Water use efficiency (WUE), measured as the ratio of plant biomass to water consumption, is an essential agronomical trait for enhancing crop production under drought. Measuring water consumption is logistically difficult, especially in field conditions. The general objective of the present Thesis is to respond to three main questions: (i) can WUE be determined by using carbon isotope discrimination (CID), easy to measure?, (ii) how WUE and CID variation analysis can contribute to the genotypic selection of sunflower subjected to drought?, and (iii) can WUE variation be revealed by the variation of plant-water relation traits. Four experiments were carried out in greenhouse across two different years: (i) on two drought scenarios, progressive soil drying and stable water-stress, and (ii) on five levels of soil water content. The main traits that have been measured include WUE, CID, as well as plant-water relation traits, i.e. control of transpiration (FTSWt), water extraction capacity (TTSW), and dehydration tolerance (OA). A highly significant negative correlation was observed between WUE and CID, and a wide phenotypic variability was observed for both WUE and CID. A wide variability was also observed for FTSWt, TTSW and OA. The results provide new insight into the genetic control of WUE and CID related-traits, which, unlike to other crops, genetic control of WUE, CID, and TTSW in sunflower have never been reported in the literature. Further, quantitative trait loci (QTL) mapping for FTSWt was never reported in any plant species. The QTL for WUE and CID were identified across different drought scenarios. The QTL for CID is considered as a ‘‘constitutive’’ QTL, because it is consistently detected across different drought scenarios. The QTL for CID co-localized with the QTL for WUE, biomass and cumulative water transpired. Co-localization was also observed between the QTL for FTSWt and TTSW, between the QTL for TTSW and WUE-CID-biomass, as well as between the QTL for FTSWt-TTSW and biomass. This study highlights that WUE is physiologically and genetically associated with CID. CID is an excellent surrogate for WUE measurement, and can be used to improve WUE by using marker-assisted selection (MAS) to achieve the ultimate goal of plant breeding at genomic level.

The goal of this project was to determine the genetic architecture of water-use efficiency (WUE) for foxtail pine, which included genomic loci, and effect sizes of this trait. Foxtail pine is a subalpine endemic conifer that inhabits two distinct regional populations separated by 500 km in the mountains of California. In order to achieve this goal, a robust linkage map containing thousands of genetic markers was created using four megagametophyte arrays ranging in size from approximately 70 to 95 megagametophytes. Quantitative trait loci (QTL) discovered for WUE were mapped along the linkage map using linear mixed models and five half-sibling families grown in a common garden. Effect sizes of these QTL were tested for differences between the two regional populations of foxtail pine.

