Deciduous sapling responses to season and large herbivores in a semi‐arid African savanna

Impacts of large herbivores (>5 kg) on woody plants in African savannas are potentially most severe among plants shorter than 1.6 m. It is well established that severe browsing leads to longer shoots, yet prevents saplings from recruiting into adult size‐classes in African savannas. Increased shoot length, indicating faster shoot growth, is often associated with reduced concentrations of tannins and increased nutrient concentrations, suggesting carbon limitation. We hypothesized that, on average, large herbivores suppress stem height or circumference, but increase shoot length. We also hypothesized that if there were concomitant positive effects on nutrients, or negative effects on tannin concentrations, they would be greatest early in the wet season. We sampled saplings of four deciduous woody species (Acacia grandicornuta, Dichrostachys cinerea, Combretum apiculatum and Grewia flavescens) at different stages of the wet season in a large‐scale, long‐term herbivore exclusion experiment in Kruger National Park, South Africa. Plant height, shoot length and stem circumference were generally not adversely affected by large herbivores, suggesting C limitation is rarely present among deciduous saplings in semi‐arid African savannas, allowing them to tolerate browsing. Time since first rainfall emerged as a predominant factor consistently affecting nutrient and tannin concentrations, rather than large herbivores. Nitrogen and phosphorus generally decreased (by 20–50%), while condensed tannin concentration increased (150–350%) during the wet season, except for one species. We postulate that A. grandicornuta is less prone than other species to accumulating tannins during the wet season because of high investment of C in spines. Although nutrient and tannin concentrations were generally not affected by large herbivores, species‐specific responses were evident very early in the wet season, which is when herbivore populations are most likely to be affected by differential forage quality among plants.     

Chemical defense production in Lotus corniculatus L. I. The effects of nitrogen source on growth,reproduction and defense

Summary The carbon to nitrogen balance theory was examined for a legume, Lotus corniculatus L., which allocates carbon to nitrogen fixation. N-fixation can influence the ratio of carbon to nitrogen in legumes by providing nitrogen in nutrient-poor habitats, and by consuming carbon for support of symbiotic N-fixation. L. corniculatus clones (genotypes) were grown under two levels of nitrogen fertilization: a treatment which suppressed nodulation with fertilization and a treatment which received no additional fertilization. These plants relied solely on symbiotic N-fixation. Plants which supported symbionts had lower biomass and lower tannin concentrations than fertilized plants; this appears to be a result of the large carbon demand on N-fixation. Plants supporting symbionts often had relatively lower protein concentrations than fertilized plants. Cyanide concentration was influenced by plant genotype but not by nitrogen source. Although symbiotic N-fixing plants were smaller, they had three times the reproductive output of fertilized plants.     

The effect of nutrient supply and light intensity on tannins and mycorrhizal colonisation in Dutch heathland ecosystems

(1) Increased atmospheric nitrogen deposition has shifted plant dominance from ericaceous plants to grass species. To elucidate the reduced competitiveness of heather, we tested the hypothesis that additions of nitrogen reduce the concentrations of phenolics and condensed tannins in ericaceous leaves and retard mycorrhizal colonisation in ericaceous plants. We also tested the negative effects of reduced light intensity on carbon-based secondary compounds and mycorrhizal colonisation in ericaceous plants. (2) We performed a field inventory at three heathland sites in the Netherlands varying in nutrient supply and light intensity. Leaves of ericaceous plants and grasses were collected and analysed for concentrations of tannins, phenolics and nutrients. Similarly, we took root samples to record mycorrhizal colonisation and soil samples to measure the soil mineralisation. In addition, we conducted two-factorial experiments with Calluna vulgaris plants, in which we varied fertiliser and shade levels under greenhouse and field conditions. (3) The field inventory revealed that nitrogen addition and shading both negatively affected the concentration of total phenolics. The total phenolics and condensed tannin concentrations were positively correlated (P < 0.001), but in the field experiment, the condensed tannins were not significantly affected by the treatments. Our results provide the first evidence that the carbon nutrient balance can be used to predict the amount of total phenolics in the dwarf shrub C. vulgaris. (4) In the field experiments, shading of plants resulted in significantly less mycorrhizal colonisation. Only in the greenhouse experiment did addition of nitrogen negatively affect mycorrhizal colonisation. (5) Our results imply that increased atmospheric nitrogen deposition can depress the tannin concentrations in ericaceous plants and the mycorrhizal colonisation in roots, thereby reducing the plants’ competitiveness with respect to grasses. Additionally, if ericaceous plants are shaded by grasses that have become dominant due to increased nitrogen supply, these effects will be intensified and competitive replacement will be accelerated.     

