New Approaches to Seaweed Aquaculture: Developing a Biosecure and Reliable Seed Stock for the Emergent Northeast Edible Seaweed Industry

Progress report for LNE21-431R

Project Type: Research Only
Funds awarded in 2021: $199,035.00
Projected End Date: 12/31/2024
Grant Recipient: Bigelow Laboratory for Ocean Sciences
Region: Northeast
State: Maine
Project Leader:
Dr. Nicole Poulton
Bigelow Laboratory for Ocean Sciences
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Project Information


Seaweed farming is the second largest aquaculture industry worldwide, with global production used in cuisine, manufactured foods, fertilizer, and animal feed. Sea vegetables (the edible subset) are a $6 billion market worldwide and a growing industry, particularly in the Northeast US where commercial species are native. In the Northeast there are >200 established seaweed farms currently in operation as well as ~6 seaweed nurseries that supply these farms. Techniques for growing sea vegetables (both brown and red algae) are well established despite a complex two-part life cycle, but the US industry has been slow to adopt the more technically advanced germ line cultivation techniques used in Asia and Northern Europe. Instead, US farmers rely upon provision of tenuous wild-sourced seed to cultivate juvenile sporophytes for product, an outdated approach subject to natural vagaries in environmental conditions and stochastic changes in reproductive timing, fecundity, and seed quality. To increase production for novel emerging US seaweed markets, an investment into the technological methods for mechanization of seed stock development and preservation are needed. Annually, sea vegetable nurseries require access to sterilized seawater and sufficient lighting, space for care and maintenance of seed stocks, and expertise, making seedling cultivation a barrier to entry for most new aquaculturists. With the development of seed or broodstock cryopreservation, this provides ‘insurance’ for common and specialized strains, minimizing manual labor associated with long-term cultivation and shortened turnaround times for the production of healthy, viable, juvenile sporophytes on ‘seeded-lines’ that can be sold to farmers.


The proposed study seeks to develop long-term maintenance technologies for current (Sugar kelp - Saccharina latissima) and emerging seaweeds (Winged kelp – Alaria esculenta, Irish moss – Chondrus crispus, and Dulse – Palmaria palmata) in the Northeast by testing the efficacy of various sterile cryopreservation techniques for storage of vegetative cultures and spores.


Laboratory experiments will be used to develop cryopreservation approaches and will be initiated by harvesting wild broodstock (spores) of commercially emergent seaweeds and subjecting these spores to different freezing and thawing rates and cryoprotectants. Initial results for sugar kelp (Saccharina latissima) spores indicate that controlled-rate-freezing (CRF) is more effective than flash freezing.  Using live-cultured spores, the traditional culturing technique will serve as a control and will be compared to revival culturing using preserved/thawed seed stocks. Accurate indication of viability is critical to developing quality control measures for nurseries seeking to offer cryopreservation services. We plan to engage both a hatchery/nursery (Atlantic Sea Farms) and a seaweed farm (Maine Sea Farms) to test the feasibility and ease of implementation of cryogenic protocols as well as provide seeded lines for grow-out tests of both brown and red algae for on-farm trials.

Project Objective:

Edible seaweeds are a $6 billion USD market worldwide. Globally, the seaweed aquaculture sector is growing 7.7% annually, and a rate twice that in the Northeast US where many commercial species are native. To increase production for novel emerging US seaweed markets, an investment into the technological methods for mechanization of seed stock development and seed preservation are required. We propose to develop and optimize a well-established method for cryopreserving (freezing), thawing and growing juvenile seaweed vegetable species in biosecure nurseries. The method maintains frozen cultures of the microscopic life stage (spores) of current farmed seaweed vegetables and emerging species.


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  • Dr. Thew Suskiewicz (Researcher)
  • Douglas Bush (Researcher)


Materials and methods:

Research goal: To develop long-term maintenance technologies for current (sugar kelp - Saccharina latissima) and emerging seaweeds (Winged Kelp – Alaria esculenta and Reds – Dulse) in the Northeast by testing the efficacy of various sterile cryopreservation techniques for storage of vegetative cultures and spores.


