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Longevity of seven species of cactophilic Drosophila and D. melanogaster on carbohydrates
J. lnsecr Physiol.
Vol. 31. No. 2. pp. 165-169, 1985 Printed in Great Britain. All rights reserved
0022-1910/85 $3.00 + 0.00 Copyright cs 1985 Pergamon Press Ltd
LONGEVITY
OF SEVEN SPECIES OF CACTOPHILIC DROSOPHILA AND D. MELANOGASTER ON CARBOHYDRATES HENRY W.
Department
of Nutrition
and Food
KIRCHER* and BEDIR AL-AZAWI* Science. College of Agriculture, University
of Arizona,
Tucson,
AZ 85721, U.S.A. (Received 31 August 1983: reaised 26 June 1984) Abstract-Adults of 6 species of Drosophila that use decaying prickly pear cactus (Opuntia sp.) as breeding and feeding sites were compared to each other and to D. nigrospiraculu. whose host is saguaro cactus, and to the cosmopolitan D. melanogaster. in their utilization of 21 sugars for longevity (time to 50% mortality). In general. the utilization of sugars by these flies for longevity followed the pattern observed with the other insects. None of the species were able to live very long on solutions of pentoses, uranic acids, inositol. rhamnose. sorbose or the P-linked disaccharides, lactose and cellobiose. Although all could use glucose, fructose, sucrose, maltose and melezitose well. their life spans on galactose, mannose, trehalose and raffinose were more variable. Two of the Opuntia feeders were also tested on a number of other carbohydrates. Ribitol, mannitol, sorbitol and xylitol significantly prolonged the life of D. arizonensis but not that of D. wheeleri. Neither species lived long on solutions of arabitol. galactitol, starch. inulin or on arabogalactan. Key Word Index: Drosophila. sugars,
longevity.
cactus
INTRODUCTION
genus Drosophila contains species having a wide variety of habitats. Adults and larvae feed on microorganisms associated with nectars, plant exudates, decaying trees, flowers, leaves, fungi, and in desert areas, decaying cacti. Several desert species are specialists, breeding in rotting tissues of only one or two kinds of large columnar cacti (Fellows and Heed, 1972), while others can feed on columnar cacti as well as the fruits and rotting stems of various prickly pear cacti (Opuntiu species) and still others are limited to the Opuntia (Barker and Starmer, 1982). A problem with desert living Drosophila is to rationalize the longevity of adults. Although the larvae of cactophilit Drosophila are able to go from eggs to pupae in the matrix of bacteria, yeasts and rotting cactus, the adults do not live as long in the laboratory on this substrate as they do on formulated media that contain sugars or on decaying cactus supplemented with glucose (Heed, personal communication). It has also been observed for 10 dipterous insects (House, 1974) and for D. melunoguster in particular, that sugars are not required for larval maturation (Sang, 1956; David, 1959) but that adults survive much longer when sugar is added to their diets (Hollingsworth and Burcombe, 1970). Recent work in our laboratory (Kircher, Mohamed, Abril, unpublished) on the methanol and 80% ethanol-soluble fractions of several fresh and decaying cacti made us aware of the status of sugars in The
*Present address: c/o Dr James Fogleman. Department Biological Sciences, University of Denver, Denver. 80208. U.S.A. *Died January 1984.
of CO
165
these plants. In two species of prickly pear cactus that are substrates for several species of Drosophila in southern Arizona, Opuntiujicus-indicu and 0. phaeucunthu, the concentration of glucose, fructose and sucrose in the stems falls from 613”/, to about 1.5% of the dry weight during the first 4-5 days of decay. The sugar concentration then remains low (1%) during the subsequent 34 week slow hydrolysis of the water soluble araboxylogalactan in the cacti and consumption of the resulting sugars by bacteria and yeasts. By contrast, fresh saguaro cactus contains 4.5% sugars (2: 1, glucose-fructose) tied up as glycosides, and this decreases to 2% of the dry matter after 8 days of decay. Organ pipe cactus contains little free sugar but a 1: 1 ratio of glucose-rhamnose is present as a tetrasaccharide attached to 3 acidic triterpenes (Kircher, 1977). These four present a varying sugar profile to adult Drosophila feeding on them and it was of interest to us to determine sugar utilization by a number of cactophilic species by using longevity of newly eclosed adults on 5% solutions of various sugars as a criterion of sugar utilization. We chose 6 species that are Opuntiu feeders, one that breeds and feeds in saguaro cactus, and compared these to the cosmopolitan species, D. melanoguster (Table 1). According to cytological studies of Wasserman (1982) and Heed (personal communication), D. uldrichi and D. wheeleri are each others closest relatives as are D. mojuvensis, D. urizonensis and D. species “N”. D. mulleri is closer to the former two than the latter three. These relationships are exemplified by groupings in Tables 1-4. Numerous papers have been published on the utilization of sugars by insects (reviews: House, 1974;
166
HENRY W. KIRCHER and BEDIR AL-AZAWI
Wyatt,
1967); this is the first to compare 7 species in the same genus and species group that share similar habitats. MATERIALS
AND METHODS
Drosophilu
The eight species used in this study (Table 1) were collected and identified by Dr W. B. Heed of the Ecology Department and were maintained until use on a standard medium (Kircher et al., 1982). Carbohydrates
The sugars and sugar alcohols (all D- except were noted), starch and inulin were obtained from Sigma. The sugars were shown to be free of contaminants by paper chromatography and the alditols free of aldoses by the phenol-sulphuric acid method (Dubois et al.. 1956) except that mannitol contained 1.3% mannose and sorbitol had 1.9% glucose. The low molecular weight arabogalactan was isolated from organ pipe cactus (Kircher, unpublished). E_rperimental procedure
Twenty newly eclosed adults, chosen at random from a large population of each species of Drosophila, were placed in a cloth-covered 1 1. wide mouth jar lined with moist filter paper that contained 7-8 ml of a So,, sugar solution in a 10ml wide-mouth cottonstoppered vial. The vials were held in a horizontal position by white tape near the tops of the jars. Each species was tested in triplicate on each sugar as well as a water control and all 21 sugars were run simultaneously for a given species. The jars were kept on a laboratory bench away from sunlight and mortalities in each jar counted daily. Paper on the bottom of the jars was kept moist and sugar solutions replaced when necessary. After completion of these experiments (8 species and 21 sugars), the same procedure was used to test 8 sugar alcohols, 3 polysaccharides and melibiose with D. arizonensis and D. wheeleri.
