Forest diversity effects on insect herbivores: do leaf traits matter?

Insect herbivore damage and abundance are often reduced in diverse plant stands. However, few studies have explored whether this phenomenon is a result of plant diversity effects on host plant traits. We explored indirect effects of tree species diversity on herbivory via changes in leaf traits in a long-term forest diversity experiment in Finland. We measured 16 leaf traits and leaf damage by four insect guilds (chewers, gall formers, leaf miners and rollers) on silver birch (Betula pendula) trees growing in one-, two-, three- and five-species mixtures. A decline in the frequency of birch in mixed stands resulted in reduced leaf area. This, in turn, mediated the reduction in chewing damage in mixed stands. In contrast, associational resistance of birch to leaf miners was not trait-mediated but driven directly by concurrent declines in birch frequency as tree species richness increased. Our results show that leaf trait variation across the diversity gradient might promote associational resistance, but these patterns are driven by an increase in the relative abundance of heterospecifics rather than by tree species richness per se. Therefore, accounting for concurrent changes in stand structure and key foliar traits is important for the interpretation of plant diversity effects and predictions of associational patterns.

Contrasting effects of tree species and genetic diversity on the leaf-miner communities associated with silver birch.

Both species and genetic diversity of plant communities can affect insect herbivores, but a few studies have compared the effects of both diversity levels within the same experimental context. We compared the effects of tree species and genetic diversity on abundance, species richness, and ß-diversity of leaf-miner communities associated with silver birch using two long-term forest diversity experiments in Finland where birch trees were planted in monocultures and mixtures of birch genotypes or other trees species. Although both abundance and species richness of leaf miners differed among birch genotypes at the tree level, birch genetic diversity had no significant effect on miner abundance and species richness at the plot level. Instead, birch genetic diversity affected leaf-miner ß-diversity with species turnover being higher among trees within genotypic mixtures than among trees within monoclonal plots. In contrast, tree species diversity had a significant negative effect on both leaf-miner abundance and species richness at plot level, but no effect on miner ß-diversity. Significant tree species diversity effects on leaf-miner abundance and species richness were found only in plots with high tree density. We have demonstrated that plant species and genetic diversity play important but contrasting roles in structuring associated herbivore communities. Tree species diversity largely affects miner abundance and species richness, whereas tree genetic diversity affects miner ß-diversity. These results have important implications for conservation and management of woodlands.

Genotypic variation in a foundation tree (Populus tremula L.) explains community structure of associated epiphytes.

Community genetics hypothesizes that within a foundation species, the genotype of an individual significantly influences the assemblage of dependent organisms. To assess whether these intra-specific genetic effects are ecologically important, it is required to compare their impact on dependent organisms with that attributable to environmental variation experienced over relevant spatial scales. We assessed bark epiphytes on 27 aspen (Populus tremula L.) genotypes grown in a randomized experimental array at two contrasting sites spanning the environmental conditions from which the aspen genotypes were collected. We found that variation in aspen genotype significantly influenced bark epiphyte community composition, and to the same degree as environmental variation between the test sites. We conclude that maintaining genotypic diversity of foundation species may be crucial for conservation of associated biodiversity.

Microsite affects willow sapling recovery from bank vole (Myodes glareolus) herbivory, but does not affect grazing risk.

BACKGROUND: Large herbivores are often removed or reduced as part of vegetation restoration programmes, but the resultant increase in vegetation biomass and changes in vegetation structure may favour small mammals. Small mammals may have large impacts on plant community composition via granivory and sapling herbivory, and increased small mammal populations may reduce any benefits of large herbivore removal for highly preferred species. This study tested the impacts of small mammal herbivory, microsite characteristics and their interaction on growth and survival of three montane willow species with differing chemical compositions, Salix lapponum, S. myrsinifolia and S. arbuscula. METHODS: In two separate years, 1-year-old saplings were planted within a 180 ha, large-mammal scrub regeneration exclosure, and either experimentally protected from or exposed to small mammals (bank voles). Saplings were planted in one of two microsite treatments, vegetation mown (to mimic a grazed sward) or disturbed (all above- and below-ground competition removed), and monitored throughout the first year of growth. KEY RESULTS: Approximately 40 % of saplings planted out in each year were damaged by bank voles, but direct mortality due to damage was very low (<2 %). There were no strong species differences in susceptibility to vole damage. Microsite treatment had no impact on the proportion of saplings attacked, but in 2004 saplings in mown microsites were more severely damaged and had smaller increases in size than those in disturbed microsites. In 2003, saplings in mown microsites had smaller increases in stem diameter following attack than those in disturbed microsites. CONCLUSIONS: Planting 1-year-old willow saplings into disturbed microsites may aid growth, reduce the severity of small mammal damage and improve recovery following sub-lethal small mammal damage. Restoration management of montane willow scrub should therefore consider manipulating the planting site to provide disturbed areas of soil.

