Scared fitless: Context-dependent response of fear to loss of predators over evolutionary time in Drosophila melanogaster

Non-consumptive effects of predators, including the effects of fear, can play an important role in animal ecology, influencing offspring development (Lardner 2000; Dahl and Peckarsky 2002; Benard 2004), adult fitness (Roitberg et al. 1979; Ylönen 1989; Dixon and Agarwala 1999; Gallagher et al. 2016), population dynamics (Lima 1998; Preisser et al. 2005; Creel et al. 2009), and even ecosystem function (Beckerman et al. 1997; Schmitz et al. 1997; Suraci et al. 2016). When risk of predation is high, animals spend more time being vigilant and less time foraging (Creel and Christianson 2008; Sheriff et al. 2009; Wirsing and Ripple 2011), leading to poorer body condition and physiological stress. Averaged across many studies, evidence suggests that the population-level effect of fear on prey mortality can be equivalent to the effect of direct predator consumption (Preisser and Bolnick 2008).

Although fear can have important ecological consequences, what is less clear is whether fear persists in the absence of predators over an evolutionary timescale. Understanding the differential evolution of naiveté might help assess the impact and success of invasive species, as the rapid evolution of fearfulness to novel, invasive predators would allow native prey to rapidly respond to such invasions (Sih et al. 2010). On the one hand, fear of predation diminishes in the absence of predators (Blumstein and Daniel 2005; Binz et al. 2014), with riskier individuals outcompeting less risky individuals for food and mates. Classic examples come from islands where naïve species are essentially fearless (Darwin 1839; Rödl et al. 2007). Indeed, Darwin’s observation that animals on remote islands were unafraid of people was an impetus for his theory of natural selection, as he posited that escape behaviour was diminished where predators were rare due to the loss of costly escape responses in the absence of strong natural selection to maintain them (Darwin 1839; Cooper et al. 2014). On the other hand, many laboratory animals, such as rats, retain innate fear of predators, such as cats, after thousands of generations without predator exposure (Blanchard et al. 1990; Zangrossi and File 1992; Dielenberg et al. 2001). A major difference between laboratory and natural populations is that the latter are often limited by food, leading to costs associated with fearfulness that are not typically present in well-fed laboratory populations. Consequently, we might expect strong selection against fear in wild, but not laboratory, populations.

To examine the evolution of fear response, we investigated how cues from a mantid predator impact the fitness of Drosophila evolved with and without predators. Drosophila were exposed to the scent of a mantid predator either in a simulated breeding season, a simulated non-breeding season, or both. Fear responses are typically multimodal, and the effect of fear from the mantid is likely due to a combination of vibrations from the mantid, the scent of the mantid, and the scent of dead flies defecated by the mantid. Previously, we demonstrated that exposing naïve Drosophila to a predator (i) caused increased abdominal lifting, an antipredator behaviour in Drosophila (Parigi et al. 2014), (ii) reduced fecundity in the following breeding season, and (iii) quicker offspring development but reduced offspring adult mass (Betini et al. 2013; Elliott et al. 2016). The impact on offspring development was due to maternal effects from stressed mothers, as offspring did not emerge until the mantids were removed (Elliott et al. 2016). Thus, fear in the non-breeding season impacted fecundity in the breeding season and fear in mothers impacted offspring development in flies that had not been exposed to predators for many generations.

To understand the evolution of fear, we examined the two endpoints highlighted by our previous study (fecundity and development time) in lineages differing in the time since they had been exposed to predators. We compared three sets of lineages: one that had been purchased from a stock population centre (stock), a population derived from the wild and evolved without mantid predators (control-evolved), and a population derived from the same ancestral base population evolved with mantid predators (predator-evolved). We predicted that the responses (fecundity and emergence time) would be higher in the predator-evolved populations than in the control-evolved or stock populations because the latter two populations had not evolved to recognize mantid cues.

Fecundity depended on the treatment (F _2,192 = 61.2, p < 0.0001) and evolutionary lineage (F _2,192 = 190.5, p < 0.0001) but not their interaction (F _3,192 = 0.92, p = 0.43). Fecundity was lower in the predator-evolved and control-evolved lineages than the stock lineage (Fig. 2a). In addition, compared with the control, fecundity in all three lineages was reduced with exposure to mantid cues during the non-breeding season. Furthermore, fecundity was also reduced by exposure to mantid cues during the breeding season for the predator-evolved and control-evolved lineages (Fig. 1a). When excluding the stock lineage, fecundity depended on treatment (F _2,84 = 5.39, p = 0.006) and evolutionary line (mantid vs. control, F _1,84 = 5.04, p = 0.03) but not their interaction (F _2,84 = 0.05, p = 0.95). Thus, fecundity was always lower by a proportionately similar amount during the non-breeding mantid treatment but not the control or breeding mantid treatments, regardless of evolutionary line, but fecundity was also lower in the evolved lines than in the stock lines.

