Jason P. Smith

Soil temperature moderates grapevine carbohydrate reserves after bud break and conditions fruit set responses to photoassimilatory stress

In cultivated grapevines (Vitis vinifera L.), suboptimal photoassimilatory conditions during flowering can lead to inflorescence necrosis and shedding of flowers and young ovaries and, consequently, poor fruit set. However, before this study it was not known whether carbohydrate reserves augment fruit set when concurrent photoassimilation is limited. Carbohydrate reserves are most abundant in grapevine roots and soil temperature moderates their mobilisation. Accordingly, we grew potted Chardonnay grapevines in soil at 15°C (cool) or 26°C (warm) from bud break to the onset of flowering to manipulate root carbohydrate reserve status. Then to induce photoassimilatory responses we subjected the plants to low (94µmolmol-1) CO2 or ambient (336µmolmol-1) CO2 atmospheres during fruit setting. Analyses of photoassimilation and biomass and carbohydrate reserve distribution confirmed that fruit set was limited by concurrent photoassimilation. Furthermore, the availability of current photoassimilates for inflorescence development and fruit set was conditioned by the simultaneous demands for shoot and root growth, as well as the restoration of root carbohydrate reserves. Results indicate that great seasonal variability in grapevine fruit set is a likely response of cultivated grapevines to photoassimilatory stresses, such as shading, defoliation and air temperature and to variations in carbohydrate reserve status before flowering.

Rapid monitoring of grapevine reserves using ATR–FT-IR and chemometrics

Predictions of grapevine yield and the management of sugar accumulation and secondary metabolite production during berry ripening may be improved by monitoring nitrogen and starch reserves in the perennial parts of the vine. The standard method for determining nitrogen concentration in plant tissue is by combustion analysis, while enzymatic hydrolysis followed by glucose quantification is commonly used for starch. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FT-IR) combined with chemometric modelling offers a rapid means for the determination of a range of analytes in powdered or ground samples. ATR-FT-IR offers significant advantages over combustion or enzymatic analysis of samples due to the simplicity of instrument operation, reproducibility and speed of data collection. In the present investigation, 1880 root and wood samples were collected from Shiraz, Semillon and Riesling vineyards in Australia and Germany. Nitrogen and starch concentrations were determined using standard analytical methods, and ATR-FT-IR spectra collected for each sample using a Bruker Alpha instrument. Samples were randomly assigned to either calibration or test data sets representing two thirds and one third of the samples respectively. Signal preprocessing included extended multiplicative scatter correction for water and carbon dioxide vapour, standard normal variate scaling with second derivative and variable selection prior to regression. Excellent predictive models for percent dry weight (DW) of nitrogen (range: 0.10-2.65% DW, median: 0.45% DW) and starch (range: 0.25-42.82% DW, median: 7.77% DW) using partial least squares (PLS) or support vector machine (SVM) analysis for linear and nonlinear regression respectively, were constructed and cross validated with low root mean square errors of prediction (RMSEP). Calibrations employing SVM-regression provided the optimum predictive models for nitrogen (R(2)=0.98 and RMSEP=0.07% DW) compared to PLS regression (R(2)=0.97 and RMSEP=0.08% DW). The best predictive models for starch was obtained using PLS regression (R(2)=0.95 and RSMEP=1.43% DW) compared to SVR (R(2)=0.95; RMSEP=1.56% DW). The RMSEP for both nitrogen and starch is below the reported seasonal flux for these analytes in Vitis vinifera. Nitrogen and starch concentrations in grapevine tissues can thus be accurately determined using ATR-FT-IR, providing a rapid method for monitoring vine reserve status under commercial grape production.

