Elevated temperature and ozone modify structural characteristics of silver birch (Betula pendula) leaves.

To study the effects of slightly elevated temperature and ozone (O3) on leaf structural characteristics of silver birch (Betula pendula Roth), saplings of four clonal genotypes of this species were exposed to elevated temperature (ambient air temperature +0.8-1.0 °C) and elevated O3 (1.3-1.4× ambient O3), alone and in combination, in an open-air exposure field over two growing seasons (2007 and 2008). So far, the impacts of moderate elevation of temperature or the combination of elevated temperature and O3 on leaf structure of silver birch have not been intensively studied, thus showing the urgent need for this type of studies. Elevated temperature significantly increased leaf size, reduced non-glandular trichome density, decreased epidermis thickness and increased plastoglobuli size in birch leaves during one or both growing seasons. During the second growing season, O3 elevation reduced leaf size, increased palisade layer thickness and decreased the number of plastoglobuli in spongy cells. Certain leaf structural changes observed under a single treatment of elevated temperature or O3, such as increase in the amount of chloroplasts or vacuole, were no longer detected at the combined treatment. Leaf structural responses to O3 and rising temperature may also depend on timing of the exposure during the plant and leaf development as indicated by the distinct changes in leaf structure along the experiment. Genotype-dependent cellular responses to the treatments were detected particularly in the palisade cells. Overall, this study showed that even a slight but realistic elevation in ambient temperature can notably modify leaf structure of silver birch saplings. Leaf structure, in turn, influences leaf function, thus potentially affecting acclimation capacity under changing climate.

Plant volatile organic compounds (VOCs) in ozone (O3) polluted atmospheres: the ecological effects.

Tropospheric ozone (O3) is an important secondary air pollutant formed as a result of photochemical reactions between primary pollutants, such as nitrogen oxides (NOx), and volatile organic compounds (VOCs). O3 concentrations in the lower atmosphere (troposphere) are predicted to continue increasing as a result of anthropogenic activity, which will impact strongly on wild and cultivated plants. O3 affects photosynthesis and induces the development of visible foliar injuries, which are the result of genetically controlled programmed cell death. It also activates many plant defense responses, including the emission of phytogenic VOCs. Plant emitted VOCs play a role in many eco-physiological functions. Besides protecting the plant from abiotic stresses (high temperatures and oxidative stress) and biotic stressors (competing plants, micro- and macroorganisms), they drive multitrophic interactions between plants, herbivores and their natural enemies e.g., predators and parasitoids as well as interactions between plants (plant-to-plant communication). In addition, VOCs have an important role in atmospheric chemistry. They are O3 precursors, but at the same time are readily oxidized by O3, thus resulting in a series of new compounds that include secondary organic aerosols (SOAs). Here, we review the effects of O3 on plants and their VOC emissions. We also review the state of current knowledge on the effects of ozone on ecological interactions based on VOC signaling, and propose further research directions.

Emissions of volatile organic compounds and leaf structural characteristics of European aspen (Populus tremula) grown under elevated ozone and temperature.

Northern forest trees are challenged to adapt to changing climate, including global warming and increasing tropospheric ozone (O(3)) concentrations. Both elevated O(3) and temperature can cause significant changes in volatile organic compound (VOC) emissions as well as in leaf anatomy that can be related to adaptation or increased stress tolerance, or are signs of damage. Impacts of moderately elevated O(3) (1.3x ambient) and temperature (ambient + 1 degrees C), alone and in combination, on VOC emissions and leaf structure of two genotypes (2.2 and 5.2) of European aspen (Populus tremula L.) were studied in an open-field experiment in summer 2007. The impact of O(3) on measured variables was minor, but elevated temperature significantly increased emissions of total monoterpenes and green leaf volatiles. Genotypic differences in the responses to warming treatment were also observed. alpha-Pinene emission, which has been suggested to protect plants from elevated temperature, increased from genotype 5.2 only. Isoprene emission from genotype 2.2 decreased, whereas genotype 5.2 was able to retain high isoprene emission level also under elevated temperature. Elevated temperature also caused formation of thinner leaves, which was related to thinning of epidermis, palisade and spongy layers as well as reduced area of palisade cells. We consider aspen genotype 5.2 to have better potential for adaptation to increasing temperature because of thicker photosynthetic active palisade layer and higher isoprene and alpha-pinene emission levels compared to genotype 2.2. Our results show that even a moderate elevation in temperature is efficient enough to cause notable changes in VOC emissions and leaf structure of these aspen genotypes, possibly indicating the effort of the saplings to adapt to changing climate.