Our research formally began in the Fall of 2021 for this project and as part of this project we established a cryogenic center with an auto-fill liquid nitrogen dewar for the storage of cryogenically preserved seaweed seed/spores.


During the fall of 2021 we targeted two phenotypes of sugar kelp (Saccharina latissima), which included skinny and wide blade phenotypes.  Sporulation and cryogenic freezing activities were conducted in October and November 2021 for each phenotype, respectively.  For each sporulation of seed the following methodology was applied:


In 2021, natural sources of sugar kelp (Saccharina latissima) reproductive tissue were obtained by the co-PIs from mature sporophytes collected from two locations along the midcoast, Maine and key morphological features were recorded (stipe length, blade length and width). For each phenotype the surface of the reproductive tissue was soaked in sterile seawater for 5–10 min, cleaned with a damp cloth, scraped with a clean razor blade then stored on paper towels in the dark for 24 hours at 10°C. The following day the meiospores were harvested following the procedures of Flavin et al 2013 and Redmond et al 2014.  Attempts were made in the fall of 2022 to obtain more natural sources of reproductive tissue for additional cryogenic tests but natural skinny kelp beds had been over harvested and were not available.  We will be working with two farmers in 2023 (see Cooperators) to obtain all reproductive sorous tissue to complete this project and to provide cryogenic seed stock training.


Cryogenic Freezing Procedure:

In 2021 two different phenotypes (skinny and wide blade kelp) of S. latissima were tested using cryogenics.  For the “skinny kelp” phenotype two cryoprotectants mixed 1:1 in 2 mL cryovials were used and the meiospores were frozen using a ThermoFisher CryoMed controlled rate freezer (CRF) at a rate of 1°C per min to a final temperature of -40°C followed by storage in liquid nitrogen vapor. For the “wide blade” phenotype three cryoprotectants were used and mixed 1:1 in 2 mL cryovials and frozen via CRF.  For each phenotype the following number of vials were collected and stored:

Skinny blade Kelp: cryoprotectant 1 = 97 vials, cryoprotectant 2 = 96 vials

Wide blade Kelp: cryoprotectant 1 = 75, cryoprotectant 2 = 76, & cryoprotectant 3 = 55.


The number of vials depended on the meiospore density for each of the sporulation event. To minimize the genetic variation between different kelp blades, the meiospores from several fronds were combined for each sporulation event and downstream cryogenic freezing. There are advantages and disadvantages to doing this, as each kelp frond with have different meiospore vitality (as per previous research), but in order to test a broad range of cryoprotectants we chose to combine meiospores from different blades in order to generate many vials for downstream testing and differentiation into sporophytes.


In 2023 our team worked in collaboration with Thew Suckiewicz at Atlantic Sea Farms (one of our cooperators/nursery farmer) to obtain meiospores for cryogenic freezing of two phenotypes of sugar kelp (Saccharina latissima - wide and skinny blade).  Please note that other species of kelp were not collected by ASF in 2023 so our main focus was to optimize cryogenic freezing protocols for S. latissima. Over the course of the fall of 2023, three sporulation events were conducted at ASF. A controlled rate freezer (CRF) with a liquid nitrogen tank was brought on site to ASF in Biddeford, ME so that meiospores could be frozen quickly after sporulation. Similar to 2021 and to minimize genetic variability, multiple fronds from one field location along midcoast Maine were combined to obtain a single mixture of meiospores for cryopreservation.  Based on our results for vitality in 2021, two cryoprotectants were used for all three sporulation events. In order to optimize the number of meiospores per cryovial the volume per cryovial was increased from 1 mL to 1.8 mL.  For each sporulation, 10 mL of live meiospores was transported back to Bigelow Laboratory for vitality testing.  The remaining meiospore material was cryopreserved via CRF. 


