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Abstract Predation involves more than just predators consuming prey. Indirect effects, such as fear responses caused by predator presence, can have consequences for prey life history. Laboratory experiments have shown that some rodents can recognize fear in conspecifics via alarm pheromones. Individuals exposed to alarm pheromones can exhibit behavioural alterations that are similar to those displayed by predator-exposed individuals. Yet the ecological and evolutionary significance of alarm pheromones in wild mammals remains unclear. We investigated how alarm pheromones affect the behaviour and fitness of wild bank voles (Myodes glareolus) in outdoor enclosures. Specifically, we compared the effects of exposure of voles living in a natural environment to a second-hand fear cue, bedding material used by predator-exposed voles. Control animals were exposed to bedding used by voles with no predator experience. We found a ca. 50% increase in litter size in the group exposed to the predator cue. Furthermore, female voles were attracted to and males were repelled by trap-associated bedding that had been used by predator-exposed voles. Movement and foraging were not significantly affected by the treatment. Our results suggest that predation risk can exert population-level effects through alarm pheromones on prey individuals that did not encounter a direct predator cue.
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Scientific REPORtS | (2018) 8:17214 | DOI:10.1038/s41598-018-35568-0
Exposure to Chemical Cues from
Predator-Exposed Conspecics
Increases Reproduction in a Wild
M. Haapakoski
1, A. A. Hardenbol2,3 & Kevin D. Matson2
Predation involves more than just predators consuming prey. Indirect eects, such as fear responses
caused by predator presence, can have consequences for prey life history. Laboratory experiments
have shown that some rodents can recognize fear in conspecics via alarm pheromones. Individuals
exposed to alarm pheromones can exhibit behavioural alterations that are similar to those displayed
by predator-exposed individuals. Yet the ecological and evolutionary signicance of alarm pheromones
in wild mammals remains unclear. We investigated how alarm pheromones aect the behaviour and
tness of wild bank voles (Myodes glareolus) in outdoor enclosures. Specically, we compared the
eects of exposure of voles living in a natural environment to a second-hand fear cue, bedding material
used by predator-exposed voles. Control animals were exposed to bedding used by voles with no
predator experience. We found a ca. 50% increase in litter size in the group exposed to the predator cue.
Furthermore, female voles were attracted to and males were repelled by trap-associated bedding that
had been used by predator-exposed voles. Movement and foraging were not signicantly aected by
the treatment. Our results suggest that predation risk can exert population-level eects through alarm
pheromones on prey individuals that did not encounter a direct predator cue.
In 1961, Robert Ardrey wrote, “sex is a sideshow in the world of the animal, for the dominant color of that world
is fear”1. Of course, the purveyors of fear in the world of the animal are predators, and their ultimate threat comes
in the form of killing and eating prey. However, predation involves more than just predators consuming prey2,3.
In fact, when facing only the risk of predation, prey can exhibit an array of anti-predatory responses, which have
arisen through co-evolution with predators4. Such awareness or perception of predators by prey can aect the
physiology, body condition, and reproduction in many prey species (reviewed by Lima 19982). At the population
level, the eects of fear of predation may be of the same magnitude or greater than the actual act of predation5.
While predation fear responses (sometimes placed under the umbrella of “stress”) are oen seen as negative or
costly for the prey, these responses can be adaptive6. For example, fear of predation can result in physiological
changes in pregnant females that prepare ospring for life under high predation pressure7.
Early predator detection is necessary for prey to maximize their own tness8. In many vertebrate predator-prey
systems, the key cue of predator presence is scent9. Prey can assess predation risk by eavesdropping on predator
scent marks le for intraspecic communication. An adaptive prey response requires recognition and correct
interpretation and response to the cue. While fresh cues may suggest predation is imminent and a change in
behaviour is needed, old cues should be ignored10. Behavioural responses by prey to avoid predation can include
reduced activity or escape to another, oen lower-quality, habitat11,12. ese responses can translate to changes in
foraging behaviours, decreased body condition13, and ultimately reduced tness. In females, poor body condition
can diminish ospring quantity and quality14; in males, predator avoidance during the mating season may lead
to missed mating opportunities15 or breeding delays. Hence, if one looks back at the world described by Andrey,
indirect interactions between the sideshow of sex and the world of fear are evident.
1Konnevesi Research Station, Department of Biological and Environmental Science, P.O. Box 35, FI-40014, University
of Jyväskylä, Jyväskylä, Finland. 2Resource Ecology Group, Environmental Sciences Department, Wageningen
University, Droevendaalsesteeg 3a, 6708 PB, Wageningen, The Netherlands. 3Present address: School of Forest
Sciences, University of Eastern Finland, P.O. Box 111, FI-80101, Joensuu, Finland. Correspondence and requests for
materials should be addressed to M.H. (email: marko.j.haapakoski@jyu.)
Received: 22 June 2018
Accepted: 1 November 2018
Published: xx xx xxxx
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Many organisms, including vertebrates, invertebrates, and plants, respond to predation threats by producing
alarm signals. (reviewed by Verheggen et al.)16. For example, some rodents are known to produce alarm phero-
mones (AP), chemical cues that warn conspecics about possible dangers17,18. Neurobiological and psycholog-
ical experiments show that individuals can recognise stressed conspecics from the AP le behind following
a stressful situation19. e receivers of this indirect signal increase their vigilance and risk assessment behav-
iour, i.e., change their behaviour in ways that are similar to individuals that encountered a direct predator cue18.
Furthermore, murine AP is biochemically related to predator-produced scent cues common to most carnivores20;
both contain similar sulfur-containing volatiles21.
In general, a role for fear in ecology of animals is recognized22; however, the ecological and evolutionary sig-
nicance of AP as a fear cue in wild mammals remains unclear. To the best of our knowledge, only two studies
have investigated the role of AP in wild mammals. In an experimental indoor arena, Cabrera voles (Microtus
cabrerae) avoided areas with the scent marks of conspecics that experienced various predation risks23. ese
predation risks included handling (simulation of capture), audio playbacks of an avian predator, and visual con-
tact with a mammalian predator. In the same study, but under eld conditions, Cabrera voles tended to decrease
their activity near sites treated with AP. In another study, black-tailed deer (Odocoileus hemionus columbianus)
produced AP when disturbed or alarmed24. In the presence of AP, female black-tailed deer became more alert
and le the site more oen than in the presence of control odours, odourless air, or deer urine24. Most other AP
studies have used lab-strain rodents and unnatural stimuli, such as electric foot-shocks17,18.
