Conifers possess a suite of physiochemical defenses that protect their subcortical tissues from bark beetle - fungal complexes. These defenses include rapid induction of terpenoids and phenolics at the site of attack. Studies of the distribution, induction, and bioactivity of conifer terpenoids have focused heavily on monoterpenes. We assessed induction of diterpene acids in white spruce (Picea glauca) and red pine (Pinus resinosa) to fungal associates of two bark beetles, and the responses of four spruce beetle (Dendroctonus rufipennis)—associated fungi to three diterpene acids. Constitutive phloem contents differed between species, in that red pine had extremely low concentrations of diterpene acids, whereas white spruce had substantial constitutive levels. Induction differed quantitatively. Both red pine and white spruce exhibited marked increases, but red pine underwent greater increases and achieved higher concentrations than white spruce. Induction also differed qualitatively in that red pine showed lower diversity and fewer compositional changes during induction than white spruce. In red pine,fungal inoculation accompanying wounding elicited greater increases than wounding alone, but in white spruce total concentrations were higher following wounding alone. Spruce beetle fungal symbiont growth varied among species and compounds. Some diterpenes elicited both stimulatory and inhibitory effects on fungi, depending on concentration. All four fungi exhibited higher tolerances compared to those associated with pine bark beetles in previous studies. Variation in tolerances to, and potentially metabolism of, diterpene acids by symbionts may reflect differences in constitutive levels between spruce and pine, and partially explain differences in concentrations achieved during induction.

Diterpene resin acids are major constituents of conifer oleoresin and play important roles in tree defense against insects and microbial pathogens. The tricyclic C-20 carboxylic acids are generally classified into two groups, the abietanes and the pimaranes. The abietanes have conjugated double bonds and exhibit characteristic UV spectra. Here, we report the analysis of abietanes by reversed-phase high-performance liquid chromatography using multiwavelength detection to optimize quantification of underivatized abietic, neoabietic, palustric, levopimaric, and dehydroabietic acids. The utility of the method is demonstrated with methanol extracts of white spruce (Picea glauca) phloem, and representative concentrations are reported.

Conifer resin and phloem tissue contain several phytochemical groups, composed primarily of monoterpenes, diterpene acids, and stilbene phenolics. The effects of monoterpenes and phenolics on stem-colonizing bark beetles and their associated microorganisms have been studied to some extent, but the roles of diterpene acids are largely unknown. Diterpene acids are known to have substantial feeding deterrent and growth inhibiting effects on a variety of insect groups and are known to inhibit a variety of fungi. We tested three diterpene acids present in red pine, Pinus resinosa, at various concentrations, on several life history components of the bark beetle Ips pini and the fungus Ophiostoma ips. No diterpene acid affected the host acceptance behavior or larval survival of Ips pini. In contrast, abietic acid and isopmaric acid strongly inhibited spore germination of O. ips, and abietic acid strongly inhibited mycelial growth. The levels of inhibition observed were higher than with any previous assays of monoterpenes or phenolics in this system. These results support the view that conifer defenses against bark beetle-fungal complexes are multifaceted, with all three phytochemical groups being important to P. resinosa, but each with varying relative activity against the beetles and fungi.

Efforts to describe the complex relationships between bark beetles and the ophiostomatoid (stain) fungi they transport have largely resulted in a dichotomous classification. These symbioses have been viewed as either mutualistic (i.e., fungi help bark beetles colonize living trees by overcoming tree Menses or by providing nutrients after colonization in return for transport to a host) or antagonistic (i.e., fungi compete for a limited resource and reduce brood development with no apparent benefit to the beetle). We investigated several components of one beetle-fungus interaction. Specifically, we addressed whether beetle entry into, and development within, a host tree vary with the degree of colonization by ophiostomatoid fungi. Ips pini (Say) transports several species of ophiostomatoid fungi, the most common being Ophiostoma ips (Rumbold) Nannfeldt, in the process of colonizing its host, Pinus resinosa Aitman. We introduced this fungus 0, 3, 7, and 10 d before beetle entry to characterize its effects on L pini colonization and development. This sequence allowed quantification of temporal effects and comparison of results with other systems. Fungal growth was greatest when inoculated before beetle colonization. Fungal colonization reduced beetle entry into logs, but increased brood production. Mate capture was not significantly affected by fungal growth. The benefits imparted by O. ips to its beetle vector during brood development are compared with results from other systems. This difference may in part be related to the exploitation of highly stressed and dead trees, rather than vigorous hosts, by I. pini.

