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Macroalgal blooms on southeast Florida coral reefs
II. Cross-shelf discrimination of nitrogen sources indicates
widespread assimilation of sewage nitrogen
Brian E. Lapointe
a,
*, Peter J. Barile
a
, Mark M. Littler
b
, Diane S. Littler
a,b
a
Harbor Branch Oceanographic Institution Inc., Division of Marine Science, 5600 US 1 North, Ft. Pierce, FL 34946, USA
b
Department of Botany, National Museum of Natural History, Smithsonian Institution, Washington, DC 20560, USA
Received 30 November 2004; received in revised form 1 June 2005; accepted 6 June 2005
Abstract
Since 1990, coral reefs off southeast Florida have experienced an unprecedented succession of macroalgal blooms and
invasions. To determine if anthropogenic land-based nitrogen (N) sources support these HABs, we collected macroalgal
tissue for stable nitrogen isotope (d
15
N) analysis at three spatially distinct depths ranging from the shallow subtidal to the
shelf break (43 m) along seven transects from Jupiter to Deerfield Beach, Florida, USA. This sampling was initiated during
a historically significant drought in the spring of 2001 when rainfall, stormwater runoff, and upwelling were at a minimum.
The sampling was repeated in the summer wet season following significant stormwater runoff and during a strong upwelling
event.
Despite significant seasonal changes in N source availability, d
15
N values did not vary between samplings. Collectively, d
15
N
values were significantly higher on inshore shallow reefs (+8.1%) compared to mid (+6.1%) and deep reefs (+6.7%). Values
were also elevated in the southern portion of the study area (e.g., Boca Raton, +8.5%) where nearly 1.5 billion l/day of
secondarily treated wastewater is discharged into the ocean via coastal outfalls. Codium isthmocladum, a macroalga that
assimilates its nutrients entirely from the water column, was the dominant bloom species in the southern study area, where tissue
d
15
N values matched source values of nearby sewage outfalls. The northern study area was dominated by species of the genus
Caulerpa, particularly the invasive Pacific native C. brachypus var. parvifolia, which are capable of accessing benthic nutrient
sources (e.g., submarine groundwater discharge, SGD) by means of root-like rhizoids. The northern area does not have sewage
outfalls but features a highly transmissive geologic zone where SGD may be enriched with septic tank leachate and effluent from
municipal deep injection wells.
Multiple lines of evidence supported the hypothesis that land-based sewage N was more important than upwelling as a N
source to these HABs: (1) d
15
N values were highest on shallow reefs and decreased with increasing depth, indicating land-based
sources of enrichment; (2) elevated d
15
N values occurred in these HABs during the dry season, prior to the onset of the summer
upwelling; (3) elevated NH
4+
concentrations occur on these reefs during both upwelling and non-upwelling periods and are
kinetically preferred by macroalgae compared to upwelled NO
3
. These findings provide a case study of a coupling between
www.elsevier.com/locate/hal
Harmful Algae 4 (2005) 1106–1122
DOI of original article: 10.1016/j.hal.2005.06.004
* Corresponding author. Tel.: +1 772 465 2400x276; fax: +1 772 465 0134.
E-mail address: lapointe@hboi.edu (B.E. Lapointe).
1568-9883/$ – see front matter #2005 Published by Elsevier B.V.
doi:10.1016/j.hal.2005.06.002
increasing anthropogenic activities and the development of macroalgal HABs, including invasive species that threaten
economically important reef resources in southeast Florida.
#2005 Published by Elsevier B.V.
Keywords: Nitrogen; Macroalgae; Stable isotopes; Coral reefs; Eutrophication
1. Introduction
The frequency, extent, and biomass of macroalgal
blooms have increased in many tropical/subtropical
coral reef communities in recent decades as a result of
increasing land-based nutrient pollution (UNEP, 1994;
ECOHAB, 1997; NRC, 2000). On coral reefs off
southeast Florida, a succession of harmful macroalgal
blooms (HABs) began in 1990 with extensive
unattached forms of Codium isthmocladum that
developed accumulations up to 2 m thick over the
reef surface and on adjacent beaches (Fig. 1A and B).
These HABs resulted in die-offs of sponges, hard
corals, and soft corals due to hypoxia/anoxia in near-
bottom waters and caused an emigration of reef fishes
from the impacted areas (Lapointe, 1997; Lapointe
and Hanisak, 1997). These initial C. isthmocladum
blooms were followed by an extensive bloom of
Caulerpa verticillata in 1997 (Lapointe, 1999;
Fig. 1C). Previously, C. verticillata had not been
observed on these reefs (Hanisak and Blair, 1988), and
this invasive alga quickly spread from reefs off Riviera
Beach near the Lake Worth Inlet northwards to reefs
off Jupiter in northern Palm Beach County (Fig. 2).
Species of the tropical green algal genus Caulerpa
are well known invaders of coastal waters, as
demonstrated by the proliferation of Caulerpa taxi-
folia in the Mediterranean Sea (Meinesz, 1999;
Verlaque et al., 2003). Evidence suggests that the
rapid invasion of C. taxifolia in the Mediterranean
(Meinesz and Hesse, 1991) was supported by land-
based sources of nutrient pollution (Chisholm et al.,
1997; Jaubert et al., 2003). Ironically, the Mediterra-
nean native, C. ollivieri, has invaded a polluted harbor
in the Bahamas where sewage has been implicated as a
primary nitrogen (N) source supporting bloom
formation (Lapointe et al., 2005a). In the Florida
Keys, C. verticillata has likewise become abundant in
sewage-polluted canal systems (Lapointe et al., 1994).
Although the appearance of HABs involving Codium
isthmocladum and C. verticillata has been considered
an indicator of escalating nutrient enrichment and
eutrophication in southeast Florida (Lapointe and
Hanisak, 1997), only limited attempts have been made
to identify the specific N sources supporting these
blooms (Lapointe, 1997). Most recently, while
conducting the initial sampling (May 2001) for this
study, we discovered a bloom of the invasive Pacific
native Caulerpa brachypus var. parvifolia (henceforth
referred to as C. brachypus, Fig. 1D and E) on reefs in
Palm Beach County.
Both natural and anthropogenic sources may
supply nutrients to HABs on reefs in southeast
Florida. Episodic summertime upwelling has histori-
cally been important in this area (Green, 1944; Taylor
and Stewart, 1958) but, because these HABs have
developed only since 1990, we hypothesized that
anthropogenic nutrient enrichment resulting from
rapid population growth and associated land-based
runoff from the watershed is the most significant
nutrient source supporting these HABs. Urbanization
of watersheds along the eastern coast of the U.S. has
significantly increased nutrient loadings to coastal
waters from a variety of sources including fertilizers,
top soils, fossil fuel combustion, and municipal
wastewaters (Howarth et al., 1996, 2000; NRC,
2000). Intensive agricultural activity in the Everglades
Agricultural Area (EAA) in the inner region of the
coastal plain enriches groundwaters and surface
waters with N and phosphorus (P) from top soil and
fertilizers. These are transported to coastal waters via
either submarine groundwater discharges (SGD; Finkl
and Charlier, 2003) or surface water discharges
through the Port Everglades, Hillsboro, Boynton,
Lake Worth, and Jupiter inlets (Fig. 2). Finkl and
Charlier (2003) estimated that groundwaters deliver
5727 metric t/year of N and 414 metric t/year of P via
SGD to the coastal reefs off Palm Beach County alone.
The watershed of southeast Florida between Palm
Beach and Dade counties now supports nearly 7 million
people. The domestic wastewater generated by this
rapidly expanding population is disposed of either via
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1107
septic tanks or by secondary treatment followed by
discharge through ocean outfalls (1.5 billion l/day or
6000 metric t of N/year; Hazen and Sawyer, 1994)or
into Class 1 injection wells (1.9 billion l/day or 7600
metric t of N/year; USEPA, 2003). Nearly 30 injection
wells in Palm Beach and Broward counties pump
nutrient-rich secondarily treated sewage effluent under
pressure into the boulder zone at 1100 m depth
(Miller, 1997; USEPA, 2003). In Palm Beach County, at
least one of these wells has been identified in a recent
USEPA (2003) wastewater risk assessment for south
Florida as having ‘‘a significant potential for vertical
migration into overlying drinking water aquifers.’’
