Uptake and metabolism of [<Superscript>14</Superscript>C]rinderine and [<Superscript>14</Superscript>C]retronecine in leaf-beetles of the genus <Emphasis Type="Italic"> Platyphora</Emphasis> and alkaloid accumulation in the exocrine defensive secretions

Summary. Sequestration and processing of pyrrolizidine alkaloids (PAs) by leaf beetles of the genus Platyphora were investigated. Tracer experiments with labeled alkaloids were performed with P. eucosma feeding on Koanophyllon panamense (Asteraceae, tribe Eupatorieae). P. eucosma catalyzes the same reactions previously demonstrated for P. boucardi specialized to Prestonia portobellensis (Apocynaceae): (i) epimerization of rinderine to intermedine; (ii) esterification of retronecine yielding insect-specific PAs; (iii) efficient transport of the PAs as free bases into the defensive secretions. P. bella feeding on Tournefortia cuspidata (Boraginaceae) shows the same sequestration behavior and ability to synthesize the specific retronecine esters. P. ligata, a species phylogenetically closely related to the PA adapted species and clustering in the same clade, but feeding on a host plant devoid of PAs, feeds easily on PA treated host-plant leaves, but does not sequester or metabolize PAs. P. kollari a species clustering outside the PA clade refused to feed on its food-plant leaves painted with PAs. The results are discussed in relation to host-plant selection of the PA adapted species and the role of PAs in chemical defense. Received 20 September 2002; accepted 18 November 2002.     

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.     

Tropane and pyrrolizidine alkaloids in the ithomiinesPlacidula euryanassa andMiraleria cymothoe (Lepidoptera: Nymphalidae)

Summary Larvae of the ithomiine butterflyPlacidula euryanassa sequester tropane alkaloids (TAs) from the host plantBrugmansia suaveolens and pass them through the pupae to freshly emerged adults. Wild caught adults also show in their tissues, variable amounts of pyrrolidizine alkaloids (PAs), probably sequestered from variable plant sources and subject to dynamics of incorporation, accumulation and utilization of PAs by ithomiine butterflies. The ratio TAs/PAs is also variable between different populations.Miraleria cymothoe, another ithomiine that feeds onB. suaveolens as larvae, does not sequester TAs from the host-plant, but sequesters PAs from plant sources visited by the adult butterflies. The main alkaloid found in both butterflies is lycopsamine, which also is the principal PA found in all genera of Ithomiinae.     

<Emphasis Type="Italic">Ceroplastes albolineatus</Emphasis>, the first scale insect shown to sequester pyrrolizidine alkaloids from its host-plant <Emphasis Type="Italic">Pittocaulon praecox</Emphasis>

Summary. The occurrence of pyrrolizidine alkaloids (PAs) in Pittocaulon (ex Senecio) praecox (Asteraceae) a species endemic to Mexico was established. The aboveground plant organs contain the 1,2-saturated monoester 7-angeloyl-dihydroxyheliotridane together with a small proportion of its 9-angeloyl isomer as major alkaloid. The monoesters are accompanied by the macrocyclic otonecine derivative senkirkine. Roots contain only related macrocyclic PAs with senecionine, senkirkine and platyphylline as major components; monoesters are absent. The broom-like succulent stems of P. praecox are infested by the scale insect Ceroplastes albolineatus conspicuously visible by its huge wax cover. All life-history stages, i.e. females, eggs, first instar nymphs (crawlers) and the wax cover were found associated with PAs. The measured PA concentrations clearly indicate sequestration. The highest PA concentrations (mg / g dry weight) reached are: mature females, 0.44; eggs, 0.58; crawlers, 0.37; wax cover, 0.08. The host plant as well as in the infesting scale insect contain the PAs exclusively as free bases. As a phloem-feeder C. albolineatus must acquire the PAs with the ingested phloem sap. This appears plausible since in Senecio species PA are transmitted and circulated through the phloem path. It is suggested that PAs may protect particularly the crawlers as the most endangered stage in the life-cycle of the scale insect.     

Detoxification of pyrrolizidine alkaloids by the harvestman <Emphasis Type="Italic">Mitopus morio</Emphasis> (Phalangidae) a predator of alkaloid defended leaf beetles

Summary. The harvestman Mitopus morio (Phalangidae) is a generalist predator. It is known to prey on larvae of the chrysomelid leaf beetle Oreina cacaliae defended by plant acquired pyrrolizidine alkaloids (PAs). Tracer feeding experiments were performed to determine how harvestmen tolerate protoxic PAs. Minced meat containing either [¹⁴C]senecionine or [¹⁴C]senecionine N-oxide was fed to M. morio and subsequently feces and bodies were analyzed. Labeled alkaloid N-oxide remained stable and was eliminated almost unaltered with the feces; only 10% was recovered as tertiary PA. In contrast, approximately 80% of labeled tertiary alkaloid (senecionine) ingested with the diet was N-oxidized and eliminated; the remaining 20% consisted of unchanged senecionine and a polar metabolite of unknown structure. Harvestmen process their diet by excreting digestive juice, indicated by bleaching of the meat color. Analysis of the processed diet revealed some N-oxidation of [¹⁴C]senecionine, suggesting the gut as the site of Noxidation. Analysis of the bodies of harvestmen 80 hours after the tracer feeding pulse revealed only trace amounts of the polar metabolite. Neither senecionine nor its N-oxide could be detected in the body extracts. The results are discussed in relation to the strategies of PA adapted insects to avoid accumulation of tertiary PAs in living tissues.     

Pyrrolizidine alkaloids and pentacyclic triterpene saponins in the defensive secretions of Platyphora leaf beetles

Summary. Field collected exocrine defensive secretions of nine neotropical Platyphora species were analyzed for the presence of plant acquired pyrrolizidine alkaloids (PAs) and pentacyclic triterpene saponins. All species secrete saponins. In addition, five species feeding on Tournefortia (Boraginaceae), Koanophyllon (Asteraceae, tribe Eupatorieae) and Prestonia (Apocynaceae) were shown to sequester PAs of the lycopsamine type, which are characteristic for species of the three plant families. The PA sequestering species commonly store intermedine, lycopsamine and their O^3′-acetyl or propionyl esters as well as O⁷- and O⁹-hydroxyisovaleryl esters of retronecine. The latter as well as the O^3′-acyl esters were not found in the beetles’ host plants, suggesting the ability of the beetles to esterify plant derived retronecine and intermedine or its stereoisomers. Despite the conformity of the beetles’ PA patterns, considerable inconsistencies exist regarding the PA patterns of the respective host plants. One host plant was devoid of PAs, while another contained only simple necines. Since the previous history of the field collected beetles was unknown this discrepancy remains obscure. In contrast to the Palearctic chrysomeline leaf beetles, e.g. some Oreina species which ingest and store PAs as their non-toxic N-oxides, Platyphora leaf beetles absorb and store PAs as the toxic free base (tertiary PA), but apparently avoid to accumulate PAs in the haemolymph. This suggests that Chrysolina and Platyphora leaf beetles developed different lines of adaptations in their parallel evolution of PA mediated chemical defense. Received 30 November 2000; accepted 5 February 2001     

Sequestration of pyrrolizidine alkaloids by larvae ofTellervo zoilus (Lepidoptera: Ithomiinae) and their role in the chemical protection of adults against the spiderNephila maculata (Araneidae)

Summary Life stages of the primitive Australian ithomiine butterflyTellervo zoilus and its larval hostplant, the apocynaceous vineParsonsia straminea, were quantitatively assayed for pyrrolizidine alkaloids (PAs). PAs were found in all stages, mainly as N-oxides, being most concentrated in larvae and freshly-emerged adults. Although adults feed at various confirmed PA sources this probably does not compensate for losses, as wild-caught adults had considerably lower concentrations of PAs. The main alkaloid present in both freshly-emerged adults and in leaves of the host-plant was lycopsamine (1b), stored by butterflies in the N-oxide form. Its presence in higher proportion, in relation to intermedine (1a), in larvae, pupae and adults ofTellervo in relation to the host-plants suggests the inversion of intermedine to lycopsamine by the insects. No 14-member ring macrocyclic PAs were detected in either food-plant or butterflies. Several other PAs were found in wild-caught adults reflecting visits to other PA sources. PAs were also found in high concentrations in freshly-emerged individuals of the danaineEuploea core bred onParsonsia straminea. Wild-caughtDanaus affinis had high PA levels acquired from adult feeding. Freshly emergedEuploea raised onIschnocarpus frutescens andDanaus raised onIschnostemma carnosum (both PA-free) were preyed on by the orb weaving spiderNephila maculata, and showed no PAs. In all cases where PAs were present, most butterflies were liberated, usually cut out of the web unharmed, byNephila. The spider's response was not closely linked to PA concentration, however, and may also depend on hunger levels and previous experience with PA-containing butterflies. All control and other non-PA containing butterflies were consumed although rejection of some body parts of freshly-emergedDanaus affinis suggests that compounds other than PAs may be involved.     

