Data were collected on several single-day cruises on small privateers out of the Virginia Institute of Marine Science, Gloucester Point, VA.

High-resolution sampling via Dataflow was performed along the lower York River estuary during two successive summers, 2020-21, before, during, and after intense blooms of Margalefidinium polykrikoides and Alexandrium monilatum for determinations of pCO2, temperature, salinity, pH, turbidity, chlorophyll-a, and dissolved oxygen (DO). High-resolution sampling was performed with a Dataflow system (Madden & Day, 1992) modified as described in Crosswell et al. (2017). The pCO2-Dataflow system is instrumented with a pCO2 analyzer, a multi-parameter datasonde (YSI 6600V2), Garmin global positioning system (GPS MAP 546S), and data acquisition system. The system continuously samples surface water (approximately every 30 meters (m) at an average speed of 20 knots) from a stern-mounted water intake located 0.5 m below the water surface with a pump, which delivers water in parallel to (1) a showerhead equilibrator and (2) a flow-through cell attached to the YSI which is configured to measure water temperature, salinity, pH, turbidity, chlorophyll-a fluorescence, and DO. pCO2 in the equilibration chamber is determined by recirculating a carrier gas at a flow of approximately 1.5 liters per minute (L/min) through the equilibrator chamber and a nondispersive infrared absorbance detection analyzer (LI-COR LI-840).

Concurrent with Dataflow sampling, grab sampling was performed at five stations within bloom patches (as determined by levels of chlorophyll-a) and at five stations outside of bloom patches for determinations of dissolved inorganic carbon (DIC), dissolved organic carbon (DOC), total dissolved nitrogen (TDN), nitrate (NO3), nitrite (NO2), ammonium (NH4), dissolved inorganic phosphorus (DIP), active chlorophyll-a (via extraction), and cell abundance of M. polykrikoides and A. monilatum. DIC samples were filtered through 0.2-micrometer (um) polycarbonate filters into 8-milliliter (mL) hungate tubes and refrigerated underwater until analysis on an Apollo SciTech AS-C3 analyzer coupled to a LI-COR LI-7000 infrared gas analyzer. DOC samples were filtered through 0.45 um polyethersulfone filters and frozen prior to analysis on a Shimadzu TOC-VCSN combustion analyzer. Nutrient samples were also filtered through 0.45 um polyethersulfone filters and frozen prior to analysis on a Lachat QuikChem 8000 automated ion analyzer (Lachat Instruments, Milwaukee, WI, USA); detection limits for NO3−, NH4+, and PO43- are 0.20, 0.36, and 0.16 micromolar (uM), respectively. Chlorophyll-a samples for extraction were filtered through 0.7 um glass fiber filters which were frozen prior to analysis following Arar and Collins (1997, EPA Method 445.0). Samples were extracted in the dark for 24 hours in 8 mL of a 45:45:10 dimethyl sulfoxide : acetone: distilled water solution with 1% diethylamine (Shoaf & Lium, 1976), and read on a 10 AU Turner Designs fluorometer before and after acidification to compute active chlorophyll-a.

Cell concentrations of M. polykrikoides (Marg) and A. monilatum (Alex) by qPCR were measured as described in Wolney et al. (2020). Briefly, 100 mL water samples were collected, and 25-100 mL were filtered onto 3 um Isopore membrane filters (Millipore Corp., Darmstadt, Germany), with the volume filtered for DNA extraction dependent on Dataflow-measured chlorophyll-a concentrations. Disposable filtration units were used to prevent cross contamination between samples. DNA was extracted from the filters using the Qiamp Fast Stool Mini Kit (QIAGEN Corp., Germantown, MD, USA) using the modified protocol as described in Wolney et al. (2020). DNA was amplified targeting Marg and Alex DNA using TaqMan qPCR assays designed in the Reece laboratory with York River Marg and Alex sequences included for assay design (Vandersea et al., 2017; Wolney et al., 2020). The cell concentrations of Marg and Alex cultures were determined by microscopy using a Sedgwick-Rafter counting cell chamber and DNA was extracted from a known number of cells. This material was used as positive control material and to generate standard curves by serially diluting the DNA to achieve a range of cell number equivalents.

Data represent means and standard errors (SE) at the five bloom and five non-bloom stations on each cruise. Dataflow values represent means and SE of all readings while sampling at each site; grab sample values represent the means and SE of three replicate samples.