Nitric oxide increases tolerance responses to moderate water deficit in leaves of Phaseolus vulgaris and Vigna unguiculata bean species.

Drought stress is one of the most intensively studied and widespread constraints, and nitric oxide (NO) is a key signaling molecule involved in the mediation of abiotic stresses in plants. We demonstrated that a sprayed solution of NO from donor sodium nitroprusside increased drought stress tolerance responses in both sensitive (Phaseolus vulgaris) and tolerant (Vigna unguiculata) beans. In intact plants subjected to halting irrigation, NO increased the leaf relative water content and stomatal conductance in both species. After cutting leaf discs and washing them, NO induced increased electrolyte leakage, which was more evident in the tolerant species. These leaf discs were then subjected to different water deficits, simulating moderate and severe drought stress conditions through polyethylene glycol solutions. NO supplied at moderate drought stress revealed a reduced membrane injury index in sensitive species. In hydrated discs and at this level of water deficit, NO increased the electron transport rate in both species, and a reduction of these rates was observed at severe stress levels. Taken together, it can be shown that NO has an effective role in ameliorating drought stress effects, activating tolerance responses at moderate water deficit levels and in both bean species which present differential drought tolerance.

A Leaf Disc Assay for Evaluating the Response of Tea (Camellia sinensis) to PEG-Induced Osmotic Stress and Protective Effects of Azoxystrobin against Drought.

Tea (Camellia sinensis), a globally cultivated beverage crop, is sensitive to drought, which can have an adverse effect on the yield and quality of tea. Azoxystrobin (AZ) is one kind of fungicide considered as an agent to relieve damage caused by stress. Initially, the response of tea plant to osmotic-gradient stress was evaluated using leaf disc assays with PEG-induced osmotic stress. The decline of the maximum quantum yield of PSII (Fv/Fm), actual photosynthetic efficiency of PS II (Y(II)), total chlorophylls, carotenoids, DPPH radical scavenging capacity, reducing power, total phenols, and the increase in MDA was observed in leaf discs treated with a gradient of PEG solutions (22.8, 33.2, 41.1% PEG, and blank). These results revealed that efficiency of photosystem II (PSII), photosynthetic pigments, and antioxidant ability in leaf discs were inhibited with an aggravated lipid peroxidation under PEG-induced osmotic stress, and indicated leaf disc assay with moderate PEG iso-osmotic condition would reflect a portion of tea plant response to drought stress. Therefore, the protective effect of AZ (0.125 and 1.25 g a.i. L-1) on tea plants suffering from drought was evaluated using leaf disc assays with 22.8% PEG iso-osmotic condition. Pretreatment of AZ (0.125 a.i. g L-1) reversed Fv/Fm, Y(II), DPPH radical scavenging capacity, and reducing power with reduced MDA in PEG-treated leaf discs, but photosynthetic pigments, total phenols, and ascorbate peroxidase activity were irresponsive to AZ. An Alleviated physiological damage in tea leaf with AZ applying was preliminarily revealed in this study. A Rapid screening of agents for tea plants against drought was developed to assist in the selection of protective agents.

Carbon and nitrogen stable isotope variations in leaves of two grapevine cultivars (Chasselas and Pinot noir): Implications for ecophysiological studies.

We investigated the within- and between-leaf variability in the carbon and nitrogen isotope composition (δ13C and δ15N) and total nitrogen (TN) content in two grapevine cultivars (Vitis vinifera cv. Chasselas and Pinot noir) field-grown under rain-fed conditions. The within-leaf variability was studied in discs sampled from base-to-tip and left and right regions from the margin to midrib. The intra- and interplant variability was studied by comparing leaves at different positions along the shoot (basal, median, apical). In leaves from both cultivars, a decrease in δ13C from base to tip was observed, which is in line with an upward gradient of stomatal density and chlorophyll concentration. Less important, but still significant differences were observed between the right and left discs. The leaf TN and δ15N values differed between cultivars, showed smaller variations than the δ13C values, and no systematic spatial trends. The intraleaf variations in δ13C, δ15N, and TN suggest that stomatal behavior, CO2 fixation, chlorophyll concentrations, and the chemical composition of leaf components were heterogeneous in the leaves. At the canopy scale, the apical leaves had less 13C and more 15N and TN than the basal leaves, indicating differences in their photosynthetic capacity and remobilizations from old, senescing leaves to younger leaves. Overall, this study demonstrates patchiness in the δ13C and δ15N values of grapevine leaves and species-specificity of the nitrogen assimilation and 15N fractionation. These findings suggest that care must be taken not to overinterpret foliar δ13C and δ15N values in studies based on fragmented material as markers of physiological and biochemical responses to environmental factors.

Rpv Mediated Defense Responses in Grapevine Offspring Resistant to Plasmopara viticola.

