Search

The spread of blood-borne pathogens by mosquitoes relies on their taking a blood meal; if there is no bite, there is no disease transmission. Although many species of mosquitoes never take a blood meal, identifying genes that distinguish blood feeding from obligate nonbiting is hampered by the fact that these different lifestyles occur in separate, genetically incompatible species. There is, however, one unique extant species with populations that share a common genetic background but blood feed in one region and are obligate nonbiters in the rest of their range: Wyeomyia smithii . Contemporary blood-feeding and obligate nonbiting populations represent end points of divergence between fully interfertile southern and northern populations. This divergence has undoubtedly resulted in genetic changes that are unrelated to blood feeding, and the challenge is to winnow out the unrelated genetic factors to identify those related specifically to the evolutionary transition from blood feeding to obligate nonbiting. Herein, we determine differential gene expression resulting from directional selection on blood feeding within a polymorphic population to isolate genetic differences between blood feeding and obligate nonbiting. We show that the evolution of nonbiting has resulted in a greatly reduced metabolic investment compared with biting populations, a greater reliance on opportunistic metabolic pathways, and greater reliance on visual rather than olfactory sensory input. W. smithii provides a unique starting point to determine if there are universal nonbiting genes in mosquitoes that could be manipulated as a means to control vector-borne disease.

Seasonal change in the temperate and polar regions of Earth determines how the world looks around us and, in fact, how we live our day-to-day lives. For biological organisms, seasonal change typically involves complex physiological and metabolic reorganization, the majority of which is regulated by photoperiodism. Photoperiodism is the ability of animals and plants to use day length or night length, resulting in life-historical transformations, including seasonal development, migration, reproduction, and dormancy. Seasonal timing determines not only survival and reproductive success but also the structure and organization of complex communities and, ultimately, the biomes of Earth. Herein, a small mosquito, Wyeomyia smithii, that lives only in the water-filled leaves of a carnivorous plant over a wide geographic range, is used to explore the genetic and evolutionary basis of photoperiodism. Photoperiodism in W. smithii is considered in the context of its historical biogeography in nature to examine the startling finding that recent rapid climate change can drive genetic change in plants and animals at break-neck speed, and to challenge the ponderous 80+ year search for connections between daily and seasonal time-keeping mechanisms. Finally, a model is proposed that reconciles the seemingly disparate 24-h daily clock driven by the invariant rotation of Earth about its axis with the evolutionarily flexible seasonal timer orchestrated by variable seasonality driven by the rotation of Earth about the Sun.

Photoperiodic time measurement regulating larval diapause in the pitcher-plant mosquito, Wyeomyia smithii, varies in a close relationship with latitude. The critical photoperiod mediating the maintenance and termination of diapause is positively correlated with latitude (r2 = 0.977) among six populations from southern (30–31° N), intermediate (40° N), and northern (46–49° N) latitudes in North America. The developmental response to unnaturally short and to unnaturally long photoperiods declines with increasing latitude, so that longer critical photoperiods are associated with a downward rather than a lateral shift in the photoperiodic response curve. Exotic light and dark cycles of varying period (T) with a short (10 h) photophase and a scotophase ranging from 14 (T = 24) to 62 (T = 72) h, reveal two geographic patterns: a decline in perturbability of the photoperiodic clock with increasing latitude, and no change with latitude in the 21-h period of rising and falling development with increasing T. These results show (1) that there is a rhythmic component to photoperiodic time measurement in W. smithii, (2) that the period of this rhythm is about 21 h in all populations, and (3) that more northern populations show decreasing responsiveness to photoperiod and increasing stability against perturbation by exotic period lengths (T > 24). Previous studies on W.␣smithii indicate that this single temperate species of a tropical and subtropical genus has evolved from south to north. We therefore conclude that the evolution of increasing critical photoperiod in W. smithii during its adaptive radiation into North America has more likely involved the amplitude and not the period of the underlying circadian pacemaker.

The mosquito, Wyeomyia smithii, enters a larval dormancy or diapause that is initiated, maintained, and terminated by photoperiod. The median or critical photoperiod regulating diapause increases from 12 h of light per day along the Gulf of Mexico, USA (30° N), to over 15 h in southern Canada (49° N). Photoperiodic time measurement in W. smithii comprises both rhythmic and hourglass (interval timer) components. Using interrupted-night and resonance experiments, we show that both the rhythmic and hourglass components are prominent in the southern (ancestral) populations and that the influence of the rhythmic component declines with increasing latitude, while the hourglass component remains strong in northern (derived) populations. Previously, it has been shown that the genetic differences in critical photoperiod between northern populations and their southern ancestors involve not only the additive (independent) effects of genes, but also gene-gene interaction (epistasis). We therefore conclude that adaptive evolution of W. smithii has probably involved the progressive epistatic masking of the ancestral rhythmic component resulting in photoperiodic time measurement in northern populations accomplished principally through a day-interval timer. A comparison of W. smithii with previous studies indicates that the decline in critical photoperiod with increasing latitude represents an overall decrease in response to light rather than a shift in the timing of photosensitivity among arthropods in general. We propose that the underlying functional components of photoperiodic time measurement, as well as the overt photoperiodic response, are either homologous or are themselves responding directly to selection over latitudinal gradients in seasonality.

Controlled laboratory experiments were used to show that Oregon and Alaskan three-spined stickleback Gasterosteus aculeatus, collected from locations differing by 18° of latitude, exhibited no significant variation in length of the polyglutamine domain of the clock protein or in photoperiodic response within or between latitudes despite the fact that male and female G. aculeatus are photoperiodic at both latitudes. Hence, caution is urged when interpreting variation in the polyglutamine repeat (PolyQ) domain of the gene clock in the context of seasonal activities or in relationship to photoperiodism along geographical gradients.

