Cold shock and cold tolerance in larvae and pupae of the blow fly, Lucilia sericata

Abstract.  Understanding the effects of low winter temperatures on mortality is essential in the development of a full understanding of the long-term population dynamics of any insect. The present study aims to examine the survival of pupae and larvae of the blow fly, Lucilia sericata , at overwintering temperatures. Groups of pupae and diapausing and nondiapausing third-stage larvae of L. sericata are maintained in cooled incubators at either 3 °C and 6 °C. Groups are removed from the incubators at 3–4-day intervals and transferred either to−8 °C or to 25 °C. After 1 h in the freezer, the larvae and pupae exposed to this cold-shock are also transferred to 25 °C. Larvae and pupae are then allowed to continue development and the number of adults emerging from each group is counted. The results demonstrate that survival decreases linearly with the period of exposure at both 3 °C and 6 °C. Mortality is higher at 3 °C than at 6 °C and, in groups that receive the cold shock, cold-shock reduces emergence by over 50%. However, there is no consistent tendency for diapausing larvae to survive prolonged cold or cold shock better than other life-cycle stages. The results suggest that the facultative development of an overwintering diapause stage in L. sericata does not appear to be an adaptation to enhance cold tolerance or resistance to cold shock. It is concluded that the survival of overwintering L. sericata is likely to be relatively less affected by low temperatures than it is by, for example, biotic factors, particularly given the buffered soil environment and short time-scales over which periods of cold act.     

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.     

Infectivity and reproductive potential of the Oswego strain of Heterorhabditis bacteriophora associated with life stages of the clover root curculio,Sitona hispidulus

The infectivity and reproductive potential of the entomopathogenic nematode Heterorhabditis bacteriophora (Oswego strain), at different concentrations, was studied. Seventy to 80.0% mortality to late instar larvae of the clover root curculio, Sitona hispidulus, and 40.0-76.0% mortality to pupae, was observed at concentrations of 15-100 infective juveniles. There were no significant differences in mortality among nematode concentrations. LC(50) levels of 4.0 and 21.4 nematodes were determined for clover root curculio larvae and pupae, respectively. Nematodes did not cause significant mortality to adult or first instar clover root curculio. H. bacteriophora was able to complete its development and reproduce in 74.0-95.0% of clover root curculio late instar larvae and pupae. Reproductive potential in curculio larvae and pupae ranged from 0 to 7040 infective juveniles per host. Larvae exposed to 100 nematodes had a reproductive potential significantly higher than in those larvae exposed to 15 and 50 nematodes. Reproductive potential in pupae decreased with an increased nematode dose, indicating potential crowding effects. Host larval and pupal mass were positively correlated with nematode progeny production.     

Mortality of eggs, larvae and pupae and larval dispersal of the cabbage moth, Mamestra brassicae, in white cabbage in south-eastern Norway

The mortality of eggs, larvae and pupae and larval dispersal of the cabbage moth, Mamestra brassicae (L.) (Lepidoptera: Noctuidae) was investigated in a series of small-scale field experiments in white cabbage, Brassicae oleracea var. capitata (L.), and in the laboratory during 1990–1992 in south–eastern Norway. The highest mortality was found in young larvae and in hibernating pupae. In 1990, larval mortality in the first instar was 80% (range 9–97% for the individual cohorts). Most larvae died within the first 1–3 days after hatching. The dispersal activity during these days was high, and failure to establish feeding sites and predation were probably the main mortality causes. Pupal mortality during winter was 90% on average for 1990–1993 (range 81–100%). The main mortality factor was probably unfavourable weather conditions, and indications of cold stress were found. The impact from parasitoids and diseases was generally low. Trichogramma semblidis (Aurivillius) (Trichogrammatidae) was reared from M. brassicae eggs in very low numbers in 1991. Larval parasitism increased from < 1% in 1990 to almost 24% in 1992, and was almost totally caused by the braconids Microplitis mediator (Haliday) and Aleiodes (Aleiodes) sp. Predation of frozen larvae on the soil surface was 75% on average (range 63–96%) during 1990–1992 in first instar larvae and decreased gradually with larval age. The consumption rates of Philonthus atratus (Gravenhorst) (Staphylinidae) and the carabids Bembidion tetracolum (Say), Pterostichus melanarius (Illiger) and Harpalus rufipes (Degeer) on M. brassicae eggs and larvae were investigated in non-choice experiments in the laboratory. A preliminary survival model based on estimates of the mortality factors identified in this study is presented.     

