Clone-specific responses in leaf phenolics of willows exposed to enhanced UVB radiation and drought stress

The effects of enhanced UVB radiation and drought stress on willow secondary phenolics were studied using the leaves of 8‐week‐old micropropagated plantlets from interspecific hybrids (Salix myrsinites L. ×S. myrsinifolia Salisb.) and pure species (S. myrsinifolia). The plantlets were subjected for 4 weeks to two levels of UVB radiation (ambient, enhanced) and two levels of watering (well‐watered, drought‐stressed) according to a 2 × 2 factorial design. Enhanced UVB radiation increased the total concentration of flavonoids and phenolic acids in all plantlets, while the total concentration of salicylates remained unaffected. Drought stress reduced the total concentration of salicylates and phenolic acids in S. myrsinifolia plantlets, while in hybrids only phenolic acids were affected. The response of phenolic acids to enhanced UVB in drought‐stressed plantlets was different from that in well‐watered ones, indicating that drought stress limited the accumulation of phenolic acids under enhanced UVB radiation. Flavonoids increased in response to enhanced UVB radiation in drought‐stressed plantlets, although drought caused serious physiological stress on growth. There were significant differences between hybrid and S. myrsinifolia plantlets with respect to the composition of phenolics and between families and clones with respect to their concentration. In addition, the response of salicylates, flavonoids and phenolic acids to enhanced UVB and drought stress was clone‐specific, which may indicate that climatic changes will alter the genetic composition of northern forests.     

THE EVOLUTION OF HOST-PLANT USE AND SEQUESTRATION IN THE LEAF BEETLE GENUS PHRATORA (COLEOPTERA: CHRYSOMELIDAE)

Leaf beetles in the genus Phratora differ in host plant use and in the chemical composition of their larval defensive secretion. Most species specialize on either poplars or willows (family Salicaceae), but two species feed on birch (family Betulaceae). Phratora vitellinae utilizes salicylates from the host plant to produce its larval secretion, which contains salicylaldehyde, while other Phratora species produce an autogenous secretion. To reconstruct the evolutionary history of host plant use and the larval secretion chemistry in this genus, we sequenced 1383 base pairs of the mt cytochrome oxidase I gene for six European and one North American Phratora species and three outgroup taxa. Bootstrap values of the complete nucleotide sequence were 99-100% for six of eight nodes in the maximum parsimony tree. They were 71% and 77% for the two other nodes. The maximum parsimony tree and the maximum likelihood tree based on nucleotide sequence showed the same relationships as a maximum parsimony tree based on the amino acid sequence. Beetle phylogeny overlapped broadly with host plant taxonomy and chemistry, and it revealed historical constraints influencing host plant use. However, there was one host shift from the willow family (Salicaceae) to the birch family (Betulaceae). The use of host plant phenol glycosides for the larval defensive secretion evolved along the lineage that led to P. vitellinae. Phratora vitellinae feeds on the taxonomically widest range of host plants, which are characterized by moderate to high levels of salicylates. The results support the hypothesis that the use of salicylates for the larval secretion evolved twice independently in chrysomeline leaf beetles.     

Influence of phenolglucosides and trichome density on the distribution of insects herbivores on willows

The effects of both trichome density and phenolglucoside content of leaves of 76 willow hybrids (Salix alba x fragilis) were measured to estimate their influence on the distribution of Phratora vitellinae (L.), Plagiodera versicolora Baly (Coleoptera: Chrysomelidae) and Pontania proxima (Lepeletier 1823) (Hymenoptera: Tenthredinidae) in a nursery at Gramont, Belgium.The willows showed differences in their phenolglucoside content and pilosity of leaves and are classified on these basis into four groups by a clustering method. Correlations and multiple regressions showed that these chemical and physical characteristics are good predictors of the abundance of insects. First, the abundance of larvae of Ph. vitellinae, adults of Pl. versicolora and galls of P. proxima is correlated positively with a high phenolglucoside content and a low pilosity of the leaves. Secondly, the distribution of adults of Ph. vitellinae and of larvae of Pl. versicolora is influenced by neither the chemical nor the physical leaves characteristics studied.     