Plant biomass partitioning and chemical defense: Response to defoliation and nitrate limitation

Summary Plant growth and allocation to root, shoot and carbon-based leaf chemical defense were measured in response to defoliation and nitrate limitation inHeterotheca subaxillaris. Field and greenhouse experiments demonstrated that, following defoliation, increased allocation to the shoot results in an equal root/shoot ratio between moderately defoliated (9% shoot mass removed) and non-defoliated plants. High defoliation (28% shoot mass or >25% leaf area removed) resulted in greater proportional shoot growth, reducing the root/shot ratio relative to moderate or non-defoliated plants. However, this latter effect was dependent on nutritional status. Despite the change in distribution of biomass, defoliation and nitrate limitation slowed the growth and development ofH. subaxillaris. Chronic defoliation decreased the growth of nitrate-rich plants more than that of nitrate-limited plants. The concentration of leaf mono- and sesqui-terpenes increased with nitrate-limitation and increasing defoliation. Nutrient stress resulting from reduced allocation to root growth with defoliation may explain the greater allocation to carbon-based leaf defenses, as well as the defoliation-related greater growth reduction of nitrate-rich plants.     

Effect of nitrogen fertilization upon the secondary chemistry and nutritional value of quaking aspen (Populus tremuloides Michx.) leaves for the large aspen tortrix (Choristoneura conflictana (Walker))

Summary We investigated the effects of nitrogen fertilization upon the concentrations of nitrogen, condensed tannin and phenolic glycosides of young quaking aspen (Populus tremuloides) leaves and the quality of these leaves as food for larvae of the large aspen tortrix (Choristoneura conflictana), a Lepidopteran that periodically defoliates quaking aspen growing in North America. Nitrogen fertilization resulted in decreased concentrations of condensed tannin and phenolic glycosides in aspen leaves and an increase in their nitrogen concentration and value as food for the large aspen tortrix. These results indicate that plant carbon/nutrient balance influences the quality of aspen leaves as food for the large aspen tortrix in two ways, by increasing the concentrations of positive factors (e.g. nitrogen) and decreasing the concentrations of negative factors (eg. carbon-based secondary metabolites) in leaves. Addition of purified aspen leaf condensed tannin and a methanol extract of young aspen leaves that contained condensed tannin and phenolic glycosides to artificial diets at high and low levels of dietary nitrogen supported this hypothesis. Increasing dietary nitrogen increased larval growth whereas increasing the concentrations of condensed tannin and phenolic glycosides decreased growth. Additionally, the methanol extract prevented pupation. These results indicate that future studies of woody plant/insect defoliator interactions must consider plant carbon/nutrient balance as a potentially important control over the nutritional value of foliage for insect herbivores.     

Growth and chemical defense in willow seedlings: trade-offs are transient

Many studies have failed to detect costs of defense and some have even found a positive correlation between growth and the concentrations of chemical defenses. These studies contradict the theoretical assumption that anti-herbivore defenses are costly—produced at the expense of growth and/or reproduction. Costs, however, may be transient and therefore difficult to detect. Here we tested the hypothesis that costs of defense would be pronounced early in development when root growth is prioritized (high percent root allocation), but not later in development. To test this hypothesis, we grew F₂ hybrid willow seedlings from five different families, and harvested cohorts of even-aged seedlings after 6, 7, 8 and 9 weeks of growth. Seedlings were divided into root and shoot tissue and shoots were analyzed for phenolics (condensed tannins and phenolic glycosides). We found evidence for transient costs of defense. The concentrations of phenolics were negatively correlated with total biomass, shoot biomass, and the proportion of biomass allocated to roots in week 6. After week 6, however, the concentrations of phenolics were positively correlated with shoot biomass and total biomass, while phenolics were uncorrelated with the proportion of biomass allocated to roots. These results, the first ever, to our knowledge, with woody plants, suggest that costs of defense were transient; specifically, costs were found in early development, when root establishment was a priority. Our findings suggest that studies should focus more on trade-offs early in plant development.     

