Estuaries and Coasts

, Volume 37, Supplement 1, pp 147–163

Interannual Variability Influences Brown Tide (Aureococcus anophagefferens) Blooms in CoastalEmbayments

Authors

    • Ocean, Earth and Atmospheric SciencesOld Dominion University
    • Department of Marine ScienceCoastal Carolina University
  • Margaret R. Mulholland
    • Ocean, Earth and Atmospheric SciencesOld Dominion University
Article

DOI: 10.1007/s12237-013-9683-3

Cite this article as:
Boneillo, G.E. & Mulholland, M.R. Estuaries and Coasts (2014) 37: 147. doi:10.1007/s12237-013-9683-3

Abstract

Blooms of Aureococcus anophagefferens in Chincoteague Bay were observed during 5 of 6 years between 2002 and 2007. In order to understand factors controlling blooms, interannual differences in nitrogen and carbon uptake and concentrations of dissolved constituents were compared at two sites in Chincoteague Bay, MD and VA over the 6-year time period. Over that time, we observed that there was no single nitrogen compound that fueled blooms each year. Instead, A. anophagefferens took up a wide range of nitrogen compounds to meet its nutritional demands. Although photosynthetic carbon fixation was the dominant form of carbon acquisition during blooms, organic carbon uptake contributed up to 30 % of the total carbon uptake. In addition to interannual variability in nitrogen and carbon uptake, we observed that there was an increase in bloom intensity and duration over the 6-year study period during which dissolved organic carbon appeared to accumulate in the system.

Keywords

Aureococcus anophagefferensBrown tideNutrientsDissolved organic matter

Introduction

Since 1985, Aureococcus anophagefferens blooms (brown tides) have occurred regularly in coastal lagoons and other embayments along the mid-Atlantic coast of North America between Cape Hatteras and Cape Cod (Milligan and Cosper 1997; Bricelj and Lonsdale 1997; Gobler et al. 2005). Surveys of A. anophagefferens abundance have shown that its geographic distribution is much broader and encompasses the entire east coast of the USA, with cells detected (1–200 cells mL−1) as far north as Maine (Anderson et al. 1993) and as far south as Florida (Popels et al. 2003). Although detected along the entire US east coast, blooms of A. anophagefferens (concentrations >35,000 cells mL−1; Gastrich and Wazniak 2002) had been confined to coastal embayments between Massachusetts and Maryland at the outset of this study where they have varied in their frequency and intensity from year to year (Gobler et al. 2005). In this study, we report on major brown tide events in both Maryland and Virginia between 2002 and 2007.

A. anophagefferens has the ability to take up a wide range of nitrogen (N) compounds to meet its N requirement for growth. Previous studies have shown that A. anophagefferens has a high affinity for ammonium (NH4+) and urea (Lomas et al. 1996; Berg et al. 1997). A. anophagefferens can also obtain a significant amount of its N through the uptake of dissolved free amino acids (DFAA) (Mulholland et al. 2002). A recent study of the A. anophagefferens genome determined that A. anophagefferens can utilize at least eight different forms of N (Berg et al. 2008). Since A. anophagefferens can utilize both dissolved inorganic N (DIN) and dissolved organic N (DON) compounds, this organism may have a competitive advantage over other phytoplankton species that can only use DIN. However, despite observations of its nutritional flexibility, blooms of A. anophagefferens have been linked to DIN depletion and high concentrations of DON relative to DIN (Lomas et al. 2004).

Previous work has demonstrated that in addition to photosynthetic carbon (C) uptake, A. anophagefferens can take up dissolved organic carbon (DOC) compounds (Dzurica et al. 1989; Mulholland et al. 2002) and organic C amendments stimulated A. anophagefferens growth in incubations of natural populations (Gobler and Sañudo-Wilhelmy 2001b). During intense monospecific blooms, there can be a significant drawdown of DOC (Gobler et al. 2004), suggesting that A. anophagefferens is taking up DOC in the environment. The ability to take up DOC may supplement autotrophic C uptake via photosynthetic CO2 fixation and may be advantageous during blooms when cell densities are high (e.g., 1.0 × 106 cells mL−1) and self-shading or depletion of dissolved inorganic C (DIC) might limit photosynthetic C uptake (Mulholland et al. 2009).

Despite the recognition that A. anophagefferens is mixotrophic and can take up DOC, the proportion of the carbon demand that is obtained through autotrophic uptake of DIC versus uptake of DOC has not been evaluated for this species or most phytoplankton mixotrophs. Similarly, it is not known how auto- and heterotrophic C uptake changes over the course of blooms. If A. anophagefferens can compensate for reduced photosynthetic C uptake with heterotrophic C uptake when conditions limit photoautrophic metabolism, they may be able to out-compete strictly autotrophic species whose growth may become light or C limited.

In this study, we compare rates of photosynthetic bicarbonate uptake and the uptake of organic and inorganic N and C over the course of several brown tide blooms over several years, between 2002 and 2007, at two sites in Chincoteague Bay, MD and VA. The two sites contain comparable bacterial and phytoplankton communities (Mulholland et al. 2009). One site had experienced brown tide blooms since at least 1999, while no blooms had been reported at the other site prior to this study. In addition to determining how nutrient and C uptake varied over the course of these blooms, we examined relative changes in the contribution of organic C and N to the total C and N nutrition of A. anophagefferens populations as blooms initiated, developed, and persisted. Finally, we compare nutrient and C and N utilization patterns over several years in order to understand nutrient controls on blooms on interannual timescales and to identify possible triggers for blooms. Results from this study may be useful for designing interventions or taking management actions to prevent the initiation of potentially damaging blooms.

Materials and Methods

We examined interannual and intersite variability in A. anophagefferens blooms within Chincoteague Bay, a coastal bay along the mid-Atlantic coast of North America. Chincoteague Bay extends from Maryland to Virginia and has two small inlets at its southern and northern ends that allow it to exchange water with the Atlantic Ocean (Fig. 1). This configuration causes the bay to have a fairly long residence time (about 63 days; Pritchard 1960). Because of the lack of riverine input, the salinity in Chincoteague Bay is closer to that of seawater than freshwater and ranged between 21 and 33 ppt during this study. The watershed for the bay is approximately 72.6 square miles and is made up mostly of forested areas and wetlands. However, the main source of freshwater coming into Chincoteague Bay is groundwater. Thirty-three percent of the watershed is made up of agricultural lands (Maryland DNR 2005). Fertilizers applied to these lands contain a high percentage of urea (Glibert et al. 2006), and these agricultural nutrients can enter the coastal bays through runoff, tidal creeks, and groundwater.
https://static-content.springer.com/image/art%3A10.1007%2Fs12237-013-9683-3/MediaObjects/12237_2013_9683_Fig1_HTML.gif
Fig. 1

Map of the study area in Chincoteague Bay, MD and VA. The two study sites at Public Landing, MD, USA and Greenbackville, VA, USA are marked with a circle

Brown tide blooms have been monitored by the Maryland Department of Natural Resources in Chincoteague Bay since 1999. For this study, weekly measurements were taken at two sites in Chincoteague Bay, MD and VA; one site that had previously experienced brown tide blooms, Public Landing, Maryland (PL), and one site where no previous blooms (as of 2002) were reported, Greenbackville, VA, USA (GB) (Fig. 1). At each site, a Hydrolab Surveyor 4a Water Quality Multiprobe equipped with sensors for temperature, salinity, pH, and photosynthetically active irradiance (PAR) was deployed to record physical parameters at the sampling depth just below the surface prior to collecting water samples. Water was then collected using a clean bucket, placed in an acid-cleaned 20 L polyethylene carboy, and transported to the Marine Science Consortium laboratory located in Greenbackville, VA, USA; this was a 30-min car ride from PL and onsite for GB. Carboys, buckets, and all other materials associated with the sampling, handling, and storage of seawater during this project were soaked in 10 % HCl between sampling events, and rinsed liberally with distilled-deionized water before each use. Because Chincoteague Bay is shallow (∼4 m) and well mixed, it is likely that the sample water collected near the surface was representative of the entire water column.

Upon arrival at the laboratory, nutrient samples were filtered using a 0.2-μm Supor filter disk (2002 and 2003) or a 0.2-μm Supor cartridge filter (2006 and 2007). The filtrate was frozen in acid-cleaned bottles for subsequent analyses of dissolved nutrient concentrations (see below). Nutrient samples were collected within 30 min of sample collection. Chlorophyll a (Chl a) samples were collected onto GF/F filters that were then frozen and analyzed within 2 weeks of their collection. Whole water samples were preserved with glutaraldehyde (1 % final concentration) in sterile polycarbonate bottles for later enumeration of bacteria and A. anophagefferens. Counts were performed within 72 h of sample collection.

Nitrate plus nitrite (NO3/NO2), phosphate, and urea concentrations were measured colorimetrically using an Astoria Pacific Autoanalyzer (Parsons et al. 1984) according to the manufacturer’s specifications. Ammonium concentrations were determined using the manual phenol hypochlorite method (Solorzano 1969). Total dissolved nitrogen (TDN) and phosphorus (TDP) were measured after persulfate oxidation (Valderrama 1981). Dissolved organic N (DON) and dissolved organic P (DOP) were calculated as the difference between TDN and DIN and TDP and dissolved inorganic P (DIP), respectively. DFAA concentrations were measured using a Shimadzu high performance liquid chromatography (HPLC) equipped with a fluorescence detector (Cowie and Hedges 1992).