Identifying and testing marker-trait associations for growth and phenology in three pine species: Implications for genomic prediction.

In tree species, genomic prediction offers the potential to forecast mature trait values in early growth stages, if robust marker-trait associations can be identified. Here we apply a novel multispecies approach using genotypes from a new genotyping array, based on 20,795 single nucleotide polymorphisms (SNPs) from three closely related pine species (Pinus sylvestris, Pinus uncinata and Pinus mugo), to test for associations with growth and phenology data from a common garden study. Predictive models constructed using significantly associated SNPs were then tested and applied to an independent multisite field trial of P. sylvestris and the capability to predict trait values was evaluated. One hundred and eighteen SNPs showed significant associations with the traits in the pine species. Common SNPs (MAF > 0.05) associated with bud set were only found in genes putatively involved in growth and development, whereas those associated with growth and budburst were also located in genes putatively involved in response to environment and, to a lesser extent, reproduction. At one of the two independent sites, the model we developed produced highly significant correlations between predicted values and observed height data (YA, height 2020: r = 0.376, p < 0.001). Predicted values estimated with our budburst model were weakly but positively correlated with duration of budburst at one of the sites (GS, 2015: r = 0.204, p = 0.034; 2018: r = 0.205, p = 0.034-0.037) and negatively associated with budburst timing at the other (YA: r = -0.202, p = 0.046). Genomic prediction resulted in the selection of sets of trees whose mean height was taller than the average for each site. Our results provide tentative support for the capability of prediction models to forecast trait values in trees, while highlighting the need for caution in applying them to trees grown in different environments.

Phenotypic trait variation in a long-term multisite common garden experiment of Scots pine in Scotland.

Multisite common garden experiments, exposing common pools of genetic diversity to a range of environments, allow quantification of plastic and genetic components of trait variation. For tree species, such studies must be long term as they typically only express mature traits after many years. As well as evaluating standing genetic diversity, these experiments provide an ongoing test of genetic variation against changing environmental conditions and form a vital resource for understanding how species respond to abiotic and biotic variation. Finally, quantitative assessments of phenotypic variation are essential to pair with rapidly accumulating genomic data to advance understanding of the genetic basis of trait variation, and its interaction with climatic change. We describe a multisite, population-progeny, common garden experiment of the economically and ecologically important tree species, Scots pine, collected from across its native range in Scotland and grown in three contrasting environments. Phenotypic traits, including height, stem diameter and budburst were measured over 14 growing seasons from nursery to field site. The datasets presented have a wide range of applications.

Location, but not defensive genotype, determines ectomycorrhizal community composition in Scots pine (Pinus sylvestris L.) seedlings.

For successful colonization of host roots, ectomycorrhizal (EM) fungi must overcome host defense systems, and defensive phenotypes have previously been shown to affect the community composition of EM fungi associated with hosts. Secondary metabolites, such as terpenes, form a core part of these defense systems, but it is not yet understood whether variation in these constitutive defenses can result in variation in the colonization of hosts by specific fungal species.We planted seedlings from twelve maternal families of Scots pine (Pinus sylvestris) of known terpene genotype reciprocally in the field in each of six sites. After 3 months, we characterized the mycorrhizal fungal community of each seedling using a combination of morphological categorization and molecular barcoding, and assessed the terpene chemodiversity for a subset of the seedlings. We examined whether parental genotype or terpene chemodiversity affected the diversity or composition of a seedling's mycorrhizal community.While we found that terpene chemodiversity was highly heritable, we found no evidence that parental defensive genotype or a seedling's terpene chemodiversity affected associations with EM fungi. Instead, we found that the location of seedlings, both within and among sites, was the only determinant of the diversity and makeup of EM communities.These results show that while EM community composition varies within Scotland at both large and small scales, variation in constitutive defensive compounds does not determine the EM communities of closely cohabiting pine seedlings. Patchy distributions of EM fungi at small scales may render any genetic variation in associations with different species unrealizable in field conditions. The case for selection on traits mediating associations with specific fungal species may thus be overstated, at least in seedlings.

