Simulating natural light and temperature cycles in the laboratory reveals differential effects on activity/rest rhythm of four Drosophilids

Recent studies under semi-natural conditions have revealed various unique features of activity/rest rhythms in Drosophilids that differ from those under standard laboratory conditions. An additional afternoon peak (A-peak) has been reported for Drosophila melanogaster and another species D. malerkotliana while D. ananassae exhibited mostly unimodal diurnal activity. To tease apart the role of light and temperature in mediating these species-specific behaviours of four Drosophilid species D. melanogaster, D. malerkotliana, D. ananassae, and Zaprionus indianus we simulated gradual natural light and/or temperature cycles conditions in laboratory. The pattern observed under semi-natural conditions could be reproduced in the laboratory for all the species under a variety of simulated conditions. D. melanogaster and D. malerkotliana showed similar patterns where as D. ananassae consistently exhibited predominant morning activity under almost all regimes. Z. indianus showed clearly rhythmic activity mostly when temperature cycles were provided. We find that gradually changing light intensities reaching a sufficiently high peak value can elicit A-peak in D. melanogaster, D. malerkotliana, and D. ananassae even at mild ambient temperature. Furthermore, we show that high mid-day temperature could induce A-peak in all species even under constant light conditions suggesting that this A-peak is likely to be a stress response.     

Natural Conditions Override Differences in Emergence Rhythm among Closely Related Drosophilids

Previous studies on adult emergence rhythm of Drosophila melanogaster (DM) done under semi-natural conditions have shown that emergence is correlated to daily changes in temperature, humidity and light at dawn. Recently we showed that under laboratory conditions D. ananassae (DA), a closely related species of DM exhibits patterns in its activity/rest rhythm distinct from the latter. Here, we report the results of a study aimed at examining whether this difference in activity/rest rhythm among species extends to other circadian behaviours such as the adult emergence rhythm under a more natural environment with multiple cyclic time cues. We monitored the adult emergence rhythm of recently wild-caught DM and DA populations in parallel with those of a related species D. malerkotliana (DK), both in the laboratory and under semi-natural conditions. We find that although DM, DK and DA showed marked difference from one another under laboratory conditions, such differences were not detectable in the emergence behaviour of these three species under semi-natural conditions, and that they respond very similarly to seasonal changes in the environment. The results suggest that seasonal changes in temperature and humidity contribute largely to the variation in adult emergence waveform in terms of gate width, phase and amplitude of the peak and day-to-day variance in the timing of the emergence peak. In all three species, seasons with cooler and wetter conditions make the rhythm less tightly gated, with low amplitude peak and high day-to-day variation in timing of the peak of emergence. We show that in nature the emergence rhythm of DM, DK and DA is strongly influenced by environmental factors such that in a given season all of them exhibit similar time course and waveform and that with the changing season, they all modify their emergence patterns in a similar manner.     

Egg-laying rhythm in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Extensive research has been carried out to understand how circadian clocks regulate various physiological processes in organisms. The discovery of clock genes and the molecular clockwork has helped researchers to understand the possible role of these genes in regulating various metabolic processes. In Drosophila melanogaster, many studies have shown that the basic architecture of circadian clocks is multi-oscillatory. In nature, different neuronal subgroups in the brain of D. melanogaster have been demonstrated to control different circadian behavioural rhythms or different aspects of the same circadian rhythm. Among the circadian phenomena that have been studied so far in Drosophila, the egg-laying rhythm is unique, and relatively less explored. Unlike most other circadian rhythms, the egg-laying rhythm is rhythmic under constant light conditions, and the endogenous or free-running period of the rhythm is greater than those of most other rhythms. Although the clock genes and neurons required for the persistence of adult emergence and activity/rest rhythms have been studied extensively, those underlying the circadian egg-laying rhythm still remain largely unknown. In this review, we discuss our current understanding of the circadian egg-laying rhythm in D. melanogaster, and the possible molecular and physiological mechanisms that control the rhythmic output of the egg-laying process.     

