47th Scientific Symposium Program of the UJNR
Aquaculture Panel

Marine Aquaculture in a Changing Environment

Okinawa Industry Support Center, Room 304
1831-1 Oroku, Naha, Okinawa
November 12th and 13th, 2019

Photo credit: Atsushi Nanami

47th Scientific Symposium of the UJNR Aquaculture Panel
Marine Aquaculture in a Changing Environment
November 12th and 13th, 2019
Okinawa Industry Support Center, Room 304
1831-1 Oroku, Naha, Okinawa
Aim of the Symposium
The UJNR Aquaculture Panel is a cooperative research exchange between the U.S. and Japan jointly
addressing environmental and technical issues that affect the aquaculture industries of both nations.
The 47th UJNR Aquaculture Symposium is the final symposium of a 3-year cycle with the theme Marine
Aquaculture in a Changing Environment. Environmental changes impact aquaculture in many ways. Nutrient
pollution is driving eutrophication and dead zones; ocean acidification is changing water chemistry, andclimate change is already influencing our food supply, freshwater availability, weather and way of life.
Aquaculture will be impacted by, and can also impact, these environmental changes over various scales.
Aquaculture of finfish, shellfish and seaweed have different threats, benefits and opportunities related toenvironmental change.
Over the last two years, we discussed the potential of aquaculture to mitigate impacts of environmental
change (e.g., carbon sequestration, bioextraction of nutrients and CO2, antacidity, and oxygen production),
impacts of environmental change on aquaculture production(e.g., effects of ocean acidification on shellfish
aquaculture), and science to mitigate these impacts (countermeasures). This is the final year of the three
year-plan, and the symposium theme is the Application of Aquaculture Technology to Provide Sustainable
Seafood and Reduce Impacts of Environmental Change. This theme includes development of technology toincrease marine aquaculture production to offset seafood deficit due to loss of capture fisheries impacted byenvironmental change, and to augment food deficits due to impacts of environmental change on inland
agriculture.

Program
Tuesday, November 12, 2019
Registration

9:00-9:15

Welcome and Aim of the Symposium
Hideaki Aono, Japan Panel Chair, Japan Fisheries Research and Education Agency

9:15 - 9:25

New techniques to assess and mitigate impacts of environmental changes on aquaculture(Moderators: Simona Augyte and Natsuki Hasegawa)
Bacterial communities in marine sediments; a biological parameter to evaluate coastal environment
Tomoko Sakami, Research Center for Aquaculture Systems, National Research Institute of Aquaculture, FRA
9:25 - 9:55
Applications of environmental DNA data in support of aquaculture
Tom Noji, Northeast Fisheries Science Center, NOAA Fisheries

9:55 - 10:25

Stable isotopic approach to investigate nitrogen pathway in coastal aquaculture area
Satoshi Watanabe, Research Center for Aquaculture Systems, National Research Institute of Aquaculture,
FRA

10:25 - 10:55

Establishment of immunological assays and baseline profile of hemocytes in the hard clam Mercenaria

mercenaria as evaluation biomarkers for environmental stresses
Huiping Yang, School of Forest Resources and Conservation, Institute of Food and Agricultural Sciences,
University of Florida

10:55 - 11:25

Purple urchin barrens: an opportunity for aquaculture and fisheries to work together to solve anenvironmental issue
Luke Gardner, California Sea Grant

11:25 - 11:55(PM: UJNR Aquaculture Panel Business Meeting)
Reception

18:00 - 20:00

Wednesday, November 13
Impact assessment of environmental changes and conservation of fisheries environment in coral reef areas(Moderators: Luke Gardner and Masakazu Hori)
Impacts of ocean acidification on Japan coastal water and marine fisheries
Haruko Kurihara (Invited speaker), University of the Ryukyus

9:15 - 9:45

Sustain seafood resources in the U.S. affiliated Pacific islands- status and strategies
Cheng-Sheng Lee, Center for Tropical and Subtropical Aquaculture, United States Department ofAgriculture

9:45 - 10:15

Towards effective coral community restoration for sustainable fishery of a coral reef grouper Epinepherus

ongus: implication of ecosystem-based management
Atsushi Nanami, Research Center for Sub-tropical Fisheries, Seikai National Fisheries Research Institute,
FRA

10:15 - 10:45

Scaling up coral restoration to meet the demands of a collapsing ecosystem
Tali Vardi, ECS for NOAA Fisheries Office of Science & Technology

10:45 - 11:15

Sustainable large-scale coral restoration by establishing "artificial spawning hotspot"
Go Suzuki, Research Center for Sub-tropical Fisheries, Seikai National Fisheries Research Institute, FRA
11:15 - 11:45
Lunch Break (75 min)Production management under environmental changes in bivalve aquaculture(Moderators: Cheng-Sheng Lee and Atsushi Nanami)
The Influence of Climate and Environment on the Growth and Survival of Pacific Oyster Seed in US West
Coast Estuaries
Brett Dumbauld, Agricultural Research Service, United States Department of Agriculture 13:00 - 13:30
Comparative Study of the Impact of Environmental Changes on Oyster Culture between USA and Japan, as
Collaborative Research under UJNRNatsuki Hasegawa, Stock Enhancement and Fisheries Oceanography Department, Hokkaido National
Fisheries Research Institute, FRA

13:30 - 14:00

Oyster aquaculture using seagrass beds as a climate change countermeasure
Masakazu Hori, National Research Institute of Fisheries and Environment of Inland Sea, FRA
14:00 - 14:30
Coffee Break (15 min)Aquaculture technologies to respond to environmental changes; Seaweed breeding and feed development(Moderators: Brett Dumbauld and Satoshi Watanabe)
Kelp, Saccharina spp, population genetics in the Northwest Atlantic for guiding a breeding program ofthermally resilient strains
Simona Augyte, Dept. of Ecology and Evolutionary Biology, University of Connecticut

14:45 - 15:15

Cell selection technique for establishment of low salinity tolerance strain in Pyropia tenuipedalis
Mahiko Abe, Applied Aquabiology, National Fisheries University, FRA

15:15 - 15:45

Improvement of dietary effect on juvenile Ruditapes philippinarum using the dietary-supplements and new
diet microalga
Yasuhiro Yamasaki, Applied Aquabiology, National Fisheries University, FRA

15:45 - 16:15

Exploration of alternative protein sources in the development of a sustainable Japanese white trevally

Pseudocaranx dentex juvenile diet
Jonas Miller, Uragami Station, Aquaculture Research Institute, Kindai University

16:15 - 16:45

Science Symposium Closing
Michael Rust, US Panel Chair, NOAA Fisheries Office of Aquaculture

16:45 - 17:00

List of ParticipantsAbe, Mahiko

Japan Fisheries Research and Education Agency (FRA), National Fisheries
University (NFU)

Aono, Hideaki

Japan Panel Chair, FRA Headquarters

Augyte, Simona

University of Connecticut

Chiba, Toshinari

Akita Prefectural Department of Agriculture, Forestry and Fisheries, Institute ofFisheries Promotion

Dumbauld, BrettUnited States Department of Agriculture, Agricultural Research Service

Gardner, Luke

California Sea Grant

Hasegawa, Natsuki

FRA, Hokkaido National Fisheries Research InstituteHori, Kazumasa

FRA, National Research Institute of Fisheries and Environment of Inland Sea

Ishikawa, Takanori

Okinawa Prefectural Fisheries Research and Extension Center（OPFREC）

Kishimoto, Kazuo

OPFREC

Kurihara, Haruko

University of the Ryukyus

Lee, Cheng-Sheng

United States Department of Agriculture, Center for Tropical and SubtropicalAquaculture(College of Tropical Agriculture and Human Resources, University of Hawaii at
Manoa)

Miller, Jonas

Kindai University

Miwa, Satoshi

FRA, National Research Institute of Aquaculture (NRIA)

Nanami, Atsushi

FRA, Seikai National Fisheries Research Institute (SNFRI)

Nakata, Yuji

OPFREC

Noji, TomNOAA Fisheries, Northeast Fisheries Science Center

Okumura, Takuji

FRA, NRIA

Oomine, Risako

OPFREC

Rust, Michael

US Panel Chair, NOAA Headquarters

Saito, Yoho

FRA, Headquarters

Sakami, Tomoko

FRA, NRIA

Suzuki, Go

FRA, SNFRI

Takano, Masatsugu

FRA, Headquarters

Vardi, TaliNOAA Fisheries Office of Science & Technology

Watanabe, Satoshi

FRA, NRIA

Yamasaki, Yasuhiro

FRA, NFU

Yang, Huiping

University of Florida

Bacterial communities in marine sediments: a biological parameter toevaluate coastal environments
Tomoko Sakami1*, Toru Udagawa2 and Hiroshi Yagi3
1*

National Research Institute of Aquaculture, Japan Fisheries Research and Education
Agency, 422-1 Nakatsuhamaura, Minami-ise, Mie, Japan 516-0193
2
National Research Institute of Fisheries Engineering, Japan Fisheries Research and
Education Agency, 7620-7 Hasaki, Kamisu, Ibaraki, Japan 314-0408
3
National Defense Academy, 1-10-20 Hashirimizu, Yokosuka, Kanagawa, Japan 2398686*E-mail: sakami at affrc.go.jp
Environmental monitoring of marine coastal areas is becoming increasingly importantbecause of the accelerated global climate change and rapid industrial development.
Understanding certain biological parameters is fundamental to assess the environmental
conditions of fisheries ground. However, much effort is required for the examination ofcommunity change because species identification must be done manually by well-trained
professionals who may not be available whenever necessary. In this context, the recently
developed metagenomic analysis of bacterial communities may be useful as a mechanism
to obtain much data with less effort. Bacterial communities in coastal sediments are
known to be affected by environmental conditions. We examined the bacterial community
composition in coastal sediments collected around Cape Inubo, where the cold Oyashio
mixing water and the warm Kuroshio current influence the northern and southern areasof the cape, respectively. The purpose of this study was to understand how bacterial
communities vary in areas under the influence of different types of ocean currents.
Surface sediments were collected from seabed at water depths of 10 m and 30 m in thenorthern and southern areas, and 16S rRNA gene profiles were examined using theIllumina Miseq platform. The bacterial community composition was different between
the two water depths, and the bacterial communities at 10 m depth was different between
the northern and southern areas. The chlorophyll a content and temperature wereindicated as the major environmental parameters influencing the composition of thebacterial communities. Characteristic taxonomic groups were identified from each area.
At 10 m depth in the southern area, operational taxonomic units (OTUs) of Bacteroides,
which is known as a high molecular weight organic matter degrader, were abundant,
whereas OTUs of Nitrosomonas and Nitrospira, which are related to the nitrification
process, were less abundant. This implies that the decomposition and mineralization
process of the sediment organic matter in this area may differ from those in the other threeareas. In fact, at 10 m depth in the southern area, the total organic matter content was highbut chlorophyll a content was low as compared to the other areas, suggesting a low input
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of fresh organic matter. In conclusion, the bacterial community composition in coastal
sediments around Cape Inubo varied with water depth. At 10 m depth, the bacterial
communities differed between the two areas. These differences were concomitant withthe differences in some environmental parameters. Moreover, the characteristic
taxonomic groups found in each area suggested that the difference seemed to be related
to properties of the sediments’ organic matter. Metagenomic information of benthic
bacterial communities is useful to monitor coastal environmental changes because not
only does it reflect the environmental conditions, but its taxonomic features indicate theecological functions of the community.
Annotated Bibliography of Key Works
Sakami, T., Kakehi, S. 2019. Distribution and community composition of ammoniaoxidizing archaea and bacteria in coastal sediments in response to sediment material
gradients at Sendai Bay, Japan. Gojobori, T. et al. (eds.), Marine Metagenomics,
Springer Nature Singapore Pte Ltd, pp 161-181.
The distributions of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing
bacteria (AOB) were determined along an environmental gradient from the coastal mud
to the offshore coarse sand at Sendai Bay, Japan. The abundance of AOA ammonia
monooxygenase alpha subunit gene (amoA) was high in the coastal muddy areas andlow in the offshore sandy areas. There was a strong positive correlation between AOAamoA abundance and ammonia content in the sediment. The distribution of AOB-amoA
was similar to that of AOA-amoA in July, but remarkably low in the muddy sediments
in December. Clone library analysis indicated that the community composition for bothtypes of organisms differed in sandy and muddy sediments and that the diversity wasconsiderably lower in the muddy sediments. These results suggest that the abundance ofammonia-oxidizing organisms was controlled by the ammonia levels in the sediment.
However, there are some inhibitive conditions for AOB: presumably, the low organic
matter supply to the surface oxic layer during autumn in the muddy sediment in Sendai
Bay.

