Stress-Hormone Levels of Wolves in Relation to Breeding Season, Pack Size, Human Activity, and Prey Density

Abstract

Human disturbance is thought to be a major source of stress for animals but breeding status, social interactions and food availability are also potential sources. Long-lasting stress may adversely affect the fitness of animals and for that reason the evaluation of stressors is important for conservation of threatened species. The aim of our study was therefore to assess which factors cause stress in wolves (Canis lupus). We evaluated the stress levels of wolves from six packs by measuring the concentration of glucocorticoid metabolites in 59 faecal samples with a Cortisol enzyme-immunoassay. During the breeding season, stress hormone concentration was higher than during the rest of the year, with two peaks around mating and begin of denning, respectively. Multiple regressions ranked by AIC showed that breeding had the highest impact on the wolves' stress levels, followed by human activity, pack size, and prey density. We conclude that human activity is only one of several factors contributing to stress in wolves and that intraspecific competition during breeding is likely to cause elevated levels of glucocorticoids.

Full text

Ann. Zool. Fennici  
 




1)Section Biology, Faculty of Science and Technology, University of Siegen, Adolf-Reichwein-
Str. 2, D-57068 Siegen, Germany (corresponding author’s e-mail: julia.eggermann@web.de)
2)Museum and Institute of Zoology, Polish Academy of Sciences, ul. Wilcza 64, PL-00-679
Warsaw, Poland
3)Bieszczady National Park, PL-38-713 Lutowiska 2, Poland
)State Forest Superintendency of Suchedniów, ul. Bodzentyńska 16, PL-26-130 Suchedniów,
Poland
Received 5 July 2012, nal version received 20 Nov. 2012, accepted 1 Feb. 2012
 
Ann. Zool.
Fennici
Human disturbance is thought to be a major source of stress for animals but breeding
status, social interactions and food availability are also potential sources. Long-lasting
stress may adversely affect the tness of animals and for that reason the evaluation of
stressors is important for conservation of threatened species. The aim of our study was
therefore to assess which factors cause stress in wolves (Canis lupus). We evaluated
the stress levels of wolves from six packs by measuring the concentration of glucocor-
ticoid metabolites in 59 faecal samples with a cortisol enzyme-immunoassay. During
the breeding season, stress hormone concentration was higher than during the rest of
the year, with two peaks around mating and begin of denning, respectively. Multiple
regressions ranked by AIC showed that breeding had the highest impact on the wolves’
stress levels, followed by human activity, pack size, and prey density. We conclude that
human activity is only one of several factors contributing to stress in wolves and that
intraspecic competition during breeding is likely to cause elevated levels of glucocor-
ticoids.
Introduction
Centuries of wolf (Canis lupus) persecution
caused wolves to avoid humans (e.g. Thurber et
al. 1994, Theuerkauf et al. 2003a, 2003b), but
human presence does not necessarily mean that
it negatively impacts wolves. The long history
of wolf persecution has evolutionarily favoured
wolves that avoided humans but, at the same
time, has forced them to adapt to live and breed
in close proximity to them. This mixture of avoid-
ance and habituation seems to be the basis of
wolf-human coexistence in areas where wolves
occupy habitats with relatively high human activ-
ity (Theuerkauf et al. 2007). In such situations, it
is difcult to discriminate which kind of human
activity actually reduces tness of wolves and can
as such be regarded as a disturbance.

