Role of arthropod communities in bioenergy crop litter decomposition†.

The extensive land use conversion expected to occur to meet demands for bioenergy feedstock production will likely have widespread impacts on agroecosystem biodiversity and ecosystem services, including carbon sequestration. Although arthropod detritivores are known to contribute to litter decomposition and thus energy flow and nutrient cycling in many plant communities, their importance in bioenergy feedstock communities has not yet been assessed. We undertook an experimental study quantifying rates of litter mass loss and nutrient cycling in the presence and absence of these organisms in three bioenergy feedstock crops-miscanthus (Miscanthus x giganteus), switchgrass (Panicum virgatum), and a planted prairie community. Overall arthropod abundance and litter decomposition rates were similar in all three communities. Despite effective reduction of arthropods in experimental plots via insecticide application, litter decomposition rates, inorganic nitrogen leaching, and carbon-nitrogen ratios did not differ significantly between control (with arthropods) and treatment (without arthropods) plots in any of the three community types. Our findings suggest that changes in arthropod faunal composition associated with widespread adoption of bioenergy feedstock crops may not be associated with profoundly altered arthropod-mediated litter decomposition and nutrient release.

Aliphatic esters as targets of esterase activity in the parsnip webworm (Depressaria pastinacella).

As a specialist on the reproductive structures of Pastinaca sativa and species in the related genus Heracleum, the parsnip webworm (Depressaria pastinacella) routinely encounters a distinctive suite of phytochemicals in hostplant tissues. Little is known, however, about the detoxification mechanisms upon which this species relies to metabolize these compounds. In this study, larval guts containing hostplant tissues were homogenized, and metabolism was determined by incubating reactions with and without NADPH and analyzing for substrate disappearance and product appearance by gas chromatography-mass spectrometry. Using this approach, we found indications of carboxylesterase activity, in the form of appropriate alcohol metabolites for three aliphatic esters in hostplant tissues-octyl acetate, octyl butyrate, and hexyl butyrate. Involvement of webworm esterases in hostplant detoxification subsequently was confirmed with metabolism assays with pure compounds. This study is the first to implicate esterases in lepidopteran larval midgut metabolism of aliphatic esters, ubiquitous constituents of flowers and fruits. In addition, this method confirmed that webworms detoxify furanocoumarins and myristicin in their hostplants via cytochrome P450-mediated metabolism, and demonstrated that these enzymes also metabolize the coumarin osthol and the fatty acid derivative palmitolactone.

Bioenergy crops Miscanthus x giganteus and Panicum virgatum reduce growth and survivorship of Spodoptera frugiperda (Lepidoptera: Noctuidae).

Large-scale cultivation of plants used as biofuels is likely to alter the ecological interactions of current agricultural crops and their insect pests in a myriad of ways. Recent evidence suggests many contemporary maize pests will be able to use potential biofuel crops such as switchgrass, Panicum virgatum L., and miscanthus as hosts. To determine how suitable these biofuels are to the maize, Zea mays L., pest and generalist graminivore, Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae), we examined host plant preference and larval performance on foliage grown for commercial biofuel production. Larvae fed leaf tissue from both field- and greenhouse-grown switchgrass and miscanthus were monitored for survival, development, and food use relative to field-grown maize. Survivorship on biofuel crops was high on greenhouse-grown leaf tissue but severely reduced for field-grown switchgrass, and no larvae survived on field-grown miscanthus. Larvae fed field-grown tissue had larger head capsules yet achieved lower pupal weights because the increased toughness of the leaf tissue prevented the assimilation of nitrogen. Given that larvae overwhelmingly preferred maize to other biofuel crop species and that survival and performance were dramatically reduced on biofuel crop species, it is likely that biofuel crops, as grown for field cultivation, will suffer reduced damage from maize pests such as S. frugiperda because of reduced suitability.

A substrate-specific cytochrome P450 monooxygenase, CYP6AB11, from the polyphagous navel orangeworm (Amyelois transitella).

