Karen M. Warkentin

Wasp predation and wasp-induced hatching of red-eyed treefrog eggs.

Eggs often suffer high levels of predation and, compared with older animals, embryos have few options available for antipredator defence. None the less, hatchlings can escape from many predators to which eggs are vulnerable. I studied early hatching as an antipredator defence of red-eyed treefrog embryos, Agalychnis callidryas, in response to egg predation by social wasps (Polybia rejecta). Red-eyed treefrogs attach their eggs to vegetation overhanging water, where they are exposed to arboreal and aerial predators. Wasps attacked half the egg clutches and killed almost a quarter of the eggs I monitored at a natural breeding site in Panama. Hatching tadpoles fall into the water, where they face aquatic predators. As predicted from improved survival of older hatchlings with aquatic predators, most undisturbed eggs hatched relatively late. However, many younger embryos directly attacked by wasps hatched immediately. Embryos attacked by wasps hatched as much as a third younger than the peak undisturbed hatching age, and most hatching embryos escaped. Thus hatching is an effective defence against wasp predation, and plasticity in hatching stage allows embryos to balance risks from stage-specific egg and larval predators. Wasp-induced hatching is behaviourally similar to the snake-induced hatching previously described in A. callidryas, but occurs in fewer eggs at a time, congruent with the scale of the risk. Individual embryos hatch in response to wasps, which take single eggs, whereas whole clutches hatch in response to snakes, which consume entire clutches. Embryos of A. callidryas thus respond appropriately to graded variation in mortality risks. Copyright 2000 The Association for the Study of Animal Behaviour.

Environmentally cued hatching across taxa: embryos respond to risk and opportunity.

Most animals begin life in eggs, protected and constrained by a capsule, shell, or other barrier. As embryos develop, their needs and abilities change, altering the costs and benefits of encapsulation, and the risks and opportunities of the outside world. When the cost/benefit ratio is better outside the egg, animals should hatch. Adaptive timing of hatching evolves in this context. However, many environmental variables affect the optimal timing of hatching so there is often no consistent best time. Across a broad range of animals, from flatworms and snails to frogs and birds, embryos hatch at different times or at different developmental stages in response to changing risks or opportunities. Embryos respond to many types of cues, assessed via different sensory modalities. Some responses appear simple. Others are surprisingly complex and sophisticated. Parents also manipulate the timing of hatching. The number and breadth of examples of cued hatching suggest that, in the absence of specific information, we should not assume that hatching timing is fixed. Our challenge now is to integrate information on the timing of hatching across taxa to better understand the diversity of patterns and how they are structured in relation to different types of environmental and developmental variation. As starting points for comparative studies, I: (1) suggest a framework based on heterokairy-individual, plastic variation in the rate, timing, or sequence of developmental events and processes-to describe patterns and mechanisms of variation in the timing of hatching; (2) briefly review the distribution of environmentally cued hatching across the three major clades of Bilateria, highlighting the diverse environmental factors and mechanisms involved; and (3) discuss factors that shape the diversity of plastic and fixed timing of hatching, drawing on evolutionary theory on phenotypic plasticity which directs our attention to fitness trade-offs, environmental heterogeneity, and predictive cues. Combining mechanistic and evolutionary perspectives is necessary because development changes organismal interactions with the environment. Integrative and comparative studies of the timing of hatching will improve our understanding of embryos as both evolving and developing organisms.

EGG-KILLING FUNGUS INDUCES EARLY HATCHING OF RED-EYED TREEFROG EGGS

Pathogens can cause substantial mortality of amphibian eggs. If the timing of hatching is phenotypically plastic, embryos could escape from otherwise lethal infections by hatching early. We tested this with the arboreal eggs of red-eyed treefrogs, Agalychnis callidryas. A filamentous ascomycete (Dothideales: Phaeosphaeriaceae) was present on ∼7% of egg clutches collected from a pond in the rain forest in Panama and, when present, killed 40% of the eggs, on average. Inoculation experiments confirmed that the fungus attacked and killed healthy embryos, establishing that this fungus is a pathogen of A. callidryas eggs. As predicted from life history theory, embryos hatched earlier from both naturally infected and inoculated clutches than from fungus-free control clutches. Within infected clutches, live embryos in contact with fungal hyphae hatched before those embryos not in contact with the fungus. Accelerated hatching allowed embryos to survive that otherwise would have been killed, and tadpoles hatched from infected clutches were themselves uninfected. Red-eyed treefrog embryos also hatch early if attacked by predators, apparently in response to vibratory cues. Because fungal infection provides no vibratory stimuli, embryos must respond to different cues in fungus-induced hatching than in predator-induced hatching. The behavioral decision of when to hatch is complex and merits further investigation. Our study indicates that pathogens can influence the timing of life history transitions, as do other stage-specific risks.