Regulation of damseifly populations: the effects of larval density on larval urvival, development rate and size in the field

SUMMARY. 1. The popuhttion density of Coenagrion pttella larvae was monitored in five populations, and of Ischntira elegans in two populations, between October 1982 and May 1983.
2. There was no measurable mortality of larvae over winter and no larval growth until April. Larvae in high density populations were smaller than those in low density populations and were more likely to have a semi- voltine life history.
3. The population density of C. ptiella was also monitored (more frequently) in two populations with differenl initial densities between July and November 1983. In the high density population there was a constant rate of larval mortality, while in the low density population there was no detectable larval mortality, indicating that larval mortality may be density dependent. Larvae in the high density population were again smaller, and more likely to be semi-voitine, than those in the low density population.
4. The role of density dependent larval growth, development and mortality in the regulation of damseifly populations is discussed.     

Depth of pupation of the wild olive fruit fly, Bactrocera (Dacus) oleae (Gmel.) (Dipt., Tephritidae), as affected by soil abiotic factors

Abstract:  The influence of four abiotic factors (temperature, soil type, compaction, moisture) on the pupation depth of the wild Bactrocera ( Dacus ) oleae (Gmel.) larvae was studied using soils sampled in the field. Two temperatures (25 and 12°C), three different soil types (alluvial deposits, soil from decaying of limestone, soil from decaying of flysch), two compaction levels (low and high) and two moisture levels (10 and 50% field capacity) were tested in a factorial experiment with a total of 96 experimental units. Five larvae were placed on the soil surface of each test container and when burrowing was completed pupae were retrieved and pupation depth was recorded. The majority of larvae pupated in the top 3 cm and the mean depth of all units was 1.16 cm. The means differed significantly depending on soil type, moisture, the temperature–soil type interaction and the soil type–moisture interaction. Larvae pupated at a greater depth in limestone than in the other two soils. Depths were greater in soils at 50% field capacity than in those at 10% field capacity. In limestone and flysch the depth was greater at 25°C whereas no differences were found in alluvial soil. Different moisture levels had diverse effects in the three soil types; in alluvial soil and in flysch the increased moisture resulted in greater values but in limestone these were slightly lower. These results can be used in developing non chemical control measures and designing efficient sampling techniques for the insect in the ground.     

蔗扁蛾生物学特性及幼虫耐寒性初步研究

蔗扁蛾在江苏地区目前已发现 5个科、8种寄主 ;在扬州 1年发生 4代 ,以幼虫越冬 ;2 5℃下 ,利用寄主发财树饲养该虫 ,卵历期为 5 7d ,幼虫期为 3 4 4d ,蛹历期为 1 4 9d ,整个世代历期为 5 7 6d;不同季节高龄幼虫的过冷却点不同 ,分别为 :冬前 -4 3 6℃ ,隆冬 -6 44℃ ,冬后 -4 89℃ ;不同低温对蔗扁蛾幼虫死亡率影响很大 ,-2℃幼虫 72h后大都死亡 ,-5℃幼虫 48h全部死亡 ;在扬州地区 ,蔗扁蛾在室外安全越冬的可能性较小     

Parasitism and the demography of wheat stem sawfly larvae, <Emphasis Type="Italic">Cephus cinctus</Emphasis>

Previous research has suggested that insect herbivores in protected environments have higher mortalities from biotic factors such as parasitism compared to herbivores in less protected situations, although overall mortalities in protected environments are often lower. However, this has not been examined using life table approaches. In this study, we used demographic methods to characterize the mortality dynamics of parasitism for pre-diapause wheat stem sawfly larvae, Cephus cintus Norton (Hymenoptera: Cephidae), a stem-mining wheat pest. Larvae were intensively sampled from commercial wheat fields at two locations in Montana, USA from 2005 through 2008. The feeding larvae present in the summer succumbed to only two mortality categories: parasitism and unknown factors. In Conrad, a location with relatively high densities of the parasitoids, mortality by parasitism was largely irreplaceable. In contrast, in Amsterdam, where parasitism did not occur to any appreciable degree, overall mortality remained relatively low.     

