The evolution and ecology of multiple antipredator defences.

Prey seldom rely on a single type of antipredator defence, often using multiple defences to avoid predation. In many cases, selection in different contexts may favour the evolution of multiple defences in a prey. However, a prey may use multiple defences to protect itself during a single predator encounter. Such "defence portfolios" that defend prey against a single instance of predation are distributed across and within successive stages of the predation sequence (encounter, detection, identification, approach (attack), subjugation and consumption). We contend that at present, our understanding of defence portfolio evolution is incomplete, and seen from the fragmentary perspective of specific sensory systems (e.g., visual) or specific types of defences (especially aposematism). In this review, we aim to build a comprehensive framework for conceptualizing the evolution of multiple prey defences, beginning with hypotheses for the evolution of multiple defences in general, and defence portfolios in particular. We then examine idealized models of resource trade-offs and functional interactions between traits, along with evidence supporting them. We find that defence portfolios are constrained by resource allocation to other aspects of life history, as well as functional incompatibilities between different defences. We also find that selection is likely to favour combinations of defences that have synergistic effects on predator behaviour and prey survival. Next, we examine specific aspects of prey ecology, genetics and development, and predator cognition that modify the predictions of current hypotheses or introduce competing hypotheses. We outline schema for gathering data on the distribution of prey defences across species and geography, determining how multiple defences are produced, and testing the proximate mechanisms by which multiple prey defences impact predator behaviour. Adopting these approaches will strengthen our understanding of multiple defensive strategies.

Negative mood affects the expression of negative but not positive emotions in mice.

Whether and to what extent animals experience emotions is crucial for understanding their decisions and behaviour, and underpins a range of scientific fields, including animal behaviour, neuroscience, evolutionary biology and animal welfare science. However, research has predominantly focused on alleviating negative emotions in animals, with the expression of positive emotions left largely unexplored. Therefore, little is known about positive emotions in animals and how their expression is mediated. We used tail handling to induce a negative mood in laboratory mice and found that while being more anxious and depressed increased their expression of a discrete negative emotion (disappointment), meaning that they were less resilient to negative events, their capacity to express a discrete positive emotion (elation) was unaffected relative to control mice. Therefore, we show not only that mice have discrete positive emotions, but that they do so regardless of their current mood state. Our findings are the first to suggest that the expression of discrete positive and negative emotions in animals is not equally affected by long-term mood state. Our results also demonstrate that repeated negative events can have a cumulative effect to reduce resilience in laboratory animals, which has significant implications for animal welfare.

Unravelling the illusion of flicker fusion.

For over 150 years, researchers have investigated the anti-predator function of animal patterns. However, this work has mainly focused on when prey remain still, and has only recently started to incorporate motion into the study of defensive coloration. As motion breaks camouflage, a new challenge is to understand how prey avoid predators while moving around their environment, and if a moving prey can ever be camouflaged. We propose that there is a solution to this, in that a 'flicker fusion effect' can change the appearance of the prey in the eyes of their predators to reduce the chances of initial detection. This effect occurs when a high contrast pattern blurs at speed, changing the appearance of the prey, which may help them better match their background. Despite being widely discussed in the literature, the flicker fusion effect is poorly described, there is no clear theoretical framework for testing how it might reduce predation, and the terminology describing it is, at best, rather confusing. Our review addresses these three key issues to enable researchers to formulate precise predictions about when the flicker fusion effect occurs, and to test how it can reduce predation.

Cognition and the evolution of camouflage.

Camouflage is one of the most widespread forms of anti-predator defence and prevents prey individuals from being detected or correctly recognized by would-be predators. Over the past decade, there has been a resurgence of interest in both the evolution of prey camouflage patterns, and in understanding animal cognition in a more ecological context. However, these fields rarely collide, and the role of cognition in the evolution of camouflage is poorly understood. Here, we review what we currently know about the role of both predator and prey cognition in the evolution of prey camouflage, outline why cognition may be an important selective pressure driving the evolution of camouflage and consider how studying the cognitive processes of animals may prove to be a useful tool to study the evolution of camouflage, and vice versa. In doing so, we highlight that we still have a lot to learn about the role of cognition in the evolution of camouflage and identify a number of avenues for future research.

