" Arabidopsis Thaliana " response to insect feeding : new components controlling defense gene expression and plant resistance

Abstract: Plants cannot run away to escape attacking herbivores, but they defend themselves by producing anti-digestive proteins and toxic compounds (for example glucosinolates). The first goal of this thesis was to study changes in gene expression after insect attack using microarrays. The responses of Arabidopsis thaliana to feeding by the specialist Pieris rapae and the generalist Spodoptera liffora is were compared. We found that the transcript profiles after feeding by the two chewing insects were remarkably similar, although the generalist induced a slightly stronger response. The second goal was to evaluate the implication of the four signals jasmonic acid (JA), salicylic acid (SA), ethylene (ET), and abscisic acid (ABA) in the control of insect-regulated gene expression. Using signaling mutants, we observed that JA was the predominant signal and that ABA modulated defense gene expression. In contrast, SA and ET appeared to control slightly gene expression, but only after feeding by S. litforalis. The third goal was to establish whether plant responses are really effective against insects. In accordance with the transcript profile, both insects were affected by the JA-dependent defenses, as they performed better on the JA-insensitive mutant. S. littoralis also performed better on ABA-deficient mutants, providing evidence for the role of ABA in defense against insects. When testing indole or aliphatic glucosinolate deficient mutants, we found that they were also more susceptible to insect feeding, providing some of the first genetic evidence for the defensive role of glucosinolates in planta. Finally, a glutathione-deficient mutant, pad2-1, was also more susceptible to insect feeding and we could attribute this phenotype to a lowered accumulation of the major indole glucosinolate. In this thesis, we provide a comprehensive list of insect-regulated genes, including many transcription factors that constitute interesting candidate genes for the further study of insect-induced expression changes. Understanding how the plant responses to insects are regulated will provide tools for a better management of insect pest in the field. Résumé: Les plantes ne peuvent s'échapper pour fuir les insectes qui les attaquent, mais elles se défendent en produisant des protéines anti-digestives et des composés toxiques (par exemple des glucosinolates). Le premier but de cette thèse était d'étudier les changements de l'expression génétique lors d'attaque par des insectes en utilisant des puces à ADN. Nous avons comparé la réponse d'Arabidopsis thaliana à deux espèces d'insectes avec des habitudes alimentaires différentes : le spécialiste Pieris rapae et le généraliste Spodoptera littoralis. Nous avons trouvé que les profils de transcription après l'attaque par les deux insectes sont remarquablement similaires, bien que le généraliste induise une réponse légèrement plus forte. Le deuxième but était de déterminer l'implication de quatre signaux dans le contrôle de la réponse :l'acide jasmonique (JA), l'acide salicylique (SA), l'éthylène (ET), et l'acide abscissique (ABA). En utilisant de mutants de signalisation, nous avons montré que l'acide jasmonique était le signal prédominant et que l'acide abscissique modulait l'expression génétique. D'autre part, l'acide salicylique et l'éthylène contrôlent à un degré moindre l'expression génétique, mais seulement après l'attaque par S. littoralís. Le troisième but était d'établir si les réponses des plantes sont efficaces contre les insectes. En accord avec le profil de transcription, les deux espèces d'insectes se sont mieux développées sur un mutant insensible au JA, indiquant que les défenses contrôlées par ce signal sont cruciales pour la plante. De plus, les larves de S. littorales se sont mieux développées sur des mutants déficients en ABA, ce qui fournit une preuve du rôle de l'acide abscissique dans la défense contre les insectes. En testant des mutants déficients en glucosinolates de type indole ou aliphatique, nous avons trouvé qu'ils étaient plus sensibles aux insectes, démontrant ainsi le rôle défensif des glucosinolates in planta. Finalement, le mutant déficient en glutathion pad2-1 était aussi plus sensible à l'attaque des insectes, et nous avons pu attribuer ce phénotype à une plus faible augmentation d'un indole glucosinolate dans ce mutant. Dans cette thèse, nous avons mis en évidence un nombre important de gènes contrôlés par les insectes, comprenant de nombreux facteurs de transcription qui constituent des candidats intéressants pour`étudier plus en détail les changements d'expression génétique induits par les insectes. Une meilleure compréhension de la réponse des plantes contre l'attaque des insectes devrait nous permettre de développer de nouvelles stratégies pour mieux gérer les ravageurs des cultures.

