This method can help answer key questions in the field of shellfish ecology such as how much susten are available as high quality food at potential aquaculture sites. The main advantage of this technique is that it is much more reliable than the water experiments because the measurements are conducted within the natural environment. This method can be applied to other systems and species as well as can be expanded to understand the ecosystem services provided by shellfish.

Individuals new to this method will struggle because the conditions at sea can make sampling challenging either through boat movement or the lack of food availability for shellfish. Visual demonstration of this method is critical because every step, especially the sequencing, needs to be done correctly to get a valid measurement. While still at the dock assemble the gimbal table to consist of 2 frames.

A gimbal table and a ballast tank. Stock a PVC cube with a removable ballast and fill the ballast tank with eighty-five kilograms of sea water. Insert a fifty kilogram zinc weight to the bottom of the ballast to act as a counterweight to the swing of the table.

And attach the ballast tank to the gimbal table. Place a fifty millimeter long T-shaped movable baffle piece on top of the feeding chamber walls for water to flow under within the chamber 1 to 2 centimeters in front of the bivalve. Fit the head chamber and feeding device on top of the gimbal table and hold the materials in place with anti-skid mats.

Then start the pump to allow the addition of sea water to the device from the appropriate experimental depth. To calibrate the flow rates place a one hundred millimeter graduated cylinder at the exit of a feeding chamber and immediately begin recording the time with a stop watch. After thirty seconds check the volume of the water collected in the cylinder, ideally one hundred millimeters of water have been collected.

If the volume collected is not within 5 millimeters of the one hundred millimeter target, close or open the valve between the head tank and the feeding chamber to adjust the flow and check the new flow rate again as demonstrated. When the device is ready, progress to the site of the experiment. Upon arrival to the field site, clean the bivalves of epibionts and other encrusting organisms to avoid filtration by other fauna.

Before placing 5 bivalves into individual basins containing three hundred millimeters of ambient unfiltered sea water. Then add 2 milliliters of tetraselmis SP monoculture to each beaker and record the time that each individual bivalve opens as signalled by a shell gate. Check each beaker every 3 to 5 minutes to ensure that the bivalves remain open and are producing brown densely packed tight-stringed feces that maintain their structure upon pipetting.

Pseudofeces, which are a result of an excess of tetraselmis SP are lightly packed cloud-like deposits of non-ingested particles that quickly resuspend when collected by pipette. Record the time for each individual bivalve as the green feces appear to allow the calculation of the mean gut transit time for all 5 bivalve replicates. While the gut transit time is being determined, collect water samples with a site specific volume of water overflowing from the head tank and from the control chambers every fifteen minutes for at least 2 hours.

Next, use hook and loop tape to attach a live mussel to each of sixteen feeding chambers of the bio deposition apparatus and one set of empty mussel shells to each of 4 control chambers. Collect particles contained in the overflow and control chamber water samples via filtration on 3 individual pre-weighted glass fiber filters per time point and rinse the filters with about 5 millimeters of isotonic imodium formate while the filters are still on the filtration manifold. When the gut transit time has been determined, set a timer to delay the onset of the biodeposit collection from the water collection by the length of the mean gut transit time.

Then clear the chambers of all of the feces pseudofeces that have been produced and shade the bivalves to increase the number of bivalves that open to feed. Use a glass pipette to separately collect and filter the feces and the pseudofeces continuously pulling each type of biodeposit into one container per bivalve throughout the collection period as demonstrated for the water samples. When all of the samples have been collected, store the filters in a cooler with ice for transport to the laboratory.

The goal of the sample collection is to obtain enough material on the filters so that the organic and inorganic fractions can be accurately weighed. Therefore, the sample collection times in volumes may vary across locations. Using this method an even particle distribution across the individual chambers within the feeding apparatus is observed indicating a consistent delivery of food particles of equal quantity and quality from the head tank to the individual chambers.

In this representative experiment 4 ship board trials were conducted with 3 mussel species in 3 locations with very different susten qualities and compositions. Bivalves adjust their feeding behavior according to differences in the amount and type of particulate matter in the water reflecting both plastics physiological responses to food quantity and quality as well as species differences across 3 of the 4 experiments. Analytical problems commonly associated with areas of low particulate matter are illustrated in these feeding behavior results from the California data collection set where some of the pseudofeces were initially mistaken for feces.

While attempting this procedure it is important to remember to accurately record all of the times, water volumes and site information. Following this procedure, other methods like nitrogen analysis of the biodeposits can be used to quantify the ecosystems services provided by shellfish. After its development this technique paved the way for researchers in the field of shellfish ecology to compare the environmental benefits provided by different species of shellfish under the same conditions.

Don't forget, that working at sea can be extremely challenging and all safety precautions should be met. Including knowing when the sea state precludes further work.

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