The navel orangeworm Amyelois transitella (Walker) (Lepidoptera: Pyralidae) is a serious pest of many tree crops in California orchards, including almonds, pistachios, walnuts and figs. To understand the molecular mechanisms underlying detoxification of phytochemicals, insecticides and mycotoxins by this species, full-length CYP6AB11 cDNA was isolated from larval midguts using RACE PCR. Phylogenetic analysis of this insect cytochrome P450 monooxygenase established its evolutionary relationship to a P450 that selectively metabolizes imperatorin (a linear furanocoumarin) and myristicin (a natural methylenedioxyphenyl compound) in another lepidopteran species. Metabolic assays conducted with baculovirus-expressed P450 protein, P450 reductase and cytochrome b(5) on 16 compounds, including phytochemicals, mycotoxins, and synthetic pesticides, indicated that CYP6AB11 efficiently metabolizes imperatorin (0.88 pmol/min/pmol P450) and slowly metabolizes piperonyl butoxide (0.11 pmol/min/pmol P450). LC-MS analysis indicated that the imperatorin metabolite is an epoxide generated by oxidation of the double bond in its extended isoprenyl side chain. Predictive structures for CYP6AB11 suggested that its catalytic site contains a doughnut-like constriction over the heme that excludes aromatic rings on substrates and allows only their extended side chains to access the catalytic site. CYP6AB11 can also metabolize the principal insecticide synergist piperonyl butoxide (PBO), a synthetic methylenedioxyphenyl compound, albeit slowly, which raises the possibility that resistance may evolve in this species after exposure to synergists under field conditions.

Impact of elevated levels of atmospheric CO2 and herbivory on flavonoids of soybean (Glycine max Linnaeus).

Atmospheric levels of carbon dioxide (CO2) have been increasing steadily over the last century. Plants grown under elevated CO2 conditions experience physiological changes, particularly in phytochemical content, that can influence their suitability as food for insects. Flavonoids are important plant defense compounds and antioxidants that can have a large effect on leaf palatability and herbivore longevity. In this study, flavonoid content was examined in foliage of soybean (Glycine max Linnaeus) grown under ambient and elevated levels of CO2 and subjected to damage by herbivores in three feeding guilds: leaf skeletonizer (Popillia japonica Newman), leaf chewer (Vanessa cardui Linnaeus), and phloem feeder (Aphis glycines Matsumura). Flavonoid content also was examined in foliage of soybean grown under ambient and elevated levels of O3 and subjected to damage by the leaf skeletonizer P. japonica. The presence of the isoflavones genistein and daidzein and the flavonols quercetin and kaempferol was confirmed in all plants examined, as were their glycosides. All compounds significantly increased in concentration as the growing season progressed. Concentrations of quercetin glycosides were higher in plants grown under elevated levels of CO2. The majority of compounds in foliage were induced in response to leaf skeletonization damage but remained unchanged in response to non-skeletonizing feeding or phloem-feeding. Most compounds increased in concentration in plants grown under elevated levels of O3. Insects feeding on G. max foliage growing under elevated levels of CO2 may derive additional antioxidant benefits from their host plants as a consequence of the change in ratios of flavonoid classes. This nutritional benefit could lead to increased herbivore longevity and increased damage to soybean (and perhaps other crop plants) in the future.

Olfactory preferences of Popillia japonica, Vanessa cardui, and Aphis glycines for Glycine max grown under elevated CO2.

Levels of atmospheric CO(2) have been increasing steadily over the last century and are projected to increase even more dramatically in the future. Soybeans (Glycine max L.) grown under elevated levels of CO(2) have larger herbivore populations than soybeans grown under ambient levels of CO(2). Increased abundance could reflect the fact that these herbivores are drawn in by increased amounts of volatiles or changes in the composition of volatiles released by plants grown under elevated CO(2) conditions. To determine impacts of elevated CO(2) on olfactory preferences, Japanese beetles (Popillia japonica Newman) and soybean aphids (Aphis glycines Matsumura) were placed in Y-tube olfactometers with a choice between ambient levels of CO(2) gas versus elevated levels of CO(2) gas or damaged and undamaged leaves and plants grown under ambient levels of CO(2) versus damaged and undamaged plants grown under elevated levels of CO(2). All plants had been grown from seeds under ambient or elevated levels of CO(2). Painted lady butterflies (Vanessa cardui L.) were placed in an oviposition chamber with a choice between plants grown under ambient and elevated levels of CO(2). A. glycines and V. cardui showed no significant preference for plants in either treatment. P. japonica showed no significant preference between ambient levels and elevated levels of CO(2) gas. There was a significant P. japonica preference for damaged plants grown under ambient CO(2) versus undamaged plants but no preference for damaged plants grown under elevated CO(2) versus undamaged plants. P. japonica also preferred damaged plants grown under elevated levels of CO(2) versus damaged plants grown under ambient levels of CO(2). This lack of preference for damaged plants grown under elevated CO(2) versus undamaged plants could be the result of the identical elevated levels of a green leaf volatile (2-hexenal) present in all foliage grown under elevated CO(2) regardless of damage status. Green leaf volatiles are typically released from damaged leaves and are used as kairomones by many herbivorous insects for host plant location. An increase in production of volatiles in soybeans grown under elevated CO(2) conditions may lead to larger herbivore outbreaks in the future.