Effect of temperature on the termination of diapause in the univoltine almond seed wasp Eurytoma amygdali

Diapausing larvae of Eurytoma amygdali Enderlein (Hymenoptera, Eurytomidae) were collected in early August and late September. They were subjected to various photoperiod and temperature regimens for up to 20 weeks, then kept at L16:D8 and 19 °C for another 14 to 26 weeks for diapause to be terminated and pupation to take place. Photoperiod did not affect diapause completion. It was confirmed that the two morphologically distinct diapause stages have different temperature requirements for their completion. The first diapause stage was completed synchronously at temperatures between 16 and 19 °C. A higher temperature of 26 °C delayed diapause development. The second stage required lower temperatures between 4 and 10 °C. Spontaneous termination of diapause was observed at constant 19 °C. When applied to the first diapause stage for 20 weeks, low temperatures made the larvae refractory to subsequent intermediate temperatures. The first stage was thus maintained until a higher temperature of 26 °C made the larvae regain their ability to respond to the intermediate temperatures and complete this stage. Larvae grown in Retsou almonds had a higher diapause intensity than larvae grown in Truoito almonds. The results suggest that, in nature, the high temperatures of late summer and early autumn are likely to maintain the first diapause stage. Subsequently, the less warm temperatures of autumn allow the completion of the first stage by late autumn, and the low temperatures of late autumn and of winter allow the completion of the second diapause stage by mid winter.     

The life histories and population dynamics of two carabid species on a Dutch heathland

Summary We deal with the causes of the synchronously fluctuating numbers of subpopulations of the carabid species Calathus melanocephalus as compared with the asynchronously fluctuating numbers of subpopulations of the carabid Pterostichus versicolor. Both species continuously occupy a large heath area, Dwingelder Veld (1600 ha), in The Netherlands, and are studied there in the same localities with the same methods. Of the adults of C. melanocephalus, 90% do not cover more than 2 ha during the entire reproductive season, while 90% of adults of P. versicolor cover no more than 12 ha. In C. melanocephalus egg production in the field is usually similar to that under optimal feeding conditions in the laboratory, but in P. versicolor egg production seems to be much lower in the field. In the field 70–80% of the eggs most probably are killed by eelworms, followed by more than 90% mortality among the remaining larvae. Comparing mortality of developmental stages in laboratory experiments with that in field experiments in enclosures, it appears that mortality of larvae is not density-dependent, even when density in the experiments is much higher than it ever is in the field. Larval mortality mainly results from the poor ability of the larvae to find prey, even when in field experiments prey density is increased far above natural densities. We discuss why these poor prey-finding abilities are not improved by natural selection. In the spring breeder P. versicolor differences between localities both in abiotic factors, soil moisture and surface temperature, and biotic factors, reactions of prey species to abiotic factors, in spring and summer when the larvae are maturing contribute to the asynchronous fluctuations of numbers between subpopulations. In the autumn breeder C. melanocephalus possible differences in biotic factors between sites are outnumbered by the effects of winters with a higher or lower than normal amount of precipitation respectively. During a wet winter mortality among the larvae is much higher than during a dry winter. As these winter conditions are similar over large areas (many km²) the fluctuations of numbers between subpopulations are synchronous.Communication No. 443 of The Biological Station, Wijster     

Natural thermal stress and heat-shock protein expression in Drosophila larvae and pupae

