EPA/600/S-21/079
Summary of benthic conditions in the Three Bays estuary (Cape Cod, MA)
as of 2019
Laura E Erban1, Donald J Cobb1, Charles S Strobel1, Casey K Tremper2, James D Hagy III1,
Timothy R Gleason1
Affiliations
1	US EPA Office of Research and Development, Center for Environmental Modeling and
Measurement, Atlantic Coastal Environmental Sciences Division, Narragansett RI
2	ORAU, Oak Ridge TN
Overview
Benthic (seafloor) habitat condition is an important indicator of the overall health of estuarine
ecosystems. Estuaries are widely degraded on Cape Cod, in large part due to excess nutrients in
wastewater, and many now have established Total Maximum Daily Loads (TMDL) for nitrogen.
The Three Bays estuary (Barnstable, MA) has a nitrogen TMDL that calls for substantial load
reductions to improve water quality and restore and maintain high quality benthic habitat
(MassDEP, 2007). As part of TMDL development, a benthic survey was conducted in the early
2000s (Howes et al., 2006) under the Massachusetts Estuaries Project (MEP). This data summary
documents a follow-up survey conducted in 2019 to update the condition assessment for Three
Bays and establish a new baseline for evaluating the impact of planned watershed interventions,
including sewering, centralized wastewater treatment and alternative technologies, to reduce the
load of nitrogen reaching the estuary.
The 2019 benthic survey of Three Bays included the prior stations as well as the TMDL sentinel
station and additional randomized locations. These stations were visited in early September to
make in-situ water quality measurements and collect sediment grabs. Grab samples were
analyzed for grain size distribution, total organic carbon (TOC) content, and taxa-specific
infaunal abundance. Infaunal data were assessed using the common ecological community
metrics used in the prior survey (density, number of species, evenness and diversity). Overall
benthic habitat condition was scored with the now widely used marine biotic index M-AMBI
(Muxika et al., 2007; Sigovini et al., 2013) and associated condition classes for US coastal
waters (Pelletier et al., 2018). Habitat condition was classified as 'poor' or worse at about 50%
of stations (13 out of 25) surveyed in 2019. The worst conditions were found in enclosed upper
areas of the estuary with better conditions in seaward subembayments.
These findings are consistent with the prior MEP survey but suggest that some additional
degradation has occurred. The prior survey found moderate to significant impairment at 7 of the
1

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11 stations sampled. These remain impaired. Most (9) of the revisited stations show no
discernable change in overall habitat condition while the remainder (2) are now assessed to be in
worse condition. This is not unexpected, given that significant nutrient load reduction has yet to
occur in the Three Bays watershed. In other local embayments, including West Falmouth Harbor,
Pleasant Bay, and Wellfleet Harbor, similar surveys are being conducted under a pilot MEP
monitoring study (see Sweeny and Rutecki, 2020). Results are being analyzed with comparable
methods, including the quantitative summary of benthic condition provided by M-AMBI. This
set of surveys will allow for assessment of changes in benthic habitat over time across Cape Cod
estuaries subject to different baseline conditions and different management actions taken to
address excess nutrients.
Methods
The Three Bays estuary spans an area just over 5 km2 on the south coast of Cape Cod, MA.
Named for three interconnected major subembayments (North, West and Cotuit Bays), the
northern end terminates in two smaller coves (Prince and Warren's). Flows from the Marstons
Mills and Little Rivers contribute focused freshwater inputs (-20%, Howes et al., 2006), with the
balance from direct precipitation and dispersed groundwater discharge. Tidal saltwater flow from
Nantucket Sound (Atlantic Ocean) is exchanged through two southern inlets to Cotuit and West
Bays. The average depth of the estuary as a whole is 2.3 m at mean tide level (computed from
1.9 m below NGVD 29 reported by Howes et al., 2006), including large areas that are more
shallow and deeper navigational
channels.
Benthic survey locations (Figure 1)
included 11 fixed stations from the prioi
MEP survey, the TMDL sentinel
station, and 13 new randomized sites.
Randomized sites were selected using
the Generalized Random Tessellation
Stratified (GRTS) method, which
generates spatially balanced,
probabilistic survey designs for a given
sample frame (Stevens and Olsen,
2004). Randomized sites, stratified by
subembayment, were selected using the
spsurvey package for R (Kincaid and
Olsen, 2019). To develop a sample
frame, or spatial representation of the
target population to be sampled, data
was acquired from NO A As Electronic
Navigational Charts
(https://encdirect.noaa.govA accessed
2019-04-18). GIS polygon layers were
r
Stations
¦ fixed
randomized
sentinel
Marstons Mills R.
Warren's Cove
Prince Cove
reqi
North Bay