How do embryos assess risk? Vibrational cues in predator-induced hatching of red-eyed treefrogs

I examined the role of vibrations in predator-induced early hatching of red-eyed treefrogs, Agalychnis callidryas. The arboreal eggs of A. callidryas hatch up to 30% early if attacked by egg-eating snakes. This induced hatching is a behavioural response that occurs once snakes begin physically disturbing the clutch, and is sufficiently rapid to allow most embryos to escape. Other intense but benign disturbances, such as tropical storms, do not induce such hatching. I used a miniature accelerometer to record vibrations in egg clutches during snake attacks and rainstorms, and analysed the recordings to identify parameters that distinguished disturbance types. Snake-induced vibrations were on average longer, more widely spaced, and of lower frequency than rain-induced vibrations. I performed three sets of vibration playbacks to examine the hatching response of embryos to different vibration patterns. (1) Playbacks of recorded snake attacks elicited more hatching than did rain recordings. (2) I edited snake and rain recordings by moving periods of stillness to clump together rain vibrations and extend intervals, and divide snake vibrations into shorter, more tightly spaced bits. In playbacks, clumped rain elicited more, and divided snake vibrations less hatching than did the original recordings. (3) Bursts of white noise with a constant vibration-to-interval ratio but different cycle durations elicited different levels of hatching. Vibrations alone were sufficient to induce early hatching, without chemical or visual cues from predators. Embryos also clearly distinguished among different vibration patterns and used cues in the gross temporal pattern in making their hatching decision.

AMPHIBIAN EMBRYO AND PARENTAL DEFENSES AND A LARVAL PREDATOR REDUCE EGG MORTALITY FROM WATER MOLD

Water molds attack aquatic eggs worldwide and have been associated with major mortality events in some cases, but typically only in association with additional stressors. We combined field observations and laboratory experiments to study egg stage defenses against pathogenic water mold in three temperate amphibians. Spotted salamanders (Ambystoma maculatum) wrap their eggs in a protective jelly layer that prevents mold from reaching the embryos. Wood frog (Rana sylvatica) egg masses have less jelly but are laid while ponds are still cold and mold growth is slow. American toad (Bufo americanus) eggs experience the highest infection levels. They are surrounded by thin jelly and are laid when ponds have warmed and mold grows rapidly. Eggs of all three species hatched early when infected, yielding smaller and less developed hatchlings. This response was strongest in B. americanus. Precocious hatching increased vulnerability of wood frog hatchlings to invertebrate predators. Finally, despite being potential toad hatchling predators, R. sylvatica tadpoles can have a positive effect on B. americanus eggs. They eat water mold off infected toad clutches, increasing their hatching success.

Plasticity of Hatching in Amphibians: Evolution, Trade-Offs, Cues and Mechanisms

Many species of frogs and salamanders, in at least 12 families, alter their timing of hatching in response to conditions affecting mortality of eggs or larvae. Some terrestrially laid or stranded embryos wait to hatch until they are submerged in water. Some embryos laid above water accelerate hatching if the eggs are dehydrating; others hatch early if flooded. Embryos can hatch early in response to predators and pathogens of eggs or delay hatching in response to predators of larvae; some species do both. The phylogenetic pattern of environmentally cued hatching suggests that similar responses have evolved convergently in multiple amphibian lineages. The use of similar cues, including hypoxia and physical disturbance, in multiple contexts suggests potential shared mechanisms underlying the capacity of embryos to respond to environmental conditions. Shifts in the timing of hatching often have clear benefits, but we know less about the trade-offs that favor plasticity, the mechanisms that enable it, and its evolutionary history. Some potentially important types of cued hatching, such as those involving embryo-parent interactions, are relatively unexplored. I discuss promising directions for research and the opportunities that the hatching of amphibians offers for integrative studies of the mechanisms, ecology and evolution of a critical transition between life-history stages.

Opposite shifts in size at metamorphosis in response to larval and metamorph predators.

Predation risk can cause organisms to alter the timing of life history switch points. Theory suggests that increased risk in an early life stage should select for switching earlier and smaller, while increased risk in the subsequent stage should select for switching later and larger. This framework has frequently been applied to metamorphosis in amphibians, with mixed results. Few studies examining the effect of larval predation risk on metamorphosis have observed the predicted pattern, and no studies, to our knowledge, have examined the effect of increased risk during and after metamorphosis on the timing of this switch point. Here we examine the effect of larval and post-metamorphic predation risk on metamorphosis in the red-eyed treefrog, Agalychnis callidryas. We raised tadpoles in the presence or absence of cues from caged water bugs fed larvae and cues from spiders fed emerging metamorphs. Water bugs are effective larval predators, while spiders are poor larval predators but prey on metamorphs. Furthermore, since spiders forage on the water surface it is possible that tadpoles could assess future risk from this predator. Predators induced opposite shifts in life history. Tadpoles emerged smaller and less developed in response to water bugs, but later and larger in response to spiders. Interestingly, predator effects on larval duration were not independent; tadpoles delayed emerging in response to spiders, but only in the absence of water bugs.