Seasonal change in the temperate and polar regions of Earth determines how the world looks around us and, in fact, how we live our day-to-day lives. For biological organisms, seasonal change typically involves complex physiological and metabolic reorganization, the majority of which is regulated by photoperiodism. Photoperiodism is the ability of animals and plants to use day length or night length, resulting in life-historical transformations, including seasonal development, migration, reproduction, and dormancy. Seasonal timing determines not only survival and reproductive success but also the structure and organization of complex communities and, ultimately, the biomes of Earth. Herein, a small mosquito, Wyeomyia smithii, that lives only in the water-filled leaves of a carnivorous plant over a wide geographic range, is used to explore the genetic and evolutionary basis of photoperiodism. Photoperiodism in W. smithii is considered in the context of its historical biogeography in nature to examine the startling finding that recent rapid climate change can drive genetic change in plants and animals at break-neck speed, and to challenge the ponderous 80+ year search for connections between daily and seasonal time-keeping mechanisms. Finally, a model is proposed that reconciles the seemingly disparate 24-h daily clock driven by the invariant rotation of Earth about its axis with the evolutionarily flexible seasonal timer orchestrated by variable seasonality driven by the rotation of Earth about the Sun.

International audience; Parasites and pathogens are increasingly recognized as significant drivers of ecological and evolutionary change in natural ecosystems. Concurrently, transmission of infectious agents among human, livestock, and wildlife populations represents a growing threat to veterinary and human health. In light of these trends and the scarcity of long‐term time series data on infection rates among vectors and reservoirs, the National Ecological Observatory Network (NEON) will collect measurements and samples of a suite of tick‐, mosquito‐, and rodent‐borne parasites through a continental‐scale surveillance program. Here, we describe the sampling designs for these efforts, highlighting sampling priorities, field and analytical methods, and the data as well as archived samples to be made available to the research community. Insights generated by this sampling will advance current understanding of and ability to predict changes in infection and disease dynamics in novel, interdisciplinary, and collaborative ways.

The National Ecological Observatory Network (NEON) intends to monitor mosquito populations across its broad geographical range of sites because of their prevalence in food webs, sensitivity to abiotic factors, and relevance for human health. We describe the design of mosquito population sampling in the context of NEON's long‐term continental scale monitoring program, emphasizing the sampling design schedule, priorities, and collection methods. Freely available NEON data and associated field and laboratory samples, will increase our understanding of how mosquito abundance, demography, diversity, and phenology are responding to land use and climate change.

Photoperiod, or length of day, has a predictable annual cycle, making it an important cue for the timing of seasonal behavior and development in many organisms. Photoperiod is widely used among temperate and polar animals to regulate the timing of sexual maturation. The proper sensing and interpretation of photoperiod can be tightly tied to an organism’s overall fitness. In photoperiodic mammals and birds the thyroid hormone pathway initiates sexual maturation, but the degree to which this pathway is conserved across other vertebrates is not well known. We use the threespine stickleback Gasterosteus aculeatus, as a representative teleost to quantify the photoperiodic response of key genes in the thyroid hormone pathway under controlled laboratory conditions. We find that the photoperiodic responses of the hormones are largely consistent amongst multiple populations, although differences suggest physiological adaptation to various climates. We conclude that the thyroid hormone pathway initiates sexual maturation in response to photoperiod in G. aculeatus, and our results show that more components of this pathway are conserved among mammals, birds, and teleost fish than was previously known. However, additional endocrinology, cell biology and molecular research will be required to define precisely which aspects of the pathway are conserved across vertebrates.

Many traits are assumed to have a causal (necessary) relationship with one another because of their common covariation with a physiological, ecological or geographical factor. Herein, we demonstrate a straightforward test for inferring causality using residuals from regression of the traits with the common factor. We illustrate this test using the covariation with latitude of a proxy for the circadian clock and a proxy for the photoperiodic timer in Drosophila and salmon. A negative result of this test means that further discussion of the adaptive significance of a causal connection between the covarying traits is unwarranted. A positive result of this test provides a point of departure that can then be used as a platform from which to determine experimentally the underlying functional connections and only then to discuss their adaptive significance.

Examination of temperate and polar regions of Earth shows that the nonbiological world is exquisitely sensitive to the direct effects of temperature, whereas the biological world is largely organized by light. Herein, we discuss the use of day length by animals at physiological and genetic levels, beginning with a comparative experimental study that shows the preeminent role of light in determining fitness in seasonal environments. Typically, at seasonally appropriate times, light initiates a cascade of physiological events mediating the input and interpretation of day length to the output of specific hormones that ultimately determine whether animals prepare to develop, reproduce, hibernate, enter dormancy, or migrate. The mechanisms that form the basis of seasonal time keeping and their adjustment during climate change are reviewed at the physiological and genetic levels. Future avenues for research are proposed that span basic questions from how animals transition from dependency on tropical cues to temperate cues during range expansions, to more applied questions of species survival and conservation biology during periods of climatic stress.

Reproductive maturation in both male and female three-spined stickleback Gasterosteus aculeatus was strongly photoperiodic in a northern population (Alaska, 61 degrees N) but not in a southern population (Oregon, 43 degrees N) from western North America. Increasing reliance on photoperiod with increasing latitude is a general phenomenon among vertebrates, and is probably due to the anticipation of a narrower window of opportunity for reproduction and development at higher latitudes.

The ubiquity of endogenous, circadian (daily) clocks among eukaryotes has long been held as evidence that they serve an adaptive function, usually cited as the ability to properly time biological events in concordance with the daily cycling of the environment. Herein we test directly whether fitness is a function of the matching of the period of an organism's circadian clock with that of its environment. We find that fitness, measured as the per capita expectation of future offspring, a composite measure of fitness incorporating both survivorship and reproduction, is maximized in environments that are integral multiples of the period of the organism's circadian clock. Hence, we show that organisms require temporal concordance between their internal circadian clocks and their external environment to maximize fitness and thus the long-held assumption is true that, having evolved in a 24-h world, circadian clocks are adaptive.

