Mechanisms underlying thermal breadth differ by species in insects from adjacent but thermally distinct streams - A test of the climate variability hypothesis.

The Climate Variability Hypothesis (CVH) predicts that ectotherms from thermally variable climates should have wider thermal tolerances than their counterparts living in stable climates. Although the CVH has been widely supported, the mechanisms underlying wider tolerance traits remain unclear. We test the CVH along with three mechanistic hypotheses that may explain how differences in tolerance limits arise: 1) Short-term Acclimation Hypothesis (mechanism: rapid, reversible plasticity), 2) Long-term Effects Hypothesis (mechanisms: developmental plasticity, epigenetics, maternal effects, or adaptation), and 3) Trade-off Hypothesis (mechanism: trade-off between short- and long-term responses). We tested these hypotheses by measuring CTMIN, CTMAX, and thermal breadths (CTMAX - CTMIN) of aquatic mayfly and stonefly nymphs from adjacent streams with distinctly different levels of thermal variation following acclimation to either cool, control, and warm conditions. In one stream, daily mean temperature varied by about 5 °C annually, whereas in the other, it varied by more than 25 °C. In support of the CVH, we found that mayfly and stonefly nymphs from the thermally variable stream had broader thermal tolerances than those from the thermally stable stream. However, support for the mechanistic hypotheses differed by species. Mayflies appear to rely on long-term strategies for maintaining broader thermal limits, whereas stoneflies achieve broader thermal limits via short-term plasticity. We found no support for the Trade-off Hypothesis.

Plasticity of salmonfly (Pteronarcys californica) respiratory phenotypes in response to changes in temperature and oxygen.

Like all taxa, populations of aquatic insects may respond to climate change by evolving new physiologies or behaviors, shifting their range, exhibiting physiological and behavioral plasticity, or going extinct. We evaluated the importance of plasticity by measuring changes in growth, survival and respiratory phenotypes of salmonfly nymphs (the stonefly Pteronarcys californica) in response to experimental combinations of dissolved oxygen and temperature. Overall, smaller individuals grew more rapidly during the 6-week experimental period, and oxygen and temperature interacted to affect growth in complex ways. Survival was lower for the warm treatment, although only four mortalities occurred (91.6% versus 100%). Nymphs acclimated to warmer temperatures did not have higher critical thermal maxima (CTmax), but those acclimated to hypoxia had CTmax values (in normoxia) that were higher by approximately 1°C. These results suggest possible adaptive plasticity of systems for taking up or delivering oxygen. We examined these possibilities by measuring the oxygen sensitivity of metabolic rates and the morphologies of tracheal gill tufts located ventrally on thoracic segments. Mass-specific metabolic rates of individuals acclimated to warmer temperatures were higher in acute hypoxia but lower in normoxia, regardless of their recent history of oxygen exposure during acclimation. The morphology of gill filaments, however, changed in ways that appeared to depress rates of oxygen delivery in functional hypoxia. Our combined results from multiple performance metrics indicate that rising temperatures and hypoxia may interact to magnify the risks to aquatic insects, but that physiological plasticity in respiratory phenotypes may offset some of these risks.

Temperature dependence of metabolic rate in tropical and temperate aquatic insects: Support for the Climate Variability Hypothesis in mayflies but not stoneflies.

A fundamental gap in climate change vulnerability research is an understanding of the relative thermal sensitivity of ectotherms. Aquatic insects are vital to stream ecosystem function and biodiversity but insufficiently studied with respect to their thermal physiology. With global temperatures rising at an unprecedented rate, it is imperative that we know how aquatic insects respond to increasing temperature and whether these responses vary among taxa, latitudes, and elevations. We evaluated the thermal sensitivity of standard metabolic rate in stream-dwelling baetid mayflies and perlid stoneflies across a ~2,000 m elevation gradient in the temperate Rocky Mountains in Colorado, USA, and the tropical Andes in Napo, Ecuador. We used temperature-controlled water baths and microrespirometry to estimate changes in oxygen consumption. Tropical mayflies generally exhibited greater thermal sensitivity in metabolism compared to temperate mayflies; tropical mayfly metabolic rates increased more rapidly with temperature and the insects more frequently exhibited behavioral signs of thermal stress. By contrast, temperate and tropical stoneflies did not clearly differ. Varied responses to temperature among baetid mayflies and perlid stoneflies may reflect differences in evolutionary history or ecological roles as herbivores and predators, respectively. Our results show that there is physiological variation across elevations and species and that low-elevation tropical mayflies may be especially imperiled by climate warming. Given such variation among species, broad generalizations about the vulnerability of tropical ectotherms should be made more cautiously.

