Journal of Student Research (2013)
Volume 2, Issue 1: pp.
43-47
Research Article
a. Department of Biology, Shippensburg University, Shippensburg, PA 17257
43
Prevalence of Strongyloides robustus in Tree Squirrels
(Sciuridae) in South-Central Pennsylvania and Potential
Impacts for the Endangered Northern Flying Squirrel,
Glaucomys sabrinus
Jessica L.
Espenshade
a
and Richard L.
Stewart
a
The northern flying squirrel is an endangered species in the state of Pennsylvania.
Many hypotheses potentially explain the
diminished population, with the most supported being parasitic infection.
While infection with the parasite Strongyloides
robustus presents with benign pathology in some squirrel species, like the southern flying squirrel (Glaucomys volans), the
eastern gray squirrel (Sciurus carolinensis) and the red squirrel (Tamiasciurus hudsonicus), S. robustus infection can result in
high mortality in the northern flying squirrel (Glaucomys sabrinus). Past research has focused primarily on the transmission
between northern and southern flying squirrels; however this project aims to better evaluate the risk to northern flying squirrels in
light of the parasite mediated competition hypothesis by surveying the prevalence of S. robustus within other sympatric reservoirs
currently found throughout Pennsylvania.
In order to survey parasite presence, fecal samples from eastern gray, red, and southern
flying squirrels were obtained from nest boxes, road kills and hunting.
Strongyloides robustus was present in 30% of the 10
southern flying squirrel samples.
The prevalence of S. robustus was 77.
3% in the 22 road-killed and hunter-killed eastern gray
squirrels.
The single hunter-killed red squirrel examined in this study demonstrated S. robustus infestation.
The results in this
study area supported the hypothesis of parasite mediated competition on a broader scale because of the high infection rate in
potential sympatric reservoirs.
Surveillance of S. robustus in localized populations should occur prior to initiating recovery plans
for the northern flying squirrel and management of this parasite in reservoir populations should be considered.
Keywords: Parasite, Squirrels, Endangered
Introduction
Glaucomys sabrinus, the northern flying squirrel, has a
population size that is considered secure nationally
(Butchkoski and Turner 2010).
However in parts of its range,
it is either a candidate for endangered listing or is already
considered endangered (USFWS 2012).
For example, the
northern flying squirrel is currently on the state endangered
species list in Pennsylvania.
A number of potential factors
may have led to this status; including climate change, habitat
destruction, insect pests, Strongyloides robustus infections,
and urban development (Krichbaum et al.
2010, Title 58,
Mahan et al.
1999).
Northern flying squirrels have unique
habitat preferences and prefer >95 year old mixed coniferous
forests adjacent to a permanent water source.
This habitat
usually also contains a significant number of overstory trees
(including spruce, northern hardwood and other mixed-
coniferous–hardwood forest types), saplings and rock cover
(Mahan et al.
2010 and Smith 2007).
However, northern
flying squirrels have been flexible in their habitat
requirements under pressure (Weigl 2007).
Historically, the range of the northern flying squirrel
covered the majority of Pennsylvania and south into parts of
Tennessee (Burt 1957).
However, by 1995 the documented
range had diminished greatly.
Habitat is being destroyed,
resulting in smaller, fragmented populations of the northern
flying squirrel (Mahan et al.
1999).
Populations are no longer
present in south-central Pennsylvania and the southern U.
S.
populations are greatly isolated on specific mountaintops
(Kurta 1995 and Sparks 2005).
When populations of the northern flying squirrel become
isolated, a number of other, potentially more dangerous,
factors arise.
Small populations exhibit lower genetic
diversity due to increased rates of inbreeding, and inbreeding
has been linked to increased potential for parasitic infection
(Sparks 2005).
Inbreeding causes a lack of genetic fitness and
consequently a lack of immune response (Stevens et al.
1997;
Allendorf and Leary 1986).
Therefore, it is possible that
reductions in genetic diversity due to inbreeding may be
associated with an increased level of parasitism by
Strongyloides robustus.
Strongyloides robustus is one of the most common
helminths in squirrels.
Members of the genus Strongyloides
are roundworms that can cause skin pathology, pulmonary
and/or intestinal distress in host species (Roberts et al.
2009).