Influences of elevated CO2 and pest damage on the allocation of plant defense compounds in Bt‐transgenic cotton and enzymatic activity of cotton aphid

Abstract Plant allocation to defensive compounds by elevated CO₂‐grown non‐transgenic and transgenic Bt cotton in response to infestation by cotton aphid, Aphis gossypii (Glover) in open‐top chambers under elevated CO₂ were studied. The results showed that significantly lower foliar nitrogen concentration and Bt toxin protein occurred in transgenic Bt cotton with and without cotton aphid infestation under elevated CO_2. However, significantly higher carbon/nitrogen ratio, condensed tannin and gossypol were observed in transgenic Bt cotton “GK‐12” and non‐transgenic Bt cotton ‘Simian‐3’ under elevated CO_2. The CO₂ level and cotton variety significantly influenced the foliar nitrogen, condensed tannin and gossypol concentrations in the plant leaves after feeding by A. gossypii. The interaction between CO₂ level × infestation time (24 h, 48 h and 72 h) showed a significant increase in cotton condensed tannin concentrations, while the interaction between CO₂ level × cotton variety significantly decreased the true choline esterase (TChE) concentration in the body of A. gossypi. This study exemplified the complexities of predicting how transgenic and non‐transgenic plants will allocate defensive compounds in response to herbivorous insects under differing climatic conditions. Plant defensive compound allocation patterns and aphid enzyme changes observed in this study appear to be broadly applicable across a range of plant and herbivorous insect interactions as CO₂ atmosphere rises.     

Tannin concentration enhances seed caching by scatter-hoarding rodents: An experiment using artificial ‘seeds’

Tannins are very common among plant seeds but their effects on the fate of seeds, for example, via mediation of the feeding preferences of scatter-hoarding rodents, are poorly understood. In this study, we created a series of artificial ‘seeds’ that only differed in tannin concentration and the type of tannin, and placed them in a pine forest in the Shangri-La Alpine Botanical Garden, Yunnan Province of China. Two rodent species (Apodemus latronum and A. chevrieri) showed significant preferences for ‘seeds’ with different tannin concentrations. A significantly higher proportion of seeds with low tannin concentration were consumed in situ compared with seeds with a higher tannin concentration. Meanwhile, the tannin concentration was significantly positively correlated with the proportion of seeds cached. The different types of tannin (hydrolysable tannin vs condensed tannin) did not differ significantly in their effect on the proportion of seeds eaten in situ vs seeds cached. Tannin concentrations had no significant effect on the distance that cached seeds were carried, which suggests that rodents may respond to different seed traits in deciding whether or not to cache seeds and how far they will transport seeds.     

Foliage acceptability to browsing ruminants in relation to seasonal changes in the leaf chemistry of woody plants in a South African savanna

Summary We investigated seasonal changes in food selection by hand-reared kudus and impalas in savanna vegetation in northern Transvaal, South Africa. The acceptability of the leaves of woody plants to these animals was compared with leaf concentrations of nutrients, fibre components and old leaf phenophases. No consistently significant correlation was found between acceptability and any single chemical factor. Based on an a priori palatability classification, discriminant function analysis separated relatively palatable species from unpalatable species in terms of a linear combination of protein and condensed tannin concentrations. The high acceptability of certain otherwise unpalatable species during the new leaf phenophase was related to elevation of protein levels relative to condensed tannin contents. Species were added to the diet during the dry season approximately in the order of their relative protein-condensed tannin difference.     

Variation in growth and ion accumulation between two selected populations of Trifolium repens L. differing in salt tolerance

Two divergent populations of T. repens cv. Haifa developed from two generations of recurrent selection for shoot chloride concentration, were grown in the greenhouse at 0 and 40 mol m^–3 NaCl. Over two harvest cycles at 40 mol m^–3 NaCl, the population selected for a low concentration of chloride in the shoot maintained a significantly lower chloride and sodium concentration compared with those plants selected for a high shoot chloride concentration. The distribution of chloride in the shoots was further examined in a subsample of plants from both populations. In all plants, concentrations of chloride were lower in the expanding and fully expanded leaves than in the older leaf tissue or petioles.While there were no significant differences in the photosynthetic rates between lines, shoot yields and relative leaf expansion rates were higher in the low chloride population. Plant death was greater in plants selected for high shoot chloride. These results suggest that selections based on measurements of low shoot chloride concentrations may be successful in developing a cultivar of T. repens with improved salt tolerance.     