Wild rodents are ideally suited for studying the ecology of predator cues. ese animals have a highly devel-
oped olfactory system, and odours play a key role in their behavioural decision-making25. We tested the hypoth-
esis that wild bank voles (Myodes glareolus), a model prey species in many studies of predator-prey interactions,
would show antipredator-like responses when exposed to AP11,26, since the murine AP shares structural simi-
larity with predator scent21. More specically, we made the following predictions. First, bank voles are expected
to decrease movement in response to AP11. Second, voles are expected to be more fearful and thus forage less
eciently in response to AP15,27. ird, these behavioural and ecological responses should lead to poorer body
condition, which in turn should lead to a smaller proportion of breeding females and a smaller litter sizes on
Litter size was signicantly greater by 1.9 ± 0.7 pups in AP enclosures compared to control enclosures (p = 0.013;
Table1, Fig.1). Previous pregnancy and the interaction between treatment and previous pregnancy were not
signicant. None of these three terms, which also comprised the pregnancy model, accounted signicantly for
variation in the proportion of pregnant females (Table1). Treatment did not have a signicant eect on the female
survival (control = 55 ± 12%, treatment = 50 ± 10%; Table1).
e interaction between treatment and sex signicantly aected bait-less trapping (p = 0.019; Table1, Fig.2a).
Females were trapped more and males less in the presence of AP. Neither treatment, sex, nor their interaction
signicantly aected trappability (Table1). Only sex and the number of trapping instances signicantly aected
movement: females moved less than males (p = 0.012, females 70 ± 13 m2, males 177 ± 45 m2; Table1, Fig.2b)
and more trapping instances correlated with greater movement areas (p < 0.001; 63.48 ± 11.33 m2 per trapping
instance, Table1).
Litter size Pregnant
females Female survival
d.f. χ2Pχ2Pχ2P
Treatment*Pregnancy 1 2.31 0.129 1.36 0.243 —
Treatment 1 6.23 0.013 0.55 0.456 0.11 0.741
Pregnancy 1<0.01 0.980 2.90 0.089
Movement area Trappability Bait-less trapping
Treatment*Sex 1 0.24 0.627 1.57 0.210 5.46 0.019
Treatment 1 0.11 0.740 0.87 0.350 —
Sex 1 6.34 0.012 0.42 0.516 —
Trappings 1 36.93 <0.001 x x x x
Giving Up Density
Treatment*Time 1 2.16 0.142
Treatment 1 0.10 0.749
Time 1 0.34 0.561
Vegetation 1 67.51 <0.001
No. of Voles 1 0.41 0.520
Table 1. A detailed summary of statistical analysis with the χ2 and P values for all xed factors.
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GUD was signicantly higher in areas where the vegetation was open compared to grass-covered areas (by
on average 38.8 ± 3.8 seeds, p < 0.001; Table1, Fig.3). None of the other model terms accounted signicantly for
variation in GUD.
Conspecic alarm cues impact tness. In the current study, exposure to conspecic AP increased litter
size by about 50% compared to control treatment. is result supports our hypothesis that voles can receive dif-
ferent information from predator-exposed and unexposed male conspecics, and it suggests that female and male
voles can respond dierently to this information. Chemical cues can serve as an important means of communica-
tion among animals, both within and between species. Some rodents AP to warn conspecics about possible dan-
gers17,18,23. AP produced by mice are known to be molecularly similar to chemical cues produced by predators21.
e observed eects of AP in voles may also be rooted in biochemical similarity, for example, to other compounds
that signal danger natural settings, but this possibility requires further investigation.
Figure 1. Litter size ± SE in the control treatment, which received bedding materials of non-predator-exposed
male bank voles, and in the AP treatment, which received beddings materials of predator- exposed male voles.
Sample size inside the bars. Letters denote statistically signicant dierence between bars.
Figure 2. Bait-less trapping proportion ± SE (a) and movement area of voles ± SE (b) in AP and control
enclosures. White bars represent females; grey bars represent males. Sample size inside the bars. Letters denote
statistically signicant dierence between bars.
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Increased investment in reproduction by female bank voles faced with a predator cue is in line with several
classic hypotheses concerning residual reproductive value29 and terminal investment30. We conducted the exper-
iment late in the breeding season with the prediction that females should heavily invest resources into current
reproduction. Such investment is expected because future reproduction opportunities are improbable for several
interacting reasons. e most important are likely the approach of winter and poor overwinter survival of females
that have already reproduced31, combined with high overall mortality risks from predation32. In comparison to
AP females, females in the control treatment did not increase their litter size and were still prepared to produce
one or two litters during ongoing breeding season. Reproduction should continue as long as there are sucient
resources for doing so; bank vole breeding season in our latitude usually lasts until September33. e litter size
eect of AP could be mediated by males (e.g., via sperm quality), but examples of such mechanisms are not cur-
rently described in the literature. Simple physiological mechanisms could allow bank voles females to increase
litter size: bank voles are induced ovulators34, meaning that females can produce more pups by mating more with
one or several dierent males35. Many animals, including sticklebacks (Gasterosteus aculeatus)36 and some song-
birds, have been found to increase investment in reproduction when predation risk is experimentally increased37.
However, such increases are not universal. In some cases, cessation or delay of reproduction may result from high
levels of predation. is phenomenon has been previously described in bank voles in laboratory studies38,39 and
in studies with challenging ecological conditions (e.g., variable food availability and predation risk) during the
rst reproductive window aer winter26. Reduced reproduction, however, is expected to be an unlikely alternative
late in a breeding season. Quantifying fear eect on prey tness has been criticized because it is dicult to assess
the magnitude of experimenter-induced predator cues40. While ecological experiments can deviate from natural
conditions in some way, we factored in vole biology as much as possible (e.g., by using naturally occurring sex
ratios and population densities and by adding scent cues produced by a single unique male per enclosure41.
Males and females respond dierently to scent cues. Chemical cues produced by prey faced with a
predator might communicate more than just alarm. For example, male lab mice exposed to either a predator (cat
urine) or a predator/competitor (rat urine) produce cues that make themselves more attractive to female mice42,43.
e element of our study involving trapping without seeds as bait oered support for this idea: female voles were
more attracted to traps in the AP enclosures, where those females were presented with bedding used by the male
voles that had encountered a predator. In contrast, male voles avoided traps in the AP enclosures and were more
frequently found in traps in the control enclosures. e fact that only male voles had been used to produce both
types of bedding materials in our study may be useful in linking our results to possible mechanisms. One possible
candidate attracting females to the bedding materials from predator-exposed AP males could be the major uri-
nary protein (MUP). Mice are able to regulate the total amount of MUP in their urine during social contacts, and
female mice advertise their reproductive state by varying the concentration of MUP during the oestrous cycle44.
