Genetic comparisons of temperature tolerances of a candidate sea vegetable crop, Alaria esculenta

Genetic comparisons of temperature tolerances of a candidate sea vegetable crop, Alaria esculenta

Summary

In 2014, the global landings of aquatic algae totaled 27.3 million tonnes, worth US$5.6 billion, the majority of which was produced through aquaculture rather than wild harvest (FAO, 2016). The kelp Alaria esculenta has potential in US markets due to its nutritious, palatable characteristics; however, it is necessary to take coastal warming attributable to climate change into consideration in order to support local aquaculture by selecting for temperature tolerant strains. This year, I tested seedstock gametophytes in thermal stress experiments to examine the direct effect of temperature on this potential crop. I also completed a reproductive phenological profile for wild Alaria esculenta throughout the Gulf of Maine, to determine when seedstock can be available to the industry. Samples were collected from the thermal stress tests for analysis of gene expression to determine the response of gametophytes to warming sea surface temperatures at a gene-specific level. The results of these experiments and surveys will help to support the developing sea vegetable industry in the Gulf of Maine and make it more resilient to climate change.

Objectives/Performance Targets

This year’s focus has been on Objectives 1, 3, and 4.

Objective 1: Identify temperature-tolerant (TT) seedstock strains of Alaria esculenta gametophytes for Maine sea vegetable aquaculture (SVA). During strain identification, regional differences can be determined.

• I released zoospores from previously cleaned sporophylls of four plants from the northern site in Lubec and four plants from the southern site in Two Lights. Cultures (n = 8) were maintained to gain vegetative mass before thermal experiments began.
• Gametophyte cultures from each source population of seedstock were exposed to each of two treatments (2×2 factor design): (1) a control treatment where seedstock was maintained at 12 C, and (2) a gradual warming treatment, where temperature of the culture chambers was increased by 1 C every 12 hours until reaching 22 C. This was done gradually to permit a realistic test of thermal tolerance. Replicate cultures were assessed daily for two parameters: (1) survival (live count) and (2) health (dead = loss of all organelles, just a cell wall remaining, unhealthy = entire gametophyte filament very pale or undergoing plasmolysis, or healthy = filament appears normal). I used an inverted microscope and returned to the same frame to track the same individual gametophytes throughout the experiment. I also took daily photographs of the gametophytes.

Objective 3: Determine which temperature-responsive genes are associated with temperature tolerance (i.e. examine differences in gene expression).

• During the thermal experiment outlined above, I harvested gametophytes onto a sterile filter and immediately flash froze samples in liquid nitrogen and transferred them to an ultracold freezer for storage (-80 C). Samples of gametophytes from both locations at 12 C, 18 C and 22 C (n=8 per temperature) will be used to examine changes in gene expression during the gradual ramp up to 22 C.
• I have extracted RNA from spare tissue to determine both quantity and quality of the isolations. This way I can ensure that I have enough RNA for high-throughput sequencing at the Center for Biotechnology and Genomics at Texas Tech. If necessary, I will modify my extraction protocol by exploring kit chemistries that are designed to recover low concentrations of RNA.
• My PI and committee have approved an experimental design for these sequencing experiments to ensure I have adequate biological and technical replication.

Objective 4: Promote sea vegetable aquaculture in Maine by attending conferences and forums and working directly with harvesters.

• I attended the 55^thAnnual Northeast Algal Society (NEAS) Symposium in Westfield, MA where I co-authored a presentation entitled “Determining Conditions for Best Sea Vegetable Crop Production on Sea Farms in Coastal Maine,” where we compared experimental grow-out of sea vegetables, including Alaria esculenta, at two different aquaculture farms at different depths.

• I attended the NSF Division of Environmental Biology: Dimensions of Biodiversity Meeting in Washington, D.C. While I presented work from another project in my dissertation, I discussed the importance of sea vegetable aquaculture with other scientists to promote interest and demonstrate the importance of aquaculture in the global economy in light of predicted worldwide food shortages.

Accomplishments/Milestones

Reproductive Phenological Surveys

• Upon completion of our bimonthly phenological surveys (full two years), I have the final results that determine the natural reproductive cycles of Alaria esculentain the Gulf of Maine. A univariate repeated reassures analysis of variance showed that Alaria was reproductive virtually year-round, although the proportion of the populations that were reproductive did vary seasonally.
• Access to seedstock does not differ across the locations studied throughout Maine (Lubec, Schoodic, and Pemaquid), but the reproductive proportions do vary throughout the year, and these differences are not consistent throughout the locations. There was also an inter-annual difference, though slight.

Laboratory Thermal Experiments: Survival versus Health

• For material from either the northern or southern seedstock source population that underwent either the control or the heated treatment, neither the source nor the treatment affected the end survival of the gametophytes (live counts). The survival rates of gametophytes changed in different ways over the days of the experiment (repeated measured MANOVA), yet not consistently due to the seedstock source population or the treatment that the gametophytes underwent. This suggests that Alaria can acclimate to the temperatures expected to become more common in the Gulf of Maine by surviving these thermal transitions. I intend to find the lethal limit in the next year.
• But in terms of seedstock health, the source of seedstock didn’t affect gametophyte health, yet there was a difference in the health of gametophytes that underwent the gradual heating treatment compared to the control treatment. This is intriguing, as one might expect seedstock from southern populations, where waters reach 22 C during summer months, to behave differently than seedstock from waters that don’t normally reach 22 C (northern population). The proportion of healthy gametophytes changed consistently over the days of the experiment (repeated measured MANOVA), based on the treatment, yet not due to the seedstock source population. Supplying healthy seedstock to farmers is important to the industry.

Impacts and Contributions/Outcomes

The findings of my recent analyses on both our phenological surveys and the thermal experiments that I performed over the past year should contribute to Maine’s aquaculture industry. Just as with land-based farming, farmers need a seed source or a nursery. Researchers and sea farmers can use results from the surveys to know what time of year is best to harvest seedstock, and may not have to worry as much about the location from where seedstock comes. While the overall survival of gametophytes does not appear to be affected by gradual temperature increases, the industry should be supplied with healthy gametophytes to contribute to higher crop yields. Thus we may have to consider how Alaria seedstocks will fair in warming waters in the future. But it seems that, at least for now, sourcing seedstock from cooler northern populations or warmer southern populations does not affect its thermal tolerance in terms of both survival and health.

I have also started a monthly meeting for graduate students at the University of Maine that are part of the Sustainable Ecological Aquaculture Network (SEANET, NSF EPSCoR project). This is an interdisciplinary group of graduate researchers spanning biology, chemistry, engineering, etc. who work together to answer aquaculturists’ questions and provide other support to the aquaculture industry in Maine. I act as a liaison between administrators and stakeholders, and the graduate student community to ensure students learn about opportunities to disseminate their research findings and to work closely with members of the industry to improve their research questions. SEANET also featured my work on Alaria esculenta on their website. The University of Maine then decided to feature this video on the main website as an example of how research at UMaine has direct impacts on local communities (https://umaine.edu/epscor/2016/08/15/new-video-highlights-quigleys-work/). This SARE grant has opened up amazing opportunities and directions for my projects and outreach.

Collaborators: