Le phloème est un tissu complexe composé de plusieurs types cellulaires, dont les cellules compagnes et les tubes criblés. Il permet le transport et l’allocation à longue distance de nombreux métabolites et de macromolécules. Il existe dans les tubes criblés des structures très spécifiques dont la fonction est inconnue. Par exemple le rôle des protéines phloémiennes PP2 (Phloem Protein 2) qui ont été de longue date décrite dans les tubes criblés, reste à définir. Les protéines PP2 présentent une activité de lectine et d’interaction avec des protéines de sève phloémienne, suggérant un rôle dans le transport de macromolécules dans le phloème.Nous avons étudié la fonction de deux membres de la famille, PP2-A1 et PP2-A2, chez l’espèce modèle Arabidopsis thaliana. Plusieurs approches ont été mises en œuvre pour étudier ces protéines ; une approche cytologique, la recherche de partenaire protéiques et la création de lignées dérégulées pour l’expression des gènes PP2. L’étude de la localisation de PP2-A1 avec des étiquettes fluorescentes dérivées de la GFP a été réalisée par microscopie confocale, dans les cellules compagnes et dans les tubes criblés ; elle a montré que cette protéine présente une localisation nucléo- cytoplasmique dans les cellules compagnes tandis qu’elle forme des agrégats fixés dans les tubes criblés. Ceci suggère que PP2-A1 est ancrée dans les tubes criblés, à la membrane plasmique ou à certains organites. Des résultats similaires ont été obtenus pour PP2-A2. Sur la base de cette première étude, nous permettant d’identifier in vivo avec précision les cellules compagnes et les tubes criblés, et en utilisant plusieurs marqueurs subcellulaires fluorescents de référence, nous avons réalisé une cartographie subcellulaire fine des cellules compagnes et des tubes criblés in vivo. Cette approche a permis de décrire in vivo l’organisation subcellulaire de ces cellules. Elle a révélé la présence de nombreux organites présents à la périphérie des tubes criblés et de nature énigmatique, suggérant une activité importante dans ces cellules, en accord avec des données récentes de protéome des tubes criblés. L’étude de lignées surexprimant des versions étiquetées de PP2-A1 et PP2-A2 nous a permis de mettre en évidence un phénotype altéré avec des plantes qui présentent un retard de floraison et une biomasse plus importante. Ces observations suggèrent que PP2-A1 et PP2-A2 pourraient avoir un rôle dans la signalisation à longue distance. Ces travaux, qui illustrent la complexité des cellules du phloème, apportent ainsi des éléments nouveaux sur les voies de signalisation à longue distance utilisées par les végétaux pour coordonner leur croissance et leur développement. / The phloem is a complex tissue, made of several cell types, including companion cells and sieve elements. It plays an important role in long-distance transport and allocation of a number of metabolites and macromolecules. The sieve elements display specific structures yet uncharacterized and of unknown function. For instance the function of the phloem specific PP2 proteins (Phloem Protein 2) that have been described for long in the sieve elements is still unclear. PP2 proteins present lectin activity and bind to phloem sap proteins, suggesting a role in the transport of macromolecules in the phloem. We have investigated the function of two members of this family, PP2-A1 and PP2-A2 in the model species Arabidopsis thaliana. Different approaches have been undertaken to study these proteins: a cytological approach, the research of partners, and the study of downregulated and upregulated lines for the PP2-A1 and PP2-A2 genes. Localization studies of PP2-A1 fused to GFP-derived fluorescent tags have been realized by confocal laser scanning microscopy in the companion cells and the sieve elements; they showed that PP2-A1 presents a nucleocytoplasmic localization in the companion cells, whereas it forms fixed aggregates in the sieve elements. It suggested that PP2-A1 is anchored to the plasma membrane or to organelles inside the sieve elements. Similar results were obtained for PP2-A2. Making use of this study to accurately identify companion cells and sieve elements in vivo, and using additional subcellular fluorescent markers, we realized a fine mapping of companion cells and sieve elements in vivo. This study revealed the presence of numerous organelles of unknown identity at the periphery of the sieve elements, suggesting an important activity in those cells consistent with recent proteomics analysis of the sieve tubes. The study of plants overexpressing tagged versions of PP2-A1 and PP2-A2 enabled to observe an altered phenotype with delayed flowering and increased biomass, suggesting that PP2-A1 and PP2-A2 may play a role in long distance signalling. This work illustrates the complexity of phloem cell organization and functions, bringing new elements on long distance signalling pathway used by the plants to coordinate their growth and development.