Post hibernation dispersal of three leaf-eating beetles (Coleoptera: Chrysomelidae) colonising cultivated willows and poplars

1 We studied the spring dispersal of three common chrysomelids, from overwintering habitats into cultivated willow and poplar coppices at four sites in southern England over 2 years. 2 Adult Galerucella lineola, Phratora vulgatissima and P. vitellinae overwintered under the bark of mature trees or in other niches that simulated this habitat, within a few hundred metres of the coppice plantation. Relatively few beetles remained in the coppice fields during the winter. 3 Phratora vitellinae at the poplar sites emerged several weeks later than G. lineola and P. vulgatissima at the willow sites, reflecting the later leafing of poplar compared to willow. For all species, dispersal was by flight, with most activity during warm periods. Dispersal continued for several weeks for the willow- feeding species but was shorter for P. vitellinae. 4 All three species initially colonized the edge of the coppice field. Typically, 80% or more of the beetles colonising a plantation were within 8 m of the edge. Both Phratora spp. accumulated in the plantation edge zone for several weeks before leaving this area and colonising the crop interior. 5 The patterns of dispersal and colonization identified by this study may facilitate chrysomelid management practices in infested short rotation coppice that avoid the need for insecticide applications over the entire plantation. Keywords Chrysomelidae, Galerucella lineola, Phratora vitellinae, Phratora vulgatissima, poplar, short rotation coppice, willow, winter dispersal.     

The influence of phenolic compounds on the suitability of three Salix species as hosts for the willow beetle Phratora vulgatissima

The phenolic glycosides salicin and salicortin were found to influence larval growth and development rates and adult feeding preference of Phratora vulgatissima in laboratory feeding studies. Salicortin was more toxic to larvae than salicin, and none of the third instar larvae fed on Salix viminalis leaves amended with 1.52% (fresh mass) salicortin pupated. Condensed tannins (proanthocyanadins) did not affect larval performance. It was concluded that Salix burjatica resistance to willow beetle is due to the high levels of salicortin which occur in leaves of this species.     

Slight tissue wounding fails to induce consistent chemical defense in three willow (Salix spp.) clones

We studied the effects of natural wounding by insects and artificial wounding by clipping with scissors on the phenolic chemistry of two willows, Salix myrsinifolia and Salix pentandra. Half of the blade of a mature leaf was removed from each experimental plant either by allowing insects (chrysomellid beetles) to feed on the leaf or by clipping off half the blade of a leaf with scissors. We also examined the ability of wounded plants to warn neighboring plants of imminent wounding by an airborne signal by maintainign one set of control plants in the room containing the wounded plants and another set of control plants in a room hermetically sealed from the room containing the wounded plants. After 48 h, the experimental leaf and the fourth leaf and eighth leaf upwards in the leaf sequence from the experimental leaf were analyzed for phenols by high-pressure liquid chromatography. The same leaves in the leaf sequence from each control plant were similarly analyzed for phenols. Only one phenol, salicortin in leaves of S. myrsinifolia, increased in concentration in response to defoliation, and the observed response was small. The type of wounding affected this increase in salicortin, with natural wounding by insects causing a greater response than artificial wounding in one S. myrsinifolia clone, and artificial wounding causing a greater response than insect wounding in the other clone. This result indicates that S. myrsinifolia cannot control the effects of diffeeent types of wounding on its leaf secondary chemistry. We also found no indication of airborne warning signals between wounded and unwounded plants that trigger an elevation of leaf defenses in unwounded plants in anticipation of herbivore attack.     

Willow genotype, but not drought treatment, affects foliar phenolic concentrations and leaf-beetle resistance

