Caroline Müller

Biology of the Plant Cuticle.

1. Introduction: Biology of the plant cuticle. Markus Riederer, Julius--von--Sachs--Institut fur Biowissenschaften, Universitat Wurzburg, Wurzburg, Germany. 2. The fine structure of the plant cuticle. Christopher E. Jeffree, Science Faculty Electron Microscope Facility, Edinburgh, UK. 3. The cutin biopolymer matrix. Ruth E. Shark and Shiying Tian, Department of Chemistry and Institute for Macromolecular Assemblies, City University of New York, College of Staten Island, 2800 Victory Boulevard, Staten Island, NY 10314--6600, USA. 4. Composition of plant cuticular waxes. Reinhard Jetter, Departments of Botany and Chemistry, University of British Columbia, Vancouver, Canada Ljerka Kunst and A. Lacey Samuels, Department of Botany, University of British Columbia, Vancouver, Canada. 5. Biosynthesis and transport of plant cuticular waxes. Ljerka Kunst, Department of Botany, University of British Columbia, Vancouver, Canada Dr Reinhard Jetter, Departments of Botany and Chemistry, University of British Columbia, Vancouver, Canada and A. Lacey Samuels, Department of Botany, University of British Columbia, Vancouver, Canada. 6. Optical properties of plant surfaces. Erhard E. Pfundel, Julius--von--Sachs--Institut fur Biowissenschaften, Universitat Wurzburg, Wurzburg, Germany Giovanni Agati, Istituto di Fisica Applicata, Firenze, Italy and Zoran G. Cerovic, LURE--CNRS, Orsay, France. 7. Transport of lipophilic non--electrolytes across the cuticle. Markus Riederer, Julius--von--Sachs--Institut fur Biowissenschaften, Universitat Wurzburg, Wurzburg, Germany and Adrian A. Friedmann, Syngenta Inc, Bracknell, Berkshire, UK. 8. Characterisation of polar paths of transport in plant cuticles. Lukas Schreiber, A-kophysiologie der Pflanzen, Botanisches Institut, Bonn, Germany. 9. Cuticular transpiration. Markus Burghardt and Markus Riederer, Julius--von--Sachs--Institut fur Biowissenschaften, Universitat Wurzburg, 082 Wurzburg, Germany. . 10. The cuticle and cellular interactions. Hirokazu Tanaka and Yasunori Machida, Division of Biological Science, Graduate School of Science, Nagoya University, Nagoya, Japan. 11. Microbial communities in the phyllosphere. Johan H. J. Leveau, Centre for Terrestrial Ecology, Heteren, The Netherlands. 12. Filamentous fungi on plant surfaces. Tim L. W. Carver, Plant Genetics and Breeding, IGER, Aberystwyth, UK and. Sarah J. Gurr, Plant Sciences, University of Oxford, Oxford, UK. 13. Plant--Insect interactions on cuticular surfaces. Caroline Muller, Julius--von--Sachs--Institut fur Biowissenschaften, Universitat Wurzburg, Wurzburg, Germany

Plant Surface Properties in Chemical Ecology

The surface of the primary aerial parts of terrestrial plants is covered by a cuticle, which has crucial autecological functions, but also serves as an important interface in trophic interactions. The chemical and physical properties of this layer contribute to these functions. The cuticle is composed of the cuticular layer and the cuticle proper, which is covered by epicuticular waxes. Whereas the cutin fraction is a polyester-type biopolymer composed of hydroxyl and hydroxyepoxy fatty acids, the cuticular waxes are a complex mixture of long-chain aliphatic and cyclic compounds. These highly lipophilic compounds determine the hydrophobic quality of the plant surface and, together with the microstructure of the waxes, vary in a species-specific manner. The physicochemical characteristics contribute to certain optical features, limit transpiration, and influence adhesion of particles and organisms. In chemical ecology, where interactions between organisms and the underlying (allelo-) chemical principles are studied, it is important to determine what is present at this interface between the plant and the environment. Several useful equations can allow estimation of the dissolution of a given organic molecule in the cuticle and its transport properties. The implementation of these equations is exemplified by examining glucosinolates, which play an important role in interactions of plants with other organisms. An accurate characterization of physicochemical properties of the plant surface is needed to understand its ecological significance. Here, we summarize current knowledge about the physical and chemical properties of plant cuticles and their role in interactions with microorganisms, phytophagous insects, and their antagonists.