Three mechanical weed control techniques in spring cereals

Selostus: Kolme viljojen mekaanista rikkakasvintorjuntamentelmää

Soil tillage affects the community structure of mycorrhizal fungi in maize roots

In this study we tested whether communities of arbuscular mycorrhizal fungi (AMF) colonizing the roots of maize (Zea mays L.) were affected by soil tillage practices (plowing, chiseling, and no-till) in a long-term field experiment carried out in Tanikon (Switzerland). AMF were identified in the roots using specific polymerase chain reaction (PCR) markers that had been developed for the AMF previously isolated from the soils of the studied site. A nested PCR procedure with primers of increased specificity (eukaryotic, then, fungal, then AMF species or. species-grouop specific) was used. Sequencing of amplified DNA confirmed that the DNA obtained from the maize roots was of AMF origin. Presence of particular AMF species or species-group was scored as a presence of a DNA product after PCR with specific primers. We also used single-strand conformation polymorphism analysis (SSCP), of amplified DNA samples to-check if the amplification of the DNA from maize roots matched the expected profile for a particular AMF isolate with a given specific primer pair. Presence of the genus Scutellospora, in maize roots was strongly reduced in plowed and chiseled soils. Fungi from the suborder Glomineae were more prevalent colonizers of maize roots growing in plowed soils, but were also present in the roots from other tillage treatments. These changes in community of AMF colonizing maize roots might be due to (1), the differences in tolerance to the tillage-induced disruption of the hyphae among the different AMF species, (2) changes in nutrient content of the soil, (3) changes in microbial activity, or (4) changes in weed populations in response to soil tillage. This is the first report on community composition of AMF in the roots of a field-grown crop plant (maize) as affected by soil tillage.

Rapid and sensitive hormonal profiling of complex plant samples by liquid chromatography coupled to electrospray ionization tandem mass spectrometry

Background Plant hormones play a pivotal role in several physiological processes during a plant's life cycle, from germination to senescence, and the determination of endogenous concentrations of hormones is essential to elucidate the role of a particular hormone in any physiological process. Availability of a sensitive and rapid method to quantify multiple classes of hormones simultaneously will greatly facilitate the investigation of signaling networks in controlling specific developmental pathways and physiological responses. Due to the presence of hormones at very low concentrations in plant tissues (10-9 M to 10-6 M) and their different chemistries, the development of a high-throughput and comprehensive method for the determination of hormones is challenging. Results The present work reports a rapid, specific and sensitive method using ultrahigh-performance liquid chromatography coupled to electrospray ionization tandem spectrometry (UPLC/ESI-MS/MS) to analyze quantitatively the major hormones found in plant tissues within six minutes, including auxins, cytokinins, gibberellins, abscisic acid, 1-amino-cyclopropane-1-carboxyic acid (the ethylene precursor), jasmonic acid and salicylic acid. Sample preparation, extraction procedures and UPLC-MS/MS conditions were optimized for the determination of all plant hormones and are summarized in a schematic extraction diagram for the analysis of small amounts of plant material without time-consuming additional steps such as purification, sample drying or re-suspension. Conclusions This new method is applicable to the analysis of dynamic changes in endogenous concentrations of hormones to study plant developmental processes or plant responses to biotic and abiotic stresses in complex tissues. An example is shown in which a hormone profiling is obtained from leaves of plants exposed to salt stress in the aromatic plant, Rosmarinus officinalis.

Response to comment on "Climatic niche shifts are rare among terrestrial plant invaders"

Webber et al. take a critical view of our findings that niche expansions are rare in plant invaders, arguing mainly that we did not include nonanalog climates in our analyses. Yet, their concerns include misunderstandings and go beyond the scope of our study, which was purposely restricted to analog climates. We further explain why our results remain robust to other factors of niche dynamics in the native range. We conclude that the implications of our findings remain valid for projections of niche models in analog climates and that projections in nonanalog climates should be undertaken with care.

Weed flora in organically grown spring cereals in Finland

Selostus: Luomukevätviljapeltojen rikkakasvillisuus 1997-1999

Plant development: should I stop or should I grow?

Plant growth is tightly controlled through the integration of environmental cues with the physiological status of the seedling. A recent study now proposes a model explaining how the plant hormone ethylene triggers opposite growth responses depending on the light environment.

