Ethology

The ability of prey to eavesdrop on predator vocalizations is expected to increase survival by reducing detection and capture. Unfortunately, most research has been conducted in vertebrates, and little is known about this ability in invertebrates. We measured latency to emerge, overall activity, and shelter visits in wild-caught fall field crickets (Gryllus pennsylvanicus) in response to acoustic playback. Stimuli included multiple predator vocalizations, non-predator vocalizations, white noise, and a control. We predicted that crickets would reduce activity, spend more time in shelter, and freeze in response to stimuli representing greater risk. Contrary to our predictions, crickets traveled greater distances, spent more time moving, and spent less time in shelter in response to predator vocalizations versus controls. We did not, however, find clear differences in responses between predator vocalizations and other treatments. Our results suggest that crickets may not differentiate between the vocalizations of predators, non-predators, and other abrupt sounds. Consequently, eavesdropping may not be a viable method of assessing predation risk for this species and its general use remains unclear.

As foraging increases exposure to predators, individuals must allocate time to vigilance in addition to food acquisition. Although those behaviors are assumed to be mutually exclusive, this is often hard to disentangle, given that multiple sensory modalities are often applied. However, when foraging, animals often have one sensory modality more impaired (e.g. vision), than others (e.g. auditory); thus, a level of compromise on vigilance capabilities is assumed while foraging. Here, we test the idea that when a food item requires manipulation, thus reducing vigilance capabilities, social foraging should promote shared costs of vigilance. We conducted video recordings to investigate whether there is a trade-off between foraging and vigilance behavior in red-legged seriemas (Cariama cristata) when foraging in pairs. We studied the social foraging behavioral patterns while seriemas were feeding on macauba palm (Acrocomia aculeata) fruit, which requires a series of throwing maneuvers to crack the hard outer shell. Our results showed that within a pair recording one individual was observed to spend more time vigilant than their partner, while the other individual spent more time throwing fruit than their partner. However, we found no evidence for synchronization or coordination of vigilance and foraging between individuals within a feeding bout. These results provide insight into the foraging ecology of red-legged seriemas and the trade-off between foraging and vigilance in social contexts.

Shorebirds provide an excellent model for investigating the relationship between bill morphology and food acquisition. Food acquisition comprises three successive behavioural stages: foraging (locomotion and prey capture), feeding (food handling and transport), and swallowing. During food transport, these birds use two non-lingual mechanisms, surface-tension transport (ST) and ballistic transport (BT), whose characteristics depends on the kinematics of head and beak movements and the physical properties of the food. We investigated the behavioural flexibility of these transport mechanisms in captive and free-ranging individuals of two species with contrasting beak morphologies: the Pied avocet (Recurvirostra avosetta) and the Black-winged stilt (Himantopus himantopus). Although these species have beaks of a similar size, avocets are distinguished by their upward-curved beaks, whereas stilts have a rather straight beak. We examined the effects of food water content and the presence or absence of water in the beak on food transport by quantifying maximum gape, maximum head displacement, and maximum head velocity. Transport kinematics were jointly influenced by food properties and water availability in the beak. Moist food improved transport performance in both species, whereas dry food required compensatory increases in gape amplitude and head movements, demonstrating that neither ST nor BT constitutes a fixed behavioural sequence. Species also differed consistently in their transport strategies: Black-winged stilts relied on slower, larger-amplitude head movements, whereas Pied Avocets exhibited faster, more precise movements, particularly when water was present. These findings demonstrate that ST and BT share common biomechanical foundations while being governed by rapid kinematic adjustments to changing environmental conditions that probably correspond to flexible motor control. This behavioural flexibility in food transport is therefore likely to enhance feeding performance and ecological resilience in the heterogeneous habitats occupied by shorebirds, suggesting that context-dependent modulation of transport behaviour represents an important adaptive feature of these shorebirds.

