Pantoea agglomerans‐associated bacteria in grape phylloxera (Daktulosphaira vitifoliae,Fitch)

1 Grape phylloxera lack intracellular symbionts, but the leaf‐galling form appears to be associated with a single microbial species. 2 16S and 18SrDNA sequences were used for identification of symbiotic material. 3 A single bacterial species, closely related to Pantoea agglomerans, was identified in adult parthenogenetic individuals, their eggs and leaf gall tissue of several populations. 4 A 16S rDNA primer pair was designed to test grape phylloxera populations more specifically for the presence of P. agglomerans. 5 16S rDNA sequences of the identified bacteria were very similar to already‐known secondary symbionts occurring in aphids, thrips and other insects. 6 The identified bacteria were culturable on simple media, which demonstrates that the relationship between grape phylloxera and P. agglomerans is not as firm as that of the obligately endosymbiotic Buchnera aphidicola and other aphids.     

An improved enzyme-linked immunoabsorbent assay protocol for the detection of small lytic peptides in transgenic grapevines (Vitis vinifera)

An enzyme-linked immunoabsorbent assay (ELISA) protocol was developed for the detection of small lytic peptides in transgenic grapevines (V. vinifera). The protocol requires a high concentration of protease inhibitor in the extraction buffer; the use of antiserum cross-absorbed with control tissue, an increased concentration of blocking reagents in the antiserum buffer, and performing all coating and/or binding processes at 37°C while reducing the time period for each step to 1 h. The procedure greatly reduced protein degradation, increased the signal-to-noise ratio, and it allowed the effective detection of the Shiva-1 lytic peptide (5 kDa) at concentrations as low as 0.1 μM. This procedure made it possible for routine analysis of transgene expression in Shiva-1 gene-containing transgenic grape plants.     

Properties of potassium uptake by seedling roots of grape cultivars

Uptake rates of (⁸⁶Rb)K⁺ by seedling roots of six cultivars were measured and compared with K⁺ content of the root, K⁺ leakage, H⁺ efflux, and K⁺-ATPase activity of a partially purified plasmalemma fraction.Different cultivars showed significantly different rates of (⁸⁶Rb)K⁺ uptake. The uptake rates of the first (0–5 min) period did not correlate with K⁺ content of the seedling roots.The rates of uptake in the 10 to 30 min period, supposed to be active, were negatively correlated with K⁺ content of the root. Roots consistently leaked K⁺ during the first 5 min. This leakage was positively correlated with the endogenous K⁺ content of the tissue.H⁺ efflux was significantly different among the cultivars and correlated with the K⁺-ATPase activity of a microsomal fraction partially purified on discontinuous (18/34%) sucrose gradient. The relationships among transport parameters are discussed.Part of the present research was carried out at the Department of Agricultural Biotechnology of University of Padova, Italy.Part of the present research was carried out at the Department of Agricultural Biotechnology of University of Padova, Italy.     

Ammonium requirement for radiation-induced accumulation of polyamines in suspension-cultured grape cells

The effects of radiation-mediated free radical production on polyamine metabolism were investigated in grape cells ( Vitis vinifera L. cv. Gamay) using a cell suspension culture. Putrescine (Put) synthesis was triggered in irradiated cells (0. 5 kGy) only when ammonium was present in the culture medium. Under these conditions. Put accumulated to a continuously high level. As also described for other kinds of stress, the level of spermidine was slightly enhanced and that of spermine unchanged. The role of ammonium was assessed by studying non-irradiated cell cultures. In the presence of ammonium, a transient increase of both arginine decarboxylase (ADC) activity and of Put synthesis was observed during the lag phase of growth. This Put enhancement was inhibited by difluoromethyl arginine and not by difluoromethyl ornithine, showing that increased Put synthesis occurs via the ADC pathway. When ammonium was withheld from the culture medium. ADC activity was still triggered though transient Put accumulation was completely suppressed. These results emphasize the importance of ammonium availability in cultured cells as a limiting factor for Put production. Polyamine synthesis, therefore, cannot be stimulated by gamma irradiation in the absence of an ammonium supply. These results support the hypothesis that Put synthesis is a detoxification process of the ammonium produced as a result of nitrogen recycling within stressed plant cells.     