Evidence for the role and fate of water-insoluble condensed tannins in the short-term reduction of carbon loss during litter decay

Warmer temperatures associated with climate change have the potential to accelerate litter decay and subsequently release large amounts of carbon stored in soils. Condensed tannins are widespread secondary metabolites, which accumulate to high concentrations in many woody plants and play key roles in forest soil nutrient cycles. Future elevated atmospheric CO₂ concentrations are predicted to reduce nitrogen content and increase tannin concentrations in plant tissues, thus reducing litter quality for microbial communities and slowing decomposition rates. How the distinct condensed tannin fractions (water-soluble, acetone:MeOH-soluble and solvent-insoluble) impact soil processes, has not been investigated. We tested the impact of condensed tannin and nitrogen concentrations on decay rates of poplar and Douglas-fir litter at sites spanning temperature and moisture gradients in coastal rainshadow forests in British Columbia, Canada. The three condensed tannin fractions were quantified using recent improvements on the butanol-HCl assay. Decay was assessed based on carbon remaining, while changes in litter chemistry were primarily observed using two methods for proximate chemical analyses. After 0.6 and 1 year of decay, more carbon remained in poplar litter with high, compared to low, condensed tannin concentrations. By contrast, more carbon remained in Douglas fir litter than poplar litter during this period, despite lower condensed tannin concentrations. Rapid early decay was especially attributed to loss of soluble compounds, including water-soluble condensed tannins. Water-insoluble condensed tannin fractions, which were transformed to acid-unhydrolyzable residues over time, were associated with reduced carbon loss in high condensed tannin litter.     

13C NMR study of pine needle decomposition

The quality of substrates in plantation forest litter, and their chemistry, can influence decomposition and N cycling. We studied the decomposition of Pinus radiata D. Don needles suspended on branches in windrows, for 3 yr after clear-cutting, using improved solid-state ¹³C NMR and chemical analysis. The NMR spectra suggested that the concentration of condensed tannins was 12–22%, and showed they were chemically altered during the period 4–12 months after clear-cutting. The spectra showed no evidence for further chemical modification of the tannins during the second or third years. Data for P. radiata needle decomposition in New Zealand indicated rapid loss of mass in the first 3 months, and condensed tannins did not appear to prevent mineralization of C or N. The tannin and lignin concentrations increased with decomposition of the needles, which was consistent with the early mineralization of readily available C compounds.     

Inducing condensed tannin production in Colophospermum mopane: Absence of response to soil N and P fertility and physical damage

The effects of available soil N and P and the effect of simulated browsing (leave removal) on foliar condensed tannin (CT) concentration were tested on young Colophospermum mopane (J. Kirk ex Benth) J. Leonard (Mopane) plants. Although clear differences in growth occurred between different levels of soil N, no differences in foliar CT concentration were found. Changes in available soil P and physical damage did not affect the plant growth or chemical composition. The complete absence of response of CT concentration to physical damage and soil nutrients may be related to the age of the trees in this study. It is proposed that existing theories on the interaction between soil properties and carbon based defences in trees are expanded, to include the potential responses of young trees to different soil nutrient levels.     

