Tradeoff Between Flight Capability and Reproduction in Male

Ecological Entomology (2012), 37, 244–251 DOI: 10.1111/j.1365-2311.2012.01361.x

Trade-off between ﬂight capability and reproduction in male Velariﬁctorus asperses crickets

YANG ZENG1 andDAO-HONG ZHU1,2 1Laboratory of Insect Behavior & Evolutionary Ecology, Central South University of Forestry and Technology, Changsha, China and 2Laboratory of Zoology, Hunan First Normal University, Changsha, China

Abstract. 1. There are numerous data that support the trade-off between flight capability and reproduction in female wing polymorphic insects, but the relationship between wing form and fitness remains poorly investigated in males. 2. In the present study, the development of flight muscle and gonads, spermatophore size, and multiple copulation ability were investigated in both long-winged (LW) and short-winged (SW) males to verify this trade-off, using a wing dimorphic cricket species Velarifictorus aspersus (Walker). 3. The LW males had better-developed wing muscles than the SW males on the day of emergence, and both of them developed wing muscles after emergence, but the peak of weight in SW males was achieved 4 days later than that of the LW males. The accessory glands (AG) of the LW males developed significantly slower than that of the SW males. These results suggest that development and maintenance of flight muscles have a cost on the development of reproductive organs in male V. asperses. 4. The SW males produced significantly heavier spermatophores in a single copulation and mated more often than LW males. This indicates the SW males have a higher mating success than the LW males, thereby increasing their chance of siring offspring.

Key words. Accessory gland, ﬂight muscle, multiple mating, spermatophore, trade- off, Velariﬁctorus aspersus, Wing dimorphism.

Introduction a trade-off exists between flight capability and reproduction in wing dimorphic insects (Roff & Fairbairn, 1991). A general Wing dimorphism occurs in many insect orders (Johnson, observation, derived from many previous studies, is the ear- 1969; Harrison, 1980; Dingle, 1985; Roff, 1986, 1990; Zera lier onset of oviposition and enhanced reproductive output of & Denno, 1997; Zera, 2004). In general, there are two dis- flightless females (Anderson, 1973; Tanaka, 1976; Walters & tinct morphs, either with long or short wings. The long-winged Dixon, 1983; Roff, 1984; Zera, 1984, 2009; Guerra, 2011), (LW) adults usually have well-developed flight muscles, and which supports this trade-off in female insects. are flight-capable. In contrast, the short-winged (SW) adults Understanding the ecological significance of dispersal poly- usually have poorly-developed flight muscles, and are flight- morphism requires that fitness differences be documented less (Zera & Denno, 1997). The advantage of flight is obvi- between the wing forms of both sexes (Langellotto et al., ous as it gives the organism the ability to move away from 2000). However, the relationship between wing form and com- unfavourable or move towards favourable habitats, and many ponents of fitness in males has been much less studied. Some insects also use flight for smaller-scale foraging and courtship published studies on male crickets have suggested the absence flights (Roff, 1990). However, there are large energetic costs of the trade-off, e.g. no paternity difference (Holtmeier & Zera, associated with the production and maintenance of flight mus- 1993; Roff & Fairbairn, 1993), similar gonad size (Roff & Fair- cles (Roff, 1986; Zera, 2009). A well-known hypothesis is that bairn, 1993; Zera & Denno, 1997), and no difference in calling song structure (Souroukis et al., 1992). However, others have Correspondence: Dao-Hong Zhu, Laboratory of Zoology, Hunan shown differences between wing morphs that are suggestive First Normal University, Changsha 410205, China. of the trade-off, e.g. differences in calling and courtship songs E-mail: [email protected] (Crnokrak & Roff, 1995, 1998a,b, 2000; Guerra & Pollack,