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Applications of environmental DNA data in support of aquaculture.
Thomas Noji1*, Daniel Wieczorek1, Beth Phelan1, Yuan Liu2, Lisa Milke2, Renee
Mercaldo-Allen2, Julie Rose2, Lauren Sassoubre3 and Bruce Nash4
1

Northeast Fisheries Science Center, NOAA Fisheries, Sandy Hook, NJ 07732, USA.
Northeast Fisheries Science Center, NOAA Fisheries, Milford, CT 06460, USA.
3
School of Engineering and Applied Sciences, University of Buffalo, Buffalo NY, 14260,
USA
4
DNA Learning Center, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724,
USA*E-mail: thomas.noji at noaa.gov
2

With advances in analytical and computational technologies the data forenvironmental DNA or eDNA are becoming rapidly and increasingly available. eDNA
data have been applied successfully to assess presence of fish species, impacts of humanactivities on benthic biota and more recently to a limited extent to assess biomass.
Because of the relative ease with which eDNA data can be collected, the number of
proposed applications is increasing rapidly; this includes applications to support
aquaculture.
The NEFSC Sandy Hook Lab was instrumental in development of a NOAA Fisheries
Strategic Initiative to explore the potential use of eDNA to assess fish stock size anddistribution. In 2017, with an expanded team of eDNA experts we began discussions to
conduct experiments to test whether environmental DNA can be used as a quantitative
tool to support fisheries stock assessments. We are the only group in NOAA performing
seawater laboratory experiments to support this research. These experiments are
conducted in a closed recirculating, to determine species-specific shedding and decay
rates of eDNA (Sassoubre et al. 2016) under different environmental conditions. This isparticularly important for the assessment of stocks which cannot easily be surveyed bystandard trawl surveys. To date we have run experiments on adult black sea bass, winterflounder, and summer flounder (Sassoubre et al. in prep) and in 2020 several new species
will be investigated. One of the important next steps to making this a quantitative survey
tool is to develop eDNA particle transport models (Andruszkiewicz et al. 2017). Thesemodels are intended to indicate the geographical origin of collected eDNA and to estimate
the biomass (Kelly et al. 2019) of the corresponding fish stock. Further, the NEFSC
Milford Laboratory is now in its third year of conducting field experiments on the effectof caged oyster on the diversity and behavior of finfish. The investigation measures
diversity with eDNA, as well as visual corroboration using underwater video and sonar.
Observed in 2018 using eDNA were 17 species of finfish; 14 species were observed
visually. Only about half of the species were recorded in both sets of observations. The
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investigation sites with caged oyster had a greater diversity of finfish than comparablesites without cages (Liu et al. submitted).
This talk will present data from these two investigations. Further the talk will explore
ways that eDNA data can help to address some of the challenges related to aquaculture.
Some of the applications to be discussed include measurement of eDNA as a surveillance
tool for shellfish and finfish pathogens and nuisance organisms such as sea lice, as anindicator of benthic impacts, as an indicator of recovery rates for augmenting natural
populations, and other uses.
Annotated Bibliography of Key Works
Sassoubre, L.M., K.M. Yamahara, L.K. Gardner, B.A. Block and A.B. Boehm.
Quantification of Environmental DNA (eDNA) Shedding and Decay Rates for Three
Marine Fish. 2016. Environ. Sci. Technol. 50: 10456−10464.
A key publication on experiments for shedding and decay rates of eDNA from marine
finfish. Also includes a much cited conceptual model for processes in the field affecting
concentrations of eDNA.
Andruszkiewicz, E.A., H.A. Starks, F.P. Chavez, L.M. Sassoubre, B.A. Block, A.B.
Boehm. 2017. Biomonitoring of marine vertebrates in Monterey Bay using eDNA
metabarcoding. PLoS ONE 12(4): e0176343. https://doi.org/10.1371/journal.
pone.0176343
This paper is the first to my knowledge to apply numerical modeling to predict theorigin of eDNA collected in the field. The outputs include levels of uncertainty for the
calculations.
Kelly, R.P., A.O. Shelton and R. Gallego. 2019. Understanding PCR Processes to Draw
Meaningful Conclusions from Environmental DNA Studies. Scientific Reports (2019)9:12133 | https://doi.org/10.1038/s41598-019-48546-x
This paper presents guidelines for the use of PCR for the successful application of
eDNA data to estimate biomass. The investigation is a modeling approach and describes
how the proportional indices of amplicon reads capture trends in taxon biomass withhigh accuracy
Liu, Y., G.H. Wikfors, J.M. Rose, R.S. McBride, L.M. Milke, R. Mercaldo-Allen.
Submitted. Application of environmental DNA metabarcoding to spatiotemporal finfish
community assessment in a temperate embayment. Frontiers in Marine Science.
Describes the field investigations addressing the beneficial effect of caged oyster on
finfish biodiversity in Long Island Sound, USA.
Sassoubre, L.M., D. Wieczorek, B. Phelan, B. Nash, A. Kirtane, T. Noji. In Preparation.
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Quantification of Environmental DNA (eDNA) Shedding and Decay Rates for Three
Marine Fish in Support of Quantitative Field Investigations.
Describes the results from laboratory experiments on one pelagic fish species and two
flatfish. Also relates these data to field surveys for these species with comparisons to
trawl data.

5

Stable isotopic approach to investigate nitrogen pathway in coastal
aquaculture area
Satoshi Watanabe
National Research Institute of Aquaculture, Japan Fisheries Research and Education
Agency, 422-1Nakatsuhamaura, Minami-ise, Mie, Japan 516-0193
E-mail: swat at affrc.go.jp
Coastal eutrophication due to increasing anthropogenic activities and consequentenvironmental deterioration, such as anoxia, harmful algal bloom and hydrogen sulfide
emission have been problematic in many parts of the world, sporadically causing massmortality of aquatic organisms. While this holds true for Japan, oligotrophication andresulting reduction in coastal fisheries and aquaculture productivity have also been arecognized problem in some parts of Japan. Some studies argue that ever-dwindlingfisheries production of coastal resources, as well as unfed aquaculture production of
inorganic and organic extractive species is partially attributable to the reduced nutrient
levels in the coastal waters in recent years. Eutrophication mitigation efforts have reduced
the load of nitrogen and phosphorus of terrestrial and aquaculture origins to the coastal
environment over the past forty years in Japan. Allegedly excessive oligotrophication has
reduced primary productivity and hence carrying capacity or coastal ecosystems
supporting coastal fisheries and aquaculture. However, complex nutrient flows withincoastal ecosystems are not necessarily well understood.
Fisheries production, inclusive of bottom culture, of the Manila clam, Ruditapes
philippinarum, for instance has reduced by 95% over the past three decades. Insufficient
food supply is one amongst many speculated factors causing the reduction. Foodavailability to the clam is often represented by chlorophyll a level in a water column
collected far from the bottom water the clams inhale and filter feed. Our stable isotopic
study has suggested that the food source of the clam may differ depending on the bottom
topography of the habitat. Many filter feeding bivalves are generally held to bephytoplanktivorous; however, they also feed on benthic microalgae. Provisional ratio of
the planktonic to benthic microalgae seems to be affected by the extent of resuspensionof the latter from the sediment to the bottom water to make them available to the clam.
As a result, clams inhabiting the same tidal flat only 10 m apart from each other can havea different carbon stable isotopic (13C) signature, indicating the presence of small-scaledifference in nutrient sources for the clam.
Anthropogenic nitrogen is known to have higher nitrogen stable isotopic (15N)
signature than naturally available nitrogen. For example, sewage treated water andagricultural fertilizers contain dissolved inorganic nitrogen (DIN) with high 15N, whichacts as an indicator of the level of anthropogenic nitrogen loads to coastal waters. The
6

15N in the soft tissues of the Manila clam was found to be positively correlated to DIN
concentration in the bottom water in tidal flats in several areas in Japan. The higher 15N
in the clam is considered to be attributable to higher 15N in the food particles, especiallythose in sediment surface, and 15N in the food particles was associated with that in DIN.
This demonstrates that elevated 15N in DIN in coastal waters due to anthropogenic
nitrogen loads is reflected in the 15N in the clam. Thus, pervasive effects of terrestrial
nutrient load to the food availability to the clam may be ascertained by 15N.
Integrated multi-trophic aquaculture (IMTA) is a technique to use unfed aquacultureof inorganic and organic extractive species to consume effluent from fed aquaculture of
finfish. Although IMTA is proposed as a mitigation measure for eutrophication, it canalternatively enhance unfed aquaculture production in an oligotrophic environment. In
order to trace nutrient flow within an IMTA system, it is necessary to know how stable
isotopic signatures differ between the aquaculture feed and fish feces, both of which are
a nutrient source for the organic extractive species, such as for example sea cucumber.
We compared the 13C and 15N of feces collected from five different finfish species(Lutjanus argentimaculatus, Chanos chanos, Siganus guttatus, Trachinotus blochii andLates calcarifer) fed the same formulated feed. While the feces 15N showed nosignificant difference among the species, 13C were different presumably depending on
the digestibility of the plant ingredients. Herbivorous and omnivorous species may beable to digest and assimilate plant ingredients that cannot be used by carnivorous species.
Therefore, stable isotopic technique may not be used to trace carbon flow within an IMTA
system.
Annotated Bibliography of Key Works
Satoshi Watanabe, Satoshi Katayama, Masashi Kodama, Naritoshi Cho, Kaoru Nakata
and Masaaki Fukuda (2009) Small-scale variation in feeding environments for the Manila
clam Ruditapes philippinarum in a tidal flat in Tokyo Bay. Fisheries Science 75: 937–
945.
The relative contribution of particulate organic matters (POMs) in water column andsediment as a food source for the Manila clam, Ruditapes philippinarum, was studied
using carbon and nitrogen stable isotopic method (13C and 15N) in a tidal flat in
Yokohama, Japan. Comparisons of 13C and 15N among R. philippinarum and POMs insurface water, bottom water, and sediment surface indicated that R. philippinarum larger
than 5 mm shell length (SL) mainly assimilated benthic POM, and individuals smaller
than 5 mm SL assimilated benthic and pelagic POM. Continuous measurements ofchlorophyll concentrations in the bottom water revealed tide-driven resuspension of thebenthic phytopigments. R. philippinarum showed differences in 13C and 15N along an
inshore–offshore transect, indicating small-scale spatial differences in POM provision in
the tidal flat. These findings suggest that POM in the bottom water, supposedly inhaled
by R. philippinarum, is a mixture of a larger proportion of resuspended benthic POM
7and a smaller proportion of pelagic POM, and that the mixing ratio of the POMs may beaffected by the hydrodynamics of flooding water associated with tidal flat topography.
Some stable isotopic studies overly emphasize the importance of benthic POM as a foodsource for benthic communities; however, relative importance of benthic and pelagic
food source should be argued quantitatively.
Satoshi Watanabe, Masashi Kodama and Masaaki Fukuda (2009) Nitrogen stable isotope
ratio in the manila clam, Ruditapes philippinarum, reflects eutrophication levels in tidal
flats. Marine Pollution Bulletin 58: 1447–1453.
Understanding the effects of anthropogenic eutrophication on coastal fisheries may help
in the enhancement of fishery production by effective utilization of sewage effluents, aswell as in the consequent reduction of eutrophication. In this study, it was revealed thatthe nitrogen stable isotope ratio (15N) in the soft tissues of the manila clam, Ruditapes
philippinarum, can be used as an indicator of anthropogenic eutrophication levels in
tidal flat environments. Investigation of 15N in dissolved inorganic nitrogen (DIN),
particulate organic matter (POM), sedimentary organic matter (SOM) and soft tissues of
the clam in five tidal flats in Japan with different levels of DIN concentration wascompleted. In addition, it was found that the acid insoluble fraction of the shell organic
matrix, conchiolin, can be used as a proxy for the soft tissues in 15N analyses. This willcontribute in easier storage handling and the expansion of chances for sample
acquisition. Oligotrophication of the coastal waters started to draw attention after
publication of this article.