Elevated atmospheric ozone increases concentration of insecticidal Bacillus thuringiensis (Bt) Cry1Ac protein in Bt Brassica napus and reduces feeding of a Bt target herbivore on the non-transgenic parent.

Sustained cultivation of Bacillus thuringiensis (Bt) transgenic crops requires stable transgene expression under variable abiotic conditions. We studied the interactions of Bt toxin production and chronic ozone exposure in Bt cry1Ac-transgenic oilseed rape and found that the insect resistance trait is robust under ozone elevations. Bt Cry1Ac concentrations were higher in the leaves of Bt oilseed rape grown under elevated ozone compared to control treatment, measured either per leaf fresh weight or per total soluble protein of leaves. The mean relative growth rate of a Bt target herbivore, Plutella xylostella L. larvae was negative on Bt plants in all ozone treatments. On the non-transgenic plants, larval feeding damage was reduced under elevated ozone. Our results indicate the need for monitoring fluctuations in Bt toxin concentrations to reveal the potential of ozone exposure for altering dosing of Bt proteins to target and non-target herbivores in field environments experiencing increasing ozone pollution.

Effects of elevated carbon dioxide and ozone on volatile terpenoid emissions and multitrophic communication of transgenic insecticidal oilseed rape (Brassica napus).

Does transgenically incorporated insect resistance affect constitutive and herbivore-inducible terpenoid emissions and multitrophic communication under elevated atmospheric CO(2) or ozone (O(3))? This study aimed to clarify the possible interactions between allocation to direct defences (Bacillus thuringiensis (Bt) toxin production) and that to endogenous indirect defences under future climatic conditions. Terpenoid emissions were measured from vegetative-stage non-Bt and Bt Brassica napus grown in growth chambers under control or doubled CO(2), and control (filtered air) or 100 ppb O(3). The olfactometric orientation of Cotesia vestalis, an endoparasitoid of the herbivorous diamondback moth (Plutella xylostella), was assessed under the corresponding CO(2) and O(3) concentrations. The response of terpenoid emission to CO(2) or O(3) elevations was equivalent for Bt and non-Bt plants, but lower target herbivory reduced herbivore-inducible emissions from Bt plants. Elevated CO(2) increased emissions of most terpenoids, whereas O(3) reduced total terpenoid emissions. Cotesia vestalis orientated to host-damaged plants independent of plant type or CO(2) concentration. Under elevated O(3), host-damaged non-Bt plants attracted 75% of the parasitoids, but only 36.8% of parasitoids orientated to host-damaged Bt plants. Elevated O(3) has the potential to perturb specialized food-web communication in Bt crops.

The significance of ectomycorrhizas in chemical quality of silver birch foliage and above-ground insect herbivore performance.

We tested whether the ectomycorrhizal (ECM) infection level of roots of silver birch (Betula pendula) affects performance of above-ground insect herbivores by increasing available plant biomass, by enhancing availability of nutrients, or by modifying concentration of defense compounds, i.e., phenolics, in birch foliage. Insect performance was determined for a phloem-feeding generalist (Lygus rugulipennis, the European tarnished plant bug), a phloem-feeding specialist (Calaphis flava, the birch aphid), and a chewing generalist (Epirrita autumnata, the autumnal moth larva). Silver birch plantlets had either natural ECM infection level (on average 24% of short roots with ECM), reduced ECM infection level with fungicide (F-, 9% ECM), or enhanced ECM infection level after inoculation with the fungus Paxillus involutus (PI+, 45% ECM) or Leccinum versipelle (LV+, 42% ECM). In general, the most pronounced effect of ECM was observed on growth of plantlets, i.e., stem growth decreased. In PI+-treated plants, leaf biomass also decreased. The effect of mycorrhizal colonization on the host plant's nitrogen (N) and phosphorous (P) concentration was dependent on the mycorrhizal species and experiment. Fungicide treatment did not cause a consistent decrease in nutrients. Finally, defense of birch against herbivory, expressed as foliar phenolic concentration in plantlets, was not modified by ECM. However, E. autumnata had a significantly higher relative growth rate on PI+ plantlets with high leaf N concentration than on LV+ plantlets with low leaf N concentration. The birch aphid C. flava produced significantly less nymphs on birches with enhanced ECM infection levels (PI+ and LV+ plantlets) than on controls. In summary, our data show that the ECM infection level mainly affects the growth parameters of plantlets, whereas effects on leaf chemical quality are minor. Our data show that effects of ECM infection of birch roots on aboveground herbivores are multifaceted and depend on the fungal species forming ectomycorrhiza and also on the degree of specialization and feeding guild of insects.