MUP can function as a pheromone and stimulate sexual attraction45 and oestrus in female mice46. e chemical
can also promote aggressive behaviour in male mice47 and extend the longevity of olfactory cues that they leave
behind48. One or more urine components (e.g., MUP), and not urine itself, most likely drive our observed dier-
ences. Control and AP bedding was collected aer being used by a vole for about 48 hours, and since bank voles
are assumed to urinate roughly constant rate, both control and AP bedding should have been thoroughly contam-
inated with urine and feces. Furthermore, prairie voles (M. ochrogaster) and woodland voles (M. pinetorum) urine
mark at the same rate with and a without predator cue49.
If the bedding material used by predator-exposed male voles contained AP that resembled direct odour
cues from predators themselves, then we predicted trappability would be higher under control conditions (i.e.,
unstressed male used bedding), since the bedding materials were distributed in the chimneys near the traps.
However, neither the two groups nor the two sexes diered signicantly in trappability when baited traps were in
use. e absence of an experimental eect might be due to the highly attractive food rewards (i.e., energetically
rich sunower seeds) that were used as a bait in the traps. During the breeding season, voles, especially female
ones, have high energy demands, which may be met (at least partially) by consuming the sunower seed bait.
Additionally, the voles in this study may have adjusted to the relatively constant presence of AP, which was present
Figure 3. Giving up density (GUD) ± SE measured as number of seed le in the seed-trays in AP and control
enclosures. All seed trays contained 80 sunower seeds mixed with two liters of sand. White bars represent
GUD boxes in dense vegetation; grey bars represent GUD boxes in open areas. Sample size inside the bars.
Letters denote statistically signicant dierence between bars.
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when the voles were added to the enclosure and regularly refreshed until the pregnant females were removed.
Ultimately, a behavioural shi favouring the gain of food rewards over the risks of predation may have occurred;
such an adjustment matches well with the risk allocation hypothesis50.
We also hypothesised that the addition of bedding materials from predator-exposed males would restrict the
movement area of voles in the experimental enclosures51. Decreasing movement area should eectively decrease
the risk of encountering and being killed by a predator4. However, we were unable to document an eect of our
experimental treatment on movement. Our results contrast with those of an earlier experiment on Cabrera voles,
which tended to decrease activity near AP patches23. In our enclosures, movement may have been suppressed
by the avian predators to such an extent that voles were unable to further reduce their movement in response
to our AP treatment52. Overall low trappability (around 50%) compared to previous summer experiments (see
for example Haapakoski et al., 201353,54) supports this nding. Not surprisingly, males were moving more than
females regardless of predation risk. Males need to move over several females territory in search of potential
mates to increase their tness55.
Perceived predation risk is expected to increase at the expense of foraging56. us, we expected a higher GUD in
enclosures with bedding materials from the predator-exposed voles; however, we were unable to document dier-
ences in GUD between treatment and control enclosures. GUD was higher in open areas compared to more densely
vegetated areas in our experimental enclosures, suggesting that the GUD method worked. With voles seeming to
avoid the avian predation risk associated with foraging in open areas, this result resembles predator facilitation57. In
fact, previous research suggests that eld voles (M. agrestis) prefer open habitats when facing a mammalian predator
(least weasel) and densely vegetated areas when facing an avian predator (kestrel falcon, Falco tinnunculus)52.
e pregnancy rate was similar in the experimental and control enclosures. Under experimentally increased
predation risk in laboratory studies, bank voles have been observed to suppress breeding with females actively
avoiding copulation38. Yet breeding suppression has never been reported during the peak of the breeding season
in the eld58, and the breeding suppression in the lab might be a laboratory artefact. Breeding suppression or
avoidance might only occur when resources are limited26,59.
e survival of females did not dier between treatment and control groups. Previous studies of bank voles
have not found an eect of predation risk on survival during summer58,60, but an inverse relationship between
predation risk and survival in winter has been reported26. In that case, the assumption was that reduced survival
was a consequence of reduced foraging. In our current study, foraging and movement were not signicantly
aected by the experimental treatment, which might explain the uniformity in survival.
e eects of predation are multifaceted and extend far beyond the death and consumption of prey. For example,
we found that bedding materials used by predator-exposed male voles contained cues AP that could exert strong
life-history eects in eld settings. Specically, indirect eects of predation risk, which here was communicated
by the presence of the scent cues, translated to 1) increased litter size and 2) an interaction with sex that aected
the attractiveness of traps. us, AP can impact the tness and behaviour of animals. Continued study of these
cues and others in both eld and lab settings will contribute to a more nuanced understanding of interactions
between predators and prey and between the sideshow of sex and the world of fear.
Material and Methods
Experimental setup. We conducted our experiment in twelve uncovered outdoor enclosures (0.25 ha per
enclosure, see Haapakoski et al.26 for complete details) and in the laboratory of Konnevesi Research Station in cen-
tral Finland (62°37N, 26°20E). Experimental status (AP vs. control, six enclosures each) was randomly assigned.
In each enclosure, 25 multiple capture traps (Ugglan Special Nr. 2, Grahnab, Hillerstorp, Sweden) were arranged
in a 5 × 5 grid with 10 m intervals between traps (Fig.4). e traps were covered with bottomless metal chimneys
(40 × 40 × 50 cm3) on which a metal lid was placed to protect voles in the traps from direct sunlight and rain.
Figure 4. Enclosures used in our experiment were 2500 m2, surrounded by a steel wall, and contained 25 traps
that had a distance of 10 m between them. At four trapping locations (circles) in each enclosure, we placed GUD
boxes, two of which had a non-altered dense vegetation surrounding the boxes and two had an mown open
areas surrounding the boxes.
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Study animals. e bank vole (Myodes glareolus) is a small common boreal microtine rodent. Vole popula-
tions cycle in Scandinavia, and specialist predators play a large role in driving these cycles61. One such predator
is the least weasel (Mustela nivalis nivalis), which specializes in hunting small mammals. Due to its small size,
the weasel is able to hunt in the burrows of voles, leaving few safe places for the voles to escape predation51. As
a consequence of high predation pressure in the wild, antipredator behaviours and physiology are important in
bank voles.
In the study area, the breeding season of bank voles usually extends from the end of April to September.
During the breeding season, female bank voles are strictly territorial and need an exclusive territory to breed.
e territories of breeding males usually overlap with the territories of several females62. Bank voles are easy to
trap in the eld and maintain and breed in laboratory, thereby making the species an ideal subject for the study
of predation risk.
