Great Lakes Restoration Initiative: Reassessment of Wildlife Reproduction and Health
Impairments in the Saginaw Bay and River Raisin Areas of Concern and Grand Traverse Bay
K. A. Grasman, D. Bouma, A. Mahn, J. Van Bruggen, Department of Biology, Calvin College, Grand Rapids, MI
J. Moore, L. Williams, U.S. Fish and Wildlife Service, Region 3, East Lansing, MI
A) Herring Gull
Introduction
Persistent organochlorines such as polychlorinated biphenyls (PCBs) and chlorinated dioxins have
been suspected to be causing reproductive problems in the Great Lakes since the late 1950s, which
was later supported in the early 1970’s when scientists noted increased mortality and deformities
such as crossed bills in fish-eating birds living in the lakes (Grasman et al., 1998). PCBs have been
shown to cause chick edema disease in chickens, and there is also strong evidence to suggest that
they cause an equivalent illness known as GLEMEDS (Great Lakes Embryo Mortality, Edema, and
Deformities Syndrome) in fish-eating birds (Gilbertson et al., 1991). Another organochlorine,
dichlorodiphenyltrichlorethane (DDT) and its breakdown product dichlorodiphenyldichlorethylene
(DDE) are known to induce eggshell thinning in Great Lakes waterbirds (Grasman et al., 1998). In
addition, organochlorines have been shown to cause decreased immune function in Great Lakes
fish-eating birds. In previous studies, scientists have shown that birds nesting in areas with higher
concentrations of organochlorines than those nesting at reference sites had suppressed t-cell
mediated immunity (Grasman et al., 1996).
In 1972, the Great Lakes Water Quality Agreement was created between the United States and
Canada; since then, it has expanded to include 14 Beneficial Use Impairments to measure the
health of the Great Lakes (Environment Canada, 2013). These guidelines are used in designating
and maintaining Areas of Concern (AOCs) which are sites with severe degradation or high
contamination in the Great Lakes. The current study monitors the River Raisin and Saginaw Bay
AOCs as part of the US Fish and Wildlife Service’s efforts under the broader Great Lakes
Restoration Initiative and reassesses two Beneficial Use Impairments: 1) Degradation of fish and
wildlife populations and 2) Bird or animal deformities or reproduction problems. In 2014, Bellow
Island in Grand Traverse Bay was added to this study due to an unusual mixture of contaminants.
This site has a high level of dioxin toxic equivalents (TEQs), a relatively low concentration of
PCBs, and a high level of residual DDE. The study compared the data obtained from these polluted
sites to reference sites at the Pipe Island Twins, Chantry Island, and Two Tree Island in Lake Huron
(Figure 1).
This study reexamined the reproduction, growth, and immunological responses of colonial
waterbirds (herring gulls, Caspian terns, and black-crowned night herons) in the Great Lakes in an
attempt to evaluate the effects of these pollutants. To this end, the study employed the endpoints of
embryonic mortality, deformities, reproductive success, chick growth, T cell-mediated immune
function (phytohemagglutanin (PHA) skin test), and antibody response (sheep red blood cell
(SRBC) test). The PHA skin test and the SRBC test are well known measures of immune function
in avian test subjects (Grasman, 2002).These tests are also known to be ecologically relevant as
studies have shown that wild songbirds demonstrating a high immune response correlates with
higher survival and establishment of new populations (Moeller and Saino 2004; Moeller and
Cassey 2004).
Materials and Methods
Study Sites (See Fig. 1):
Reference
Pipe Island Twins (gulls 2011-15)
Two Tree Island (terns 2011-13)
Chantry Island (herons 2010)
River Raisin AOC
Monroe (gulls 2010-2015)
Grand Traverse Bay
Bellow Island (2014-15)
Saginaw Bay AOC
Confined Disposal Facility (CDF)
(gulls 2010-15, terns 2010-14, herons 2010)
Little Charity Island (gulls 2010,
2012-15 and terns 2015)
Charity Reef (terns 2010, 2014)
Embryonic Nonviability
• Three egg herring gull nests were marked during laying. During mid incubation (21 days),
embryonic viability was assessed using an Avitronics Digital Egg Monitor (Cornwall England),
which measures the embryo's heart rate. Nonviable eggs were opened to determine infertility,
stage of development, and deformities.
Growth and Reproductive Success
• Body mass and size measurements were taken at 3 and 4 weeks of age in order to measure the
rates of growth (or degradation) of chicks in the colonies.
• Chick productivity in enclosed nests was measured by counting the number of chicks alive at 3
weeks divided by the number of nests.
Immune Function
• T-cell mediated immune test - To test the T cell-mediated immunity, 3 week gulls and terns or 2
week herons were injected with PHA (a lymphocyte activator) in phosphate buffered saline
(PBS) in one wing web and a placebo of PBS in the other wing web. Wing web thickness
measurements were taken with pressure sensitive calipers before and one day after injections.
The PHA stimulation index was calculated as the difference between the change in the PHA
wing web thickness and the change in the control wing web thickness.
