAA/NMFS Chesapeake Bay Program Office
Virginia Sea Grant College Program
Chesapeake Bay Foundation
US Army Corps of Engineers
JAC ARSs
Jan F.A. Cordes
Jens A. Carlsson
Research Assistant Scientists