Values of M,, (days to 50% mortality) were obtained from the data gathered from each replicate and average values of this parameter together with standard errors (SE) were calculated from the 3 replicates of each test. RESULTS
Values of M,, ranged from O-3 days for the eight species when given only water. These were subtracted from the values of M,, for the various sugars to give “enhanced longevities” which can be attributed to the nutritive values of the sugars for the insects. It is Table Species D. aldrichr D.
w,her/err
D. mulleri D. mojawnsis D. orizonensis D.
species “N”
D. nigrospiracula D. melanogaster
*Cacti names.
1. Drosophila
Species group Repleta Repleta Repleta Repleta Repleta Repleta Repleta Melanogaster
possible that some sugars were nutritionally useless to the flies, but their flavour induced more water consumption and thereby retarded mortality. It is also possible that oesophageal and crop microorganisms were able to utilize some sugars and therefore allow their host to live longer by digestion of the multiplying microorganisms. We were not able to gauge the importance of these two factors from the present experiments. The results give in Tables 2-4 simply show how much longer eight species of Drosophila were able to live compared to water alone on 5”/; solutions of pentoses, hexoses, oligosaccharide sugar alcohols and three polysaccharides, and these data were compared. where applicable, to those by Hassett (1948) with D. melanogaster.
DISCUSSION
The ability of individual carbohydrates to prolong the life of adult insects is a complex function of at least 4 factors: (I) The amount of substrate eaten, which is a function of its concentration and taste (Dethier, 1976). (2) The presence of enzymes in the gut and haemolymph to hydrolyze complex sugars and polysaccharides (Webber, 1957). (3) Absorption from the gut into the haemolymph. (4) Metabolism of sugars in the haemolymph and in the tissues (Wyatt, 1967). The longevity of adult Diptera caged with various sugars and polysaccharides as the only nutrient has been determined for numerous species: Calliphora erythrocephala (Haslinger, 1935; Fraenkel 1940), Drosophila melanogaster (Hassett, 1948; Ohsawa and Tsuda, 1956), Phormia regina (Hassett et al., 1950), Lucilia cuprina (Webber, 1957), Aedes aegypti, Sarcophaga bullata and Musca domestica (Galun and Fraenkel, 1957) Eroischia brassicae (Finch and Coaker, 1969), Ceratatis capitata (Gothilf et al., 1971) and Dacus oleae (Tsiropoulos, 1980). In our study, the sugars that provided for maxima1 longevity were also those that were the most nutritive in all of these other cases. Of these, glucose, fructose and sucrose, which are the most effective in prolonging life, are also the most widely distributed in nature and elicit the strongest gustatory responses (Dethier, 1976; Falk, 1979; Miyakawa et af., 1980). There were, however, important differences in sugar utilization between the eight species of Drosophila used in this study and also with other insects. These differences will be discussed by groups of sugars.
species used m longevity
experiments
Complex Mulleri Mulleti Mulleri Mulleti Mulleri Mldleri Anceps
Natural Opuntia Opuntia Opuntia Opuntia, Opuntia Opuntia Saguaro. decaying
substrates*
cacti cacti cacti barrel, agria and organ pipe cacti and cina cacti, citrus fruit cacu hecho. cardon cacti vegetable matter
Drosophila
and sugars
167
Table 2. Enhanced longevity” of Drosophila on 5% solutions of carbohydrates that either are not eaten or are poorly absorbed and metabolized bv most of the 8 soecies Species of Drosophilab
ST; Solutions of ____. __ o-arabinose L-atabinose Lyxose Ribose Xylose r-rhamnose Sorbose Inosltol Galacturonic acid Glucuronic acid Cellobiose
D. aldr.
D. wheel.
0
2 I 0 2 3 2 2 3 0 2 I
D. melanogaster
D. mull.
D. ark.
D. moja.
0
0
-I
t 0 0 0 0 0
4il 6 0 0 3 0 2
452 2
-I -1
-I 0 0
D. sp “N” -I 0 0
0
3
0
-1 0 0 0
-1 0 -I
-I -2
-I
-3
““Enhanced longevity” equals days to 50”b mortality on the SOjO carbohydrate each species. SE c 1.0 unless noted. bSee Table I for complete names of the Drosophila species. c7.59; solutions. d 1.8”; solutions. ‘3.6”, solutions.
