OUR GOAL IS: To assess the intra-seasonal health and condition of farmed mussels in Casco Bay, Maine to provide valuable information to farmers on when their crop is the most healthy, stressed, and nutritious.
OUR QUESTION IS: How do fatty acids, storage tissue abundance, reproduction and pathology vary in farmed blue mussels in Casco Bay, Maine both temporally and spatially?
OUR OBJECTIVE IS: Through a partnership with Bang’s Island Mussel Farm in Casco Bay to use a combination of environmental monitoring, histopathology (a cost effective tool used to assess health and condition of mussels) and fatty acid analysis (measure of energy storage vs. expenditure due to metabolic maintenance) to determine how storage tissue quantity and quality (Adipogranular and Vesicular Connective Tissues coupled with fatty acid quantity and profile) as well as the severity and presence of pathological indicators vary seasonally across farm sites with different environmental conditions and within the same site.
This will serve as an important baseline for monitoring farmed mussel populations in the future allow for the development of best management practices to mitigate product loss either through limited handling during periods during stress, optimization of stocking densities, raft spacing.
The use of general condition indices (CI) have been established and assessed for M. edulis and used in research and commercial practices for over 50 years. Davenport & Chen (1987) conducted a comparison of seven different methods of calculating CI, and suggest the following as the best assay, one not affected by prior freezing of samples:
CI is an accessible and inexpensive tool for assessing condition of mussels but does not provide insight to the cellular, pathological, and biochemical condition of the organism.
Several studies have employed histology as a tool to assess the pathology, reproduction and condition of mussels. Bignell et al. (2008) histologically assessed extensively 29 different specific pathological health parameters, as well as the presence and abundance of adipogranular cells (ADG) within vesicular connective tissues (VCT) of wild populations of M. edulis and Mytilus galloprovincialis in the UK for example. Duinker et al. (2008) similarly used an assessment of ADG and VCT in an assessment of health, condition and spawning in mussels in Norway. ADG and VCT are the primary storage tissues for the mussel; VCT is associated with storage of glycogen and ADG with lipid and proteins (Mathieu & Lubet 1993). Thus, ADG, VCT and reproductive status are all valuable indicators of energy organismal investment and potential resiliency in the face of stressors.
Especially informative to our proposal is a study by Sunila et al. (2004) in which the reproduction and pathology of M. edulis on an experimental mussel farm in Long Island Sound, Connecticut, USA was conducted. Long Island Sound (LIS) is considered the a southern extent for the species and prior to this study the pathology, reproduction and aquaculture potential for M. edulis in the LIS region was unknown. Results of the study found that despite exceptionally high growth rates, condition was greatly affected by high prevalence of the diagenetic trematode parasite Proctoeces maculatus, which is considered typically a tropical to sub-tropical species. The GoM is currently considered one of the fastest warming portions of the oceans and as warming continues there is high probability of sub-tropical parasites, akin to Proctoeces maculatus moving into the GoM. However there is little if any available information on the pathology and condition of farmed mussels in the GoM.
In response to a mass mortality event on mussel farms in Casco bay in 2016, the UNE Ocean Food Systems Group conducted a limited case study led by Center for Excellence in the Marine Sciences (UNE-CEMS) Research Scientist Adam St. Gelais, on M. edulis histopathology of farmed mussels impacted by the event. This has allowed the research team to perfect the methodology for conducting histology in this species. Moreover, preliminary results suggest the mussels affected in the mortality event were extremely gravid, but appeared to lack both VCT and ADG storage tissues (figure 1) suggesting a low resiliency to stress recovery due to inadequate energy stores at a time of year (late summer) when food is in limited supply.
Economic and environmental sustainability of Maine working waterfronts has declined as capture fisheries that support them have consolidated or collapsed. Adoption of sustainable aquaculture has become critical to the diversification of the working waterfront in Maine. Emerging as a foundational aquaculture species in Maine are rope-grown farmed blue mussels (Mytilus Edulis). Considered among the highest quality mussels on the market they command a premium yielding market prices upwards of $8.00/lb with a seemingly insatiable market demand.
Any yet, the blue mussel in the Gulf of Maine (GoM) is struggling in the face of climate change. The GoM is rapidly warming (Pershing et al., 2015), producing physiological disruptions (Lesser, 2016) and bringing with it acidic waters, invasive predators, competitor species. The result is an alarming decline in wild intertidal populations (Sorte et al, 2016.) Furthermore, in summer 2016 an unexplained mortality event of farmed mussels led to a loss of ~$60,000 worth of mussel stock on one farm in Casco Bay, Maine.
Despite this, there has been no assessment of the condition, pathology, or parasitology of farmed blue mussels in Maine. We propose an intra-seasonal health and condition assessment through combined histopathological and biochemical analysis of pathology, reproduction and fatty acids in mussels farmed in Casco bay, Maine.
