Expressão transiente do gene gus, sob regulação de quatro promotores, em diferentes tecidos de mamoeiro (Carica papaya L.) e videira (Vitis sp.)

O mamoeiro (Carica papaya L.) e a videira (Vitis vinifera L.) destacam-se entre as fruteiras produzidas no Brasil por serem plantadas em quase todo o território nacional e apresentarem importância econômica e social. A tecnologia de produção de organismos geneticamente modificados, também conhecidos como "transgênicos", tem grande potencial de uso no desenvolvimento de fruteiras melhoradas. Porém, questões de propriedade intelectual limitam o uso da engenharia genética por países em desenvolvimento, que normalmente não detêm direitos sobre processos ou produtos necessários ao uso desta. Neste contexto, o presente estudo buscou avaliar promotores de expressão gênica alternativos ao CaMV 35S, que é o mais utilizado no desenvolvimento de transgênicos, mas é patenteado. Para tanto, construções gênicas com o gene gus sob a regulação de diferentes promotores foram testadas para expressão transiente em diversos tecidos de mamoeiro e videira. Expressão transiente foi avaliada em embriões somáticos, folhas, caules, raízes e frutos. O promotor do gene UBQ3, que é constitutivo e se encontra em domínio publico, mostrou ser uma alternativa promissora para futuros trabalhos de transformação genética de mamoeiro, mas não de videira.

Efeito de agrotóxicos sobre Calepitrimerus vitis (Nalepa, 1905) (Acari: Eriophyidae) na região da campanha do Rio Grande do Sul

O Rio Grande do Sul apresenta a maior área de cultivo de videiras no Brasil; neste Estado, a Região da Campanha possui características edafoclimáticas adequadas à produção de cultivares viníferas de origem europeia e apresentou, na primeira década deste milênio, significativa expansão de área. A partir de 2005, o ácaro-da-ferrugem-da-videira, Calepitrimerus vitis, passou a ser encontrado com frequência nos vinhedos da Campanha. Nos países onde esta espécie está estabelecida, reduções significativas na produção de uva são observadas, e a necessidade de controle é constante, não havendo, até o momento, produtos autorizados no Brasil para o controle de C. vitis. O presente trabalho objetivou avaliar diversos agrotóxicos quanto à eficiência de controle de C. vitis em um vinhedo comercial, na região da Campanha do Rio Grande do Sul. O experimento foi conduzido durante os anos de 2008 e 2009, comparando o nível de infestação nas plantas através do emprego de armadilhas adesivas. No ano de 2008, a eficiência de controle durante o outono não diferiu da testemunha. O emprego de enxofre em uma única pulverização no outono, ou em duas pulverizações, sendo uma no outono e outra na primavera, controlou eficientemente C. vitis na primavera. Os acaricidas abamectina e espirodiclofeno foram eficientes no controle de C. vitis na primavera, tanto com uma única pulverização, como com duas pulverizações, sendo uma no outono e outra na primavera. No outono de 2009, os tratamentos com espirodiclofeno e cihexatina foram eficientes no controle de C. vitis. O ácaro-da-ferrugem-da-videira é controlado eficientemente com pulverizações de cihexatina, enxofre ou espirodiclofeno no outono ou durante a primavera com abamectina, enxofre ou espirodiclofeno.

Dinâmica populacional de Calepitrimerus vitis (Nalepa) (Acari: eriophyidae) em cultivares de videira na região da campanha do Rio Grande do Sul

Na Região da Campanha do Rio Grande do Sul, o ácaro-da-ferrugem-da-videira, Calepitrimerus vitis (Nalepa) (Acari: Eriophyidae), é encontrado com frequência em vinhedos de cultivares europeias, desde a safra de 2004/2005, causando bronzeamento nas folhas. A dinâmica populacional de C. vitis nas cultivares Chardonnay e Merlot foi avaliada em vinhedo comercial localizado no município de Dom Pedrito, na região da Campanha, durante os anos agrícolas de 2005/2006 e 2006/2007, por meio de amostragem realizada em folhas das posições basal, intermediária e apical de ramos de produção. O pico populacional de C. vitis ocorre entre o final de fevereiro e o início de março, sendo seguido de forte declínio populacional. A infestação variou de intensidade entre as cultivares de acordo com o ano, sendo a cultivar Chardonnay mais infestada no primeiro ano, e Merlot, no segundo. Folhas na posição basal, mediana e apical apresentam níveis similares de infestação. Uma correlação positiva foi encontrada entre o número de C. vitis na face abaxial das folhas e o percentual de folhas com infestação.

Functional genomics of O-glucosyltransferases from Concord grape (Vitis labrusca)

Grape (Vitis spp.) is a culturally and economically important crop plant that has been cultivated for thousands of years, primarily for the production of wine. Grape berries accumulate a myriad of phenylpropanoid secondary metabolites, many of which are glucosylated in plantae More than 90 O-glucosyltransferases have been cloned and biochemically characterized from plants, only two of which have been isolated from Vitis spp. The world-wide economic importance of grapes as a crop plant, the human health benefits associated with increased consumption of grape-derived metabolites, the biological relevance of glucosylation, and the lack of information about Vitis glucosyltransferases has inspired the identification, cloning and biochemical characterization of five novel "family 1" O-glucosyltransferases from Concord grape (Vitis labrusca cv. Concord). Protein purification and associated protein sequencIng led to the molecular cloning of UDP-glucose: resveratrollhydroxycinnamic acid O-glucosyltransferase (VLRSGT) from Vitis labrusca berry mesocarp tissue. In addition to being the first glucosyltransferase which accepts trans-resveratrol as a substrate to be characterized in vitro, the recombinant VLRSGT preferentially produces the glucose esters of hydroxycinnamic acids at pH 6.0, and the glucosides of trans-resveratrol and flavonols at 'pH 9.0; the first demonstration of pH-dependent bifunctional glucosylation for this class of enzymes. Gene expression and metabolite profiling support a role for this enzyme in the bifuncitonal glucosylation ofstilbenes and hydroxycinnamic acids in plantae A homology-based approach to cloning was used to identify three enzymes from the Vitis vinifera TIGR grape gene index which had high levels of protein sequence iii identity to previously characterized UDP-glucose: anthocyanin 5-0-glucosyltransferases. Molecular cloning and biochemical characterization demonstrated that these enzymes (rVLOGTl, rVLOGT2, rVLOGT3) glucosylate the 7-0-position of flavonols and the xenobiotic 2,4,5-trichlorophenol (TCP), but not anthocyanins. Variable gene expression throughout grape berry development and enzyme assays with native grape berry protein are consistent with a role for these enzymes in the glucosylation of flavonols; while the broad substrate specificity, the ability of these enzymes to glucosylate TCP and expression of these genes in tissues which are subject to pathogen attack (berry, flower, bud) is consistent with a role for these genes in the plant defense response. Additionally, the Vitis labrusca UDP-glucose: flavonoid 3-0-glucosyltransferase (VL3GT) was identified, cloned and characterized. VL3GT has 96 % protein sequence identity to the previously characterized Vitis vinifera flavonoid 3-0-glucosyltransferase (VV3GT); and glucosylates the 3-0-position of anthocyanidins and flavonols in vitro. Despite high levels of protein sequence identity, VL3GT has distinct biochemical characteristics (as compared to VV3GT), including a preference for B-ring methylated flavonoids and the inability to use UDP-galactose as a donor substrate. RT-PCR analysis of VL3GT gene expression and enzyme assays with native grape protein is consistent with an in planta role for this enzyme in the glucosylation of anthocyanidins,but not flavonols. These studies reveal the power of combining several biochemistry- and molecular biology-based tools to identify, clone, biochemically characterize and elucidate the in planta function of several biologically relevant O-glucosyltransferases from Vitis spp.

Impact of different irrigation strategies on grapes and wine quality of four grapevine cultivars (Vitis sp.) in cool climate conditions. An investigation into the relationships among ABA, water status, grape cultivar and wine quality

