Jack C. Schultz

Major Signaling Pathways Modulate Arabidopsis Glucosinolate Accumulation and Response to Both Phloem-Feeding and Chewing Insects

Plant responses to enemies are coordinated by several interacting signaling systems. Molecular and genetic studies with mutants and exogenous signal application suggest that jasmonate (JA)-, salicylate (SA)-, and ethylene (ET)-mediated pathways modulate expression of portions of the defense phenotype in Arabidopsis (Arabidopsis thaliana), but have not yet linked these observations directly with plant responses to insect attack. We compared the glucosinolate (GS) profiles of rosette leaves of 4-week-old mutant and transgenic Arabidopsis (Columbia) plants compromised in these three major signaling pathways, and characterized responses by those plants to feeding by two phloem-feeding aphids (generalist Myzus persicae and specialist Brevicoryne brassicae) and one generalist caterpillar species (Spodoptera exigua Hubner). Blocked JA signaling in coronatine-insensitive (coi1) and enhanced expression of SA-signaled disease resistance in hypersensitive response-like (hrl1) mutants reduced constitutive GS concentrations, while blocking SA signaling at the mediator protein npr1 mutant (NPR) increased them. There was no significant impact on constitutive GS contents of blocking ET signaling (at ET resistant [etr1]) or reducing SA concentrations (nahG transgene). We found increased GS accumulation in response to insect feeding, which required functional NPR1 and ETR1 but not COI1 or SA. Insect feeding caused increases primarily in short-chain aliphatic methylsulfinyl GS. By contrast, responses to exogenous JA, a frequent experimental surrogate for insect attack, were characterized by an increase in indolyl GS. Insect performance, measured as population increase or weight increase, was negatively related to GS levels, but we found evidence that other, ET-regulated factors may also be influential. Plant resistance to (consumption by) S. exigua was not related to insect growth because some plant chemistries inhibited growth while others inhibited feeding. These major signaling pathways modulate Arabidopsis GS accumulation and response to both phloem-feeding and chewing insects, often antagonistically; NPR appears to be central to these interactions. Our results indicate that exogenous signal application and plant consumption measures may not provide useful measures of plant responses to actual insect feeding.

Growth responses of tropical shrubs to treefall gap environments

To investigate the effects of differences in light and nutrient availability on growth, we planted seven species of shrubs in two genera, Miconia (Melastomataceae) and Piper (Piperaceae), into the centers, edges, and adjacent forest understory of four natural treefall gaps (275-335 M2) in the tropical, premontane rain forest of Costa Rica. We used rooted cuttings of species typical of forest understory environments on the one hand and large clearings or disturbed areas on the other. We also compared growth rates of three Miconia species grown in shade houses under 2, 20, and 40% full sunlight. Both light and nutrient availability in newly formed gaps of these sizes were strongly buffered by the canopy and root systems of the surrounding forest. Total incident radiation was higher in gap centers (9-23% full sunlight) than in gap-forest edges (3-11%) or under intact forest canopy (0.4-2%), but varied among similar microhabitats from different sites. Relative stem growth rates (RGRS) of all field-grown plants were significantly greater in gap centers than at edges or beneath forest understories. Fertilization did not significantly affect growth rate in any light environment. Light appears to be the most critical resource limiting growth at these gap sizes. In general, shade-tolerant species were less plastic than light-demanding species, but at these gap sizes grew as fast or faster in the gap centers. In shade-houses, the shade-tolerant species grew fastest at 20% full sunlight and light-demanding species grew fastest at 40% full sunlight. We found no evidence of a trade-off between growth and foliar phenolic concentration in these species.

