Source: TRIPLE N OYSTER FARM LLC submitted to NRP
INLAND PRODUCTION OF OYSTER SEED UTILIZING AN ARTIFICIAL SEAWATER CLOSED RECIRCULATING AQUACULTURE SYSTEM.
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
Reporting Frequency
Annual
Accession No.
1023381
Grant No.
2020-33610-32015
Cumulative Award Amt.
$105,090.80
Proposal No.
2020-04792
Multistate No.
(N/A)
Project Start Date
Jul 15, 2020
Project End Date
Feb 14, 2023
Grant Year
2020
Program Code
[8.7]- Aquaculture
Recipient Organization
TRIPLE N OYSTER FARM LLC
1265 STEPHENS AVENUE
BATON ROUGE,LA 708083766
Performing Department
(N/A)
Non Technical Summary
Inland oyster seed production and R&D in recirculating aquaculture systems is being developed in Baton Rouge, LA at the LSU Innovation Park. The Gulf Coast oyster industry faces numerous challenges including increased exposure to anthropogenic pollution, climate change, ocean acidification, harmful algal blooms, predation, disease outbreaks, and low salinity events. This has prompted Triple N Oysters to culture oysters in a closed system away from the coast of Louisiana. This move has several potential benefits including the production of oyster seed in a controlled environment where temperature, salinity, feed, and nutrients can be optimized for a reliable production of seed for the oyster industry. These inland operations will also be resilient in the face of tropical storms and hurricanes.
Animal Health Component
40%
Research Effort Categories
Basic
30%
Applied
40%
Developmental
30%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
13508111060100%
Knowledge Area
135 - Aquatic and Terrestrial Wildlife;

Subject Of Investigation
0811 - Shellfish;