Niagara Peninsula of Ontario is the largest viticultural area in Canada. Although it is considered to be a cool and wet region, in the last decade many water stress events occurred during the growing seasons with negative effects on grape and wine quality. This study was initiated to understand and develop the best strategies for water management in vineyards and those that might contribute to grape maturity advancement. The irrigation trials investigated the impact of time of initiation (fruit set, lag phase and veraison), water replacement level based on theoretical loss through crop evapotranspiration (ETc; 100,50 and 25%) and different irrigation strategies [partial root zone drying (PRD) versus regulated deficit irrigation (RD!)] on grape composition and wine sensory profiles. The irrigation experiments were conducted in a commercial vineyard (Lambert Vineyards Inc.) located in Niagara-on-the-Lake, Ontario, from 2005 through 2009. The two experiments that tested the combination of different water regimes and irrigation time initiation were set up in a randomized block design as follows: Baco noir - three replicates x 10 treatments [(25%, 50% and 100% of ETc) x (initiation at fruit set, lag phase and veraison) + control]; Chardonnay - three replicates x seven treatments [(25%, 50% and 100% of ETc) x (initiation at fruit set and veraison) + control]. The experiments that tested different irrigation strategies were set up on two cultivars as follows: Sauvignon blanc - four replicates x four treatments [control, fully irrigated (100% ETc), PRD (100% ETc) and RDI (25% ETc)]; Cabemet Sauvignon - four replicates x five treatments [control, fully irrigated (100% ETc), PRD (100% ETc), RDI (50% ETc) and RDI (25% ETc)]. The controls in each experiment were nonirrigated. The irrigation treatments were compared for many variables related to soil water status, vine physiology, berry composition, wine sensory profile, and hormone composition [(abscisic acid (ABA) and its catabolites]. Soil moisture profile was mostly affected by irrigation treatments between 20 and 60 em depth depending on the grapevine cultivar and the regime of water applied. Overall soil moisture was consistently higher throughout the season in 100 and 50% ETc compare to the control. Transpiration rates and leaf temperature as well as shoot growth rate were the most sensitive variables to soil water status. Drip irrigation associated with RDI treatments (50% ETc and 25% ETc) had the most beneficial effects on vine physiology, fruit composition and wine varietal typicity, mainly by maintaining a balance between vegetative and reproductive parts of the vine. Neither the control nor the 100 ETc had overall a positive effect on grape composition and wine sensory typicity. The time of irrigation initiation affected the vine physiology and grape quality, the most positive effect was found in treatments initiated at lag phase and veraison. RDI treatments were overall more consistent in their positive effect on grape composition and wine varietal typicity comparing to PRD treatment. The greatest difference between non-irrigated and irrigated vines in most of the variables studied was found in 2007, the driest and hottest season of the experimental period. Soil water status had a greater and more consistent effect on red grapevine cultivars rather than on white winegrape cultivars. To understand the relationships among soil and plant water status, plant physiology and the hormonal profiles associated with it, abscisic acid (ABA) and its catabolites [phaseic acid (PA), dihydrophaseic acid (DPA), 7-hydroxy-ABA (TOH-ABA), 8' -hydroxy-ABA, neophaseic acid and abscisic acid glucose ester (ABA-GE)] were analyzed in leaves and berries from the Baco noir and Chardonnay irrigation trials over two growing seasons. ABA and some of its catabolites accurately described the water status in the vines. Endogenous ABA and some of its catabolites were strongly affected in Baco noir and Chardonnay by both the water regime (i.e. ET level) and timing of irrigation initiation. Chardonnay grapevines produced less ABA in both leaves and berries compared to Baco noir, which indicated that ABA synthesis is also cultivar dependant. ABA-GE was the main catabolite in treatments with high water deficits, while PA and DPA were higher in treatments with high water status, suggesting that the vine produced more ABA-GE under water deficits to maintain rapid control of the stomata. These differences between irrigation treatments with respect to ABA and catabolites were particularly noticeable in the dry 2007 season. Two trials using exogenous ABA investigated the effect of different concentrations of ABA and organs targeted for spraying, on grape maturation and berry composition of Cabemet Sauvignon grapevines, in two cool and wet seasons (2008-2009). The fIrst experiment consisted of three replicates x three treatments [(150 and 300 mg/L, both applications only on clusters) + untreated control] while the second experiment consisted in three replicates x four treatments [(full canopy, only clusters, and only leaves sprayed with 300 ppm ABA) + untreated control]. Exogenous ABA was effective in hastening veraison, and improving the composition of Cabemet Sauvignon. Ability of ABA to control the timing of grape berry maturation was dependant on both solution concentration and the target organ. ABA affected not only fruit composition but also yield components. Berries treated with ABA had lower weight and higher skin dry mass, which constitutes qualitative aspects desired in the wine grapes. Temporal advancement of ripening through hormonal control can lead to earlier fruit maturation, which is a distinct advantage in cooler areas or areas with a high risk of early frost occurrence. Exogenous ABA could provide considerable benefits to wine industry in terms of grape composition, wine style and schedule activities in the winery, particularly in wet and cool years. These trials provide the ftrst comprehensive data in eastern North America on the response of important hybrid and Vitis vinifera winegrape cultivars to irrigation management. Results from this study additionally might be a forward step in understanding the ABA metabolism, and its relationship with water status. Future research should be focused on ftnding the ABA threshold required to trigger the ripening process, and how this process could be controlled in cool climates.

Influência do frio e do défice hídrico nas características fisiológicas e bioquímicas em videiras (Vitis vinífera L.) cv. Thompson seedless

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Respuesta de Vitis vinífera L. cv. Malbec a UV-B y ABA

Malbec es la variedad de uva tinta más importante en la vitivinicultura argentina, especialmente en Mendoza, donde se produce más del 80% del total nacional. Las zonas ubicadas a mayor altitud están experimentando un aumento en la superficie cultivada, ya que en ellas se producen uvas con altos contenidos de compuestos fenólicos, responsables de muchas de las características deseadas en los vinos tintos, por su valor organoléptico y capacidad antioxidante. A partir de estos antecedentes, se plantea como objetivo caracterizar la radiación UV-B que reciben los viñedos ubicados a mayor altitud en Mendoza, estudiando los efectos de UV-B, las aplicaciones de ABA y de su interacción, sobre aspectos fisiológicos y bioquímicos que afectan el crecimiento de las bayas (rendimiento cuantitativo) y la calidad enológica de Vitis vinifera L. cv. Malbec.

