44th Scientific Symposium of the UJNR
Aquaculture Panel
Genetics and Breeding in Aquaculture
NOAA Northwest Fisheries Science Center
2725 Montlake Blvd. East
Seattle, WA
November 1st and 2nd, 2016

Photo credit: Mark Drawbridge/Hubbs-SeaWorld Research Institute44th Scientific Symposium of the UJNR Aquaculture Panel
Genetics and Breeding in Aquaculture
November 1st and 2nd, 2016
NOAA Northwest Fisheries Science Center
2725 Montlake Blvd. East
Seattle, WA
Aim of the Symposium
Exciting new developments in genetics and genomics contribute significantly to advances inaquaculture production today and will be of even greater importance in the future. Genetic
improvements through selective breeding, genetics and health management, understanding
genetic interactions of wild and cultured stocks and genetics and climate change are all research
priorities of the Japanese Fisheries Research Agency, the National Oceanic and Atmospheric
Administration and the United States Department of Agriculture. The primary focus of this
symposium will be on solving production problems faced by the aquaculture industries of thetwo nations using genetic and genomic approaches. This will facilitate development of morecompetitive new and existing aquaculture industries. Selective breeding has a long history of
improving production traits in many livestock species and is having progressively greater
impacts in aquaculture; enhancing genetic improvement through application of genomics cansignificantly accelerate this process. In health management genetic approaches to improve
resistance, enhance immune response, better understand pathogens and improve vaccines are alltools to strengthen the aquaculture industry. Climate change will present many challenges toaquacultured species and genetics and genomics will prove to be valuable tools in addressingthese. We hope to encourage broad ranging discussions of some of the new approaches being
developed and how they might be applied in collaborative research efforts to resolve key
bottlenecks and facilitate aquaculture industry expansion in Japan and the United States.
Program
Tuesday, November 1, 2016
Registration

13:00 - 13:30

Opening Session and Orientation to the Northwest Fisheries Science Center
WelcomeJohn Stein, Director, NOAA Northwest Fisheries Science Center

13:30 - 13:40

Aim of the Symposium
Michael Rust, US Panel Chair, NOAA Fisheries Office of Aquaculture

13:40-13:50

Ecosystem Approach to Aquaculture(Moderators: Paul Olin and Takuro Shibuno)
Integrated Multi-Trophic Aquaculture (IMTA) as a countermeasure for coastal
oligotrophication
Satoshi Watanabe, National Research Institute of Aquaculture
Fisheries Research and Education Agency
13:50-14:15
Interactions between shellfish aquaculture and the environment in the Northeastern US
Matt Poach, NOAA Northeast Fisheries Science Center
14:15-14:40
Land-based aquaculture for red-spotted grouper
Masatsugu Takano, Fisheries Research and Education Agency Headquarters

14:40-15:05

Using physiological tools to assess and optimize aquaculture of California Yellowtail,
Seriola dorsalis
Nick Wegner, NOAA Southwest Fisheries Science Center
15:05-15:30
Wednesday, November 2, 2016
Breeding Part 1(Moderators: Andrew Severin and Masatsugu Takano)Paradigm shift in fish breeding; marker-assisted selection to genomic selection
Akiyuki Ozaki, National Research Institute of Aquaculture,
Fisheries Research and Education Agency
9:00-9:25
Genomic analyses to inform sablefish aquaculture research
Krista Nichols, NOAA Northwest Fisheries Science Center

9:25-9:50

Mutagenesis and genome editing for aquaculture fish species
Hiroyuki Okamoto, National Research Institute of Aquaculture,
Fisheries Research and Education Agency

9:50-10:15

Break

10:15-10:45
Breeding Part 2(Moderators: Caird Rexroad and Satoshi Watanabe)

Genome-enabled selection doubles the accuracy of predicted breeding values for bacterial
cold water disease resistance compared to traditional family-based selection in rainbow
trout aquaculture
Roger Vallejo, US Department of Agriculture, Agriculture Research Service
10:45-11:10

Genetic improvement of Atlantic salmon (Salmo salar L.) and Eastern oyster (Crassostrea
virginica Gmelin 1791) at the USDA-ARS National Cold Water Marine Aquaculture
CenterGary Burr, US Department of Agriculture, Agriculture Research Service
11:10-11:35
The Molluscan Broodstock Program: twenty years of selective breeding of the Pacificoyster on the West Coast of the US
Romain Morvezen, Oregon State University
11:35-12:00
Lunch Break

12:00-13:00
Genomics and Epigenetics, Part 1(Moderators: Krista Nichols and Hiroyuki Okamoto)

Seriola genomics and the knowledge repository Serioladb.org
Andrew Severin, Iowa State University

13:00-13:25

The first meeting of Seriola genomics and breeding consortium
Akiyuki Ozaki, National Research Institute of Aquaculture,
Fisheries Research and Education Agency

13:25-13:50

Uses of genetic parentage analysis in cultured California Yellowtail (Seriola dorsalis)
Matthew Craig, NOAA Southwest Fisheries Science Center
13:50-14:15
Break

14:15-14:45
Genomics and Epigenetics, Part 2(Moderators: Roger Vallejo and Akiyuki Ozaki)

Past, present and future research on the Ostreid herpes virus - 1 infections in the Pacificoyster
Caroline Friedman, University of Washington
14:45-15:10
Type 1 Ostreid Herpesvirus (OsHV-1) variants in Japan
Satoshi Watanabe, National Research Institute of Aquaculture,
Fisheries Research and Education Agency

15:10-15:35

Characterization of genetic and epigenetic variation in hatchery and natural-origin
steelhead, Oncorhynchus mykiss
Mackenzie Gavery, University of Washington
15:35-16:00
Towards a functional understanding of DNA methylation in shellfish and implications foraquaculture
Steve Roberts, University of Washington
16:00-16:25
Open Discussion: Development and support of research collaborations
Moderators: Mike Rust and Fuminari Ito

16:25-16:50

Science Symposium Closing
Fuminari Ito, Japan Panel Chair, Fisheries Research and Education Agency
Symposium Reception: One on one discussions continue
Ivar’s Salmon House (see below)

16:50-17:00
17:30-19:00

List of ParticipantsBurr, Gary
Craig, Matthew
Dumbauld, BrettFriedman, Carolyn
Gavery, Mackenzie
Ito, Fuminari
Morvezen, Romain
Nichols, Krista
Okamoto, Hiroyuki
Okumura, Takuji
Olin, Paul
Otoshi, Clete
Ozaki, Akiyuki
Poach, Matt
Rexroad, Caird
Roberts, Steve
Rust, Mike
Severin, Andrew
Shibuno, Takuro
Takano, Masatsugu
Vallejo, Roger
Watanabe, Satoshi
Wegner, Nick

USDA Agricultural Research Service (ARS)NOAA Southwest Fisheries Science Center (SWFSC)
ARS
University of Washington (UW)UW
Japan Panel Chair, Japan Fisheries Research and Education
Agency, National Research Institute of Aquaculture (FRA)
Oregon State University
NOAA Northwest Fisheries Science Center
FRA
FRA
California Sea Grant
NOAA Fisheries Office of Aquaculture (AQC)
FRA
NOAA Northeast Fisheries Science Center
ARSUW
US Panel Chair, AQC
Iowa State University
FRA
FRA, Headquarters
ARS
FRA
SWFSC

44th Scientific Symposium of the UJNR Aquaculture Panel

Genetics and Breeding in Aquaculture
Symposium Abstracts
1. Integrated multi-trophic Aquaculture (IMTA) as a countermeasure for coastal
oligotrophication
Satoshi Watanabe*1, Natsuki Hasegawa1, Yuka Ishihi1, Tomomi Mizuno2 and Junya Higano3
Presenting author*
1

National Research Institute of Aquaculture, Japan Fisheries Research and Education Agency,
422-1 Nakatsuhamaura, Minamiise, Mie, 516-0193, Japan, *E-mail: swat at affrc.go.jp
2
Mie Prefecture Fisheries Research Institute, 3564-3 Hamajima, Shima, Mie, 517-0404, Japan,
3
National Research Institute of Fisheries and Environment of Inland Sea, Japan Fisheries
Research and Education Agency, 2-17-5 Maruishi, Hatsukaichi, Hiroshima 739-0452, Japan
Abstract
Intensification of aquaculture production has caused environmental deterioration associated withwater and sediment eutrophication and consequent harmful algal bloom, hypoxia and sulfide
production, resulting in sporadic mass fish kills in some parts of the world. In contrast,
eutrophication has been subsiding in coastal waters of Japan due to reduced production andimproved feeding efficiency of finfish aquaculture and/or enforcement of the total pollutant load
control system (TPLCS, issued by the Ministry of the Environment in 1979) that regulates theallowable amount of terrestrial nitrogen/phosphorus discharge to the sea. Red seabream, Pagrus
major, aquaculture production, for instance, peaked in the early 1990’s and has been in adecreasing trend since 2000 in Japan. The levels of dissolved inorganic nitrogen (DIN, i.e. sumof NH4+, NO2-, NO3-) have gradually decreased, while phosphorus levels have been rather
constant in Ise Bay since the 1990’s, possibly causing a nutrient imbalance for primary
production. Surface chlorophyll a levels have been in a decreasing trend since the 1990’s, andthe occurrence frequency of red tide has decreased by 90% since 1979 in Ise Bay. The
oligotrophication (relative to the past) of coastal waters is considered to have brought about adecline in carrying capacity of the coastal environment, causing, in part, a continuous decrease insome coastal fishery resources and reduced productivity in molluscan and algal aquaculture. The
national production of the Manila clam, Ruditapes philippinarum, for example, has continuously
declined for thirty years, falling below 10% of the peak value marked in the mid 1980’s. In
Ariake Bay, DIN deficiency causes bleaching of cultured nori, Pyropia yezoensis, severely
reducing the market value especially towards the end of the culture season in early spring.
Integrated multi-trophic aquaculture (IMTA) is an aquaculture approach that combines culture of
economically important species from different trophic levels, typically finfish, organic extractive
species (e.g. bivalve) and inorganic extractive species (e.g. seaweed). Along with reinforcementof economic stability, one of the important goals of IMTA is to mitigate effluent load from

finfish culture for environmental integrity. In this study, however, we are trying to develop
techniques to use IMTA to compensate for the reduced aquaculture productivity of bivalves andseaweeds associated with the coastal oligotrophication. We found that chlorophyll a level washigh in the areas affected by red seabream aquaculture in the semi-closed Hasamaura Cove in
Mie Prefecture. Cultured green alga, Monostroma nitidum (Japanese common name Hitoegusa),
was found to have twice as high a nitrogen content in the cove than in those cultured in nearby
areas without finfish aquaculture. The amount of NH4+-N excretion from the red seabream
culture in the cove was estimated to be 7.5 t/yr. This is equivalent to the amount of N in 1872 t/yr.
(wet weight) of Manila clam suspended culture production. The amount of N extracted by M.
nitidum culture in the cove (264 kg/yr.) is equal to 3.3% of the fish NH4+-N. Thus, IMTA has a
big potential to enhance aquaculture production by using what would otherwise be fish waste.
Annotated Bibliography of Key Works
Chopin, T., Yarish C., Wilkes, R., Belyea, E., Lu, S. and Mathieson, A. 1999. Developing
Porphyra/salmon integrated aquaculture for bioremediation and diversification of the aquacultureindustry. J. Appl. Phycol. 11: 463–472.
This is one of the first papers on IMTA (referred to as integrated culture) combining salmon cage
culture and Porphyra (currently Pyropia) species culture to alleviate the seasonal nutrient
depletion by using the significant loading of salmon farms, which is then valued (wastes become
fertilizers) and managed (competition for nutrients between desirable algal crops and problem
species associated with severe disturbances). The development of integrated aquaculture systems
is a positive initiative for optimizing the efficiency of aquaculture operations, while maintaining
the health of coastal waters.
Watanabe S., Kodama, M. and Fukuda, M. 2009. Nitrogen stable isotope ratio in the manila clam,
Ruditapes philippinarum, reflects eutrophication levels in tidal flats. Mar. Poll. Bull. 58: 1447–
1453.
The authors revealed that the nitrogen stable isotope ratio (15N) in the soft tissues of the manila
clam, Ruditapes philippinarum, could be used as an indicator of anthropogenic eutrophicationlevels in tidal flat environments. In addition, they found that the acid insoluble fraction of theshell organic matrix, conchiolin, could be used as a proxy for the soft tissues in 15N analyses,
which will result in easier storage handling and the expansion of chances for sample acquisition.
Understanding the effects of anthropogenic eutrophication on coastal fisheries may help in theenhancement of fishery production by effective utilization of sewage effluents, as well as in theconsequent reduction of eutrophication.
2. Interactions between shellfish aquaculture and the environment in the Northeastern US
Matthew Poach1*, Bob Alix2, Paul Clark2, April Croxton2, Mark Dixon2, Cathy Kuropat2, Judy
Li2, Renee Mercaldo-Allen2, Shannon Meseck2, Lisa Milke2, Dylan Redman2, Julie Rose2, Barry
Smith2, Gary Wikfors2.
1

US Department of Commerce, National Oceanic and Atmospheric Administration, National
Marine Fisheries Service, James J. Howard Marine Sciences Laboratory, Highlands, NJ 07732,
USA.