Host Genetics and Environment Drive Divergent Responses of Two Resource Sharing Gall-Formers on Norway Spruce: A Common Garden Analysis.

A central issue in the field of community genetics is the expectation that trait variation among genotypes play a defining role in structuring associated species and in forming community phenotypes. Quantifying the existence of such community phenotypes in two common garden environments also has important consequences for our understanding of gene-by-environment interactions at the community level. The existence of community phenotypes has not been evaluated in the crowns of boreal forest trees. In this study we address the influence of tree genetics on needle chemistry and genetic x environment interactions on two gall-inducing adelgid aphids (Adelges spp. and Sacchiphantes spp.) that share the same elongating bud/shoot niche. We examine the hypothesis that the canopies of different genotypes of Norway spruce (Picea abies L.) support different community phenotypes. Three patterns emerged. First, the two gallers show clear differences in their response to host genetics and environment. Whereas genetics significantly affected the abundance of Adelges spp. galls, Sacchiphantes spp. was predominately affected by the environment suggesting that the genetic influence is stronger in Adelges spp. Second, the among family variation in genetically controlled resistance was large, i.e. fullsib families differed as much as 10 fold in susceptibility towards Adelges spp. (0.57 to 6.2 galls/branch). Also, the distribution of chemical profiles was continuous, showing both overlap as well as examples of significant differences among fullsib families. Third, despite the predicted effects of host chemistry on galls, principal component analyses using 31 different phenolic substances showed only limited association with galls and a similarity test showed that trees with similar phenolic chemical characteristics, did not host more similar communities of gallers. Nonetheless, the large genetic variation in trait expression and clear differences in how community members respond to host genetics supports our hypothesis that the canopies of Norway spruce differ in their community phenotypes.

Growth and mortality in mountain hares: the effect of sex and date of birth.

Current theory suggests that high juvenile growth effort, may result in higher mortality. This prediction is tested in mountain hares (Lepus timidus), by examination of post-weaning growth and mortality of males and females. Dates of birth were estimated from weight at first capture and growth in body weight and hind foot length was described by the logistic growth equation. Although adult female hares are on average larger than males, this results from a longer period of growth and not from a faster growth rate. There was no clear sex bias in mortality, a slight but not significantly greater proportion of males suffered mortality during the growth period. Predictions as to sex difference in mortality should specify which aspects of growth (rate or duration) are the agents of mortality, since the two parameters are not necessarily positively associated. Overall asymptotic body size decreased and rate of growth increased from early through to late-born young. Rate of growth in hind foot length was greater in late-born males but not in late-born females as compared with those born earlier. It is hypothesized that late-born males are more tightly constrained to complete growth and subsequent sexual maturation earlier than late-born females.

The flexibility of an intermediate feeder: dietary selection by mountain hares measured using faecal n-alkanes.

Herbivores with an intermediate feeding strategy either vary their diet between a grazing (bulk roughage feeders) or browsing (concentrate selectors) strategy on a seasonal basis or select a mixed diet at any one time. The underlying ecological causes of the seasonal dietary shift in a small non-ruminant intermediate feeder - the mountain or arctic hare (Lepus timidus L.) were determined. Diet composition and selection relative to availability were investigated for 41 individual free-ranging mountain hares (of which 18 female hares were radio-collared) occupying an upland mosaic landscape in north-east Scotland. Diet composition was determined using faecal n-alkane analysis. Radio-collared hares were designated as pasture, woodland or moorland hares according to the habitat that predominated their home-range. In common with previous studies, mountain hares switched from a browse-dominated diet during winter to a Gramineae-dominated diet in summer, although it was only significant for reproductively active females during the peak breeding season. Diet composition remained consistent regardless of habitat occupied. However, the diet of radio-tracked hares differed significantly from the biomass available in the individual home-ranges; Gramineae were preferentially selected over browse species throughout the year. During winter and in particular during the early breeding season, intermediate feeders, such as mountain hares, ate browse material when the availability of higher quality was restricted. The ability to browse or graze represents a flexible foraging strategy permitting survival and production through periods of changing or unpredictable forage quality and availability.

Genetic and ontogenetic variation in an endangered tree structures dependent arthropod and fungal communities.