Chl a concentrations were measured using standard fluorometric methods (Welschmeyer 1994). Bacteria and A. anophagefferens cells were enumerated using epifluorescent microscopy. Heterotrophic bacteria were first stained with 4′,6-di-amidinophenyl-indole as outlined by Porter and Feig (1980). A. anophagefferens were enumerated using the immunofluorescence (fluorescein isothiocyanate) method of Anderson et al. (1989). The protocol was modified by doubling the amount of primary and secondary antibody (Mulholland et al. 2009). Samples were gently (<5 kPa) filtered onto 0.8-μm black polycarbonate filters for counting (Anderson et al. 1989). A minimum of 100 cells were counted per sample in at least 10 fields to yield a relative standard deviation of 9 % for replicate counts of the same sample (n = 6)at cell densities of 2 × 105 cells mL−1, within the range of average A. anophagefferens cell densities during blooms. Blooms were defined as A. anophagefferens concentrations >35,000 cells mL−1 (Gastrich and Wazniak 2002).

The amount of Chl a contributed by A. anophagefferens was estimated by assuming a constant Chl a content per cell for A. anophagefferens (0.035 ± 0.003 pg/cell), as has been done previously (Gobler and Sañudo-Wilhelmy 2001a; Gobler et al. 2002; Mulholland et al. 2002). Variability in cellular Chl a concentrations could potentially bias these calculations.

Nutrient uptake experiments were conducted from March–October (2002) (Mulholland et al. 2009) and from May–July (2003, 2006, and 2007) during daylight hours. Whole water was placed into acid-cleaned polycarbonate bottles and experiments to estimate N and C uptake were initiated by adding highly enriched (96–99 %) 15 N- and 13C-labeled substrates (NH4+, NO3, urea, bicarbonate, glucose, and leucine) to incubation bottles. Urea and amino acids were dually labeled with 15 N and 13C. Additions of labeled substrate were 0.03 μmol L−1. This represented an atom percent enrichment ranging from 1 to 94 %. However, for the majority of the incubations, atom percent enrichment ranged from 5 to 10 %; enrichments between 1 and 10 % yield accurate estimates of in situ uptake rates (Mulholland et al. 2009). Higher enrichment levels may overestimate actual uptake rates if additions stimulated nutrient uptake, but because nutrient concentrations were rarely below <0.1 μM N for all of the N substrates used for uptake experiments, high atom percent enrichments were rarely measured in our incubations. Incubation bottles were placed in incubators where temperatures were maintained within 2 °C of ambient levels in Chincoteague Bay under ambient light conditions. The average incoming solar radiation during light incubations ranged from 49 to 2,234 μE m−2 s−1 (measured using the Hydrolab dual-PAR sensor).

After 15–30 min, N uptake incubation experiments were terminated by filtering the sample onto a precombusted (450 °C for 2 h) GF/C filter (nominal pore size of 1.2 μm), rinsed with filtered seawater, and filters were stored frozen until analysis. Light and dark bicarbonate incubations were terminated after 2–3 h. The frozen filters were dried at 50 °C for 48 h in a drying oven and pelletized in tin disks for isotopic analysis. The isotopic composition of samples was determined using a Europa Scientific isotope ratio mass spectrometer, equipped with an automated nitrogen and carbon analyzer. Uptake rates were calculated using the equations from Mulholland et al. (2009). Isotope dilution was not measured in these short incubations; substantial isotope dilution during incubations would result in underestimates of uptake rates.

N and C content of the DFAA pool was calculated based on the C/N ratio of the ambient DFAA pool from individual HPLC runs during 2002, as described by Mulholland et al. (2002). We established that, on average, there was 1.2 (±0.2) μmol N (μmol DFAA)−1 and 4.4 (±0.5) μmol C (μmol DFAA)−1. The ambient DIC concentrations were calculated based on salinity assuming that CO2 concentrations were saturating in water samples. Glucose concentrations were estimated as 2 % of the ambient DOC pool, the lower end of the range estimated by Benner (2002; 2–6 %) for marine surface waters.

Results

Physical Parameters

Seasonal temperature patterns did not show much variation from year to year (Tables 1 and 2). Typically, there was a period of rapid warming between April and May with maximum water temperatures at the end of July. Salinity, however, did show a high degree of interannual variability, varying inversely with rainfall. In 2002, a drought year, salinities were >30 throughout the sampling period (Simjouw et al. 2004; Mulholland et al. 2009). During 2003, a near record wet year, salinity ranged from 21.5 to 27.1 between April and June. In 2006 and 2007, salinities were generally lower than in 2002 and higher than 2003, our two extreme years, ranging from 25.0 to 32.8 (Tables 1 and 2). Typically, salinities were higher at Greenbackville than Public Landing, likely because GB is closer to Chincoteague Inlet where water is exchanged with the ocean. After large rain events, however, the salinity at Greenbackville was lower than that at Public Landing due to its proximity to Swans Gut Creek, which drains directly into Chincoteague Bay just south of that sampling site (Tables 1 and 2).
Table 1

Physical, biological, and chemical parameters at Public Landing in Chincoteague Bay, MD and VA, during 2003, 2006, and 2007

Date

Salinity

Temperature (°C)

pH

Chl a (μg Chl L−1)

A. anophagefferens (cells mL−1) × 105

Bacteria (cells mL−1) × 105

PC (μmol C L−1)

PN (μmol N L−1)

NH4+ (μmol L−1)

NO3/NO2 (μmol L−1)

DIP (μmol L−1)

DIN/DIP

2003

 07 Mar

24.3

4.9

7.9

5.10 (0.1)

BDL

3.86 (0.02)

  

0.23 (0.01)

0.77 (0.01)

0.33 (0.00)

3.5

 14 Apr

24.9

12.8

7.9

4.35 (0.3)

BDL

5.15 (0.02)

  

0.18 (0.02)

1.40 (0.02)

0.20 (0.00)

8.2

 14 May

26.0

18.8

7.9

0.83 (0.5)

1.59 (0.17)

5.61 (0.04)

  

0.25 (0.02)

0.93 (0.02)

0.72 (0.00)

8.4

 29 May

26.5

16.9

8.0

7.15 (0.7)

2.50 (0.31)

13.34 (0.08)

  

0.17 (0.03)

0.89 (0.00)

0.59 (0.00)

3.8

 04 Jun

25.2

18.4

8.0

6.87 (1.3)

4.91 (0.53)

12.14 (1.06)

  

0.28 (0.03)

0.65 (0.01)

0.67 (0.09)

2.2

 12 Jun

25.9

19.1

8.0

9.66 (0.7)

0.06 (0.01)

11.13 (0.14)

  

0.36 (0.08)

0.54 (0.02)

0.58 (0.04)

5.1

 18 Jun

25.7

24.6

7.9

4.52 (0.2)

3.51 (0.56)

12.76 (0.08)

  

0.15 (0.01)

0.64 (0.00)

0.50 (0.35)

3.2

 27 Jun

24.5

21.9

7.9

21.8 (0.5)

0.89 (0.16)

15.67 (0.15)

  

0.14 (0.07)

0.73 (0.00)

0.56 (0.21)

3.3

 10 Jul

26.0

26.5

8.0

11.2 (0.5)

BDL

10.98 (0.03)

  

0.29 (0.06)

0.84 (0.02)

0.45 (0.02)

2.5

 07 Aug

25.8

29.3

7.8

8.04 (0.2)

BDL

   

0.49 (0.04)

1.03 (0.10)

0.43 (0.04)

3.0

2006

 10 May

30.4

17.1

8.8

6.4 (0.3)

0.17 (0.09)

7.97 (0.59)

126 (6)

14 (0.8)

0.61 (0.03)

0.08 (0.01)

0.06 (0.05)

0.7

 18 May

30.3

21.0

8.8

8.6 (0.1)

4.88 (0.69)

9.67 (0.38)

191 (11)

15 (1.1)

1.08 (0.02)

0.13 (0.01)

0.11 (0.01)

0.5

 23 May

31.1

18.3

8.8

8.7 (0.5)

5.03 (0.79)

12.07 (0.88)

261 (12)

20 (0.8)

0.93 (0.05)

0.17 (0.02)

0.03 (0.01)

1.0

 31 May

31.4

19.3

8.9

12.9 (0.5)

7.11 (0.95)

16.43 (0.80)

291 (15)

22 (0.8)

0.55 (0.01)

0.19 (0.08)

0.07 (0.04)

0.7

 07 Jun

31.2

21.8

8.7

25.7 (0.4)

13.1 (1.19)

16.19 (0.66)

380 (13)

32 (1.2)

0.64 (0.01)

0.11 (0.01)

0.66 (0.01)

0.4

 14 Jun

31.8

21.7

8.8

17.6 (0.6)

12.0 (0.58)

17.74 (1.50)

347 (17)

31 (1.5)

0.42 (0.00)

0.21 (0.09)

0.55 (0.08)

0.3

 21 Jun

30.7

26.5

8.9

28.5 (0.4)

5.96 (0.45)

22.13 (1.81)

397 (17)

33 (1.1)

0.46 (0.08)

0.11 (0.05)

0.46 (0.07)