Increased predation of nutrient-enriched aposematic prey.

Avian predators readily learn to associate the warning coloration of aposematic prey with the toxic effects of ingesting them, but they do not necessarily exclude aposematic prey from their diets. By eating aposematic prey 'educated' predators are thought to be trading-off the benefits of gaining nutrients with the costs of eating toxins. However, while we know that the toxin content of aposematic prey affects the foraging decisions made by avian predators, the extent to which the nutritional content of toxic prey affects predators' decisions to eat them remains to be tested. Here, we show that European starlings (Sturnus vulgaris) increase their intake of a toxic prey type when the nutritional content is artificially increased, and decrease their intake when nutritional enrichment is ceased. This clearly demonstrates that birds can detect the nutritional content of toxic prey by post-ingestive feedback, and use this information in their foraging decisions, raising new perspectives on the evolution of prey defences. Nutritional differences between individuals could result in equally toxic prey being unequally predated, and might explain why some species undergo ontogenetic shifts in defence strategies. Furthermore, the nutritional value of prey will likely have a significant impact on the evolutionary dynamics of mimicry systems.

Better the devil you know: avian predators find variation in prey toxicity aversive.

Toxic prey that signal their defences to predators using conspicuous warning signals are called 'aposematic'. Predators learn about the toxic content of aposematic prey and reduce their attacks on them. However, through regulating their toxin intake, predators will include aposematic prey in their diets when the benefits of gaining the nutrients they contain outweigh the costs of ingesting the prey's toxins. Predators face a problem when managing their toxin intake: prey sharing the same warning signal often vary in their toxicities. Given that predators should avoid uncertainty when managing their toxin intake, we tested whether European starlings (Sturnus vulgaris) preferred to eat fixed-defence prey (where all prey contained a 2% quinine solution) to mixed-defence prey (where half the prey contained a 4% quinine solution and the other half contained only water). Our results support the idea that predators should be more 'risk-averse' when foraging on variably defended prey and suggest that variation in toxicity levels could be a form of defence.

A synthesis of deimatic behaviour.

Deimatic behaviours, also referred to as startle behaviours, are used against predators and rivals. Although many are spectacular, their proximate and ultimate causes remain unclear. In this review we aim to synthesise what is known about deimatic behaviour and identify knowledge gaps. We propose a working hypothesis for deimatic behaviour, and discuss the available evidence for the evolution, ontogeny, causation, and survival value of deimatic behaviour using Tinbergen's Four Questions as a framework. Our overarching aim is to direct future research by suggesting ways to address the most pressing questions in this field.

Sporting contests: seeing red? Putting sportswear in context.

The shirt colour worn by sportsmen can affect the behaviour of the competitors, but Hill and Barton show that it may also influence the outcome of contests. By analysing the results of men's combat sports from the Athens 2004 Olympics, they found that more matches were won by fighters wearing red outfits than by those wearing blue; they suggest that red might confer success because it is a sign of dominance in many animal species and could signal aggression in human contests. Here we use another data set from the 2004 Olympics to show that similar winning biases occur in contests in which neither contestant wears red, indicating that a different mechanism may be responsible for these effects.

Predators' toxin burdens influence their strategic decisions to eat toxic prey.