The physiological function of the plant enzyme alternative oxidase has long been a topic of debate. The cyanide-resistant alternative oxidase (AOX), along with the cytochrome c oxidase (COX), catalyzes the reduction of oxygen to water in the electron transport chain of mitochondrial respiration. Although respiration via the alternative pathway (AP) results in approximately one third of the ATP production as respiration via the cytochrome pathway (CP), the AP is utilized by all plants and some fungi and animals. This “energy wasteful” pathway has been proposed to reduce oxidative stress in plant cells under a variety of stressful conditions. Virtually all previous work on the AP has been performed on laboratory-grown plants in controlled environment conditions; thus, there is little knowledge of how the AP responds to unstable conditions and multiple environmental stresses in the field. This thesis presents new methodology for studying AP respiration and the AOX protein in field-grown plants, and investigates how the AP responds to natural changes in environmental conditions in the field in several plant species grown in diverse ecosystem types. The experimental work presented here also investigates how AP activity is related to changes in total rates of respiration, and questions whether abundance of the AOX protein determines electron partitioning to the AP. AP partitioning (or relative changes in AP partitioning) varied over seasonal timescales in each of the experimental studies. Chapter 3 reports on two species of Chionochloa, a native New Zealand tussock grass growing along an altitudinal gradient. In Chapter 4, seasonal variation was studied in two tree types: Populus x canadensis, a deciduous angiosperm, and Pinus radiata, an evergreen gymnosperm. Quercus rubra trees were studied along an urban-rural gradient originating in New York City in Chapter 5. In a highly exposed and variable environment, relative changes in AP partitioning in two species of Chionochloa were correlated with the previous day’s integrated light. In Quercus rubra, the AP was instead related to temperature changes: relative AP partitioning increased in response to seasonally low temperature in trees grown at colder, more rural field sites, while at the warmer, urban sites, it increased in response to high summer temperatures. Each of these environmental conditions that were related to increases in the AP (high light, low temperatures, and heat) are potentially stressful to plants. Thus, it is possible that the increases in AP respiration observed in these studies served to oxidize excess reducing equivalents generated through stressful conditions. In Chapter 4, although AP partitioning in Populus x canadensis and Pinus radiata varied seasonally, these changes were not directly related to environmental parameters. However, AP partitioning in Populus x canadensis was clearly shown to be dependent on measurement temperature. In each of the studies presented here, changes in the AP were not related to abundance of the AOX protein. AOX protein abundance showed consistent seasonal patterns in the two deciduous angiosperms, Populus sp. and Quercus sp, and was correlated with seasonal changes in temperature in Chionochloa spp. However, the lack of correlation between protein abundance and AP partitioning indicates that the AP is subject to post-translational control and likely varies more rapidly than protein levels. In each of Chapters 3 – 5, there was no clear impact of changes in AP partitioning on rates of total respiration. As the AP produces less ATP than the CP, I hypothesized that increases in AP activity would lead to higher respiration rates in order to meet a plant’s energy demands. However, in Populus x canadensis and Quercus rubra, respiration rates remained stable during sharp increases in AP partitioning, indicating that, at least under certain conditions, increases in AP activity are accompanied by a decrease in the CP. In some of the first research studying AP partitioning in field-grown plants, this thesis illuminates possible mechanisms, functions, and implications of the AP. Over a range of plant taxa and environmental settings, this work shows that the AP does respond to stressful conditions in the wild, but that this does not result in increased respiration. Lastly, the methods presented here to study AP activity and AOX proteins in the field enable future studies to further probe the specific responses of AOX to natural stresses.

Cette thèse visait (i) à quantifier la variabilité de l’efficience d’utilisation de l’eau (WUE) chez le peuplier noir (Populus nigra L.), (ii) à juger de sa plasticité phénotypique en réponse à des contraintes hydrique et thermique et (iii) à juger des liens entre WUE, croissance et survie. A cette fin, des études à partir de semis, en chambre de croissance et en milieu naturel, ainsi qu’à partir de populations installées en dispositifs de pépinière ont été réalisées. Nos travaux ont permis de montrer une importante variabilité et une importante plasticité du caractère ‘efficience d’utilisation de l’eau’ quels que soient l’échelle d’étude, le fond génétique et les conditions de croissance. Une diminution de WUE a été observée en réponse à la contrainte thermique alors qu’une augmentation de WUE a été observée en réponse à un déficit hydrique modéré. Nos travaux démontrent également un lien entre WUE et la survie sous de fortes températures suggérant que sous ces conditions, une faible efficience d’utilisation de l’eau conférerait un avantage pour les individus. De plus in situ, un lien négatif entre WUE et la densité de régénération a été observé démontrant que les plus faibles valeurs de WUE sont observées pour les plus fortes densités. Le caractère ‘efficience d’utilisation de l’eau’ pourrait donc s’avérer un caractère limitant de la régénération de l’espèce dans des conditions d’augmentation de température ; une telle augmentation pourrait alors avoir une conséquence directe sur la structuration génétique des populations futures. Ces travaux suggèrent enfin une structuration géographique de la variabilité de WUE qui reste à confirmer à partir d’un plus grand nombre de populations. Ce travail ouvre des perspectives intéressantes pour l’identification des bases physiologiques à l’origine des variations de WUE, de sa plasticité et de sa structuration géographique. / This work aimed (i) to quantify variability of water-use efficiency (WUE) on black poplar (Populus nigra L.), (ii) to quantify the phenotypic plasticity in response to water deficit and increase of temperature and (iii) to precise the relationships between WUE, growth and leaf survival. Studies on seedlings in growth chambers and in natural conditions, and on populations established in common garden studies were realized. Significant variations and significant plasticity of water-use efficiency were observed on seedlings and adults whatever growth conditions and genetic background. A significant decrease of WUE was observed in response to increase of temperature while an increase of WUE was observed in response to moderate water deficit. Under elevated temperature a significant relationship was observed between WUE and leaf survival suggesting that low WUE confers an advantage for individuals under these conditions. Moreover in situ, a negative relationship between WUE and seedlings density was detected suggesting that low WUE are observed under high seedlings densities. The complex trait ‘water-use efficiency’ should be a limiting character of species establishment under increase of temperatures; this temperature elevation should have consequences on genetic structuration of populations in the future. Our results suggest a geographical structuration of WUE variability but results must be confirmed on several others populations. This present work opens prospects for identification of physiological bases of WUE variations, its plasticity and its geographical structuration.

