SUSCEPTIBILITY OF WINTER TICK LARVAE AND EGGS TO
ENTOMOPATHOGENIC FUNGI - BEAUVERIA BASSIANA, BEAUVERIA
CALEDONICA, METARHIZIUM ANISOPLIAE, AND SCOPULARIOPSIS
BREVICAULIS

Jay A. Yoder1, Peter J. Pekins2, Blake W. Nelson1, Christian R. Randazzo1, and
Brett P. Siemon1

1Department of Biology, Wittenberg University, Springfield, Ohio 45501, USA; 2Department of Natural
Resources and the Environment, University of New Hampshire, Durham, New Hampshire 03824, USA

ABSTRACT: An isolate of the soil fungus Scopulariopsis brevicaulis was identified from the surface
of female winter ticks (Dermacentor albipictus) collected from recently dead moose (Alces alces) calves
in New Hampshire in the northeastern United States. It was the sole isolate, and it matched with 98%
nucITS similarity (molecular systematics Blast match) to S. brevicaulis species from soil and other
tick species. Inoculation of tick larvae and eggs with 108 spores/mL + 0.05% Tween (aqueous inoculum)
resulted in mortality, reduced survival time, and recovery of S. brevicaulis from within tick tissues.
Rapid water loss and death from dehydration were the pathogenic consequences of the fungal infection.
Three entomopathogenic fungal isolates from laboratory culture (Beauveria bassiana, B. caledonica,
and Metarhizium anisopliae) inoculated concurrently at the same dose, were slightly less pathogenic
to eggs than larvae of winter ticks. We conclude that S. brevicaulis imposes a limitation on the free-
living stages of the winter tick population in specific environmental conditions, but commercial fungal
treatments as used in local situations to control ticks, are impractical as a means of controlling winter
tick density across moose habitats.

ALCES VOL. 53: 41–51 (2017)

Key words: Alces alces, Dermacentor albipictus, moose, pathogenic fungi, Scopulariopsis brevicaulis,
survival, water balance, winter ticks

INTRODUCTION
Winter ticks (Dermacentor albipictus)

periodically cause high calf mortality in
moose (Alces alces) populations in the north-
eastern United States and southern Canada
due to numerous interactive stressors, in-
cluding extreme blood loss, problems with
thermoregulation, and incidence of patho-
genic bacteria (McLaughlin and Addison
1986, Campbell et al. 1994, Samuel 2004,
Musante et al. 2007, Addison and McLaughlin
2014, Jones 2016). As a one-host tick, it
feeds, molts, and mates on the same individual
moose from approximately late September to

mid-April. Once a female has mated and fully
engorged, she drops to and crawls on the
ground to lay eggs in a suitable, moisture-rich
reprieve in soil and leaf litter. Eggs hatch in
about a month and the larvae enter a resting
period during summer, regaining activity and
questing for a host during autumn (Drew
and Samuel 1985, Addison and McLaughlin
1988). The moisture-rich microhabitats in the
northeastern forest preferred for oviposition,
hatching, and quiescence (Yoder et al. 2016)
expose adult females, eggs, and unfed larvae
(the sole transmission stage) to numerous fila-
mentous soil fungi (Tuininga et al. 2009) that

Corresponding author: Jay Yoder, Department of Biology, Wittenberg University, Ward Street at
North Wittenberg Avenue, PO. Box 720, Springfield, Ohio 45501-0720, USA, jyoder@wittenberg.edu

41

mailto:jyoder@wittenberg.edu


are principally saprobes and agents of decay,
but could be infective to ticks.

Fungal infection is a primary source of
mortality in ticks and is often the basis
for their biological control as with the ento-
mopathogenic fungi Beauveria bassiana,
Metarhizium anisopliae, and Scopulariopsis
brevicaulis (Kirkland et al. 2004, Suleiman
et al. 2013). These fungi are regular soil sap-
robes that produce copious amounts of spores
(Barnett and Hunter 2003), live free in soil,
and can invade a perfectly healthy tick. Pre-
sumably, ticks come into contact and are
infected by way of spores that adhere to the
cuticular surface, germinate, and then colon-
ize via hyphae that gain internal access via
the mouth, anus, genital pore, cuticular gland
openings, and between leg segments. Infec-
tion often results from a single fungal species
that exploits the tick (Yoder et al. 2008), such
that an infected tick is essentially a pure cul-
ture of the infectious agent. Once the infec-
tion proliferates, the fungal hyphae typically
protrude from the cuticular glands around
the body. After death, the tick dries out, the
body flattens, and fungal hyphae typically
protrude from the mouth spreading over
the front pair of legs, eventually enveloping
the carcass within the fungal mycelium.
An increased rate of water loss serves as an
indicator of establishment and progression
of the infection (Cradock and Needham
2011). Not all entomopathogenic fungi are
universally pathogenic to all ticks; for ex-
ample, B. bassiana and M. anisopliae are
particularly ineffective against the American
dog tick (Dermacentor variabilis; Kirkland
et al. 2004), a close relative of the winter tick.

We suspected that a fungal mortality
agent existed for winter ticks during routine
handling of specimens used in related studies
with engorged females collected from dead
moose in New Hampshire (Yoder et al. 2016).
Under relatively moist and warm conditions
(93% RH, 25 °C), some of the engorged
females died and became moldy during storage.

Further, we observed healthy ticks die and
became covered with a whitish mold when
housed in the same storage containers with
moldy ticks. The objectives of this study were
to 1) isolate and determine the fungus that
infected the engorged female ticks, 2) deter-
mine if the fungus was pathogenic to winter
tick eggs and larvae (ground-dwelling stages),
and 3) compare the relative pathogenicity of
the isolated fungus with the entomopathogenic
fungi B. bassiana and M. anisopliae in the con-
text of biological control agents.

METHODS
Study area

Winter ticks were collected from dead
moose in eastern Coos County, New Hamp-
shire, an area considered the best habitat and
of highest moose density in New Hampshire
(see Jones 2016). The area is dominated
by mountainous terrain (elevation 300 to
1200 m) bordered by lowland valleys con-
taining a myriad of lakes, ponds, and river
systems. The dominant cover type is northern
hardwood forest with a mix of American
beech (Fagus grandifolia), yellow birch
(Betula alleghaniensis), and sugar maple
(Acer saccharum); balsam fir (Albies balsa-
mea), red spruce (Picea rubens), and white
pine (Pinus strobus) occur on more poorly
drained sites. Monthly precipitation, mean am-
bient temperature, precipitation, snow depth,
and other weather variables were available
from the National Climatic Data Center
(44827 'N, 71811 'W) weather station in Berlin,
New Hampshire (#270690/99999) located
centrally in the study area at 283 m elevation.
Annual ambient temperature ranged from
30 to �30 °C, annual precipitation from 91
to 123 cm, and maximum snow depth from
50 to 70 cm.

Tick collection
Aseptic technique was followed in the

laboratory using materials that were auto-
clave-sterilized (121 °C, 19 psi, 15 min),

42

ENTOMOPATHOGENIC FUNGI TO MOOSE TICKS – YODER ET AL. ALCES VOL. 53, 2017



flamed off using a Bunsen burner, treated
with 95% ethanol, or purchased sterile from
the manufacturer. Sterile, powder-free gloves
were worn when necessary (Microlex Co.,
Reno, Nevada, USA).

Ticks were collected from 5, 10.5 month-
old moose that had been captured and radio-
collared in January 2015; death occurred
24-36 h prior to collection. All moose were
emaciated and mortality was attributed to an-
emia and hypoproteinemia from excessive
blood loss associated with >30,000 winter
ticks/animal (Jones 2016). Ticks were col-
lected from the neck, shoulder, abdomen,
and rump of moose, and included nymphs
and adults in various stages of feeding and
unfed specimens. All ticks were identified
as Dermacentor albipictus from keys (Brinton
et al. 1965). Fed females were placed individu-
ally into 50 mL polypropylene centrifuge tubes
(Fisher Scientific, Pittsburgh, Pennsylvania,
USA) within Whirl-Pak bags (Nasco, Salida,
California, USA) and transported to the labora-
tory in 5 L coolers kept at ~15 °C with cold
packs (Koolit; FDC Packaging, Medfield,
Massachusetts, USA).

In the laboratory, each fed female was
transferred to a fresh tube and stored at
93% RH (SD ± 0.5% RH; Winston and
Bates 1960) in 3000 mL glass desiccators at
25 ± 0.5 °C, 10L:14D (programmable incu-
bator; Fisher). Subsequent hatched larvae
were identified as D. albipictus using keys
(20 slide-mounted larvae); no other species
of larvae was identified. At the time of collec-
tion, all fed females were in healthy condition
in that their body was plump and blood-filled,
and they could crawl 5 body lengths.

Isolation and identification of fungi from
tick cadavers

The methods used to isolate fungi from
moldy ticks, the preparation and spore con-
centration of inoculum, use of Tween as an
emulsifier (dispersing agent), treatment of
larvae and eggs, and reisolation of fungi

were modified from previous studies with
ticks and entomopathogenic fungi (Fernandes
et al. 2004, Kirkland et al. 2004, Tuininga
et al. 2009, Suleiman et al. 2013). Moldy
engorged females were used to culture fungi
from pieces of hyphae scraped from their
carcasses. Individual hyphae were plated indi-
vidually onto solidified potato dextrose agar
(PDA; Fisher) in disposable 100 � 15 mm
Petri plates (Fisher) that were incubated in dark-
ness at 25 °C. The fungus was purified with
3 rounds of subculturing hyphal tips, each util-
izing the advancing edge of a 3-4 week old
mycelium. Pure cultures were identified at the
University of Alberta Microfungus Collection
and Herbarium (UAMH) Centre for Global
Microfungal Biodiversity at the Gage Re-
search Institute (Toronto, Ontario, Canada).
The gene sequenced for identification was
nucITS (internal transcribed spacer region)
and primers ITS5/ITS4 were used for amplifi-
cation. ClustalX software in MEGA5 was
used for aligning nucITS sequences (Gen
Bank) to compare with other species at the
Department of Biological Sciences, University
of Cincinnati (Cincinnati, Ohio, USA).

Preparation of fungal inoculum
An aqueous inoculum was prepared from

1 month-old sporing PDA cultures in phos-
phate buffered saline (PBS, pH 7.5) + 0.05%
Tween 20 (Fisher). Spores were scraped from
the plates into PBS, purified, and the concentra-
tion was adjusted to 1.4 � 108 spores/mL with a
0.1% dye exclusion (AO Spencer Bright-Line
Hemocytometer, St. Louis, Missouri, USA).
Identical preparation of aqueous inocula (each
at 1.4 � 108 spores/mL in PBS + Tween) was
performed for Beauveria bassiana, B. caledo-
nica, and Metarhizium anisopliae from the
Agricultural Research Service Collection of
Entomopathogenic Fungal Cultures (ARSEF)
associated with the United States Depart‐
ment of Agriculture-Agricultural Research Ser-
vice (USDA-ARS) (Ithaca, New York, USA)
(see Table 1 for isolate number information).

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43



We used these entomopathogenic fungi as posi-
tive controls; PBS + 0.05% Tween served as the
negative control.

Treatment with inoculum
Ten larvae were placed into a 1.5 mL

microcentrifuge tube (Fisher) containing
1 mL of inoculum that was gently agitated
for 2 min, and then poured onto filter paper
(No. 3, Whatman, Hillsboro, Oregon, USA).
Actively crawling larvae were collected and
placed into a clean 1.5 mL microcentrifuge
tube; a hole was punched through the tube
lid and covered with mesh. It was placed at
80% RH (Winston and Bates 1960), 25 °C,
and 10h:14h L:D cycle in a sealed glass des-
iccator. Dead larvae were identified with a
40� light microscope; i.e., larvae with curled
legs, deflated opisthosoma, and no move-
ment. The identical inoculum treatment was
used with eggs and death was assumed
when the eggshell chorion showed sign of
collapse; eggs in such condition fail to hatch
based on water balance studies (Yoder et al.
2016). The experiment was complete after
all larvae died, including those in the PBS +
Tween control, and after egg hatching occurred.

Reisolation of fungi
In accordance with Koch's postulates

as confirmation of pathogenicity, dead larvae

and eggs were prepared for reisolation by
fungus culturing. Briefly, each was surface
sterilized twice for 1 min in a mild bleach
solution (18:1:1 ratio of deionoized water:
absolute ethanol:5.25% NaOCl by volume),
with a final rinse in water. It was then halved
by scalpel and the portions were embedded
in PDA, each in its own plate, and incubated
in darkness at 25 °C. Tips of hyphae that
could be traced as originating from the inter‐
nal body contents (40/45� microscopy) were
removed as a 1 cm3 block for sub-culturing
and identification. The fungus was identified
with standard keys (Barnett and Hunter 2003)
and pure culture comparison to the original
fungus isolates used to prepare the inoculum.
Defining characteristics were identified by
using colony obverse and reverse, and spore
size and shape under oil (1000�).

Larvae were 4-6 weeks old and eggs
were 2–3 weeks post-oviposition; all were
healthy at the time of treatment. Eggs were
full and rounded, with a visible accumulation
of white guanine through the eggshell that
is a developmental landmark of regular em-
bryonic development. Larvae crawled about
actively, could self-right, and crawl 5 body
lengths.

Data are the mean ± SE of 10 replicates
from 10 specimens each (n = 100). An ana-
lysis of covariance was used to test data

Table 1. Entomopathogenic fungi used in this study. The Scopulariopsis brevicaulis isolate 11903 is the new
isolate collected from dead winter ticks originating from New Hampshire, USA.

Fungus and isolate# Host Origin Deposited

Beauveria bassiana Leptinotarsa decemlineata France, Europe ARSEF1

149 (Colorado potato beetle)

Beauveria caledonica Hadenoecus cumberlandicus Kentucky, USA UAMH2

11821 (Cave cricket)

Metarhizium anisopliae Conoderus sp. North Carolina, USA ARSEF

23 (Click beetle)

Scopulariopsis brevicaulis Dermacentor albipictus New Hampshire, USA UAMH

11903 (Winter tick)

1ARSEF, USDA-ARS Collection of Entomopathogenic Fungal Cultures, Ithaca, New York, USA.
2 UAMH, UAMH Centre for Global Microfungal Biodiversity, Toronto, Ontario, Canada.

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ENTOMOPATHOGENIC FUNGI TO MOOSE TICKS – YODER ET AL. ALCES VOL. 53, 2017



(ANCOVA; P = 0.05; SPSS 14.0, Microsoft
Excel and Minitab, Chicago, Illinois, USA).
Survival times were compared with the t stat-
istic utilizing a Kaplan-Meier survival curve
with a log rank test. An Abbott correction
for mortality data and logit-transformation
for percentage data were used prior to analysis.

Water balance experiments
Eggs and larvae were analyzed similarly.

After treatment with inoculum (4 d post-
treatment), each specimen was weighed indi-
vidually with a microbalance (SD ± 0.2 μg
precision, ± 6 μg accuracy at 1 mg; Cahn
Ventron Co., Cerritos, California, USA).
This measurement was made in <1 min with-
out enclosures or anesthesia. Standard kinetic
model equations were used to determine water
balance characteristics based on the mass mea-
surements (Yoder et al. 2016). All specimens
were predesiccated by 4-6% so that the change
in mass reflected the change in body water
content. The percent water content was deter-
mined by weighing the specimen (initial,
fresh mass, f ), drying it to constant mass (dry
mass, d) at 90 °C in a drying oven (< ± 2 °
C; Blue M Electric Co., Chicago, Illinois,
USA), and calculating the difference between
these measurements: 100% (f � d)/f, where f
� d is the water mass, m.

The dehydration tolerance limit was de‐
termined by weighing the specimen, placing
the specimen at 33% RH (Winston and Bates
1960) in a glass desiccator at 25 °C, and then
reweighing the specimen at time intervals.
The critical mass measurement for larvae
was the point at which a larva was unable
to self-right and crawl 5 body lengths, and
for eggs when the eggshell chorion began to
collapse. Specimens at their critical mass
were transferred to a 90 °C drying oven to de-
termine dry mass (d). The difference between
critical mass and dry mass was defined as the
critical water mass (mc). The amount of water
loss that was sustained between the initial
water mass (m0) and mc was defined as the

dehydration tolerance limit expressed as a
percentage: 100% (mc � m0)/m0.

The water loss rate (respiratory + integu-
mental water loss) was measured at 0%
RH because this is the only condition where
the rate is exponential, allowing it to be
derived from the slope of a regression line.
The 0% RH condition was maintained with
anhydrous calcium sulfate at 25 °C in a glass
desiccator (Drierite; 1.5 x 10�2% RH; W. A.
Hammond Drierite Company, Xenia, Ohio,
USA). The specimen was weighed, placed
at 0% RH, and reweighed 5 times at various
intervals. The specimen was then transferred
to a 90 °C drying oven to achieve dry mass
(d) and water mass (m) was calculated by
subtraction. The water loss rate (�kt) was
determined from mt = m0exp-kt, where mt is
the water mass at any time t, and m0 is the
initial water mass.

Each water balance characteristic was
based on a sample size of 100; 10 replicates
of 10 specimens each. Data (the mean ± SE)
were tested using ANCOVA (P = 0.05). Per-
centage data were logit-transformed prior to
analysis, and regression lines were compared
with a test to compare characteristics and
slopes from multiple regression lines (SPSS
14.0, Microsoft Excel and Minitab).

RESULTS
Identification of fungus (S. brevicaulis)

The fungus that was scraped and isolated
from a moldy, fed female was identified as
Scopulariopsis brevicaulis (Sacc.) Bainier
(UAMH isolate 11903); this isolate origi-
nated from a single tick. The identification
was based on sequence analysis and 99%
similarity of the ITS region with other S. bre-
vicaulis strains and the basic morphological
characters of the group. The S. brevicaulis
isolate 11903 is deposited at The UAMC
Centre in Toronto, Ontario, Canada (Table 1).
Scopulariopsis brevicaulis was the most com-
mon isolate and was recovered in pure culture
from 21 moldy females (identification based

ALCES VOL. 53, 2017 YODER ET AL. – ENTOMOPATHOGENIC FUNGI TO MOOSE TICKS

45



on pure culture comparison to authentic S. bre-
vicaulis strain 11903). Not all of the collected
fed female ticks from moose had a fungal in‐
fection (n = 21 of >110), not all fed female ticks
were collected from the same moose, and not
all fed female ticks were collected from moose
during the same week. The inoculums for test-
ing were made from a single isolate culture
(i.e., not batched from all 21 positive ticks).
Other fungi recovered less frequently (<15%)
from fed female ticks included Aspergillus
spp., Penicillium spp., and Paecilomyces spp.

Effect of S. brevicaulis 11903 on survival
on larval ticks and eggs

Survival was reduced in S. brevicaulis-
treated larvae (P < 0.05; Fig. 1) that lasted
14.0 d (7.5 d for 50% of larvae) versus 18.0 d
for control larvae (11.2 d for 50% of larvae).
The recovery of S. brevicaulis from dead larvae
at the end of the experiment was 90.4 ± 2.2% in
the treated group and 6.3 ± 2.9% in the control
group (P < 0.05). Similarly, treated eggs had
lower survival and hatching rate, and higher
recovery of S. brevicaulis from unhatched
eggs than the control group (Table 2).

Effect of S. brevicaulis 11903 on water loss
on larval ticks and eggs

Scopulariopsis brevicaulis-treated larvae
lost water >2 � faster (P < 0.05) than con‐
trol larvae (4.39 ± 0.07%/h vs. 1.84 ±
0.04%/h; Fig. 2). The recovery of S. brevi-
caulis was ~5� higher (P < 0.05) in the trea-
ted (86.7 ± 4.7%) than control group (17.8 ±
3.9%). Scopulariopsis brevicaulis was re‐
covered internally from dead larvae in the
treatment group, whereas only dead larvae
tested positive for S. brevicaulis in the control
group (P < 0.05). Similarly, eggs had higher
(P < 0.05) water loss rate in the treatment
(1.12 ± 0.019%/h) than control group (0.71
± 0.033%/h) (Fig. 2). Dead eggs also had
higher (P < 0.05) internal recovery of S. bre-
vicaulis in the treated (71.1 ± 2.9%) than con-
trol group (4.4 ± 3.9%). There was greater
recovery of S. brevicaulis in larvae than
eggs (P < 0.05).

After 4 days post-treatment, the fresh mass,
water mass, and % water content were similar
for eggs and larvae in the S. brevicaulis-
treated and control groups (Table 3), which
reflected the similarity of the water:dry mass

Control M. anisopliae S. brevicaulis

0

20

40

60

80

100

0 2 4 6 8 10 12 14 16 18
0

20

40

60

80

100

0 2 4 6 8 10 12 14 16 18

B. caledonica B. bassiana

Time (days)

Su
rv

iv
al

 o
f 

�
ck

 la
rv

ae
 (

%
)

Fig. 1. Survivorship curves for unfed larvae of Dermacentor albipictus after
treatment with Metarhizium anisopliae, Scopulariopsis brevicaulis, Beauveria
caledonica, or B. bassiana ( in order from left to right on the graph). The control
on both plots is the solid black line for comparison with the treatments. Each point
is the mean of 100 larvae (± SE ≤ 2.1).

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ENTOMOPATHOGENIC FUNGI TO MOOSE TICKS – YODER ET AL. ALCES VOL. 53, 2017



ratios (m/d): 1.44 and 1.28 for larvae and 1.81
and 1.96 for eggs in the S. brevicaulis-treated
and control groups, respectively. In all cases,
water mass was a positive correlate of dry
mass in larvae (R ≥ 0.90 for control and
≥ 0.94 for S. brevicaulis-treated) and eggs
(R ≥ 0.89 for control and ≥ 0.91 for S.
brevicaulis-treated) (P < 0.001).

Comparative observations with
other entomopathogenic fungi on
larval ticks and eggs

Larval survival was reduced (P < 0.05)
by treatment with B. bassiana, B. caledonica,
and M. anisopliae: 8.1, 7.0, and 8.3 d, re-
spectively, compared to 11.2 days for 50%
of control larvae (Fig. 1). At the end of the

Table 2. Mortality characteristics associated with Beauveria bassiana, B. caledonica, Metarhizium
anisopliae, or Scopulariopsis brevicaulis to eggs of Dermacentor albipictus. Values (the mean ± SE; n =
100 eggs) followed by the same superscript letter within a column are not significantly different (P <
0.05). Eggs in the control had an incubation time of 44.6 ± 2.1 days.

Days elapsed before

Treatment
chorion collapsed
(50% of eggs)

%/100 eggs
that hatched

% dead eggs positive
for test fungus

Control (PBS + Tween) Not observed (0.0)a 82.6 ± 5.5a 11.8 ± 2.0a

Test (108 spores/mL)

B. bassiana 16.1 ± 3.1b 43.7 ± 5.2b 73.2 ± 2.0b

B. caledonica 13.2 ± 1.7b 46.2 ± 3.0b 62.9 ± 1.4c

M. anisopliae 11.0 ± 2.2c 44.1 ± 4.7b 80.4 ± 3.2d

S. brevicaulis 8.4 ± 2.1d 36.6 ± 4.3c 76.2 ± 2.8b

–0.25

–0.2

–0.15

–0.1

–0.05

0
0 2 4 6 8 10

ln
(m

t/
m

0)

Control +S. brevicaulis

Egg Larva

Time (hours)

–0.25

–0.2

–0.15

–0.1

–0.05

0
0 1 2 3 4 5

Fig. 2. Water loss rate of unfed larvae and eggs of Dermacentor albipictus after
treatment with Scopulariopsis brevicaulis. The water loss rate is derived from the
slope of the regression line; mt = water mass at time t; m0 = initial water mass. Each
point is the mean of 100 specimens.

