Upregulation of two actin genes and redistribution of actin during diapause and cold stress in the northern house mosquito, Culex pipiens

Two actin genes cloned from Culex pipiens L. are upregulated during adult diapause. Though actins 1 and 2 were expressed throughout diapause, both genes were most highly expressed early in diapause. These changes in gene expression were accompanied by a conspicuous redistribution of polymerized actin that was most pronounced in the midguts of diapausing mosquitoes that were exposed to low temperature. In nondiapausing mosquitoes reared at 25 degrees C and in diapausing mosquitoes reared at 18 degrees C, polymerized actin was clustered at high concentrations at the intersections of the muscle fibers that form the midgut musculature. When adults 7-10 days post-eclosion were exposed to low temperature (-5 degrees C for 12 h), the polymerized actin was evenly distributed along the muscle fibers in both nondiapausing and diapausing mosquitoes. Exposure of older adults (1 month post-eclosion) to low temperature (-5 degrees C for 12 h) elicited an even greater distribution of polymerized actin, an effect that was especially pronounced in diapausing mosquitoes. These changes in gene expression and actin distribution suggest a role for actins in enhancing survival of diapausing adults during the low temperatures of winter by fortification of the cytoskeleton.     

Physical and physiological factors in diapause initiation of two hyperparasites of the tobacco hornworm,Manduca sexta

Hypopteromalus tabacum diapauses as a last larval instar and Catolaccus aeneoviridis as a non-pharate pupa within cocoons of the primary parasite, Apanteles congregatus. Four factors (photoperiod, temperature, maternal age, and physiological state of the host) were tested for their significance in diapause initiation of both species. A 10L : 14D photoperiod produced a higher incidence of diapausing individuals than a 13L : 11D photoperiod. In the temperature range from 18 to 24°C, the incidence of diapause was inversely related to temperature. The incidence of progeny entering diapause varied with the age of the mother, younger ones producing fewer diapausing offspring. An increase in the number of C. aeneoviridis entering diapause was observed when the host, A. congregatus, was in the diapausing, rather than the nondiapausing condition. Analysis of the data indicated that interactions between these four factors also played an important rôle in diapause initiation of both species.     

Effects of temperature of adults and eggs on the induction of embryonic diapause in the band-legged ground cricket, Dianemobius nigrofasciatus

Abstract.  Eggs laid by adult female Dianemobius nigrofasciatus , reared under long-day (LD 16 : 8 h, 25 °C) or short-day (LD 12 : 12 h, 25 °C) conditions from the nymphal stage, are kept at several constant temperatures. At 22.5–30.0 °C, eggs laid by long-day adults show lower incidences of diapause than those laid by short-day adults. In both eggs laid by adults under long-day conditions and those under short-day conditions, the higher the temperature at which the eggs are kept, the lower the incidence of diapause. When eggs of long-day adults are exposed to a low-temperature pulse (10 °C, 24 h) on the day of deposition (day 0), the incidence of diapause increases. The low-temperature pulse on day 1 does not increase the incidence of diapause. By contrast, when the eggs of short-day adults are exposed to a high-temperature pulse (35 °C, 24 h) on day 0 or day 1, the incidence of diapause decreases. The temperature pulses on day 0 are more effective at diapause prevention. Staining of diapause eggs by the Feulgen–Rossenbeck method shows that the eggs enter diapause at the blastoderm stage, which is on day 1 or day 2 at 25 °C. The exposure of adults to long days and higher temperatures prevents the eggs from entering diapause. In D. nigrofasciatus , embryonic diapause is controlled by maternal effects, adult photoperiod and temperature, and egg temperature before or at diapause.     

Reproductive and developmental consequences of a diapause maternal effect in the flesh fly, Sarcophaga bullata

Abstract Progeny of Sarcophaga bullata produced from mothers with a history of short day will not enter pupal diapause even if they are reared in a strong diapause-inducing environment (LD 12:12 h at 20^oC). Short-day exposure and diapause commitment are normally inseparable, but this maternal effect provides a tool for examining separately the effect of photoperiod and diapause commitment. Duration of the wandering period of the third instar is longer in diapause-destined larvae than in non-diapause-destined larvae, and fecundity of flies that have experienced pupal diapause is lower than in long-day flies that have not been through diapause. The puparia of diapausing pupae contain more hydrocarbons than puparia of nondiapausing pupae, and this contributes to higher rates of net transpiration for the nondiapausing flies. Flies showing the maternal effect (short-day experience but no diapause) show an intermediate response: length of wandering, fecundity rate and quantities of puparial hydrocarbon are between the extremes observed in the other two groups of flies. Thus, the maternal effect switches the developmental programme to nondiapause, but the progeny retain some characteristics of diapause. Evidence from reciprocal crosses indicates that the photoperiodic history of the female, rather than the male, is responsible for the influence on fecundity.     

