Principles of experimental design for ecology and evolution.

Good experimental design is critical for sound empirical ecology and evolution. However, many contemporary studies fail to replicate at the appropriate biological or organizational level, so causal inference might have less vigorous support than often assumed. Here, I provide a guide for how to identify the appropriate scale of replication for a range of common experimental designs in ecological and evolutionary studies. I discuss the merits of replicating multiple scales of biological organization. I suggest that experimental design be discussed in terms of the scale of replication relative to the scale at which inferences are sought when designing, discussing and reviewing experiments in ecology and evolution. I also suggest that more conversations about experimental design are needed, and I hope this piece stimulates such conversation.

Per capita sperm metabolism is density dependent.

From bacteria to metazoans, higher density populations have lower per capita metabolic rates than lower density populations. The negative covariance between population density and metabolic rate is thought to represent a form of adaptive metabolic plasticity. A relationship between density and metabolism was actually first noted 100 years ago, and was focused on spermatozoa; even then, it was postulated that adaptive plasticity drove this pattern. Since then, contemporary studies of sperm metabolism specifically assume that sperm concentration has no effect on metabolism and that sperm metabolic rates show no adaptive plasticity. We did a systematic review to estimate the relationship between sperm aerobic metabolism and sperm concentration, for 198 estimates spanning 49 species, from protostomes to humans from 88 studies. We found strong evidence that per capita metabolic rates are concentration dependent: both within and among species, sperm have lower metabolisms in dense ejaculates, but increase their metabolism when diluted. On average, a 10-fold decrease in sperm concentration increased per capita metabolic rate by 35%. Metabolic plasticity in sperm appears to be an adaptive response, whereby sperm maximize their chances of encountering eggs.

Fertilization Mode Covaries with Body Size.

AbstractThe evolution of internal fertilization has occurred repeatedly and independently across the tree of life. As it has evolved, internal fertilization has reshaped sexual selection and the covariances among sexual traits, such as testes size, and gamete traits. But it is unclear whether fertilization mode also shows evolutionary associations with traits other than primary sex traits. Theory predicts that fertilization mode and body size should covary, but formal tests with phylogenetic control are lacking. We used a phylogenetically controlled approach to test the covariance between fertilization mode and adult body size (while accounting for latitude, offspring size, and offspring developmental mode) among 1,232 species of marine invertebrates from three phyla. Within all phyla, external fertilizers are consistently larger than internal fertilizers: the consequences of fertilization mode extend to traits that are only indirectly related to reproduction. We suspect that other traits may also coevolve with fertilization mode in ways that remain unexplored.

How and Why Does Metabolism Scale with Body Mass?

Most explanations for the relationship between body size and metabolism invoke physical constraints; such explanations are evolutionarily inert, limiting their predictive capacity. Contemporary approaches to metabolic rate and life history lack the pluralism of foundational work. Here, we call for reforging of the lost links between optimization approaches and physiology.

Metabolic evolution in response to interspecific competition in a eukaryote.

Competition drives rapid evolution, which, in turn, alters the trajectory of ecological communities. These eco-evolutionary dynamics are increasingly well-appreciated, but we lack a mechanistic framework for identifying the types of traits that will evolve and their trajectories. Metabolic theory offers explicit predictions for how competition should shape the (co)evolution of metabolism and size, but these are untested, particularly in eukaryotes. We use experimental evolution of a eukaryotic microalga to examine how metabolism, size, and demography coevolve under inter- and intraspecific competition. We find that the focal species evolves in accordance with the predictions of metabolic theory, reducing metabolic costs and maximizing population carrying capacity via changes in cell size. The smaller-evolved cells initially had lower population growth rates, as expected from their hyper-allometric metabolic scaling, but longer-term evolution yielded important departures from theory: we observed improvements in both population growth rate and carrying capacity. The evasion of this trade-off arose due to the rapid evolution of metabolic plasticity. Lineages exposed to competition evolved more labile metabolisms that tracked resource availability more effectively than lineages that were competition-free. That metabolic evolution can occur is unsurprising, but our finding that metabolic plasticity also co-evolves rapidly is new. Metabolic theory provides a powerful theoretical basis for predicting the eco-evolutionary responses to changing resource regimes driven by global change. Metabolic theory needs also to be updated to incorporate the effects of metabolic plasticity on the link between metabolism and demography, as this likely plays an underappreciated role in mediating eco-evolutionary dynamics of competition.