Only two out of the 35 different Strongyloides species, S.
robustus and S. papillosus, are known to parasitize squirrels.
For most squirrel species, there are no known deleterious
effects from S. robustus. That being said, these squirrels can
serve as reservoirs for infection.
Historically, S. papillosus
was recorded from eastern gray squirrels in Tennessee by
Reiber and Byrd (1942) but has not since been documented in
the northern United States or Canadian provinces.
Sparks (2005) suggested the hypothesis of parasite
mediated competition for sympatric flying squirrel species.
The parasite mediated competition hypothesis predicts that
when multiple species are hosts to the same parasite, one host
will become more tolerant of the parasite while the others do
not acquire the same resistance (Price et al.
1988).
Krichbaum
Journal of Student Research (2013)
Volume 2, Issue 1: pp.
43-47
Research Article
ISSN: 2167-1907
www.jofsr.
com
44
et al.
(2010) tested this prediction and collected data that
partially supported the parasite mediated hypothesis in
sympatric populations of northern and southern flying
squirrels.
Although no ill effects from S. robustus infection in
southern flying squirrel have been identified; when S.
robustus infects northern flying squirrels, it seems to suppress
the squirrel’s ability to put on winter weight or even maintain
their current weight.
The result of this infection can be fatal
(Wetzel and Weigl 1994).
Focusing research on the transmission between the
northern and southern flying squirrels is limited in its
application.
The vulnerable northern flying squirrel is in
contact with other squirrel species such as the red squirrel, fox
squirrel and the eastern gray squirrel throughout its range.
Little evidence has been published examining the abundance
of this parasite in these other common sympatric squirrel
species.
The exclusion of potentially important information
relating to sources for parasite transmission could be a very
important reason explaining decline in the northern flying
squirrel.
To better understand the abundance and prevalence
of S. robustus in potential reservoir populations, we
conducted a survey for this parasite in squirrel species located
in south-central Pennsylvania using a simplified fecal-float
method of detection.
Our goal was to add information to the
previous records of S. robustus parasitism in order to evaluate
the parasite mediated competition hypothesis on a broader
scale.
Pennsylvania has seven squirrel species (ASM 2012) and
all but the northern flying squirrels have established
populations near Shippensburg (Stewart et al.
2008).
Within
their distributions, many of these squirrel territories overlap
and therefore a high rate of competition between the squirrels
may exist.
Nest sharing could provide the necessary habitat
overlap to inoculate the nest with fecal material and therefore
introduce parasites into the habitat.
If there is a high
prevalence of S. robustus in any squirrel species, it could
potentially reduce or eliminate the northern flying squirrel
populations when the parasite is transmitted between species.
In this survey, three species of squirrels were evaluated
for the presence of S. robustus. These species include: the
eastern gray squirrel (Sciurus carolinensis), the red squirrel
(Tamiasciurus hudsonicus), and the southern flying squirrel
(Glaucomys volans). No information is available concerning
the prevalence of S. robustus throughout most of
Pennsylvania including Franklin and Cumberland counties.
These results will aid in our understanding of the parasite
mediated competition hypothesis providing detail to the
relationship of S. robustus and the declining populations of
the northern flying squirrel.
We hypothesized that S. robustus
will be prevalent in all three squirrel species being tested
which may have resulted in the absence of northern flying
squirrels in the study area.
Materials and Methods
Fecal samples were collected from eastern gray, red and
southern flying squirrels.
The flying squirrel samples were
obtained from 40 previously placed nest boxes on land
adjacent to Pennsylvania State Game Lands 169 near
Newville, PA.
The eastern gray and red squirrel samples were
collected from road kills and hunter-harvested squirrels.
The
locations where these squirrels were found include
Letterkenny Army Depot (Franklin County, PA), and the area
surrounding Newburg, PA and Shippensburg, PA (both
Cumberland County, PA).
Dried fecal samples were taken from the flying squirrel
nest boxes.
The eastern gray and red squirrel fecal samples
were removed directly from the squirrel’s colon.
To
concentrate eggs within the feces, the samples were placed in
test tubes and mixed with a modified Sheather’s sugar
flotation solution (Dryden et al.
2005).
