Insect Photoperiodism Research Articles

Time measurement in insect photoperiodism: external and internal coincidence

The identity and nature of the photoperiodic photoreceptors are now quite well known, as is the nature of the endocrine regulation of the resulting diapauses. The central problem of time measurement—how the photoperiodic clock differentiates long from short days—however, is still obscure, known only from whole-animal experiments and abstract models, although it is clearly a function of the insect circadian system. This review describes some of these experiments in terms of oscillator entrainment and two widely applicable photoperiodic clock models, external and internal coincidence, mainly using data from experiments on flesh flies (Sarcophaga spp) and the parasitic wasp, Nasonia vitripennis.

Circadian and Neuroendocrine Basis of Photoperiodism Controlling Diapause in Insects and Mites: A Review.

The photoperiodic system is concealed in the highly complex black-box, comprising four functional subunits: 1) a photo/thermo-sensitive input unit, 2) a photoperiodic clock based on a circadian system, 3) a condenser unit counting the number of inductive signals, and 4) a neuroendocrine switch that triggers a phenotypic shift. This review aims to summarize the research history and current reach of our understanding on this subject to connect it with the molecular mechanism of the circadian clock rapidly being unveiled. The review also focuses on the mode of intersubunit information transduction. It will scan the recent advancement in research on each functional subunit, but special attention will be given to the circadian clock–endocrine conjunct and the role of melatonin signaling in the regulation of insect photoperiodism. Prothoracicotropic hormone (PTTH) probably plays the most crucial role in the regulation of pupal diapause, which is the simplest model system of diapause regulation by hormones investigated so far, particularly in the Chinese oak silkmoth (Antheraea pernyi). A search for the trigger to release the PTTH found some candidates, that is, indoleamines. Indolamine metabolism is controlled by arylalkylamine N-acetyltransferase (aaNAT). Indolamine dynamics and aaNAT enzymatic activity changed according to photoperiods. aaNAT activity and melatonin content in the brain showed not only a photoperiodic response but also a circadian fluctuation. aaNAT had multiple E-boxes, suggesting that it is a clock-controlled gene (ccg), which implies that cycle (cyc, or brain–muscle Arnt-like 1 = Bmal1)/Clock (Clk) heterodimer binds to E-box and stimulates the transcription of aaNAT, which causes the synthesis of melatonin. RNAi against transcription modulators, cyc, or Clk downregulated aaNAT transcription, while RNAi against repressor of cyc/Clk, per upregulated aaNAT transcription. Immunohistochemical localization showed that the circadian neurons carry epitopes of melatonin-producing elements such as aaNAT, the precursor serotonin, HIOMT, and melatonin as well as clock gene products such as cyc-ir, Per-ir, and dbt-ir, while PTTH-producing neurons juxtaposed against the clock neurons showed hMT2-ir in A. pernyi brain. Melatonin probably binds to the putative melatonin receptor (MT) that stimulates Ca2+ influx, which in turn activates PKC. This induces Rab 8 phosphorylation and exocytosis of PTTH, leading to termination of diapause. All the PTTH-expressing neurons have PKC-ir, and Rab8-ir. When diapause is induced and maintained under short days, serotonin binding to 5HTR1B suppresses PTTH release in a yet unknown way. RNAi against this receptor knocked out photoperiodism; short day response is blocked and diapause was terminated even under the short day condition. The result showed that a relatively simple system controls both induction and termination in pupal diapause of A. pernyi: the circadian system regulates the transcription of aaNAT as a binary switch, the enzyme produces a melatonin rhythm that gates PTTH release, and 5HTR1B and MT are probably also under photoperiodic regulation. Finally, we listed the remaining riddles which need to be resolved, to fully understand this highly complex system in future studies.