Honey bees can swim on the water surface toward dark regions, a behavior known as scototaxis that may facilitate escape from water. Although this behavior has been reported in both honey bees and solitary bees, variation among honey bee species remains poorly understood. We compared scototaxis during swimming in four honey bee species representing two nesting types: open-nesting (Apis florea and A. dorsata) and cavity-nesting (A. cerana and A. mellifera). Individual bees were released into a water-filled arena containing a dark sector, and their landing angles were recorded. All species exhibited significant orientation toward the dark sector. However, open-nesting species showed significantly stronger orientation than cavity-nesting species. No significant differences were detected between replicate colonies within species or between species within the same nesting type, whereas differences between nesting types were highly significant. Hierarchical clustering based on orientation strength placed Osmia, a solitary cavity-nesting bee from our previous study, in the same behavioral cluster as the two open-nesting Apis species rather than the cavity-nesting honey bees. We also measured swimming duration, distance, and velocity, but found no consistent differences between nesting types. These results demonstrate substantial interspecific variation in swimming scototaxis. The behavioral clustering is consistent with the hypothesis that strong scototaxis represents an ancestral trait that has been reduced in the derived A. cerana/A. mellifera lineage.

Animal personality is the expression of consistent among-individual variation in a behavioural trait across time and context. The application of this theory to behavioural research provides a valuable framework to investigate the among- and within-individual variation in behaviour. The animal personality theory is particularly relevant to understand and diagnostic fish welfare. It can be integrated within the nature-based welfare framework, which emphasizes the expression of species-specific behaviours and the maintenance of consistent behavioural patterns over time. Because behavioural trait such as boldness may be closely linked to other functional phenotypes, such associations reflect the broader concept of animal personality, whereby a behavioural trait can covary with another phenotypic trait to support an adaptive responses to environmental conditions. Although these relationships are both species and context dependent, they are consistently shaped by environmental conditions and environmental complexity tend to promote species specific behaviours and reduce maladaptive traits. Here we examined how structural environmental complexity shapes personality in Arctic charr and their covarying functional phenotypes, specifically growth rate and brain size. We propose that environmental complexity promotes alternative behavioural and functional phenotypes through multivariate phenotypic plasticity. We found that environmental complexity did not influence mean boldness between both treatments, but repeatability of boldness in the complex environment was remarkably consistent over a longer-term period, while estimates collapse after seven days in the plain treatment. Our findings are a clear evidence that environmental complexity foster stable behavioural trait expression and that a plain environment may suppress personality. Our results provide compelling evidence that behavioural structure and dynamics are embedded within patterns of behavioural variance. Although we found no support for behavioural covariation or associations with growth rate and brain size, we suggest that the animal personality framework may offer a valuable approach for diagnosing fish welfare issues through the partitioning of behavioural variance.

When food resources provide reward while simultaneously imposing risk, animals can regulate their behaviour to continue exploiting them while reducing associated costs. Angling presents such a feeding situation, in which obtaining bait can result in hooking and capture. Although fish become less vulnerable to angling following capture experience, the behavioural changes underlying this reduced vulnerability remain poorly understood. Here, we used fine-scale video observations to examine feeding behaviour of red sea bream (Pagrus major) toward angling gear before and after capture experience. Naive fish rapidly swallowed the bait and were subsequently captured, whereas capture-experienced fish continued to approach and feed on the gear but shifted from swallowing to biting. This feeding-mode shift may allow fish to maintain exploitation of the bait while reducing capture risk. Our observations provide a behavioural explanation for reduced angling vulnerability and suggest that direct experience with capture risk can alter how fish exploit a rewarding food resource.

Offspring phenotypes are often influenced by maternal age at reproduction. However, the persistence of such effects across multiple generations remains poorly understood, especially in rare cases where offspring of older mothers exhibit enhanced performance. Using the predatory mite Amblyseius herbicolus, we experimentally tested whether maternal age at reproduction influences offspring life history across two generations under contrasting dietary conditions. We found that grandmaternal (F0) and maternal (F1) age affected offspring (F2) developmental time and body size, but not hatching success, survival, or oviposition. The effect of maternal age on body size depended on grandmaternal age, suggesting that grandmaternal age mediates how mothers adjust offspring provisioning. Diet strongly influenced offspring traits but showed limited interaction with age effects. To our knowledge, this is the first study to demonstrate the persistence of an inverse Lansing effect across generations, whereby enhanced immature development is maintained without detectable trade-offs in reproduction or adult survival. These results identify maternal age as a key driver of transgenerational phenotypic variation, with important implications for population dynamics and responses to environmental change at the population level.

