A trade-off between desiccation resistance and developmental humidity for pupation height in the North Indian seasonal population of Drosophilid-Zaprionus indianus.

Drosophila larvae and pupae are vulnerable to seasonal abiotic stressors such as humidity and temperature. In wild low-humidity habitats, desiccation stress can occur as Drosophila larvae forsake wet food in search of a drier pupation site. Henceforth, the hypothesis that developmental humidity impacts pupation height, affecting larval and pupae water balance and fitness-related traits, was examined. Accordingly, warm-adapted Drosophilid- Zaprionus indianus from two seasons were reared under season-specific simulated conditions, with significantly varying relative humidity (summer RH: 40%; rainy RH: 80%), but nearly identical temperatures. A trade-off between pupation height and developmental humidity was observed. Drier summer conditions lead to pupae wandering farther from drier glass surfaces, resulting in higher pupation height (17.3 cm) while rainy pupae prefer wet food, resulting in lower pupation height (7.12 cm). Additionally, density-dependent pupation height was developmental humidity-specific, with most rainy-season pupae pupated on wetter food, while dry summer pupae pupated on glass surfaces or cotton. Nevertheless, flies from far pupation exhibited greater desiccation resistance, fecundity, and copulation duration than those from near pupation. The cuticular lipid mass of larvae and pupae was higher during far-than-near pupation, indicating decreased water loss rates compared to near-pupation. Finally, pupae eclosion (%) was unaffected by greater humidity (85%) in either season. Still, it considerably decreased at lower humidity (RH: 0% and 38%) for rainy pupae, further supporting the selection of low-humidity desiccation resistance in pupae. In conclusion, low humidity is crucial for survival of pre-adult stages of Zaprionus indianus under desiccation stress and for preference of pupation site.

Adaptive changes in energy reserves and effects of body melanization on thermal tolerance in Drosophila simulans.

Seasonally polyphenic types have been documented in many Drosophilids, which differ significantly during thermal stress. Although Drosophila simulans is a sibling species to Drosophila melanogaster, both thrive in the temperate and tropical climates, but various climatic factors are expected to impact their distribution and abundance. As a result, D. simulans may use phenotypic plasticity to adapt to colder and drier circumstances in temperate zones, although such studies are less known. In the present study, our main aim was to find a link between adaptive plasticity and thermal tolerance in D. simulans. We characterized two morphs in D. simulans flies based on the abdominal melanization collected from the same locality and season, as this trait is highly associated with the larval developmental conditions. Our results suggested that flies reared from dark and light morph showed significant differences in the basal level of proline, carbohydrates (trehalose, glycogen), and lipids (cuticular lipids and total body lipids) within simulated seasons and morph lineages in D. simulans flies. We further showed that D. simulans reared from dark morph are better adapted to cold conditions, whereas light flies are more adapted to warm conditions. The flies, both from light and dark morph lineages, when reared at 15 °C, showed an increase in the level of total body lipids after acclimation at 0 °C but a decrease in the level of proline and carbohydrates (trehalose, glycogen). Heat acclimation increases glycogen levels in the flies from light morph lineage while decreases trehalose and proline.

Plastic changes in cold and drought tolerance of Drosophila nepalensis correlate with sex-specific differences in body melanization, cuticular lipid mass, proline accumulation, and seasonal abundance.

Autumn-collected flies of Himalayan Drosophila nepalensis differ in body color phenotypes (males more melanized relative to females) and in their behavior (males abundant in the open sites vs. shelters-seeking females). In contrast, winter-collected flies of both sexes are equally melanized and abundant in the open sites. We tested developmental and adult plasticity changes in cold or drought tolerance in D. nepalensis flies reared under winter or autumn simulated conditions. In D. nepalensis flies reared at 21 °C, male flies were more cold tolerant (as shown by shorter chill-coma recovery time and lower cold-shock mortality). Further, male flies also exhibited greater drought tolerance (increased levels of desiccation resistance, cuticular lipid mass, melanization, hydration level, and dehydration tolerance) as compared to females. We observed sex-specific differences in the adult plasticity responses due to rapid cold or drought hardening (RCH or RDH); and for the persistence of cold acclimation effects. RCH or RDH-induced changes in the level of proline accumulations are negatively correlated with a decrease in the chill-coma recovery time. Therefore, cold or drought hardening treatments are likely to influence cold tolerance through proline accumulation. Developmental acclimation and adult hardening responses revealed significant interaction effects between sexes and thermal treatments. Thus, sex-specific differences in morphological traits (body melanization and cuticular lipid mass) and physiological traits (adult plasticity changes in cold tolerance and proline accumulation) correlate with behavioral divergence (habitat usage) across sexes.