L'objectif de cette thèse était d'élucider le rôle des transporteurs de saccharose dans les racines d’A. thaliana pour mieux comprendre la répartition du carbone dans la plante entière.Nous nous sommes focalisés sur l’étude des deux principaux transporteurs de saccharose exprimés dans les racines : AtSUC1 et AtSUC2. Une étude du mutant KO suc1 et une étude des plantes greffées présentant la mutation KO du gène AtSUC2 uniquement dans les racines ont été réalisées sur des plantes au stade adulte (30 à 32 jours après semis) cultivées en rhizobox. De plus, les localisations de l’expression des gènes et des protéines de ces transporteurs ont été réalisées dans les racines de plantes adultes pour la première fois. La localisation de l’expression d’AtSCU1 dans les pointes racinaires, nous a permis de conclure sur un rôle potentiel d’AtSUC1 dans le déchargement du saccharose du phloème dans les zones de croissance des racines et/ou un rôle potentiel de senseur du signal glucidique ou du déficit hydrique, probablement en lien avec l’ABA. Les résultats montrent qu’AtSUC2 aurait un rôle dans le développement racinaire, certainement via le déchargement du saccharose du phloème dans les zones d’élongation des racines et dans le rechargement du phloème le long des racines (notamment dans les départs de racines latérales). De plus, AtSUC2 pourrait avoir un rôle dans l’augmentation locale d’hexoses liée à l’émergence des racines latérales. Enfin, nos résultats indiquent une contradiction entre la localisation de l’expression du gène AtSUC2 retrouvée dans le parenchyme cortical et la localisation de la protéine qui n’est pas retrouvée dans ces cellules. Néanmoins les deux approches s’accordent sur la localisation dans les cellules compagnes. / The aim of this thesis was to elucidate the role of sucrose transporters in the roots of Arabidopsis thaliana to have a better comprehension of the carbon partitioning in the whole plant.We focused on the study of the two main sucrose transporters expressed in the roots: AtSUC1 and AtSUC2. A study of the KO mutant suc1 and a study on grafted plants with the KO mutation of the AtSUC2 gene only in the roots (micro-graft) were carried out on adult plants (30 to 32 days after sowing) grown in rhizobox. In addition, the localization of the expression of the genes and proteins of transporters were carried out in the roots of adult plants for the first time. The localization of AtSCU1 expression in the root tips allowed us to conclude on a potential role of AtSUC1 in the sucrose phloem unloading in root growth areas and/or a potential role of sensor of carbohydrate signal or of water deficit, probably related to ABA. The results also show that AtSUC2 has a role in root development and certainly via sucrose phloem unloading in root elongation’s areas and in the phloem reloading along the roots (especially in sites of emergence of lateral roots). In addition, AtSUC2 may have a role in the local increase of hexoses associated with the emergence of lateral roots. Finally, our results indicate a contradiction between the localization of the AtSUC2 gene expression found in the cortical parenchyma and the localization of the protein that is not found in these cells. Nevertheless the two approaches agree on the localization in the companion cells.

Caesalpinia echinata Lam (Leguminosae) é a espécie símbolo do Brasil, muito importante historicamente para o país. A árvore de pau brasil foi intensamente explorada no período do descobrimento, durante o século XVI, para a extração de corante e, atualmente, a sua madeira é utilizada na fabricação de arcos para instrumentos de corda. Neste trabalho foi estudada a espécie em três áreas: Mogi Guaçu, em São Paulo, num bosque experimental; em Ilhéus, na Bahia, em uma fazenda de cacau, e em Campo Grande no Mato Grosso do sul, na Reserva do campus da Universidade Federal. Pretendeu se neste trabalho comparar a sazonalidade da atividade cambial, o xilema e floema secundários, assim como descrever a formação e a diferenciação dos tecidos vasculares secundários. Para isso, um método não destrutivo de coleta foi usado, além de várias técnicas de observação do material, como microscopia de luz, eletrônica de varredura e transmissão, e confocal de varredura a laser. Foi realizado o acompanhamento fenológico da espécie assim como dados ambientais, tal como pluviosidade, temperatura e fotoperíodo. A atividade cambial foi observada a partir da presença de figuras mitóticas e fragmoplastos nas células cambiais. O câmbio é ativo em Mogi Guaçu por seis meses, enquanto que em Ilhéus por cerca de dez meses. A atividade cambial é principalmente influenciada pela pluviosidade, e os períodos de inatividade estão relacionados com o déficit hídrico no solo. No desenvolvimento do xilema secundário, os elementos de vaso são as primeiras células a se diferenciarem. Após a expansão celular, há o rompimento do tonoplasto, e posteriormente, ocorre a deposição da parede secundaria e lignificação, com formação das pontoações e das guarnições, e delimitação da placa de perfuração. A última etapa é a lise do núcleo e hidrólise do conteúdo citoplasmático. A anatomia comparada do xilema coletado em Mogi Guaçu e em Ilhéus mostrou diferenças significativas para os seguintes parâmetros: diâmetro tangencial do elemento de vaso, relação diâmetro e comprimento do elemento de vaso, freqüência dos elementos de vaso por mm2, espessura da parede da fibra, comprimento da fibra. Os raios mostram uma tendência a serem mais largos nos espécimes de Ilhéus, embora esta diferença não seja estatisticamente significativa. Quanto às guarnições das pontoações intervasculares tem se que na abertura externa da pontoação são similares, sendo ramificadas e longas nos espécies das três regiões analisadas; porém na abertura interna da pontoação as guarnições são mais abundantes e ramificadas em indivíduos coletados em Mogi Guaçu e Campo Grande, onde há um maior período de déficit hídrico no solo por ano. O floema é composto por uma porção não condutora e uma porção condutora, que corresponde a 1/4 a 1/6 do floema secundário. Os elementos de tubo crivado e as células do parênquima floemático apresentam um arranjo tangencial alternado em muitos dos espécimes analisados. No floema não condutor são observadas esclereides. No início de sua diferenciação, ainda possuem núcleo, e a proteína P não dispersa. No floema de pau brasil não foi detectada a presença de marcadores de crescimento, como já visto em algumas espécies tropicais. / Caesalpinia echinata Lam (Leguminosae) is the species symbol of Brazil, historically very important to the country. The Pernambuco wood tree was intensely exploited in the period of the discovery, during XVI century, for the pigment extraction and, nowadays, the wood is extensively used in the production of bows for string instruments. In this work it was studied the species in three areas, in Mogi Guaçu, São Paulo; in an experimental forest; in Ihéus, Bahia, in a cocoa farm, and in Campo Grande, Mato Grosso do Sul, in the Reserve of the Federal University campus. The goal of this work was to compare the seasonality of cambium activity, secondary xylem and phloem, as well as describing the formation and differentiation of secondary vascular tissues. For that, a non destructive collection method was employed and, several techniques were used to analyze the samples, including optic, scanning, transmission and confocal scanning laser microscopy. The phenology of the species was followed correlated to environmental variables, such as precipitation, temperature and photoperiod. Cambium activity was observed from the presence of mitotic figures and phragmoplasts in cambium cells. Cambium is active in Mogi Guaçu for six months, while in Ilhéus is for about ten months. Cambial activity is mostly influenced by precipitation, and inactivity periods are related to water deficit in the soil. In the secondary xylem development, vessel elements are the first cells to differentiate. After cellular expansion, there is the breakdown of the tonoplast, and later, the walls undergo lignification with pits and vestures formation, and the delimitation of the perforation plate. The last stage is the nucleus lise of the nucleus and hidrolysis of cytoplasm contents. Compared anatomy of xylem between Mogi Guaçu and Ilhéus showed significant differences regarding the following parameters: vessels tangential diameter, vessels diameter and length rate, vessel elements frequency mm2, fiber walls thickness and length Rays show a tendency to be wider in the specimens of Ihéus, although not statistically significant Regarding vessels, there is no difference between vestures of the external pit aperture; however vestures are more abundant in the internal apertures in samples collected in Mogí Guaçu and Campo Grande, where there is a longer period of water deficit in the soil per year. Phloem is composed of conductive and non conductive portions, corresponding to 1/4 up to 1/6 of secondary phloem. Sieve tube elements and parenchyma cells have a tangential alternate arrangement. In non conductive phloem sclereids are observed In the beginning of its differentiation, sieve tube elements still possess nucleus, and non disperse protein P. In Pernambuco wood phloem there is no growth markers, as already detected some other tropical species.