2

US Department of Commerce, National Oceanic and Atmospheric Administration, National
Marine Fisheries Service, Milford Laboratory, Milford, CT 06460, USA.
Abstract
Opposition to new shellfish aquaculture operations in the Northeastern US frequently is based
upon the premise that these commercial operations have negative environmental consequences.
This perception is not consistent with the myriad of ecosystem services that are attributed toshellfish, including improving water clarity, reducing excess nutrients, providing habitat,stabilizing sediments, and serving as a food for wildlife and humans. Researchers at the Milford
Laboratory have worked to quantify the complex interactions between shellfish aquaculture andthe natural environment across the Northeastern United States region and beyond.
From 2008-2010 experiments were conducted in East Creek, Long Island, New York to quantify
the environmental footprint of a floating upweller system (FLUPSY) within the surrounding
embayment. Metrics such as dissolved nutrients, phytoplankton abundance and community
composition, chlorophyll a, temperature, salinity, and dissolved oxygen were measured. Resultsof the multi-year study indicate that the FLUPSY had a very small role in ecosystem function
compared with the range of natural environmental variation within the East Creek System. From2009 to 2013 studies were conducted to measure the consequences of hydraulic dredging forclam harvest upon sediment chemistry and the benthic community in coastal Milford
Connecticut. Milford Laboratory data indicated that hydraulic dredging caused initial, shortlived changes in sediment chemistry that resolved within a few weeks. Disturbance of thebenthic community was minimal, and the community that inhabited this nearshore zone wasnaturally resilient to a dynamic physical environment.
Milford research also has quantified the environmental benefits of shellfish aquaculture. Startingin 2011, the biodeposition method was employed to measure nitrogen removal and water clarity
improvements by shellfish throughout New England and on the west coast of the Korean
peninsula. US study locations have included NY, CT, RI and MA and species included ribbed
mussels, blue mussels, clams and oysters. Research indicated that shellfish are able to adapt to awide range of environmental conditions, but careful site selection is needed to maximize
environmental benefits of aquaculture operations. Ultimately this work demonstrates thatshellfish aquaculture can have a positive role in eutrophic ecosystems. We plan to continue ourwork on interactions between aquaculture and the environment by expanding into multi-trophic
and off-shore aquaculture systems in New England.
Annotated Bibliography of Key Works
Li, Y., Meseck, S. L., Dixon, M. S., Rivara, K., and Wikfors, G. H. (2012). Temporal
Variability in Phytoplankton Removal by a Commercial, Suspended Eastern Oyster Nursery andEffects on Local Plankton Dynamics. Journal of Shellfish Research, 31(4): 1077-1089.
Meseck, S.L., Li, Y., Dixon, M.S., Rivara, K., Wikfors, G.H. and Luther, G.I. (2012). Effects ofa Commercial, Suspended Eastern Oyster Nursery upon Nutrient and Sediment Chemistry in a
Temperate, Coastal Embayment. Aquaculture and Environment Interactions 3: 65-79.

Li, Y., Meseck, S., Dixon, M., Rose, J., Smith, B., and Wikfors, G. (2013). Short Term Effects
of a Commercial Eastern Oyster Nursery upon Nutrient and Plankton Dynamics of a Coastal
Embayment: Observations from Mesocosm Experiments. Aquaculture Research, In press.
Mercaldo-Allen, R., Goldberg, R., Clark, P., Kuropat, C., Meseck, S.L., Rose, J. M.
2016. Benthic Ecology of Northern Quahog Beds with Different Hydraulic Dredging Histories
in Long Island Sound. Journal of Coastal Research 32(2):408-415.
Meseck, S.L., Mercaldo-Allen, R., Rose, J.M., Clark, P., Kuropat, C., Pereira, J.J., Goldberg, R.
2014. Impacts of Hydraulic Dredging for Mercenaria mercenaria, Northern Quahog, on
Sediment Biogeochemistry. Journal World Aquaculture Society 45(3): 301-311.
Goldberg, R., Rose, J.M., Mercaldo-Allen, R., Meseck, S.L., Clark, P., Kuropat, C., and Pereira,
J.J. 2014. Effects of hydraulic dredging on the benthic ecology and sediment chemistry on acultivated bed of the hard clam, Mercenaria mercenaria. Aquaculture 428–429: 150–157
Goldberg, R., R. Mercaldo-Allen, J. M. Rose, P. Clark, C. Kuropat, S. Meseck, and J. Pereira.
2012. Effects of hydraulic shellfish dredging on the ecology of a cultivated clam bed.
Aquaculture Environment Interactions 3:11-21.
Galimany, E., Rose, J.M., Dixon, M.S., Wikfors, G.H., 2013. Quantifying Feeding Behavior of
Ribbed Mussels (Geukensia demissa) in Two Urban Sites (Long Island Sound, USA) withDifferent Seston Characteristics. Estuaries and Coasts, 1-9.
Rose, J.M., Bricker, S.B., Tedesco, M.A., Wikfors, G.H., 2014. A Role for Shellfish Aquaculturein Coastal Nitrogen Management. Environmental Science & Technology 48, 2519-2525.
Rose, J.M., Bricker, S.B., Ferreira, J.G., 2015. Comparative analysis of modeled nitrogen
removal by shellfish farms. Marine Pollution Bulletin 91, 185-190.
3. Land-based aquaculture of red-spotted grouper（Epinephelus akaara）using the closed
recirculation system
Masatsugu Takano1*, Tetsuo Morita2, Tadashi Imai2 and Yoshihisa Yamamoto2
1

Headquarters, Japan Fisheries Research and Education Agency, Minato-mirai 2-3-3, Queen
Tower B 15F, Nakaku, Yokohama, Kanagawa 221-6115, Japan
2
National Research Institute of Fisheries and Environment of Inland Sea, Japan Fisheries
Research and Education Agency, Yashima-higashimathi 234, Takamatsu, Kagawa 761-0111,
Japan
Abstract
The red-spotted grouper, Epinephelus akaara, is a commercially expensive fish in Japan. This
species is expected to be a new target fish for aquaculture. However, its culture is thought to bedifficult because of its slow growth (four or five years until the fish are of commercial size).

To achieve faster growth and higher feed efficiency for the red-spotted grouper, we
experimented with different water temperatures using a closed recirculation system. In higher
water temperatures between 16 and 31 degrees Celsius, growth was faster; best feed efficiency
was obtained at 25 degrees Celsius. We carried out similar experiments with salinity. These
results reveal that it is possible to rear the red-spotted grouper to commercial size inapproximately two years.
The more traditional flow-through system is expensive to maintain in optimum condition. The
closed recirculation system solves this problem by reusing drainage. However, it needs toremove harmful ammonia, suspended materials and the residual material. We have improved theefficiency of two filter devices (the biological filter and the bubble separation device); thesedevices were built into the system. The closed recirculation system is expected to assist the
expansion of land-based aquaculture and find applications in breeding research.
Annotated Bibliography of Key Works
Yamamoto, Y. and Hayase, S., 2008. Japan, Thematic regional reviews, In The future ofmariculture: a regional approach forresponsible development in the Asia-Pacific region. FAO Fisheries Proceedings, 11, 189-198.
Yamamoto, Y. and Masaaki Kamoshida and Toshio Takeuchi, 2013. Examination of Suitable
Condition of Recirculation Rate in a Closed Recirculating System for Larviculture of Red Sea
Bream, Pagrus major. Eco-Engineering, 25(2), 49-58.
Tetuso Morita, 2014. Potential for circulating land farming business of red spotted grouper
Epinephelus akaara. Aqua Culture Business, Volume: 51, Issue: 13, 14-16.
Tomohide Nambu, 2014. II-7. Seed production and aquaculture technology of red spottedgrouper Epinephelus akaara. Nippon Suisan Gakkaishi, 80(6), 1000.
4. Using physiological tools to assess and optimize aquaculture of California Yellowtail,
Seriola dorsalis
Nicholas Wegner*1, Laura Schwebel2, and John Hyde3
1

