Non-diapausing Pupae Research Articles

Comparison of the circadian eclosion rhythm between non-diapause and diapause pupae in the onion fly, Delia antiqua

The influence of pupal diapause on adult eclosion rhythm of Delia antiqua was investigated. When non-diapause and diapause pupae were exposed to various photoperiods at 15, 20 and 25 °C, both of them emerged as adults close to the light-on time, but the phase of eclosion varied with photoperiod and temperature. Moreover, there was a significant difference in the eclosion time between non-diapause and diapause pupae; the eclosion peak of diapause pupae was earlier than that of non-diapause pupae. When non-diapause and diapause pupae were transferred to constant darkness (DD) after having experienced LD 12:12 at 15, 20 and 25 °C, both showed circadian rhythmicity in eclosion. Although the free-running period ( τ) decreased slightly as temperature increased in both non-diapause and diapause pupae, the latter tended to show shorter τ than the former. This observation suggests that the observed difference in eclosion time in LD cycles between non-diapause and diapause pupae is due to differences in τ.

Effect of Cold Storage on Emergence, Longevity, Fertility, and Survival of Cotesia marginiventris (Hymenoptera: Braconidae)2

The effect of short-term cold storage on non-diapausing pupae and adults of Cotesia marginiventris (Cresson) was evaluated for developing protocols for the mass production of this parasitoid. When stored at 10°C, successful emergence of adults from cocoons occurred within 6 d and was not affected by storage time (0 d, 4 d, 12 d, or 20 d). Longevity of males and females was not affected by cold storage, but fewer males and females remained alive per consecutive 8 d interval after emergence at 27°C. A fertility experiment revealed that fewer progeny (F1 cocoons, F1 females) were produced by parental females previously cold-stored as pupae in cocoons for 20 d rather than 4 d. A within-storage adult survival experiment demonstrated that males and females were not affected by any of the storage times (8 d, 16 d, 24 d, 32 d). This survival rate was achieved by removing the adults from the cold at 8 d intervals, placing them at 23 ± 1 °C for ~ 2 h and providing honey. Another fertility experiment revealed that more F1 males, but fewer F1 females, were produced when the parental females (which had been cold-stored as adults) were stored for 32 d rather than for 16 d. This study suggests that adults are more amenable to short-term storage than pupae if the adults are periodically removed for feeding. Nevertheless, cold storage of cocoons (pupae) for 20 d or adults for 32 d can limit the production of female progeny.

Physiology of diapause and cold hardiness in the overwintering pupae of the fall webworm Hyphantria cunea (Lepidoptera: Arctiidae) in Japan

The fall webworm Hyphantria cunea Drury, which was accidentally introduced to Japan in 1945, overwinters on the ground in pupal diapause. Diapause termination, as indicated by the respiration rate and the period required for adult emergence, began in March and ended in April. Cold hardiness (the ability to survive exposure to −15°C) decreased linearly with diapause development from November to the following April under field conditions. Cold hardiness of diapause pupae (DP) decreased as the acclimation temperature decreased from 15 to −10°C, whereas cold hardiness of non-diapause pupae (NDP) remained high as the acclimation temperature decreased from 5 to −5°C. However, H. cunea in Japan can survive exposure to −5°C for two weeks, whether it is in a diapause or non-diapause state. Trehalose was the main sugar detected in the body, but its level was less than 0.8%. Trehalose levels increased in field-collected pupae from January to March. DP accumulated less trehalose than NDP, as the acclimation temperature was decreased from 5 to −5°C. The alanine content in field-collected pupae increased from November to February. Both diapause and low temperature caused an accumulation of alanine. These results suggest that under field conditions, overwintering pupae of H. cunea in Japan do not accumulate high levels of sugars and polyols and do not develop a high level of cold hardiness. Furthermore, DP do not accumulate high levels of sugars and polyols and their ability to survive exposure to −15°C is not greater than that of NDP. The physiological and biochemical bases of diapause in H. cunea from Japan are discussed.

Dynamic changes in cold hardiness, high-temperature tolerance and trehalose content in the onion maggot, Delia antiqua (Diptera: Anthomyiidae), associated with the summer and winter diapause.