Cold‐induced expression of diapause in Praon volucre: fitness cost and morpho‐physiological characterization

Cold exposure (2°C for 7 days) in constant darkness at mummy stage induces diapause expression in 9% of the Praon volucre Haliday population. Diapausing parasitoids show a significant delay in emergence time compared with nondiapausing counterparts. A diapause‐mediated polyphenism is observed in mummy colour, with diapausing mummies being clearly darker than nondiapausing ones. The diapause status of dark mummies is confirmed by a significant reduction in metabolic rate. Diapausing parasitoids also display specific morphological characteristics: they are heavier (fresh and dry mass) and accumulate larger fat reserves than nondiapausing counterparts. The diapause status is associated with a fitness cost in terms of adult longevity. There is no evidence of diapause‐related change in supercooling ability.     

Life-History Variation inChrysoperla carnea:Implications for Rearing and Storing a Mexican Population

A population ofChrysoperla carneafrom San Pedro, Coahuila, Mexico, showed phenological responses that differ from those of previously studied North American biotypes. Short daylengths that induce diapause in other biotypes resulted in low incidences of diapause in the San Pedro population. Diapause induction required a decrease in daylength (from long to short days). Constant long days produced variable reproductive responses, whereas an increase in daylength (from short to long day) resulted in high incidences of reproduction. Providing prey to adults that had been reared under short days consistently increased the incidences of reproduction. Storage of either diapausing or nondiapausing adults at low temperature (5°C) for 13 weeks yielded better survivorship and reproductive performance than in unstored (24°C) controls. For efficient production, the San Pedro population ofC. carneamay be reared either with an increase in daylength or with the regular intervention of a storage period. To induce diapause in individuals destined for storage, larvae may be reared under long-day photoperiod and the prepupae transferred to short days.     

Cold hardiness and sugar content in photo‐insensitive individuals of the cotton bollworm Helicoverpa armigera

Abstract A proportion of Helicoverpa armigera collected from fields in Okayama Prefecture (Western Japan; 34.6°N, 134.1°E) does not enter diapause when reared under a short days at 20 °C during the larval stages. However, diapause in such photo‐insensitive individuals can be induced when they are reared at moderately low temperatures, such as 15 °C, regardless of photoperiod. To determine whether such photo‐insensitive individuals can survive overwintering in fields, the present study compares the cold hardiness and sugar content between nondiapausing and diapausing pupae of photo‐insensitive individuals selected over several generations at 20 °C under a short day photoperiod (LD 10 : 14 h). Diapausing and nondiapausing pupae are obtained under the short days by rearing at 15 and 20 °C, respectively, during larval and pupal stages. These pupae are stepwise acclimated at a reduction of 5 °C every 5 days to 0 °C. Maximum survival periods of nondiapausing and diapausing pupae at 0 °C are approximately 30 and 90 days, respectively. Trehalose content in diapausing pupae increases, reaches a maximum level (1.95 mg 100 mg^?1 in males and 2.1 mg 100 mg^?1 in females) 28 days after exposure to 0 °C and then decreases. On the other hand, glucose content in diapausing pupae increases (maximum level: 0.32 mg 100 mg^?1 in males and 0.21 mg 100 mg^?1 in females) with decreasing trehalose content 42 days after exposure to 0°C. The decrease in trehalose content and the increase in glucose content may be linked to termination of diapause in H. armigera. These results suggest that, in Japan, the photo‐insensitive individuals can only survive in the mild winters of southern regions, and not in the severe winters of northern regions.     

Induction of reproductive diapause in Habrobracon hebetor (Hymenoptera: Braconidae) when reared at different photoperiods at low temperatures

Development of the parasitoid Habrobracon hebetor (Say) (Hymenoptera: Braconidae) at low temperatures was determined to identify rearing conditions that might result in adults that were in reproductive diapause. Diapausing adults would be expected to survive cold storage longer than nondiapausing adults for use in biological control programs. Only a few eggs were found in the ovaries when H. hebetor females were reared during the immature stages at 17.5 and 20°C with a 16-h photoperiod, and the ovaries were poorly developed and contained no eggs when females were reared with a 10-h photoperiod in these low temperatures. Rearing H. hebetor at 17.5 and 20°C did not result in diapause of immature stages, but did appear to result in possible adult reproductive diapause when the immature stages were reared with a 10-h photoperiod. Females reared during the immature stages at 17.5°C with a10-h photoperiod lived longer and took longer to lay their first eggs and to lay 50% of their eggs than those females reared at 17.5°C with a16-h photoperiod. Females reared during the immature stages at 20°C with a10-h photoperiod took longer to lay their first eggs and to lay 50% of their eggs, and they had a lower respiration rate, than those females reared at 20°C with a16-h photoperiod. Females that were reared in conditions that appeared to induce reproductive diapause resumed oviposition and their respiration rate increased soon after being transferred to a higher temperature (27.5°C). Thus, females reared at a 10-h photoperiod at 17.5 and 20°C appear to enter reproductive diapause.     

