Control of peach phenolic compounds content

Phenolic compounds play a central role in peach fruit colour, flavour and health attributes. Phenolic profiles of several peaches and nectarines and most of the structural genes leading to the anthocyanin synthesis in peach fruit have been studied. Moreover, crosses of red and non-red peaches suggested that a major gene controls skin colour of the extreme phenotypes ‘highlighter’ and ‘full-red’. However, there is no data about either the ‘flavan-3-ols specific genes’ (ANR and LAR) or the regulation of the flavonoid metabolism in this crop. In the present study, we determined the concentration of phenolic compounds in the yellowfleshed nectarine Prunus persica cv. ‘Stark Red Gold’ during fruit growth and ripening. We examined the transcript levels of the main structural genes of the flavonoid pathway. Gene expression of the biosynthetic genes correlated well with the concentration of flavan-3-ols, which was very low at the beginning of fruit development, strongly increased at mid-development and finally decreased again during ripening. In contrast, the only gene transcript which correlated with anthocyanin concentration was PpUFGT, which was high at the beginning and end of fruit growth, remaining low during the other developmental stages. These patterns of gene expression could be explained by the involvement of different transcription factors, which up-regulate anthocyanin biosynthesis (PpMYB10 and PpbHLH3), or repress (PpMYBL2) the transcription of the structural genes. These transcription factors appeared to be involved also in the regulation of the lightinduced anthocyanin accumulation in ‘Stark Red Gold’ nectarines, suggesting that they play a critical role in the regulation of flavonoid biosynthesis in peaches and nectarines in response to both developmental and environmental stimuli. Phenolic profiles and expression patterns of the main flavonoid structural and regulatory genes were also determined for the extreme phenotypes denominated ‘highlighter’ and ‘full-red’ and hypotheses about the control of phenolic compounds content in these fruit are discussed.

Expression profiling in developing peach seed and mesocarp as affected by growth regulators

Jasmonates (JAs) and spermidine (Sd) influence fruit (and seed) development and ripening. In order to unravel their effects in peach fruit, at molecular level, field applications of methyl jasmonate (MJ) and propyl dihydrojasmonate (PDJ), and Sd were performed at an early developmental stage (late S1). At commercial harvest, JA-treated fruit were less ripe than controls. Realtime RT-PCR analyses confirmed a down-regulation of ethylene biosynthetic, perception and signaling genes, and flesh softening-related genes. The expression of cell wall-related genes, of a sugar-transporter and hormone-related transcript levels was also affected by JAs. Seeds from JA-treated fruit showed a shift in the expression of developmental marker genes suggesting that the developmental program was probably slowed down, in agreement with the contention that JAs divert resources from growth to defense. JAs also affected phenolic content and biosynthetic gene expression in the mesocarp. Levels of hydroxycinnamic acids, as well as those of flavan-3-ols, were enhanced, mainly by MJ, in S2. Transcript levels of phenylpropanoid pathway genes were up-regulated by MJ, in agreement with phenolic content. Sd-treated fruits at harvest showed reduced ethylene production and flesh softening. Sd induced a short-term and long-term response patterns in endogenous polyamines. At ripening the up-regulation of the ethylene biosynthetic genes was dramatically counteracted by Sd, leading to a down-regulation of softening-related genes. Hormone-related gene expression was also altered both in the short- and long-term. Gene expression analyses suggest that Sd interfered with fruit development/ripening by interacting with multiple hormonal pathways and that fruit developmental marker gene expression was shifted ahead in accord with a developmental slowing down. 24-Epibrassinolide was applied to Flaminia peaches under field conditions early (S1) or later (S3) during development. Preliminary results showed that, at harvest, treated fruit tended to be larger and less mature though quality parameters did not change relative to controls.

Antimicrobial activity of peach and grapevine defensins

Antimicrobial peptides (AMPs) are an important component of the innate immune system of the plants. Plant defensins are a large family of antimicrobial peptides with several interesting features, such as small dimension, high stability and broad spectrum of action. The discovery of new molecules and the study of their mechanism of action allow to consider them attractive for biotechnological applications. In this PhD thesis a defensin from Prunus persica (PpDFN1) and four novel DEFensin Like (DEFL) peptides from Vitis vinifera have been studied. In order to characterize the antimicrobial activity of these molecules, the recombinant mature peptides have been expressed in Escherichia coli and purified to homogeneity by chromatography techniques. PpDFN1 is able to inhibit the growth of B. cinerea, P. expansum and M. laxa with different intensity. The recombinant peptide is capable of membrane permeabilization as demonstrated by SYTOX green fluorescence uptake in treated mycelia. Its interaction with membranes containing sphingolipid species has been shown by artificial lipid monolayers. Furthermore, PpDFN1 displays stronger interaction with monolayers composed by lipids extracted from sensitive fungi with the highest interaction against P. expansum, the most sensitive fungi to PpDFN1 action. DEFL 13, a defensin from grapevine, resulted the strongest antibotrytis peptides. It is electrostatically attracted to the fungal membranes as shown by the antagonist effect of the cations and is able to membrane permeabilization in B. cinerea hyphae. DEFL 13 is internalized in fungal cells and leads to fungal death by activation of some signaling pathways as demonstrated by screening of a mutant collection of B. cinerea