We used sexually mature bank voles that were either enclosure- or laboratory-born. These voles were
rst-generation descendants of wild bank voles that had been originally trapped in the Konnevesi region. Prior
to the experiment, these voles were housed individually under standard conditions60. Voles grouped together in
an outdoor enclosure were unrelated. Enclosure groups were balanced based on vole body size and number of
females who had previously given birth.
Production of alarm pheromone and control bedding materials. To produce AP and control bed-
ding materials, we used six pairs of bank vole brothers from litters produced by six dierent parental pairs. Each
of these twelve male ospring was placed individually into a standard rodent cage containing 3 l of wood shavings.
A randomly selected vole from each parental pair was used for AP production, and these six were housed in the
same room. e six that produced the control bedding were housed in a separate room.
In order to induce AP production, we exposed the six AP voles to a weasel. For the rst two weeks, a caged
weasel was placed daily on top of the cage of each AP vole for 2 minutes. Weasel cage was equipped with solid
bottom in order to prevent weasel urine and feces mixing in to AP bedding. Aer this procedure, we captured
each AP vole with our hands and released it back into its cage to simulate predator capture. e AP production
procedure was changed two weeks aer the start of the experiment in order to minimize stress for the weasel.
With this new method, a bank vole was rst captured by hand from its cage and placed individually inside a small
wire mesh cage, which was then placed inside the weasel’s cage for 2 minutes. Aerwards, each vole was released
back into its original cage. e weasel used in this procedure was housed under standard conditions60 in a cage
that was kept in a dierent building from all voles.
About 48 h old bedding material originating from one specic male vole (either AP or control, see below)
was assigned to a specic outdoor enclosure for the duration of the experiment. In all enclosures, 0.1 l of bedding
material was placed in each trap chimney and around the giving-up density tray (see below). Bedding material
(AP or control) was placed at the start of the experiment and replaced every other day during the experiment.
e experiment started with the spreading AP and control bedding materials into the enclosures at the end of
July 2015. On the same day bedding materials were added to an enclosure, four females were released to allow for
habituation to the environment and its cue of indirect predation risk or not. ree days aer adding the females,
four males were released into each enclosure. In order to determine female survival, the proportion of pregnant
females, parturition date, and litter size, females were trapped and moved to the laboratory around 18 days aer
initiation of the experiment.
Giving-up density. To measure the voles’ perception of predation risk in each enclosure, giving-up density
(GUD56) was measured using four boxes (20 × 20 × 15 cm3 covered with a transparent lids) accessible via two
2 cm diameter entrances. Two GUD boxes were placed in open vegetation, and two were placed under the grass
and were thus more protected from avian predators (see Fig.1). Open vegetation areas were created by mowing
the grass in a 1 m diameter area where the GUD boxes were placed. GUD boxes contained 80 sunower seeds
mixed with 2 L of sand. GUD, quantied here the as the number of seeds remaining, was recorded aer two
consecutive 24 hr periods, which started on the aernoon of day 4 and ending on the aernoon of day 6. If voles
perceive that predation risk outweighs foraging gains, then they will give up searching for seeds, and GUD will be
high. In situations where they feel safer, the voles will spend more time searching for the diminishing seeds, and
GUD will be lower.
Movement and trappability. To quantify movement area, all traps were activated and regularly checked,
and the location of any captured individual was recorded before release at the point of capture. Movement area
trapping was conducted on days 10 (evening only), 11 and 12 (both morning and evening), and 13 (morning
only). e rst movement area trapping (day 10 evening) was conducted without bait (hereaer “bait-less trap-
ping”) to measure the eect of AP treatment, since the food reward might have otherwise lured voles into traps.
In bait-less trapping traps were activated immediately aer distributing AP and control bedding materials in
the chimneys. Bait-less trapping is possible since voles are using trap chimneys and traps as hiding places when
traps are deactivated. Movement areas (100% convex polygons) were calculated from the trapping data with the
program Ranges VI (Anatrack ltd. Wareham, UK). Movement area calculations excluded individuals that were
caught only once. Trappability was calculated per vole as the number of captures divided by six (the number of
baited trap checks). Bait-less trapping, used for the calculation of the movement areas but not for trappability.
At the start of the intensive trapping period, two individuals were found dead in one trap in one enclosure; these
individuals were replaced.
Statistical analysis. R version 3.2.263 with package lme464 was used to perform linear mixed eects anal-
yses, which accounted for the eight voles per enclosure. We used log-likelihood ratio tests and χ2 statistics to
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evaluate statistical signicance (α = 0.05). Litter size was analysed by using a model that included treatment (AP/
control), previous pregnancy (yes/no) and the interaction between the two. Movement area, trappability and
bait-less trapping were analysed by using a model that included treatment (AP/control), sex (male/female), and
the interaction between the two. In addition, the number of captures per individual was used as a covariate in the
movement area analysis to account for dierences in movement area due trappability. Female survival and female
pregnancy were analysed with generalized linear mixed models (GLMM) with binomial error distribution. In the
analysis of survival, treatment was used as a xed factor. e pregnancy model, like the litter size model, included
as xed treatment, previous pregnancy (yes/no) and the interaction between the two. GUD was analysed using
repeated measures analyses because GUD was measured on two days. For GUD, models included treatment, time
(day one/two), their interaction, vegetation (open/covered), and the minimum number of voles known to be
alive in the enclosure. Enclosure identity was included as a random factor in all analyses. Residuals were visually
inspected for deviations from homoscedasticity and normality. In the movement area analysis, data were cube
root transformed to meet the assumptions. For unknown reasons, four voles in one control enclosure died during
the movement area trapping. erefore, we excluded this enclosure from all analyses of data from aer the point
of mortality. (Data from this enclosure were used in analyses of GUD and bait-less trapping, which were collected
before the mortality event.) We also excluded two individuals (IDs 4885 and 4886) in an AP treatment enclosure
from our movement analyses, since their tag numbers were indistinguishable in the eld.
Ethics. All work was conducted according Finnish legislation with the animal experimentation permission
from Jyväskylä University No. ESAVI/6370/04.10.07/2014.
Data Accessibility
Data will be deposited to Dryad Digital Repository aer acceptance.
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We thank the mammalian ecology and life histories research group lead by Prof. Hannu Ylönen for providing
all of the facilities for our study and commenting the manuscript. Technicians at the Konnevesi Research Station
did an excellent job of maintaining the enclosures. Alexandra iel’s help in the lab and in the eld was valuable.