VitiCanopy: A Free Computer App to Estimate Canopy Vigor and Porosity for Grapevine

Leaf area index (LAI) and plant area index (PAI) are common and important biophysical parameters used to estimate agronomical variables such as canopy growth, light interception and water requirements of plants and trees. LAI can be either measured directly using destructive methods or indirectly using dedicated and expensive instrumentation, both of which require a high level of know-how to operate equipment, handle data and interpret results. Recently, a novel smartphone and tablet PC application, VitiCanopy, has been developed by a group of researchers from the University of Adelaide and the University of Melbourne, to estimate grapevine canopy size (LAI and PAI), canopy porosity, canopy cover and clumping index. VitiCanopy uses the front in-built camera and GPS capabilities of smartphones and tablet PCs to automatically implement image analysis algorithms on upward-looking digital images of canopies and calculates relevant canopy architecture parameters. Results from the use of VitiCanopy on grapevines correlated well with traditional methods to measure/estimate LAI and PAI. Like other indirect methods, VitiCanopy does not distinguish between leaf and non-leaf material but it was demonstrated that the non-leaf material could be extracted from the results, if needed, to increase accuracy. VitiCanopy is an accurate, user-friendly and free alternative to current techniques used by scientists and viticultural practitioners to assess the dynamics of LAI, PAI and canopy architecture in vineyards, and has the potential to be adapted for use on other plants.

Shifts in biomass and nitrogen allocation of tree seedlings in response to root-zone temperature

Root-zone warming of trees can result in an increase in biomass production but the mechanisms for this increase may differ between evergreen and deciduous species. The leaf gas exchange, carbohydrate and nitrogen (N) partitioning of two Australian evergreens, Acacia saligna and Eucalyptus cladocalyx, were compared to the deciduous Populus deltoides and Acer negundo after exposure to cool or warm soil during spring. The warm treatment stimulated aboveground biomass production in all four species; however, the form of this increase was species dependent. Compared with the evergreens, soluble sugars were mobilised from the above- and belowground components to a greater extent in the deciduous species, especially during root-zone warming. Photosynthesis, stomatal conductance and transpiration were increased in the warm soil treatment for the two evergreens and P. deltoides only. In P. deltoides and A. saligna the new fine roots contained greater starch concentrations when grown in warm soil but only in A. negundo was new root growth greater. Compared with the other three species, the leguminous A. saligna contained the highest N and most of this was concentrated in the phyllodes of warmed plants with no apparent mobilisation from the existing biomass. In the other evergreen, E. cladocalyx, the existing leaves and stems were a N source for new growth, while in the two deciduous species N was derived from the woody components and structural roots. These data show that the carbohydrate movement and N partitioning patterns in response to soil warming differ between perennial and deciduous plants and are likely responsible for the different forms of biomass accumulation in each of these species.

Implications of the Presence of Maturing Fruit on Carbohydrate and Nitrogen Distribution in Grapevines under Postveraison Water Constraints

Grapevine (Vitis vinifera) berries are sugar and nitrogen (N) sinks between veraison and fruit maturity. Limited photoassimilation, often caused by water constraints, induces reserve total nonstructural carbohydrate (TNC) remobilization, contributing to berry sugar accumulation, while fruit N accumulation can be affected by vine water supply. Although postveraison root carbohydrate remobilization toward the fruit has been identified through C tracing studies, it is still unclear when this remobilization occurs during the two phases of berry sugar accumulation (rapid and slow). Similarly, although postveraison N reserve mobilization toward the fruit has been reported, the impact of water constraints during berry N accumulation on its translocation from the different grapevine organs requires clarification. Potted grapevines were grown with or without fruit from the onset of veraison. Vines were irrigated to sustain water constraints, and fortnightly root, trunk, shoot, and leaf structural biomass, starch, soluble sugar, total N, and amino N concentrations were determined. The fruit sugar and N accumulation was also assessed. Root starch depletion coincided with root sucrose and hexose accumulation during peak berry sugar accumulation. Defruiting at veraison resulted in continuous root growth, earlier starch storage, and root hexose accumulation. Leaf N depletion coincided with fruit N accumulation, while the roots of defruited vines accumulated N reserves. Root growth, starch, and N reserve accumulation were affected by maturing fruit during water constraints. Root starch is an alternative source to support fruit sugar accumulation, resulting in reserve starch depletion during rapid fruit sugar accumulation, while root starch refills during slow berry sugar accumulation. On the other hand, leaf N is a source toward postveraison fruit N accumulation, and the fruit N accumulation prevents root N storage. Grapevine berries are sinks for the incorporation of both carbohydrates (Davies and Robinson, 1996) and N (RoubelakisAngelakis and Kliewer, 1992) between veraison and fruit maturity. Restricted TNC availability, induced by limited leaf photoassimilation, can cause starch redistribution from the roots during berry sugar accumulation (Candolfi-Vasconcelos et al., 1994), while N concentrations in the berries and roots are affected by abiotic conditions, such as vine water availability, during the growing season (Araujo et al., 1995). Furthermore, apart from also being affected by vine N supply, the N reserve accumulation in the roots is restricted by the presence of fruit before and after veraison (Rodriguez-Lovelle and Gaudillere, 2002). It is still unclear how the postveraison distribution of TNC and N reserves among the different organs, are affected by a combination of fruit presence and sustained water constraints. A further question remains on how this distribution contributes to, or inhibits, TNC and N reserve storage, or the contents of TNC and N in the fruit during berry maturation. In plant roots, TNC are mainly stored as starch, which can be hydrolyzed, yielding osmotic active soluble sugars (Regier et al., 2009). Apart from the possible remobilization of sugars via phloem sucrose transportation (Ruan et al., 2010) between Received for publication 23 Nov. 2016. Accepted for publication 9 Jan. 2017. This work was supported by the National Wine and Grape Industry Centre, and the Australian grapegrowers and winemakers through their investment body, Wine Australia, with matching funds from the Australian Government. Current address: Institut f€ur Allgemeinen und €okologischen Weinbau, Hochschule Geisenheim University, Geisenheim, 65366, Germany Current address: Montpellier SupAgro, Montpellier, 34060, France Corresponding author. E-mail: grossouw@csu.edu.au. J. AMER. SOC. HORT. SCI. 142(2):71–84. 2017. 71 the perennial vine parts (the roots and trunk) and the ripening berries, thereby contributing to berry sugar accumulation (Candolfi-Vasconcelos et al., 1994), sugars also accumulate in various tissues of water stressed plants, to aid in osmotic regulation (Regier et al., 2009). Therefore, upon water constraints during berry sugar accumulation, the accumulation of soluble sugars in different vine parts could theoretically contribute to various functions; e.g., to facilitate TNC remobilization, and to improve abiotic stress tolerance. As grapevines are perennial plants, the storage of starch reserves at the end of the season is essential for reserve TNC utilization the following season, required for vegetative and reproductive development from budburst (Holzapfel et al., 2010). Depleted root starch concentration can then lead to limited initiation and development of the inflorescence, and decreased fruit set and fruit yield the following season (Smith and Holzapfel, 2009). The N allocation to grapevine berries, and subsequent accumulation during berry maturation is, from a wine quality perspective, essential as it determines the juice yeast assimilable N content, influencing fermentation and wine composition. However, root N accumulation late in the growing season is important for its overwintering storage (Cheng et al., 2004). Similar to TNC, N reserves are used toward the initiation of early season vegetative growth, where their mobilization regulate spring growth and account for most of the N distribution until around flowering, as N soil uptake is usually still insufficient at this stage (Zapata et al., 2004). Nitrogen accumulation in the perennial vine parts usually initiates before berry maturity, and the reserves continue to increase until leaf fall (RoubelakisAngelakis and Kliewer, 1992). The presence of fruit reduces N assimilation in grapevine roots (Morinaga et al., 2003). Nitrogen is mostly stored in the roots, and these reserves consists of a range of amino acids and proteins (Zapata et al., 2004). Amino acids in plants are involved in the regulation of N metabolism, and play essential roles in N transport and storage (Roubelakis-Angelakis and Kliewer, 1992). The metabolic pathway related to the a-ketoglutarate family of amino acids, yields glutamic acid, glutamine, arginine, and proline. These amino acids are abundant in plants, and have distinct roles in N metabolism (Verma et al., 1999). Glutamic acid is the intermediate product of nitrate and ammonium assimilation, and a precursor for the synthesis of glutamine, arginine, and proline (Berg et al., 2002). Arginine is considered the main N-storage amino acid in grapevines (Xia and Cheng, 2004), while glutamine is an essential N transporter (Coruzzi and Last, 2000). Proline accumulation is linked to osmotic adjustment following abiotic plant stresses (Hare and Cress, 1997). The metabolism of the a-ketoglutarate-derived amino acids is therefore essential to regulate plant N partitioning and distribution, particularly during abiotic constraints. The aim of this experiment was to determine the effect of fruit presence during sustained postveraison water constraints, on the TNC and N distribution within grapevines. The first goal was to investigate the response in the structural development of the leaves, shoots, trunk, and roots, based on the presence of fruit, during sustained water constraints. The second goal was to determine how fruit presence affects TNC accumulation in the different organs, during the sustained water constraints, and to assess which individual sugars accumulate in the grapevine roots during the two phases of berry maturation (rapid and slow sugar accumulation). The final goal was to determine how the presence of maturing fruit affects the allocation of N between the grapevine organs, and to identify potential contributions of amino N, and especially the amino acids yielded from a-ketoglutarate metabolism, toward N storage or translocation. Materials and Methods EXPERIMENTAL DESIGN. Own-rooted ‘Shiraz’ grapevines, grown in 50-L pots containing commercial potting mix, were used for this experiment in the 2014–15 growing season. The grapevines were grown in an outside bird-proof cage in the warm to very warm climate Riverina region of New South Wales, Australia. The 3-year-old grapevines were winter pruned to 10 spurs with two buds each, and arranged in four rows of nine vines each. From just after budburst, the grapevines were fertilized every 3 weeks with 250 mL of 50:1 diluted complete liquid fertilizer (MEGAMIX Plus; Rutec, Tamworth, Australia). In total, 2.6 g N was applied to each vine through fertilization, and the fertilization events were ceased 1 month before the start of the experiment, aiming to limit soil N uptake during the experiment. The grapevines were well watered between budburst and veraison, when irrigation was supplied three times a day to the point of visual free drainage from the pots. Vines were shoot thinned so as to leave 17 shoots per vine from fruit set, and at the onset of veraison, 2 d after the first sign of berry softening was observed, the treatments were initiated. Four randomly selected vines, one from each row, were destructively harvested on the day when then the treatments were initiated, to represent the population of grapevines before the implementation of the treatments. After removal of the four initial vines, the eight remaining vines per row were evenly spaced out in the row. All bunches were removed on half the vines, to have 16 vines with, and 16 vines without fruit. Two vines in each row (one with fruit and one without fruit) were used as a visual reference to control irrigation scheduling, and received double the irrigation volume than the other vines. Irrigation was scheduled three times per day (0800, 1400, and 1800 HR) for all vines, and the vines receiving double the irrigation, were rewatered each day just to the point of visual free drainage from the pots during the 1400 HR irrigation event, through two irrigation emitters per pot. The remaining 24 vines were irrigated throu

Vitis vinifera root and leaf metabolic composition during fruit maturation: implications of defoliation

Grapevine (Vitis vinifera) roots and leaves represent major carbohydrate and nitrogen (N) sources, either as recent assimilates, or mobilized from labile or storage pools. This study examined the response of root and leaf primary metabolism following defoliation treatments applied to fruiting vines during ripening. The objective was to link alterations in root and leaf metabolism to carbohydrate and N source functioning under conditions of increased fruit sink demand. Potted grapevine leaf area was adjusted near the start of véraison to 25 primary leaves per vine compared to 100 leaves for the control. An additional group of vines were completely defoliated. Fruit sugar and N content development was assessed, and root and leaf starch and N concentrations determined. An untargeted GC/MS approach was undertaken to evaluate root and leaf primary metabolite concentrations. Partial and full defoliation increased root carbohydrate source contribution towards berry sugar accumulation, evident through starch remobilization. Furthermore, root myo-inositol metabolism played a distinct role during carbohydrate remobilization. Full defoliation induced shikimate pathway derived aromatic amino acid accumulation in roots, while arginine accumulated after full and partial defoliation. Likewise, various leaf amino acids accumulated after partial defoliation. These results suggest elevated root and leaf amino N source activity when leaf N availability is restricted during fruit ripening. Overall, this study provides novel information regarding the impact of leaf source restriction, on metabolic compositions of major carbohydrate and N sources during berry maturation. These results enhance the understanding of source organ carbon and N metabolism during fruit maturation.

Vitis vinifera berry metabolic composition during maturation: implications of defoliation.

Leaves are an important contributor toward berry sugar and nitrogen (N) accumulation, and leaf area, therefore, affects fruit composition during grapevine (Vitis vinifera) berry ripening. The aim of this study was to investigate the impact of leaf presence on key berry quality attributes in conjunction with the accumulation of primary berry metabolites. Shortly after the start of véraison (berry ripening), potted grapevines were defoliated (total defoliation and 25% of the control), and the accumulation of berry soluble solids, N and anthocyanins were compared to that of a full leaf area control. An untargeted approach was undertaken to measure the content in primary metabolites by gas chromatography/mass spectrometry. Partial and full defoliation resulted in reduced berry sugar and anthocyanin accumulation, while total berry N content was unaffected. The juice yeast assimilable N (YAN), however, increased upon partial and full defoliation. Remobilized carbohydrate reserves allowed accumulation of the major berry sugars during the absence of leaf photoassimilation. Berry anthocyanin biosynthesis was strongly inhibited by defoliation, which could relate to the carbon (C) source limitation and/or increased bunch exposure. Arginine accumulation, likely resulting from reserve translocation, contributed to increased YAN upon defoliation. Furthermore, assessing the implications on various products of the shikimate pathway suggests the C flux through this pathway to be largely affected by leaf source limitation during fruit maturation. This study provides a novel investigation of impacts of leaf C and N source presence during berry maturation, on the development of key berry quality parameters as underlined by alterations in primary metabolism.

Circadian regulation of grapevine root and shoot growth and their modulation by photoperiod and temperature

Some plant species demonstrate a pronounced 24 h rhythm in fine root growth but the endogenous and exogenous factors that regulate these diel cycles are unclear. Photoperiod and temperature are known to interact with diel patterns in shoot growth but it is uncertain how these environmental factors are interrelated with below-ground growth. In this particular study, the fine root system of two grapevine species was monitored over a period of ten days with a high resolution scanner, under constant soil moisture and three different photoperiod regimes. Pronounced diel rhythms in shoot and root growth rates were apparent under a fixed 14 h photoperiod. Maximal root growth rate occurred 1-2 h prior to- and until 2 h after the onset of darkness. Subsequently, during the latter part of the dark period, root growth rate decreased and reached minimal values at the onset of the light period. Relative to 22 °C, exposure to a 30 °C air and soil temperature halved root growth but stimulated shoot growth. Notably, the shoot extension rate peak shifted from late afternoon to midnight at this higher temperature zone. When plants were exposed to a delayed photoperiod or progressively shortening photoperiod, the diel changes in root growth rate followed the same pattern as in the fixed photoperiod, regardless of whether the plant was in light or dark. This suggests that light was not the predominant trigger for stimulating root elongation. Conversely, shoot growth rates were not fixed to a clock, with minimum growth consistently at the completion of the dark period regardless of the time of day. In summary, fine root growth of grapevines was found to have a pronounced diel pattern and an endogenous circadian clock appears to orchestrate this rhythm. Soil temperature modified the amplitude of this pattern, but we argue here that, as evidenced from exhausted starch reserves within root tips by early morning, carbon supply from photosynthesis is also required to maintain maximum root growth.