In a greenhouse experiment we examined the effect of willow genotype and irrigation regime (moderate drought and well‐watered) on plant growth parameters, foliar nitrogen, and phenolic concentrations, as well as on the preference and performance of the blue leaf beetle, Phratora vulgatissima (L.) (Coleoptera: Chrysomelidae). The 10 vegetatively propagated willow genotypes in the experiments were F2 full‐sibling hybrids, originated from a cross between Salix viminalis (L.) (Salicaceae) (high in condensed tannins) and Salix dasyclados (L.) (Salicaceae) (rich in phenolic glycosides). Insect bioassays were conducted on detached leaves in Petri dishes as well as with free‐living insects on intact potted plants. The 10‐week long irrigation treatments caused statistically significant phenotypic differences in the potted willow saplings. Total biomass was somewhat higher in the well‐watered treatment. The root to total biomass ratio was higher in the drought‐treatment plants. There was significant genotypic variation in foliar nitrogen concentrations, and they were higher in the drought‐treatment plants. There was also a strong genotypic variation in each of the phenolic substances analyzed. Condensed tannins, which accounted for the greatest proportion of total phenolic mass, were higher in the well‐watered treatment. There was, however, no difference in levels of the other phenolics (salicylates, cinnamic acid, flavonoids, and chlorogenic acid) between irrigation treatments. The sum of these phenolics was higher in the well‐watered treatment. There was a strong variation in P. vulgatissima larval development on different willow genotypes, and larval performance was negatively correlated with levels of salicylates and cinnamic acid. There was, however, no effect of irrigation treatment on larval performance. Phratora vulgatissima preferred to feed on well‐watered plants, and we found a preference for oviposition there, but neither feeding nor oviposition site preference was affected by willow genotype. Adult feeding and oviposition preferences were not correlated with larval performance.     

Nitrate Supply and the Biophysics of Leaf Growth in Salix viminalis

The influence of nitrogen on leaf area development and the biophysicsof leaf growth was studied using clonal plants of the shrubwillow, Salix viminalis grown with either optimal (High N) orsub-optimal (Low N) supplies of nitrate. Leaf growth rate andfinal leaf size were reduced in the sub-optimal treatment andthe data suggest that in young rapidly growing leaves, thiswas primarily due to changes in cell wall properties, sincecell wall extensibility (% plasticity) was reduced in the LowN plants. The biophysical regulation of leaf cell expansion also differedwith nitrogen treatment as leaves aged. In the High N leaves,leaf cell turgor pressure (P) increased with age whilst in theLow N leaves P declined with age, again suggesting that foryoung leaves, cell wall plasticity limited expansion in theLow N plants. Measurements of cell wall properties showed thatcell wall elasticity (%E) was not influenced by nitrogen treatmentand remained constant regardless of leaf age. Key words: Salix, cell wall extensibility, nitrogen nutrition, biophysics of leaf growth     

Dynamics of Carbon Photosynthetically Assimilated in Nodulated Soya Bean Plants under Steady-state Conditions 2. The Incorporation of 13C into Carbohydrates, Organic Acids, Amino Acids and some Storage Compounds

Nodulated soya bean (Glycine max L.) plants at the early floweringstage were allowed to assimilate ¹³CO₂ under steady-state conditions,with a constant ¹³C abundance, for 8 h in the light. The plantswere either harvested immediately or 2 d after the end of the¹³CO₂ feeding, divided into young leaves (including flower buds),mature leaves, stems+petioles, roots and nodules; the ¹³C abundancein soluble carbohydrates, organic acids, amino acids, starchand poly-ß-hydroxybutyric acid was determined witha gas chromatography-mass spectrometry. The rapid turnover of ¹³C in the sucrose pools observed in allorgans of the plants showed that sucrose was the principal materialin the translocation stream of primary products of photosynthesis.At the end of the ¹³CO₂ exposure, sucrose in the mature leavesas the major source organs and in the stems+petioles was labelledwith currently assimilated carbon to about 75 per cent, whereasa much higher labelling of sucrose was found in the roots andin the nodules. This suggests the existence of two or more compartmentedpools of sucrose in mature leaves and also in stems+petioles. The relative labelling patterns of individual organic acidsand amino acids were similar in various plant organs. However,the rapid turnover of succinate and glycine was characteristicof nodules. Treatment with a high concentration of nitrate inthe nutrient media increased the turnover rate of amino acidcarbon in shoot organs and roots, while it markedly decreasedthe labelling of amino acids in nodules. The cyclitols, exceptfor D-pinitol, were significantly labelled with assimilated¹³C in mature leaves, but in nodules, the labelling was verymuch less. In the nodules, which were actively fixing atmospheric nitrogen,a large proportion (80–90 per cent) of currently assimilatedcarbon was found as sucrose and starch at the end of the ¹³CO₂feeding. This was also true of the roots. On the other hand,in young growing leaves, the distribution of currently assimilatedcarbon into sucrose, starch and other soluble compounds wasmuch less. This suggests that a large amount of carbon assimilatedby and translocated to young leaves was used to make up structuralmaterials, mainly protein and cell wall polymers synthesis,during the light period. Glycine max L., soya bean, ¹³CO₂ assimilation, carbon metabolism in nodules     

