Dataset: Detection of polysaccharide enzymatic hydrolysis using a novel detection method from waters taken aboard the R/V Endeavor in the Western North Atlantic (EN683 in May and June, 2022), and from a freshwater lake in North Carolina in 2025

FLA-laminarin linked beads preparation:
Fluorescently-labeled laminarin (FLA-laminarin) was synthesized using previously described methods (Arnosti 2003). Laminarin was used because it is commonly found throughout the surface water of the ocean (Becker et al. 2020). Subsequently, epoxy-activated beads (MC LAB) were covalently linked to lyophilized FLA-laminarin. One week prior to the cruise, beads were aliquoted into incubation vials to ensure a consistent starting number of beads in each vial. A bead slurry was prepared by mixing 1 volume of beads to 10 volumes of bicarbonate buffer. Then, 100 microliters (µL) of the bead slurry was aliquoted into 1.5-milliliter (mL) GC vials and stored at 4 degrees Celsius (°C) in the dark. Directly prior to the start of each experiment, the bicarbonate was removed, and the beads were washed using seawater or autoclaved seawater.

Collection:
Seawater was collected from 13 stations in the western North Atlantic on R/V Endeavor cruise EN683. Water samples were taken at the depth of the deep chlorophyll maximum (determined via CTD; ca 35 meters (m) and 152m, respectively) and at the bottom, 4092m and 5305 m, respectively.

Twelve experiments used seawater that was collected from the ship's underway intake, located ~3 m below the surface, and one experiment used bottom water collected at 4100 m from a Niskin bottle at station 22-B. At each site, temperature and salinity parameters were recorded. For Sargassum mesocosm experiments, an acid-washed 50 mL Falcon tube was submerged into tanks in which Sargassum was incubating to collect surrounding seawater. For freshwater experiments, University Lake (North Carolina) water was collected on August 8th, 2025 from the University of North Carolina (UNC) rowing dock at 09:55. A separate glass Duran bottle was filled with seawater, Sargassum incubation water, or filtered (0.2-mirometer (μm) pore size) freshwater and sterilized in an autoclave for 20-30 minutes to serve as a killed control for microbial activity measurements.

Incubation and measurement:
For seawater and Sargassum experiments, aliquoted beads were pre-washed, using either live or autoclaved seawater, before the start of each experiment. This pre-washing step was done to remove any leftover bicarbonate and reduce the potential salt shock effect on bacterial communities. After the pre-wash, the supernatant was removed, and 300 µL of fresh seawater was added to the vial and then gently mixed together. Once the beads settled, 70 µL of overlying liquid was measured on a mini-fluorimeter using the micro-cuvette adapter (Fig. 2). After measuring the fluorescence, the liquid was placed back into the corresponding vial it came from. Incubations were stored in the dark at room temperature; note that for Stn. 7B, the incubation was stored in the dark in a 4°C fridge, which was close to the in-situ temperature. The signal of the overlying solution was taken every 24 hours for 96 hours.

For freshwater experiments, beads were washed with bicarbonate buffer and kept suspended to maintain a homogeneous slurry. A 1 mL pipettor and a 1.25 mL pipette tip with approximately 5 millimeters (mm) of the tip cut off was used to maintain a homogeneous slurry and transfer 150 µL of bead slurry to triplicate 3 mL exetainer vials per incubation set. The bead slurry was mixed several times between each transfer. Reverse pipetting technique was used to improve precision. To each exetainer vial, 850 µL of either bicarbonate, bulk, 105 µm screened, or killed control water was added. A fourth vial per set containing only 1 mL of the incubation water was used as a background control. Vials were capped, and an initial (t0) measurement was taken. Additionally, vials were fitted with a 1/2 inch interior diameter (ID) rubber grommet just below the cap to keep vials securely and precisely positioned in the mini-fluorometer. Incubations were stored in the dark at room temperature. Measurements were taken at 0.75, 16.25, and 22.42 hrs, and ~ 2, 3, 4, 7, 10, 15, 20, 25, and 30 days.