8

Establishment of immunological assays and baseline profile ofhemocytes in the hard clam Mercenaria mercenaria as evaluation
biomarkers for environmental stresses
Yangqing Zeng and Huiping Yang*
School of Forest Resources and Conservation, Institute of Food and Agricultural Sciences,
University of Florida, Gainesville, FL 32653, USA*E-mail: huipingyang at ufl.edu
Environmental stresses, such as temperature, salinity, dissolved oxygen, pollutions,and algal toxins and are important factors to affect shellfish aquaculture by disrupting
physical and physiological conditions. In general, vertebrate animals respond toenvironmental stresses (and diseases pathogens) through physical barriers, such as skinfur system, and a complex network of circulating cells and molecular regulations by
neurological, endocrinological, and immunological systems. As invertebrates,
aquaculture shellfish bivalves can usually close their shells tight or move away (forspecies with this capability) as immediate responses to environmental stresses. On
neurological or endocrinological levels, the responses are limited due to their primitive
nervous system and endocrine organs. On immunological level, shellfish bivalves possess
an open circulatory system with hemolymph circulating through vessels and sinuses
throughout soft tissues and the internal organs to deliver nutrients and oxygen. Throughhemocytes in circulatory system, shellfish bivalves can use innate immunity, a primitive
nonspecific immunological system which all animals have, to provide internal defensivefunctions against various pathogen and environmental stresses, and may also use adaptive
immunity, an acquired sophisticated immunological system which was used to recognize
only in vertebrates but now in bacteria and invertebrates, to recognize and destroy specificinvaders.
Shellfish aquaculture in the U. S. is a $325 million industry and involves in several
hundreds of farms along the coastal states. Environmental stresses often occur in theinshore farming sites, such as high temperatures in hot summers, especially during low
tides with shallow water, dramatic salinity changes with river rundown in rainy season,and harmful algal bloom. While facing these stresses, hemocytes immune system ofshellfish would make responses. With prolonged exposure, shellfish could be
subsequently susceptible to pathogens, increase disease outbreaks and eventually suffer
heavy mortality. Similar to the blood tests as diagnostic tool for health evaluation inhuman and livestock, it is hypothesized that hemocyte assays in shellfish could be used
as effective parameters to evaluate the impact of the environmental stresses, serve as
criteria for genetic breeding, and provide diagnosis tools to guidance the aquaculture
operation management.
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The goal of this study was to develop hemocyte immunological assays and establish
baseline profile for hemocyte in the hard clam Mercenaria mercenaria, one major
aquaculture species. The immunological assays established in this study will include
hemolymph osmotic pressure, pH, hemocyte count, cell type, viability, apoptosis,
phagocytosis, and reactive oxygen species (ROS) production. Cell count showed aconcentration of 0.809-5.310 × 106/ml (n = 10) with three different types – agranulocytes,
granulocytes and blast-like agranulocytes. Cell viability stained with SYBR green I andpropidium iodide indicated a percentage of 71.6 ± 12.4% alive cells (n =10). This study
is currently ongoing, and more data will be further reported.
Annotated Bibliography of Key Works
Anisimova, A. A. (2013). Morphofunctional parameters of hemocytes in the assessmentof the physiological status of Bivalves. Russian Journal of Marine Biology 39(6): 381391.
This is the comprehensive review publication on shellfish hemocyte immunology. The
basic morphology, cell type, cell function and dynamic changes of hemocytes inshellfish bivalves were summarized. The effects of temperature, season, salinity, annual
cycle, food quality availability, toxin algae, bacteria and virus, pollutions, and heavy
metals on hemocyte morphology, number and function were reviewed and documented.
Donaghy, L., et al. (2009). Flow cytometry studies on the populations and immune
parameters of the hemocytes of the Suminoe oyster, Crassostrea ariakensis. Fish &Shellfish Immunology 27(2): 296-301.
This publication studied the immunological activities and morphology of hemocytes in
the Suminoe oyster Crassostrea ariakensis using flow cytometry and light microscopy.
Three types of hemocyte types were identified, including hyalinocyte, granulocyte andblast-like cells. Cell count, survival, mortality, phagocytosis, and reactive oxygen
species (ROS) production were evaluated using flow cytometer with different staining
methods. It revealed that the granulocytes are most active in the cell phagocytosis andthe hyalinocytes showed a certain level of the phagocytosis and oxidative activity, andthe blast-like cells did not show any phagocytosis or oxidative activity.
Vieira, G. C., et al. (2017). Morphological and functional characterization of thehemocytes from the pearl oyster Pteria hirundo and their immune responses against
Vibrio infections. Fish & Shellfish Immunology 70: 750-758.
This paper tested most hemocyte parameters including morphological characterization
through light & electron microscopy and flow cytometry. Same as that in Sumino
oysters, three types of the hemocytes were identified. Assays of phagocytosis andreactive oxygen species (ROS) production was performed by use of flow cytometer.
Furthermore, hemocyte responses with exposure to a Vibrio pathogen was evaluated.
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Hegaret, H., et al. (2003). Flow cytometric analysis of haemocytes from eastern oysters,
Crassostrea virginica, subjected to a sudden temperature elevation II. Haemocyte
functions: aggregation, viability, phagocytosis, and respiratory burst. Journal of
Experimental Marine Biology and Ecology 293(2): 249-265.
This paper studied the effects of temperature increase on hemocyte functions of easternoysters, including aggregation, viability, phagocytosis, and ROS production. This is oneof a series of publications on oyster hemocyte functions and detailed protocols were
documented for readers to understand the analysis procedure. The results indicated thattemperature increase (from 20 to 28 degree) caused no significant change in hemocyte
aggregation, decreased the phagocytosis of all hemocyte types, induced significant
hemocyte mortality in all hemocyte types, and increased, although not significantly, theROS production.
Jauzein, C., et al. (2013). Flow cytometric characterization of hemocytes of the sunrayvenus clam Macrocallista nimbosa and influence of salinity variation. Fish & Shellfish
Immunology 35(3): 716-724.
Salinity is one the most important factor potentially affecting shellfish physiology,
especially in the inshore area with river flows. This paper estimated the hemocyte types
and cellular parameters (oxidative activity, lysosomal content, phagocytosis capacity) insunray venus clams, a potential aquaculture species in Florida. After exposure tosalinities of 18, 21, 25, 30, 35 and 38 ppt for 7 days, hemocyte samples were collected
and analyzed the parameters along with estimation of physiological status of clams,
including mortality, valve closure, and filtration activity. It was interestingly found that
hemocytes of sunray venus clam appeared as a unique population, both in terms ofmorphology and intracellular parameters. Clams after transferring to 18 and 21 ppt
resulted in valve closure, mortality and decreased filtration activity. Low salinities
highly impacted hemocyte functions as follows: increased cell and lysosomal
compartment volumes, decreased phagocytosis capacity, and increased oxidative stressand mortality.

11

Purple urchin barrens: an opportunity for aquaculture and fisheries towork together to solve an environmental issue
Luke Gardner1,3*, Katherine Neylan1, Walan Chang2, Katrina Herrmann2, Helaina
Lindsey1, Max Rintoul1, Katherine Roy2
1

Moss Landing Marine Laboratories. 8272 Moss Landing Rd, Moss Landing, CA 95039,
USA
2
California State University, Monterey Bay. 5108 Fourth Avenue, Marina, CA 93933,
USA
3
California Sea Grant, University of California, San Diego. 9500 Gilman Dr. La Jolla, CA
92093-0232, USA*E-mail: lgardner at ucsd.edu
Sea urchin barrens and can stretch over 1000s of kilometers and last decades at a time.
They are characterized by a predominance of urchins and coralline algae where kelp
forests once existed. In contrast to barrens, kelp forests provide habitat supporting
thousands of vertebrates, invertebrates and plant species. Because kelp forests are
keystone hosts, their presence is vital to sustaining commercial and recreational industries,
including fishing and tourism. However, these kelp forests can collapse and shift toalternate stable states whereby urchin barrens persist. Over the last 4 decades, transitions
between kelp beds and sea urchin barrens have been widely reported along temperate
coastlines globally. During a kelp forest phase, urchin predation is the primary mechanism
keeping sea urchin populations in check. However, due to various factors including
climate change, predator densities can be reduced leading to shifts toward urchin barrens.
Development of urchin fisheries has been implicated several times in recent history as adriver to return urchin barrens to kelp forests. However, this driver most recently has notworked in California where a large barren is persisting. California already had a urchinfishery, but it has been uneconomical for the fishery to operate given urchins in the barrens
had little gonad development or undesirable human consumption traits necessary forcommercialization. Aquaculture in the US has potential to restore kelp forests bycollaborating with fisheries to harvest wild urchins from barrens and fatten them in an
aquaculture setting prior to sale. This is of particular interest in California given U.S
fisheries and environmental groups have long opposed aquaculture development due toconcern of competition and environmental damage respectively. A collaboration between
aquaculture and fisheries to positively impact the environment via urchin barren
restoration would help to develop aquaculture in the US and California. Although urchinranching operations exist, there are still technical limitations to the activity, primarily
being availability of macroalgae diets given seasonality and the propensity of urchinbarrens to deplete kelp forests. Development of sustainable alternative diets for urchins12is necessary for future commercial urchin aquaculture. In this student lead study, apreliminary replicated diet trial was performed for ranching purple sea urchins(Strongylocentrotus purpuratus) collected from California barrens using 4 diet treatments
including giant kelp (Macrocystis pyrifera), ogo (Gracilaria pacifica), formulatedcommercial diet (Urchinomics) and an unfed control. During the 10 week study duration,
gonadal somatic index (GSI) was measured in a subset of urchins from each replicate tank
every 2 weeks. Baseline GSI at the beginning of the trial was <0.5%. A GSI of 10% wasreached most rapidly in the formulated diet treatment at 6 weeks, followed by ogo andkelp at 8 and 17 weeks, respectively. This study was a preliminary examination of thefeasibility of urchin ranching in California, showing biological potential for alternative
diets to develop urchin gonads with a view to restore kelp forests and develop a nascentaquaculture industry in California.
Annotated Bibliography of Key Works
Filbee-Dexter, K., Scheibling, R.E., 2014. Sea urchin barrens as alternative stable statesof collapsed kelp ecosystems. Marine Ecology Progress Series 495: 1–25.
https://doi.org/10.3354/meps10573
This paper provides an excellent description of sea urchin barrens as to what they arecomprised of and their extent both temporally and spatially across the world. The authors
examine and list the drivers of phase shifts between barrens and kelp forests. They
describe different thresholds for forward (to barrens) and reverse (to kelp beds) shifts, inaccordance with alternative stable-state dynamics. They surmise that accelerating climatechange and increasing anthropogenic impacts play important roles in altering alternative
stable-state dynamics and triggering phase shifts.
Heflin, L.E., Makowsky, R., Taylor, J.C., Williams, M.B., Lawrence, A.L., Watts, S.A.,
2016. Production and economic optimization of dietary protein and carbohydrate in theculture of juvenile sea urchin Lytechinus variegatus. Aquaculture 463: 51–60.
https://doi.org/10.1016/j.aquaculture.2016.05.023
This manuscript is a through collection of urchin feeding experiments to understand thenutrient requirements of urchins for aquaculture. The paper creates predictive models of
growth, production and efficiency outcomes and generates economic analysis models in
relation to these dietary outcomes for urchins held in culture. The models compare
dietary requirements and growth outcomes in relation to economic costs and provide
insight for future commercialization of sea urchin aquaculture
Unuma, T., Sakai, Y., Agatsuma, Y., Kayaba, T., 2015. Sea Urchin Aquaculture in Japan,
in: Echinoderm Aquaculture. John Wiley & Sons, Ltd, pp. 75–126.
https://doi.org/10.1002/9781119005810.ch5
This is the most recent review of urchin aquaculture in Japan. Japan is the foremost
13consumer of urchins and significant producers of urchins both from ranching and closed
life cycle aquaculture. The chapter details the history of urchin fisheries in Japan andthe rise of urchin barrens and urchin aquaculture. The review discusses the divermediated destruction of urchins to bring back kelp beds as well as reseeding efforts torestore overfished urchin grounds. The review also discusses the development of fulllife-cycle aquaculture to meet both reseeding and commercial production requirements.
Also the movement of urchins from barrens to other kelp grounds and aquaculture
facilities for commercial fattening are detailed.

14

Impacts of ocean acidification on Japan coastal waterand marine fisheries
Haruko Kurihara
University of the Ryukyus, 1 Senabaru, Nishihara, Okinawa, 903-0213, Japan
E-mail: harukoku at sci.u-ryukyu.ac.jp
Increase of anthropogenic atmospheric carbon dioxide (CO2) is now causing changeof seawater carbon chemistry, which is known as ocean acidification (OA, IPCC 2007).
As the excess atmospheric CO2 dissolves into the seawater, seawater pCO2 rises while
seawater pH and calcium carbonate saturation state (Ω) decrease (Orr et al. 2000). Several
studies have demonstrated that change of seawater carbonate chemistry can affect thephysiology of marine organisms, particularly reducing calcification of marine calcifiers
such as mollusks, echinoderms and corals (Fabry et al. 2008, Kurihara 2008, Doney et al.
2009). In addition to these global climate impacts, principally in coastal waters, land
derived stresses such as eutrophication and hypoxia can affect coastal ecosystems andmarine organisms (Gattuso et al. 2015). Hence, there is now a strong need forunderstanding the coastal seawater chemistry and its potential impact on marine resources.
Considering the high reliance on marine resource as a food source for tourism, here inthis study, I will highlight present scientific knowledge and gaps of ocean acidification
impacts in coastal waters of Japan, including Okinawan reefs.
Annotated Bibliography of Key Works
Kurihara H (2008) Effects of CO2-driven ocean acidification on the early developmental
stages of invertebrates. Marine Ecology Progress Series 373: 275-284This paper first reviewed the effect of ocean acidification (OA) on the early
developmental stages of marine calcifiers including mollusks, sea urchins and corals.
Results highlight that future changes in ocean acidity will potentially impact the
population size and dynamics, as well as the community structure of calcifiers, and will
therefore have negative impacts on marine ecosystems.
Doney SC, Fabry VJ, Feely RA, Kleypas JA (2009) Ocean acidification: The other CO2
problem. Annual Review of Marine Science 1: 169-192
This paper has highlighted that global warming and the increase of atmospheric CO2
can cause another problem in the ocean called ocean acidification. This paper reviewed
the effect of increase atmospheric CO2 on ocean carbon system, the biological effects ofocean acidification on marine organisms and the potential impacts on marine ecology
and biogeochemistry.
15

Gattuso J-P, Magnan A, Bille R, Cheung WWL, Howes EL, Joos F, Bopp L, Cooley SR
et al. (2015) Contrasting futures for ocean and society from different anthropogenic CO2
emissions scenarios. Science 349 (6243)
This paper reviewed the potential impacts of climate change including ocean
acidification on the ocean ecosystem and its services under high and stringent emission
scenario (RCP 8.5 and 2.6). Results suggest that services, including coastal protection
and fish capture, will be in high risk at RCP 8.5 scenario.