Heat- and humidity-induced plastic changes in body lipids and starvation resistance in the tropical fly Zaprionus indianus during wet and dry seasons.

Insects in tropical wet or dry seasons are likely to cope with starvation stress through plastic changes (developmental as well as adult acclimation) in energy metabolites. Control and experimental groups of Zaprionus indianus flies were reared under wet or dry conditions, but adults were acclimated at different thermal or humidity conditions. Adult flies of the control group were acclimated at 27°C and low (50%) or high (60%) relative humidity (RH). For experimental groups, adult flies were acclimated at 32°C for 1 to 6 days and under low (40%) or high (70%) RH. For humidity acclimation, adult flies were acclimated at 27°C but under low (40%) or high (70%) RH for 1 to 6 days. Plastic changes in experimental groups as compared with the control group (developmental as well as adult acclimation) revealed significant accumulation of body lipids owing to thermal or humidity acclimation of wet season flies, but low humidity acclimation did not change the level of body lipids in dry season flies. Starvation resistance and body lipids were higher in the males of dry season flies but in the females of wet season flies. Adults acclimated under different thermal or humidity conditions exhibited changes in the rate of utilization of body lipids, carbohydrates and proteins. Adult acclimation of wet or dry season flies revealed plastic changes in mean daily fecundity; and a reduction in fecundity under starvation. Thus, thermal or humidity acclimation of adults revealed plastic changes in energy metabolites to support starvation resistance of wet or dry season flies.

Cross-tolerance effects due to adult heat hardening, desiccation and starvation acclimation of tropical drosophilid-Zaprionus indianus.

Some insect taxa from polar or temperate habitats have shown cross-tolerance for multiple stressors but tropical insect taxa have received less attention. Accordingly, we considered adult flies of a tropical drosophilid-Zaprionus indianus for testing direct as well as cross-tolerance effects of rapid heat hardening (HH), desiccation acclimation (DA) and starvation acclimation (SA) after rearing under warmer and drier season specific simulated conditions. We observed significant direct acclimation effects of HH, DA and SA; and four cases of cross-tolerance effects but no cross-tolerance between desiccation and starvation. Cross-tolerance due to heat hardening on desiccation showed 20% higher effect than its reciprocal effect. There is greater reduction of water loss in heat hardened flies (due to increase in amount of cuticular lipids) as compared with desiccation acclimated flies. However, cross-tolerance effect of SA on heat knockdown was two times higher than its reciprocal. Heat hardened and desiccation acclimated adult flies showed substantial increase in the level of trehalose and proline while body lipids increased due to heat hardening or starvation acclimation. However, maximum increase in energy metabolites was stressor specific i.e. trehalose due to DA; proline due to HH and total body lipids due to SA. Rapid changes in energy metabolites due to heat hardening seem compensatory for possible depletion of trehalose and proline due to desiccation stress; and body lipids due to starvation stress. Thus, observed cross-tolerance effects in Z. indianus represent physiological changes to cope with multiple stressors related to warmer and drier subtropical habitats.

Cold and desiccation stress induced changes in the accumulation and utilization of proline and trehalose in seasonal populations of Drosophila immigrans.

Changes in the levels of energy metabolites can limit survival ability of Drosophila species under stressful conditions but this aspect has received less attention in wild populations collected in different seasons. We tested cold or desiccation triggered changes in the accumulation or utilization of two energy metabolites (trehalose and proline) in Drosophila immigrans flies reared under season specific environmental conditions. Such D.immigrans populations were subjected to different durations of cold (0°C) or desiccation stress (5% RH) or dual stress. We found stress induced effects of cold vs desiccation on the levels of trehalose as well as for proline. Different durations of cold stress led to accumulation of trehalose while desiccation stress durations revealed utilization of trehalose. In contrast, there was accumulation of proline under desiccation and utilization of proline with cold stress. Since accumulation levels were higher than utilization of each energy metabolite, the effects of dual stress showed additive effect. However, there was no utilization of total body lipids under cold or desiccation stress. We observed significant season specific differences in the amount of energy metabolites but the rate of metabolism did not vary across seasons. Stress triggered changes in trehalose and proline suggest possible link between desiccation and cold tolerance. Finally, stress specific (cold or desiccation) compensatory changes in the levels of trehalose and proline suggest possible energetic homeostasis in D.immigrans living under harsh climatic conditions of montane localities.