Stable nitrogen isotope ratios (d
15
N) can be used to
effectively discriminate among natural and anthro-
pogenic sources in marine food webs when the
signatures of the various N sources are known (see
reviews by Peterson and Fry, 1987; Owens, 1987;
Lajtha and Michener, 1994). Atmospheric N and
nitrogen fixation have baseline d
15
N values of 0%
(Heaton, 1986; Owens, 1987). Enrichment of d
15
Nin
aquatic systems can result from N transformations that
occur prior to, during, or following the treatment and
discharge of wastewater. Volatilization of ammonia
and isotopic fractionation by microbes during nitri-
fication and denitrification produce residual DIN with
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221108
Fig. 1. Images of various green macroalgae that have formed harmful and invasive blooms on coral reefs off southeast Florida between 1990 and
2001: (A) Codium isthmocladum smothering octocorals at the ‘‘Football Field,’’ Lake Worth, FL., August 1990; (B) massive beach drift of
Codium isthmocladum at Boynton Beach, FL., August 1992; (C) Caulerpa verticillata overgrowing sponge at North Colonel’s Ledge, off Juno
Beach, FL.; (D) Caulerpa brachypus var. parvifolia near the Princess Anne off Riviera Beach, FL.; (E) Codium isthmocladum surrounded by the
invasive Caulerpa brachypus var. parvifolia at North Colonel’s Ledge, Juno Beach, FL; (F) Caulerpa racemosa at North Colonel’s Ledge, Juno
Beach, FL.
elevated d
15
N values of +6%to +22%(Heaton, 1986;
Lindau et al., 1989). Globally, many case studies have
used d
15
N as a tool to discriminate between natural
and anthropogenic N sources supporting macroalgal
growth (Hobbie et al., 1990; Lapointe, 1997; France
et al., 1998; McClelland and Valiela, 1998; Rogers,
1999; Costanzo et al., 2001; Wayland and Hobson,
2001; Umezawa et al., 2002; Lapointe and Thacker,
2002; Gartner et al., 2002; Barile, 2004; Savage and
Elmgren, 2004). Several studies have successfully
utilized d
15
N values in coral reef ecosystems to assess
the extent of land-based N enrichment along gradients
into the coastal ocean, on scales from several
kilometers (Umezawa et al., 2002; Lapointe et al.,
2004) to nearly 40 km across the Great Barrier Reef
lagoon (Sammarco et al., 1999).
We hypothesized that, if sewage rather than
upwelling was the primary DIN source supporting
macroalgal HABs, then the highest d
15
N values
would occur on the shallow reefs most influenced by
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1109
Fig. 2. Map of southeast Florida study area showing grid of sample stations. Sample stations where Caulerpa brachypus was discovered in May
2001 are denoted.
land-based wastewater discharges. Macroalgae that
rely on N-fixation have low d
15
N values near the
atmospheric signature of 0%(France et al., 1998;
Table 1) in contrast to those using sewage N, which
become increasingly enriched in d
15
N with increasing
sewage N contributions over a range from +3%to
+16%(Lapointe, 1997; Costanzo et al., 2001). This
range includes secondary-treated wastewater dis-
charges from sewage outfalls in the study area (Hoch
et al., 1995;Table 1), as well as shallow (<10 m)
groundwaters contaminated by septic tanks in the
Palm Beach County watershed (Lapointe and Krupa,
1995;Table 1). In comparison, both fertilizers and
organic peat from agricultural areas in the western
region of the watershed have d
15
N values ranging from
0%to +3%(Heaton, 1986;Table 1) and can therefore
be effectively discriminated from the wastewater N
signature. The d
15
N value of upwelled NO
3
from
adjacent North Atlantic deep water is +4.8%
(Sigman et al., 2000; Montoya et al., 2002;
Table 1), which coincides with the lower end of the
sewage d
15
N range. Hence, additional evidence of the
temporal and spatial presence of upwelling as a
potential N and P source was needed. Accordingly, we
collected seawater samples from a variety of reef sites
during upwelling periods in this study for determina-
tion of DIN (=NH
4+
+NO
3
+NO
2
) and soluble
reactive phosphorus (SRP), and recorded sample
depths and water temperatures.
Because the spring of 2001 was a period when
climatological conditions were not favorable for
upwelling and when a record drought occurred in
south Florida (Abtew et al., 2002), we hypothesized
that N availability from upwelling and stormwater
runoff was minimal compared to wastewater N
loading in the study area prior to our initial May
sampling. In contrast, the August sampling followed
considerable wet season rainfall (60 cm between May
and August, 2001; South Florida Water Management
District), which coupled with strong upwelling during
the August sampling, hypothetically reflected sig-
nificant N contributions from both of these sources.
2. Materials and methods
2.1. Study sites and rationale for collection of
macroalgae
To provide comprehensive spatial and temporal
discrimination of N sources to macroalgal HAB
communities in Palm Beach and northern Broward
counties (Fig. 1A–F), tissue samples of abundant
macroalgae were collected over a grid of 21 stations
within seven transects extending offshore of Jupiter,
Juno Beach, Riviera Beach, Lake Worth, Boynton
Beach, Boca Raton, and Deerfield Beach in the dry
(10–30 May) and wet (20–28 August) seasons of 2001
(Fig. 2). The sampling design involved depth
stratification across the southeast Florida shelf in
order to assess spatial variability among shallow
subtidal (<5 m), mid-depth (25–30 m) and deep
(40–43 m) reefs. This sampling network allowed
us to quantify possible influences of anthropogenic
versus natural N sources to the reef macroalgae. The
HAB species Codium isthmocladum and Caulerpa
brachypus, as well as other abundant macroalgae
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221110
Table 1
Source d
15
N values for the southeast Florida study area
Source and location d
15
N(%) Reference
Ocean sewage outfall
N. Broward County +8.6 Hoch et al. (1995)
Septic tank effluent
Jupiter Creek Monitor Well #4 +7.3 Lapointe and Krupa (1995a)
Jupiter Creek Monitor Well #5 +19.5 Lapointe and Krupa (1995a)
Tequesta Monitor Well #6 +4.6 Lapointe and Krupa (1995b)
Tequesta Monitor Well #10 +11.8 Lapointe and Krupa (1995b)
Upwelled Nitrate
North Atlantic Ocean +4.8 Sigman et al. (2000)
Inorganic fertilizer 0 to +3 Owens (1987)
Peat 0 to +3 Heaton (1986)
Atmospheric nitrogen 0 Owens (1987)
(particularly other Caulerpa spp.), were sampled and
analyzed for tissue d
15
N(Table 2).
2.2. Collection and preparation of macroalgae
SCUBA was used to sample macroalgae from
the 21 reef sites between 10 and 30 May 2001, and
between 20 and 28 August 2001. We collected
samples of Codium isthmocladum,Caulerpa spp.,
and other abundant macroalgae into nylon mesh bags.
Water temperatures and depths at the reef sites were
measured using Oceanic Datamax Pro Plus
TM
dive
computers. Immediately following collection, macro-
algae were identified (Littler and Littler, 2000), sorted
to species, cleaned of debris, transferred to plastic
Ziploc
TM
baggies, and held in a cooler for transport to
the Marine Nutrient Dynamics Lab at HBOI. In the
lab, the samples were rinsed briefly in deionized water
to further remove debris and salt and then randomly
sorted into five replicate composite samples per
species (3–6 individual thalli per composite). The
cleaned tissues were placed in plastic drying dishes
and dried in a Fisher Scientific Isotemp
TM
oven at
60 8C for 48 h. Dried macroalgal thalli were ground to
a fine powder using a mortar and pestle and stored in
plastic vials until analysis.