Southwest Fisheries Science Center, NOAA Fisheries, 8901 La Jolla Shores Dr., La Jolla, CA
92037, nick.wegner at noaa.gov
2Southwest Fisheries Science Center, NOAA Fisheries, 8901 La Jolla Shores Dr., La Jolla, CA
92037, lschwebel at sandiego.edu
3John Hyde, Ph.D., Southwest Fisheries Science Center, NOAA Fisheries, 8901 La Jolla ShoresDr., La Jolla, CA 92037, john.hyde at @noaa.gov
Abstract:
The California Yellowtail (Seriola dorsalis) is a promising candidate for offshore commercial
aquaculture in Southern California. However, best rearing practices for this and other Seriolaspecies are still under development, and our work has shown that yellowtail spawned and rearedin captivity are typically less healthy than their wild-caught counterparts. Our research thus
seeks to use various physiological tools and metrics to monitor fish health, fitness, and growth in
an attempt to enhance rearing techniques. Over the past few years our work has focused on 1:
Examining fitness differences between wild-caught and aquaculture-reared individuals throughmeasures of critical swimming speed and metabolic rate, 2. Determining the adverse effects of an
extremely common deformity (an uninflated gas bladder) in hatchery-reared yellowtail on theircritical swimming speed, metabolic rate, growth rate, and feed conversion ratio, and 3.
Investigating the potential growth and health benefits of forced exercise on early juvenile-stage
yellowtail. Our results suggest that although wild-caught fish tend to show better health andmetabolic efficiency, great strides in yellowtail health and fitness in aquaculture have occurred
during the duration of our research, and eliminating deformities such as uninflated gas bladders
greatly increases growout efficiency. While our swimming exercise studies are still underway,
preliminary data suggest that a short duration of exercise (2-4 weeks) followed by growout instandard rearing tanks produces a sustained growth advantage. Such exercise regimes may thus
represent not only a mechanism for improving fish health and fitness, but also a non-genetic andnon-hormonal treatment to accelerate growth of fish for food or wild stock enhancement.
Annotated Bibliography of Key Works
Brown, E.J., M. Bruce, S. Pether, and N.A. Herbert (2011) Do swimming fish always grow fast?
Investigating the magnitude and physiological basis of exercise-induced growth in juvenile New
Zealand yellowtail kingfish, Seriola lalandi. Fish Physiol. Biochem., 37: 327-336.
This study investigated the effect of swimming exercise on the growth of cultured New Zealandyellowtail kingfish, Seriola lalandi at different water temperatures and swimming speeds. Two
growth trials exposed fish to different exercise levels (determined by current speed) at 14.9° and
21.1°C. Lengths and weights were measured for each fish before and post exercise, along withmetabolic rates using a respirometer. Results of these trials showed that exercise yielded a 10%
increase in growth but only at low swimming speeds (0.75 BL s-1) and at a temperature of 21.1°C.
Experiments using a swim tunnel respirometer indicated that exercise training had no effect on
metabolic scope or critical swimming speeds but it did improve swimming efficiency.
Palstra, A. P., et al. (2015). Forced sustained swimming exercise at optimal speed enhances
growth of juvenile yellowtail kingfish (Seriola lalandi). Frontiers in Physiology 5: 506.
The swimming exercise trials in this study for Seriola lalandi resulted in 46% greater increases
in mass and 92% larger increases in length compared to non-exercised controls. This study thus
showed greater exercise benefits than those observed in the Brown et al (2012) study (above),
and these differences were likely related to using more optimal exercise swimming speeds. The
exercise and control groups in this study were fed equivalent rations, thus indicating theexercised fish had more efficient resource allocation, resulting in a lower feed conversion ratio(1.21 vs 1.74 for non-exercised yellowtail). These researchers thus suggest that growth rate canbe greatly enhanced without increased feeding.
Palstra, A.P. and J.V. Planas (eds). (2013) Swimming Physiology of Fish: Towards Using
Exercise to Farm a Fit Fish in Sustainable Aquaculture. Springer-Verlag: Berlin.
This multi-chaptered volume reviews the relevant advances in understanding the physiology offish swimming and examines topics ranging from fish biomechanics and migrations to thepotential benefits of swimming exercise in aquaculture resulting in changes in fish growth andflesh quality. It brings together both field and laboratory studies to better understand thephysiology of fish swimming with the intent to apply such findings to enhance aquaculture
practices. A chapter by Davison and Herbert summarizes research activities and advances onswimming-enhanced growth.
5. Paradigm shift in fish breeding; marker-assisted selection to genomic selection
Akiyuki Ozaki1*, Kazuo Araki1, Jun-ya Aoki1, Yukinori Shimada1, Hiroyuki Okamoto1,
Hironori Usuki 1, Koichi Okuzawa1, Kazunori Yoshida2, Tsutomu Noda2
1

National Research Institute of Aquaculture, Fisheries Research Agency, 422-1, Nakatsuhamaura,
Minamiise-cho, Watarai-gun, Mie, 516-0193, Japan. *E-mail: aozaki at affrc.go.jp
2
Seikai National Fisheries Research Institute, Fisheries Research Agency, 122-7, Nunoura,
Tamanoura-machi, Goto-shi, Nagasaki, 853-0508, Japan
Abstract:
The essential conditions for DNA marker-assisted selection is development of useful resource
families to evaluate phenotypes and information about genetic linkages and a large number ofpolymorphic genetic markers. Some of the cases have already reached a practical stage, and havebeen used as genetic improvement productions. This research and development are being applied
to other kinds of aquaculture fish. High density SNP arrays have become the tool of choice for
QTL mapping, Genome-wide association studies, marker-assisted selection (MAS). Earlier
mapping studies have identified QTL for important commercial traits including disease
resistance, and combining the resources of a high density genetic map with genome sequence
data will facilitate the fine mapping of these loci. Otherwise traditional MAS did not result in awidespread use of DNA information in animal breeding. The main reason was that the traits ofinterest in livestock production were much more complex than expected: they were determinedby thousands of genes with small effects on phenotype. These effects were usually too small to
be statistically significant and therefore were ignored. Genomic selection (GS) assumes that all
markers might be linked to a gene affecting the trait and concentrates on estimating their effectrather than testing its significance. There are technological breakthroughs that have resulted in
the current wide-spread use of DNA information in animal breeding: the development of the
genomic selection technology, the discovery of massive numbers of genetic markers (SNPs), andhigh-throughput technology to genotype animals for thousands of SNPs in a cost-effective
manner.
In aquaculture species, these very stringent tests result in only the largest QTL being found. For
disease resistance traits, such large QTL were detected, e.g., Japanese flounder resistant tolymphocystis disease, Atlantic salmon resistant to infectious pancreatic necrosis (IPN),
yellowtail resistant to Benedenia disease. Those large QTL results were limited to disease
resistance. Growth and diet fish meal feed resistant traits might be difficult to find the largest
QTL, and difficult to apply for MAS. But GS have a possibility to improve such traits which are
dominated by polygenic QTL. It is anticipated that future genetic breeding application will shift
to GS from MAS in aquaculture.

Keywords: marker-assisted selection (MAS); Genomic selection (GS); polygenic QTL.
Annotated Bibliography of Key Works
Fuji, K., Hasegawa, O., Honda, K., Kumasaka, K., Sakamoto, T., Okamoto, N., (2007) Markerassisted breeding of a lymphocystis disease-resistant Japanese flounder (Paralichthys olivaceus).
Aquaculture 272, 291-295.
An allele of a microsatellite, Poli9-8TUF, has a dominant effect at a single major locus and is
responsible for resistance to lymphocystis disease (LD-R) in Japanese flounder. We developed a
new population of Japanese flounder produced by marker assisted breeding using this allele. A
female that originated from the KP-B inbred line with LD-R that was homozygous for thefavorable allele (B-favorable) and a male from a commercial stock bred for higher growth rate
and good body shape were selected as parents. A female was selected as the LD-R-bearing
parent because the recombination rate of females is lower in the region where the LD-R locus islocated. As expected, the B-favorable allele was transmitted as a heterozygote to the progeny(LD-R+ population). The LD-R+ population, when tested at two commercial fish farms that had
LD outbreaks, showed no incidence of LD at either farm, while a control population without Bfavorable alleles (LD-R−) had incidences of 4.5% and 6.3% at the two farms. These results showthat marker-assisted breeding using molecular markers linked to an economically important trait
is an efficient strategy for breeding.
Moen, T., Baranski, M., Sonesson, A.K., Kjøglum, S., (2009) Confirmation and fine-mapping ofa major QTL for resistance to infectious pancreatic necrosis in Atlantic salmon (Salmo salar):population-level associations between markers and trait. BMC Genomics 10:368.
Infectious pancreatic necrosis (IPN) is one of the most prevalent and economically devastating
diseases in Atlantic salmon (Salmo salar) farming worldwide. The disease causes large
mortalities at both the fry- and post-smolt stages. Family selection for increased IPN resistance isperformed through the use of controlled challenge tests, where survival rates of sib-groups arerecorded. However, since challenge-tested animals cannot be used as breeding candidates, withinfamily selection is not performed and only half of the genetic variation for IPN resistance isbeing exploited. DNA markers linked to quantitative trait loci (QTL) affecting IPN resistancewould therefore be a powerful selection tool. The aim of this study was to identify and fine-map
QTL for IPN-resistance in Atlantic salmon, for use in marker-assisted selection to increase therate of genetic improvement for this trait. The QTL confirmed in this study represents a case of amajor gene explaining the bulk of genetic variation for a presumed complex trait. QTL
genotypes were deduced within most parents of the 2005 generation of a major breedingcompany, providing a solid framework for linkage-based MAS within the whole population insubsequent generations. Since haplotype-trait associations valid at the population level were
found, there is also a potential for MAS based on linkage disequilibrium (LD). However, in order
to use MAS across many generations without reassessment of linkage phases between markers
and the underlying polymorphism, the QTL needs to be positioned with even greater accuracy.
This will require higher marker densities than are currently available.
Akiyuki Ozaki, Kazunori Yoshida, Kanako Fuji, Satoshi Kubota, Wataru Kai, Jun-ya Aoki,
Yumi Kawabata, Junpei Suzuki, Kazuki Akita, Takashi Koyama, Masahiro Nakagawa, Takurou
Hotta, Tatsuo Tsuzaki, Nobuaki Okamoto, Kazuo Araki, Takashi Sakamoto. (2013) Quantitative

Trait Loci (QTL) Associated with Resistance to a Monogenean Parasite (Benedenia seriolae) in
Yellowtail (Seriola quinqueradiata) through Genome Wide Analysis. PLoS ONE 8(6): e64987.
Benedenia infections caused by the monogenean fluke ectoparasite Benedenia seriolae seriously
impact marine finfish aquaculture. Genetic variation in host has been inferred to play asignificant role in determining the susceptibility to this parasitic disease. To evaluate the genetic
basis of Benedenia disease resistance in yellowtail (Seriola quinqueradiata), a genome-wide andchromosome-wide linkage analyses were initiated using F1 yellowtail families (n = 90 per
family) based on a high density linkage map with 860 microsatellite and 142 single nucleotide
polymorphism (SNP) markers. Two major quantitative trait loci (QTL) regions on linkage
groups Squ2 (BDR-1) and Squ20 (BDR-2) were identified. These QTL regions explained 32.9–
35.5% of the phenotypic variance. On the other hand, the relationship between QTL forsusceptibility to B. seriolae and QTL for fish body size were investigated. The QTL related to
growth was found on another linkage group (Squ7). As a result, the authors present first genetic
evidence that contributes to detailing phenotypic resistance to Benedenia disease, and the results
will help resolve the mechanism of resistance to this important parasitic infection of yellowtail.
Meuwissen TH1, Hayes BJ, Goddard ME. (2001) Prediction of total genetic value using
genome-wide dense marker maps. Genetics;157(4):1819-1829.
Recent advances in molecular genetic techniques will make dense marker maps available andgenotyping many individuals for these markers feasible. Here we attempted to estimate theeffects of approximately 50,000 marker haplotypes simultaneously from a limited number ofphenotypic records. A genome of 1000 cM was simulated with a marker spacing of 1 cM. The
markers surrounding every 1-cM region were combined into marker haplotypes. Due to finite
population size N(e) = 100, the marker haplotypes were in linkage disequilibrium with the QTL
located between the markers. Using least squares, all haplotype effects could not be estimatedsimultaneously. When only the biggest effects were included, they were overestimated and theaccuracy of predicting genetic values of the offspring of the recorded animals was only 0.32.
Best linear unbiased prediction of haplotype effects assumed equal variances associated to each
1-cM chromosomal segment, which yielded an accuracy of 0.73, although this assumption was
far from true. Bayesian methods that assumed a prior distribution of the variance associated witheach chromosome segment increased this accuracy to 0.85, even when the prior was not correct.
It was concluded that selection on genetic values predicted from markers could substantially
increase the rate of genetic gain in animals and plants, especially if combined with reproductive
techniques to shorten the generation interval.
6. Genomic analyses to inform Sablefish aquaculture research
Krista M. Nichols1*, Andrew Jasonowicz2, Giles Goetz3, J. Adam Luckenbach3, Frederick W.
Goetz2
1

Conservation Biology Division, Northwest Fisheries Science Center, National Marine FisheriesService, NOAA, Seattle, WA. Email: krista.nichols at noaa.gov
2
Environmental and Fisheries Sciences Division, Northwest Fisheries Science Center, National
Marine Fisheries Service, Manchester Research Station, NOAA, Manchester, WA