Effects of saturation deficit on desiccation resistance and water balance in seasonal populations of the tropical drosophilid Zaprionus indianus.

Seasonally varying populations of ectothermic insect taxa from a given locality are expected to cope with simultaneous changes in temperature and humidity through phenotypic plasticity. Accordingly, we investigated the effect of saturation deficit on resistance to desiccation in wild-caught flies from four seasons (spring, summer, rainy and autumn) and corresponding flies reared in the laboratory under season-specific simulated temperature and humidity growth conditions. Flies raised under summer conditions showed approximately three times higher desiccation resistance and increased levels of cuticular lipids compared with flies raised in rainy season conditions. In contrast, intermediate trends were observed for water balance-related traits in flies reared under spring or autumn conditions but trait values overlapped across these two seasons. Furthermore, a threefold difference in saturation deficit (an index of evaporative water loss due to a combined thermal and humidity effect) between summer (27.5 mB) and rainy (8.5 mB) seasons was associated with twofold differences in the rate of water loss. Higher dehydration stress due to a high saturation deficit in summer is compensated by storage of higher levels of energy metabolite (trehalose) and cuticular lipids, and these traits correlated positively with desiccation resistance. In Z. indianus, the observed changes in desiccation-related traits due to plastic effects of simulated growth conditions correspond to similar changes exhibited by seasonal wild-caught flies. Our results show that developmental plastic effects under ecologically relevant thermal and humidity conditions can explain seasonal adaptations for water balance-related traits in Z. indianus and are likely to be associated with its invasive potential.

Divergent strategies for adaptations to stress resistance in two tropical Drosophila species: effects of developmental acclimation in D. bipectinata and the invasive species D. malerkotliana.

Previous studies on two tropical Drosophila species (D. malerkotliana and D. bipectinata) have shown lower resistance to stress-related traits but the rapid colonization of D. malerkotliana in the past few decades is not consistent with its sensitivity to desiccation and cold stress. We tested the hypothesis that developmental acclimation at two growth temperatures (17 and 25°C) can confer adaptations to desiccation and thermal stresses. We found divergence in developmental plastic effects on cuticular traits, i.e. a significant increase of body melanisation (~2-fold) and of cuticular lipid mass (~3-fold) in D. malerkotliana but only 1.5-fold higher cuticular lipid mass in D. bipectinata when grown at 17°C compared with 25°C. A comparison of the water budget of these two species showed significantly higher effects of developmental acclimation on body water content, rate of water loss and dehydration tolerance resulting in higher desiccation resistance in D. malerkotliana than in D. bipectinata. When grown in cooler conditions (17°C), D. malerkotliana had greater resistance to cold as well as desiccation stress. In contrast, heat resistance of D. bipectinata was higher when grown at 25°C. These laboratory observations are supported by data on seasonally varying populations. Furthermore, adult D. malerkotliana acclimated to different stresses showed greater resistance to those stresses than D. bipectinata adults. Thus, significant increase in stress resistance of D. malerkotliana through developmental acclimation may be responsible for its invasion and ecological success on different continents compared with D. bipectinata.

Sex-specific divergence for body size and desiccation-related traits in Drosophila hydei from the western Himalayas.