2.3. Analysis of macroalgal d
15
N
Samples of dried, powdered macroalgae were
analyzed for stable nitrogen isotope ratios with a
Carlo-Erba N/A 1500 Elemental Analyzer and a
VG Isomass mass spectrometer using Dumas com-
bustion. The standard used for stable nitrogen
isotope analysis was N
2
in air. d
15
N values (%) were
calculated as [(R
sample
/R
standard
)1] 10
3
, with R
equal to
15
N/
14
N. Values were statistically compared
using one- and two-way ANOVA for main effects,
after heterogeneity of variance of the data were tested
using an F-test. Fisher’s PLSD multiple comparisons
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1111
Table 2
Macroalgal species collected for d
15
N analysis in the wet and dry seasons, 2001
Species Location
Jupiter Juno Beach Riviera
Beach
Lake Worth Boynton
Beach
Boca Raton Deerfield
Beach
May August May August May August May August May August May August May August
Chlorophyta
Caulerpa brachypus D M,D M,D M,D M,D D
Caulerpa mexicana S
Caulerpa racemosa D S S,D S D S S,D D
Caulerpa verticillata MM M MM
Caulerpa spp. D
Codium isthmocladum M,D M,D S,M,D M,D M,D M,D D D S,D M,D D M,D D M,D
Rhodophyta
Bryothamnion triquetrum SS
Ceramium sp. M M
Galaxaura oblongata MMM
Halymenia elongata DDM
Halymenia echinophysa D
Hypnea musciformis S
Laurencia poiteaui SSSSSSS SS
Wrangelia sp. S
Phaeophyta
Dictyota cronulata S M,D M M
Dictyota menstralis S
Lobophora variegata MM MM
Padina sanctae-crucis SSS
Spatoglossum shroederi S
S = shallow reefs (<5 m), M = mid-depth reefs (25–30 m), D = deep reefs (40–43 m).
test was used to compare groups (e.g. sites, transects)
within main treatment effects of ANOVA. The
analyses facilitated comparison of discrete N sources
assimilated by macroalgae among the study sites,
transects, and between dry and wet season samplings.
Results were considered significant when the prob-
ability ( p) of the null hypothesis was less than 0.05
(p<0.05).
2.4. Analysis of dissolved inorganic nutrients in
upwelled water
One liter polyethylene HDPE bottles were used to
collect samples (n= 2) of cold (<25 8C), upwelled
bottom water between 20 and 25 August 2001 at the
following deep stations: Jupiter, Juno Beach, Riviera
Beach, Lake Worth, and Deerfield Beach. Aliquots of
the water samples were filtered through 0.45 mm
Whatman GF/F filters and analyzed for NH
4+
–N,
NO
3
–N, NO
2
–N, and PO
43
–P (SRP) by the
Nutrient Analytical Services Laboratory, Chesapeake
Biological Laboratory, Solomons, MD. NO
3
and
SRP were analyzed using a Technicon Auto Analyzer
II whereas NH
4+
and NO
2
were analyzed using a
Technicon TRAACS 800. Detection limits for these
analyses were 0.21 mM for NH
4+
, 0.01 mM for
NO
3
+NO
2
, 0.01 mM for NO
2
, and 0.02 mM for
SRP (see D’Elia et al., 1997).
3. Results
3.1. d
15
N of macroalgae
A total of 431 tissue samples of macroalgae were
analyzed for d
15
N in this study. Results are presented
below for the main effects of transect location and
depth within the dry season, the wet season, and of
interactions among seasons, locations, and genera.
3.2. Dry season sampling
A total of 209 samples of macroalgae (Table 2)
were processed and analyzed for d
15
N from the May
sampling of the 21 reef sites. Overall, there was a
significant (ANOVA, F= 11.12, p<0.0001) effect of
location on the d
15
N values of macroalgae during the
dry season (Fig. 3) – the lowest d
15
N values occurring
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221112
Fig. 3. d
15
N values for reef macroalgae at three depths along the seven transects in the dry (May) vs. wet (August) seasons, 2001. Values
represent mean S.E. (n= 5–15).
at Juno Beach (+5.66 1.32%,n= 32) and the
highest at Boca Raton (+8.50 1.27%,n= 24) –
reflecting a general trend of increasing d
15
N values
from Juno Beach southward to Boca Raton (Fig. 3).
The d
15
N values in macroalgae at Juno Beach were
significantly lower (Fisher’s post hoc test) than those
at Jupiter ( p= 0.0005), Riviera Beach ( p<0.0001),
Lake Worth ( p= 0.0018), Boynton Beach ( p=
0.0073), Boca Raton ( p<0.0001), and Deerfield
Beach ( p<0.0001). Conversely, the d
15
N values in
macroalgae at Boca Raton were significantly higher
than those at Jupiter ( p= 0.0003), Juno Beach
(p= 0.0005), Riviera Beach ( p<0.0001), Lake
Worth ( p<0.001), Boynton Beach ( p<0.001),
and Deerfield Beach ( p= 0.0045).
The mean d
15
N values of reef macroalgae were
significantly (F= 47.54, p<0.0001) affected by depth
during the dry season. At all locations, except Boca
Raton (an outfall site), the highest d
15
N values occurred
on shallow reef sites rather than deep reef sites (Fig. 3).
Overall, the mean d
15
N of macroalgae on shallow reefs
(+8.06 1.10%,n= 56) was significantly ( p<0.001)
higher than that of mid-depth (+6.05 1.22%,n= 80)
and deep reefs (+ 6.65 1.76%,n= 74). The mean
d
15
N of macroalgae on the mid-depth reefs was
significantly ( p<0.0008) lower than that of the deep
reef sites.
3.3. Wet season sampling
A total of 222 samples of macroalgae (Table 2)
were processed and analyzed for d
15
N from the
August sampling of the 21 stations. There was a
significant (ANOVA, F= 8.96, p<0.0001) effect of
location on the d
15
N values of reef macroalgae. As in
the dry season, the lowest d
15
N values occurred at
Juno Beach (+6.44 1.49%,n= 40) and the highest
at Boca Raton (+7.86 1.69%,n= 30) with a trend
of increasing d
15
N values southward from Juno Beach
to Boca Raton (Fig. 3). The d
15
N values of macroalgae
at Juno Beach were significantly lower (Fisher’s post
hoc test) than those at Lake Worth ( p= 0.0045),
Boynton Beach ( p= 0.0013), and Boca Raton
(p<0.0001). The highest d
15
N values of macroalgae
were at Boca Raton, which were significantly higher
than those at Jupiter ( p= 0.0004), Juno Beach
(p= 0.0001), Riviera Beach ( p<0.0001), and Deer-
field Beach ( p= 0.011).
There was a significant (ANOVA, F= 87.23,
p<0.0001) effect of reef depth on the d
15
N values
of macroalgae in the wet season. At all locations, the
highest d
15
N values occurred on the shallow reef sites
rather than the mid-depth or deep reef sites (Fig. 3).
Overall, the mean d
15
N of macroalgae on shallow
reefs (+8.58 0.87%) was significantly ( p<0.0001)
higher than that of mid-depth reefs (+6.7 0.89%),
which was significantly ( p<0.0004) higher than that
of the deep reef sites (+6.3 1.26%).
3.4. Interactions among seasons, locations and
genera
Two-way ANOVA indicated significant effects of
location (F= 52.36, p<0.001), genera (F= 13.10,
p<0.001), and the location x genera interaction
(F= 2.45, p= 0.047) on d
15
N values. The overall
mean d
15
N value of Codium isthmocladum (+6.95
0.97%,n= 45) was significantly higher than that of
Caulerpa spp. (+5.52 0.88%,n= 55) and the mean
for all macroalgae (+6.44 1.12%,n= 169, Fig. 4).
For the shallow reefs, the mean d
15
N value of C.
isthmocladum (+ 8.37 1.0%,n= 7) was statistically
similar to that of Caulerpa spp. (+8.36 1.47%,
n= 15) and to the overall mean for all macroalgae
(+8.31 1.03%,n=106, Fig. 4). On the mid-depth
reefs, however, the mean d
15
N value of C. isthmocla-
dum (+6.95 0.97%,n= 45) was significantly higher
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1113
Fig. 4. d
15
N values for Codium spp., Caulerpa spp., and all macro-
algae grouped by depth strata from all transects for the dry (May)
and wet (August) season sampling in 2001. Values represent mean
S.E. (n= 30–35).
than that of Caulerpa spp. (+5.52 0.88%,n= 55,
p<0.001) and higher than the overall mean for all
macroalgae (+6.44 1.12%,n=169, p<0.047;
Fig. 4). Similarly, on the deep reefs the mean d
15
N
value of C. isthmocladum (+ 7.16 1.26%,n= 65)
was significantly higher than that of Caulerpa spp.