Metabolism of Inorganic Carbon Taken Up by Roots in Salix Plants

The metabolic products of inorganic carbon taken up throughthe roots from nutrient solution were studied in willow plants.Willow cuttings (Salix cv. Aquatica gigantea) were suppliedwith unlabelled or ¹⁴C-labelled NaHC0₃ for 1, 5, 10, and 24h in light or in darkness. After feeding, the plants were dividedinto six samples (upper and lower leaves and corresponding stems,cuttings and roots), which were frozen in liquid N₂. Freeze-driedground samples were extracted into water-soluble, chloroform-solubleand insoluble fractions. The water-soluble fraction was furtherseparated into basic, acidic, and neutral fractions by ion-exchangechromatography. In the light experiment pronase treatment wasused to separate the insoluble fraction into proteins and insolublecarbohydrates. After I h feeding time, most of the ¹⁴C was fixed into organicacids and amino acids both in light and in darkness in all partsof the plants. In the roots a large part of the ^l4C-carbon wasincorporated into the protein and insoluble fractions alreadyduring short feeding times, and the amounts incorporated increasedwith time. In the leaves, after 1 and 5 h the main labelledcompounds were the organic acids and amino acids, but after10 h about half of the total ¹⁴C was in protein and in the insolublefraction. A further analysis of amino acids and organic acidswith HPLC showed that C-4 acids were labelled initially andthat over time the proportion of different acids changed. These results indicate that the metabolism of carbon in rootsmight take place via ß-carboxylation of PEP. Partof the fixed ¹⁴C is transported from the roots, probably asamino acids and organic acids, to the shoot. In roots the C-4acids are metabolized further into structural compounds (proteinsand insoluble carbohydrates). Key words: DIC, Salix, roots, metabolism, HPLC     

Origin of Amino Acids in Datura stramonium Seeds

¹⁴C isotope studies show that the seeds of Datura stramoniumL. can produce a number of amino acids (particularly alanine,glutamate, phenylalanine, and tyrosine) from a supply of sucroseand nitrate. These amino acids can be incorporated into theseed protein. The bulk of the amino acids incorporated into the seed proteinmust, however, be supplied by adult leaves in the proximityof the fruit, either as the amino acids themselves, or theirimmediate precursors. The major free-amino-acid products of Datura leaves are theamides asparagine and glutamine.     

High herbivore pressure favors constitutive over induced defense

Theoretical and empirical studies show that, when past or current herbivory is a reliable cue of future attack and defenses are costly, defenses can be induced only when needed and thereby permit investment in other functions such as growth or reproduction. Theory also states that, in environments where herbivory is constantly high, constitutive defenses should be favored. Here, we present data to support the second aspect of the induced resistance hypothesis. We examined herbivore‐induced responses for four species of Inga (Fabaceae), a common canopy tree in Neotropical forests. We quantified chemical defenses of expanding leaves, including phenolic, saponin and toxic amino acids, in experimental field treatments with and without caterpillars. Because young leaves lack fiber and are higher in protein than mature leaves, they typically lose >25% of their leaf area during the few weeks of expansion. We predicted that the high rates of attack would select for investment in constitutive defenses over induction. Our data show that chemical defenses were quite unresponsive to herbivory. We demonstrated that expanding leaves showed no or only small increases in investment in secondary metabolites, and no qualitative changes in the phenolic compound profile in response to herbivory. The proteinogenic amino acid tyrosine, which can be toxic at high concentrations, showed the greatest levels of induction. Synthesis: These results provide some of the first support for theoretical predictions that the evolution of induced vs. constitutive defenses depends on the risk of herbivory. In habitats with constant and high potential losses to herbivores, such as tropical rainforests, high investments in constitutive defenses are favored over induction.     

