Male size and mating success in Drosophila melanogaster: the roles of male and female behaviour

Anim. Behav. 1987, 35, 555 562

Male size and mating success in Drosophila melanogaster: the roles of male and female behaviour LINDA PARTRIDGE, ARTHUR EWING & AMANDA CHANDLER

Department of Zoology, Edinburgh University, West Mains Rd, Edinburgh EH9 3JT, U.K.

Abstract. Larger D. melanogaster males delivered more courtship to virgin females, produced more courtship song and sang more loudly than smaller males. They also moved around more when not courting. All courtships were terminated by the female decamping or while she was moving. Females decamped equally often when courted by large and small males but were more likely to be moving while courted by large males, and this appeared to be a consequence of the faster running and courtship tracking speeds of larger males. The results suggest that scramble competition between males to deliver courtship is important in determining mating success, but not that females discriminate between males of different sizes. However, the higher audibility of louder sounds to females and their tendency to move while courted both have the effect of favouring larger males.

Ever since Darwin (1871) first proposed his theory of sexual selection, the role of female discrimination in producing variation in male mating success has been contentious. Well-documented cases where success in various forms of male rivalry is important in determining male mating success are now numerous (e.g. Cox & le Boeuf 1977; Davies & Halliday 1978, 1979; Appleby 1982; Johnson 1982; Otronen 1984). However, few studies have examined female discrimination, and good evidence for female preferences for particular male phenotypes is rare (but see Andersson 1982; Majerus et al. 1982 a, b; O'Donald & Majerus, in press). Part of the reason for the paucity of data on female preferences may lie in the very prevalence of male rivalry, which can obscure the effects of female behaviour (Borgia 1981; Halliday 1983; Partridge & Halliday 1984). One way to begin to overcome this problem is to find cases where non-random mating persists in the absence of any opportunity for male rivalry, and this is the approach used in the present study. In an evolutionary sense female preference can include any behaviour that affects the identity of her mate, and we investigate both these and male behaviours producing the higher mating success of large D. melanogaster males. A correlation between male size and mating success has been a common finding in studies of sexual selection (e.g. Davies & Halliday 1978, 1979; Berven 1981; Borgia 1981; Johnson 1982). In Drosophila melanogaster male size has been shown

to correlate with short-term measures of mating success in the field and laboratory (Ewing 1961, 1964; Ewing 1978; Partridge et al. 1987) and with lifetime measures in the laboratory (Partridge & Farquhar 1983; Partridge, in press). Part of the reason for the higher mating success of larger males may lie in their ability to win fights with smaller males (Partridge & Farquhar 1983), since success in fights has been shown to be a determinant of male mating success (Jacobs 1960; Dow &von Schilcher 1975). However, this cannot be the whole explanation because larger males achieve mating more quickly when allowed individually to court single virgin females (Partridge & Farquhar 1983). A result of this kind need not imply that female discrimination is involved. For instance, females might have a constant probability of mating with any courting male and, if larger males delivered more courtship, females would inevitably mate more often with them. To demonstrate that females prefer large males it would be necessary to show either that females behave differently towards males of different size classes in a way that affected their mating success or that females behave in a way that has the effect of favouring matings by larger males. The aim of the present study was to examine the roles of male and female behaviour in producing the higher courtship success of larger males. To do this we quantified the visual and acoustic aspects of the courtship of males of different sizes, and

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observed female behaviour towards courting males. We also investigated the ability of males of different sizes to keep in contact with a moving female during courtship.

thorax lengths were measured on the day before the courtship observations, again using carbon dioxide for immobilization.

Courtship Song M A T E R I A L S AND M E T H O D S The stock used was an outbred one collected in Dahomey in 1970 and maintained since in population cage culture (see Partridge & Farquhar 1983 for details). The size variation in the flies was produced by the decline in mean size and increase in size variability as the adult emergence proceeds in the population cage culture bottles, and was therefore mainly environmental in origin.