(+ 5.59 0.89%,n= 59, p<0.001) and higher than
the mean for all macroalgae (+6.47 1.53%,n= 158,
p<0.047, Fig. 4).
Although the overall effect of season was not
significant among all transects and depths in the study
(p= 0.111), significant differences in mean d
15
N
values between the dry and wet seasons of 2001 were
apparent in both Codium isthmocladum and Caulerpa
spp. from mid-depth and deep reefs in the northern
study area off Juno Beach and Jupiter (Fig. 5). Two-
way ANOVA of these data indicated significant effects
of location (F= 55.06, p<0.001), genera (F= 99.38,
p<0.0001), season (F= 9.58, p= 0.003), the sea-
son genera interaction (F= 26.59, p<0.0001), the
season location interaction (F= 25.41, p<0.0001)
and the season depth interaction (F= 51.06,
p<0.0001). d
15
N values were generally higher in
C. isthmocladum than in Caulerpa spp. and increased
from the dry season to wet season at both mid-depth
and deep reefs off Jupiter, and at the mid-depth reef off
Juno Beach (Fig. 5). The d
15
N values of Caulerpa spp.
also increased from the dry to wet season on the mid-
depth reef off Jupiter, but decreased at the deep reefs
off both Jupiter and Juno Beach (Fig. 5).
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221114
Fig. 5. d
15
N values of Codium isthmocladum and Caulerpa bra-
chypus at mid and deep stations in the dry (May) and wet (August)
seasons at Jupiter and Juno Beach. Values represent mean S.E.
(n= 5–10).
Fig. 6. Linear regressions for temperature vs. concentrations of
nitrate (NO
3
, A) and soluble reactive phosphorus (SRP, B) in the
study area during the upwellings of August 2001. Values represent
mean S.D. (n= 2).
3.5. Nutrient concentrations of upwelled water
During the wet season sampling we observed
elevated NO
3
and SRP concentrations associated
with a cold-water upwelling event. NO
3
concentra-
tions at the deep sites of five transects ranged from 2 to
9mM during these upwellings, with the highest
concentrations measured at the Juno Beach deep reef
site. These elevated NO
3
concentrations correlated
significantly and negatively (R
2
= 0.90) with water
temperature (Fig. 6A). SRP concentrations were
elevated up to 0.75 mM in the upwelled waters
and also correlated significantly and negatively
(R
2
= 0.84) with water temperature (Fig. 6B).
4. Discussion
Results of this study support the hypothesis that
anthropogenic N sources have contributed to the
development of macroalgal HABs and the explosive
invasion of Caulerpa brachypus on southeast Florida’s
coral reefs in recent years. This study also represents
the first report on the distribution of C. brachypus var.
parvifolia in the coastal waters of southeast Florida
(Fig. 2). More broadly, these findings contribute to a
growing recognition of the role of anthropogenic
nutrient enrichment in relieving nutrient limitation
while enhancing the productivity, biomass, and
ultimate success of invasive HAB species, particularly
chlorophytes in the genus Caulerpa (see also:
Chisholm et al., 1997; Jaubert et al., 2003; Lapointe
et al., 2005a).
4.1. The role of upwelling in nutrient enrichment
of reefs
Although previous studies have described tem-
perature anomalies associated with summer upwelling
along Florida’s southeast coast (Green, 1944; Taylor
and Stewart, 1958; Smith, 1982), this study provides
one of the first reports of NO
3
and SRP concentra-
tions associated with these phenomena in this study
area. Coastal upwelling was not apparent during
previous studies of the Codium isthmocladum blooms
in the summer of 1994 when mean NO
3
concentra-
tions were relatively low (0.66 0.33 mM) compared
to NH
4+
(0.96 0.76 mM) and low temperatures
associated with upwelling did not occur (Lapointe,
1997). However, strong upwelling did occur during
the present study in August 2001 when increasing
NO
3
concentrations up to 9 mM correlated signifi-
cantly with decreasing temperature (to 19 8C) and reef
depth (to 43 m). These upwelling events typically
occur during summer months when southerly winds
(parallel to shore) and onshore movement of the
Florida Current produce conditions favorable for
upwelling (Smith, 1982), which can persist for a
period of 7–14 days (Lee and Mayer, 1977). At Conch
Reef in the Upper Florida Keys, Leichter et al. (2003)
reported NO
3
concentrations of 4mM associated
with internal tidal bores (which they referred to as
upwelling) but there was no significant correlation
with temperature. Leichter et al. (2003) also reported
relatively high concentrations of NH
4+
(1.0–2.2 mM)
associated with the elevated NO
3
concentrations but
did not consider the importance of SGD at Conch Reef
(see Simmons, 1992)orNH
4+
as a preferred N source
to C. isthmocladum. At three stations along an 18 km
transect extending from Big Pine Key offshore to the
Looe Key Sanctuary Protection Area in the Lower
Florida Keys, Lapointe et al. (2004) found that NH
4+
concentrations increased significantly from the winter
dry season to the summer wet season; this pattern
correlated significantly with seasonal increases in
rainfall, offshore advection of sewage-contaminated
stormwater runoff, increased macroalgal biomass, and
corresponding enrichment of macroalgal d
15
N.
Previous nutrient kinetic studies with macroalgae
provide mechanistic evidence as to why episodic,
upwelled NO
3
has not historically supported bloom
formation and excessive biomass of Codium isthmo-
cladum and Caulerpa spp. in southeast Florida. In
controlled laboratory studies, uptake of DIN by
Codium fragile subsp. tomentosoides was highly
dependant upon light, temperature, and the source
of DIN – i.e., NO
3
versus NH
4+
(Hanisak and Harlin,
1978). At temperatures of 20–25 8C, the uptake rate of
NH
4+
was 7-fold greater than that of NO
3
and the
presence of NH
4+
inhibited uptake of NO
3
. Similar
preferences for NH
4+
over NO
3
have been reported
for red macroalgae (D’Elia and DeBoer, 1978) as well
as for natural phytoplankton communities (Conway,
1977). Because an average of 0.91 mMNH
4+
was
present in the near-bottom waters during the upwelling
in August 2001, it is unlikely that NO
3
was a major
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1115
DIN source to either Codium isthmocladum or
Caulerpa spp. In a seasonal upwelling system similar
to that of southeast Florida, Fujita et al. (1989)
reported that relatively low concentrations of NH
4+
(1.22 mM) supplied >100% of the N required for
maximum growth of the chlorophyte Ulva rigida, even
in the presence of much higher NO
3
concentrations
(10.8 mM).
The turbid, low light conditions and low tempera-
tures associated with upwelling events could further
limit the ability of Codium isthmocladum and
Caulerpa spp. to assimilate upwelled NO
3
.Codium
fragile subsp. tomentosoides assimilated NH
4+
more
slowly in the dark than in the light, but the uptake rate
in the dark (and under low irradiance) was still higher
than the uptake rate for NO
3
in the light (Hanisak and
Harlin, 1978). Furthermore, N uptake was suppressed
as temperatures decreased (Hanisak and Harlin, 1978).
We observed these sub-optimal growth conditions
during the summer 2001 upwelling event, which
appeared to coincide with diminished bloom devel-
opment. These observations support a conceptual
model that predicts bloom formation to be highly
dependant on the presence of 1mMNO
4+
, high
downwelling irradiance, and temperatures of 25–
29 8C(Lapointe and Hanisak, 1997), conditions that
do not occur during cold, plankton-rich upwelling
events in the southeastern United States (Atkinson
et al., 1984; Atkinson, 1985).