Modeling systems and vis/NIR device to improve peach and nectarine pre and post-harvest fruit maturity management

During the last decade peach and nectarine fruit have lost considerable market share, due to increased consumer dissatisfaction with quality at retail markets. This is mainly due to harvesting of too immature fruit and high ripening heterogeneity. The main problem is that the traditional used maturity indexes are not able to objectively detect fruit maturity stage, neither the variability present in the field, leading to a difficult post-harvest management of the product and to high fruit losses. To assess more precisely the fruit ripening other techniques and devices can be used. Recently, a new non-destructive maturity index, based on the vis-NIR technology, the Index of Absorbance Difference (IAD), that correlates with fruit degreening and ethylene production, was introduced and the IAD was used to study peach and nectarine fruit ripening from the “field to the fork”. In order to choose the best techniques to improve fruit quality, a detailed description of the tree structure, of fruit distribution and ripening evolution on the tree was faced. More in details, an architectural model (PlantToon®) was used to design the tree structure and the IAD was applied to characterize the maturity stage of each fruit. Their combined use provided an objective and precise evaluation of the fruit ripening variability, related to different training systems, crop load, fruit exposure and internal temperature. Based on simple field assessment of fruit maturity (as IAD) and growth, a model for an early prediction of harvest date and yield, was developed and validated. The relationship between the non-destructive maturity IAD, and the fruit shelf-life, was also confirmed. Finally the obtained results were validated by consumer test: the fruit sorted in different maturity classes obtained a different consumer acceptance. The improved knowledge, leaded to an innovative management of peach and nectarine fruit, from “field to market”.

Life Cycle Assessment of Peach Nectar: a comparative analysis between conventional and bioenergy-from-waste integrated food chains

Modern food systems are characterized by a high energy intensity as well as by the production of large amounts of waste, residuals and food losses. This inefficiency presents major consequences, in terms of GHG emissions, waste disposal, and natural resource depletion. The research hypothesis is that residual biomass material could contribute to the energetic needs of food systems, if recovered as an integrated renewable energy source (RES), leading to a sensitive reduction of the impacts of food systems, primarily in terms of fossil fuel consumption and GHG emissions. In order to assess these effects, a comparative life cycle assessment (LCA) has been conducted to compare two different food systems: a fossil fuel-based system and an integrated system with the use of residual as RES for self-consumption. The food product under analysis has been the peach nectar, from cultivation to end-of-life. The aim of this LCA is twofold. On one hand, it allows an evaluation of the energy inefficiencies related to agro-food waste. On the other hand, it illustrates how the integration of bioenergy into food systems could effectively contribute to reduce this inefficiency. Data about inputs and waste generated has been collected mainly through literature review and databases. Energy balance, GHG emissions (Global Warming Potential) and waste generation have been analyzed in order to identify the relative requirements and contribution of the different segments. An evaluation of the energy “loss” through the different categories of waste allowed to provide details about the consequences associated with its management and/or disposal. Results should provide an insight of the impacts associated with inefficiencies within food systems. The comparison provides a measure of the potential reuse of wasted biomass and the amount of energy recoverable, that could represent a first step for the formulation of specific policies on the integration of bioenergies for self-consumption.

Phenotype and genotype characterization of Monilinia spp. isolates and preformed antifungal compounds in peach peel fruit at different developmental stages

The brown rot fungi belong to a group of fungal pathogens that causes considerable damage to cultivated fruits trees, particularly stone fruits and apples in the temperate regions of the World and during the postharvest with an important economic impact. In particular in Italy, it is important to monitor the Monilinia population to control economic losses associated to the peach and nectarine market. This motivates the research steps presented in this dissertation on Monilinia Italian isolates. The Monilinia species collected from stone fruits have been identified using molecular analysis based on specific primers. The relevant role of M. fructicola was confirmed and, for the first time, it was found also on apple fruits. To avoid the development of resistant strains and implement valid treatment strategies, the understanding of the fruit natural resistance during different developmental stages and the assessment of the Monilinia sensitivity/resistance to fungicides are required. The relationship between the inhibition spots and the phenolic compounds in peach fruit peel was highlighted in this research. Three methods were used to assess isolate resistance/sensitivity, the amended medium, the Spiral Gradient Endpoint Method (SGD) and the Alamar Blue method. The PCR was used to find possible mutation points in the b-tubulin gene that is responsible for fungicide resistance. Interestingly, no mutation points were observed in resistant M. laxa isolates, suggesting that the resistance could be stimulated by environmental factors. This lead to the study of the effect of the temperature on the resistance and the preliminary results of in vitro tests showed that maximum inhibition was observed at 30°C.