Constitutive and herbivore-inducible glucosinolate concentrations in oilseed rape (Brassica napus) leaves are not affected by Bt Cry1Ac insertion but change under elevated atmospheric CO2 and O3.

Glucosinolates are plant secondary compounds involved in direct chemical defence by cruciferous plants against herbivores. The glucosinolate profile can be affected by abiotic and biotic environmental stimuli. We studied changes in glucosinolate patterns in leaves of non-transgenic oilseed rape (Brassica napus ssp. oleifera) under elevated atmospheric CO2 or ozone (O3) concentrations and compared them with those from transgenic for herbivore-resistance (Bacillus thuringiensis Cry1Ac endotoxin), to assess herbivory dynamics. Both elevated CO2 and O3 levels decreased indolic glucosinolate concentrations in transgenic and non-transgenic lines, whereas O3 specifically increased the concentration of an aromatic glucosinolate, 2-phenylethylglucosinolate. The herbivore-inducible indolic glucosinolate response was reduced in elevated O3 whereas elevated CO2 altered the induction dynamics of indolic and aliphatic glucosinolates. Herbivore-resistant Bt plants experienced minimal leaf damage after target herbivore Plutella xylostella feeding, but exhibited comparatively similar increase in glucosinolate concentrations after herbivory as non-transgenic plants, indicating that the endogenous glucosinolate defence was not severely compromised by transgenic modifications. The observed differences in constitutive and inducible glucosinolate concentrations of oilseed rape under elevated atmospheric CO2 and O3 might have implications for plant-herbivore interactions in Brassica crop-ecosystems in future climate scenarios.

Effect of long-term forest fertilization on Scots pine xylem quality and wood borer performance.

We tested whether changes in long-term nutrient availability would affect the xylem quality and characteristics of Scots pine trees as a food source for the larvae of the xylophagous wood borer Hylotrupes bajulus L. (Cerambycidae). We looked for an effect of host plant growth and xylem structural traits on H. bajulus larval performance, and looked for delayed effects of long-term forest fertilization on xylem chemical quality. In general, larval performance was dependent on larval developmental stage. However, the growth of larvae also varied with host plant quality (increases in the concentration of nitrogen and carbon-based secondary compounds of xylem were correlated with a decrease in the larval growth rate). The greater annual growth of trees reduced tracheid length and correlated positively with second-instar H. bajulus growth rate. This is consistent with the hypothesis that intrinsic growth patterns of host plants influence the development of the xylophagous wood borer H. bajulus.

The role of ozone-reactive compounds, terpenes, and green leaf volatiles (glvs), in the orientation of Cotesia plutellae.

The emission of inducible volatile organic compounds (VOCs), i.e., inducible terpenes, and green leaf volatiles (GLVs), is a common response of plants to herbivore attack. These VOCs are involved in the orientation of natural enemies, i.e., predators and parasitoids, toward their herbivore prey or hosts (indirect defense of plants). Terpenes and some GLVs are readily oxidized by ozone (O(3)), an important oxidant of the low atmosphere and predicted to increase as a result of anthropogenic activity. It has been recently reported that O(3) degradation of terpenes and GLVs does not affect signaling in two selected tritrophic systems. Natural enemies may have learned to use oxidation products that are more stable in nature to locate their prey. To understand the role of these compounds on the tritrophic system Brassica oleracea-Plutella xylostella-Cotesia plutellae, we assessed the preference of wasps to different combinations of cabbage VOCs (intact vs. herbivore-induced and herbivore-induced vs. herbivore-induced VOCs) in the presence or absence of O(3). We found that C. plutellae preferred P. xylostella-damaged plants at 0 and 120 nl l(-1) O(3) to intact plants at 0 nl l(-1) O(3). However, wasps preferred P. xylostella-damaged plants at 0 nl l(-1) to P. xylostella-damaged plants at 120 nl l(-1) O(3). The results suggest that compounds other than terpenes and GLVs are crucial for the orientation of the wasps, but terpenes and GLVs contribute to the behaviorally active VOC blend of herbivore-damaged cabbages by increasing their attraction to them. The products resulting from oxidation of terpenes and GLVs do not seem to play a role in the host location process as speculated previously.