ASF Sporulation Event 1 - October 3, 2023, Sugar Kelp - wide blade, source - midcoast Maine. ASF sporulation Lot # 275A1-23. For this sporulation event, we attempted to seed spooled lines (for eventual outplanting) using the standard ASF procedure. The initial spore density was ~ 1.6 million per mL, 220 vials frozen (110 per cryoprotectant) using CRF (at a rate of 1°C per min to a final temperature of -40°C followed by storage in liquid nitrogen vapor). 


ASF Sporulation Event 2 - October 18, 2023, Sugar Kelp - wide blade, source midcoast Maine. ASF sporulation Lot #291A1-23. For this sporulation we froze vials using 2 cryoprotectants for return to the laboratory for long term vitality testing and meiospore differentiation.  The initial spore density was ~1.7 million per mL, 302 vials were frozen (151 per cryoprotectant) using CRF (at a rate of 1°C per min to a final temperature of -40°C followed by storage in liquid nitrogen vapor). 


ASF Sporulation Event 3 - October 29, 2023, Sugar Kelp - skinny blade, source midcoast Maine. ASF sporulation Lot #302B1-23.  For this sporulation we froze vials using 2 cryoprotectants for return to the laboratory for long term vitality testing and meiospore differentiation.  The  initial spore density was ~400,000 per mL, ~176 vials were frozen (88 per cryoprotectant) using CRF (at a rate of 1°C per min to a final temperature of -40°C followed by storage in liquid nitrogen vapor). 


Thaw Procedure(s):

For cyrogenic revival of meiospores both the freezing process and thawing process are important aspects of meiospore survival and differentiation.  It is vitally important to remove / dilute the cryoprotectants as soon as possible. Two thawing techniques were examined for each cryoprotectant used for our two Saccharina latissima phenotypes (skinny and wide blade), 1) rapid thaw in water bath and 2) CryoThaw: a simple tube adapter to expedite and automate thawing.


Rapid thaw in water bath: Each frozen meiospore vial was held in a 20 C water bath until a pea sized pellet appears and then the vial content is placed in 9 mL of chilled (4 C) Prov50/2 media, and inverted 5 times the evaluated for vitality using Fluorescein Diacetate (FDA - see protocol below in the next section). 


CryoThaw (Beddall et. al, 2016 DOI: 1016/j.jim.2016.08.009): Using a small tube adaptor (commercially available) the frozen uncapped meiospore cryovial was placed at the entrance of a 15 mL centrifuge tube containing 9 mL Prov50/2 media (chilled to 4 C), the centrifuge is programmed to 22 C, 1500 RPM, for 5 min. Once the vial thaws the meiospore pellet is in the media within minutes and was diluted rapidly and within a controlled environment. The 15 mL vial was removed and inverted 5 times prior to analysis for vitality (as in step 1). The goal of this method is to provide a more controlled environment for thawing and diluting meiospores with the goal to remove the cryoprotectant as soon as possible. 


In 2023 for optimization when working with ASF, only the rapid thaw procedure was used, as obtaining a large centrifuge would likely be challenging for farmers and nurseries. Since all frozen meiospore vials generated at ASF contained 1.8 mL each frozen meiospore vial was held in a 20 C water bath until a pea sized pellet appeared and then the vial contents was placed into 17.2 mL of Pro50/2 media (chilled to 4 C). The optimal dilution for all experimental vials was 1:10.


Vitality Testing:

After thawing, 1 mL of each thawed solution (from different cryoprotectants) is stained with 10 uL of Fluorescein Diacetate (FDA) working stock and incubated at 10 C for 20 mins prior to flow cytometric analysis on a BioRad ZE5 Cell Analyzer. All vital meiospores will fluorescence green with blue laser excitation. The total number of meiospores is determined using red fluorescence (as an indicator of chlorophyll) as well as the number of vital cells. The percent of vital cells is determined and recorded for each vial and cryoprotectant.  As a negative control, 1 mL of thawed solution is heat killed at 50 C for 30 min and stained using the FDA protocol above.