Decision-making is key to survival, with choices and behaviour typically shaped by evolutionary pressures to enhance fitness and minimize loss. Inequity aversion, the tendency to respond negatively to unequal reward distributions, is therefore unexpected, as acts of fairness may be costly to the individual in the short term. In social, cooperative species, however, fairness may enhance long-term benefits through stable social interactions. Inequity aversion is believed to promote cooperative social structures and has only been demonstrated in certain social species. Yet, solitary species have not been tested, and whether this behaviour depends on social cooperative structures is unknown. Examining solitary species could clarify whether inequity aversion reflects an adaptation to cooperation or a general cognitive capacity linked to social comparison and reward evaluation. We studied inequity aversion in polar bears (Ursus maritimus), a largely solitary species, using three complementary, non-invasive cognition tests on seven bears from two zoos in the Netherlands. (1) An impunity experiment assessing effort-based inequity, in which bears performed a standardized action to obtain food rewards. (2) A choice-based experiment testing active selection between equal and unequal reward distributions. (3) A no-task control experiment assessing responses to unequal reward distribution without effort. Under effort-based inequity, bears reduced task performance and increased task latency when a partner received a superior reward. Unequal reward distributions did not affect reward acceptance independent of effort. In the choice-based paradigm, bears preferentially selected equal reward distributions with a partner present. Although generalizability to wild populations is limited, these findings demonstrate behavioural aversion of social inequity in a non-cooperative species. Inequality aversion may therefore not be restricted to social, cooperative animals but instead reflect a broadly distributed cognitive mechanism that emerges under relevant social conditions. These findings challenge current theories and advance our understanding of the cognitive foundations of fairness-related behaviour in animals.

Female-female competition is increasingly recognized as a key driver of female ornamentation, including birdsong, which often functions in intrasexual competition. However, the specific resources females use elaborate traits to compete for remain unclear. In addition, few studies have simultaneously investigated the use of multiple competitive traits in females, despite growing independent interest in these traits (e.g., female song and aggression). We investigated the competitive contexts of female song, aggression and calling behavior in northern house wrens (Troglodytes aedon) to determine which resources females compete for across the breeding season. We simulated conspecific territorial intrusions using female song at three breeding stages representing different contexts: arrival (mate and territory acquisition), nest building (nest site and breeding status defense), and egg laying (brood defense). We tested whether female song and physical aggression varied as reproductive resources shifted across the breeding cycle. Females were significantly more aggressive during nest building, showing 5.8 times greater odds of a higher-intensity aggressive response during nest building compared to arrival. Female song output was similar across early stages but declined during egg laying, though this was not statistically significant after correction for multiple comparisons and individuals varied substantially in overall singing propensity. Non-song vocalizations varied by call type and breeding stage. Calls associated with aggression occurred most frequently during nest building, consistent with peak physical aggression responses. Together, these results identify nest building as the stage of highest female aggression, consistent with heightened competition over nest cavities and associated breeding status in this cavity-nesting species. In contrast, female song occurred across all stages and appears to function in multiple competitive contexts. This study provides evidence for context and mode-specific female signaling in a temperate songbird and highlights that females strategically use aggression, calls, and song to mediate social conflict across breeding contexts.

Species exhibit variation in their circadian activity rhythms, including being active during the day (diurnal), the night (nocturnal), or the twilight periods (crepuscular). While the influence of environmental factors such as light on entraining and controlling activity patterns is well known, it is unclear how social and abiotic factors affect circadian behaviors. The daily activity patterns of Lake Tanganyikan cichlid fish are diverse, and may be associated with species diversification. Intriguingly, some shell-dwelling cichlid species, which anecdotal observation suggests are diurnal, displayed strong nocturnal activity when assayed in a "common garden" lab setup. Here, by integrating field and lab-based studies, we provide three lines of evidence that social and environmental cues mask underlying circadian rhythms in shell-dwelling cichlids. First, we demonstrate that shell-dwelling cichlids are diurnal in their natural habitat, but convert to nocturnal activity when assayed alone in a common garden. Second, we identify that in the presence of a shell and conspecifics the highly social N. multifasciatus becomes diurnal. In contrast, the circadian activity pattern of a closely related, but sub-social species, N. brevis, is masked only by a shell, and unaffected by the presence of conspecifics. Third, we demonstrate that the masking effect of shells and conspecifics is circadian and continues in the absence of light. Fourth, we identify that the response to these cues is influenced by sex, with greater effects in female fish, and by pedigree, with stronger effects in individuals bred and raised in captivity compared to those captured in the wild. Together, these experiments reveal new relationships between circadian rhythms and sociality in these fishes, offering broader insights into the ecological and evolutionary drivers of these behaviours.