Sheather’s sugar
solution has been successfully used in other experiments to
identify helminthes in squirrels including: Eimeria (Fuller and
Duszynski 1997), Strongyloides robustus and Citellinemia
bifurcatum (Pedder et al.
2009).
Sheather’s solution was prepared with a specific gravity
of 1.27 and a wooden applicator stick was used to mix in the
fecal sample.
Additional Sheather’s solution was added until a
meniscus formed on top of the test tube at which point a
microscope slide was placed on top of the meniscus.
After 5
minutes the slide was removed and inverted before a coverslip
was placed on the area with Sheather’s solution to determine
parasite egg presence.
Slides were observed under 40X
magnification using a student Leica CM E microscope.
The
parasites were identified by being the only strongyloid that
infects squirrels and by egg dimensions of 45-72µm by 27-
42µm (Chandler 1942; Sloss et al.
1994; Bartlett 1995).
Results
Samples from thirty three squirrels were collected
including; 10 southern flying squirrels, 22 eastern gray
squirrels and 1 red squirrel.
Live flying squirrels were not
documented in the nest boxes; however, their presence was
noted by gray underbelly pelage (Krichbaum et al.
2010) and
one occurrence of three deceased infant flying squirrels.
Ten
of the 40 nest boxes contained southern flying squirrel
evidence including fur.
Fecal pellets deposited within the nest
were collected.
Eggs of S. robustus were identified in 21 out of the 33
samples.
Three of the 10 southern flying squirrel (30%)
samples were positive.
Seventeen of the 22 eastern gray
squirrel (77.
3%) road-killed and hunter-killed samples were
positive.
The single red squirrel examined in this study
exhibited S. robustus infection and was collected through
hunter-harvest (Table 1.
)
Journal of Student Research (2013)
Volume 2, Issue 1: pp.
43-47
Research Article
ISSN: 2167-1907
www.jofsr.
com
45
Table 1: Number of individuals sampled and prevalence of Strongyloides robustus infestation identified from eggs from species
in Pennsylvania and New York.
Essex County (NY)
Krichbaum et al.
2010
Carbon, Pike, Warren
Counties (PA)
Krichbaum et al.
2010
Northeastern
Pennsylvania
from Pedder et al.
2009
Nolo, PA (Indiana
County) from Patrick
1991
South-central
Pennsylvania
Current Study
2012
Squirrel
Species
Number
sampled
(N)
Prevalence
(%)
Number
sampled
(N)
Prevalence
(%)
Number
sampled
(N)
Prevalence
(%)
Number
sampled
(N)
Prevalence
(%)
Number
sampled
(N)
Prevalence
(%)
Glaucomys
volans
*
---
20
45
9
11.1
10
100
10
30
Glaucomys
sabrinus
7
0
4
75
2
50
---
---
---
---
Sciurus
carolinensis
---
---
---
---
6
0
---
---
22
77.3
Tamiasciurus
hudsonicus
---
---
---
---
---
---
---
---
1
100
Tamias
striatus
---
---
---
---
3
0
---
---
Total
7
24
20
10
33
* No Glaucomys volans were present.
Discussion
We compared our results in south-central Pennsylvania
to previous experiments in other parts of Pennsylvania and
New York where the northern flying squirrel was historically
(Burt 1957) or is currently found (Table 1) to better illustrate
how parasite distribution may affect northern flying squirrels.
In Essex county New York, where there is a healthy
population of northern flying squirrels and no southern flying
squirrels, no Strongyloides robustus infection was observed in
the 7 specimens tested (Krichbaum et al.
2010).
However, in
the Pennsylvania counties of Carbon, Pike and Warren where
the southern and northern flying squirrels occur sympatrically,
45% (n=20) of southern flying squirrels and 75% (n=4) of
northern flying squirrels were infected with S.
robustus
(Krichbaum et al.
2010).
Additionally another unidentified
location in northeastern Pennsylvania showed similar results
with an unhealthy population of northern flying squirrels
occurring sympatrically with an infected population of
southern flying squirrels (Pedder et al.
2009).
The only other information detailing the infection of
squirrels by S. robustus in the northeastern historical range of
the northern flying squirrel in Pennsylvania was Patrick
(1991), who observed a 100% (n=10) infection rate in
southern flying squirrels from Indiana County Pennsylvania
where no northern flying squirrels were reported.