4.2. Land-based nutrient enrichment in southeast
Florida
Multiple lines of evidence from these studies support
the hypothesis that N derived from land-based sources,
rather than coastal upwelling, was the primary N source
supporting the blooms of Codium isthmocladum and
Caulerpa spp. on reefs in southeast Florida. First, the
blooms of C. isthmocladum and Caulerpa spp. were
well developed in mid-May 2001, prior to the onset
of the summer upwelling period (Taylor and Stewart,
1958). Extensive blooms of C. brachypus were
overgrowing sponges, hard corals, octocorals, and
other macroalgal HAB species on reefs in northern
Palm Beach County, whereas reef sites in southern Palm
Beach County were primarily impacted by drift
populations of C. isthmocladum (see Table 2). These
observations in May 2001 support the conceptual model
proposed by Lapointe and Hanisak (1997) that drought
periods may be an important environmental factor
contributing to the formation of macroalgal blooms on
deep reefs off southeast Florida. Like the initial blooms
of C. isthmocladum that developed in southern Palm
Beach County during the drought of 1989–1990, the
blooms we observed in May 2001 occurred during
one of the most severe droughts on record in South
Florida (Abtew et al., 2002). Drought conditions
reduce terrestrial stormwater runoff and increase
water transparency in the coastal ocean, resulting in
maximal downwelling irradiance, which we suspect is
critical for bloom formation on these deep reefs
(Lapointe and Hanisak, 1997). Water transparency
during the May 2001 sampling was very high, most
reefs having >33 m vertical visibility.
Secondly, the lack of a significant change in d
15
N
values of macroalgae between the non-upwelling
(temperatures >25 8C) May 2001 sampling and the
August 2001 upwelling period supports the hypothesis
that upwelled NO
3
was not a major DIN source
supporting the macroalgalblooms in the study area. The
upwelling in August 2001 was most pronounced on the
Juno Beach deep reefs where temperatures of 19 8C
coincided with peak NO
3
concentrations of 9.29 mM.
The macroalgae from this deep site had the lowest d
15
N
value of all sites during the August sampling and closely
matched the d
15
N signature (+4.8%) of North Atlantic
deep-water NO
3
(Sigman et al., 2000; Montoya et al.,
2002). This pattern suggests that upwelled NO
3
was
reducing, not enriching, the higher background d
15
N
values of macroalgae in the study area. This pattern can
best be explained by the dominance of another N source
with a higher d
15
N content than N associated with
upwelling.
The significantly higher d
15
N values in macroalgae
on the shallow reefs during this study clearly indicate a
land-based source of N enrichment to the HABs. The
magnitude of the d
15
N values obtained from the
shallow reefs are in the range of human sewage and
could be delivered to coastal waters of the study area
via a variety of sources and pathways. An estimated
77,391 septic tanks occur in Palm Beach County with
another 106,254 in Broward County (www.doh.state.-
fl.us). These on-site sewage disposal systems con-
taminate shallow groundwaters and downstream
surface waters with elevated concentrations of
NH
4+
,NO
3
, and SRP (Lapointe and Krupa, 1995).
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221116
The d
15
N values of these contaminated groundwaters
range from +4%to +18%, increasing with distance
from the source as a result of fractionation associated
with volatization of ammonia and nitrification
(Lapointe and Krupa, 1995a,b).
Other sources of sewage N to reefs off southeast
Florida are ocean outfalls, which deliver NO
4+
-
enriched wastewaters directly into the water column
in the southern part of our study area. Six ocean outfalls
located off of southeast Florida include: Delray Beach
(89 MLD), Boca Raton (65 MLD), North Broward (244
MLD), Hollywood (163 MLD), North Dade (370
MLD), and Central Dade (529 MLD), which collec-
tively deliver 1462 MLD of secondarily treated
wastewater with high NH
4+
concentrations (900 mM)
into coastal waters (Hazen and Sawyer, 1994). Three of
these outfalls, Delray Beach, Boca Raton, and North
Broward, directly influence the water column in the
southern part of the study area (Fig. 2).
Additional pathways for transport of sewage-
derived NH
4+
(and NO
3
) to the mid-depth and deep
reefs are via Classes I and V injection wells and SGD.
Palm Beach and Broward counties have nearly 30
Class I injection well facilities that dispose of NH
4+
-
rich secondarily treated wastewater under pressure to
1100 m depths (USEPA, 2003). One of these
facilities along the coast in Palm Beach County has
been identified in a recent EPA Wastewater Risk
Assessment Report (USEPA, 2003) as a facility with
upward vertical migration of effluent into a federally
protected and regulated drinking water aquifer. Meyer
(1989) described how upward vertical movement of
groundwater from these depths occurs as a result of
thermal convection, density gradients, and buoyant
flow. Class V wells discharge into depths <30 m and
are used primarily for stormwater runoff rather than
for sewage disposal. Swayze and Miller (1984)
reported a highly permeable surficial aquifer unit
between 20 and 33 m depths that could be enriched by
the upward flow of wastewater and stormwater N from
deeper zones. This geological unit extends offshore to
the reef outcrops where the macroalgal HABs occur.
4.3. d
15
N source assimilation and fractionation in
reef macroalgae
One of the challenges of using stable nitrogen
isotopes in ecological studies is the ‘‘fractionation
effect’’ whereby assimilation of N at higher trophic
levels leads to volitilization of the lighter
14
N,
resulting in stepwise enrichment of
15
N through food
webs (Minagawa and Wada, 1984). Macroalgae are
useful for discriminating specific nutrient sources in
marine ecosystems because they do not fractionate
d
15
N values of their N sources in N-limited systems
(France et al., 1998; Waser et al., 1999). Because the
summer blooms of macroalgae on southeast Florida
reefs are N limited (Lapointe, 1997; Lapointe et al.,
2005b; water column N:P <16:1, macroalgal tissue
N:P <35), fractionation of d
15
N source values by
macroalgae would not be expected. Where fractiona-
tion has been documented between the N source
(groundwater NO
3
) and coral reef macroalgal tissue,
enrichment in tissue d
15
N was slight (0.2–1.4%; see
Umezawa et al., 2002). For zooxanthellate reef corals,
irradiance can confound the fractionation process
when increased photosynthesis under high irradiance
favors reduced fractionation of d
15
N(Muscatine and
Kaplan, 1994; Heikoop et al., 1998). Fortunately, this
is not the case with macroalgae. Low d
15
N values
(+0.5%) are typical of macroalgae growing on
shallow, oligotrophic reefs that experience high
irradiance and natural N sources, such as nitrogen
fixation (see France et al., 1998), whereas elevated
d
15
N values of macroalgae occur in shallow sites
receiving sewage N enrichment (Umezawa et al.,
2002; Lapointe and Thacker, 2002).
High human population density and associated
wastewater NH
4+
loadings from sewage outfalls were
associated with elevated d
15
N values of macroalgae in
the southern study area where blooms of C. isthmo-
cladum were centered. The d
15
N value of effluent from
the North Broward outfall just south of Deerfield Beach
was reported to be +8%(Hoch et al., 1995), similar to
the range of values of +8%to +12%widely reported
for secondarily treated effluent (Lindau et al., 1989;
Costanzo et al., 2001; Savage and Elmgren, 2004). The
warm, buoyant fresh water effluents discharged from
the sewage outfalls at 27 m depths quickly rise to the
surface, potentially affecting shallow, mid, and deep
reefs. The reported outfall d
15
N values closely matched
the d
15
N signature of macroalgae on the mid and
deep reefs near the Boca Raton outfall, as well as the
shallow reefs throughout the study area.
In comparison, the northern study area had a lower
population density, more diffuse anthropogenic N
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1117
enrichment (no outfalls), and the reefs were dominated
by Caulerpa spp. The lowest d
15
N values (+5.7%)
in macroalgae occurred at the most offshore station –
Juno Beach in northern Palm Beach County –
suggesting relatively low-level enrichment of sewage
d
15
N at this site. In addition to effects of increased
distance from the sewage outfalls in the southern
portion of the study area, relatively lower d
15
N values
in northern Palm Beach County may also reflect the
contribution of more diffuse sewage N enrichment via
SGD. These factors would tend to lower d
15
N values
closer to that of Atlantic subsurface water (+4.8%,
Sigman et al., 2000; Montoya et al., 2002). Umezawa
et al. (2002) reported a similar dilution of d
15
N values
of macroalgae associated with deep-water NO
3
moving onto coral reef systems with high rates of
nitrogen-fixation.