Pentoses und L-rhamnose
Although no insects can survive very long on these sugars. adults of a few species are able to live 14 days longer on ribose and xylose solutions than on water alone (Hassett et al., 1950; Finch and Coaker, 1969: Tsiropoulos. 1980). This was also observed
D. nigro.
This work._ __. 0 -I 0 0 0 0 0 0 0 0 0
Hassett ( 1948) I< 0
0’
solutions minus days to 50% mortality on water alone for
with Drosophila in the present study (Table 2). D and L-arabinose were of little value to any of the species and lyxose and L-rhamnose to only I or 2 whereas ribose and/or xylose could be used by the majority of the flies to prolong their lives from 24 days longer than on water alone. D. aldrichi and D. arizonensis were able to utilize ribose and xylose the most
Table 3. Enhanced longevity’of Drosophila on 5”, solutions of carbohydrates species
that slgnilicantly prolong the adult lifespans of most of the
Species of Drosophila h
D. melanogas&r
Hasset So, Solutions of Glucose Fructose Galactose Mannose Sucrose Maltose Melezitose Trehalose Raffinose
D. aldr.
D. wheel.
I8 20 II I3 I8 16 I4 IO IO
19 29_+ 3.5 8 22& 3 34 16 21*7 30 2
.__-
D. mull.
18+4 20 8i3 I2 24 22 20 15 f 2.5 13
D. ark. __~~
25 25 8 1323 24k2.5 23 21 5 8
rp “N” ~_ ~~ D. ____~~
D. moja.
28 28 2 I8 29 25 I9 23f4 6
24 31 I5 I4 23 19 20 i 5 13 1
D. nigro.
~_~
This work ~_ (1948) ~____.
24 25 8 6 I8 I6 13 I6 8
24 28 I5 24 27 25 20 20 I6 + 3
“Enhanced longevity = SE $2.0 unless noted. “See Table I for complete names of each species. c~.8U, solutions. d3.6°:, solutions. Table 4. Enhanced longevity (in days) of Drosophila on 54” solutions of sugar alcohols, polysaccharides and melibiosea,b D. arironensis
Carbohydrate Adonitol (Ribotol) Ribose o-Arabitol L-Arabitol Dulcitol (Galactitol) Fructose Mannitol Glucitol Xylitol Xylose Maltitol Maltose Melibiose Arabogalactan Inulin Starch “Unusual values are underlined. bSE < I unless noted. ‘Hassett (1948). 1.80,, solutions.
_-~~ MS0 ~~~~22 + 1.5 4 2 2 1 25 w 34 lo 6 31 * 1.5 23 20 2 1 4
D. wheeleri
MS0 3 2 0
D. melanogaster’
_~_.
~?? 2 0
I 0 29 + 3 2 4 4 3 I1 I6 6 0 1 2
0 I6 4 3
I5 0 2
14’ 16’ 9 I? 22d 15d I2 19’ 13’
168
HENRY W. KIRCHER and BEDIR AL-AZAWI
efficiently. Their modest utilization of the two pentoses may reflect the ability of the flies to reduce them to ribitol and xylitol and then reoxidize the alditols to the metabolizable ribulose and xylulose. The lower longevity of the insects on D or t_-arabinose and lyxose may be due to their inability either to reduce these pentoses to arabitol and lyxitol or to oxidize the two pentitols to ribulose and xylulose respectively. This hypothesis is supported by several pieces of evidence from work with other insects. An aldehyde sugar reductase in the silkworm is able to reduce triose. tetrose and pentose but not hexose sugars to the corresponding alditols (Faulkner. 1958) and galactose to galactitol reductase occurs in the bollweevil (Nettles and Burks, 1971). The oxidation of sorbitol to fructose by an alditol oxidase occurs in mosquitos (Van Handel, 1969) as well as nine other insects (Wenzl, 1969). Therefore, reductions of aldoses such as ribose and xylose to the alditols and the oxidation of these to metabolizable ribulose and xylulose has a precedent in the literature. Unfortunately, the two ketopentoses are too expensive to be used in feeding trials. The utilization of pentoses and t-rhamnose by D. nigrospiracufa may reflect the chemical composition of its host cactus. Saguaro contains no free glucose. fructose or sucrose as the Opuntia do and instead contains glucose and rhamnose as glycosides. D. nigrospiraculu adults may have adapted to the utilization of unusual sugars because the common ones are not available to this species as they are to flies living on Opuniia cacti. Hexoses and inositol The values of M,, for glucose, fructose, galactose. and mannose are listed in Table 3. Those for L-sorbose. inositol, and the uranic acid derivatives of galactose and glucose are shown in Table 2. Of the first four hexoses, galactose was the least used and mannose was also poorer than glucose or fructose in prolonging life. D. mojauensis was unusual in that it lived only slightly longer on galactose than water alone and D. nigrospiracula could use mannose onehalf or less as well as the other seven species. Of the remaining 4 compounds (Table 2) L-sorbose. which prolonged the life of only one other insect to a small extent (Galun and Fraenkel, 1957) was of some benefit to D. nigrospiracula and D. wheeleri. It is metabolically inert because it cannot be reduced to sorbitol (Faulkner, 1958). Inositol could also be used to a small extent by four of the species of Drosophila and has been reported to be of some benefit to three other Diptera (Galun and Fraenkel. 1957: Finch and Coaker, 1969; Tsiropou10s. 1980). Uranic acids are usually not tested as energy sources for insects. Of the two, glucuronic acid supported three species while galacturonic acid supported only 1 for 24 days longer than the water controls. Di- and trisuccharides Data obtained with these sugars are shown Tables 2 and 3. Sucrose. maltose and melezitose clearly those that prolong life of Drosophila longest. Trehalose and raffinose are less well used cellobiose and lactose not all. Although lactose
in are the and can