Time measurement in insect photoperiodism: The role of photophase duration and light intensity

The initial stages of diapause induction - as summer gives way to autumn - involve a process of time measurement in which the duration of daylength (or nightlength) is determined by a photoperiodic 'clock' based upon the circadian system. In many insects so far examined, a photophase of sufficient duration and illuminance resets a photoperiodic oscillator to a constant phase equivalent to the beginning of the 'subjective night' (Circadian time, CT 12 h) whereupon it proceeds to measure nightlength in a clock of the external coincidence type. A possible exception may be found in the linden bug, Pyrrhocoris apterus, which - in laboratory studies using relatively low light intensity - suggests that daylength is measured rather than the night. Earlier studies of photoperiodic responses (pupal diapause induction) in the flesh fly Sarcophaga argyrostoma, however, showed that 'weak' or short light pulses giving rise to Type 1 phase response curves could be converted by increasing light intensity to Type 0 responses that phase set the oscillation to the beginning of the subjective night (CT 12 h) whereupon it could begin to measure the night. Based upon these data it is therefore suggested that the photoperiodic clock in P. apterus might also measure nightlength if the bugs were exposed to photophases of higher irradiance simulating daytime exposure to the light intensity experienced by these diurnally active insects in their natural environment.

Circadian clock outputs regulating insect photoperiodism: A potential role for glutamate transporter

Many temperate ectotherms survive winter by entering diapause – a state of developmental (or reproductive) suppression or arrest – in response to short autumnal day lengths. Day lengths are assessed by the circadian clock, the biological time-keeping system that governs biological rhythms with a period of approximately 24 h. However, clock output molecules controlling this photoperiodic response are largely unknown for many insects. To identify these molecules in Hemiptera, we performed RNAi knockdowns of several candidate genes in the bean bug Riptortus pedestris to determine whether their silencing affects photoperiodic regulation of ovarian development (reproductive diapause). Knockdown of diuretic hormone 31, short neuropeptide F, neuropeptide F, ion transport peptide, neuropeptide-like precursor 1, and choline acetyltransferase had no effect on ovarian development and were therefore ruled out as regulators of the photoperiodic response. However, knockdown of vesicular glutamate transporter promoted ovarian development under diapause-inducing short days, and this is the first report of the functional involvement of glutamate signalling in insect photoperiodism. Improved knockdown of this transporter (or receptor) and RNAi of other genes involved in glutamate signal transduction is required to verify its role as an output of the circadian clock.

Insect photoperiodism: Bunning's hypothesis, the history and development of an idea

In insects, the photoperiodic system comprises a linked sequence of events from photoreception to final seasonally-appropriate phenotypes such as overwintering diapause. The first and last of these events are reasonably well known, but central phenomena such as those distinguishing short from long days (time measurement) and the nature, accumulation and transfer of this information through development, metamorphosis and sometimes across generations remains obscure. BĂźnning's intuitive suggestion that photoperiodic time measurement was a function of the circadian system, made eight decades ago, however, has provided a framework for numerous studies investigating these connections. This review examines the development of BĂźnning's hypothesis from its origin in plants to the physiology of diapause in insects. Despite considerable inter-species differences, a close and probably causal relationship between circadian rhythmicity and photoperiodism is indicated.

Insect photoperiodism: Seasonal development on a revolving planet

This review starts by comparing photoperiodic diapause with non-photoperiodic quiescence in four representative species, paying particular attention to overwintering in Drosophila melanogaster. In the second part it describes dormancy strategies of insects from the equator to the polar regions, addressing topics such as the role of the circadian system in photoperiodic time measurement, latitudinal clines in diapause-related traits, hourglass-like photoperiodic clocks based on dampening circadian oscillators, and the dormancy strategies of insects close to the equator or at high latitudes where seasonal changes in photoperiod are unreliable or absent.

Dormancy, Diapause, and the Role of the Circadian System in Insect Photoperiodism.

Whole-animal experiments devised to investigate possible association between photoperiodic time measurement and the circadian system (Bünning's hypothesis) are compared with more recent molecular investigations of circadian clock genes. In Sarcophaga argyrostoma and some other species, experimental cycles of light and darkness revealed a photoperiodic oscillator, set to constant phase at dusk and measuring night length repeatedly during extended periods of darkness. In some species, however, extreme dampening revealed an unrepetitive (i.e., hourglass-like) response. Rhythms of clock gene transcript abundance may also show similar phase relationships to the light cycle, and gene silencing of important clock genes indicates that they play a crucial role in photoperiodism either alone or in concert. However, the multiplicity of peripheral oscillators in the insect circadian system indicates that more complex mechanisms might also be important.