Toxic prey advertise their unprofitability to predators via conspicuous aposematic coloration [1]. It is widely accepted that avoidance learning by naive predators is fundamental in generating selection for aposematism [2, 3] and mimicry [4, 5] (where species share the same aposematic coloration), and consequently this cognitive process underpins current evolutionary theory [5, 6]. However, this is an oversimplistic view of predator cognition and decision making. We show that predators that have learned to avoid chemically defended prey continue to attack defended individuals at levels determined by their current toxin burden. European starlings learned to discriminate between sequentially presented defended and undefended mealworms with different color signals. Once birds had learned to avoid the defended prey at a stable asymptotic level, we experimentally increased their toxin burdens, which reduced the number of defended prey that they ingested in the subsequent trial. This was due to the birds making strategic decisions to ingest defended prey on the basis of their visual signals. Birds are clearly able to learn about the nutritional benefits and defensive costs of eating defended prey, and they regulate their intake according to their current physiological state. This raises new perspectives on the evolution of aposematism, mimicry, and defense chemistry.

Birds learn to use distastefulness as a signal of toxicity.

Aposematic prey advertise their toxicity using conspicuous visual signals that predators quickly learn to avoid. However, it is advantageous for predators not to simply avoid toxic prey, but to learn about the amount of toxin that prey contain, and include them in their diets when the nutritional gains are high relative to the costs of ingesting the toxin. Therefore, when foraging on a defended prey population where individuals vary in their toxin concentration, predators should learn to use cues which distinguish prey with different levels of toxicity in order to include less defended individuals in their diets. In this experiment, we found that European starlings (Sturnus vulgaris) could learn to use a bitter taste to predict the amount of toxin that individual prey contained, and use that information to preferentially ingest less toxic prey to maximize their nutrient intake relative to the amount of toxin ingested. Our results suggest that bitter tastes could evolve as reliable signals of toxicity, and can help to explain why many toxins taste bitter. They also highlight the need to develop new mathematical simulations of the evolution of prey defences which incorporate the adaptive decision-making processes underlying nutrient and toxin management.

Taste-rejection behaviour by predators can promote variability in prey defences.

The evolution and maintenance of toxicity in a prey population is a challenge to evolutionary biologists if the investment in toxin does not benefit the individual. Recent experiments suggest that taste-rejection behaviour enables predators to selectively ingest less toxic individuals, which could stabilize investment in defences. However, we currently do not know if taste rejection of defended prey is accurate across different contexts, and that prey always benefit according to their investment. Using avian predators, we show that the rejection probability does not solely depend on the investment in defence by an individual, but also on the investment by other individuals in the same population. Therefore, taste rejection by predators could lead to destabilization in the investment in defences, and allow variability in prey defences to exist.

Using preferred fluids and different reward schedules to motivate rhesus macaques (Macaca mulatta) in cognitive tasks.

Rhesus macaques (Macaca mulatta) used in behavioural neuroscience are often required to complete cognitively complex tasks, for which a high level of motivation is essential. To induce motivation, researchers may implement fluid-restriction protocols, whereby freely available water is limited, such that fluid can be used as a reward in the laboratory. A variety of different rewards and schedules are used, but there exists a lack of data assessing their effectiveness. In this study, we aimed to quantify fluid preference in rhesus macaques and to use these preferences to compare the motivational quality of different reward schedules: the monkey's previous reward (i.e. the fluid used to reward them in past studies), their new preferred reward, a variable schedule of previous and preferred reward, and a choice between the previous and preferred rewards. We found that it may be possible to reduce the level of restriction if an adequately motivating preferred reward is identified, but that this is dependent on the animal. Each monkey responded differently to both the fluid-preference assessments and to the different reward schedules. As such, monkeys should not be subject to 'blanket' protocols but should be assessed individually to maintain adequate scientific data collection at the least severe level of fluid restriction.

Pattern and Speed Interact to Hide Moving Prey.