2007), differences in gonad size (Tanaka, 1999; Crnokrak & Materials and methods Roff, 2002), difference in aggression (Guerra & Pollack, 2010), and differences in male nuptial gift size (Sakaluk, 1997). Vary- Insects and rearing method ing conditions under which the insects were raised to assess Adults of V. aspersus were collected from Taian City, fitness differences between wing forms may have contributed ◦ ◦ to the conflicting evidence of the trade-off (Denno, 1994; Shandong Province (35 38 N, 116 20 E) in August 2007, and × × Zera & Denno, 1997). In addition, any reproductive penal- maintained in plastic containers (30 cm 18 cm 20 cm) ties selectively imposed on macropterous males may be more to establish a laboratory strain. Eggs, nymphs, and adults were handled as described in previous studies (Zeng et al., difficult to detect (Ott, 1994; Zera & Denno, 1997) because ◦ 2010). The eggs collected were exposed to 8 C for 60 days reproduction costs are generally much lower in males than in ◦ females (Trivers, 1972). However, males often spend a great to terminate their diapause, and then incubated at 25 C deal of energy on different reproductive behaviours besides for hatching. Newly hatched nymphs were raised in groups (50 nymphs) in plastic containers (30 cm × 18 cm × 20 cm) gonad development. For example, calling requires a 10-fold ◦ increase in metabolic rate (Crnokrak & Roff, 1998a). Thus, under LD 16:8 h and 25 C. After emergence, the adults × × different results would also arise when different reproductive were moved to another container (10 cm 10 cm 10 cm) traits are examined. For instance, the testes size and paternity and kept separately under the same condition for the next of the LW and SW Gryllus firmus males are not significantly experiments. different (Roff & Fairbairn, 1993), but SW males do call longer and attract more females than the LW male (Crnokrak Development of flight muscles and reproductive organs in LW & Roff, 1998a). Generally, male reproduction includes three and SW adults steps: the first step is pre-copulatory behaviour, such as calling and courtship songs, (Hedrick, 1986; Simmons, 1988; Nelson Every 2 days for 14 days and then on the 17 and 20 day & Nolen, 1997), aggression towards the rival males (Langel- after emergence, LW and SW males (10 males for each) were ◦ lotto et al., 2000; Guerra & Pollack, 2010), and pre-copulatory frozen at −20 C for 12 h. The frozen insects were thawed at mounting (Zhu & Tanaka, 2002). The second step is copula- room temperature for 5 min. The dorso-longitudinal muscles tion, including prolonged copulation (Carroll, 1991; Michiels, (DLM) and testes of the males were dissected out under a 1992) and spermatophores transformation (Hayashi, 1998; binocular dissecting microscope (Leica Camera AG, Solms, Wedell & Ritchie, 2004). The last step is the post-copulatory Germany), and weighed using an electronic balance (Mettler- behaviour, such as post-copulatory guarding (Parker, 1970; Toledo Group, Zurich, Switzerland; 0.0001 g). Because the Tanaka & Zhu, 2003) and multiple copulation (Evans, 1987; accessory gland is important for crickets and other insects Hissmann, 1990; Fox, 1993). Therefore, all energy investments to form spermatophores (Kaulenas, 1992), the weights of in those three aspects should be considered when verifying reproductive accessory glands (AG) were also examined. To whether the trade-off really exists in male insects. Moreover, examine the effect of histolysis of flight muscles on the in some wing dimorphic insects, LW adults histolyse their expression of the trade-off, the LW males were classified flight muscles, and have increased investment into reproduc- by status of flight muscles and the weights of gonads were tion (Guerra & Pollack, 2007; Mitra et al., 2011). Thus, the compared between LW with fully developed flight muscles status of flight muscles may also influence the expression of (LWF), LW with histolysed flight muscles (LWH), and the trade-off. SW males. Velarifictorus aspersus (Walker) is a common cricket species found in fields, and is widely distributed throughout China (Yin & Liu, 1995), which means that flight may play a Comparative mating success in males between two wing forms very important role in dispersal of this species. This cricket species displays distinct wing dimorphism, and exposure to To examine the effects of wing morphs and age on the long-day, high-temperature, and high-density conditions induce spermatophore size of males, copulation experiments were macroptery (Zeng et al., 2010). Like the other wing dimorphic conducted on days 3, 5, 7, and 9 after male emergence. An insects, the flight capable female V. aspersus has a longer pre- individual male of either morph was placed with a virgin oviposition period and less fecundity than the SW female (Zeng 12- to 15-day-old adult SW female. The spermatophores were et al., 2012). Thus, this trade-off may underlie the differenti- removed and weighed immediately after they were transferred ation of life history strategy that is the LW individuals are to the females. capable of flight and SW ones benefit more in reproduction. To determine whether SW males have a higher capacity To verify the trade-off between flight capability and reproduc- for multiple copulation than LW males, an individual male of tioninmaleV. asperses, we investigated: (i) development of either morph was placed with a 12- to 15-day-old virgin adult testes and reproductive accessory glands that reflects the sex- SW female on day 9 after emergence. Their mating behaviour ual maturity and development of wing muscles; (ii) effects of was observed continuously. The spermatophore was removed wing morph and age on spermatophore size, which is an indi- and weighed immediately after copulation, and another vir- cation of the copulation ability; and (iii) the multiple mating gin SW female was immediately placed into the container to behaviour that is the major post-copulatory behaviour of the replace the previous one. The observation was continued for males between the two wing forms. 24 h, and the chamber was illuminated using a red light during

