A Novel Alkaline α-Galactosidase from Melon Fruit with a Substrate Preference for Raffinose1

The cucurbits translocate the galactosyl-sucrose oligosaccharides raffinose and stachyose, therefore, α-galactosidase (α-d-galactoside galactohydrolase, EC 3.2.1.22) is expected to function as the initial enzyme of photoassimilate catabolism. However, the previously described alkaline α-galactosidase is specific for the tetrasaccharide stachyose, leaving raffinose catabolism in these tissues as an enigma. In this paper we report the partial purification and characterization of three α-galactosidases, including a novel alkaline α-galactosidase (form I) from melon (Cucumis melo) fruit tissue. The form I enzyme showed preferred activity with raffinose and significant activity with stachyose. Other unique characteristics of this enzyme, such as weak product inhibition by galactose (in contrast to the other α-galactosidases, which show stronger product inhibition), also impart physiological significance. Using raffinose and stachyose as substrates in the assays, the activities of the three α-galactosidases (alkaline form I, alkaline form II, and the acid form) were measured at different stages of fruit development. The form I enzyme activity increased during the early stages of ovary development and fruit set, in contrast to the other α-galactosidase enzymes, both of which declined in activity during this period. In the mature, sucrose-accumulating mesocarp, the alkaline form I enzyme was the major α-galactosidase present. We also observed hydrolysis of raffinose at alkaline conditions in enzyme extracts from other cucurbit sink tissues, as well as from young Coleus blumei leaves. Our results suggest different physiological roles for the α-galactosidase forms in the developing cucurbit fruit, and show that the newly discovered enzyme plays a physiologically significant role in photoassimilate partitioning in cucurbit sink tissue.

Modulation of higher-plant NAD(H)-dependent glutamate dehydrogenase activity in transgenic tobacco via alteration of beta subunit levels

Glutamate dehydrogenase (GDH; EC 1.4.1.2-1.4.1.4) catalyses in vitro the reversible amination of 2-oxoglutarate to glutamate. In vascular plants the in vivo direction(s) of the GDH reaction and hence the physiological role(s) of this enzyme remain obscure. A phylogenetic analysis identified two clearly separated groups of higher-plant GDH genes encoding either the alpha- or beta-subunit of the GDH holoenzyme. To help clarify the physiological role(s) of GDH, tobacco (Nicotiana tabacum L.) was transformed with either an antisense or sense copy of a beta-subunit gene, and transgenic plants recovered with between 0.5- and 34-times normal leaf GDH activity. This large modulation of GDH activity (shown to be via alteration of beta-subunit levels) had little effect on leaf ammonium or the leaf free amino acid pool, except that a large increase in GDH activity was associated with a significant decrease in leaf Asp (similar to 51%, P=0.0045). Similarly, plant growth and development were not affected, suggesting that a large modulation of GDH beta-subunit titre does not affect plant viability under the ideal growing conditions employed. Reduction of GDH activity and protein levels in an antisense line was associated with a large increase in transcripts of a beta-subunit gene, suggesting that the reduction in beta-subunit levels might have been due to translational inhibition. In another experiment designed to detect post-translational up-regulation of GDH activity, GDH over-expressing plants were subjected to prolonged dark-stress. GDH activity increased, but this was found to be due more likely to resistance of the GDH protein to stress-induced proteolysis, rather than to post-translational up-regulation.

Developing pineapple fruit has a small transcriptome dominated by metallothionein

In a first step toward understanding the molecular basis of pineapple fruit development, a sequencing project was initiated to survey a range of expressed sequences from green unripe and yellow ripe fruit tissue. A highly abundant metallothionein transcript was identified during library construction, and was estimated to account for up to 50% of all EST library clones. Library clones with metallothionein subtracted were sequenced, and 408 unripe green and 1140 ripe yellow edited EST clone sequences were retrieved. Clone redundancy was high, with the combined 1548 clone sequences clustering into just 634 contigs comprising 191 consensus sequences and 443 singletons. Half of the EST clone sequences clustered within 13.5% and 9.3% of contigs from green unripe and yellow ripe libraries, respectively, indicating that a small subset of genes dominate the majority of the transcriptome. Furthermore, sequence cluster analysis, northern analysis, and functional classification revealed major differences between genes expressed in the unripe green and ripe yellow fruit tissues. Abundant genes identified from the green fruit include a fruit bromelain and a bromelain inhibitor. Abundant genes identified in the yellow fruit library include a MADS box gene, and several genes normally associated with protein synthesis, including homologues of ribosomal L10 and the translation factors SUI1 and eIF5A. Both the green unripe and yellow ripe libraries contained high proportions of clones associated with oxidative stress responses and the detoxification of free radicals.