Roles of the circadian clock and endocrine regulator in the photoperiodic response of the brown‐winged green bug Plautia stali

AbstractThe physiological mechanisms underlying photoperiodism in insects have been studied extensively, although the associated molecular machinery remains largely unknown. In the present study, we investigate the roles of the circadian clock gene cycle (cyc) and the endocrine regulator gene myoinhibitory peptide (Mip) in the photoperiodic response of the brown‐winged green bug Plautia stali Scott (Hemiptera, Pentatomidae). Typically, adult females of this species develop their ovaries under long‐day conditions, whereas they suppress its development under short‐day conditions. We find that RNA interference (RNAi) directed against cyc causes malfunction of the circadian clock governing the locomotor activity rhythm and yields abnormal activity profiles not only under constant darkness, but also under light/dark conditions. RNAi directed against cyc and Mip disrupts the photoperiodic response in ovarian development. cyc RNAi suppresses the ovarian development even under long‐day conditions, whereas Mip RNAi induces it even under short‐day conditions. We propose that the core circadian clock gene cyc regulates the photoperiodic response and that Mip is the causal regulator of juvenile hormone biosynthesis in the corpus allatum. Neither photoperiod, nor cyc RNAi affect Mip mRNA levels, and therefore it remains unknown how the photoperiodic information is processed and mediated by Mip.

Convergent photoperiodic plasticity in developmental rate in two species of insects with widely different thermal phenotypes

Growth and development rates in many insects are affected by photoperiod, which enables insects to synchronize their life histories with seasonal events, but this aspect of insect photoperiodism remains understudied. Here we use several experimental combinations of constant day length and temperature to determine whether there are quantitative developmental responses to photoperiod in the bug Scantius aegyptius and leaf beetle Timarcha tenebricosa. The thermal ecology of these two species is strikingly different: the former is thermophilic and active throughout summer and the latter is spring-active and avoids the hottest time of the year. In accordance with their contrasting natural thermal environments, S. aegyptius survives better and achieves a larger final body mass at the high experimental temperatures, while T. tenebricosa survives better and is heavier at the low experimental temperatures. Despite this polarity, long-day conditions accelerate larval development relative to a short-day photoperiod in both species, and this developmental response is stronger at low temperatures. Our re-visitation of previous literature in light of the new findings indicates that this similarity in photoperiodic response is superficial and that relatively faster development in midsummer is likely to have a different ecological role in summer- and spring-active species. In the former, it may allow completion of an additional generation during the favourable season, whereas in the latter, this acceleration likely ensures that the larval stage, which is vulnerable to heat, is completed before the onset of hot weather.

Insect Arylalkylamine N-Acyltransferases: Mechanism and Role in Fatty Acid Amide Biosynthesis.

Arylalkylamine N-acyltransferases (AANATs) catalyze the formation of an N-acylamide from an acyl-CoA thioester and an amine. One well known example is the production of N-acetylserotonin from acetyl-CoA and serotonin, a reaction in the melatonin biosynthetic pathway from tryptophan. AANATs have been identified from a variety of vertebrates and invertebrates. Considerable efforts have been devoted to the mammalian AANAT because a cell-permeable inhibitor specifically targeted against this enzyme could prove useful to treat diseases related to dysfunction in melatonin production. Insects are an interesting model for the study of AANATs because more than one isoform is typically expressed by a specific insect and the different insect AANATs (iAANATs) serve different roles in the insect cell. In contrast, mammals express only one AANAT. The major role of iAANATs seem to be in the production of N-acetyldopamine, a reaction important in the tanning and sclerotization of the cuticle. Metabolites identified in insects including N-acetylserotonin and long-chain N-fatty acyl derivatives of dopamine, histidine, phenylalanine, serotonin, tyrosine, and tryptophan are likely produced by an iAANAT. In vitro studies of specific iAANATs are consistent with this hypothesis. In this review, we highlight the current metabolomic knowledge of the N-acylated aromatic amino acids and N-acylated derivatives of the aromatic amino acids, the current mechanistic understanding of the iAANATs, and explore the possibility that iAANATs serve as insect “rhymezymes” regulating photoperiodism and other rhythmic processes in insects.

