Plant Latex and First-Instar Monarch Larval Growth and Survival on Three North American Milkweed Species

First-instar larvae of the monarch butterfly, Danaus plexippus, a milkweed specialist, generally grew faster and survived better on leaves when latex flow was reduced by partial severance of the leaf petiole. The outcome depended on milkweed species and was related to the amount of latex produced. The outcome also may be related to the amount of cardenolide produced by the plants as a potential chemical defense against herbivory. Growth was more rapid, but survival was similar on partially severed compared with intact leaves of the high-latex/low-cardenolide milkweed, Asclepias syriaca, whereas both growth and survival were unaffected on the low-latex/low-cardenolide milkweed A. incarnata. On the low-latex/low-cardenolide milkweed A. tuberosa, both growth and survival of larvae were only marginally affected. These results contrast sharply to previous results with the milkweed, A. humistrata, in Florida, which has both high latex and high cardenolide. Larval growth and survival on A. humistrata were both increased by partially severing leaf petioles. Larval growth rates among all four milkweed species on leaves with partially severed petioles were identical, suggesting that latex and possibly the included cardenolides are important in first-instar monarch larval growth, development, and survivorship.     

Seasonal and intraplant variation of cardenolide content in the California milkweed,Asclepias eriocarpa,and implications for plant defense

Root, stem, leaf, and latex samples ofAsclepias eriocarpa collected from three plots in one population at 12 monthly intervals were assayed for total cardenolide content by spectroassay and for individual cardenolides by thin-layer chromatography. From May to September mean milligram equivalents of digitoxin per gram of dried plant were: latices, 56.8 stems, 6.12 > leaves, 4.0 > roots, 2.5. With the exception of the roots, significant changes in gross cardenolide content occurred for each sample type with time of collection during the growing season, whereas variation within this population was found to be small. Labriformin, a nitrogen-containing cardenolide of low polarity, predominated in the latices. Leaf samples contained labriformin, labriformidin, desglucosyrioside, and other unidentified cardenolides. In addition to most of the same cardenolides as the leaves, the stems also contained uzarigenin. The roots contained desglucosyrioside and several polar cardenolides. The results are compared with those for other cardenolide-containing plants, and discussed in relation to anti-herbivore defense based on plant cardenolide content. Arguments are advanced for a central role of the latex in cardenolide storage and deployment which maximizes the defensive qualities of the cardenolides while preventing toxicity to the plant.Research supported by National Science Foundation Grants DEB 75-14266 and DEB 78-15419 (U.C. Davis) and DEB 75-14265, DEB 78-10658, and DEB 80-40388 (U.F.)     

Selective sequestration of milkweed (Asclepias sp.) cardenolides inOncopeltus fasciatus (Dallas) (Hemiptera: Lygaeidae)

The cardenolide content of the gut, wings, and fat body ofOncopeltus fasciatus was examined. The female fat body contained 4–5% of the total cardenolide content of the insect. The cardenolide content of male fat body, and gut and wings of both sexes was below the detection limit of the cardenolide assay. Thin-layer chromatography was used to determine the cardenolide array of various tissues and secretions ofO. fasciatus reared on seeds of a single species of milkweed (A. Speciosa) and adult extracts and dorsolateral space fluid ofO. fasciatus reared on seeds of two species of milkweed with different cardenolide arrays (A. speciosa andA. syriaca). Our results indicate that cardenolides are not sequestered in the insect simply on the basis of polarity and that metabolism and differential excretion of cardenolides are involved in the sequestration of cardenolides inO. fasciatus. The similarities in the cardenolide profiles ofO. fasciatus reared on different food sources, and tissues ofO. fasciatus reared on a single food source indicates that there is regulation of the cardenolide array inO. Fasciatus.     