Summary The impacts of elevated atmospheric CO2 and/or O3 have been examined over 4 years using an open‐air exposure system in an aggrading northern temperate forest containing two different functional groups (the indeterminate, pioneer, O3‐sensitive species Trembling Aspen, Populus tremuloides and Paper Birch, Betula papyrifera, and the determinate, late successional, O3‐tolerant species Sugar Maple, Acer saccharum). The responses to these interacting greenhouse gases have been remarkably consistent in pure Aspen stands and in mixed Aspen/Birch and Aspen/Maple stands, from leaf to ecosystem level, for O3‐tolerant as well as O3‐sensitive genotypes and across various trophic levels. These two gases act in opposing ways, and even at low concentrations (1·5 × ambient, with ambient averaging 34–36 nL L−1 during the summer daylight hours), O3 offsets or moderates the responses induced by elevated CO2. After 3 years of exposure to 560 µmol mol−1 CO2, the above‐ground volume of Aspen stands was 40% above those grown at ambient CO2, and there was no indication of a diminishing growth trend. In contrast, O3 at 1·5 × ambient completely offset the growth enhancement by CO2, both for O3‐sensitive and O3‐tolerant clones. Implications of this finding for carbon sequestration, plantations to reduce excess CO2, and global models of forest productivity and climate change are presented.

We investigated the effects of CO2 enrichment on fine root chemical composition of two tree species common to northern hardwood forests. Two-year-old Betula papyrifera and 3-year-old Acer saccharum saplings were grown under ambient (400 µmol·mol–1) and elevated (700 µmol·mol–1) CO2 in a glasshouse experiment. In both species, root/shoot ratios and fine root percentages (of total biomass) were unaltered by CO2 enrichment. Tissue nitrogen concentrations decreased in the fine roots, and consequently, C/N ratios increased with elevated CO2. In birch, only condensed tannins increased with CO2 enrichment, while root starch levels were conserved. In maple, neither condensed tannins nor hydrolysable tannins were positively influenced by elevated CO2. Both fine root biomass and chemistry responses of the tree saplings may be related to their successional status.

Nature 420, 403–407 (2002). In this Letter, the conversion to SI units led to several errors. On page 404, left column, lines 16 and 17, the values should read 10–15 and 30–40 nanolitres per litre. On page 405, right column, lines 3 and 4, the values should read 360 microlitres per litre and 36.0–38.8 nanolitres per litre. On page 407, left column, lines 3 and 4, the values should read 560 microlitres per litre, and 46.4 to 55.5 nanolitres per litre. The conclusions of the paper are not affected.

I This research was conducted at the Aspen FACE (Free Air CO2 Enrichment) site located in northern Wisconsin, U.S.A. where trembling aspen (Populus tremuloides Michaux) trees were exposed to one of four atmospheric treatments: elevated carbon dioxide (CO2; 560 muL/L), elevated ozone (O-3; ambient x 1.5), elevated CO2 and 03, or ambient air. We evaluated the effects of these fumigants on aspen foliar quality and the performance of aspen blotch leafminer (Phyllonorycter tremuloidiella Braun).2 CO2 and O-3 each affected foliar quality, with the major changes consisting of an 11% reduction in nitrogen under elevated CO2 and a 20% reduction in tremulacin under elevated O-3. In the CO2+O-3 treatment, nitrogen levels were reduced by 15% and CO2 ameliorated the O-3-mediated reduction in tremulacin levels.3 Phyllonorycter tremuloidiella were allowed to colonize trees naturally. Elevated CO2 and O-3 reduced colonization rates by 42 and 49% relative to ambient CO2 and O-3, respectively. The only effect of fumigation treatments on larval performance occurred under elevated O-3, where male development time and larval consumption increased by 8 and 28%, respectively, over insects reared under ambient O-3.4 These data demonstrate that the individual and combined effects Of CO2 and O-3 can alter aspen foliar chemistry and that these alterations in foliar chemistry produce little to no change in larval performance. However, both CO2 and O-3 greatly reduced oviposition. In order to ascertain the full effects Of CO2 and O-3 on insect performance, future studies should address both population- and individual-level characteristics.

Nature 420, 403–407 (2002). In this Letter, the conversion to SI units led to several errors. On page 404, left column, lines 16 and 17, the values should read 10–15 and 30–40 nanolitres per litre. On page 405, right column, lines 3 and 4, the values should read 360 microlitres per litre and 36.0–38.8 nanolitres per litre.