Dedicated issue: photoperiodic induction of diapause and seasonal morphs

Whether transitions from warm to cold in temperate regions or dry to wet in areas nearer the equator, the changing of the seasons is a major stress for most organisms and as such, a major organizer of organismal lifecycles. To mitigate stressful periods and to exploit periods with high resource availability that are favorable for growth and development, insects and other arthropods have evolved a suite of seasonal adaptations including programmed seasonal dormancy, diapause. Yet, climate change, urbanization, and other alterations in land use are increasingly exposing insects and other organisms to changes in seasonality, especially decreased winter cold periods, lengthened growing seasons, and increased climatic variability on the seasonal shoulders in the fall and spring where warm and cold spells are frequently interspersed. Furthermore, increasing globalization has led to many insects expanding into new areas with very different patterns of seasonality than in their home ranges, whether due to inadvertent introductions of invasive species or managed introductions of beneficial pollinators or natural enemies. We must continue to expand our understanding of insect seasonal responses to meet these increasing challenges in seasonal adaption in our ever-changing world. Building greater understanding of the seasonal responses of both pest and beneficial insects will require an inherently interdisciplinary approach that includes a range of expertise from studying organismal behavior and field ecology to identifying the genetic architecture and regulatory physiology of diapause. A recent symposium at the XXV International Congress of Entomology held in Orlando, Florida, USA from 25–30 September 2016 titled ‘Photoperiodic Induction of Diapause and Seasonal Morphs’ provided the forum for such a broad and interdisciplinary discussion. Many symposium participants and several additional investigators generously agreed to contribute papers to this Dedicated Issue of Physiological Entomology. Reflective of the breadth in approaches and perspectives within the sub-field of insect seasonal biology, this issue contains a range of article types. One particularly broad article leverages the rapidly accumulating transcriptomic datasets available for a wide variety of insects and performs a meta-analysis to test the degree to which a common ‘genetic toolkit’ underlies insect diapause responses. Another article reviews the classic literature on diapause in spider mites and puts forth a modern perspective on the genetics of spider mite reproductive diapause with a focus on the essential role of carotenoid compounds. Several more primary data articles follow that range in their approach and perspectives from whole-brain transcriptomics, to focused transcript abundance work on clock genes and photoperiodic responses, and even classical genetic architecture of diapause inheritance patterns. Additional articles cover a gamut of topics from evolutionary ecology of bet-hedging strategies in diapause length and the timing of diapause termination to endocrine changes associated with progression through diapause development, again reflecting the integrative nature of studying insect photoperiodism and seasonal responses. We thank all of our contributors for their timely submissions and efforts that have made it possible to continue the long tradition of both symposia on photoperiodism and seasonal biology at the International Congress of Entomology and producing a published series of papers allied with that session. We expect that this issue will serve to stimulate discussion of past, contemporary, and future issues in the study of insect seasonal biology for both specialist practitioners within the field and among insect biologists in general.

Both the anterior and posterior eyes function as photoreceptors for photoperiodic termination of diapause in the two-spotted spider mite

Photoreceptors involved in photoperiodism in insects and mites can be either the retinal photoreceptors in the visual system or nonvisual extraretinal photoreceptors. Mites with no eyes have a clear photoperiodic response, suggesting the involvement of extraretinal photoreceptors in mite photoperiodism. In mites equipped with eyes, however, it is not known whether the retinal or extraretinal photoreceptors are involved in photoperiodism. The two-spotted spider mite Tetranychus urticae possesses two pairs of eyes. Adult females of this species terminate diapause in response to long days. To investigate whether the eyes function as photoperiodic photoreceptors in T. urticae, their eyes were ablated using a laser ablation system. Mites with their eyes intact terminated diapause under long days after low temperature exposure, whereas they remained in diapause under short days. Under constant darkness, they did not terminate diapause. When all eyes were removed, the mites remained in diapause even when they were maintained under long days. In contrast, the mites showed clear photoperiodic response when only the anterior or posterior eyes were removed. These results indicate that both the anterior and posterior eyes function as photoreceptors in photoperiodic termination of diapause in T. urticae.