Assimilation of gaseous ammonia and the transport of its products in barley and spinach leaves

The interactions between the assimilation and transport of nitrogenand carbon were investigated in barley and spinach leaves. Bothplants were fumigated with NH₃ (1 mg m^–3 and the contentof amino acids, sucrose and carbon intermediates of amino acidmetabolism were analysed in the leaves, apoplast and phloemsap. The following changes took place in the C- and N-metabolismof barley leaves during 5 h of fumigation with NH₃ (a) The contentsof amino acids, especially glutamine, largely increased andthe contents of sucrose, 2-oxoglutarate, phosphoenolpyruvate,and glycerate-3-phosphate declined. (b) A decrease in the phophoenolpyruvatecontent was accompanied by an increased activity of phosphoenolpyruvatecarboxylase. (c) The altered cytosolic concentrations of aminoacids and sucrose during NH₃ fumigation correlated with similarchanges in the apoplast and phloem sap. The altered percentageof each amino acid relative to the total amino acid concentrationin the cytosol, caused by NH₃ fumigation, is reflected in theapoplast and the phloem sap. The results indicate that the concentrations of amino acids in the cytosol determine their concentrationsin the phloem. Key words: Amino acids, ammonia fumigation, barley leaves, C: N partitioning, phosphoenolpyruvate carboxylase, phloem sap, spinach leaves     

Auxin-induced Conjugation Systems in Peas

Pretreatment of pea (Pisum sativum var. Alaska) sections with any active auxin induces an enzyme which forms aspartate conjugates of exogenously supplied indoleacetic acid, naphthaleneacetic acid, or benzoic acid. Whereas induction of this system is an absolutely auxin-specific process, another enzyme, which forms benzoylmalic acid, is induced both by auxins and by physiologically inactive aromatic carboxylic acids. Induction of both enzymes is abolished by low levels of RNA and protein synthesis inhibitors. The induction specificities and other characteristics of the two systems are compared.     

Intra- and interspecific effects of larval secretions in some chrysomelids (Coleoptera)

The larval secretions of the Chrysomelinae (Coleoptera, Chrysomelidae) are generally considered to be chemical defense substances against predators and parasites. The experiments presented in this paper provide evidence that the range of activity of these larval secretions also extends to interactions between different conspecific developmental stages and between competing phytophagous species. Four chrysomeline species were tested: Gastrophysa viridula De Geer, Phaedon cochleariae (F.), Plagiodera versicolora (Laich.), and Phratora vitellinae (L.). In the latter species, we did not test the intraspecific effect of the natural larval secretion, but the activity of its purchasable main component, salicylaldehyde (synthetic larval secretion). In G. viridula, Ph. cochleariae, and Ph. vitellinae the (synthetic) larval secretions act as oviposition deterrents against conspecific females. An investigation of the persistence of the oviposition-deterring activity of the larval secretion in G. viridula revealed that the deterrent effect is lost after 20 to 26 hours. In each of the four tested species the (synthetic) larval secretion deters feeding of conspecific adults. Since P. versicolora and Ph. vitellinae may occur on the same host plant (e.g. the willow Salix fragilis L.), the interspecific effect of the larval secretion between competing phytophagous species was examined in these two willow leaf beetles. The larval secretions of both species and salicylaldehyde, respectively, act as interspecific repellent and feeding deterrent against adults of the other species.