A citricultura brasileira é a maior produtora e exportadora de citros e tem sido afetada por doenças que causam sérios prejuízos a produção e a qualidade dos frutos. No entanto, a cultura apresenta grandes problemas, entre eles, os fatores fitossanitários, que vem dizimando milhares de plantas e afetando a produtividade e a competitividade do setor. Atualmente, o huanglongbing (HLB), associado às bactérias de floema Candidatus Liberibacter spp., é considerado uma das mais destrutivas doenças de citros. A inexistência de cultivares de laranja doce resistentes ao HLB torna a transformação genética de citros uma ferramenta importante no controle desta doença. Para se defender do ataque de pragas e patógenos as plantas desenvolveram, durante o processo evolutivo, uma série de mecanismos de defesa, no qual pode-se incluir a produção de peptídeos com atividade antimicrobiana. As defensinas vegetais são peptídeos pequenos relacionadas à patogênese (PR), que possuem atividade antimicrobiana associada aos mecanismos de defesa das plantas. Assim, o objetivo deste trabalho foi a obtenção de plantas transgênicas de laranja doce (Citrus sinensis L.) cvs. \'Hamlin\', \'Natal\', \'Valência\' e \'Pera\', via Agrobacterium tumefaciens, superexpressando o gene codificador de defensina (def), isolado de Citrus sinensis cv. \'Valência\', dirigido pelo promotor com expressão preferencial no floema AtSUC2 (transportador de sacarose, clonado de Arabidopsis thaliana) ou pelo promotor constitutivo CaMV 35S (clonado do vírus do mosaico da couve-flor). Os explantes utilizados na transformação genética foram segmentos de epicótilo obtidos de plantas germinadas in vitro. A identificação das plantas transgênicas foi realizada por meio da análise da PCR, utilizando-se primers para a detecção do fragmento do gene de seleção nptII. As plantas PCR+ foram aclimatizadas e transferidas para casa-de-vegetação. A análise de Southern blot confirmou a integração do transgene em 36 plantas. Foram obtidas 7 plantas transgênicas da cultivar \'Hamlin\', 9 da cultivar \'Natal\', 1 da cultivar \'Pera\' e 9 da cultivar \'Valência\' contendo a construção gênica pC35S/def, e 3 plantas transgênicas da cultivar \'Hamlin\', 6 da cultivar \'Natal\' e 1 da cultivar \'Valência\' contendo a construção gênica pcAtSUC2/def. Os resultados obtidos neste trabalho serão importantes para futura avaliação e estudo visando o controle de Candidatus Liberibacter spp.. / The Brazilian citrus industry is the world\'s largest producer and exporter of citrus, however, it has been affected by diseases that cause serious production losses and damages to fruit quality. However, the culture faces problems, namely phytosanitary issues that have been damaging thousands of plants, affecting yield and competitiveness of the sector. Currently, Huanglongbing (HLB), associated with phloem bacteria Candidatus Liberibacter spp., is considered one of the most destructive citrus diseases. The lack of sweet orange cultivars resistant to HLB makes genetic transformation an important tool in the disease control. To defend from pest and pathogen attack, plants developed a series of defense mechanisms during the evolutionary process, which may include the production of peptides with antimicrobial activity. Plant defensins are small peptides related to pathogenesis (PR) which have antimicrobial activities, associated with plant defense mechanisms . The objective of this study was to obtain transgenic plants of sweet orange (Citrus sinensis L.) cultivars \'Hamlin\', \'Natal\', \'Valência\' and \'Pera\' with Agrobacterium tumefaciens overexpressing the defensin gene (def), isolated from Citrus sinensis cv. \'Valência\', controlled by the promoter with preferential expression in the phloem AtSUC2, (sucrose transporter, cloned from Arabidopsis thaliana) or by the constitutive promoter CaMV 35S (cloned from the mosaic virus of cauliflower). The explants used in genetic transformation were epicotyl segments obtained from germinated plants in vitro. The identification of transgenic plants was accomplished by PCR analysis using primers for the detection of nptII gene fragment. The PCR+ plants were acclimatized and transferred to greenhouse. The analysis of Southern blot confirmed the transgene integration in 36 plants. Seven transgenic plants were obtained for the cultivar \'Hamlin\', nine for \'Natal\', one for \'Pera\' and nine for \'Valência\' containing the gene construct pC35S/def and three transgenic plants for \'Hamlin\', six for \'Natal\' and one for \'Valência\' containing the gene construct pcAtSUC2/def. The results obtained in this work are important for future evaluation of the plants for resistance to Candidatus Liberibacter spp..

