Recent increases in the frequency and intensity of thermal anomalies can result in recurrent stressful weather events. Among annual insects, this phenomenon may expose parents and offspring to a succession of different weather anomalies (e.g. summer heatwaves and late spring frosts). Parental stress can alter offspring physiology and performance, potentially affecting their tolerance of environmental variation. We investigated the transgenerational effects of thermal anomalies on a univoltine wetland butterfly. We induced parental thermal stress by exposing pupae to a constant temperature, ranging from stressfully cold to stressfully hot (10 °C, 26 °C, 28 °C, 32 °C), for three days. Offspring were then subjected to either winter heatwaves or spring thermal anomalies. In the parental generation, pupal exposure to heat stress (32 °C) reduced adult longevity by 46%, while exposure to 28 °C maximized female reproductive output. Parental exposure to thermal stress did not affect offspring larval mass at the onset of diapause, but it did reduce offspring survival during winter and spring. Under winter heatwaves, survival was significantly higher among offspring of parents exposed to 26 °C than those exposed to 10 °C. Parental thermal stress was associated with decreased offspring survivorship in spring under control conditions and after a late frost. We observed anticipatory parental effects, as offspring of heat-stressed parents had increased survival under winter heatwaves, and negative transgenerational effects, as offspring of both heat and cold-stressed parents exhibited lower spring survivorship. We demonstrate a mechanism by which altered weather patterns can result in butterfly declines, highlighting the challenges that warming temperatures pose for insects.
Recent increases in the frequency and intensity of thermal anomalies can result in recurrent stressful weather events. Among annual insects, this phenomenon may expose parents and offspring to a succession of different weather anomalies (e.g. summer heatwaves and late spring frosts). Parental stress can alter offspring physiology and performance, potentially affecting their tolerance of environmental variation. We investigated the transgenerational effects of thermal anomalies on a univoltine wetland butterfly. We induced parental thermal stress by exposing pupae to a constant temperature, ranging from stressfully cold to stressfully hot (10 °C, 26 °C, 28 °C, 32 °C), for three days. Offspring were then subjected to either winter heatwaves or spring thermal anomalies. In the parental generation, pupal exposure to heat stress (32 °C) reduced adult longevity by 46%, while exposure to 28 °C maximized female reproductive output. Parental exposure to thermal stress did not affect offspring larval mass at the onset of diapause, but it did reduce offspring survival during winter and spring. Under winter heatwaves, survival was significantly higher among offspring of parents exposed to 26 °C than those exposed to 10 °C. Parental thermal stress was associated with decreased offspring survivorship in spring under control conditions and after a late frost. We observed anticipatory parental effects, as offspring of heat-stressed parents had increased survival under winter heatwaves, and negative transgenerational effects, as offspring of both heat and cold-stressed parents exhibited lower spring survivorship. We demonstrate a mechanism by which altered weather patterns can result in butterfly declines, highlighting the challenges that warming temperatures pose for insects.
Molecular and genomic tools are increasingly used in biosecurity to support early detection, diagnostics and reconstruction of invasion pathways. However, selecting an appropriate marker for the question being addressed is critical, and translating molecular evidence into operational decisions can be challenging. Over-interpretation of commonly used markers may lead to misleading conclusions about population structure, invasion history and management priorities. DNA barcoding remains valuable for species identification and initial haplotype assignment, but questions concerning invasion pathways, admixture and phenotypic variation require additional molecular and ecological evidence. Here, we use coconut rhinoceros beetle (Oryctes rhinoceros) incursions in the Pacific as a case study to highlight interpretive pitfalls and lessons for applying molecular and genomic tools in biosecurity and biological control. We synthesise evidence on the strengths and limitations of mitochondrial markers, particularly CoxI, for haplotyping and inference of population relationships, and show how incomplete sampling and simplified assays can obscure cryptic diversity and bias interpretation. We then examine factors that complicate interpretation of OrNV-based biocontrol outcomes, including diagnostic sensitivity, study design and environmental modulation of host-pathogen interactions. Overall, these examples show that molecular evidence is most informative when marker choice, biological validation and phenotypic data are aligned with the biosecurity question.