Evaluación de tolerancia al NaCl en cultivares viníferas y portainjertos híbridos del género Vitis

El objetivo fue evaluar la supervivencia al riego con soluciones de NaCl en 28 genotipos del género Vitis para identificar su grado de tolerancia salina. Se estudiaron 11 híbridos de Vitis americanos y 17 variedades de Vitis vinifera. Plantas de 1 y 2 años se regaron durante 66 días con una solución 100 mM de NaCl. Para clasificar las variedades de acuerdo con su grado de tolerancia a la salinidad, se calculó el día en que el 25% de la población había muerto (1er cuartil de la muestra). Se consideraron como genotipos sensibles aquellos en que al menos 25% había muerto antes del día 30 (3309 Couderc, 161-49 Couderc, Fercal, Freedom, 1103 Paulsen, 99 Richter, SO4, Torrontés Sanjuanino), como poco tolerantes cuando al menos 25% murió entre los días 30 y 60 (Cereza, Colombard, Criollas Blanca y Ballista, Palomino, 110 Richter, 140 Ruggeri, Syrah, Torrontés Riojano) y como tolerantes cuando el 75% o menos sobrevivió más de 60 días (101-14 Millardet-Grasset, 196-17 Castel, Criollas Chica y Sanjuanina, Moscatel de Alejandría, Pedro Giménez). Aun en los genotipos más tolerantes cuando las plantas crecieron bajo salinidad, la integridad de membranas celulares se redujo un 17% y el contenido de clorofila total disminuyó un 52%.

Poliploidia en Vitis Vinífera L.

Se detallan métodos originales para inducir a la poliploidía en variedades de la Vitis Vinífera, en base a tratamientos con colchicina sobre yemas de vid en vías de formación. Se describe un poliploide (tetraploide) de la variedad Malbeck (Cot rouge) y las ventajas que su cultivo puede representar en nuestra zona. Se han efectuado las pruebas citológicas correspondientes.

Mineral stress response of Vitis cell wall transcriptome: identification, characterization and expression of genes from key families involved in wall biosynthesis and modification

Doutoramento em Engenharia Agronómica - Instituto Superior de Agronomia - UL

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

2016

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.

High-throughput SNP genotyping in the highly heterozygous genome of Eucalyptus: assay success, polymorphism and transferability across species

Background: High-throughput SNP genotyping has become an essential requirement for molecular breeding and population genomics studies in plant species. Large scale SNP developments have been reported for several mainstream crops. A growing interest now exists to expand the speed and resolution of genetic analysis to outbred species with highly heterozygous genomes. When nucleotide diversity is high, a refined diagnosis of the target SNP sequence context is needed to convert queried SNPs into high-quality genotypes using the Golden Gate Genotyping Technology (GGGT). This issue becomes exacerbated when attempting to transfer SNPs across species, a scarcely explored topic in plants, and likely to become significant for population genomics and inter specific breeding applications in less domesticated and less funded plant genera. Results: We have successfully developed the first set of 768 SNPs assayed by the GGGT for the highly heterozygous genome of Eucalyptus from a mixed Sanger/454 database with 1,164,695 ESTs and the preliminary 4.5X draft genome sequence for E. grandis. A systematic assessment of in silico SNP filtering requirements showed that stringent constraints on the SNP surrounding sequences have a significant impact on SNP genotyping performance and polymorphism. SNP assay success was high for the 288 SNPs selected with more rigorous in silico constraints; 93% of them provided high quality genotype calls and 71% of them were polymorphic in a diverse panel of 96 individuals of five different species. SNP reliability was high across nine Eucalyptus species belonging to three sections within subgenus Symphomyrtus and still satisfactory across species of two additional subgenera, although polymorphism declined as phylogenetic distance increased. Conclusions: This study indicates that the GGGT performs well both within and across species of Eucalyptus notwithstanding its nucleotide diversity >= 2%. The development of a much larger array of informative SNPs across multiple Eucalyptus species is feasible, although strongly dependent on having a representative and sufficiently deep collection of sequences from many individuals of each target species. A higher density SNP platform will be instrumental to undertake genome-wide phylogenetic and population genomics studies and to implement molecular breeding by Genomic Selection in Eucalyptus.

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.