O sistema vascular das lianas, em especial o xilema, mostrou−se repetidas vezes distinto nas lianas, com aspectos compartilhados mesmo dentre linhagens distantemente relacionadas, tais como a presença de variações cambiais, vasos mais largos e longos, parênquima axial mais abundante − frequentemente não−lignificado − raios mais altos e largos − geralmente heterocelulares. Não obstante todo esse conhecimento, poucos trabalhos investigaram o impacto da evolução do hábito lianescente no sistema vascular em linhagens cujos ancestrais não são lianas e sim plantas auto−suportantes. Portanto, nesta tese exploramos o lenho, o floema e a anatomia caulinar como um todo em linhagens que contêm lianas e plantas auto−suportantes, utilizando filogenias bem sustentadas e investigações anatômicas detalhadas. Em Bignoniaceae (Lamiales), investigamos em detalhe a anatomia do lenho, delimitando caracteres e estados de caráter e mapeando-os na filogenia mais recente do grupo, encontrando que modificações eco−fisiológicas e transições de hábito tiveram grande impacto na evolução do lenho na família. Anéis porosos e semi−porosos, bem como espessamento espiralado foram encontrados em plantas crescendo em latitudes mais altas ou em regimes hídricos fortemente sazonais, ao passo que fibras septadas apareceram correlacionadas com a presença de parênquima axial escasso. A evolução de lianas, por sua vez, parece ter levado a um aumento no diâmetro dos vasos, contudo dimórficos, células perfuradas de raio, parênquima axial mais escasso e surgimento de variações cambiais. Apesar da enorme diversidade dentro de Bignoniaceae, os grandes clados possuem uma anatomia bastante preditiva e 9 possíveis sinapomorfias morfológicas são sugeridas para clados delimitados somente com base em dados moleculares. Dentro das traqueófitas, investigamos 26 pares filogeneticamente controlados de lianas espécies auto−suportantes relacionadas pertencentes a todas as principais linhagens de traqueófitas (exceto licófitas), a fim de buscar caracteres que tenham evoluído em correlação com o hábito lianescente. Encontramos que os elementos crivados e os poros das placas crivadas têm sempre maior calibre nas lianas, e que os raios são mais altos e heterocelulares. Contudo, as principais características do floema das lianas se mantêm conservadas em relação às espécies auto−suportantes relacionadas, evidenciando que as lianas teriam evoluído um sistema de condução de fotossintetatos mais eficiente, porém preservando um alto sinal filogenético. Em Malpighiaceae, lianas são abundantes, tal como as variações cambiais. Contudo, pouco se sabe sobre o número de variações presentes na família ou como elas estariam distribuídas. Aqui delimitamos 6 diferentes tipos de variação cambial, que teriam evoluído independentemente 8 vezes na família, cujo ancestral é reconstruído como tendo caule simples. Muitas dessas variações compartilham estágios de desenvolvimento, ao passo que variações anatomicamente muito similares derivam de trajetórias ontogenéticas distintas. Dentro dos gêneros as variações se mostraram conservadas e mesmo dentre grupos irmãos do novo e velho mundo, evidenciando que as variações cambiais seriam bons indicadores de relações na família. De maneira geral, podemos concluir que lianas impactam significativamente o sistema vascular nas linhagens onde ocorrem e que tais modificações em geral resultam em um sistema de condução hídrico e de fotossintetatos mais eficiente e também mais