Heredity 72 (1994) 508-514 Received 22 September 1993 Genetical Society of Great Britain
Variation in wing length in Eurasian natural populations of Drosophila melanogaster
ALEXANDRA G. IMASHEVA*, OLEG A. BUBLI & OLEG E. LAZEBNY Vavilov Institute of General Genetics, Gubkin Street 3, 117809 GSP- 1, Moscow 8-333, Russia
Astudy of 16 natural populations of Drosophila melanogaster from Eastern Europe, the Caucasus and Central Asia has revealed a dine in wing length associated with geographical position of the populations. Wing length was shown to be positively correlated with temperature. The coefficient of variation in wing length was significantly different in town and orchard populations. The existence of a dine in wing length in the northern part of the species range and in the region where migration must be substantial suggests strong selection pressure acting in natural populations of D. melanogaster.
Keywords:dines,Drosophila melanogaster, genetic variation, natural populations, wing.
Introduction temperatures and dates of collection are given in Table 1. Since the size of the wing in natural populations of Amongnatural populations of Drosophila melano- Drosophila is subject to seasonal changes (see e.g. gaster considerable genetic differentiation in quantita- Tantawy & Mallah, 1961), we have restricted the tive characters has been demonstrated (Lemeunier et sampling period to the two hottest summer months at., 1986; David & Capy, 1988; Singh, 1989). Some (from mid-July to early September) when D. melano- morphological and physiological characters exhibit gaster is most abundant in the surveyed region. Collec- latitudinal dines in this species. One of them is a well- tions were made in the large commercial apricot or documented dine for numerous characters, including apple orchards situated in rural areas ('orchard' wing length, which extends from France to tropical samples), or on fruit stalls in city markets and in Africa (David & Bocquet, 1975a, b; Allemand & garbage bins outside wineries or fruit-processing facto- David, 1976; David etal., 1977; Cohet&David, 1980; ries ('town' samples). About 30 isofemale lines werç Cohet et at., 1980). Another dine for wing length and started for each of the localities with females taken abdominal bristle number has been reported in popula- from the sample at random. After oviposition the tions of the east coast of the U.S.A. (Coyne & females of the parental generation were removed from Beecham, 1987). vials and their left wings were measured. In this paper we present evidence for clinal variation From the progeny of each isofemale line five pairs of in wing length in wild populations of D. melanogaster flies were put into a separate vial, left for three days to that are located in the Palearctic zoogeographic region, lay eggs and discarded. To make the population density i.e. in the northern part of the species range. The exist- for the analysed flies uniform for all cultures, 60 eggs ence of three parallel dines in wing length is a strong from each vial were removed and placed in a vial with argument for an adaptive significance of variation in fresh medium. After emergence of adults they were this character. collected, five females chosen at random, their left wings removed, mounted on a slide and measured with Materialsand methods an ocular micrometer. This gave a total sample size of about 150 wings per locality. The length of the wing Theflies were collected in 16 localities of Eastern was measured as the distance from the outer margin of Europe, the Caucasus region and Central Asia (Fig. 1). the anterior crossvein along the third longitudinal vein The list of collection sites, their latitudes, longitudes, to the wing tip and given in micrometer units (1 unit=0.014 mm). All cultures were kept on standard Drosophila *Correspondence medium at 2 5°C, the physiologically optimal tempera- 508 WING LENGTH VARIATION IN DROSOPHILA 509
Fig. 1 Collection sites. For explanation, see Table 1.