Sex-specific-differences are a widespread source of genetic variation in various Drosophila species. In the present study, we have examined desiccation survival in males and females of Drosophila hydei from colder and drier montane conditions of the western Himalayas (altitudinal populations; 600-2202 m). In contrast with most other studies in drosophilids, D. hydei males exhibited comparatively higher desiccation resistance despite smaller body size compared to females. Accordingly, we tested the physiological basis of such adaptations in both sexes of D. hydei. Body size traits (wing length, wet weight and dry weight) were ~1.2 fold higher in females than males. However, desiccation resistance was 10 to 13 h higher in males than females. These differences matched enhanced storage of trehalose content (~1.2 fold), higher hemolymph content (~1.2 fold) and enhanced cuticular lipid mass (~1.5 fold) in males than females. Water loss before succumbing to death (dehydration tolerance) was much higher in males (~81%) than females (~64%). A greater loss of hemolymph water until death under desiccation stress was associated with higher desiccation resistance in males. Further, there were lacks of differences in the rate of water loss, rate of trehalose utilization and rate of hemolymph depletion between the sexes in D. hydei. Therefore, sex-specific differences in desiccation resistance of D. hydei were independent of body size as well as the exhaustion of metabolite reserves and rather were caused by the higher dehydration tolerance as well as higher acquisition of hemolymph and trehalose contents.

Seasonal changes in humidity impact drought resistance in tropical Drosophila leontia: testing developmental effects of thermal versus humidity changes.

Drosophila leontia is native to highly humid equatorial tropical habitats but its desiccation sensitivity (~10h) is not consistent with its abundance during the drier autumn season in the subtropical regions. We have tested the effects of developmental acclimation on desiccation resistance and water balance related traits of D. leontia collected during rainy and autumn seasons. The isofemale lines of seasonal populations were reared under ecologically relevant growth temperatures (18 or 26 °C) or humidity conditions (35 or 85% RH) but tested at different times under identical experimental conditions. The larvae as well as flies reared under two thermal conditions (18 or 26 °C) showed no effect on desiccation related traits as well as storage and utilization of energy metabolites. In contrast, for D. leontia reared under low humidity (35% RH), significant changes at larval as well adult stages include increase in the desiccation resistance as well as cuticular lipid quantity, reduced levels of rate of body water loss, higher storage of carbohydrates but lower rate of utilization of carbohydrates as compared with flies reared at high humidity (85% RH). D. leontia has responded to rearing under low humidity conditions by increasing its desiccation resistance but not due to changes in the growth temperatures. These laboratory observations on seasonal populations highlight differences due to rearing conditions but not due to seasons. Further, direct analysis of wild-caught seasonal populations has shown trends similar to developmental acclimation effects. For wild caught flies, there are significant seasonal differences i.e. higher desiccation resistance as well as cuticular lipid quantity but reduced rate of water loss for autumn than rainy season flies. Thus, our laboratory observations are relevant for understanding seasonal adaptations of natural populations of tropical D. leontia to wet-dry conditions in the wild.

Developmental acclimation to low or high humidity conditions affect starvation and heat resistance of Drosophila melanogaster.

Several Drosophila species originating from tropical humid localities are more resistant to starvation and heat stress than populations from high latitudes but mechanistic bases of such physiological changes are largely unknown. In order to test whether humidity levels affect starvation and heat resistance, we investigated developmental acclimation effects of low to high humidity conditions on the storage and utilization of energy resources, body mass, starvation survival, heat knockdown and heat survival of D. melanogaster. Isofemale lines reared under higher humidity (85% RH) stored significantly higher level of lipids and showed greater starvation survival hours but smaller in body size. In contrast, lines reared at low humidity evidenced reduced levels of body lipids and starvation resistance. Starvation resistance and lipid storage level were higher in females than males. However, the rate of utilization of lipids under starvation stress was lower for lines reared under higher humidity. Adult flies of lines reared at 65% RH and acclimated under high or low humidity condition for 200 hours also showed changes in resistance to starvation and heat but such effects were significantly lower as compared with developmental acclimation. Isofemale lines reared under higher humidity showed greater heat knockdown time and heat-shock survival. These laboratory observations on developmental and adult acclimation effects of low versus high humidity conditions have helped in explaining seasonal changes in resistance to starvation and heat of the wild-caught flies of D. melanogaster. Thus, we may suggest that wet versus drier conditions significantly affect starvation and heat resistance of D. melanogaster.

Sex-specific divergence for adaptations to dehydration stress in Drosophila kikkawai.