Plant chemistry and insect sequestration

Most plant families are distinguished by characteristic secondary metabolites, which can function as putative defence against herbivores. However, many herbivorous insects of different orders can make use of these plant-synthesised compounds by ingesting and storing them in their body tissue or integument. Such sequestration of putatively unpalatable or toxic metabolites can enhance the insects’ own defence against enemies and may also be involved in reproductive behaviour. This review gives a comprehensive overview of all groups of secondary plant metabolites for which sequestration by insect herbivores belonging to different orders has been demonstrated. Sequestered compounds include various aromatic compounds, nitrogen-containing metabolites such as alkaloids, cyanogenic glycosides, glucosinolates and other sulphur-containing metabolites, and isoprenoids such as cardiac glycosides, cucurbitacins, iridoid glycosides and others. Sequestration of plant compounds has been investigated most in insects feeding or gathering on Apocynaceae s.l. (Apocynoideae, Asclepiaoideae), Aristolochiaceae, Asteraceae, Boraginaceae, Fabaceae and Plantaginaceae, but it also occurs for some gymnosperms and even lichens. In total, more than 250 insect species have been shown to sequester plant metabolites from at least 40 plant families. Sequestration predominates in the Coleoptera and Lepidoptera, but also occurs frequently in the orders Heteroptera, Hymenoptera, Orthoptera and Sternorrhyncha. Patterns of sequestration mechanisms for various compound classes and common or individual features occurring in different insect orders are highlighted. More research is needed to elucidate the specific transport mechanisms and the physiological processes of sequestration in various insect species.

Sequestration of Host Plant Glucosinolates in the Defensive Hemolymph of the Sawfly Athalia rosae

Interactions between insects and glucosinolate-containing plant species have been investigated for a long time. Although the glucosinolate–myrosinase system is believed to act as a defense mechanism against generalist herbivores and fungi, several specialist insects use these secondary metabolites for host plant finding and acceptance and can handle them physiologically. However, sequestration of glucosinolates in specialist herbivores has been less well studied. Larvae of the turnip sawfly Athalia rosae feed on several glucosinolate-containing plant species. When larvae are disturbed by antagonists, they release one or more small droplets of hemolymph from their integument. This “reflex bleeding” is used as a defense mechanism. Specific glucosinolate analysis, by conversion to desulfoglucosinolates and analysis of these by high-performance liquid chromatography coupled to diode array UV spectroscopy and mass spectrometry, revealed that larvae incorporate and concentrate the plants characteristic glucosinolates from their hosts. Extracts of larvae that were reared on Sinapis alba contained sinalbin, even when the larvae were first starved for 22 hr and, thus, had empty guts. Hemolymph was analyzed from larvae that were reared on either S. alba, Brassica nigra, or Barbarea stricta. Leaves were analyzed from the same plants the larvae had fed on. Sinalbin (from S. alba), sinigrin (B. nigra), or glucobarbarin and glucobrassicin (B. stricta) were present in leaves in concentrations less than 1 μmol/g fresh weight, while the same glucosinolates could be detected in the larvaes hemolymph in concentrations between 10 and 31 μmol/g fresh weight, except that glucobrassicin was present only as a trace. In larval feces, only trace amounts of glucosinolates (sinalbin and sinigrin) could be detected. The glucosinolates were likewise found in freshly emerged adults, showing that the sequestered phytochemicals were transferred through the pupal stage.