Pyrrolizidine alkaloids on three trophic levels – evidence for toxic and deterrent effects on phytophages and predators

Summary. Pyrrolizidine alkaloids (PAs) present a model system in the investigation of tritrophic interactions mediated by plant secondary compounds. However, their toxicity for insect herbivores has never been experimentally proven. Here, we demonstrate the toxic effects of a PA on growth and survival of the eri silk moth Philosamia ricini. In a feeding experiment, larvae of this generalist herbivore fed with an artificial PA diet gained weight significantly slower than control animals, and died as pupae. We suggest that derivatives of the ingested PA N-oxide damage developmental functions during metamorphosis. A tracer test with [¹⁴C]senecionine N-oxide revealed that the caterpillars lack adaptations that would prevent conversion of the chemical into the pro-toxic free base. In contrast, the PA adapted leaf beetle Longitarsus anchusae accumulates PAs as N-oxides. We tested the purpose of sequestration in this species as defence against predators. Through a series of prey choice experiments with three carabid predator species, chemically non-protected bark beetle pupae were chosen almost uniformly over L. anchusae pupae. In a following choice test with one of these predators, artificially PA-treated mealworm segments deterred the predator from feeding. Overall the study corroborates the immediate toxic effect of PAs on non-adapted herbivores and the protective effect that adapted insects may gain by sequestering them. It thereby underlines the potential for PAs to play a central role in multitrophic interactions between plants, phytophages and their predators.     

Pyrrolizidine alkaloids between plants and insects: A new chapter of an old story

Summary Among alkaloids the pyrrolizidine alkaloids (PAs) play a unique role in the interactions between plants and adapted insects. InSenecio spp. (Asteraceae) PAs are synthesized in the roots as alkaloidN-oxides which are specifically translocated into shootsvia the phloem-path and channeled to the preferred sites of storage (e.g. inflorescences) where they are stored in the cell vacuoles. In differentSenecio spp. senecionineN-oxide is produced as the common product of biosynthesis, which subsequentlyvia a number of simple but specific reactions is transformed into typical speciesspecific PA-patterns. Insects from diverse taxa sequester PAs for their own defense. Lepidopterans (e.g. arctiids such asTyria jacobaeae andCreatonotos transiens) may hydrolyze plant acquired ester-PAs and convert the resulting necine base into insect-specific PAs by esterification with an acid of their own metabolism. Adapted arctiids and the grasshopperZonocerus take up PAs in the state of the tertiary amine.N-Oxides are reduced in the guts prior to uptake. In the bodies the tertiary PAs are rapidlyN-oxidized by a specific mixed-function oxigenase and are maintained in theN-oxide state. The importance of the reversible interconversion of the nontoxicN-oxide (pro-toxine) into the toxic tertiary alkaloid is discussed as the specific feature of PAs in plant-insect interactions.     

Female sex pheromone in immature insect males—a case of pre-emergence chemical mimicry?

Sexual selection by competition for mates is a formidable force that has led to extraordinary adaptations in males. Here we present results suggesting a novel case of pheromone mimicry in males of Lariophagus distinguendus, a parasitic wasp of beetle larvae that develop in stored grain. Females of L. distinguendus produce a pheromone even before they emerge from a grain. Males are attracted to the parasitised grain and wait for females to emerge. Males emerging later than others are under enormous selection pressure since females mate only once. We show evidence that developing males fool their earlier emerging competitors by mimicking the female pheromone. Males exposed to pupae of either sex exhibit typical courtship behaviour. Searching males are not only arrested by grains containing developing females but spend as much time on grains containing developing males. Hence, by distracting their competitors away from receptive females late males may increase their own chance to mate with these females. After emergence, males decompose the active compounds within 32 h probably to decrease molestation during their own search for mates. Chemical analyses of active pheromone extracts and bioassays using fractions demonstrate that the active compounds are among the cuticular hydrocarbons.     