Several studies on diverse Drosophila species have reported higher desiccation resistance of females, but the physiological basis of such sex-specific differences has received less attention. We tested whether sex-specific differences in cuticular traits (melanic females and non-melanic males) of Drosophila kikkawai correspond with divergence in their water balance mechanisms. Our results are interesting in several respects. First, positive clinal variation in desiccation resistance was correlated with cuticular melanisation in females but with changes in cuticular lipid mass in males, despite a lack of differences between the sexes for the rate of water loss. Second, a comparative analysis of water budget showed that females of the northern population stored more body water as well as hemolymph content and exhibited greater dehydration tolerance than flies from the southern tropics. In contrast, we found no geographical variation in the males for water content and dehydration tolerance. Third, an ~10-fold increase in the rate of water loss after organic solvent treatment of male D. kikkawai suggested a role of cuticular lipids in cuticular transpiration, but had no effect in the females. Fourth, geographical differences in the storage of carbohydrate content (metabolic fuel) were observed in females but not in males. Interestingly, in females, the rate of utilization of carbohydrates did not vary geographically, but males from drier localities showed a 50% reduction compared with wetter localities. Thus, body melanisation, increased body water, hemolymph, carbohydrate content and greater dehydration tolerance confer greater desiccation resistance in females, but a reduced rate of water loss is the only possible mechanism to cope with drought stress in males. Finally, acclimated females showed a significant increase in drought resistance associated with higher trehalose content as well as dehydration tolerance, while males showed no acclimation response. Thus, sex-specific differences in desiccation resistance of D. kikkawai are associated with divergence in some water balance strategies, despite a lack of differences in the rate of water loss between the two sexes.

Invasion and adaptation of a warm-adapted species to montane localities: effect of acclimation potential.

Drosophila ananassae has successfully invaded the cold and dry montane localities of the Western Himalayas in recent years. The ability of this desiccation- and cold-sensitive tropical species to evolve in response to seasonal changes in montane localities is largely unknown. Here, we investigated how this sensitive species adapt to seasonally varying environmental conditions that are lethal to its survival. We observed change in the frequency of dark and light morphs of D. ananassae in five mid-altitude localities during the last decade (2000-2010). We document invasion of D. ananassae to montane localities and increase in frequency of the dark morph. The stress tolerance of morphs (dark and light) remained unaffected of developmental acclimation. However, adult acclimation has shown significant effects on tolerance to various environmental stresses in morphs and effect of this acclimation persist for long durations. Desiccation and cold stress tolerance was increased after adult acclimation for respective stress in the dark morph; while tolerance of the light morph was not affected. Further, heat tolerance of the light morph was increased after adult heat acclimation with no influence on heat tolerance of the dark morph. Our results suggest a possible role of adult acclimation in successful invasion and adaptation of D. ananassae to montane localities. Future experiments should be carried out to determine whether the survival in adverse conditions of low versus high temperature and humidity during seasonal changes is assisted by different acclimation abilities of the two morphs of D. ananassae.

Direct and correlated responses to laboratory selection for body melanisation in Drosophila melanogaster: support for the melanisation-desiccation resistance hypothesis.

For Drosophila melanogaster, cuticular melanisation is a quantitative trait, varying from no melanin to completely dark. Variation in melanisation has been linked with stress resistance, especially desiccation, in D. melanogaster and other species. As melanism has a genetic component, we selected melanic and non-melanic phenotypes of D. melanogaster in order to confirm the association of desiccation resistance and rate of water loss with cuticular melanisation previously reported for this species. A bidirectional selection experiment for dark (D1-D4) and light (L1-L4) body colour in D. melanogaster was conducted for 60 generations. We observed a 1.6-fold increase in abdominal melanisation in selected dark strains and a 14-fold decrease in selected light strains compared with control populations. Desiccation resistance increased significantly in the dark-selected morphs as compared with controls. The observed increase in desiccation resistance appeared as a consequence of a decrease in cuticular permeability. Our results show that traits related to water balance were significantly correlated with abdominal melanisation and were simultaneously selected bidirectionally along with melanisation.

Rapid effects of humidity acclimation on stress resistance in Drosophila melanogaster.