© 2012 The Authors Ecological Entomology © 2012 The Royal Entomological Society, Ecological Entomology, 37, 244–251 246 Yang Zeng and Dao-Hong Zhu the dark period. The mating frequencies and spermatophore 60 (a) *** weights of each male were recorded. ** 50 *** *** *** *** * 40 *** Results ** 30 Developmental proﬁles of ﬂight muscles and reproductive organs in male adults 20 10

The LW males had pink and well-developed flight muscles Weight of DLM (mg) on the day of emergence, which weighed 33.23 ± 6.76 mg 0 ± = (mean SD, n 10), making up 11.9% of the total body 20 weight. During early adulthood, their wing muscles developed (b) fast, and by day 8, the weight increased to 45.69 ± 8.71 mg. * The wing muscles of the SW males were white and poorly 15 developed on the day of emergence, weighing only 16.79 ± 3.72 mg (n = 10), and making up 6.6% of the body weight. 10 Their wing muscles also developed after emergence, and ± increasedupto27.09 4.61 mg by day 12, which was signif- 5 icantly higher than that of the males at day 0 (t-test, d.f. = 18, t = 5.53, P<0.001). The peak of the SW males’ weight was Weight of testis (mg) achieved 4 days later than that of the LW males. ancova 0 with body weight as a covariate indicated the wing muscles of 30 the LW males were significantly heavier than that of the SW (c) males during the observation period (day 2: F1,18 = 110.74, 25 * P<0.001; day 4: F1,18 = 114.68, P<0.001; day 6: F1,18 = ** 33.75, P<0.001; day 8: F1,18 = 138.23, P<0.001; day 10: 20 F1,18 = 120.71, P<0.001; day 12: F1,18 = 15.30, P<0.01; *** = = 15 ** day 14: F1,18 7.10, P<0.05; day 20: F1,18 9.29, P< *** 0.01), except for day 17 (F1,18 = 2.03, P = 0.17) (Fig. 1a). 10 The testes were well developed in both the LW and SW males on the day of emergence, and they were getting smaller 5 from 12 days on. No significant differences in weight were 0 found between the LW and SW males during the whole 024 6 8 10 12 14 17 20 Weight of accessory glands (mg) observation period except on day 12 (ancova with body Days after emergence weight as a covariate, F1,18 = 7.31, P<0.05), which might be caused by the small sample size (Fig. 1b). However, in Fig. 1. Temporal changes in mean (± SD; n = 10) fresh weight of both the LW and SW males, the AG were poorly developed the dorso-longitudinal muscles (DLM) (a), testes (b), and accessory on the day of emergence, weighing 3.58 ± 0.70 mg and glands (c) in long-winged (LW) (black bars) and short-winged (SW) 3.63 ± 1.22 mg, respectively, with no significant difference males (white bars) of Velarifictorus aspersus. *, **, and *** indicate between the two wing morphs (ancova with body weight a significant difference between the groups at the 5%, 1%, and 0.1% level, respectively, ancova with body weight as a covariate. as a covariate, F1,18 = 0.57, P = 0.46). After emergence, the AG of both male wing morphs developed gradually, but the SW males showed more rapid development than the (ancova with body weight as a covariate, day 14: F1,8 = 6.56, LW males, and the AG of the SW males were significantly P<0.05; day 17: F1,8 = 7.38, P<0.05) (Fig. 2a). ancova heavier than those of the LW males within 2–10 days with body weight as a covariate indicates that there was a after emergence (ancova with body weight as a covariate, significant difference in AG weight between SW and LWF day 2: F1,18 = 23.06, P<0.001; day 4: F1,18 = 16.7, P< males (day 14: F1,14 = 6.52, P<0.05; day 17: F1,12 = 5.67, 0.01; day 6: F1,18 = 21.67, P<0.001; day 8: F1,18 = 9.62, P<0.05), but no significant difference between LWH and SW P<0.01; day 10:F1,18 = 8.35, P<0.05) (Fig. 1c). males (day 14: F1,12 = 0.02, P = 0.89; day 17: F1,14 = 0.04, Some LW males had histolysed flight muscles from 12 days P = 0.85). Although the accessory glands of LWH males after emergence, and the proportions of LWH males were were apparently heavier than that of the LWF males, the 10%, 40%, 60%, and 80% on days 12, 14, 17, and 20, difference was not statistically significant (day 14: F1,8 = 2.54, respectively. Considering the sample size was too small, only P = 0.16; day 17: F1,8 = 1.05, P = 0.34), which might be one LWH male on day 12 and two LWF males on day 20, as a result of the small sample size (Fig. 2c). Thus, the we analysed the difference in organ mass between LWF, histolysis of flight muscles might promote the development LWH, and SW males on days 14 and 17. The colour of LWH of accessory glands but it had no effect on the development of males’ wing muscles was white and the weights of the wing testis (day 14: F2,17 = 0.86, P = 0.44; day 17: F2,17 = 0.43, muscles were significantly lighter than that of the LWF males P = 0.66) (Fig. 2b).