Quercetin-metabolizing CYP6AS enzymes of the pollinator Apis mellifera (Hymenoptera: Apidae).

Although the honey bee (Apis mellifera) genome contains far fewer cytochrome P450 genes associated with xenobiotic metabolism than other insect genomes sequenced to date, the CYP6AS subfamily, apparently unique to hymenopterans, has undergone an expansion relative to the genome of the jewel wasp (Nasonia vitripennis). The relative dominance of this family in the honey bee genome is suggestive of a role in processing phytochemicals encountered by honey bees in their relatively unusual diet of honey (comprising concentrated processed nectar of many plant species) and bee bread (a mixture of honey and pollen from many plant species). In this study, quercetin was initially suggested as a shared substrate for CYP6AS1, CYP6AS3, and CYP6AS4, by its presence in honey, extracts of which induce transcription of these three genes, and by in silico substrate predictions based on a molecular model of CYP6AS3. Biochemical assays with heterologously expressed CYP6AS1, CYP6AS3, CYP6AS4 and CYP6AS10 enzymes subsequently confirmed their activity toward this substrate. CYP6AS1, CYP6AS3, CYP6AS4 and CYP6AS10 metabolize quercetin at rates of 0.5+/-0.1, 0.5+/-0.1, 0.2+/-0.1, and 0.2+/-0.1 pmol quercetin/ pmol P450/min, respectively. Substrate dockings and sequence alignments revealed that the positively charged amino acids His107 and Lys217 and the carbonyl group of the backbone between Leu302 and Ala303 are essential for quercetin orientation in the CYP6AS3 catalytic site and its efficient metabolism. Multiple replacements in the catalytic site of CYP6AS4 and CYP6AS10 and repositioning of the quercetin molecule likely account for the lower metabolic activities of CYP6AS4 and CYP6AS10 compared to CYP6AS1 and CYP6AS3.

Tritrophic effects of xanthotoxin on the polyembryonic parasitoid Copidosoma sosares (Hymenoptera: Encyrtidae).

Plant chemistry can have deleterious effects on insect parasitoids, which include the reduction in body size, increased development time, and increased mortality. We examined the effects of xanthotoxin, a linear furanocoumarin, on the polyembryonic encyrtid wasp Copidosoma sosares, a specialist parasitoid that attacks the parsnip webworm, Depressaria pastinacella, itself a specialist on furanocoumarin-producing plants. Furanocoumarins, allelochemicals abundant in the Apiaceae and Rutaceae, are toxic to a wide range of herbivores. In this study, we reared parasitized webworms on artificial diets containing no xanthotoxin (control) or low or high concentrations of xanthotoxin. Clutch sizes of both male and female C. sosares broods were more than 20% smaller when they developed in hosts fed the diet containing high concentrations of xanthotoxin. Xanthotoxin concentration in the artificial diet had no effect on the development time of C. sosares, nor did it have an effect on the body size (length of hind tibia) of individual adult male and female C. sosares in single-sex broods. Webworms fed artificial diets containing low or high concentrations of xanthotoxin were not significantly smaller, and their development time was similar to that of webworms fed a xanthotoxin-free diet. Mortality of webworms was not affected by xanthotoxin in their artificial diet. Therefore, dietary xanthotoxin did not appear to affect C. sosares via impairment of host health. However, unmetabolized xanthotoxin was found in D. pastinacella hemolymph where C. sosares embryos develop. Hemolymph concentrations were fourfold greater in webworms fed the high-xanthotoxin-containing diet than in webworms fed the low-xanthotoxin-containing diet. We failed to detect any xanthotoxin metabolism by either C. sosares embryos or precocious larvae. Therefore, the observed tritrophic effects of xanthotoxin are likely to be due to the effects of xanthotoxin after direct contact in the hemolymph rather than to the effects of compromised host quality.