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47



experiment, 86.4 ± 2.6% larvae tested posi-
tive for B. bassiana, 88.6 ± 3.1% tested posi-
tive for B. caledonica, and 92.9 ± 1.9% tested
positive for M. anisopliae in their respective
treatment groups; none was detected in the
control groups.

The 3 fungal treatment groups reduced
egg survival and hatching compared to the
control group (Table 2). The S. brevicaulis
treatment had more detrimental effect on
hatching than the other fungal treatments.
No dead eggs in the control tested positive
for B. bassiana, B. caledonica, or M. aniso-
pliae; however, 11.8% were positive for S.
brevicaulis. The recovery of B. bassiana,
B. caledonica, and M. anisopliae was con-
sistently lower (P < 0.05) in treated eggs
than treated larvae.

DISCUSSION
This study produced 2 novel findings:

1) that larvae and eggs of the winter tick
are susceptible to fungal isolates of B. bassi-
ana, B. caledonica, and M. anisopliae, and
2) that larvae and eggs of the winter tick are
susceptible to S. brevicaulis, a common soil
fungus. Both B. bassiana and M. anisopliae
are approved for tick control in the United
States under various commercial formulation
trademarks (Stafford and Allan 2010). Beau-
veria caledonica is a pathogen of forest
beetles and is used in formulated appli‐
cations for biological control of bark beetles
(Hylastes ater and Hylurgus ligniperda) in

New Zealand (Brownbridge et al. 2010).
This is the first instance that an isolate of
B. caledonica has been tested and shown to
be pathogenic against ticks, suggesting prom-
ise for biological tick control. It is apparent
that S. brevicaulis is a pathogen in the study
area given its origin from host moose.

Although B. bassiana, M. anisopliae,
B. caledonica, and S. brevicaulis were patho-
genic against both winter tick larvae and
eggs, larvae were more vulnerable to infec-
tion. It follows that application of an entomo-
pathogenic agent would probably be most
effective in the autumn when larvae are
active. Similarly, other investigators noted
that eggs of other tick species are more resist-
ant to entomopathogenic fungal infection
than later life stages, and attribute this to the
resistant properties of the eggshell chorion
(Fernandes et al. 2004). The entomopatho-
genic fungi tested were consistent in shared
features with other ticks challenged with ento‐
mopathogenic fungi in laboratory studies
(see Fernandes et al. 2012). Specifically,
under warm temperature (25 °C) and mois-
ture levels >80% RH favorable to ticks,
there was: 1) confirmation of pathogenicity
by Koch's postulates, 2) suitable infection
from a topical application, 3) high mortality
with a 108 spores/mL concentration, and 4)
post-treatment infection with 108 spores/mL
causing death in approximately 10-12 days.

Scopulariopsis brevicaulis is distributed
worldwide as a common mold in soil and

Table 3. Water content and dehydration tolerance of unfed larvae and eggs of Dermacentor albipictus that
were treated with Scopulariopsis brevicaulis. Values (the mean ± SE) followed by the same superscript
letter within a column are not significantly different (P < 0.05); n = 100 specimens each.