3

Environmental and Fisheries Sciences Division, Northwest Fisheries Science Center, National
Marine Fisheries Service, NOAA, Seattle, WA
Abstract:
The marine fish species, Sablefish (Anoplopoma fimbria, black cod), are long-lived, migratory
species native to the west coast of the U.S. Sablefish is an ideal species for aquaculture becauseof their high market value and fast growth in culture. The development of the species foraquaculture will benefit from genomic technologies for a number of objectives, including: 1) an
understanding of the genetic stock structure in the wild, from which progenitor broodstock isdeveloped, 2) an understanding of the genetic basis of important production traits, and 3) the
development of comprehensive genotyping assays for screening sablefish in culture. We discussin this presentation our findings of genetic stock structure in wild populations, and future
directions in genome sequencing and genomics of quantitative traits. Using Restriction Site
Associated DNA sequencing (RADseq), we identified more than 100,000 SNPs in >400
individuals collected from a U.S. West Coast surveys from the Bering Sea in Alaska to southern
California. After filtering for genetic analyses, 2661 SNPs were used to assess population
structure and test for signatures of natural selection and association with environmental variables.
Our results show a lack of population structure and adaptive variation in Sablefish, and aresuggestive of a single panmictic population that is likely the result of a complex juvenile life
history and long range movements as adults. Our current studies now focus on understanding thegenetic basis of sexually dimorphic growth in Sablefish, and the development of a refined
genome assembly to provide a reference for genomic studies in this species.
Annotated Bibliography of Key Works
Jasonowicz, A.J., F.W. Goetz, Giles W. Goetz and K.M. Nichols. 2016. Love the one you’re
with: genomic evidence of panmixia in the sablefish (Anoplopoma fimbria). Canadian Journalof Fisheries and Aquatic Sciences (in press)
This study examines genetic stock structure in Sablefish along the U.S. West Coast and Alaska
using RADseq, and finds no evidence for population differentiation.
Luckenbach, J.A., W.T. Fairgreve. 2016. Gonadal sex differentiation and effects of dietary
methyltestosterone treatment in Sablefish (Anoplopoma fimbria). Fish Physiology andBiochemistry 42: 233-248.
An important characterization of sex differentiation in Sablefish, and identification of thedevelopmental time period sensitive to sex reversal by dietary testosterone.
Rondeau, E.B., A.M. Messmer, D.S. Sanderson, S.G. Jantzen, von Schalburg, K.R., Minkley,
D.R., Leong, J.S., Macdonald, G.M., Davidsen, A.E., Parker, W.A., Mazzola, R.S., Campbell B.
and B.F. Koop. 2013. Genomics of sablefish (Anoplopoma fimbria): expressed genes,
mitochondrial phylogeny, linkage map, and identification of a putative sex gene. BMC
Genomics 14: 452.
An important paper providing evidence for the sablefish sex determining gene, which will be
important in understanding genetic sex control in sablefish. This paper also provides the firstdraft genome sequence that we are currently improving with additional sequence data, andprovides the first comprehensive genomic resource for this fish species.

7. Mutagenesis and genome editing for aquaculture fish species
Hiroyuki Okamoto
National Research Institute of Aquaculture, Japan Fisheries Research and Education Agency,
422-1 Nakatsuhamaura, Minami-Ise, Mie, 516-0193, Japan. E-mail: hiokamot at affrc.go.jp
Abstract:
There is increasing interest in manipulating the function of target genes to achieve production
gains in agriculture and aquaculture. Although transgenic techniques are often used, there areseveral alternatives. Of these, the two primary methods are chemical mutagenesis, including
TILLING (Targeting Induced Local Lesion IN Genomes), and genome editing, also termed new
breeding technology (NBT). Chemical mutagenesis has been widely used in agriculture, butthere have been few reported successes in aquaculture. TILLING has been applied toexperimental fish such as zebrafish and medaka. In these instances, researchers have subjected
adult fish to a bath of strong alkylating agents (e.g., N-ethyl-N-nitrosourea (ENU)). Exposure to
the chemical mutagen induces single nucleotide substitutions in the genomes of all cells, butparticularly in the gametes. Because the body size of most intensively cultured food fish is muchlarger than these experimental species, it is not practical to use whole-body baths of ENU
solution. To address this, methods have been developed to induce mutagenesis by soaking eggs
or sperm in ENU solution or injecting ENU solution into the abdominal cavity of ripe males.
Whereas chemical mutagens induce genome-wide random mutations, artificial nuclease forgenome editing technology such as CRISPR/Cas can target mutations to a specific locus withrandom-length deletion or specific nucleotide sequence replacement. Genome editing ispreferred when genes or candidate genes that are associated with desirable traits have been
identified. In the absence of this information, or if there is a need to survey for alternative
phenotypes, chemical mutagenesis is the preferred method. To date, few candidate genes havebeen identified for commercially important traits such as growth, disease resistance, and foodconversion efficiency. This is because such traits are controlled by complex gene networks, witheach gene having a very small effect individually. Thus, it is expected that improvements in our
understanding of gene functions related to commercial traits could be made by further
investment in mutagenesis and gene editing research.
Keywords: mutagenesis, TILLING, genome editing, NBT, CRISPR/Cas
Annotated Bibliography of Key Works
Satoshi Ansai, Masato Kinoshita (2014) Targeted mutagenesis using CRISPR/Cas system inmedaka, Biology Open , 3: 362-371; doi:10.1242/bio.20148177
Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas)system-based RNA-guided endonuclease (RGEN) has recently emerged as a simple and efficient
tool for targeted genome editing. In this study, we showed successful targeted mutagenesis using
RGENs in medaka, Oryzias latipes. Somatic and heritable mutations were induced with highefficiency at the targeted genomic sequence on the DJ-1 gene in embryos that had been injectedwith the single guide RNA (sgRNA) transcribed by a T7 promoter and capped RNA encoding a
Cas9 nuclease. The sgRNAs that were designed for the target genomic sequences without the 5′

end of GG required by the T7 promoter induced the targeted mutations. This suggests that the
RGEN can target any sequence adjacent to an NGG protospacer adjacent motif (PAM) sequence,
which occurs once every 8 bp. The off-target alterations at 2 genomic loci harboring double
mismatches in the 18-bp targeting sequences were induced in the RGEN-injected embryos.
However, we also found that the off-target effects could be reduced by lower dosages of sgRNA.
Taken together, our results suggest that CRISPR/Cas-mediated RGENs may be an efficient andflexible tool for genome editing in medaka.
M Kuroyanagi, T Katayama, T Imai, Y Yamamoto, S Chisada, Y Yoshiura, T Ushijima, T
Matsushita, M Fujita, A Nozawa, Y Suzuki, K Kikuchi and H Okamoto (2013) New approachfor fish breeding by chemical mutagenesis: establishment of TILLING method in fugu (Takifugu
rubripes) with ENU mutagenesis, BMC Genomics, 14:786
Background: In fish breeding, it is essential to discover and generate fish exhibiting an effective
phenotype for the aquaculture industry, but screening for natural mutants by only depending on
natural spontaneous mutations is limited. Presently, reverse genetics has become an important
tool to generate mutants, which exhibit the phenotype caused by inactivation of a gene.
TILLING (Targeting Induced Local Lesions IN Genomes) is a reverse genetics strategy thatcombines random chemical mutagenesis with high-throughput discovery technologies forscreening the induced mutations in target genes. Although the chemical mutagenesis has beenused widely in a variety of model species and also genetic breeding of microorganisms and crops,
the application of the mutagenesis in fish breeding has been only rarely reported.
Results: In this study, we developed the TILLING method in fugu with ENU mutagenesis andhigh-resolution melting (HRM) analysis to detect base pair changes in target sequences. Fugu
males were treated 3 times at weekly intervals with various ENU concentrations, and then thecollected sperm after the treatment was used to fertilize normal female for generating themutagenized population (F1). The fertilization and the hatching ratios were similar to those of
the control and did not reveal a dose dependency of ENU. Genomic DNA from the harvested F1
offspring was used for the HRM analysis. To obtain a fish exhibiting a useful phenotype (e.g.
high meat production and rapid growth), fugu myostatin (Mstn) gene was examined as a target
gene, because it has been clarified that the mstn deficient medaka exhibited double-muscle
phenotype in common with MSTN knockout mice and bovine MSTN mutant. As a result, ten
types of ENU-induced mutations were identified including a nonsense mutation in theinvestigated region with HRM analysis. In addition, the average mutation frequency in fugu
Mstn gene was 1 mutant per 297 kb, which is similar to values calculated for zebrafish andmedaka TILLING libraries.
Conclusions: These results demonstrate that the TILLING method in fugu was established. We
anticipate that this TILLING approach can be used to generate a wide range of mutant alleles,
and be applicable to many farmed fish that can be chemically mutagenized.
Jiang X-Y, Sun C-F, Zhang Q-G, Zou S-M (2011) ENU-Induced Mutagenesis in Grass Carp
(Ctenopharyngodon idellus) by Treating Mature Sperm. PLoS ONE 6(10): e26475.
doi:10.1371/journal.pone.0026475
N-ethyl-N-nitrosourea (ENU) mutagenesis is a useful approach for genetic improvement ofplants, as well as for inducing functional mutants in animal models including mice and zebrafish.
In the present study, mature sperm of grass carp (Ctenopharyngodon idellus) were treated with arange of ENU concentrations for 45 min, and then wild-type eggs were fertilized. The results
indicated that the proportion of embryos with morphological abnormalities at segmentation stage
or dead fry at hatching stage increased with increasing ENU dose up to 10 mM. Choosing a dosethat was mutagenic, but provided adequate numbers of viable fry, an F1 population was
generated from 1 mM ENU-treated sperm for screening purposes. The ENU-treated F1
population showed large variations in growth during the first year. A few bigger mutants withmorphologically normal were generated, as compared to the controls. Analysis of DNA from 15
F1 ENU-treated individuals for mutations in partial coding regions of igf-2a, igf-2b, mstn-1,
mstn-2, fst-1and fst-2 loci revealed that most ENU-treated point mutations were GC to AT or AT
to GC substitution, which led to nonsense, nonsynonymous and synonymous mutations. The
average mutation rate at the examined loci was 0.41%. These results indicate that ENU treatmentof mature sperm can efficiently induce point mutations in grass carp, which is a potentially
useful approach for genetic improvement of these fish.
W. L. Russell, P. R. Hunsicker, D. A. Carpenter, C. V. Cornett and G. M. Guinn (1989) Effect ofdose fractionation on the ethylnitrosourea induction of specific-locus mutations in mouse
spermatogonia, Proc. Nat’l Acad. Sci. USA Vol. 79, pp. 3592-3593
As measured by' specific-locus mutations in mouse spermatogonia, fractionating a dose of 100
mg of ethylnitrosourea per kg of body weight into doses of 10 mg/kg injected intraperitoneally atweekly intervals greatly reduces the mutation frequency compared with that from a single doseof 100 mg/kg. Because there is independent evidence that the doses of 10 and 100 mg/kg reachthe germ cells in amounts directly proportional to the injected dose, the lower mutational
response with the fractionated dose is attributed to repair. The induced mutation rate expectedfrom a single 10-mg/kg dose (on the assumption that this would be 1/10th the rate observed after
10 such doses) would be only 75% of the spontaneous mutation rate. Mouse spermatogonia
apparently have an efficient repair system that is effective even against a potent mutagen.
W. L. Russell, Liane Brauch Russell, Elizabeth M. Kelly (1958) Radiation Dose Rate andMutation Frequency, Science, Vol. 128, No. 3338 (Dec. 19, 1958), pp. 1546-1550
New data have clearly confirmed the earlier finding that specific locus mutation rates obtainedwith chronic gamma irradiation of spermatogonia are lower than those obtained with acute xrays. Since this result is in contrast to classical findings for Drosophila spermatozoa, andapparently contradicts one of the basic tenets of radiation genetics, it was important to determine
what factors were responsible for it. Experiments undertaken for this purpose reveal thefollowing: (i) the lower mutation frequency is due mainly to difference in dose rate of radiation,rather than quality; (ii) a dose-rate effect is not obtained in experiments with mouse spermatozoa,
confirming classical findings for spermatozoa, and indicating that the ex-planation for intensity
dependence in spermatogonia resides in some characteristic of gametogenic stage; and (iii) adose-rate effect is found not only in spermatogonia but also in oocytes, where cell selection is
improbable, indicating that the radiation intensity effect is on the mutation process itself. A
threshold response for all mutations in spermatogonia and oocytes is not a necessaryconsequence of the findings. Plausible hypotheses consistent with the present results can lead toother predictions. From a practical point of view, the results indicate that the genetic hazards, atleast under some radiation conditions, may not be as great as those estimated from the mutation
rates obtained with acute irradiation. However, it should not be forgotten that even the lower
mutation rates obtained with the present intensity levels are still appreciable (16).