0.2

 28 Jun

30.3

25.4

8.6

26.8 (1.6)

 

25.38 (0.99)

  

0.51 (0.05)

0.15 (0.05)

2.32 (0.00)

0.2

 05 Jul

30.8

27.8

8.7

11.2 (0.3)

6.19 (0.51)

   

0.65 (0.02)

0.11 (0.00)

1.58 (0.12)

0.2

2007

 15 May

25.6

18.4

8.8

10.8 (0.1)

4.04 (0.54)

12.80 (0.74)

269 (25)

20 (2.1)

0.25 (0.02)

1.33 (0.01)

0.04 (0.00)

4.6

 19 May

26.7

19.3

8.9

20.7 (0.3)

5.02 (0.29)

11.54 (0.82)

244 (13)

25 (1.3)

1.07 (0.01)

1.64 (0.01)

0.25 (0.00)

6.8

 29 May

26.9

24.0

8.8

19.5 (0.2)

8.94 (1.28)

20.13 (1.42)

371 (26)

31 (2.2)

0.33 (0.02)

2.00 (0.08)

0.10 (0.00)

2.1

 05 Jun

26.9

25.3

8.9

27.7 (0.4)

10.7 (0.87)

22.30 (1.07)

421 (40)

35 (2.5)

0.25 (0.02)

1.31 (0.01)

0.13 (0.00)

5.6

 12 Jun

27.9

24.3

8.9

34.4 (0.7)

10.9 (1.10)

26.25 (0.78)

498 (26)

38 (2.7)

0.23 (0.01)

1.03 (0.01)

0.36 (0.00)

2.5

 19 Jun

28.3

25.0

8.9

28.7 (1.0)

13.2 (1.10)

28.07 (1.20)

428 (37)

34 (1.6)

0.18 (0.02)

0.80 (0.01)

0.29 (0.01)

3.8

 26 Jun

28.7

25.2

8.9

33.6 (0.3)

14.1 (1.59)

28.73 (1.32)

450 (33)

38 (4.0)

0.51 (0.01)

0.02 (0.07)

0.29 (0.01)

0.9

 03 Jul

28.3

24.3

8.9

21.1 (0.7)

5.15 (0.43)

31.34 (0.98)

222 (24)

27 (1.7)

1.07 (0.01)

0.94 (0.01)

0.78 (0.00)

1.1

 10 Jul

28.6

28.5

8.8

15.6 (0.9)

9.51 (0.54)

28.79 (1.51)

271 (16)

27 (4.8)

0.82 (0.06)

1.13 (0.00)

0.30 (0.01)

1.1

Standard deviations are in parentheses and BDL indicates that analytes were below the limits of analytical detection (about 0.03 μmol L−1 for nutrients and 0.2 × 103 cell mL−1 for A. anophagefferens abundance). Dissolved inorganic N was calculated as the sum of the average NH4+ and NO3/NO2 concentrations and ratios were calculated from average values. Salinity, temperature, Chl a and A. anophagefferens abundance from 2003 are also reported in Minor et al. (2006). Empty fields indicate that there were no data

Table 2

Physical, biological, and chemical parameters at Greenbackville in Chincoteague Bay, MD and VA, during 2003, 2006, and 2007

Date

Salinity

Temperature (°C)

pH

Chl a (μg Chl L−1)

A. anophagefferens (cells mL−1) × 105

Bacteria (cells mL−1) × 105

PC (μmol C L−1)

PN (μmol N L−1)

NH4+ (μmol L−1)

NO3/NO2 (μmol L−1)

DIP (μmol L−1)

DIN/DIP

2003

 07 Mar

22.5

6.0

7.8

5.11 (0.0)

BDL

2.67 (0.15)

  

0.27 (0.00)

1.19 (0.01)

0.42 (0.04)

3.5

 14 Apr

21.5

12.0

8.1

6.90 (0.0)

BDL

10.78 (0.32)

  

1.34 (0.04)

0.79 (0.00)

0.26 (0.00)

8.2

 14 May

26.7

19.0

7.8

0.57 (0.0)

BDL

2.42 (0.21)

  

0.20 (0.01)

1.48 (0.07)

0.20 (0.17)

8.4

 29 May

23.6

17.9

7.9

8.25 (0.6)

2.90 (0.33)

8.09 (1.12)

  

1.01 (0.03)

0.70 (0.00)

0.45 (0.01)

3.8

 04 Jun

24.2

18.8

8.0

10.89 (0.4)

6.77 (0.40)

13.30 (0.77)

  

0.21 (0.02)

0.64 (0.02)

0.39 (0.03)

2.2

 12 Jun

25.1

24.3

8.0

14.88 (2.0)

7.24 (0.51)

9.86 (0.47)

  

1.31 (0.01)

0.78 (0.04)

0.41 (0.02)

5.1

 18 Jun

25.7

22.4

8.0

9.51 (0.3)

3.58 (0.43)

9.22 (0.25)

  

0.27 (0.01)

1.19 (0.03)

0.46 (0.05)

3.2

 27 Jun

27.1

27.5

8.1

13.73 (1.1)

1.11 (0.06)

   

0.25 (0.02)

1.47 (0.20)

0.52 (0.22)

3.3

 10 Jul

27.5

29.3

7.8

15.61 (0.2)

BDL

   

1.07 (0.01)

1.81 (0.01)

1.15 (0.00)

2.5

 07 Aug

29.7

25.9

7.9

11.17 (0.9)

BDL

   

0.33 (0.02)

1.65 (0.00

0.67 (0.00)

3.0

2006

 10 May

31.8

18.3

8.7

7.1 (0.23)

0.10 (0.02)

10.37 (0.77)

82 (8.1)

7 (0.7)

0.40 (0.01)

0.10 (0.06)

0.69 (0.10)

0.7

 18 May

31.8

21.9

8.6

7.9 (0.5)

0.22 (0.04)

12.06 (0.50)

199 (12)

10 (1.2)

0.47 (0.05)

0.09 (0.01)

1.23 (0.21)

0.5

 23 May

32.4

19.2

8.5

2.7 (0.1)

0.29 (0.04)

9.56 (1.30)

52 (5.2)

6 (0.6)

0.48 (0.03)

0.34 (0.02)

0.83 (0.10)

1.0

 31 May

32.8

20.3

8.6

12.5 (0.2)

2.19 (0.22)

12.68 (1.78)

195 (11)

20 (1.1)

0.73 (0.02)

0.31 (0.09)

1.46 (0.08)

0.7

 07 Jun

32.0

21.4

8.7

24.5 (0.4)

7.69 (0.60)

19.65 (1.98)

141 (8.6)

11 (0.4)

0.48 (0.00)

0.27 (0.04)

1.76 (0.16)

0.4

 14 Jun

31.2

21.1

8.7

31.4 (0.3)

7.31 (0.55)

19.01 (1.41)

949 (44)

79 (3.2)

0.44 (0.02)

0.18 (0.05)

2.20 (0.20)

0.3

 21 Jun

31.7

26.4

8.8

26.8 (0.8)

12.7 (0.91)

23.76 (3.75)

359 (20)

32 (0.6)

0.31 (0.00)

0.18 (0.08)

3.24 (0.24)

0.2

 28 Jun

25.0

25.8

8.5

19.6 (0.2)

7.90 (1.22)

18.32 (2.39)

  

0.56 (0.01)

0.15 (0.13)

3.21 (0.03)

0.2

 05 Jul

27.7

27.7

8.5

12.8 (0.2)

    

0.53 (0.06)

0.20 (0.07)

4.28 (0.79)

0.2

2007

 15 May

26.6

19.2

8.7

15.3 (0.1)

2.76 (0.18)

11.44 (0.98)

275 (14)

22 (2.5)

0.27 (0.00)

1.43 (0.01)

0.37 (0.00)

4.6

 19 May

28.2

18.2

8.6

10.4 (0.1)

4.12 (0.36)

12.93 (0.67)

113(6)

12 (1.5)

1.34 (0.04)

2.13 (0.02)

0.51 (0.00)

6.8

 29 May

28.4

24.1

8.7

25.3 (0.5)

8.27 (1.00)

21.65 (0.92)

430 (17)

38 (3.1)

0.20 (0.01)

1.19 (0.01)

0.66 (0.00)

2.1

 05 Jun

30.0

25.4

8.6

12.2 (0.2)

5.02 (0.42)

24.93 (0.88)

205 (13)

17 (2.6)

1.01 (0.03)

2.10 (0.05)

0.56 (0.00)

5.6

 12 Jun

30.2

24.9

8.7

30.5 (0.2)

15.1 (0.67)

25.97 (1.33)

866 (20)

37 (1.9)

0.21 (0.02)

1.06 (0.01)

0.50 (0.01)

2.5

 19 Jun

30.2

25.9

8.8

29.9 (1.0)

11.4 (1.50)

22.72 (1.33)

479 (14)

40 (3.1)

3.64 (0.25)

0.81 (0.03)

1.18 (0.00)

3.8

 26 Jun

30.5

25.7

8.8

45.3 (1.0)

13.3 (1.42)

32.15 (1.34)

400 (25)

40 (3.6)

0.51 (0.02)

0.86 (0.01)

1.53 (0.01)

0.9

 03 Jul

30.2

24.8

8.7

33.2 (0.3)

5.23 (0.37)