Field Of Science
1060 - Biology (whole systems);
Goals / Objectives
The major goal is to develop innovative protocols at an inland hatchery to produce oyster larvae and seed for the oyster industry and coastal restoration efforts in a significantly more reliable fashion than is currently taking place in coastal regions of the USA.The Phase I specific goals will be attained from results of experiments designed to yield data during the 8 month duration of the award:Determine broodstock maintenance and conditioning protocols.Determine larvae rearing conditions.Determine seed setting and grow-out conditions.Task 1: experimental determination of broodstock maintenance (months 1-6) For this task we will attempt to condition 900 oysters in a controlled environment. The oysters will be split into 10 separate coolers that have been plumbed to control the water flow through rates. Therefore, 90 oysters will be maintained in each cooler at a specified temperature, water chemistry, and feeding regime. Water temperatures will be set to duplicate the onset of winter in Southern Louisiana. A cold winter scenario will hold the water temperature at a constant 7 C. A warm winter will use a water temp of 12 C. An intermediate temperature for winter of 17 C for an average winter. Table 1 shows the different temperatures that will be used for each treatment.Table 1. Temperature for each of the coolers.Cooler #Temperature17 C27 C37 C412 C512 C612 C717 C817 C917 CA modified protocol of Buchanan et al. (1998) will be utilized to condition the oysters. After a period of approximately 3 months the temperature will be gradually increased over a period of 2 weeks to simulate the end of winter and the beginning of spring. A spring temperature of 25 C will be maintained for an additional 6 weeks. The oysters will be monitored for Perkinus marinus (dermo) infection (Dr Jerome La Pyre, LSU) when mortality occurs, and quality of gonads at the end of the 6 weeks of conditioning. If the oysters appear ripe after the 6 weeks they will be induced to spawn using thermal fluctuations to release their gametes. Weekly water parameters will be analyzed for alkalinity, calcium, dissolved oxygen, nitrate, nitrite, pH, salinity, temperature, and total ammonia. Dissolved oxygen and pH will be recorded daily at the hatchery. Additional water analyses will be performed by Wetland Biogeochemistry Analytical Services (WBAS) at Louisiana State University.From these experiments we predict that one of the conditioning temperatures will yield more oysters that have mature gonads. These initial results will be expanded upon in the Phase 2 effort to more precisely define the conditioning temperature regimes and begin to look at feed effects on oyster conditioning. Feed quality and quantity likely play a major role in conditioning (Ingle, 1951; Buchanan et al., 1989; Kuhns et al, 2013).Task 2: determine optimal oyster larvae rearing conditions in artificial seawater in a closed recirculating aquaculture system (months 5-8)Spawning of oysters from Task 1 or of oysters obtained from Grand Isle, LA or Cocodrie, LA will produce larvae for these experiments. Oyster larvae will be equally split into five 200 L cone bottom tanks setup for flow-through culture (FAO, 2004). In the Phase 1 effort the effect of flow rates on larvae growth will be tested (see Table 2).Tank 1 will have no flow through and will operate as a static tank with a total water change every two days. Tank 2 will have a "fast" rate of 200 L per hour, equivalent to one tank volume per hour. Tank 3 will have a rate equivalent to Tank 1, but spread over a 2 day period of 4.17L per hour. Tank 4 will be set at 50L per hour flow-through, and Tank 5 will be set at 100L per hour flow-through. All tanks will be stocked at an initial concentration of 20 D-hinge oyster larvae per mL.Table 2. Flow-through rates for larvae rearing tanks 1 to 5.Tank #Flow-throughRate (L/hr)10220034.174505100Dissolved oxygen, salinity, and pH will be recorded daily in each tank. Additional water analyses will be performed by Wetland Biogeochemistry Analytical Services (WBAS) at Louisiana State University. Specifically, ammonia, nitrite, nitrate, and alkalinity will be monitored every 4 days in the 5 tanks, with samples being taken from Tank #1 before and after the water is changed.As many replicates as possible will be performed before the end of the Phase 1 period. If there is a significant difference in the growth of larvae amongst the 5 tanks the experimental range of flow-through rates will be adjusted to attempt to narrow our results if time permits. Once an optimum flow rate is observed, additional experiments will be performed to examine the stocking densities of the larvae in each tank at a set flow rate, once again if time permits; otherwise this would be examined in a Phase 2 project. Feed quality and quantity is another variable that would be investigated further during a Phase 2 project.Task 3: Development of a land-based oyster setting and bottle nursery to produce oyster seed ranging in size from 1mm to 3mm. (Months 6-8)When oyster larvae are 12-18 days old they go through metamorphosis to become eyed-larvae. At this stage they are ready to undergo a final metamorphism to become an oyster seed or spat. Setting tables will be utilized in our inland hatchery to "set" these larvae on micro cultch. Micro cultch is finely ground oyster shell size selected for particle between 250 and 300 microns in size. Artificial seawater will be utilized in a closed recirculating aquaculture system to all the larvae a period of up to 5 days to transition in to oyster seed. After setting is completed the newly formed oyster seed will be transferred to an inland bottle nursery where they will be grown from approximately 350 microns in size up to 3 mm in size for sale to commercial oyster farms across the US Gulf Coast. Artificial seawater will be utilized for all aspect of growth in the nursery. Feed will be produced from algae ponds that are maintained on site at the LSU Innovation Park immediately adjacent to the hatchery/nursery operations. Between the months of May and October of 2019 Triple N Oyster Farm successfully grew algae in artificial seawater to concentrations exceeding 2 million cells per mL. Of significance: we are able to maintain high concentration of algae and exponential growth even through the hottest periods of the summer with temperatures exceeding 36C in the algal pools. We have a total algae pool capacity of 45,000L.The two variables of concern are water chemistry and feed quality and quantity. Initial experiments will attempt to grow from 1 to 12 million oyster seed from 350 microns to 3 mm in size. During this grow out phase water analysis will be performed every three days to measure alkalinity, calcium, dissolved oxygen, nitrate, nitrite, pH, salinity, temperature, and total ammonia. Dissolved oxygen and pH will be recorded daily at the hatchery. Additional water analyses will be performed by Wetland Biogeochemistry Analytical Services (WBAS) at Louisiana State University. We hypothesize that calcium in the form of aragonite may fall in concentration as the oysters utilize aragonite as a principle component of their shells. We intend to supplement the seawater with calcium carbonate (aragonite) to maintain high growth rates in the closed recirculating system.If successful our growth rates should approach 1mm per week. If time permits we will conduct additional experiments with a commercial oyster nursery located in Grand Isle, LA (Grand Isle Sea Farms, Boris Guerrero). In these experiments 2 million 350 micron oyster seed will be split equally between the inland nursery in Baton Rouge and the Grand Isle nursery to compare growth rates.To meet the demands of the industry data from Task 3 will be utilized to design a large scale inland oyster nursery with much larger algal ponds and a larger nursery footprint.
Project Methods
Experiments designed on previous observations will be conducted to reach the project goals. Statistical analyses will be used to determine signifigant dofferences exist. Results will be communicated through publication in peer reviewed journals, presentations at conferences, and hatchery to hatchery communications.