8. Genome-enabled selection doubles the accuracy of predicted breeding values for bacterial
cold water disease resistance compared to traditional family-based selection in rainbow trout
aquaculture
Roger L. Vallejo1*, Timothy D. Leeds1, Guangtu Gao1, James E. Parsons2, Kyle E. Martin2,
Jason P. Evenhuis1, Breno O. Fragomeni3, Gregory D. Wiens1 and Yniv Palti11National Center for Cool and Cold Water Aquaculture, Agricultural Research Service, UnitedStates Department of Agriculture, Kearneysville, WV, USA. Email: roger.vallejo at ars.usda.gov
2
Troutlodge, Inc., P.O. Box 1290, Sumner, WA, USA
3
Animal and Dairy Science Department, University of Georgia, Athens, GA, USA
Abstract
We have shown previously that bacterial cold water disease (BCWD) resistance in rainbow trout
can be improved using traditional family-based selection, but progress has been limited to
exploiting only between-family genetic variation. Genomic selection (GS) is a new alternative
enabling exploitation of within-family genetic variation. We compared three GS models to
predict genomic-enabled breeding values (GEBVs) for BCWD resistance in a commercial
rainbow trout population, and compared the accuracy of GEBVs to traditional breeding values(EBVs) estimated with a pedigree-based BLUP model. For these comparisons, we used BCWD
survival phenotypes recorded on 7893 training fish from 102 families, from which 1473 fishfrom 50 families had genotypes (57K SNP array). Naïve siblings of the training fish (n = 930
testing fish) were genotyped to predict their GEBVs, of which 193 were mated to produce 138
progeny testing families (PTFs). In the following generation, 9968 progeny from the PTFs were
BCWD phenotyped to empirically assess the accuracy of GEBV predictions made on their nonphenotyped parents. The accuracy of GEBVs from all three GS models were substantially higher
than the BLUP model EBVs, with the increase in accuracy ranging from 83.3% to 108.8%
depending on the GS model and survival phenotype used. Reducing the training sample size to n
= ~1000 had no negative impact on the accuracy (0.67−0.72), but with n = ~500 the accuracy
dropped to 0.53−0.61 if the training and testing fish were full-sibs, and even substantially lower
to 0.22−0.25 when they were not full-sibs. Thus using progeny performance data, we haveshown that the accuracy of genomic predictions with GS models (0.63−0.71) is substantiallyhigher than the traditional pedigree-based BLUP model (0.34−0.36). We also found that using amuch smaller training sample size compared to similar studies in livestock, GS can substantially
improve the selection accuracy and genetic gains for BCWD resistance in commercial rainbow
trout aquaculture.
Annotated Bibliography of Key Works
Meuwissen, T. H. E., B. J. Hayes and M. E. Goddard. 2001. Prediction of total genetic value
using genome-wide dense marker maps. Genetics 157: 1819-1829.
This is a landmark publication on whole genome-enabled selection. Using computer simulateddata, the authors proposed the method of genomic selection (GS) when high density SNP chips
were not yet available for any plant and livestock species. The authors attempted to estimate theeffects of ~50000 marker haplotypes simultaneously from a limited number of phenotypic

records. A genome of 1000 cM was simulated with a marker spacing of 1 cM. The markers
surrounding every 1-cM region were combined into marker haplotypes. Due to finite population
size (Ne =100), the marker haplotypes were in linkage disequilibrium with the QTL locatedbetween the markers. Then using least squares, all haplotype effects could not be estimatedsimultaneously. When only the biggest effects were included, they were overestimated and theaccuracy of predicting genetic values of the offspring of the recorded animals was only 0.32.
Next, with best linear unbiased prediction of haplotype effects assuming equal variances
associated to each 1-cM chromosomal segment, the accuracy of genomic predictions increased to
0.73. They also used Bayesian methods that assumed a prior distribution of the varianceassociated with each chromosome segment and increased the accuracy to 0.85. The authors
concluded that selection on genetic values predicted from high density marker maps could
substantially increase the rate of genetic gain in animals and plants. With this publication, theauthors proposed a GS method that today has revolutionized traditional animal and plant
breeding programs. The authors laid out concepts and methods that are used in our GS researchin rainbow trout aquaculture.
Odegard, J., T. Moen, N. Santi, S. A. Korsvoll, S. Kjoglum and T. H. Meuwissen. 2014.
Genomic prediction in an admixed population of Atlantic salmon (Salmo salar), Frontiers in
Genetics 5: 402.
The authors for the first time tested the reliability of genomic selection (GS) models in an
admixed population of Atlantic salmon. The models included ordinary genomic BLUP models(GBLUP), using genome-wide SNP markers of varying densities(1–220K), a genomic identityby-descent model (IBD-GS), using linkage analysis of sparse genome-wide markers, as well as aclassical pedigree-based model. The authors compared the reliability of the models using 5-foldcross-validation, and the studied traits were salmon lice resistance (LR) and fillet color (FC) withheritability of 0.14 and 0.43, respectively. Overall, the authors found that all genomic models
outperformed the classical pedigree-based model, for both traits and at all marker densities.
However, the relative improvement differed considerably between traits, models and marker
densities. For the highly heritable FC, the IBD-GS had similar reliability as GBLUP at highmarker densities (>22K). In contrast, for the lowly heritable LR, IBD-GS was clearly inferior to
GBLUP, irrespective of marker density. Hence, GBLUP was robust to marker density for the
lowly heritable LR, but sensitive to marker density for the highly heritable FC. The authors
hypothesized that this phenomenon may be explained by historical admixture of different
founder populations, expected to reduce short-range linkage disequilibrium (LD) and induce
long-range LD. The authors also highlighted that using ordinary GBLUP, the typical long-range
LD of an admixed population may be effectively captured by sparse marker density, while
efficient utilization of relationship information may require denser markers (e.g., >22K).
Vallejo, R. L., T. D. Leeds, B. O. Fragomeni, G. Gao, A. G. Hernandez, I. Misztal, T. J. Welch,
G. D. Wiens and Y. Palti. 2016. Evaluation of Genome-Enabled Selection for Bacterial Cold
Water Disease Resistance Using Progeny Performance Data in RainbowTrout: Insights on
Genotyping Methods and Genomic Prediction Models, Frontiers in Genetics 7: 96.
The authors evaluated the potential utility of implementing GS in traditional family-based
selective breeding programs using training and testing fish sampled from the first generation of
the NCCCWA’s BCWD resistance breeding line. The authors used GS models to predictgenomic breeding values (GEBVs) for BCWD resistance, compared the predictive ability (PA)of GEBVs to pedigree-based estimated breeding values (EBVs), and compared the impact of two
SNP genotyping methods on the accuracy of GEBV predictions. The best GEBV predictions
were similar to EBV with PA values of 0.49 and 0.46 vs. 0.50 and 0.41 for DAYS and STATUS,
respectively. Among the GEBV prediction models, ssGBLUP consistently had the highest PA.
The RAD genotyping platform had GEBVs with similar PA to those of GEBVs from the Chip
platform. The overall GEBV accuracy in this study was low to moderate, likely due to the small
training sample used. In this study, the authors explored the potential of GS for improving
resistance to BCWD in rainbow trout using, for the first time, progeny testing data to assess theaccuracy of GEBVs, and it provided the basis for further investigation on the implementation of
GS in commercial rainbow trout populations.
9. Genetic improvement of Atlantic salmon (Salmo salar L.) and Eastern oyster (Crassostrea
virginica Gmelin 1791) at the USDA-ARS National Cold Water Marine Aquaculture Center
Gary S. Burr1, Michael R. Pietrak1, Dina A. Proestou2, Brian C. Peterson1
1

USDA Agricultural Research Service, National Cold Water Marine Aquaculture Center, 25
Salmon Farm Road, Franklin, ME 04634, USA
2
USDA Agricultural Research Service, National Cold Water Marine Aquaculture Center, 469
CBLS, 120 Flagg Road, Kingston, RI 02881, USA
Abstract:
The USDA-ARS National Cold Water Marine Aquaculture Center (NCWMAC) focuses on the
coldwater marine aquaculture industry’s highest priority research needs including developmentof improved genetic stocks. Coldwater aquaculture production has potential for expansion, andboth Atlantic salmon and Eastern oysters are widely accepted by American consumers.
Commercial salmon and oyster producers predominantly utilize stocks that are not many
generations removed from wild, unselected strains. North America salmon producers are legallyrequired to culture certified stocks of North American salmon. The NCWMAC is the USDAARS facility that has been supporting the U.S. coldwater marine aquaculture industry by
developing genetically improved salmon stocks for the past thirteen years. Aquaculture of theEastern oyster is a large segment of shellfish aquaculture in the U.S. and minimal selective
breeding has been accomplished. In both species, there is a need to improve the performance ofexisting stocks. Our research meets this need through the following objectives: 1) definephenotypes, estimate genetic and phenotypic parameters, and develop a selection index in
Atlantic salmon for important traits such as carcass weight, cold tolerance, fillet color, fat
content, and sea lice resistance; 2) evaluate and validate the usefulness of incorporating genomic
information into the salmon breeding program; and 3) establish links between disease susceptible
and resistant phenotypes and genotype for the Eastern oyster. Identification of genes associatedwith oyster disease will provide markers that can be used to enhance and accelerate thedevelopment of high-performing oyster lines through selective breeding and will support the East
Coast shellfish aquaculture industry. Research accomplished with salmon will result in thedevelopment of genetically improved Atlantic salmon for release to U.S. producers andconsumers.