Table 1 Populations of Drosophila melanogaster used in this study
Locality Country Latitude (N) Longitude (E) Tmaxt Tyeart Habitat Timeof sampling
Tartu (Tar) Estonia 58°23 26°44' 17.4° 5.0° Town Aug. 1989 Rybnoe(Ryb) Russia 54°43' 39°30' 18.4° 3.8° Orchard Jul.1989 PyanyRog(Pya) Russia 52°54' 33°35' 19.0° 5.3° Orchard Aug. 1990 Gomel(Gom) Byelorussia 52°26' 30°59' 18.5° 6.1° Town Sep.1991 Khristinovka (Khr) Ukraine 48°49' 29°58' 19.2° 7.3° Orchard Aug. 1991 Kishinev (Kis) Moldavia 47°01' 28°50' 21.7° 9.5° Orchard Aug. 1990 Zabuzan(Zab) Russia 46°32' 48°18' 25.5° 9.4° Orchard Sep.1991 Astrakhan(Ast) Russia 46°22' 48°03' 25.3° 9.4° Town Sep.1990 Vilino(Vil) Ukraine 44°52' 33°36' 23.0° 11.3° Orchard Jul.1989 Alma-Ata (Aim) Kazakhstan 43°16' 77°57' 23.4° 8.7° Town Jul. 1991 Akzhar (Akz) Kazakhstan 43°07' 71°38' 23.6° 9.1° Orchard Jul. 1992 Jambul (Jam) Kazakhstan 42°54' 71°23' 23.6° 9.1° Town Aug. 1991 Gergebil(Ger) Russia 42°31' 47°04' 18.3° 7.2° Orchard Jul.1992 Makhinjauri (Mak) Georgia 41°40' 41°42' 22.0° 14.5° Orchard Jul. 1990 Cheptura (Che) Tajikistan 38°32' 68°21' 27.9° 14.2° Town Sep. 1991 Bezmein (Bez) Turkmenistan 38°03' 58°1 1' 30.3° 16.4° Town Aug. 1992
Average temperature of the hottest calendar month. 4Average year temperature. 510 A. G. IMASHEVA ETAL. ture for this species (David et at., 1983). Although in ANOVA for laboratory-reared flies are presented in the present study there was not much danger of con- Table 3. fusing D. melanogaster with its sibling species D. Variation among populations is highly significant, simulans, since the latter is rarely found in the region showing that populations are geographically differen- covered by the survey (Imasheva, A. G. et al., tiated according to wing length. The isofemale lines unpublished data), all F1 cultures were checked by also contribute significantly to the variation. This is to examining males; no D. simulans individuals were be expected since genetic determination of wing found. length in Drosophila is a firmly established fact, and Temperature data for weather stations adjacent to heritability of this character estimated in laboratory collection sites were obtained from the Committee for conditions is high, varying from 0.2 to 0.6 (see e.g. Meteorology of the Russian Federation (Rosgidromet). Robertson & Reeve, 1952; Reeve & Robertson, 1953; The distance from collection sites to weather stations Latter & Robertson, 1962). was in all cases less than 50 km (20 km on average). The data in Table 2 clearly show that the average Two temperature variables were used: average year wing length in the populations under study decreases temperature (Tyear) and average temperature of the from north to south and from west to east, thus forming hottest calendar month (Tmax; July for all localities a dine. This is confirmed by regression analysis, which except Makhinjauri for which it was August); for both demonstrated a significant relationship between wing parameters, 10-year averages (1980—1990) were used. length and latitude and longitude of populations Temperature values are listed in Table 1. (Fig. 2a, b). Regression slopes were significantly Statistical analysis was done using SYSTAT (version different from zero both in wild-caught parents 4.1). (latitude: b=0.782, P<0.01, r2=0.498; longitude: b= —0.294, P<0.001, r2=0.618) and in labora- tory-reared offspring (latitude: b =0.188,P <0.001, = longitude: b =— 0.062,P< 0.001, r = Results 0.595; 0.574). As can be seen from Fig. 2, the regression slope is In Table 2 average wing lengths and coefficients of steeper in the wild-caught flies compared with the variation are given for wild-caught (P) and laboratory- laboratory-reared ones, which must be due to the fact reared F2 flies from the 16 studied populations (to that in nature flies live in a variety of habitats while in eliminate the dependence of variance on the mean laboratory cultures environmental conditions are value of wing length we have used coefficients of varia- uniform. tion rather than variance as the measure of within- No significant association with altitude was found population variation). The results of the nested for either wild-caught or laboratory-reared flies (the