prolong the life of 2 insects (Galun and Fraenkel, 1957; Finch and Coaker, 1969) the two p-linked disaccharides appear to be toxic to D. mojarrensis. These results are generally in accord with those obtained with other insects (Table 2; House, 1974). In our study, D. arizonensis lived for a considerably shorter time and D. wheeleri for a longer time on trehalose than the other 7 species. D. species “N” and D. wheeleri were exceptional in their poor usage of raffinose and all of the cactophilic species were less able than D. melnnogaster to use this sugar as an energy source. Melibiose, polwaccharides, multitol und other ulditols Data in Table 4 show longevities of D. arizonensis and D. wheeleri compared to D. melanogaster on solutions of these compounds. The two cactophilic species used maltitol as readily as maltose and their relative use of melibiose paralleled that of raffinose. Maltitol has not been previously tested with insects; melibiose is readily used by many (Fraenkel, 1940; Hassett, 1948; Webber, 1957: Galun and Fraenkel, 1957; Finch and Coaker. 1969 and Tsiropolous, 1980); an exception being the blowfly (Hassett et al., 1950). The ability of polysaccharides to sustain the life of adult insects varies with the polysaccharide and insect tested. Whereas dextrin could prolong the life of 7 insects (Fraenkel, 1940; Hassett. 1948; Webber, 1957: Galun and Fraenkel, 1957) and starch, the life of 3 (Webber, 1957; Gallun and Fraenkel, 1957). inulin and glycogen (except for I case. Tables 23) were generally inert. None of the 3 Drosophila in Table 4 were able to use the polysaccharides to significantly prolong their lives. Performance on sugar alcohols is also variable (Table 4). Those that can prolong the life of some insects, such as mannitol. sorbitol, adonitol and to a lesser extent, xylitol. as with D. urkonensis. are probably oxidized to fructose. ribulose and xylulose for subsequent utilization for energy (Van Handel, 1969; Wenzl, 1969). These alditols are also readily metabolized by mammals (Hood et al., 1977; Sipple and McNutt. 1974). whereas II- and t-arabitol and dulcitol. not usable by any insects (Fraenkel. 1940; Hassett et ul., 1950: Webber, 1957; Galun and Fraenkel, 1957: Finch and Coaker, 1969; Tsiropolous, 1980). are also poorly utilized by mammals. These findings also correlate with the modest usage of ribose and xylose by Drosophilu compared to that of II- and L-arabinose (Table 2). The longer life of D. urkonensis on adonitol and xylitol than on the respective aldoses, ribose and xylose, lends credence to the hypothesis that metabolism of these pentoses goes by the route: aldose-alditolLketose. Comparison of the three species in Table 4 shows a strong resemblance between D. wheeleri and D. melanogaster: they both differ considerably from n. arizonensis. This relationship is also reflected in the relative utilization of pentoses (Table 2). mannose (Table 3) and trehalose (Table 3) by the three species. Taken in their entirety, the data obtained in this study show that adults of eight species of Drosophilu resemble other insects in their utilization of carbohydrates to prolong their lives. In addition. the ability of the largest in size but shortest lived species, D.
Drosophila and sugars nigrospiracula, to use L-rhamnose, L-sorbose, inositol, galacturonic and glucuronic acids better than the other 6 cactophilic species (Table 2) may be related
to the fact that it is the only one that does not breed in Opuntiu cacti (Table I). The natural host of D. nigrospiracula, saguaro cactus, contains no detectable free sugars (glucose, fructose, sucrose) before decay, whereas the concentration of these sugars in prickly pear cacti is 612%. This may have forced D. nigrospiraculu to adapt to “less utilizable” sugars to sustain adult life. Acknowledgements-We thank Drs W. B. Heed, David Baldwin and J. C. Fogleman for fruitful discussions concerning this work and the National Science Foundation for grant DEB-8207056 for partial support. Arizona Agricultural Experiment Station Journal article no. 3795.
REFERENCES Barker J. S. F. and Starmer W. T. eds (1982) Ecological Genetics and Etlolution. The Cactus- Yeast-Drosophila Model System. Academic Press, Australia, North Ryde, N.S.W. David J. (1959) Etude Quantitative du Developpement de la Drosophfle eleoee en Milieu Axenique. Bull. Biol. Fr. Belg. (Paris) 93, 472-505. Dethier V. Ci. (1976) The Hungry Fly. Harvard Univ. Press. Cambridge, MS. Dubois M., Gillis K., Hamilton J. K., Rebers P. A. and Smith F. (1956) Calorimetric method for determinations of sugars and related substances. Analyt. Chem. 28, 35C-356. Falk R. (1979) Taste responses of Drosophila melanogaster J. Insect Physiol. 25, 87-9 I. Faulkner P. (1958) Polyol dehydrogenase of the silkworm. Biorhem. J. 68, 374-380. Fellows D. P. and Heed W. B. (1972) Factors affecting host plant selection in desert adapted cactiphilic Drosophila. Ecology 53, 85&858. Finch S. and Coaker T. H. (1969) Comparison of the nutritive values of carbohydrates and related compounds to Eroischia brassicae. Ent. exp. Appl. 12, 44453. Fraenkel G. (1940) Utilization and digestion of carbohydrates by the adult blowfly. J. exp. Biol. 17, 18-28. Galun R. and Fraenkel G. (1957) Physiological effects of carbohydrates in the nutrition of a mosquito Aedes aegypti and two flies. Sarcophaga bullata and Musca domestica. J. Cell. Comp. Physiol. 50, l-23. Gibson A. C. and Horak K. E. (1978) Systematic anatomy and phylogeny of Mexican columnar cacti. Ann. Missouri Bot. Gard. 65, 999-1057. Gothilf S.. Galun R. and Bar-Zeev M. (1971) Taste recep-
169
