Factors that Affect Reproductive Success in Male and Female Monarchs
Do Males and Females Benefit From Spermatophores in the Same Way?Abstract | Introduction | Methods | Results | Discussion | Acknowledgments | References | Karen's Research Questions After mating, spermatophores transferred by male monarch butterflies are broken down within the female bursa copulatrix; mass decreased at a constant rate until little material remained. The time required for breakdown of large spermatophores is longer than the average intermating interval of females that receive large spermatophores. Because there is last male sperm precedence in monarch butterflies, a significant portion of a male's investment could thus be used to benefit offspring from other males. This suggests that there is conflict over the fate of monarch spermatophores, with females benefiting most by digesting their contents as quickly as possible in order to maximize nutrient gain, and males benefiting most when their spermatophores remain in the bursa copulatrix as long as possible, thus delaying female remating. Like other Lepidoptera, male monarch butterflies produce a spermatophore during mating that often represents a large material investment, comprising up to 10% of the male’s body mass (Oberhauser 1988, 1992, What factors affect the size and composition of monarch spermatophores?). Accessory gland products contained in spermatophores have been shown to function in sperm activation and stimulation of oogenesis and oviposition (Leopold 1976). However, spermatophores often contain more material than is necessary for these functions; even the very small spermatophores transferred by recently-mated males result in egg fertilization and oviposition (Rutowski et al. 1987, Oberhauser 1989, What factors affect the number of eggs that female monarchs lay?). Spermatophores contain significant quantities of nitrogen (Oberhauser 1992, What factors affect the size and composition of monarch spermatophores?), largely in the form of protein (Marshall 1982). Because the adult food of monarch butterflies, nectar, is a poor nitrogen source, most spermatophore nitrogen must come from larval reserves, and could represent an important limiting resource for males. Spermatophore production presents an opportunity to study the benefits of reproductive investment by males, and possible conflicts between the sexes. Two potential benefits of producing large spermatophores have been suggested. First, spermatophore constituents may be used by females to increase offspring quantity or quality, thereby increasing male reproductive success as well. We will call this benefit the nutrient function. The female digests the spermatophore with enzymes secreted in the bursa copulatrix, and the nutrients that are released during this process can leave the bursa and go to other parts of the female’s body. In this way, the bursa is similar to a human stomach. Several studies have shown that the contents of lepidopteran spermatophores are incorporated into both eggs and female somatic tissue (Boggs and Gilbert 1979, Boggs 1981, 1990, Boggs and Watt 1981, Greenfield 1982, Wells et al 1993), and in many species there is a positive relationship between the amount of spermatophore material received and fecundity (Rutowski et al 1987, Watanbe 1988, Oberhauser 1989, 1992, Tamhankar et al. 1993, Wiklund et al. 1993, What factors affect the number of eggs that female monarchs lay?). The second potential benefit of producing large spermatophores is increasing the proportion of the eggs laid by a given female that are fertilized by sperm from the donating male. We will call this benefit the delay function. Several studies have indicated that large ejaculates delay female remating (Labine 1964, Sugawara 1979, Rutowski 1980, 1984, Rutowski et al. 1981, Oberhauser 1989, 1992, How often do males and females mate, and what factors affect the intervals between mating?). Many female Lepidoptera mate more than once, and there is a pattern of sperm precedence of the last male to mate with the female (Gwynne 1984, Whose sperm fertilize the female’s eggs if she mates more than once?). Thus, sperm from males that produce large spermatophores could fertilize a higher proportion of a female’s eggs without necessarily increasing her lifetime reproductive success. These two benefits are not mutually exclusive; males may benefit both by increasing female reproductive output and by delaying female remating. However, since a spermatophore that has been digested will not be as effective at delaying remating, there must