16

Sustain seafood resources in the U.S. affiliated Pacific islands- status
and strategies
Cheng-Sheng Lee
College of Tropical Agriculture and Human Resources, University of Hawaii at Manoa,
1955 East West Road, Ag. Sci. 216 Honolulu, HI 96822, USA
E-mail: chenglee at hawaii.edu
The U.S.-affiliated Pacific Islands (U.S.A. PI) include American Samoa, the Republicof the Marshall Islands (RMI), the Federated States of Micronesia (FSM), theCommonwealth of the Northern Mariana Islands (CNMl), Guam, and the Republic of
Palau (Palau). This region, composed of thousands of tiny islands widely spread between
the latitudes of 15° N to 14° S and the longitudes of 134° E to 170° W, extends across anarea as large as the continental United States. With less than 2558 km2 in total land massand extended Exclusive Economic Zone (EEZ), the primary source of dietary protein for
Pacific islanders has come from the Ocean. The per capita seafood consumption exceeded
global average. In 2015, per capita seafood consumption in Oceania was 25.0 kg/year vs15.5 kg/year worldwide (FAO 2018).
Seafood comes from capture fisheries and aquaculture but capture fisheries are themain seafood source for the U.S.A. PI. Climate change and overfishing are two major
threats to the sustainability of yield from capture fisheries. Although there were optimistic
reports on status of tuna stocks in the Pacific, foreign fishing companies have exported
majority of their catch to consumers outside the islands. Islanders have relied on
subsistence catch to meet their demand. As the harvest from nearshore fishery declined,
they have to consider secure their seafood supplies via aquaculture. However, the islandsof the Pacific is still the least developed region in terms of aquaculture worldwide asconcluded at 2010 FAO Global Conference on Aquaculture in Thailand. On the other
hand, this region enjoys superior natural resources, such as pristine water, year-roundwarm weather, and isolated condition for disease prevention. However, it has to overcome
constraints to aquaculture development such as small land area, natural hazards for someislands, lack of knowledge base, short of available capital, distant markets, and poortransportation systems. Other than natural hazards for some islands, the other constraints
are solvable. A good strategic development plan is essential to reveal aquaculture
potential in the region.
This report discusses the potential threat of climate change to fisheries, reviews thecurrent status and challenges of aquaculture, and finally presents some suggestions on
future development.
Annotated Bibliography of Key Works
17

Adams, T., Bell, J. and Labrosse, P. 2001. Current status of aquaculture in the PacificIslands. In: R.P. Subasinghe, P. Bueno, M.J. Phillips, C. Hough, S.E. McGladdery & J.R.
Arthur, eds. Aquaculture in the Third Millennium. Technical Proceedings of theConference on Aquaculture in the Third Millennium, Bangkok, Thailand, 20-25 February
2000. pp. 295-305. NACA, Bangkok and FAO, Rome.
In the book “Aquaculture in the Third Millennium”, this chapter “Current status ofaquaculture in the Pacific Islands” gave an overview the status of aquaculture in the
Pacific Islands at the turn of 20th century. It served as the base to assess any new
development in 21st century. Through the new wave of international cooperation, it isexpected to increase sustainable use of aquatic resources to meet the goal of foodsecurity. Food and Agriculture Organization of the UN (FAO) and Network of
Aquaculture Centres in Asia-Pacific (NACA) co-organized this conference in 2000.
Lee, C.S. and Awaya, K. 2003. Viable aquaculture development in the U.S. affiliatedislands – lessons from giant clam and sponge farming. Aquaculture Economics &Management 7 (1&2):125-135.
This paper reviewed the farming technology for giant clam and bath sponges. Then, it
used giant clam and bath sponges as an example to discuss the challenges of technology
transfer, which include biological, technological, environmental, and socioeconomic, toPacific Islands. Finally, some recommendation was made for the successful technology
transfer.
FAO. 2018. The State of World Fisheries and Aquaculture 2018 - Meeting the sustainable
development goals. Rome. 227pp.
An important of aquaculture and fisheries status report published by FAO every two
years. It highlights the critical importance of fisheries and aquaculture for the food,nutrition and employment of millions of people, many of whom struggle to maintainreasonable livelihoods. Data and graphics presented in this publication are widely usedby research groups to assess the progress and to propose future works.
Bell, J.D., Johnson, J.E., Ganachaud A.S., Gehrke, P.C., Hobday A.J., Hoegh-Guldberg,
O., Le Borgne, R., Lehodey, P., Lough, J.M., Pickering, T., Pratchett, M.S. and Waycott,M. 2011. Vulnerability of Tropical Pacific Fisheries and Aquaculture to Climate Change:
Summary for Pacific Island Countries and Territories. Secretariat of the PacificCommunity, Noumea, New Caledonia. 386pp.
The book entitled Vulnerability of Tropical Pacific Fisheries and Aquaculture to Climatechange provides the region with the understanding needed, and the adaptations, policies
and investments recommended to reduce the likely impacts of climate change on
fisheries and aquaculture. It also gives the sector a roadmap for capitalising on theopportunities expected to arise from the changing climate. This book is the product of a18

partnership that started between the Australian Agency for International Development(AusAID) and the Secretariat of the Pacific Community (SPC), and then grew toembrace contributions from 36 institutions. This summary, which is a companion to thebook, provides this vital information in an accessible form for each Pacific Island
country and territory. It is a quick access to understand projected changes to surface
climate and the ocean, to oceanic fisheries, to coastal fisheries, to freshwater andestuarine fisheries, to aquaculture, economic and social implications, and adaptations
and suggested policies. It is a valuable document to have a quick overview of potential
climate change impacts and suggested remedy policy.
Charlton, K. E., Russell, J., Gorman, E., Hanich, Q., Delisle, A., Campbell, B., and Bell,J. 2016. Fish, food security and health in Pacific Island countries and territories: asystematic literature review. BioMedCentrak Public Health 16: 285
This paper discussed the importance of fish to Pacific Island Countries and territories(PICTs) in both food security and health related concerns based on the review of 29
studies. However, there is a paucity of research aimed at assessing how maintaining
and/or improving fish consumption benefits the diets and health of Pacific Islanders.
Instead of fresh seafood, there is an increasing demand for packaged imported foods,
such as canned meats, instant noodles, cereals, rice, and sugar-sweetened beverages,
with subsequent decreased consumption of locally-produced plants and animals.
Gillett, R., and Tauati, M. I. 2018. Fisheries in the Pacific Regional and national
information. FAO Fisheries and Aquaculture Technical Paper No. 625. Apia, FAO. 401pp.
This paper discusses the important species, the status of the resources, and the fisheries
management under offshore fishing and coastal (or nearshore) fishing. This report also
provides information on the fisheries in each of the 14 independent Pacific Island
countries (including Federated States of Micronesia, Marshall Islands, and Palau) in thefollowing categories:• Overview and main indicators• Production sector• Post-harvest sector• Socio-economic contribution of the fishery sector• Trends, issues and development• Institutional framework• Legal framework

19

Towards effective coral community restoration for sustainable fisheryof a coral reef grouper Epinepherus ongus: implication of
ecosystem-based management
Atsushi Nanami
Research Center for Sub-tropical Fisheries, Seikai National Fisheries Research Institute,Japan Fisheries Research and Education Agency, Fukai-Ota 148, Ishigaki, Okinawa 9070451, Japan
E-mail: nanami at fra.affrc.go.jp
Spawning aggregation of coral reef fishes are very vulnerable to fishing since only
conspecific individuals gather at particular sites in restricted seasons and lunar phases.
Since capturing spawning aggregation has a significant negative impact for both local
stock and reproductive success, protection of the spawning aggregation has been a
recently urgent action in coral reefs all over the world. Furthermore, conservation of
habitat in non-spawning season is also essential for the species that form spawning
aggregation since habitat loss at their home ground would also lead to reduce their
population.
In the present presentation, I will focus on a coral reef grouper, Epinephelus ongus,that is a very important fishery species and forms spawning aggregations in an Okinawancoral reef and discuss effective coral community restoration to establish a sustainablefishery of the species. My previous studies have shown: (1) their home ground reachedseveral kilometers from the spawning ground; (2) their home range was very limited
around a coral colony in non-spawning period; (3) juveniles preferentially used corals
with fine structures (e.g. bottle-brushed acroporid corals), whereas adults were mainly
found at corals with coarse structures (e.g. massive corals and staghorn acroporid corals);(4) very precise returning ability was observed after spawning migration, i.e., the species
could return to the coral colonies that were used before the spawning migration.
These results suggest that: (1) conservation of coral community around the spawning
ground is indispensable; (2) coral species that is preferentially used by E. ongus should
be selected for coral community restoration; (3) the area that is several kilometers aroundthe spawning ground should be the proposed area for coral community restoration. Since
the spawning ground of the species has been already assigned as a marine protected area,
coral community restoration around the spawning ground would be useful to enhance theE. ongus stock.
Annotated Bibliography of Key Works
Almany GR et al. 2013. Dispersal of grouper larvae drives local resource sharing in acoral reef fishery. Current Biology 23: 626-630.
20

This study examined the dispersal distance of larvae of a coral reef grouper,
Plectropomus areolatus, from the spawning ground. The parentage analysis revealed
that 50% of the larvae settled within 14 km of the spawning ground. The noteworthy
point of the study is that the spawning ground protection would be effective at a local
scale (within several-tens of kilometers) for larval dispersal. The method described in
the present study is greatly helpful to test the actual manner of larval dispersal in mystudy site.
Nanami A et al. 2013. Microhabitat association in white-streaked grouper Epinephelus
ongus: importance of Acropora spp. Marine Biology 160: 1511-1517.
Microhabitat associations are considered to be important for juvenile survivorship andgrowth of coral reef fishes. The aim of this study was to quantify microhabitat
associations for juvenile and adult white-streaked grouper Epinephelus ongus.
Underwater observations revealed that most juveniles were found in bottlebrush
Acropora spp., staghorn Acropora spp. and coral rubble and there was a significant
positive use of bottlebrush Acropora spp. and a significant negative use of coral rubble.
For adults, most individuals were found in bottlebrush Acropora spp. and staghorn
Acropora spp., and there was a significant positive use of staghorn Acropora spp. andsignificant negative use of coral rubble. A habitat choice experiment by using presettlement individuals revealed that both bottlebrush Acropora spp. and staghorn
Acropora spp. were used as settlement sites, whereas coral rubble was rarely used as asettlement site. Field results of the study suggest that juvenile and adult E. ongus
showed significantly positive microhabitat associations with bottlebrush Acropora spp.
and staghorn Acropora spp., respectively. Bottlebrush Acropora spp. has smaller
interbranch spaces than staghorn Acropora spp., which could drive patterns of
microhabitat associations. In addition, post-settlement processes such as predation may
influence the spatial distribution of juveniles.
Nanami A et al. 2015. Estimation of spawning migration distance of the white-streaked
grouper (Epinephelus ongus) in an Okinawan coral reef system using conventional tagand-release. Environmental Biology of Fishes 98: 1387-1397.
The white-streaked grouper (Epinephelus ongus) is an important fisheries species in theregion around Okinawa. This species forms spawning aggregations at specific spawning
grounds in the last quarter of the moon in April and/or May. Using a tag-and releasemethod, we estimated the migration distance and the degree of unified movement ofgroupers associated with the spawning migration. In total, 1157 E. ongus individuals
were tagged and released at their home grounds in the non-spawning period and 350
were tagged at a known spawning ground (Yonara Channel) in the spawning period. For
the fish that were released at the home grounds, 23 individuals were recaptured at the
spawning ground during the spawning periods. For the individuals that were released at21

the spawning ground, six individuals were recaptured outside of the spawning ground.
The estimated migration distances from the home ground to the spawning ground
ranged from 2.2 to 8.8 km.
Nanami A et al. 2017. Spawning aggregation of white-streaked grouper Epinephelus
ongus: spatial distribution and annual variation in the fish density within aspawning ground. PeerJ 5: e3000
The aims of the present study were to investigate the ecological characteristics of annual
spawning aggregations such as 1) spatial variations in the density of E. ongus at the
spawning ground, 2) the relationship between fish density and environmental variables,3) inter-annual variations in the spawning aggregation, 4) the proportion of males tofemales at the spawning ground for several days pre and post-spawning and 5) the
relationship between male density and female density at the protected spawning ground,based on observations over five years at an Okinawan coral reef. Although the protected
spawning ground area was large (ca. 2500 m  700 m), high density of E. ongus (over
25 individuals per 100 m2) was found in a limited area (within c.a. 750 m  50 m).
Current velocity and coverage of rocks had significant positive effects on the spatial
distribution of E. ongus at the spawning ground. Inter-annual variation in the degree of
aggregation was found and this variation was explained by the annual variation of mean
seawater temperature 40 days before the spawning day. The male-female ratio (male :female) at the spawning ground was ca. 3 : 1 for three years (May 2012, May 2014 andMay 2015) whereas >13 : 1 for one year (May 2013). Significant positive relationships
between male density and female density were found at the aggregation sites. It issuggested that E. ongus use aggregation sites with greater current velocity to reduce therisk of egg predation and seawater temperature is one of the main factors that is
responsible for determining the degree of aggregation. It is also suggested that females
possibly select sites with a greater density of males and this selection behavior might be
the reason why females arrived at the spawning ground after the arrival of the males.
For effective management of spawning grounds, precise site selection as well as theduration of the protection period are suggested to be key aspects to protect the spawning
aggregations of E. ongus, which have been currently achieved at the spawning ground.
Nanami A et al. 2018. Diel variation in home range size and precise returning ability
after spawning migration of a coral reef grouper Epinephelus ongus: implications for
effective marine protected area design. Marine Ecology Progress Series 606: 119-132.
Marine protected areas (MPAs) are regarded as effective tools for protection of marineorganisms. Precise estimation of home range size as well as diel differences in homeranges are essential to consider appropriate MPA size. In addition, for setting MPAs,
behavioral characteristics of the spawning migration should also be considered forspecies that form spawning aggregations. The purpose of the present study is to clarify
22the diel variation in home range size and the degree of precision for returning ability ofwhite-streaked grouper Epinephelus onus by acoustic telemetry. Seventeen individuals
were studied, and night-time home range sizes that were calculated by 50% and 95%
kernel utilization distributions were 5.9-times and 5.5-times greater than the respective
daytime home ranges. The average inter-center distance of home ranges between the
two time periods ranged from 3.0 m to 67.9 m (22.5 m on average), suggesting that daynight home range shift in the home ground varied individually. Returning ability for 10
individuals that showed clear spawning migration behaviour was also analysed andaverage inter-center distance of home ranges between the period before and after the
spawning date was 8.1 m. Eight out of the 10 individuals showed precise returning after
the spawning migration to the patchy coral substrates that were used before thespawning migration. The present study suggests that appropriate setting position of thehome ground can establish long-term protection of the species due to their precise
returning ability after the spawning migration.