Interspecific Variation Within the Genus <Emphasis Type="Italic">Asclepias</Emphasis> in Response to Herbivory by a Phloem-feeding Insect Herbivore

Induced plant responses to leaf-chewing insects have been well studied, but considerably less is known about the effects of phloem-feedings insects on induction. In a set of laboratory experiments, we examined density-dependent induction by the milkweed-oleander aphid, Aphis nerii, of putative defenses in four milkweed species (Asclepias incarnata, Asclepias syriaca, Asclepias tuberosa, and Asclepias viridis). We hypothesized that high aphid density would lead to increased cardenolide expression in species with low constitutive levels of cardenolides (e.g., A. tuberosa), but that there would be no induction in high constitutive cardenolide species (e.g., A. viridis). Based on previous studies, we did not expect cardenolide induction in A. incarnata. Contrary to our predictions, we observed feeding-induced declines of cardenolide concentrations in A. viridis. Cardenolide concentrations did not respond to aphid feeding in the other three milkweed species. Aphids also caused reductions in biomass accumulation by two of four Asclepias species, A. viridis and A. incarnata. High aphid density led to a decrease in A. viridis foliar nitrogen concentration. However, aphids had no effect on the defensive chemistry, growth, or nutritional quality of either A. syriaca or A. tuberosa. Our results highlight that congeneric plant species may respond differently to the same levels of herbivore damage.     

Density-Dependent Reduction and Induction of Milkweed Cardenolides by a Sucking Insect Herbivore

The effect of aphid population size on host-plant chemical defense expression and the effect of plant defense on aphid population dynamics were investigated in a milkweed-specialist herbivore system. Density effects of the aposematic oleander aphid, Aphis nerii, on cardenolide expression were measured in two milkweed species, Asclepias curassavica and A. incarnata. These plants vary in constitutive chemical investment with high mean cardenolide concentration in A. curassavica and low to zero in A. incarnata. The second objective was to determine whether cardenolide expression in these two host plants impacts mean A. nerii colony biomass (mg) and density. Cardenolide concentration (microgram/g) of A. curassavica in both aphid-treated leaves and opposite, herbivore-free leaves decreased initially in comparison with aphid-free controls, and then increased significantly with A. nerii density. Thus, A. curassavica responds to aphid herbivory initially with density-dependent phytochemical reduction, followed by induction of cardenolides to concentrations above aphid-free controls. In addition, mean cardenolide concentration of aphid-treated leaves was significantly higher than that of opposite, herbivore-free leaves. Therefore, A. curassavica induction is strongest in herbivore-damage tissue. Conversely, A. incarnata exhibited no such chemical response to aphid herbivory. Furthermore, neither host plant responded chemically to herbivore feeding duration time (days) or to the interaction between herbivore initial density and feeding duration time. There were also no significant differences in mean colony biomass or population density of A. nerii reared on high cardenolide (A. curassavica) and low cardenolide (A. incarnata) hosts.     

Variation in cardenolide content of the lygaeid bugs,Oncopeltus fasciatus andLygaeus kalmii kalmii and of their milkweed hosts (Asclepias spp.) in central California

A colorimetric assay was used to quantify the amount of cardenolides in the lygaeid bugsOncopeltus fasciatus andLygaeus kalmii kalmii and their milkweed host plants (Asclepias spp.) in central California. The cardenolide content of individual insects, determined in microgram equivalents of digitoxin, varied from zero to over 300 g per insect. Sources of variation of cardenolide content in the insects include interspecific and intraspecific differences in the content of the host plant species and also differences in the content of plant organs on which insects were feeding. This last source of variability may explain temporal variation in the cardenolide content of the insects. Adults ofO. fasciatus, which migrate into California in the late spring and early summer, and adults ofL. k. kalmii, which emerge from winter hibernacula in the early spring, contained small to immeasureable amounts of cardenolides. The colonization pattern ofO. fasciatus on species ofAsclepias in north central California suggests that this species does not maximize its opportunities to sequester large quantities of cardenolides from potential hosts. The emetic potential of lygaeids in California to vertebrate predators is briefly discussed.     

Cardenolide fingerprint of monarch butterflies reared on common milkweed,Asclepias syriaca L.

Monarch butterfly,Danaus plexippus (L.), larvae were collected during August 1983 from the common milkweed,Asclepias syriaca L., across its extensive North American range from North Dakota, east to Vermont, and south to Virginia. This confirms that the late summer distribution of breeding monarchs in eastern North America coincides with the range of this extremely abundant milkweed resource. Plant cardenolide concentrations, assayed by spectrophotometry in 158 samples from 27 collection sites, were biased towards plants with low cardenolide, and ranged from 4 to 229 g/ 0.1 g dry weight, with a mean of 50 g/0.1 g. Monarch larvae reared on these plants stored cardenolides logarithmically, and produced 158 adults with a normally distributed concentration range from 0 to 792 g/0. l g dry butterfly, with a mean of 234 g/0.1 g. Thus butterflies increased the mean plant cardenolide concentration by 4.7. The eastern plants and their resultant butterflies had higher cardenolide concentrations than those from the west, and in some areas monarchs sequestered more cardenolide from equivalent plants. Plants growing in small patches had higher cardenolide concentrations than those in larger patches, but this did not influence butterfly concentration. However, younger plants and those at habitat edges had higher cardenolide concentrations than either older, shaded, or open habitat plants, and this did influence butterfly storage. There were no apparent topographical differences reflected in the cardenolides of plants and butterflies. Twenty-eight cardenolides were recognized by thin-layer chromatography, with 27 in plants and 21 in butterflies. Butterflies stored cardenolides within the more polar 46% of the plantR _d range, these being sequestered in higher relative concentrations than they occurred in the plants. By comparison with published TLC cardenolide mobilities, spots 3, 4, 9, 16, 24 or 25, 26, and 27, may be the cardenolides syrioside, uzarin, syriobioside, syriogenin, uzarigenin, labriformidin, and labriformin, respectively. Cochromatography with cardenolide standards indicated that desglucosyrioside did not occur in the plants but did occur in 70% of the butterflies, and aspecioside was in 99% of the plants and 100% of the butterflies. The polar aspecioside was the single most concentrated and diagnostic cardenolide in both plants and butterflies. ButterflyR _d values were dependent on those of the plant, and both showed remarkable uniformity over the range of areas sampled. Thus contrary to previous reports,A. syriaca has a biogeographically consistent cardenolide fingerprint pattern. The ecological implications of this for understanding the monarch's annual migration cycle are significant.     

Genetic and Environmental Sources of Variation in the Autogenous Chemical Defense of a Leaf Beetle

Chemical defense plays a central role for many herbivorous insects in their interactions with predators and host plants. The leaf beetle genus Oreina (Coleoptera, Chrysomelidae) includes species able to both sequester pyrrolizidine alkaloids and autogenously produce cardenolides. Sequestered compounds are clearly related to patterns of host-plant use, but variation in de novo synthesized cardenolides is less obviously linked to the environment. In this study, intraspecific variation in cardenolide composition was examined by HPLC–MS analysis in 18 populations of Oreina speciosa spanning Europe from the Massif Central to the Balkans. This revealed the defense secretion to be a complex blend of up to 42 compounds per population. There was considerable geographical variation in the total sample of 50 compounds detected, with only 14 found in all sites. The environmental and genetic influences on defense chemistry were investigated by correlation with distance matrices based on habitat factors, host-plant use, and genetics (sequence data from COI, COII, and 16s rRNA). This demonstrated an influence of both genetics and host-plant use on the overall blend of cardenolides and on the presence of some of the individual compounds. The implications of this result are discussed for the evolution of defense chemistry and for the use of cardenolide compounds as markers of the evolutionary history of the species. Triponez and Naisbit contributed equally to this work and are considered joint first authors. An erratum to this article can be found at     

Cardenolide sequestration by the dogbane tiger moth (Cycnia tenera; Arctiidae)

Cycnia tenera adults, reared as larvae onAsclepias humistrata, had 10 times higher cardenolide concentrations, and contained 15 times more total cardenolide, than did moths reared onA. tuberosa. Thin-layer chromatography confirmed that each individual cardenolide visualized in the adult moths reared on the former host plant corresponds to one present in the plant, thus demonstrating that the insects' cardenolides are indeed derived from the larval food. Adult weights were significantly greater when the larvae had been fed upon the higher cardenolide plant species,A. humistrata. Similar results for other milkweed-feeding insects have been interpreted by some authors as evidence against a metabolic cost of handling cardenolides. However, such interpretations confound cardenolide differences among milkweed species with other differences in plant primary and secondary chemistry that affect insect growth and development. While the cooccurrence inC. tenera of other noxious chemicals (e.g., alkaloids) is not precluded, cardenolides sequestered from larval host plants have probably contributed to the evolution of visual and auditory aposematism in this species. As the eggs are laid in large clutches and larvae are gregarious, such aposematism may have evolved via kin selection.     

Induced Responses to Herbivory and Jasmonate in Three Milkweed Species

We studied constitutive and induced defensive traits (latex exudation, cardenolides, proteases, and C/N ratio) and resistance to monarch caterpillars (Danaus plexippus) in three closely related milkweed species (Asclepias angustifolia, A. barjoniifolia and A. fascicularis). All traits showed significant induction in at least one of the species. Jasmonate application only partially mimicked the effect of monarch feeding. We found some correspondence between latex and cardenolide content and reduced larval growth. Larvae fed cut leaves of A. angustifolia grew better than larvae fed intact plants. Addition of the cardenolide digitoxin to cut leaves reduced larval growth but ouabain (at the same concentration) had no effect. We, thus, confirm that latex and cardenolides are major defenses in milkweeds, effective against a specialist herbivore. Other traits such as proteases and C/N ratio additionally may be integrated in the defense scheme of those plants. Induction seems to play an important role in plants that have an intermediate level of defense, and we advocate incorporating induction as an additional axis of the plant defense syndrome hypothesis.     

Chemical constituents ofErysimum cheiranthoides deterring oviposition by the cabbage butterfly,Pieris rapae

Avoidance ofErysimum cheiranthoides for oviposition byPieris rapae has been attributed to the presence of water-soluble deterrents. The active material was extracted inton-butanol and isolated by a series of HPLC separations. TLC of the active fraction and visualization of individual constituents with Kedde's reagent indicated that cardenolides are responsible for deterring oviposition. UV spectra were also characteristic of cardenolides. Bioassays of selected known cardenolides revealed a general lack of activity, except for cymarin, which was as strongly deterrent as the most prominent cardenolide isolated in pure form fromE. cheiranthoides. The results suggest that cardenolides in this plant can explain its escape from cabbage butterflies, but specific structural features of the glycosides are necessary for oviposition-deterring activity.     

Cardenolide connection between overwintering monarch butterflies from Mexico and their larval food plant,Asclepias syriaca

The majority (85%) of 394 monarch butterflies sampled from overwintering sites in Mexico contain the same epoxy cardenolide glycosides, including most conspicuously a novel polar glycoside with a single genin-sugar bridge (aspecioside), as occur in the milkweedsAsclepias speciosa andA. syriaca. This cardenolide commonality was established by isolating aspecioside and syriobioside from the wings of overwintering monarchs and the two plant species, and comparing Chromatographie and NMR spectrometric characteristics of the isolates. When combined with the migratory pattern of monarchs and the distribution of these two milkweed species, this chemical evidence lends strong support to the hypothesis thatA. syriaca is the major late summer food plant of monarchs in eastern North America. This finding may be of ecological importance, forA. syriaca contributes less cardenolide and cardenolides of lower emetic potency to monarchs than most milkweeds studied to date.Research supported by National Science Foundation Grants DEB 81-19391 (UC Davis) and BSR-81-19382 (Univ. of Florida).     

Plant-determined variation in the cardenolide content,thin-layer chromatography profiles,and emetic potency of monarch butterflies,Danaus plexippus L. Reared on milkweed plants in California: 2.Asclepias speciosa