Progress 07/15/20 to 02/15/23

Outputs
Target Audience: The target audience included three academic hatcheries: AUSL, USM, and LSU hatcheries. The majority of these intereactions helped both Triple N Oyster Farm and the 3 university hatcheries. The interactions specifically increased the hatchery experimentation and development of protocols related to operating an inland hatchery. After further development, the principles used during this study will also benefit the commercial oyster hatchery industry in the USA. Many hours were spent discussing findings, both positive and negative, to both new hatchery managers and senior managers and investigators across the USA. Changes/Problems:Early in the grant period a decision was made to utilize static growth versus flow-through growth to grow oyster larvae. This change was not problematic, and the hatchery was able to produce record numbers of pediveligers in the second year of the grant research period. In future research Triple N Oysters plans to revisit a flow-through system of growing larvae to increase the capacity of the hatchery. What opportunities for training and professional development has the project provided?Several undregraduate students from LSU were trained in phycological methods to perfom all tasks required in algae production at a commercial shellfish hatchery. How have the results been disseminated to communities of interest?Many of these results have been communicated directly to communities of interest. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? This NIFA funded project has successfully proven that an artificial seawater recirculating artificial seawater system can by used to condition oyster broodstock, spawn oysters, raise oyster larvae, set the larvae, and grow the seed to 3mm in size. The advancement is signifigant in that production of oysters for oyster farms, coastal restoration, and research can take place away from the coast where water pollution, variations in water chemistry, and tropical storms are major issues for a consistent supply of oyster seed. Triple N Oyster Farm successfullycompleting all three tasks. Task 1: The inland hatchery wassuccessfulin developing a broodstock conditioning protocol for diploid and tetraploid Eastern oysters in artificial seawater using a closed recirculating aquaculture system. The ideal conditions were achieved by replicating the natural seasonal changes of water in Grand Isle, LA. This is the first report of successful tetraploid oyster conditioning in the Gulf region. Task 2: A static (no flowthrough) system was highlysuccessfulin producing record numbers of oyster larvae in the artificial seawater closed recirculating system. Egg quality, water chemistry and biology, and algae quality and quantity were optimized for the 6000 L capacity for each brood. See figure 3 for a typical growth curve. Task 3: The hatcherysuccessfullyproduced 3 mm oyster seed 65 miles away from the coast in artificial seawater.

Publications


    Progress 07/15/21 to 03/14/22

    Outputs
    Target Audience:Target audience included three academic hatcheries: AUSL, USM, and LSU hatcheries. The majority of these intereactions helped both Triple N Oyster Farm and the 3 university hatcheries. The interactions specifically increased the hatchery experimentation and development of protocols related to operating an inland hatchery. Changes/Problems: Task 2 issues: The clumping of algae in the artifical seawater initially lead to the cloggin of flow-through filters. This problem lead to the use of a static larval rearing system to first experimentally develop larvae rearing protocols in artificial seawater. Triple N Oysters plans to experimentally assess the possibility of utilizing a flow-through system in the months ahead. Task 3 issues: Additional time is required to experimentally develop protocols to grow largers numbers of oyster seed. A large quantity of algae is required to grow seed to 3mm in size. To achieve this level of feed we will experiment with the use of algal ponds to prove that this concept is achievable in large numbers far away from the coast. What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? The results were communicated with several oyster hatchery managers across the USA. The results have also been communicted to the oyster industry in Louisiana at the public meetings help by the Louisiana Oyster Task Force and at the Louisiana Oyster Aquaculture Association. What do you plan to do during the next reporting period to accomplish the goals? Task 1: Triple N Oyster Farm will further develop through experimentation a broodtscok conditioning protocol for diploid and tetraploid eastern oysters to produce ripe oysters 2-3 month in advance of the natural occuring populations. Task 2: Continue monitoring change in the artifial seawter chemitry to better predict changes in the levels of micronutrients as the water is reused for over 12 months. Task 3: Experimentat will contiue to fine tune the production of large quatities of oyster seed in artificial seawater. This will include experimenting with varied algae diets, flowrates, and development of water quality protocol through experimentation.