be some tradeoffs between the two effects. This could lead to conflict between male and female interests in the fate of spermatophore contents and the rate of spermatophore digestion. In assessing potential benefits to males of providing females with nutrients, we need to know not only whether the nutrients increase female fecundity, but also whether they are used in offspring of the donating male. This would be true if the female did not remate until after she had used the nutrients from a previous mate. The nutrient function predicts that the average female intermating interval should be greater than the time required for spermatophore degradation. If this interval is shorter than degradation time, the delay function is probably more important. In this study, I estimate the proportion of a male's nutrient contribution that is available exclusively to his own offspring by measuring the proportion of spermatophore nutrients that have been removed from the bursa copulatrix by the time the female remates. Boggs and Gilbert (1979) showed that male-derived nutrients were present in monarch butterfly eggs the day after mating and increased in level for approximately five days after mating, at which point their levels declined sharply. Since this period corresponds roughly to the time over which spermatophores are degraded (see below), I assume that male-derived nutrients are used soon after being removed from the bursa copulatrix. General methods Monarch butterflies were reared and maintained as described previously (Oberhauser 1988, What factors affect the size and composition of monarch spermatophores?). Experiments were carried out during the summer of 1988 in central Wisconsin. Butterflies were kept in outdoor screen cages (2m x 2m x 2m or 2m x 3m x 2m), and fed a 30% honey solution to satiation daily. All butterflies were offspring of females captured from the wild.
Rate of spermatophore degradation In order to measure the rate at which spermatophores were degraded, 28 males in each of five groups were allowed to mate with virgin females aged five to eight days. The five male groups included three groups of unmated males (aged five, eight and 12-15 days), and two groups of previously-mated males (mated one and four days prior to the experimental mating). After mating, I kept females in outdoor screen cages and provided them with fresh common milkweed (Asclepias syriaca) on which to oviposit. At three-day intervals one to 19 days after mating, four females from each group were randomly selected for dissection. I placed the female’s abdomen in insect saline solution, removed the spermatophore from the bursa copulatrix, blotted it to uniform dryness on tissue paper, and then weighed it to the nearest 0.01 mg on a Mettler semi-micro analytical balance.
Spermatophore masses transferred by wild males To estimate the mass of spermatophores that wild females receive, I captured wild males and allowed them to mate with females in captivity. Males were put with unmated females as soon as possible after capture. When males mated within two days of capture, their mates were dissected to determine the spermatophore mass transferred. Rate of Spermatophore Degradation Figure 1 illustrates the rate at which spermatophores decreased in mass after they had been transferred to the female. Those from males mated four days previously are not shown, because they were so similar to those from five-day-old unmated males. I used a linear regression model to determine that spermatophore mass decreases at a relatively constant rate (about 3.3 mg per day), but stops decreasing when about 5 mg of material remains. The remaining material consists of part of the tough outer covering, and the spermatophore neck, or collum.
Figure 1: Spermatophore mass remaining at different times after mating from unmated males of different ages and males that had mated one day previously. Marks on the graph represent the average of all of the spermatophores dissected from a given category at a given time after transfer. Older males transfer spermatophores that are initially larger, and these spermatophores take about ten days to break down. Smaller spermatophores (from five and eight day old males) take about seven days to break down. By two days after mating, spermatophores from males that mated one day previously are completely degraded.
Spermatophore sizes transferred by wild males Ten males captured from the wild mated within two days of capture. Figure 2 shows the distribution of spermatophore masses from these males. Mean spermatophore mass was 17.9 mg.