   Stress hormones in wolves 
An elevated level of glucocorticoids is a
physiological reaction allowing animals to
efciently hunt or ee. Prolonged elevation of
glucocorticoids, however, has serious negative
effects and is dened as chronic stress (McEwen
& Sapolsky 1995, Wingeld & Sapolsky 2003).
It has a considerable impact on virtually all
bodily functions and can disrupt reproduction,
alter the animal’s behaviour and cognition, and
degrade the performance of the animal’s immune
system, resulting in reduced resistance to disease
(McEwen & Sapolsky 1995, Wingeld & Sapol-
sky 2003). It is unknown which type of human
activity within the wolf environment can cause
chronic stress and adversely affect their tness.
Human recreational activity and snow sports
were shown to increase faecal glucocorticoids
in several mammalian species (Creel et al. 2002,
Taylor & Knight 2003, Arlettaz et al. 2007).
Such stress response suggests that human pres-
ence may cause prolonged stress in animals and
hence adversely affect their tness.
Wolves deal with human activity by tem-
porarily avoiding areas used by humans at that
particular moment (spatio-temporal segregation
as dened in Theuerkauf et al. 2003b). Another
possibility that allows wolves to live in the prox-
imity of humans without experiencing the nega-
tive consequence of permanent stress may be
habituation to human presence and subsequent
reduced stress response. However, empirically
little is known if these assumptions apply in
the wild, because no study so far measured the
actual levels of stress response by wolves in rela-
tion to varying intensities of human presence.
Glucocorticoids in scats were used as physi-
ological indicators of stress in a variety of spe-
cies (Touma & Palme 2005). As capture and han-
dling of the animals is omitted, stress measure-
ments from scats are unaffected by the observer
and thus reect the actual stress level of the
animal more accurately (Kotrschal et al. 1998).
Moreover, glucocorticoid metabolites in scats
are pooled over a certain period, determined
by gut passage time and dynamics of excre-
tion (Scheiber et al. 2005). Thus, hormone con-
centrations in scats represent an assessment of
chronic stress. For these reasons, we measured
faecal glucocorticoid metabolite concentrations
to assess stress levels of wolves noninvasively.
We aimed at assessing the effect of breeding,
pack size, prey abundance, and human activity
within six wolf pack home ranges on the stress
levels of wolves. We hypothesised that human
activity would not be the major factor inuenc-
ing stress in wolves.
Methods
This study was conducted in two distinct areas.
One was situated in the southeast of Poland,
in the Bieszczady Mountains (49°19´–49°50´N,
22°15´–22°45´E). We took faecal samples of
ve wolf packs that inhabited an area of about
1000 km2 (Eggermann et al. 2009). The second
region (250 km2) was situated in the Holy
Cross Forest (Puszcza Świętokrzyska) in cen-
tral Poland (51°02´N, 20°44´E). Here, we took
faecal samples of a wolf pack that was discov-
ered only in 2006, which was the rst record of
wolves in this region since they were extirpated
in 1953 (Gula 2008a, 2008b).
We collected faecal samples incidentally in
2004 and 2005 (n = 4), but carried out a system-
atic scat survey from March 2006 to March 2007
(n = 55). We searched for scats along randomly
selected transects on roads or tracks throughout
the study area. During summer and autumn,
only fresh (still humid) scats were collected to
avoid microbial fermentation (Khan et al. 2002).
Samples were frozen within a few hours after
collection and stored at –20 °C until analysis, as
recommended by Hunt and Wasser (2003). As
we did not assign faecal samples to individual
wolves, we might have sampled some wolves
more often than others.
We extracted metabolites from the scats fol-
lowing the protocol by Schatz and Palme (2001).
They yielded the highest amount of metabolites
(about 70%) with 80% methanol. For the meas-
urement of glucocorticoid metabolite concentra-
tions, we used a cortisol enzyme immunoassay
developed by Palme and Möstl (1997), which
was validated in the red wolf Canis rufus (Young
et al. 2004) and the dog Canis lupus familiaris
(Schatz & Palme 2001).
To detect potential seasonal variation, we
grouped faecal samples in ve periods and cal-
culated mean glucocorticoid metabolite concen-