1. Whether Drosophila larvae and pupae naturally experience temperatures that can cause heat damage or death is poorly understood, but bears directly on numerous investigations of the thermal biology and heat-shock response in Drosophila . Accordingly, the temperatures of necrotic fruit, which Drosophila larvae and pupae inhabit, the temperatures of larvae and pupae outside the laboratory, and the levels of the heat-shock protein hsp 70 expressed by larvae in nature were examined.
2. When necrotic fruit was sunlit, internal temperatures rose to levels that can harm indwelling insects. Fruit size and evaporative water loss affected these temperatures. Temperatures of larvae and pupae in the field commonly exceeded 35 °C, with living larvae recorded at >44°C and pupae at >41°C. Natural mortality was evident, presumably because of heat.
3. In the laboratory, these temperatures kill larvae rapidly, with LT₅₀s (time taken for half the sample to be killed) of 30 min at 39 °C, 15 min at 40 °C and 8·5 min at 41 °C. Gradual transfer from 25°C to these temperatures resulted in no lesser mortality than did direct transfer.
4. Hsp 70 levels in lysates of whole larvae were measured by ELISA (enzyme-link immunosorbent assay) with an hsp 70-specific antibody. For larvae within necrotic apples experimentally transferred from shade to sun and within necrotic fruit in situ , hsp 70 levels equalled or exceeded levels detected in parallel laboratory studies of whole larvae or cells in culture.
5. These data provide an ecological context for studies of thermal stress and the heat-shock response in Drosophila that has heretofore been lacking.     

Seasonal variation in natural mortality factors of Tuta absoluta (Lepidoptera: Gelechiidae) in open‐field tomato cultivation

The seasonal variation in natural mortality of phytophagous insects is determined by the relative importance of biotic and abiotic factors in agroecosystems. Knowledge regarding these factors throughout the year represents a key concern for IPM programmes. Seasonal population fluctuations of tomato pinworm, Tuta absoluta, led to an investigation of its natural mortality factors during the rainy season when the population level is low and during the dry season when population peaks occur. The aim of this study was to verify the seasonal variation in T. absoluta mortality factors in tomato crops. Immature stages of T. absoluta were obtained from laboratory‐rearing in the laboratory. These were taken to the field and monitored over two years. The mortality causes for each stage of insect development from egg to adult were assessed daily. Multiple biotic and abiotic mortality factors affected the immature T. absoluta stages such as rainfall, physiological disturbances, diseases, parasitoids and predators. The key T. absoluta mortality factor during summer–spring was predation. In addition, larvae predation correlated positively with temperature, wind velocity, photoperiod and rainfall. Nevertheless, during winter–fall, the key mortality factor was parasitism. Therefore, the critical stage for mortality was 3rd‐ and 4th‐instar larvae, being more vulnerable to natural control factors. Finally, the results showed the importance of vertical and horizontal action on natural mortality factors.     

Factors Influencing Survival of Ditylenchus dipsaci (Kühn, 1857) in Soil

Ditylenchus dipsaci larvae survived in soil without a host plant for at least 242 days when held at 15 C and 21 C. Larvae held at 15 C remained infective for 212 days. Moisture levels within both clayey and sandy soils did not appreciably affect recovery of larvae. Active nematodes recovered from soil are not necessarily infective. Temperatures of -12, 0 and 4 C had little adverse effect on larvae in infected leaf tissues in soil. Larvae in soil exposed to 0 C for short periods of time were not affected adversely. Recovery of larvae from sandy soil by Baermann funnels was significantly better at 24 C than at 4 C. Differences in recovery from clay soil were not significant at these temperatures.     

Influence of low temperature on pupation induction in Costelytra zealandica (Coleoptera: Scarabaeidae)

Costelytra zealandica (White) lives within the soil throughout its life cycle, except for brief, crepuscular excursions to the surface by the adults to mate and feed. Larvae collected more than 18 weeks before 50% field pupation did not pupate unless they were artificially chilled. Such chilling had no significant effect on the percentage pupation of larvae collected after this time; it is assumed that these larvae had received sufficient chilling from their environment to induce pupation. Larger larvae were more likely to pupate than smaller larvae during the period 13–7 weeks before 50% field pupation. It is postulated that larvae remain quiescent until the soil temperature increases in the spring, and pupate only then. Such a process would explain how the adults of a population emerge at about the same time.     