Courtship Observations We made observations of the courtship behaviour of 76 males of different sizes. To do this, we introduced a single pair of flies by aspiration into a circular Perspex courtship chamber 38 mm in diameter and 16 mm deep, with a disc of moistened filter paper, 21 mm in diameter, in the centre of the floor. We chose this size of cell both to allow females to escape from courting males (Ewing & Ewing 1984) and to ensure that the pair came into reasonably frequent contact. We then used an electronic bleeper to time a taped vocal list of the behaviour of the male and the female every 2 s for a total of 3 min. We included in courtship the following behaviour elements which correspond to the descriptions of Connolly & Cook (1973) and Ewing & Ewing (1984): (1) male: orientate, follow, vibrate, lick, attempted copulation; (2) female: kick, extrude, decamp (jump or fly off). For both sexes, we categorized behaviour during those periods of time when the pair were out of contact as either still or moving. Courtships were always observed in the first 2 h after lights on (the flies were kept on an L : D 12 : 12 cycle), and observations on large (greater than 0.84 mm thorax length) and small (less than 0.76 mm thorax length) males were alternated within each observation period to control for any effect of day and time, because both of these variables are known to affect fly behaviour. All females were large (greater than 0'92 mm thorax length), and the effects of female size on courtship behaviour were not investigated. All flies were virgins, the females 2 days old, the males 3 days old, and were collected under carbon dioxide anaesthesia more than 3 h after eclosion. The

The songs of 41 males of differing sizes were recorded. Flies were collected and measured as described for courtship observations above. One 3day old male and one large 2-day old virgin female were aspirated into a copper wire mesh cell, measuring 20 mm x 8 mm x 5 mm, covered with a Perspex lid which was placed over a ribbon microphone connected via an amplifier to a tape-recorder (see Bennet-Clark & Ewing 1968 and Ewing 1979 for further details of the apparatus). The flies were allowed 1 min to settle and, provided some courtship had occurred during that time, 2 min of courtship were then recorded. The tapes were played back through an oscilloscope, and the trace filmed onto Kodak linagraph paper. The pulse song (Bennet-Clark & Ewing 1968) of all males during the 2-min period was filmed at a speed of 80 mm/s. Occasional pulses were higher than the width of the film, and these were re-filmed at half the gain. Since sine song (von Schilcher 1976) is hard to detect reliably on traces filmed in this way, we re-filmed 1 min of the song of 18 males at 100 mm/s and higher gain. The filmed song bursts were measured to the nearest 1 mm using a ruler. A pulse song burst was considered to end when the distance to the next pulse was more than twice that between the previous two, while a sine song burst consisted of any unbroken period of song. The number of pulses in each burst of pulse song was counted, and used to calculate the mean interpulse interval for each song burst. The median of these means was then calculated for each male (median interpulse interval). The courtship songs are produced by the wings of the male, and in males where the size of the wing is surgically reduced there is an association between wing area and mating success (Ewing 1964). To determine whether larger males, because of their larger wings, sing more loudly than small males, the height of the highest pulse in each pulse song burst and the maximum height of each sine song burst were measured to give the maximum amplitude of song in each burst, and the median of these measurements was again calculated for each male (the median maximum amplitude). No attempt was made to measure intrapulse frequency of pulse song or the frequency of sine song.

Partridge et al.: Mating success in Drosophila

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Figure l. The apparatus used to measure courtship tracking and runniqg speeds, from above on the left and from the side on the right. The female model is suspended by a wire (W) waxed to her thorax.

Running and Courtship Tracking Speeds The apparatus used to measure running and courtship tracking speed is illustrated in Fig. 1. It consisted of an annulus milled from Perspex covered with a transparent Perspex lid. The annulus was rotated by a D.C. motor whose speed was controlled by varying the voltage. A digital voltmeter showed the applied voltage from which the speed of rotation could be calculated. For measurement of courtship tracking speed, a female with wings and legs amputated was suspended from a fine wire inside the annulus as shown, and a male was introduced into the annulus. The annulus was rotated slowly and the male usually started courting soon after contacting the female. Once courtship was established the speed of rotation was increased until (1) the male stopped courting and following the female (this usually happened quite abruptly) or (2) the male fell behind and out of the view of the microscope, an angle of about 3 0 (see Fig. 1). Under these circumstances males courted and sang vigorously. Black spokes were painted on the annulus to provide visual feedback of movement for the fly. A similar method was used to measure the maximum running speeds of non-courting flies of both sexes, but here we were measuring a phototactic response, the flies running towards a light source at one side of the annulus. The same criteria as for courtship tracking were used for maximum running speed. In one experiment, three running speeds were measured for 39 males of differing sizes and for 15 large females, while in a second we measured courtship tracking and running speeds from 29 large males, attempting to obtain five measurements of each although the number sometimes fell below this.