Metabolism of myristicin by Depressaria pastinacella CYP6AB3v2 and inhibition by its metabolite.

Although methylenedioxyphenyl (MDP) compounds, such as myristicin, are useful in the management of insecticide-resistant insects, the molecular mechanisms for their action in mammals and insects have not been elucidated. In this study, GC-MS analyses of methanol extracts of foliage of wild parsnip (Pastinaca sativa) have identified myristicin as a substrate for CYP6AB3v2, an imperatorin-metabolizing cytochrome P450 monooxygenase from Depressaria pastinacella (parsnip webworm). In contrast with its strong inhibitory effects on many mammalian P450s, myristicin is effectively metabolized by CYP6AB3v2 (V(max) and K(m) of 97.9 pmol/min/pmol P450 and 17.9 microM, respectively) at a rate exceeding that recorded previously for imperatorin, the only other known substrate for this highly specialized enzyme. The myristicin metabolite of CYP6AB3v2 is 1-(3',4'-methylenedioxy-5'-methoxyphenyl)-2,3-epoxypropane. Molecular dockings have indicated that, unlike other epoxide metabolites of furanocoumarins, this epoxide metabolite is likely to remain in the CYP6AB3v2 catalytic site due to its low binding energy (-31.0 kcal/mol). Inhibition assays indicate that myristicin acts as a mixed inhibitor of this insect P450 and suggest that the epoxide metabolite may be an intermediate involved in the formation of P450-methylenedioxyphenyl complexes.

Longevity and fecundity of Japanese beetle (Popillia japonica) on foliage grown under elevated carbon dioxide.

Atmospheric levels of carbon dioxide (CO(2)) have been increasing steadily over the last century. Plants grown under elevated CO(2) experience physiological changes that influence their suitability as food. Previous studies have found increased insect herbivory on plants grown under elevated CO(2). To determine effects of consuming foliage of soybean (Glycine max) grown under elevated CO(2) on adult survivorship and fecundity, Japanese beetles (Popillia japonica Newman) were fed for the duration of their adult lives leaves grown under elevated CO(2) (550 mumol/mol), under ambient atmosphere (370 mumol/mol), or grown under ambient atmosphere but supplemented with a solution of sugars. To determine effects of a diet of foliage grown under elevated ozone (O(3)), another anthropogenic gaseous pollutant, beetles in the laboratory were fed soybean leaves grown under elevated CO(2), elevated O(3), or a combination of both elevated gases. Leaf tissue was also analyzed for longevity-enhancing antioxidants, because increases in dietary antioxidants can increase lifespan. Lifespan of Japanese beetles was prolonged by 8-25% when fed foliage developed under elevated CO(2), but consuming foliage that had taken up sugars to approximately the same level as foliage grown under elevated CO(2) had no effect on fecundity or longevity. Females consuming elevated CO(2) foliage laid approximately twice as many eggs as females fed foliage grown under ambient conditions. Consuming foliage grown under elevated O(3) had no effect on fecundity. No significant differences in total antioxidant content of foliage from ambient and elevated CO(2) conditions were detected. Although the precise mechanism is unclear, by altering components of leaf chemistry other than sugar content, elevated CO(2) may increase populations of Japanese beetles and their impact on crop productivity.

Foliage of oaks grown under elevated CO2 reduces performance of Antheraea polyphemus (Lepidoptera: Saturniidae).

To understand how the increase in atmospheric CO2 from human activity may affect leaf damage by forest insects, we examined host plant preference and larval performance of a generalist herbivore, Antheraea polyphemus Cram., that consumed foliage developed under ambient or elevated CO2. Larvae were fed leaves from Quercus alba L. and Quercus velutina Lam. grown under ambient or plus 200 microl/liter CO2 using free air carbon dioxide enrichment (FACE). Lower digestibility of foliage, greater protein precipitation capacity in frass, and lower nitrogen concentration of larvae indicate that growth under elevated CO2 reduced the food quality of oak leaves for caterpillars. Consuming leaves of either oak species grown under elevated CO2 slowed the rate of development of A. polyphemus larvae. When given a choice, A. polyphemus larvae preferred Q. velutina leaves grown under ambient CO2; feeding on foliage of this species grown under elevated CO2 led to reduced consumption, slower growth, and greater mortality. Larvae compensated for the lower digestibility of Q. alba leaves grown under elevated CO2 by increasing the efficiency of conversion of ingested food into larval mass. Despite equivalent consumption rates, larvae grew larger when they consumed Q. alba leaves grown under elevated compared with ambient CO2. Reduced consumption, slower growth rates, and increased mortality of insect larvae may explain lower total leaf damage observed previously in plots in this forest exposed to elevated CO2. By subtly altering aspects of leaf chemistry, the ever-increasing concentration of CO2 in the atmosphere will change the trophic dynamics in forest ecosystems.