Perennial ryegrass (Lolium perenne L.) is a cool-season turfgrass susceptible to low temperature injury. Wilt-based (WB) irrigation is a common practice in scheduling turf irrigation as an alternative to well-watered (WW). Moisture stress has been shown to promote cold hardiness but this has not been investigated in response to WB irrigation. Measurements of 13C isotope discrimination (DELTA) are useful predictors of water use efficiency (WUE), drought resistance, evapotranspiration (ET) and salinity tolerance but the relevance to turfgrass cold hardiness has not been determined. DELTA analyses may enable more efficient screening protocols in breeding for improved cold hardiness. Objectives of this study were to examine perennial ryegrass genotypes in relation to cold hardiness, DELTA and WUE in response to WW and WB irrigation schedules, to compare genetic diversity between top-performing (TP) and bottom-performing (BP) perennial ryegrass genotypes, and to assess the predictive value of DELTA of for cold hardiness. Six genotypes were selected based on turf quality from the most northern NTEP location (Orono, ME) and included three TP (‘All Star 2’, ‘Mach I’ and ‘Sunkissed’) and three BP (‘APR-1234’, ‘Buccaneer’ and ‘WVPB-R-82’) genotypes. ET, yield, WUE, shoot water content, rooting potential, wilting tendency, DELTA and median lethal temperatures (LT50) using whole-plant survival were measured from greenhouse samples grown in weighing lysimeters in 2007 and 2008. Plant measurements in both years were based on sampling conducted at the last cycle after 68-d of irrigation with 100% of ET applied at leaf-roll (WB) versus ET replacement every 4-d (WW). Lower LT50 values were generally associated with low yield, low WUE and low shoot water content, whether the result of irrigation treatment or genotypic variation. TP genotypes demonstrated significantly lower LT50 temperatures (greater cold hardiness) in comparison to BP genotypes in both years. Modest cold hardiness enhancement with WB irrigation was highest for TP genotypes. Wilting tendency and DELTA were not reliable predictors of cold hardiness, although individual TP genotypes exhibited responses distinctly different than some BP genotypes. Further research is needed to investigate the physiological mechanisms of enhanced turfgrass cold hardiness in response to moisture stress.

No description available.