Effect of pre-bloom leaf defoliation on cluster morphology and disease pressure

Mestrado Vinifera Euromaster - Instituto Superior de Agronomia - UL

Canopy management effects on Syrah grapevines under tropical semi-arid conditions.

The aim of this study was to evaluate the effects of shoot topping and leaf removal practices on vegetative vigor, yield and physicochemical characteristics of the 'Syrah' grape in the semi-arid tropical climate conditions of the São Francisco River Valley in the northeast of Brazil. The experiment was conducted over five growing cycles from 2010 to 2012 in a 'Syrah' commercial vineyard in the municipality of Casa Nova, Bahia, Brazil. Treatments consisted of leaf removal once or twice during fruit set or at the beginning of bunch closure, and one or two shoot toppings of the berry when pea-sized or 10 days after berry growth had commenced. Leaf removal and shoot topping did not affect yield, plant vigor components nor the main attributes of fruit quality. In contrast, there was a seasonal effect with higher yield and better balance between production and vigor in grapes produced in the first half of 2012. The Syrah grape quality was favored in cycles in the first half of the year when its ripening phase coincided with the period of mild temperatures.

Assessment of Injuries Caused by Anastrepha fraterculus (Wied.) (Diptera: Tephritidae) on the Incidence of Bunch Rot Diseases in Table Grape.

Resumo: Anastrepha fraterculus (. Wied) é a principal praga de uvas de mesa (Vitis vinifera) na Região Sul do Brasil. Neste estudo, o objetivo foi investigar o efeito da punção de frutas por fêmeas adultas e infestação larvária por A. fraterculus na ocorrência da doença podridões na uva (cultivar "Itália"). Abstract: Anastrepha fraterculus (Wied.) is the main insect pest of table grapes (Vitis vinifera) in the Southern Region of Brazil. In this study, we aimed to investigate the effect of fruit puncturing by adult females and larval infestation by A. fraterculus on the occurrence of bunch rot disease in the grape (cultivar ?Itália?) by evaluating grapes (a) punctured for oviposition by females of A. fraterculus, sterilized in laboratory with novaluron (40 mg L−1) and further spray-inoculated separately with Botrytis cinerea (1 × 106 conidia mL−1), Glomerella cingulata (1 × 106 conidia mL−1), and bacteria and yeast that cause sour rot (1 × 105 cells mL−1), (b) grapes punctured for oviposition by non-sterilized females with pathogen spraying, (c) grapes with mechanical wounds and pathogen spraying, (d) grapes with no wounds and with pathogen spraying, (e) grapes punctured for oviposition by A. fraterculus chemically sterilized in laboratory with novaluron, (f) grapes punctured for oviposition by A. fraterculus non-sterilized in laboratory with novaluron, (g) grapes with mechanical wounds, and (h) grapes with no sterilization or pathogen spraying. Our data indicated that the mechanical and oviposition wounds caused by A. fraterculus increased the percentage of grapes infected by B. cinerea, G. cingulata, and microorganisms of acid rot. The grape puncturing by A. fraterculus and the mechanical wound allows the penetration of B. cinerea and microorganisms leading to acid rot. We conclude that the fruit fly A. fraterculus may facilitate phytopathogens penetration leading to bunch rots in the table grape Itália.

Desenvolvimento de marcadores para a seleção assistida de ausência de sementes em videiras no programa de melhoramento da Embrapa Uva e Vinho.

desenvolvimento de novas cultivares de uvas sem sementes é uma das prioridades dos programas de melhoramento de uvas de mesa do mundo. Em trabalho anterior o nosso grupo detectou um QTL (quantitative trait locus) para ausência de sementes no cromossomo 18 no locus SDI (seed development inhibitor). Evidências adicionais demonstraram que o gene VvAGL11, localizado neste locus, possui papel fundamental na morfogênese de sementes em videira. O objetivo deste trabalho foi genotipar acessos apirêincos e pirênicos com nove marcadores do tipo SNP e INDEL únicos para o alelo associado a ausência de sementes em Vitis vinifera e verificar se a metodologia de genotipagem baseada em KASP? tem potencial de uso em seleção assistida.

Effect of acaricides on Calepitrimerus vitis (Napela, 1905) (Acari: Eriophyidae) and on the production os vineyards.

2016

Suscetibilidade de bagas de genótipos de videira pela infestação por Drosophila suzukii (Diptera: Drosophilidae).