In 2023, we wanted to determine if vitality changed over time while being stored in LN2 vapor.  For this testing two phenotypes of Sugar Kelp (wide and skinny blade) were examined ~ every 15 days for up to ~ 60 days after cryopreservation using two cryoprotectants. 


Motility Testing:

After thawing, 50 uL of each thawed solution is observed under the microscope using a hemocytometer to determine if any of the thawed cells are motile – an indication of vitality.


Meiospore Differentiation examination:

After thawing the diluted mixture (1:10 dilution) was placed into a 6-well tissue culture plate and incubated on a 12/12 light/dark cycle and monitored for meiospore differentiation into gametophytes and eventual sporophyte formation over a period of 4-6 weeks.  The 6 plates were monitored for differentiation approximately 1-2 times per week after thawing.  This was done in triplicate using both cryoprotectants.


Other Kelp species examined to date: In fall of 2022 a protocol for releasing Winged Kelp (Alaria esculenta) spores from reproductive fronds was developed in order to test spore cryopreservation techniques (TBD in 2023). The sporophylls were collected from natural sources (Pemaquid Point, ME), and returned to the lab on ice for processing. The sporophylls were removed from the fronds and scraped with a razor blade to remove mucus and contaminates from the surface. Afterwards the sporophylls were divided into two treatments to induce spore release. 1) Betadine dip: sporophylls were dipped in a Betadine solution (500 uL of Betadine to 500 mL of DIW) for 2 mins then removed into a tray of filtered seawater (FSW) and scraped with a razor blade again, sandwiched between paper towels, and incubated overnight in the dark at 12 C. 2) FSW treatment: sporophylls were dipped in FSW and scraped with a razor blade to remove mucus and surface contaminants, afterwards all scraped sporophylls were dipped in FSW for 30 seconds then removed and scraped again with a razor blade. All sporophylls were sandwiches between paper towels and incubated overnight at 12 C. Of the two treatments the one with the most success was the Betadine dip. Unfortunately, all the reproductive tissue in the field had been harvested when we attempted a sporulation for cryopreservation. To date (both 2022 and 2023) we have been unable to obtain additional reproductive fronds of Winged Kelp and have diverted our focus to two phenotypes of Sugar Kelp.

Research results and discussion:

Preliminary results are available from our first cryopreservation tests of sugar kelp (Saccharina latissima) using three cryoprotectants and two thawing methods.  Of the three cryoprotectants tested, cryoprotectant 3 (cryo 3) had the best % vitality results with the overall highest FDA fluorescence after thawing (Figure 1 - Percent vitality of Slatissima_wide blade). These results were significant using a Tukey T-Test.  For both phenotypes of sugar kelp, cryoprotectant 2 was better than cryoprotectant 1 (Figure 1 - Percent vitality of Slatissima_wide blade  & Figure 2 - Percent vitality of Slatissima_skinny blade).  However, differences between the two thawing methods were inconclusive and require more replication. Very limited motility was observed immediately after thawing.


In 2023 during our collaboration with ASF, we attempted to seed spooled lines using the standard meiospore seeding protocol developed and used at ASF.  After the first sporulation on Oct 3rd, 2023, spooled lines were seeded at a density of ~ 8,000-10,000 meiospores per mL (using live and not cryopreserved material). Separately, meiospores were also frozen with a CRF using two cryoprotectants and thawed following the methods described previously.  The thawed meiospores were used to seed spooled lines in aquaria with 15 liters of water containing diluted cryogenically preserved meiospores at a density of 10,000 spores per mL.  After 3-4 weeks normal sporophyte growth was observed on the spools that contained non-cryogenically preserved material, however, no sporophyte growth was observed on the spools seeded with cryogenically preserved meiospores.  It is our hypothesis that due to the lack of motility (after cryopreservation and thawing) that the meiospores were unable to adhere to the spools prior to media transfer.  We decided for the remaining sporulation events in 2023 (Lots #291A1-23 & #302B1-23)  to focus on methods for seeding line post cryopreservation in the laboratory.