Annotated Bibliography of Key Works
Gjedrem, T. (ed). 2005. Selection and breeding programs in Aquaculture. Springer. Dordecht,
The Netherlands. 364 pp.
T. Gjedrem and M. Baranski. 2009. Selective breeding in Aquaculture: An Introduction.
Reviews: Methods and Technologies in Fish. Biology and Fisheries 10, Springer.
The books referenced above describes selective breeding programs for a wide range of cultured
species. The editor and many authors are from Norway which has the longest running Atlantic
salmon selective breeding program in the world.
Gjerde, B., Odegard, J. & Thorland, I. 2011. Estimates of genetic variation in the susceptibilityof Atlantic salmon (Salmo salar) to the salmon louse Lepeophtheirus salmonis. Aquaculture, 314,
66-72.
The paper above provides a brief overview of previous research looking at the heritability of sea
lice resistance, while importantly demonstrating the need to normalize lice infection counts to afactor, such as lice density, that accounts for fish size at the time of infection.
Proestou, D.A., Vinyard, B.T., Corbett, R.J., Piesz, J., Allen, S.K., Small, J.M., Li, C., Liu, M.,
DeBrosse, G., Guo, X. and Rawson, P., 2016. Performance of selectively-bred lines of easternoyster, Crassostrea virginica, across eastern US estuaries. Aquaculture. 464:17-27.
The article referenced above looked at the performance of selectively-bred lines of easternoysters in five representative grow-out sites and detected significant effects of selection and large
and significant genotype by environment interactions. This information will be useful inevaluating current breeding strategies for the eastern oyster.
Ermgassen, P., Spalding, M., Blake, B. et al. 2012. Historical ecology with real numbers: pastand present extent and biomass of an imperiled estuarine habitat. Proceedings of the Royal
Society of London B: Biological Sciences. 279:3393-3400.
The article referenced above analyzed historical shellfish landing and biomass data along theEastern U.S. coast and correlated it to various anthropogenic stressors including overexploitation,
habitat degradation, climate change and the resultant spread of disease.
10. The Molluscan Breeding Program: 20 years of selective breeding of the Pacific oyster,
Crassostrea gigas, on the West Coast of the US
Romain Morvezen1*, Claudio de Melo2, Blaine Schoolfield1, Evan Durland1, Brett Dumbauld3,
Chris Langdon1
1

Department of Fisheries and Wildlife and Coastal Oregon Marine Experiment Station, Hatfield
Marine Science Center, Oregon State University, 2030 Marine Science Drive, Newport, OR
97365, US. Email: romain.morvezen at oregonstate.edu
2Laboratory of Marine Mollusks, Department of Aquaculture, Federal University of Santa
Catarina, Florianópolis, SC, 88061-600, Brazil
3
USDA-ARS, Hatfield Marine Science Center, Oregon State University, 2030 Marine Science
Drive, Newport, OR 97365, US
Abstract:

The Molluscan Broodstock Program (MBP) was initiated in 1996 by Oregon State University(OSU) and the US Dept. of Agriculture with the goal of increasing the performance of Pacificoysters (Crassostrea gigas) for use in aquaculture on the West Coast, USA. MBP utilizes afamily-based selection breeding design and was founded by broodstock collected from wild
populations in Washington State and British Columbia, Canada. Breeding design and hatcheryproduction of larvae and spat is undertaken by OSU at the Hatfield Marine Science Center in
Newport, Oregon and families are planted out for field trait estimates at industry sponsored fieldsites along the West Coast. The selection process utilizes an index based on family performances
for three traits of interest: family yield, average individual harvest weight and survival. Resultsof five generations of selective breeding show that substantial genetic gains have been attained.
Realized gains are lower than expected, however, and we suspect this might be caused bycompetition for food in dense culture conditions and a changing climate, including intensified
upwelling events which bring deep, acidified and nutrient-rich water to the surface. Heritability
estimates have declined in the last two generations of the program, which corroborate theseresults and further suggest a changing environment. Significant genetic by environment (GxE)interactions have been detected between Puget Sound and coastal estuarine plant-out sites,
indicating major environmental differences between the two types of sites. Taken together, theseresults are being used to develop modified breeding approaches for MBP, including a more
careful indexing approach to take into account GxE interactions. Further considerations for MBP
include breeding for new traits in response to emerging issues such as ocean acidification anddiseases.
Annotated Bibliography of Key Works
Langdon, C., Evans, F., Jacobson, D., & Blouin, M. (2003). Yields of cultured Pacific oysters
Crassostrea gigas Thunberg improved after one generation of selection. Aquaculture, 220(1),227-244.
This publication marks the first evaluation of the genetic gain obtained by the Molluscan
Broodstock Program, after one generation of selection. The authors demonstrate significant
improvement in yields of selected lines compared to unselected controls. However, they foundonly weak correlation between family performances in intertidal and subtidal environments,
which indicated possible interactions between family performance and environment.
Evans, F., Matson, S., Brake, J., & Langdon, C. (2004). The effects of inbreeding on
performance traits of adult Pacific oysters (Crassostrea gigas).Aquaculture, 230(1), 89-98
The effect of inbreeding was evaluated in the context of a commercial growing operation cby
measuring survival, family yield and individual growth after one or two growing season. Results
showed that families with a high (F=0.2) inbreeding coefficient performed worse than families
with a low (F=0.006) inbreeding coefficient, and this effect was stronger after 2 growing seasons.
The result called for a strict monitoring of inbreeding in a selective broodstock program such as
the MBP.
Melo, C. M. R., Durland, E., & Langdon, C. (2016). Improvements in desirable traits of the
Pacific oyster, Crassostrea gigas, as a result of five generations of selection on the West Coast,
USA. Aquaculture, 460, 105-115.
Authors evaluated the heritability, genetic correlations and genetic gain for survival, individual
weight and yield over 5 generations of selective breeding in the MBP. Substantial genetic gainwas observed throughout generations except for individual weight, where gains were limited
after the second generation. Realized gains were less than expected however given the genetic
gain, perhaps due to poor control over the selection of broodstock for controlled crosses.
Heritabilities seemed to decline in the 4th and 5th generations, and the authors suspect this wascaused by changing environmental conditions such as the intensification of upwelling events due
to climate change.
11. Seriola genomics and the knowledge repository Serioladb.org
Andrew Severin2*, Catherine Purcell1, Margaret Carter2, Rick Masonbrink2, Arun Seetharam2,
John Hyde1
1

NOAA Fisheries, Southwest Fisheries Science Center (SWFSC), La Jolla, CA, USA
Genome Informatics Facility, Office of Biotechnology, Iowa State University, Ames Iowa,
USA. Email: severin at iastate.edu
2

Abstract:
Due to the high value of Seriola in the seafood industry, Seriola aquaculture is a great interest in
the US and Japan. Hatchery production of Seriola species is accelerating but has been hindered
by a propensity for deformities and growth heterogeneity developed during larval and early
juvenile stages that limit the production capacity and efficiency.
To advance Seriola aquaculture, we are developing genomic resources for Seriola dorsalis andSeriola rivoliana. The starting point for this resource includes a draft genome assembly withgene annotations for S. dorsalis. In addition, we have resequenced 90 fish from three locations(Cedros Island, La Paz, and San Diego) and generated an F2 population to explore the genetic
component in malformations in juvenile fish (swim bladder inflation, several head and jaw
deformities and malformations in the operculum and branchiostegal structures).
To consolidate this data into a single knowledge repository, we are creating an international web
resource Serioladb.org. As yellowtail culture grows, the genetic tools on the website will bemade available to producers and other groups working to improve culture for this and other
Seriola species. This research may also serve as a model for genetic resource development foraquaculture species or conservation dependent species with limited genomic resources, and may
help reveal genomic regions and genes contributing to trait variation in wild populations.
Annotated Bibliography of Key Works
Martinez-Takeshita, N, Purcell CM, Chabot CL, Craig MT, Paterson CN, Hyde JR, and Allen
LG. 2015. A Tale of Three Tails: Cryptic Speciation in a Globally Distributed Marine Fish of theGenus Seriola. Copeia: Vol. 103, No. 2, pp. 357-368.
Authors sampled 42 Seriola lalandi from South Africa, Japan, New Zealand, California, Mexico
Pacific, Mexico Gulf and Chile to better understand regional populations and evolutionary
patterns. Two mitochondrial genes (CR and COI) and four nuclear genes (RAG2, EHHADH,
UBE3A, and MLL) were used to generate a phylogenetic tree with Seriola dumerili as an
outgroup. Based on this data, it was concluded that fish currently named as Seriola lalandi in fact

comprise three species. The authors propose that Seriola lalandi in the Northwest Pacific revert
to the name Seriola aureovittata and the Northeast pacific fish should revert to the name Seriola
dorsalis while the fish in the southern hemisphere should remain as Seriola lalandi.
Westesson O, Skinner M, and Holmes I. 2012. Visualizing next-generation sequencing data with
JBrowse. Briefings in Bioinformatics: Vol. 14, No. 2, pp. 172-177.
The authors describe a web-based browser for displaying genetic sequencing data at thenucleotide level. JBrowse is the successor to the very successful GBrowse genome browser.
The most significant advance is the user-side processing of tracks for visualization, which allows
for greater scalability without significant investment on the server side. Other improvements
include Google maps-style browser interface with smooth transitions. JBrowse is actively
maintained and can be downloaded from https://github.com/GMOD/jbrowse. JBrowse is an
important tool for the visualization of high throughput sequencing data and should be included inany aquaculture genomic resources toolkit.
Ficklin SP, Sanderson LA, Cheng CH, Staton ME, Lee T, Cho IH, Jung S, Bett KE and Main D.
2011. Tripal: a construction toolkit for online genome databases. Database: Vol 2011,
doi:10.1093/database/bar044.
In this work, the authors describe web based content management system named Tripal. Once a
research group starts to accumulate high throughput sequencing data such as a genome assembly,
annotation, SNPs, a genetic map, and epigenetic data etc., Tripal provides a web interface thatorganizes this data in a database for easy display on web pages. It is widely used by modelorganisms (FlyBase, WormBase, Genome Database for Rosaceae, TreeGenes and so on). The
Chado database that serves to store and distribute the data to the webpages can also be used by
JBrowse to visualize genomic data.
12. The first meeting of Seriola genomics and breeding consortium
Akiyuki Ozaki1*, Kazuharu Nomura1, Catherine Purcell2, John Hyde2, Victor Martinez3, Andrew
Severin4
1

National Research Institute of Aquaculture, Fisheries Research Agency, 422-1, *E-mail: aozaki
at affrc.go.jp. Nakatsuhamaura, Minamiise-cho, Watarai-gun, Mie, 516-0193, Japan
2
Southwest Fisheries Science Center, National Marine Fisheries Service, 8901 La Jolla Shores
Drive
La Jolla, CA 92037-1508, U.S.A
3
Universidad de Chile, FAVET-INBIOGEN, Unidad de Genetica y Genomica Animal Facultadde Ciencias Veterinarias y Pecuarias.
4
Iowa State University, Office of Biotechnology Genome Informatics Facility 207 science I
Ames, Iowa 50011-3260
Abstract:
The first Seriola Genomics Symposium was held at the NOAA Fisheries, Southwest Fisheries
Science Center in La Jolla, California on January 7th and 8th of 2016. 35 people attended this
symposium from six countries (USA, Japan, Mexico, Brazil, Chile, and Australia), and the list of

participants included both scientists and Seriola producers. Specifically, the participants
represented eight universities, five government agencies, four Seriola producers, and twobiotechnology companies. During the two-day symposium, presenters described both the natureof their research and the progress of the research being conducted; these presentations outlined
development of genetic/genomic resources to improve aquaculture for five different Seriola
species (S. quinqueradiata, S. dumerili, S. lalandi, S. dorsalis, and S. rivoliana). Presenters werealso asked to identify research needs and collaborative potential they could identify from theirwork. Discussions on the second day described the process of making a web-based Seriola
genetic/genomic resource and addressed concerns in collaborative projects in terms ofintellectual property. Notably, there was a long discussion with the Seriola producers identifying
key needs and how to address those needs. By the end of the workshop, plans were formed for
future workshops to occur at regular intervals (every other year) and steps were generally
outlined for individual research groups to contribute to the genetic/genomic resources for Seriola.
Participants agreed that this meeting would serve to identify them as part of a more formal
Seriola Consortium. Several collaborations have already been formed from the Seriola
Symposium, including with the NOAA Fisheries, SWFSC and Iowa State University to share
genomic sequencing data, and with the University of Chile for the development of a S. lalandi
SNP array. The National Research Institute of Aquaculture, FRA Japan, agreed to supply the
SNP data of S. quinqueradiata and S. dumerili, to development of a Seriola genotyping array.
Keywords: Yellowtail Seriola species (S. quinqueradiata, S. dumerili, S. lalandi, S. dorsalis, andS. rivoliana).
Annotated Bibliography of Key Works
Akiyuki Ozaki., Kazunori Yoshida., Kanako Fuji., Satoshi Kubota., Wataru Ka.i, Jun-ya Aoki.,
Yumi Kawabata., Junpei Suzuki., Kazuki Akita., Takashi Koyam.a, Masahiro Nakagawa.,
Takurou Hott., Tatsuo Tsuzaki., Nobuaki Okamoto., Kazuo Araki., Takashi Sakamoto. (2013)
Quantitative Trait Loci (QTL) Associated with Resistance to a Monogenean Parasite (Benedenia
seriolae) in Yellowtail (Seriola quinqueradiata) through Genome Wide Analysis. PLoS ONE
8(6): e64987.
Benedenia infections caused by the monogenean fluke ectoparasite Benedenia seriolae seriously
impact marine finfish aquaculture. Genetic variation in host has been inferred to play asignificant role in determining the susceptibility to this parasitic disease. To evaluate the genetic
basis of Benedenia disease resistance in yellowtail (Seriola quinqueradiata), a genome-wide andchromosome-wide linkage analyses were initiated using F1 yellowtail families (n = 90 per
family) based on a high density linkage map with 860 microsatellite and 142 single nucleotide
polymorphism (SNP) markers. Two major quantitative trait loci (QTL) regions on linkage
groups Squ2 (BDR-1) and Squ20 (BDR-2) were identified. These QTL regions explained 32.9–
35.5% of the phenotypic variance. On the other hand, the relationship between QTL forsusceptibility to B. seriolae and QTL for fish body size were investigated. The QTL related to
growth was found on another linkage group (Squ7). As a result, the authors present first genetic
evidence that contributes to detailing phenotypic resistance to Benedenia disease, and the results
will help resolve the mechanism of resistance to this important parasitic infection of yellowtail.