EVOLUTION OF ADAPTIVE PLASTICITY: RISK-SENSITIVE HATCHING IN NEOTROPICAL LEAF-BREEDING TREEFROGS

Adaptive plasticity at switch points in complex life cycles (e.g., hatching, metamorphosis) is well known, but the evolutionary history of such plasticity is not. Particularly unclear is how a single plastic response (e.g., shifts in hatching timing) evolves to respond to different threats and cues (e.g., abiotic and biotic). We conducted a comparative phylogenetic study of hatching plasticity in a group of frogs with arboreal embryos to determine when risk-accelerated hatching evolved in the clade, whether responses to two common egg-stage risks (snake predation and flooding) evolved independently, and whether the overall capacity for hatching plasticity was evolutionarily more conservative than responses to specific cues. Red-eyed treefrogs (Agalychnis callidryas) hatch early to escape from several egg-stage risks but otherwise hatch later, improving larval survival with predators. We reconstructed a phylogeny for Agalychnis and related genera based on three mitochondrial and four nuclear genes. We quantified onset of hatching competence, spontaneous hatching timing, responses to egg-stage risks, and costs of premature hatching in Agalychnis and Pachymedusa. We also assessed hatching plasticity in a basal phyllomedusine, Cruziohyla calcarifer. The capacity to hatch ;30% before the spontaneous hatching age appears ancestral for phyllomedusines, with little change over ;34-50 million years among the species examined. A strong hatching response to flooding, with no mortality of hatching-competent eggs, is similarly ancient and conserved. Premature hatchlings of Agalychnis and Pachymedusa are more vulnerable to fish predation than are full-term hatchlings, indicating a conserved risk trade-off across hatching that would make plasticity advantageous. In contrast, the hatching response to snake attack has undergone major changes at least twice in the Agalychnis-Pachymedusa clade, with two species showing substantially lower escape success than the others. Responses to different threats have thus evolved independently. The capacity for switch point plasticity may be evolutionarily more stable than the response to individual stage-specific threats.

Reproductive mode plasticity: Aquatic and terrestrial oviposition in a treefrog

Diversification of reproductive mode is a major theme in animal evolution. Vertebrate reproduction began in water, and terrestrial eggs evolved multiple times in fishes and amphibians and in the amniote ancestor. Because oxygen uptake from water conflicts with water retention in air, egg adaptations to one environment typically preclude development in the other. Few animals have variable reproductive modes, and no vertebrates are known to lay eggs both in water and on land. We report phenotypic plasticity of reproduction with aquatic and terrestrial egg deposition by a frog. The treefrog Dendropsophus ebraccatus, known to lay eggs terrestrially, also lays eggs in water, both at the surface and fully submerged, and chooses its reproductive mode based on the shade above a pond. Under unshaded conditions, in a disturbed habitat and in experimental mesocosms, these frogs lay most of their egg masses aquatically. The same pairs also can lay eggs terrestrially, on vegetation over water, even during a single night. Eggs can survive in both aquatic and terrestrial environments, and variable mortality risks in each may make oviposition plasticity adaptive. Phylogenetically, D. ebraccatus branches from the basal node in a clade of terrestrially breeding species, nested within a larger lineage of aquatic-breeding frogs. Reproductive plasticity in D. ebraccatus may represent a retained ancestral state intermediate in the evolution of terrestrial reproduction.

The shape of things to come: linking developmental plasticity to post‐metamorphic morphology in anurans

Development consists of growth and differentiation, which can be partially decoupled and can be affected by environmental factors to different extents. In amphibians, variation in the larval environment influences development and causes changes in post‐metamorphic shape. We examined post‐metamorphic consequences, both morphological and locomotory, of alterations in growth and development. We reared tadpoles of two phylogenetically and ecologically distant frog species (the red‐eyed treefrog Agalychnis callidryas and the African clawed frog Xenopus laevis) under different temperatures with ad libitum food supply and under different food levels at a constant temperature. Low temperature and low food levels both resulted in similarly extended larval periods. However, low temperature yielded relatively long‐legged frogs with a lower degree of ossification than warm temperature, whereas low food yielded relatively short‐legged frogs with a higher degree of ossification than high food levels. Such allometric differences had no effect on locomotor performance of juveniles. Our results provide a basis for understanding the relationship between growth, differentiation and post‐metamorphic shape in anurans and help explain many of the discrepancies reported in previous studies.