50 a Table 1. Analysis of variance of the effects of wing morph and age (a) x group on spermatophore weight in Velariﬁctorus aspersus. 40 Age of male adults y b 30 b y Day 3 Day 5 Day 7 Day 9 Long-winged 0.29 ± 0.03† 0.35 ± 0.07† 0.34 ± 0.05† 0.40 ± 0.07† 20 Short-winged 0.33 ± 0.05 0.47 ± 0.07 0.46 ± 0.08 0.52 ± 0.08 Source d.f. SS FP 10 Wing 1 0.210 48.029 <0.001 Weight of DLM (mg) Age 3 0.237 18.086 <0.001 0 Wing × age 3 0.026 2.010 0.120 15 Error 72 0.315 – – (b) a x a a x x † Mean ± SD, base unit is milligrams and 10 individuals for each 10 group.

morph and age group (anova followed by Tukey’s test, F = 5 2.01, P = 0.12). Weight of testis (mg) 0 Mating frequency and its effect on spermatophore size y (c) Both the LW and SW males mated several times with the xy ± a b ab x female. Mating occurred 7.9 1.3 times among LW males and 9.6 ± 1.7 times among SW males within 24 h; the difference is statistically significant (t-test, d.f. = 18, t = 2.60, P<0.05) (Table 2). The weights of spermatophores were 0.54 ± 0.07 mg among the SW males and 0.43 ± 0.09 mg among LW males at first copulation. As the mating frequencies increased, the spermatophore size decreased gradually, but the SW males produced significantly heavier spermatophores than the LW males in each mating until the mating frequency reached eight times = = 14 days 17 days (t-test, first mating: t 2.55, d.f. 18, P<0.05; second Weight of accessory glands (mg) mating: t = 2.57, d.f. = 18, P<0.05; third mating: t = 3.53, Adult age range d.f. = 18, P<0.01; fourth mating: t = 2.76, d.f. = 18, P< = = Fig. 2. Comparison of fresh weight of dorso-longitudinal muscles 0.05; fifth mating: t 7.07, d.f. 18, P<0.001; sixth mat- (DLM) (a), testes (b), and accessory glands (c) between long-winged ing: t = 3.22, d.f. = 18, P<0.01; seventh mating: t = 3.79, (LW) males with fully developed muscles (LWF) (black bars), LW d.f. = 15, P<0.01; eighth mating: t = 2.26, d.f. = 11, P< males with histolysed muscles (LWH) (grey bars), and short-winged 0.05) (Fig. 3). Given that very few males mated more than (SW) males (white bars) of Velarifictorus aspersus. Different letters eight times, the differences in spermatophore weight between indicate a significant difference between the groups at the 5% level, the LW and SW males was not analysed after they had mated ancova with body weight as a covariate. For LWF males, n = 6on eight times. The total spermatophore weights produced by the = = day 14 and n 4 on day 17; for LWH males, n 4onday14and SW males and the LW males were 4.26 ± 0.91 and 3.04 ± = = n 6 on day 17; and for SW males, n 10 both on days 14 and 17. 0.73 mg, respectively, and the difference was statistically significant (t-test, d.f. = 18, t = 4.18, P<0.01) (Table 2). Effects of male wing morph and age on spermatophore size Table 2. Comparison of mating frequency and total spermatophore As shown in Table 1, age has a significant effect on sper- weight in 24 h between LW and SW male Velarifictorus aspersus of matophore size. Both the LW and SW males produced heav- 9 days’ age. ier spermatophores as their age increased (anova followed by Tukey’s test, F = 18.09, P<0.001). The wing morph Mating frequency Total weight of the also influenced the spermatophore size (anova followed by Wing morph (times) spermatophore (mg) Tukey’s test, F = 48.03, P<0.001). The spermatophore was LW 7.9 ± 1.3†a‡ 3.04 ± 0.73†a‡ significantly heavier in the SW males than in the LW males SW 9.6 ± 1.7b 4.26 ± 0.91 b in all of the four age groups (t-test, day 3: t = 2.19, d.f. = 18, P<0.05; day 5: t = 3.88, d.f. = 18, P<0.01; day 7: † Mean ± SD, 10 individuals for each group. t = 3.84, d.f. = 18, P<0.01; day 9: t = 3.67, d.f. = 18, P< ‡ Different letters indicate a significant difference between LW and SW 0.01). No significant interaction was observed between wing males by t-test, P < 0.05.