Table 2 Mean wing length ()andcoefficient of variation (CV) in 16 Eurasian populations of Drosophila inelanogaster
Wild-caught flies (P) Laboratory-rearedflies (F2)
Population Habitatn S.E, CV (%) n S.E.CV (%)
Tartu Town 55116.820.70 4.44 150114.030.30 3.26 Rybnoe Orchard53115.530.95 5.98 150113.500.33 3.53 PyanyRog Orchard50119.740.61 3.60140112.860.23 2.43 Gomel Town60116.850.95 6.32 150113.150.28 3.02 KhristinovkaOrchard60116.630.99 6.57150114.550.23 2.46 Kishinev Orchard45114.641.07 6.24150112.680.22 2.37 Zabuzan Orchard55102.931.03 7.44150111.450.24 2.67 Astrakhan Town 33113.640.55 2.77150112.020.28 3.07 Vilino Orchard53110.811.30 8.57150111.200.28 3.11 Alma-Ata Town 60105.071.32 9.75150111.490.25 2.80 Akzhar Orchard60106.730.82 5.94150110.290.22 2.45 Jambul Town 5598.241.41 10.65135111.310.30 3.12 Gergebil Orchard29106.380.99 5.00145112.860.23 2.47 MakhinjauriOrchard27114.001.68 7.67135112.660.20 2.09 Cheptura Town60108.451.20 8.57150109.850.41 4.56 Bezmein Town50101.780.87 6.02150109.940.29 3.25 WING LENGTH VARIATION IN DROSOPHILA 511 results of regression analysis are not presented), of laboratory-reared populations and the hottest calen- probably because none of the populations under study dar month temperature, Tmax (b =0.330,P
a b l2 120 o/ 120' 0 0 0/0 0 0 E115 0 E 8 —4 110 0 / 110 0 o ••". 0 II 105 0 105 0 0 0 0 0 -4100 100 0 0
--cc 95 35 40 45 50 55 20 30 40 50 60 70 80 Latitude (°North) E Longitude (°East) C d L) IIC l2
120 0 120 0 .a-(2 O 000 115 0 1l5". • 0 0
110 110 bJ ...' 0 Fig. 2 The relationship of wing length 0 0 0 in wild-caught (open circles) and 105 0 105 0 laboratory-reared (solid circles) popu- 0 •' 0 lations of Drosophila melanogaster to 0 100 100 latitude (a), longitude (b), temperature 0 0 of the hottest calendar month (c) and average year temperature (d). Regres- Qc Qc sion lines: dotted, wild-caught; solid, 16 1820 2224 '26 28 30 32 2 4 6 8 10 12 14 16 18 laboratory-reared. Hottest month temperature (° C) Average year temperature (°C) 512 A. G. IMASHEVA ETAL.
Table 4 Linear multiple regression of wing length on Table 5 Kruskal—Wallis test for differences in mean wmg geographical and temperature variables length ()andcoefficient of variation (CV) of wild-caught(P) and laboratory-reared (F2) flies from different habitats Laboratory-reared Wild-caught flies (P) flies (F2) Average rank Average rank off ofCV Variables b R2 b Sample Habitat size P F2 P F2 Latitude 0.638 0.019 Longitude —0.127 0.700** —0.026 0.843*** Orchards 9 9.33 9.44 7.94 6.00 Tmaxl —0.663 Towns 7 7.43 7.29 9.21 11.71 Tyeart 0.851 —0.001 Test statistic, H 0.63 0.81 0.28 5.67* b, partial regression coefficient; R2, coefficient of determination. * <0.05. *<0.05,**<0.01,***<0.001. tAverage temperature of the hottest calendar month. tAverage year temperature. The selective factor that forms these dines is almost certainly temperature. Whenever it is colder flies' wings are larger; this is evident in seasonal variation (Tantawy A comparison of populations occupying different & Mallah, 1961) and in association with altitude habitats showed that average wing length was not (Stalker & Carson, 1948; Louis etal., 1982). This view influenced by habitat either in wild-caught or in labora- is also confirmed by laboratory selection experiments tory-reared flies. The same was shown for variation in where flies kept at a colder temperature for many wild-caught flies. In laboratory-reared flies, however, generations eventually acquirebigger wings coefficients of variation were dependent on the habitat, (Anderson, 1966, 1973; Powell, 1974). being on average higher in populations sampled in In our experiment, wing length showed significant fruit-processing plants and in markets, and lower in correlation with both temperature parameters used in those sampled in fruit orchards. The difference the analysis, but the association with the temperature of between them was significant as shown by the Kruskal- the hottest month, Tmax, was much stronger than that Wallis