Flow increases tolerance of heat and hypoxia of an aquatic insect.

Recent experiments support the idea that upper thermal limits of aquatic insects arise, at least in part, from a lack of sufficient oxygen: rising temperatures typically stimulate metabolic demand for oxygen more than they increase rates of oxygen supply from the environment. Consequently, factors influencing oxygen supply, like water flow, should also affect thermal and hypoxia tolerance. We tested this hypothesis by measuring the effects of experimentally manipulated flows on the heat and hypoxia tolerance of aquatic nymphs of the giant salmonfly (Plecoptera: Pteronarcys californica), a common stonefly in western North America. As predicted, stoneflies in flowing water (10 cm s-1) tolerated water that was approximately 4°C warmer and that contained approximately 15% less oxygen than did those in standing water. Our results imply that the impacts of climate change on streamflow, such as changes in patterns of precipitation and decreased snowpack, will magnify the threats to aquatic insects from warmer water temperatures and lower oxygen levels.

Narrow thermal tolerance and low dispersal drive higher speciation in tropical mountains.

Species richness is greatest in the tropics, and much of this diversity is concentrated in mountains. Janzen proposed that reduced seasonal temperature variation selects for narrower thermal tolerances and limited dispersal along tropical elevation gradients [Janzen DH (1967) Am Nat 101:233-249]. These locally adapted traits should, in turn, promote reproductive isolation and higher speciation rates in tropical mountains compared with temperate ones. Here, we show that tropical and temperate montane stream insects have diverged in thermal tolerance and dispersal capacity, two key traits that are drivers of isolation in montane populations. Tropical species in each of three insect clades have markedly narrower thermal tolerances and lower dispersal than temperate species, resulting in significantly greater population divergence, higher cryptic diversity, higher tropical speciation rates, and greater accumulation of species over time. Our study also indicates that tropical montane species, with narrower thermal tolerance and reduced dispersal ability, will be especially vulnerable to rapid climate change.

Insects in high-elevation streams: Life in extreme environments imperiled by climate change.

Climate change is altering conditions in high-elevation streams worldwide, with largely unknown effects on resident communities of aquatic insects. Here, we review the challenges of climate change for high-elevation aquatic insects and how they may respond, focusing on current gaps in knowledge. Understanding current effects and predicting future impacts will depend on progress in three areas. First, we need better descriptions of the multivariate physical challenges and interactions among challenges in high-elevation streams, which include low but rising temperatures, low oxygen supply and increasing oxygen demand, high and rising exposure to ultraviolet radiation, low ionic strength, and variable but shifting flow regimes. These factors are often studied in isolation even though they covary in nature and interact in space and time. Second, we need a better mechanistic understanding of how physical conditions in streams drive the performance of individual insects. Environment-performance links are mediated by physiology and behavior, which are poorly known in high-elevation taxa. Third, we need to define the scope and importance of potential responses across levels of biological organization. Short-term responses are defined by the tolerances of individuals, their capacities to perform adequately across a range of conditions, and behaviors used to exploit local, fine-scale variation in abiotic factors. Longer term responses to climate change, however, may include individual plasticity and evolution of populations. Whether high-elevation aquatic insects can mitigate climatic risks via these pathways is largely unknown.

Mountain stoneflies may tolerate warming streams: Evidence from organismal physiology and gene expression.

Rapid glacier recession is altering the physical conditions of headwater streams. Stream temperatures are predicted to rise and become increasingly variable, putting entire meltwater-associated biological communities at risk of extinction. Thus, there is a pressing need to understand how thermal stress affects mountain stream insects, particularly where glaciers are likely to vanish on contemporary timescales. In this study, we measured the critical thermal maximum (CTMAX ) of stonefly nymphs representing multiple species and a range of thermal regimes in the high Rocky Mountains, USA. We then collected RNA-sequencing data to assess how organismal thermal stress translated to the cellular level. Our focal species included the meltwater stonefly, Lednia tumana, which was recently listed under the U.S. Endangered Species Act due to climate-induced habitat loss. For all study species, critical thermal maxima (CTMAX  > 20°C) far exceeded the stream temperatures mountain stoneflies experience (<10°C). Moreover, while evidence for a cellular stress response was present, we also observed constitutive expression of genes encoding proteins known to underlie thermal stress (i.e., heat shock proteins) even at low temperatures that reflected natural conditions. We show that high-elevation aquatic insects may not be physiologically threatened by short-term exposure to warm temperatures and that longer-term physiological responses or biotic factors (e.g., competition) may better explain their extreme distributions.