The proportion of individuals with early emergence depended on the treatment (F _2,192 = 34.5, p < 0.0001) and the interaction between the treatment and lineage (F _3,192 = 7.05, p = 0.0002) but not lineage itself (F _2,192 = 2.37, p = 0.10). Within lineages, the proportion of individuals with early emergence was greater in the breeding treatment relative to the non-breeding and control treatments (Fig. 2b). For the predator manipulation during the breeding season, the proportion of individuals with early emergence was greater for the predator-evolved lineage relative to the stock and control-evolved lineages (Fig. 2b). The proportion of individuals with early emergence was also higher in the control treatment for the control-evolved and stock lineages than in the predator-evolved lineages, and during the non-breeding mantid treatment for the stock lineage relative to the predator-evolved lineage. When excluding the stock lineage, the proportion of individuals with early emergence depended on treatment (F _2,84 = 43.7, p < 0.00001) and the interaction between the treatment and line (F _2,84 = 13.04, p = 0.00001) but not the line itself (F _1,84 = 0.38, p = 0.54).

Activity was independent of the treatment for the entire 96 h (F _3,25 = 0.72, p = 0.62) and when we only examined the first 24 h (F _3,25 = 0.55, p = 0.65; Fig. 3).

Although we predicted that the carryover effect on both fecundity and development time would be higher in the predator-evolved populations than in the control-evolved or stock populations, we only observed a higher response in development time. We suggest that predator-evolved flies likely had greater earlier emergence than the other two groups in response to a perceived high cost to reduced foraging opportunities. In other words, this context-dependent (i.e., age-dependent) response could be associated with fearful behavior (i.e., reduced foraging) at early-life stages. In captivity, adult Drosophila are given food ad libitum. Under typical captive conditions, an adult Drosophila that is slightly more vigilant will still be able to feed to satiety and have no worse fecundity than one that is less vigilant, potentially explaining why we found no effect of lineage on fecundity and no difference in fecundity between the breeding mantid and control treatments. For instance, in the presence of mantids, the Drosophila showed similar levels of activity as during the absence of mantids. However, an earlier study demonstrated that they spent more time lifting their abdomens as a signal of vigilance (Parigi et al. 2014). Unless more time spent lifting led to higher energy expenditure, similar levels of activity implies that the reduced body mass at the end of the non-breeding season is due to reduced foraging associated with increased vigilance, which would have little cost to a typical laboratory Drosophila fed ad libitum (unlike our non-breeding season). In contrast, larval Drosophila in captivity must eat voraciously to develop rapidly, attain maximum adult body size to gain mates or produce eggs (Mueller and Joshi 2000; Betini et al. 2013, 2014), and avoid being cannibalized under high densities (Vijendravarma et al. 2013). Even a minor reduction in foraging efficiency due to false fear could have negative consequences, explaining why development time, and consequently adult body size (Elliott et al. 2016), were affected by exposure to predators. Given the differences in development time among lineages, differences in evolutionary background may explain why some studies find strong responses to fear and others only weak responses (Preisser et al. 2005; Preisser and Bolnick 2008).

The different evolutionary histories impacted the relative fecundity of the different lineages. The control lineage consisted of flies that had evolved for thousands of generations in an environment where they had only 3 d to reproduce before being destroyed (and the offspring used for the next generation). In contrast, the control-evolved and predator-evolved lineages were maintained in overlapping generations (fresh food added every 5 d but no flies experimentally culled). Therefore, the control lineage was potentially selected for high immediate fecundity compared with the evolved lineages. Similarly, the predator-evolved flies tended to have higher fecundity than the control-evolved flies. Those lineages with higher extrinsic mortality (e.g., predator-evolved) tend to evolve more rapid senescence and higher fecundity (Zwaan et al. 1995; Stearns et al. 2000). Similarly, predator-evolved flies evolved delayed emergence in the absence of predators compared with control-evolved flies. Possibly, because the larvae were not consumed by the mantids, they were selected to emerge later and larger (more fecund). Despite the very different evolutionary histories of our lines, we found that fear impacted fecundity and development time for all three lineages.