O objetivo deste trabalho foi avaliar a suscetibilidade de genótipos de videira à drosófila?da?asamanchada, Drosophila suzukii (Diptera: Drosophilidae), em bagas de uva intactas e em bagas infestadas após a ocorrência de puncturas, causadas pela oviposição da mosca?das?frutas sul?americana [Anastrepha fraterculus (Diptera: Tephritidae)] ou por danos mecânicos (alfinetes). Os experimentos foram realizados em laboratório, a 22±1°C, umidade relativa de 65±10% e fotófase de 12 horas. A suscetibilidade foi avaliada para 18 genótipos de videira, em bagas intactas submetidas às fêmeas de D. suzukii. O potencial de interação foi verificado em bagas de uva 'Italia', cuja epiderme foi danificada por puncturas de A. fraterculus ou por alfinete, em comparação a frutos de morango 'Albion'. As cultivares de Vitis labrusca 'Niagara Rosada' e 'Concord' não foram infestadas por D. suzukii, e cinco dos oito genótipos que foram infestados são cultivares melhoradas. A infestação de D. suzukii em bagas de uva 'Italia' com danos mecânicos, feitos com um alfinete ou pela oviposição de A. fraterculus, foi semelhante à de bagas íntegras. Há baixa adequação hospedeira de videiras a D. suzukii, mesmo com a presença de danos. As cultivares 'Benitaka', 'BRS Vitória' e 'BRS Morena' são as mais suscetíveis a D. suzukii.

Potenziali effetti della consociazione tra genotipi di Vitis: osservazioni in vivaio

È necessario intraprendere una via maggiormente ecosostenibile e vantaggiosa per gestire le problematiche presenti in vivaio, e in generale in vigneto, adottando tecniche ammesse in agricoltura biologica o, meglio ancora, con una visione agroecologica della gestione. Le osservazioni condotte presso i Vivai Maiorana hanno indagato la diversa suscettibilità di genotipi di vite ai principali patogeni fungini e gli effetti della consociazione sui sintomi causati da tali patogeni. In primo luogo si è osservata la differenza di suscettibilità tra le cultivars di Merlot e la varietà portinnesto Kober 5BB. I genotipi di Vitis vinifera analizzati si sono dimostrati suscettibili, a differenza del portinnesto Kober 5BB che è risultato completamente esente da sintomi. In seguito, le osservazioni si sono concentrate nel lotto del barbatellaio, in cui tre filari di Calabrese e tre filari di Nerello Mascalese sono separati da un filare di Isabella. Le osservazioni svolte hanno permesso di indagare gli effetti della consociazione, sfruttando l’attuale impostazione del vivaio, in cui diversi genotipi di vite vengono coltivate a stretto contatto. Si è osservata una variazione dei sintomi di peronospora su Vitis vinifera in relazione alla distanza dalla varietà Isabella. L’effetto è presumibilmente mediato da VOCs specifici. Questi benefici, se opportunamente sfruttati, consentirebbero di ridurre sensibilmente la necessità di intervenire con prodotti esogeni nella gestione del barbatellaio, aumentando la possibilità di impostare una produzione biologica che soddisfi le necessità della viticoltura moderna, con vantaggi agronomici ed ecologici.

Impacts of trace element pollution on sewage sludge and soil-plant systems: developing remediation techniques and assessing plant uptake prevention measures

Trace Elements (TEs) pollution is a significant environmental concern due to its toxic effects on human and ecosystem health and its potential to bioaccumulate in the food chain and to threaten species survival, leading to a decline in biodiversity. Urban areas, industrial and mining activities, agricultural practices, all contribute to the release of TEs into the environment posing a significant risk to human health and ecosystems. Several techniques have been developed to control TEs into the environment. This work presents the findings of three-year PhD program that focused on research on TEs pollution. The study discusses three fundamental aspects related to this topic from the perspective of sustainable development, environmental and human health. (1) High levels of TEs contamination prevent the use of sewage sludge (SS) as a fertilizer in agriculture, despite its potential as a soil amendment. Developing effective techniques to manage TEs contamination in SS is critical to ensure its safe use in agriculture and promote resource efficiency through sludge reuse. Another purpose of the study was to evaluate different strategies to limit the TEs uptake by horticultural crops (specifically, Cucumis Melo L.). This study addressed the effect of seasonality, Trichoderma inoculation and clinoptilolite application on chromium (Cr), copper (Cu) and lead (Pb) content of early- and late-ripening cultivars of Cucumis Melo L.. Finally, the accumulation of copper and the effect of its bioavailable fraction on bacterial and fungal communities in the rhizosphere soil of two vineyards, featuring two different varieties of Vitis vinifera grown for varying lengths of time, were evaluated.