Changes in cold hardiness, high-temperature tolerance and trehalose content in the onion maggot, Delia antiqua (Diptera: Anthomyiidae), associated with the summer and winter diapause were investigated. Survival of summer- and winter-diapausing pupae after a 15 day exposure to −15°C was greater than 80%, while less than 5% of non-diapausing pupae survived the same treatment. Greater than 80% of winter-diapausing pupae survived −23°C for 15 days, but survival of summer-diapausing pupae at the same procedure was less than 30%. More than 75% of summer- and winter-diapausing pupae, but not non-diapausing pupae, tolerated 35°C for 15 days. Trehalose concentrations in summer- and winter-diapausing pupae were not high at the beginning of diapause (≈5 μg/mg), but increased gradually and reached 10 μg/mg in 40 days, although they were kept at relatively high temperatures of 25°C (summer diapause) or 15°C (winter diapause). Cold hardiness of pupae, however, was not proportional to their trehalose content; although trehalose concentrations in winter- and summer-diapausing pupae at day 40 (WD40 and SD40) were similar (≈10 μg/mg), cold hardiness of these pupae differed largely (WD40>SD40).

Relationship between cold hardiness and northward invasion in the great mormon butterfly, Papilio memnon L. (Lepidoptera: Papilionidae) in Japan.

To investigate factors involved in the recent northward invasion of the great mormon butterfly, Papilio memnon L. (Lepidoptera: Papilionidae), cold hardiness of pupae was compared among 4 populations including those in a subtropical and a northernmost area of Japan. The mean supercooling points (SCP) of diapausing and non-diapausing pupae were lower than −20°C in all populations. The SCP was not affected by an acclimation period nor by pupal weight, although it became higher with inoculation. Diapausing pupae were placed in 4 sites at different altitudes in southern Osaka Prefecture during the winter, then survival rates were compared in the spring. All individuals died at altitudes of 1, 100 and 800 m, while more than half of the individuals survived the winter at 400 and 30 m. There was no difference in the survival rate at each site among populations. Even the subtropical population showed cold hardiness to survive winter in Osaka, which is located near the northern edge of the distribution of this species. These results show that the recent northward invasion has occurred without any substantial changes in the cold hardiness of this species. Based on the results, we inferred that the climatic lethal limits during the winter lie between −3.8 and −7.3°C for the minimum temperature, and between 52 and 68 frost days. We discuss the effect of climatic warming on the northward invasion of this species.

Diapausing pupae of the flesh fly Sarcophaga crassipalpis (Diptera: Sarcophagidae) are more resistant to inoculative freezing than non-diapausing pupae

Summary The resistance of diapausing (overwintering) and non-diapausing (summer) Sarcophaga crassipalpis (Diptera: Sarcophagidae) pupae to inoculative freezing was examined. Although both types of pupae resisted inoculative freezing after 24-h submergence in water, diapausing pupae were overall significantly more resistant than non-diapausing pupae. Exposing the thin pupal cuticle by removing the ends of the puparial case eliminated the capacity of both pupal types to resist inoculative freezing, indicating that resistance to inoculative freezing resides with the puparium. Pupae submerged in surfactant solution were significantly less resistant to inoculative freezing than those submerged in water. Furthermore, the puparial water content of pupae submerged in surfactant solution was significantly greater than that of puparia from pupae submerged in water. Surfactant may have promoted inoculative freezing by facilitating the spread of water over the surface of and into the puparium, thereby creating bridges between external ice and pupal body fluids. Extracting puparial surface lipids with chloroform/methanol (2 : 1, v:v) decreased the resistance of non-diapausing pupae to inoculative freezing but did not significantly affect that of diapausing pupae. This finding indicates that the puparium of diapausing pupae contains protection against inoculative freezing separate from its surface lipids. This barrier may be important in protecting the freezing-intolerant overwintering pupae against inoculative freezing within their soil hibernaculum.