flexível para a escalada. / The vascular system of lianas, especially the xylem, has been repeatedly been shown to be different, with lianas having a set of features shared among even distantly related lineages, such as the presence of cambial variants, wide and long vessels, more abundant axial parenchyma, frequently non−lignified, taller and wider rays, which are generally heterocellular. In spite of this amount of knowledge, few works have investigated the impact for the vascular system of the evolution of this habit within lineages whose ancestors are not lianas, but self-supporting plants. Therefore, in this dissertation we explored wood, phloem and overall stem−anatomy evolution in lineages that contain lianas and self−supporting plants, using well−supported phylogenies and detailed anatomical investigations. Within Bignoniaceae (Lamiales), we thoroughly investigated the wood anatomy, delimiting character states and mapping them onto the last phylogeny for the group, encountering that eco−physiological and habit transition were the main drivers of modifications in the wood anatomy in the family. Ring−porous and semi−ring porous woods and helical thickening was found in plants either growing in higher latitudes or with marked seasonal water regimes, and septate fibres correlated with scanty axial parenchyma, which are eco−physiological drivers. Evolution of lianas, in turn, drove an increase in vessel diameter, wide vessels accompanied by very narrow ones, presence of perforated ray cells, scanty axial parenchyma and cambial variants. Despite the great wood anatomical diversity within the family, major clades have quite predictive wood anatomy and 9 possible anatomical synapomorphies were raised in this work to clades previously delimitated exclusively by molecular characters. Within the tracheophytes, we investigated 26 phylogenetically controlled pairs of lianas and their self−supporting relatives within all major lineages of tracheophytes (except lycophytes), in order to seek characters evolving in correlation with the lianescent habit. We found that the sieve elements and sieve pores were always wider in the lianas, and that the rays were always taller. However, all the main characters of the phloem of the lianas remained conserved with that of their self−supporting relatives. This evidenced that although a more efficient photosynthetic conductive system evolved in the phloem of lianas, overall anatomy conserved a high phylogenetic signal. Within Malpighiaceae, lianas are abundant and many cambial variants are present. However, nothing was known regarding how many types of cambial variants there were in the family and how they were distributed. We were able to delimit 6 different types of cambial variants that evolved at least 8 times independently in the family, which ancestrally lacks a cambial variant. Many of these types share common stages of development and some variants that are anatomically very similar derive from different ontogenetic trajectories. Within the genera, the variants are conserved, and even between sister groups in the new and old world, evidencing that cambial variants may be a good indicator of relationships within the family. Overall, we conclude that lianas greatly impact the evolution of the vascular system in the lineages where they have evolved, and these modifications normally result in a more efficient water and photosynthates conduction system and an increased flexibility for climbing.

