Glues or poisons: which triggers vein cutting by monarch caterpillars?

Summary. Late instar larvae of the monarch butterfly (Danaus plexippus L., Lepidoptera: Nymphalidae) deactivate the latex defense of milkweeds by chewing a furrow in the leaf midrib or petiole. The larva then feeds beyond the cut where latex outflow is minimal. If a larva does encounter latex exudate during feeding, it often returns to its initial cut to damage the midrib or petiole more extensively before resuming feeding. We used this response to latex as an assay for testing what cue triggers vein cutting. A sticky solution of polyethylene glycol and a mixture of cardenolides both were ineffective; drops applied to the mouthparts of feeding monarchs failed to elicit renewed vein cutting. Activity resided instead within a methanol extract of the supernatant obtained from centrifuged latex of the milkweed, Asclepias curassavica L. (Asclepiadaceae). Treatment with proteinase K did not eliminate activity documenting that the active compound is unlikely to be proteinaceous. Our results indicate that latex adhesiveness and low polarity cardenolides are relatively unimportant releasers of vein cutting. We propose that milkweeds contain noncardenolide stimulants of vein cutting; these compounds presumably serve a defensive role for milkweeds. Over 50 species of insects are known to employ vein-cutting before feeding on plants with canal-borne exudates; most of these species are dietary specialists like the monarch. Our behavioral assay provides a novel approach for identifying ecologically-significant compounds in the exudates of their diverse host plants.     

Potential direct fitness consequences of ornament-based mate choice in a butterfly

Female mate choice has been shown to provide direct mating benefits in several animal groups. In butterflies, for which there are increasing reports of fine-scale color-based mate choice, the evolutionary benefits that accrue from such mating biases, if any, are largely unknown. We addressed this issue in the butterfly Colias eurytheme, a species in which females choose mates on the basis of iridescent ultraviolet (UV) wing ornamentation and in which males donate reproductively beneficial nuptial gifts. In the first experiment, we assessed the mass of gifts donated to 77 virgin females by males sampled directly from a field encounter site. Despite large variance in the male adult phenotype and ejaculate, no single aspect of dorsal wing coloration, including UV brightness, chroma, or hue, was related to ejaculate mass. There was, however, an interesting interaction between the effects of male body size and copula duration upon ejaculate mass, with size scaling positively with ejaculate mass among males involved in shorter copulations (those lasting <70 min) but negatively among males in longer copulations. In the second experiment, we assessed the lifetime fecundity, fertility, and longevity of 85 females mated under similar circumstances to free-flying wild males. Although several wing color parameters proved subtly informative in more sophisticated multivariable models, no model predicted more than about 20% of the variation in any single female fitness parameter. The duration of copulation, which ranged from 35 min to over 16 h and which carries putative costs for females, was, again, only very weakly predicted by male wing color parameters (i.e., R ² = 0.089). Given the overall minor predictive power of male wing coloration in general and of UV brightness in particular, our results do not strongly support the hypothesis that female C. eurytheme prefer bright UV males to obtain direct benefits or to minimize the costs associated with lengthy copulations.     

Susceptibility of Eucalyptus globulus to Mnesampela privata defoliation in relation to a specific foliar wax compound

Summary. Increased abundance of several aliphatic benzyl and phenylethyl alkanoates were previously associated with reduced defoliation of Eucalyptus globulus Labill. by the autumn gum moth (Mnesampela privata Guenée) in two clonally replicated F₂ families. We examine the robustness of this association by comparing the abundance of the implicated foliar wax compounds with defoliation levels in three plantation sites and a genetics trial in northern Tasmania. At all locations, the aliphatic alkanoate, benzyl n-tetracosanoate (C₂₄), was found in significantly higher abundance in the foliar wax of E. globulus trees that had displayed low M. privata damage (designated resistant) compared to trees that had displayed high damage (designated susceptible). Further, when sites were combined, benzyl n-tetracosanoate (C₂₄) accounted for 16 % of variation in field defoliation. Field defoliation was also positively correlated to M. privata oviposition both in the field and on foliage sprigs in a cage. In the cage bioassay, oviposition accounted for 20 % of variation in field defoliation making it a better predictor of tree defoliation in the field than either field oviposition, which accounted for 9 % of variation, or screening foliage for the abundance of benzyl n-tetracosanoate (C₂₄). Despite both benzyl n-tetracosanoate (C₂₄) and oviposition being related to field defoliation, there was no statistically significant relationship between the abundance of benzyl n-tetracosanoate (C₂₄) and oviposition in the field or cage bioassay. Further work is therefore required to determine if increased levels of this compound in the foliar wax operates as a direct deterrent to M. privataoviposition.     