The thermal breadth of temperate and tropical freshwater insects supports the climate variability hypothesis.

Climate change involves increases in mean temperature and changes in temperature variability at multiple temporal scales but research rarely considers these temporal scales. The climate variability hypothesis (CVH) provides a conceptual framework for exploring the potential effects of annual scale thermal variability across climatic zones. The CVH predicts ectotherms in temperate regions tolerate a wider range of temperatures than those in tropical regions in response to greater annual variability in temperate regions. However, various other aspects of thermal regimes (e.g. diel variability), organisms' size and taxonomic identity are also hypothesised to influence thermal tolerance. Indeed, high temperatures in the tropics have been proposed as constraining organisms' ability to tolerate a wide range of temperatures, implying that high annual maximum temperatures would be associated with tolerating a narrow range of temperatures. We measured thermal regimes and critical thermal limits (CTmax and CTmin) of freshwater insects in the orders Ephemeroptera (mayflies), Plecoptera (stoneflies) and Trichoptera (caddisflies) along elevation gradients in streams in temperate and tropical regions of eastern Australia and tested the CVH by determining which variables were most correlated with thermal breadth (T br = CTmax - CTmin). Consistent with the CVH, T br tended to increase with increasing annual temperature range. T br also increased with body size and T br was generally wider in Plecoptera than in Ephemeroptera or Trichoptera. We also find some support for a related hypothesis, the climate extreme hypothesis (CEH), particularly for predicting upper thermal limits. We found no evidence that higher annual maximum temperature constrained individuals' abilities to tolerate a wide range of temperatures. The support for the CVH we document suggests that temperate organisms may be able to tolerate wider ranges of temperatures than tropical organisms. There is an urgent need to investigate other aspects of thermal regimes, such as diel temperature cycling and minimum temperature.

An Integrative Perspective On the Mechanistic Basis of Context- Dependent Species Interactions.

It has long been known that the outcome of species interactions depends on the environmental context in which they occur. Climate change research has sparked a renewed interest in context-dependent species interactions because rapidly changing abiotic environments will cause species interactions to occur in novel contexts and researchers must incorporate this in their predictions of species' responses to climate change. Here, we argue that predicting how the environment will alter the outcome of species interactions requires an integrative biology approach that focuses on the traits, mechanisms, and processes that bridge disciplines such as physiology, biomechanics, ecology, and evolutionary biology. Specifically, we advocate for quantifying how species differ in their tolerance and performance to both environmental challenges independent of species interactions, and in interactions with other species as a function of the environment. Such an approach increases our understanding of the mechanisms underlying outcomes of species interactions across different environmental contexts. This understanding will help determine how the outcome of species interactions affects the relative abundance and distribution of the interacting species in nature. A general theme that emerges from this perspective is that species are unable to maintain high levels of performance across different environmental contexts because of trade-offs between physiological tolerance to environmental challenges and performance in species interactions. Thus, an integrative biology paradigm that focuses on the trade-offs across environments, the physiological mechanisms involved, and how the ecological context impacts the outcome of species interactions provides a stronger framework to understand why species interactions are context dependent.

Powerhouses in the cold: mitochondrial function during thermal acclimation in montane mayflies.

Mitochondria provide the vast majority of cellular energy available to eukaryotes. Therefore, adjustments in mitochondrial function through genetic changes in mitochondrial or nuclear-encoded genes might underlie environmental adaptation. Environmentally induced plasticity in mitochondrial function is also common, especially in response to thermal acclimation in aquatic systems. Here, we examined mitochondrial function in mayfly larvae (Baetis and Drunella spp.) from high and low elevation mountain streams during thermal acclimation to ecologically relevant temperatures. A multi-substrate titration protocol was used to evaluate different respiratory states in isolated mitochondria, along with cytochrome oxidase and citrate synthase activities. In general, maximal mitochondrial respiratory capacity and oxidative phosphorylation coupling efficiency decreased during acclimation to higher temperatures, suggesting montane insects may be especially vulnerable to rapid climate change. Consistent with predictions of the climate variability hypothesis, mitochondria from Baetis collected at a low elevation site with highly variable daily and seasonal temperatures exhibited greater thermal tolerance than Baetis from a high elevation site with comparatively stable temperatures. However, mitochondrial phenotypes were more resilient than whole-organism phenotypes in the face of thermal stress. These results highlight the complex relationships between mitochondrial and organismal genotypes, phenotypes and environmental adaptation. This article is part of the theme issue 'Linking the mitochondrial genotype to phenotype: a complex endeavour'.