Ines Klemme provided comment to the manuscript. Financial support came from the Jenny and Antti Wihuri
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Author Contributions
M.H. and A.A.H. conceived the study; A.A.H. and M.H. carried out eld experiment; M.H., K.D. M. and A. A. H.
performed statistical analyses; and M.H., K.D.M. and A.A.H. wrote the manuscript.
Additional Information
Competing Interests: e authors declare no competing interests.
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... These factors can also impact each other (Pianka, 1976;Koskela & Ylönen, 1995), so the behavioural response of animals exposed to any combination of these variables may be different than that observed during exposure to only one of these variables. Three hypotheses describe how prey behaviour may be altered by food deprivation or the threat of predation: the safety in numbers hypothesis (SIN) (Daan & Slopsema, 1978;Gerkema & Verhulst, 1990;Jochym & Halle, 2012), the terminal investment hypothesis (TIN) (Clutton-Brock, 1984;Gliwicz, 2007;Zhang et al., 2008;Haapakoski et al., 2018;Gu et al., 2020), and the avoidance hypothesis . The SIN hypothesis predicts that aggregation of prey provides protection from predators by diluting the risk of predation for each individual (Daan & Slopsema, 1978;Jochym & Halle, 2012). ...
... Similarly, the TIN hypothesis also predicts an increase in conspecific association when animals experience a threat to their survival, but only within the context of mating. Specifically, the TIN hypothesis predicts that prey will increase their reproductive investment when their future reproductive success is compromised due to a threat to their survival (Clutton-Brock, 1984;Gliwicz, 2007;Zhang et al., 2008;Haapakoski et al., 2018;Gu et al., 2020). Finally, the avoidance hypothesis predicts that animals will cease interaction with conspecifics when facing the stress of predation or food deprivation Jochym & Halle, 2012). ...
... However, elk congregations only occurred in the absence of wolves (Canis lupus), suggesting that the risk, or cost, of predation may be too great for congregations of foraging elk when wolves were present. Research on the impact of predation pressure on mating, grouping, foraging, and the interactions of these variables on prey behaviour is vital to the understanding of the complex behavioural responses of prey during a predation threat (Pianka, 1976;Koskela & Ylönen, 1995;Bleicher et al., 2018Bleicher et al., , 2019aBleicher et al., , b, 2020Haapakoski et al., 2018;Beckmann et al., 2022). ...
The risk of predation and food deprivation may alter the degree to which animals associate with conspecifics. We examined if food deprivation, the risk of predation, or simultaneous exposure to both altered meadow voles’ preference for odour cues in a way that adheres to the terminal investment, safety in numbers, or avoidance hypotheses. Satiated or food-deprived meadow voles were given the choice to investigate either opposite-sex conspecific bedding, same-sex conspecific bedding, clean bedding, or self-bedding when exposed to mink urine or olive oil. Mink urine and food deprivation did not impact the amount of time meadow voles spent with each type of bedding, but meadow voles did begin investigating more quickly when experiencing either or both stressors. However, food deprivation and mink urine did not have an additive impact on any measured variable. Further research is needed to determine if the terminal investment hypothesis is the hypothesis that best describes the mating behaviour of meadow voles facing one or multiple stressors.
... Alternatively, the production of larger eggs by mothers experiencing an elevated death risk may be a manifestation of terminal investment. Females who perceive a high risk of being killed may invest more resources in current breeding attempts because of decreased chances of survival in the next reproductive event (Haapakoski et al. 2018;Sievert et al. 2019). ...
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Predators affect prey by killing them or inducing changes in their physiology and behaviour through a fear effect associated with predation risk. In birds, perceived predation risk influences reproductive decisions, such as the reduction of parental investment in offspring during both egg production and nestling rearing. Visual and vocal cues of predator presence have been widely used to test the direct effects of predation risk. However, few studies have examined the indirect cues of predator activity such as dead avian prey or their remains. In this study, for the first time, we experimentally studied whether piles of feathers, simulating the remains of avian prey, induce changes in the reproductive decisions of adult birds. Before and during egg laying, great tit, Parus major, pairs were exposed to piles of bright down and cover feathers from domestic goose (treatment), woodchips (procedural control), or were not exposed (control). Our experiment affected maternal investment in individual eggs, but did not influence other reproductive parameters. Females from the treatment group laid larger and more asymmetrical (pointed) eggs than control females. Moreover, females from the procedural control group laid larger eggs than those from the control group, but without differences in egg shape. However, the eggs from the treatment and procedural control groups did not differ. This indicates that great tit females can perceive feathers and woodchips as informative cues, such as potential predation risk or habitat suitability, or as novel items in the environment. Importantly, females respond to such cues by changing their maternal investment in eggs, which may result from an adaptive mechanism aimed at increasing offspring fitness in the face of specific environmental conditions experienced by a female. Our study contributes to the understanding of how female songbirds adjust their maternal reproductive investment in response to publicly available social and environmental cues.
... Researchers in these types of projects are not seeking answers to pathological questions requiring physiological data. Instead, they are examining responses such as: reduced foraging time or effort (Mills and Wajnberg, 2008), the amount of food left in a food patch (GUD; giving up density (Haapakoski et al., 2018)), or the amount of time spent in an agriculture area that is 'guarded' with a predator scent. Such responses might simply be due to repugnance, distaste, or aversion and have little relation to actual fear (Nolte et al., 1994;Kimball and Nolte, 2006). ...
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Mismatches between highly-standardized laboratory predatory assays and more realistic environmental conditions may lead to different outcomes. Understanding rodents’ natural responses to predator scents is important. Thus, field studies on the same and related species are essential to corroborate laboratory findings to better understand the contexts and motivational drives that affect laboratory responses to predator scents. However, there are too few field assays to enable researchers to study factors that influence these responses in genetically variable populations of wild rodents. Therefore, we placed laboratory-style chambers and remote-sensing devices near multiple colonies of two species of wild mice (Apodemus agrarius and Apodemus flavicollis) to test dual-motivational drives (appetitive and aversive) in a ‘familiar’, yet natural environment. A highly-palatable food reward was offered daily alongside scents from coyotes, lions, rabbits, and both wet and dry controls. In all but two instances (n = 264), animals entered chambers and remained inside for several minutes. Animals initiated flight twice, but they never froze. Rather, they visited chambers more often and stayed inside longer when predatory scents were deployed. The total time spent inside was highest for lion urine (380% longer than the dry control), followed by coyote scent (75% longer), dry control and lastly, herbivore scents (no difference). Once inside the chamber, animals spent more time physically interacting with predatory scents than the herbivore scent or controls. Our findings support the common assumption that rodents fail to respond as overtly to predatory scents in the field compared to what has been observed in the laboratory, possibly due to their varying motivational levels to obtain food. More time spent interacting with scents in the field was likely a function of ‘predator inspection’ (risk assessment) once subjects were in a presumed safe enclosure. We conclude this sort of chamber assay can be useful in understanding the contexts and motivational drives inherent to field studies, and may help interpret laboratory results. Our results also suggest more attention should be given to subtle behaviors such as scent inspection in order to better understand how, and when, environmental stimuli evoke fear in rodents.
