Control of reproduction and a survival cost to mating in female Bicyclus anynana butterflies

Abstract.  1. Given that sexual conflict is all pervasive, investigating potential costs to mating and the control of female reproduction are important issues.
2. Here, female reproductive output and longevity are investigated in relation to mating status (virgin vs. once-mated females) and host-plant availability in the tropical butterfly Bicyclus anynana .
3. Both factors significantly affected realised fecundity in a manner demonstrating that female reproduction is not entirely under control of the female herself, but is rather subject to additive contributions of the female, her male partner, and intrinsic physiological processes. As evident from the deposition of significant egg numbers by virgin, host-deprived females, B. anynana is effectively unable to completely inhibit oogenesis.
4. Mated females suffered a reduction in adult life span, which cannot be explained as a side-effect of variation in egg size, lifetime, or early fecundity.
5. Such detrimental effects of mating per se are indicative of the cooperation–conflict balance between sexes being shifted towards conflict in B. anynana .     

Effects of adult nutrition on female reproduction in a fruit-feeding butterfly: the role of fruit decay and dietary lipids

It was generally believed that butterflies and other holometabolous insects rely primarily on reserves accumulated during the larval stage for reproduction. Recent studies, however, highlight the often fundamental importance of adult nutrition to realize the full reproductive potential. While the importance of carbohydrates is fairly well understood, the role of most other adult-derived substances is only partially resolved. We here focus on the effects of dietary lipids (cholesterol, polyunsaturated fatty acids) and fruit decay (dietary yeast, ethanol) on female reproduction in the tropical, fruit-feeding butterfly Bicyclus anynana (Nymphalidae). We found that banana-fed control females outperformed all other groups fed on sucrose-based diets. Lipids, yeast or ethanol added to a sugar solution did not yield a similarly high reproductive output compared to fruit-fed females. Groups fed fresh or decaying banana showed no differences in reproductive performance. As we could not identify a single pivotal substance, we conclude that resource congruence (the use of nutrient types in a specified ratio) rather than any specific nutrient component is of key importance for maximum reproductive output. Further, dietary quality may affect egg hatching success in spite of no obvious effects on egg size and number. Thus, any implications about potential fitness effects of different diets need to consider egg (and hatchling) viability in addition to fecundity.     

Enhancing offspring quality or quantity? Different ways for using nectar amino acids in female butterflies

Butterfly-pollinated flowers offer nectar with higher amino acid concentrations than most flowers pollinated by other animals, and female butterflies of some species prefer to consume amino acid-rich nectar. However, for over 30 years, there has been an ongoing discussion about whether nectar amino acids benefit butterfly fitness. A clear positive effect was only shown for the nectar-feeding Araschnia levana, and females of the fruit-feeding Bicyclus anynana also increased offspring quality when they were fed amino acids as adults. Thus, severe doubts remain about the general significance of these single positive results. We therefore tested a further species from a phylogenetically different butterfly subfamily, the small heath (Coenonympha pamphilus L., Satyrinae), taking into account feeding conditions over the whole life cycle of this species. C. pamphilus females receiving nectar amino acids as adults, irrespective of larval food quality, produced heavier larvae and also increased the hatching success of their eggs over the oviposition period. Furthermore, females raised under nitrogen-poor larval conditions tended to use nectar amino acids to increase the number of eggs laid. Thus, C. pamphilus females used nectar amino acids primarily to increase their offspring quality, and secondly tended to increase offspring quantity, if larval resources were scarce, showing a resource allocation pattern differing from both B. anynana and A. levana. Our study supports the old postulate that nectar amino acids generally enhance butterfly fitness.     

Larval starvation reduces responsiveness to feeding stimuli and does not affect feeding preferences in a butterfly