Populus tremuloides Michx. (trembling aspen) is a tree species native to much of North America, characterized by an excurrent crown with horizontal to ascending branches and a dominant terminal leader. An unusual clone of trembling aspen was discovered in the 1940s near Hafford, Saskatchewan. This clone demonstrates abnormal crown morphology, in which vigorous shoots bend down, ultimately leading to an overall twisted or crooked appearance. The objectives of the present study were to investigate the mechanism of shoot bending by (1) characterizing the process and timing of bending, (2) evaluating structural aspects of developing wild-type and crooked aspen shoots, and (3) comparing anatomical features of bending shoots with wild-type shoots. L-system reconstruction models of 3-D digitized shoot development revealed dramatic bending midway through the growing season. Morphological analyses revealed that crooked aspen shoots had greater taper compared to the wild-type, typically known to create shoots resist deflection and bending. However, preliminary strength analyses indicated that crooked aspen shoots were less rigid, with smaller values of Young’s modulus compared to wild-type shoots. Anatomical investigations revealed differences in several structural tissues between developing wild-type and crooked aspen shoots, and differences within crooked aspen shoots. Primary phloem fibres on the upper side of bending shoots maintained relatively large lumens while those on the lower side were fully lignified, similar to those of mature vertically oriented wild-type leader shoots. These differences may result in differential extension growth early in development, and/or uneven mechanical support later on, ultimately resulting in bending due to self-weight. Gelatinous fibres (G-fibres), characteristic of tension wood (TW), were found throughout older wild-type and vertically oriented crooked aspen shoots; however, G-fibres were only found on the lower side of crooked aspen shoots. These lateral differences could have contributed to shoot bending by actively bending shoots downwards, or lack of TW on the upper side may not have prevented biomechanical bending from self weight. Nevertheless, shoot bending stops at the end of the growing season, suggesting that the mechanisms involved in creating bent shoots are only functional during the first growing season. / February 2006

Most studies of under-bark microclimate have been restricted to observations of a few coniferous trees in wooded southern latitudes. This limitation is worrying because of emerging wood-boring pests that specialize on deciduous trees in Canada, such as emerald ash borer (Agrilus planipennis). Using a large data set that includes 60 ash trees spread across both urban and woodlot sites in 6 different Ontario locations, I found that the under-bark microclimate of deciduous trees can provide wood-boring beetles with an environment in which temperatures which differ from air temperature. On average, daily minimum under-bark temperatures are significantly warmer than air temperatures in the winter months. At temperatures low enough to cause substantial cold-temperature mortality to emerald ash borer, the difference between under-bark and air temperature can be large. In addition, I observed that the difference between daily minimum under-bark and air temperature can vary, and consequently that assumptions of a constant level of difference between the two are not valid. In the spring season, I found that daily under-bark temperature maxima on the south side of the tree are significantly warmer than air temperature maxima. This difference lead to faster predicted development times for beetles in the southern under-bark microclimate of urban trees as compared to predictions based on air temperature, suggesting that city trees may impact overall population dynamics. While it is clear that under-bark temperatures differ from air temperatures, and are important to predicting possible range and population growth of wood-boring insects, large scale measurements of microclimate conditions are not feasible. I tested the ability of a simple Newtonian cooling model to predict under-bark temperature extremes using weather station data. While the model did not predict daily under-bark temperature maxima accurately, predictions of minima were quite accurate (1.31˚C average root mean squared error), especially when compared to the errors from assuming under-bark temperature is the same as air temperature (3.20˚C average root mean squared error). I recommend use of the Newtonian cooling model to predict under-bark temperature minima of deciduous and coniferous trees.

Populus tremuloides Michx. (trembling aspen) is a tree species native to much of North America, characterized by an excurrent crown with horizontal to ascending branches and a dominant terminal leader. An unusual clone of trembling aspen was discovered in the 1940s near Hafford, Saskatchewan. This clone demonstrates abnormal crown morphology, in which vigorous shoots bend down, ultimately leading to an overall twisted or crooked appearance. The objectives of the present study were to investigate the mechanism of shoot bending by (1) characterizing the process and timing of bending, (2) evaluating structural aspects of developing wild-type and crooked aspen shoots, and (3) comparing anatomical features of bending shoots with wild-type shoots. L-system reconstruction models of 3-D digitized shoot development revealed dramatic bending midway through the growing season. Morphological analyses revealed that crooked aspen shoots had greater taper compared to the wild-type, typically known to create shoots resist deflection and bending. However, preliminary strength analyses indicated that crooked aspen shoots were less rigid, with smaller values of Young’s modulus compared to wild-type shoots. Anatomical investigations revealed differences in several structural tissues between developing wild-type and crooked aspen shoots, and differences within crooked aspen shoots. Primary phloem fibres on the upper side of bending shoots maintained relatively large lumens while those on the lower side were fully lignified, similar to those of mature vertically oriented wild-type leader shoots. These differences may result in differential extension growth early in development, and/or uneven mechanical support later on, ultimately resulting in bending due to self-weight. Gelatinous fibres (G-fibres), characteristic of tension wood (TW), were found throughout older wild-type and vertically oriented crooked aspen shoots; however, G-fibres were only found on the lower side of crooked aspen shoots. These lateral differences could have contributed to shoot bending by actively bending shoots downwards, or lack of TW on the upper side may not have prevented biomechanical bending from self weight. Nevertheless, shoot bending stops at the end of the growing season, suggesting that the mechanisms involved in creating bent shoots are only functional during the first growing season.