10
Cotuit Bay
22
25
11
29
West Bay
1 km	|	Nantucket Sound
ft I Leaflet | © OpenStreetMap contributors © CARTO
Figure 1. The Three Bays estuary, subembayments,
and locations of 2019 benthic survey stations.
2

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downloaded for the 'Harbor' scale band for the extent of the estuary, which includes a depth
contour for zero meters below Mean Lower Low Water (MLLW). The sample frame was limited
to areas that are submerged (non-exposed) at MLLW.
Samples were collected from September 9-11, 2019 according to protocols previously developed
for national coastal condition surveys (US EPA, 2015). At each station, in-situ water quality
measurements (see Table 1) were made with a multi-parameter YSIEX02 sonde at shallow and
deep depths in the water column (or a single shallow depth, where the water column was <2m).
Water depth recorded during sampling was used to calculate the mean tide level (MTL) depth by
referencing levels recorded by the NOAA tide gauge at Nantucket adjusted by tidal phase lags
from Howes et al. (2006) and local tidal datums obtained from the NOAA VDATUM tool
(http://vdatum.noaa.gov). Secchi depth was recorded and used to estimate the fraction of incident
light reaching sediments at MTL. Sediment grabs were recovered with a 0.04 m2 Young
modified Van Veen grab sampler. Acceptable grabs were at least 7 cm deep with a level surface.
The first sediment grab, collected for infauna, was sieved on board using ambient seawater and a
0.5 mm screen. All material retained was transferred to a 1 L Nalgene bottle and fixed in 10%
buffered formalin with Rose Bengal stain. The second grab, collected for sediment properties,
was subsampled (-100 cm3) and stored at 4°C prior to analysis.
Infaunal community composition and sediment properties were analyzed by Normandeau
Associates, Inc. and Alpha Analytical, respectively, following established procedures (US EPA,
1995). Briefly, the sieved infaunal sample was sorted under a dissecting microscope and
macroinvertebrates were enumerated and identified by taxonomic group, to the species level
when possible. Sediments were sorted to determine grain size distribution with a set of standard
sieves, and hydrometer for fine fractions, according to ASTM D6913/D7928, and classified as
per the Unified Soil Classification System (USCS). Acidified sediment samples were analyzed
for total organic carbon (TOC) content as in US EPA 9060A, using a Perkin Elmer 2400 Series
II CHNS/O Analyzer with a thermal conductivity detector. Additional details of the analyses
performed by both contractors and their results are provided in the supplemental materials,
available at doi: 10.23719/1520968. Field and laboratory data will also be made available in the
Water Quality Portal to support their broader discoverability.
Computational analysis is coded in the supplemental html notebook, which includes interactive
versions of the maps shown here and R packages used. Specialized packages oce (Kelley and
Richards, 2020) and vegan (Oksanen et al., 2019) were used to calculate relative seawater
densities and infaunal community metrics. Infaunal taxa were assigned to ecological groups for
US coastal waters (hybrid values from Gillett et al., 2015). The AZTI Marine Biotic Index
(AMBI; Boija et al., 2000) was calculated according to the representation of individuals in each
group. Multivariate AMBI scores (M-AMBI; Muxika, et al., 2007) for the salinity bin
(polyhaline) for these samples were calculated following Sigovini et al., (2013). M-AMBI was
not calculated for any station where more than 50% of the sample abundance could not be
assigned to an ecological group (these were typically oligochaetes, or worms). Established values
(see Pelletier et al., 2018) were used for the reference anchor points for the Northeast USA and
cut-points for condition classes (Bad, Poor, Moderate, Good, High).
3

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Table 1. Water quality observations at surveyed stations by type and subembayment. Depths are
for the measured water column at the time of sampling. A Secchi depth measurement of "NA"
indicates that the disk could be seen on the bottom.
Subembayment Station
Type
Date
Time
Latitude Longitude
Depth
(m)
Secchi
depth
(m)
Obs.
depth
(m)
Temp.
(°C)
Salinity
(PPt)
DO
(mg/L)
DO
("osat)
Warren's Cove
1
fixed
9/11/2019
12:27
41.644967 -70.40557
1.0
1
0.5
22.3
22.8
9.2
120.3
Prince Cove
2

9/11/2019
11:29
41.644417 -70.41062
2.4
1.2
1.0
2.0
22.0
22.0
23.1
23.4
9.1
9.0
119.6
117.1