tion in the Mediterranean fruit fly: electrophysiological and behavioral studies. J. lnsecf Ph_vsiol. 17, 1371- 1384. Haslinger F. (1935) Uber den Geschmack Sinn Von Calliphora erythrocephala Meigen und uber die ververtung von Zuckern und Zuckeralkohole druch diese Fliege. Zeit. f. vergl. Physiol. 22, 614640. Hassett C. C. (1948) The utilization of sugars and other substances by Drosophila. Bioi. Bull. mar. biol. Luh., Woo& Hole 95, 114123. Hassett C. C.. Dethier B. G. and Gans J. (I 950) A comparison of nutritive values and taste thresholds of carbohydrates for the blowfly. Biol. Bull. mar. biol. Lab., Woods Hole 99, 446-453. Hollingsworth M. J. and Burcombe J. V. (1970) The nutritional requirements for longevity in Drosophila. J. Insect Physiol. 16, 1017-1025. House H. L. (1974) P/2ysiology of‘ Insecta. Vol. V, p. 26. Academic Press, NY. Kircher H. W. (1977) Triterpene glycosides and quereteroic acid in organ pipe cactus. Phytochemistry. 16, 1078- 1080. Kircher H. W., Phariss R. L.. Rosenstein, F. U.. Baldwin D. and Fogleman J. C. (1982) Response of Drosophila to cis- and trans-22-dehydrocholesterol: I. A survey. Lipids 17, 209-214. Miyakawa Y., Fujishiro N.. Kijima H. and Morita H. (1980) Differences in feeding response to sugars between adults and larvae in Drosophila melanogaster. J. Insect. Physiol. 26, 685-688. Nettles W. C. Jr. (1972) Toxic sugars and longevity of Anthonomus grandis. J. Insect Physiol. 18, I 107-I I 13. Nettles W. C. and Burks M. L. (1971) Absorption and metabolism of galactose and galactitol in Anthonus grandis. J. Insect Physiol. 17, 1615-1623. Ohsawa W. and Tsukuda H. (1956) Some notes on the utilization of sugars by Drosophila melanogaster. J Inst. Polvtech. Osaka Citv, Unio. Ser. D. 7, 163-11 I. Sang J. H. (1956) The quantitative nutritional requirements of Drosophila melanogaster. J. exp. Biol. 33, 45-72. Tsiropoulos G. J. (1980) Carbohydrate utilization by normal and sterilized Dacus oleae. J. Insert Ph?siol. 26, 633-631. Van Handel E. (1969) The equilibrium reaction sorbitol fructose in the intact mosquito. Camp. Biochem. P/lJ,.siol. 29, 1023-1030. Wasserman M. (1982) Ed. by Barker J. S. F. and Starmer W. T. Ecological Genetics and Ettolution. The CuctusYeast-Drosophila Model System. Academic Press, Australia, North Ryde, N.S.W. Webber L. G. (1957) Utilization and digestion of carbohydrates by the Australian sheep blowfly Lucilia cuprina. Ausf. J. Zool. 5, 164-172. Wenzl H. (1969) Sorbitol dehydrogenase in insects. Z. F. Vergl. Physiol. 62, 41 l-412. Wyatt G. R. (1967) The biochemistry of sugars and polysaccharides in insects. Adll. Insect. Physiol. 4, 287-460.
Vol. 31. No. 2. pp. 165-169, 1985 Printed in Great Britain. All rights reserved
0022-1910/85 $3.00 + 0.00 Copyright cs 1985 Pergamon Press Ltd
LONGEVITY
OF SEVEN SPECIES OF CACTOPHILIC DROSOPHILA AND D. MELANOGASTER ON CARBOHYDRATES HENRY W.
Department
of Nutrition
and Food
KIRCHER* and BEDIR AL-AZAWI* Science. College of Agriculture, University
of Arizona,
Tucson,
AZ 85721, U.S.A. (Received 31 August 1983: reaised 26 June 1984) Abstract-Adults of 6 species of Drosophila that use decaying prickly pear cactus (Opuntia sp.) as breeding and feeding sites were compared to each other and to D. nigrospiraculu. whose host is saguaro cactus, and to the cosmopolitan D. melanogaster. in their utilization of 21 sugars for longevity (time to 50% mortality). In general. the utilization of sugars by these flies for longevity followed the pattern observed with the other insects. None of the species were able to live very long on solutions of pentoses, uranic acids, inositol. rhamnose. sorbose or the P-linked disaccharides, lactose and cellobiose. Although all could use glucose, fructose, sucrose, maltose and melezitose well. their life spans on galactose, mannose, trehalose and raffinose were more variable. Two of the Opuntia feeders were also tested on a number of other carbohydrates. Ribitol, mannitol, sorbitol and xylitol significantly prolonged the life of D. arizonensis but not that of D. wheeleri. Neither species lived long on solutions of arabitol. galactitol, starch. inulin or on arabogalactan. Key Word Index: Drosophila. sugars,
longevity.
cactus
INTRODUCTION
genus Drosophila contains species having a wide variety of habitats. Adults and larvae feed on microorganisms associated with nectars, plant exudates, decaying trees, flowers, leaves, fungi, and in desert areas, decaying cacti. Several desert species are specialists, breeding in rotting tissues of only one or two kinds of large columnar cacti (Fellows and Heed, 1972), while others can feed on columnar cacti as well as the fruits and rotting stems of various prickly pear cacti (Opuntiu species) and still others are limited to the Opuntia (Barker and Starmer, 1982). A problem with desert living Drosophila is to rationalize the longevity of adults. Although the larvae of cactophilit Drosophila are able to go from eggs to pupae in the matrix of bacteria, yeasts and rotting cactus, the adults do not live as long in the laboratory on this substrate as they do on formulated media that contain sugars or on decaying cactus supplemented with glucose (Heed, personal communication). It has also been observed for 10 dipterous insects (House, 1974) and for D. melunoguster in particular, that sugars are not required for larval maturation (Sang, 1956; David, 1959) but that adults survive much longer when sugar is added to their diets (Hollingsworth and Burcombe, 1970). Recent work in our laboratory (Kircher, Mohamed, Abril, unpublished) on the methanol and 80% ethanol-soluble fractions of several fresh and decaying cacti made us aware of the status of sugars in The
*Present address: c/o Dr James Fogleman. Department Biological Sciences, University of Denver, Denver. 80208. U.S.A. *Died January 1984.