 Eggermann et al.   
trations for each of them. During the breed-
ing season we chose shorter intervals, to reveal
potential peaks connected to breeding. Based on
their sampling date, we classied faecal samples
as collected during the breeding period (Febru-
ary–May) or the non-breeding period (June–
December). We considered pack size as the max-
imum number of wolves seen or snow-tracked
(Gula 2008c) during the same year we collected
the scats. We calculated prey abundance within
the home ranges of each pack as the sum of
harvest densities of ungulates (red deer Cervus
elaphus, roe deer Capreolus capreolus and wild
boar Sus scrofa; data provided by local hunting
authorities), which are the main prey of wolves
in the study areas (Gula 2004). We calculated
home ranges as MCP (minimum convex poly-
gon) of multiannual radio telemetry locations or
multiannual snow tracking data (Gula 2008c).
A magnetic trafc counter (NC-30 NU-met-
rics, Uniontown, Pennsylvania, USA) recorded
the trafc on roads within the home ranges.
We assigned roads in each MCP to four classes
(1–500, 501–5000, 5001–10 000 and more than
10 000 vehicles per week) according to the
recorded intensity of trafc. We then calculated
the road density of each class (km km–2) for
every home range and multiplied it by the aver-
age trafc in each class. We calculated a trafc
index for each pack by summing up the calcu-
lated trafc values for each road class (Table 1).
We used multiple linear regression models
(SPSS ver. 19), with the level of stress hormones
in individual wolf scats as a response variable
and breeding, pack size, prey density, and traf-
c index as predictor variables. Breeding was
entered as a nominal (0/1) variable whereas the
other three parameters were metric. We then
ranked the models by Akaike weights (w) (Burn-
ham & Anderson 2002) to assess which of the
four parameters inuenced stress hormone levels
the most. We log-transformed hormone concen-
trations for the regression analyses, because they
were not normally distributed. All means are
provided with 95% condence intervals (CI).
Results
The mean glucocorticoid metabolite concentra-
tion of individual scats was 11.4 ± 2.8 (CI) ng g–1
of fresh faecal mass and ranged from 0.6 ng g–1
to 53.9 ng g–1. Average hormone concentrations
varied among wolf packs from 8.5 ng g–1 to
16.3 ng g–1 (Table 1). Glucocorticoid metabolite
concentrations peaked during mating and at the
beginning of denning (Fig. 1). Breeding was the
highest-ranking factor (Table 2) inuencing log-
transformed levels of glucocorticoid metabolites
(sum of Akaike weights: 0.60), followed by
trafc index (0.53), pack size (0.40), and prey
density (0.35).
Discussion
Despite the intensive trafc, high density of roads
and a human density of 36 inhabitants per km
(Theuerkauf et al. 2007) within the home ranges
of the studied packs, the levels of glucocorticoids
in wolf scats were most inuenced by breed-
ing. Breeding is a period of elevated aggression
among members of a wolf pack (Rabb et al.
1967, Zimen 1976). During this period, sexual
Table 1.

      
  
      
      
      
      
      
      


   Stress hormones in wolves 
0
5
10
15
20
25
Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec
Cortisol concentration (ng g–1)
Table 2.Δ
  wΔ 
    
    
    
    
    
    
    
    
    
    
    
    
    
    
    
competition and the related aggression can even
trigger dispersal of young, but sexually mature,
wolves (Mech & Boitani 2003). Our data sug-
gest that social interactions among wolves cause
more stress than human presence in wolf habitat.
Therefore, we argue that human activity is not
the major factor inducing stress in wolves. These
ndings support earlier conclusions that wolves
living in areas with higher human densities might
habituate to human activity (Theuerkauf et al.
2003b, 2007). Habituation and a reduced stress
response is another indication for the behavioural
plasticity of wolves, such as high variation in
their daily activity patterns (Eggermann et al.
2009) and spatio-temporal avoidance of humans
(Theuerkauf et al. 2003b). We believe that behav-
ioural plasticity is a decisive adaptation, which
allowed wolves to survive in close proximity of
people as long as they are not persecuted.
Although our sample size was too small to
produce conclusive results, it seems that wolves
experience the highest stress in two periods of
the year: during the mating period in February
and at the start of the denning period. While
stress during the rst period is likely caused by
sexual competition and the resulting conicts,
it is more difcult to explain why the denning
induces stress. An explanation might be that
wolves need more effort than usual for hunting
to provide enough food for nursing females,
and later for the growing pups. Prey availability
along with pack size and human activity was one
of the factors that also contributed to the level of
stress hormones. While the impact of pack size
is likely to be explained by the intensity of social
interactions, with higher stress levels in larger
packs, the inuence of prey availability is related
to the level of difculties in hunting and subse-
Fig. 1.  

    n 
 n 
nn
n 
  
    
  
   


 Eggermann et al.   
quent food stress. Prey abundance is also known
to determine the habitat selection by wolves (i.e.
Eggermann et al. 2011) and their activity pat-
terns (Theuerkauf 2009). However, under the
study conditions, it does not seem that prey
availability was an important factor for stress in
wolves. We conclude that mostly intrinsic factors
inuence the level of stress in wolves and that
human activity is less important.
Acknowledgements
This study was part of the Bieszczady Wolf Project funded
by the Polish National Committee for Scientic Research
(KBN 6P04F 006), budget of the Museum and Institute of
Zoology (Polish Academy of Sciences), SAVE — Wildlife
Conservation Fund and scholarships of the “Allgemeines
Promotionskolleg” of the Ruhr University of Bochum and
the Ruhr University Research School. We thank anonymous
reviewers for useful comments.
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