33.43 (1.83)

169 (22)

26 (2.6)

0.94 (0.11)

1.04 (0.02)

1.88 (0.01)

1.1

 10 Jul

29.4

29.2

8.7

22.0 (1.0)

2.23 (0.28)

38.67 (1.93)

143 (12)

19 (1.8)

0.74 (0.03)

0.70 (0.01)

1.37 (0.01)

1.1

Standard deviations are in parentheses and DL indicates that analytes were below the limit of analytical detection (about 0.03 μmol L−1 for nutrients and 0.2 × 103 cell mL−1 for A. anophagefferens abundance). Dissolved inorganic N was calculated as the sum of the average NH4+ and NO3/NO2 concentrations and ratios were calculated from average values. Salinity, temperature, Chl a and A. anophagefferens abundance from 2003 are also reported in Minor et al. (2006). Empty fields indicate that there were no data

Microbial Biomass

In contrast to 2002, when there was no brown tide bloom and only low A. anophagefferens cell densities at GB (Mulholland et al. 2009), there were blooms at both sites in 2003 (Minor et al. 2006), 2004 (Fig. 2), 2006, and 2007 (Fig. 2 and Tables 1 and 2). During 2003, A. anophagefferens abundance at PL increased during the spring and reached a peak concentration of 4.9 × 105 cells mL−1 in June (Fig. 2a and Table 1). Despite there being no previous reports of brown tide blooms at GB, there were higher overall A. anophagefferens cell concentrations during the bloom there than at PL (up to 7.2 × 105 cells mL−1 on June 12) during 2003, and cell concentrations remained high at GB through the end of June (Fig. 2b and Table 2). In 2004, there were again blooms at both sampling sites, and A. anophagefferens cell concentrations reached 6.4 × 105cells mL−1 at GB on June 18 and 8.8 × 105 cells mL−1 at PL on June 26 (Fig. 2). In 2005, there were no brown tide blooms at either site (data not shown). In 2006 and 2007, both sites again experienced intense brown tide blooms with A. anophagefferens cell numbers in excess of 105 cells mL−1 at both sites (Fig. 2). During 2006 and 2007, we observed the highest A. anophagefferens cell densities, and intense bloom conditions persisted longer than during other years with blooms initiating in May and lasting through early July (Fig. 2; Tables 1 and 2).
https://static-content.springer.com/image/art%3A10.1007%2Fs12237-013-9683-3/MediaObjects/12237_2013_9683_Fig2_HTML.gif
Fig. 2

A. anophagefferens abundance during 2002, 2003, 2004, 2006, and 2007 blooms in Chincoteague Bay at a Greenbackville, VA, USA and b Public Landing, MD, USA. Figure includes data reported in Simjouw et al. (2004), Minor et al. (2006), and Mulholland et al. (2009)

Nutrient Dynamics

Unlike 2002, when NH4+concentrations were below detection limits on several occasions at both sites (Mulholland et al. 2009), NH4+ was always measureable during blooms at PL and GB in 2003, 2006, and 2007 (Tables 1 and 2; no nutrient data were collected in 2004). In 2003, NH4+concentrations ranged from 0.20 to 1.34 μmol L−1, and concentrations at GB were usually higher than those at PL. In 2006, NH4+ concentrations were always detectable at both sites but <1.00 μmol L−1 on all but one sampling date. In 2007, NH4+concentrations ranged from 0.18 to 3.64 μmol L−1 but were usually <1.00 μmol L−1.

During 2002, NO3/NO2 concentrations at PL and GB were below analytical detection (0.03 μmol L−1) during and after the bloom (Mulholland et al. 2009). In contrast, during 2003, 2006, and 2007, NO3/NO2 concentrations were detectable throughout the bloom period at both sites in Chincoteague Bay (Tables 1 and 2). In 2003, NO3/NO2 concentrations ranged from 0.54 to 1.81 μmol L−1. In 2006, NO3/NO2 concentrations were lower, but always measureable, ranging from 0.09 to 0.48 μmol L−1. In 2007, NO3/NO2 concentrations were higher in excess of 1.00 μmol L−1 even during the initiation of brown tide blooms in May and reaching concentrations of 2.13 μmol L−1.

During 2002, urea concentrations were generally higher (0.24–2.30 μmol L−1; Mulholland et al. 2009) than during subsequent sampling years when concentrations were consistently <1.0 μmol L−1 (Tables 3 and 4). During 2003, urea concentrations ranged from 0.10 to 0.89 μmol L−1, higher than the range observed during 2006 and 2007 (0.02–0.63 μmol L−1). A notable exception was in July 2007 when urea concentrations were 3.61 and 3.65 μmol L−1 at GB and PL, respectively. DFAA concentrations were fairly consistent across and within years. Typically, DFAA concentrations were <1.0 μmol L−1 (Table 4). The average C and N concentration for the DFAA pool during 2002 was 1.2 μmol N (μmol DFAA)−1 and 4.4 μmol C (μmol DFAA)−1 (data not shown). The most abundant amino acids were serine, glycine, and histidine.
Table 3

Concentrations and ratios of organic nutrients at Public Landing in Chincoteague Bay, MD, USA during 2003, 2006, and 2007

Date

DOP (μmol L−1)

Urea (μmol L−1)

DFAA (μmol L−1)

DON (μmol L−1)

DOC (μmol L−1)

DOC/DON

DOC/DOP

DON/DOP

TDN/TDP

2003

 07 Mar

0.32

0.33 (0.00)

0.27 (0.05)

24.1

276 (1.3)

12

873

76

39.4

 14 Apr

0.26

0.20 (0.00)

0.38 (0.13)

28.0

268 (9.2)

10

1,021

107

66.8

 14 May

BDL

0.72 (0.00)

0.71 (0.04)

26.8

260 (1.4)

10

  

152

 29 May

0.07

0.67 (0.09)

0.58 (0.02)

36.4

313 (4.0)

9

4,611

536

78.4

 04 Jun

0.15

0.58 (0.04)

0.63 (0.38)

29.0

294 (1.3)

10

1,954

193

59.8

 12 Jun

0.25

0.50 (0.35)

0.27 (0.03)

34.3

286 (6.7)

8

1,154

138

93.1

 18 Jun

0.19

0.56 (0.21)

0.53 (0.03)

36.5

382 (11.5)

11

2,011

192

88.9

 27 Jun

0.14

0.45 (0.02)

0.94 (0.06)

40.1

353 (2.5)

9

2,562

291

108

 10 Jul

0.28

0.43 (0.04)

0.66 (0.01)

45.7

397 (2.7)

9

1,418

163

86.7

 07 Aug

0.16

0.67 (0.00)

0.40 (0.04)

47.7

376 (1.2)

8

2,350

298

100

2006

 10 May

0.41

BLD

0.25 (0.00)

33.9

352

10

859

82

73.4

 18 May

0.26

0.02 (0.02)

0.13 (0.02)

29.6

529

18

2,035

114

84.7

 23 May

0.49

0.36 (0.12)

0.43 (0.01)

36.2

466

13

951

73

70.9

 31 May

0.76

0.07 (0.01)

0.49 (0.02)

41.1

577

14

759

54

50.6

 07 Jun

0.29

0.12 (0.03)

0.40 (0.02)

35.7

1188

33

4,097

124

38.8

 14 Jun

0.37

0.22 (0.13)

0.35 (0.01)

38.0

580

15

1,568

102

42.3

 21 Jun

0.42

0.10 (0.05)

0.31 (0.01)

34.9

652

19

1,552

83

40.7

 28 Jun

BDL

0.14 (0.01)

0.31 (0.03)

41.9

710

17

  

19.5

 05 Jul

0.38

0.20 (0.03)

0.56 (0.05)

45.4

751

17

1,976

118

19.2

2007

 15 May

0.36

0.63 (0.01)

0.44 (0.05)

30.0

473

16

1,314

82

78.5

 19 May

0.32

0.21 (0.01)

0.83 (0.00)

27.8

378

14

1,181

88

54.2

 29 May

0.29

0.07 (0.01)

0.25 (0.02)

31.3

459

15

1,583

108

86.0

 05 Jun

0.34

0.15 (0.01)

0.24 (0.03)

36.1

630

17

1,853

105

79.4

 12 Jun

0.39

0.41 (0.01)

0.63 (0.01)

34.1

500

15

1,282

89

47.6

 19 Jun

0.42

0.09 (0.02)

0.41 (0.00)

32.1

476

15

1,133

76

46.8

 26 Jun

0.31

0.20 (0.02)

0.26 (0.01)

35.3

524

15

1,690

113

59.6

 03 Jul

0.37

3.65 (0.00)

0.24 (0.01)

43.7

561

13

1,516

119

40.0

 10 Jul

0.51

0.16 (0.03)

0.20 (0.03)

35.3

1,638

46

3,212

69

45.9

Standard deviations are in parentheses. DOC concentrations from 2003 are also reported in Minor et al. (2006). BDL indicates concentrations below the limit of analytical detection (<0.03 μmol L−1). Empty fields indicate that there were no data.