Stage Fresh mass (mg) Water mass (mg) Water content (%) Dehydration tolerance (%)

Larva

Control 0.041 ± 0.008a 0.023 ± 0.006a 56.10 ± 1.43a 21.73 ± 0.62a

+ S. brevicaulis 0.044 ± 0.010a 0.026 ± 0.008a 59.09 ± 1.28a 24.06 ± 0.49a

Egg

Control 0.073 ± 0.012b 0.047 ± 0.005b 64.38 ± 1.23b 36.19 ± 0.82b

+ S. brevicaulis 0.068 ± 0.009b 0.045 ± 0.008b 66.18 ± 1.37b 38.44 ± 0.57b

48

ENTOMOPATHOGENIC FUNGI TO MOOSE TICKS – YODER ET AL. ALCES VOL. 53, 2017



forest leaf litter, and is a common saprobe
found on fur, hooves, and horns of small
and large mammals (Shubina et al. 2013).
Thus, seed ticks come into contact with S.
brevicaulis spores when crawling or as eggs
on the ground, or as larvae on host fur. Sco-
pulariopsis brevicaulis has also been isolated
from moose dung in black spruce (Picea
mariana) forests in Alberta, Canada (listed
as teleomorph M. brevicaulis UAMH num-
ber 9458; isolator S. P. Abbott), indicating
a linkage among S. brevicaulis, moose, and
moose habitat.

Results here for S. brevicaulis show at
this inoculum dose (108 spores/mL), it kills
winter tick. An extended period of wetness
can trigger S. brevicaulis to proliferate and
possibly induce tick mortality. For example,
certain engorged specimen females died
from S. brevicaulis infection after prolonged
storage under high moisture (93% RH); ef-
fectively, S. brevicaulis proliferated and
killed the ticks. Exposure to a large inoculum
of 108 spores/mL of the study set of fungi
(B. bassiana, B. caledonica, M. anisopliae,
and S. brevicaulis) and moisture >80% RH
is a lethal combination to eggs, larvae, and
adult female winter ticks. The biological sig-
nificance of S. brevicaulis as a limitation on
winter tick populations would presumably
occur only during optimal environmental
conditions (e.g., extended period of high
moisture and temperature to enhance spore
production), and the likelihood or frequency
of such is unknown.

CONCLUSIONS
Due to the potential in mammalian nail

and skin infections by S. brevicaulis (Lee
et al. 2012), precautionary measures should
be taken if it is considered as a biological con-
trol agent. Biological control of winter ticks
is theoretically possible with B. bassiana
and M. anisopliae at 108 spores/mL or higher
formulations that are consistent with com-
mercially available products. However,

commercial applications are typically local
(e.g., backyards), and arguably, impractical
across moose habitat. Fernandes et al.
(2012) discuss concerns and issues concern-
ing application of entomopathogenic fungi
in natural settings for tick control.

ACKNOWLEDGEMENTS
This project was made possible by grants

from M. A. Senich (Midland, Texas) to
BWN and S. McWilliam to BPS, a gift from
E. E. Powelson (Springfield, Ohio) to the
Department of Biology at Wittenberg, and
the New Hampshire Fish and Game Depart-
ment (Concord, New Hampshire) to PJP.
We thank Y. Guardiola and J. Scott (UAMH,
Toronto, Ontario) for fungus identification,
J. Benoit (University of Cincinnati, Cincinnati,
Ohio) for sequence analysis, and H. Jones
and D. Ellingwood of the University of New
Hampshire (Durham, New Hampshire) for
collecting specimen ticks.

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	SUSCEPTIBILITY OF WINTER TICK LARVAE AND EGGS TO ENTOMOPATHOGENIC FUNGI -BEAUVERIA BASSIANA, BEAUVERIA CALEDONICA, METARHIZIUM ANISOPLIAE, AND SCOPULARIOPSIS BREVICAULIS 
	INTRODUCTION
	METHODS
	Study area
	Tick collection
	Isolation and identification of fungi from tick cadavers
	Preparation of fungal inoculum
	Treatment with inoculum
	Reisolation of fungi
	Water balance experiments

	RESULTS
	Identification of fungus (S. brevicaulis) 
	Effect of S. brevicaulis 11903 on survival on larval ticks and eggs
	Effect of S. brevicaulis 11903 on water loss on larval ticks and eggs
	Comparative observations with other entomopathogenic fungi on larval ticks and eggs

	DISCUSSION
	Conclusions
	ACKNOWLEDGEMENTS
	REFERENCES