In the malaria vector Anopheles gambiae s.l. Toll and Imd immune pathways signal through Re1l and Rel2 NF-κB transcription factors, respectively. Toll/Rel1 and Imd/Rel2 have been characterized mostly in the context of defense against malaria parasites, exhibiting Plasmodium species-specific roles. There is also evidence associating Rel2 with mosquito resistance to bacterial infections, however, the role of Rel1 in this context remains questionable, especially that there have been no rigorous functional genetic studies assessing Rel1 and Rel2 contributions side-by-side to mosquito defense during systemic microbial infections. Here, we conducted this analysis in An. gambiae using systemic infections with five bacterial species and the fungus Beauveria bassiana. We show that Rel2 is largely responsible for controlling the bacterial burden and for mediating mosquito tolerance to bacterial infections, whereas Rel1 contributes mainly to tolerating infections with certain bacteria, specifically those exhibiting high virulence. Rel2 is also largely responsible for regulating the expression of Cecropin 1 and Defensin 1 antimicrobial peptide genes in the fat body after bacterial systemic infections, yet in certain infections a synergistic role for Rel1 and Rel2 was also evident. In the context of fungal infections, Re1l but not Rel2 controlled the fungal burden in mosquitoes, yet resistance and tolerance to fungal infections were largely attributed to TEP1. In sum, these results advance our understanding of Imd/Rel2 and Toll/Rel1 contributions to mosquito systemic immunity and suggest that they most likely control distinct transcriptional programs in the fat body that differentially impact mosquito resistance and tolerance to systemic microbial infections.
Globally, the area of protected cropping has grown exponentially in the past 25 years. Concomitantly, the number of greenhouse crop species has increased, and a substantial proportion of these rely on, or benefit from, bee pollination. Both in crops grown outside and under cover, diversity in pollinating species could create resilience in this important service. Here we review, for the first time, the utility and use of solitary bee species for crop pollination in protected cropping environments. An increasing number of bee species, including solitary bees, are managed for pollination of crops grown outside, and our review shows that there is no shortage of solitary species able to pollinate crops grown under protective covers. Despite this, only four solitary species are used on a small commercial scale, and only in orchard crops, while pollination in greenhouses worldwide is delivered by a few, often introduced, bumble bee species. We argue that there are two impediments to the use of solitary bees for pollination in protective cropping environments: (a) Bumble bees satisfy most current greenhouse requirements, and (b) There are intrinsic difficulties in relation to husbandry and reliable large-scale supply of solitary bees, in particular when they are required year-round. We explore the possibility to overcome the second impediment by selecting species with suitable life-history traits or by deploying multiple species.Structured AbstractPurpose To review the utility and use of solitary bee species for pollination of crops grown under covers, and to investigate what could be done to enhance the use of solitary bees in such environments.Basic Procedures Using a systematic literature search, we review the solitary bee species that have been experimentally tested for their efficacy as pollinators of greenhouse and protected orchard crops. Studies included measured efficacy as increased seed/fruit set, fruit quality, pollen deposition or, where appropriate, buzz pollination activity, relative to controls. We also searched the literature for evidence of use of solitary bees in the pollination of crops under cover.Main findings The pollination efficacy of 45 solitary bee species had been tested in 98 bee-crop species combinations, with 87% of tests yielding positive outcomes. Representatives of different bee families differed in their efficacy for the pollination of different crop species. Despite their utility, only four species are used commercially in orchards, and none in greenhouses, because (a) bumble bees satisfy most current requirements; (b) there are intrinsic difficulties in relation to husbandry and reliable year-round supplies of solitary bees in sufficient numbers.New and important aspects We provide a global review and juxtapose utility and use of solitary bees. We also explore, for the first time, the difficulties posed by life-history traits on the deployment of solitary bees for the pollination of greenhouse crops and discuss whether and how these could be overcome.