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Zusammenfassung Die Sekrete der Larven der Chrysomelinae (Fam. Chrysomelidae) werden im allgemeinen als chemische Abwehrstoffe gegen Prädatoren und Parasiten betrachtet. Die hier dargestellten Untersuchungen zeigen, daß die Wirkung der Larvensekrete nicht nur auf die Feindabwehr begrenzt ist. Das Wirkungsspektrum umfaßt darüber hinaus auch Interaktionen zwischen verschiedenen Entwicklungsstadien einer Art bzw. zwischen konkurrierenden phytophagen Arten. Folgende Chrysomelinenarten wurden untersucht: Gastrophysa viridula De Geer, Phaedon cochleariae (F.), Phratora vitellinae (L.) und Plagiodera versicolora (Laich.). Da das Larvensekret von Ph. vitellinae Salicylaldehyd als Hauptkomponente enthält, wurde hier-wenn nicht anders erwähnt-die Wirkung von synthetischem Salicylaldehyd getestet. Bei G. viridula, Ph. cochleariae und Ph. vitellinae wirkt das Larvensekret bzw. die authentische synthetische Substanz als hochwirksames Eiablage-Deterrens auf konspezifische Weibchen. Die eiablagehemmende Wirkung des Larvensekretes von P. versicolora wurde nicht getestet. Eine Untersuchung der Wirkungsdauer der eiblagehemmenden Aktivität des Larvensekretes von G. viridula ergab, daß die Hemmwirkung nach 20 bis 26 Stunden nicht mehr nachweisbar ist. Bei allen oben genannten vier Arten hemmt das. Larvensekret bzw. dessen authentische synthetische Substanz den Fraß von Adulten derselben Art. Diese fraßhemmende Aktivität ist bei G. viridula, Ph. cochleariae und Ph. vitellinae mindestens 18 Stunden wirksam. Das Larvensekret von P. versicolora zeigte bei 18-stündiger Versuchsdauer keine fraßhemmende Wirkung auf Adulte derselben Art; erst bei Reduktion der Versuchsdauer auf drei Stunden konnte hier eine signifikant fraßhemmende Wirkung nachgewiesen werden. P. versicolora und Ph. vitellinae können als Weidenblattkäfer an derselben Wirtspflanze auftreten. Deshalb wurde auch die interspezifische Wirkung der Larvensekrete dieser beiden Chrysomelinenarten geprüft. Das Larvensekret von Ph. vitellinae sowie auch dessen authentische synthetische Substanz (Salicylaldehyd) wirkt auf adulte P. versicolora als Repellent und Fraßhemmstoff. Ebenso wirkt umgekehrt das Larvensekret von P. versicolora abschreckend und fraßhemmend auf adulte Ph. vitellinae.

     

Effects of elevated CO2 and temperature on plant growth and herbivore defensive chemistry

Concentration of atmospheric CO₂ and temperature have both been rising for the last three decades. In this century, the temperature has been predicted to rise by 2–5 °C and the CO₂ concentration to double. These changes may affect the primary and secondary metabolism of plants and thus have implications for other trophic levels. However, the biotic interactions in changing climate conditions are poorly known. In this study, two questions were addressed: (i) How will climate change affect growth and the amounts of secondary compounds in flexible plant species? and (ii) How will this affect herbivores living on this species. Four clones of the dark‐leaved willow (Salix myrsinifolia (Salisb.)) seedlings were grown in closed‐top chambers with two controlled factors: concentration of atmospheric CO₂ and temperature (T). There were four combinations of these factors, each combination replicated four times (total of 16 chambers): (i) Control CO₂ (350 ppm) and control T, (ii) Elevated CO₂ (700 ppm) and control T, (iii) Control CO₂ and elevated T (2 °C), and (iv) Elevated CO₂ and elevated T. Stem growth and aerial biomass of the plants were determined; and the leaf phenolics, nitrogen and water concentrations were analysed. In addition the growth rate of larvae and feeding preference of adults of a specialist herbivore, the chrysomelid beetle Phratora vitellinae (L.), on the treated willow leaves were measured. Elevated temperature and CO₂ concentration increased the stem biomass and elevated CO₂ increased leaf biomass and total aerial biomass of the willows. Patterns of biomass allocation were different in different temperature treatments. At elevated temperature there was less branch and leaf material in relation to stems than at the control temperature. Moreover, patterns of biomass allocation differed among clones. CO₂ enhancement increased the specific leaf weight (SLW) and reduced both water and nitrogen content of the leaves, however, leaf area was unaffected by the treatments. Carbon dioxide (CO₂) and T enhancement reduced the concentrations of several phenolic compounds in the leaves. Phenolic compounds, nutrients, and water in the leaves might be diluted partly due to increased carbon allocation to different structures (e.g. thickening of cell wall and increase of trichomes, etc.). In some cases plant clones showed specific responses to treatments. The CO₂ enhancement reduced the relative growth rate (RGR) of the beetle larvae, and in contrast, temperature elevation increased it. Adult beetles did not clearly discriminate between willow leaves grown in different T and CO₂ environments, but tended to eat more leaf material from chambers with doubled CO₂ concentration. At elevated CO₂ adult beetles may need to eat more leaf material in order to reproduce, which may in turn prolong the life cycles, increasing the risk of being eaten and possibly affecting ability to overwinter successfully. Overall, climate change may significantly modify the dynamic interaction between willow and beetle populations.     