Sequestration of plant pyrrolizidine alkaloids by chrysomelid beetles and selective transfer into the defensive secretions

Summary Oreina cacaliae andO. speciosissima (Coleoptera, Chrysomelidae) sequester in their elytral and pronotal defensive secretions pyrrolizidine alkaloids (PAs) as Noxides (PA N-oxides). The PA N-oxide patterns found in the beetles and their host plants were evaluated qualitatively and quantitatively by capillary gas chromatography/mass spectrometry (GC-MS). Of the three host plantsAdenostyles alliariae (Asteraceae) is the exclusive source for PA N-oxide sequestration in the defensive secretions of the beetles. With the exception of O-acetylseneciphylline the N-oxides of all PAs ofA. alliariae, i.e. senecionine, seneciphylline, spartioidine, integerrimine, platyphylline and neoplatyphylline were identified in the secretion. PA N-oxides typical ofSenecio fuchsii (Asteraceae) were detected in the bodies of the beetles but not in their secretion. No PAs were found in the leaves of the third host plant,Petasites paradoxus (Asteraceae). The results suggest the existence of two distinctive storage compartments for PA N-oxides in the beetle: (1) the defensive secretion, containing specifically PA N-oxides acquired fromA. alliariae; (2) the body of the beetle, sequestering additionally but less selectively PA N-oxides from other sources,e.g. S. fuchsii or monocrotaline N-oxide fed in the laboratory. The concentration of PA N-oxides in the defensive secretion is in the range of 0.1 to 0.3 mol/1, which is more than 2.5 orders of magnitude higher than that found in the body of the beetle. No significant differences exist in the ability of the two species of beetles to sequester PA N-oxides fromA. alliariae, althoughO. speciosissima, but notO. cacaliae, produces autogenous cardenolides. A negative correlation seems to exist between the concentrations of plant-derived PA N-oxides andde novo synthesized cardenolides in the defensive secretion ofO. speciosissima.     

Storage and metabolism of radioactively labeled pyrrolizidine alkaloids by butterflies and larvae of Mechanitis polymnia (Lepidoptera: Nymphalidae,Ithomiinae)

Summary. Tracer feeding studies with radioactively labeled pyrrolizidine alkaloids (PAs) were performed to attain experimental information about the specificity and mechanisms of uptake, metabolism and storage of PAs in the alkaloid sequestering ithomiine butterfly Mechanitis polymnia. Adult butterflies easily ingest the tracers offered dissolved in a saturated sugar solution. Feeding of [¹⁴C]rinderine (free base) confirmed that M. polymnia is well adapted to sequester and maintain PAs of the lycopsamine type. Approximately 80% of the ingested radioactivity can be recovered in methanol extracts of the butterflies over a period of at least 6 hours. Labeled rinderine is efficiently N-oxidized and transformed into a metabolite of still unknown structure. These two metabolites are formed in almost equal amounts and account for more than 90% of total radioactivity. After four hours the toxic free base is only detectable in traces. Radioactively labeled senecioylretronecine (free base), a PA that often accompanies PAs of the lycopsamine type in plants, is metabolized in a different manner. The toxic free base disappeared as fast as the tertiary rinderine, but the final products which accumulated in a stable ratio after 12 hours were mainly two polar metabolites of unknown structure; senecioylretronecine N-oxide accounts for less than 10% of total PAs. Labeled senecionine a macrocyclic PA, which never has been found in wild caught M. polymnia is only slowly N-oxidized. In females ca 50% of the ingested senecionine is still present as free base after 24 hours, whereas under the same conditions in males this percentage is only ca 20%. This difference in N-oxidation was the only significant sex-specific difference observed in various experiments. Larvae of M. polymnia which feed on Solanum tabacifolium, a plant that does not contain PAs, are able to sequester and partly N-oxidize labeled senecioylretronecine and senecionine. However, the storage is not very efficient; with the two tracers less than 5% of radioactivity remained in the bodies after 24 hours. Received 19 October 1999; accepted 24 November 1999     