0.7 In Gryllodes sigillatus, SW males had bigger gonads and * produced heavier spermatophores than LW males (Sakaluk, 0.6 * ** 1997). Therefore, the difference in gonad size may contribute * *** 0.5 ** ** * to a difference in spermatophore size. We found the wing 0.4 muscles of SW male V. aspersus also developed significantly 0.3 within the first 12 days. The male crickets usually rub their fore-wings to make songs, including fighting, calling, and 0.2 courtship songs, so the fore-wing muscles are very important 0.1 for reproduction. For the LW males, they need to flap their hind-wings very quickly to fly, so the hind-wing muscles may

Weight of spermatophore (mg) 0 0123456789101112 be the main part of their wing muscles. For the SW males, Mating times in 24h as they have non-functional, short hind wings, they may have fewer muscles related to hind wings and their wing muscle ± = Fig. 3. Comparison of the mean ( SD; n 10) spermatophore fibers may be mostly related to moving the fore wings. Thus, weight between long-winged (LW) (black bars) and short-winged (SW) we may infer that development of LW males’ wing muscles (white bars) male Velarifictorus aspersus during multiple copulation. *, **, and *** indicate a significant difference between the groups at are for flight, and development of SW males’ wing muscles the 5%, 1%, and 0.1% level, respectively, by t-test. are for reproduction. Crnokrak and Roff (1998a) found that G. firmus SW males had longer calling songs and attracted more females than LW males, and SW male Gryllus texensis Discussion could also produce courtship songs at a significantly higher Although a trade-off between dispersal capability and repro- probability than LW male (Guerra & Pollack, 2007). These ductive success has been well documented in females of many differences in pre-copulatory behaviour between LW and SW wing-dimorphic insects, the relationship between wing form males may be associated with this functional difference in their and fitness remains poorly investigated in males (Denno, 1994; wing muscles. Ott, 1994; Zera & Denno, 1997; Roff & Fairbairn, 2007; Most male cricket species produce spermatophores dur- Guerra, 2011). Published studies on male wing-dimorphic ing copulation (Gwynne, 1984). Some cricket species, termed insects have provided conflicting evidence for the existence as gift-giving species, produce big spermatophores that con- of the trade-off. Certain studies on planthoppers and other tain a large spermatophylax and a sperm-containing ampulla, wing-dimorphic insects have shown a fitness advantage for whereas non-gift-giving species do not produce a large brachyptery in males (Fujisaki, 1992; Crnokrak & Roff, spermatophylax (Sakaluk, 1986). In the gift-giving species 1995). Other studies failed to detect a significant difference G. sigillatus, the spermatophores of SW males were signifi- in some of the components of reproductive success between cantly heavier than those of LW males, which consequently male wing forms (Roff & Fairbairn, 1991; Holtmeier & required more time for the females to consume. Given that the Zera, 1993). sperm ampulla is removed soon after the spermatophylax is In the present study, the LW male V. aspersus had better- consumed, the SW males could transfer more sperm into the developed wing muscles and their wing muscles developed female’s reproductive tract, which suggests a higher insemina- faster than the SW males after emergence, which indicates tion success for SW males (Sakaluk, 1997). The V. aspersus is that the LW male V. aspersus may need a great deal of energy a non-gift-giving species, and the present results showed that for the development and maintenance of the flight muscles in the SW males produced a significantly heavier spermatophore early adulthood just like other male wing dimorphic insects than LW males. In non-gift-giving species, the difference in (Tanaka, 1999; Socha & Sula, 2008). The testes of these two spermatophore size between the SW and LW males and its wing morphs were well developed on the day of emergence influence on male mating success have not previously been and their weights were not significantly different. However, the studied. However, this effect may be different from that of gift- accessory glands of the SW males developed faster than that of giving species because the spermatophore does not contain a the LW males in early adulthood. These results suggest that the big spermatophylax for the females to consume, which means development and maintenance of flight muscles has a cost on the females cannot decide when to remove the sperm ampulla. the development of reproductive organs in male V. asperses. In non-gift-giving cricket species, females also showed great In Callosobruchus maculatus, the well-developed testes were propensity to eat the spermatophores after mating (Alexander observed only among SW males on the day of emergence, & Otte, 1967; Sakaluk & Cade, 1980), and we observed that indicating that the SW male sexually matures faster than the the female V. aspersus eat the spermatophore after sperm trans- LW males (Utida, 1972). However, in cricket species, such fer. Thus, the heavier spermatophores may correlate with more as Modicogryllus confirmatus and G. firmus, the testes were sperm or nutrients in this species, but it remains to be seen if well developed after emergence both in LW and SW males there is a positive correlation between spermatophore weight (Roff & Fairbairn, 1993; Tanaka, 1999), suggesting that both and sperm number or nutrients. In some insects, the nutrients wing-morphed males may have the ability to reproduce very contained in the spermatophore are often incorporated into the soon after emergence. Indeed, the production of calling song, female’s developing oocytes, thereby increasing egg produc- which is one of the indications of sexual maturity, begun at a tion (Friedel & Gillott, 1977; Boggs & Gilbert, 1979; Gwynne, similar age in LW and SW G. firmus (Crnokrak & Roff, 1998a). 1984; Butlin et al., 1987).