23

Scaling up coral restoration to meet the demands ofa collapsing ecosystem
Tali Vardi
ECS for NOAA Fisheries Office of Science & Technology, 1315 East-West Highway,
Silver Spring Maryland, USA
E-mail: tali.vardi at noaa.gov
Coral reefs feed millions of people on earth and are critical to the livelihoods ofmillions more. They house approximately a quarter of the ocean’s biodiversity and are
critical to fisheries worldwide. Their importance and fragility to humans cannot be
overstated. According to climate models, in about thirty years half of all coral reefs are
predicted to disappear due to bleaching caused by unprecedented ocean warming. If
political and economic forces begin to seriously address emissions, corals will still needhelp maintaining enough abundance and diversity to rebuild reefs by the end of the
century when ocean temperatures may begin to stabilize. With the tacit assumption thatemissions will be curbed, an explosion of coral restoration and research into coral“interventions” has taken place over the past two years. Coral aquaculture is a pivot pointof any advancement in scaling-up restoration or implementing a new intervention - as any
and all applications involve the delicate care and propagation of corals in the water or on
land. NOAA has taken a lead role in coordinating coral restoration efforts throughout the
US and the globe, primarily via the Coral Restoration Consortium and by commissioning
the National Academies of Sciences to review coral interventions. I will highlight recent
restoration and intervention successes in the U.S. and globally and describe NOAA’s
proposed research and action plan on coral interventions. I will also describe my own
research on coral population modelling, highlighting how results can be incorporated into
decision-making processes that help managers and practitioners implement and increase
the success and scale of coral restoration programs.
Annotated Bibliography of Key WorksNational Academies of Sciences, Engineering, and Medicine. 2019. A Research Review
of Interventions to Increase the Persistence and Resilience of Coral Reefs. Washington,
DC: The National Academies Press AND National Academies of Sciences, Engineering,and Medicine. 2019. A Decision Framework for Interventions to Increase the Persistence
and Resilience of Coral Reefs. Washington, DC: The National Academies Press.
NOAA commissioned the U.S. National Academies to review the science behind next
generation coral restoration techniques. Most of these techniques rely on coralaquaculture in one form or another. The Research Review groups 23 interventions types(e.g. assisted evolution, marine shading to prevent bleaching, genetic engineering) into24

four categories and describes the benefits, potential scale of application, current
feasibility, risks, limitations, and infrastructure needs for each. The Decision Frameworkoutlines an adaptive management strategy by which interventions can be evaluatedagainst each other, additional research needs, and recommendations for the Caribbean.
Research on coral interventions is progressing very rapidly and these reports thoroughly
capture the current state of the science and make some solid recommendations about
how the field should move forward in the near future.
Mary Hagedorn, Christopher A. Page, Keri O’Neil, Daisy M. Flores, Lucas Tichy, Valérie
F. Chamberland, Claire Lager, Nikolas Zuchowicz, Kathryn Lohr, Harvey Blackburn, Tali
Vardi, Jennifer Moore, Tom Moore, Mark J. A. Vermeij, Kristen L. Marhaver. 2018.
Successful Demonstration of Assisted Gene Flow in the Threatened Coral Acropora
Palmata Across Genetically-Isolated Caribbean Populations using Cryopreserved Sperm.
BioRxiv.
Rescuing genetically depauperate coral populations by increasing genetic diversity fromnearby populations, is one of the most feasible coral interventions proposed. This study
demonstrated for the first time that viable juveniles of an endangered coral can becreated by artificially inseminating cryo-preserved sperm from one population, witheggs of another regionally-distinct population. This project highlighted the importanceand need for additional expertise in coral larval rearing and early life-stage aquaculture.
Currently, there are only a handful of such experts worldwide. As corals continue to bethreatened with extinction primarily due to climate change, these techniques will
become increasingly necessary for the long-term persistence of coral reefs which feed
hundreds of millions of people and provide habitat to 25% of marine fisheries globally.
Darling, E. S., McClanahan, T. R., Maina, J., Gurney, G. G., Graham, N. A., JanuchowskiHartley, F., ... & Puotinen, M. (2019). Social–environmental drivers inform strategic
management of coral reefs in the Anthropocene. Nature Ecology & Evolution.
Data from more than 2500 reefs across the Indo-Pacific Ocean were analyzed todelineate key drivers and suggest appropriate management at three different levels:
protection, recovery, and transformation. Over 50% of surveyed reefs would benefit
from recovery which includes some form of restoration in addition to traditional
management, and, most importantly, reduction in ocean warming. Using comprehensive
studies like this and the Global Coral Reef Monitoring Network’s reports, should help
nations prioritize coral research, restoration, and management efforts.
Iliana B. Baums, Andrew C. Baker, Sarah W. Davies Andréa G. Grottoli, Carly D. Kenke,lSheila A. Kitchen, Ilsa B. Kuffner, Todd C. LaJeunesse, Mikhail V. Matz, Margaret W.
Miller, John E. Parkinson, Andrew A. Shant. 2019. Considerations for maximizing theadaptive potential of restored coral populations in the western Atlantic. Ecological25

Applications.
This paper is a succinct guide to maximizing genetic diversity using various coralaquaculture techniques for restoration. Despite differences in species and environmental
conditions, restoration is happening throughout the globe. Although this paper uses a
Caribbean example, the recommendations serve as a helpful starting point for coralaquaculture and restoration elsewhere. The paper explicitly recommends the number ofgenets per area that should be planted to maintain or maximize diversity. Because
bleaching, coral gardening, and assisted evolution have the potential to reduce genetic
diversity, having clear guidelines that managers and practitioners can understand isparamount for proper aquaculture and coral reef restoration techniques.

26

Sustainable large-scale coral restoration by establishing “artificial
spawning hotspots”
Go Suzuki
Research Center for Subtropical Fisheries, Seikai National Fisheries Research Institute,Ishigaki, Okinawa, 907-0451, Japan
Email: gosuzu at fra.affrc.go.jp
Coral reefs have degraded over time and, recently, severe bleaching events during
2015-2017 caused further damage to coral communities worldwide. Because coral reefs
are an important source of coastal fish and invertebrates in tropical coasts, these fisheries
resources are decreasing as well. For sustainable use of these resources, therefore, it isimportant to preserve coral communities that act as fisheries grounds and nurseries. Coraltransplantation is known as a positive restoration method. However, large-scaletransplantation requires a great deal of labor and, even if it succeeds once, the transplanted
corals are vulnerable to extermination by only a single disturbance such as bleaching oran outbreak of crown-of-thorns starfish (COTS). Hence, we need to enhance annual larval
recruitment for sustainable large-scale coral restoration.
Two key factors are crucial for sustainable coral restoration by enhancing
reproduction. The first factor is establishment and maintenance of “artificial spawning
hotspots” that consist of densely populated conspecific adult colonies. The second factor
is improvement of initial survivability by collecting eggs and sperms at spawning andrearing larvae until settlement. For “artificial spawning hotspots”, safeguards are required
against predation by COTS and the use of shading is desirable against bleaching during
the high seawater temperature season. In addition, a special gamete and larval collector,“larval cradle”, was developed to consistently perform bundle collection, fertilization,and larval rearing. Eventually, we aim to develop simple techniques that can be easily
handled by any users such as fishermen and leisure divers.
Annotated Bibliography of Key Works
Zayasu Y, Suzuki G (2019) Comparisons of population density and genetic diversity inartificial and wild populations of an arborescent coral, Acropora yongei: implications forthe efficacy of “artificial spawning hotspots”. Restoration Ecology: 27: 440-446
The authors assessed population density and genetic diversity of a wild, arborescentcoral, Acropora yongei, and compared these parameters with those of an artificiallyestablished A. yongei population in the field. The population density of wild arborescentcorals was only 0.27% of that in the artificial population, even in a high-coverage area.
Genetic diversity was also low in the wild population compared with the artificial
population, and approximately 10% of all wild colonies were clones. Based on these27results, the larval supply in the artificial population was estimated to be at least 1,400
times higher than that in wild A. yongei populations for the same area of adult
population.
Suzuki G, Yamashita H, Kai S, et al. (2013) Early uptake of specific symbionts enhances
the post-settlement survival of Acropora corals. Marine Ecology Progress Series 494:
149-158.
The authors tested the hypothesis that early acquisition of symbionts enhances postsettlement survival. Symbiotic and aposymbiotic Acropora larvae were prepared in thelaboratory and settled on experimental plates in the field. The survival of settlers wasmonitored for 15 months and the results showed that more larval-stage settlers
harbouring symbionts survived than those without. The higher survival rate of ‘early
uptake’ corals was more pronounced on shaded plates. These results suggest that theearly uptake of specific symbionts enhances post-settlement survival in dark places suchas reef crevices, which are sites commonly settled by coral larvae.
Suzuki G, Kai S, Yamashita H, et al. (2011) Narrower grid structure of artificial reefenhances initial survival of in situ settled coral. Marine Pollution Bulletin 62: 2803-2812.
The authors demonstrated through field experiments that the design of artificial gridplates may influence the initial survival of Acropora corals, with narrower grids being
the most effective. In fact, grid plates with a 2.5-cm mesh presented the highest
recorded survival rate (14%) at 6 months after settlement (representing approximately
50 corals per 0.25 m2 of plate). This was the first study where such high survival rates,
matching those of cultures under aquarium conditions, were obtained in the field
without using additional protective measures, such as guard nets against fish grazing
after seeding. Therefore, their results provide a foundation for establishing new andeffective coral restoration techniques for larval seeding.

28

The influence of climate and environment on the growth and survivalof Pacific oyster seed in US west coast estuaries
Brett Dumbauld1*, Evan Durland2, Konstantin Divilov2, Kelly Muething2, Anna Bolm2,
Ylva Durland2 and Chris Langdon2
1

US Department of Agriculture, Agriculture Research Service, Hatfield Marine Science
Center, Newport, OR 97365, USA
2
Oregon State University, Fisheries and Wildlife Dept., Hatfield Marine Science Center,
Newport, OR 97365, USA*E-mail: brett.dumbauld at usda.gov
Pacific oysters Crassostrea gigas were introduced to the US west coast in the early
1900’s and were initially raised directly from seed (juveniles that had set naturally on
cultch shell) imported from Japan. Oysters regularly spawned and became “naturalized”
in only several selected West coast locations, such as Willapa Bay, Washington, whereconditions allowed for both adult oyster spawning and larval survival, retention andsettlement. The shellfish industry relied on “natural or wild” caught seed from theselocations or continued seed imports until the advent of hatchery technology in the late1970’s. Recent larval mortality events in hatcheries have been linked directly to changes
in seawater chemistry with high pCO2 conditions and acidified water associated withseasonal upwelling along the U.S. West Coast. These conditions may have also resulted
in reduced natural oyster sets in Willapa Bay, but estuarine gradients in water chemistry
and temperature add complexity making this more difficult to assess. Shellfish hatcheries
have adapted to these conditions by measuring seawater carbonate chemistry, buffering
incoming water, and adjusting the timing of larval production cycles. While there appear
to be larval fitness traits that are genotype-dependent, the potential of breeding programs
to improve OA specific traits remains uncertain. Experiments have rarely been conductedthat distinguish success at important physiological transitions in the larval life history andpotentially also at the juvenile seed stage when these oysters are out-planted to estuaries
with variable conditions.
We present results from experiments where survival of oyster larvae raised fromcrosses of “wild” parental broodstock collected in Willapa Bay was compared to that oflarvae raised from controlled crosses with improved lines created by the Molluscan
Broodstock Program (MBP) at Oregon State University. The MBP breeding program wasdesigned to enhance valuable field traits including growth and survival of juvenile andadult oysters, but to date has not explicitly addressed larval traits. Nonetheless, MBP
larvae produced more than twice the number of settled spat compared with wild larvae
under commercial hatchery conditions (with buffered seawater, pH ~8.3 and arag >2).
This advantage also occurred, but to a lesser degree, under unfavorable high pCO2)
29

conditions. Separate experiments were conducted to evaluate survival of juvenile seedalong the estuarine gradient and inside and outside eelgrass (Zostera marina) which also
has the potential to modify local seawater chemistry. Growth and survival of juvenile
oyster seed varied along the estuarine gradient with fastest growth, but lower survival
occurring near the estuary mouth and there was no apparent effect of seed source. Resultsfrom experiments where MBP seed was planted both inside and outside eelgrass along
these same gradients, suggest that this plant can reduce seed growth, especially at
locations away from the estuary mouth.
Annotated Bibliography of Key Works
Barton, A., G.G. Waldbusser, R.A. Feely, S.B. Weisberg, J.A. Newton, B. Hales, S. Cudd,
B. Eudeline, C.J. Langdon, I. Jefferds, T. King, A. Suhrbier, and K. McLaughlin. 2015.
Impacts of Coastal Acidification on the Pacific Northwest Shellfish Industry and
Adaptation Strategies Implemented in Response. Oceanography 28: 146-159.
This is a review of the history and science underpinning the effects of changing
seawater chemistry on bivalve shellfish larvae and the impacts that have already takenplace to the commercial shellfish aquaculture industry on the US West Coast. Multiple
authors contributed to this review which addresses a broad audience but covers theleading research on direct effects to bivalve larvae as well as monitoring seawaterconditions and adapting to these changes.
Durland, E., G. Waldbusser, and C. Langdon. 2019. Comparison of larval development
in domesticated and naturalized stocks of the Pacific oyster Crassostrea gigas exposed tohigh pCO2 conditions. Marine Ecology Progress Series 621: 107-125.
Two replicated experiments were conducted where Pacific oyster larvae from “wild”
parent crosses and those from selected stocks (OSU Molluscan Broodstock Program)
were raised and set under favorable hatchery conditions (buffered seawater, pH ~7.8 andarag ~2) and unfavorable (pH ~7.4 and arag ~1) conditions. Early larval development
was inhibited by acidified seawater and it affected the timing, but not magnitude oflarval mortality. The effect on metamorphosis (setting) was variable, but MBP larvae
produced more and larger spat in ambient and high pCO2 seawater, respectively.
Hales, B., A. Suhrbier, G.G. Waldbusser, R.A. Feely, and J.A. Newton. 2017. The
carbonate chemistry of the "Fattening Line," Willapa Bay, 2011-2014. Estuaries andCoasts 40: 173-186.
The authors present detailed data on seawater chemistry (especially PCO2 and aragonite
saturation state) for Willapa Bay, Washington where Pacific oysters have been themainstay of the oyster aquaculture industry for almost 100 years and there is a long term
record of spawning and setting. They reconstruct this record for a longer historicalperiod and their data suggest that recent conditions provide a smaller window of optimal
30