Most studies of under-bark microclimate have been restricted to observations of a few coniferous trees in wooded southern latitudes. This limitation is worrying because of emerging wood-boring pests that specialize on deciduous trees in Canada, such as emerald ash borer (Agrilus planipennis). Using a large data set that includes 60 ash trees spread across both urban and woodlot sites in 6 different Ontario locations, I found that the under-bark microclimate of deciduous trees can provide wood-boring beetles with an environment in which temperatures which differ from air temperature. On average, daily minimum under-bark temperatures are significantly warmer than air temperatures in the winter months. At temperatures low enough to cause substantial cold-temperature mortality to emerald ash borer, the difference between under-bark and air temperature can be large. In addition, I observed that the difference between daily minimum under-bark and air temperature can vary, and consequently that assumptions of a constant level of difference between the two are not valid. In the spring season, I found that daily under-bark temperature maxima on the south side of the tree are significantly warmer than air temperature maxima. This difference lead to faster predicted development times for beetles in the southern under-bark microclimate of urban trees as compared to predictions based on air temperature, suggesting that city trees may impact overall population dynamics. While it is clear that under-bark temperatures differ from air temperatures, and are important to predicting possible range and population growth of wood-boring insects, large scale measurements of microclimate conditions are not feasible. I tested the ability of a simple Newtonian cooling model to predict under-bark temperature extremes using weather station data. While the model did not predict daily under-bark temperature maxima accurately, predictions of minima were quite accurate (1.31˚C average root mean squared error), especially when compared to the errors from assuming under-bark temperature is the same as air temperature (3.20˚C average root mean squared error). I recommend use of the Newtonian cooling model to predict under-bark temperature minima of deciduous and coniferous trees.

Populus tremuloides Michx. (trembling aspen) is a tree species native to much of North America, characterized by an excurrent crown with horizontal to ascending branches and a dominant terminal leader. An unusual clone of trembling aspen was discovered in the 1940s near Hafford, Saskatchewan. This clone demonstrates abnormal crown morphology, in which vigorous shoots bend down, ultimately leading to an overall twisted or crooked appearance. The objectives of the present study were to investigate the mechanism of shoot bending by (1) characterizing the process and timing of bending, (2) evaluating structural aspects of developing wild-type and crooked aspen shoots, and (3) comparing anatomical features of bending shoots with wild-type shoots. L-system reconstruction models of 3-D digitized shoot development revealed dramatic bending midway through the growing season. Morphological analyses revealed that crooked aspen shoots had greater taper compared to the wild-type, typically known to create shoots resist deflection and bending. However, preliminary strength analyses indicated that crooked aspen shoots were less rigid, with smaller values of Young’s modulus compared to wild-type shoots. Anatomical investigations revealed differences in several structural tissues between developing wild-type and crooked aspen shoots, and differences within crooked aspen shoots. Primary phloem fibres on the upper side of bending shoots maintained relatively large lumens while those on the lower side were fully lignified, similar to those of mature vertically oriented wild-type leader shoots. These differences may result in differential extension growth early in development, and/or uneven mechanical support later on, ultimately resulting in bending due to self-weight. Gelatinous fibres (G-fibres), characteristic of tension wood (TW), were found throughout older wild-type and vertically oriented crooked aspen shoots; however, G-fibres were only found on the lower side of crooked aspen shoots. These lateral differences could have contributed to shoot bending by actively bending shoots downwards, or lack of TW on the upper side may not have prevented biomechanical bending from self weight. Nevertheless, shoot bending stops at the end of the growing season, suggesting that the mechanisms involved in creating bent shoots are only functional during the first growing season.