The pathway by which chronic anthropogenic
nutrients enter coastal waters can confer competitive
advantages upon Codium and/or Caulerpa and may
contribute to their success in eutrophic coastal waters.
The siphonaceous Codium isthmocladum relies
entirely on the water column for nutrients, compared
to the Caulerpa spp. that can access sediment and/or
reef pore water nutrient pools with their root-like
rhizoids. With these structures, Caulerpa spp.
compete successfully with tropical seagrasses for
sediment NH
4+
in the Caribbean (Williams, 1984;
Williams and Fisher, 1985). Different nutrient
acquisition strategies may explain why C. isthmocla-
dum blooms are best developed in southern Palm
Beach County, where sewage from ocean outfalls is a
primary source of N enrichment of the water column.
In contrast, SGD is likely to be a more important N
source to the reefs in northern Palm Beach County,
which lacks sewage outfalls and where C. brachypus,
C. verticillata, and C. racemosa accounted for
extensive cover on the reefs (Fig. 1C–F; up to 80%
cover, Lapointe et al., 2004). The ability of Caulerpa
spp. to attach firmly to the reefs with their root-like
rhizoids would also be an asset in these turbulent flow
fields and would be a competitive advantage in
accessing N pools in reef interstitial waters. Because
thick mats of macroalgae may attenuate NH
4+
flux
from sediments to the overlying water column
(McGlathery et al., 1997), dense Caulerpa blooms
could intercept nutrient supplies which would other-
wise be available to C. isthmocladum and other
macroalgae that depend on the water column for
nutrients. This hypothesis is supported by the
significantly higher d
15
N values of C. isthmocladum,
compared to Caulerpa spp., on mid-depth and deep
reefs, and may explain the prevalence of C.
isthmocladum on reefs in southern Palm Beach
County where the water column is chronically
enriched by sewage effluent from ocean outfalls.
Whereas C. isthmocladum was most prevalent in the
southern portion of the study area, both C. isthmo-
cladum and Caulerpa spp. were found throughout the
study area (see Table 2), which facilitated this
comparison.
The conclusion that sewage was a primary N source
to these macroalgal blooms is supported by several
other studies from the scientific literature reporting
elevated d
15
N values in the range of those reported in
this study. The d
15
N ratio of reef macroalgae ranged
from +5.7%off Juno Beach to +8.5%off Boca Raton,
values within the range reported for macroalgae
growing on sewage N in both temperate and tropical
coastal waters (Lapointe, 1997; McClelland and
Valiela, 1998; France et al., 1998; Costanzo et al.,
2001; Lapointe and Thacker, 2002; Umezawa et al.,
2002; Barile, 2004; Savage and Elmgren, 2004;
Table 1). The significant cross-shelf pattern of
elevated d
15
N associated with land-based sources of
pollution reported here parallels the findings of
Sammarco et al. (1999) who reported that the
influence of terrestrial N sources extended seaward
across nearly 40 km of the Great Barrier Reef lagoon.
4.4. Cultural eutrophication of southeast Florida
coral reefs
The SEFLOE II investigations (Hazen and Sawyer,
1994), funded by local wastewater utilities and
performed in collaboration with the NOAA Atmo-
spheric, Oceanographic and Meterological Laboratory
(AOML) in Miami, FL, assessed the environmental
impacts of sewage outfalls in the southern portion of
our study area. The SEFLOE II study reported that
ambient concentrations of NH
4+
on the reefs in north
Broward County averaged 6mM and that maximum
ambient concentrations were 36 mM. Those back-
ground NH
4+
concentrations are some 6- to 36-fold
higher than the mean NH
4+
concentrations reported by
Lapointe (1997) and Lapointe et al. (2005b) for this
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221118
area, which reflect not only the actual background
concentration, but also the contribution of the waste-
water loadings. Hazen and Sawyer (1994) used these
erroneous NH
4+
concentrations in a dilution model
that supported their ‘‘finding of no significant impact’’
(FONSI) for the north Broward County outfall, which
discharges 1100 metric t/year of NH
4+
into the water
column over coral reefs in 27 m depths. Section 403
of the USEPA (1994) Clean Water Act concluded:
‘‘Because of the relatively short term of the SEFLOE
studies (several years), the long term or cumulative
risks of nutrient loading and loading of other effluent
constituents cannot be evaluated.’’
The Ocean Regulatory Programs section of the
USEPA (1994) Clean Water Act (CWA, Section 403)
specifically addresses impacts on the marine environ-
ment from point source discharges. The section’s
requirements are intended to ensure that no unreason-
able degradation of the marine environment will
occur as a result of the discharge and that sensitive
ecological communities are protected. Unreasonable
degradation is defined as ‘‘significant adverse changes
in ecosystem diversity, productivity, and stability of
the biological community within the area of discharge
and surrounding biological community.’’ Sensitive
ecological communities include unique species or
communities, endangered or threatened species,
species critical to the structure or function of the
ecosystem, nursery/forage areas, and coral reefs. In
our study, the d
15
N values in macroalgae (+8%)on
coral reefs off Deerfield Beach closely matched the
values reported by Hoch et al. (1995) for the NH
4+
-
rich effluent from the north Broward County outfall
located ‘‘upstream’’ several kilometers to the south.
While the SEFLOE II Report utilized empirical
modeling estimates to suggest no impact of outfall
sewage to the continental shelf biota, our study
provides source-sink evidence of sewage N assimila-
tion from outfalls into reef HABs. The southeast
Florida reefs provide habitat to a number of
endangered species (e.g. sea turtles) and finfish that
have been affected by diseases associated with toxin
production, possibly from epiphytes of these macro-
algal HABs (Landsberg, 1995).
In summary, these results provide multiple lines of
evidence that macroalgal assimilation of sewage N
from ocean outfalls and other land-based sources
(septic tanks, injection wells) supports HABs and their
degradation of coral reef habitats. If coral reef habitats
are to be protected in the study area, we recommend
that our results be considered in future National
Pollution Discharge Elimination System (NPDES)
permitting of the sewage outfalls, particularly as the
NPDES process addresses the concept of ‘‘unreason-
able degradation’’ of coral reef habitats. USEPA states
that ‘‘if section 403 requirements for protection of the
ecological health of marine waters are not met, a
NPDES permit will not be issued.’’
This evidence, that anthropogenic nutrient enrich-
ment from land-based sewage discharges supports
blooms of native Codium isthmocladum and the
invasive Caulerpa brachypus var. parvifolia on coral
reefs in southeast Florida, parallels recent reports that
land-based sources of pollution, specifically human
sewage, supported the invasion of C. taxifolia in the
northwestern Mediterranean (Chisholm et al., 1997;
Jaubert et al., 2003) as well as that of the
Mediterranean native Caulerpa ollivieri in the
Bahamas (Lapointe et al., 2005a). These findings
suggest that current wastewater management practices
in southeast Florida may be in conflict with federal and
state initiatives to reduce the effects of land-based
sources of pollution on sensitive ecosystems, such as
coral reef habitats in southeast Florida. Implementa-
tion of advanced wastewater treatment (AWT) for the
removal of N and P from sewage effluent, along with
increased wastewater reuse for beneficial purposes, is
critical to mitigate the impacts of these wastewater
discharges on southeast Florida’s reefs. This study
supports the conclusion of the Joint Group of Experts
on the Scientific Aspects of Marine Pollution
(GESAMP, see Windom, 1992) that nutrient pollution
from sewage and other land-based sources poses the
greatest present and future threat to the coastal marine
environment (Windom, 1992).
Acknowledgements
We thank Jupiter Dive Center, S.S. Minnow
Charters, Jim Abernathy’s Scuba Adventures, Splash
Down Divers, and Bill Parks for providing boat
support for this research. We are grateful to Connie
Gasque who assisted as a volunteer diver during these
studies. Michael Wynne (University of Michigan)
provided positive identification of C. brachypus var.
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1119
parvifolia. Funding was provided by the State of
Florida Harmful Algal Bloom Task Force through sub-
agreement # 4710173L3A with the Florida Institute of
Oceanography. This is contribution # 1596 from the
Harbor Branch Oceanographic Institution and con-
tribution # 613 from the Smithsonian Marine Station
at Fort Pierce, FL. [SES]
References
Abtew, W., Huebner, R.S., Sunderland, S., 2002. Part I. Hydro-
logical analysis of the 2000–2001 drought in South Florida.