3

9/11/2019
11:08
41.642533 -70.41185
2.2
1
1.0
2.0
22.0
21.8
23.6
24.0
8.8
7.8
115.7
103.5
North Bay
4

9/10/2019
13:48
41.635167 -70.4099
4.2
1.5
1.0
3.0
22.5
21.7
26.0
26.8
9.5
7.8
127.4
103.9

5

9/10/2019
14:13
41.630767 -70.40547
3.6
1.7
1.0
3.0
22.4
21.6
26.1
27.1
9.4
7.8
126.2
100.5

6

9/11/2019
9:39
41.62805 -70.40173
2.0
1.7
1.0
21.7
26.3
8.7
115.4
West Bay
7

9/10/2019
11:11
41.618117 -70.39522
1.4
NA
1.0
21.6
27.9
8.3
111.4

8

9/10/2019
10:09
41.614833 -70.3991
1.4
NA
1.0
21.1
27.9
7.9
104.4
Eel River
9

9/10/2019
12:25
41.612217 -70.39
2.4
2.3
1.0
2.0
22.1
21.7
27.8
28.0
8.9
9.0
119.4
120.7
Cotuit Bay
10

9/9/2019
9:20
41.622867 -70.41848
2.5
2.3
0.5
2.0
20.4
21.0
26.4
27.6
6.8
6.1
88.2
80.3

11

9/9/2019
11:17
41.612967 -70.42712
1.7
NA
1.0
21.1
27.9
7.2
95

12
sentinel
9/11/2019
8:25
41.62925 -70.41192
2.8
1.8
1.0
2.5
21.5
21.5
26.5
26.6
7.4
7.4
97.4
97.3
Prince Cove
13
randomized
9/11/2019
10:46
41.640767 -70.41375
1.7
1
1.0
22.1
24.2
8.8
115.1
Warren's Cove
15

9/11/2019
12:54
41.6433 -70.40618
2.4
1.7
1.0
2.0
22.2
22.1
22.7
24.3
9.6
8.9
125.3
117.8
North Bay
17

9/11/2019
10:10
41.629233 -70.39833
1.9
1.9
1.0
21.7
27.1
8.1
107.6

18

9/10/2019
14:35
41.629617 -70.40332
1.9
1.8
1.0
22.2
26.3
9.5
127.2

19

9/11/2019
9:05
41.6284 -70.4069
1.6
1.5
1.0
21.8
26.6
8.2
109.7
West Bay
22

9/10/2019
9:07
41.618183 -70.40535
1.4
NA
1.0
21.2
28.2
7.9
104.4

23

9/10/2019
10:43
41.619733 -70.39637
1.4
NA
1.0
21.2
28.3
7.9
104.6

24

9/10/2019
11:41
41.61675 -70.39278
1.3
NA
1.0
22.0
28.6
9.0
121.2

25

9/10/2019
9:38
41.616417 -70.40482
1.7
NA
1.0
21.1
27.9
7.9
104.1
Cotuit Bay
28

9/9/2019
12:38
41.6148 -70.43288
2.6
2.2
0.5
2.0
21.3
21.3
27.2
27.2
7.4
7.4
97.9
97.4

29

9/9/2019
10:25
41.611183 -70.42737
1.8
NA
1.0
20.6
27.8
7.2
94

30

9/9/2019
13:18
41.616833 -70.42497
3.1
2.3
1.0
2.6
21.0
20.7
28.0
28.0
7.3
7.3
96.7
95.6

31

9/9/2019
14:04
41.624 -70.42278
2.6
1.9
0.5
2.0
21.7
21.6
27.0
26.9
7.8
7.8
103.4
103.2
4

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Results and Discussion
Water quality data from surveyed stations (Table 1) indicate generally well-mixed, euphotic
conditions. Surface salinities of 22.7-28.6 ppt reflect significant tidal flushing relative to
freshwater input. The freshwater fraction in the estuary was limited to between 5 and 25%, given
that offshore salinity measured at the Waquoit Bay National Estuarine Research Reserve was
30.2 ppt during the survey. Salinity in the Three Bays increased linearly from north to south as
did water clarity (Figures 2-3). Secchi depth (s.d.) measurements ranged from 1 to 2.3m where
they could be made; the disk was visible on the bottom at 8 of 25 sites despite the water level
being above mean tide level (MTL) when most stations were sampled. Assuming the measured
Secchi depth was a lower limit when the disk was visible on the bottom, the minimum light
fraction (assuming ^=1.45/s.d., Batiuk et al., 1992) reaching sediments at those sites at MTL
was 31 to 37%. At sites where the Secchi disk was not visible on the bottom, the light fraction
reaching sediments at MTL was 2-40% (mean=18%). Across all stations the median light
fraction reaching sediments at MTL was 24%.
lat
-70.43 -70.40
' ¦ J, ¦ ¦
!_j!	*
*	•—
*
* •**
Ion
. *
~ * ~
* * * *
** *
I I I I I I I
41.610 41.630
~ * *
20.5 21.5 22.5
1 1 1
*** * *
6.0 7.5 9.0
		