Post-copulatory behaviours, such as guarding, mounting, live longer than SW males. Considering that male reproduc- and multiple copulation, are found in many insect species. tive behaviour begins at 4–6 days of age (Cade & Wyatt, In Ischnura graellsii, males continued to mount the females 1984; Crnokrak & Roff, 1995), and that the testes begin to after copulation until the female lays eggs, making sure their degenerate several days after emergency (Tanaka, 1999), the sperm successfully fertilise the ova (Cordero, 1990). Multiple early adulthood may be the most active period of the males’ copulations occur in many species of Coleoptera, Lepidoptera, reproduction. Hemiptera, and so on; they are believed to be beneficial to both The present findings in male V. aspersus strongly support males and females (Drummond, 1984; Page, 1986; Fox, 1993). the hypothesis of trade-off between flight capability and repro- The mating frequency in non-gift-giving crickets is very high. duction. However, we did not consider the effects of actual For example, the male Gryllus veletis could mate 6.8 ± 0.8 flight behaviour on reproduction in this study. In G. texen- times a day (Burpee & Sakaluk, 1993). In the present study, sis, a short-time flight has been shown to increase courtship both of the SW and LW male V. aspersus mated several times behaviour (Guerra & Pollack, 2009), but in G. rubens, a 1–8 h within 24 h. In Drosophila melanogaster, the courtship alone flight could further decrease the reproductive output of the could reduce the longevity of the male (Cordts & Partridge, female (Zera & Rankin, 1989). So, the time of flight may have 1996), and Paukku and Kotiaho (2005) also found virgin males an effect on the expression of the trade-off. We are currently lived longer than mated males in Callosobruchus maculatus. conducting studies addressing this issue in V. aspersus. Thus, the multiple copulations may cost the males a large amount of energy. The mating frequency and total weight of the spermatophores were both significantly higher among Acknowledgements SW males than LW males. This suggests that the SW male V. aspersus has a higher ability in multiple copulation than This work was supported by the National Nature Science the LW male. In some other insects, such as chinch bugs Foundation of China (Grant nos. 31070586 and 30771740). and water striders, SW males also mated more frequently than their macropterous counterparts (Crespi, 1988; Fujisaki, 1992). References In Prokelisia dolus, when each male was placed and allowed to copulate with 10 virgin females for 8 h, the brachyterous Alexander, R.D. & Otte, D. (1967) The evolution of genitalia and male sired twice as many offspring as macropterous males, mating behavior in crickets (Gryllidae) and other Orthoptera. 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