Choosing and using diversity indices: Insights for ecological applications from the German Biodiversity Exploratories

Biodiversity, a multidimensional property of natural systems, is difficult to quantify partly because of the multitude of indices proposed for this purpose. Indices aim to describe general properties of communities that allow us to compare different regions, taxa, and trophic levels. Therefore, they are of fundamental importance for environmental monitoring and conservation, although there is no consensus about which indices are more appropriate and informative. We tested several common diversity indices in a range of simple to complex statistical analyses in order to determine whether some were better suited for certain analyses than others. We used data collected around the focal plant Plantago lanceolata on 60 temperate grassland plots embedded in an agricultural landscape to explore relationships between the common diversity indices of species richness (S), Shannon’s diversity (H’), Simpson’s diversity (D1), Simpson’s dominance (D2), Simpson’s evenness (E), and Berger–Parker dominance (BP). We calculated each of these indices for herbaceous plants, arbuscular mycorrhizal fungi, aboveground arthropods, belowground insect larvae, and P. lanceolata molecular and chemical diversity. Including these trait-based measures of diversity allowed us to test whether or not they behaved similarly to the better studied species diversity. We used path analysis to determine whether compound indices detected more relationships between diversities of different organisms and traits than more basic indices. In the path models, more paths were significant when using H’, even though all models except that with E were equally reliable. This demonstrates that while common diversity indices may appear interchangeable in simple analyses, when considering complex interactions, the choice of index can profoundly alter the interpretation of results. Data mining in order to identify the index producing the most significant results should be avoided, but simultaneously considering analyses using multiple indices can provide greater insight into the interactions in a system.

Plant invasions, generalist herbivores, and novel defense weapons

One commonly accepted mechanism for biological invasions is that species, after introduction to a new region, leave behind their natural enemies and therefore increase in distribution and abundance. However, which enemies are escaped remains unclear. Escape from specialist invertebrate herbivores has been examined in detail, but despite the profound effects of generalist herbivores in natural communities their potential to control invasive species is poorly understood. We carried out parallel laboratory feeding bioassays with generalist invertebrate herbivores from the native (Europe) and from the introduced (North America) range using native and nonnative tetraploid populations of the invasive spotted knapweed, Centaurea stoebe. We found that the growth of North American generalist herbivores was far lower when feeding on C. stoebe than the growth of European generalists. In contrast, North American and European generalists grew equally well on European and North American tetraploid C. stoebe plants, lending no support for an evolutionary change in resistance of North American tetraploid C. stoebe populations against generalist herbivores. These results suggest that biogeographical differences in the response of generalist herbivores to novel plant species have the potential to affect plant invasions.

Testing Predictions of the ‘Evolution of Increased Competitive Ability’ Hypothesis for an Invasive Crucifer

The successful spread of invasive plants may result from an evolutionary shift in resource allocation from defence to growth due to release from enemies, as proposed by the ‘evolution of increased competitive ability’ hypothesis (EICA). The crucifer Lepidium draba was used to test this hypothesis, measuring growth and levels of glucosinolates and myrosinase of leaves as constitutive defence parameters. Individuals from 21 populations of the native (Europe) and the invasive range (North-America) were grown under common greenhouse conditions. According to the EICA hypothesis it was predicted that plants from the invasive range might show stronger growth and have lower levels of defence as a result of selection favouring such genotypes. There was significant variation between populations in shoot, root, total biomass, and number of ramets of 3-month-old plants but no difference due to origin from both continents. The main glucosinolate p-hydroxybenzyl glucosinolate was significantly higher in seedlings of the invasive range while myrosinase activity was higher in old plants of the invasive range. Therefore, the EICA hypothesis does not hold, however, alternatively there is evidence for selection favouring stronger defence in the invasive range. The binary defence system of this crucifer is discussed with respect to the degree of specialisation of potential herbivores.