V*
. t •
*•* # A
7T-
v
secchi
temp
* \ • •
salinity
» *
. .r
• -
A .
I II II II	I I I II I
1.0 1.6 2.2	23 25 27
• shallow • deep

( . frf
" < *

*• »• •"
. * .
DO
+ * * •
—^
• A

~ * * i
, t .
DO%sat
i i i i r
80 100 120
Figure 2. Scatterplots of water quality parameters at sampled stations. Shallow measurements
(0.5-lm) are plotted above the diagonal, deep measurements (2-3m) are plotted below it. Units
for each parameter correspond to those given in Table 1.
Vertical differences in salinity, temperature, and density were minimal; maximum absolute
values were S: 1.6 ppt, T: 0.9°C, Aat: 1.2. Vertical differences (shallow-deep) in dissolved
5

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oxygen (DO) varied from -0.2 to 1.6 mg/L (-1.3 to 26%) and were largest at the two deepest
sites, both in North Bay. The minimum observed DO was 6.1 mg/L; actual minimum values
likely occurred overnight, however, not during the day when these measurements were made
DO saturation ranged from 80-127% (median: 105%). Supersaturation of oxygen at shallow
water sites (0.5-lm), likely due to in large part to benthic primary production, was observed
throughout the northern subembayments and West Bay.
Salinity (ppt)
22-24
124-26
26-28
28-30
o0*

Leaflet | © OpenStreetMap contributors © CARTO
Secchi
depth (m)
1.0-1.5
¦ 1.5-2.0
12.0-2.5
NA
O
o°
p
° o%o
: V
Leaflet | © OpenStreetMap contributors © CARTO
DO%sat
80-90
90-100
1100-110
110-120
120-130
O
O

o

o
o
• ®
o°*
Leaflet | © OpenStreetMap contributors © CARTO
Figure 3. Maps of selected water quality measures. The salinity and dissolved oxygen saturation
(IX)'w) maps were made with the shallow (0.5-lm) observations, which were made at all
stations. Secchi depths shown in gray indicate that the disk could be seen on the bottom. Fixed
stations are outlined in black.
Grabbed sediments were typically black or brown with significant fines. More than half of the
samples had a combined silt and clay (i.e., fines) content greater than 60% (see Table 2). The
balance was largely sand, with minimal gravels (max: 5.2% of sample). The sediment had a
sulfidic odor at 9 out of 25 sites with no clear spatial pattern (Figure 4). The total organic carbon
(TOC) content in sediments, which varied inversely with salinity, ranged from <1% to 9.5%
(median: 2.8%). These levels are considered intermediate (>1% TOC) and high (>3.5% TOC) at
76% and 32% of stations, respectively, compared with sediments globally (Flyland et al., 2005).
TOC enrichment can also be assessed using a region-specific relationship with grain size (see
Pelletier et al., 2011, Virginian province). Measured TOC was higher than predicted by this
relationship at most stations (24 of 25), by a factor of 2.6, on average.
6

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Fines (%)
0-20
20-40
B 40-60
60-80
80-100

° %»
o
*o~°°
Leaflet | © OpenStreetMap contributors © CARTO
TOC (%)
0.0-1.0
1.0-3.5
¦ 3.5-5.0
| 5.0-10.0
O
O
o
o <%o

o
Leaflet | © OpenStreetMap contributors © CARTO
Sulfidic odor
No
| Yes	O
I 1 km	|
| 3000 ft
o
° o°*o»
o
•	% *o
° ^	O
P	o
Leaflet | © OpenStreetMap contributors © CARTO
Figure 4. Maps of selected sediment properties: percent fines (silt and clay), total organic carbon
(TOC) content, and field-noted sulfidic odor. Fixed stations are outlined in black.
Density
(1000s/m2>
0-10
10-20
20-30
30-40
40-50
I
O

o
o
o
O
• *;0°

o
Leaflet | © OpenStreetMap contributors © CARTO
Number
of species
0-10
| 10-20
120-30
30-40
40-50
O
O
o0^
o
• oV
$ ^
o
o
Leaflet | © OpenStreetMap contributors © CARTO
M-AMBI
condition