Schooling is a widespread behaviour among pelagic fish that reduces locomotion costs of migration and predation risk. It is characterised by coordinated movements, speed and orientation of group members. Although schooling can vary both between populations and among individuals of the same population, these levels of variation are seldom considered together. Here, we investigated whether schooling tendencies in three-spined sticklebacks (Gasterosteus aculeatus) vary between populations and among individuals within-populations depending on their phenotype. We first compared the schooling behaviour of anadromous (migrants) and recently landlocked (residents) populations and tested whether individual sociability explained within-population variation in schooling. We then studied whether schooling behaviour is modulated when migrants and residents are in mixed groups. Migrant groups displayed better schooling performance than residents with higher alignment. Within migrant groups, less social individuals displayed more leadership, were better aligned, swam closer to others and fatigued less than more social fish. These patterns were absent in residents. When the two populations were mixed, overall schooling performance declined, particularly in migrants, with schools exhibiting reduced compaction and alignment, resembling resident groups. Thus, individuals did not modulate their schooling behaviour to the group social composition and phenotype-dependent schooling patterns were disrupted in migrants when mixed. These findings suggest that migration-related behaviours can disappear quickly post-isolation. They also highlight the importance of individual sociability in self-organisation and shaping collective behaviour, especially in populations that retain migratory behaviour. Considering behavioural variation across multiple hierarchical levels is thus key to understanding complex social systems.

Social animals must identify, evaluate, and appropriately respond to other individuals, often relying on multiple sources of information to guide context-dependent behavior. While individual recognition is thought to be widespread and critical for many social interactions (e.g. territoriality, dominance hierarchies), convincing demonstrations across animals beyond class-level recognition remain rare. Further, the cues animals use to recognize individuals remains poorly understood, especially in natural environments where they are exposed to numerous cues which may require cue integration. Here, we examine how socially dominant males of the African cichlid fish, Astatotilapia burtoni, use individual recognition to modulate aggression and whether this process depends on the spatial context. Using naturalistic experimental setups, we exposed males to familiar and unfamiliar neighbors while systematically manipulating spatial location and presenting a neighbor either singly or simultaneously to simulate competing social cues. Additionally, we measured circulating testosterone to assess endocrine correlates of recognition and aggression. We found that focal males habituated to familiar neighbors only when they remained in a consistent territorial location, demonstrating that spatial context is important for recognition. Further, when familiar and unfamiliar neighbors were presented simultaneously, males preferentially aggressed toward unfamiliar rival and adjust their aggressive response to the perceived threat of their neighbors, indicating that identity-based threat assessment is maintained under competing social demands. Together, our results show that recognition is a flexible, multi-faceted process that depends on the integration of identity and contextual information to guide territorial decision-making under dynamic social conditions.

Budgerigars (Melopsittacus undulatus) are vocal-learning birds with well-developed auditory abilities, but how they behaviorally evaluate melodic and rhythmic structure in sound sequences remains unclear. We examined whether budgerigars show preferences for these acoustic features and whether such preferences differ between the sexes. Three male and three female budgerigars were presented with four 8-s sound sequences in a preference apparatus: Simple (no pitch or temporal variation), Melody (pitch variation only), Rhythm (temporal variation only), and Complex (both pitch and temporal variation). Preference was quantified as the time spent in the area associated with each stimulus. No statistically significant differences among the four stimuli were detected within individuals. However, effect-size estimates indicated that two females spent more time with sequences containing rhythmic structure, whereas males showed no consistent preference related to either melodic or rhythmic components. Multidimensional scaling further suggested greater separation among stimulus conditions in females than in males. Consistent with this pattern, condition differentiation indices were higher in all three females than in all three males, although the sex difference was not statistically significant. These results suggest a possible sex-related difference in how budgerigars behaviorally weight temporal structure, with females showing greater differentiation among auditory sequence types under the present testing conditions.