Chemical and mechanical defense against herbivory in two sympatric species of desert Acacia

Abstract. Most African Acacia trees occur in semi-arid savannas, but some species grow in deserts at the periphery of their range in northern Africa and the Middle East. We studied Acacia raddiana and A. tortilis in the southern Negev desert of Israel. According to the resource availability hypothesis, plants in poor habitats invest heavily into anti-herbivore defense because losses to browsers are costly when growth rates are low. Few plants invest simultaneously in different categories of defense. This suggests that trade-offs exist, although little is known about how individuals allocate resources to defence in natural populations. In a field experiment, we studied chemical and mechanical defences as a response to differential browsing for more than 10 yr. Both Acacia species increased spinescence but there was no clear response in the production of total polyphenols, condensed tannins, and protein-precipitating tannins (only A. raddiana may increase secondary compounds under very high browsing levels). There was an age effect in spinescence but not in secondary compounds for both species, with younger trees investing more into anti-herbivore defense than older individuals. We were unable to find negative correlations between traits of chemical and mechanical defense. We conclude that inducibility and age effects suggest a cost for mechanical but not necessarily for chemical defense. Contrary to the predictions of the resource availability hypothesis, concentrations of secondary compounds were not higher in this desert environment than in savannas.     

Phenolic compounds in Norway spruce as affected by boron nutrition at the end of the growing season

The increase in concentrations of phenolic compounds in boron (B) deficiency has been demonstrated in many herbaceous plant species, but information on woody plants is scarce. It has been suggested that accumulation of phenolic compounds plays a role in the development of cold hardiness in herbaceous plants but also that B deficiency decreases winter hardiness. Here we study the effects of B nutrition on phenolic compounds in Norway spruce (Picea abies L.) in the course of cold acclimation. Phenolic compounds were analysed in Norway spruce seedlings from three different B-fertilisation treatments in two harvests: non-acclimated and cold-acclimated seedlings. Norway spruce phenolic compounds consisted mainly of condensed tannins. During B deficiency, condensed tannins and monocoumaroyl–astragalin der. 1 increased in non-acclimated seedlings. The increase in tannins was 21%, which was nearly significant. However, the effect of B on phenolic compounds was almost absent in cold-acclimated seedlings. The condensed tannin concentration increased much more with time in the simulated autumn than due to B deficiency, and we conclude that the B effect was probably not large enough to be important for the hardening of the seedlings. The total phenolic concentrations more than doubled during the course of cold hardening suggesting that phenolics have a role in the winter hardiness in Norway spruce.     

Costs and benefits of defense by tannins in a neotropical tree

Summary The costs and benefits of defense by tannins were investigated for a neotropical tree, Cecropia peltata L. (Moraceae). Seedlings of equal age were grown under uniform conditions in a greenhouse for 18 months. Within a plant, leaf tannin concentrations measured in different years were highly correlated. Tannin concentrations differed substantially among individuals; plants with high tannin content had lower damage levels in herbivory experiments. The effects of tannin on herbivory appeared to be dosage dependent. There was, however, a cost associated with tannin production in terms of reduced leaf production.     

Condensed Tannin Formation by Agrobacterium rhizogenes Transformed Root and Shoot Organ Cultures of Lotus corniculatus

Root cultures of Lotus corniculatus L. cv. Leo transformed withAgrobacterium rhizogenes (C58Cl-pRi15834) grew rapidly in liquidmedium when cultured in the dark and produced large numbersof shoots when illuminated. The shoots, which could be regeneratedto produce fertile plants, were maintained in liquid mediumas shoot-organ cultures The accumulation and cellular distribution of condensed tanninswas determined during the growth of these root and shoot organcultures and in primary callus from non-transformed explants.Root and shoot cultures predominantly accumulated insolublepolymeric tannins which yielded both cyanidin and delphinidinon hydrolysis at ratios equivalent to control plants. Methanol-solublevanillin-positive compounds were isolated but no free oligomericproanthocyanidins, monomeric flavans or dihydroflavonols weredetected in these extracts. Condensed tannin accumulation waslinearly related to root growth and had a similar spatial distributionin ‘tannin’ cells in roots and leaves as comparedto control plants. Tannin-containing cells were absent frommeristematic cells of the root tip and root/shoot interface.Primary callus cultures failed to accumulate condensed tanninson media containing auxins, and exogenously supplied auxinswere found to inhibit tannin accumulation by transformed rootand shoot cultures Key words: Lotus corniculatus, Agrobacterium rhizogenes, hairy roots, condensed tannins, shoot and root cultures.     