Delayed density-dependent parasitism of eggs and pupae as a contributor to the cyclic population dynamics of the autumnal moth

Many populations of forest Lepidoptera exhibit 10-year cycles in densities, with impressive outbreaks across large regions. Delayed density-dependent interactions with natural enemies are recognized as key factors driving these cyclic population dynamics, but emphasis has typically been on the larval stages. Eggs, pupae and adults also suffer mortality from predators, parasitoids and pathogens, but little is known about possible density relationships between mortality factors and these non-feeding life stages. In a long-term field study, we experimentally deployed autumnal moth (Epirrita autumnata) eggs and pupae to their natural enemies yearly throughout the 10-year population cycle in northern Norway. The abundance of another geometrid, the winter moth (Operophtera brumata), increased in the study area, permitting comparisons between the two moth species in predation and parasitism. Survival of autumnal moth eggs and pupae was related to the moth abundance in an inverse and delayed manner. Egg and pupal parasitoids dominated as density-dependent mortality factors and predicted the subsequent growth rate of the host population size. In contrast, effects of egg and pupal predators were weakly density dependent, and generally predation remained low. Parasitism rates did not differ between the autumnal and winter moth pupae, whereas predators preferred winter moth pupae over those of the autumnal moth. We conclude that parasitism of the autumnal moth by egg and pupal parasitoids can be related to the changes of the moth density in a delayed density-dependent manner. Furthermore, egg and pupal parasitoids cannot be overlooked as causal factors for the population cycles of forest Lepidoptera in general.     

Pupal colour dimorphism in a desert swallowtail (Lepidoptera: Papilionidae) is driven by changes in food availability,not photoperiod

1. The swallowtail butterfly Battus polydamas archidamas Boisduval, 1936, exhibits polyphenism for pupal coloration (green and brown). It is distributed across arid regions with winter rains and is monophagous on Aristolochia plants, which emerge after the winter rains and dry out the during summer. Thus, day length does not covary positively with host plant productivity. It was hypothesised that pupal colour was driven by food availability, not photoperiod. The benefits of pupal coloration matching the colour of pupation sites in terms of field survival were also investigated to evaluate the adaptive value of pupa colour. 2. Larvae were reared under a factorial array of two photoperiods (LD 10:14 h and LD 14:10 h) and two food availability regimes (leaves ad libitum and available every other day) to assess the frequency of green and brown pupae. Field survival of green and brown pupae was quantified in three commonly used habitats that differ in background coloration (cacti, rocks and shrubs). 3. Food availability determined pupal colour. Larvae in the ad libitum regime resulted mostly in green pupae, while those with restricted food were mostly brown. In contrast, photoperiod did not influence pupal colour. Survival probability of pupae placed on cacti was higher than those placed on rocks and shrubs, and the lowest predation risk across habitats was for green pupae on cacti. 4. Food availability plays a major role in the seasonal polyphenism for pupal colour of specialist butterflies inhabiting arid environments with winter rains.     

Evaluation of green lacewings, Chrysoperla plorabunda (Fitch) (Neurop., Chrysopidae), for augmentative release against Toxoptera citricida (Hom., Aphididae) in citrus

Larvae of the lacewing Chrysoperla plorabunda (Fitch) were evaluated in laboratory and field tests for potential to control the brown citrus aphid, Toxoptera citricida (Kirkaldy) in inundative releases. Larvae surviving to pupation consumed an average of 1676, 1297, 392, 165 and 130 1st–4th instar T. citricida nymphs and apterous adults, respectively, and the mean developmental time was 27.0, 23.0, 16.5, 13.8 and 18.3 days, respectively. The average pupation rate was 37.5%, but only 6.3% of pupae yielded adults. Developmental time and survival to pupation varied with the life stage of aphids fed to larvae, 4th instars yielding the highest survival and shortest developmental time. Development of C. plorabunda was compared on diets of T. citricida and Aphis spiraecola . Only 6.3% of larvae completed development on T. citricida , whereas 37.5% yielded adults on A. spiraecola . Larvae consumed more of the brown T. citricida than the green A. spiraecola in a choice test on a white background, but no preference was evident when aphids were offered on a black background. Two separate field trials were performed in citrus groves with T. citricida infestations in which an average of 275 and 116 C. plorabunda larvae/tree were released, respectively. The rate of T. citricida colony maturation (= survival to alate production) was very low in the first trial, and relatively high in the second trial, but did not differ between control and release trees in either trial. The most apparent cause of aphid colony mortality was predation by the coccinellids Cycloneda sanguinea L. and Harmonia axyridis Pallas. Despite acceptance of T. citricida in the laboratory, very few C. plorabunda larvae were observed feeding on aphid colonies on release trees.     