Evolutionary biologists have long been fascinated by camouflage patterns that help animals reduce their chances of being detected by predators [1-4]. However, patterns that hide prey when they remain stationary, such as those that match their backgrounds [5, 6], are rendered ineffective once prey are moving [7-10]. The question remains: can a moving animal ever be patterned in a way that helps reduce detection by predators? One long-standing idea is that high-contrast patterns with repeated elements, such as stripes, which are highly visible when prey are stationary, can actually conceal prey when they move fast enough [11-14]. This is predicted by the "flicker fusion effect," which occurs when prey move with sufficient speed that their pattern appears to blur, making them appear more featureless and become less conspicuous against the background [2, 8]. However, although this idea suggests a way to camouflage moving prey, it has not been empirically tested, and it is not clear that it would work at speeds that are biologically relevant to a predator [13]. Combining psychophysics and behavioral approaches, we show that speed and pattern interact to determine the detectability of prey to the praying mantis (Sphodromantis lineola) and, crucially, that prey with high-contrast stripes become less visible than prey with background-matching patterns when moving with sufficient speed. We show that stripes can reduce the detection of moving prey by exploiting the spatiotemporal limitations of predator perception, and that the camouflaging effect of a pattern depends upon the speed of prey movement.

Testing the feasibility of the startle-first route to deimatism.

Many prey species perform deimatic displays that are thought to scare or startle would-be predators, or elicit other reflexive responses that lead to attacks being delayed or abandoned. The form of these displays differs among species, but often includes prey revealing previously-hidden conspicuous visual components. The evolutionary route(s) to deimatism are poorly understood, but it has recently been suggested that the behavioural component of the displays evolves first followed by a conspicuous visual component. This is known as the "startle-first hypothesis". Here we use an experimental system in which naïve domestic chicks forage for artificial deimatic prey to test the two key predictions of this hypothesis: (1) that movement can deter predators in the absence of conspicuously coloured display components; and, (2) that the combination of movement and conspicuously coloured display components is more effective than movement alone. We show that both these predictions hold, but only when the movement is fast. We thus provide evidence for the feasibility of 'the startle-first hypothesis' of the evolution of deimatism.

Handling method alters the hedonic value of reward in laboratory mice.

Mice are the most widely used model species for drug discovery and scientific research. Consequently, it is important to refine laboratory procedures and practices to ensure high standards of welfare and scientific data quality. Recent studies have identified that the standard practice of handling laboratory mice by their tails increases behaviours indicative of anxiety, which can be overcome by handling mice using a tunnel. However, despite clear negative effects on mice's behaviour, tunnel handling has yet to be widely implemented. In this study, we provide the first evidence that tail handling also reduces mice's responses to reward. Anhedonia is a core symptom of clinical depression, and is measured in rodents by assessing how they consume a sucrose solution: depressed mice consume less sucrose and the size of their licking bouts when drinking (their 'lick cluster sizes') also tend to be smaller. We found that tail handled mice showed more anhedonic responses in both measures compared to tunnel handled mice, indicative of a decreased responsiveness to reward and potentially a more depressive-like state. Our findings have significant implications for the welfare of laboratory mice as well as the design and interpretation of scientific studies, particularly those investigating or involving reward.

A critique of comparative studies of brain size.

In recent years, there have been over 50 comparative analyses carried out in which social or ecological variables have been used to explain variation in whole brain size, or a part thereof, in a range of vertebrate species. Here, we review this body of work, pointing out that there are a number of substantial problems with some of the assumptions that underpin the hypotheses (e.g. what brain size means), with the data collection and with the ways in which the data are combined in the analyses. These problems are particularly apparent in those analyses in which attempts are made to correlate complex behaviour with parts of the brain that carry out multiple functions. We conclude that now is the time to substantiate these results with data from experimental manipulations.

The use of preferred social stimuli as rewards for rhesus macaques in behavioural neuroscience.

Macaques are often motivated to perform in neuroscientific experiments by implementing fluid restriction protocols. Daily access to water is controlled and the monkeys are rewarded with droplets of fluid for performing correct trials in the laboratory. Although these protocols are widely used and highly effective, it is important from a 3Rs perspective to investigate refinements that may help to lessen the severity of the fluid restriction applied. We assessed the use of social stimuli (images of conspecifics) as rewards for four rhesus macaques performing simple cognitive tasks. We found that individual preferences for images of male faces, female perinea and control stimuli could be identified in each monkey. However, using preferred images did not translate into effective motivators on a trial-by-trial basis: animals preferred fluid rewards, even when fluid restriction was relaxed. There was no difference in the monkeys' performance of a task when using greyscale versus colour images. Based on our findings, we cannot recommend the use of social stimuli, in this form, as a refinement to current fluid restriction protocols. We discuss the potential alternatives and possibilities for future research.