• Antibody response test- A sheep red blood cell (SRBC) suspension was injected into the wing
of 3 week old chicks to stimulate antibody production. The hemagglutination assay was
performed with plasma from 4 week old chicks by serial dilutions with addition of SRBCs.
Total antibody response titer value was determined as the highest dilution where agglutination
formed between the antigen and antibody. IgG antibody response was assessed by denaturing
IgM antibodies with 2-mercaptoethanol prior to the addition of SRBCs.
B) Caspian Turn
OLD
OLD
Areas of Concern
(A)
Other Polluted Sites
(B)
(N/A)
( N/A )
(N/A)(N/A)(N/A)(N/A)
(N/A)
(N/A)
(N/A) (N/A) (N/A)
SITE
Figure 1. Colonial waterbird colonies studied in Great Lakes
AOCs, other contaminates sites, and reference areas for this
assessment of health and reproduction in fish-eating birds during
2010-15.
A) Herring Gull
Figure 2. Embryonic nonviability in herring gulls at the
Saginaw Bay and River Raisin AOCs and Grand Traverse
Bay during 2010, 2012-14. Numbers on bars are egg
totals. “Undetermined” categorizes infertile or failed eggs
that were difficult to distinguish due to addling or rot.
Two-way ANOVA
site: p < 0.0001
year: p < 0.0001
B) Caspian Tern
Figure 3. Crossed-billed herring gull chicks (3 weeks old) found at
the Monroe colony in the River Raisin AOC during 2012 (A) and
2013 (B).
A) Herring Gull
Two-way ANOVA
site: p = 0.0262
year: p =0.1342
Figure 5. Mean growth in body mass between 3 and 4 weeks of age in pre-fledgling fish-eating birds at Great Lakes AOCs, Grand
Traverse Bay, and reference sites in 2010-15. Normal average growth rates are 4-18 g/day for Caspian terns and 14-20 g/day for
herring gulls (Grasman et al. 1996). Numbers on bars are sample sizes. Error bars indicate standard error.
Two-way ANOVA
site: p < 0.0001
year: p < 0.0015
B) Caspian Tern
Figure 4. Reproductive productivity (number of 3 week old chicks surviving per nest) at AOCs, Grand Traverse
Bay, and reference sites in 2010-15. For herring gulls, productivity of 0.75 chicks/nest or greater is necessary to
maintain a stable population (Kadlec and Drury 1968).
Two-way ANOVA
site: p <0.0001
year: p < 0.094
OLD
Reproductive Success
Reproductive success at AOCs was variable across years, but consistently strong at
reference colonies. Gull populations that produce at least 0.75 chicks/nest are usually
stable (Kadlec & Drury 1968). In 2010-14, gulls in the Saginaw Bay AOC reproduced at
sustainable rates (0.75-2.18 chicks/nest) (Fig. 4A). Productivity at the Monroe colony
fluctuated between the years, with good productivity in 3 out of 5 years, but complete
failure in 2010. Gulls in Grand Traverse Bay had a good productivity, comparable to
that of the reference site. Caspian terns on Charity R. had low productivity in 2010 (0.3
chicks/nest), followed by good productivity in 2014 (0.95 chicks/nest). Higher
productivities were observed on the SB CDF (Fig. 4B) until 2014, when productivity
fell to 0.2 chicks/nest. Previous studies of this state threatened species have shown low
recruitment of young terns raised in Saginaw Bay and other highly contaminated
ecosystems (Mora et al. 1993). In 2014, the reference colony nests were lost due to
flooding from and higher water levels and weather.
OLD
T-test
p < 0.05
Figure 6. Mean phytohemagglutinin (PHA) stimulation index, a measure of the T cell-mediated immunity, in pre-fledging
fish-eating birds at AOCs, Grand Traverse Bay, and reference sites in 2010-15. Bars sharing the same letter were not
statistically different by Tukey’s test (p<0.05). Numbers on bars indicate sample sizes. Error bars indicate the standard error.
Results and Discussion
Embryonic Nonviability and Deformities
Embryonic nonviabiliy rates in late incubation herring gull eggs were elevated at
Monroe in the River Raisin AOC (multi-year mean of 7.7%), SB CDF (4.5%) and L.
Charity Is. (7.4%) in the Saginaw Bay AOC, and Bellow Is. in Grand Traverse Bay
(5.6%) compared to reference sites (2.6% at Pipe Is. Twins, <2% at other reference sites
in studies by Craig Herbert, Environment Canada, personal communication; Fig. 2).
Infertility was the primary cause of nonviability at the Pipe Is. Twins reference site and
contributed substantially to nonviability at all AOC colonies and Grand Traverse Bay.
Embryonic death (failed development) also accounted for a sizable portion of total
nonviability at Monroe in the River Raisin AOC and L. Charity Is. in the Saginaw Bay
AOC. Crossed-bill deformities were found in two herring gull chicks at the River Raisin
AOC during 2012 (7mm offset of the upper and lower bill) and 2013 (4mm offset; Fig
3). Embryonic nonviability and deformities such as crossed bills are characteristic of
GLEMEDS and are associated with exposure to PCBs and dioxins (Gilbertson et al.
