Seasonal and daily thermal variation can limit species distributions because of physiological tolerances. Low temperatures are particularly challenging for ectotherms, which use both basal thermotolerance and acclimation, an adaptive plastic response, to mitigate thermal stress. Both basal thermotolerance and acclimation are thought to be important for local adaptation and persistence in the face of climate change. However, the evolutionary independence of basal and plastic tolerances remains unclear. Acclimation can occur over longer (seasonal) or shorter (hours to days) time scales, and the degree of mechanistic overlap is unresolved. Using a midlatitude population of Drosophila melanogaster, we show substantial heritable variation in both short-and long-term acclimation. Rapid cold hardening (short-term plasticity) and developmental acclimation (long-term plasticity) are positively correlated, suggesting shared mechanisms. However, there are independent components of these traits, because developmentally acclimated flies respond positively to short-term acclimation. A strong negative correlation between basal cold tolerance and developmental acclimation suggests that basal cold tolerance may constrain developmental acclimation, whereas a weaker negative correlation between basal cold tolerance and short-term acclimation suggests less constraint. Using genome-wide association mapping, we show the genetic architecture of rapid cold hardening and developmental acclimation responses are nonoverlapping at the SNP and corresponding gene level. However, genes associated with each trait share functional similarities, including genes involved in apoptosis and autophagy, cytoskeletal and membrane structural components, and ion binding and transport. These results indicate substantial opportunity for short-term and long-term acclimation responses to evolve separately from each other and for short-term acclimation to evolve separately from basal thermotolerance.

Regulatory variation in gene expression can be described by cis-and trans-genetic components. Here we used RNA-seq data from a population panel of Drosophila melanogaster test crosses to compare allelic imbalance (AI) in female head tissue between mated and virgin flies, an environmental change known to affect transcription. Indeed, 3048 exons (1610 genes) are differentially expressed in this study. A Bayesian model for AI, with an intersection test, controls type I error. There are 200 genes with AI exclusively in mated or virgin flies, indicating an environmental component of expression regulation. On average 34% of genes within a cross and 54% of all genes show evidence for genetic regulation of transcription. Nearly all differentially regulated genes are affected in cis, with an average of 63% of expression variation explained by the cis-effects. Trans-effects explain 8% of the variance in AI on average and the interaction between cis and trans explains an average of 11% of the total variance in AI. In both environments cis-and trans-effects are compensatory in their overall effect, with a negative association between cis-and trans-effects in 85% of the exons examined. We hypothesize that the gene expression level perturbed by cis-regulatory mutations is compensated through trans-regulatory mechanisms, e. g., trans and cis by trans-factors buffering cis-mutations. In addition, when AI is detected in both environments, cis-mated, cis-virgin, and trans-mated-trans-virgin estimates are highly concordant with 99% of all exons positively correlated with a median correlation of 0.83 for cis and 0.95 for trans. We conclude that the gene regulatory networks (GRNs) are robust and that trans-buffering explains robustness.

Stored products represent an enormous economic output, but insects regularly immigrate into stored products from the surrounding landscape throughout the postharvest supply chain. Long-lasting insecticide-incorporatednetting (LLIN), which usually contains an incorporated pyrethroid, has been used as part of a strategy to reduce the spread of malaria in tropical regions since the 1990s and has only recently been considered for its application in pre- and postharvest agricultural contexts. The goal of this study was to determine how short-term exposure to LLINs in the laboratory impacts the locomotory behavior and mortality for adult Tribolium castaneum (Herbst), (Coleoptera: Tenebrionidae) red flour beetle, and Rhyzopertha dominica (Fauvel), (Coleoptera: Bostrichidae) lesser grain borer, at different periods of time after exposure. Exposure to LLINs resulted in multiple-fold reductions in the distance moved and elevated angular velocity in both species that quickly took effect and persisted even after 168 h compared with adults exposed to control netting. R. dominica was somewhat more susceptible than T. castaneum to LLINs. Finally, the dispersal capacity of both species, measured as ability to move to a remote resource patch, was significantly impaired or absent after exposure to LLINs compared to adults exposed to control netting. Our results demonstrate that LLINs are a promising new technology for reducing infestation by stored product insects since even short exposures limit movement and ultimately lead to knockdown and death.

Artificial selection can be used to create populations with extreme phenotypic responses to environmental stressors. When artificial selection is applied to a single component of a stress response, this selection may result in correlated responses in other stress responses, a phenomenon called cross-tolerance, which is ultimately controlled by the genetic correlations among traits. We selected for extreme responses to cold tolerance by selecting for chill-coma recovery time from a single temperate population of Drosophila melanogaster. Chill-coma recovery time is a common metric of low, but non-lethal, cold temperature tolerance. Replicated divergent artificial selection was applied to a genetically variable base population for 31 generations, resulting in two cold resistant, two cold susceptible, and two unselected control lines. To quantify the relationship between selection on chill-coma recovery and other metrics of thermal performance, we also measured survivorship after acute cold exposure, survivorship after chronic cold exposure, survivorship after cold exposure following a pre-treatment period (rapid cold hardening), starvation tolerance, and heat tolerance. We find that chill-coma recovery time is heritable within this population and that there is an asymmetric response to increased and decreased chill-coma recovery time. Surprisingly, we found no cross-tolerances between selection on chill-coma recovery time and the other environmental stress response traits. These results suggest that although artificial selection has dramatically altered chill-coma recovery time, the correlated response to selection on other stress response phenotypes has been negligible. The lack of a correlated response suggests that chill-coma recovery time in these selection lines is likely genetically independent from measures of cold survivorship tested here.