Plant genetic and ontogenetic variation can significantly impact dependent fungal and arthropod communities. However, little is known of the relative importance of these extended genetic and ontogenetic effects within a species. Using a common garden trial, we compared the dependent arthropod and fungal community on 222 progeny from two highly differentiated populations of the endangered heteroblastic tree species, Eucalyptus morrisbyi. We assessed arthropod and fungal communities on both juvenile and adult foliage. The community variation was related to previous levels of marsupial browsing, as well as the variation in the physicochemical properties of leaves using near-infrared spectroscopy. We found highly significant differences in community composition, abundance and diversity parameters between eucalypt source populations in the common garden, and these were comparable to differences between the distinctive juvenile and adult foliage. The physicochemical properties assessed accounted for a significant percentage of the community variation but did not explain fully the community differences between populations and foliage types. Similarly, while differences in population susceptibility to a major marsupial herbivore may result in diffuse genetic effects on the dependent community, this still did not account for the large genetic-based differences in dependent communities between populations. Our results emphasize the importance of maintaining the populations of this rare species as separate management units, as not only are the populations highly genetically structured, this variation may alter the trajectory of biotic colonization of conservation plantings.

Do multiple herbivores maintain chemical diversity of Scots pine monoterpenes?

A central issue in our understanding of the evolution of the diversity of plant secondary metabolites (PSMs) is whether or not compounds are functional, conferring an advantage to the plant, or non-functional. We examine the hypothesis that the diversity of monoterpene PSMs within a plant species (Scots pine Pinus sylvestris) may be explained by different compounds acting as defences against high-impact herbivores operating at different life stages. We also hypothesize that pairwise coevolution, with uncorrelated interactions, is more likely to result in greater PSM diversity, than diffuse coevolution. We tested whether up to 13 different monoterpenes in Scots pine were inhibitory to herbivory by slugs (Arion ater), bank voles (Clethrionomys glareolus), red deer (Cervus elaphus) and capercaillie (Tetrao urogallus), each of which attack trees at a different life stage. Plants containing more α-pinene were avoided by both slugs and capercaillie, which may act as reinforcing selective agents for this dominant defensive compound. Herbivory by red deer and capercaillie were, respectively, weakly negatively associated with δ(3)-carene, and strongly negatively correlated with the minor compound ß-ocimene. Three of the four herbivores are probably contributory selective agents on some of the terpenes, and thus maintain some, but by no means all, of the phytochemical diversity in the species. The correlated defensive function of α-pinene against slugs and capercaillie is consistent with diffuse coevolutionary processes.

Assessment and implications of intraspecific and phenological variability in monoterpenes of Scots pine (Pinus sylvestris) foliage.

Scots pine populations contain individuals with widely differing amounts and composition of monoterpenes and exist as one of two chemotypes: with or without delta3-carene. We investigated the significance for ecological studies of two types of variation in monoterpenes: (1) the inherent variability in the concentration of monoterpenes or their relative amounts in needles of seedlings, saplings, and mature trees; and (2) phenological variation in developing needles. The relative composition of needle monoterpenes in 5-year-old saplings changed during the needle development period until the final composition was reached upon needle maturity. Changes in composition depended on chemotype. Needles of the "no-delta3-carene" chemotype had higher absolute concentrations of alpha-pinene, beta-pinene, camphene, and total monoterpenes than "delta3-carene" chemotype. For the "delta3-carene" chemotype, the relative concentration of delta3-carene during the needle growing season and immediately after emergence of seedlings was higher compared to that reached at needle maturity. Repeated removal of single needles (at weekly intervals during growth) from 5-year-old saplings did not influence the composition of monoterpenes. Within a natural Scots pine dominated woodland, 18% of mature Scots pines (N=574) belonged to the "no-delta3-carene" chemotype. Chemotypic variation within populations means that the statistical power with which differences in monoterpene concentrations can be detected is lower when sampling from the whole population compared to sampling within chemotypes. Reduction of this background variation and accounting for chiral variation if present, would significantly aid efficiency, interpretation, and understanding of processes in chemical and ecological research. One method for achieving this is the screening of plants for chemotypes before the establishment of experiments or field sampling regimes. We present a summary of suitable analytical methods for needle tissue that facilitates this prior screening.

The role of genetic and chemical variation of Pinus sylvestris seedlings in influencing slug herbivory.