... As expected, we found that male disturbance odors can trigger an avoidance response by females, demonstrating that disturbed male odors of K. odontotarsus can act as an alarm cue for conspecifics. Many vertebrates use signals or cues to warn con-or heterospecifics of potential predation risk (Schmitt et al. 2016;Oliveira et al. 2017;Haapakoski et al. 2018;Suzuki 2018). For example, Ruiz-Monachesi and Labra (2020) reported that conspecific distress calls triggered a reduction in activity in weeping lizard (Liolaemus chiliensis). ...
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Mate choice in frogs depends heavily on acoustic communication, and females in many species possess an inherent preference for longer and/or more complex calls. Recently, it has become clear that conspecific chemical cues can also be useful in attracting potential mates in anuran species. However, how conspecific chemical cues influence mate choice decisions when paired with acoustic signals remains less clear. In the present study, we conducted three experiments to investigate whether and how male odor cues affect female choice decisions in the absence or presence of acoustic signals in serrate-legged small treefrogs (Kurixalus odontotarsus). We found that disturbance odors alone can trigger avoidance behavior of females, suggesting that the odors of disturbed K. odontotarsus can act as an alarm cue for conspecifics. Females also avoided disturbance odors in the presence of advertisement calls, except for calls with five notes. In addition, females preferred calls with two notes to calls with four notes when the latter was paired with disturbance odors. In contrast, female choice decisions were not affected by the odors of undisturbed males, either in the absence or the presence of advertisement calls. Our results indicate that disturbance odors can influence, or even reverse, mate choice by females for acoustic signals. Significance statement In addition to acoustic signals, chemical cues can also be used in close-range communication in some anuran species. In the present study, we investigated how female Kurixalus odontotarsus use male chemical cues and/or acoustic signals during the mate choice process. We demonstrate that male disturbance odors can act as an alarm cue to trigger the avoidance behavior of females. Furthermore, this alarm cue can even reverse mate choice by females for acoustic signals. Our study highlights the importance of integrating multisensory signals and cues in mate choice decisions and has important implications for understanding chemical communication in anurans.
... The general consensus is that extended exposure to predator odors can lead to breeding suppression, as demonstrated in grey-sided voles (Myodes rufacanus) and field voles (Microtus agrestis), and lead to lower offspring birth weights (Koskela and Ylönen 1995;Perrot-Sinal et al. 2000;Fuelling and Halle 2004). Conversely, Haapakoski et al. (2018) demonstrated that secondary exposure of wild bank voles (Myodes glareolus) to bedding used by individuals who had been directly exposed to predator odors increased the former's subsequent litter size. Sievert et al. (2019) also found that in bank voles, exposure to predator odors and conspecific alarm pheromones led to higher pregnancy rates and increased offspring birth weights. ...
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In rodents, defensive behaviors increase the chances of survival during a predator encounter. Observable rodent defensive behaviors have been shown to be influenced by the presence of predator odors and nearby environmental cues such as cover, odors from conspecifics and food availability. Our experiment tested whether a predator scent cue influenced refuge preference in meadow voles within a laboratory setting. We placed voles in an experimental apparatus with bedding soaked in mink scent versus olive oil as a control across from four tubes that either contained (a) a dark plastic covering, (b) opposite-sex conspecific odor, (c) a food pellet, or (d) an empty, unscented space. A three-way interaction of tube contents, subject sex, and the presence of mink or olive oil on the preference of meadow voles to spend time in each area of the experimental apparatus and their latency to enter each area of the apparatus revealed sex differences in the environmental preference of meadow voles facing the risk of predation. The environmental preference of female, but not male, meadow voles was altered by the presence of mink urine or olive oil. A similar trend was found in the latency of males and females to enter each area of the experimental apparatus. These differences suggest that each sex utilizes different methods to increase their fitness when experiencing a predation risk. The observed sex differences may be explained by the natural history of voles owing to the differences in territorial range and the dynamics of evasion of terrestrial predators.
... Predation risk impacts the activity of mammals, particularly rodents, with an increase in perceived risk causing animals to alter their behavior to reduce that perceived risk (Lima and Dill 1990;Kats and Dill 1998;Caro 2005). These behavioral alterations are diverse and include changes in habitat usage (Borowski and Owadowska 2001), foraging activity (Koivisto and Pusenius 2003;Hinkelman et al. 2012), activity levels (Borowski 1998), defensive behaviors (Dielenberg and McGregor 2001;Ayon et al. 2017), and reproduction (Apfelbach et al. 2005;Haapakoski et al. 2018). To assess risk, mammals rely on a variety of indirect and direct predator cues (Orrock and Danielson 2009;Sivy et al. 2011;Crego et al. 2018). ...
Rodents use direct and/or indirect cues of predators to assess predation risk. The responses to these cues are well studied with regard to mammalian predators, but less understood with regard to reptilian predators. These responses are of particular importance in tropical and subtropical regions where reptile diversity is high and the likelihood of establishment of invasive reptilian predators also is high. We hypothesized that rodents would respond to direct scent cues of snake predators and that rodents would show greater aversion to scents of native snake predators than non-native snake predators. To assess this, scents of three snake species, two native and one non-native, and a non-snake control odor were distributed in Sherman live traps using a randomized block design. A total of 69 rodents representing four species were captured. Responses varied by species reinforcing that some species utilize indirect cues to assess predation risk, whereas others use direct cues. Moreover, one species (Neotoma floridana) showed a preference for non-native Python scent, indicating a lack of the appropriate anti-predator behavior, suggesting that some native rodents are more at risk of attack from invasive snakes than other native rodents.
... After perceiving increased predation risk, multiple mechanisms and adaptations by prey animals are possible, from simple immediate behavioural responses to long-term physiological or even intergenerational adaptations (Abrams 2000). Anti-predatory behaviours employed in prey range from simple avoidance of high-risk areas (Ferrero et al. 2011;Clinchy et al. 2013;Pérez-Gómez et al. 2015) and freezing to decrease detectability (Wallace and Rosen 2000;Sundell and Ylönen 2004), over changes in vigilance and foraging (Brown 1999;Ylönen and Brown 2007;Embar et al. 2011), to drastic changes in the reproductive behaviours (Ylönen and Ronkainen 1994;Sih 1994;Mappes and Ylönen 1997;Mönkkönen et al. 2009;Haapakoski et al. 2012Haapakoski et al. , 2018Sievert et al. 2019). ...