The social complexity hypothesis for communication posits that species with elaborate social systems require more sophisticated vocal communication to navigate social life. Geladas (Theropithecus gelada), who live in complex multilevel societies and form diverse social bonds, are ideal for testing this hypothesis. Males produce species-specific, or derived, calls that reflect social status and function in facilitating male-female bonding. However, although derived call use also occurs in male-male social interactions, their function in male-male communication remains unexamined. In this pilot study we investigate the use of derived calls within a captive all-male bachelor group and test two hypotheses: (1) derived calls signal bachelor dominance status, and (2) derived calls function to maintain male-male social bonds. Despite a limited sample size, our results show that bachelor leaders produce higher proportions of derived calls than bachelor followers, mirroring patterns found in harem groups. Furthermore, bachelor followers were most likely to use derived calls in affiliative and resting contexts, suggesting a role in social bonding. In contrast, bachelor leaders used them more broadly. They additionally produced derived calls during agonism and travel, thereby possibly reflecting their more elaborate leadership duties. These findings provide the first preliminary evidence that derived calls serve functions beyond male-female communication and may be integral to both status signalling and affiliative interactions among bachelor males. Our results on the vocal dynamics in male geladas shed further light on the co-evolution of social and vocal complexity and offer broader implications for the origins of human language.

Reliable morphological criteria for sexing birds without the need for invasive procedures are scientifically and ethically valuable. Smith et al (2005) reported that iris colour assigns the genetic sex of Starlings Sturnus vulgaris with 98% accuracy, and bill base colour with 100% accuracy when birds are in breeding condition, but these figures have not been tested independently. We applied their criteria to 31 captive Starlings in breeding condition, using three observers blind to genetic sex, two of them new to Starling sexing. Bill base colour correctly predicted genetic sex for all 29 birds scored, with complete observer agreement. Mean iris score was correct for 27 of 31 (87.1%), fewer than the 94 of 96 reported by Smith et al (2005) for birds in breeding condition (Fisher exact test, P = 0.031), although with so few errors in either study this comparison is imprecise. Observer agreement was high, so these errors reflect iris colouration conflicting with genetic sex rather than inconsistent scoring; bill base colour was correct for all four misassigned birds. Body mass and tarsus length classified 77% and 58% of birds correctly, their published thresholds missing in opposite directions. We conclude that bill base colour should be the primary criterion for sexing Starlings in breeding condition; the accuracy of iris colour should not be assumed to be 98%.

Many social insects, such as ants, change their behavior stereotypically from nursing to foraging as they age, a concept known as temporal polyethism. Biogenic amines are associated with these specific behaviors, but much about them remains poorly understood in these animals. Many aminergic systems lack anatomical characterization, and little is known about the expression of genes required for the aforementioned behavioral phenotypes. Here, we studied Pogonomyrmex barbatus brains and identified the neurons that produce two relevant amines, dopamine and octopamine, by detecting their synthesis enzymes, tyrosine hydroxylase and tyramine beta-hydroxylase, and their transcripts. We also compared the expression of both genes in young nurses and mature foragers by measuring the fluorescence intensity. Dopaminergic and octopaminergic neurons are predominantly located in the protocerebrum and in the subesophageal zone. Neurons that produce dopamine are also present in the optic lobes, whereas octopamine-producing ones show clusters in the antennal lobes. Both genes are downregulated as the organism ages. A reduction in the expression of these genes might be correlated with the mature workers increased propensity to perform extranidal tasks, such as foraging.

Human-influenced environments pose challenges but also provide wildlife with anthropogenic resources. Individuals vary widely in their ability to exploit such resources, often as a function of behavioural type. However, we lack a clear understanding of how variation in behavioural traits influences stages of resource exploitation required to use anthropogenic resources. Using fully automated foraging puzzles, we examined how boldness and sociability influenced three aspects of resource exploitation - discovery, problem-solving, and overall performance - in two wild populations of California ground squirrels (Otospermophilus beecheyi). Bolder individuals discovered the resource earlier, solved the task faster and achieved higher performance, indicating that boldness promotes efficient exploitation of anthropogenic resources across multiple stages in the process. Greater sociability and more opportunities to observe conspecifics solving the task led to faster problem-solving, consistent with evidence for observational learning. Squirrels in the recreational-use population - with regular exposure to humans and anthropogenic food - were faster to discover the resource than those in a trail-use population - where human exposure was transient and no anthropogenic food available. Problem-solving latency and performance were consistent between populations. Our findings highlight how individual variation in behavioural traits drives performance in novel ecological contexts, providing a mechanistic understanding of behavioural plasticity in human-influenced environments.