It is commonly assumed that holometabolic insects such as Lepidoptera rely primarily on larval storage reserves for reproduction. Recent studies though have documented a prominent role of adult-derived carbohydrates for butterfly reproduction. Moreover, a few studies have shown that adult butterflies may also benefit from adult-derived amino acids, at least when larval storage reserves are reduced. Given that in holometabolous insects larval deficiencies are carried over into the adult stage, reduced storage reserves have the potential to modulate adult feeding preferences and responses in order to allow for a successful compensation. We tested this hypothesis here in the fruit-feeding butterfly Bicyclus anynana using larval food stress to manipulate storage reserves. Alcohols (methanol, ethanol, butanol, propanol), sugars (maltose, glucose, fructose, sucrose), and acetic acid acted as feeding stimuli, while butterflies did not respond to other substances such as amino acids, yeast, salts, or vitamins. Contrary to expectations, stressed butterflies showed a weaker response than controls to several feeding stimuli. In preference tests, butterflies preferred sugar solutions containing proline, arginine, glutamic acid, acetic acid, or ethanol over plain sugar solutions, but discriminated against salts. However, there were no general differences among starved and control butterflies. We conclude that larval food-stress does not elicit compensatory feeding behavior such as a stronger preference for amino acids or other essential nutrients in B. anynana. Instead, the stress imposed by a period of starvation yielded negative effects.     

Introduced fire ants can exclude native ants from critical mutualist-provided resources

After over 30 years of research, it was recently shown that nectar amino acids increase female butterfly fecundity. However, little attention has been paid to the effect of nectar amino acids on male butterfly reproduction. Here, we show that larval food conditions (nitrogen-rich vs. nitrogen-poor host plants) and adult diet quality (nectar with or without amino acids) affected the amount of consumed nectar in Coenonympha pamphilus males. Furthermore, amino acids in the nectar diet of males increased progeny’s larval hatching mass, irrespective of paternal larval reserves. Our study takes the whole reproductive cycle of male butterflies into account, and also considers the role of females in passing male nutrients to offspring, as males’ realized reproduction was examined indirectly via nuptial gifts, by female performance. With this comprehensive approach, we demonstrate for the first time that nectar amino acids can improve male butterfly reproduction, supporting the old postulate that nectar amino acids generally enhance butterfly fitness.     

Adult diet affects lifespan and reproduction of the fruit‐feeding butterfly Charaxes fulvescens

Fruit‐feeding butterflies are among the longest lived Lepidoptera. While the use of pollen‐derived amino acids by Heliconius butterflies has been interpreted as important for the evolution of extended lifespans, very little is known about the life‐history consequences of frugivory. This issue is addressed by investigating effects of four adult diets (sugar, sugar with amino acids, banana, and moistened banana) on lifespan and reproduction in the fruit‐feeding butterfly Charaxes fulvescens Aurivillius (Lepidoptera: Nymphalidae). Female butterflies were collected from Kibale National Park, Uganda, and kept individually in cages near their natural habitat and data were collected on lifespan, oviposition, and hatching of eggs. Lifespan in captivity was longer for the sugar and the amino acid cohort, than for the banana cohorts. The longitudinal pattern of oviposition was erratic, with many days without oviposition and few periods with high numbers of eggs laid. Butterflies typically did not lay eggs during their 1st week in captivity and the length of the period between capture and first reproduction was significantly shorter for butterflies fed moistened banana. The length of the reproduction period (first reproduction–last reproduction in captivity) and the reproduction rate (total number of eggs/length of the reproduction period) did not differ significantly between the diet treatments. Those fed with amino acid and moistened banana had significantly higher egg hatchability than those fed with sugar and banana. We found no evidence for a lifespan cost of reproduction. Our results show that (1) female C. fulvescens can use amino acids in their diet for laying fertile eggs, (2) more wing‐wear does correlate with lower survival in captivity (indicating aging in the wild), but not with intensity of reproduction (providing no evidence for reproductive aging), and (3) fruit‐feeding butterflies may be dietary restricted in the field.     

Effects of adult-derived carbohydrates, amino acids and micronutrients on female reproduction in a fruit-feeding butterfly

It is generally believed that butterflies (and other holometabolous insects) rely primarily on reserves accumulated during the larval stage for reproduction, whereas the carbohydrate-rich adult diet is thought to mainly cover energy requirements. In at least some species though, realization of the full reproductive potential is extensively affected by post-eclosion nutrition. While the importance of carbohydrates is fairly well understood, the role of adult-derived amino acids and micronutrients is controversial and largely unknown, respectively. We here focus on the effects of different adult diets on female reproduction in the tropical, fruit-feeding butterfly Bicyclus anynana (Nymphalidae). Carbohydrates were the most important adult-derived nutrients affecting reproduction. Adding amino acids, vitamins or minerals to sucrose-based solutions did not yield a reproductive output equivalent to that of fruit-fed females, which showed the highest performance throughout. This suggests that either not yet identified compounds of fruit substantially contribute to reproduction, or that resource congruence (the use of nutrient types in a specified ratio) rather than any specific nutrient component is of key importance. Apart from adult income, realized fecundity depended on egg size and longevity, with the former dominating when dietary quality was low, but the latter when quality was high. Thus, the egg size-number trade-off seems to be affected by female nutrition.     