The primary nonbiological result of recent rapid climate change is warming winter temperatures, particularly at northern latitudes, leading to longer growing seasons and new seasonal exigencies and opportunities. Biological responses reflect selection due to the earlier arrival of spring, the later arrival of fall, or the increasing length of the growing season. Animals from rotifers to rodents use the high reliability of day length to time the seasonal transitions in their life histories that are crucial to fitness in temperate and polar environments: when to begin developing in the spring, when to reproduce, when to enter dormancy or when to migrate, thereby exploiting favourable temperatures and avoiding unfavourable temperatures. In documented cases of evolutionary (genetic) response to recent, rapid climate change, the role of day length (photoperiodism) ranges from causal to inhibitory; in no case has there been demonstrated a genetic shift in thermal optima or thermal tolerance. More effort should be made to explore the role of photoperiodism in genetic responses to climate change and to rule out the role of photoperiod in the timing of seasonal life histories before thermal adaptation is assumed to be the major evolutionary response to climate change.

Photoperiodism is the ability of organisms to assess and use the day length as an anticipatory cue to time seasonal events in their life histories. Photoperiodism is especially important in initiating physiological and developmental processes that are typically irrevocable and that culminate at a future time or at a distant place; the further away in space or time, the more likely a seasonal event is initiated by photoperiod. The pervasiveness of photoperiodism across broad taxa, from rotifers to rodents, and the predictable changes of photoperiodic response with geography identify it as a central component of fitness in temperate and polar seasonal environments. Consequently, the role of day length cannot be disregarded when evaluating the mechanisms underlying life-historical events, range expansions, invasions of novel species, and response to climate change among animals in the temperate and polar regions of the world.

A wide variety of temperate animals rely on length of day (photoperiodism) to anticipate and prepare for changing seasons by regulating the timing of development, reproduction, dormancy, and migration. Although the molecular basis of circadian rhythms regulating daily activities is well defined, the molecular basis for the photoperiodic regulation of seasonal activities is largely unknown. We use geographic variation in the photoperiodic control of diapause in the pitcher-plant mosquito Wyeomyia smithii to create the first QTL map of photoperiodism in any animal. For critical photoperiod (CPP), we detect QTL that are unique, a QTL that is sex linked, QTL that overlap with QTL for stage of diapause (SOD), and a QTL that interacts epistatically with the circadian rhythm gene, timeless. Results presented here confirm earlier studies concluding that CPP is under directional selection over the climatic gradient of North America and that the evolution of CPP is genetically correlated with SOD. Despite epistasis between timeless and a QTL for CPP, timeless is not located within any detectable QTL, indicating that it plays an ancillary role in the evolution of photoperiodism in W. smithii. Finally, we highlight one region of the genome that includes loci contributing to CPP, SOD, and hormonal regulation of development.

Background Internal circadian (circa, about; dies, day) clocks enable organisms to maintain adaptive timing of their daily behavioral activities and physiological functions. Eukaryotic clocks consist of core transcription-translation feedback loops that generate a cycle and post-translational modifiers that maintain that cycle at about 24 h. We use the pitcher-plant mosquito, Wyeomyia smithii (subfamily Culicini, tribe Sabethini), to test whether evolutionary divergence of the circadian clock genes in this species, relative to other insects, has involved primarily genes in the core feedback loops or the post-translational modifiers. Heretofore, there is no reference transcriptome or genome sequence for any mosquito in the tribe Sabethini, which includes over 375 mainly circumtropical species. Methods We sequenced, assembled and annotated the transcriptome of W. smithii containing nearly 95 % of conserved single-copy orthologs in animal genomes. We used the translated contigs and singletons to determine the average rates of circadian clock-gene divergence in W. smithii relative to three other mosquito genera, to Drosophila, to the butterfly, Danaus, and to the wasp, Nasonia. Results Over 1.08 million cDNA sequence reads were obtained consisting of 432.5 million nucleotides. Their assembly produced 25,904 contigs and 54,418 singletons of which 62 % and 28 % are annotated as protein-coding genes, respectively, sharing homology with other animal proteomes. Discussion The W. smithii transcriptome includes all nine circadian transcription-translation feedback-loop genes and all eight post-translational modifier genes we sought to identify (Fig. 1). After aligning translated W. smithii contigs and singletons from this transcriptome with other insects, we determined that there was no significant difference in the average divergence of W. smithii from the six other taxa between the core feedback-loop genes and post-translational modifiers. Conclusions The characterized transcriptome is sufficiently complete and of sufficient quality to have uncovered all of the insect circadian clock genes we sought to identify (Fig. 1). Relative divergence does not differ between core feedback-loop genes and post-translational modifiers of those genes in a Sabethine species (W. smithii) that has experienced a continual northward dispersal into temperate regions of progressively longer summer day lengths as compared with six other insect taxa. An associated microarray platform derived from this work will enable the investigation of functional genomics of circadian rhythmicity, photoperiodic time measurement, and diapause along a photic and seasonal geographic gradient. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-1937-y) contains supplementary material, which is available to authorized users.

The two major rhythms of the biosphere are daily and seasonal; the two major adaptations to these rhythms are the circadian clock, mediating daily activities, and the photoperiodic timer, mediating seasonal activities. The mechanistic connection between the circadian clock and the photoperiodic timer remains unresolved. Herein, we show that the rhythmic developmental response to exotic light:dark cycles, usually used to infer a causal connection between the circadian clock and the photoperiodic timer, has evolved independently of the photoperiodic timer in the pitcher‐plant mosquito Wyeomyia smithii across the climatic gradient of eastern North America from Florida to Canada and from the coastal plain to the mountains. We conclude that the photoperiodic timing of seasonal events can evolve independently of the daily circadian clock.