Table 4

Concentrations and ratios of organic nutrients at Greenbackville in Chincoteague Bay, VA, USA during 2003, 2006, and 2007

Date

DOP (μmol L−1)

Urea (μmol L−1)

DFAA (μmol L−1)

DON (μmol L−1)

DOC (μmol L−1)

DOC/DON

DOC/DOP

DON/DOP

TDN/TDP

2003

 07 Mar

0.38

0.36 (0.01)

0.56 (0.00)

37.3

344 (2.0)

9

911

99

48.6

 14 Apr

0.15

0.10 (0.04)

0.18 (0.03)

28.9

310 (4.4)

11

2,066

193

75.7

 14 May

0.74

0.88 (0.17)

0.74 (0.01)

33.7

242 (2.2)

7

326

45

37.5

 29 May

0.30

0.88 (0.02)

0.25 (0.00)

34.6

350 (2.7)

10

1,167

115

48.4

 04 Jun

0.22

0.67 (0.03)

0.74 (0.03)

36.7

315 (2.5)

9

1,430

167

61.5

 12 Jun

0.15

0.54 (0.02)

0.23 (0.01)

26.8

295 (5.0)

11

1,985

180

51.6

 18 Jun

0.06

0.22 (0.10)

1.53 (0.53)

34.8

336 (3.9)

10

5,600

580

69.7

 27 Jun

0.32

0.66 (0.04)

0.38 (0.11)

37.9

330 (1.1)

9

1,031

118

47.1

 10 Jul

BDL

0.82 (0.05)

0.42 (0.02)

65.6

338 (11.1)

5

  

59.1

 07 Aug

0.25

0.89 (0.12)

0.47 (0.03)

64.2

264 (0.5)

4

1,056

257

72.0

2006

 10 May

0.29

0.13 (0.03)

0.40 (0.01)

31.6

304

10

1,048

109

32.9

 18 May

BDL

0.29 (0.15)

0.29 (0.00)

33.2

324

10

  

31.5

 23 May

0.23

0.08 (0.04)

0.18 (0.03)

28.9

325

11

1,413

123

28.0

 31 May

BDL

0.11 (0.02)

0.27 (0.01)

26.5

374

14

  

24.5

 07 Jun

BDL

0.10 (0.07)

0.31 (0.01)

27.6

362

13

  

17.0

 14 Jun

BDL

0.03 (0.01)

0.60 (0.01)

36.4

442

12

11,050

845

16.5

 21 Jun

BDL

0.05 (0.02)

0.65 (0.03)

34.9

492

14

  

15.7

 28 Jun

BDL

0.22 (0.00)

0.33 (0.02)

41.3

597

14

  

14.4

 05 Jul

BDL

0.28 (0.18)

0.32 (0.04)

28.6

402

14

  

10.5

2007

 15 May

0.51

0.20 (0.04)

0.32 (0.13)

32.2

356

11

698

63

38.8

 19 May

0.25

0.27 (0.02)

0.40 (0.01)

31.3

257

8

1,028

126

46.3

 29 May

0.49

0.32 (0.02)

0.43 (0.03)

31.1

437

14

892

63

28.2

 05 Jun

0.26

0.31 (0.00)

 

23.3

254

11

977

88

32.6

 12 Jun

0.42

0.09 (0.01)

0.18 (0.01)

30.0

499

17

1,188

71

33.9

 19 Jun

0.53

0.25 (0.06)

0.37 (0.01)

29.6

513

17

968

56

20.0

 26 Jun

1.47

0.19 (0.01)

0.70 (0.01)

39.1

678

17

461

27

13.5

 03 Jul

0.53

3.61 (0.07)

0.27 (0.03)

40.3

484

12

913

76

17.6

 10 Jul

0.83

0.44 (0.02)

0.38 (0.05)

31.4

841

27

1,013

38

15.0

Standard deviations are in parentheses. DOC concentrations from 2003 are also reported in Minor et al. (2006). BDL indicates concentrations below the limit of analytical detection (<0.03 μmol L−1). Empty fields indicate that there were no data

DIP concentrations were always detectable and ranged from 0.03 to 4.28 μmol L−1 over the study period. As during 2002 (Mulholland et al. 2009), DIN/DIP ratios were <16, the Redfield ratio, at both sites (Tables 1 and 2), suggesting N limitation, with only three exceptions at PL (Table 1); low concentrations of DIP (0.03–0.10 μmol L−1) were observed at these times. The lowest DIN/DIP ratios (0.2–1.0) were observed at GB during the 2006 bloom and were due to high DIP concentrations. Low DIN/DIP ratios were also observed in 2007 at GB at the end of the bloom when DIP concentrations were again high (Table 2).

In 2002, bulk DON concentrations in Chincoteague Bay ranged from about 5.5 to 49.9 μmol N L−1 at PL; concentrations were lower in the spring and increased in the fall. DON concentrations were up to 2.5 times higher at PL than at GB where there was no bloom (Mulholland et al. 2009). During subsequent years (2003, 2006, and 2007), bulk DON concentrations were more similar among sites, ranging from 23.3 to 65.6 μmol N L−1 (Tables 3 and 4). In 2003, DON concentrations at PL ranged from 24.1 to 47.7 μmol N L−1 (Table 3). GB had a larger range in DON concentrations (26.8–65.8 μmol N L−1) than PL. In 2006, DON concentrations at PL were similar to 2003 and ranged from 29.6 to 45.4 μmol N L−1 (Table 3). In contrast, at GB, DON concentrations were lower in 2006, ranging from 26.5 to 41.3 μmol N L−1 (Table 4). In 2007, DON concentrations ranged from 27.8 to 43.7 μmol N L−1 at PL (Table 3) and 23.3 to 40.3 μmol N L−1 at GB (Table 4).

DOC concentrations were higher at PL than at GB during 2002, and concentrations were always <400 μmol C L−1 (Simjouw et al. 2004, Mulholland et al. 2009). In the 2002 study, differences in the DOC concentrations and characteristics between GB and PL were attributed to the brown tide bloom at PL (Simjouw et al. 2004). However, during 2003, both GB and PL experienced brown tide blooms and DOC concentrations were similar at both sites (within 25 % of each other; Tables 3 and 4). During 2006, both sites experienced blooms, but DOC concentrations were always greater at PL (mean, 661 μmol C L−1) than at GB (mean, 393 μmol C L−1) (Tables 3 and 4). During 2007, DOC concentrations were highest and showed more variation over the sampling period than in any other year, ranging from 254 to 841 μmol C L−1 at GB and 378–1,638 μmol C L−1 at PL. Overall, an increase in DOC concentrations in Chincoteague Bay was observed over the 6-year sampling period increasing from a mean of 296 μmol C L−1 and 461 μmol C L−1 at GB and PL, respectively, during 2002 (Simjouw et al. 2004), to a mean of 480 and 627 μmol C L−1 at GB and PL, respectively, during 2007. Throughout the study period, DOC/DON ratios were >6.6, the Redfield ratio, with the exception of the post-bloom period (July–August) at GB during 2003.

In 2002, DOP concentrations were similar among sites (Mulholland et al. 2009). In 2003, DOP concentrations ranged from below the detection limit (BDL) to 0.74 μmol L−1 with a lower range at PL. During 2006, DOP concentrations were below detection at GB on 8 of 10 sampling days with a maximum concentration of 0.29 μmol L−1. DOP concentrations at PL ranged from BDL to 0.76 μmol L−1. During 2007, DOP concentrations were always measurable and higher than during previous years, ranging from 0.25 to 1.47 μmol L−1 at GB and 0.29 to 0.51 μmol L−1at PL.

The TDN/TDP ratios ranged from 16.1 to 23.7 between May and July at GB, where there was no bloom, and from 16.3 to 48.5 at PL during the same period during 2002 (Mulholland et al. 2009). In contrast, TDN/TDP ratios ranged from 37.5 to 69.7 at GB and 59.8 to 152.0 at PL between May and July 2003. High TDN/TDP ratios were also observed during 2006 and 2007 at PL (range of 38.8–86.0, excluding two post bloom dates in 2006), but not at GB (range of 10.5–46.3).

Nitrogen and Carbon Uptake

During most of 2002, total N uptake was much higher (almost an order of magnitude) at the PL site than at GB, consistent with the higher biomass during and after the bloom at that site (Mulholland et al. 2009). Urea was the dominant form of N taken up during most of the year at both bloom and nonbloom sites. While photosynthetic uptake of HCO3 provided the bulk of the measured C uptake during the 2002 bloom, urea, DFAA and glucose contributed to the microbial C demand, particularly at the end of and subsequent to the bloom (Mulholland et al. 2009).