Divergence and constraint in the thermal sensitivity of aquatic insect swimming performance.

Environmental temperature variation may play a significant role in the adaptive evolutionary divergence of ectotherm thermal performance curves (TPCs). However, divergence in TPCs may also be constrained due to various causes. Here, we measured TPCs for swimming velocity of temperate and tropical mayflies (Family: Baetidae) and their stonefly predators (Family: Perlidae) from different elevations. We predicted that differences in seasonal climatic regimes would drive divergence in TPCs between temperate and tropical species. Stable tropical temperatures should favor the evolution of "specialists" that perform well across a narrow range of temperatures. Seasonally, variable temperatures in temperate zones, however, should favor "generalists" that perform well across a broad range of temperatures. In phylogenetically paired comparisons of mayflies and stoneflies, swimming speed was generally unaffected by experimental temperature and did not differ among populations between latitudes, suggesting a maintenance of performance breadth across elevation and latitude. An exception was found between temperate and tropical mayflies at low elevation where climatic differences between latitudes are large. In addition, TPCs did not differ between mayflies and their stonefly predators, except at tropical low elevation. Our results indicate that divergence in TPCs may be constrained in aquatic insects except under the most different thermal regimes, perhaps because of trade-offs that reduce thermal sensitivity and increase performance breadth.

High elevation insect communities face shifting ecological and evolutionary landscapes.

Climate change is proceeding rapidly in high mountain regions worldwide. Rising temperatures will impact insect physiology and associated fitness and will shift populations in space and time, thereby altering community interactions and composition. Shifts in space are expected as insects move upslope to escape warming temperatures and shifts in time will occur with changes in phenology of resident high-elevation insects. Clearly, spatiotemporal shifts will not affect all species equally. Terrestrial insects may have more opportunities than aquatic insects to exploit microhabitats, potentially buffering them from warming. Such responses of insects to warming may also fuel evolutionary change, including hitchhiking of maladaptive alleles and genetic rescue. Together, these considerations suggest a striking restructuring of high-elevation insect communities that remains largely unstudied.

Thermal Acclimation Ability Varies in Temperate and Tropical Aquatic Insects from Different Elevations.

It has long been recognized that populations and species occupying different environments vary in their thermal tolerance traits. However, far less attention has been given to the impact of different environments on the capacity for plastic adjustments in thermal sensitivity, i.e., acclimation ability. One hypothesis is that environments characterized by greater thermal variability and seasonality should favor the evolution of increased acclimation ability compared with environments that are aseasonal or thermally stable. Additionally, organisms under selection for high heat tolerance may experience a trade-off and lose acclimation ability. Few studies have tested these non-mutually exclusive hypotheses at both broad latitudinal and local elevation scales in phylogenetically paired taxa. Here, we measure short-term acclimation ability of the critical thermal maximum (CTMAX) in closely related temperate and tropical mayflies (Ephemeroptera) and stoneflies (Plecoptera) from mountain streams at different elevations. We found that stream temperature was a good predictor of acclimation ability in mayflies, but not in stoneflies. Specifically, tropical mayflies showed reduced acclimation ability compared with their temperate counterparts. High elevation tropical mayflies had greater acclimation ability than low elevation mayflies, which reflected the wider temperature variation experienced in high elevation streams. In contrast, temperate and tropical stoneflies exhibited similar acclimation responses. We found no evidence for a trade-off between heat tolerance and acclimation ability in either taxonomic order. The acclimation response in stoneflies may reflect their temperate origin or foraging mode. In combination with previous studies showing tropical taxa have narrower thermal breadths, these results demonstrate that many lower elevation tropical aquatic insects are more vulnerable to climate warming than their temperate relatives.