THE GROWTH-DEFENSE TRADE-OFF AND HABITAT SPECIALIZATION BY PLANTS IN AMAZONIAN FORESTS

Tropical forests include a diversity of habitats, which has led to specialization in plants. Near Iquitos, in the Peruvian Amazon, nutrient-rich clay forests surround nutrient-poor white-sand forests, each harboring a unique composition of habitat specialist trees. We tested the hypothesis that the combination of impoverished soils and herbivory creates strong natural selection for plant defenses in white-sand forest, while rapid growth is favored in clay forests. Recently, we reported evidence from a reciprocal-transplant experiment that manipulated the presence of herbivores and involved 20 species from six genera, including phylogenetically independent pairs of closely related white-sand and clay specialists. When protected from herbivores, clay specialists exhibited faster growth rates than white-sand specialists in both habitats. But, when unprotected, white-sand specialists outperformed clay specialists in white-sand habitat, and clay specialists outperformed white-sand specialists in clay habitat. Here we test further the hypothesis that the growth defense trade-off contributes to habitat specialization by comparing patterns of growth, herbivory, and defensive traits in these same six genera of white-sand and clay specialists. While the probability of herbivore attack did not differ between the two habitats, an artificial defoliation experiment showed that the impact of herbivory on plant mortality was significantly greater in white-sand forests. We quantified the amount of terpenes, phenolics, leaf toughness, and available foliar protein for the plants in the experiment. Different genera invested in different defensive strategies, and we found strong evidence for phylogenetic constraint in defense type. Overall, however, we found significantly higher total defense investment for white-sand specialists, relative to their clay specialist congeners. Furthermore, herbivore resistance consistently exhibited a significant trade-off against growth rate in each of the six phylogenetically independent species-pairs. These results confirm theoretical predictions that a trade-off exists between growth rate and defense investment, causing white-sand and clay specialists to evolve divergent strategies. We propose that the growth-defense trade-off is universal and provides an important mechanism by which herbivores govern plant distribution patterns across resource gradients.

Relationships among Defoliation, Red Oak Phenolics, and Gypsy Moth Growth and Reproduction

This research assessed (1) the relationship between defoliation and phenolic chemistry of red oak trees, and (2) the relationship between food quality, measured in terms of oak phenolic chemistry and defoliation, and aspects of gypsy moth biology that can influence population dynamics. In a field experiment, larvae were reared in mesh bags on red oaks. Measurements of phenolics were made weekly on foliage inside and outside bags of 60 trees, which experienced 7 to 58% tree—wide defoliation by the end of the larval feeding period. We found that the phenological pattern of tree phenolic concentration over 8 wk was unrelated to defoliation or bagging. Bagging itself caused no change in foliar phenolic chemistry. Greater defoliation was associated with higher levels of total phenolics, hydrolyzable tannins, and protein—binding capacity of leaves. Pupal mass and fecundity were negatively correlated with both defoliation and the concentration of total phenolics, hydrolyzable tannins, and protein—binding capacity of the host tree. Additionally, egg mass was negatively correlated with the protein—binding capacity of the mothers diet. Statistical investigation of the relationship between components of food quality (both related and unrelated to damage) and insect performance, indicated that gypsy moth pupal mass was influenced by constitutive phenolic variation (unrelated to defoliation), by damage—induced nonphenolic variation, and by damage—induced phenolic variation. Variation in defoliation and in phenolic concentrations were related to variation in fecundity and egg mass.

Limitations of folin assays of foliar phenolics in ecological studies

We examined the response of the widely used Folin-Denis assay to purified tannins from 16 woody plant species and to three commercial polyphenol preparations often used as standards. The reagents response to these chemical mixtures differed significantly among sources (tree species, commercial preparations) and sampling dates, even though the mixtures contained the same total dry weight of tannins. Response to commercial standards usually did not resemble response to actual plant tannin and produced estimates that differed from actual concentrations by as much as twofold. Species-based and seasonal differences in polyphenol composition are evidently responsible for these variable results. Reagents that depend on redox reactions, such as the Folin-Denis, do not produce reliable absolute or relative quantification of phenolics when different species or samples from different dates are compared, and use of commercial standards does not resolve this problem.

Induced sink strength as a prerequisite for induced tannin biosynthesis in developing leaves of Populus

Induced defenses occur predominately in young, developing plant tissues that rely upon carbohydrate import to support their growth and development. To test the hypothesis that the induced production of carbon-based defenses is dependent upon photoassimilate import, we examined the response of developing leaves of hybrid poplar (Populus deltoides × P. nigra) saplings to wounding by gypsy moth caterpillars (Lymantria dispar L.) and exogenous jasmonic acid (JA). Growth rates, condensed tannin contents and acid invertase activities were measured for individual leaves and the translocation of 13C-labeled resources between orthostichous source-sink pairs was quantified. Results showed a substantial increase in the activity of cell wall invertase in sink leaves wounded by gypsy moth caterpillars and treated with JA. JA-induced sink leaves also imported 3–4 times as much 13C-labeled carbon from orthostichous source leaves relative to controls and allocated a significant portion of this imported 13C to condensed tannin biosynthesis. Reduced carbohydrate flow to these leaves, caused by source leaf removal, resulted in reduced condensed tannin levels and the emergence of a growth–defense tradeoff. These results indicate that (1) induced sink strength is elicited by insect wounding and JA application in hybrid poplar foliage, (2) imported resources are allocated to the production of carbon-based defenses, and (3) the level of induced defense in leaves can be constrained by the ability of leaves to import carbohydrates from source tissues. Together, these results suggest that within-canopy variations in induced resistance may arise in part because of uneven distribution of resources to induced foliage.