In contrast to 2002, N uptake at PL was dominated byNH4+ in 2006 (Fig. 3 and Table 5) and NO3 in 2007 (Fig. 4 and Table 5). In 2006, NH4+ uptake averaged 52 % (±15 %) of the total N uptake at PL and urea, NO3, and DFAA accounted for 20 % (±12 %), 18 % (±11 %), and 10 % (±9 %) of the N uptake, respectively, and DIN uptake was higher than DON uptake (Fig. 3). At GB, DON compounds (urea plus DFAA) accounted for 56 % (±15 %) of the total nitrogen uptake. Urea, NH4+, DFAA, and NO3 contributed 29 % (±18 %), 30 % (±9), 27 % (±21 %), and 15 % (±9 %), respectively, of the total N uptake, respectively.
https://static-content.springer.com/image/art%3A10.1007%2Fs12237-013-9683-3/MediaObjects/12237_2013_9683_Fig3_HTML.gif
Fig. 3

Carbon uptake at a Public Landing, MD, USA and b Greenbackville, VA, USA and nitrogen uptake at c Public Landing, MD, USA and d Greenbackville, VA, USA during 2006

Table 5

Carbon uptake rates during the 2006 and 2007 blooms: standard deviations are in parentheses

Date

Site

Bicarbonate (μmol C L−1 h−1)

Urea (μmol C L−1 h−1)

Glucose (μmol C L−1 h−1 L)

DFAA (μmol C L−1 h−1 L)

Total (μmol C L−1 h−1 L)

2006

 10 May

GB

2.27 (0.11)

0.07 (0.01)

0.25 (0.00)

0.14 (0.01)

2.83

 18 May

GB

3.23 (0.22)

0.92 (0.01)

1.65 (0.07)

0.57 (0.07)

6.37

 23 May

GB

4.05 (0.04)

0.01 (0.00)

0.06 (0.01)

0.00 (0.00)

4.12

 31 May

GB

13.17 (0.34)

0.00 (0.00)

0.10 (0.03)

0.62 (0.08)

13.90

 07 Jun

GB

7.05 (0.16)

0.05 (0.00)

0.00 (0.00)

0.31 (0.01)

7.36

 14 Jun

GB

33.83 (0.54)

0.10 (0.00)

0.00 (0.00)

5.00 (0.46)

38.82

 21 Jun

GB

27.02 (0.20)

0.10 (0.00)

0.00 (0.00)

1.97 (0.13)

28.99

2006

 10 May

PL

3.62 (0.10)

0.00 (0.00)

0.35 (0.03)

0.11 (0.01)

4.09

 18 May

PL

6.59 (0.26)

0.00 (0.00)

0.00 (0.00)

0.03 (0.01)

6.62

 23 May

PL

10.48 (0.32)

0.00 (0.00)

0.00 (0.00)

0.00 (0.00)

10.48

 31 May

PL

10.35 (0.30)

0.00 (0.00)

0.29 (0.46)

0.00 (0.00)

10.64

 07 Jun

PL

17.90 (0.00)

0.06 (0.01)

0.73 (0.08)

1.35 (0.16)

20.15

 14 Jun

PL

7.16 (0.38)

0.12 (0.01)

2.53 (0.17)

1.03 (0.04)

10.84

 21 Jun

PL

23.08 (0.26)

0.12 (0.06)

2.01 (0.16)

1.58 (0.13)

26.80

2007

 15 May

GB

7.57 (0.44)

0.00 (0.00)

2.39 (1.13)

2.20 (0.08)

12.17

 19 May

GB

8.81 (3.02)

0.03 (0.00)

0.14 (0.02)

0.52 (0.03)

9.51

 29 May

GB

14.57 (0.08)

0.08 (0.01)

2.87 (0.16)

3.38 (0.08)

20.89

 05 Jun

GB

10.15 (2.35)

0.03 (0.00)

0.84 (0.06)

0.08 (0.01)

11.10

 12 Jun

GB

48.33 (0.75)

0.11 (0.03)

7.22 (0.06)

2.38 (0.09)

58.04

 19 Jun

GB

23.60 (0.28)

0.02 (0.00)

3.57 (0.24)

0.72 (1.08)

27.92

 26 Jun

GB

30.04 (1.09)

0.07 (0.01)

4.57 (0.12)

3.09 (1.23)

37.77

 03 Jul

GB

10.04 (0.33)

0.19 (0.00)

3.36 (0.09)

1.27 (0.04)

14.86

 10 Jul

GB

13.37 (2.30)

0.06 (0.00)

2.84 (0.08)

0.80 (0.06)

17.07

2007

 15 May

PL

5.25 (0.19)

0.00 (0.00)

0.02 (0.06)

0.68 (0.02)

5.94

 19 May

PL

10.31 (0.34)

0.02 (0.00)

0.35 (0.25)

1.33 (1.10)

12.00

 29 May

PL

12.51 (0.44)

0.04 (0.00)

1.44 (0.40)

1.14 (0.09)

15.13

 05 Jun

PL

14.28 (0.35)

0.01 (0.00)

1.44 (0.10)

0.55 (0.02)

16.28

 12 Jun

PL

14.45 (0.50)

0.03 (0.01)

2.19 (0.06)

1.56 (0.14)

18.24

 19 Jun

PL

9.05 (0.26)

0.02 (0.00)

1.19 (0.15)

1.06 (0.06)

11.31

 26 Jun

PL

14.15 (2.13)

0.04 (0.00)

1.51 (0.17)

0.73 (0.31)

16.43

 03 Jul

PL

8.75 (0.63)

0.07 (0.00)

2.51 (0.25)

1.49 (0.27)

12.82

 10 Jul

PL

11.94 (1.21)

0.05 (0.00)

6.51 (0.65)

0.00 (0.00)

18.50

https://static-content.springer.com/image/art%3A10.1007%2Fs12237-013-9683-3/MediaObjects/12237_2013_9683_Fig4_HTML.gif
Fig. 4

Carbon uptake at a Public Landing, MD, USA and b Greenbackville, VA, USA and nitrogen uptake at c Public Landing, MD, USA and d Greenbackville, VA, USA during 2007

During 2007, NO3 was the dominant form of N taken up during the bloom representing 50 % (±29 %) of the total N uptake at PL (Fig. 4); NO3/NO2 concentrations in 2007 were also the highest observed over the 6-year study period (Table 1). Higher NO3/NO2 concentrations were also observed at GB during 2007 (Table 2), but NO3 uptake only comprised 30 % (±13 %) of the total N uptake at that site (Fig. 4). At GB, urea uptake was 52 % (±15 %) of the total measured N uptake and DON uptake was >50 % of the total N uptake on nine of the sampling days.

As in 2002, photosynthetic bicarbonate uptake was the main form of carbon taken at both sites during 2006 (Fig. 3 and Table 6) and 2007 (Fig. 4 and Table 6). On average, photosynthetic uptake of HCO3 accounted for 89 % (±14 %) and 86 % (±16 %) of the total measured C uptake at PL and GB, respectively, during 2006 (Fig. 3). Similar to 2002, urea C was a small fraction of the total measured carbon at both sites during 2006 and 2007, averaging <2 % of the total C uptake.
Table 6

Nitrogen uptake rates during the 2006 and 2007 blooms: standard deviations are in parentheses

Date

Site

NO3 (μmol N L−1 h−1)

Urea (μmol N L−1 h−1)

NH4+ (μmol N L−1 h−1)

DFAA (μmol N L−1 h−1)

Total (μmol N L−1 h−1)

2006

 10 May

GB

0.01 (0.00)

0.17 (0.01)

0.09 (0.00)

0.06 (0.00)

0.32

 18 May

GB

0.00 (0.00)

0.02 (0.00)

0.02 (0.01)

0.08 (0.00)

0.13

 23 May

GB

0.03 (0.00)

0.12 (0.00)

0.13 (0.01)

0.00 (0.00)

0.27

 31 May

GB

0.52 (0.01)

1.47 (0.00)

0.83 (0.06)

0.48 (0.06)

3.30

 07 Jun

GB

0.24 (0.01)

0.31 (0.00)

0.43 (0.01)

0.14 (0.00)

1.12

 14 Jun

GB

1.43 (0.10)

0.36 (0.00)

1.56 (0.03)

2.48 (0.18)

5.83

 21 Jun

GB

0.45 (0.01)

0.22 (0.11)

0.63 (0.01)

0.77 (0.07)

2.07

2006

 10 May

PL

0.10 (0.02)

0.13 (0.00)

0.73 (0.01)

0.04 (0.00)

1.00

 18 May

PL

0.21 (0.01)

0.16 (0.00)

0.78 (0.01)

0.03 (0.00)

1.18

 23 May

PL

0.31 (0.04)

1.76 (0.00)

1.79 (0.07)

0.09 (0.01)

3.94

 31May

PL

0.37 (0.02)

0.35 (0.02)

0.69 (0.03)

0.04 (0.01)

1.44

 07 Jun

PL

0.31 (0.02)

0.55 (0.00)

1.58 (0.03)

0.53 (0.07)

2.97

 14 Jun

PL

0.44 (0.07)

0.20 (0.00)

0.37 (0.01)

0.27 (0.01)

1.27

 21 Jun

PL

0.29 (0.29)

0.27 (0.13)

0.91 (0.02)

0.41 (0.01)

1.89

2007

 15 May

GB

0.51 (0.55)

0.31 (0.02)

0.08 (0.00)

0.45 (0.02)

1.36

 19 May

GB

0.02 (0.00)

0.38 (0.02)

0.07 (0.00)

0.14 (0.01)

0.62

 29 May

GB

1.79 (0.06)

2.45 (0.01)

0.09 (0.02)

1.09 (0.10)

5.42

 05 Jun

GB

0.13 (0.00)

0.36 (0.01)

0.08 (0.00)

0.02 (0.00)

0.60

 12 Jun

GB

2.06 (0.10)

2.24 (0.19)

0.14 (0.00)

0.36 (0.01)

4.80

 19 Jun

GB

1.84 (0.06)

2.33 (0.06)

0.10 (0.00)

0.22 (0.34)

4.49

 26 Jun

GB

1.59 (0.04)

2.06 (0.06)