Control of diapause and supercooling by the retrocerebral complex inPyrrhocoris apterus

Effects of external (photoperiod, temperature) and internal (retrocerebral complex, diapause condition) factors on supercooling were studied inPyrrhocoris apterus (L.) (Heteroptera: Pyrrhocoridae) adults. An increase in supercooling capacity is associated with the induction of diapause by short-day photoperiod or cardiacallatectomy of long-day non-diapause insects in spite of a high temperature of 26°C. The induction of diapause is a prerequisite for a further increase in supercooling capacity by cold acclimation. Post-diapause adults show low values of the supercooling point also in late January or early February, although their developmental potential is fully restored. Evidently the ability to supercool associated with diapause induction is maintained by a low ambient temperature in spite of diapause termination. The supercooling point increases rapidly when these adults are transferred to a high temperature of 26°C and cold re-acclimation is then no longer possible. The inhibition of morphogenesis (maturation of gonads) by cardiacallatectomy does not prevent the supercooling point from increasing in post-diapause adults at the high temperature.     

Diapause and cold tolerance in Asian species of the parasitoid Leptopilina (Hymenoptera: Figitidae)

Diapause and cold tolerance are essential for temperate insects to pass the winter, with the mechanisms controlling these two traits varying considerably among insects. In the present study, diapause and cold tolerance are compared among three Leptopilina species: Leptopilina japonica Novkovi? & Kimura, Leptopilina victoriae Nordlander and Leptopilina ryukyuensis Novkovi? & Kimura, all larval parasitoids of frugivorous drosophilid flies, with the aim of understanding their climatic adaptations. The first species is divided into the temperate (Leptopilina japonica japonica) and subtropical subspecies (Leptopilina japonica formosana), and the latter two species are distributed in the tropical and subtropical regions. The temperate subspecies of L. japonica enters prepupal diapause at low temperatures (15 or 18 °C), irrespective of photoperiod, and some individuals enter diapause when exposed to 0 °C for 1 or 2 day(s) or when placed at low humidity. Leptopilina victoriae also shows signs of diapause initiation at 15 °C, although L. ryukyuensis and L. j. formosana from the subtropical regions do not. Preimaginal viability at low temperature (13, 14 or 15 °C) is usually lower in L. victoriae from the tropical regions compared with L. japonica or L. ryukyuensis from the temperate or subtropical regions. Diapausing prepupae of the temperate subspecies appear to be cold tolerant. However, the cold tolerance of nondiapausing prepupae, pupae and adult females varies little among the tropical, subtropical and temperate species or subspecies, and adult males of the temperate subspecies of L. japonica are less cold tolerant than those of the tropical or subtropical species or subspecies. Cold tolerance may be unnecessary, except for diapausing individuals of the temperate species, because nondiapausing individuals appear in warmer seasons.     

Haemolymph ecdysteroid titres in diapause and nondiapause larvae of the European corn borer, Ostrinia nubilalis

The ecdysteroid titres of last-instar prediapausing, diapausing and nondiapausing larvae of Ostrinia nubilalis were determined by radioimmunoassay. In the nondiapause larvae a major peak of ecdysteroid activity preceded pupation by 24 h and continued through the pupal ecdysis. This peak was correlated with head and thorax critical periods as well as with changes in behaviour and physiology marking the transition from feeding larva to prepupa. Nondiapause larvae also displayed a rise in ecdysteroid titre during the feeding phase of development. This rise was approx one tenth that of the major peak and lasted 32 h. It was not correlated with any overt changes in larval physiology or behaviour. The diapause ecdysteroid profile was distinctive in that the levels measured were all lower than the lowest of the nondiapause curve. No peaks were observed in the diapause titres. Prepupal changes such as spinning and the cessation of feeding were not correlated with any increase in ecdysteroid levels. During diapause termination, under a long-day photoperiod, no increases in ecdysteroid titre were observed for the first 10 days. After 12 days individuals began to show ecdysteroids above the diapause levels. Pupation started after 16 long days.     