Insecticide-resistant Aedes aegypti (Linnaeus) poses a growing global challenge, reducing the effectiveness of conventional chemical control and raising the need for sustainable alternatives. Targeting the larval stage remains a key strategy in mosquito management. Here, the effects of the microalga Chlorella sorokiniana (Shihira & R.W. Krauss) strain KU.B2 on larval development, pupation, adult emergence, and mortality of Ae. aegypti were investigated, together with gut ultrastructural and histological observations. Larvae were reared under five dietary regimes: (i) rabbit food suspension alone (10 mg/mL; control); (ii-iv) rabbit food suspension maintained at 10 mg/mL supplemented with 25%, 50%, or 75% (v/v) microalgal suspension; and (v) 100% microalgal suspension. Larvae exposed to microalgae showed significantly reduced growth compared with controls. While control larvae completed development within 10 days, pupation and adult emergence were delayed and reduced in the lower microalgal proportions (25% and 50%). Conversely, larvae exposed to higher microalgal proportions (75% and 100%) remained predominantly arrested at the L2 stage (day 30), with mortality rates of 89.5% and 100%, respectively. Ultrastructural and histological analyses revealed persistence of intact and partially degraded microalgal cells in the larval gut of selected groups over time suggesting that prolonged retention of microalgal materials within the gut lumen might be associated with developmental inhibition. Fractionation experiments performed using a selected microalgal supplementation proportion (62.5%) further showed that whole microalgae, disrupted cell materials, and soluble fractions were all associated with larval developmental inhibition. Overall, C. sorokiniana KU.B2 exhibits larval development-inhibitory effects against Ae. aegypti under these laboratory conditions.
Climate change impacts species interactions with cascading effects on ecosystem functions, including seed dispersal. Studies show that frugivore behaviors and associated seed dispersal are altered under climate change. However, little is known about how climate-induced behavioral shifts of non-frugivores, such as secondary seed dispersers, impact seed fate. Dung beetles are important secondary seed dispersers in many ecosystems because they bury seed-containing dung as either food caches for adult feeding or brood balls for reproduction. We utilized field manipulations with mini-greenhouses to investigate the effects of increased temperature on the seed dispersal behavior of a tunneling dung beetle species, Phanaeus vindex, in Tennessee, USA. Using beads as a proxy for seeds, we found that food caches contained more beads and were buried at shallower depths than brood balls. This suggests that food caches, which have largely been overlooked in previous studies, may play a key role in plant recruitment. Under warmer conditions in mini-greenhouses, we found that beetles buried food caches deeper than under control conditions, but food caches experienced similar temperatures regardless of treatment. For brood balls, we did not find a difference in burial depth or experienced temperatures between treatments. The greater burial depth of seeds in food caches under warmer conditions could adversely impact germination and emergence but could also buffer seeds in seed banks from the adverse effects of warmer surface temperatures. We conclude that climate change can alter secondary seed disperser behavior and thus plant-animal interactions, with ramifications for plant recruitment, community structure, and ecosystem functioning.
Rhodnius prolixus is an insect vector of the protozoan Trypanosoma cruzi , the causative agent of debilitating Chagas disease, which is transmitted to humans during blood feeding. Identifying germline markers is a critical step in advancing vector control and transgenic technologies of these medically important insects. Transmission of genetic traits to the next generation requires proper differentiation of the germline that gives rise to gametes. Germline precursors are established during early stages of development as the primordial germ cell (PGC) population. Among the genes required for this process, vasa homologues exert a conserved role in germline specification. Here, we characterize and validate the genomic structure of the R. prolixus Rp-vasa locus and assess its expression during early embryogenesis. We observe widespread Rp-vasa expression in preblastoderm embryos. Later, during the cellular blastoderm and at the beginning of gastrulation, Rp-vasa and Rp-piwi2 expression is restricted to PGCs, morphologically identifiable as a cluster of cells at the posterior of the embryo. We also report, for the first time, the use of R. prolixus regulatory sequences to drive the expression of exogenous genes. We identify the Rp-vasa regulatory region and show that these cis-regulatory sequences are sufficient to drive Cas9 and dsRed expression in the early embryo. Together, these findings demonstrate that Rp-vasa has great potential for use as a PGC marker and as a driver for gene expression in transgenic and gene editing approaches for Triatomine vectors.
Chironomus riparius midge larvae are ubiquitous in freshwater ecosystems throughout the Northern Hemisphere. Recently, freshwater in temperate regions has experienced a threat from salinization due to the use of road de-icers. As ambient temperatures fluctuate, snow and ice melt carry the de-icers into nearby freshwater and significantly raise salt levels. Recently, organic de-icers including brine beet juice de-icer (BBJD) have been implemented as an “ecofriendly” alternative to traditional road salt. Not much is known about the effects of BBJD on freshwater invertebrates. Aquatic insects respond to salinity by adjusting the ion transport functions of osmoregulatory organs. A key component in insect osmoregulation is the presence of water channel proteins known as aquaporins (AQPs) that allow movement of water along an osmotic gradient. To date, there is limited knowledge on the effects of salinity on AQP function in aquatic insects. In this study, we characterized a water-specific AQP known as CrAQP2 (a PRIP homolog) in the osmoregulatory organs of C. riparius larvae. CrAQP2 was immunolocalized in osmoregulatory organs with greatest transcript abundance in the Malpighian tubules. NaCl caused differential Craqp2 transcript expression in some of the organs, whereas BBJD had little effect on Craqp2 transcript levels. Craqp2 knockdown decreased total body water regardless of treatment and reduced survival of larvae in BBJD and NaCl. Therefore, CrAQP2 appears to be important in maintaining total body water levels stable and likely plays a role in the ability of midge larvae to respond to salinity.