Carbon allocation to defense, storage, and growth in seedlings of two temperate broad-leaved tree species

Optimal carbon allocation to growth, defense, or storage is a critical trait in determining the shade tolerance of tree species. Thus, examining interspecific differences in carbon allocation patterns is useful when evaluating niche partitioning in forest communities. We hypothesized that shade-tolerant species allocate more carbon to defense and storage and less to growth compared to shade-intolerant species. In gaps and forest understory, we measured relative growth rates (RGR), carbon-based defensive compounds (condensed tannin, total phenolics), and storage compounds (total non-structural carbohydrate; TNC) in seedlings of two tree species differing in shade tolerance. RGR was greater in the shade-intolerant species, Castanea crenata, than in the shade-tolerant species, Quercus mongolica var. grosseserrata, in gaps, but did not differ between the species in the forest understory. In contrast, concentrations of condensed tannin and total phenolics were greater in Quercus than in Castanea at both sites. TNC pool sizes did not differ between the species. Condensed tannin concentrations increased with increasing growth rate of structural biomass (GRstr) in Quercus but not in Castanea. TNC pool sizes increased with increasing GRstr in both species, but the rate of increase did not differ between the species. Accordingly, the amount of condensed tannin against TNC pool sizes was usually higher in Quercus than in Castanea. Hence, Quercus preferentially invested more carbon in defense than in storage. Such a large allocation of carbon to defense would be advantageous for a shade-tolerant species, allowing Quercus to persist in the forest understory where damage from herbivores and pathogens is costly. In contrast, the shade-intolerant Castanea preferentially invested more carbon in growth rather than defense (and similar amounts in storage as Quercus), ensuring establishment success in gaps, where severe competition occurs for light among neighboring plants. These contrasting carbon allocation patterns are closely associated with strategies for persistence in these species’ respective habitats.     

Mitigation of growth retardation effect of plant defense activator,acibenzolar-<Emphasis Type="Italic">S</Emphasis>-methyl,in amaranthus plants by plant growth-promoting rhizobacteria

Four rhizobacterial strains and acibenzolar-S-methyl (ASM), a chemical activator, which suppressed foliar blight of amaranthus (Amaranthus tricolor L.) caused by Rhizoctonia solani Kühn were evaluated for their effect on plant growth. The experiments were performed both under sterile and non-sterile soil conditions, in the presence or absence of the pathogen. In all cases, plants treated with ASM showed significant reduction in growth, as determined by shoot length, and shoot and root dry weight when compared to other treatments. The growth retardation effect of ASM was more profound with respect to shoot length. Reduction in shoot length was least when plants were treated with a combination of the chemical activator and Pseudomonas putida 89B61 under non-sterile soil conditions in the absence of the pathogen. Both under sterile and non-sterile soil conditions, in the presence of the pathogen, reduction in shoot length due to application of ASM was diminished significantly when plants were treated with rhizobacterial strain Pseudomonas fluorescens PN026R. Combined use of plant growth-promoting rhizobacteria (PGPR) and ASM was found to be beneficial as the growth retardation effect of the plant defense activator was reduced by the growth-promoting ability of the rhizobacteria.     

Genetic Manipulation of Condensed Tannins in Higher Plants : II. Analysis of Birdsfoot Trefoil Plants Harboring Antisense Dihydroflavonol Reductase Constructs

We have produced and analyzed transgenic birdsfoot trefoil (Lotus corniculatus L.) plants harboring antisense dihydroflavonol reductase (AS-DFR) sequences. In initial experiments the effect of introducing three different antisense Antirrhinum majus L. DFR constructs into a single recipient genotype (S50) was assessed. There were no obvious effects on plant biomass, but levels of condensed tannins showed a statistical reduction in leaf, stem, and root tissues of some of the antisense lines. Transformation events were also found, which resulted in increased levels of condensed tannins. In subsequent experiments a detailed study of AS-DFR phenotypes was carried out in genotype S33 using pMAJ2 (an antisense construct comprising the 5′ half of the A. majus cDNA). In this case, reduced tannin levels were found in leaf and stem tissues and in juvenile shoot tissues. Analysis of soluble flavonoids and isoflavonoids in tannin down-regulated shoot tissues indicated few obvious default products. When two S33 AS-DFR lines were outcrossed, there was an underrepresentation of transgene sequences in progeny plants and no examples of inheritance of an antisense phenotype were observed. To our knowledge, this is the first report of the genetic manipulation of condensed tannin biosynthesis in higher plants.