Use of human urine fertilizer in cultivation of cabbage (Brassica oleracea)--impacts on chemical, microbial, and flavor quality.

Human urine was used as a fertilizer in cabbage cultivation and compared with industrial fertilizer and nonfertilizer treatments. Urine achieved equal fertilizer value to industrial fertilizer when both were used at a dose of 180 kg N/ha. Growth, biomass, and levels of chloride were slightly higher in urine-fertilized cabbage than with industrial-fertilized cabbage but clearly differed from nonfertilized. Insect damage was lower in urine-fertilized than in industrial-fertilized plots but more extensive than in nonfertilized plots. Microbiological quality of urine-fertilized cabbage and sauerkraut made from the cabbage was similar to that in the other fertilized cabbages. Furthermore, the level of glucosinolates and the taste of sauerkrauts were similar in cabbages from all three fertilization treatments. Our results show that human urine could be used as a fertilizer for cabbage and does not pose any significant hygienic threats or leave any distinctive flavor in food products.

Ozone degrades common herbivore-induced plant volatiles: does this affect herbivore prey location by predators and parasitoids?

Inducible terpenes and lipoxygenase pathway products, e.g., green-leaf volatiles (GLVs), are emitted by plants in response to herbivory. They are used by carnivorous arthropods to locate prey. These compounds are highly reactive with atmospheric pollutants. We hypothesized that elevated ozone (O(3)) may affect chemical communication between plants and natural enemies of herbivores by degrading signal compounds. In this study, we have used two tritrophic systems (Brassica oleracea-Plutella xylostella-Cotesia plutellae and Phaseolus lunatus-Tetranychus urticae-Phytoseiulus persimilis) to show that exposure of plants to moderately enhanced atmospheric O(3) levels (60 and 120 nl l(-1)) results in complete degradation of most herbivore-induced terpenes and GLVs, which is congruent with our hypothesis. However, orientation behavior of natural enemies was not disrupted by O(3) exposure in either tritrophic system. Other herbivore-induced volatiles, such as benzyl cyanide, a nitrile in cabbage, and methyl salicylate in lima bean, were not significantly reduced in reactions with O(3). We suggest that more atmospherically stable herbivore-induced volatile compounds can provide important long-distance plant-carnivore signals and may be used by natural enemies of herbivores to orientate in O(3)-polluted environments.

Emission of Plutella xylostella-induced compounds from cabbages grown at elevated CO2 and orientation behavior of the natural enemies.

Several plant species defend themselves indirectly from herbivores by producing herbivore-induced volatile compounds that attract the natural enemies of herbivores. Here we tested the effects of elevated atmospheric CO(2) (720 micromol mol(-1)) concentration on this indirect defense, physiological properties, and constitutive and induced emissions of white cabbage (Brassica oleracea ssp. capitata, cvs Lennox and Rinda). We monitored the orientation behavior of the generalist predator Podisus maculiventris (Heteroptera: Pentatomidae) and the specialist parasitoid Cotesia plutellae (Hymenoptera: Braconidae) to plants damaged by Plutella xylostella (Lepidoptera: Plutellidae) in the Y-tube olfactometer. Elevated CO(2) levels did not affect stomatal densities but reduced specific leaf area and increased leaf thickness in cv Lennox. In addition to enhanced constitutive monoterpene emission, P. xylostella-damaged cabbages emitted homoterpene (E)-4,8-dimethyl-1,3,7-nonatriene, sesquiterpene (E,E)-alpha-farnesene, and (Z)-3-hexenyl acetate. Growth at elevated CO(2) had no significant effect on the emissions expressed per leaf area, while minor reduction in the emission of homoterpene (E)-4,8-dimethyl-1,3,7-nonatriene and (E,E)-alpha-farnesene was observed at elevated CO(2) in one of two experiments. The generalist predator P. maculiventris discriminated only between the odors of intact and P. xylostella-damaged cv Rinda plants grown at ambient CO(2) concentration, preferring the odor of the damaged plants. The specialist parasitoid C. plutellae preferred the odor of damaged plants of both cultivars grown at ambient CO(2) but did not detect damaged cv Lennox plants grown at elevated CO(2). The results suggest that elevated atmospheric CO(2) concentration could weaken the plant response induced by insect herbivore feeding and thereby lead to a disturbance of signaling to the third trophic level.

Ozone exposure triggers the emission of herbivore-induced plant volatiles, but does not disturb tritrophic signalling.