Seasonal growth of "algodão-bravo" (Ipomoea carnea spp. fistulosa)

Ipomoea carnea spp. fistulosa, a native woody perennial, is capable of spreading rapidly over seasonally flooded grassland in the Brazilian Pantanal, South America's largest wetland, thus conflicting with the local cattle ranching. I. carnea is controlled by mowing at the onset of the rainy season, as close as possible before the seasonal flooding. Often, however, flooding begins after the plant has had enough time to re-sprout enabling it to survive. The objective of this study was to verify if Ipomoea carnea plant's production follows a seasonal cycle, and, if so, at which point in this cycle, the plant is most vulnerable to mechanical control measures. Seasonal dynamics of stem and leaf production of I. carnea were studied. The results showed that growth of I. carnea is fastest at the onset of the rainy season in November/December. Production declines when seasonal flooding commences in January/February and almost ceases towards the begin of the dry season in May/June. This leads to the proposal that I. carnea could be controlled more effectively if the weed were mown in the early dry season when its production and its capability to re-sprout is lowest, and if any new sprouts were cut by hand when the seasonal flooding starts.

Determination of the critical period of weed control in corn using a thermal basis

Field studies were conducted over 3 years in southeast Buenos Aires, Argentina, to determine the critical period of weed control in maize (Zea mays L.). The treatments consisted of two different periods of weed interference, a critical weed-free period, and a critical time of weed removal. The Gompertz and logistic equations were fitted to relative yields representing the critical weed-free and the critical time of weed removal, respectively. Accumulated thermal units were used to describe each period of weed-free or weed removal. The critical weed-free period and the critical time of weed removal ranged from 222 to 416 and 128 to 261 accumulated thermal units respectively, to prevent yield losses of 2.5%. Weed biomass proved to be inverse to the crop yield for all the years studied. When weeds competed with the crop from emergence, a large increase in weed biomass was achieved 10 days after crop emergence. However, few weed seedlings emerged and prospered after the 5-6 leaf maize stage (10-20 days after emergence).

Decision criteria for optimizing postemergence johnsongrass control in soybean crops in Argentina

Field studies were established in Zavalla and Oliveros, Argentina, during four years in order to optimize Johnsongrass (Sorghum halepense (L.) Pers.) chemical control by means of the thermal calendar model in comparison with other criteria (weed height or days after sowing). The effect of three application dates of postemergence herbicides was determined by visual control, density of tillers originated from rhizome bud regrowth, and from crown and shoot bud regrowth, and soybean yield. Following the thermal calendar model criterion, applications during the second date afforded the best control. Weed height for the first date showed little variability between experiments but was highly variable in the second and third application dates, achieving in some cases values greater than 120 cm. For all years, no significant differences were detected for crop yield between the first and second application dates, and yields were always lower for the third date. The greatest rhizome bud regrowth was observed for the earliest application date and the highest crown and shoot bud regrowth was determined for the last application date. Parameters associated with control efficiency showed the best behaviour for the second date. However, plant height at this moment may interfere with herbicide application and the variability exhibited by this parameter highlights the risk of determining control timing using only one decision criterion.

Detection of plant-modulated alterations in antifungal gene expression in Pseudomonas fluorescens CHA0 on roots by flow cytometry.

The biocontrol activity of the root-colonizing Pseudomonas fluorescens strain CHA0 is largely determined by the production of antifungal metabolites, especially 2,4-diacetylphloroglucinol. The expression of these metabolites depends on abiotic and biotic environmental factors, in particular, elements present in the rhizosphere. In this study, we have developed a new method for the in situ analysis of antifungal gene expression using flow cytometry combined with green fluorescent protein (GFP)-based reporter fusions to the phlA and prnA genes essential for the production of the antifungal compounds 2,4-diacetylphloroglucinol and pyrrolnitrin, respectively, in strain CHA0. Expression of phlA-gfp and prnA-gfp in CHA0 cells harvested from the rhizosphere of a set of plant species as well as from the roots of healthy, leaf pathogen-attacked, and physically stressed plants were analyzed using a FACSCalibur. After subtraction of background fluorescence emitted by plant-derived particles and CHA0 cells not carrying the gfp reporters, the average gene expression per bacterial cell could be calculated. Levels of phlA and prnA expression varied significantly in the rhizospheres of different plant species. Physical stress and leaf pathogen infection lowered phlA expression levels in the rhizosphere of cucumber. Our results demonstrate that the newly developed approach is suitable to monitor differences in levels of antifungal gene expression in response to various plant-derived factors. An advantage of the method is that it allows quantification of bacterial gene expression in rhizosphere populations at a single-cell level. To our best knowledge, this is the first study using flow cytometry for the in situ analysis of biocontrol gene expression in a plant-beneficial bacterium in the rhizosphere.