We tested the hypothesis whether developmental acclimation at ecologically relevant humidity regimes (40% and 75% RH) affects desiccation resistance of pre-adults (3rd instar larvae) and adults of Drosophila melanogaster Meigen (Diptera: Drosophilidae). Additionally, we untangled whether drought (40% RH) acclimation affects cold-tolerance in the adults of D. melanogaster. We observed that low humidity (40% RH) acclimated individuals survived significantly longer (1.6-fold) under lethal levels of desiccation stress (0-5% RH) than their counter-replicates acclimated at 75% RH. In contrast to a faster duration of development of 1st and 2nd instar larvae, 3rd instar larvae showed a delayed development at 40% RH as compared to their counterparts grown at 75% RH. Rearing to low humidity conferred an increase in bulk water, hemolymph content and dehydration tolerance, consistent with increase in desiccation resistance for replicates grown at 40% as compared to their counterparts at 75% RH. Further, we found a trade-off between the levels of carbohydrates and body lipid reserves at 40% and 75% RH. Higher levels of carbohydrates sustained longer survival under desiccation stress for individuals developed at 40% RH than their congeners at 75% RH. However, the rate of carbohydrate utilization did not differ between the individuals reared at these contrasting humidity regimes. Interestingly, our results of accelerated failure time (AFT) models showed substantial decreased death rates at a series of low temperatures (0, -2, or -4°C) for replicates acclimated at 40% RH as compared to their counter-parts at 75% RH. Therefore, our findings indicate that development to low humidity conditions constrained on multiple physiological mechanisms of water-balance, and conferred cross-tolerance towards desiccation and cold stress in D. melanogaster. Finally, we suggest that the ability of generalist Drosophila species to tolerate fluctuations in humidity might aid in their existence and abundance under expected changes in moisture level in course of global climate change.

Divergence of desiccation-related traits in two Drosophila species of the takahashii subgroup from the western Himalayas.

Drosophila nepalensis is more abundant under colder and drier montane habitats in the western Himalayas compared with Drosophila takahashii, but the mechanistic basis of such a climatic adaptation is largely unknown. We tested the hypothesis that divergence in the physiological basis of desiccation-related traits is consistent with species-specific adaptations to climatic conditions. Drosophila nepalensis showed approximately twofold higher desiccation resistance, hemolymph content as well as carbohydrate content than D. takahashii despite a modest difference in rate of water loss (0.3% h(-1)). Water loss before succumbing to death (dehydration tolerance) was much higher in D. nepalensis (82.32%) than in D. takahashii (∼50%). A greater loss of hemolymph water under desiccation stress until death is associated with higher desiccation resistance in D. nepalensis. In both species, carbohydrates were utilized under desiccation stress, but a higher level of stored carbohydrates was evident in D. nepalensis. Further, we found increased desiccation resistance in D. nepalensis through acclimation whereas D. takahashii lacked such a response. Thus, species-specific divergence in water-balance-related traits in these species is consistent with their adaptations to wet and dry habitats.

Trade-off of energy metabolites as well as body color phenotypes for starvation and desiccation resistance in montane populations of Drosophila melanogaster.

Storage of energy metabolites has been investigated in different sets of laboratory selected desiccation or starvation resistant lines but few studies have examined such changes in wild-caught populations of Drosophila melanogaster. In contrast to parallel selection of desiccation and starvation tolerance under laboratory selection experiments, opposite clines were observed in wild populations of D. melanogaster. If resistance to desiccation and starvation occurs in opposite directions under field conditions, we may expect a trade-off for energy metabolites but such correlated changes are largely unknown. We tested whether there is a trade-off for storage as well as actual utilization of carbohydrates (trehalose and glycogen), lipids and proteins in D. melanogaster populations collected from different altitudes (512-2500 m). For desiccation resistance, darker flies (>50% body melanization) store more body water content and endure greater loss of water (higher dehydration tolerance) as compared to lighter flies (<30% body melanization). Based on within population analysis, we found evidence for coadapted phenotypes i.e. darker flies store and actually utilize more carbohydrates to confer greater desiccation resistance. In contrast, higher starvation resistance in lighter flies is associated with storage and actual utilization of greater lipid amount. However, darker and lighter flies did not vary in the rate of utilization of carbohydrates under desiccation stress; and of lipids under starvation stress. Thus, we did not find support for the hypothesis that a lower rate of utilization of energy metabolites may contribute to greater stress resistance. Further, for increased desiccation resistance of darker flies, about two-third of total energy budget is provided by carbohydrates. By contrast, lighter flies derive about 66% of total energy content from lipids which sustain higher starvation tolerance. Our results support evolutionary trade-off for storage as well as utilization of energy metabolites for desiccation versus starvation resistance in D. melanogaster.