We evaluated the similarities between ozone-induced and mite-induced emission of volatile organic compounds (VOCs) from lima beans, and tested the response of the natural enemies of herbivores to these emissions using trophic system of two-spotted spider mites and predatory mites. The acute ozone-exposure and spider mite-infestation induced the emission of two homoterpenes, (E)-4,8-dimethyl-1,3,7-nonatriene and (E,E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene, and (Z)-3-hexenyl acetate. Only plants with spider mite-infestation emitted the monoterpene (E)-beta-ocimene. Predatory mites were equally attracted to ozone-exposed and unexposed plants, but discriminated between spider mite-infested and uninfested plants, when both were exposed to ozone. The similarities between ozone and herbivore-induced VOCs suggest that plant defence against phytotoxic ozone and the production of VOCs for attraction of the natural enemies of herbivores may have adaptive coevolution. However, the expected elevated ozone concentrations in future may not disturb tritrophic signalling, unless herbivore-induced VOCs are lost in the process of aerosol formation.

Elevated atmospheric CO(2) affects the chemical quality of brassica plants and the growth rate of the specialist, Plutella xylostella, but not the generalist, Spodoptera littoralis.

Cabbage, Brassica oleracea subsp. capitata (cv. Lennox and Rinda), and oilseed rape, Brassica rapa subsp. oleifera (cv. Valo and Tuli), plants were grown under ambient CO(2) (360 ppm) or elevated CO(2) (720 ppm) at 23/18 degrees C and under a photoperiod of 22/2 h light (250 micromol m(-)(2) s(-)(1))/dark regime for up to 5 weeks. Afterward, the performance of the crucifer specialist Plutella xylostella (Lepidoptera: Plutellidae) and the generalist Spodoptera littoralis (Lepidoptera: Noctuidae) on those plants was studied. The mean relative growth rate (RGR) of P. xylostella larvae, feeding on both cultivars of oilseed rape or on the Lennox cultivar of cabbage leaves grown at an elevated CO(2) concentration, was significantly reduced as compared to ambient CO(2). A negative larval growth rate at elevated CO(2) was observed for P. xylostella on both oilseed rape cultivars, but the growth rate was reduced but positive on cabbage. Conversely, the RGR of S. littoralis on either plant species was not affected by CO(2) treatment but was lower on cabbage cv. Rinda than on cv. Lennox. The mortality of the larvae was not affected by CO(2) treatment either. At the same time, elevated CO(2) significantly decreased the concentrations of leaf phytochemical constituents in oilseed rape, i.e., total phenolics and total nitrogen, but not in cabbage. The effect of elevated CO(2) on the leaf glucosinolate concentrations of both plant species was marginal. In addition, the observed significant changes in individual glucosinolate concentrations of oilseed rape leaves were not consistent among cultivars. However, our results demonstrate for the first time quite strong effects of CO(2) enrichment on the larval performance of P. xylostella, which is an important pest of Brassica plants around the world. Further studies are still required to increase our understanding of why elevated CO(2) differently affects the performance of specialist and generalist insect herbivores on Brassica plants.

Chemical changes induced by methyl jasmonate in oilseed rape grown in the laboratory and in the field.

The effect of methyl jasmonate (MJ) spraying on the chemistry of Brassica plants was investigated. Glucosinolates (GLS) in the leaves, stems, and roots of laboratory-grown oilseed rape (Brassica rapa subsp. oleifera cv. Tuli and Valo) 3 and 7 days after MJ treatment were analyzed. Volatile organic compounds (VOCs) from whole oilseed rape plants were collected 3 days after MJ treatment. GLS were also analyzed from field-grown oilseed rape (cv. Valo) treated with MJ. The production of indolyl GLS in laboratory-grown oilseed rape, especially the concentration of 4-hydroxy-3-indolylmethyl (4-OH-glucobrassicin) in leaves, stems, and roots, 3-indolylmethyl (glucobrassicin) in stems, and 4-methoxy-3-indolylmethyl (4-methoxyglucobrassicin) in roots, was induced after MJ treatment. The VOC emission profile changed after MJ treatment, and homoterpene (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) was detected only in MJ-treated plants. The GLS concentration in the field-grown plants was significantly higher in MJ-treated plants than in control plants. These results suggest that spraying with MJ induces the production of secondary compounds, that is, GLS and VOCs, in Brassica plants. The induction of VOC emissions in oilseed rape is comparable to that caused by insect feeding damage. Thus, MJ-treated crop plants may become less palatable to insect herbivores and more attractive to natural enemies of herbivores.