Red flesh fruit in european pear (Pyrus communis): genetic sources, QTLs, candidate genes and tools for breeding

Red flesh fruit is a character which interest is increasing in several commercial species. Following a review of the research on the biosynthesis and accumulation of anthocyanin in pears (Chapter 1) the general aim of the project is reported in Chapter 2. Chapter 3 reports the results of a molecular analysis of 33 red-fleshed pear accessions, genotyped with 18 SSR markers with the aim of improving germplasm conservation strategies to support ongoing breeding programs. The molecular profiles revealed both cases of synonymy and homonymy and 6 unique genotypes were identified. The S-allele were established to highlight the genetic relationships among these landraces. Four of the unique genotypes have been clustered based on pomological data. In the Chapter 4, the work was directed to identify the putative genomic regions involved in the appearance of this character in pear fruit. A crossing population (‘Carmen’ x ‘Cocomerina Precoce’) segregating for the trait was phenotyped for 2 consecutive years and used for QTL analysis. A strong QTL was identified in a small genomic region related to the red flesh fruit trait at 27 Mb from the start of LG5. Two candidate genes were detected in this genomic region: ‘PcMYB114’ and ‘PcABCC2’. SSR marker SSR114 was found able to detect the red flesh phenotype segregation in all the red-fleshed pear accessions and segregating progenies tested. Chapter 5 focuses on examining the trend of anthocyanin synthesis and accumulation during the fruit development, from fruit set to ripening time. Three different trials were planned: qPCR and HPLC methods were performed to correlate the genes expression with the anthocyanin accumulation in ‘Cocomerina Precoce’ and six progenies. Total transcriptome sequencing was used to compare the differential genes expression between red and white-fleshed fruit. Chapter 6 reviews and analyses all the earlier study findings while providing new potential future perspectives.

TWO NEW SPIDER MITES (ACARI: TETRANYCHIDAE) FROM BRAZIL: A MONOCERONYCHUS MCGREGOR (BRYOBIINAE) FROM FINGERGRASS AND AN OLIGONYCHUS BERLESE (TETRANYCHINAE) FROM GRAPE

Two new Tetranychidae (Prostigmata) mites are described from Brazil Monoceronychus tchecensis n. sp., a bryobiine collected from weeping fingergrass, Eustachys distichophylla (Lag.) Nees (Poaceae), in the State of Rio Grande do Sul; and Oligonychus fileno n. sp., a tetranychine collected from grape, Vitis vinifera L. (Vitaceae), in the State of Minas Gerais. Monoceronychus tchecensis n. sp. is the second species in this genus described from South America. In addition to the description of these new species, the tetranychine Eotetranychus smithi Pritchard and Baker, 1955 was recorded for the first time for South America.

Phenolic composition of South American red wines classified according to their antioxidant activity, retail price and sensory quality

In this study, 73 South American red wines (Vitis vinifera) from 5 varietals were classified based on sensory quality, retail price and antioxidant activity and characterised in relation to their phenolic composition. ORAC and DPPH assays were assessed to determine the antioxidant activity, and sensory analysis was conducted by seven professional tasters using the Wine Spirits Education Trust`s structured scales. The use of multivariate statistical techniques allowed the identification of wines with the best combination of sensory characteristics, price and antioxidant activity. The most favourable varieties were Malbec, Cabernet Sauvignon, and Syrah produced in Chile and Argentina. Conversely, Pinot Noir wines displayed the lowest sensory characteristics and antioxidant activity. These results suggest that the volatile compounds may be the main substances responsible for differentiating red wines on the basis of sensory evaluation. (C) 2011 Elsevier Ltd. All rights reserved.

Differential display and suppression subtractive hybridization analysis of the pulp of ripening banana

The fruit of banana undergoes several important physico-chemical changes during ripening. Analysis of gene expression would permit identification of important genes and regulatory elements involved in this process. Therefore, transcript profiling of preclimacteric and climacteric fruit was performed using differential display and Suppression subtractive hybridization. Our analyses resulted in the isolation of 12 differentially expressed cDNAs, which were confirmed by dot-blots and northern blots. Among the sequences identified were sequences homologous to plant aquaporins, adenine nucleotide translocator, immunophilin, legumin-like proteins, deoxyguanosine kinase and omega-3 fatty acid desaturase. Some of these cDNAs correspond to newly isolated genes involved in changes related to the respiratory climacteric, or stress-defense responses. Functional characterization of ripening-associated genes could provide information useful in controlling biochemical pathways that would have an impact on banana quality and shelf life. (C) 2009 Elsevier B.V. All rights reserved.