conditions (low aragonite saturation state and warm enough temperatures for oyster
spawning) than occurred historically. While they did not sample larvae and therefore
cannot confirm effects, they substantiate the complexity of measuring these effects andattributing them to a single cause in an estuary where conditions are variable.
Lowe, A.T., J. Kobelt, M. Horwith, and J. Ruesink. 2019. Ability of eelgrass to alter oyster
growth and physiology is spatially limited and offset by increasing predation risk.
Estuaries and Coasts 42: 743-754.
The authors planted juvenile seed oysters and measured water chemistry inside andoutside of eelgrass (Zostera marina) at several locations in Willapa Bay and in the
Salish Sea in Washington state. They demonstrate that while eelgrass modified
carbonate chemistry, it did not appear to be the primary variable influencing juvenile
oyster growth and survival and, instead, predation was higher and thus survival lower
inside eelgrass beds where surviving oysters also grew slower.
Ruesink, J.L., G.C. Roegner, B.R. Dumbauld, J.A. Newton, and D.A. Armstrong. 2003.
Contributions of coastal and watershed energy sources to secondary production in a
Northeastern Pacific estuary. Estuaries 26: 1079-1093.
These authors measured growth of juvenile Pacific oyster seed deployed at numerous
locations throughout Willapa Bay and used stable isotopes to distinguish marine versus
terrestrial energy sources. Results demonstrated a very distinct along estuary gradientwith oysters growing faster near the mouth and slower at greater distance from themouth and away from the mouth. Oysters also displayed distinct stable isotope ratiosthat reflected a strong marine signal near the mouth (-18 δ13C) and a stronger terrestrial
signature at distance (-22 δ13C). Oysters grown just off the bottom grew slower thanthose off-bottom at any given tidal elevation.

31

Comparative study of the impact of environmental changes on oyster
culture between USA and Japan, as collaborative research under
UJNR
Natsuki Hasegawa*1, Brett R. Dumbauld2, Masakazu Hori3, Satoshi Watanabe4,
Michael Rust5, and Zachary Forster6
1

Hokkaido National Fisheries Research Institute, Japan Fisheries Research and Education
Agency, Kushiro Laboratory: 116 Katsurakoi, Kushiro, Hokkaido, 085-0802, Japan
2
USDA-ARS, Hatfield Marine Science Center, Newport, OR 97365, USA
3
National Research Institute of Fisheries and Environment of Inland Sea, Fisheries
Research Agency, 2-17-5, Maruishi, Hatsukaichi, Hiroshima, 739-0452, Japan
4
National Research Institute of Aquaculture, Japan Fisheries Research and Education
Agency, 422-1 Nakatsuhamaura, Minamiise, Mie, 516-0193, Japan
5
NOAA Office of Aquaculture, 1315 East-West Highway, Silver Spring MD. 20910,
USA
6
Washington Dept. of Fish and Wildlife, Ocean Park, WA. 98640, USA*E-mail: hasena at affrc.go.jp
Several oyster species are cultured globally, and the Pacific oyster, Crassostrea gigas,
is the most widely cultured species both in Japan and the USA. In Japan, aquaculture
production of the Pacific oyster is decreasing slightly for multiple reasons including
large die-offs of adults during and after the reproductive season due to a delay in the
reproductive season and poor post-spawning recovery, poor wild spat collection, and alabor shortage for both operation of aquaculture and post-harvest processing. These
problems may be aggravated by environmental changes such as global warming andoligotrophication around Japan’s coastal areas. FRA is investigating the causes of the dieoff during the reproductive season and attempting to establish countermeasures. The
Pacific oyster is native to Japan but was introduced to the USA for aquaculture in theearly 1900’s. Aquaculture production in the USA has now exceeded that in Japan.
Nonetheless there are large differences in the culture system, habitats and environmental
conditions between the two countries. A comparative study was initiated to evaluate
oyster reproduction in the two countries in order to understand the effects of habitat andenvironment on future success of aquaculture given predicted environmental change.
Oyster culture experiments were conducted in intertidal and subtidal zones just inside andoutside seagrass habitat in Hiroshima Bay in Japan and Willapa Bay in USA during the
reproductive season (March to June 2019 and February to July 2019, respectively). We
focused on elucidating the effects of habitat and environment on the energy allocation of
the oyster between reproduction and somatic growth.
32

Annotated Bibliography of Key Works
Akashige, A., Hrata, Y., Takatsuji, Y., Aida, S. 2006. Occurrence of mass mortality inoyster culture with relation to seawater temperature and rainfalls in Hiroshima. Bulletinof the Hiroshima Pref. Fisheries & Marine Technology Center 1: 9-13.
Relationships between mass mortality in Pacific oyster, Crassostrea gigas, occurred in
1979, 1994, 2001 and 2002 and surface water temperature or rainfall in Hiroshima Bay
were analyzed using a dataset from 1970 to 2004. Mass mortality occurred in years withhigh water temperature (i.e. the number of days over 20℃ was greater than the mean +SD) and small rainfall (i.e. cumulative total rainfall from July to October was smaller
than the mean - SD) except in 2001. Extraordinary mass mortality of spat occurred in2001 which were introduced from Miyagi Prefecture which has colder climate than
Hiroshima Prefecture. Therefore, there is a high risk of mass mortality in local oyster inyears with high water temperature and small rainfall.
Akashige, A., Hrata, Y., Takayama, K., Sorammoto, K. 2005. Seasonal change in oxygen
consumption rates and filtration rates of the cultured Pacific oyster Crassostrea gigas,
Nippon Suisan Gakkaishi 71(5): 762-767.
The oxygen consumption rate (OCR) and filtration rate (FR) of cultured Pacific oyster,
Crassostrea gigas, of different sizes were measured in still water systems at different
seawater temperature in various seasons. The OCR had clear relationships with watertemperature (t, ℃) and dry body flesh weight (Wd g) expressed by the formula: OCR(mg O2 / hr / ind) = (0.072t - 0.64) Wd0.75 and there was no difference among seasons.
The FR was expressed by the formula: FR (L / h / ind) = (0.70t - 6.6) Wd in nonspawning seasons. However, in the spawning season, the FR were expressed by theformula: FR = 4.9 Wd, which was lower than the value expected by the formula applied
on non-spawning season. These results clearly showed that feeding activities dropped
during the spawning season, but oxygen demand (weight base and temperaturedependent) was constant throughout the year. Oysters would be especially vulnerable
during the spawning season when metabolic activities would be devoted to gamete
production.
Cheney, D.P., B.F. MacDonald, and R.A. Elston. 2000. Summer mortality of Pacificoysters, Crassostrea gigas (Thunberg): Initial findings on multiple environmental
stressors in Puget Sound, Washington, 1998. Journal of Shellfish Research 19: 353-359.
Summer mortality has been a recognized problem in Pacific Coast oyster aquaculture on
the west coast of the USA since the mid 1950's, and these authors related some of theseincidences to stress during the reproductive season combined with environmental factors.
Nonetheless triploid oysters experienced higher mortality than diploids.

33

Oyster aquaculture using seagrass beds as a climate change
countermeasure
Masakazu Hori*1, Franck Lagarde2, Masami Hamaguchi1, Valerie Derolez2 andMarion Richard2
1

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
2
MARBEC Ifremer, IRD, Univ. Montpellier, CNRS, 34200 Sete, France*E-mail: mhori at affrc.go.jp
In the framework of the Sustainable Development Goals (SDGs) led by the United
Nations, it is required to develop coastal management methods to achieve bothsustainable food production and environmental conservation as a climate change
countermeasure. Oyster farming is an important food production method now being
developed in coastal areas around the world though, in certain instances, oyster farming
has caused local environmental degradation. As a typical example, local ecosystem
functioning of the coastal area has been often overused by oyster aquaculture at an
excessive oyster density derived from economic priority without consideration forecosystem sustainability. Recently, climate change has caused several negative effects on
oyster aquaculture such as poor spat collection due to oligotrophication, ocean
acidification, and poor spat growth and survival due to frequent anoxic events derived
from high seawater temperature. These effects gradually shifted the oyster farmers’
concern from high production aquaculture to sustainable aquaculture. In response to therecent consideration for environmentally friendly aquaculture, collaborative research
between France and Japan has started to achieve both sustainable oyster aquaculture andenvironmental conservation which aims for climate change mitigation and adaptation.
The oysters cultivated in many regions of the world are intertidal species inhabiting
intertidal zones such as sandy/muddy tidal flats and estuaries, where seagrass beds areoften distributed in adjacent subtidal areas. Seagrass vegetation is one of the mostimportant ecosystems functioning as a countermeasure for global climate change. Not
only does it mitigate green-house gas emission by sequestration and storage of blue
carbon derived from atmospheric CO2, but it also functions as an adaptation measure
utilizing the buffer function against ocean acidification and moreover as water quality
improvement.
Based on returning to aquaculture using the natural ecosystem interactions between
oysters and seagrass beds, our project is now demonstrating whether aquaculture
techniques that take into account both mitigation and adaptation to climate change are
effective for both sustainable use of coastal areas and coastal environmental conservation.
34

We will present the results from field experiments conducted in both the French
Mediterranean Sea and the Seto Inland Sea of Japan, to clarify the effect of eelgrass bedson (1) natural oyster spat collection and (2) growth and survival of oyster spat.
The results of our experiments revealed that spat recruitment was significantly higher in
the intertidal zone where there was no eelgrass distribution in the subtidal zone, while
spat growth and survival rate after settlement was significantly higher in eelgrass beds
even when anoxic events occurred. In addition, the organic carbon storage in the sediment
was larger in the eelgrass meadows with oyster aquaculture, suggesting that oyster
aquaculture using seagrass meadows would be a good practice for the climate change
adaptation and mitigation.
Annotated Bibliography of Key Works
Hori, M, Hamaoka, H, Hirota, M, Lagarde, F, Vaz, S, Hamaguchi, M, Hori, J, Makino, M.
2018. Application of the coastal ecosystem complex concept toward integrated
management for sustainable coastal fisheries under oligotrophication. Fisheries Science
84 (2): 283-292.
Harmonizing coastal fisheries with water-quality improvement has become an essential
factor for the sustainable use of coastal ecosystem services. Here, we present the scopeof our study based on an interdisciplinary approach including ecological actions, socioeconomic actions and socio-psychological actions. We chose to focus on the interaction
between oyster aquaculture and seagrass vegetation as a typical ecological action using
the coastal ecosystem complex (CEC) concept. Coastal organisms have adapted theirtraits to the environment over a long period of time, so that restoration of the CEC
represents reconstruction of the original process of coastal production. Subtidal seagrass
vegetation with intertidal oyster reefs is the original CEC in Japan, which would be
expected to enhance coastal production by improving the production efficiency without
adding nutrients. A simple field experiment examining carbon and nitrogen contents andstable isotope ratios revealed that oyster spats cultivated on a tidal flat adjacent toseagrass beds had higher nitrogen contents and higher δ13C ratios than spats cultivatedin an offshore area using only pelagic production. This result suggests that utilization of
the CEC, which enables oysters to use both pelagic and benthic production, has
potential to sustain a food provisioning service for humans, even in oligotrophic
conditions.
Lagarde, F, Richard, M, Bec, B, Roques, C, Mortreux, S, Bernard, I, Chiantell, C,
Messiaen, G, Nadalini, J, Hori, M, Hamaguchi, M, Pouvreau, S, d’Orbcastel, ER,
Trenblay, R. 2018. Strophic environments influence size at metamorphosis andrecruitment performance of Pacific oysters. Marine Ecology Progress Series 602: 135153.
Reproduction and recruitment of benthic invertebrates are influenced by the climate and35

by the ecological structure of marine ecosystems, along with local anthropogenicpressures such as eutrophication or oligotrophication. Using the Pacific oyster,
Crassostrea gigas, as a biological model, we tested the hypothesis that the variability inprodissoconch II (PII) size (i.e. size at metamorphosis) depends on ecological
functioning. Settlement and recruitment were assessed at 5 sampling sites in the French
Mediterranean shellfish farmed Thau lagoon during the main summer recruitment
events in 3 consecutive years (2012−2014). Hydrobiological and planktonic analyses
were conducted at 3 sampling sites. Our results showed that recruitment was extremely
heterogeneous, ranging from 0 to 260 ± 27 SE ind. dm−2 throughout the ecosystem andwas linked with variability in PII size, which ranged from 180 to 296 μm. The annual
temporal pattern of PII sizes appeared to be controlled by temperature during thesettlement period, whereas the spatial pattern depended on phytoplankton biomass andon the trophic functioning of the ecosystem. Smaller PII sizes were significantly
correlated with the highest phytoplankton biomass, while larger PII sizes werepositively correlated with mixotrophic cryptophyte abundance. We found an inverse
relationship between PII size and survival after metamorphosis, showing that
recruitment success was associated with smaller PII sizes. Regional climate conditions
and local trophic functioning appear to be key factors in metamorphosis andconsequently contribute to recruitment heterogeneity. Further studies should be
performed in other ecosystems following an oligotrophication trajectory to generalize
this result.