Allelic variation in the Depressaria pastinacella CYP6AB3 protein enhances metabolism of plant allelochemicals by altering a proximal surface residue and potential interactions with cytochrome P450 reductase.

CYP6AB3v1, a cytochrome P450 monooxygenase in Depressaria pastinacella (parsnip webworm), is highly specialized for metabolizing imperatorin, a toxic furanocoumarin in the apiaceous host plants of this insect. Cloning and heterologous expression of CYP6AB3v2, an allelic variant identified in D. pastinacella, reveals that it metabolizes imperatorin at a rate (V(max) of 10.02 pmol/min/pmol of cytochrome P450 monooxygenase (P450)) significantly higher than CYP6AB3v1 (V(max) of 2.41 pmol/min/pmol) when supplemented with even low levels of cytochrome P450 reductase. Comparisons of the NADPH consumption rates for these variants indicate that CYP6AB3v2 utilizes this electron source at a faster rate than does CYP6AB3v1. Molecular modeling of the five amino acid differences between these variants and their potential interactions with P450 reductase suggests that replacement of Val(92) on the proximal face of CYP6AB3v1 with Ala(92) in CYP6AB3v2 affects interactions with P450 reductase so as to enhance its catalytic activity. Allelic variation at this locus potentially allows D. pastinacella to adapt to both intraspecific and interspecific variation in imperatorin concentrations in its host plants.

Comparison of photosynthetic damage from arthropod herbivory and pathogen infection in understory hardwood saplings.

Arthropods and pathogens damage leaves in natural ecosystems and may reduce photosynthesis at some distance away from directly injured tissue. We quantified the indirect effects of naturally occurring biotic damage on leaf-level photosystem II operating efficiency (Phi(PSII)) of 11 understory hardwood tree species using chlorophyll fluorescence and thermal imaging. Maps of fluorescence parameters and leaf temperature were stacked for each leaf and analyzed using a multivariate method adapted from the field of quantitative remote sensing. Two tree species, Quercus velutina and Cercis canadensis, grew in plots exposed to ambient and elevated atmospheric CO(2) and were infected with Phyllosticta fungus, providing a limited opportunity to examine the potential interaction of this element of global change and biotic damage on photosynthesis. Areas surrounding damage had depressed Phi(PSII )and increased down-regulation of PSII, and there was no evidence of compensation in the remaining tissue. The depression of Phi(PSII) caused by fungal infections and galls extended >2.5 times further from the visible damage and was approximately 40% more depressed than chewing damage. Areas of depressed Phi(PSII) around fungal infections on oaks growing in elevated CO(2) were more than 5 times larger than those grown in ambient conditions, suggesting that this element of global change may influence the indirect effects of biotic damage on photosynthesis. For a single Q. velutina sapling, the area of reduced Phi(PSII) was equal to the total area directly damaged by insects and fungi. Thus, estimates based only on the direct effect of biotic agents may greatly underestimate their actual impact on photosynthesis.

Cytochrome P450-mediated metabolism of xanthotoxin by Papilio multicaudatus.