The foraging cost of being fearful explains why captive animals may retain fear responses to predators, whereas other animals, such as those on islands, often do not (Blanchard et al. 1990; Zangrossi and File 1992; Dielenberg et al. 2001; Rödl et al. 2007). Captive animals fed ad libitum, such as is common in Drosophila experimental cultures, face few fitness costs if they occasionally delay eating because they detect a scent reminiscent of a predator. In contrast, animals living on islands are limited by competition for food and exhibit classic K-selected traits such as low reproductive output and increased longevity (Tamarin 1977; Austad 1993). Thus, it is unsurprising that island species such as the Falkland Islands fox (Dusicyon australis) and dodo (Raphus cucullatus) evolved to have no fear of predators—an evolutionary trajectory that famously doomed these species once predators arrived. The evolution of naiveté also occurs in invertebrates, such as deaf moths living on bat-free islands (Fullard 1994) and zooplankton living in freshwater ponds that are eliminated by introduced predators (Cox and Lima 2006; Amundsen et al. 2009). We would predict that island species in which foraging competition during adulthood plays less of a role would retain fear in the absence of predators.

One potential criticism of our system is that the breeding season was shorter than the non-breeding season. However, this was purposeful as we attempted to create an ecologically relevant experimental system, and breeding seasons are shorter than non-breeding seasons for many wild animals (e.g., most migrating birds, and insects in northern temperate environments). Consequently, the flies in the breeding season were exposed to the predator for a shorter period than the flies in the non-breeding season, which may account for the differences in sensitivity to the predator treatments. A longer breeding season would be unlikely to change the impact of the predator on fecundity as ∼80% of eggs are laid within 6 h of exposure of the females to the medium (depending on when the females are added relative to light–dark cycles). In addition, larvae hatch after ∼22 h, and as they grow they cannibalize eggs that are subsequently laid. The control treatment followed the same protocols as the other two treatments (see also the next section) but did not have a mantid added in either season (Fig. 1). An additional potential criticism is that the form of mantid cue was different between non-breeding and breeding seasons, with the mantid being continually present during the non-breeding season, and only the scent remaining in the breeding season. However, the response in both cases was similar along a gradient of mantid sizes and presumably chemical cues associated with the mantids, meaning that the response to the cue was likely some form of threshold function (Elliott et al. 2016). Therefore, we must have been near the maximum threshold response in both cases, and the responses were thus apparently similar. Future studies should examine the relationship between the fear-like response and exposure to predator cues.

We provide evidence that fear impacted carryover effects across three different evolutionary contexts. Indeed, given that we measured a direct reduction in fecundity of 20%–25% and a likely reduction of subsequent fecundity of ∼10% (based on advancement in development time leading to reductions in adult size; Elliott et al. 2016), the total effect of fear on an equilibrium population size of ∼200 flies in our environment was likely similar to the effect of direct mortality to predators (∼4 flies/d; I. Dworkin, personal observation, 2014). Several recent studies have documented population declines of species in seasonal environments following increases in predator abundance (Ydenberg et al. 2004; Creel and Christianson 2008). We speculate that cross-seasonal carryover effects of fear on fecundity or offspring growth may play a role in such circumstances.

We add to the growing consensus that fear can impact fitness within and across seasons. Combined with our earlier experiments (Elliott et al. 2016), there is repeatable evidence that the fear of a predator in the non-breeding season leads to lower condition of individuals at the end of the non-breeding season, which then impacts subsequent fecundity. There is also clear evidence that the fear of a predator in the breeding season leads to earlier emergence and smaller flies. Thus, fear amplifies carryover effects across seasons and maternal effects from mother to offspring. Despite those fitness costs, the seasonal carryover effects are maintained for many generations in the absence of mantids, presumably because increased vigilance comes at little cost in typical well-fed lab environments (in contrast to our non-breeding season). In contrast, the reduced fitness costs associated with maternal effects presumably led to the evolution of fearlessness by larvae in the absence of mantids. The context-dependent results may explain why, for instance, some prey species remain resilient to invasive predators while others rapidly go extinct. Regardless, additional studies examining how different contexts impact the evolution of emotions, including fear, would be fruitful avenues for investigation.

We thank B. Felipe, R. Scarfone, L. Teeger, A. Van Natto, and R. Wolach for assistance in the lab. This research was funded by both a postdoctoral fellowship to KHE and Discovery grants to DRN and ID from the Natural Sciences and Engineering Research Council, and a University of Guelph Research Chair to DRN.