Nematode and Grape Rootstock Interactions Including an Improved Understanding of Tolerance

Sixteen cultivars of grape were screened over a two-year period in the presence or absence of 10 different nematode populations. Populations of Meloidogyne spp., Xiphinema index, and Mesocriconema xenoplax developed more rapidly and caused greater damage than populations of X. americanum and Tylenchulus semipenetrans. Populations of mixed Meloidogyne spp. having a history of feeding on grape were among the fastest developing populations. Tolerance to nematode parasitism appeared to be based on different mechanisms. Slow developing, less pathogenic nematode populations often stimulated vine growth, thus vines appeared to possess tolerance. Likewise, cultivars selected for nematode resistance often stimulated vine growth when fed upon by the nematode. However, tolerance sources that resulted from nematode resistance are vulnerable due to the occurrence of populations that break resistance mechanisms. Growth of cultivars with phylloxera (Daktalospharia vitifoliae) resistance was unchanged by the presence of nematodes, indicating that phylloxera resistance may provide a useful source of nematode relief. These and several additional sources of specific tolerance are discussed.     

Potassium transport in developing fleshy fruits: the grapevine inward K+ channel VvK1.2 is activated by CIPK–CBL complexes and induced in ripening berry flesh cells

The grape berry provides a model for investigating the physiology of non‐climacteric fruits. Increased K⁺ accumulation in the berry has a strong negative impact on fruit acidity (and quality). In maturing berries, we identified a K⁺ channel from the Shaker family, VvK1.2, and two CBL‐interacting protein kinase (CIPK)/calcineurin B‐like calcium sensor (CBL) pairs, VvCIPK04–VvCBL01 and VvCIPK03–VvCBL02, that may control the activity of this channel. VvCBL01 and VvCIPK04 are homologues of Arabidopsis AtCBL1 and AtCIPK23, respectively, which form a complex that controls the activity of the Shaker K⁺ channel AKT1 in Arabidopsis roots. VvK1.2 remained electrically silent when expressed alone in Xenopus oocytes, but gave rise to K⁺ currents when co‐expressed with the pairs VvCIPK03–VvCBL02 or VvCIPK04–VvCBL01, the second pair inducing much larger currents than the first one. Other tested CIPK–CBL pairs expressed in maturing berries were found to be unable to activate VvK1.2. When activated by its CIPK–CBL partners, VvK1.2 acts as a voltage‐gated inwardly rectifying K⁺ channel that is activated at voltages more negative than –100 mV and is stimulated upon external acidification. This channel is specifically expressed in the berry, where it displays a very strong induction at veraison (the inception of ripening) in flesh cells, phloem tissues and perivascular cells surrounding vascular bundles. Its expression in these tissues is further greatly increased upon mild drought stress. VvK1.2 is thus likely to mediate rapid K⁺ transport in the berry and to contribute to the extensive re‐organization of the translocation pathways and transport mechanisms that occurs at veraison.     

Impact of leaf removal as a source of stresses on grapevine yields,chemical characteristics,and anthocyanin content in the grapevine variety Babica

Leaf removal (LR) treatments improve the photosynthetic capacity of the remaining leaves and induce flavonoid synthesis as a stress response in the common grapevine (Vitis vinifera L.). However, excessive exposure of grape berries to UV-B radiation as a result of cultural practices in the Mediterranean climate may have negative effects on berry composition. This 2-year study determined the effects of defoliation on the autochthonous red grape variety ‘Babica’ in a Mediterranean climate (wine-growing region Dalmatia, Croatia). Six leaves were removed before flowering (FLR) and at the end of véraison (the onset of grape ripening; VerLR) and were compared to the untreated control. Yield parameters, sugar content, grape must pH, total polyphenols (TP), total anthocyanin (TA) content, and individual anthocyanin compounds were measured in grape skin extracts and wines. However, the greater mean daily temperature during the vegetation period and lesser rainfall before harvest in 2018 increased yield per vine, average cluster weights, density, and total acidity, compared to 2017. Both defoliation treatments significantly reduced TP in grape extracts, but these differences were not observed in wine. Compared to the control (NLR), VerLR treatment significantly reduced TA in grape skin extracts and wine. Significantly lesser TP concentrations, in grape skin extracts and wine, as well as TA were noticed during the 2017 season. VerLR treatment reduced the concentration of nine individual anthocyanins compared to the control in grape skin extracts, while this effect was not observed in wine. Season year was a statistically significant source of variability of the individual anthocyanin contents in wine. Under specific environmental conditions LR can decrease polyphenols, especially anthocyanins, and negatively impact grape and wine quality.