Within the genus Papilio, the P. glaucus group contains the most polyphagous Papilio species within the Papilionidae. The majority of Papilio species are associated with hostplants in the families Rutaceae and Apiaceae, and characterizing most are secondary metabolites called furanocoumarins. Recent phylogenetic studies suggest that furanocoumarin metabolism is an ancestral trait, with the glaucus group derived from ancestors associated with furanocoumarin-containing Rutaceae. In this study, we examined this relationship by conducting a gravimetric analysis of growth that used various concentrations of the furanocoumarin xanthotoxin. Papilio multicaudatus, the putative ancestor of the glaucus group, includes at least one furanocoumarin-containing rutaceous species among its hostplants; this species can consume leaf tissue containing up to 0.3% xanthotoxin with no detectable effect on relative growth rate, relative consumption rate, or efficiency of conversion of ingested food. As is the case for other Papilio species, xanthotoxin metabolism is mediated by cytochrome P450 monooxygenases (P450s). Ingestion of xanthotoxin by ultimate instar P. multicaudatus increases activity up to 30-fold in a dose-dependent fashion. Midguts of induced larvae can also effectively metabolize six other furanocoumarins, including both linear (bergapten, isopimpinellin, imperatorin) and angular (angelicin, sphondin) forms. A metabolite of xanthotoxin in the frass from xanthotoxin-treated larvae, identified as 6-(7-hydroxy-8-methoxycoumaryl)-acetic acid by MS-MS and NMR analyses, is identical to one from the frass of P. polyxenes. The occurrence of this metabolite in two swallowtails and the presence of a second metabolite of xanthotoxin, 6-(7-hydroxy-8-methoxycoumaryl)-hydroxyethanol in the frass of both P. polyxenes and Depressaria pastinacella are consistent with the suggestion that lepidopterans share as the first step of xanthotoxin metabolism the P450-mediated epoxidation of the furan ring 2'-3' double bond.

The differential effects of herbivory by first and fourth instars of Trichoplusia ni (Lepidoptera: Noctuidae) on photosynthesis in Arabidopsis thaliana.

The effect of different feeding behaviours of 1st and 4th instar Trichoplusia ni on photosynthesis of Arabidopsis thaliana var. Columbia was characterized using spatially resolved measurements of fluorescence and leaf temperature, as well as leaf gas exchange,. First instars made small holes with a large perimeter-to-area ratio and avoided veins, while 4th instars made large holes with a low perimeter-to-area ratio and consumed veins. Herbivory by 1st instars reduced photosynthesis more strongly in the remaining leaf tissue than that by 4th instars. Photosystem II operating efficiency (PhiPSII) was correlated with the rate of CO2 exchange, and reductions in PhiPSII in areas around the missing tissues contributed to a 15.6% reduction in CO2 assimilation on the first day following removal of 1st instars. The corresponding increases in non-photochemical quenching and greater rates of non-stomatal water loss from these regions, as well as the partial reversal of low PhiPSII by increasing the ambient CO2 concentration, suggests that localized water stress and reduced stomatal conductance contributed to the inhibition of photosynthesis. Damage by 1st but not 4th instars reduced the maximum quantum efficiency of photosystem II photochemistry (Fv/Fm) by 4-8%. While herbivory by both 1st and 4th instars increased dark respiration rates, the rates were too low to have contributed to the observed reductions in CO2 exchange. The small holes produced by 1st instars may have isolated patches of tissue from the vascular system thereby contributing to localized water stress. Since neither 1st nor 4th instar herbivory had a detectable effect on the expression of the Rubisco small subunit gene, the observed differences cannot be attributed to changes in expression of this gene. The mode of feeding by different instars of T. ni determined the photosynthetic response to herbivory, which appeared to be mediated by the level of water stress associated with herbivore damage.

Parsnip webworms and host plants at home and abroad: trophic complexity in a geographic mosaic.

Due to differences in the structure of communities in which interactions are embedded, the intensity of interactions between species may vary with location; thus, what results from differences in outcomes and in degree of specialization is a geographic mosaic, which provides the raw material for divergent coevolutionary trajectories. Where selection intensity is great, reciprocal responses are likely in so-called "hotspots"; in contrast, where selection pressures are relaxed, reciprocal responses in "coldspots" are far less likely to occur. There are few if any studies examining how a gradient of increasing trophic complexity might influence the probability of phenotype matching and, correspondingly, the "temperature" of the coevolutionary interaction. This study was conducted to compare outcomes of the interaction between wild parsnip (Pastinaca sativa) and parsnip webworm (Depressaria pastinacella) in its indigenous area, Europe, to its area of introduction, the midwestern United States. Specifically, we tested the hypothesis that increasing trophic complexity, represented by alternate host plants or the presence of natural enemies, reduces the selective impact of parsnip webworms and hence diminishes linkage between host plant chemistry and webworms that would be expected in coevolutionary hotspots. This comparison of a two-species interaction in its area of introduction and its area of indigeneity revealed common patterns that are more reflective of interaction temperature than of continental origin. Where webworms are rare, parsnips produce lower levels of xanthotoxin and bergapten in both the midwestern United States and Netherlands populations. However, the most striking result from this intercontinental comparison is that what is a ubiquitous two-species interaction in North America is in fact exceptional in Europe; webworms could more reliably be found infesting H. sphondylium even where P. sativa was available as well. This preference for H. sphondylium exists despite the comparatively high probability of parasitism associated with this host plant and may reflect the overall lower furanocoumarin content of H. sphondylium. The interaction of parsnip webworms and wild parsnips at home and away demonstrates clearly the potential for rapid contemporary evolution of chemical traits upon re-association with a coevolved enemy, a potential evolutionary outcome that should be considered in the design and implementation of future weed biological control programs.