Technical Report EMA-405, South Florida Water Management
District, West Palm Beach, FL.
Atkinson, L.P., 1985. Hydrography and nutrients of the southeastern
U.S. continental shelf. Oceanogr. Southeast U.S. Cont. Shelf
(AGU) 2, 77–91.
Atkinson, L.P., O’Malley, P.G., Yoder, J.A., Paffenhoffer, G.A.,
1984. The effect of summertime shelf break upwelling on
nutrient flux in southeastern United States continental shelf
waters. J. Mar. Res. 42, 969–993.
Barile, P.J., 2004. Evidence of anthropogenic nitrogen enrichment of
the littoral waters of east central Florida. J. Coast. Res. 20 (4),
1237–1245.
Chisholm, J.R.M., Fernex, F.E., Mathieu, D., Jaubert, J.M., 1997.
Wastewater discharge, seagrass decline and algal proliferation
on the Cote d’ Azur. Mar. Poll. Bull. 34, 78–84.
Conway, H.L., 1977. Interaction of inorganic nitrogen in the uptake
and assimilation by marine phytoplankton. Mar. Biol. 39, 221–
232.
Costanzo, S.D., O’Donohue, M.J., Dennison, W.C., Loneragan,
N.R., Thomas, M., 2001. A new approach for detecting and
mapping sewage impacts. Mar. Poll. Bull. 42, 149–156.
D’Elia, C.F., DeBoer, J.A., 1978. Nutritional studies of two red
algae. 2. Kinetics of ammonium and nitrate uptake. J. Phycol.
14, 266–272.
D’Elia, C.F., Connor, E.E., Kaumeyer, N.L., Keefe, C.W., Wood,
K.V., Zimmerman, C.F., 1997. Nutrient Analytical Services
standard operating procedures. Technical Report Series No.
158-97. Chesapeake Biological Laboratory, Solomons, MD.
ECOHAB, 1997. The Ecology and Oceanography of Harmful Algae
Blooms. A National Research Agenda. Anderson, D.M. (Ed.),
WHOI, 66 p.
Finkl, C.W., Charlier, R.H, 2003. Sustainability of subtropical
coastal zones in southeastern Florida: challenges for urbanized
coastal environments threatened by development, pollution,
water supply, and storm hazards. J. Coast. Res. 19 (4), 934–943.
France, R., Holmquist, J., Chandler, M., Cattaneo, A., 1998. Evi-
dence for nitrogen fixation associated with macroalgae from a
seagrass–mangrove–coral reef system. Mar. Ecol. Progr. Ser.
167, 297–299.
Fujita, R.M., Wheeler, P.A., Edwards, R.L., 1989. Assessment of
macroalgal nitrogen limitation in a seasonal upwelling region.
Mar. Ecol. Progr. Ser. 53, 293–303.
Gartner, A., Lavery, P., Smit, A.J., 2002. Use of d
15
N signatures of
different functional forms of macroalgae and filter feeders to
reveal temporal and spatial patterns in sewage dispersal. Mar.
Ecol. Progr. Ser. 235, 63–73.
Green, C., 1944. Summer upwelling-northeast coast of Florida.
Science 100, 546–547.
Hanisak, M.D., Blair, S.M., 1988. The deep water macroalgal
community of the East Florida continental shelf (USA). Helgo-
lan. Meeres. 42, 133–163.
Hanisak, M.D., Harlin, M.M., 1978. Uptake of inorganic nitrogen by
Codium fragile subsp. tomentosoides (Chlorophyta) J. Phycol.
14, 450–454.
Hazen and Sawyer, 1994. SEFLOE II Final Report: Broward
County Office of Environmental Services North Regional
Wastewater Treatment Plant; City of Hollywood Utilities
Department Southern Region Wastewater Treatment Plant;
Miami-Dade Water and Sewer Department North District Was-
tewater Treatment Plant; Miami-Dade Water and Sewer
Department Central District Wastewater Treatment Plant
Hollywood (FL): Hazen and Sawyer. Submitted to the
National Oceanic and Atmospheric Administration (AOML,
Miami, FL).
Heaton, T.H.E., 1986. Isotopic studies of nitrogen pollution in the
hydrosphere and atmosphere: a review. Chem. Geol. (Isotope
Geoscience Section) 59, 87–102.
Heikoop, J.M., Dunn, J.J., Risk, M.J., Sandeman, I.M., Schwarcz,
H.P., Waltho, N., 1998. Relationship between light and the d
15
N
of coral tissue: Examples from Jamaica and Zanzibar. Limnol.
Oceanogr. 43, 909–920.
Hobbie, J.E., Larsson, U., Elmgren, R., Fry, B., 1990.
Sewage derived
15
N in the Baltic traced in Fucus. EOS 71,
190.
Hoch, M.P., Cifuentes, L.A., Coffin, R., 1995. Assessing geochem-
ical and biological fate for point source loads of sewage-derived
nitrogen and organic carbon in coastal waters of southern
Florida. Final Report to the USEPA.
Howarth, R.W., Billen, G., Swaney, D., Townsend, A., Jaworski, N.,
Downing, J.A., Elmgren, R., Caraco, N., Lajtha, K., 1996.
Regional nitrogen budgets and riverine N & P fluxes for the
drainages to the North Atlantic ocean: natural and human
influences. Biogeochemistry 35, 75–139.
Howarth, R., Anderson, D., Cloern, J., Elfring, C., Hopkinson, C.,
Lapointe, B., Malone, T., Marcus, N., McGlathery, K., Sharpley,
A., Walker, D., 2000. Nutrient pollution of coastal rivers, bays,
and seas. Issues Ecol. 7, 1–15.
Jaubert, J.M., Chisholm, J.R.M., Minghelli-Roman, A., Marchior-
etti, M., Morrow, J.H., Ripley, H.T., 2003. Re-evaluation of the
extent of Caulerpa taxifolia development in the northern Med-
iterranean using airborne spectrographic sensing. Mar. Ecol.
Progr. Ser. 263, 75–82.
Lajtha, K., Michener, H.H., 1994. Stable isotopes in ecology
and environmental science. In: Lawton, J.H., Likens, G.E.
(Eds.), Methods in Ecology. Blackwell Scientific Publications,
Oxford, UK.
Landsberg, J.H., 1995. Tropical reef-fish disease outbreaks and mass
mortalities in Florida USA: what is the role of dietary biological
toxin? Dis. Aquat. Org. 22, 83–100.
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221120
Lapointe, B.E., 1997. Nutrient thresholds for bottom-up control of
macroalgal blooms on coral reefs in Jamaica and southeast
Florida. Limnol. Oceanogr. 42 (5, part 2), 1119–1131.
Lapointe, B.E., 1999. Simultaneous top-down and bottom-up forces
control macroalgal blooms on coral reefs (reply to the comment
by Hughes et al.) Limnol. Oceanogr. 44 (6), 1586–1592.
Lapointe, B.E., Hanisak, M.D., 1997. Algal blooms in coastal
waters: Eutrophication on coral reefs of southeast Florida. Final
Report-Florida Sea Grant Project R/C-E-34.
Lapointe, B.E., Krupa, S., 1995a. Jupiter Creek septic tank water
quality investigation. Final Report to the Loxahatchee River
Environmental Control District, Jupiter, FL, p. 96.
Lapointe, B.E., Krupa, S., 1995b. Tequesta Peninsula septic tank
water quality investigation. Final Report to the Loxahatchee
River Environmental Control District, Jupiter, FL, p. 93.
Lapointe, B.E., Thacker, K., 2002. Community-based water qual-
ity and coral reef monitoring in the Negril Marine Park,
Jamaica: Land-based nutrient inputs and their ecological
consequences. In: Porter, J.W., Porter, K.G. (Eds.), The Ever-
glades, Florida Bay, and Coral Reefs of the Florida Keys: An
Ecosystem Sourcebook. CRC Press, Boca Raton, FL, pp. 939–
963.