| Bad


Poor

Moderate


Good

¦
High

o
o
c #
o
o
g %
o
Leaflet | © OpenStreetMap contributors © CARTO
Figure 5. Maps of benthic infaunal density, number of species, and M-AMBI condition. Fixed
stations are outlined in black. M-AMBI was not calculated at the stations shown in gray due to
overrepresentation of individuals that could not be assigned to an ecological group. M-AMBI
scores were assigned to condition classes for US coastal waters as follows (from Pelletier et al.,
2018): Bad (<0.2), Poor (0.2-0.39), Moderate (0.39-0.53), Good (0.53-0.77), High (>0.77).
Locations where habitat condition has worsened since the prior survey are indicated with a
double outline.
7

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Table 2. Sediment properties at surveyed stations. ND = not detected.
Subembayment
Station
Gravel
Sand
Fines
TOC
Sulfidic


(%)
(%)
(%)
(%)
odor
Warren's Cove
1
ND
25.6
74.4
9.5
Yes
Prince Cove
2
ND
12.5
87.5
6.4
No

3
ND
25.1
74.9
6.7
Yes
North Bay
4
ND
5.3
94.7
5.3
No

5
ND
7.2
92.8
4.9
No

6
ND
24.2
75.8
3.8
No
West Bay
7
ND
74.3
25.7
2.7
Yes

8
ND
94.1
5.9
0.3
No
Eel River
9
1.6
28.8
69.6
3.4
No
Cotuit Bay
10
ND
65.6
34.4
1.6
No

11
ND
81.7
18.3
1.2
No

12
0.6
95.6
3.8
0.3
No
Prince Cove
13
1.4
27.4
71.2
5.8
Yes
Warren's Cove
15
0.4
91.7
7.9
1.5
No
North Bay
17
ND
56.3
43.7
3.1
Yes

18
ND
14.4
85.6
4.3
Yes

19
ND
60.9
39.1
1.0
No
West Bay
22
ND
39.7
60.3
3.3
No

23
5.2
89.5
5.3
0.2
No

24
ND
83.3
16.7
0.8
No

25
ND
13.9
86.1
3.3
Yes
Cotuit Bay
28
ND
20.7
79.3
2.4
No

29
0.2
88.6
11.2
0.5
No

30
0.4
51.6
48
1.7
Yes

31
0.6
31.4
68
2.8
Yes
A few of the sediment grabs recovered macroalgae, but no submerged vascular plants (including
eel grass) were encountered. Laboratory determined infaunal counts ranged from 12 to 1996
individuals per 0.04 m2 grab, or an extrapolated density of 300 to 49,900 organisms per m2
The number of species varied between 2 and 50 per grab, with Shannon-Weiner diversity
ranging from 0.1 to 2.3 and Pielou's evenness scores of 0.15-0.83. M-AMBI scores ranged from
0.06-1 (mean: 0.33), or nearly the full spectrum (0-1). See Table 3 for values of each community
metric by station and Figure 5 for maps of selected metrics. M-AMBI scores were significantly
(p < 0.05) and negatively correlated with latitude, water temperature and dissolved oxygen, as
well as the amount of fine materials and organic carbon in sediments. M-AMBI scores were
significantly and positively correlated with sand content and Secchi depth. Several of these
variables were themselves correlated. Pairwise correlation coefficients and their statistical
significance are plotted in Figure 6.
8

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Table 3. Benthic infaunal community metrics and overall habitat condition. Type F = Fixed,
S = Sentinel, R = Randomized. A value of NA for M-AMBI score and habitat condition indicates
that the index was not calculated due to overrepresentation (>50%) of individuals in a sample
that could not be assigned to an ecological group.
Subembayment Station Type Density Number of Shannon Pielou's M-AMBI Habitat



(per m2)
species
(S)
diversity
(H') "
evenness
(E)
score
condition
Cotuit Bay
10
F
12125
32
1.87
0.54
0.72
good

11
F
7400
26
2.26
0.69
0.60
good

12
S
49900
48
2.06
0.53
NA
NA

28
R
11925
22
1.74
0.56
0.50
moderate

29
R
10375
41
2.34
0.63
NA
NA

30
R
300
2
0.56
0.81
NA
NA

31
R
2025
14
1.71
0.65
0.33
poor
mean


13436
26
1.79
0.63
0.54
good
Eel River
9
F
3500
6
0.38
0.21
0.14
bad
North Bay
4
F
525
3
0.83
0.76
0.10
bad