Results will inform farmers of when their crop is healthy, stressed, and nutritious, and hopefully lead to the development of best management practices to mitigate product loss, and serve as an important pathological baseline for monitoring farmed mussel populations as the GoM and climate continue to change.
Maine rope-grown farmed blue mussels (Mytilus Edulis) have garnered a reputation as some of the highest quality mussels on the market commanding a premium farm-gate value and yielding market prices upwards of $8.00/lb. The industry is currently incapable of filling market demand and is poised for sustainable expansion. However, in the face of climate change the GoM is among the most rapidly warming portions of our global oceans (Pershing et al., 2015), making farming blue mussels in the Gulf of Maine not without its challenges and risks. In fact, mussels appear to be already struggling in the GoM; a recent alarming study reported on the severe long term decline in this species within the intertidal zone along the entire 3,500 mile coast of Maine (Sorte et al., 2016) and evidence suggests severe sub-lethal impacts and metabolic depression in response to climate change stressors as the organismal level (Lesser, 2016). In fact, farmers in the mid-coast Maine region during the 2016 growing season reported spontaneous mass mortality of market size product following spawning out of synch with normal reproductive cycles. This reproduction-mortality coupled phenomenon is indicative of severe stress in sessile marine invertebrates and has been documented previously in farmed mussels (Myrand et al., 2000). Despite these alarming events, there is paucity of data pertaining to the condition, pathology and parasitology of mussels in Maine. A histopathological and biochemical baseline for the region is needed in order to effectively monitor for changes in the health of farmed mussels in Maine in the face of a changing climate and a warming GoM.
(Map of region studied and full citations are attached in Research section below.)
Citations and regional map document.ONE17-306-St.-Gelais_Maps-and-References
Our approach leverages the farmer and farm crew to collect samples and deploy environmental monitoring equipment. This allows for greater sample sizes than the research team would be able to collect alone due to logistics of working at sea and the expense of boat time. Twice monthly, beginning in February, 2017 through the end of the sampling window for the project (December 2018) our partner farmer has collected mussels from two separate farms sites, in two size classes: juvenile (shell length 30-50mm) and market size (>50mm shell length). During each collection, enough mussels are harvested from each size class to be parsed across three different laboratory assays: Histology, Fatty Acid Analysis (FAA), and Stable Isotope Analysis (SIA). This large sample set will allows for a robust comparison between each farm site, especially with regards to histological analysis.
In addition to the site level sampling conducted by the farmer, researchers travel to the farm site at Clapboard Island in order to conduct more extensive sampling. During these occasions mussels are collected in the same size classes as are collected by the farmer: juvenile and market size. For these sampling events, mussels are collected at each raft present at the farm site in order to assess raft to raft differences within a site. These sampling events have been opportunistic in nature as field logistics allowed. Due to their limited nature, these samples are not robust enough to make the raft-by-raft comparisons initially intended as the outset of the project.
Overall we now have a 24 month data set across two site representing some 637 mussels sampled for histology (resulting in over 3,000 individual histology slides) , 269 archived lipid samples, and 284 archived for stable isotope analysis. Analyzing all lipid and stable isotope samples is outside the scope of this award but our team hope to find funding to analyze backlogged samples in the near future.
Environmental Data Collection
At each farm site is deployed an array of environmental monitoring equipment & data loggers. Parameters measured include temperature, light, dissolved oxygen and salinity. In addition to these data, we have leveraged datasets of opportunity available via regional weather and tide stations as well as oceanographic buoys deployed in the bay as a part of the Maine EPSCOR Sustainable Ecological Aquaculture (SEANET) project (http://maine.loboviz.com/). Importantly, we also have a detailed chlorophyll data set collected via ongoing water quality monitoring efforts of Friend of Casco Bay, a local water-keeper organization.
Immediately after harvest, mussels are put on ice and transported to the UNE Marine Science Center where they are worked up for each analysis. For every mussel collected, basic biometric data are collected including Shell length, width, and depth, total weight, shell weight, and meat weight (wet), as well as a colorimetric estimate of gender.
Mussels destined for histology are fixed in 10% zinc buffered formalin in seawater solution for 24 hrs prior to rinsing and storage in 70% ethanol. Prior to histological processing, mussels are cut dorsoventrally in the center of the mussel, just anterior of the foot and byssal gland. Mussels are sectioned at 5um thickness at 1mm intervals, a total of 5 sections per mussel, and stained with Hematoxylin.
