Matti Rousi

Chemical diversity of several Betulaceae species: comparison of phenolics and terpenoids in northern birch stems

Abstract Phenolics and terpenoids characteristic for silver birch (Betula pendula Roth) were screened in current-growth twigs of seedlings (1-growing-season-old individuals) and current-growth twigs of saplings (3 to 5-year-old individuals) of several birch species. Seedlings were grown in a greenhouse for 1 growing season. Saplings were field cultivated or wild-stand individuals. Chemicals were methanol-extracted, purified and HPLC-diode array detector-, gas chromatograph-flame ionizaton detector- and gas chromatograph-mass selective-analyzed. Species-specific qualitative and quantitative variation of secondary chemicals in birch stems were considerable and dependent on the age of the individual. The chemical diversity and amount were lower in seedlings than in saplings. In saplings, (+)-catechin and its derivatives were found in nearly all species, while arylbutanoids (±rhododendrol glucosides), an arylheptanoid (platyphylloside) and dammaraneterpenoids (papyriferic acid, deacetylpapyriferic acid and pendulic acid) showed a more restricted distribution. The chemicals analyzed in birch stems are a useful tool for recognition of species.

Variation in Phenolic Compounds within a Birch (Betula pendula) Population

In previous studies, the qualitative and quantitative variation found in defense chemistry among birch populations and even among individual clones has been considerable. However, information about variation among adult, naturally regenerated birch trees from natural populations is still lacking. In this study, the phenolic composition of leaves of 30 naturally regenerated 20-year-old birch (Betula pendula) trees was analyzed for two successive years in order to characterize the chemical composition of individual trees, analyze the annual variation, and determine chemical similarities among individual trees within a population. The main phenolic compounds were flavonoid glycosides, myricetin, and quercetin derivatives. Annual variation in concentration among leaves was large. In most trees, concentrations were markedly higher in 1998 than in 1997; for certain compounds, the detected increase was as much as a 50%. However, for some individual trees, there were no differences between years in chemical quantity. Thus, when selection or grouping of trees is based on secondary chemistry, quantitative variation should be considered carefully. With the qualitative UPGMA method of classification, four chemotypes were found. The grouping was similar for both years, and qualitatively the results of an individual tree seem to be independent of sampling year. The stability in chemical profile of individual trees suggests that quality is tightly controlled by genotype, which provides a recognition tool for chemotaxonomy. The high within-population variation found in leaf defense chemistry may provide protection against different types of insects (generalists or specialists) and, thus, have positive effects on population survival.

Induced Accumulation of Foliage Phenols in Mountain Birch: Branch Response to Defoliation?

Plant phenols are frequently considered unspecific defensive substances that protect plants against herbivores (Feeny 1970, 1976). Although the lack of specificity of phenolic compounds (Zucker 1983) and their protein-precipitation efficiency in vivo (Bernays 1981) have been questioned, ecological evidence suggests that phenolic compounds can function as a potential chemical defense against some folivorous insects. These compounds have adverse effects on lepidopteran larvae (Feeny 1968; Lincoln et al. 1982), and they are abundant in tissues most susceptible to attacks by herbivores (McKey 1979). Phenolic compounds can also be induced to accumulate in foliage in response to leaf damage and defoliation (Levin 1971; Schultz and Baldwin 1982; Wagner and Evans 1985; Bergelson et al. 1986; Faeth 1986). In mouintain birch, Betula pubescens ssp. tortuosa (Ledeb.) Nyman, phenols have been observed to accumulate in current foliage shortly after adjacent leaves have sustained damage (Niemela et al. 1979) and as long as 3-4 yr after the entire tree had been defoliated (Tuomi et al. 1984). Both kinds of inducible responses in mountain birch foliage are also associated with retarded larval growth of the natural defoliator Epirrita autumnata (Haukioja and Niemela 1977, 1979; Haukioja 1980, 1982); foliage phenols are therefore at least potentially defensive characteristics of birch trees (Haukioja et al. 1985a). A tacit assumption in plant-herbivore studies seems to be that the presence of phenols and other potentially defensive secondary compounds in plant tissues is an adaptation for maximizing the fitness of whole individual plants (Janzen 1974; Rhoades and Cates 1976; Haukioja and Niemela 1976; Rhoades 1979). Since individual plants should therefore be the actively responding units, the cues indicating an increased risk of herbivory should trigger an induced production of secondary substances in the entire plant (Rhoades 1979; Haukioja and Neuvonen 1985). In this study, we tested whether phenolic accumulation can be induced separately in individual branches of mountain birch. The study was carried out at the Kevo Subarctic Research Institute of the University of Turku in northern Lapland. We examined separate branches on 10 birch trees (2.5-3.5 m in height) growing on the same hill, Puksalskaidi. Five branches per tree were manually defoliated after leaf flush (June 20, 1985) and the other five branches later toward the end of the growing season (August 17, 1985). Foliage phenols (Folin-Denis method) and total nitrogen (H-N-C analyzer) were determined for samples of mature leaves collected (July 26, 1986) from the recovered, previously defoliated branches and untreated control branches. We made a pairwise comparison of branches of the same trees, and the mean con-

A Mechanism of Resistance to Hare Browsing in Winter-Dormant European White Birch (Betula pendula)

The palatability of the birch Betula pendula to the snowshoe hare Lepus timidus differed considerably depending on geographical origin and family. The seedlings of Betula platyphylla were more resistant than any tested seed lot of B. pendula. The preference of hares for birch seed lots and individual seedlings was strongly and negatively correlated with the number of resin droplets on the surface of the bark. There were no indications of a trade-off between growth and defense; on the contrary, the tallest seedlings were also the most resistant. Nor did our material provide any support for the hypothesis of a carbon-nutrient balance; we found no significant decrease in resistance after fertilization of seedlings.