0.11 (0.02)

0.66 (0.50)

4.42

 03 Jul

GB

1.41 (0.05)

6.91 (0.13)

0.10 (0.00)

0.37 (0.01)

8.79

 10 Jul

GB

0.76 (0.04)

1.08 (0.05)

0.09 (0.01)

0.22 (0.13)

2.15

2007

 15 May

PL

0.00 (0.00)

0.42 (0.00)

0.13 (0.00)

0.31 (0.06)

0.86

 19 May

PL

1.67 (0.18)

1.53 (0.03)

0.12 (0.00)

0.46 (0.28)

3.78

 29 May

PL

6.42 (0.20)

1.79 (0.01)

0.13 (0.00)

0.48 (0.02)

8.82

 05 Jun

PL

6.33 (0.13)

1.29 (0.02)

0.13 (0.04)

0.30 (0.02)

8.06

 12 Jun

PL

5.18 (0.21)

1.98 (0.00)

0.13 (0.01)

0.82 (0.08)

8.10

 19 Jun

PL

4.42 (0.26)

1.52 (0.19)

0.07 (0.02)

0.48 (0.02)

6.50

 26 Jun

PL

0.13 (0.02)

1.83 (0.06)

0.13 (0.00)

0.31 (0.03)

2.41

 03 Jul

PL

5.36 (0.07)

1.58 (0.04)

0.15 (0.00)

0.68 (0.29)

7.76

 10 Jul

PL

3.87 (1.23)

1.11 (0.01)

0.10 (0.00)

0.00 (0.00)

5.08

Carbon uptake from DFAA and glucose accounted for a substantial fraction of the total measured C uptake during 2006 and 2007 (Figs. 3 and 4). In 2006, glucose uptake accounted for 9 % (±2 %) of the C uptake on 10 May and 26 % (±4 %) on 18 May at the beginning of the A. anophagefferens bloom (Fig. 3). At PL and GB, glucose uptake averaged 15 % (±10 %) and 12 % (±10 %) of the total measured C, respectively, during 2007. The highest rates of glucose uptake were measured during the peak and at the end of the bloom (Fig. 4).

Discussion

High A. anophagefferens concentrations have been shown to result in negative impacts to shellfish, seagrasses, and planktonic organisms (Bricelj and Lonsdale 1997). Several shellfish aquaculture facilities are located in or on Chincoteague Bay and could be negatively affected by blooms. In addition to aquaculture facilities, ∼250 acres of bay bottom have been leased for potential use in raising hard clams (Tarnowski 1997). Growth rates of the hard clam Mercenaria mercenaria can be negatively impacted at A. anophagefferens concentrations as low as 20,000 cells mL−1 (Wazniak and Glibert 2004). Both the 2010 (http://ian.umces.edu/pdfs/ian_report_card_318.pdf) and 2011 (http://ian.umces.edu/pdfs/ian_report_card_385.pdf) Coastal Bays report cards give low scores for hard clams in Chincoteague Bay, and this has been related to the recurrent brown tide blooms.

Bloom Dynamics

Between 2002 and 2007, Chincoteague Bay experienced brown tide blooms (>35,000 cells mL−1; Gastrich and Wazniak 2002) every spring except during 2005. In each of the bloom years, there was a gradual warming trend during which the A. anophagefferens bloom initiated; during 2005, the water warmed abruptly in the spring, likely preventing bloom development (http://www.dnr.state.md.us/coastalbays/bt_results.html). While only one of our study sites had documented a brown tide bloom prior to 2002, during 2003, blooms occurred at both study sites and blooms appeared to have spread to previously unimpacted areas in Chincoteague Bay. However, the lack of monitoring at the VA study site may have precluded detection of blooms prior to the onset of this study.

The duration and intensity of brown tide blooms in Chincoteague Bay also appear to have increased over our study period (Fig. 2). Although the 2003 bloom at PL was not as intense (lower A. anophagefferens cell densities) as the 2002 bloom, the 2003 bloom lasted longer and spread to our previously unimpacted site at GB in Chincoteague Bay. During 2006 and 2007, blooms at both sites reached higher densities and lasted longer than in previous years (Fig. 2). For the entire 2007 sampling period (15 May–10 July), A. anophagefferens concentrations were above the category 3 threshold for brown tide blooms (>200,000 cells mL−1). This differs from the 2002 bloom at PL where A. anophagefferens concentrations were above the category 3 threshold only from 5/30 to 6/12 (Mulholland et al. 2009).

Nitrogen Dynamics

Previous studies have shown that A. anophagefferens blooms only after NO3 concentrations have been depleted (Gobler and Sañudo-Wilhelmy 2001a) or when DIN concentrations are low (Lomas et al. 2004). In Chincoteague Bay, these criteria were met in 2002, when NO3/NO2 concentrations were below the limit of analytical detection from April through the end of June at PL, and DIN concentrations were near or at the limit of analytical detection during the bloom period (Mulholland et al. 2009). However, NO3/NO2 and DIN concentrations were low or at the analytical detection limit at both the bloom and nonbloom sites in Chincoteague Bay, suggesting that low DIN was not sufficient for bloom initiation since A. anophagefferens cells were present at both sites prior to the bloom (Mulholland et al. 2009). During our subsequent multiyear study, neither NO3/NO2 nor DIN was depleted during brown tide blooms at GB and PL in 2003, 2006, and 2007. DIN concentrations did not vary greatly between the three bloom years, ranging from 0.5 to 4.5 μmol L−1 and averaging 1.4 μmol L−1 (Tables 1 and 2). These concentrations were within the ranges previously observed in Chincoteague Bay during nonbloom years (Glibert et al. 2007).

As for dissolved N concentrations, during this 6-year study, N uptake during blooms varied greatly and NH4+, urea, NO3, and DFAA all contributed to the total measured N uptake (Figs. 3 and 4). Previously, it was shown that A. anophagefferens has a high affinity for NH4+ and urea (Lomas et al. 1996, Berg et al. 1997). Consistent with those observations, these two compounds accounted for the majority of the N uptake in Chincoteague Bay during 2002 (Mulholland et al. 2009) and 2006 (Figs. 3, 4). Similarly, in Long Island, NY, USA, coastal bays, NH4+ and DFAA were the primary N compounds taken up during brown tide blooms there (Mulholland et al. 2002). In contrast, high NO3/NO2 concentrations in Chincoteague Bay during 2007 were accompanied by high uptake rates of this compound during that year (Fig. 4 and Table 5). This was unexpected since studies have shown that the growth of A. anophagefferens relative to other competing phytoplankton can be suppressed with the addition of NO3 (Gobler and Sañudo-Wilhelmy 2001a; Taylor et al. 2006) and that blooms of this organism typically occur in years with low NO3 concentrations (LaRoche et al. 1997). Interestingly, cultures of A. anophagefferens grow equally well on NO3 and urea (Pustizzi et al. 2004; MacIntyre et al. 2004), and some blooms have been observed during years with high nitrate inputs to other coastal embayments (Gobler and Sañudo-Wilhelmy 2001b). Together, these results suggest that blooms of A. anophagefferens can be supported by a variety of organic and inorganic N compounds.

Carbon Dynamics

A major goal of this study was to determine the degree to which brown tide organisms augment autotrophic uptake of DIC with heterotrophic uptake of DOC and to determine how this changes over the course of blooms as cell densities increase, potentially self-shade, and populations draw down DIC. Overall, DIC uptake was the dominant form of C taken up during 2006 and 2007 (Figs. 3 and 4); however, C from glucose, DFAA, and urea was also taken up during blooms and DOC uptake accounted for as much as 49 % of the total measured C uptake. On average, measured DOC uptake accounted for 16 % of the total C uptake increasing at the end of the blooms during 2006, but similar during all stages of the blooms in 2007. These results are similar to what was observed during a 2002 bloom at PL where organic C uptake in whole water accounted for 17–71 % of the total carbon uptake, with the percentage generally increasing as the bloom progressed (Mulholland et al. 2009). C uptake from DFAA was also similar to rates observed in a 2000 bloom in Quantuck Bay, NY, USA (Mulholland et al. 2002).

There are several factors that could have biased our DIC and DOC uptake results. For DIC uptake, we estimated DIC concentrations from salinity assuming that DIC was saturated. However, in 2006 and 2007, pH was as high as 8.9 by the end of the blooms (Tables 1 and 2), presumably due to the photosynthetic drawdown of inorganic C. If DIC concentrations were less than saturating, than our additions were likely >10 % of the ambient pool, and this would result in lower calculated DIC uptake rates. To calculate DIC and DOC uptake, we multiplied the specific uptake rate (h−1) by the particulate C concentration. If significant amounts of particulate inorganic C or particulate detrital C contributed to the particulate C concentration, this would cause us to overestimate the absolute C uptake in our samples. Another complicating factor is that most DOC is uncharacterized (Benner 2002), and isotopic tracers are not available for bulk DOC; therefore, we measured the uptake of only a small subset of labile compounds present in the natural DOC pool. Consequently, we may have underestimated DOC uptake in the environment. Finally, all of our incubations in this study were done during the middle of the day, when photosynthetic uptake rates were likely at or near maximum levels. If A. anophagefferens are also able to take up organic C at night or when light is limiting, this would result in higher uptake rates during other portions of the day. Theoretically, taking up DOC at night or when light or DIC is limiting could give populations a large advantage over competing phytoplankton that can only use the inorganic C pool during the day in conjunction with photosynthesis.