Mussels of each size class are chosen and cleaned of fouling organisms. The mussel is then shucked and removed from the shell. The meat is then placed in a lipid-cleaned vial -80ºC. because of the expensive nature of lipid and fatty acid elucidation, lipid samples will be archived and analyzed based upon storage tissue inferences observed via the histological analysis. Selected samples have been analyzed for fatty acid content and profile by Bigelow Laboratory for Ocean Sciences via extraction and methylation to Fatty Acid Methyl Esters (FAMEs) with internal standard (C19:0) using approach of Breuer et al. (2013) modified for fish tissue, based on Parrish et al. (2015). Quantification will be by GC/MS using Supelco 37 component FAME standard, to confirm compound identity and for individual compound calibration relative to the internal standard.
To date, 15 mussel samples have been analyzed based on fluctuations in storage tissue identified via histology. 3 samples across 5 dates spread across the months of August & November, 2017 and January, March and May, 2018. Three more samples are planned to be analyzed from December or November of 2018.
Accessory data: Stable Isotope Analysis
While not included in the initial award, our partner laboratory, the Byron Marine Ecology Lab, routinely conducts stable isotopic analysis (SIA) of marine invertebrates for assessment of trophic station and food web analysis via carbon and nitrogen. We collect mussels for SIA in the hope their analysis may shed greater light on the food and food web connections to mussel storage tissue ratios, fatty acids, and reproduction. The meat is then placed in a labeled and ashed 20mL vial and stored in a freezer. Frozen samples are freeze dried for 24 hours (more time may be necessary for larger mussels) then either placed back in the freezer or crushed using a mortar and pestle. Samples are crushed to a fine powder, returned to their respective vials and can be frozen indefinitely (OR placed in a Tupperware with desiccant). 1 (microgram) of powder is weighed out and packaged in foil. Samples are then sent to an outside laboratory for analysis of carbon and nitrogen.
Histopathology has been monitored continually as samples have been processed. As part of this section, included are two student reports (pdf format) completed during the past year that will be built upon and unified with environmental, lipid and isotope data as well as with samples collected in months 12-20 of this 24 month project.
Parker_Katherine_2018 (please ignore NSF language at end of report. NSF/Main EPSCoR funds were leveraged as student stipends to conduct this work)
In depth analysis continues by the research team. Over the next 5 months as we approach the conclusion of this award, Connor Jones and myself will be working to interpret the data and draft at least one, but perhaps two manuscripts for publication and to fill out complete the results section. However, in generalities we can propose some general trends.
Pathogens and Physiological Stress Response
Overall pathogen loads in terms of presence and intensity are relatively low across both sites, across both years. Most notable is documentation of multiple life stages of an as yet unidentified trematode species.
More prevalent however, across both years, are indicators of physiological stress response, namely that of oocyte atresia and resorption (the apoptotic death of oocytes and subsiquent reallocation of lipids and other oocytic materials to non reproductive processes) as well as digestive glad atrophy (the thinning of the wall of digestive gland canals). While some level of physiological stress response is normal in any population, the levels seen in our samples, especially that of oocyte atresia, suggest pathological levels of stress, especially in summer months.
Reproduction and Energy Investment
Via analysis and staging of gametogenesis over multiple years, it can be inferred that farmed blue mussels in the region can exhibit either single or dual spawning events across a season. Mussels in 2017 suggest a disjointed spawn in late June and early October, 2017. in 2018 however mussels appear to have a single strong spawning, again in late June. Also noteworthy is an increased investment in reproduction in 2018 over 2017, indicating that energy investment in reproduction is variable across years, likely mediated by environmental factors.
As expected, an inverse relationship was observed in terms of total mantle tissue area occupied by either gametes (reproductive energy investment) and adipogranular tissue (lipid storage). Vesicular connective tissue (glycogen: short term energy storage) did not appear to be related to either reproductive or ADG tissue investment.
Results of the histo-referenced samples analyzed for lipid classes and fatty acid profiles have only just arrived to us from the laboratory we contracted to run these analyses (Memorial University, Newfoundland, CA). Results will be interpreted between the time of this reporting and the conclusion of the project in April, 2019.
Thus far, we have established the dominant pathological indicators and agents the present in blue mussels on our partner farm. In addition we now have a detailed window into the reproductive cycle and timing at the farm sites as well as refined methods for assessing storage tissue ratios using image analysis software. Over the course of the next year, we will continue intensively sampling, send out lipid samples for analysis sand begin to build the picture of health and reproduction in farmed blue mussels in Casco Bay, Maine.