Characterization of the genetic control of fruit flesh color in peach

The analysis of a carotenoid cleavage dioxygenase gene in a pool of peach cultivars revealed the existence of a functional allele (W1), associated with the white flesh trait, and three independent mutations associated with the yellow phenotype: a 2 bp insertion within a repetitive sequence (y1), a large transposable element within the intron (y2) and a single base substitution generating a premature stop codon (y3). Based on these evidences, the yellow flesh phenotype seems to have arisen from at least three independent mutational events.

Leaf level VOC emissions of single plants from Amazonian and Mediterranean ecosystems: Ontogeny and flooding as stress factor for VOC emissions

Die Vegetation ist die wichtigste Quelle von organischen flüchtigen Verbindungen (auf Englisch volatile organic compounds,VOCs), die einen bemerkenswerten Einfluss auf der Chemie und Physik der Atmosphäre haben. VOCs beeinflussen die oxidative Kapazität der Atmosphäre und tragen zu der Bildung und zum Wachstum von sekundären organischen Aerosolen bei, welche einerseits eine Streuung und Reflektierung der Energie verursachen und andererseits sich an der Bildung und Entwicklung von Wolken beteiligen. Ziel dieser Arbeit war die Beschreibung und der Vergleich von VOC Emissionen aus Pflanzen aus zwei verschiedenen Ökosystemen: Mediterranes Ökosystem und Tropisches Ökosystem. Für diese Aufgabe wurden gewöhnliche Pflanzen von beiden Ökosystemen untersucht. Siebzehn Pflanzenspezies aus der Mittelmeergebiet, welches bekannt ist für seine Vielfalt an VOC emittierenden Pflanzen, wurden in die Untersuchungen einbezogen. Im Gegensatz zum mediterranen Ökosystem sind nur wenig Information verfügbar über VOC Emissionen aus Blättern tropischer Baumspezies. Vor diesem Hintergrund wurden sechsundzwanzig Baumspezies aus verschiedenen Ökotypen des Amazonasbeckens (Terra firme, Várzea und Igapó) wurden auf VOC Emissionen auf Blattebene mit einem Küvetten-System untersucht. Analysen von flüchtigen organischen Verbindungen wurden online mit PTR-MS und offline mittels Sammlung auf entsprechenden Adsorbern (Kartuschen) und nachfolgender GC-FID Analyse untersucht. Die höchsten Emissionen wurden für Isoprene beobachtete, gefolgt durch Monoterpene, Methanol und Aceton. Die meisten Mittelmeer Spezies emittierten eine hohe Vielfalt an Monoterpenspezies, hingegen zeigten nur fünf tropische Pflanzenspezies eine Monoterpene mit einen sehr konservativen Emissionsprofil (α-Pinen>Limonen>Sabinen >ß-Pinen). Mittelmeerpflanzen zeigten zusätzlich Emissionen von Sesquiterpenen, während bei der Pflanzen des Amazonas Beckens keine Sesquiterpenemissionen gefunden wurden. Dieser letzte Befund könnte aber auch durch eine niedrigere Sensitivität des Messsystems während der Arbeiten im Amazonasgebiet erklärt werden. Zusätzlich zu den Isoprenoidemissionen waren Methanolemissionen als Indikator für Wachtumsvorgänge sehr verbreitet in den meisten Pflanzenspezies aus tropischen und mediterranen Gebieten. Einige Pflanzenspezies beider Ökosystemen zeigten Acetonemissionen. rnrnVOC Emissionen werde durch eine große Vielfalt an biotischen und abiotischen Faktoren wie Lichtintensität, Temperatur, CO2 und Trockenheit beeinflusst. Ein anderer, öfter übersehener Faktor, der aber sehr wichtig ist für das Amazonas Becken, ist die regelmäßige Überflutung. In dieser Untersuchung wir fanden heraus, dass am Anfang einer Wurzelanoxie, die durch die Überflutung verursacht wurde, Ethanol und Acetaldehyd emittiert werden können, vor allem in Pflanzenspezies, die schlechter an eine unzureichende Sauerstoffversorgung bei Flutung adaptiert sind, wie z.B. Vatairea guianensis. Die Spezies Hevea spruceana, welche besser an Überflutung adaptiert ist, könnte möglicherweise der gebildete Ethanol sofort remetabolisieren ohne es zu emittieren. Nach einer langen Periode einer Überflutung konnte allerdings keine Emission mehr beobachtet werden, was auf eine vollständige Adaptation mit zunehmender Dauer schließen lässt. Als Reaktion auf den ausgelösten Stress können Isoprenoidemissionen ebenfalls kurzfristig nach einigen Tage an Überflutung zunehmen, fallen dann aber dann nach einer langen Periode zusammen mit der Photosynthese, Transpiration und stomatäre Leitfähigkeit deutlich ab.rnrnPflanzen Ontogenese ist anscheinend von Bedeutung für die Qualität und Quantität von VOC Emissionen. Aus diesem Grund wurden junge und erwachsene Blätter einiger gut charakterisierten Pflanzen Spezies aus dem Mittelmeerraum auf VOC Emissionen untersucht. Standard Emissionsfaktoren von Isopren waren niedriger in jungen Blättern als in erwachsene Blätter. Hingegen wurden höhere Monoterpen- und Sesquiterpenemissionen in jungen Blätter einiger Pflanzenspezies gefunden. Dieser Befund deutet auf eine potentielle Rolle dieser VOCs als Abwehrkomponenten gegen Pflanzenfresser oder Pathogene bei jungen Blätter hin. In einigen Fällen variierte auch die Zusammensetzung der Monoterpen- und Sesquiterpenspezies bei jungen und erwachsenen Blättern. Methanolemissionen waren, wie erwartet, höher in jungen Blättern als in ausgewachsenen Blättern, was mit der Demethylierung von Pectin bei der Zellwandreifung erklärt werden kann. Diese Befunde zu Änderungen der Emissionskapazität der Vegetation können für zukünftige Modellierungen herangezogen werden. rn