Cyclic Presence and Absence of Conspecifics Alters Circadian Clock Phase But Does Not Entrain the Locomotor Activity Rhythm of the Fruit Fly Drosophila melanogaster

Circadian clocks use a wide range of environmental cues, including cycles of light, temperature, food, and social interactions, to fine-tune rhythms in behavior and physiology. Although social cues have been shown to influence circadian clocks of a variety of organisms including the fruit fly Drosophila melanogaster, their mechanism of action is still unclear. Here, the authors report the results of their study aimed at investigating if daily cycles of presence and absence (PA) of conspecific male visitors are able to entrain the circadian locomotor activity rhythm of male hosts living under constant darkness (DD). The results suggest that PA cycles may not be able to entrain circadian locomotor activity rhythms of Drosophila. The outcome does not change when male hosts are presented with female visitors, suggesting that PA cycles of either sex may not be effective in bringing about stable entrainment of circadian clocks in D. melanogaster. However, in hosts whose clock phase has already been set by light/dark (LD) cycles, daily PA cycles of visitors can cause measurable change in the phase of subsequent free-running rhythms, provided that their circadian clocks are labile. Thus, the findings of this study suggest that D. melanogaster males may not be using cyclic social cues as their primary zeitgeber (time cue) for entrainment of circadian clocks, although social cues are capable of altering the phase of their circadian rhythms. (Author correspondence: vsharma@jncasr.ac.in, vksharmas@gmail.com)     

Circadian Dysfunction Reduces Lifespan in Drosophila melanogaster

Circadian clocks regulate physiological and behavioral processes in a wide variety of organisms, and any malfunction in these clocks can cause significant health problems. In this paper, we report the results of our study on the physiological consequences of circadian dysfunction (malfunctioning of circadian clocks) in two wild‐type populations of fruit flies (Drosophila melanogaster). We assayed locomotor activity behavior and lifespan among adult flies kept under constant dark (DD) conditions of the laboratory, wherein they were categorized as rhythmic if their activity/rest schedules followed circadian (approximately 24 h) patterns, and as arrhythmic if their activity/rest schedules did not display any pattern. The rhythmic flies from both populations lived significantly longer than the arrhythmic ones. Based on these results, we conclude that circadian dysfunction is deleterious, and proper functioning of circadian clocks is essential for the physiological well being of D. melanogaster.     

Rhythmic egg-laying behaviour in virgin females of fruit flies Drosophila melanogaster

Fruit fly Drosophila melanogaster females display rhythmic egg-laying under 12:12?h light/dark (LD) cycles which persists with near 24?h periodicity under constant darkness (DD). We have shown previously that persistence of this rhythm does not require the neurons expressing pigment dispersing factor (PDF), thought to be the canonical circadian pacemakers, and proposed that it could be controlled by peripheral clocks or regulated/triggered by the act of mating. We assayed egg-laying behaviour of wild-type Canton S (CS) females under LD, DD and constant light (LL) conditions in three different physiological states; as virgins, as females allowed to mate with males for 1?day and as females allowed to mate for the entire duration of the assay. Here, we report the presence of a circadian rhythm in egg-laying in virgin D. melanogaster females. We also found that egg-laying behaviour of 70 and 90% females from all the three male presence/absence protocols follows circadian rhythmicity under DD and LL, with periods ranging between 18 and 30?h. The egg-laying rhythm of all virgin females synchronized to LD cycles with a peak occurring soon after lights-off. The rhythm in virgins was remarkably robust with maximum number of eggs deposited immediately after lights-off in contrast to mated females which show higher egg-laying during the day. These results suggest that the egg-laying rhythm of D. melanogaster is endogenously driven and is neither regulated nor triggered by the act of mating; instead, the presence of males results in reduction in entrainment to LD cycles.     

Temperature can entrain egg laying rhythm of Drosophila but may not be a stronger zeitgeber than light

In Drosophila multiple circadian oscillators and behavioral rhythms are known to exist, yet most previous studies that attempted to understand circadian entrainment have focused on the activity/rest rhythm and to some extent the adult emergence rhythm. Egg laying behavior of Drosophila females also follows circadian rhythmicity and has been seen to deviate substantially from the better characterized rhythms in a few aspects. Here we report the findings of our study aimed at evaluating how circadian egg laying rhythm in fruit flies Drosophila melanogaster entrains to time cues provided by light and temperature. Previous studies have shown that activity/rest rhythm of flies entrains readily to light/dark (LD) and temperature cycles (TC). Our present study revealed that egg laying rhythm of a greater percentage of females entrains to TC compared to LD cycles. Therefore, in the specific context of our study this result can be taken to suggest that egg laying clocks of D. melanogaster entrains to TC more readily than LD cycles. However, when TC were presented along with out-of-phase LD cycles, the rhythm displayed two peaks, one occurring close to lights-off and the other near the onset of low temperature phase, indicating that upon entrainment by TC, LD cycles may be able to exert a greater influence on the phase of the rhythm. These results suggest that temperature and light associatively entrain circadian egg laying clocks of Drosophila.     