Figure 2: Spermatophore masses (+ 2.5 mg) transferred by 10 males captured from the wild and mated with captive females within two days of their capture. (Mean = 17.86 mg)
Two things must be known to estimate how much of a male's material investment is available exclusively to his offspring: (1) the time between his mating with a female and her next mating and (2) the amount of spermatophore material that has been removed from the female bursa copulatrix at this time. Any material remaining at this point could potentially be used in the offspring of another male, depending on the degree of sperm precedence. Both (1) and (2) are a function of initial spermatophore size (Oberhauser 1989, How often do males and females mate, and what factors affect the timing of mating?, present results). Spermatophores of approximately 15 mg or less are likely to be degraded before a female remates; most of the material is removed from the bursa copulatrix within two or three days, which is close to mean and median intermating intervals for females receiving small spermatophores (How often do males and females mate, and what factors affect the timing of mating?). Thus the predictions suggested by the nutrient function are met when males transfer small spermatophores; intermating intervals are greater than or equal to spermatophore degradation time. However, it is likely that a significant proportion of larger spermatophores will remain in the bursa copulatrix after the female remates; females that receive large spermaotphores remate after three to four days (Oberhauser 1989, 1992, How often do males and females mate, and what factors affect the timing of mating?). At this point, only half of the mass of larger spermatophores has been removed (figure 1). In this case, the intermating interval is shorter than degradation time, which suggests that the delay effect is most important to males. Wild males appear to transfer both large and small spermatophores (figure 2). Thus, some spermatophores within size ranges likely to be transferred by wild males meet predictions of the nutrient effect, and some meet predictions of the delay effect. Can these results help us to draw any conclusions as to the function of the large material investment by male monarch butterflies? Several factors suggest that the delay effect is likely to be more important in this species. First, because males transfer all of their available accessory gland material during a mating (Oberhauser 1988, What factors affect the size and composition of monarch spermaotphores?), it is probably advantageous for males to transfer large spermatophores. Second, because it is likely that females will remate before they have used all of the material in a large spermatophore, the nutrient function suggests that male monarchs often transfer suboptimally large spermatophores. Finally, when females receive either one large or one small spermatophore, there is no detectable difference in fecundity (Oberhauser 1989, What factors affect the number of eggs that female monarchs lay?). This suggests that the benefits of the nutrient effect are not very important to males. If males benefit by delaying female remating, and females benefit by using spermatophore nutrients, there is potential for conflict between the sexes in monarchs and other Lepidoptera in which females mate more than once. I think that it is most likely that the primary function of large spermatophores (the way in which they benefit the individuals making the investment) is to delay female remating, but that females have evolved to take advantage of the male investment by digesting and using its constituents. Conflict between the sexes on how spermatophores are used is likely, with females benefiting most by digesting their contents as rapidly as possible and then remating, thus gaining the maximum amount of nutrients from males. Males maximize their reproductive success by delaying both spermatophore breakdown and female remating. Svärd and Wiklund (1989) presented comparative data showing a positive correlation between the degree of female multiple mating in butterflies and male ejaculate mass, suggesting that males do not invest as heavily when there is less advantage to delayed female remating. However, because spermatophores do not remain intact within the female bursa copulatrix, it is also important to compare rates of degradation when determining the function of this investment. If it is to delay female remating, there should be a negative correlation between degradation rates and the degree of polyandry across species; the selection pressure to make long-lasting spermatophores should be stronger in more polyandrous species. There are few data on the rates at which spermatophores are degraded, but there is some evidence of this negative correlation. Boggs (1981) measured spermatophore degradation in two heliconiine butterflies, one monandrous and one polyandrous, and found slower degradation in the polyandrous species. While detailed studies of spermatophore content and degradation rates are scarce (see Rutowski (1984) for an exception) there is evidence of a great deal of variability within the Lepidoptera that could provide an interesting basis for comparative study. Spermatophores of Heliothis zea contain chitin (Callahan 1958), which is presumably difficult for females to break down; spruce budworms (Outram 1971), and possibly some skippers (Dana 1989), produce spermatophores that may not be degraded at all. On the other extreme, gypsy moth spermatophores are completely degraded within hours after mating (Taylor 1967, Loerch and Cameron 1984). I thank D. Alstad, R. Rutowski, C. Wiklund, P. Oberhauser, S. Oberhauser, and P. Abrams for help and advice during various stages of this research and writing. Financial support was provided by the NSF (BSR 8805884), the University of Minnesota Graduate School, and the Dayton and Wilkie Funds for the Study of Natural History, administered by the Bell Museum of Natural History at the University of Minnesota. return to Karen's Research Questions
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