Induced plant defenses breached ? Phytochemical induction protects an herbivore from disease

Although wound-induced responses in plants are widespread, neither the ecological nor the evolutionary significance of phytochemical induction is clear. Several studies have shown, for example, that induced responses can act against both plant pathogens and herbivores simultaneously. We present the first evidence that phytochemical induction can inhibit a pathogen of the herbivore responsible for the defoliation. In 1990, we generated leaf damage by enclosing gypsy moth larvae on branches of red oak trees. We then inoculated a second cohort of larvae with a nuclear polyhedrosis virus (LdNPV) on foliage from the damaged branches. Larvae were less susceptible to virus consumed on foliage from branches with increasing levels of defoliation, and with higher concentrations of gallotannin. Defoliation itself was not related to any of our chemistry measures. Field sampling supported the results of our experiments: death from virus among feral larvae collected from unmanipulated trees was also negatively correlated with defoliation. In 1991, defoliation and gallotannin were again found to inhibit the virus. In addition, gallotannin concentrations were found to be positively correlated with defoliation the previous year. Compared with previous results that demonstrated a delecterious effect of induction on gypsy moth pupal weight and fecundity, the inhibition of the virus should confer an advantage to the gypsy moth. Since leaf damage levels increase as gypsy moth density increases, and since leaf damage inhibits the gypsy moth virus, there is the potential for positive feedback in the system. If phytochemical induction in red oak can inhibit an animal pathogen such as LdNPV, it suggests to us that induction in red oak is a generalized response to tissue damage rather than an adaptive defense against herbivores.

Hostplant, larval age, and feeding behavior influence midgut pH in the gypsy moth (Lymantria dispar)

SummaryThe midgut pH of late instar gypsy moth (Lymantria dispar L.) larvae is strongly alkaline, and varies with diet, larval stadium, and time since feeding. Midgut pH rises with time since feeding, and does so more quickly, reaching greater maximum values, on some diets than others. Leaf tissues of 23 tree species resist increases in alkalinity differentially; this trait and differing initial leaf pH may explain the impact of diet on gut pH. Third instar larvae may have gut conditions favorable for tannin-protein binding shortly after ingesting certain foods, but with time midgut alkalinity becomes great enough to dissociate tannin-protein complexes. Older instars rarely exhibit gut pHs low enough to permit tannin activity. Alkaline gut conditions may explain the gypsy moths ability to feed on many tanniniferous plant species, especially in later instars. Consequences for pathogen effectiveness are discussed.

Plant responses induced by herbivores.

Physiological and chemical traits of many plant species change in response to real or simulated herbivory. These changes often have significant impacts on behavior, growth, survivorship, feeding and oviposition of insects. However, evidence that plants gain direct or indirect protection from insect enemies thereby is equivocal at present. Evidence is lacking for an impact of induced defenses on insect population dynamics, but few studies have sought it. More detailed studies of plant physiology, biochemistry, genetics and net benefit to individual plants are needed to identify the adaptive significance of induced defenses.

Chemical defense production in Lotus corniculatus L. II. Trade-offs among growth, reproduction and defense.

SummaryEcological trade-offs between growth, reproduction and both condensed tannins and cyanogenic glycosides were examined in Lotus corniculatus by correlating shoot (leaves and stem) size and reproductive output with chemical concentrations. We found that cyanide concentration was not related to shoot size, but that condensed tannin concentrations were positively correlated with shoot size; larger plants contained higher tannin concentrations. Both tannin and cyanide concentrations were depressed when plants produced fruits. Defense costs change as plants mature and begin to reproduce. These trade-offs indicate that cost of defense chemical production cannot be predicted merely on the basis of molecular size, composition or concentration.