36

Kelp, Saccharina spp, population genetics in the Northwest Atlantic for
guiding a breeding program of thermally resilient strains
Simona Augyte*1,2, Jean-Luc Jannink3, Xiaowei Mao3,4, Kelly Robbins3, Matt Hare5,
Schery Umanzor1, Michael Marty-Rivera1, Scott Lindell6, David Bailey6, Charles
Yarish11

Dept. of Ecology and Evolutionary Biology, University of Connecticut, 1 University
Place, Stamford, CT 06901, USA2
Symbrosia Inc., 73-984 Makako Bay Drive, Kailua-Kona, HI 96740, USA
3
Plant Breeding and Genetics Section, Cornell University, Ithaca, NY 14853, USA
4
Key Laboratory of Vertebrate Evolution and Human Origins of Chinese Academy of
Sciences, Institute of Vertebrate Paleontology and Paleoanthropology, Chinese Academyof Sciences, Beijing 100044, China
5
Department of Natural Resources, Cornell University, Ithaca, NY 14853, USA
6
Applied Ocean Physics and Engineering Department, Woods Hole Oceanographic
Institution, Woods Hole MA 02543, USA*E-mail: simona.augyte at uconn.edu
The cold-water sugar kelp, Saccharina latissima has a circumboreal distribution andin the Northwest Atlantic is at its southern distributional limits in Long Island Sound. An
understanding of genetic diversity of natural kelp populations is critical for making
recommendations for breeding and cultivation efforts of the growing seaweed aquaculture
sector in the US. An important component of the ARPA-E’s MARINER project isselectively breeding Saccharina spp. in order to improve overall productivity for biofuels,
feeds and food.
Historical records indicate the presence of regional kelp ecotypes based on
physiological tolerance, specifically temperature. We made collections of 13 wild
Saccharina spp. populations via SCUBA along the New England coast. These parental
populations were also used to make over 300 hybrid crosses that were planted at several
farm locations. We then used genome-wide single nucleotide polymorphism data to
explore the biogeographic diversity of the kelp.
An assessment of the sequence diversity revealed distinct genetic variation between
the Gulf of Maine and Southern New England, confirming that Cape Cod acts as a barrier
to kelp gene flow. Furthermore, based on the analysis of molecular variance (AMOVA),
we found the largest variance (58%) within sites. We also observed admixture among fivesub-populations and isolation by distance in the Gulf of Maine.
Future steps for this project include skim sequencing the haploid phase of the kelp
life cycle to identify trait heritability and phenotypic diversity observed for bothmorphological traits and tissue composition. Furthermore, we plan to place our sequence37

data into a larger context to include samples from sites in Europe and Asia.
Annotated Bibliography of Key Works
Langton, R., S. Augyte, N. Price, J. Forster, T. Noji, G. Grebe, A. St. Gelais and C. J.
Byron. 2019. An Ecosystem Approach to the Culture of Seaweed. NOAA Technical
Memo. NMFS-F/SPO-195, 24 pp. url: https://spo.nmfs.noaa.gov/content/techmemo/ecosystem-approach-culture-seaweed
Seaweeds are a significant component of current marine aquaculture production andwill play an increasing role in global food security as the human population increases
rapidly over the next 30 years. Seaweed farming is analogous to plant-based agriculture
except that the crop is cultured in a marine environment. It differs from agriculture inthat seaweeds do not require tillable land, fertilization or freshwater, which areresources that may ultimately constrain the expansion of agriculture. Seaweeds areconverted into a variety of goods, such as food and nutritional supplements for humans
and livestock, fertilizer, unique biochemical and biofuels. Wild and cultured seaweedalso offer multiple ecosystem services, such as bioremediation for coastal pollution,
localized control of ocean acidification, mitigation of climate change and habitat forother marine organisms. Incorporation of seaweeds into marine aquaculture farms in the
United States (U.S.) is, however, not without its challenges. Seaweed is an
unconventional food which necessitates establishing product acceptability, creating asustained market and then balancing demand with a consistent supply for long term
economic profitability. Seaweed farms also need to be developed in a manner that iscompatible with wild capture fisheries, marine mammal migrations and other users ofthe marine environment. A comprehensive understanding of the role that cultured
seaweeds play in the marine ecosystem is necessary in order to determine not only the
economic value of the goods produced but also the ecosystem services offered bymarine farming activities. This will result in a better understanding of how an ecosystemapproach to aquaculture incorporates the role and need for both the goods and services
these macroalgae will provide.
Augyte, S., Umanzor, S., Yarish, C., and S. Lindell. 2018. Enhancing marine ecosystemservices via kelp aquaculture. International Society of Applied Phycology News l.
December-2018;10–14.
https://docs.wixstatic.com/ugd/e9f50b_f3edc9d188f248a18061e09cfa25b5d9.pdf
For centuries, humans have harvested different seaweeds for food, medicinal purposes,
feed, and more recently as raw materials for industrial processes. Currently, wild
harvested seaweed account for less than 5% of the total worldwide supply (FAO 2018).
The majority of seaweed production is provided via aquaculture, with 99% of the
production taking place in Asia including China, Korea, Japan, Indonesia, and thePhilippines, worth US $11.7 billion annually (FAO 2018). Although seaweed38

aquaculture is a fast expanding industry, the global demand for seaweed-based products
is surpassing the supply. Such demands necessitate either domesticating new species orfurther expanding the productivity of the existing leading seaweeds. According to the
FAO, only a few species dominate seaweed farming, including two brown kelps,
Saccharina japonica and Undaria pinnatifida (Buschmann et al., 2018). To meet thepresent market requirements and to contribute in reducing the over-exploitation of wild
stocks, experimental trials worldwide have assessed the performance of newly farmed
seaweeds as potential products, particularly kelp species that have been traditionallyharvested from wild populations.
Augyte, S., C. Yarish, and C. D. Neefus. 2019. Thermal and light impacts on the early
growth stages of the kelp Saccharina angustissima (Laminariales, Phaeophyceae). Algae
34: 1-10. https://doi.org/10.4490/algae.2019.34.5.12.
Anthropogenic disturbances, including coastal habitat modification and climate change
are threatening the stability of kelp beds, one of the most diverse and productive marine
ecosystems. To test the effect of temperature and irradiance on the microscopic
gametophyte and juvenile sporophyte stages of the rare kelp, Saccharina angustissima,from Casco Bay, Maine, USA, we carried out two sets of experiments using atemperature gradient table. The first set of experiments combined temperatures between
7-18°C with irradiance at 20, 40, and 80 μmol photons m-2 s-1. The second set combined
temperatures of 3-13°C with irradiance of 10, 100, and 200 μmol photons m-2 s-1. Overtwo separate 4-week trials, in 2014 and again in 2015, we monitored gametogenesis, theearly growth stages of the gametophytes, and early sporophyte development of thiskelp. Gametophytes grew best at temperatures of 8-13°C at the lowest irradiance of 10μmol photons m-2 s-1. Light had a significant effect on both male and female
gametophyte growth only at the higher temperatures. Temperatures of 8-15°C andirradiance levels of 10-100 μmol photons m-2 s-1 were conditions for the highest
sporophyte growth. Sporophyte and male gametophyte growth were reduced at bothtemperature extremes—the hottest and coldest temperatures tested. S. angustissima is aunique kelp species known only from a very narrow geographic region along the coastof Maine, USA. The coupling of global warming with high light intensity effects mightpose stress on the early life-history stages of this kelp, although, as an intertidal species,
it could also be better adapted to temperature and light extremes than its subtidal
counterpart, Saccharina latissima.

39

Cell selection technique for establishment of low salinity tolerance
strain in Pyropia tenuipedalis
Mahiko Abe*1, Tomomi Ohashira1, Noboru Murase1 and Masanobu Kishioka2
1

National Fisheries University, FRA, Shimonoseki, Yamaguchi 759-6595, Japan
Yamaguchi Prefectural Fisheries Research Center, 2861-3, Senzaki, Nagato,
Yamaguchi, 759-4106, Japan*E-mail: abemahi at fish-u.ac.jp

2

Pyropia tenuipedalis is characterized by reddish thalli and direct budding from theshell substratum. The distribution of this species is limited to coastal areas in Seto InlandSea, Ise Bay and Tokyo Bay in Japan. In Yamaguchi Prefecture, this species has beenused for direct human consumption for its nutritional value. Alanine and glutamic acidcontents of P. tenuipedalis are three times and two times higher, respectively, than thoseof P. yezoensis which is commonly referred to as “nori”. Yamaguchi Prefectural Fisheries
Research Center began development of mariculture techniques for P. tenuipedalis in 2002
and, in 2007, they succeeded in commercial cultivation of this species. However,production levels have continued to decrease since 2012. More recently, extensive
disappearance of thallus has been observed from January to February since 2016. One of
the reasons for the production decrease is thought to be low salinity seawater due to recent
heavy rain fall supposedly associated with global climate change. The aim of this study
was to develop low salinity tolerance strains of P. tenuipedalis with cell selection
techniques. Disks of thallus were cultured in a 10% seawater medium diluted with
distilled water with 1/2SWM-III for one to four months. Cells that survived in thismedium were then cultured in a 100% seawater medium with 1/2SWM-III. Surviving
cells divided and regrew to thallus, and stock culture strains were established with selffertilization. A field culture trial of the established low salinity tolerance strain and theconventional strain was carried out in Koto River estuary from December 2018 to
February 2019. The low salinity tolerance strain showed better growth than theconventional strain, suggesting the efficacy of the cell selection technique for breeding.
However, some of the thalli growing on the culture plates suddenly shortened or
disappeared in early January 2019. Time-lapse observation with an underwater camera
revealed that the disappearance of the thalli was due to predation by blackhead seabream,
Acanthopagrus schlegelii. Immediate establishment of not only strains resistant to thechanging environment but also effective measures for predation are necessary to increase
production of P. tenuipedalis.
Annotated Bibliography of Key Works
Abe, M., Murase, N., Hatama, T., Shikano, Y., Kanai, T. 2015. New record of Pyropia
40

tenuipedalis from Koto river estuary, Yamaguchi prefecture. J. Nat. Fish. Univ. 63, 244248.
We collected reddish foliose thalli from Koto River estuary, Yamaguchi Prefecture. In
order to identify the species, we carried out ﬁeld collections of the foliose thalli,
morphological observation in culture and PCR-RFLP analysis using the two regions of
the partial mitochondrial DNA. As for the morphological survey, the spherical cells
were formed at the tips of conchocelis and developed to foliose thalli. The fragment
patterns of this species in the PCR-RFLP analysis matched with Pyropia tenuipedalis.
In the present study, the foliose thalli collected at Koto River were identified as P.
tenuipedalis, which is an endangered species distributed along the coast of Japan. This
species was newly recorded from Koto River estuary, Yamaguchi Prefecture.
Nakayama, T., Abe, M., Murase, N., Shikano, Y. 2017. Influence of salinity on growthof red alga Pyropia tenuipedalis and Pyropia yezoensis foliose thallus. Aquacult. Sci.
65, 321-330.
Pyropia tenuipedalis is a new culture species in Yamaguchi Prefecture, Japan. The
habitat of this species is more brackish as compared with that of P. yezoensis, which isthe common species in Japanese nori mariculture. In this paper, the authors investigated
the relationship between salinity and growth of P. tenuipedalis and P. yezoensis. It was
revealed that P. tenuipedalis tolerates lower salinity in comparison with P. yezoensis.
Abe, M., Murase, N., Hatama, T., Shikano, Y., Kanai, T. 2017. Environmental
characteristics of Pyropia tenuipedalis (Miura) Kikuchi et Miyata growing at
Yamaguchi Bay, Yamaguchi Prefecture. J. Nat. Fish. Univ. 65, 19-29.
This study surveyed environmental characteristics of Pyropia tenuipedalis foliose
thallus growing in Yamaguchi Bay, Yamaguchi Prefecture from November to March,
2010-2014. Young foliose thallus of this species appeared in December and grew tomaximum length from January to March. In March, mature thalli were observed. Water
temperature from November to March was usually within the range of 6 - 16°C, but thetemperature was lower in early December 2012 than other years. Light reaching thegrowing depth of this species was 6.2 ± 3.1% of the level at water surface at high tide.
The concentration of dissolved inorganic nitrogen at the sampling site was within therange of 8.8 - 68.3 μmol L-1, that was approximately 10 times higher than the level thatis assumed to cause the discoloration of P. yezoensis. Water temperature falling ratefrom November to December in 2012-2013 was 0.33°C day-1, which was higher thanthose in other years. In this study, it was suggested that the growth of thalli fromspherical cell was inhibited by a long or short term rapid water temperature decrease,which occurred from November to December.
Abe, M., Murase, N., Nakae, M., Nakayama, T., Nakagawa, M., Shikano, Y. 2018.
41

Water temperature characteristics in growth of filamentous thallus and formations ofspherical cell, uniseriate and foliose thallus of Pyropia tenuipedalis (Miura) Kikuchi et
Miyata. J. Nat. Fish. Univ. 66, 81-88.
We investigated the water temperature effects on growth of filamentous thallus andformations of spherical cell, uniseriate and foliose thallus of Pyropia tenuipedalis with
culture experiments. Optimal growth of filamentous thalli was observed at 20ºC.
Moreover, optimal water temperatures for formations of spherical cells, uniseriate andfoliose thalli were 20ºC, 15–20ºC and 15ºC, respectively. Optimal water temperatures ofeach life stage were different. At 10ºC, the formations of uniseriate and foliose thalliwere suppressed. At 25ºC, almost all thalli had morphological abnormalities.
Furthermore, decreasing temperature to 15ºC from 20ºC enhanced formation ofuniseriate and foliose thalli. It was thought that low production in 2012 resulted fromsuppressed formation of uniseriate and foliose thalli at less than 10ºC.
Murase, N., Abe, M., Fukudome, K., Nakagawa, M., Shikano, Y. 2018. Influence oftemperature on the growth of red alga Pyropia tenuipedalis thalli. J. Nat. Fish. Univ. 66,
215-220.
This study was designed to clarify the optimal temperatures for growth in uniseriate
thalli and foliose thalli of Pyropia tenuipedalis in the laboratory culture at 5ºC intervalfrom 10ºC to 25ºC or 30ºC. The optimal temperature for uniseriate thalli developed
from a spherical cell at two-cell stage were 15ºC and 20ºC. The optimal temperature forfoliose thalli allowed to develop from young blade with the length about 6 cm was
15ºC. It was suggested that the shift in the optimal temperatures from uniseriate tofoliose thalli stages was related to the seasonal reduction of water temperature fromautumn to winter.