of CO
165
these plants. In two species of prickly pear cactus that are substrates for several species of Drosophila in southern Arizona, Opuntiujicus-indicu and 0. phaeucunthu, the concentration of glucose, fructose and sucrose in the stems falls from 613”/, to about 1.5% of the dry weight during the first 4-5 days of decay. The sugar concentration then remains low (1%) during the subsequent 34 week slow hydrolysis of the water soluble araboxylogalactan in the cacti and consumption of the resulting sugars by bacteria and yeasts. By contrast, fresh saguaro cactus contains 4.5% sugars (2: 1, glucose-fructose) tied up as glycosides, and this decreases to 2% of the dry matter after 8 days of decay. Organ pipe cactus contains little free sugar but a 1: 1 ratio of glucose-rhamnose is present as a tetrasaccharide attached to 3 acidic triterpenes (Kircher, 1977). These four present a varying sugar profile to adult Drosophila feeding on them and it was of interest to us to determine sugar utilization by a number of cactophilic species by using longevity of newly eclosed adults on 5% solutions of various sugars as a criterion of sugar utilization. We chose 6 species that are Opuntiu feeders, one that breeds and feeds in saguaro cactus, and compared these to the cosmopolitan species, D. melanoguster (Table 1). According to cytological studies of Wasserman (1982) and Heed (personal communication), D. uldrichi and D. wheeleri are each others closest relatives as are D. mojuvensis, D. urizonensis and D. species “N”. D. mulleri is closer to the former two than the latter three. These relationships are exemplified by groupings in Tables 1-4. Numerous papers have been published on the utilization of sugars by insects (reviews: House, 1974;
166
HENRY W. KIRCHER and BEDIR AL-AZAWI
Wyatt,
1967); this is the first to compare 7 species in the same genus and species group that share similar habitats. MATERIALS
AND METHODS
Drosophilu
The eight species used in this study (Table 1) were collected and identified by Dr W. B. Heed of the Ecology Department and were maintained until use on a standard medium (Kircher et al., 1982). Carbohydrates
The sugars and sugar alcohols (all D- except were noted), starch and inulin were obtained from Sigma. The sugars were shown to be free of contaminants by paper chromatography and the alditols free of aldoses by the phenol-sulphuric acid method (Dubois et al.. 1956) except that mannitol contained 1.3% mannose and sorbitol had 1.9% glucose. The low molecular weight arabogalactan was isolated from organ pipe cactus (Kircher, unpublished). E_rperimental procedure
Twenty newly eclosed adults, chosen at random from a large population of each species of Drosophila, were placed in a cloth-covered 1 1. wide mouth jar lined with moist filter paper that contained 7-8 ml of a So,, sugar solution in a 10ml wide-mouth cottonstoppered vial. The vials were held in a horizontal position by white tape near the tops of the jars. Each species was tested in triplicate on each sugar as well as a water control and all 21 sugars were run simultaneously for a given species. The jars were kept on a laboratory bench away from sunlight and mortalities in each jar counted daily. Paper on the bottom of the jars was kept moist and sugar solutions replaced when necessary. After completion of these experiments (8 species and 21 sugars), the same procedure was used to test 8 sugar alcohols, 3 polysaccharides and melibiose with D. arizonensis and D. wheeleri.
Values of M,, (days to 50% mortality) were obtained from the data gathered from each replicate and average values of this parameter together with standard errors (SE) were calculated from the 3 replicates of each test. RESULTS
Values of M,, ranged from O-3 days for the eight species when given only water. These were subtracted from the values of M,, for the various sugars to give “enhanced longevities” which can be attributed to the nutritive values of the sugars for the insects. It is Table Species D. aldrichr D.
w,her/err
D. mulleri D. mojawnsis D. orizonensis D.
species “N”
D. nigrospiracula D. melanogaster
*Cacti names.
1. Drosophila
Species group Repleta Repleta Repleta Repleta Repleta Repleta Repleta Melanogaster
possible that some sugars were nutritionally useless to the flies, but their flavour induced more water consumption and thereby retarded mortality. It is also possible that oesophageal and crop microorganisms were able to utilize some sugars and therefore allow their host to live longer by digestion of the multiplying microorganisms. We were not able to gauge the importance of these two factors from the present experiments. The results give in Tables 2-4 simply show how much longer eight species of Drosophila were able to live compared to water alone on 5”/; solutions of pentoses, hexoses, oligosaccharide sugar alcohols and three polysaccharides, and these data were compared. where applicable, to those by Hassett (1948) with D. melanogaster.
DISCUSSION
The ability of individual carbohydrates to prolong the life of adult insects is a complex function of at least 4 factors: (I) The amount of substrate eaten, which is a function of its concentration and taste (Dethier, 1976). (2) The presence of enzymes in the gut and haemolymph to hydrolyze complex sugars and polysaccharides (Webber, 1957). (3) Absorption from the gut into the haemolymph. (4) Metabolism of sugars in the haemolymph and in the tissues (Wyatt, 1967). The longevity of adult Diptera caged with various sugars and polysaccharides as the only nutrient has been determined for numerous species: Calliphora erythrocephala (Haslinger, 1935; Fraenkel 1940), Drosophila melanogaster (Hassett, 1948; Ohsawa and Tsuda, 1956), Phormia regina (Hassett et al., 1950), Lucilia cuprina (Webber, 1957), Aedes aegypti, Sarcophaga bullata and Musca domestica (Galun and Fraenkel, 1957) Eroischia brassicae (Finch and Coaker, 1969), Ceratatis capitata (Gothilf et al., 1971) and Dacus oleae (Tsiropoulos, 1980). In our study, the sugars that provided for maxima1 longevity were also those that were the most nutritive in all of these other cases. Of these, glucose, fructose and sucrose, which are the most effective in prolonging life, are also the most widely distributed in nature and elicit the strongest gustatory responses (Dethier, 1976; Falk, 1979; Miyakawa et af., 1980). There were, however, important differences in sugar utilization between the eight species of Drosophila used in this study and also with other insects. These differences will be discussed by groups of sugars.