Roles of aorta, ostia and tracheae in heartbeat and respiratory gas exchange in pupae of Troides rhadamantus Staudinger 1888 and Ornithoptera priamus L. 1758 (Lepidoptera, Papilionidae)

In non-diapausing pupae of the two birdwing butterfly species Troides rhadamantus and Ornithoptera priamus (Lepidoptera, Papilionidae) heart activity and CO 2 release rates were measured simultaneously within the initial half of pupal development. Heartbeat patterns in these pupae consist of three different types of activity: Continuous forward-pulse periods of different duration with a frequency range of about 0.25–0.52 s −1, continuous backward-pulse periods with lower frequencies (0.15–0.29 s −1) and intermittent backward-pulse periods when short series of three to 10 single heartbeats at frequencies of 0.12–0.35 s −1 alternated with heart pauses of 2–10 min. CO 2 release was discontinuous (CFO-type) from about four to 12 days after pupation in Troides rhadamantus and from about four to 18 days in Ornithoptera priamus. Mean CO 2 release rates were very low in both species (10–30 nmol g −1 min −1). After this period, heart pauses occurred more frequently, probably indicating the onset of metamorphosis and the beginning partial histolysis of the heart. Infrared-optical and thermometrical measurements of heartbeat indicated that haemolymph transport within the dorsal vessel in forward direction is more effective than in backward direction. This is deduced from the higher heartbeat frequency and heartbeat amplitude of the forward pulsations. Results from ultrasonic doppler velocimetry suggest that haemolymph flow velocity is highest during the relatively long diastasis of 2–3 s (30–40 mm s −1), while minimum particle speed (about 20 mm s −1) is at the end of systole and the beginning of diastole. This would mean that haemolymph velocity is highest between two consecutive peristaltic waves. In contrast to the haemolymph velocity, the speed of the peristaltic wave measured with the infrared transmission technique was lower (about 8.4–22 mm s −1 in Troides, 10–23 mm s −1 in Ornithoptera) and remained constant during forward pulse periods. During backward beating the speed was lower (8–20 mm s −1 in Troides, 9–17 mm s −1 in Ornithoptera) and decreased during backward pulse periods. During day two to seven in Troides and day three to nine in Ornithoptera, spiracular opening periods coincided with changes in heartbeat direction from backward to forward pulsations. A possible influence is the more efficient convective haemolymph mixing in the haemocoel during forward heartbeat. The mixing allows to bring the haemolymph in close contact with the tracheal system where the discharge of CO 2 takes place. Heartbeat may therefore serve for shortening the diffusion pathways for a rapid transition into the tracheal system during the open period of the spiracles.

Melatonin in the Testis of the Cabbage Armyworm, Mamestra brassicae

N-Acetyl-5-methoxytryptamine (melatonin; MEL) was detected in the testis of non-diapausing pupae and in the testis of post-diapause pharate adults of the cabbage armyworm, Mamestra brassicae, by use of a threedimensional HPLC system with multiple coulometric electrochemical detectors. Intermediates providing evidence of various metabolic pathways were identified, as follows: tryptophan (TRP) → 5-hydroxytryptophan (5-HTP) → 5-hydroxytryptamine (5-HT) → N-methyl-5-hydroxytryptamine (N-MET) ↳5-hydroxyindoleacetic acid (5-HIAA) ↳N-acetyl-5-hydroxytryptamine (N-ACET-5-HT) ↳melatonin (MEL). The possible physiological roles of melatonin in the testis of both diapausing and non-diapausing pupae are discussed.

Upregulation of a 23 kDa small heat shock protein transcript during pupal diapause in the flesh fly, Sarcophaga crassipalpis

A diapause upregulated cDNA clone was isolated from a cDNA library generated from brain mRNA of diapausing Sarcophaga crassipalpis pupae. The clone hybridized to a 1600 bp transcript on a northern blot. The insert is 823 bp in length, has a tentative open reading frame of 615 bp, and codes for a 23 kDa protein. The clone has a high level of identity at the amino acid level with the four small heat shock proteins of Drosophila melanogaster. Northern analysis revealed no detectable expression of the transcript in diapause- or nondiapause-programmed wandering larvae, and only trace expression in nondiapausing pupae. But, the transcript was highly expressed beginning at the onset of diapause and continuing throughout diapause. Expression promptly decreased when diapause was terminated. In nondiapausing individuals the transcript was highly expressed in response to cold shock or heat shock, but temperature stress did not cause greater expression in diapausing pupae. The results imply that expression of this small heat shock protein, a response elicited by temperature stress in nondiapausing individuals, is a normal component of the diapause syndrome. The upregulation of this gene during diapause suggests that it plays an essential role during this overwintering developmental arrest.