    Impacts
    What was accomplished under these goals? This NIFA funded project has successfully proven that an artificial seawater recirculating artificial seawater system can by used to condition oyster broodstock, spawn oysters, raise oyster larvae, set the larvae, and grow the seed to 3mm in size. The advancement is signifigant in that production of oysters for oyster farms, coastal restoration, and research can take place away from the coast where water pollution, variations in water chemistry, and tropical storms are major issues for a consistent supply of oyster seed. Triple N Oyster Farm was successful in moving foraward on all three tasks. Task 1: The inland hatchery was successful in developing a broodstock conditioning protocol for diploid and tetraploid Eastern oysters in artifical seawater using a closed recirculating aquaculture system. The ideal conditions were achieved by following the natural seasonal changes of water in Grand Isle, LA. Task 2: A static (no flowthrough) system was highly successful in producing record numbers of oyster larvae in the artificial seawater closed recirculating system. Task 3: The hatchery successfully produced 3 mm oyster seed in small quantities (less than 10,000 seed). We hope to achieve greater numbers before the end of this award.

    Publications


      Progress 07/15/20 to 07/14/21

      Outputs
      Target Audience:Target audience included three academic hatcheries: AUSL, USM, and LSU hatcheries. The majority of these intereactions help both Triple N Oyster Farm and the 3 university hatcheries. The interactions specifically increased the hatchery experimentation and development of protocols. Changes/Problems:Task 2 issues: The clumping of algae in the artifical seawater initially lead to the cloggin of flow-through filters. This problem lead to the use of a static larval rearing system to first experimentally develop larvae rearing protocols in artificial seawater. Triple N Oysters plans to experimentally assess the possibility of utilizing a flow-through system in the months ahead. Task 3 issues: Additional time is required to experimentally develop protocols to grow largers numbers of oyster seed. A large quantity of algae is required to grow seed to 3mm in size. To achieve this level of feed we will experiment with the use of algal ponds to prove that this concept is achievable in large numbers far away from the coast. What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest?The results were communicated with several oyster hatchery managers across the USA. The results have also been communicted to the oyster industry in Louisiana at the public meetings help by the Louisiana Oyster Task Force and at the Louisiana Oyster Aquaculture Association. What do you plan to do during the next reporting period to accomplish the goals?Task 1: Triple N Oyster Farm will further develop through experimentation a broodtscok conditioning protocol for diploid and tetraploid eastern oysters to produce ripe oysters 2-3 month in advance of the natural occuring populations. Task 2: Continue monitoring change in the artifial seawter chemitry to better predict changes in the levels of micronutrients as the water is reused for over 12 months. Task 3: Experimentat will contiue to fine tune the production of large quatities of oyster seed in artificial seawater. This will include experimenting with varied algae diets, flowrates, and development of water quality protocol through experimentation.

      Impacts
      What was accomplished under these goals? This NIFA funded project has successfully proven that an artificial seawater recirculating artificial seawater system can by used to condition oyster broodstock, spawn oysters, raise oyster larvae, set the larvae, and grow the seed to 3mm in size. The advancement is signifigant in that production of oysters for oyster farms, coastal restoration, and research can take place away from the coast where water pollution, variations in water chemistry, and tropical storms are major issues for a consistent supply of oyster seed. Triple N Oyster Farm was successful in moving foraward on all three tasks. Task 1: The inland hatchery was successful in developing a broodstock conditioning protocol for diploid and tetraploid Eastern oysters in artifical seawater using a closed recirculating aquaculture system. The ideal conditions were achieved by following the natural seasonal changes of water in Grand Isle, LA. Task 2: A static (no flowthrough) system was highlysuccessful in producing record numbers of oyster larvae in the artificial seawater closed recirculating system. Task 3: The hatchery successfully produced 3 mm oyster seed in small quantities (less than 10,000 seed). We hope to achieve greater numbers before the end of this award.

      Publications