species used m longevity
experiments
Complex Mulleri Mulleti Mulleri Mulleti Mulleri Mldleri Anceps
Natural Opuntia Opuntia Opuntia Opuntia, Opuntia Opuntia Saguaro. decaying
substrates*
cacti cacti cacti barrel, agria and organ pipe cacti and cina cacti, citrus fruit cacu hecho. cardon cacti vegetable matter
Drosophila
and sugars
167
Table 2. Enhanced longevity” of Drosophila on 5% solutions of carbohydrates that either are not eaten or are poorly absorbed and metabolized bv most of the 8 soecies Species of Drosophilab
ST; Solutions of ____. __ o-arabinose L-atabinose Lyxose Ribose Xylose r-rhamnose Sorbose Inosltol Galacturonic acid Glucuronic acid Cellobiose
D. aldr.
D. wheel.
0
2 I 0 2 3 2 2 3 0 2 I
D. melanogaster
D. mull.
D. ark.
D. moja.
0
0
-I
t 0 0 0 0 0
4il 6 0 0 3 0 2
452 2
-I -1
-I 0 0
D. sp “N” -I 0 0
0
3
0
-1 0 0 0
-1 0 -I
-I -2
-I
-3
““Enhanced longevity” equals days to 50”b mortality on the SOjO carbohydrate each species. SE c 1.0 unless noted. bSee Table I for complete names of the Drosophila species. c7.59; solutions. d 1.8”; solutions. ‘3.6”, solutions.
Pentoses und L-rhamnose
Although no insects can survive very long on these sugars. adults of a few species are able to live 14 days longer on ribose and xylose solutions than on water alone (Hassett et al., 1950; Finch and Coaker, 1969: Tsiropoulos. 1980). This was also observed
D. nigro.
This work._ __. 0 -I 0 0 0 0 0 0 0 0 0
Hassett ( 1948) I< 0
0’
solutions minus days to 50% mortality on water alone for
with Drosophila in the present study (Table 2). D and L-arabinose were of little value to any of the species and lyxose and L-rhamnose to only I or 2 whereas ribose and/or xylose could be used by the majority of the flies to prolong their lives from 24 days longer than on water alone. D. aldrichi and D. arizonensis were able to utilize ribose and xylose the most
Table 3. Enhanced longevity’of Drosophila on 5”, solutions of carbohydrates species
that slgnilicantly prolong the adult lifespans of most of the
Species of Drosophila h
D. melanogas&r
Hasset So, Solutions of Glucose Fructose Galactose Mannose Sucrose Maltose Melezitose Trehalose Raffinose
D. aldr.
D. wheel.
I8 20 II I3 I8 16 I4 IO IO
19 29_+ 3.5 8 22& 3 34 16 21*7 30 2
.__-
D. mull.
18+4 20 8i3 I2 24 22 20 15 f 2.5 13
D. ark. __~~
25 25 8 1323 24k2.5 23 21 5 8
rp “N” ~_ ~~ D. ____~~
D. moja.
28 28 2 I8 29 25 I9 23f4 6
24 31 I5 I4 23 19 20 i 5 13 1
D. nigro.
~_~
This work ~_ (1948) ~____.
24 25 8 6 I8 I6 13 I6 8
24 28 I5 24 27 25 20 20 I6 + 3
“Enhanced longevity = SE $2.0 unless noted. “See Table I for complete names of each species. c~.8U, solutions. d3.6°:, solutions. Table 4. Enhanced longevity (in days) of Drosophila on 54” solutions of sugar alcohols, polysaccharides and melibiosea,b D. arironensis
Carbohydrate Adonitol (Ribotol) Ribose o-Arabitol L-Arabitol Dulcitol (Galactitol) Fructose Mannitol Glucitol Xylitol Xylose Maltitol Maltose Melibiose Arabogalactan Inulin Starch “Unusual values are underlined. bSE < I unless noted. ‘Hassett (1948). 1.80,, solutions.
_-~~ MS0 ~~~~22 + 1.5 4 2 2 1 25 w 34 lo 6 31 * 1.5 23 20 2 1 4
D. wheeleri
MS0 3 2 0
D. melanogaster’
_~_.