Divergence of water balance mechanisms in two melanic Drosophila species from the western Himalayas.

Drosophila busckii is more abundant under colder and drier montane habitats in the western Himalayas as compared to Drosophila melanogaster but the mechanistic basis of such climatic adaptations is largely unknown. We tested the hypothesis whether genetic variation or phenotypic plasticity of cuticular traits confer adaptive protection against desiccation stress in two melanic Drosophila species living under drier montane localities. For D. melanogaster, changes in melanisation are known to be associated with reduced water loss but there are no data on D. busckii. We investigated changes in body melanisation, cuticular lipids, desiccation resistance, water loss, extractable hemolymph volume (%), and dehydration tolerance in six sympatric populations of D. busckii and D. melanogaster over an altitudinal range of 640-2236 m. D. busckii is a melanic species but changes in cuticular water loss are negatively correlated with cuticular lipid mass and not with body melanisation. In D. melanogaster, there are no plastic effects (14-28 °C) for cuticular lipid mass but variation in body melanisation is associated with desiccation-related traits. Effects of organic solvents (hexane or chloroform: methanol), developmental plasticity and seasonal variation in cuticular lipids affect body water loss in D. busckii but no such changes occur in D. melanogaster. Thus, sympatric populations of D. busckii and D. melanogaster have evolved different water balance mechanisms under shared environmental conditions in the western Himalayas. Multiple measures of desiccation resistance in these species show clinal variation with altitude, consistent with adaptation to increased desiccation stress.

Developmental and adult acclimation impact cold and drought survival of invasive tropical Drosophila kikkawai.

Narrow distribution patterns of tropical Drosophila species are limited by lower resistance to cold or drought. In the invasive tropical Drosophila kikkawai, we tested whether developmental and adult acclimations at cooler temperatures could enhance its stress resistance level. Adult acclimation of winter collected body color morphs revealed a significant increase in the level of cold resistance. For light morph, its abundance during winter is not consistent with thermal-melanism hypothesis. However, higher cold acclimation capacity, as well as storage of energy metabolites could support its winter survival. In the wild-caught light and intermediate morphs, there is a lack of trade-off between cold and heat resistance but not in the case of dark morph. Developmental plasticity (15°C) resulted in the fivefold increase of cold survival at 0°C; and a twofold increase in desiccation resistance but a modest reduction (∼28-35%) in heat resistance as compared to morph strains reared at 25°C. Drought acclimation changes were significantly higher as compared with cold or heat pretreatment. We observed a trade-off between basal resistance and acclimation capacity for cold, heat, or drought resistance. For homeostatic energy balance, adult acclimation responses (cold versus drought; heat versus drought) caused compensatory plastic changes in the levels of proline or trehalose (shared patterns) but different patterns for total body lipids. In contrast, rapid cold or heat hardening-induced changes in energy metabolites were different as compared to acclimation. The ability of D. kikkawai to significantly increase stress tolerance through plasticity is likely to support its invasion potential.

Sexual dimorphism for water balance mechanisms in montane populations of Drosophila kikkawai.

Conservation of water is critical to the ecological success of Drosophila species living in the drier montane localities of the Western Himalayas. We observed clinal variation in desiccation resistance for both sexes of Drosophila kikkawai from an altitudinal transect (512-2226 m above sea level). Since more than 90 per cent of body water is lost through cuticular transpiration, the target of selection may be cuticular lipids or cuticular melanization. We tested whether melanic females and non-melanic males of D. kikkawai have similar mechanisms of desiccation resistance. There is clinal variation in the amount of cuticular lipids per fly in males, but not in females. By contrast, for females, elevational increase in melanization is positively correlated with desiccation resistance and negatively with cuticular water loss, but there is no variation in the amount of cuticular lipids. Thus, sexual dimorphism for the mechanism of desiccation resistance in D. kikkawai matches the water proofing role of body melanization as well as cuticular lipids.