Environmental temperature can modify not only rates of ectotherm growth, development, and reproduction, but also, at the extremes, temperature can limit survivorship and ultimately fitness. We studied ectotherm populations from a latitudinal gradient to understand how alterations in the thermal environment (e.g., rapid climate change) may affect the persistence of populations experiencing stress-induced mortality. Populations of the mosquito, Wyeomyia smithii, from 30-50' N in North America were subjected to heat and cold stress based on observed field temperatures during the simulated passage of a typical 4-5 d warm- or cold-weather front. Cold but not heat tolerance corresponded to latitude of origin, reflecting previously observed patterns in year-long fitness. Both heat and cold stress resulted in an average of 44% mortality relative to unstressed controls but did not result in a significant loss of fitness (Ro) at the population level. We conclude that individuals most likely to survive during periods of stress are also the individuals most likely to make the greatest contribution to the next generation in the absence of stress. Since individual survivorship must, at some level, become limiting to population viability, these results imply that apparently viable populations may be pushed rapidly to extinction by a small increment in environmental stress.

1. The effects of resource levels, thermal microclimate, and seasonal oviposition patterns on fecundity and survivorship in the pitcher-plant mosquito, Wyeomyia smithii (Coq.), were examined at a northern Wisconsin bog over the course of 2 years. Wyeomyia smithii are bivoltine at this locality, thereby enabling the study of summer and overwintering generations separately. 2. Nutrient resources of W. smithii were not limiting and there was no indication of density-dependent survivorship or fecundity. 3. Oviposition rates were highest in young, large pitchers and individual mosquitoes appeared to allocate only a few eggs to any one leaf. 4. Winter was the harsh season, and the principal manifestation of seasonal harshness was reduced survivorship. 5. Overwintering W. smithii that had been oviposited later in the summer had a higher odds of survival than those oviposited earlier in the summer. 6. It was concluded that dispersal of eggs among many pitchers serves to spread the risk of encountering lethal winter temperatures among spatially unpredictable patches.

Protandry is the emergence or arrival into a seasonal population of males before females and is widespread among both plants and animals. Conceptually, protandry should be subject to stabilizing selection because early emerging males risk death before mating and late-emerging males miss opportunities to mate. However, for any given male, the optimal emergence time may depend upon the mean and distribution of emergence of other males in the population. Using lifetime offspring sired as our criterion of reproductive success, we found that, in laboratory populations of the pitcher-plant mosquito, Wyeomyia smithii, there is, indeed, stabilizing selection acting on the optimal time for male emergence, and that the specific optimum does depend upon emergence time of other males in the population. In addition, male cohorts achieved greater reproductive success when their emergence was spread over several days rather than occurring in a single day. However, reproductive success did not differ between normal and superdispersed emergence distri- butions, suggesting that, under more variable, natural conditions, the specific pattern of male emergence may be far less important than variation in emergence time, per se.

Seasonal change in daylength (photoperiod) is widely used by insects to regulate temporal patterns of development and behaviour, including the timing of diapause (dormancy) and migration. Flexibility of the photoperiodic response is critical for rapid shifts to new hosts, survival in the face of global climate change and to reproductive isolation. At the same time, the daily circadian clock is also essential for development, diapause and multiple behaviours, including correct flight orientation during long-distance migration. Although studied for decades, how these two critical biological timing mechanisms are integrated is poorly understood, in part because the core circadian clock genes are all transcription factors or regulators that are able to exert multiple effects throughout the genome. In this chapter, we discuss clocks in the wild from the perspective of diverse insect groups across eco-geographic contexts from the Antarctic to the tropical regions of Earth. Application of the expanding tool box of molecular techniques will lead us to distinguish universal from unique mechanisms underlying the evolution of circadian and photoperiodic timing, and their interaction across taxonomic and ecological contexts represented by insects. This article is part of the themed issue ‘Wild clocks: integrating chronobiology and ecology to understand timekeeping in free-living animals’.

A wide variety of higher plants, vertebrates, and arthropods use the length of day to synchronize growth, development, reproduction, dormancy, and migration with the changing seasons. In the pitcher-plant mosquito, Wyeomyia smithii, critical photoperiod mediating the onset and maintenance of larval diapause has evolved about 10 standard deviations in mean critical photoperiod between the ancestral, Gulf Coast populations, and the derived, Canadian populations. We are seeking to understand how this evolution has been accomplished at both the genetic and the physiological levels. At the genetic level, average heterozygosity at protein-coding loci decreases with latitude of origin, while genetic variation for photoperiodic response increases with latitude of origin, particularly within the formerly glaciated regions of North America. Hybridization experiments reveal widespread genetic differences in critical photoperiod due to epistasis. We ascribe the increase in genetic variation in photoperiodic response, despite directional and stabilizing selection to the contrary, to the release of additive from epistatic variance during successive founder events in W. smithii’s northward dispersal following recession of the Laurentide Ice Sheet, and to the resulting genetic drift and reorganization of genetic architectures in descendent populations. At the physiological level, northern populations of W. smithii, as well as northern populations of spider mites, flies, moths, and beetles in both North America and Europe show a declining expression of the rhythmic component of photoperiodic response. In Drosophila melanogaster, when the epistatic coupling between the period locus and photoperiodic response is disrupted, the critical photoperiod is shifted towards shorter daylengths; analogously in W. smithii, when epistatic interactions are disrupted in the recombining generations of hybrid populations, the critical photoperiod is shifted towards shorter daylengths. The implications here are (1) that in W. smithii, it is the epistatic modification of the photoperiodic response curve by the circadian clock that is being disrupted in the recombining generations and (2) that post-glacial range expansion into the North-Temperate Zone by arthropods in general may involve uncoupling of the circadian and photoperiodic clocks. Despite the tremendous advances that have been made in understanding circadian rhythmicity at the molecular level, virtually nothing is known about how or whether any of the downstream ‘clock-controlled genes’ connect with photoperiodic time measurement. Except in W. smithii, little is known about how this connection changes with seasonal adaptation of photoperiodic response through evolutionary time. It is our goal and desire that our top–down approach to the evolution of photoperiodic time measurement will meet and mesh with the bottom–up approach that is being developed so fruitfully for circadian clocks.  2001 Elsevier Science Ltd. All rights reserved.