The Arabidopsis PHYTOCHROME KINASE SUBSTRATE2 protein is a phototropin signaling element that regulates leaf flattening and leaf positioning

In Arabidopsis (Arabidopsis thaliana), the blue light photoreceptor phototropins (phot1 and phot2) fine-tune the photosynthetic status of the plant by controlling several important adaptive processes in response to environmental light variations. These processes include stem and petiole phototropism (leaf positioning), leaf flattening, stomatal opening, and chloroplast movements. The PHYTOCHROME KINASE SUBSTRATE (PKS) protein family comprises four members in Arabidopsis (PKS1-PKS4). PKS1 is a novel phot1 signaling element during phototropism, as it interacts with phot1 and the important signaling element NONPHOTOTROPIC HYPOCOTYL3 (NPH3) and is required for normal phot1-mediated phototropism. In this study, we have analyzed more globally the role of three PKS members (PKS1, PKS2, and PKS4). Systematic analysis of mutants reveals that PKS2 (and to a lesser extent PKS1) act in the same subset of phototropin-controlled responses as NPH3, namely leaf flattening and positioning. PKS1, PKS2, and NPH3 coimmunoprecipitate with both phot1-green fluorescent protein and phot2-green fluorescent protein in leaf extracts. Genetic experiments position PKS2 within phot1 and phot2 pathways controlling leaf positioning and leaf flattening, respectively. NPH3 can act in both phot1 and phot2 pathways, and synergistic interactions observed between pks2 and nph3 mutants suggest complementary roles of PKS2 and NPH3 during phototropin signaling. Finally, several observations further suggest that PKS2 may regulate leaf flattening and positioning by controlling auxin homeostasis. Together with previous findings, our results indicate that the PKS proteins represent an important family of phototropin signaling proteins.

Monte Carlo implementation, validation, and characterization of a 120 leaf MLC

Recently, the new high definition multileaf collimator (HD120 MLC) was commercialized by Varian Medical Systems providing high resolution in the center section of the treatment field. The aim of this work is to investigate the characteristics of the HD120 MLC using Monte Carlo (MC) methods.

Leaf press juice from Bryophyllum pinnatum (Lamarck) Oken induces myometrial relaxation

The use of preparations from Bryophyllum pinnatum (Lamarck) Oken (Kalanchoe pinnata (Lamarck) Persoon) in tocolysis is supported by clinical evidence. We studied here the effect of B. pinnatum leaf press juice and its chemical fractions on the response of human myometrial strips. No data are available if the influence on myometrial strips of the juice differs from that of its components in the chemical fractions, in order to increase the pharmacological effect.