Varietal forestry may increase the productivity of loblolly pine (Pinus taeda L.) in the Southern United Sates. However, the effects of these genetic x environment interactions are still poorly understood. In this study we examined the responses in growth, stand uniformity and leaf level physiology of loblolly pine clonal varieties and families to silvicultural intensity and site effects. We also looked for patterns in observed traits that were consistent between crown ideotypes. Two varieties of each crown ideotype (narrow vs broad crowns) and two families (controlled mass pollinated (CMP) and open pollinated (OP) family) were tested on the Virginia Piedmont (VA) and the North Carolina Coastal Plain (NC) under different silvicultural intensities (operational vs intensive), and planting density (617, 1235 and 1852 trees per hectare). Data were collected during the first four growing season after establishment. At NC, intensive silviculture increased crown-width, height and dbh by 33%, 14%, and 23%, respectively. At VA, intensive silviculture increased crown-width, height and dbh by 41%, 10%, and 23%, respectively. Intensive silviculture also increased slightly but significantly the stand uniformity of stem growth. However, the differences in productivity between silvicultural treatments were not explained by differences in leaf-level physiology. Across all treatments and sites the varieties generally grew faster than the OP family, but the differences were higher at VA. Varieties did not differ in stem growth, but the broadest crown variety had greater stand uniformity, photosynthetic rate (Asat), carbon isotope discrimination (∆¹³) and lower fascicle size than the OP family. None of the traits assessed inthis study was consistent within the ideotypes. Varieties classified in the same crown-ideotypes may respond differently to the environmental effects of site and silviculture, which reinforces the need of matching varietal forestry with precision silviculture to achieve gains in productivity. / Ph. D.

Resumo: Artemisia annua L. (Asteraceae) é uma planta herbácea altamente aromática, nativa da Ásia e aclimatada no Brasil. As folhas são fonte abundante de artemisinina, uma lactona sesquiterpênica que, conjuntamente aos seus derivados semi-sintéticos, apresentam ação efetiva contra as cepas resistentes das espécies de Plasmodium causadoras da malária. Os objetivos deste trabalho foram identificar o mecanismo fotossintético e avaliar o efeito de diferentes condições de temperatura e fotoperíodo no crescimento e desenvolvimento do acesso CPQBA 2/39x1V de A. annua. Para identificar o mecanismo fotossintético foram realizados experimentos para determinar composição dos isótopos do carbono (d‰13C) e a anatomia foliar associada a testes histoquímicos. Para avaliar o efeito da temperatura e fotoperíodo no crescimento e desenvolvimento de A. annua foram realizados dois experimentos em câmaras fotoperiódicas (o primeiro com temperaturas médias máx. de 37ºC e mín. de 19ºC e o segundo com máx. de 29ºC e mín. de 13ºC) e um experimento com 6 épocas de plantio em campo no município de Pato Branco-PR (26º11' S, 52º36' W e 760 m de altitude), sul do Brasil. Um último experimento realizado, foi a aplicação exógena de artemisinina em plantas de A. annua para verificar o papel da molécula na indução do florescimento. Como resultados, A. annua apresentou uma d‰13C - 31.76 l 0.07, valor típico de espécies com mecanismo fotossintético C3, que apresentam em média valores de d‰13C - 28. Os estudos da anatomia foliar e testes histoquímicos confirmaram os resultados encontrados para a d‰13C, onde, a despeito da existência de células parenquimáticas formando um anel ao redor do feixe vascular, estas não apresentaram cloroplastos e amido, confirmando ser A. annua uma espécie de mecanismo... / Abstract: Leaves of Artemisia annua L. are a plentiful source of artemisinin, a drug with proven effectiveness against malaria. One of the objectives of this work was to identify the photosynthetic type of A. annua through studies of the carbon isotope composition (d‰13C) and the leaf anatomy. We also verified the growth and development of the CPQBA 2/39x1V accession under moist subtropical climate. Two experiments were carried out in greenhouse using photoperiodic chambers. The plants were submitted to photoperiods of 7, 9, 11, 13, 15 and 17 h, in two natural conditions of temperature: spring/summer (maximum of 37ºC and minimum of 19ºC) and autumn/winter (maximum of 29ºC and minimum of 13ºC). Another experiment with different planting dates at field was carried out in Pato Branco, PR (26º11' S, 52º36' W and 760 m), Southern part of Brazil. The last experiment was the application of artemisinin (0, 500, 5000, and 10000 mg L-1) in A. annua plants to verify the role of the molecule on the flowering induction. A. annua presented a d‰13C - 31.76 l 0.07, what characterizes it as typical species with a C3 photosynthetic mechanism, with an average of d‰13C- 28, while C4 species possess an average of d‰13C - 14. The study of A. annua leaf anatomy confirms the results of d‰13C, where, in spite of the existence of parenchymatic cells forming a different sheath surrounding of the vascular tissue, these cells do not have chloroplast or starch. These features do not describe the Kranz anatomy typical of C4 species. The application of artemisinin did not induce the flowering of A. annua at any of the concentrations used. The results suggest that the accumulation of the artemisinin in the pre-flowering and flowering phases in A. annua is not the physiological signal of floral induction in this species... / Orientador: Fernando Broetto / Coorientador: Rita Maria Moraes / Banca: João Domingos Rodrigues / Banca: Lin Chau Ming / Banca: Cícero Deschamps / Banca: Vera Lúcia Garcia Rehder / Doutor