Effects of larval starvation and adult diet-derived amino acids on reproduction in a fruit-feeding butterfly

Availability of adequate nutrition is among the most important factors affecting growth, development, and reproduction in animals. In holometabolous insects, diets and nutritional needs change between life stages, with larval storage, and adult feeding and reproduction being linked to one another. In several butterfly species, adult feeding is of fundamental importance to realize the full reproductive potential, primarily due to a prominent role of adult diet-derived carbohydrates. In contrast, the role of adult diet-derived amino acids is still under debate, despite the fact that butterflies were often found to preferentially feed on amino acid-rich substrates. Recently it was found that amino acids from adult income could compensate for adverse effects of larval food quality. In our study on the tropical butterfly Bicyclus anynana (Butler) (Lepidoptera: Nymphalidae), we used larval starvation to investigate corresponding effects on female reproductive output. Short periods of larval starvation prolonged development time and reduced larval survival, larval growth rate, pupal mass, and egg size. Regardless of the degree of larval starvation, access to amino acids in the adult diet increased egg size, whereas egg number remained largely unaffected. Thus, although there was some evidence for adult diet-derived amino acids being overall beneficial to reproduction, there was no indication that they can compensate for larval starvation.     

Linking nectar amino acids to fitness in female butterflies

Nectar of butterfly-pollinated flowers contains generally higher levels of amino acids than does nectar of flowers pollinated by most other animal types. One proposed explanation is that these amino acids promote butterfly fitness, although the evidence has been equivocal. In a new study, Mevi-Schütz and Erhardt showed that nectar amino acids enhanced fecundity in the butterfly Araschnia levana, but only when the larval diet was poor. Their results support the hypothesis that butterflies are agents of selection for higher nectar amino acid production, suggest that the larval food plant has a key role in the evolution of the flower-butterfly mutualism, and demonstrate that the importance, to butterfly reproduction, of different nutrient sources varies with butterfly nutritional state.     

Host strain specific sex pheromone variation in Spodoptera frugiperda

Background

In the Lepidoptera it was historically believed that adult butterflies rely primarily on larval-derived nutrients for reproduction and somatic maintenance. However, recent studies highlight the complex interactions between storage reserves and adult income, and that the latter may contribute significantly to reproduction. Effects of adult diet were commonly assessed by determining the number and/or size of the eggs produced, whilst its consequences for egg composition and offspring viability were largely neglected (as is generally true for insects). We here specifically focus on these latter issues by using the fruit-feeding tropical butterfly Bicyclus anynana, which is highly dependent on adult-derived carbohydrates for reproduction.

Results

Adult diet of female B. anynana had pronounced effects on fecundity, egg composition and egg hatching success, with butterflies feeding on the complex nutrition of banana fruit performing best. Adding vitamins and minerals to a sucrose-based diet increased fecundity, but not offspring viability. All other groups (plain sucrose solution, sucrose solution enriched with lipids or yeast) had a substantially lower fecundity and egg hatching success compared to the banana group. Differences were particularly pronounced later in life, presumably indicating the depletion of essential nutrients in sucrose-fed females. Effects of adult diet on egg composition were not straightforward, indicating complex interactions among specific compounds. There was some evidence that total egg energy and water content were related to hatching success, while egg protein, lipid, glycogen and free carbohydrate content did not seem to limit successful development.

Conclusion

The patterns shown here exemplify the complexity of reproductive resource allocation in B. anynana, and the need to consider egg composition and offspring viability when trying to estimate the effects of adult nutrition on fitness in this butterfly and other insects.     