Amino Acid Content of Leaves in Mycorrhizal and Non-mycorrhizal Citrus Rootstocks

Nitrogen status was examined in leaves of sour orange and roughlemon citrus rootstocks grown in a low phosphorus sand inoculatedwith Glomus etunicatus, in sand amended with superphosphateat a rate of 2240 kg ha^–1, and in a sand control leftuntreated. Sour orange was 3.1- and 3.5-fold taller and roughlemon was 1.8- and 2.0-fold taller than the controls in theinoculated and phosphorus treatments, respectively. In the controls,leaf N was up to 2.5-fold higher than in the other treatments.Both total and free amino acids accumulated in leaves of bothrootstocks to higher levels in the control than in the othertreatments. Most total amino acids in the control were lowerthan in the treatments, with the exceptions of arginine (upto 12-fold increase), proline (up to 1.8-fold increase), lysine,and free ammonia. Twenty-two free amino acids, urea, and ammoniawere detected. Both rootstocks grown in control sand had significantincreases in citrulline, ornithine, lysine, histidine, arginineand ammonia. Levels of total and most free amino acids in theinoculated and phosphorus treatments were similar to one another.It is suggested that mineral deficiency caused by the absenceof G. etunicatus causes a reorganization of N-metabolism witha shift to a greater synthesis of ornithine cycle intermediates. Citrus aurantium L., Citrus limon (L.) Burm, amino acid content, citrus rootstocks, mycorrhiza, nitrogen metabolism     

Phenolic glucosides as feeding cues for willow-feeding leaf beetles

The effects of individual phenolic glucosides and total glucoside fractions on the feeding behaviour of three willowfeeding leaf beetles (Coleoptera: Chrysomelidae) were tested in the laboratory. Feeding preferences of the tested leaf beetles were strongly influenced by certain phenolic glucosides which are typical secondary compounds of willows (Salicaceae:Salix). Two of the tested leaf beetles,Galerucella lineola andLochmaea capreae showed rather similar responses to glucoside treatments. Both of them were strongly stimulated by total glucoside fractions fromSalix triandra and by its major glucoside salidroside. The third species,Phatora vitellinae, was attracted most by the fractions fromS. myrsinifolia andS. pentandra, and by two related salicylate glucosides, tremulacin and salicortin. Food selection pattern of the tested beetles in the laboratory concords fairly well with their distributions in the field and with the occurrence of phenolic glucosides in their host willows. Phenolic glucoside extracts stimulated more feeding than individual pure glucosides. This indicates that different compounds have synergistic effects in the feeding behaviour of leaf beetles. Our results clearly show that willow leaf beetles select their food based on phenolic glucosides of their host plants.     

Induction of phenylalanine ammonia-lyase activity by tryptophan in Ustilago maydis.

To understand the regulation of phenylalanine ammonia-lyase (PAL) activity in the corn smut fungus, Ustilago maydis, we examined the effects of different media, metabolic effectors (including aromatic amino acids), and environmental factors on induction and repression of PAL activity. PAL was detected only in cell extracts and not in the culture medium. U. maydis PAL is constitutively produced at a low level in all media tested but its regulation can be influenced by aromatic amino acids. L-Tryptophan (0.3 mM) induces PAL activity 3- to 5-fold but tryptophan analogs and tryptophan-related metabolites do not. The enzyme is most readily induced during the early stationary phase of growth and the induced activity remains relatively constant during stationary stage. No induction or inhibition of PAL activity was observed as a function of culture temperature, pH or light. PAL induction was repressed by glucose but not by its reaction product, t-cinnamic acid. Induction did not require de novo protein synthesis, suggesting that some form of post-translational protein modification or a metabolic effect may be involved. This study shows that the regulation of U. maydis PAL is very different from the patterns known for plants and other fungi.