We are concerned with the climatic factors that affect the adaptive radiation of species into and within the temperate zone. As one progresses northward, there is a decrease in summer temperatures and a concomitant increase in the duration and intensity of winter cold. For the pitcher-plant mosquito, Wyeomyia smithii, it has been shown previously that (1) range expansion and evolution has proceeded from south to north, and (2) either realistically hot summer temperatures or a simulated 7-mo winter reduce fitness by 60% or more in two New Jersey (40° N) populations. Herein, we compare the relative fitness of nine populations of W. smithii from 30° to 50° N when exposed to these same stressful summer temperatures and simulated winter. We determined the survivorship, fecundity, and fertility of five cohorts from each population at each temperature in both environments, and we calculated the cohort replacement rate (R0) as the product of these three component traits. Survivorship declined with increasing lati...

We performed truncation selection for increased fitness (rc) under conditions of chronic stress from the combined effects of low nutrients and high temperature, representative of extremes likely to be encountered in nature by the pitcher-plant mosquito, Wyeomyia smithii. We performed selection on geographical parental populations and their hybrids to determine whether hybridization would facilitate or constrain adaptation under our selection protocol. The stressful environment decreased fitness (rc) by 54% averaged across all populations relative to near-optimal conditions. After approximately 10 generations of selection under chronically stressful conditions, exactly one-half of the parental and one-half of the hybrid populations had gone extinct. Thus hybridization had no effect on the likelihood of population persistence. Fitness (rc) of the surviving populations did not show any response to selection. Despite initial hybrid vigour under stressful conditions, the fitness (rc) of surviving hybrid populations was either equal to, or worse than, the fitness (rc) of surviving parental populations after approximately 10 generations of selection. These results suggest that outcrossing populations to augment genetic variation and facilitate adaptation to a rapidly changing environment may not be useful over longer time scales, even in cases where hybridization does initially increase fitness. Although we detected no direct response to selection for increased fitness (rc) under lifetime chronic stress, selected populations showed a strong correlated response for survivorship through transient, acute heat and desiccation shock. In evaluating how organisms might respond to future climate change, biologists must maintain a clear distinction between lifelong performance in chronically stressful environments and short-term survivorship through transient, acute stress.

The mosquito, Wyeomyia smithii, develops only within the water-filled leaves of the purple pitcher plant, Sarracenia purpurea, where it overwinters in larval diapause. We reared uniform-aged cohorts in the leaves of intact plants in three environments: near-optimal summer conditions, intentionally stressful summer conditions, and a 23-wk simulated winter. Overwintering W. smithii suffered a 60% loss in fitness (R0), comparable to the 68% loss in fitness imposed by the summer stress. Overwintering cohorts lost fitness through reduced survivorship, fecundity, fertility, adult longevity, and mass-specific fecundity. We argue that very real trade-offs occur between summer performance and winter performance among temperate insects in general. Survivorship and reproduction of the overwintering generation should therefore be an important source of selection shaping not only the seasonal expression of dormancy, but also the evolution of summer life-history syndromes as well.

We determine the contribution of composite additive, dominance, and epistatic effects to the genetic divergence of photoperiodic response along latitudinal, altitudinal, and longitudinal gradients in the pitcher-plant mosquito, Wyeomyia smithii. Joint scaling tests of crosses between populations showed wide-spread epistasis as well as additive and dominance differences among populations. There were differences due to epistasis between an alpine population in North Carolina and populations in Florida, lowland North Carolina, and Maine. Longitudinal displacement resulted in differences due to epistasis between Florida and Alabama populations separated by 300 km but not between Maine and Wisconsin populations separated by 2000 km. Genetic differences between New Jersey and Ontario did not involve either dominance or epistasis and we estimated the minimum number of effective factors contributing to a difference in mean critical photoperiod of 5 SD between them as nE = 5. We propose that the genetic similarity of populations within a broad northern region is due to their more recent origin since recession of the Laurentide Ice Sheet and that the unique genetic architecture of each population is the result of both mutation and repeated migration-founder-flush episodes during the dispersal of W. smithii in North America. Our results suggest that differences in composite additive and dominance effects arise early in the genetic divergence of populations while differences due to epistasis accumulate after more prolonged isolation.

Models of protandry (1) assume implicitly or explicitly that independent evolution of male and female preadult development times is possible and (2) assume or assert that protandry should increase in populations that are univoltine or at least composed of discrete, nonoverlapping generations. Herein we examine these assumptions in the pitcher-plant mosquito, Wyeomyia smithii. Heritability and additive genetic variance of development time are higher in female than in male W. smithii. Protandry is higher in lines selected for slow development than in lines selected for fast development or in unselected control lines. Protandry is therefore capable of evolving. Contrary to predictions based on sexual selection, southern populations with multiple, overlapping generations are just as protandrous as northern bi- and univoltine populations with discrete generations. Voltinism and developmental synchrony of the population do not appear to have been major selective factors in the evolution of protandry in W. smithii. We propose that protandry can be maintained by natural selection in multivoltine populations with overlapping generations as a consequence of sexually dimorphic fitness criteria. Selection should minimize development time in males but maximize growth rate in females. In W. smithii, females achieve higher growth rate than males but also harbor greater genetic variation for development time, indicating that selection has, indeed, minimized development time to a greater extent in males than in females. We conclude that if both natural and sexual selection are involved in the maintenance of protandry in populations of W. smithii, then their relative importance changes with the degree of generation overlap.