Summary Modified White's solution (1 g HgCl 2 /l H 2 O) is widely used to surface disinfest bark beetles of their phoretic fungi. We investigated the effectiveness of this solution at disinfesting adult Ips pini from its associated ophiostomatoid fungi. A treatment for 1, 4 or 8 min does not completely rid beetles of phoretic fungi, but does substantially reduce the amount of fungi they carry externally. Sterilizing with modified White's solution caused limited mortality (<16%).

This research tested the effects of paper birch, Betula papyrifera Marshall, condensed tannin on larval performance of the whitemarked tussock moth, Orgyia leucostigma (J. E. Smith). We conducted laboratory bioassays on fifth stadium larvae. Larvae were reared on one of three diets: control (no condensed tannin), moderate condensed tannin (8.8% dry mass), and high condensed tannin (17.6% dry mass). Although survivorship was not different between the treatments, larvae fed diets amended with condensed tannin exhibited increased stadium duration, decreased relative growth rate, and decreased food conversion efficiencies. Prolonged development times enabled larvae to compensate for low consumption and growth rates such that insects on tannin diets ate more and grew larger than insects on the control diet. Analysis of tannin levels in food, frass, and body tissue indicated that larvae do not metabolize condensed tannin, but concentrate and egest it. Our results show that paper birch condensed tannin has both positive and negative effects on the performance indices of whitemarked tussock moths. However, whether the benefits of increased final size (and possibly fecundity) outweigh the risks of increased development time and prolonged exposure to natural enemies remains unclear.

1 Genetic variation in the phytochemical responses of plants to CO2 enrichment is likely to alter trophic dynamics, and to shift intraspecific selection pressures on plant populations. We evaluated the independent and interactive effects of atmospheric CO2 and quaking aspen (Populus trernuloides Michx.) genotype on chemical composition of foliage and performance of the whitemarked tussock moth (Orgyia leucostigrna J. E. Sm.).2 This research was conducted at the Aspen FACE (Free Air CO2 Enrichment) site in northern Wisconsin, U.S.A. Leaf samples were collected periodically from each of three genetically variable aspen genotypes growing under ambient and elevated CO2, and analysed for levels of primary and secondary metabolites. Tussock moth larvae were reared in situ on experimental trees, and development times and pupal masses were recorded.3 Foliar chemical composition varied among aspen genotypes and in response to CO2 enrichment. However, chemical responses of trees to elevated CO2 were generally consistent across genotypes.4 Larval development times varied among host genotypes and increased slightly for insects on high-CO2 plants. Enriched CO2 tended to reduce insect pupal masses, particularly for females on one of the three aspen genotypes.5 CO2 x genotype interactions observed for plant chemistry and insect performance in this study with a small number of genotypes are probably too few, and too weak, to shift selection pressures in aspen populations. These results differ, however, from earlier work in which more substantial CO2 x genotype interactions were observed for plant chemistry.

Human activity causes increasing background concentrations of the greenhouse gases CO2 and O-3(1). Increased levels of CO2 can be found in all terrestrial ecosystems(2). Damaging O-3 concentrations currently occur over 29% of the world's temperate and subpolar forests but are predicted to affect fully 60% by 2100 (ref. 3). Although individual effects of CO2 and O-3 on vegetation have been widely investigated, very little is known about their interaction, and long-term studies on mature trees and higher trophic levels are extremely rare(4). Here we present evidence from the most widely distributed North American tree species(5), Populus tremuloides, showing that CO2 and O-3, singly and in combination, affected productivity, physical and chemical leaf defences and, because of changes in plant quality, insect and disease populations. Our data show that feedbacks to plant growth from changes induced by CO2 and O-3 in plant quality and pest performance are likely. Assessments of global change effects on forest ecosystems must therefore consider the interacting effects of CO2 and O-3 on plant performance, as well as the implications of increased pest activity.

The purpose of this study was to assess the independent and interactive effects of CO2, O3, and plant genotype on the foliar quality of a deciduous tree and the performance of a herbivorous insect. Two trembling aspen (Populus tremuloides Michaux) genotypes differing in response to CO2 and O3 were grown at the Aspen FACE (Free Air CO2 Enrichment) site located in northern Wisconsin, USA. Trees were exposed to one of four atmospheric treatments: ambient air (control), elevated carbon dioxide (+CO2; 560 µl/l), elevated ozone (+O3; ambient ×1.5), and elevated CO2+O3. We measured the effects of CO2 and O3 on aspen phytochemistry and on performance of forest tent caterpillar (Malacosoma disstria Hübner) larvae. CO2 and O3 treatments influenced foliar quality for both genotypes, with the most notable effects being that elevated CO2 reduced nitrogen and increased tremulacin levels, whereas elevated O3 increased early season nitrogen and reduced tremulacin levels, relative to controls. With respect to insects, the +CO2 treatment had little or no effect on larval performance. Larval performance improved in the +O3 treatment, but this response was negated by the addition of elevated CO2 (i.e., +CO2+O3 treatment). We conclude that tent caterpillars will have the greatest impact on aspen under current CO2 and high O3 levels, due to increases in insect performance and decreases in tree growth, whereas tent caterpillars will have the least impact on aspen under high CO2 and low O3 levels, due to moderate changes in insect performance and increases in tree growth.