Declines in photosynthesis and growth rates observed in coastal and oceanic marine diatoms and a natural assemblage of phytoplankton from Naragansett Bay when pH was >8.8 (Chen and Durbin 1994) and pH values of 8.9 and higher have been shown to affect the growth rate of some heterotrophic protists (Pedersen and Hansen 2003) and dinoflagellates (Hansen et al. 2007). While there was some uptake of organic C during the 2006 bloom at PL and GB, uptake was much higher during 2007, when pH was consistently higher at both sites and DIC may have become limiting. This suggests that A. anophagefferens might be able to augment DIC uptake with uptake of organic carbon when DIC is limiting. DOC uptake also increased as blooms progressed during both 2006 and 2007 (Figs. 3, 4), consistent with the idea that DOC can supplement photosynthetic C uptake when cell densities are high and photosynthesis is light or limited. Previous studies found that glucose additions stimulated brown tide growth relative to other algae and that the DOC pool was drawn down during brown tide blooms (Gobler et al. 2004, Minor et al. 2006).

Despite their ability to take up DOC compounds, A. anophagefferens appears to be a net source of DOC to Chincoteague Bay (Simjouw et al. 2004; Mulholland et al. 2009). We observed a 50 % increase in DOC concentrations over the 6-year study period. Accumulation of material in this system may be due to the long residence time of this coastal lagoon (63 days; Pritchard 1960); however, the DOC accumulated during and after the nearly annual brown tide blooms appears to have carried over into subsequent years. As for DOC, maximum bacterial abundances increased over time in Chincoteague Bay and this may be related to the high DOC concentrations supporting their growth. If blooms continue to occur in this lagoon and organic carbon concentrations continue to accumulate, this may drive Chincoteague Bay towards a net heterotrophic system. Not only might this favor the growth of heterotrophic bacteria and phytoplankton mixotrophs such as A. anophagefferens, but this system change may also favor other mixotrophic harmful algal species (HABs) that tend to flourish in eutrophic estuaries (Burkholder et al. 2008). Elevated bacterial production might also provide increased prey for bactivorous protists and may negatively impact the system by depleting dissolved oxygen.

A. anophagefferens blooms may also have a positive feedback on this HAB species' growth (Sunda et al. 2006). Because grazing is reduced during A. anophagefferens blooms (Gobler et al. 2002), there are fewer recycled nutrients available (Sunda et al. 2006) to support the growth of other species. As A. anophagefferens blooms intensify, light penetration is reduced, which limits the growth of benthic algae (MacIntyre et al. 2004). Without benthic algae intercepting nutrients coming from the sediments, there may be a greater flux of nutrients coming out of the sediments and into the water column (MacIntyre et al. 2004; Sunda et al. 2006), and this may further stimulate the growth of A. anophagefferens. Wazniak (2004) found that Chincoteague Bay has a considerable benthic microalgae population, with summertime benthic chlorophyll concentrations averaging 38.69 mg m−2 in 2002 and 28.6 mg m−2 in 2003. If these blooms block light reaching the sediments on a regular basis, the benthic algal community could also be impacted by dense blooms. These positive feedbacks could result in increases in brown tide bloom intensity in Chincoteague Bay in the future.

Nutrient Ratios

During this study, DIN/DIP ratios were consistently below 16. Although the DIN/DIP ratio ranged from 0.2 to 39.5, the average ratio was 5.0 (±7.5). The low DIN/DIP ratios suggest that the system was depleted in N relative to P, as has been shown previously for coastal and estuarine systems (Fisher et al. 1992). This was also observed during 2002 (Mulholland et al. 2009) at both the bloom and nonbloom sites. However, TDN/TDP ratios were usually in excess of the Redfield ratio, suggesting P limitation if DON is present in bioavailable forms (Fig. 5). DOP concentrations were generally low, resulting in high DOC/DOP and DON/DOP ratios. DON concentrations were similar to what has been observed in other brown tide prone estuaries, including those on Long Island (Lomas et al. 2001, 2004).It has been suggested that brown tide blooms are associated with high DOC/DON and low DON/DOP (Lomas et al. 2001). However, this was not observed in Chincoteague Bay in 2002 (Mulholland et al. 2009) or in the subsequent bloom years reported here (Fig. 6). Thus, while it has been suggested that organic nutrient ratios and DIN depletion are causative agents promoting brown tide bloom formation, results reported here suggest that relating blooms to nutrient concentrations and ratios may be more complicated than previously thought.
https://static-content.springer.com/image/art%3A10.1007%2Fs12237-013-9683-3/MediaObjects/12237_2013_9683_Fig5_HTML.gif
Fig. 5

Total dissolved nitrogen (TDN) versus total dissolved (TDP) phosphorus for all the blooms (2002, 2003, 2006, and 2007) sampled as part of this project (including data reported in Simjouw et al. 2004; Minor et al. 2006; and Mulholland et al. 2009). The black line is the 16:1 line (Redfield)

https://static-content.springer.com/image/art%3A10.1007%2Fs12237-013-9683-3/MediaObjects/12237_2013_9683_Fig6_HTML.gif
Fig. 6

Comparison of DOC/DON and DON/DOP ratios during brown tide blooms in 2003, 2006, and 2007 at both locations, GB and PL. The 2002 PL and GB data are added from Mulholland et al. 2009)

The total measured C/N uptake ratio estimated from the short-term incubation experiments averaged 7.6, but the range was quite high (1–44), suggesting that there may be short-term uncoupling between C and N uptake during blooms. The short-term C/N uptake ratios at PL were below Redfield for most of the study, suggesting short-term imbalances in C and N uptake or unquantified C sources supporting the growth of bloom organisms. We estimated uptake of DOC using glucose, urea, and amino acids during mid-day incubations. A combination of these compounds represents a very small fraction of the DOC pool (Benner 2002). Uptake of DOC during dark periods or uptake of compounds not measured here may also have contributed to A. anophagefferens growth and may alleviate this imbalance.

Urea was taken up primarily as a nitrogen source during this study, and cells did not incorporate much of the carbon associated with the urea. This had been observed previously during brown tide blooms in Quantuck Bay, NY, USA (Lomas 2003). Urea carbon was taken up at a higher rate than urea nitrogen at only one time point during the 2006 GB bloom (Fig. 3). Excluding this one exception, the C/N uptake ratio for urea averaged just 0.06. In contrast, the C/N ratio for DFAA averaged 2.6. This is similar to what was observed during a 2000 bloom in Quantuck Bay, NY, USA and the 2002 bloom at PL when C/N uptake for DFAA was about 2 (Mulholland et al. 2002; Mulholland et al. 2009). It is unclear why urea C was not assimilated. Urea degradation can produce ammonium and CO2 in the environment (Kamennaya et al. 2008) and urease catalyzes the degradation of urea to two ammonium ions and CO2 within cells. When DIC is not limiting, cells may simply release CO2 from urea back into the environment. Amino acids, on the other hand, may be assimilated directly into cellular proteins or other materials thereby conserving the C in intermediate cell metabolites.

Conclusions

During this 6-year study of brown tide blooms in Chincoteague Bay, MD and VA, we found an increase in bloom intensity and duration over time and an overall accumulation of DOC in this lagoonal system. This has important implications for this systems' productivity and may lead to changes in ecosystem structure and metabolism, trophic status, and food web interactions. A. anophagefferens is nutritionally versatile and are able to use a wide range of nitrogen and carbon sources to meet their nutritional demands. Consequently, any strategy for managing nutrient loads to prevent blooms should take into account the ability of both inorganic and organic C and N to be used by this organism. Because no single N compound is responsible for fueling brown tide growth, the total N load and retention of that load within the system may be key factors contributing to brown tides rather than inputs of any particular form of N. During blooms, organic C uptake subsidized C acquisition from photosynthesis. Although bicarbonate uptake was higher than organic carbon uptake, sampling and rate measurements were made mid-day when PAR was at its peak. Further investigations are needed to determine the contribution of DOC to A. anophagefferens bloom maintenance and proliferation, particularly when light or C may be limiting.

Acknowledgments

We wish to thank P. W. Bernhardt, J.-P. Simjouw, A. M. Watson, A. Rocha, E. Cornfeld, and S. Reynolds for help collecting and analyzing samples and the staff at the Wallops Island Marine Science Consortium Laboratory and Virginia Institute of Marine Sciences Eastern Shore Laboratory for providing us with boats and laboratory space during this study. M. Luckenbach, P. G. Ross, and G. Arnold provided assistance during field sampling and important information regarding the coastal bays of Virginia. C. Wazniak (MD Department of Natural Resources) and B. Sturges (MD National Park Service) also contributed time and intellectual support for the Chincoteague Bay studies. We also thank two anonymous reviewers and the editor for comments on an earlier version of this manuscript. This study was funded by a grant from the US ECOHAB Program to M. R. Mulholland. The ECOHAB Program is sponsored by the National Oceanographic and Atmospheric Administration, Environmental Protection Agency, National Science Foundation, National Aeronautics and Space Administration, and Office of Naval Research. This is contribution number 764 from the US ECOHAB Program.

Copyright information

© Coastal and Estuarine Research Federation 2013