Copepod life history evolution under high- and low-food regimes.

Copepods play a critical role in the carbon cycle of the planet - they mediate the sequestration of carbon into the deep ocean and are the trophic link between phytoplankton and marine food webs. Global change stressors that decrease copepod productivity create the potential for catastrophic positive feedback loops. Accordingly, a growing list of studies examine the evolutionary capacity of copepods to adapt to the two primary stressors associated with global change: warmer temperatures and lower pH. But the evolutionary capacity of copepods to adapt to changing food regimes, the third major stressor associated with global change, remains unknown. We used experimental evolution to explore how a 10-fold difference in food availability affects life history evolution in the copepod, Tisbe sp. over 2 years, and spanning 30+ generations. Different food regimes evoked evolutionary responses across the entire copepod life history: we observed evolution in body size, size-fecundity relationships and offspring investment strategies. Our results suggest that changes to food regimes reshape life histories and that cryptic evolution in traits such as body size is likely. We demonstrate that evolution in response to changes in ocean productivity will alter consumer life histories and may distort trophic links in marine foodchains. Evolution in response to changing phytoplankton productivity may alter the efficacy of the global carbon pump in ways that have not been anticipated until now.

Fundamental Niche Narrows through Larval Stages of a Filter-Feeding Marine Invertebrate.

AbstractOntogenetic niche theory predicts that resource use should change across complex life histories. To date, studies of ontogenetic shifts in food niches have mainly focused on a few systems (e.g., fish), with less attention on organisms with filter-feeding larval stages (e.g., marine invertebrates). Recent studies suggest that filter-feeding organisms can select specific particles, but our understanding of whether niche theory applies to this group is limited. We characterized the fundamental niche (i.e., feeding proficiency) by examining how niche breadth changes across the larval stages of the filter-feeding marine polychaete Galeolaria caespitosa. Using a no-choice experimental design, we measured feeding rates of trochophore, intermediate-stage, and metatrochophore larvae on the prey phytoplankton species Nannochloropsis oculata, Tisochrysis lutea, Dunaliella tertiolecta, and Rhodomonas salina, which vary 10-fold in size, from the smallest to the largest. We formally estimated Levins's niche breadth index to determine the relative proportions of each species in the diet of the three larval stages and also tested how feeding rates vary with algal species and stage. We found that early stages eat all four algal species in roughly equal proportions, but niche breadth narrows during ontogeny, such that metatrochophores are feeding specialists relative to early stages. We also found that feeding rates differed across phytoplankton species: the medium-sized cells (Tisochrysis and Dunaliella) were eaten most, and the smallest species (Nannochloropsis) was eaten the least. Our results demonstrate that ontogenetic niche theory describes changes in fundamental niche in filter feeders. An important next step is to test whether the realized niche (i.e., preference) changes during the larval phase as well.

Optimisation and constraint: explaining metabolic patterns in biology.

Constraint-based explanations have dominated theories of size-related patterns in nature for centuries. Explanations for metabolic scaling - the way in which metabolism changes with body mass - have been based on the geometry of circulatory networks through which resources are distributed, the need to dissipate heat produced as a by-product of metabolic processes, and surface-area-to-volume constraints on the flux of nutrients or waste. As an alternative to these constraint-based approaches, we recently developed a new theory that predicts that metabolic allometry arises as a consequence of the optimisation of growth and reproduction to maximise fitness within a finite life. Our theory is free of physical geometric constraints that limit the possibilities available to evolution, and we therefore argue that metabolic allometry can be explained without the need to invoke any of the assumed constraints traditionally imposed by metabolic theories. Our findings also suggest that metabolism, growth and reproduction have co-evolved to maximise fitness (i.e. lifetime reproduction) and that the observed patterns in these fundamental characteristics of life can similarly be explained by optimisation rather than constraint. In this Centenary Commentary, we present an overview of our approach and a critique of its limitations. We propose a suite of empirical tests that we hope will move the field forward, discuss the dangers of model overparameterisation and highlight the need to remain open to non-adaptive hypotheses for the origin of biological patterns.