Diapause-specific gene expression in pupae of the flesh fly Sarcophaga crassipalpis.

Several cDNAs isolated from brains of diapausing pupae of the flesh fly, Sarcophaga crassipalpis, show expression patterns unique to diapause. To isolate such cDNAs a diapause pupal brain cDNA library was screened by using an elimination hybridization technique, and cDNAs that did not hybridize with cDNA probes constructed from the RNA of nondiapausing pupae were selected for further screening. The 95 clones that did not hybridize in the initial library screen were selected for further characterization. These clones were then screened against diapause and nondiapause pupal poly(A)+ Northern blots. The secondary screen identified 4 diapause-up-regulated clones, 7 diapause-down-regulated clones, 8 clones expressed equally in both diapause and nondiapause, and 75 clones without detectable expression. The diapause-up-regulated and down-regulated clones were further characterized by partial DNA sequencing and identity searches by using GenBank. Identities between our cloned cDNAs and other genes included those linked to cell cycle progression, stress responses, and DNA repair processes. The results suggest that insect diapause is not merely a shutdown of gene expression but is a unique, developmental pathway characterized by the expression of a novel set of genes.

Stage-Dependent Changes of the Prothoracicotropic Hormone (PTTH) Activity of Brain Extracts and of the PTTH Sensitivity of the Prothoracic Glands in the Cabbage Armyworm, Mamestra brassicae, before and during Winter and Aestival Pupal Diapause

The cabbage armyworm, Mamestra brassicae, enters winter and aestival diapause in the pupal stage under 10L-14D and 16L-8D, respectively, at 23°C. It develops without pupal dormancy under 14L-10D at 23°C. In 6th-instar larvae, the hemolymph ecdysteroid titers as well as the activity and sensitivity of the prothoracic glands (PGs) were nearly the same in the diapause- and nondiapause-programmed larvae. In non-diapause pupae (ND-pupae), the ecdysteroid titer peaked in the first half of the pupal stage. By contrast, in winter and aestival diapause pupae (WD- and AD-pupae), the ecdysteroid titer remained low after pupation. The ecdysteroid titer in diapausing AD-pupae was 8- to 10-times higher than that in diapausing WD-pupae. PGs from 5th-instar and 6th-instar larvae, as well as those from pupae, secreted ecdysteroids and showed a far stronger response to big prothoracicotropic hormone (PTTH) than to small PTTH in vitro. PG activity and its sensitivity to big PTTH fluctuated in a stage-dependent manner, and the pattern was nearly the same for 6th-instar larvae committed to develop into ND-, WD- or AD-pupae. When ecdysteroids for larval-pupal metamorphosis appears in the hemolymph of 6th-instar larvae, the activity and sensitivity of the PGs may be suppressed quickly. After pupation, the titer of hemolymph ecdysteroids as well as the activity and sensitivity of the PGs may be related to the strength (or duration) of pupal dormancy which differs between WD-pupae and AD-pupae.

Reproductive, developmental and nutritional biology of the tomato moth,Lacanobia oleracea(Lepidoptera: Noctuidae) reared on artificial diet