Nectar sugars enhance fitness in male Coenonympha pamphilus butterflies by increasing longevity or realized reproduction

The principal components of floral nectar are water and the sugars sucrose, fructose and glucose. Several studies have shown the importance of nectar sugars for female butterfly fecundity, whereas to date little attention has been paid to the effect of nectar sugars on male butterfly reproduction. Clear evidence for an effect of nectar sugars on male realized reproductive success is still missing. In this study, we fed male Coenonympha pamphilus butterflies nectar mimics with low (5%), medium (20%) or high (30%) total sugar concentrations with a sucrose:glucose:fructose ratio of 2.7:1.1:1. Sugar solutions were made mimicking Knautia arvensis, an essential nectar plant for C. pamphilus and many other European butterflies. Realized male reproductive success for each treatment was measured indirectly via nuptial gifts, by recording reproductive parameters and by characterizing time patterns over the oviposition period of their female partner. Male butterflies fed high‐concentrated nectar sugars had a longer lifespan than males fed low‐concentrated nectar sugars. In contrast, offspring of males fed medium‐concentrated nectar sugars had a higher hatching mass than progeny of males fed low‐concentrated nectar sugars, indicating a tradeoff between somatic maintenance and reproduction in the use of nectar sugars. Thus, allocation patterns of nectar sugars differed according to sugar concentrations in adult food. The method used in this experiment took into account the indispensable role of female butterflies in passing male nutrients to offspring. With this comprehensive approach, we can show the general importance of nectar sugars for male butterfly fitness and support previous findings suggesting a coevolutionary process between butterflies and flowers dependent on butterfly pollination.     

Longevity-fertility trade-offs in the tephritid fruit fly, Anastrepha ludens, across dietary-restriction gradients

Although it is widely known that dietary restriction (DR) not only extends the longevity of a wide range of species but also reduces their reproductive output, the interrelationship of DR, longevity extension and reproduction is not well understood in any organism. Here we address the question: ‘Under what nutritional conditions do the longevity‐enhancing effects resulting from food restriction either counteract, complement or reinforce the mortality costs of reproduction? To answer this question we designed a fine‐grained DR study involving 4800 individuals of the tephritid fruit fly, Anastrepha ludens, in which we measured sex‐specific survival and daily reproduction in females in each of 20 different treatments (sugar : yeast ratios) plus 4 starvation controls. The database generated from this 3‐year study consisted of approximately 100 000 life‐days for each sex and 750 000 eggs distributed over the reproductive lives of 2400 females. The fertility and longevity‐extending responses were used to create contour maps (X‐Y grid) that show the demographic responses (Z‐axis) across dietary gradients that range from complete starvation to both ad libitum sugar‐only and ad libitum standard diet (3 : 1 sugar : yeast). The topographic perspectives reveal demographic equivalencies along nutritional gradients, differences in the graded responses of males and females, egg production costs that are sensitive to the interaction of food amounts and constituents, and orthogonal contours (equivalencies in longevity or reproduction) representing demographic thresholds related to both caloric content and sugar : yeast ratios. In general, the finding that lifespan and reproductive maxima occur at much different nutritional coordinates poses a major challenge for the use of food restriction (or a mimetic) in humans to improve health and extend longevity in humans.     

Adult life spans of butterflies (Lepidoptera: Papilionoidea + Hesperioidea): broadscale contingencies with adult and larval traits in multi-species comparisons

Adult life spans of insects are part of supposedly adaptive life-history syndromes, which involve a variety of reproductive strategies with regard to mating, egg maturation and egg laying. Contingencies of life span with many other morphological, behavioural, and ecological traits are to be expected. We used a data-mining approach, incorporating controls for phylogeny, to uncover broad patterns of trait contingencies in a literature-derived data set of adult life spans for 350 butterfly species. We found that adult feeding habit has strong links with life spans, with pollen-feeders living longer than fruit feeders, which in turn live longer than nectar feeders. Furthermore, traits reducing susceptibility to predators (i.e. aposematism, eye-spots) are associated with longer life. Mud-puddling behaviour as a means of gaining access to minerals, and possibly nitrogen, was generally associated with a short life. We also found strong impacts of the region of occurrence of species (i.e. whether they were tropical or temperate taxa), but received conflicting results depending on whether we analysed field data (longer life in the tropics) or data from caged individuals (shorter life in the tropics). Adult longevity was associated with the growth form of larval host plants (higher on lianas as opposed to herbs, shrubs, or trees), whereas other larval host plant traits did not have strong links with species' life spans. We discuss the hypothetical evolutionary mechanisms explaining those patterns and suggest predictions that could be tested in further research.  © 2009 The Linnean Society of London, Biological Journal of the Linnean Society , 2009, 96 , 166–184.     