A synomone imparting distinct sex pheromone communication channels for Choristoneura rosaceana (Harris) and Pandemis limitata (Robinson) (Lepidoptera: Tortricidae)

Summary. Male obliquebanded leafrollers, Choristoneura rosaceana (Harris), were induced to respond to a pheromone source tainted with a behavioural antagonist, Z9-tetradecenyl acetate, when a source releasing the antagonist was placed 10 cm upwind of the tainted source in a wind tunnel. However, placement of the antagonist upwind of an attractive pheromone source did not interrupt pheromone-mediated responses. Placement of a source releasing Z9-tetradecenyl acetate, a minor pheromone component of the sympatric species, the threelined leafroller, Pandemis limitata (Robinson), upwind of a calling P. limitata female, reduced conspecific male pheromone-mediated response but resulted in upwind flight by male C. rosaceana and contact with heterospecific females. Male P. limitata locked on and flew upwind to but did not contact heterospecific females when a source releasing Z9-tetradecenyl acetate was positioned upwind of a calling C. rosaceana female. In the field, adaptation or habituation to Z9-tetradecenyl acetate caused by atmospheric treatment with this compound apparently resulted in reciprocal heterospecific pheromone responses. More C. rosaceana males were captured in traps baited with their pheromone and the behavioural antagonist in small field plots treated atmospherically with Z9-tetradecenyl acetate than in nontreated control plots. Fewer male P. limitata were captured in traps baited with their own pheromone, or with C. rosaceana pheromone tainted with Z9-tetradecenyl acetate in plots treated atmospherically with Z9-tetradecenyl acetate than in nontreated control plots. We argue that Z9-tetradecenyl acetate is an important synomone which assists in partitioning the sexual chemical communication channels of C. rosaceana and P. limitata. Received 9 February 1999; accepted 22 March 1999.     

Wind tunnel attraction of grapevine moth females, Lobesia Botrana, to natural and artificial grape odour

Summary. Host plant volatiles which attract insect herbivores for egg-laying are of principal interest with respect to insect ecology and evolution. Direct applications concern population monitoring and control through behavioural manipulation. Identification of behaviourally active plant secondary metabolites is essential also for plant breeding for insect resistance. Grapevine moth females Lobesia botrana are attracted by upwind flight to green grape berries Vitis vinifera. The headspace of grape berries was collected on air filters. A solvent extract of these filters, released from a sprayer, attracted females in the wind tunnel. The results demonstrate that volatile cues mediate attraction of grapevine moth females to grape berries, and that headspace collections capture the essence of this odour signal. The air filter extracts were examined by gas chromatography coupled with electroantennographic detection, and the compounds eliciting a consistent antennal response in L. botrana females were identified by mass spectrometry. The headspace collection apparatus was calibrated for collection efficiency of the active compounds. Their recovery rate ranged from 35 % for methyl salicylate to 83 % for (E,E)-α-farnesene. A synthetic ten-component blend was then formulated. The blend consisted of compounds eliciting an antennal response, formulated in a blend ratio corrected for differences in collection efficiency. Subsequent wind tunnel tests showed that female attraction to this synthetic ten-component blend was not significantly different from attraction to grape berries, or to headspace collections of the same berries. At a release rate of 35 ng/h of the most abundant compound (E)-β-caryophyllene, 20 % of the test females approached the source of sprayed headspace collections and the ten-component synthetic blend, respectively. In comparison, 100 g of green berries, releasing the main compound (E)-β-caryophyllene at a rate of ca. 4.7 ng/h, attracted 10 % of the females by upwind flight followed by source contact.     

Cardenolide content,emetic potency,and thin-layer chromatography profiles of monarch butterflies,Danaus plexippus,and their larval host-plant milkweed,Asclepias humistrata,in Florida

Summary This paper is the fourth in a series on cardenolide fingerprints of monarch butterflies (Danaus plexippus, Danainae) and their host-plant milkweeds (Asclepiadaceae) in the eastern United States. Cardenolide concentrations ofAsclepias humistrata plants from north central Florida ranged from 71 to 710 µg/0.1 g dry weight, with a mean of 417 µg/0.1 g. Monarchs reared individually on these plants contained cardenolide concentrations ranging from 243 to 575 µg/0.1 g dry weight, with a mean of 385 µg/0.1 g. Cardenolide uptake by butterflies was independent of plant concentration, suggesting that sequestration saturation occurs in monarchs fed cardenolide-rich host plants. Thinlayer chromatography resolved 19 cardenolides in the plants and 15 in the butterflies. In addition to humistratin,A. humistrata plants contained several relatively non-polar cardenolides of the calotropagenin series which are metabolized to more polar derivatives in the butterflies. These produced a butterfly cardenolide fingerprint clearly distinct from those previously established for monarchs reared on otherAsclepias species. In emetic assays with the blue jay,Cyanocitta cristata, the 50% emetic dose (ED₅₀) per jay was 57.1 µg, and the average number of ED₅₀ units per butterfly was 13.8, establishing that this important south eastern milkweed produces highly emetic, chemically defended monarchs. Our data provide further support for the use of cardenolide fingerprints of wild-caught monarchs to make ecological predictions concerning defence against natural enemies, seasonal movement and larval host-plant utilization by monarch butterflies during their annual cycle of migration, breeding and overwintering.     