Lapointe, B.E., Barile, P.J., Matzie, W.R., 2004. Anthropogenic
nutrient enrichment of seagrass and coral reef communities in
the lower Florida keys: discrimination of local versus regional
nitrogen sources. J. Exp. Mar. Biol. Ecol. 308 (1), 23–58.
Lapointe, B.E., Tomasko, D.A., Matzie, W.R., 1994. Eutrophication
and trophic state classification of seagrass communities in the
Florida keys. Bull. Mar. Sci. 54, 696–717.
Lapointe, B.E., Barile, P.J., Wynne, M.J., Yentsch, C.S., 2005a.
Reciprocal Invasion: Mediterranean native Caulerpa ollivieri in
the Bahamas supported by human nitrogen enrichment. Aq.
Invad. 16 (2), 2–5.
Lapointe, B.E., Barile, P.J., Littler, M.M., Littler, D.S., Bedford, B.,
Gasque, C., 2005b. Macroalgal blooms on southeast Florida
coral reefs. I. Nutrient stoichiometry of the invasive green alga
Codium isthmocladum in the wider Caribbean indicates nutrient
enrichment. Harmful Algae 4, 1092–1105.
Lee, T.N., Mayer, D.A., 1977. Low-frequency current variability and
spin-off eddies along the shelf off southeast Florida. J. Mar. Res.
35 (1), 193–220.
Leichter, J.J., Stewart, H., Miller, S.L., 2003. Episodic nutrient
transportto Florida coral reefs. Limnol. Oceanogr. 48, 1394–1407.
Lindau, C.W., Delaune, R.D.,Patrick, W.H.,Lambremont, E.N.,1989.
Assessment of stable isotopes in fingerprinting surface water
inorganic nitrogen sources. Water Air Soil Poll. 48, 489–496.
Littler, D.S., Littler, M.M., 2000. Caribbean Reef Plants. Offshore
Graphics, Washington, DC.
McClelland, J.W., Valiela, I., 1998. Changes in food web structure
under the influence of increased anthropogenic nitrogen inputs
to estuaries. Mar. Ecol. Progr. Ser. 168, 259–271.
McGlathery, K.J., Krause-Jensen, D., Rysgaard, S., Christensen,
P.B., 1997. Patterns of ammonium uptake within dense mats of
the filamentous macroalga Chaetomorpha linum. Aquat. Bot. 59,
99–115.
Meinesz, A., 1999. Killer Algae. The University of Chicago Press,
Chicago, IL.
Meinesz, A., Hesse, B., 1991. Introduction and invasion of the
tropical alga Caulerpa taxifolia in the Mediterranean. Oceanol.
Acta 14 (4), 415–426.
Meyer, F.W., 1989. Hydrogeology, ground-water movement, and
subsurface storage in the Floridan aquifer system in southern
Florida. U.S. Geological Survey Professional Paper 1403-G.
Miller, J.A., 1997. Hydrogeology of Florida. In: Randazzo, A.F.,
Jones, D.S. (Eds.), The Geology of Florida. University Press,
Gainesville, FL, pp. 69–88.
Minagawa, M., Wada, E., 1984. Stepwise enrichment of
15
N along
food chains: further evidence and the relation between d
15
N and
animal age. Geochem. Cosmochem. Acta 48, 1135–1140.
Montoya, J.P., Carpenter, E.J., Capone, D.G., 2002. Nitrogen fixa-
tion and nitrogen isotope abundances in zooplankton of the
oligotrophic North Atlantic. Limnol. Oceanogr. 47 (6), 1617–
1628.
Muscatine, L., Kaplan, I.R., 1994. Resource partitioning by reef
corals as determined from stable isotope composition. II. d
15
Nof
zooxanthellae and animal tissue versus depth. Pacific Sci. 48,
454–460.
National Research Council, 2000. Clean Coastal Waters: Under-
standing and Reducing the Effects of Nutrient Pollution. Ocean
Studies Board, Water Science and Technology Board, 391 p.
Owens, N.P.J., 1987. Natural variations in
15
N in the marine
environment. Adv. Mar. Biol. 24, 389–451.
Peterson, B.J., Fry, B., 1987. Stable isotopes in ecosystem studies.
Annu. Rev. Ecol. Syst. 18, 293–320.
Rogers, K.M., 1999. Effects of sewage contamination on macro-
algae and shellfish at Moa Point New Zealand using stable
carbon and nitrogen isotopes. New Zeal. J. Mar. Freshwater
Res. 33, 181–188.
Sammarco, P.W., Risk, M.J., Schwarcz, H.P., Heikoop, J.M., 1999.
Cross-continental shelf trends in coral d
15
N on the Great Barrier
Reef: further consideration of the reef nutrient paradox. Mar.
Ecol. Progr. Ser. 180, 131–138.
Savage, C., Elmgren, R., 2004. Macroalgal d
15
N values trace
decrease in sewage influence. Ecol. Appl. 14 (2), 517–526.
Sigman, D.M., Altabet, M.A., McCorkle, D.C., Francois, R., Fisher,
G., 2000. The d
15
N of nitrate in the southern ocean: Nitrogen
cycling and circulation in the ocean interior. J. Geophys. Res. C
105, 19599–19614.
Simmons Jr., G.M., 1992. Importance of submarine groundwater
discharge (SGWD) and seawater cycling to material flux across
sediment/water interfaces in marine environments. Mar. Ecol.
Prog. Ser. 84, 173–184.
Smith, N.P., 1982. Upwelling in Atlantic shelf waters of South
Florida. Florida Sci. 45 (2), 117–125.
Swayze, L.J., Miller, W.L., 1984. Hydrogeology of a zone of
secondary permeability in the surficial aquifer of eastern Palm
Beach County, FL. U.S. Geology Survey, Tallahassee, FL, p. 38.
Taylor, C., Stewart, H., 1958. Summer upwelling along the east
coast of Florida. J. Geophys. Res. 64, 33–39.
Umezawa, Y., Miyajima, T., Yamamuro, M., Kayanne, H., Koike, I.,
2002. Fine-scale mapping of land-derived nitrogen in coral reefs
by d
15
N in macroalgae. Limnol. Oceanogr. 47 (5), 1405–1416.
UNEP, 1994. Regional overview of land-based sources of pollution
in the wider Caribbean region. Caribbean Environment Program
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–1122 1121
Technical Report 33, UNEP Caribbean Environment Program,
Kingston, Jamaica.
USEPA, 1994. Clean Water Act, Section 403 Ocean Regulatory
Programs, Phase II - Point Source Discharges Inside the Base-
line. Report to Congress. EPA 842-R-94-001.
USEPA, 2003. Relative risk assessment of management options for
treated wastewater in south Florida. Office of Water. EPA 816-R-
03-010.
Verlaque, M., Durand, C., Huisman, J.M., Boudouresque, C.-F., La
Parco, Y., 2003. On the identity and origin of the Mediterranean
invasive Caulerpa racemosa (Caulerpales Chlorophyta). Eur. J.
Phycol. 38 (4), 325–339.
Waser, N.A., Zhiming, Y., Yin, K., Nielsen, B., Harrison, P.J.,
Turpin, D.H., Calvert, S.E., 1999. Nitrogen isotopic fractiona-
tion during a simulated diatom spring bloom: importance of N-
starvation in controlling fractionation. Mar. Ecol. Progr. Ser.
179, 291–296.
Wayland, M., Hobson, K.A., 2001. Stable carbon, nitrogen, and
sulfur isotope ratios in riparian food webs on rivers receiving
sewage and pulp-mill effluents Canada. J. Zool. 79, 5–15.
Williams, S.L., 1984. Uptake of sediment ammonium and translo-
cation in a marine green macroalga Caulerpa cupressoides.
Limnol. Oceanogr. 29, 274–279.
Williams, S.L., Fisher, T.R., 1985. Kinetics of nitrogen-15 labelled
ammonium uptake by Caulerpa cupressoides. J. Phycol. 21,
287–296.
Windom, H., 1992. Contamination of the marine environment from
land-based sources. Mar. Poll. Bull. 35, 32–36.
B.E. Lapointe et al. / Harmful Algae 4 (2005) 1106–11221122