Herein we tested the repeatability of phylogenetic inference based on high throughput sequencing by increased taxon sampling using our previously published techniques in the pitcher-plant mosquito, Wyeomyia smithii in North America. We sampled 25 natural populations drawn from different localities nearby 21 previous collection localities and used these new data to construct a second, independent phylogeny, expressly to test the reproducibility of phylogenetic patterns. Comparison of trees between the two data sets based on both maximum parsimony and maximum likelihood with Bayesian posterior probabilities showed close correspondence in the grouping of the most southern populations into clear clades. However, discrepancies emerged, particularly in the middle of W. smithii's current range near the previous maximum extent of the Laurentide Ice Sheet, especially concerning the most recent common ancestor to mountain and northern populations. Combining all 46 populations from both studies into a single maximum parsimony tree and taking into account the post-glacial historical biogeography of associated flora provided an improved picture of W. smithii's range expansion in North America. In a more general sense, we propose that extensive taxon sampling, especially in areas of known geological disruption is key to a comprehensive approach to phylogenetics that leads to biologically meaningful phylogenetic inference.

Protandry, the early emergence of males into a seasonally breeding pop- ulation, has been described mathematically as a mating strategy resulting from sexual selection on both sexes. A key implicit assumption in protandry models is that all matings contribute equally to a male's reproductive success. We build a simulation model of pro- tandry based on field censuses and investigate the consequences of size-specific temporal variation in female fecundity for the optimal timing of protandry in the western treehole mosquito, Aedes sierrensis. We show that theoretical predictions of protandry are robust when differential female fecundity is incorporated into a model of protandry in A. sierrensis. In addition, we utilize field data and laboratory experiments to elucidate the selective forces acting on both sexes of this mosquito. Under conditions of reduced per capita resources, males minimized development time by pupating at lower mass; females maximized mass by delaying pupation. These gender-specific, homeostatic adjustments to food and density, which result in density-dependent protandry, indicate that protandry is the result of selection on independent fitness criteria in each sex.

Identifying regions of the genome contributing to phenotypic evolution often involves genetic mapping of quantitative traits. The focus then turns to identifying regions of ‘major’ effect, overlooking the observation that traits of ecological or evolutionary relevance usually involve many genes whose individual effects are small but whose cumulative effect is large. Herein, we use the power of fully interfertile natural populations of a single species of mosquito to develop three quantitative trait loci (QTL) maps: one between two post-glacially diverged populations and two between a more ancient and a post-glacial population. All demonstrate that photoperiodic response is genetically a highly complex trait. Furthermore, we show that marker regressions identify apparently ‘non-significant’ regions of the genome not identified by composite interval mapping, that the perception of the genetic basis of adaptive evolution is crucially dependent upon genetic background and that the genetic basis for adaptive evolution of photoperiodic response is highly variable within contemporary populations as well as between anciently diverged populations.

The genetic relationship between the daily circadian clock and the seasonal photoperiodic timer remains a subject of intense controversy. In Wyeomyia smithii, the critical photoperiod (an overt expression of the photoperiodic timer) evolves independently of the rhythmic response to the Nanda–Hamner protocol (an overt expression of the daily circadian clock) over a wide geographical range in North America. Herein, we focus on these two processes within a single local population in which there is a negative genetic correlation between them. We show that antagonistic selection against this genetic correlation rapidly breaks it down and, in fact, reverses its sign, showing that the genetic correlation is due primarily to linkage and not to pleiotropy. This rapid reversal of the genetic correlation within a small, single population means that it is difficult to argue that circadian rhythmicity forms the necessary, causal basis for the adaptive divergence of photoperiodic time measurement within populations or for the evolution of photoperiodic time measurement among populations over a broad geographical gradient of seasonal selection.

Many traits are assumed to have a causal (necessary) relationship with one another because of their common covariation with a physiological, ecological, or geographical factor. Herein we demonstrate a straightforward test for inferring causality using residuals from regression of the traits with the common factor. We illustrate this test using the covariation with latitude of a proxy for the circadian clock and a proxy for the photoperiodic timer in Drosophila and salmon. A negative result of this test means that further discussion of the adaptive significance of a causal connection between the covarying traits is unwarranted. A positive result of this test provides a point of departure that can then be used as a platform from which to determine experimentally the underlying functional connections and only then to discuss their adaptive significance.

Demographic and phenological traits compose the basic elements of an insect's life-history strategy in a seasonal environment. An insect's long-term fitness depends on its ability to exploit favorable conditions, to avoid unfavorable conditions, and to convert from one life style to the other. For the pitcher-plant mosquito, Wyeomyia smithii, we show that genetic variation exists in both development time (a demographic trait) and critical photoperiod (a phenological trait) in six populations spanning much of the species' geographical range. During the northward range expansion of W. smithii in North America, these traits have evolved independently under strong directional and stabilizing selection. The correlated response in critical photoperiod to divergent selection on development time reveals significantly positive genetic correlations in five populations and a negative correlation in one population. The positive correlations form a genetically coordinated phenotype: faster developing individuals use a shorter photoperiodic switch point and are able to exploit the late favorable season; slower developing individuals use a longer photoperiodic switch point and are able to avoid extending development into the unfavorable season. This genetic coordination of demography and phenology has not, however, prevented their independent evolution. We propose that evolutionary flexibility in W. smithii may arise in part from the reorganization of their genetic architecture following repeated founder events during their northward invasion of North America.