The pattern of variation in gross cardenolide concentration of 111Asclepias speciosa plants collected in six different areas of California is a positively skewed distribution which ranges from 19 to 344 g of cardenolide per 0.1 g dry weight with a mean of 90 g per 0.1 g. Butterflies reared individually on these plants in their native habitats ranged from 41 to 547 g of cardenolide per 0.1 g dry weight with a mean of 179 g. Total cardenolide per butterfly ranged from 54 to 1279 g with a mean of 319 g. Differences in concentrations and total cardenolide contents in the butterflies from the six geographic areas appeared minor, and there were no differences between the males and the females, although the males did weigh significantly more than females. The uptake of cardenolide by the butterflies was found to be a logarithmic function of the plant concentration. This results in regulation: larvae which feed on low-concentration plants produce butterflies with increased cardenolide concentrations relative to those of the plants, and those which feed on high-concentration plants produce butterflies with decreased concentrations. No evidence was adduced that high concentrations of cardenolides in the plants affected the fitness of the butterflies. The mean emetic potencies of the powdered plant and butterfly material were 5.62 and 5.25 blue jay emetic dose fifty units per milligram of cardenolide and the number of ED₅₀ units per butterfly ranged from 0.28 to 6.7 with a mean of 1.67. Monarchs reared onA. speciosa, on average, are only about one tenth as emetic as those reared onA. eriocarpa. UnlikeA. eriocarpa which is limited to California,A. speciosa ranges from California to the Great Plains and is replaced eastwards byA. syriaca L. These two latter milkweed species appear to have a similar array of chemically identical cardenolides, and therefore both must produce butterflies of relatively low emetic potency to birds, with important ecological implications. About 80% of the lower emetic potency of monarchs reared on A. speciosa compared to those reared onA. eriocarpa appears attributable to the higher polarity of the cardenolides inA. speciosa. Thin-layer Chromatographie separation of the cardenolides in two different solvent systems showed that there are 23 cardenolides in theA. speciosa plants of which 20 are stored by the butterflies. There were no differences in the cardenolide spot patterns due either to geographic origin or the sex of the butterflies. As when reared onA. eriocarpa, the butterflies did not store the plant cardenolides withR _f values greater than digitoxigenin. However, metabolic transformation of the cardenolides by the larvae appeared minor in comparison to when they were reared onA. eriocarpa. AlthoughA. eriocarpa andA. speciosa contain similar numbers of cardenolides and both contain desglucosyrioside, the cardenolides ofA. speciosa overall are more polar. ThusA. speciosa has no or only small amounts of the nonpolar labriformin and labriformidin, whereas both occur in high concentrations inA. eriocarpa. A. speciosa plants and butterflies also contain uzarigen, syriogenin, and possibly other polar cardenolides withR _f values lower than digitoxin. The cardenolide concentration in the leaves is not only considerably less than inA. eriocarpa, but the latex has little to immeasurable cardenolide, whereas that ofA. eriocarpa has very high concentrations of several cardenolides. Quantitative analysis ofR _f values of the cardenolide spots, their intensities, and their probabilities of occurrence in the chloroform-methanol-formamide TLC system produced a cardenolide fingerprint pattern very different from that previously established for monarchs reared onA. eriocarpa. This dispels recently published skepticism about the predictibility of chemical fingerprints based upon ingested secondary plant chemicals.Lepidoptera: Danaidae.Apocynales: Asclepiadaceae.This study was supported by U.S. National Science Foundation grants DEB 75-14265 and 78-10658 to Amherst College and BSR-8119382 to the University of Florida with L.P. Brower as Principal Investigator and DEB75-14266, DEB78-15419, and DEB-81-19391 to the University of California at Davis with J.N. Seiber as Principal Investigator.     

Disruption of web structure and predatory behavior of a spider by plant-derived chemical defenses of an aposematic aphid

Two toxic and bitter-tasting cardenolides (cardiac-active steroids) were sequestered by the brightly colored oleander aphid,Aphis nerii B. de F., from the neotropical milkweed host plantAsclepias curassavica L. After feeding on milkweed-reared aphids, the orb-web spiderZygiella x-notata (Clerck) built severely disrupted webs and attacked fewer nontoxic, control aphids, whereas the webs of spiders fed only nontoxic aphids remained intact. The regularity and size of the prey-trapping area of webs were reduced significantly in proportion to the amount of toxic aphids eaten. The effects of toxic aphids on spider web structure were mimicked by feeding spiders the bitter-tasting cardenolide digitoxin, a cardenolide with similar steroidal structure and pharmacological activity to the two aphid cardenolides. These results show that the well-known effects of psychoactive drugs on spider web structure are more than interesting behavioral assays of drag activity. Similar effects, produced by plant-derived chemicals in the spider's aphid prey, are relevant to the ecology and evolution of interactions between prey defense and predator foraging.     