Paul Whatmore., Nguyen Hong Nguyen., Adam Miller., Rob Lamont., Dan Powell., Trent
D'Antignana., Erin Bubner., Abigail Elizur., Wayne Knibb. (2013) Genetic parameters for
economically important traits in yellowtail. Kingfish Seriola lalandi., Auaculture 400-401 77-84
The aim of the present study was to estimate genetic parameters for body and carcass traits,
visual condition score, and deformity in yellowtail kingfish Seriola lalandi, an emerging
aquaculture species in Australia. These novel data and genetic parameters are required to solve
the problem of how to conduct efficient selection in this and related species. Analyses were
performed on a total of 400 data records collected from a yellowtail kingfish breeding population.
These results suggest that selection for high growth would result in concomitant increase in fillet
weight, a carcass trait of paramount importance. It is concluded that there is substantial potential
for genetic improvement of economically important traits especially growth performance andfillet weight in the current population of yellowtail kingfish.
Catherine M., Chris L. Chabot., Matthew T. Craig., Natalie Martinez-Takeshita., Larry G. Allen.,
John R. Hyde. (2015) Developing a genetic baseline for the yellowtail amberjack species
complex, Seriola lalandi sensu lato, to assess and preserve variation in wild populations of these
globally important aquaculture species. Conservation Genetics 16-6,1475–1488
Recent study suggest the globally distributed yellowtail amberjack, Seriola lalandi sensu lato, isa complex of three closely related species. Together, these and three other species of Seriola
comprise an important component of global aquaculture production with an estimated annual
value of $1.3 billion. As yellowtail aquaculture grows, the impact of unintentional releases onwild populations has become an increasingly important issue, particularly in light of international
trade of hatchery seed. To create a genetic baseline, we examined spatial genetic structure in 260
specimens collected from seven locations over a wide geographical range using 15 nuclear
microsatellites and mitochondrial control region sequences. Overall genetic differentiation
among locations, as revealed by microsatellite data, was highly significant (FST = 0.085, DEST
= 0.382, P < 0.001), and pairwise estimates of divergence derived from mitochondrial andmicrosatellite data support the presence of four significantly differentiated populations
corresponding to the N.E. Pacific, N.W. Pacific, S. Pacific, and South Atlantic. Based on thegenetic differentiation detected in this study, and recently published sequence data, these
populations more accurately reflect the presence of at least three cryptic species of Seriola.
Especially strong genetic differentiation between hemispheres indicates that the equatorial region
is a significant dispersal barrier for yellowtail. This study represents the broadest geographic
investigation of genetic population structure conducted, to date, for specimens of the S. lalandi
complex.
A. Patel., P. Dettleff., E. Hernandez., V. Martinez (2015) A comprehensive transcriptome ofearly development in yellowtail kingfish (Seriola lalandi) Molecular Ecology Resources 16-1,
364-376
Seriola lalandi is an ecologically and economically important species that is globally distributed
in temperate and subtropical marine waters. The aim of this study was to identify large numbersof genic single nucleotide polymorphisms (SNPs) and differential gene expression (DGE) related
to the early development of normal and deformed S. lalandi larvae using high-throughput RNAseq data. A de novo assembly of reads generated 40 066 genes ranging from 300 bases to 64 799
bases with an N90 of 788 bases. Homology search and protein signature recognition assigned
gene ontology (GO) terms to a total of 15 744 (39.34%) genes. A search against the Kyoto

Encyclopedia of Genes and Genomes Pathway database (KEGG) retrieved 6808 KEGG
orthology (KO) identifiers for 10 520 genes (26.25%), and mapping of KO identifiers generated
337 KEGG pathways. Comparisons of annotated genes revealed that 1262 genes were
downregulated and 1047 genes were upregulated in the deformed larvae group compared to thenormal group of larvae. Additionally, we identified 6989 high-quality SNPs from the assembled
transcriptome. These putative SNPs contain 4415 transitions and 2574 transversions, which will
be useful for further ecological studies of S. lalandi. This is the first study to use a global
transcriptomic approach in S. lalandi, and the resources generated can be used further forinvestigation of gene expression of marine teleosts to investigate larval developmental biology.
The results of the GO enrichment analysis highlight the crucial role of the extracellular matrix innormal skeleton development, which could be important for future studies of skeletal deformities
in S. lalandi and other marine species.
13. Uses of Genetic Parentage Analysis in Cultured California Yellowtail (Seriola dorsalis)
Matthew Craig1*, Catherine Purcell1. Kevin Stuart2, Mark Drawbridge, John Hyde1
1

NOAA National Marine Fisheries Service, La Jolla, California, USA. *Email: matthew.craig at
noaa.gov
2
Hubbs SeaWorld Research Institute, San Diego, California, USA
Abstract
The genus Seriola is an important group of cultured species representing a more than USD$1.5
million industry. With the demand for seafood continually rising, this popular group of fishes isa prime candidate for culture thus supplementing supply and relieving harvest pressure on wild
populations. In a culture setting, we have the ability to obtain a complete set of genetic markers
for the brood stock, allowing for determination of the parental contribution to each spawning
event. In order to maximize output and quality of offspring, we explored the various usages ofgenetic parentage data in a cultured population of the California Yellowtail, Seriola dorsalis. It
was determined that parentage was informative in not only allowing for an estimate of parental
contribution numerically, but was also able to provide linkages to egg quality and may provide
linkages to the presence/absence of deformities in offspring.
Annotated Bibliography of Key Works
Purcell, C., Chabot, C., Craig, M., Martinez-Takeshita, N., Allen, L., Hyde. J. 2015. Developing
a genetic baseline for the yellowtail amberjack species complex, Seriola lalandi sensu lato, toassess and preserve variation in wild populations of these globally important aquaculture species.
Con. Gen. 16(6):1475-1488
The authors present a global description of genetic variation in the Seriola lalandi species
complex. Overall genetic differentiation among locations, as revealed by microsatellite data, washighly significant (FST = 0.085, DEST = 0.382, P < 0.001), and pairwise estimates of divergencederived from mitochondrial and microsatellite data support the presence of four significantly
differentiated populations corresponding to the N.E. Pacific, N.W. Pacific, S. Pacific, and South
Atlantic. Based on the genetic differentiation detected in this study, and recently publishedsequence data, these populations more accurately reflect the presence of at least three cryptic
species of Seriola. Especially strong genetic differentiation between hemispheres indicates thatthe equatorial region is a significant dispersal barrier for yellowtail. This study represents thebroadest geographic investigation of genetic population structure conducted, to date, for
specimens of the S. lalandi complex.
Baskett, M., Waples, R. 2013. Evaluating alternative strategies for minimizing unintended fitness
consequences of cultured individuals on wild populations. Con. Biol. 27(1):83-94
The authors discuss important considerations for the maintenance of adaptively positive traits incultured populations in order to minimize impact on wild populations due to accidental or
intentional release into the wild. The authors quantitatively assessed two strategies: 1. Reduce
selection in captivity, or 2. Maintaining a second population to minimize captive/wildinteractions. They concluded that the appropriate approach depends on the goals of the culture
activity.
Martinez-Takashita, N., Purcell, N., Chabot, C., Craig, M., Paterson, C., Hyde, J., Allen, L. 2015.
A Tale of three tails: Cryptic speciation in a globally distributed marine fish of the genus Seriola.
Copeia 103(2):357-368.
This paper demonstrates that the nominal species Seriola lalandi is actually comprised of three
distinct species. The authors present compelling evidence for genetic separation of populations.
This information is critical in culture applications for this species in that what were once
considered populations of the same species will now be separated in new broodstock groups.
14. Past, present and future research on Ostreid herpesvirus 1 infections of the Pacific oyster
in Tomales Bay, California
Colleen A. Burge1 and Carolyn S. Friedman2
1

Institute of Marine and Environmental Technology, University of Baltimore County, 701 E Pratt
Street, Baltimore, MD 21224 USA
2School of Aquatic & Fishery Sciences, University of Washington, Box 355020, Seattle,
Washington 98195 USA
Abstract
Large-scale sporadic mortalities of juvenile Pacific oyster, Crassostrea gigas have occurred in
Tomales Bay, California for 20 years. The Ostreid herpesvirus 1 (OsHV-1) is the identified
causative agent consistently associated with mortalities, with a confirmed infectious etiology.
OsHV-1 is a global pathogen of bivalve molluscs, although detection in the United States islimited to Tomales Bay and nearby Drakes Bay. OsHV-1 is a member of the Order
Herpesvirales, sharing common morphological criteria with vertebrate herpesviruses, althoughsequence data indicates a tenuous relationship. OsHV-1 was classified as the first member of an
invertebrate herpesvirus family, Family Malacoherpesviridae. Sequence data indicate thatmultiple global variants of OsHV-1 exist, and the virus detected in Tomales Bay is not identicalto any one variant. Elevated water temperatures are consistently associated with oyster
mortalities in Tomales Bay, and may trigger viral replication and/or transmission of OsHV-1 tonaïve juvenile oysters. Laboratory trials indicate qPCR and RT qPCR can be used to demonstratevirus replication and gene expression. Survival of young Pacific oysters in Tomales Bay isdependent on outplant time, size, and oyster stock indicating genetic improvement anddevelopment of biomarkers for improved survival of Pacific oysters infected with OsHV-1 ispossible. Since 2008, an economically devastating increase in C. gigas mortality in France has
been associated with a new genetic variant OsHV-1 µvar, which is lethal to all life history stages.
OsHV-1 µvar continues to spread in Europe and a similar variant causes losses in Australia, New
Zealand, and Asia. OsHV-1 µvar’s ability to kill seed and adults heightens concern over thisvariant relative to its progenitor, OsHV-1, which is lethal to larvae and seed only. OsHV-1
resistance has been shown to confer resistance to µvar. Recent studies demonstrated the ability toselect for resistance to OsHV-1 and we propose studies to evaluate selection of resistance to both
OsHV-1 µvars and their progenitor in US oyster lines.