Animals often display active decision-making while navigating noisy habitats, for varied purposes like foraging, finding shelter, evading predators or finding suitable mates. While some responses are hardwired, following fixed action patterns, others are more flexible, and display context-dependent plasticity. Animals might weigh behavioural consistency against adaptive plasticity when facing recurrent ecological difficulties, but the nuanced mechanisms orchestrating this balance is not well understood. We investigated this question by repeatedly exposing 47 free-ranging dogs to chicken contaminated with lemon solution, an aversive but edible and familiar food source, across four consecutive trials. By integrating behavioural frequencies, latency measures, feeding efficiency, interaction-time and location effects, repeatability analyses, and behavioural sequence structure, we examined how repeated experience influenced foraging behaviour. Contrary to expectations of broad behavioural reorganisation, most behaviours remained stable. Initial approach latency was strongly repeatable within individuals with no detectable influence of sex or trial number across the population. This suggests that how an animal first assesses risk is primarily shaped by stable personality traits rather than momentary external factors. Eating behaviour and bowl nudging also changed selectively across exposures, with modest sex differences in several behaviours. Behavioural sequences became increasingly similar over time, while several behaviours retained consistent individual differences, indicating persistent behavioural tendencies. Our results collectively demonstrated that repeated aversive exposure selectively modified a few critical decision points rather than the entire behavioural repertoire. This implies that animals can adapt by making small adjustments to specific behaviours while maintaining the stability of their overall behavioural architectures. Animals seem to assess risk separately from how well they use resources. That separation may help explain how stable personalities can coexist with the fast, flexible decisions animals need in unpredictable environments shaped by humans.

Japanese quail (Coturnix japonica) rely on an extensive vocal repertoire to coordinate social behaviour and respond to predation risk, yet how individual call types are modulated by predator identity remains poorly characterized. In this pilot case-series, two sexually mature Japanese quail (each contributing more than 2,000 classified vocalizations across repeated predator-exposure trials) were exposed to simulated aerial (hawk) and terrestrial (cat) predator stimuli across four social contexts, before, during, and after stimulus presentation. Vocalizations were classified, via blind dual-observer review, into 18 spectrographically distinct call types (an exploratory repertoire estimate, not independently statistically validated), of which 10 were acoustically characterized. Chi-square analyses of the full call-type composition revealed highly significant differences between hawk and cat exposure in both individuals (both p < 0.0001); call types 5 and 10 were significantly cat-biased and call type 2 was significantly hawk-biased in both individuals. An exploratory correlation suggested that call types with a higher maximum frequency tended to be relatively hawk-biased (Spearman rho = -0.67, p = 0.033, uncorrected for multiple comparisons). Independently, call-type composition shifted significantly between the pre- and post-exposure phases in all four individual-by-predator combinations (all p < 0.0001), driven substantially by a consistent post-exposure increase in call type 9 across both individuals and both predator classes. Additional exploratory analyses of social-context (mirror and concealed-conspecific) effects on call-type composition are presented in the supporting information. Given the small sample size (n = 2), these findings are presented as case-based, hypothesis-generating observations intended to justify and inform a fully powered follow-up study, rather than as population-level conclusions.

Chorus howls play a central role in wolves' (Canis lupus) social communication, shaping both intra- and inter-pack interactions. While synchrony has been shown to influence perceived threat from outgroups in birds and humans, whether the synchrony of an outside pack's chorus howl plays a role in wolf pack behaviour remains unclear. This study investigated behavioural responses of 19 zoo-housed wolf packs to playbacks of synchronous and asynchronous chorus howls from unfamiliar packs, in comparison to a dove control call. We hypothesized that packs would show stronger intra-pack cohesion and greater spatial withdrawal in response to more synchronous, and thus cohesive - and potentially more formidable - sounding vocalisations. Alternatively, asynchronous howls might elicit stronger responses due to their ambiguity and the "Beau Geste" effect (where asynchronous signals make it harder to infer the number of callers). Synchronous howls resulted in more distance from the sound source and less activity compared with asynchronous howls. This could be interpreted as a more cautious response towards a perceived threat. However, since there was no effect on pack-directed behaviour and no difference between the synchronous and control sound, lower interest or motivation to engage with the stimulus cannot be excluded. In contrast, wolves spent more time near the sound-source after hearing asynchronous howls, approached it more, and oriented towards it for longer compared to synchronous playbacks, a pattern consistent with threat-investigation. In how far these reactions result from a more difficult localisation of synchronous howls or more ambiguity in pack size in the asynchronous playbacks, as well as how this translates to wild wolf behaviour, remains to be investigated.