A maternal effect that eliminates pupal diapause in progeny of the flesh fly, Sarcophaga bullata

Flesh flies that have experienced pupal diapause produce progeny that will not enter diapause even when reared in a strongly diapause-inducing environment. The effect is determined, not by diapause itself, but by the short days previously received by the larvae during the programming of pupal diapause. Reciprocal cross matings indicate that the effect is transmitted solely by the female parent. Though the embryos develop within the uterus of the female, the maternal effect is transmitted prior to the onset of embryogenesis, probably during oögenesis. Only by rearing a generation in long-day (nondiapausing) conditions can the capacity for pupal diapause be restored in the progeny. The effect is likely to provide an adaptive mechanism for preventing an untimely diapause response among the progeny of overwintering females that emerge early in the spring.     

A nondiapausing variant of the flesh fly, Sarcophaga bullata, that shows arrhythmic adult eclosion and elevated expression of two circadian clock genes, period and timeless

We describe a variant of the flesh fly, Sarcophaga bullata, which fails to enter pupal diapause in response to short daylength and low temperatures. This fly also has an arrhythmic adult eclosion pattern: rather than eclosing in early photophase, the variant ecloses arrhythmically throughout the photophase and scotophase. The loss of both diapause (photoperiodic response) and the gating of adult eclosion (presumably a circadian response) suggests that the same clock system is involved in these two responses. An examination of the expression patterns of the clock genes period and timeless demonstrates that both genes are present in the nondiapausing variant, but surprisingly, both genes are expressed at higher levels. This abnormality we observe, possibly the consequence of an upstream clock gene malfunction or a malfunction of the autoregulatory loop, results in disruption of a component of the clock system that is apparently needed for both photoperiodism and circadian rhythmicity.     

Regulation of heat shock proteins in the apple maggot Rhagoletis pomonella during hot summer days and overwintering diapause

Abstract Developing larvae of the apple maggot Rhagoletis pomonella are frequently exposed to summertime apple temperatures that exceed 40 °C and, during their overwintering diapause, pupae are exposed to sub‐zero soil temperatures for prolonged periods. To investigate the potential involvement of heat shock proteins (Hsps) in response to these environmental extremes, the genes encoding Hsp70 and Hsp90 in R. pomonella are cloned and expression monitored during larval feeding within the apple and during overwintering pupal diapause. Larvae reared in the laboratory at constant temperatures of 25, 28 or 35 °C express Hsp90 but very little Hsp70. Larvae do not survive rearing at 40 °C. The temperature cycles to which larvae were exposed inside apples in the field, ranging 16–46.9 °C over a 24‐h period, elicit strong Hsp70 and Hsp90 expression, which begins at mid‐day and reaches a peak in late afternoon, coinciding with peak air and apple temperatures. Heat shock proteins are also expressed strongly by pupae during their overwintering diapause. Hsp70 is not expressed in nondiapausing pupae but is highly expressed throughout diapause. Hsp90 is constitutively expressed in both diapausing and nondiapausing pupae. Rhagoletis pomonella thus strongly expresses its Hsps during pupal diapause, presumably as a protection against low temperature injury, and during larval development to cope with natural temperature cycles prevailing in late summer.     

Age‐dependent changes in tolerance to cold and accumulation of cryoprotectants in overwintering and non‐overwintering larvae of European corn borer Ostrinia nubilalis

Abstract The age‐dependent cold hardiness profile of Ostrinia nubilalis is compared between nondiapausing and diapausing larvae, as well as with field‐collected larvae. The results suggest that both cold tolerance and accumulation of cryoprotectants depends upon the age of O. nubilalis larva. Late fifth‐instar nondiapausing larvae are more cold tolerant than younger fifth‐instars because they show enhanced ability to withstand sub‐zero temperatures. No appreciable difference is observed between the experimental groups of diapausing larvae as far as their supercooling ability and tolerance at sub‐zero temperatures above the supercooling point. In general, both field‐collected and diapausing larvae are more cold tolerant than nondiapausing larvae, indicating a direct link between diapause and cold hardiness. The age of diapausing larvae affects the ability to accumulate glycerol. Glycerol levels of 45‐day‐old diapausing larvae are significantly higher (2.7‐fold) compared with 90‐day‐old diapausing larvae. Moreover, diapausing larvae display a five‐ to 13‐fold higher glycerol content compared with nondiapausing larvae. There is a trend for an age‐dependent cold hardiness profile in O. nubilalis and further tests that could demonstrate a causal relationship between age and cold tolerance are needed.