We measured the additive genetic variance within populations and the composite additive, dominance, and epistatic effects contributing to differentiation of photoperiodic response between two southern (ancestral) and each of four progressively more northern (derived) populations of the pitcher-plant mosquito, Wyeomyia smithii. Critical photoperiod and its additive genetic variance but not its heritability increased with latitude. Directional selection on critical photoperiod during the northward divergence of W. smithii has therefore not eroded the additive genetic variance underlying this trait. Joint scaling tests of crosses between populations showed that epistatic effects, especially additive x additive and dominance x dominance interactions, overwhelm composite additive and dominance effects on critical photoperiod. The presence of substantial epistasis suggests that multiple founder events during the northward divergence of W. smithii may have been responsible for the release of progressively greater additive genetic variance in derived populations, despite directional and stabilizing selection to reduce it. If epistasis makes a similar contribution to the genetic differentiation of populations in other species, then current models of adaptive evolution that consider only additive genetic variation and covariation within populations may be of limited value in predicting how natural populations differentiate in life history.

To date, all altered patterns of seasonal interactions observed in insects, birds, amphibians, and plants associated with global warming during the latter half of the 20th century are explicable as variable expressions of plastic phenotypes. Over the last 30 years, the genetically controlled photoperiodic response of the pitcher-plant mosquito, Wyeomyia smithii , has shifted toward shorter, more southern daylengths as growing seasons have become longer. This shift is detectable over a time interval as short as 5 years. Faster evolutionary response has occurred in northern populations where selection is stronger and genetic variation is greater than in southern populations. W. smithii represents an example of actual genetic differentiation of a seasonality trait that is consistent with an adaptive evolutionary response to recent global warming.

The distinction between model and nonmodel organisms is becoming increasingly blurred. High-throughput, second-generation sequencing approaches are being applied to organisms based on their interesting ecological, physiological, developmental, or evolutionary properties and not on the depth of genetic information available for them. Here, we illustrate this point using a low-cost, efficient technique to determine the fine-scale phylogenetic relationships among recently diverged populations in a species. This application of restriction site-associated DNA tags (RAD tags) reveals previously unresolved genetic structure and direction of evolution in the pitcher plant mosquito, Wyeomyia smithii , from a southern Appalachian Mountain refugium following recession of the Laurentide Ice Sheet at 22,000–19,000 B.P. The RAD tag method can be used to identify detailed patterns of phylogeography in any organism regardless of existing genomic data, and, more broadly, to identify incipient speciation and genome-wide variation in natural populations in general.

The daily rhythm of 24 h and the annual rhythm of 12 mo constitute the 2 major, highly predictable rhythms of the biosphere. The internal circadian clock enables organisms to track daily changes in their environment; the photoperiodic timer, alone or in concert with a circannual clock, enables organisms to anticipate and prepare in advance for seasonal changes in their environment. The circadian clock entrains to dawn and dusk and tracks light and temperature on a day-to-day basis, while the photoperiodic timer serves as a long-term, physiological go/no-go switch that commits an animal to development, reproduction, dormancy, or migration on a seasonal or even lifetime basis. In 1936, Erwin Bünning proposed that circadian rhythms formed the basis ( Grundlage) for photoperiodic response to day length. Historical inertia generated by correlative evidence from early physiological studies and a proliferating number of descriptive models has resulted in the widespread assumption that the circadian clock constitutes the necessary, causal basis of photoperiodism in general. This historical inertia has also restricted the search for genes involved in insect photoperiodism to genes central to the circadian clock in Drosophila and has led investigators to conclude that any behavior, process, or gene expression that varies with day length represents photoperiodism or a gene involved in photoperiodism. The authors discuss how blinders imposed by the circadian imperative have retarded progress toward identifying the genetic basis of photoperiodism and how the insights gained from geographic variation in photoperiodic response have been used to show the independent evolution of the circadian clock and photoperiodism. When geographic variation is found in circadian genes, the most immediate and parsimonious search for adaptive significance should be in circadian function, not in extrapolation to photoperiodism. Finally, the authors propose that circadian-unbiased, forward genetic approaches should be used to identify genes involved in photoperiodism within extant populations and among populations over evolutionary time.

Background The mosquito Wyeomyia smithii overwinters in a larval diapause that is initiated, maintained and terminated by day length (photoperiod). We use a forward genetic approach to investigate transcriptional events involved in the termination of diapause following exposure to long-days. Methods/Principal Findings We incorporate a novel approach that compares two populations that differentially respond to a single day length. We identify 30 transcripts associated with differential response to day length. Most genes with a previously annotated function are consistent with their playing a role in the termination of diapause, in downstream developmental events, or in the transition from potentially oxygen-poor to oxygen-rich environments. One gene emerges from three separate forward genetic screens as a leading candidate for a gene contributing to the photoperiodic timing mechanism itself (photoperiodic switch). We name this gene photoperiodic response gene 1 (ppdrg1). WsPpdrg1 is up-regulated under long-day response conditions, is located under a QTL for critical photoperiod and is associated with critical photoperiod after 25 generations of recombination from a cross between extreme phenotypes. Conclusions Three independent forward genetic approaches identify WsPpdrg1 as a gene either involved in the photoperiodic switch mechanism or very tightly linked to a gene that is. We conclude that continued forward genetic approaches will be central to understanding not only the molecular basis of photoperiodism and diapause, but also the evolutionary potential of temperate and polar animal populations when confronted with rapid climate change.

The insect fauna of water-filled tree holes in southern Britain consists primarily of the mosquitoes Aedes geniculatus, Anopheles plumbeus, Culex torrentium, and a benthic detritivorous fauna that includes primarily the scirtid beetle Prionocyphon serricornis and the chironomid midge Metriocnemus martinii. Culex torrentium has been documented only relatively recently in tree holes but all three species of mosquitoes partition the resource in space and time. When mosquito larvae were forced to coexist in natural tree holes at limiting densities and at higher than natural levels of interspecific encounter, there was no evidence that Aedes geniculatus or Anopheles plumbeus affected pupation success, pupal weight, or development time of the other or that either Aedes geniculatus or C. torrentium affected the survivorship, pupation success, pupal weight, and biomass yield of the other. When A. geniculatus at limiting densities were forced in natural tree holes to live without or to coexist with natural or twice natural densities of P. serricornis and M. martinii, the presence, absence, or superabundance of the benthic insects did not affect pupation success or pupal weight of A. geniculatus; development time of A. geniculatus was faster when a superabundance of the benthic fauna was present. Effects of the benthic fauna on A. geniculatus are slight and the only significant interaction is facilitative, not competitive. The pattern of habitat segregation among treehole mosquitoes in southern Britain is characteristic of their respective genera and we propose that this pattern is more likely (but not certain) to have arisen through a process of independent evolution than through competitively driven niche shifts among already coexisting species.