~?? 2 0
I 0 29 + 3 2 4 4 3 I1 I6 6 0 1 2
0 I6 4 3
I5 0 2
14’ 16’ 9 I? 22d 15d I2 19’ 13’
168
HENRY W. KIRCHER and BEDIR AL-AZAWI
efficiently. Their modest utilization of the two pentoses may reflect the ability of the flies to reduce them to ribitol and xylitol and then reoxidize the alditols to the metabolizable ribulose and xylulose. The lower longevity of the insects on D or t_-arabinose and lyxose may be due to their inability either to reduce these pentoses to arabitol and lyxitol or to oxidize the two pentitols to ribulose and xylulose respectively. This hypothesis is supported by several pieces of evidence from work with other insects. An aldehyde sugar reductase in the silkworm is able to reduce triose. tetrose and pentose but not hexose sugars to the corresponding alditols (Faulkner. 1958) and galactose to galactitol reductase occurs in the bollweevil (Nettles and Burks, 1971). The oxidation of sorbitol to fructose by an alditol oxidase occurs in mosquitos (Van Handel, 1969) as well as nine other insects (Wenzl, 1969). Therefore, reductions of aldoses such as ribose and xylose to the alditols and the oxidation of these to metabolizable ribulose and xylulose has a precedent in the literature. Unfortunately, the two ketopentoses are too expensive to be used in feeding trials. The utilization of pentoses and t-rhamnose by D. nigrospiracufa may reflect the chemical composition of its host cactus. Saguaro contains no free glucose. fructose or sucrose as the Opuntia do and instead contains glucose and rhamnose as glycosides. D. nigrospiraculu adults may have adapted to the utilization of unusual sugars because the common ones are not available to this species as they are to flies living on Opuniia cacti. Hexoses and inositol The values of M,, for glucose, fructose, galactose. and mannose are listed in Table 3. Those for L-sorbose. inositol, and the uranic acid derivatives of galactose and glucose are shown in Table 2. Of the first four hexoses, galactose was the least used and mannose was also poorer than glucose or fructose in prolonging life. D. mojauensis was unusual in that it lived only slightly longer on galactose than water alone and D. nigrospiracula could use mannose onehalf or less as well as the other seven species. Of the remaining 4 compounds (Table 2) L-sorbose. which prolonged the life of only one other insect to a small extent (Galun and Fraenkel, 1957) was of some benefit to D. nigrospiracula and D. wheeleri. It is metabolically inert because it cannot be reduced to sorbitol (Faulkner, 1958). Inositol could also be used to a small extent by four of the species of Drosophila and has been reported to be of some benefit to three other Diptera (Galun and Fraenkel. 1957: Finch and Coaker, 1969; Tsiropou10s. 1980). Uranic acids are usually not tested as energy sources for insects. Of the two, glucuronic acid supported three species while galacturonic acid supported only 1 for 24 days longer than the water controls. Di- and trisuccharides Data obtained with these sugars are shown Tables 2 and 3. Sucrose. maltose and melezitose clearly those that prolong life of Drosophila longest. Trehalose and raffinose are less well used cellobiose and lactose not all. Although lactose
in are the and can
prolong the life of 2 insects (Galun and Fraenkel, 1957; Finch and Coaker, 1969) the two p-linked disaccharides appear to be toxic to D. mojarrensis. These results are generally in accord with those obtained with other insects (Table 2; House, 1974). In our study, D. arizonensis lived for a considerably shorter time and D. wheeleri for a longer time on trehalose than the other 7 species. D. species “N” and D. wheeleri were exceptional in their poor usage of raffinose and all of the cactophilic species were less able than D. melnnogaster to use this sugar as an energy source. Melibiose, polwaccharides, multitol und other ulditols Data in Table 4 show longevities of D. arizonensis and D. wheeleri compared to D. melanogaster on solutions of these compounds. The two cactophilic species used maltitol as readily as maltose and their relative use of melibiose paralleled that of raffinose. Maltitol has not been previously tested with insects; melibiose is readily used by many (Fraenkel, 1940; Hassett, 1948; Webber, 1957: Galun and Fraenkel, 1957; Finch and Coaker. 1969 and Tsiropolous, 1980); an exception being the blowfly (Hassett et al., 1950). The ability of polysaccharides to sustain the life of adult insects varies with the polysaccharide and insect tested. Whereas dextrin could prolong the life of 7 insects (Fraenkel, 1940; Hassett. 1948; Webber, 1957: Galun and Fraenkel, 1957) and starch, the life of 3 (Webber, 1957; Gallun and Fraenkel, 1957). inulin and glycogen (except for I case. Tables 23) were generally inert. None of the 3 Drosophila in Table 4 were able to use the polysaccharides to significantly prolong their lives. Performance on sugar alcohols is also variable (Table 4). Those that can prolong the life of some insects, such as mannitol. sorbitol, adonitol and to a lesser extent, xylitol. as with D. urkonensis. are probably oxidized to fructose. ribulose and xylulose for subsequent utilization for energy (Van Handel, 1969; Wenzl, 1969). These alditols are also readily metabolized by mammals (Hood et al., 1977; Sipple and McNutt. 1974). whereas II- and t-arabitol and dulcitol. not usable by any insects (Fraenkel. 1940; Hassett et ul., 1950: Webber, 1957; Galun and Fraenkel, 1957: Finch and Coaker, 1969; Tsiropolous, 1980). are also poorly utilized by mammals. These findings also correlate with the modest usage of ribose and xylose by Drosophilu compared to that of II- and L-arabinose (Table 2). The longer life of D. urkonensis on adonitol and xylitol than on the respective aldoses, ribose and xylose, lends credence to the hypothesis that metabolism of these pentoses goes by the route: aldose-alditolLketose. Comparison of the three species in Table 4 shows a strong resemblance between D. wheeleri and D. melanogaster: they both differ considerably from n. arizonensis. This relationship is also reflected in the relative utilization of pentoses (Table 2). mannose (Table 3) and trehalose (Table 3) by the three species. Taken in their entirety, the data obtained in this study show that adults of eight species of Drosophilu resemble other insects in their utilization of carbohydrates to prolong their lives. In addition. the ability of the largest in size but shortest lived species, D.
Drosophila and sugars nigrospiracula, to use L-rhamnose, L-sorbose, inositol, galacturonic and glucuronic acids better than the other 6 cactophilic species (Table 2) may be related
to the fact that it is the only one that does not breed in Opuntiu cacti (Table I). The natural host of D. nigrospiracula, saguaro cactus, contains no detectable free sugars (glucose, fructose, sucrose) before decay, whereas the concentration of these sugars in prickly pear cacti is 612%. This may have forced D. nigrospiraculu to adapt to “less utilizable” sugars to sustain adult life. Acknowledgements-We thank Drs W. B. Heed, David Baldwin and J. C. Fogleman for fruitful discussions concerning this work and the National Science Foundation for grant DEB-8207056 for partial support. Arizona Agricultural Experiment Station Journal article no. 3795.
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