RESULTS In all statistical tests two-tailed probability values are given.

Courtship Observations The data were analysed as matched pairs of consecutive courtships by large and small males, to minimize variance caused by the time at which observations were made. The large male delivered more courtship than the small male in 29 of the 38 comparisons (sign test, P=0.002). This difference may have reflected the higher levels of movement of the larger males when not involved in courtship (more movement by the large male in 23 out of 38 pairs, five cases were omitted because of a tie or because one male was always involved in courtship, sign test P = 0"04), To determine whether the difference in the amount of courtship delivered by males of different sizes could be accounted for in terms of female behaviour towards them, we established that every bout of courtship was terminated either by the female decamping or while she was moving, and v~c checked if females showed either of these behaviours more when courted by small males. For decamps we compared the ratio of number of decamps to amount of time spent courting for each of the inatched pairs of large and small males. In 16 pairs there were either no decamps to either male or there was a tie because the observed and expected number of decamps for one male were zero. In the remaining 22 matched pairs the female decamped fi'om the larger male at a higher rate than the small one in l0 and at a lower rate in t2 (sign test, Ns). Females therefore did not decamp very frequently, and they did so from large and small males at

Animal BehaL'iour, 35, 2

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similar rates. We investigated female movement by comparing matched males for the proportion of their courtship during which the female was moving. In 25 out of 38 pairs (one tie) the female was more likely to be moving when courted by a large male (sign test, P=0.0488). This result does not necessarily mean that female movement was a more likely response to the courtship of large males; these may simply have been more effective at tracking a moving female, as is implied by our results for tracking speeds (see below).

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Figure 2. (a) The percentage of the 2-min recording period spent producing pulse song by males of different sizes. (b) The number of pulse song bursts produced in 2 min by males of different sizes. (c) The median burst length (ms) of pulse song produced by males of different sizes. (d) The median maximum amplitude (mV) of pulse song produced by males of different sizes.

Partridge et al.. Mating success in Drosophila

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There was a significant positive correlation between male thorax length and mean maximum running speed (Fig. 4; r~= 0.383, P < 0.02). Mean maximum courtship tracking speeds were without exception lower than mean maximum running speeds in the same individual males (sign test, P < 0.001), and the two showed a significant positive correlation with each other (Fig. 5; r~ = 0461,

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P<0.001) by larger males, There was also a significant correlation between male thorax length and the median maximum amplitude of pulse song (Fig. 2d; r~-- 0.591, P < 0.001 ) but not with median interpulse interval (r~ = 0.128, Ns). There was a positive correlation between male thorax length and amount of time spent producing sine song (Fig. 3a; r~=0.805, P<0"02), and this association was a consequence mainly of a greater number of bursts of sine song by larger males (Fig. 3b; r~=0.814, P<0.02), the correlation between thorax length and burst length being non-significant (r~=0-449). The correlation between thorax

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P<0-02). The running speeds of courting males were probably lower because they engaged in almost continuous wing vibration, although exactly how this lowers running speed is unknown. The mean maximum running speeds of full-sized females were significantly greater than those of males (female median=18-8 mm/s, male median = 15.8 mm/s, P < 0.001 median test).