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Chemical communication plays an important role in mammalian life history decisions. Animals send and receive information based on body odour secretions. Odour cues provide important social information on identity, kinship, sex, group membership or genetic quality. Recent findings show, that rodents alarm their conspecifics with danger-dependent body odours after encountering a predator. In this study, we aim to identify the chemistry of alarm pheromones (AP) in the bank vole, a common boreal rodent. Furthermore, the vole foraging efficiency under perceived fear was measured in a set of field experiments in large outdoor enclosures. During the analysis of bank vole odour by gas chromatography–mass spectrometry, we identified that 1-octanol, 2-octanone, and one unknown compound as the most likely candidates to function as alarm signals. These compounds were independent of the vole’s sex. In a field experiment, voles were foraging less, i.e. they were more afraid in the AP odour foraging trays during the first day, as the odour was fresh, than in the second day. This verified the short lasting effect of volatile APs. Our results clarified the chemistry of alarming body odour compounds in mammals, and enhanced our understanding of the ecological role of AP and chemical communication in mammals.
... EE2 removal by wastewater treatment, which is estimated to be between 36 and 98%, is incomplete [6,7]. Therefore, EE2 is a major contaminant in aquatic environments and can be detected in wastewater effluents [8][9][10][11]. Based on directive 2013/39/UE, EE2 was added to the first "Watch list" of substances that may pose a significant risk to or via the aquatic environment [12], and a recent report recommended that EE2 contamination should remain under close monitoring [13]. ...
Objective17α-Ethinylestradiol (EE2) is a synthetic compound used as an oral contraceptive in birth control pills. EE2 is a major contaminant in aquatic environments. It is considered an endocrine-disrupting chemical due to its oestrogenic activity. This study investigates the effect of repeated exposure to EE2 at a dose of 1 µg/kg·(body weight)/day on reproductive function and metabolism. Methods Female Sprague–Dawley rats were subcutaneously injected with EE2 at a dose of 1 µg/kg·(body weight)/day from postnatal day 1 until postnatal day 5, which corresponds to a highly sensitive period of neuroendocrine system organisation. ResultsThe anogenital distance, body weight and blood levels of insulin and glucose of EE2-treated female rats were not different from those of control rats. These results indicate that growth and glucose metabolism were unaffected by treatment. In contrast, vaginal opening occurred significantly earlier in EE2-treated females than in control females. In adulthood, these females present an alteration of their oestrous cyclicity, characterised by persistent vaginal oestrus. Moreover, treated females showed altered uterine and ovarian morphology associated with lower circulating levels of oestradiol and progesterone. Conclusion These results indicate precocious onset of puberty and dysfunction of the reproductive organs. Therefore, early postnatal exposure to EE2 at a dose of 1 µg/kg·bw/d has delayed adverse effects on reproductive function.
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Prey animals can assess the risks predators present in different ways. For example, direct cues produced by predators can be used, but also signals produced by prey conspecifics that have engaged in non-lethal predator-prey interactions. These non-lethal interactions can thereby affect the physiology, behavior, and survival of prey individuals, and may affect offspring performance through maternal effects. We investigated how timing of exposure to predation-related cues during early development affects offspring behavior after weaning. Females in the laboratory were exposed during pregnancy or lactation to one of three odor treatments: (1) predator odor (PO) originating from their most common predator, the least weasel, (2) odor produced by predator-exposed conspecifics, which we call conspecific alarm cue (CAC), or (3) control odor (C). We monitored postnatal pup growth, and we quantified foraging and exploratory behaviors of 4-week-old pups following exposure of their mothers to each of the three odour treatments. Exposure to odors associated with predation risk during development affected the offspring behavior, but the timing of exposure, i.e., pre-vs. postnatally, had only a weak effect. The two non-control odors led to different behavioral changes: an attraction to CAC and an avoidance of PO. Additionally, pup growth was affected by an interaction between litter size and maternal treatment, again regardless of timing. Pups from the CAC maternal treatment grew faster in larger litters; pups from the PO maternal treatment tended to grow faster in smaller litters. Thus, in rodents, offspring growth and behavior are seemingly influenced differently by the type of predation risk perceived by their mothers.
Past experiences are known to affect average behavior but effects on animal personality and plasticity are less well studied. To determine whether experience with predators influences these aspects, we compared the behavior of Gryllodes sigillatus before and after exposure to live predators. We found that emergence from shelter and distance moved during open‐field trials (activity) changed after exposure, with individuals becoming less likely to emerge from shelters but more active when deprived of shelter. We also found that behavioral plasticity in activity increased after exposure to predators. Our results demonstrate that experience with predators affects not only the average behavior of individuals but also how individuals differ from their own prior behavior.
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In recent years, it has been argued that the effect of predator fear exacts a greater demographic toll on prey populations than the direct killing of prey. However, efforts to quantify the effects of fear have primarily relied on experiments that replace predators with predator cues. Interpretation of these experiments must consider two important caveats: 1) the magnitude of experimenter‐induced predator cues may not be realistically comparable to those of the prey's natural sensory environment, and 2) given functional predators are removed from the treatments, the fear effect is measured in the absence of any consumptive effects, a situation which never occurs in nature. We contend that demographic consequences of fear in natural populations may have been overestimated because the intensity of predator cues applied by experimenters in the majority of studies has been unnaturally high, in some instances rarely occurring in nature without consumption. Furthermore, the removal of consumption from the treatments creates the potential situation that individual prey in poor condition (those most likely to contribute strongly to the observed fear effects via starvation or reduced reproductive output) may have been consumed by predators in nature prior to the expression of fear effects, thus confounding consumptive and fear effects. Here, we describe an alternative treatment design that does not utilize predator cues, and in so doing, better quantifies the demographic effect of fear on wild populations. This treatment substitutes the traditional cue experiment where consumptive effects are eliminated and fear is simulated with a design where fear is removed and consumptive effects are simulated through the experimental removal of prey. Comparison to a natural population would give a more robust estimate of the effect of fear in the presence of consumption on the demographic variable of interest. This approach represents a critical advance in quantifying the mechanistic pathways through which predation structures ecological communities. Discussing the merits of both treatments will motivate researchers to go beyond simply describing the existence of fear effects and focus on testing their true magnitude in wild populations and natural communities. This article is protected by copyright. All rights reserved.