AbstractSeveral aspects of the biology of the tomato moth,Lacanobia oleracea(Linnaeus) were investigated when these insects were fed an artificial diet and kept under laboratory conditions. The six larval stadia occupied a total period of 35.6 days at 20°C and 65% r.h. Larvae of each stadium could be separated by their head capsule widths, but not weight ranges, which overlapped. Feeding studies carried out on larvalL. oleracearevealed instar dependent changes in the indices of food consumption and utilization. Larvae increased in weight from the fourth to sixth instar due to a combination of the increased efficiency with which ingested food was converted to body tissue, and increased food consumption by older larvae. Sixth (final) instar larvae exhibited three distinct behavioural phases; during one of these, the feeding phase, larvae consumed a greater amount of diet than either the fourth or fifth instars. The mean pupal period of non-diapausing pupae was 25 days. Pupal diapause was induced when larvae were reared under short day conditions. The male to female sex ratio of adult moths was 1:1.3. Mating generally began during the first scotophase after adult eclosion, and stopped soon after the start of the next photophase. Females usually mated only once, but males were capable of mating at least seven times. Oviposition normally commenced on day 2 of adult life, and the mean total egg production was 1186 eggs/female. Sex pheromone components were not present in solvent extracts of the pheromone glands of 20-day-old female pupae, but low levels were detected in pharate adults. A further increase in pheromone levels in newly-emerged moths was detected and, in isolated virgin moths, pheromone levels increased each day of adult life until day 9. Pheromone levels were significantly lower in two-day-old mated adult females, than in two-day-old virgins.

Cuticular lipid composition of Heliothis virescens and Helicoverpa zea pupae

The cuticular lipids of laboratory-reared tobacco budworms, Heliothis virescens (Fabricius), and corn earworms, Helicoverpa zea (Boddie), pupae were characterized. Quantities of cuticular lipid were greater for pupae in diapause than for non-diapausing pupae. Major cuticular lipids of H. virescens pupae were longchain n-alcohols (42%) and aldehydes (41%) with chain lengths of mainly C 26, C 28 and C 24 in decreasing order of abundance. Lesser amounts (2–5%) of wax esters and hydrocarbons were identified. The wax ester fraction was a complex mixture of C 32-C 52 components composed mainly of C 24-C 28 n-alcohol and saturated, monounsaturated and diunsaturated fatty acid (C 16-C 22) moieties. For H. zea, the major fractions were wax esters (38%), hydrocarbons (19%), n-alcohols (17%), diols (16%) and lesser amounts (2–5%) of aldehydes, acetate esters of alcohols and oxoalcohols. Triacontanyl hexadecanoate comprised 85% of the wax ester fraction. The H. zea alcohols, alcohol ester moieties and aldehydes consisted of a homologous series of C 32-C 52 compounds with the 30 carbon chain length as major constituents. The H. zea diols were C 30-C 36 even-chain n-alcohols with hydroxyl groups on carbon numbers 11, 12, 13, 14 or 15. Mass spectral analysis indicated the presence of unsaturation in the alkyl chain of the major diol components.

Preadult Changes of Ecdysteroid and Juvenile Hormone Titers in Relation to Diapause and Pigmental Variations in two Lepidopteran Species, Cerura vinula and Araschnia levana (Lepidoptera: Notodontidae / Nymphalidae)

Ecdysteroid and juvenile hormone titers were determined through preadult development and set in relation to changes and variations in pigmentation in 2 lepidopteran species : Cerura vinula (Linnaeus 1758) a species with different color patterns in successive larval instars and Araschnia levana (Linnaeus 1758) a species with facultative pupal diapause and adult seasonal diphenism of wing coloration. In C vinula larvae, the black melanin in the color pattern is stepwise reduced at successive molts. The last instar larva [L 5 ] turnes dark red at the wandering stage. Peaks in hemolymph ecdysteroids [ES] correspond to larval molts. The reddening prior to pupation corresponds to a small ES peak followed by the large pupation peak. - The juvenile hormone [JH] titer decreases with every larval instar and altogether becomes lower in every successive instar. At the middle of the L 5 stage, it decreases to an undetectable level and increases again immediately before pupation. The results are in accordance with the assumption that morphological differences between successive larval instars can be controlled by a stepwise decrease of the JH level, while the reddening at the wandering stage is caused by a small ES peak during entire absence of JH. In A levana, short-day larvae develop to diapause pupae and the red spring-morph adults. Long-day larvae become non-diapause pupae and black and white summer-morph adults. In last instar larvae determined for non-diapause an earlier increase of ES than in diapause prepupae corresponds to an earlier pupation. Their adult development is initiated by an ES increase one day after pupation and leads to the black and white summer-morph. By contrast, in diapause pupae, ES remain low during pupal diapause and only after months rises inducing adult development to the red spring-morph. JH titers are high in the middle of the penultimate larval instar and then decrease in all larvae. In the L 5 , JH titer decreases within 2 days to an undetectable level. In diapause larvae, it remains low until pupation. Only in non-diapause larvae, it increases again before pupation. This prepupation JH peak and the earlier ES release are the main differences to diapause development. Presumably, one of them or both may determine non-diapause development and thereby finally the summer-morph adult.