Increase in toxicity of an invasive weed after reassociation with its coevolved herbivore.

The ability of weeds to proliferate into nonindigenous habitats has been attributed to escape from their native natural enemies, allowing reallocation of resources from chemical defense into growth and reproduction. Many invasive weeds, however, eventually encounter their native, coevolved enemies in areas of introduction. Examination of herbarium specimens of an invasive phototoxic European weed, Pastinaca sativa, through 152 years reveals phytochemical shifts coincident in time with the accidental introduction of a major herbivore, the parsnip webworm, Depressaria pastinacella. Plants collected before the introduction of webworms in North America and during the earliest stages of establishment (1850-1889) are lower in toxic furanocoumarins than all plants subsequently collected in North America and lower than European plant samples collected before 1889. Thus, introduction of a major specialist herbivore can increase noxiousness of a species in its area of introduction, illuminating a potential consequence of classical biocontrol programs involving insect herbivores and poisonous weeds.

Elevated CO2 reduces leaf damage by insect herbivores in a forest community.

By altering foliage quality, exposure to elevated levels of atmospheric CO(2) potentially affects the amount of herbivore damage experienced by plants. Here, we quantified foliar carbon (C) and nitrogen (N) content, C : N ratio, phenolic levels, specific leaf area (SLA) and the amount of leaf tissue damaged by chewing insects for 12 hardwood tree species grown in plots exposed to elevated CO(2) (ambient plus 200 microl l(-1)) using free-air CO(2) enrichment (FACE) over 3 yr. The effects of elevated CO(2) varied considerably by year and across species. Elevated CO(2) decreased herbivore damage across 12 species in 1 yr but had no detectable effect in others. Decreased damage may have been related to lower average foliar N concentration and SLA and increased C : N ratio and phenolic content for some species under elevated compared with ambient CO(2). It remains unclear how these changes in leaf properties affect herbivory. Damage to the leaves of hardwood trees by herbivorous insects may be reduced in the future as the concentration of CO(2) continues to increase, perhaps altering the trophic structure of forest ecosystems.

Genetic variation in primary metabolites of Pastinaca sativa; can herbivores act as selective agents?

Although insect herbivory has been shown to act as a selective agent on plant secondary metabolism, whether primary metabolites contribute to resistance and can respond to selection by herbivores remains untested. In the wild parsnip (Pastinaca sativa), its principal herbivore, Depressaria pastinacella, acts as a selective agent on furanocoumarin resistance factors. In this study, we determined whether webworms can, by causing differential reductions in fitness, act as selective agents on parsnip primary metabolites. Estimates of narrow-sense heritabilities were significantly different from zero for C18 fatty acids in buds and developing fruits, fructose and sorbitol in buds, fructose, myo-inositol, bergapten, and psoralen in fruits. Wild parsnips protected from webworms by insecticide produced 2.5 times as much seed biomass as unsprayed plants; that webworms accounted for this difference in plant fitness was indicated by a significant negative relationship between reproductive effort and an index of webworm damage. Only a handful of metabolites influenced resistance to webworms; these included osthol, sorbitol, and protein in developing fruits as well as previously documented furanocoumarins. Osthol, a coumarinic compound, enhanced resistance, as did protein content, while sorbitol lowered resistance. Other primary metabolites may affect resistance to webworms, but their effect was context-dependent, that is, their effect depended on concentrations of other metabolites (epistasis). Susceptible plant phenotypes were found to have average chemical compositions. Although there was genetic variation in some of the primary metabolites in parsnips, the epistatic nature of their involvement in resistance and the lack of genetic variation in some suggest that selection on them from webworms will be either inconsistent or ineffective.