Effects of Shading and Fertilization on Resistance of Winter-Dormant Birch (Betula Pendula) to Voles and Hares

Seedlings representing four families of European white birch (Betulapendula) were grown through their first growing season in two fertilization and three shading treat- ments. The growth responses of seedlings were measured, and after complete winter hard- ening the basal parts of seedlings were offered to voles (Microtus agrestis) and the top parts of the same seedlings were fed to hares (Lepus timidus). The growth response of 1-yr-old birch seedlings to fertilization and shading was family specific. Nitrogen fertilization gen- erally increased the biomass accretion and the palatability of the seedlings to voles. Fer- tilization significantly decreased resistance of seedlings to hares in only one out of the four birch families. Shading decreased the biomass accretion of seedlings and increased their palatability to hares but tended to decrease the palatability to voles. The differing prefer- ences of the two mammalian herbivores for seedlings grown under different environmental conditions may be due to vertical differences in the defense of birch seedlings to herbivore feeding. No trade-off was found between seedling defense and growth.

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.

Taxonomic implications of phenolic variation in leaves of birch (Betula L.) species

Abstract The phenolic composition of the seedling leaves of 10 birch species was compared. The results fit rather well to the infrageneric classification proposed for birches, except for B. schmidtii , which exhibited a flavonoid composition similar to white birches. Among white birches, differences in phenolic profiles were small, except for B. pubescens , which contained several flavonoids not detected in other species. Qualitative chemotypes were distinguished from five species, and in white birches they were mostly based on presence or absence of the same compounds. The results support the view that the diploid white birches form a continuum of closely related taxa.

Chemical determinants of resistance in winter-dormant seedlings of European white birch (Betula pendula) to browsing by the mountain hare

SummaryA sample of one-year-old seedlings of European white birch (Betula pendula) was analyzed to determine the content of sugars, phenolics and terpenoid compounds. Two vertical segments of each seedling were analyzed separately. The number of resin droplets, which correlates strongly and negatively with feeding by the mountain hare, was also counted on the bark of experimental seedlings. The variation in the palatability of birch seedlings to mountain hare was determined primarily by the most abundant terpenoid, papyriferic acid. On the other hand, the resistance to hare feeding at the seedling bases, which are frequently attacked by voles, appeared to be dependent on other components, apparently phenolic substances. Sugars did not affect the resistance of the tested seedlings. The variation among experimental seedlings was much greater for secondary substances, especially terpenoid compounds, than for sugars. It is suggested that this high variation in protective compounds may be an adaptive trait selected for by the feeding of generalist herbivores.

Elevation of night-time temperature increases terpenoid emissions from Betula pendula and Populus tremula

Volatile organic compounds (VOCs) are expected to have an important role in plant adaptation to high temperatures. The impacts of increasing night-time temperature on daytime terpenoid emissions and related gene expression in silver birch (Betula pendula) and European aspen (Populus tremula) clones were studied. The plants were grown under five different night-time temperatures (6, 10, 14, 18, and 22 °C) while daytime temperature was kept at a constant 22 °C. VOC emissions were collected during the daytime and analysed by gas chromatography–mass spectrometry (GC-MS). In birch, emissions per leaf area of the C11 homoterpene 4,8-dimethy1-nona-1,3,7-triene (DMNT) and several sesquiterpenes were consistently increased with increasing night-time temperature. Total sesquiterpene (SQT) emissions showed an increase at higher temperatures. In aspen, emissions of DMNT and β-ocimene increased from 6 °C to 14 °C, while several other monoterpenes and the SQTs (Z,E)-α-farnesene and (E,E)-α-farnesene increased up to 18 °C. Total monoterpene and sesquiterpene emission peaked at 18 °C, whereas isoprene emissions decreased at 22 °C. Leaf area increased across the temperature range of 6–22 °C by 32% in birch and by 59% in aspen. Specific leaf area (SLA) was also increased in both species. The genetic regulation of VOC emissions seems to be very complex, as indicated by several inverse relationships between emission profiles and expression of several regulatory genes (DXR, DXS, and IPP). The study indicates that increasing night temperature may strongly affect the quantity and quality of daytime VOC emissions of northern deciduous trees.

Fungi, including Ophiostoma karelicum sp. nov., associated with Scolytus ratzeburgi infesting birch in Finland and Russia.

Several elm-infesting bark beetles belonging to the genus Scolytus (Coleoptera: Scolytinae) are vectors of Ophiostoma spp., most notably the Dutch elm disease fungi. A related bark beetle species, Scolytus ratzeburgi, is known to infest birch in various parts of Europe, but it is unknown whether fungi are associated with this beetle. The aim of this study was to identify several fungal species isolated from S. ratzeburgi. Beetles and their galleries were collected from Betula pendula at three different sites in the boreal forests of the Karelia region, on both the Finnish and Russian sides of the border. Three ophiostomatoid fungi were isolated from the beetles and their galleries. One Penicillium and one Bionectria species were isolated only from the Finnish material and, based on DNA sequences, were identified as P. brevicompactum and a species close to the anamorph of B. zelandianovae. Two Ophiostoma species present in low numbers included O. quercus and a species closely related to O. catonianum. Only one Ophiostoma species was isolated consistently from all the galleries and beetles considered in the study. Comparison of DNA sequences and morphological characterization showed that this fungus represents an undescribed taxon, described here as O. karelicum sp. nov.