In insects, juvenile hormone (JH) is essential for orchestrating reproductive events. For example, in the male moth Agrotis ipsilon, the behavioral response to female sex pheromone is linked to neuronal sensitivity in the primary olfactory centers (antennal lobes, ALs), and the maturation of accessory sex glands (ASGs) are known to be age- and JH-dependent. The molecular basis of this regulatory action of JH is not fully deciphered, and we show here that the heterodimerizing partner of Methoprene-tolerant called Taiman (Tai) is essential for the sexual maturation of male A. ipsilon. Tai expression in ALs and ASGs is elevated from the third day of adult life and is responsible for the acquisition of behavioral responsiveness to the sex pheromone and ASG maturation. Tai-deficient old males exhibited altered sexual behavior and delayed ASG maturation. Moreover, the expression levels of Tai and Krüppel homolog 1 (Kr-h1), an early JH-induced transcription factor, were reduced in ALs and ASGs of JH-deprived and Tai-deficient old males, respectively. Exogenous JH injection into young males resulted in precocious sexual maturation and this JH induction was suppressed by Tai silencing. Our results demonstrate that Tai is an actor of the JH signaling pathway that operates in ALs and ASGs to promote pheromone information processing and consequently the display of sexual behavior in synchrony with ASG maturation, ultimately optimizing male reproductive success. Thus, this study provides additional insights into the molecular mechanisms underlying hormonal regulation of sexual maturation in insects.
The house cricket, Acheta domesticus, is commonly reared for food and feed purposes and is often infected with the Acheta domesticus densovirus (AdDV). This single-stranded DNA virus can cause high mortality in crickets resulting in colony collapse. AdDV disease outbreaks in cricket mass-rearing can be prevented by either obtaining virus-free crickets or by reducing virus spread. Therefore, insight into viral levels in cricket developmental stages and into viral transmission routes is needed. Viral levels were monitored using quantitative PCR on samples collected 1) simultaneously from different life stages present in the rearing room and 2) weekly from a single rearing container during the successive developmental stages. To study viral tissue tropism and to infer the route of virus transmission, viral levels were measured in cricket tissues and in mated and non-mated adult crickets. Results showed that viral levels increased when developing from nymphs into adults and that unmated individuals had significantly lower viral levels than mated individuals. Furthermore, AdDV was present in every tested tissue and the gut and ovaries of females showed the highest viral levels. Our results suggest that AdDV is both horizontally and vertically transmitted among house crickets and provide relevant information to establish virus-free cricket lines.
Parnassius smintheus is an alpine butterfly that overwinters as a first-instar caterpillar within its egg and often beneath the snow. While extreme temperatures in early winter appear pivotal to year-to-year population change of P. smintheus in the Rocky Mountains of Canada, the sources of mortality for these eggs are unclear. Here we tested three hypotheses about the vulnerability of eggs to warming and extreme weather in early winter (i.e. upper quartile of daily maximum and minimum microclimate temperatures in November): (1) warming disrupts the acquisition of cold tolerance, making eggs susceptible to subsequent cold snaps; (2) warming drives premature development and hatch, such that precocious hatchlings either starve or freeze; and (3) warming depletes the energy stores of dormant eggs. We then used these hypotheses to guide a simulation of the risk of winter mortality for eggs over the last half-century (1971 - 2020) in the Rocky Mountains of Canada. Early winter warming did not interrupt the acquisition of cold tolerance by freeze-avoidant eggs. Eggs did risk lethal freezing in simulated winters when extreme low temperatures coincided with an absence of snow cover. Early winter warming increased the risk of subsequent hatching, and precocious larvae were less cold-tolerant than eggs. Our simulation found that precocious larvae risked freezing during snow-free cold snaps in spring. Early winter warming did not appreciably drawdown energy stores, and we found that P. smintheus could not only fuel overwintering but tolerate several days of starvation after hatch. We conclude that eggs risk precocious development after early winter warming and are likely vulnerable in winters that lack persistent snow cover. Together, these sources of winter mortality may explain year-to-year change in P. smintheus populations. Identifying unidirectional thresholds, such as hatching and freezing, may be important for predicting the susceptibility of some alpine insects to future winter warming.