Insect photoperiodism: effects of temperature on the induction of insect diapause and diverse roles for the circadian system in the photoperiodic response

AbstractThis review considers the effects of temperature on insect diapause induction and the photoperiodic response, and includes constant temperature, temperature cycles, pulses and steps in daily light–dark cycles, constant darkness and in constant light, all with reference to various circadian‐based “clock” models. Although it is a comparative survey, it concentrates on two species, the flesh fly Sarcophaga argyrostoma and its pupal parasite Nasonia vitripennis, which possess radically different photoperiodic mechanisms, although both are based upon the circadian system. Particular attention is given to the effects of daily thermoperiod in darkness and to low and high temperature pulses in conjunction with a daily light–dark cycle, treatments that suggest that S. argyrostoma “measures” night length with a “clock” of the external coincidence type. However, N. vitripennis responds to seasonal changes in photoperiod with an internal coincidence device involving both “dawn” and “dusk” oscillators. Other species may show properties of both external and internal coincidence. Although the precepts of external coincidence have been well formulated and supported experimentally, those for internal coincidence remain obscure.

The involvement of the brain region containing pigment-dispersing factor-immunoreactive neurons in the photoperiodic response of the bean bugRiptortus pedestris

The concept of insect photoperiodism based on a circadian clock has been supported by many studies demonstrating that the behavioural circadian rhythm and the photoperiodic response are driven by the same circadian clock genes. However, the neuronal mechanism of the circadian clock underlying photoperiodism is poorly understood. To examine whether circadian rhythm and photoperiodism share a neuronal mechanism, we focused on the neurons that express neuropeptide pigment-dispersing factor (PDF) in the bean bug, Riptortus pedestris. PDF has been identified as an important regulator of the insect circadian rhythm and is expressed in circadian clock neurons of various insect species. In R. pedestris, PDF immunoreactivity was detected in some clusters of cells and their fibres in the optic lobe and the protocerebrum. cDNA encoding a PDF precursor protein was highly conserved between R. pedestris and many other insects. Differences between day and night were not observed in the immunolabelling intensity in cell bodies of PDF-immunoreactive neurons and pdf mRNA expression levels in the head. Surgical removal of the region containing PDF-immunoreactive cell bodies at the medulla disrupted the photoperiodic regulation of diapause. However, gene suppression of pdf by RNA interference did not affect the photoperiodic response. These results suggest that the region containing PDF-immunoreactive somata is important for the photoperiodic response in R. pedestris, but pdf mRNA expression is probably not required for the response.

Roles of circadian clock genes in insect photoperiodism

AbstractFunctional involvement of a circadian clock in photoperiodism for measuring the length of day or night had been proposed more than 70 years ago, and various physiological experiments have supported the idea. However, the molecular basis of a circadian clock has remained veiled in insects. Nevertheless, our knowledge of the functional elements of a circadian clock governing circadian rhythmicity has advanced rapidly. Since both circadian rhythms and photoperiodism depend on the daily cycles of environmental changes, it is easy to assume that the same clock elements are involved in both processes. Recently, the RNA interference (RNAi) technique clarified that the molecular machinery of a circadian clock governing photoperiodism is identical to that governing circadian rhythmicity. Here, I review the theoretical background of photoperiodic responses incorporating a circadian clock(s) and recent progress on the molecular clockwork involved in photoperiodism in the bean bug Riptortus pedestris and other insect species. I have focused on the intense controversy regarding the involvement of a circadian clock in insect photoperiodism.

Unity and diversity in the insect photoperiodic mechanism*

AbstractThis review examines some of the models to account for time measurement in insect photoperiodism. It considers the supporting evidence for these models and the attempts to discriminate among them. Although hourglass timers may exist, it is suggested that most photoperiodic mechanisms, including many hourglass‐like timers, are circadian‐based, making Bünning's original hypothesis, that the circadian system somehow provides the essential “clockwork” for photoperiodic timing, the most persuasive unifying principle. The apparent diversity among modern species in their modes of time measurement is probably the result of differences between the underlying circadian systems that were adopted for seasonal night length measurement as the insects, or groups of insects, moved northwards into areas with a pronounced winter season. Photoperiodic time measurement, therefore, exhibits both unity (in their common circadian basis) and diversity in detail. Attention to this diversity may provide invaluable insights into the problem of photoperiodic time measurement at comparative, and molecular, levels.