Education & Outreach Activities and Participation Summary
Parker, K., Byron, C., St. Gelais, A. Histopathological Analysis of Parasites and Environmental Stress Responses of Farmed Blue Mussels (Mytilus edulis) In Casco Bay, Maine. New England Estuarine Research Society (NEERS), Portsmouth, NH, April 26, 2018. [Oral]
Jones, C., St. Gelais, A., Byron, C., Costa-Pierce, B., Smolowitz, R. ‘A histopathological health and condition assessment of farmed blue mussels (Mytilus edulis) in a changing Gulf of Maine’. American Fisheries Society (AFS), Atlantic City, NJ, August 2018. [Oral]
Parker, K., Condon, M., Jones, C., Byron, C., St. Gelais, A. ‘A histopathological health survey or farmed blue mussels (Mytilus edulis) in the Gulf of Maine’. Maine-North Atlantic & Arctic Education Consortium, Portland, ME, April 2018. [Poster]
Jones, C., Condon, M., Jane, A., Parker, K., St. Gelais, A., Byron, C. ‘A histopathological health survey of farmed blue mussels (Mytilus edulis) in the Gulf of Maine’. 2018 RARGOM Annual Science Meeting, Portland, ME, October 2018. [Poster]
Parker, K., Byron, C., St. Gelais, A. ‘Histopathological Analysis of Parasites and Environmental Stress Responses of Farmed Blue Mussels (Mytilus edulis) in Casco Bay, Maine’. University of New England College of Arts and Sciences 19th Annual Spring Research Symposium, Biddeford, ME, May 2018. [Oral]
Condon, M., Byron, C., St. Gelais, A. ‘Analysis of Reproduction and Energy Investment within a population of Farmed Blue Mussels (Mytilus edulis) in Casco Bay, Maine’. University of New England College of Arts and Sciences 19th Annual Spring Research Symposium, Biddeford, ME, May 2018. [Oral]
Condon, M., Parker, K., St. Gelais, A., Byron, C., Jones, C. ‘A Histopathological Health Survey of Farmed Blue Mussels (Mytilus edulis) in the Gulf of Maine’. SEANET 2018 All Hands Annual Meeting, Orono, ME, May 2018. [Poster]
Condon, M., Parker, K., St. Gelais, A., Byron, C., Jones, C. ‘A Histopathological Health Survey of Farmed Blue Mussels (Mytilus edulis) in the Gulf of Maine’. Milford Aquaculture Seminar, Shelton, CT, January 2018. [Poster]
Condon, M., St. Gelais, A., Byron, C., Jones, C. ‘Interannual Analysis of Reproduction and Energy Investment within a Population of Farmed Blue Mussels (Mytilus edulis)’. SEA Fellows Symposium at the Darling Marine Center, Walpole, ME, August 2018. [Poster]
Condon, M., St. Gelais, A., Byron, C., Jones, C. ‘Interannual Analysis of Reproduction and Energy Investment within a Population of Farmed Blue Mussels (Mytilus edulis)’. University of New England College of Arts and Sciences Annual Fall Research Symposium, Biddeford, ME, September 2018. [Poster]
Jane, A., Parker, K., Jones, C., St. Gelais, A., Byron, C. ‘Preliminary Assessment of Trematode Infection in Farmed Blue Mussels (Mytilus edulis) in Casco Bay, Maine’. University of New England College of Arts and Sciences 19th Annual Spring Research Symposium, Biddeford, ME, May 2018. [Poster].
Jane, A., Jones, C., St. Gelais, A., Byron, C., Parker, K. ‘An Assessment of Trematode Infection in Farmed Blue Mussels (Mytilus edulis) in Casco Bay, Maine’. Sustainble Ecological Aquaculture Network 2018 All Hands Annual Meeting, Orono, ME, May 2018. [Poster].
Jane, A., Jones, C., St. Gelais, A., Byron, C., Parker, K. ‘An Assessment of Trematode Infection in Farmed Blue Mussels (Mytilus edulis) in Casco Bay, Maine’. SEA Fellows Symposium at the Darling Marine Center, Walpole, ME, August 2018. [Poster].
Jane, A., Jones, C., St. Gelais, A., Byron, C., Parker, K. ‘An Assessment of Trematode Infection in Farmed Blue Mussels (Mytilus edulis) in Casco Bay, Maine’. University of New England College of Arts and Sciences Annual Fall Research Symposium, Biddeford, ME, September 2018. [Poster].
Farmers have expressed the value of increased awareness of the physiological health and condition of their crop. An important finding for the farmer is that traditional condition index (a ratio of meat yield to shell size) is an inadequate metric for crop health. In fact, in the summer of 2018, our research team was able to give advance warning to the farmer that the conditions that precipitated significant crop loss in 2016 were again presenting themselves in summer 2018; namely, an over-investment in reproduction and low short and long term energy stores, coupled with a very hot and dry period of weather.
Subsequently, the farmer reported crop moralities as hot, dry conditions persisted. The farmer was able to take the physiological information we provided and make more informed farm management decisions as to whether to harvest the crop prematurely to prevent loss of revenue, or to hold back on harvesting product to avoid handling and additional stress to the crop populations.