42

Improvement of dietary effect on juvenile Ruditapes philippinarum
using the dietary-supplements and a new diet microalga
Yasuhiro Yamasaki
National Fisheries University, Nagata-Honmachi, Shimonoseki, Yamaguchi 759-6595,
JapanE-mail: yamasaky at fish-u.ac.jp
The Manila clam, Ruditapes philippinarum (Adams and Reeve, 1850), is not only acommercially important bivalve but also one of the most ecologically important bivalvesin the world. However, the annual catch of this species in Japan continues to decrease
drastically, and several causative factors have been reported, such as the degradation ofbottom sediment, mass depletion of egg production and larvae, predation, and overfishing.
In addition, these factors may be related in some way to the relatively rapid global
environmental change in recent years. To conserve the clam resources, there have been
many studies investigating the development of seed production and juvenile clam culture.
In these circumstances, a persistent problem is insufficient food supply for juvenile clams
exceeding a shell length of 1 mm, which is a size when the clams have high foodrequirements, especially during winter and spring where low temperature slows thegrowth of the diet microalgae. Therefore, the development of dietary-supplements and/or
new species of diet microalgae that contain the essential nutritional properties and arecapable of rapid growth under low water temperatures will lead to a breakthrough forjuvenile clam culture and consequently to the conservation of the clam resource in the
coastal environment. This talk introduces the results of enhancement of dietary effect onjuvenile R. philippinarum using alginate, an acidic polysaccharide known as a dietarysupplement, and the euglenophyte Eutreptiella eupharyngea as a new diet microalga.
Future challenges on development of a process for practical utilization of the dietarysupplements and the new diet microalga for clam culture will also be discussed.
Annotated Bibliography of Key Works
Yamasaki, Y., Taga, S., Kishioka, M. 2015. Preliminary observation of growth-promoting
effects of alginate hydrolysates on juvenile Manila clams, Ruditapes philippinarum.
Aquaculture Research 46, 1013-1017. DOI: 10.1111/are.12246.
Several studies have been suggested that certain types of sugars are potentially a good
supplement for growth of bivalves such as Ruditapes philippinarum. We observed the
dietary effects of a harmful raphidophyte Heterosigma akashiwo on juvenile clams andsuggested that the acidic sugars in phytoplankton might be an important factor
determining the shell-length growth of clams because total sugar and acidic sugar
content of H. akashiwo were higher than other diet microalgae. Therefore, we focused43

on alginate known as one of the acidic polysaccharides, and showed that shell-length
growth of juvenile clams (average shell length: 432 to 507 µm) was significantly
promoted by supplementing the diatom Chaetoceros neogracile (40,000 to 80,000
cells/mL) with alginate-hydrolysates (AHs) of at least the concentration of 1 mg/L. In
addition, the most effective concentration of AHs was 2 to 4 mg/L.
Yamasaki, Y., Taga, S., Kishioka, M., Kawano, S. 2016. A metabolic profile in Ruditapes
philippinarum associated with growth-promoting effects of alginate hydrolysates. Sci.
Rep. 6, 29923. DOI: 10.1038/srep29923.
We demonstrated that shell-length growth of Ruditapes philippinarum (average shelllength: 15.7 mm) was significantly promoted by supplementing the diatom Chaetoceros
neogracile (80,000 cells/mL) with alginate-hydrolysates (AHs) at the concentration of 4
mg/L. Furthermore, metabolomics indicates that clams in the groups given C.
neogracile with AHs at the concentration of 4 mg/L actively utilized excesscarbohydrate for the development of reproductive tissue. On the other hand, clams in
the groups given C. neogracile only were actively growing through the use of theiradequate carbohydrate resources. Thus, supplementation of AHs with the algal diet may
be an effective way to shorten the rearing period of clams.
Yamasaki, Y., Ishii, K., Taga, S., Kishioka, M. 2018. Enhancement of dietary effect ofNannochloropsis sp. on juvenile Ruditapes philippinarum clams by alginate hydrolysates.
Aquaculture Rep. 9, 31-36. DOI: https://doi.org/10.1016/j.aqrep.2017.11.006
The eustigmatophyte Nannochloropsis sp. is widely used in the aquaculture industry
because this species can be produced on a large scale at low cost. However,Nannochloropsis sp. has less dietary effect on juvenile bivalves compared with otherdiet algae such as the diatom Chaetoceros neogracile and the haptophyte Diacronema(=Pavlova) lutheri. In this study, the use of alginate-hydrolysates (AHs) to enhance the
dietary effect of Nannochloropsis sp. on juvenile Ruditapes philippinarum (averageshell length: 1,090 µm) was attempted. As a result, enhancement of the dietary effect onshell-length growth of juvenile clams was observed in the groups given
Nannochloropsis sp. (300,000 cells/mL) with AHs at the concentration of 4 mg/L.
Hence, the enhanced dietary effect of a combination of Nannochloropsis sp. and AHs
will be useful to shorten the rearing period of R. philippinarum.
Yamasaki, Y., Ishii, K., Hikihara, R., Ishimaru, M., Sato, F., Taga, S., Kishioka, M.,
Matsunaga, S., Shikata, T., Abe, M., Kato, S., Tanaka, R., Murase, N. 2019. Usefulnessof the euglenophyte Eutreptiella eupharyngea as a new diet alga for clam culture. Algal
Res. 40, 101493. DOI: https://doi.org/10.1016/j.algal.2019.101493.
Microalgae are an essential feed source for seed production of bivalves such as
Ruditapes philippinarum. However, there is a deficiency of microalgae that can provide
44

a stable supply of nutrient-rich feed at low water temperatures during the winter andspring. To develop a new diet of microalga that can grow well outdoors at low watertemperatures and possesses the essential nutritive constituents, we focused on theeuglenophyte Eutreptiella eupharyngea, which was isolated from a pond used forextensive phytoplankton cultivation at the Yamaguchi Prefectural Fisheries ResearchCenter (Yamaguchi, Japan) in January 2013. As a result, this species grew well at watertemperatures of 10–25 °C, but could not grow at 30 °C. Furthermore, the dietary effectof E. eupharyngea per dry weight on juvenile Ruditapes philippinarum (average shelllength: 1,426 µm) exceeded that of the diatom Chaetoceros neogracile. These findings
are attributable to the high nutritional value of E. eupharyngea as typified by its highprotein and sugar content and high content ratio of n-3 fatty acids such as
eicosapentaenoic and docosahexaenoic acid and n-6 fatty acids such as arachidonic acid.

45

Exploration of alternative protein sources in the development of asustainable Japanese white trevally Pseudocaranx dentex juvenile diet
Jonas Miller1*, Shuhei Tanaka1, Hiroki Kihara1, Shinichi Yamada1, Fumiaki Takakuwa1,
Keitaro Kato2, Amal Biswas1 and Hideki Tanaka1
1

Uragami Station, Aquaculture Research Institute, Kindai University, 468-3 Uragami,
Nachi-katsuura, Higashimuro, Wakayama 649-5145, Japan
2
Shirahama Station, Aquaculture Research Institute, Kindai University, 1-5 Shirahama,Nishimuro, Wakayama 649-2211, Japan*E-mail: Jm6254 at gmail.com
White trevally (Pseudocaranx dentex) is a novel marine finfish species that is being
cultured commercially in Japan. Few nutritional studies have been conducted on whitetrevally using alternative protein sources to replace the marine ingredients fish meal (FM),the primary protein source in marine fish diets, and the flavor attractant krill meal (KM).
The plant protein soybean meal (SBM) is widely produced and has a good amino acid(AA) balance for aquaculture diet formulations. Poultry byproduct meal (PBM) is madefrom combining the byproducts originating from poultry processing plants and also
contains a strong AA balance. Three feeding trials were conducted to evaluate thepotential of SBM as a replacement of FM and PBM as a replacement for KM in an effort
to develop an eco-friendly juvenile diet for white trevally.
In trial 1, six iso-energetic diets were formulated; FM 560 g kg-1 diet (Control, diet C), andFM protein in diet C was replaced at 10 (S-10), 20 (S-20), 30 (S-30), 40 (S-40) and 50%(S-50) by SBM, and compared to a commercial reference. Fifteen juveniles (42.7 g) wererandomly distributed into each of twenty-one 500 L tanks, set in triplicate and fed for 10
weeks. In trial 2 seven iso-energetic diets were formulated, and FM protein was replaced
at 30 (S-30), 35 (S-35), 40 (S-40), 45 (S-45) and 50% (S-50) by SBM. Another diet was
designed by omitting binders carboxymethyl cellulose and guar gum from diet C andreferred to as C-CG. Thirty juveniles (3.68 g) were randomly distributed into each oftwenty-one 300 L tanks, set in triplicate and fed for 6 weeks. In trial 3, diet C-CG fromtrial 2 was used as the control. Five iso-energetic diets were formulated as follows: FMprotein from diet C-CG was replaced at 50 (S-50) and 70% (S-70) and KM from diets S50 and S-70 was replaced by PBM and referred to as P-50 and P-70, respectively, and a
FM-free diet (FMF). Fifteen juveniles (31.8 g) were randomly distributed into eighteen
500 L tanks, set in triplicate and fed for 8 weeks.
In trial 1, all SBM-based diets outperformed the control and commercial diet in termsof final weight gain and feed efficiency. The results of trial 2 revealed no significant
differences between diets C and C-CG, indicating that binder was not the cause of poor
growth performance of diet C in trial 1. Although there were no significant differences46

among SBM-based diets (P>0.05), S-50 exhibited the greatest final weight gain out of alltreatments. In trial 3, there were no significant differences in the growth performance
among SBM-based diets (P>0.05). While there were no statistical differences between
diets C-CG and FMF, fish fed with diet S-70 showed significantly higher growth
performance than those fed with both diets (P<0.05). The results of all three feeding trials
indicate that SBM can effectively replace FM up to 70% with the addition of KM,
methionine, lysine, taurine and phytase supplementation. Additionally, PBM has theability to effectively replace KM in up to 50% FM replacement diets by SBM.
Annotated Bibliography of Key Works
Biswas, A., H. Araki, T. Sakata, T. Nakamori, K. Kato, and K. Takii. 2017. Fish meal
replacement by soy protein from soymilk in the diets of red sea bream (Pagrus major).
Aquaculture Nutrition 23: 1379-1389.
The results of this study introduce the ecological benefits of replacing FM by soy-based
products in juvenile red sea bream, such as reduced phosphorus discharge to theenvironment. This feeding experiment was also carried out at the Aquaculture ResearchInstitute of Kindai University, Japan using the same methods that were utilized in the
research presented in our abstract.
Jirsa, D., A. Davis, K. Stuart, and M. Drawbridge. 2011. Development of a practical soybased diet for California yellowtail, Seriola lalandi. Aquaculture Nutrition 17: e869-e875.
This study was carried out in an effort to develop an environmentally sustainable andcost-effective diet for California yellowtail juveniles using soy products to replace fishmeal. This research used a similar approach in terms of feed formulation, specificallyinvestigating the utility of soybean meal and soy protein concentrate. Interestingly, fish
fed diets containing soybean meal outperformed fish fed soy protein concentrate, whichfurther influenced our choice of testing the utility of conventional soybean meal on whitetrevally juveniles. This research was carried out in the United States.
Kader, A.Md., M. Bulbul, S. Koshio, M. Ishikawa, S. Yokoyama, B.T. Nguyen, and C.F.
Komilus. 2012. Effect of complete replacement of fishmeal by dehulled soybean meal
with crude attractants supplementation in diets for red sea beam, Pagrus major.
Aquaculture 350-353: 109-116.
The authors found that in red sea bream juveniles, nutrient utilization, body composition
and blood parameters were improved or not significantly affected by replacing fish meal
with soybean meal in combination with flavor attractant supplementation. Growth
performance results from this experiment express a similar trend to the results presented
in our abstract, in which soybean meal diets outperformed the fish meal control diets inwhite trevally. This research was carried out in Japan.
47

Novriadi, R., G. Salze, A. Abebe, T. Hanson and D.A. Davis. 2019. Partial or total
replacement of fish meal in the diets of Florida pompano Trachinotus carolinus.
Aquaculture Research 2019: 1-12.
This research demonstrates the high utility of soybean meal in combination with otheralternative protein sources other than fish meal. The results of this study revealed that 466
g/kg conventional soybean meal in combination with enzyme treated soybean meal and
80 g/kg corn protein concentrate can be used to reduce dietary fish meal from 15 to 9 g/kg
in practical diets for Florida pompano. This research was carried out in the United States.
Waldemar, R. Jr. and D.A. Davis. 2012. Replacement of fish meal with poultry by-productmeal in the diet of Florida pompano Trachinotus carolinus L. Aquaculture 338-341: 160166.
The authors of this research found that poultry byproduct meal in combination withsoybean meal proved to be favorable, even in diets containing up to 50% conventional
soybean meal, 9.8% poultry byproduct meal, and 5% fish meal. This research influenced
our decision to explore the use of poultry by-product meal as replacement for krill meal,in combination with soybean meal in the feed formulations used in trial 3. This research
was carried out in the United States.

48