Per-/polyfluoroalkyl substances (PFAS) have been commonly used over several decades for a variety of products and are very persistent in the environment. However, not much is known about their direct effects on aquatic invertebrates and their ecosystems. We examined the survival, behavior, development, and predation susceptibility of mosquito larvae Culex quinquefasciatus exposed to concentrations of perfluorooctanesulfonic acid (PFOS) ranging from 0.02 to 453.7 µg/L. PFOS exposure resulted in reduced larval survival, with a 48 hour LC50 (concentration with 50 % mortality) of 255.99 µg/L. PFOS exposure also resulted in reduced developmental success and slower maturation to adulthood (and thus slower emergence from the water) compared to control larvae. PFOS also resulted in delays in reaction to prodding stimuli, which were meant to simulate a predator attack, and longer reactions to prodding. Larvae exposed to PFOS also spent more time at the bottom of the water column, rather than at the surface where respiration takes place. Effects of larval mosquito PFOS exposure on predation by unexposed damselfly naiads (genus Ischnura) were not detected. Overall, this research suggests that PFOS impacts mosquito larvae survival, behavior, development, and adult emergence, which could have important implications for food webs or public health given the role of mosquitos as disease vectors.
Extracellular vesicles (EVs) are produced by cells of all domains of life and current research shows their involvement in intercellular transfer of information. However, a major drawback to the progress of the field relates to the fact that isolating pure EV fractions from complex biofluids is a challenging task. Isolation of EVs is often compromised by the presence of contaminating protein and lipoprotein particles, which has recently led to the establishment of guidelines for analyzing and reporting on EVs. In insects, reports on EV studies are starting to emerge, with several techniques being used without consideration of contaminant co-isolation. To address this, we optimized and validated a robust procedure for the isolation of EVs from insect hemolymph.
Tsetse flies are the primary vectors of African trypanosomes, which are transmitted through blood feeding. To supplement this nutritionally limited diet, tsetse evolved an obligate mutualism with the bacterium Wigglesworthia glossinidia, housed within a specialized organ called the bacteriome. While the functional contributions of this symbiosis towards tsetse fitness have been studied, host morphological changes that accommodate this relationship remain less understood. In pregnant flies, variable expression of microRNAs (miRNAs) regulates protein expression, but the specific impacts are unknown. During pregnancy, high expression of fatty acyl-CoA reductase (far) within the bacteriome is indirectly correlated with miR-31 abundance and coincides with bacteriome size increase. We explored the roles of far and miR-31 towards this morphological change. Although RNAi effectively reduced far expression, bacteriome size still increased, suggesting its expansion is independent of far. In contrast, disrupting miR-31 activity resulted in significantly enlarged bacteriomes in virgin flies, resembling those of mated females. These results suggest that gene(s) other than far are regulated by miR-31 and may contribute to bacteriome remodeling during pregnancy, potentially to meet increased symbiosis demands. Ultimately, disrupting this obligate mutualism may present a promising target for future vector control strategies.
The Mediterranean fruit fly (medfly) is a serious frugivorous Afro-tropical pest common in the Mediterranean Region in Europe, which, despite the requirements of Directive 2009/128/EC and the availability of biological methods, is still widely controlled using pesticides. Our objective was to reassess the suitability of the classical Integrated Pest Management (IPM) paradigm for medfly control and the feasibility of its mission to promote biological methods as an alternative to pesticides. For this purpose, different IPM scenarios were simulated for three hypothetical farms located along the 1000 km latitudinal transect in Italy using the PESTonFARM model. The simulations revealed a fundamental flaw in the classical IPM paradigm. By disregarding the ability of medfly to inconspicuously increase its population before the economic (action) threshold is reached, it alerts the farmer too late. By then, the medfly population enters the phase of exponential growth, when gradually acting biological methods are unable to cope. The guidelines of classical IPM reinforce the tendency of farmers to focus on large medfly populations during summer, strengthening the reliance on pesticides as the most effective option at the time. We hereby propose to move away from principles of monitoring-based and threshold-initiated control and target the sparse population of overwintering medflies before they can be detected by monitoring. The control starts on earliest fruits, even if not usually infested to a noticeable extent, and gradually continues to protect subsequent fruits. Our results confirmed that the proposed approach is highly effective, allows for the reliable use of non-pesticide methods of fruit fly control, and therefore ensures that the key ambitions of IPM can be achieved.