Protandry in Delia antiqua (Diptera: Anthomyiidae)

Sex differences in the duration of egg-larval and pupal stages, and in eclosion curves were examined under 3 different temperature regimes in the laboratory to assess the likelihood of protandry as an adaptive strategy for the onion maggot, Delia antiqua (Meigen). Significant sex differences in development time of larval and pupal stages were evident when eggs were reared at constant (22°C) or alternating (12-22°C) temperature. When male egg development was delayed 24 h by storage at 4°C, followed by 22°C, mean larval development time was significantly greater, but pupal biomass and development time were significantly lower, suggesting variation in relative larval growth rates between the sexes. Under nondiapause conditions, male pupae weighed significantly less than females, but pupal weight was only weakly correlated with larval development. Eclosion of diapause male pupae occurred over a shorter interval (4 d) than from nondiapause pupae (9-12 d), but protandry resulted from disparate pupal development times between the sexes alone, because sexual dimorphism of pupal weight was not apparent. The hypothesis that protandry is an adaptive strategy providing reproductive benefits to either sex is equivocal for D. antiqua because eclosion curves of pupae were normally distributed and mean time lags in eclosion peaks between the sexes were too short (<4 d) to optimize any reproductive benefits from early sexual maturation or mating. The alternative hypothesis is presented that protandry is a nonadaptive outcome of selection for other life history traits that is unbiased by female size dimorphism.

Dynamics of cold hardiness, supercooling and cryoprotectants in diapausing and non-diapausing pupae of the cabbage root fly, Delia radicum L.

The cabbage root fly, Delia radicum L., overwinters as a phanerocephalic pupa (PCP) in a diapause state and its development may be temporarily stopped at the PCP stage even by high summer temperatures. Cold acclimation (5 days at 5 °C) of non-diapausing PCP resulted in an increased cold hardiness (survival of exposure to − 15 °C for 1 or 20 days) and was associated with an increase of trehalose content (from 6.6 to 10.0 μg/mg wet weight). Non-diapausing pupae in a high-temperature quiescence had a higher osmotic pressure of haemolymph (623 mOsm) and a higher glucose content (3.4 μg/mg) in comparison to normal non-diapausing pupae (550 mOsm, 1.4 μg/mg). Diapausing PCP at the start of diapause showed cold hardiness levels equal to non-diapausing PCP. Their cold hardiness rapidly increased during the first 10–30 days of diapause development attaining a maximum between days 30 and 50 followed by an obvious decrease. Osmotic pressure increased from about 550 mOsm at the start of diapause to 860 mOsm by day 80 and then dropped slowly. Trehalose increased from 4.7 to 11.2 μg/mg during the first 80 days and then declined too. Glucose showed transitional increase of concentration during the first 20 days of diapause development followed by an abrupt drop until about day 50 and repeated the increase toward the diapause termination. Myo-inositol level increased slowly during diapause development showing a plateau after 80 days. Supercooling capacity of the PCP stage was similar irrespective of the physiological state or age of the pupa.

Electrical properties of the cerebral prothoracicotropic hormone cells in diapausing and non-diapausing pupae of the tobacco hornworm, Manduca sexta.