Fluorescence patterns of heterochromatin in mitotic and polytene chromosomes in seven members of three sub-groups of the melanogaster species group of Drosophila

A comparative study of fluorescence patterns of heterochromatin in mitotic and polytene chromosomes of seven species belonging to 3 subgroups melanogaster sub-group: D. melanogaster and D. simulans; montium sub-group: D. kikkawai and D. jambulina; ananassae sub-group: D. ananassae, D. malerkotliana and D. bipectinata) of the melanogaster species group of Drosophila (Sophophora) has been made. Hoechst 33258 (H) fluorescence patterns of mitotic chromosomes reveal differences correlated to the taxonomic groupings of these species. The melanogaster sub-group species have H-bright regions on heterochromatin of all chromosomes; the montium subgroup species have H-bright regions mainly on the 4th and Y-chromosomes; in the ananassae sub-group, while D. ananassae chromosomes do not show any H-bright regions, D. malerkotliana and D. bipectinata have small H-bright segments only on their 4th chromosomes. The H-and quinacrine mustard (QM) fluorescence patterns of larval salivary gland polytene chromocentre in these species, however, do not show the same taxonomic correlation. While D. ananassae and D. kikkawai polytene nuclei lack any H-or QMbright region in the chromocentre, the remaining species have prominent H-and/or QM-bright region(s). In D. jambulina, the QM-bright regions are generally bigger than H-bright regions, while in D. malerkotliana and D. bipectinata the situation is reversed. Actinomycin D counterstaining prior to H-staining of polytene preparations of each species confirms that the H-bright region/s in the chromocentre are composed of A-T rich sequences. In vivo labelling of salivary gland polytene nuclei with 5-bromodeoxyuridine for 24 to 48 h and subsequent H-staining reveals that in all the species, the H-bright regions do not replicate in 3rd instar stage and presumably represent the non-replicating alpha heterochromatin. Significantly, in all the species (excepting D. kikkawai and D. ananassae), the size, location and the number of H-and/or QM-bright regions were seen to vary in different polytene nuclei in the same gland. It seems that the organization and the extent of under-replication of alpha heterochromatin varies in different polytene nuclei. Present studies also show that even closely related species differ in the content and organization of H-bright heterochromatin. The 81 F band at the base of 3 R in D. melanogaster, but not in D. simulans, appears to contain non-replicating H-bright sequences in addition to replicating chromatin.     

Entrainment of Eclosion Rhythm in Drosophila melanogaster Populations Reared for More Than 700 Generations in Constant Light Environment

In this paper, we report the results of our extensive study on eclosion rhythm of four independent populations of Drosophila melanogaster that were reared in constant light (LL) environment of the laboratory for more than 700 generations. The eclosion rhythm of these flies was assayed under LL, constant darkness (DD) and three periodic light‐dark (LD) cycles (T20, T24, and T28). The percentage of vials from each population that exhibited circadian rhythm of eclosion in DD and in LL (intensity of approximately 100 lux) was about 90% and 18%, respectively. The mean free‐running period (τ) of eclosion rhythm in DD was 22.85 ± 0.87 h (mean ± SD). Eclosion rhythm of these flies entrained to all the three periodic LD cycles, and the phase relationship (ψ) of the peak of eclosion with respect to “lights‐on” of the LD cycle was significantly different in the three periodic light regimes (T20, T24, and T28). The results thus clearly demonstrate that these flies have preserved the ability to exhibit circadian rhythm of eclosion and the ability to entrain to a wide range of periodic LD cycles even after being in an aperiodic environment for several hundred generations. This suggests that circadian clocks may have intrinsic adaptive value accrued perhaps from coordinating internal metabolic cycles in constant conditions, and that the entrainment mechanisms of circadian clocks are possibly an integral part of the clockwork.     

Correlated changes in circadian clocks in response to selection for faster pre-adult development in fruit flies Drosophila melanogaster

Although, circadian clocks are believed to be involved in the regulation of life-history traits such as pre-adult development time and lifespan in fruit flies Drosophila melanogaster, there is very little unequivocal evidence either to support or refute this. Here we report the results of a long-term study aimed at examining the role of circadian clocks in the temporal regulation of pre-adult development in D. melanogaster. We employed laboratory selection protocol for faster pre-adult development on four large, outbred, random mating populations of Drosophila. We assayed pre-adult development time and circadian period of locomotor activity rhythm of these flies at regular intervals of 5–10 generations. After 50 generations of selection, the overall egg-to-adult duration in the selected stocks was reduced by ~29 h (~12.5 %) relative to controls, with the selected populations showing a concurrent reduction in time taken to hatching, pupation and wing pigmentation, by ~2, ~16, and ~25.2 h, respectively. Furthermore, selected populations showed a concomitant reduction in the circadian period of locomotor activity rhythm, implying that circadian clocks and development time are correlated. Thus, our study provides the first ever unequivocal evidence for the evolution of circadian clocks as a correlated response to selection for faster pre-adult development, suggesting that circadian clocks and development are linked in fruit flies D. melanogaster.     