In response to high temperatures, insect metabolic rates increase, favoring the release of higher amounts of reactive oxygen species (ROS). These ROS need to be counteracted by antioxidants to avoid oxidative stress, which can lead to cell damage and death. In this manuscript, we review evidence in insects showing the effects of high temperatures on ROS production, the antioxidant defenses reported in insects in response to high and extremely high temperatures and the extent to which they contribute to preventing oxidative damage. Endogenously produced antioxidants can be enzymatic or non-enzymatic and are involved in heat responses in at least seven insect orders. Our review indicates that evidence is very limited for the effect of high temperature on ROS production, but it clearly shows that at least one antioxidant is upregulated during short-term heat exposure. However, the effects of antioxidants in effectively reducing oxidative damage in biomolecules are still poorly supported by evidence. Dietary-dependent antioxidants show strong potential for coping with heat stress, but evidence is limited, although numerous plants produce antioxidant compounds and a great number of insect species feed on plants. The role of antioxidants in heat acclimation and adaptation is promising but evidence is still very limited in insects. Antioxidants also protect from other prooxidant conditions such as pesticide exposure, nutrient stress, or new biotic interactions, which often act in combination. Potential trade-offs between antioxidant use to different functions could define insect survival and pace of life in response to multiple stressors, including high temperatures. Our literature review indicates that there is only limited evidence of the role of antioxidants in preventing oxidative damage caused by heat, opening the possibility that ROS production might be mitigated by the action of uncoupling proteins or degradation of mitochondria. Finally, we conclude by proposing promising research avenues to gain a deeper understanding of the role of ROS and antioxidants in the oxidative balance of insects exposed to mild and extreme heat.
Adaptive evolution requires both natural selection and genetic variation. In introduced species, the selective dynamics of range expansion are predicted by theory to lead to differences between the core and the leading edge, with edge individuals evolving to be more fecund (under r-selection) and have greater dispersal ability than core individuals. In arthropods, both fecundity and dispersal ability are often positively correlated with body size. Here, we quantify genetic variation available for evolution of body size in a beetle (Diorhabda carinulata) introduced into North America as a biological control agent. Previously, we found that females at the edge of the range expansion have evolved to be larger than those at the core as predicted by theory, while male body size has not clearly changed, despite the evolution of increased dispersal capacity. Using a half-sib mating design, we measure genetic variation in mass at eclosion and thorax width of female and male beetles from a single introduced population at the core of the range expansion. We find significant heritable genetic variation in females in both traits, but not in males. Thus, lack of genetic variation in body size may preclude evolution of size in males along this expansion front.
As ectotherms, many insects spend the winter months in a state of suspended animation (i.e., diapause), lowering their metabolic rates to subsist on a limited store of energy reserves. The ability to lower metabolic rate during diapause relies, in part, on cold winter temperatures to intrinsically lower metabolic rate. Winter warming associated with global climate change may pose a challenge to diapausing insects by intrinsically increasing metabolic rate, potentially leading to the exhaustion of energetic reserves. We used stop-flow respirometry to measure oxygen consumption in response to temperatures representative of both acute and chronic winter warming scenarios in diapausing Pieris rapae pupae. Metabolic rate increased with increasing temperature in diapausing pupae, but metabolic rate depended on both pupal age and warming severity, with older pupae having lower metabolic rates overall. Despite the increases in metabolic rate, pupae recovered metabolic rate within 24-hours after short-term acute-warming exposure. In contrast, chronic exposure to warming over weeks and months led to significant decreases in metabolic rate later in diapause, as well as reductions in pupal mass. These results demonstrate that while respiration was thermally responsive, warming did not lead to sustained increases in metabolic rate. Instead, diapausing P. rapae appear to acclimate to higher temperature by lowering their metabolic rates in response to months of chronic warming. Overall, these patterns suggest that this species could be resilient to winter warming, at least in the context of energetics. However, the precise mechanisms underlying these responses remain to be characterized. Thus, future research-e.g., on the genetic underpinnings of energetics in the context of warming-could further elucidate the relative vulnerability of diapausing insects to future winter warming.