For the two remaining sporulation events in 2023 the longevity of vital meiospores over a period of 50-60 days was examined after cryopreservation for two ASF sporulation Lots (#291A1-23 & #302B1-23).  For live meiospores (prior to cryopreservatio) both Lots had high vitality ranging from 89 - 85 % vital (Figure 3_Percent vitality_Slatissima_wide-blade overtime & Figure 4_Percent vitality_Slatissima_skinny-blade overtime).  For each cryoprotectant the percent vitality did not change over time significantly, however, there was a slight decrease observed.  When comparing the two cryoprotectants to each other there was a significant difference in vitality.  Overall, cryoprotectant 2 was on average more vital ranging between 42 and 30% for both ASF Lots whereas cryoprotectant 1 ranged from 7.5 - 5.5 % vital (Figure 3 and 4).

Research conclusions:

No formal conclusions at this time.

Participation Summary
2 Farmers participating in research

Education & Outreach Activities and Participation Summary

Educational activities:

8 Published press articles, newsletters
3 Tours
7 Webinars / talks / presentations
1 Workshop field days
1 Other educational activities

Participation Summary:

Outreach description:


USDA Agricultural Research Services Interagency Working Group for Farming Seaweeds and Seagrasses stakeholder engagement listening sessions (four, scheduled throughout March 2022) received input from over 178 attendees to collect input on topics to be included in a report to US Congress on the state of seaweed research.

Maine Blue Carbon Symposium (hosted by the Maine Blue Carbon Network; Price is a steering committee member) in Portland, ME 03/07/2023 reviewed the state of the science in measurement approaches for blue carbon.

Hakai Magazine kelp farming dialogue and subsequent session with early career ocean professionals (ECOPs) on 04/11/2023; 1,039 registered ahead of the event; 463 participants joined the Zoom; 31 watched live on YouTube; 81 participants also joined the ECOP session.

National Seaweed Symposium in Portland, ME – 04/26/2023 project representation at Seaweed Showcase tabling event

Dig Deeper with World Wildlife Fund on Seaweed Farming at Wolfe’s Neck Center in Freeport, ME 09/20/2023

Builder’s Initiative Grantees Workshop, various locations on midcoast Maine and Portland 10/10-12/2023

Virtual “Listening Sessions” for seaweed farming across U.S. Territories hosted by Bigelow Laboratory for Ocean Sciences (10/23/23, 10/30/23, 11/6/23, 11/13/23); >1,000 registered. In service of development of a report for Congress due May 2024 to reflect emerging knowledge and resource gaps for the growing farmed seaweed industry in the US.




Civil Eats story released on 03/16/2022 titled “Can Small Seaweed Farms Help Kelp Scale Up?”

Northeast Aquaculture Conference & Exposition (NACE) on April 27-29, 2022: Center for Seafood Solutions’ Nichole Price hosted a field trip of ~20 members of aquaculture industry and the general public to highlight aquaculture research projects and analytical services available at Bigelow.


NPR MPBN Maine Calling radio panel with Nichole Price on 9/7/2022 titled “Maine’s role in the future of the global seaweed industry”


Country Folks web article released on 9/23/2022 titled “Looking to the seas for feed”


News Center Maine web article release on 9/26/2022 titled “Kelp could play a crucial role in trapping carbon emissions from our atmosphere”


The Science Writer web article released on 12/15/2022 titled “The Seaweed Solution: A climate change treatment for coastal Maine?”



Aquaculture Research Institute web article release on 03/07/2023 titled “Sustainable Aquaculture in Maine: Research, Innovation, and Workforce Development”


Portland Press Herald on seaweed as blue carbon source for Maine, 09/22/2023

Global Seafood Alliance article on selective breeding of seaweed, 11/06/2023

Portland Press Herald on multitrophic aquaculture 11/05/2023

Any opinions, findings, conclusions, or recommendations expressed in this publication are those of the author(s) and do not necessarily reflect the view of the U.S. Department of Agriculture or SARE.