Both Patrick
(1991) and Krichbaum et al.
(2010) failed to assess the
reservoir potential of other sympatric squirrel species and
although Krichbaum et al.
(2010) demonstrated results that
were consistent with the predictions of the parasite mediated
competition hypothesis they did not survey other squirrel
species that were present in the area.
The lack of potential
reservoir inclusion leaves out a key component toward our
understanding of the parasite mediated competition
hypothesis because it omits a consistent risk factor of
transmission.
Pedder et al.
(2009) surveyed eastern gray
squirrels and eastern chipmunks (Tamias striatus) in an
attempt to assess risk without detecting infection, however,
their sample size (6 gray squirrels) was limited and included
the eastern chipmunk, which is not a known reservoir
(Anderson, 2000).
Our survey hypothesized that a high
prevalence of S. robustus infection would be observed in
south-central Pennsylvania where northern flying squirrels are
extirpated.
These data support the parasite mediated
competition hypothesis and lend increased explanation toward
the that explanation for northern flying squirrel decline by
demonstrating that the infection rate of other squirrel species
can be high in reservoir sympatric squirrels, i.
e. gray squirrels
(77%, n=22) in this study.
As Weigl (2007) notes, there has
been concern for the northern flying squirrel over much of its
range in North America and therefore many studies have
searched for the “magic” factor that might explain its biology
and ensure its survival.
Unfortunately, there may not be a
“magic” factor as the parasite mediated competition
hypothesis may be as important as habitat fragmentation or
loss of suitable habitat.
While the data contained within this report are a
significant contribution towards evaluating the parasite
mediated competition hypothesis, the numbers are small,
including the evaluation of only one red squirrel.
Additional
data needs to be contributed so that a more complete picture
can be obtained that details the prevalence and distribution
throughout the northeastern range of the northern flying
squirrel.
To further develop the pattern of parasitic infections
in Pennsylvania and surrounding areas, the authors
recommend that Sheather’s solution (Dryden et al.
2005)
should be used as the standard floatation method to evaluate
the presence of S. robustus.
Other isolation techniques require
direct observation of intestinal presence after the death of the
specimen (Bartlett 1995, Patrick et al.
1991) or the use of a
modified formalin-ethyl acetate sedimentation technique
(Pauli et al.
2004) that is more difficult to make and use.
Sheather’s sugar flotation solution was a vital part of this
experiment and that of Pedder et al.
(2009).
Sheather’s sugar
solution was used because it has a quick preparation time,
uses a simple design and inexpensive ingredients, as well as
Journal of Student Research (2013)
Volume 2, Issue 1: pp.
43-47
Research Article
ISSN: 2167-1907
www.jofsr.
com
46
possessing the highest specific gravity (1.
27) of other
common floatation solutions (Dryden et al.
2005).
These
attributes increase the opportunity for easy use for other
undergraduate or graduate students and should result in more
data generated detailing the geographical range of the
parasite.
Most importantly, the design followed here is a
noninvasive survey method, relying on deposited fecal matter.
Survivorship is the key when working with any endangered
species.
Since S. robustus has deleterious effects for the northern
flying squirrel, future research should focus on both
increasing the resistance to the parasite, as well as decreasing
the presence of the parasite in sympatric squirrel species.
To
increase resistance, conservationists should increase gene
flow by decreasing
fragmentation.
Consequently,
conservation efforts should focus on developing corridors
between the forest fragments to increase out-breeding and this
may result in an increase resistance to the deleterious effects
of S. robustus. However, even if suitable habitats exist in
south-central Pennsylvania or other areas within the northern
flying squirrel’s historical range, it is unlikely that a
reintroduction effort would be successful without treatment
for the parasite in the squirrel community.
Baiting with
ivermectin medicated bait, such as peanut butter or corn, may
be considered to eliminate the risk of parasitic infection,
similar to the way it has been successfully used to reduce tick
parasitism (Pound et al.
1996).
Unidentified parasite reservoirs for S. robustus have been
historically overlooked by researchers trying to understand
the decline of the northern flying squirrel and this lack of data
does not permit researchers to effectively evaluate the parasite
mediated competition hypothesis.