Responses of three mouse species to deterrent chemicals in the monarch butterfly. I. Taste and toxicity tests using artificial diets laced with digitoxin or monocrotaline

Summary Of three common mouse species at the Mexican overwintering sites of the monarch butterfly, onlyPeromyscus melanotis eats monarchs. We hypothesized thatP. aztecus andReithrodontomys sumichrasti reject monarchs because they are more sensitive to the bitter taste and/or toxic effects of the cardiac glycosides (CGs) and pyrrolizidine alkaloids (PAs) in the butterflies. Two-choice preference tests revealed no difference in taste avoidance thresholds to free base and N-oxide forms of the PA, monocrotaline, but very different avoidance thresholds to the CG, digitoxin. Avoidance thresholds forR. sumichrasti andP. aztecus were, in respective order, 1020 and 34 times less than that forP. melanotis. We also tested the toxic sensitivity of juvenile mice by chronically feeding diets containing digitoxin or monocrotaline at concentrations similar to those used in the preference tests. No species developed CG toxicity, but bothP. melanotis andP. aztecus developed moderate PA toxicity (R. sumichrasti was not tested for PA toxicity).P. aztecus grew more slowly and manyP. melanotis had hepatic metabolic lesions. Thus, the three mouse species responded very differently to the taste and toxic properties of CGs and PAs at ecologically relevant concentrations: 1) CGs were taste rejected by all species exceptP. melanotis, while PAs were not; and 2) PAs were toxic, while CGs were not.     

Indirect evidence indicates female semiochemicals release male precopulatory behaviour in the snow crab,Chionoecetes opilio (Brachyura: Majidae)

Summary Experiments were conducted in aquaria to test the hypothesis that females of the snow crab,Chionoecetes opilio, release a sex pheromone to attract mates. Males exhibited significantly increased activity to water from a source aquarium containing recently-moulted pubescent females, egg-stripped multiparous females and recently-moulted immature females than to water from an aquarium containing berried multiparous females, eggs alone, adolescent males or an empty aquarium. Males without their cephalic chemoreceptors maintained low activity levels in the presence of recently-moulted pubescent females, whereas maxilla-ablated males reacted as strongly as intact males. These findings are discussed within the context of the ecdysteroid-sex pheromone hypothesis, proposed by Kittredgeet al. (1971) and Kittredge & Takahashi (1972), but subsequently rejected. We propose that ecdysteroids from both pubescent and multiparous females may elicit male search-and-clasp behaviour inC. opilio and that the reproductive biology of species used to refute the hypothesis was inappropriate to test the role of ecdysteroids as a cue in the mating process.     

Apportionment of nuptial alkaloidal gifts by a multiply-mated female moth (Utetheisa ornatrix): eggs individually receive alkaloid from more than one male source

Summary. In the moth Utetheisa ornatrix the female is promiscuous and receives a nuptial gift of pyrrolizidine alkaloid (PA) by seminal infusion from each mate. The alkaloidal gifts are transmitted by the female to the eggs, which are protected as a result. We here show that individual eggs may receive PAs from more than one male source and that individual males have no assurance that the PA they themselves contribute to the female will find its way exclusively to eggs of their siring. Received 28 June 2002; accepted 28 August 2002.     

Green leaf volatiles enhance sex attractant pheromone of the tobacco budworm,Heliothis virescens (Lep.: Noctuidae)

Summary Components of the green leaf volatile complex (Z-3-hexenyl acetate andE-2-hexenyl acetate) were shown to enhance responses of tobacco budworm,Heliothis virescens, males to the sex attractant pheromone of conspecific females in the field. The results are discussed with regard to green leaf volatiles which enhance the attractant pheromone of a cohabiting species, and serve as attractants of a parasitoid of conspecific larvae.     