5
F
20900
21
1.48
0.49
0.48
moderate

6
F
1200
2
0.1
0.15
0.06
bad

17
R
3600
18
1.39
0.48
0.41
moderate

18
R
625
5
1.33
0.83
0.15
bad

19
R
2175
8
1.54
0.74
0.22
poor
mean


4838
10
1.11
0.58
0.24
poor
Prince Cove
2
F
850
1
1.29
0.66
0.18
bad

3
F
2975
1
0.64
0.33
0.17
bad

13
R
2525
4
0.38
0.28
0.10
bad
mean


2117
6
0.77
0.42
0.15
bad
Warren's Cove
1
F
3100
10
1.6
0.7
0.25
poor

15
R
950
9
1.46
0.66
NA
NA
mean


2025
9.5
1.53
0.68
0.25
poor
West Bay
7
F
400
6
1.12
0.63
0.17
bad

8
F
11450
29
1.65
0.49
0.64
good

22
R
4300
10
1.15
0.5
0.24
poor

23
R
34775
50
2.11
0.54
1.00
high

24
R
12100
18
1.73
0.6
0.42
moderate

25
R
2125
11
1.33
0.56
0.26
poor
mean


10858
21
1.52
0.55
0.46
moderate
9

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The 2019 infaunal community data suggest that benthic habitat condition has not improved since
the prior survey (Howes et al., 2006) and there is some evidence of further degradation. Among
the set of revisited, or fixed, stations, the recent survey found a lower density and diversity of
organisms, and lower species evenness (Table 4). Note that densities have been extrapolated to a
common area (1 m2) to facilitate comparison; the prior survey reported data as the average of
three grabs per station, each of which sampled a larger surface area (0.0625 m2). Changes in the
total number of species sampled were less definitive, with a higher mean and lower median
among fixed stations compared with the prior survey. Overall change in habitat condition over
time was assessed by comparing condition classes used in the past under MEP and those
associated with US M-AMBI (see Table 4). Habitat condition shows little to no change at 9 of
the 11 fixed stations, while conditions at the remaining 2 have worsened.
Table 4. Benthic infaunal community metrics and change in overall habitat condition over time.
S = number of species, H' = Shannon diversity, E = Pielou's evenness. Change in overall
condition was assessed by making equivalences between the condition classes used in the prior
MEP survey ("Significant Impairment (SI)", "Moderate Impairment (MI)", "Healthy habitat
(H)"), and the respective groupings of M-AMBI classes ("Bad/Poor", "Moderate",
"Good/High").

prior MEP survey (early 2000s)
current survey (2019)

Station &
Density
£
H'
jj
Infaunal
Density
s
H'
jj
M-AMBI
Habitat
Change in
overall
condition
subembayment
(perm 2 )


Indicators
(perm 2 )


score
condition
1. Warren's Cove
112
4.7
2.01
0.86
SI
3100
10
1.60
0.70
0.25
poor
no change
2. Prince Cove
800
9.3
2.49
0.82
SI
850
7
1.29
0.66
0.18
bad
no change
3. Prince Cove
688
4.7
0.9
0.43
SI
2975
7
0.64
0.33
0.17
bad
no change
4. North Bay
176
4.7
1.9
0.85
SI
525
3
0.83
0.76
0.11
bad
no change
5. North Bay
13136
14.3
1.35
0.36
MI
20900
21
1.48
0.49
0.48
moderate
no change
6. North Bay
112
3
1.91
0.92
SI
1200
2
0.10
0.15
0.06
bad
no change
7. West Bay
8016
17.3
3.39
0.82
H/MI
400
6
1.12
0.63
0.18
bad
worse
8. West Bay
30320
26.3
2.02
0.42
H/MI
11450
29
1.65
0.49
0.65
good
no change
9. Eel River
7456
11
2.28
0.67
MI
3500
6
0.38
0.21
0.14
bad
worse
10. CotuitBay
8560
16.3
2.99
0.75
H/MI
12125
32
1.87
0.54
0.71
good
no change
11. Cotuit Bay
3728
16
3.26
0.82
H
7400
26
2.26
0.69
0.60
good
no change
mean
6646
12
225
0.70