test (Table 5). with the average year temperature, Tyear. In view of the interpretation involving selection, these results seem Discussion quite reasonable. Tmax is an indicator of the tempera- ture of the warm season when selection must operate Geographicaldines similar to the one we have most strongly, since it is at this time that D. melano- revealed theoretically could result from two causes gaster populations are most abundant and live out- (Endler, 1977). One is the pressure of natural selection doors. On the other hand, the Tyear parameter includes associated with some climatic variable, which in turn temperatures of colder months when in northern depends on the geographical position of the popula- regions D. melanogaster overwinters indoors and is not tion. The alternative explanation is the interaction of subjected to factors of the external environment. genetic drift and migration. At present there is little Though temperature seems to be generally accepted doubt that latitudinal dines in wing length —andin as a selective agent in this case, the actual mechanism body size in general —inDrosophila are caused byof selection is not yet understood. Since wing size is selection. The dine demonstrated in the present study positively correlated with body size (Reeve & Robert- (as has been) is the third reported in D. melanogaster. son, 1953), it is presumed that selection works directly Similiar dines more or less well documented have been on body size. Bergmann's rule (body size increases with demonstrated for several other Drosophila species: D. latitude) is usually invoked though it does not really robusta (Stalker & Carson, 1947), D. subobscura clarify the issue, since in poikilothermic animals like (Prevosti, 1955; Misra & Reeve, 1964; Pfriem, 1983), Drosophila it is not clear how it works. A review of D. virilis (David & Kitagawa, 1982) and D. simulans possible mechanisms of selection for size by tempera- (Hyytia et al., 1985). ture is given by Coyne and Beecham (1987). One In all of them there was positive correlation of wing hypothetical explanation discussed by them is selection size with latitude: the wings in the northern populations connected with developmental time of flies. were larger. It is extremely unlikely that such a number Since at higher temperatures flies develop faster and of parallel dines can result from chance events. are smaller, selection favouring rapid development WING LENGTH VARIATION IN DROSOPHILA 513
could lead to smaller fly size. This hypothesis is con- laboratory-reared flies was about 12, which is close to firmed by the results of Cohet et al. (1980) who a previous estimate of 15 for a French natural popula- demonstrated faster developmental rate for African tion of D. melanogaster (David, 1979). The low esti- populations of D. melanogaster compared with Euro- mate of this ratio reported by Coyne & Beecham pean ones. However, our preliminary experiments have (1987) for populations of the east coast of North not shown any differences between time of develop- America (between 1.5 and 3, depending on the ment of Central Asian and European populations temperatures at which laboratory flies were raised) (Bubli & Imasheva, 1994). There is also a possibility seems puzzling as it is not obvious what differences that selection might act on wing length in a way could exist between the ecology of the sampling sites in connected with efficiency of flight (Stalker, 1980) or their case and ours. with thermoregulation via the wing surface (Kingsolver One further point to consider is the difference in &Koehl, 1985). variation in populations occupying different habitats. The two previously reported dines for D. melano- Although mean wing length did not differ in this case, gaster (David et al., 1977; Coyne & Beecham, 1987) the coefficient of variation was higher in populations concerned populations inhabiting much more southern sampled in the town areas (wineries, fruit-processing areas with climatic conditions ranging from mild to hot. factories and markets) compared with large gardens Our survey includes localities