Our data demonstrates that
S. robustus can be locally abundant in reservoir sympatric
squirrels, which will assist in later conservation efforts.
Understanding the relationship between sympatric squirrel
species and their reservoir roles for S. robustus where the last
remnants of the endangered northern flying squirrel exist will
aid in the development of an effective species recovery plan.
Acknowledgement
We would like to thank several unnamed hunters, personnel at
Letterkenny Army Depot, Jen Wysocki for collecting road-
killed squirrels, Brett Loski for placing the flying squirrel
boxes and Dr.
Tim Maret for allowing access to the boxes
located on his property and Dr.
Nathan Thomas for squirrels
obtained with the Pennsylvania Game Commission permit
SAL00580.
Funding for this project was provided by the
Shippensburg Undergraduate Research Advisory Committee
UGR2011/12-18.
Literature Cited
Allendorf, F.
W., and R.
F. Leary.
1986. Heterozygosity and
fitness in natural populations of animals.
Pp. 57-76 in M.
E. Soule, ed.
Conservation biology: the science of
scarcity and diversity.
Sinauer, Sunderland, MA.
American Society of Mammalogists (ASM) 2012.
Mammals
of Pennsylvania. Retrieved June 5, 2012 from
http://www.
mammalogy.
org/mammals-pennsylvania
Anderson, R.
C. 2000.
Nematode Parasites of Vertebrates:
Their Development and Transmission. Second edition.
Guelph, Ontario: CABI Publishing.
Bartlett, C.
M. 1995.
Morphology , homogonic development,
and lack of a free-living generation in Strongyloides
robustus (Nematoda, Rhabditoidea), a parasite of North
American sciurids.
Folia Parasitologica 42:102-114.
Burt, W.
H. 1957.
Mammals of the Great Lakes Region.
First
edition.
Michigan: The University of Michigan Press.
Butchkoski, E.
and G.
Turner.
2010. Northern flying squirrel,
Glaucomys sabrinus macrotis. Retrieved September 25,
2012, from
http://www.
google.
com/url?sa=t&rct=j&q=&esrc=s&fr
m=1&source=web&cd=2&sqi=2&ved=0CCUQFjAB&u
rl=http%3A%2F%2Fwww.
portal.
state.
pa.us%2Fportal%
2Fserver.
pt%2Fdocument%2F775666%2Fnorthern_flyin
g_squirrel_pdf&ei=0mRjUJetFM6x0QHzooDIBQ&usg
=AFQjCNG56asoMCJJDvij66Ibf2oA4eLXvw&sig2=x1
ID8kxwScpu5Tx8k4oJWw&cad=rja
Chandler, A.
C. 1942.
Helminths of tree squirrels in southeast
Texas.
Journal of Parasitology 28(2): 135-140.
Dryden, M.
W., P.
A. Payne, R.
Ridley, and V.
Smith.
2005.
Comparison of common fecal flotation techniques for the
recovery of parasite eggs and oocysts.
Veterinary
Therapeutics 6(1):15-28.
Fuller ,C.
A. and D.
W. Duszynski.
1997. Eimera
(Protozoa:Eimeriidae) from North American sciurids,
Glaucomys sabrinus and Tamias townsendii: with a
description of a new species.
Faculty Publications from
the Harold W.
Manter Laboratory of Parasitology. 168.
Krichbaum, K.
, C.G.
Mahan, M.
A. Steele, G.
Turner, and P.
J.
Hudson.
2010.The potential role of Strongyloides
robustus on parasite-mediated competition between two
species of flying squirrels (Glaucomys). Journal of
Wildlife Disease 46(1): 229-235.
Kurta, A.
1995. Mammals of the Great Lakes Region.
Revised.
Michigan: The University of Michigan Press.
Mahan, C.
G, J.A.
Bishop, M.
A. Steele, G.
Turner, and W.
L.
Myers.
2010. Habitat characteristics and revised gap
landscape analysis for the northern flying squirrel
(Glaucomys sabrinus), a state endangered species in
Pennsylvania.
American Midland Naturalist 164: 283-
295.
Mahan, C.
G., M.
A. Steele, M.
J. Patrick, and G.
L. Kirkland Jr.
1999. The status of the northern flying squirrel
(Glaucomys sabrinus) in Pennsylvania.