Nectar Production in Louisiana Iris Hybrids

Nectar is an important attractant for pollinators, and a plant's success in sexual reproduction can be influenced by the amount and concentration of nectar produced by its flowers. We studied nectar production over flower lifetime in Iris fulva, Iris brevicaulis, and four classes of hybrids-reciprocal F1's and backcrosses-between these species. Iris fulva produced less concentrated nectar than did I. brevicaulis, whereas I. brevicaulis flowers had a shorter life span. Hybrids were not intermediate, but they had the high nectar concentration of I. brevicaulis combined with the long life span of I. fulva flowers. Nectar production and concentration declined after the first day in all classes, but flowers continued to produce nectar until they were completely wilted. Backcrosses did not show a shift in mean or increased variation for the characters that distinguished the parental species; backcrosses toward I. fulva retained the high nectar concentration of I. brevicaulis, and backcrosses toward I. brevicaulis did not have a reduced flower life span. Overall, F1 hybrid flowers produced the highest amounts of nectar and nectar sugar over their life spans. These results, together with previously obtained data on pollinator choice in mixed arrays of the same flower classes, show that F1 hybrids between these species do not suffer from reduced attractiveness to pollinators. F1 individuals produced more nectar and nectar sugar than did their parents, and thus, they are possibly even more attractive to pollinators that forage for nectar.     

The importance of amino acids in the adult diet of male tropical rainforest butterflies

Adult diet is a major factor contributing to life history variation in the Lepidoptera. Amino acids derived from adult feeding, in particular, are suspected to be a major contribution to fitness, but this could be proven in only few species so far. Effects of amino acids in the diet of butterflies were experimentally investigated in 13 species from a Borneo rainforest community (using caged males without mating opportunity). Four species lived substantially longer when given a mix of amino acids additionally to water, sodium and sugar solutions. No significant phylogenetic pattern was found for effects of amino acid feeding, although none of six pierid species were among the taxa with significant effects. Species that do react to amino acids tend to be among the most long-lived taxa in the community, suggesting that amino acids are a key variable to attain long life spans. Weaker effects, or impacts on other fitness-related variables, cannot be excluded in the remaining taxa. Results indicate that adult amino acid intake may not be a rare strategy of few exotic taxa, but is, at least in non-seasonal tropical regions, a common life history trait in a substantial number of butterfly species.     

Carbon source for reproduction in a spring ephemeral herb, Corydalis ambigua (Papaveraceae)

1.  The carbon source for reproduction in plants may differ between flowering and fruiting stages. To clarify how spring ephemerals use current photosynthetic products for reproduction, the allocation patterns of photosynthate at flowering and fruiting and the effects of resource limitation on reproductive performance in Corydalis ambigua were assessed.
2.  A ¹³C tracing experiment revealed that about 20% of the current photosynthetic carbon was used for reproduction at both flowering and fruiting. The proportion of ¹³C allocated to fruits was constant irrespective of the light level. In contrast, ¹³C translocation to tubers increased at fruiting, and this trend was accelerated when plants were shaded.
3.  Defoliation treatment significantly reduced nectar production and tuber mass, while seed production was not affected. Therefore, when carbon assimilation was limited, carbon was preferentially allocated to current reproduction (seeds) rather than to pollinator attraction (nectar) or storage (tuber).
4.  If seed production is partly supported by carbohydrate reserved in the old tissue of tubers, nectar and seed production may not compete strongly for carbon sources. In contrast to the ability of high seed production, the susceptibility of nectar production to current photosynthesis indicates that seed production of this species is basically limited by pollen capture.
5.  Therefore, temporal separation of resource pool for reproduction may mitigate the joint limitation of seed production between pollinator attraction and resource availability. Temporal variation of the sink–source balance of storage organ is crucial to understand the cost of reproduction in perennial plants.     