The Impact of Detoxification Costs and Predation Risk on Foraging: Implications for Mimicry Dynamics.

Prey often evolve defences to deter predators, such as noxious chemicals including toxins. Toxic species often advertise their defence to potential predators by distinctive sensory signals. Predators learn to associate toxicity with the signals of these so-called aposematic prey, and may avoid them in future. In turn, this selects for mildly toxic prey to mimic the appearance of more toxic prey. Empirical evidence shows that mimicry could be either beneficial ('Mullerian') or detrimental ('quasi-Batesian') to the highly toxic prey, but the factors determining which are unknown. Here, we use state-dependent models to explore how tri-trophic interactions could influence the evolution of prey defences. We consider how predation risk affects predators' optimal foraging strategies on aposematic prey, and explore the resultant impact this has on mimicry dynamics between unequally defended species. In addition, we also investigate how the potential energetic cost of metabolising a toxin can alter the benefits to eating toxic prey and thus impact on predators' foraging decisions. Our model predicts that both how predators perceive their own predation risk, and the cost of detoxification, can have significant, sometimes counterintuitive, effects on the foraging decisions of predators. For example, in some conditions predators should: (i) avoid prey they know to be undefended, (ii) eat more mildly toxic prey as detoxification costs increase, (iii) increase their intake of highly toxic prey as the abundance of undefended prey increases. These effects mean that the relationship between a mimic and its model can qualitatively depend on the density of alternative prey and the cost of metabolising toxins. In addition, these effects are mediated by the predators' own predation risk, which demonstrates that, higher trophic levels than previously considered can have fundamental impacts on interactions among aposematic prey species.

The benefits of being toxic to deter predators depends on prey body size.

Many prey have evolved toxins as a defense against predation. Those species that advertise their toxicity to would-be predators with conspicuous warning signals are known as "aposematic." Investment in toxicity by aposematically signaling prey is thought to underpin how aversive prey are to predators; increasing toxicity means that predators learn to avoid prey faster and attack them at lower rates. However, predators' foraging decisions on aposematic prey are determined not only by their toxicity, but also by their nutrient content: predators can trade-off the costs of ingesting toxin with the benefits of acquiring nutrients. Prey body size is a cue that positively correlates with nutrient content, and that varies within and between aposematic species. We predicted that a dose of quinine (known to be toxic to birds) would be a more effective deterrent to avian predators when prey were small compared with when they were large, and that the benefits of possessing toxin would be greater for small-bodied prey. Using an established laboratory protocol of European starlings (Sturnus vulgaris) foraging on mealworms (Tenebrio molitor), we found evidence for increased protection from a dose of quinine for small-bodied compared with large-bodied prey. This shows that larger prey need more toxin to attain the same level of defense as smaller prey, which has implications for the evolution of aposematism and mimicry.

Physiological, Behavioral, and Scientific Impact of Different Fluid Control Protocols in the Rhesus Macaque (Macaca mulatta).

Rhesus macaques are an important model in behavioral neuroscience due to their advanced cognitive abilities. To motivate animals to engage in complex tasks, fluid rewards, in conjunction with fluid control protocols, are often used. The impact of these protocols on animal welfare is controversial. We compared two fluid control protocols against a protocol providing free access to water and evaluated the impacts on physiological states of hydration, behavioral measures of welfare, and scientific output. Blood physiology did not significantly differ between any of the protocols, and urine measures were indicative of well functioning, healthy kidneys. Changes in behaviors were limited, the main one being an increase in motivation to drink on the stricter fluid control protocol, and improved task performance early in the week. Overall, fluid control protocols had little measurable impact on the welfare of rhesus macaques while ensuring that scientific data of high quality could be obtained.