Effect of nitrogen and water treatment on leaf chemistry in horsenettle (Solanum carolinense), and relationship to herbivory by flea beetles (Epitrix spp.) and tobacco hornworm (Manduca sexta)

We studied the interaction between plants (horsenettle; Solanum carolinense) and herbivorous insects (flea beetles; Epitrix spp., and tobacco hornworm; Manduca sexta) by focusing on three questions: (1) Does variation in nitrogen availability affect leaf chemistry as predicted by the carbon-nutrient balance (CNB) hypothesis? (2) Does variation in plant treatment and leaf chemistry affect insect feeding? (3) Is there an interaction between the insect herbivores that is mediated by variation in leaf chemistry? For three successive years (1998-2001), we grew a set of clones of 10 maternal plants under two nitrogen treatments and two water treatments. For each plant in the summer of 2000, we assayed herbivory by hornworms in both indoor (detached leaf) and outdoor (attached leaf) assays, as well as ambient flea beetle damage. Estimates of leaf material consumed were made via analysis of digitized leaf images. We also assayed leaves for total protein, phenolic, and glycoalkaloid content, and for trypsin inhibitor, polyphenol oxidase, and peroxidase activity. Despite strong effects of nitrogen treatment on growth and reproduction, only total protein responded as predicted by CNB. Leaf phenolic levels were increased by nitrogen treatment, polyphenol oxidase activity was decreased, and other leaf parameters were unaffected. Neither hornworm nor flea beetle herbivory could be related to plant treatment or genotype or to variation in any of the six leaf chemical parameters. A negative relationship between flea beetle and hornworm herbivory was found, but was not apparently mediated by any of the measured leaf chemicals. Because leaf resistance was maintained in low nitrogen plants at the apparent expense of growth and reproduction, our results support the concept of a fitness cost of defense, as predicted by the optimal defense hypothesis.     

Volatile Emissions from an Odorous Plant in Response to Herbivory and Methyl Jasmonate Exposure

Induced volatile terpenes have been commonly reported among diverse agricultural plant species, but less commonly investigated in odorous plant species. Odorous plants synthesize and constitutively store relatively large amounts of volatiles, and these may play a role in defense against herbivores. We examined the effect of herbivory and methyl jasmonate (MeJA) exposure on the release of volatile organic compounds (VOCs) in the marsh elder, Iva frutescens, which contains numerous constitutive VOCs, mainly mono- and sesquiterpenes. Our specific goal was to test for the presence of inducible VOCs in a naturally occurring plant already armed with VOCs. The abundant, native specialist leaf beetle Paria aterrima was used in herbivore induction trials. VOCs were sampled from herbivore wounded and unwounded, and from MeJA treated and untreated I. frutescens. Total VOC emissions were significantly greater in response to herbivory and MeJA treatment compared to unwounded controls. Herbivore wounding caused a substantial shift in the emission profile (42 VOCs from wounded, compared to 8 VOCs from unwounded I. frutescens), and MeJA had a similar yet less substantial influence on the emission pattern (28 VOCs from MeJA treated compared to 8 VOCs from untreated I. frutescens). Constitutive VOC emissions predominated, but some VOCs were detected only in response to herbivory and MeJA treatment, suggesting de novo synthesis. Several VOCs exhibited a delayed emission profile in contrast to the rapid release of constitutive VOCs, and principal components analysis revealed they were not associated with constitutive emissions. While I. frutescens contains many constitutive VOCs that are released immediately in response to herbivory, it also produces novel VOCs in response to feeding by the specialist P. aterrima and MeJA treatment.     

The Effects of Milkweed Induced Defense on Parasite Resistance in Monarch Butterflies, <Emphasis Type="Italic">Danaus plexippus</Emphasis>