The amino acids used in reproduction by butterflies: a comparative study of dietary sources using compound-specific stable isotope analysis

It is a nutritional challenge for nectar-feeding insects to meet the amino acid requirements of oviposition. Here we investigate whether egg amino acids derive from larval diet or are synthesized from nectar sugar in four species of butterfly: Colias eurytheme, Speyeria mormonia, Euphydryas chalcedona, and Heliconius charitonia. These species exhibit a range of life history and differ in degree of shared phylogeny. We use 13C differences among plants to identify dietary sources of amino acid carbon, and we measure amino acid 13C using compound-specific stable isotope analysis. Egg essential amino acids derived solely from the larval diet, with no evidence for metabolic carbon remodeling. Carbon in nonessential amino acids from eggs derived primarily from nectar sugars, with consistent variation in amino acid turnover. There was no relationship between the nonessential amino acids of eggs and host plants, demonstrating extensive metabolic remodeling. Differences between species in carbon turnover were reflected at the molecular level, particularly by glutamate and aspartate. Essential amino acid 13C varied in a highly consistent pattern among larval host plants, reflecting a common isotopic "fingerprint" associated with plant biosynthesis. These data demonstrate conservative patterns of amino acid metabolism among Lepidoptera and the power of molecular stable isotope analyses for evaluating nutrient metabolism in situ.     

Nectar sugar and amino acid preferences of Battus philenor (Lepidoptera, Papilionidae)

Abstract.

• 1 Butterflies of Battus philenor were tested for their preferences for nectar sugars and amino acids in an outdoor cage experiment.
• 2 The butterflies clearly preferred both sucrose and fructose over glucose. They also preferred sucrose over fructose.
• 3 No other preferences were found to be statistically significant, although male butterflies tended to prefer a plain sugar solution over a sugar solution containing a mixture of amino acids: females consumed both of these solutions in almost equal proportions.
• 4 The results are discussed with respect to nectar composition of butterfly pollinated flowers, flower preferences, physiological and reproductive aspects of butterflies.

     

Effect of Adult Feeding on Male Mating Behaviour in the Butterfly, <Emphasis Type="Italic">Bicyclus anynana</Emphasis> (Lepidoptera: Nymphalidae)

Many species of lepidopterans supplement their nectar diet with foods rich in nitrogen and minerals, which are present only in trace amounts in nectar. We examined the effect of adult diet on mating behaviour and spermatophore characteristics in male Bicyclus anynana (Butler, 1879) butterflies, which feed on rotten fruits as adults. We found little effect of adult diet on male reproduction in terms of mating rate and sperm production, although males fed on fruit produced larger spermatophores on their first mating compared to males fed sugar only. We also examined how males allocate sperm across matings. Males ejaculate larger spermatophores during their first mating, and produce spermatophores containing decreasingly fewer non-fertile sperm with number of matings performed. Males that produced more non-fertile sperm on their first mating had reduced lifespan possibly indicating a trade-off between sperm production and adult longevity. It is suggested that adult diet has little affect on male ejaculate production and males feed on fruit to supplement their energetic carbon requirements.     

The effect of developmental nutrition on life span and fecundity depends on the adult reproductive environment in Drosophila melanogaster

Both developmental nutrition and adult nutrition affect life‐history traits; however, little is known about whether the effect of developmental nutrition depends on the adult environment experienced. We used the fruit fly to determine whether life‐history traits, particularly life span and fecundity, are affected by developmental nutrition, and whether this depends on the extent to which the adult environment allows females to realize their full reproductive potential. We raised flies on three different developmental food levels containing increasing amounts of yeast and sugar: poor, control, and rich. We found that development on poor or rich larval food resulted in several life‐history phenotypes indicative of suboptimal conditions, including increased developmental time, and, for poor food, decreased adult weight. However, development on poor larval food actually increased adult virgin life span. In addition, we manipulated the reproductive potential of the adult environment by adding yeast or yeast and a male. This manipulation interacted with larval food to determine adult fecundity. Specifically, under two adult conditions, flies raised on poor larval food had higher reproduction at certain ages – when singly mated this occurred early in life and when continuously mated with yeast this occurred during midlife. We show that poor larval food is not necessarily detrimental to key adult life‐history traits, but does exert an adult environment‐dependent effect, especially by affecting virgin life span and altering adult patterns of reproductive investment. Our findings are relevant because (1) they may explain differences between published studies on nutritional effects on life‐history traits; (2) they indicate that optimal nutritional conditions are likely to be different for larvae and adults, potentially reflecting evolutionary history; and (3) they urge for the incorporation of developmental nutritional conditions into the central life‐history concept of resource acquisition and allocation.