This study investigated the genetic and chemical basis of resistance of Pinus sylvestris seedlings to herbivory by a generalist mollusc, Arion ater. Using feeding trials with captive animals, we examined selective herbivory by A. ater of young P. sylvestris seedlings of different genotypes and correlated preferences with seedling monoterpene levels. We also investigated the feeding responses of A. ater to artificial diets laced with two monoterpenes, Delta(3)-carene and alpha-pinene. Logistic regression indicated that two factors were the best predictors of whether seedlings in the trial would be consumed. Individual slug variation (replicates) was the most significant factor in the model; however, alpha-pinene concentration (also representing beta-pinene, Delta(3)-carene and total monoterpenes due to multicollinearity) of needles was also a significant factor. While A. ater did not select seedlings on the basis of family, seedlings not eaten were significantly higher in levels of alpha-pinene compared to seedlings that were consumed. We also demonstrated significant genetic variation in alpha-pinene concentration of seedlings between different families of P. sylvestris. Nitrogen and three morphological seedling characteristics (stem length, needle length and stem diameter) also showed significant genetic variation between P. sylvestris families. Artificial diets laced with high (5 mg g(-1) dry matter) quantities of either Delta(3)-carene or alpha-pinene, were eaten significantly less than control diets with no added monoterpenes, supporting the results of the seedling feeding trial. This study demonstrates that A. ater selectively feed on P. sylvestris seedlings and that this selection is based, in part, on the monoterpene concentration of seedlings. These results, coupled with significant genetic variation in alpha-pinene concentration of seedlings and evidence that slug herbivory is detrimental to P. sylvestris fitness, are discussed as possible evidence for A. ater as a selective force on the evolution of defensive chemistry in P. sylvestris.

Behavioral strategies of mammal herbivores against plant secondary metabolites: the avoidance-tolerance continuum.

We review the evidence for behavioral avoidance of plant secondary metabolites (PSMs) and identify how, and the circumstances under which it occurs. Behavioral strategies of avoidance of PSM can only be fully understood in relation to the underlying physiological processes or constraints. There is considerable evidence that animals learn to avoid PSMs on the basis of negative postingestive effects. The extent to which this process determines foraging choices is limited by the ability of animals to experience the consequences of their behaviors and associate particular cues in foods with their specific effects in the body. The proposed learning mechanisms require that animals must at least "sample" plants that contain PSMs. They do not completely avoid PSMs, but there is evidence that they restrict their ingestion to within limits that they are physiologically able to tolerate, and that the amounts of PSM ingested result from a balance between toxicological considerations and the nutrient content of the plant material. These limits are influenced by the kinetics of PSM elimination, which underlies patterns of bite and patch selection from plant parts to landscapes. We suggest that altering spatial location of feeding (to alternative food patches or alternative foods within patches, including plant parts), and temporal distribution of feeding activity, by either cessation of feeding or by continuing to feed, but on alternative foods, can both lead to reduction of the intake and toxic effects of PSMs. We propose that the strategy of avoidance or reduction of intake of PSMs coevolved with the animal's ability to physiologically tolerate their ingestion, and that avoidance and tolerance are inversely related (the avoidance-tolerance continuum). The animals' propensity and ability to seek alternatives also vary with the dispersion of their food resources. Further work is required to test these proposals and integrate temporal and spatial aspects of foraging behavior and its nutritional consequences in relation to PSMs.

The role of plant secondary metabolites in mammalian herbivory: ecological perspectives.

Plant secondary metabolites (PSM) have many ecological functions, but have long been considered as defences against pathogens or herbivores (vertebrate or invertebrate), reducing the likelihood and extent of attack. However, mammalian herbivores ingest many foods containing PSM and use both behavioural methods and physiological strategies to limit their negative effects. Most physiological counter-adaptations are inducible in response to ingested PSM, providing efficient protection against toxic effects. Possible positive effects of PSM include antioxidant and anthelminthic properties and complex formation between protein and condensed tannins that protects dietary protein from degradation by the symbiotic microflora of foregut fermenters, increasing its utilisation by the animal. This protein effect is probably only beneficial to animals under a narrow range of nutrient-rich conditions found mainly in agricultural systems. There are many examples of PSM causing food avoidance or reducing food intake, but there is as yet relatively little evidence for positive selection of them by herbivores. Although the feedback mechanisms relating the post-ingestive consequences of PSM to subsequent foraging behaviour are beginning to be understood, knowledge of the integration of behavioural and physiological strategies for regulating the effects of PSM is relatively poor. The opportunities for learned avoidance of PSM may be restricted in animals with complex diets that cannot associate a particular feedback signal with a given food type. A greater emphasis on the study of subclinical effects of PSM rather than acute effects, on pharmaco-kinetic studies in relation to behavioural studies and on the use of realistic experimental models is advocated.