Adult behaviors of the regal fritillary, Speyeria idalia (Drury), were studied in Kansas tallgrass prairie. Our objective was to determine nectar resources, adult emergence patterns, and behaviors throughout the flight period, which extends from early June to early October, Availability of nectar sources changed throughout the adult flight period. Adults exhibited extreme protandry; males emerged in early June (ca. 2 weeks before females) and showed flight patterns suggestive of searching for females, even though none were present. Mating occurred soon after females emerged, and most males were gone by July 10. Females reduced flight activity and remained hidden in clumps of plants from the time they eclosed (mid-June) until late August/early September. Females resumed flight activity a week prior to oviposition.

Atmospheric chemical composition affects foliar chemical composition, which in turn influences the dynamics of both herbivory and decomposition in ecosystems. We assessed the independent and interactive effects of CO2 and O-3 fumigation on foliar chemistry of quaking aspen (Populus tremuloides) and paper birch (Betula papyrifera) at a Free-Air CO2 Enrichment (FACE) facility in northern Wisconsin. Leaf samples were collected at five time, periods during a single growing season, and analyzed for nitrogen, starch and condensed tannin concentrations, nitrogen resorption efficiencies (NREs), and C:N ratios. Enriched CO2 reduced foliar nitrogen concentrations in aspen and birch; O-3 only marginally reduced nitrogen concentrations. NREs were unaffected by pollution treatment in aspen, declined with O-3 exposure in birch, and this decline was ameliorated by enriched CO2- C:N ratios of abscised leaves increased in response to enriched CO2 in both tree species. O-3 did not significantly alter C:N ratios in aspen, although values tended to be higher in + CO2 + O-3 leaves. For birch, O-3 decreased C:N ratios under ambient CO2 and increased C:N ratios under elevated CO2. Thus, under the combined pollutants. the C:N ratios of both aspen and birch leaves were elevated above the averaged responses to the individual and independent trace gas treatments. Starch concentrations were largely unresponsive to CO2 and O-3 treatments in aspen, but increased in response to elevated CO2 in birch. Levels of condensed tannins were negligibly affected by CO2 and O-3 treatments in aspen, but increased in response to enriched CO, in birch. Results from this work suggest that changes in foliar chemical composition elicited by enriched CO2 are likely to impact herbivory and decomposition, whereas the effects Of O-3 are likely to be minor, except in cases where they influence plant response to CO2. (C) 2001 Elsevier Science Ltd. All rights reserved.

Reproductive characteristics of a northeastern Kansas population of the fritillary Speyeria idalia (Drury) were studied. Hemolymph juvenile hormone (JH) titers, ovarian development, and fat body utilization were monitored weekly in adult females over their entire 1997 flight period, which extended from mid-June to early October. Dissections of female reproductive systems revealed that S. idalia females mate just once, soon after they emerge in mid- to late June. Gas chromatographic-mass spectrometric determinations of juvenile hormone suggested that they undergo a reproductive diapause through mid-August related to the absence of or very low titers of juvenile hormone. Oogenesis and fat body. depiction do not commence until late August/early September, soon after which oviposition occurs. The onset of oogenesis coincides with a rapid rise in hemolymph titers of JH I, JH II, and JH III. The predominant juvenile hormone homolog was IH II, but both JH I and JH III exhibited smaller, concomitant peaks. Four fundamentally different lepidopteran reproductive strategies have been recognized based on various reproductive characteristics and the type of gonadotropic hormones used to stimulate oogenesis. Speyeria idalia exhibits a type of reproductive strategy that has not been documented in Lepidoptera, typified by protandry, female monandry, long-lived (>8 wk) females that feed throughout their adult lives, greatly delayed oogenesis that occurs late in adult life, and apparent juvenile hormone control of gonadotropic processes. This reproductive strategy appears to be an adaptation to the phenology of larval host plants, namely coordinating the life cycle with that of the seasonally restricted violets on which larval survival of this monophagous species depends.