DISCUSSION Large males showed more courtship activity than did small ones, and they delivered more of both types of courtship song. This appeared to be at least in part a purely male effect, because large males were more active generally in that they moved around more when out of contact with the female, and this probably also brought them back into contact with the female sooner after breaks in courtship. In addition, during well over half our courtship observations the female was moving, and breaks in courtship were most likely to occur during these times. Large males were able to run faster than small ones, males ran more slowly when courting and females could run faster than males. It therefore seems overwhelmingly likely that part of the reason large males deliver more courtship is that they are better able to keep up with the female when she moves. Scramble-type competition between males to find, pursue and court females is therefore implicated in producing the higher mating success of larger males, and there is hence almost certainly selection on males to deliver high levels of courtship. A reduction in the amount of courtship delivered has been implicated in the lower mating success of mutant yellow males (Bastock 1956) while the higher mating success of males heterozygous for the mutant ebony is associated with elevated levels of courtship (Kyriacou et al. 1978). In species such as some Orthoptera and anuran amphibians where females approach singing males for matings, the amount of calling by individual males is often associated with their mating success (e.g. Whitney & Krebs 1975a, b; Butlin et al. 1985), perhaps because frequent calling makes the male more likely to be singing alone and hence easier for the female to locate (Whitney & Krebs 1975a, b). The role of the female in producing the higher mating success of larger males is far more difficult to disentangle. For instance, larger males produced

louder pulse song than did small ones and this could affect the female in two ways. The song is perceived by the female as particle velocity which attenuates extremely rapidly, as the sixth power of distance from the source (Bennet-Clark 1975). As the distance of the male from the female varies during courtship, large males are therefore likely to be audible to the female for more of the time. We would then be dealing with another case of scramble competition between males, this time to deliver audible song. On the other hand, females may be more likely to mate in response to louder sounds, and would then be exerting some discrimination between males on the basis of their song volume. To discover the relative roles of these two processes one would have to relate song loudness to both song perception in the female and her probability of mating, which would be extremely difficult with existing techniques. Where females will approach one of several simultaneously singing males it is a much simpler matter to investigate female discrimination of sound volume. For instance, experiments where female anurans are placed equidistant from two loudspeakers producing different volumes of male song have established that females selectively approach the speaker producing the higher volume (Fellers 1979; Arak 1983). A similar difficulty occurs in interpreting the data on female movement when courted by large and small males. Females may discriminate in favour of larger males by being more likely to move in response to courtship by smaller males, but detecting this against a background of differences in tracking ability between males is problematic. It is even difficult to establish whether females respond to courtship by moving, because female movement is a stimulus to male courtship (Tompkins et al. 1982), so that a transition from immobility to movement by the female at the time courtship commences or an association between movement by the female and courtship could occur for either reason. Because small males are less able to track the female, movement by her during courtship increases the likelihood that she will be courted by larger males, without any need for her to discriminate between males. In an evolutionary sense, movement would therefore qualify as a potential mechanism of female preference for large males. However, the evolution of female movement during courtship may have been unrelated to its effect on male mating success, and could instead indicate that she is unreceptive or in search of food.

Partridge et al.: Mating success in Drosophila In the present study, the only female b e h a v i o u r which could have produced unequivocal evidence for discrimination, namely decamping, failed to do so, so our results do not support any involvement of female discrimination a l t h o u g h they c a n n o t dismiss it. The difficulties in establishing the exact role of the female evcn in this very simplified system highlight some of the problems associated with attempts to detect female discrimination. The greater audibility of louder sounds favours males with large wings and, since there is a strong genetic and e n v i r o n m e n t a l correlation between thorax and wing length ( R o b e r t s o n & Reeve 1952), an increase in thorax length and p r o b a b l y other aspects of body size will be favoured. It is not clear exactly why larger flies can run faster, but again large body size in males is p r o b a b l y thereby favoured. Genetic models of sexual selection by female choice all involve a genetic correlation between the female preference a n d the preferred male character ( O ' D o n a l d 1967, 1980; Lande 1981 ; Kirkpatrick 1982), a n d an investigation of this might prove profitable by examining the genetic correlation between, for instance, female movement and male body size. However, the present behavioural evidence does not suggest that an explanation of female b e h a v i o u r in terms of sexual selection is needed, and the scope for female preference for large males is greatly limited by the effects of male behaviour. ACKNOWLEDGMENTS We t h a n k the S E R C and the Royal Society of L o n d o n for financial support.

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(Receit'ed 10 Fehrual3' 1986," rez'ised 3 April 1986," MS. number: 2810)