from as far north as situated in the rural areas. This might be due to a Estonia, which is probably the extreme northern edge greater genetic heterogeneity of the town samples, of the range of the species (David & Capy, 1988). The which could consist of a mixture of flies transported fact that a stable dine exists so far north and in such there from different neighbouring localities, or to the drastic temperature conditions supports the view that fact that the town habitats are less uniform. populations of D. melaongaster living there are not a result of frequent (each season) recolonization from warmer regions, but are continuously living and repro- Acknowledgements ducing there. Wethank Jerry Coyne, Boris Kalabushkin, Yuri A noteworthy fact is the existence of a dine over a Dubrova and anonymous reviewers for critical rather small geographical distance in a cosmopolitan comments on the earlier version of the manuscript. domestic species like D. melanogaster. Even supposing The work is supported by a research grant from the that by active migration D. melanogaster does not Programme 'Frontiers in Genetics' of the Russian cover hundreds of miles (though there is evidence to Federation. the contrary: see Coyne et al., 1982; Coyne & Milstead, 1987; Coyne et al., 1987), the passive trans- portation (with food products, etc.) must be substantial, References especially in the region in question, where rotten fruits and vegetables are often transported by train in mass ALLEMAND,R. AND DAVID, J. R. 1976. The circadian rhythm of oviposition in Drosophila melanogaster: A genetic latitu- quantities. The existence of a dine in such conditions dinal dine in wild populations. Experientia, 32, proves the existence of strong selection pressure. 1403-1404. The regressions of wing length on geographical and ANDERSON, w. w. 1966. Genetic divergence in M. Vetukhiv's temperature parameters in wild-caught flies were experimental populations of Drosophila pseudoobscura. 3. stronger than those in laboratory ones, i.e. the dines in Divergence in body size. Genet. Res., 7, 255—266. the wild-caught flies were much steeper. This is to be ANDERSON, W. w. 1973. Genetic divergence among experi- expected because of the effect of local temperature on mental populations of Drosophila pseudoobscura kept at individual development. Flies from northern parts of different temperatures. Evolution, 27, 278—294. the range have an additional source of variation since BUBLI, 0. A. AND IMASHEVA, A, G. 1994. Genetic differentiation they develop in a cold habitat which causes larger body of three quantitative traits in Drosophila melanogaster size (and vice versa for flies from southern parts of the natural populations of Eastern Europe and Central Asia. Russian J. Genet., 30, 57—63. range). Note, though, that in D. melanogaster popula- COHET, V. AND DAVID, J. R. 1980. Geographic divergence and tions of the east coast of North America, the opposite sexual behaviour: Comparison of mating systems in trend was found: the latitudinal dine in wing length was French and Afrotropical populations of Drosophila shallower in wild-caught flies compared with their melanogaster. Genetica, 54, 161—165. laboratory progeny (Coyne & Beecham, 1987). COHET, Y., VOUIDIBIO, J. AND DAVID, J. R. 1980. Thermal tolerance As expected, phenotypic variation of wing length and geographic distribution: A comparison of cosmopoli- was much higher in wild-caught than in laboratory- tan and tropical endemic Drosophila species. J. Therm. reared flies. The ratio of the variances of wild-caught to Biol., 5,69—74. 514 A. G. IMASHEVA ETAL.