No description available.

C4 plants have a biochemical carbon concentrating mechanism (CCM) that increases CO2 concentration around Rubisco in the bundle sheath (BS). Maize CCM has two CO2 delivery pathways to the Bundle Sheath (BS) (respectively via malate, MAL or aspartate, ASP); rates of PGA reduction, carbohydrate synthesis and PEP regeneration vary between BS and Mesophyll (M) cells. For these anatomical and biochemical complexities, C4 plants are highly sensitive to light conditions. Under limiting light, the activity of the CCM generally decreases, causing an increase in leakiness, (Φ), the ratio of CO2 retrodiffusing from the BS relative to C4 carboxylation processes. This increase in Φ had been theoretically associated with a decrease in biochemical operating efficiency (expressed as ATP cost of gross assimilation, ATP / GA) under low light and, because a proportion of canopy photosynthesis is carried out by shaded leaves, to potential productivity losses at field scale. In C4 leaves, because of the concentric anatomy, light reaches M cells before the deeper BS (Evans et al., 2007), and could alter the energetic partitioning balance between BS and M and potentially cause efficiency losses. In this experimental programme I investigated strategies deployed by C4 plants to adjust operating efficiency under different illumination conditions. Firstly, maize plants were grown under high and low light regimes (respectively HL, 600 vs LL, 100 μE m-2 s-1). Short term acclimation of Φ was compared from isotopic discrimination (Δ), gas exchange and photochemistry using an improved modelling approach which does not suffer from elements of circularity. Long term acclimation to low light intensities brought about physiological changes which could potentially increase the operating efficiency under limiting ATP supplies. Secondly, profiles of light penetration across a leaf were used to derive the potential ATP supply for M and BS cells induced by changing light quality. Empirical measurements of net CO2 uptake, ATP production rate and carbon isotope discrimination were made on plants under a low light intensity. The overall conversion efficiency was not affected by light quality. A comprehensive metabolic model highlighted the importance of both CO2 delivery pathways in maize. Further, metabolic plasticity allowed the balancing of ATP and NADPH requirements between BS and M. Finally, I tested the hypothesis that plants can modify their physiology so as to reach a status of higher operating efficiency when exposed to high light and then to low light, so as to mimic the transition which leaves undergo when shaded by newly emerging leaves in a crop canopy. Plants were grown under high light and low light for three weeks, then, HL plants were transferred to low light for a further three weeks. Re-acclimation was very effective in reducing ATP cost of net assimilation under low light intensities. In addition, the hyperbolic leakiness increase observed under low light intensities was not associated to operating efficiency loss. Overall, in the three experimental Chapters I showed compelling theoretical and empirical evidence proving the hypothesis that C4 plants deal with low light conditions and with different light qualities without losing operating efficiency.