Overview: We performed dissolution assays to explore the consequences of two levels of abrasion, used to remove the periostracum, on shell dissolution rates using dead shells of sacrificed adult Mytilus californianus mussels. For the current study, adult mussels (42 - 64 mm in length) were collected from Marshall Gulch, California (38.369738 °N, -123.073921 °W) between August 2021 and March 2022 and transported immediately to the University of California Davis’ Bodega Marine Laboratory (< 30 min distance), in Bodega Bay, California. Mussels were held in filtered, flow-through seawater and fed ad libitum until used in experiments. Because we were interested in the effects of the periostracum in protecting the exterior surface of the shell, we sealed the inner, nacre layer of the shell with silicone (Loctite marine silicone sealant) to prevent its contact with seawater.
Mussel valve preparation: To prepare mussel valves for the incubation, shell periostracum was abraded using sanding sponges (Gator brand) of two levels of grit coarseness (P50-60: coarse (n = 11), and P120-150: fine (n = 12)). We included control mussel valves (15% of daily sample size), collected with no periostracum cover, to compare dissolution rates between abraded and periostracum that was removed naturally in the field.
Dissolution Incubations and Analysis: We incubated each mussel valve in a sealed 250 mL Nalgene bottle filled with modified seawater for 48 hours in a temperature-controlled room, recording seawater properties in each bottle before and at the end of each incubation, including temperature, salinity, and pH (Yellow Springs Instruments Professional Plus Sonde). YSI sonde pH values were calibrated to the total scale based on pH spectrophotometric measurements of m-cresol dye absorbance at the incubation temperature. Incubation bottles were agitated gently once every 8 hours over the course of the incubations to reduce the establishment of strong chemical gradients of their fluid contents. We took discrete seawater samples at the onset and end of each incubation for determination of total alkalinity concentration (TA); these “before-after” measurements of TA enabled quantification of the increase in alkalinity within each bottle over the duration of the incubation, and thereby the rate of dissolution of calcium carbonate (CaCO3) shell material for individual valves of known periostracum cover. Shell dissolution rates were quantified using standard alkalinity anomaly techniques (analyzing seawater samples in triplicate and selecting median TA for dissolution quantification), which relate calcium carbonate shell loss to an increase in the total alkalinity (TA) of surrounding seawater, per unit time. Across the dissolution trials, control incubations (15% of daily sample size) of modified seawater were conducted to verify minimal background changes in TA.
Seawater manipulation: We used direct chemical modification to the seawater carbonate system in order to create seawater treatments (pH = 7.4). To modify, equimolar additions of 1 M sodium bicarbonate (NaHCO3) and 1 M hydrochloric acid (HCl) were added to filtered seawater, capped, and were well mixed. This method results in an increase of dissolved inorganic carbon (DIC) and reduction of seawater pH without changing total alkalinity (TA).