Cohort fitness and individual female pupal weight in a northern (40°N) U.S. population of Wyeomyia smithii (Coquillett) were inversely proportional to larval density on a fixed resource. Larger females lived longer and realized higher absolute and weight-specific fecundity than did smaller individuals. Females produced a large initial batch of eggs early during their reproductive period and several smaller batches later. Early fecundity was correlated with pupal weight but not longevity. Late fecundity increased with both pupal weight and longevity but there was no significant tradeoff between late and early weight-specific fecundity. Fertility (percent hatch) was higher in earlier than later batches of eggs but there were no significant trade-offs between late fertility and either early fertility or early weight-specific fecundity. Variation in larval density results in reduced weight at pupation which, in turn, results in reduced fecundity and longevity. Consequences of increased larval density thus persist into adulthood as unrealized reproductive potential.

Drosophila melanogaster from Australia, Europe and North America enter an adult ovarian dormancy in response to short days and low temperatures. The independent effects of temperature and day length in the determination of dormancy have been examined only in one long-established laboratory line (Canton-S). In all other studies of natural or laboratory populations, dormancy has been assessed at either a single short day or a single moderately low temperature. Herein, we determine the relative roles of temperature, photoperiod, and their interaction in the control of ovarian dormancy in D. melanogaster from two natural populations representing latitudinal extremes in eastern North America (Florida at 27 degrees N and Maine at 44 degrees N). In both natural populations, temperature is the main determinant of dormancy, alone explaining 67% of the total variation among replicate isofemale lines, whereas photoperiod has no significant effect. We conclude that ovarian dormancy in D. melanogaster is a temperature-initiated syndrome of winter-tolerant traits that represents an adaptive phenotypic plasticity in temperate seasonal environments.

Understanding gene interaction and pleiotropy are long-standing goals of developmental and evolutionary biology. We examine the genetic control of diapause in insects and show how the failure to recognize the difference between modular and gene pleiotropy has confounded our understanding of the genetic basis of this important phenotype. This has led to complications in understanding the role of the circadian clock in the control of diapause in Drosophila and other insects. We emphasize three successive modules – each containing functionally related genes – that lead to diapause: photoperiodism, hormonal events and diapause itself. Understanding the genetic basis for environmental control of diapause has wider implications for evolutionary response to rapid climate change and for the opportunity to observe evolutionary change in contemporary time.

1 Populations of three mosquitoes, Aedes geniculutus (Olivier), Anopheles plumbeus Stephens and Culex torrentium Martini, were found in three types of tree holes: rot holes penetrating the bark of the host tree and exposed, or deeper, more cryptic, bark-lined buttress holes. Aedes geniculatus occupied all three tree-hole types but reached their greatest abundance in exposed buttress holes; A.plumbeus predominated in rot holes; C.torrentium occupied buttress holes exclusively. 2 After a 1 month period of winter freezing, larvae of A.plumbeus survived better than larvae of A. genicularus. Freeze resistance of A.plumbeus increased from cryptic buttress to rot holes. Freeze resistance of A.geniculatus declined from exposed buttress to cryptic buttress to rot holes and was markedly higher in more protected than in more exposed macrohabitats. 3 Among manipulated larval cohorts of A.geniculutus and A.plumbeus, survivorship, pupation success, pupal weight, and/or biomass yield did not differ among tree-hole types or was the reverse of freeze-related survivorship. Despite seasonal differences in fitness correlates, annual fitness may be similar among tree-hole types for both species. Manipulations carried out at a single census or during a single season are therefore likely to produce misleading information about fitness variation among sub-habitats. 4 Both species should exploit a broad range of tree-hole types but A.plumbeus occupies more restricted types of tree holes than does A.geniculutus. The restricted habitat usage by A.plumbeus does not parallel their fitness among tree-hole types in southern Britain but does resemble that of other tree-hole anophelenes. We therefore propose that habitat selection by A.plumbeus is more likely due to their phylogenetically determined physiological tolerances and behaviour than to competition from coexisting A.geniculatus.

For over 70 years, researchers have debated whether the ability to use day length as a cue for the timing of seasonal events (photoperiodism) is related to the endogenous circadian clock that regulates the timing of daily events. Models of photoperiodism include two components: (1) a photoperiodic timer that measures the length of the day, and (2) a photoperiodic counter that elicits the downstream photoperiodic response after a threshold number of days has been counted. Herein, we show that there is no geographical pattern of genetic association between the expression of the circadian clock and the photoperiodic timer or counter. We conclude that the photoperiodic timer and counter have evolved independently of the circadian clock in the pitcher-plant mosquito Wyeomyia smithii and hence, the evolutionary modification of photoperiodism throughout the range of W. smithii has not been causally mediated by a corresponding evolution of the circadian clock.

A wide diversity of organisms use photoperiod (daylength) as an environmental cue to anticipate the changing seasons and to time various life-history events such as dormancy and migration. Photoperiodic time measurement consists of two main components, (1) the photoperiodic timer that discriminates between long and short days, and (2) the photoperiodic counter that accumulates and stores information from the timer and then induces the phenotypic output. Herein, we use extended night treatments to show that light is necessary to accumulate photoperiodic information across the geographic range of the mosquito, Wyeomyia smithii and that the photoperiodic counter counts extrinsic (external) light:dark cycles and not endogenous (internal) circadian cycles.