COYNE, J. A. AND BEECHAM, E. 1987. Heritability of two ential scaling and evolutionary change. Evolution, 39, morphological characters within and among natural 488—504. populations of Drosophila melanogaster. Genetics, 117, LATFER, B. 0. H. AND ROBERTSON, A. 1962. The effects of 727—737. inbreeding and artificial selection on reproductive fitness. COYNE, J. A. AND MILSTEAD, B. 1987. Long-distance migration of Genet. Res.,3, 110—138. Drosophila. 3. Dispersal of D. melanogaster alleles from a LEMEUNIER, F., DAVID, J. R., TSACAS, L. AND ASHBURNER, M. 1986. Maryland orchard. Am. Nat., 130, 70—82. The melanogaster species group. In: Ashburner, M., COYNE, J. A., BOUSSY, J. A., PROUT, T., BRYANT, S. N., JONES, J. S. AND Carson, H. L. and Thompson, J. N. (eds) The Genetics and MOORE, j.A.1982. Long-distance migration of Drosophila. Biology of Drosophila vol. 3, pp. 147—256. Academic Am. Nat., 119, 589—595. Press, London. COYNE,J. A., BRYANT, S. N. AND TURELLI, M. 1987. Long-distance LOUIS, J., DAVID, J. R., R0UAuLT, I. AND CAPY, P. 1982. Altitudinal migration of Drosophila. 2. Presence in desolate sites and variations of Afro-tropical D. melongaster populations. dispersal near a desert oasis. Am. Nat., 129, 847—861. Dros. Inf Serv., 58, 100—101. DAVID, J. R. 1979. Utilization of morphological traits for the MISRA, R. K. AND REEVE, E. C. R. 1964. Clines in body dimensions analysis of genetic variability in wild populations. Aquilo in populations of Drosophila subobscura. Genet. Res., 5, Ser. Zool., 20, 41—6 1. 240—256. DAVID, J. R. AND BOCQUET, c. 1975a. Evolution in a cosmopoli- PFRIEM, p 1983. Latitudinal variation in wing size in Droso- tan species: genetic latitudinal dines in Drosophila phila subobscura and its dependence on polygenes of melanogaster wild populations. Experientia, 31, 164—166. chromosome 0. Genetica, 61, 221—232. DAVID,J. R. AND BOCQUET, c. 1975b. Similarities and differences POwELL, j.R.1974. Temperature related genetic divergence in in latitudinal adaptation of two Drosophila sibling species. Drosophila body size. J. Hered., 65, 257—258. Nature, 257, 588—590. PREvO5TI, A. 1955. Geographical variability in quantitative DAVID, j.R.AND KITAGAwA, a. 1982. Possible similarities in traits in populations of Drosophila subobscura. Cold ethanol tolerance and latitudinal variations between Spring Harbor Symp. Quant. Biol., 20, 294—299. Drosophila virilis and D. melanogaster. Jpn J. Genet., 57, REEVE, E. C. R. AND ROBERTSON, F. w. 1953. Studies in quantita- 89—95. tive inheritance. II. Analysis of a strain of Drosophila DAVID, J. R. AND CAPY, P. 1988. Genetic variation of Drosophila melanogaster selected for long wings. J. Genet., 51, melanogaster natural populations. Trends Genet., 4, 276—316. 106—111. ROBERTSON, F. W. AND REEVE, E. C. R. 1952. Studies in quantita- DAVID, J., BOCQUET, C. AND DE SCHEEMAEKER-LOUI5, M. 1977. tive inheritance. I. The effects of selection on wing and Genetic latitudinal adaptation of Drosophila melanogaster: thorax length in Drosophila. J. Genet., 50, 4 14—448. new discriminative biometrical traits between European SINCH, R. s. 1989. Population genetics and evolution of species and equatorial African populations. Genet. Res., 30, related to Drosophila melanogaster. Ann. Rev. Genet., 23, 247—255. 425—453. DAVID, J R., ALLEMAND, R., VAN HERREWEGE, J. AND COT-JET, V. STALKER, H. D. 1980. Chromosome studies in wild populations 1983. Ecophysiology: abiotic factors. In: Ashburner, M., of D. melanogaster. II. Relationship of inversion frequen- Carson, H. L. and Thompson, J. N. (eds) The Genetics and cies to latitude, season, wing-loading and flight activity. Biology of Drosophila, vol. 3, pp. 105—170. Academic Genetics, 95, 211—223. Press, London. STALKER, I-I. D. AND CARSON, H. L. 1947. Morphological variation ENDLER, .1. A. 1977. Geographic Variation, Speciation and in natural populations of Drosophila robusta Sturtevant. Clines. Princeton University Press, Princeton, NJ. Evolution, 1, 237—248. HYYTIA, P., CAPY, P., DAvID, J. R. AND SINGH, R. s. 1985. Enzymatic STALKER, H. D. AND CARSON, H. L. 1948. An altitudinal transect and quantitative variation in European and African of D. robusta Sturtevant. Evolution, 2, 295—305. populations of Drosophila simulans. Heredity, 54, TANTAWY, A. 0. AND MALLAH, G. s. 1961. Studies in natural 209—2 17. populations of Drosophila. I. Heat resistance and KINOSOLVER, J. G. AND KOEHL, M. A. R. 1985. Aerodynamics, geographical variation in Drosophila melanogaster and D. thermoregulation, and the evolution of insect wings: differ- simulans. Evolution, 15, 1—14.