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Habitat fragmentation affects individual movements between favorable resource patches. In many small mammal species, an important intrinsic factor affecting recruitment of young is infanticide, committed especially by males. We predict that habitat fragmentation hinders movements of males between patches due to predation risk in the open areas. Thus, fragmentation reduces the number of males to which young litters are exposed to and decreases risk of infanticide in isolated habitat patches. Nonfragmented habitat provides not only breeding possibilities for more females but also safe movements to mate or commit infanticide for males. In a replicated enclosure experiment, we tested how infanticidal status of male bank voles (Myodes glareolus) affects their movements in fragmented vs. nonfragmented habitats with same total area and how this affects female spacing and offspring recruitment into population.We found no difference in the number of offspring recruited per female between infanticidal and fragmentation treatments. Females in the fragmented enclosures had smaller movement areas and stayed closer to their nests, suggesting better protection of pups against intruders. Infanticidal males moved more in general but especially in fragmented enclosures, whilst noninfanticidal males were moving more in nonfragmented enclosures. Our results suggest that behavior of females is affected by the habitat fragmentation, as we expected, but males searching for mates move similarly in safe and risky habitats. Thus, the threat of infanticide was not reflected in the recruitment of young into the population as probably the male–female interactions and effective nest protection by the mothers remained similar in both habitats.
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Maximum likelihood or restricted maximum likelihood (REML) estimates of the parameters in linear mixed-effects models can be determined using the lmer function in the lme4 package for R. As for most model-fitting functions in R, the model is described in an lmer call by a formula, in this case including both fixed- and random-effects terms. The formula and data together determine a numerical representation of the model from which the profiled deviance or the profiled REML criterion can be evaluated as a function of some of the model parameters. The appropriate criterion is optimized, using one of the constrained optimization functions in R, to provide the parameter estimates. We describe the structure of the model, the steps in evaluating the profiled deviance or REML criterion, and the structure of classes or types that represents such a model. Sufficient detail is included to allow specialization of these structures by users who wish to write functions to fit specialized linear mixed models, such as models incorporating pedigrees or smoothing splines, that are not easily expressible in the formula language used by lmer.
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The invasion of alien species into areas beyond their native ranges is having profound effects on ecosystems around the world. In particular, novel alien predators are causing rapid extinctions or declines in many native prey species, and these impacts are generally attributed to ecological naïveté or the failure to recognise a novel enemy and respond appropriately due to a lack of experience. Despite a large body of research concerning the recognition of alien predation risk by native prey, the literature lacks an extensive review of naïveté theory that specifically asks how naïveté between novel pairings of alien predators and native prey disrupts our classical understanding of predator–prey ecological theory. Here we critically review both classic and current theory relating to predator–prey interactions between both predators and prey with shared evolutionary histories, and those that are ecologically ‘mismatched’ through the outcomes of biological invasions. The review is structured around the multiple levels of naïveté framework of Banks & Dickman (2007), and concepts and examples are discussed as they relate to each stage in the process from failure to recognise a novel predator (Level 1 naïveté), through to appropriate (Level 2) and effective (Level 3) antipredator responses. We discuss the relative contributions of recognition, cue types and the implied risk of cues used by novel alien and familiar native predators, to the probability that prey will recognise a novel predator. We then cover the antipredator response types available to prey and the factors that predict whether these responses will be appropriate or effective against novel alien and familiar native predators. In general, the level of naïveté of native prey can be predicted by the degree of novelty (in terms of appearance, behaviour or habitat use) of the alien predator compared to native predators with which prey are experienced. Appearance in this sense includes cue types, spatial distribution and implied risk of cues, whilst behaviour and habitat use include hunting modes and the habitat domain of the predator. Finally, we discuss whether the antipredator response can occur without recognition per se, for example in the case of morphological defences, and then consider a potential extension of the multiple levels of naïveté framework. The review concludes with recommendations for the design and execution of naïveté experiments incorporating the key concepts and issues covered here. This review aims to critique and combine classic ideas about predator–prey interactions with current naïveté theory, to further develop the multiple levels of naïveté framework, and to suggest the most fruitful avenues for future research.
Predators play a critical, top–down role in shaping ecosystems, driving prey population and community dynamics. Traditionally, studies of predator-prey interactions have focused on direct effects of predators, namely the killing of prey. More recently, the non-consumptive effects of predation risk are being appreciated; e.g. the ‘ecology of fear’. Prey responses to predation risk can be morphological, behavioural, and physiological, and are assumed to come at a cost to prey fitness. However, few studies have examined the relationship between predation risk and survival in wild animals. We tested the hypothesis that predation risk itself could reduce survival in wild-caught snowshoe hares. We exposed female snowshoe hares to a simulated predator (a trained dog) during gestation only, and measured adult survival and, in surviving females, their ability to successfully wean offspring. We show for the first time in a wild mammal that the risk of predation can itself be lethal. Predation risk reduced adult female survival by 30%, and had trans-generational effects, reducing offspring survival to weaning by over 85% – even though the period of risk ended at birth. As a consequence of these effects the predator-exposed group experienced a decrease in number, while the control group substantially increased. Challenges remain in determining the importance of risk-induced mortality in natural field settings; however, our findings show that non-lethal predator encounters can influence survival of both adults and offspring. Future work is needed to test these effects in free-living animals.
Rats are predators of mice in nature. Nevertheless, it is a common practice to house mice and rats in a same room in some laboratories. In this study, we investigated the behavioral and physiological responsively of mice in long-term co-species housing conditions. Twenty-four male mice were randomly assigned to their original raising room (control) or a rat room (co-species-housed) for more than 6 weeks. In the open-field and light-dark box tests, the behaviors of the co-species-housed mice and controls were not different. In a 2-choice test of paired urine odors [rabbit urine (as a novel odor) vs. rat urine, cat urine (as a natural predator-scent) vs. rabbit urine, and cat urine vs. rat urine], the co-species-housed mice were more ready to investigate the rat urine odor compared with the controls and may have adapted to it. In an encounter test, the rat-room-exposed mice exhibited increased aggression levels, and their urines were more attractive to females. Correspondingly, the levels of major urinary proteins were increased in the co-species-housed mouse urine, along with some volatile pheromones. The serum testosterone levels were also enhanced in the co-species-housed mice, whereas the corticosterone levels were not different. The norepinephrine, dopamine, and 5-HT levels in the right hippocampus and striatum were not different between the 2. Our findings indicate that chronic co-species housing results in adaptation in male mice; furthermore, it appears that long-term rat-odor stimuli enhance the competitiveness of mice, which suggests that appropriate predator-odor stimuli may be important to the fitness of prey animals.