Many plants express induced defenses against herbivores through increasing the production of toxic secondary chemicals following damage. Phytochemical induction can directly or indirectly affect other organisms within the community. In tri-trophic systems, increased concentrations of plant toxins could be detrimental to plants if herbivores can sequester these toxins as protective chemicals for themselves. Thus, through trophic interactions, induction can lead to either positive or negative effects on plant fitness. We examined the effects of milkweed (Asclepias spp.) induced defenses on the resistance of monarch caterpillars (Danaus plexippus) to a protozoan parasite (Ophryocystis elektroscirrha). Milkweeds contain toxic secondary chemicals called cardenolides, higher concentrations of which are associated with reduced parasite growth. Previous work showed that declines in foliar cardenolides caused by aphid attack render monarch caterpillars more susceptible to infection. Here, we ask whether cardenolide induction by monarchs increases monarch resistance to disease. We subjected the high-cardenolide milkweed A. curassavica and the low-cardenolide A. syriaca to caterpillar grazing, and reared infected and uninfected caterpillars on these plants. As expected, monarchs suffered less parasite growth and disease when reared on A. curassavica than on A. syriaca. We also found that herbivory increased cardenolide concentrations in A. curassavica, but not A. syriaca. However, cardenolide induction in A. curassavica was insufficient to influence monarch resistance to the parasite. Our results suggest that interspecific variation in cardenolide concentration is a more important driver of parasite defense than plasticity via induced defenses in this tri-trophic system.     

Cardenolide content and thin-layer chromatography profiles of monarch butterflies,danaus plexippus L., and their larval host-plant milkweed,asclepias viridis walt., in northwestern louisiana

This paper is the first in a series on cardenolide fingerprinting of monarch butterflies and their host-plant milkweeds in the eastern United States. Spectrophotometric determinations of the gross cardenolide content of 60Asclepias viridis plants in northwestern Louisiana indicate a positively skewed variation ranging from 95 to 432 g/0.1 g dry weight with a mean of 245 g/0.1 g. Butterflies reared individually on these plants contained a normal cardenolide distribution ranging from 73 to 591 g/0.1 g dry weight with a mean of 337 g/0.1 g. The uptake of cardenolide by the butterflies best fit a logarithmic function of the plant concentration. Female monarchs (385 g/0.l g) contained significantly greater mean cardenolide concentrations than did males (287 g/0.1 g). No indications of a metabolic cost for either cardenolide ingestion or storage were adduced from size or dry weight data. Thin-layer chromatograms of 24 individual plant-butterfly pairs developed in two solvent systems resolved 21 individual spots in the plants and 15 in the butterflies.A. viridis plants appear to contain several relatively nonpolar cardenolides of the calotropagenin series which are metabolized to the more polar 3'-hydroxy derivatives calactin and calotropin as well as to calotropagenin in the butterflies. The epoxy cardenolides labriformin and labriformidin were absent, although desglucosyrioside (a 3'-hydroxy derivative) appeared present in both plants and butterflies. Quantitative evaluation of theR _f values, spot intensities, and probabilities of occurrence in the chloroform-methanol—formamide TLC system produced a cardenolide fingerprint clearly distinct from those previously established for monarchs reared on otherAsclepias species, supporting the use of fingerprints to make ecological predictions concerning larval host-plant utilization.A. viridis is the predominant early spring milkweed throughout most of the south central United States and may be important in providing chemical protection to spring and early summer generation monarchs in the eastern United States.Lepidoptera: Danaidae.Apocynales: Asclepiadaceae.     

Differences and similarities in cardenolide contents of queen and monarch butterflies in florida and their ecological and evolutionary implications

Florida queen butterflies are highly variable in cardenolide content and, in three populations studied, contained less cardenolide than did a sample of sympatric Florida monarchs. The possibility that queens stored a more potent set of cardenolides from their host plants (and therefore were as well protected as monarchs, even at lower concentrations) is refuted by Chromatographic analysis of wild butterflies, as well as controlled laboratory rearings. It therefore appears that, with respect to cardenolides, monarchs are better defended than are queens. Consequently, cardenolides are unlikely to explain the apparent shift in Florida viceroy mimicry away from resemblance of the monarch, toward mimicry of the queen. Other hypotheses to explain this mimetic phenomenon are suggested. Adult monarchs exhibit significant negative correlations between the concentration of cardenolide stored in their tissues and both body size and weight, whereas queens show no such correlations. The implications of these results for the study of metabolic costs of allelochemic storage are discussed. Chromatographic evidence is provided that monarchs do breed in south Florida during the winter months and that the likely host plant employed by the population studied wasAsclepias curassavica. This represents the first practical application of cardenolide fingerprinting to identify the larval host plants of wild danaid butterflies.