5857
14
1.20
0.51
0.32
poor

median
3728
11
2.02
0.82

3100
7
1.29
0.54
0.18
bad

The longitudinal comparison of habitat condition is less than ideal for several reasons. The dates
of the prior benthic survey and coordinates of the surveyed locations were not published. Here
fixed station locations were georeferenced from the published map (Howes et al., 2006). There
are some differences in sampling methods (discussed above) and assessment methodology. The
11
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summary ratings for habitat condition used in the prior survey (see "Infaunal Indicators" in
Table 4) were not described in enough quantitative detail to apply to the current one. The
correspondence over time in overall assessed condition at most stations, however, suggests a
signal that is stronger than these sources of noise. The probability of finding no overall
improvement in benthic habitat at any of the fixed stations is less than 1% if 7 of the 11 sites
could have shown improvement due to random chance. Conversely, 6 of the revisited sites had
the potential to get worse, and 2 did (see Figure 7), indicating that the classification is at least
subject to change. The direction of change (worse) is consistent with expectations, given the
limited scope of contaminant mitigation measures. The relative influence of nutrient loading
versus other factors, like climate, on changes in benthic condition in Three Bays has not been
evaluated here.
Prior survey
(early 2000s)
¦	HorH/MI
Ml
¦	si
Revisited sites
(2019)
| Good
Moderate
| Poor or Bad
Leaflet ® OpenStreetMap contributors © CARTO	Leaflet I © OpenStreetMap contributors © CARTO
Figure 7. Comparison of habitat condition assessments from prior and current benthic surveys.
Colors reflect equivalences that were made between the condition classes used in the prior MEP
survey ("Significant Impairment (SI)", "Moderate Impairment (MI)", "Healthy habitat (H)"), and
the respective groupings of US M-AMBI classes ("Bad/Poor", "Moderate", "Good/High"). None
of the revisited sites had "High" quality benthic habitat condition in the 2019 survey, though one
of the randomized stations did (see Figure 5).
The current survey also included locations that were not previously assessed, yielding a more
nuanced perspective on current conditions. The TMDL sentinel station has regulatory
significance; its location was chosen such that meeting water quality goals there would indicate
an expectation of goal attainment throughout the Three Bays system (MassDEP, 2007).
12
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Interestingly, the sentinel had the highest density of organisms and second highest number of
species of any station. An overabundance of organisms that could not be assigned to an
ecological group precluded determination of the M-AMBI score and condition class at the
sentinel and three of the randomized stations. The latter were added to the 2019 survey to avoid
selection biases that may be present in the fixed set of stations. Randomized stations had a higher
average density, species count, diversity, and M-AMBI score than fixed stations. M-AMBI
scores for randomized stations also showed greater spatial variability (see Figure 5), as was
previously discussed. Both fixed stations in Cotuit Bay were classified as good, for example,
whereas two nearby randomized locations were assessed to be in moderate to poor condition.
Use of an established benthic index supports analysis of habitat condition in Three Bays as well
as comparison with other estuaries. The M-AMBI score provides a quantitative summary of
infaunal community data that can be used to standardize assessment. Widely used around the
world (see, e.g., Sigovini et al., 2013), M-AMBI is also currently being applied to other estuaries
on Cape Cod as part of a MEP review of benthic monitoring procedures. In nearby West
Falmouth Harbor, a recent benthic survey applied M-AMBI and showed that habitat conditions
had remained largely unchanged over a 16-year period at some of the revisited stations (Sweeny
and Rutecki, 2020). Other stations showed improvements, including re-established eelgrass beds,
that may be related to upgraded nutrient control at a nearby wastewater treatment plant. Pending
results for Pleasant Bay and Wellfleet Harbor may be considered alongside these to learn how
estuarine habitats across Cape Cod respond to differences in nutrient loading and interventions to
manage it.
Acknowledgments
The authors appreciate assistance with planning and executing the 2019 benthic survey provided
by Marty Chintala and Zee Crocker, guidance on the application of M-AMBI provided by Peg
Pelletier and Giancarlo Cicchetti. They are grateful for comments from internal reviewers that
have improved this data summary. The work was funded by the US EPA Office of Research and