Life history optimisation drives latitudinal gradients and responses to global change in marine fishes.

Within many species, and particularly fish, fecundity does not scale with mass linearly; instead, it scales disproportionately. Disproportionate intraspecific size-reproduction relationships contradict most theories of biological growth and present challenges for the management of biological systems. Yet the drivers of reproductive scaling remain obscure and systematic predictors of how and why reproduction scaling varies are lacking. Here, we parameterise life history optimisation model to predict global patterns in the life histories of marine fishes. Our model predict latitudinal trends in life histories: Polar fish should reproduce at a later age and show steeper reproductive scaling than tropical fish. We tested and confirmed these predictions using a new, global dataset of marine fish life histories, demonstrating that the risks of mortality shape maturation and reproductive scaling. Our model also predicts that global warming will profoundly reshape fish life histories, favouring earlier reproduction, smaller body sizes, and lower mass-specific reproductive outputs, with worrying consequences for population persistence.

Response to Comments on "Metabolic scaling is the product of life-history optimization".

Froese and Pauly argue that our model is contradicted by the observation that fish reproduce before their growth rate decreases. Kearney and Jusup show that our model incompletely describes growth and reproduction for some species. Here we discuss the costs of reproduction, the relationship between reproduction and growth, and propose tests of models based on optimality and constraint.

Carry-over effects and fitness trade-offs in marine life histories: The costs of complexity for adaptation.

Most marine organisms have complex life histories, where the individual stages of a life cycle are often morphologically and ecologically distinct. Nevertheless, life-history stages share a single genome and are linked phenotypically (by "carry-over effects"). These commonalities across the life history couple the evolutionary dynamics of different stages and provide an arena for evolutionary constraints. The degree to which genetic and phenotypic links among stages hamper adaptation in any one stage remains unclear and yet adaptation is essential if marine organisms will adapt to future climates. Here, we use an extension of Fisher's geometric model to explore how both carry-over effects and genetic links among life-history stages affect the emergence of pleiotropic trade-offs between fitness components of different stages. We subsequently explore the evolutionary trajectories of adaptation of each stage to its optimum using a simple model of stage-specific viability selection with nonoverlapping generations. We show that fitness trade-offs between stages are likely to be common and that such trade-offs naturally emerge through either divergent selection or mutation. We also find that evolutionary conflicts among stages should escalate during adaptation, but carry-over effects can ameliorate this conflict. Carry-over effects also tip the evolutionary balance in favor of better survival in earlier life-history stages at the expense of poorer survival in later stages. This effect arises in our discrete-generation framework and is, therefore, unrelated to age-related declines in the efficacy of selection that arise in models with overlapping generations. Our results imply a vast scope for conflicting selection between life-history stages, with pervasive evolutionary constraints emerging from initially modest selection differences between stages. Organisms with complex life histories should also be more constrained in their capacity to adapt to global change than those with simple life histories.

Mapping the correlations and gaps in studies of complex life histories.