Time course of pyrrolizidine alkaloid sequestration in <Emphasis Type="Italic">Longitarsus</Emphasis> flea beetles (Coleoptera,Chrysomelidae)

Summary. Several species of Longitarsus take up, metabolize and store pyrrolizidine alkaloids (PAs) from their host plants. In feeding experiments using radioactively labeled PAs of different types we examined the time course of the sequestration process in L. jacobaeae and L. aeruginosus. We found that adapted species efficiently store PAs for at least two weeks without major losses. During that time, there is virtually no change in the ratio of tertiary alkaloids to stored non-toxic N-oxides, regardless of chemical form fed to the beetles. This implies a transient N-oxidation process where the alkaloids are only temporarily accessible to the enzyme. A dissection experiment with L. aeruginosus six days after uptake of labeled PAs showed that the tertiary alkaloids are not found in the hemolymph but are stored in the elytra and other body compartments. This conforms with earlier experiments that localized the enzymes site of action in the hemolymph. Furthermore we show that different total alkaloid doses in the diet of L. jacobaeae and the potentially less adapted L. succineus do not affect the ratio of recovered N-oxides to tertiary molecules. Thus, the efficiency of the N-oxidizing enzyme is not dependent on the concentration of alkaloids offered.     

Chemical defense of the ladybird beetle Epilachna paenulata

Summary. The defensive chemistry of the ladybird beetle Epilachna paenulata (Coleoptera: Coccinellidae) was characterized as a mixture of piperidine, homotropane and pyrrolidine alkaloids. Whole body extracts of adult beetles contain four major alkaloids: 1-(6-Methyl-2,3,4,5-tetrahydro-pyridin-2-yl)-propan-2-one; 1-(6-methyl-2-piperidyl)-propan-2-one; 9-aza-1-methyl-bicyclo[3.3.1]nonan-3-one and 1- (2′′- hydroxyethyl)-2-(12′-aminotridecyl)-pyrrolidine. Comparative studies of the defensive chemistry of eggs, larvae, pupae and adults showed both qualitative and quantitative differences in alkaloid composition among the four life stages, and also within adult age. Laboratory predation bioassays with wolf spiders showed that the adults are better protected than the larvae and pupae. Field tests showed the adult alkaloid extract to be deterrent to ants.     

Allelochemical effects of ergoline alkaloids fromIpomoea parasitica onHeliothis virescens

Lysergol, elymoclavine and three other van Urk-positive alkaloids were identified by 2D chromatography in the seed extract ofIpomoea parasitica. The presence of the same ergoline alkaloids was also demonstrated in vegetative tissue ofI. parasitica. Heliothis virescens larvae reared on a diet containing an alkaloid extract ofI. parasitica showed an increase in the consumption index and a reduction in the effiency of conversion of food. No difference was observed in the approximated digestibility. The percentages of pupation and emergence were reduced and the malefemale ratio was altered in insects fed on a diet containing the alkaloid extract ofI. parasitica or lysergol. The increases in the concentration of ergoline alkaloids inI. parasitica at the seedling stage and at flowering support the theory that these compounds play a defensive role against herbivory in the plant.     

Quantitative analysis of pyrrolizidine alkaloids sequestered from diverse host plants in Longitarsus flea beetles (Coleoptera, Chrysomelidae)

Summary. Several species of the flea beetles genus Longitarsus are able to sequester pyrrolizidine alkaloids (PAs) from their host plants. In five Longitarsus species we compare the concentration of PAs present in their host plants belonging to the Asteraceae or Boraginaceae with those found in the beetles. To get an estimate of the intrapopulation variability, three samples of five beetles each and five individual plants were analyzed for each comparison. A strong intrapopulation variability could be detected both among plant and beetle samples. The total concentration found in the beetles varied strongly between species. The local host plant and its phenology influence the concentrations present in the beetles as evidenced in comparisons of a single beetle species from two different hosts and of one beetle species collected at the same site at different times of the year. In addition, different beetle species apparently vary in their capacity to sequester the alkaloids, at the lowest extreme the mean PA concentration in the beetles (0.034 μg PA/mg dry weight) was 1/30 of the mean concentration found in the plant leaves (L. aeruginosus from Eupatorium cannabinum), at the highest extreme (2.098 μg PA/mg dw) the concentration in the beetles was a 1000 fold higher than in the plant leaves (L. nasturtii from Symphytum officinale). The highest mean concentration found in the beetles was 3.446 μg/mg dw (L. exoletus from Cynoglossum officinale). The absolute concentrations found in the beetles are comparable to other insects which have been shown to be effectively defended against their potential predators. Received 22 June 1999; accepted 25 August 1999