Robustness of Circadian Timing Systems Evolves in the Fruit Fly Drosophila melanogaster as a Correlated Response to Selection for Adult Emergence in a Narrow Window of Time

Robustness is a fundamental property of biological timing systems that is likely to ensure their efficient functioning under a wide range of environmental conditions. Here we report the findings of our study aimed at examining robustness of circadian clocks in fruit fly Drosophila melanogaster populations selected to emerge as adults within a narrow window of time. Previously, we have reported that such flies display enhanced synchrony, accuracy, and precision in their adult emergence and activity/rest rhythms. Since it is expected that accurate and precise circadian clocks may confer enhanced stability in circadian time-keeping, we decided to examine robustness in circadian rhythms of flies from the selected populations by subjecting them to a variety of environmental conditions comprising of a range of photoperiods, light intensities, ambient temperatures, and constant darkness. The results revealed that adult emergence and activity/rest rhythms of flies from the selected stocks were more robust than controls, as they displayed enhanced stability under a wide variety of environmental conditions. These results suggest that selection for adult emergence within a narrow window of time results in the evolution of robustness in circadian timing systems of the fruit fly D. melanogaster. (Author correspondence: vsharma@jncasr.ac.in or vksharmas@gmail.com)     

Selection for early and late adult emergence alters the rate of pre-adult development in Drosophila melanogaster

Background  

Circadian clocks have been implicated in the regulation of pre-adult development of fruit flies Drosophila melanogaster. It is believed that faster clocks speed up development and slower clocks slow it down. We established three sets of D. melanogaster populations (early, control and late). The early and late populations were raised by selecting for flies that emerged either in the morning or in the evening under 12:12 hr light/dark (LD) cycles. After 75 generations of selection, the time course and waveform of the adult emergence and activity rhythms of the early and the late populations diverged from each other as well as from the controls. In this paper, we report the consequence of this selection on the rate of pre-adult development.     

Evolution of circadian rhythms in Drosophila melanogaster populations reared in constant light and dark regimes for over 330 generations

Organisms are believed to have evolved circadian clocks as adaptations to deal with cyclic environmental changes, and therefore it has been hypothesized that evolution in constant environments would lead to regression of such clocks. However, previous studies have yielded mixed results, and evolution of circadian clocks under constant conditions has remained an unsettled topic of debate in circadian biology. In continuation of our previous studies, which reported persistence of circadian rhythms in Drosophila melanogaster populations evolving under constant light, here we intended to examine whether circadian clocks and the associated properties evolve differently under constant light and constant darkness. In this regard, we assayed activity-rest, adult emergence and oviposition rhythms of D. melanogaster populations which have been maintained for over 19 years (~330 generations) under three different light regimes – constant light (LL), light–dark cycles of 12:12 h (LD) and constant darkness (DD). We observed that while circadian rhythms in all the three behaviors persist in both LL and DD stocks with no differences in circadian period, they differed in certain aspects of the entrained rhythms when compared to controls reared in rhythmic environment (LD). Interestingly, we also observed that DD stocks have evolved significantly higher robustness or power of free-running activity-rest and adult emergence rhythms compared to LL stocks. Thus, our study, in addition to corroborating previous results of circadian clock evolution in constant light, also highlights that, contrary to the expected regression of circadian clocks, rearing in constant darkness leads to the evolution of more robust circadian clocks which may be attributed to an intrinsic adaptive advantage of circadian clocks and/or pleiotropic functions of clock genes in other traits.     