Prothoracicotropic hormone (PTTH) is an insect brain neuropeptide that is a primary factor regulating an insect development. Curtailment of its release is thought to be responsible for the pupal diapause of tobacco hornworm, Manduca sexta. The cell synthesizing and secreting the PTTH has been identified as a pair of neurosecretory cells in the pars lateralis on each brain hemisphere. Using intracellular recording techniques, we have demonstrated electrical properties of the PTTH cells in different physiological status, i.e., diapausing and developing pupae. In diapausing pupae, they showed threshold value increasing and input resistance decreasing with the progress of diapausing state, indicating that they were getting unexcitable. Spontaneous action potentials and excitatory postsynaptic potentials (EPSPs) were rarely observed in deeply diapausing state. Non-diapausing PTTH cells were almost silent except day-2, showing rather constant values of electrical properties. On day-2, a significant proportion of the cells had spontaneous action potentials, showing less negative membrane potential values than inactive cells. Exclusively inhibitory postsynaptic potentials (IPSPs) were observed in significant numbers of the cells during the period from day-2 to day-5. On the basis of the results obtained, we proposed a working hypothesis that electrical activities of the PTTH cell may be primarily regulated by its membrane properties which are further modulated by the synaptic mediation.

Development of the overwintering stages of the carrot fly, Psila rosae (Diptera: Psilidae)

AbstractThe overwintering stages of the carrot fly (Psila rosae (Fabricius)) were sampled at Wellesbourne, Warwickshire, during the winters of 1987, 1988, 1989 and 1990. Between 20% and 55% of the insects overwintered as pupae in diapause. The remaining insects stayed in the larval stage during the winter period and gave rise to non-diapausing pupae in the early spring. In May, adults emerged earlier, and over a shorter period, from insects that overwintered as larvae than those that overwintered as pupae. Diapause was most intense during late December and early January. In the laboratory, diapause was completed at temperatures of 5°C and above but was not completed at 0°C. When the overwintering stages were sampled in early February and maintained at constant temperatures in the laboratory, the overwintering pupae required from 114 days at 9°C to 44 days at 21.5°C to give rise to adults. In contrast, the overwintering larvae required only 91 days (9°C) and 27 days (21.5°C), respectively. Populations of overwintering P. rosae collected from Warwickshire and Norfolk had similar requirements for spring emergence but diapause pupae collected in Lancashire emerged over a longer period of time than those collected in Warwickshire. Comparisons with published data indicated that North American and European pupae had different requirements for the completion of diapause. Accumulation of weather data for a P. rosae forecast should start when temperatures are low and any date in early February would be suitable.

Cellular differences in ring glands of flesh fly pupae as a consequence of diapause programming

The ultrastructure of the ring gland (corpus cardiacum (CC), prothoracic gland (PG) and corpus allatum (CA)) was examined in diapausing and nondiapausing flesh fly pupae. The diapause developmental state, which is environmentally regulated and coordinated by the brain-ring gland complex, is associated with differences in the ultrastructure of PG and CA cells but not in the CC. During diapause the PG and CA cells have extensive arrays of rough endoplasmic reticulum and spherical mitochondria. The PG cells also contain lipid droplets surrounded by an electron dense amorphous coat not seen in PG cells from nondiapausing pupae. In nondiapausing pupae, the PG and CA cells contain large amounts of ribosomes throughout the cytoplasm but very little rough endoplasmic reticulum and elongated mitochondria. The fact that ring glands from diapausing pupae readily incorporate 35S-methioninc indicates that the gland is actively synthesizing proteins, thus the contrasts in ring gland ultrastructure are not due to cellular quiescence during diapause but reflect fundamental cellular and physiological differences between the diapause and nondiapause developmental program.

Evidence for a Brain Factor That Stimulates Deposition of Puparial Hydrocarbons in Diapausing Flesh Flies

ABSTRACT Puparia from diapausing pupae of the flesh fly Sarcophaga crassipalpis are lined with twice as much hydrocarbon as puparia from nondiapausing pupae (J. A. Yoder, D. L. Denlinger, M, W. Dennis and P. E. Kolattukudy, unpublished observations). The additional hydrocarbon favors water conservation during diapause by reducing water loss, enhancing water vapor absorptivity and elevating the critical transition temperature (Yoder and Denlinger, 1991). In this study we seek to identify the source of a hormonal modulator that would account for this difference in hydrocarbons. Our results suggest that a factor unique to the brains of diapause-programmed larvae is responsible for increasing the quantity of hydrocarbon deposited on the puparium. Cyclic AMP elicits the same effect, suggesting that the factor is probably a neuropeptide or another modulator that uses cyclic AMP as a second messenger.