Development's (ORD) Safe and Sustainable Water Resources program. It is identified by
ORD-041256.
Disclaimer
This document and the associated supplemental materials include contributions from The U.S.
Environmental Protection Agency (EPA) and individuals outside the United States Government.
EPA, through its Office of Research and Development, funded this effort and managed the
contracted analyses described herein. This data summary has been subjected to the Agency's
review and has been approved for publication. Mention of trade names, products, or services
does not convey EPA approval, endorsement, or recommendation. The views expressed in this
summary are those of the author(s) and do not necessarily represent the views or the policies of
the U.S. Environmental Protection Agency.
13
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References
Batiuk R. A., Orth R. J., Moore K. A., Dennison W. C., Stevenson J. C., Staver L. W., Carter V.,
Rybicki N. B., Hickman R. E., Kollar S., Bieber S., Heasly P. 1992. Chesapeake Bay submerged
aquatic vegetation habitat requirements and restoration targets: A technical synthesis. US
Environmental Protection Agency, Annapolis, MD.
Boija A., Franco J., Perez, V. 2000. A marine biotic index to establish the ecological quality of
soft-bottom benthos within European estuarine and coastal environments. Mar. Pollut. Bull. 40,
1100-1114.
Gillett D.J., Weisberg S. B., Grayson T., Hamilton A., Hansen V., Leppo E. W., Pelletier
M. C., Borja A., Cadien D., Dauer D., Diaz R., Dutch M., Hyland J. L., Kellogg M.,
Larsen P. F., Levinton J. S., Llanso R., Lovell L. L., Montagna P. A., Pasko D.,
Phillips C. A., Rakocinski C., Ranasinghe J. A., Sanger D. M., Teixeira H., VanDolah
R. F., Velarde R. G., Welch K. I. 2015. Effect of ecological group classification
schemes on performance of the AMBI benthic index in US coastal waters. Ecol. Indie.
50, 99-107.
Howes B., Kelley S.W., Ramsey J. S., Samimy R., Schlezinger D., Eichner E. 2006. Linked
Watershed-Embayment Model to Determine Critical Nitrogen Loading Thresholds for Three
Bays, Barnstable, Massachusetts. Massachusetts Estuaries Project, Massachusetts
Department of Environmental Protection. Boston, MA.
Hyland J., Balthis L., Karakassis I., Magni P., Petrov A., Shine J., Vestergaard O., Warwick R.
2005. Organic carbon content of sediments as an indicator of stress in marine benthos. Mar.
Ecol. Prog Ser. 295: 91-103.
Kelley D. and Richards C. 2020. oce: Analysis of Oceanographic Data. R package version 1.2-0.
Kincaid T. M., Olsen, A. R. 2019, Weber M. H. spsurvey. Spatial Survey Design and Analysis. R
package version 4.1.0.
MassDEP. 2007. Three Bays System Total Maximum Daily Loads for Total Nitrogen.
Massachusetts Department of Environmental Protection. Report # 96-TMDL-10 Control # 242.0.
Muxika I., Borja A., Bald J. 2007. Using historical data, expert judgement and multivariate
analysis in assessing reference conditions and benthic ecological status, according
to the European water framework directive. Mar. Poll. Bull. 55, 16-29.
Oksanen J., Blanchet., F.G., Friendly M., Kindt R., Legendre P., McGlinn D., Minchin P.R.,
O'Hara R. B., Simpson G. L., Solymos P., Henry M., Stevens H., Szoecs E., Wagner H. 2019.
vegan: Community Ecology Package. R package version 2.5-6.
Pelletier M. C., Campbell D. E., Ho K. T., Burgess R. M., Audette C. T., Detenbeck N. E. 2011.
Can sediment total organic carbon and grain size be used to diagnose organic enrichment in
estuaries? Environ. Toxicol. Chem. 30(3), 538-547.
14
-------
Pelletier M. C., Gillett D. J., Hamilton A., Grayson T., Hansen V., Leppo E. W., Weisberg S. B.,
Boija A. 2018. Adaptation and application of multivariate AMBI (M-AMBI) in US coastal
waters. Ecol. Indie. 89, 818-827.
Sigovini M., Keppel E., Tagliapietra D. 2013. M-AMBI revisited: looking inside a widely-used
benthic index. Hydrobiologia 111, 41-51.
Stevens D. L., Olsen A. R. 2004. Spatially Balanced Sampling of Natural Resources, J. Am. Stat.
Assoc., 99:465, 262-278, DOI: 10.1198/016214504000000250
Sweeny M., Rutecki D. A. 2020. Benthic monitoring report: West Falmouth Harbor 2019.
Prepared for the MassDEP Massachusetts Estuaries Project. June 2020. 49pp.
US EPA. (U.S. Environmental Protection Agency). 1995. Environmental Monitoring and
Assessment Program (EMAP): Laboratory Methods Manual - Estuaries, Volume 1: Biological
and Physical Analyses. United States Environmental Protection Agency, Office of Research and
Development, Narragansett, RI. EPA/620/R-95/008.
US EPA (U.S. Environmental Protection Agency). 2015. National Coastal Condition
Assessment: Field Operations Manual. Version 1.0, May 2015. EPA-841-R-14-007. U.S.
Environmental Protection Agency, Office of Water, Washington, DC. 166 pp.
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