Cyclic presence and absence of conspecifics alters circadian clock phase but does not entrain the locomotor activity rhythm of the fruit fly Drosophila melanogaster

Circadian clocks use a wide range of environmental cues, including cycles of light, temperature, food, and social interactions, to fine-tune rhythms in behavior and physiology. Although social cues have been shown to influence circadian clocks of a variety of organisms including the fruit fly Drosophila melanogaster, their mechanism of action is still unclear. Here, the authors report the results of their study aimed at investigating if daily cycles of presence and absence (PA) of conspecific male visitors are able to entrain the circadian locomotor activity rhythm of male hosts living under constant darkness (DD). The results suggest that PA cycles may not be able to entrain circadian locomotor activity rhythms of Drosophila. The outcome does not change when male hosts are presented with female visitors, suggesting that PA cycles of either sex may not be effective in bringing about stable entrainment of circadian clocks in D. melanogaster. However, in hosts whose clock phase has already been set by light/dark (LD) cycles, daily PA cycles of visitors can cause measurable change in the phase of subsequent free-running rhythms, provided that their circadian clocks are labile. Thus, the findings of this study suggest that D. melanogaster males may not be using cyclic social cues as their primary zeitgeber (time cue) for entrainment of circadian clocks, although social cues are capable of altering the phase of their circadian rhythms.     

Possible Evidence for Shift Work Schedules in the Media Workers of the Ant Species Camponotus compressus

The locomotor activity rhythm of the media workers of the ant species Camponotus compressus was monitored under constant conditions of the laboratory to understand the role of circadian clocks in social organization. The locomotor activity rhythm of most ants entrained to a 24 h light/dark (12:12 h; LD) cycle and free-ran under constant darkness (DD) with circadian periodicities. Under entrained conditions about 75% of media workers displayed nocturnal activity patterns, and the rest showed diurnal activity patterns. In free-running conditions these ants displayed three types of activity patterns (turn-around). The free-running period (τ) of the locomotor activity rhythm of some ants (10 out of 21) showed period lengthening, and those of a few (6 out of 21) showed period shortening, whereas the locomotor activity rhythm of the rest of the ants (5 out of 21) underwent large phase shifts. Interestingly, the pre-turn-around τ of those ants that showed nocturnal activity patterns during earlier LD entrainment was shorter than 24 h, which became greater than 24 h after 6–9 days of free-run in DD. On the other hand, the pre-turn-around τ of those ants, which exhibited diurnal patterns during earlier LD entrainment, was greater than 24 h, which became shorter than 24 h after 6–9 days of free-run in DD. The patterns of activity under LD cycles and the turn-around of activity patterns in DD regime suggest that these ants are shift workers in their respective colonies, and they probably use their circadian clocks for this purpose. Circadian plasticity thus appears to be a general strategy of the media workers of the ant species C. compressus to cope with the challenges arising due to their roles in the colony constantly exposed to a fluctuating environment.     

Molecular evolution of sex-biased genes in the Drosophila ananassae subgroup

Background  

Genes with sex-biased expression often show rapid molecular evolution between species. Previous population genetic and comparative genomic studies of Drosophila melanogaster and D. simulans revealed that male-biased genes have especially high rates of adaptive evolution. To test if this is also the case for other lineages within the melanogaster group, we investigated gene expression in D. ananassae, a species that occurs in structured populations in tropical and subtropical regions. We used custom-made microarrays and published microarray data to characterize the sex-biased expression of 129 D. ananassae genes whose D. melanogaster orthologs had been classified previously as male-biased, female-biased, or unbiased in their expression and had been studied extensively at the population-genetic level. For 43 of these genes we surveyed DNA sequence polymorphism in a natural population of D. ananassae and determined divergence to the sister species D. atripex and D. phaeopleura.     

Drosophila ezoana uses an hour‐glass or highly damped circadian clock for measuring night length and inducing diapause

Insects inhabiting the temperate zones measure seasonal changes in day or night length to enter the overwintering diapause. Diapause induction occurs after the duration of the night exceeds a critical night length (CNL). Our understanding of the time measurement mechanisms is continuously evolving subsequent to Bünning's proposal that circadian systems play the clock role in photoperiodic time measurement (Bünning, 1936). Initially, the photoperiodic clocks were considered to be either based on circadian oscillators or on simple hour‐glasses, depending on ‘positive’ or ‘negative’ responses in Nanda–Hamner and Bünsow experiments (Nanda & Hammer, 1958; Bünsow, 1960). However, there are also species whose responses can be regarded as neither ‘positive’, nor as ‘negative’, such as the Northern Drosophila species Drosophila ezoana, which is investigated in the present study. In addition, modelling efforts show that the ‘positive’ and ‘negative’ Nanda–Hamner responses can also be provoked by circadian oscillators that are damped to different degrees: animals with highly sustained circadian clocks will respond ‘positive’ and those with heavily damped circadian clocks will respond ‘negative’. In the present study, an experimental assay is proposed that characterizes the photoperiodic oscillators by determining the effects of non‐24‐h light/dark cycles (T‐cycles) on critical night length. It is predicted that there is (i) a change in the critical night length as a function of T‐cycle period in sustained‐oscillator‐based clocks and (ii) a fixed night‐length measurement (i.e. no change in critical night length) in damped‐oscillator‐based clocks. Drosophila ezoana flies show a critical night length of approximately 7 h irrespective of T‐cycle period, suggesting a damped‐oscillator‐based photoperiodic clock. The conclusion is strengthened by activity recordings revealing that the activity rhythm of D. ezoana flies also dampens in constant darkness.     