For species with complex life histories, phenotypic correlations between life-history stages constrain both ecological and evolutionary trajectories. Studies that seek to understand correlations across the life history differ greatly in their experimental approach: some follow individuals ("individual longitudinal"), while others follow cohorts ("cohort longitudinal"). Cohort longitudinal studies risk confounding results through Simpson's Paradox, where correlations observed at the cohort level do not match that of the individual level. Individual longitudinal studies are laborious in comparison, but provide a more reliable test of correlations across life-history stages. Our understanding of the prevalence, strength, and direction of phenotypic correlations depends on the approaches that we use, but the relative representation of different approaches remains unknown. Using marine invertebrates as a model group, we used a formal, systematic literature map to screen 17,000+ papers studying complex life histories, and characterized the study type (i.e., cohort longitudinal, individual longitudinal, or single stage), as well as other factors. For 3315 experiments from 1716 articles, 67% focused on a single stage, 31% were cohort longitudinal and just 1.7% used an individual longitudinal approach. While life-history stages have been studied extensively, we suggest that the field prioritize individual longitudinal studies to understand the phenotypic correlations among stages.

Macroevolutionary patterns in marine hermaphroditism.

Most plants and many animals are hermaphroditic; whether the same forces are responsible for hermaphroditism in both groups is unclear. The well-established drivers of hermaphroditism in plants (e.g., seed dispersal potential, pollination mode) have analogues in animals (e.g., larval dispersal potential, fertilization mode), allowing us to test the generality of the proposed drivers of hermaphroditism across both groups. Here, we test these theories for 1153 species of marine invertebrates, from three phyla. Species with either internal fertilization, restricted offspring dispersal, or small body sizes are more likely to be hermaphroditic than species that are external fertilizers, planktonic developers, or larger. Plants and animals show different biogeographical patterns, however: animals are less likely to be hermaphroditic at higher latitudes-the opposite to the trend in plants. Overall, our results suggest that similar forces, namely, competition among offspring or gametes, shape the evolution of hermaphroditism across plants and three invertebrate phyla.

Metabolic scaling is the product of life-history optimization.

Organisms use energy to grow and reproduce, so the processes of energy metabolism and biological production should be tightly bound. On the basis of this tenet, we developed and tested a new theory that predicts the relationships among three fundamental aspects of life: metabolic rate, growth, and reproduction. We show that the optimization of these processes yields the observed allometries of metazoan life, particularly metabolic scaling. We conclude that metabolism, growth, and reproduction are inextricably linked; that together they determine fitness; and, in contrast to longstanding dogma, that no single component drives another. Our model predicts that anthropogenic change will cause animals to evolve decreased scaling exponents of metabolism, increased growth rates, and reduced lifetime reproductive outputs, with worrying consequences for the replenishment of future populations.

Long-term experimental evolution decouples size and production costs in Escherichia coli.

Body size covaries with population dynamics across life's domains. Metabolism may impose fundamental constraints on the coevolution of size and demography, but experimental tests of the causal links remain elusive. We leverage a 60,000-generation experiment in which Escherichia coli populations evolved larger cells to examine intraspecific metabolic scaling and correlations with demographic parameters. Over the course of their evolution, the cells have roughly doubled in size relative to their ancestors. These larger cells have metabolic rates that are absolutely higher, but relative to their size, they are lower. Metabolic theory successfully predicted the relations between size, metabolism, and maximum population density, including support for Damuth's law of energy equivalence, such that populations of larger cells achieved lower maximum densities but higher maximum biomasses than populations of smaller cells. The scaling of metabolism with cell size thus predicted the scaling of size with maximum population density. In stark contrast to standard theory, however, populations of larger cells grew faster than those of smaller cells, contradicting the fundamental and intuitive assumption that the costs of building new individuals should scale directly with their size. The finding that the costs of production can be decoupled from size necessitates a reevaluation of the evolutionary drivers and ecological consequences of biological size more generally.

Predicting the response of disease vectors to global change: The importance of allometric scaling.