Lack of sequestration of host plant glucosinolates in Pieris rapae and P. grarricae

Summary. Sequestration of plant toxins in herbivores is often correlated with aposematic coloration and gregarious behaviour. Larvae of Pieris brassicae show these conspicuous morphological and behavioural characteristics and were thus suggested to sequester glucosinolates that are characteristic secondary metabolites of their host plants. P. rapaeare camouflaged and solitary, and are thus not expected to sequester. To test this hypothesis and to check the repeatabi-lity of a study that did report the presence of the glucosinolate sinigrin in P. brassicae, larvae were reared on three species of Brassicaceae (Sinapis alba, Brassica nigra and Barbarea stricta), and different leaf and insect samples were taken for glucosinolate analysis. The major host plant glucosinolates could only be found in traces or not at all in larval haemolymph, bled or starved larvae, faeces or pupae of both species or P. brassicae regurgitant. Haemolymph of both Pieris spp. was not rejected by the ant Myrmica rubra in dual-choice assays; the regurgitant of P. brassicae was rejected. This suggests the presence of compounds other than glucosinolates that might be sequestered in or produced by P. brassicae only. In faeces of both Pieris spp. a compound which yielded 4-hydroxybenzylcyanide (HBC) upon incubation with sulfatase was detected in high concentrations when larvae had been reared on S. alba. This compound may be derived from hydrolysis of sinalbin, the main glucosinolate of that plant. The unidentified HBC progenitor was apparently not sequestered in the two Pieris spp., and was not detected in faeces of larvae reared on B. nigra or B. stricta.

Interactions between glucosinolate- and myrosinase-containing plants and the sawfly Athalia rosae

Several insects have specialised on using Brassicaceae as host plants. Therefore, they evolved metabolic pathways to cope with the defensive glucosinolate–myrosinase system of their diet. Larvae of the turnip sawfly, Athalia rosae L. (Hymenoptera: Tenthredinidae), incorporate various glucosinolates from their hosts into their haemolymph. The ability to sequester these metabolites makes A. rosae a useful model system to study mechanisms of glucosinolate metabolism in this species compared to other specialists, and to study effects of sawfly feeding on levels of glucosinolates and their hydrolysing enzymes in plants. The levels of plant metabolites might in turn directly affect the performance of the insect. On the one hand, costs for glucosinolate uptake and avoidance of myrosinase activity were postulated. On the other hand, sequestration of glucosinolates can be part of the insect’s defence against several predators. Here, the findings on glucosinolate metabolic pathways are compared between different herbivores and the sawfly. The impact of different glucosinolate levels and myrosinase activities on the performance of A. rosae is discussed. Furthermore, effects of feeding by A. rosae larvae on the chemical composition and enzyme activities of various Brassicaceae species are summarised. Induction patterns vary not only between different plant species and cultivars but also due to the inducing agent. Finally, the plant–herbivore interactions are discussed with regard to the sawflies’ defence abilities against different carnivore guilds.

Host plant derived feeding deterrence towards ants in the turnip sawfly Athalia rosae

Larvae of the sawfly Athalia rosae ruficornis Jakovlev (Hymenoptera: Tenthredinidae) feed on several glucosinolate‐containing plants and have been shown to sequester the main glucosinolates of different hosts, namely sinalbin (p‐hydroxybenzylglucosinolate) from Sinapis alba L., sinigrin (allylglucosinolate) from Brassica nigra (L.) Koch, and glucobarbarin ((S)‐2‐hydroxy‐2‐phenylethylglucosinolate) from Barbarea stricta Andrz. (Brassicaceae). These plant metabolites are stored in the haemolymph, which is readily released when larvae are attacked by predators. In a dual‐choice bioassay the bio‐activity of sawfly haemolymph collected from larvae reared on different host plants (S. alba, B. nigra, and B. stricta) was tested against the ant Myrmica rubra L. (Hymenoptera: Formicidae). The haemolymph had a stronger deterrence effect when the corresponding sawfly larvae were reared on S. alba than when reared on B. nigra and B. stricta. Haemolymph of caterpillars of Pieris rapae L. (Lepidoptera: Pieridae) that had fed on S. alba was not deterrent to the ants. No sinalbin could be detected in their haemolymph. The glucosinolates sinalbin and sinigrin, offered in a concentration comparable to that in the sawfly haemolymph, were deterrent to the ants, but not as strongly as the corresponding haemolymph samples. This suggests, that glucosinolates are not the only compounds involved in the chemical defence of A. rosae. However, the presence of sequestered glucosinolates is already a sufficient defence towards predators such as ants, and their effectiveness is modulated by the host plant chemistry.