Egg protein insolubility inLymantria dispar versus other forest Lepidoptera

Summary A standard buffer (5 mM phosphate at pH 7) which is used to extract protein from insect eggs provided complete protein solubility for eggs from three of four tree-feeding lepidopteran species: obliquebanded leaf roller (Choristoneura rosaceana), forest tent caterpillar (Malacosoma disstria), and the eastern tent caterpillar (Malacosoma americanum). Under the same extraction protocol, egg proteins from the gypsy moth (Lymantria dispar), remained nearly insoluble. An array of methods typically used to solubilize insect egg proteins were tried and all but the most denaturing (2% SDS) were ineffective. Extraction buffers with typically high pH values were then evaluated. The results indicated that 1) solubility of gypsy moth egg proteins was pH dependent, and full solubility of most egg proteins required the extraction buffer to have a pH of 12 or more prior to the addition of eggs. We also determined that 2) the gypsy moth egg has a buffering capacity which must be surpassed for complete protein extraction, 3) low salt/high pH buffers gave slightly higher total protein values than did high salt/high pH buffers, 4) parental nutritional history (host species utilized) and egg developmental state (pre-embryonatedvs postembryonated/pre-hatch) were unrelated to the requirements for complete egg protein solubilization, and 5) the presence of soluble phenolics, compounds that have the potential to bind to protein and cause insolubility, was confirmed for the gypsy moth egg with 2-D paper chromatography and several other tests. Based on these results, we present a hypothesis about the cause of egg protein insolubility in the gypsy moth.     

Male mating history and female fecundity in the Lepidoptera: do male virgins make better partners?

In insects, large ejaculate and associated materials, including spermatophores, appear to have evolved via sexual selection acting on males to either delay female remating or to increase the rate of egg-laying. It is also possible, however, that females use nutrients transferred during mating to increase their lifetime fecundity. If so, male ejaculate size may also have evolved under natural selection as a form of paternal investment. In Lepidoptera, males with a greater number of prior matings tend to produce smaller spermatophores. However, the reported effects of male mating history on female fecundity vary widely among species. We therefore performed a meta-analysis using data from 29 studies of 25 species. Overall, the reproductive output of females mated to virgin males was significantly higher than that of females mated to sexually experienced males (Hedges d=0.33, P<0.01). A sample size of around 145 females per male mating type is required to detect an effect of this size with 80% statistical power at =0.05 (two-tailed). There was no difference in mean effect size between butterflies/skippers and moths. After controlling for any effect of taxonomic group, however, the mean effect size for polyandrous species was significantly greater than that for monandrous species (Hedges d=0.45 vs 0.25, P=0.01). We then discuss possible reasons why male mating history, presumably acting through its effect on spermatophore size, might have a stronger effect in polyandrous than monandrous species.Communicated by A. Cockburn     

Variation of pyrrolizidine alkaloids in Ithomiinae: A comparative study between species feeding on Apocynaceae and Solanaceae

Summary The primitive, Apocynaceae-feeding Ithomiine,Tithorea harmonia, incorporates dehydropyrrolizidine alkaloids (PAs) from its larval foodplant (Prestonia acutifolia), rarely visiting PA sources pharmacophagously in the adult; females show higher concentrations of PAs than males, with similar variance. The close relativeAeria olena (feeding onP. coalita, without PAs) shows similar PA concentrations in both sexes and greater variation in males, like more advanced Solanaceae-feeding Ithomiine such asMechanitis polymnia, which likeA. olena obtain PAs by pharmacophagy in the adult (mainly males). This difference is due to the dynamics of PA incorporation in these species. Little variation in PA content was found among allopatric populations of the same species, but variation in available PA sources in different months was correlated with different average storage levels in the butterflies.     

Sequestration of plant secondary compounds by butterflies and moths

Summary A number of aposematic butterfly and moth species sequester toxic substances from their host plants. Some of these insects can detect the toxic compounds during food assessment. Some pipevine swallowtails use aristolochic acids among the host finding cues during oviposition and larval feeding and accumulate the toxins in the body tissues throughout all life stages. Likewise, a danaine butterfly,Idea leuconoe, which sequesters high concentrations of pyrrolizidine alkaloids in the body, lays eggs in response to the specific alkaloid components contained in the apocynad host. Insect species sharing the same poisonous host plants may differ in the degree of sequestration of toxins. Two closely ralated aposematic geometrid moth species,Arichanna gaschkevitchii andA. melanaria, sequester a series of highly toxic diterpenoids (grayanotoxins) in different degrees, while a cryptic geometrid species,Biston robstus, does not sequester the toxins, illustrating the diversity in adaptation mechanisms even within the same subfamily. By contrast, a number of lepidopteran species store the same compounds though feeding upon taxonomically diverse plant species. A bitter cyanoglycoside, sarmentosin, was characterised from several moth species in the Geometridae, Zygaenidae and Yponomeutidae, and from the apollo butterflies,Parnassius spp. (Papilionidae), although each species feeds on different groups of plants.Interspecific similarities and differences in life history and ecology are discussed in relation to variable characteristics of sequestration of plant compounds among these lepidopteran insects.