Anhydrobiosis increases survival of trichostrongyle nematodes

This study demonstrates that infective-stage larvae of 2 trichostrongyle ruminant gastrointestinal nematodes, Haemonchus contortus and Trichostrongylus colubriformis, can enter into anhydrobiotic states when completely desiccated. Larvae of control trichostrongyle species, Heligmosomoides polygyrus and Nippostrongylus brasiliensis, that infect mice were unable to survive desiccation or to enter into anhydrobiosis. Ruminant larvae were able to survive up to 7 desiccation/rehydration cycles, and, during anhydrobiosis, metabolic activity was decreased and survival of the larvae was prolonged both in the laboratory and in the field. Relative humidity had no effect on ruminant larval survival after anhydrobiosis compared with controls. Temperature had a significant effect, 85.8 +/- 2.3% of larvae in anhydrobiosis could survive low temperatures (0 C) that killed all control larvae. Metabolic activity, measured by changes in lipid content and CO2 respiration, was significantly lower in larvae that entered anhydrobiosis compared with controls (P < 0.05). In field experiments using open-meshed chambers under ambient environmental conditions, larvae in anhydrobiosis had significantly higher survival rates in the field compared with controls (P < 0.05) during summer and winter trials. These data suggest that anhydrobiosis in ruminant larvae promotes survival at freezing temperatures, decreases metabolic activity, and prolongs survival under natural field conditions.     

Relating the ontogeny of functional morphology and prey selection with larval mortality in Amphiprion frenatus

Survival during the pelagic larval phase of marine fish is highly variable and is subject to numerous factors. A sharp decline in the number of surviving larvae usually occurs during the transition from endogenous to exogenous feeding known as the first feeding stage in fish larvae. The present study was designed to evaluate the link between functional morphology and prey selection in an attempt to understand how the relationship influences mortality of a marine fish larva, Amphiprion frenatus, through ontogeny. Larvae were reared from hatch to 14 days post hatch (DPH) with one of four diets [rotifers and newly hatched Artemia sp. nauplii (RA); rotifers and wild plankton (RP); rotifers, wild plankton, and newly hatched Artemia nauplii (RPA); wild plankton and newly hatched Artemia nauplii (PA)]. Survival did not differ among diets. Larvae from all diets experienced mass mortality from 1 to 5 DPH followed by decreased mortality from 6 to 14 DPH; individuals fed RA were the exception, exhibiting continuous mortality from 6 to 14 DPH. Larvae consumed progressively larger prey with growth and age, likely due to age related increase in gape. During the mass mortality event, larvae selected small prey items and exhibited few ossified elements. Cessation of mass mortality coincided with consumption of large prey and ossification of key elements of the feeding apparatus. Mass mortality did not appear to be solely influenced by inability to establish first feeding. We hypothesize the interaction of reduced feeding capacities (i.e., complexity of the feeding apparatus) and larval physiology such as digestion or absorption efficiency contributed to the mortality event during the first feeding period. J. Morphol., 2010. © 2010 Wiley‐Liss, Inc.     

Interactions of environmental factors influencing pupal coloration in swallowtail butterfly Papilio xuthus

The swallowtail butterfly Papilio xuthus Linné [Lepidoptera: Papilionidae] exhibits pupal protective color polyphenism. Interactions of various environmental factors on pupal coloration were analyzed in non-diapausing individuals. Under sufficient light (200lux), most pupating larvae became green pupae when the surface of the pupation site was smooth, while they became brown when the surface was rough. Tactile signals are the positive environmental factors causing induction of the brown pupal coloration. In dark boxes, the induction of the brown pupal coloration was easily induced even on a smooth surface, suggesting that light suppresses induction of brown coloration. Different colors of pupation sites did not affect pupal coloration under sufficient light. Environmental factors received during a critical period both before girdling and after girdling affected pupal coloration. When tactile signals received from rough surfaces reach threshold levels during pupation, brown pupal coloration is determined. Larvae reared under a daily periodicity of natural light formed a girdle at midnight, subsequently, the prepupae received strong daylight the following day. Under natural light most larvae produced brown pupae on rough surfaces and green pupae on smooth surfaces.