Possible role of eclosion rhythm in mediating the effects of light-dark environments on pre-adult development in <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>

Background  

In insects, circadian clocks have been implicated in affecting life history traits such as pre-adult development time and adult lifespan. Studies on the period (per) mutants of Drosophila melanogaster, and laboratory-selected lines of Bactrocera cucurbitae suggested a close link between circadian clocks and development time. There is a possibility of clock genes having pleiotropic effects on clock period and pre-adult development time. In order to avoid such pleiotropic effects we have used wild type flies of same genotype under environments of different periodicities, which phenotypically either speeded up or slowed down the eclosion clock of D. melanogaster.     

Effects of Temperature,Photoperiod, and Light Intensity on the Eclosion Rhythm of the High-Altitude Himalayan Strain of Drosophila ananassae

Eclosion rhythm of the high-altitude Himalayan strain of Drosophila ananassae from Badrinath (altitude 5123 m) was temperature-dependent and at 21°C, it was entrained by cycles of 12 h light: 12 h darkness (LD 12:12) and free-ran in constant darkness, however, it was arrhythmic at 13°C or 17°C under identical experimental conditions (Khare, P. V., Barnabas, R. J., Kanojiya, M., Kulkarni, A. D., Joshi, D. S. (). Temperature dependent eclosion rhythmicity in the high altitude Himalayan strains of Drosophila ananassae. Chronobiol. Int. 19:1041–1052). The present studies were designed to see whether or not these strains could be entrained at 13°C, 17°C, and 21°C by two types of LD cycles in which the photoperiod at 100 lux intensity varied from 6 h to 18 h, and the light intensity of LD 14:10 cycles varied from 0.001 lux to 1000 lux. All LD cycles entrained this strain at 21°C but not at 13°C or 17°C. These results demonstrate that the entrainment of eclosion rhythm depends on the ambient temperature and not on the photoperiod or light intensity of LD cycles. Thus the temperature has taken precedence over the light in the entrainment process of eclosion rhythm of the high altitude Himalayan strain of D. ananassae. This may be the result of natural selection in response to the environmental temperature at Badrinath that resembles that of the sub-Arctic region but the photoperiod or light intensity are of the subtropical region.     

Impact of body melanisation on contrasting levels of desiccation resistance in a circumtropical and a generalist <Emphasis Type="Italic">Drosophila</Emphasis> species

We investigated the role of cuticular lipids, body melanisation and body size in conferring contrasting levels of desiccation resistance in latitudinal populations of Drosophila melanogaster and Drosophila ananassae on the Indian subcontinent. Contrary to the well known role of cuticular lipids in water proofing in diverse insect taxa, there is lack of geographical variations in the amount of cuticular lipids per fly in both the species. In D. ananassae, quite low levels of body melanisation are correlated with lower desiccation resistance. By contrast, increased levels of desiccation resistance are correlated with quite high melanisation in D. melanogaster. Thus, species specific cuticular melanisation patterns are significantly correlated with varying levels of desiccation resistance within as well as between populations and across species. Role of body melanisation in desiccation resistance is further supported by the fact that assorted dark and light flies differ significantly in cuticular water loss, hemolymph and dehydration tolerance. However, similar patterns of body size variation do not account for contrasting levels of desiccation resistance in these two Drosophila species. Climatic selection is evidenced by multiple regression analysis with seasonal amplitude of thermal and humidity changes (Tcv and RHcv) along latitude on the Indian subcontinent. Finally, the contrasting levels of species specific distribution patterns are negatively correlated with RHcv of sites of origin of populations i.e. a steeper negative slope for D. ananassae corresponds with its desiccation sensitivity as compared with D. melanogaster. Thus, evolutionary changes in body melanisation impact desiccation resistance potential as well as distribution patterns of these two Drosophila species on the Indian subcontinent.