The distribution of disease vectors such as mosquitoes is changing. Climate change, invasions and vector control strategies all alter the distribution and abundance of mosquitoes. When disease vectors undergo a range shift, so do disease burdens. Predicting such shifts is a priority to adequately prepare for disease control. Accurate predictions of distributional changes depend on how factors such as temperature and competition affect mosquito life-history traits, particularly body size and reproduction. Direct estimates of both body size and reproduction in mosquitoes are logistically challenging and time-consuming, so the field has long relied upon linear (isometric) conversions between wing length (a convenient proxy of size) and reproductive output. These linear transformations underlie most models projecting species' distributions and competitive interactions between native and invasive disease vectors. Using a series of meta-analyses, we show that the relationship between wing length and fecundity are nonlinear (hyperallometric) for most mosquito species. We show that whilst most models ignore reproductive hyperallometry (with respect to wing length), doing so introduces systematic biases into estimates of population growth. In particular, failing to account for reproductive hyperallometry overestimates the effects of temperature and underestimates the effects of competition. Assuming isometry also increases the potential to misestimate the efficacy of vector control strategies by underestimating the contribution of larger females in population replenishment. Finally, failing to account for reproductive hyperallometry and variation in body size can lead to qualitative errors via the counter-intuitive effects of Jensen's inequality. For example, if mean sizes decrease, but variance increases, then reproductive outputs may actually increase. We suggest that future disease vector models incorporate hyperallometric relationships to more accurately predict changes in mosquito distribution in response to global change.

Reproductive hyperallometry and managing the world's fisheries.

Marine fisheries are an essential component of global food security, but many are close to their limits and some are overfished. The models that guide the management of these fisheries almost always assume reproduction is proportional to mass (isometry), when fecundity generally increases disproportionately to mass (hyperallometry). Judged against several management reference points, we show that assuming isometry overestimates the replenishment potential of exploited fish stocks by 22% (range: 2% to 78%) for 32 of the world's largest fisheries, risking systematic overharvesting. We calculate that target catches based on assumptions of isometry are more than double those based on assumptions of hyperallometry for most species, such that common reference points are set twice as high as they should be to maintain the target level of replenishment. We also show that hyperallometric reproduction provides opportunities for increasing the efficacy of tools that are underused in standard fisheries management, such as protected areas or harvest slot limits. Adopting management strategies that conserve large, hyperfecund fish may, in some instances, result in higher yields relative to traditional approaches. We recommend that future assessment of reference points and quotas include reproductive hyperallometry unless there is clear evidence that it does not occur in that species.

Metabolism drives demography in an experimental field test.

Metabolism should drive demography by determining the rates of both biological work and resource demand. Long-standing "rules" for how metabolism should covary with demography permeate biology, from predicting the impacts of climate change to managing fisheries. Evidence for these rules is almost exclusively indirect and in the form of among-species comparisons, while direct evidence is exceptionally rare. In a manipulative field experiment on a sessile marine invertebrate, we created experimental populations that varied in population size (density) and metabolic rate, but not body size. We then tested key theoretical predictions regarding relationships between metabolism and demography by parameterizing population models with lifetime performance data from our field experiment. We found that populations with higher metabolisms had greater intrinsic rates of increase and lower carrying capacities, in qualitative accordance with classic theory. We also found important departures from theory-in particular, carrying capacity declined less steeply than predicted, such that energy use at equilibrium increased with metabolic rate, violating the long-standing axiom of energy equivalence. Theory holds that energy equivalence emerges because resource supply is assumed to be independent of metabolic rate. We find this assumption to be violated under real-world conditions, with potentially far-reaching consequences for the management of biological systems.

Larger cells have relatively smaller nuclei across the Tree of Life.

Larger cells have larger nuclei, but the precise relationship between cell size and nucleus size remains unclear, and the evolutionary forces that shape this relationship are debated. We compiled data for almost 900 species - from yeast to mammals - at three scales of biological organisation: among-species, within-species, and among-lineages of a species that was artificially selected for cell size. At all scales, we showed that the ratio of nucleus size to cell size (the 'N: C' ratio) decreased systematically in larger cells. Size evolution appears more constrained in nuclei than cells: cell size spans across six orders of magnitude, whereas nucleus size varies by only three. The next important challenge is to determine the drivers of this apparently ubiquitous relationship in N:C ratios across such a diverse array of organisms.