Photoperiodic response requires mammalian-type cryptochrome in the bean bug Riptortus pedestris

The hypothesis that a circadian clock comprised of circadian clock genes is causally involved in insect photoperiodism has been supported by several studies. However, there remains a possibility that the effects of the circadian clock genes on photoperiodism are exerted through pleiotropic (non-circadian) functions provided by each gene independently from its role in the circadian clock. In the present study, we investigated the involvement of the circadian clock gene mammalian-type cryptochrome ( cry-m) in photoperiodic regulation of ovarian development in the bean bug Riptortus pedestris by using RNA interference (RNAi). Injection of cry-m double-stranded RNA (dsRNA) induced expression of period ( per), whereas did not affect expression of cycle ( cyc), showing that CRY-m functions as a negative element on CYC-mediated transcription in the circadian clock. If the circadian clock is indeed involved in photoperiodism, a phenotype produced by RNAi of cry-m will be the same as that produced by RNAi of per, another negative element. The intact insects and insects injected with control dsRNA were found to enter diapause when kept under short-day conditions after adult emergence, while they developed ovaries when kept under long-day conditions after adult emergence. However, cry-m RNAi significantly increased the incidence of reproductive individuals under diapause-inducing short-day conditions, as per RNAi did, in accordance with our expectation.

What Season Is It Anyway? Circadian Tracking vs. Photoperiodic Anticipation in Insects

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.

Circadian rhythms and the evolution of photoperiodic timing in insects

Abstract.This review discusses possible evolutionary trends in insect photoperiodism, mainly from a chronobiological perspective. A crucial step was the forging of a link between the hormones regulating diapause and the systems of biological rhythms, circadian or circannual, which have independently evolved in eukaryotes to synchronize physiology and behaviour to the daily cycles of light and darkness. In many of these responses a central feature is that the circadian system resets to a constant phase at the beginning of the subjective night, and then ‘measures’ the duration of the next scotophase. In ‘external coincidence’, one version of such a clock, light now has a dual role. First, it serves to entrain the circadian system to the stream of pulses making up the light/dark cycle and, second, it regulates the nondiapause/diapause switch in development by illuminating/not illuminating a specific light sensitive phase falling at the end of the critical night length. Important work by A. D. Lees on the aphid Megoura viciae using so‐called ‘night interruption experiments' demonstrates that pulses falling early in the night lead to long‐day effects that are reversible by a subsequent dark period longer than the critical night length and also show maximal sensitivity in the blue–green range of the spectrum. Pulses falling in the latter half of the night, however, produce long‐day effects that are irreversible by a subsequent long‐night and show a spectral sensitivity extending into the red. With movement to higher latitudes, insects develop genetic clines in various parameters, including critical night length, the number of long‐night cycles needed for diapause induction, the strength of the response, and the ‘depth’ or intensity of the diapause thus induced. Evidence for these and other types of photoperiodic response suggests that they provided strong selective advantages for insect survival.

Clock genes period and timeless are rhythmically expressed in brains of newly hatched, photosensitive larvae of the fly, Sarcophaga crassipalpis

While roles of the clock genes period ( per) and timeless ( tim) are relatively well understood in relation to circadian clocks, their potential roles in insect photoperiodism remain enigmatic. In this study, the expression of per and tim genes under two contrasting photoperiods is described in the central nervous system of photoperiodically sensitive, newly hatched first instar larvae of the flesh fly, Sarcophaga crassipalpis. Using qPCR, diel oscillations were observed in the mRNA levels of both genes under long-day (15 h light:9 h dark, promotes direct development) and short-day conditions (11 h light:13 h dark, induces pupal diapause). Peak per and tim mRNA oscillations were closely associated with the light/dark transition. The conspicuous difference between the two photoperiodic conditions was that the sharp increase in per and tim mRNA abundance occurred during the light phase under long days but during the dark phase under short days. The diel oscillations were, at least in part, driven by an endogenous component, as demonstrated by transferring larvae to continuous darkness. The cells displaying Tim- and Per-like immunoreactivities (Tim- and Per-LIRs) were localized using anti- Drosophila-Per and anti- Chymomyza-Tim antibodies. Per-LIR and Tim-LIR co-localized in three groups of cells in each brain hemisphere. Two other groups, one in the brain hemispheres and the other in the fused ventral nerve ganglion, expressed only the Per-LIR.