15. Type 1 Ostreid Herpesvirus (OsHV-1) variants in Japan
Yoshiko Shimahara*1, Satoshi Watanabe2, Jun Kurita3, Ikunari Kiryu3, Toyohiro Nishioka1, Kei
Yuasa2, Morihiko Kawana4, Takashi Kamaishi5, Norihisa Oseko4
1

National Research Institute of Aquaculture, Japan Fisheries Research and Education Agency,
Tsuiura, Kamiura, Saiki, Oita 879-2602, Japan, *E-mail: shimahara at affrc.go.jp

2

National Research Institute of Aquaculture, Japan Fisheries Research and Education Agency,
Nakatsuhamaura 422-1, Minamiise, Mie 516-0193,
3
National Research Institute of Aquaculture, Japan Fisheries Research and Education Agency,
Hiruda 224-1, Tamaki, Mie 519-0423,
4
Fisheries Agency, Tokyo 100-8907, Japan,
5
Hokkaido National Fisheries Research Institute, Japan Fisheries Research and Education
Agency, Nakanoshima 2-2, Toyohira-ku, Sapporo-shi, Hokkaido 062-0922, Japan
Abstract:
Ostreid herpesvirus 1 (OsHV-1) μVar is a variant of OsHV-1 newly reported from France(GenBank accession no. HQ842610), and it is suspected of being the causative agent of acutemass mortality events of Pacific oysters Crassostrea gigas during summers in Europe since 2008.
OsHV-1 μVar differed from reference OsHV-1 by nucleotide mutations in the C2/C6 fragment
including ORF4, and in the IA2/IA1 fragment including ORF42/43 (Segarra et al., 2010). Japanis one of the major producers of Pacific oyster. Our previous study indicated that, 23 types of
OsHV-1 variant, showing 96% to 99% similarity to the reference OsHV-1, were obtained from
Pacific oyster, kumamoto oyster, C. sikamea and suminoe oyster, C. ariakensis collected in 2007
and from Pacific oyster in 2011 in Japan (Shimahara et al., 2012). Although 18 variants among
the obtained 23 possessed a microsatellite deletion in C2/C6 unique to OsHV-1 μVar, thenucleotide sequence was not identical to the OsHV-1 μVar. In this study, further surveillance of
OsHV-1 variants was conducted for Pacific oysters in 2012. Nine hundreds of spats, or juveniles
of Pacific oyster collected in the 4 main oyster producing areas were subjected to PCR assay
using the primer pairs C2 and C6 (Renault and Arzul 2001). PCR products were amplified from40 out of 900 oysters, and 13 different nucleotide sequences, showing 96% to 99% similarity to
the reference OsHV-1, were obtained. Although 11 sequences among the obtained 13 possesseda microsatellite deletion unique to OsHV-1 μVar, all PCR products contained 2 conserved
nucleotides that were shared with the reference OsHV-1 and not with OsHV-1 μVar. Here, we
found variable types of OsHV-1 in oysters in Japan, which had different nucleotide sequencesfrom that of OsHV-1 μVar in France (HQ842610).
Keywords: Ostreid herpesvirus, OsHV-1, Crassostrea gigas, Japan, Pacific oyster
Annotated Bibliography of Key Works
Shimahara, Y., Kurita, J., Kiryu, I., Nishioka, T., Yuasa, K., Kawana, M., Kamaishi, T., and N.
Oseko. 2012. Surveillance of Type 1 Ostreid Herpesvirus (OsHV-1) variants in Japan. Fish
Pathology, Vol: 47: 4. Pp 129-136
Ostreid herpesvirus 1 (OsHV-1) μVar is a variant of OsHV-1 that is suspected of being thecausative agent of acute mass mortality events of Pacific oysters during summers in Europe since2008. The authors investigated a distribution of OsHV-1 variants in six main oyster-producing
areas in Japan by PCR to determine whether OsHV-1 μVar was present in Japan. PCR products
were amplified from 123 out of 1,714 oysters, and 23 different nucleotide sequences wereobtained. Although 18 sequences among the 23 obtained possessed a microsatellite deletion
unique to OsHV-1 μVar, all PCR products contained two conserved nucleotides that were sharedwith reference OsHV-1 and not OsHV-1 μVar in ORF4. It was found that variable types of
OsHV-1 distribute throughout Japan, but their nucleotide sequences were not completely
identical to OsHV-1 μVar.

16. Characterization of genetic and epigenetic variation in hatchery and natural-origin
steelhead, Oncorhynchus mykiss
Mackenzie Gavery1*, Krista Nichols2, Giles Goetz1, Mollie Middleton1, Penny Swanson3
1

School of Aquatic and Fishery Sciences, University of Washington, Seattle, WA 98195. Email:mackenzie.gavery at noaa.gov
2

Conservation Biology Division, Northwest Fisheries Science Center, National Marine Fisheries Service,
NOAA, 2725 Montlake Blvd East, Seattle, WA 98112
3
Environmental and Fisheries Sciences Division, Northwest Fisheries Science Center, National Marine
Fisheries Service, NOAA, 2725 Montlake Blvd East, Seattle, WA 98112

Abstract:
Relative reproductive success studies have documented substantial fitness loss for wild steelhead after asingle generation of rearing in the hatchery, but the relative contribution of genetic selection and/or
environmentally-induced heritable epigenetic changes passed through the germline are unknown. Here,as a step toward understanding mechanisms of fitness loss, we describe genetic and epigenetic variation in
adult hatchery and natural-origin steelhead from the Methow River, Washington USA. Our main
objectives were to determine if hatchery and natural-origin fish from this stock could be distinguishedgenetically by examining SNPs across the entire genome, and whether differences in epigenetic
programming (DNA methylation) in somatic and germ cells could be detected between the two groups.
Although genetic analysis using RAD-Seq did not reveal differences between the hatchery and natural
fish, we found significant differences in epigenetic programming in both somatic (red blood cells) andgerm cells (sperm). Because hatchery fish experience similar environmental conditions as their wildconspecifics once they leave the hatchery, our results raise the possibility that these DNA methylation
changes occurred during the first year in the hatchery and persisted into adulthood in the form of an‘epigenetic memory’ of the hatchery environment.

Annotated Bibliography of Key Works
Baerwald M. R., Meek M. H., Stephens M. R., Nagarajan, R. P., Goodbla, et al. 2015 Migration-related
phenotypic divergence is associated with epigenetic modifications in rainbow trout. Mol. Ecol. 25: 17851800.
This study investigates the role of epigenetics (DNA methylation) in migration-related life history traits in
Oncorhynchus mykiss. The authors used reduced representation bisulfite sequencing to perform
comparative DNA methylation analysis between juvenile resident and smolt F2 siblings generated from across between steelhead (migratory) and rainbow trout (nonmigratory). Fifty-seven differentiallymethylated regions, many of which were in gene regulatory regions, were identified between residents
and smolts, suggesting a relationship between epigenetic variation and variation in migration-related
phenotypes.
Gavery MR and Roberts SB. 2013 Predominant intragenic methylation is associated with gene expression
characteristics in a bivalve mollusc. PeerJ. 1:e215.
This dataset, generated using methylation-enriched high-throughput bisulfite sequencing, represents thefirst high-resolution methylome in any mollusc. DNA methylation data were compared to gene
expression datasets and a positive relationship between intragenic methylation and gene expression levels
was identified. These data suggest that DNA methylation patterns may play a role in regulating gene
expression in molluscs.
Navarro-Martín L., Viñas J., Ribas L., Díaz N., Gutiérrez A., et al. 2011 DNA methylation of the gonadal
aromatase (cyp19a) promoter is involved in temperature-dependent sex ratio shifts in the European sea
bass. PLoS Genet. 7: e1002447

The authors report that in European sea bass (Dicentrarchus labrax), which exhibits
temperature-dependent sex determination, exposure to high temperature in early development
was associated with increased DNA methylation in the promoter of the aromatase gene
(cyp19a1a) and a higher proportion of phenotypic males. Furthermore, in vitro methylation of
the aromatase promoter was sufficient to suppress transcription of the gene, supporting a role for
DNA methylation as a mechanistic link between temperature and sex ratios in species exhibiting
temperature-dependent sex determination.
Potok M. E., Nix D. A., Parnell T. J., Cairns B. R., 2013 Reprogramming the maternal zebrafish genome
after fertilization to match the paternal methylation pattern. Cell 153:759-72.
Genome-wide DNA methylation patterns in zebrafish gametes, various stages of embryos and a somatic
tissue (muscle) were analyzed using whole genome bisulfite sequencing. This high-resolution approach
identified dynamic and unique patterns of DNA methylation during development in zebrafish. Results
suggest that the functional significance of sperm DNA methylation patterns in fish is to provide
transcriptional competency to the early embryo, which ‘inherits’ the DNA methylation pattern in the
sperm.
Shao C., Li Q., Chen S., Zhang P., Lian J., et al., 2014 Epigenetic modification and inheritance in sexual
reversal of fish. Genome Res. 24: 604–615.
The half-smooth tongue sole (Cynoglossus semilaevis) was used as a model to investigate the role of
epigenetic regulation in species with environmental sex determination. Using genome-wide bisulfite
sequencing of normal male, female and pseudomale fish (generated by exposing genetic females to hightemperature during a sensitive developmental window), the authors showed that sex-reversed genetic
females exhibit methylation patterns consistent with genetic males, both of which differ from themethylome of normal females. Furthermore, it was reported that global methylation patterns are inheritedby F1 pseudomale offspring generated by crosses between temperature-induced sex-reversed
pseudomales and normal females, suggesting transgenerational epigenetic inheritance of sex reversal inthis species.

17. Towards a functional understanding of DNA methylation in shellfish and implications foraquaculture.
Mackenzie Gavery, Claire Olson, Steven Roberts*
School of Aquatic and Fishery Sciences, University of Washington, Seattle WA.
Email sr320 at uw.edu
Abstract:
There is an amazing amount of diversity incorporated into the genome of oysters and othermarine invertebrates including vastly expanded gene families, high mutation rates, and numerous
mobile elements. These are certainly a benefit to broadcast spawners living in fluctuating
environments. Recent work examining DNA methylation is revealing new insight into similar
diversity at the epigenetic level. The function of DNA methylation in species such as bivalves
where the limited amount of DNA methylation is predominantly found in gene bodies is notcompletely understood. One emerging possible explanation is that the role of gene body DNA
methylation is dependent on gene function, a potential phenomenon that has arisen fromselective pressure on lineage-specific life history traits.
With respect to commercially important traits, we know that in other taxa epigenetic marks areassociated with phenotypes independent of genetic variation, the environment can influence
DNA methylation, and epigenetic marks can be inherited. In shellfish, we are still learning aboutrole of epigenetic processes such as DNA methylation in controlling the phenome. However itdoes appear that epigenetic processes will be important to consider in future efforts to advanceaquaculture, particularly in changing environments.
Annotated Bibliography of Key Works
Gavery MR and Roberts SB. (2014) A context specific role for DNA methylation in bivalves
Briefings in Functional Genomics. doi:10.1093/bfgp/elt054 https://goo.gl/YrnzXi
A review of current knowledge of DNA methylation in bivalves. A primary conclusion is that thefunctional role of the gene could influence the role of DNA methylation in influencing
expression.
Claire E. Olson, Steven B. Roberts. Indication of family-specific DNA methylation patterns indeveloping oysters bioRxiv doi: http://dx.doi.org/10.1101/012831
This study provides the first single-base pair resolution DNA methylomes for both oyster sperm
and larval samples from multiple crosses. While sample sizes are very low, this work suggests
DNA methylation patterns could be inherited.

Roberts, Steven (2015): Compilation of DNA Methylation Genome Feature Tracks (Crassostrea
gigas). figshare. https://dx.doi.org/10.6084/m9.figshare.1456226.v2
Genome feature tracks and accompanying IGV session file to visualize DNA methylation data
for the Pacific oyster (Crassostrea gigas).