Journal of the
Pennsylvania Academy of Science 73(1): 15-21.
Pauli, J.
N., S.
A. Dubay, E.
M. Anderson, and S.
J. Taft.
2004.
Strongyloides robustus and the northern sympatric
populations of northern (Glaucomys sabrinus) and
southern (G. volans) flying squirrels.
Journal of Wildlife
Diseases 40(3): 579-582.
Patrick, M.
J. 1991.
Distribution of enteric helminths in
Glaucomys volans L.
(Sciurdiae): A test for competition.
Ecology 72(2): 755-758.
Pedder, S.
, C.G.
Mahan, and A.
VanKuren.
2009. Prevalence
of parasites including Strongyloides robustus and
Citellinemia bifurcatum in the endangered northern
flying squirrel and other members of the squirrel family.
[Abstract] In: Proceedings of The Pennsylvania Chapter
of the Wildlife Society pg.
28. Retrieved April 28, 2012
from
Journal of Student Research (2013)
Volume 2, Issue 1: pp.
43-47
Research Article
ISSN: 2167-1907
www.jofsr.
com
47
http://joomla.
wildlife.
org/PA/images/2009%20conf%20p
rogram-%20final.
doc
Pound, J.
M., J.
A. Miller, J.
E. George, D.
D. Oehler, and D.
E.
Harmel.
1996. Systemic Treatment of White-tailed Deer
with Ivermectin-Medicated Bait To Control Free-Living
Populations of Lone Star Ticks (Acari: Ixodidae).
Journal of Medical Entomology 33(3): 385-394.
Price, P.
W., M.
Westoby, and B.
Rice. 1988.
Parasite-
Mediated Competition: Some Predictions and Tests.
The
American Naturalist 131(4): 544-555.
Reiber, R.
J., and E.
E. Byrd.
1942. Some nematodes from
mammals of Reelfoot Lake in
Tennessee.
Jour. Tenn.
Acad. Sci.
17: 78-89.
Roberts, L.
S. and J.
Janovy Jr.
2009. Gerald D.
Schmidt and
Larry S.
Roberts’s Foundations of Parasitology (8
th
ed.).
(pp.414-417).
McGraw Hill, Boston, MA.
Sloss, M.
W., R.
L. Kemp and A.
M. Zajac.
1994. Veterinary
Clinical Parasitology.
Blackwell Publishing.
Ames, IA,
4-96.
Smith, W.
P. 2007.
Ecology of Glaucomys sabrinus: habitat,
demography, and community relations.
Journal of
Mammalogy 88(4): 862-881.
Sparks, J.
L. Jr.
2005. Genetic variability, pathogen
susceptibility, subspecies identity, and conservation of
the endangered northern flying squirrel (Glaucomys
sabrinus) in Virginia.
Master Thesis, Virginia
Commonwealth University, Richmond, Virginia,73pp.
Stevens, L.
, G. Yan, and L.
A. Pray.
1997. Consequences of
Inbreeding on Invertebrate Host Susceptibility to
Parasitic Infection.
Evolution 51(6):2032-2039.
Stewart, R.
L. Jr.
, P.R.
Delis, and C.
M. Kindlin.
2008. An
inventory of the small mammals of Letterkenny Army
Depot, Franklin county Pennsylvania.
Journal of the
Pennsylvania Academy of Sciences 81(2):59-65.
Title 58, Pennsylvania Code, Section 133.
41.
U.S. Fish and Wildlife Service, Virginia Field Office
(USFWS).
2012. Federally Endangered, Threatened,
Proposed, and Candidate Species in Virginia. Retrieved
September 25, 2012, from
http://www.
fws.gov/northeast/virginiafield/pdf/endspecie
s/state_list/VaSpeciesList.
pdf
Weigl, P.
D. 2007.
The northern flying squirrel (Glaucomys
sabrinus): A conservation challenge.
Journal of
Mammalogy 88(4):897-907.
Wetzel, E.
J. and P.
D. Weigl.
1994. Ecological implications
for flying squirrels (Glaucomys spp.) of effects of
temperature on the in vitro development and behavior of
Strongyloides robustus. American Midland Naturalist
131(1): 43-54.