1991, Grasman et al. 1998).
C) Black-crowned
Night Heron
Growth
Growth in herring gull chicks varied by year and site (Two-way ANOVA for site and year:
p<0.0001) (figure ?). Gulls at Monroe and Little Charity Island had lower growth than the
reference sites during 2011-2012 and 2014 and the overall mean growth for Little Charity was
significantly lower than Pipe Island Twins (Tukey p<0.05). In 2013 and 2015 the growth
trends differed in that the mean growth for the reference site was lower than the other sites
with the exception of Bellow Island in 2015. Bellow Island had lower growth than the
reference site in both 2014 and 2015 (Figure 5A).
The normal mean growth rate for herring gulls is 14-20 g/day (Grasman et al., 1996). In our
study, normal growth rates were achieved in 2010 at the CDF, in 2011 and 2014 at the Pipe
Island Twins, and in 2013 and 2015 at Monroe, CDF, and Little Charity; weight loss was seen
at the CDF in 2014 and at Little Charity Island in 2012 and 2014 (Figure 5A).
The normal mean growth rate for Caspian terns is 4.0-18.0 g/day (Grasman et al., 1996) In
2011 through 2013 at the reference site, terns ranged from 3.6-10.8 g/day in mean growth rate.
However, at the CDF between 2010-2012 and in 2014, terns only averaged between 2.0-5.0
g/day for mean growth rate (Figure 5B).
T Cell-Mediated Immunity
Herring gulls and Caspian terns had a lower T-cell mediated immune response at all AOC sites
as well as at Grand Traverse Bay compared to the reference sites at Pipe Island Twins and Two
Tree Island (Two-way ANOVA p<0.0001) (Figure ?). The mean PHA skin response of gulls
was 53-57% lower at AOCs compared to the Pipe Island Twins. Gulls in Grand Traverse Bay
had a 50% lower mean PHA skin response than the reference site at Pipe Island Twins (Tukey
p<0.05). Caspian terns at AOCs had a 46-50% lower response as compared to Two Tree
Island. Black-crowned night herons on the CDF showed a 39% lower T-cell response
compared to herons at the Chantry Island reference site (Figure 6)
Antibody Titer of Plasma Samples
Herring gulls at the AOC sites and Grand Traverse Bay had significantly lower total antibody
response and lower IgG response than the herring gulls at the Pipe Island Twins reference site
(Two-way ANOVA p<0.0001, Tukey p<0.05). At AOCs mean titer values for total antibodies
(4.3-4.9) and IgG responses (2.1-3.0) were significantly lower than values for the reference
site (Total: 5.6, IgG: 4.1). For Caspian Terns, total and IgG responses differed by year and no
significant differences between site were found (Two-way ANOVA Total: p =0.09, IgG:
p=0.066) (Figure 7).
Two-way
ANOVA
Site: p<0.001
Year: p=
0.0015
Figure 7. Mean total and IgG antibody titer in pre-fledging
herring gulls at AOCs, Grand Traverse Bay, and reference site
in 2010-2015. Bars sharing the same case and letter were not
statistically different by Tukey’s test (p<0.05). Numbers on bars
indicate sample sizes. Error bars indicate the standard error.
Conclusions
Herring gulls, Caspian terns, and black-crowned night herons in the Saginaw Bay
AOC exhibited adverse health effects, including a decreased immune responses and
(or) decreased productivity, which were consistent with past studies:
• Embryonic non viability was higher at the AOCs
• Chick growth was significantly less at Little Charity
• AOC has less reproductive success
• Suppressed T- cell mediated immunity in herring gulls, Caspian terns, and blackcrowned night herons
• Suppressed total antibody and IgG response in herring gulls
Herring gulls at the River Raisin AOC showed impairments in immune responses
and reproductive success, consistent with past studies
• Suppressed T cell-mediated immune response
• Suppressed total antibody and IgG response
• Decreased growth rates for chicks, excluding 2013 and 2015
• Higher than normal embryonic nonviability
• Complete reproductive failure in 2010 and decreased productivity in 2012
Herring gulls at Grand Traverse Bay, a site with high concentrations of TEQs and
DDE, showed impairments in immune responses and growth
• Suppressed T cell-mediated immune response
• Suppressed total antibody response
• Low growth rate between three and four weeks of age
Acknowledgements
This project was funded by the Great Lakes Restoration Initiative Funding for Area of Concern and Remediation and
Restoration of Contaminated Sediments through the U. S. Fish and Wildlife Service, Region 3. Ted Ludwig, Jim
Ludwig, Dave Best, and Therese Best provided logistical support in the field while Calvin VanOpstall provided
contributed greatly to lab work. Research stipends from Calvin College were provided to Jenna Van Bruggen by the Star
Fellowship and to Alaina Mahn by the Dornbush De Vries Family Student Research Fellowship.
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