Downy mildew, caused by the biotrophic oomycete Plasmopara viticola, is one of the most serious grapevine diseases. The development of new varieties, showing partial resistance to downy mildew, through traditional breeding provides a sustainable and effective solution for disease management. Marker-assisted-selection (MAS) provide fast and cost-effective genotyping methods, but phenotyping remains necessary to characterize the host-pathogen interaction and assess the effective resistance level of new varieties as well as to validate MAS selection. In this study, the Rpv mediated defense responses were investigated in 31 genotypes, encompassing susceptible and resistant varieties and 26 seedlings, following inoculation of leaf discs with P. viticola. The offspring differed in Rpv loci inherited (none, one or two): Rpv3-3 and Rpv10 from Solaris and Rpv3-1 and Rpv12 from Kozma 20-3. To improve the assessment of different resistance responses, pathogen reaction (sporulation) and host reaction (necrosis) were scored separately as independent features. They were differently expressed depending on Rpv locus: offspring carrying Rpv3-1 and Rpv12 loci showed the strongest resistance response (scarce sporulation and necrosis), those carrying Rpv3-3 locus showed the highest levels of necrosis while Rpv10 carrying genotypes showed intermediate levels of both sporulation and necrosis.

Co-Acquired Nanovirus and Geminivirus Exhibit a Contrasted Localization within Their Common Aphid Vector.

Single-stranded DNA (ssDNA) plant viruses belong to the families Geminiviridae and Nanoviridae. They are transmitted by Hemipteran insects in a circulative, mostly non-propagative, manner. While geminiviruses are transmitted by leafhoppers, treehoppers, whiteflies and aphids, nanoviruses are transmitted exclusively by aphids. Circulative transmission involves complex virus-vector interactions in which epithelial cells have to be crossed and defense mechanisms counteracted. Vector taxa are considered a relevant taxonomic criterion for virus classification, indicating that viruses can evolve specific interactions with their vectors. Thus, we predicted that, although nanoviruses and geminiviruses represent related viral families, they have evolved distinct interactions with their vector. This prediction is also supported by the non-structural Nuclear Shuttle Protein (NSP) that is involved in vector transmission in nanoviruses but has no similar function in geminiviruses. Thanks to the recent discovery of aphid-transmitted geminiviruses, this prediction could be tested for the geminivirus alfalfa leaf curl virus (ALCV) and the nanovirus faba bean necrotic stunt virus (FBNSV) in their common vector, Aphis craccivora. Estimations of viral load in midgut and head of aphids, precise localization of viral DNA in cells of insect vectors and host plants, and virus transmission tests revealed that the pathway of the two viruses across the body of their common vector differs both quantitatively and qualitatively.

Production of herbicide-resistant medicinal plant Salvia miltiorrhiza transformed with the bar gene.

In this study, we successfully performed Agrobacterium-mediated genetic transformation of Salvia miltiorrhiza and produced herbicide-resistant transformants. Leaf discs of S. miltiorrhiza were infected with Agrobacterium tumefaciens EHA105 harboring pCAMBIA 3301. The pCAMBIA 3301 includes an intron-containing gus reporter and a bar selection marker. To increase stable transformation efficiency, a two-step selection was employed which consists of herbicide resistance and gus expression. Here, we put more attention to the screening step of herbicide resistance. The current study provides an efficient screening system for the transformed plant of S. miltiorrhiza harboring bar gene. To determine the most suitable phosphinothricin concentration for plant selection, non-transformed leaf discs were grown on selection media containing six different phosphinothricin concentrations (0, 0.2, 0.4, 0.6, 0.8, and 1.0 mg/l). Based on the above results of non-transformed calluses, the sensitivity of phosphinothricin (0, 0.4, 0.8, 1.2, 1.6 mg/l) was tested in the screening of transgenic S. miltiorrhiza. We identified that 0.6 mg/l phosphinothricin should be suitable for selecting putatively transformed callus because non-transformed callus growth was effectively inhibited under this concentrations. When sprayed with Basta, the transgenic S. miltiorrhiza plants were tolerant to the herbicide. Hence, we report successful transformation of the bar gene conferring herbicide resistance to S. miltiorrhiza.

Investigating changes in the redox state of Photosystem I at low pH.

Changes in the redox state of Photosystem I (PSI) were studied in spinach leaf discs suspended in buffers of different pH (pH 7.5, 6.5, 5.5 and 4.5). By measuring absorbance changes at 820 nm, it was observed that under normal conditions, the electrons were supplied by Photosystem II (PSII) for the photo-oxidation of P700 while in the presence of DCMU when electrons coming from PSII are blocked, cyclic electron flow (CEF) around PSI was the major source for the absorbance changes observed at 820 nm. This was supported by complete inhibition in the reduction of both single turnover (ST) area and multiple turnover (MT) area, in the presence of DCMU, which is generally filled up by the electrons coming from PSII. In the absence of DCMU, the intersystem electron pool or plastoquinone (PQ) pool was increased at low pH which was probably due to enhanced cyclic electron flow around PSI. Our results also suggest that at low pH, in the absence of DCMU, the major contribution for faster dark re-reduction of P700(+) is attributed mainly by PSII and CEF PSI while in the presence of DCMU, the significant contribution is provided by CEF PSI and other stromal components.

Non-destructive models to estimate leaf area on bell pepper crop / Modelos não-destrutivos para estimar a área foliar na cultura do pimentão

ABSTRACT: The leaf area determination are extremely important in agronomic and physiological studies, as they provide valuable information in vegetable metabolism, methods used for this purpose should be accurate. This study aimed to develop polynomial models to estimate leaf area (LA). The methods used were leaf discs and ImageJ(r) software, with measures of length (L), width (W), and relationship between length times width (LW) in leaves of the bell pepper crop in two field trials with two hybrids: Arcade and Impacto, as well as, the correlation of values found when using the two methods. Through a random sampling, 560 leaves from each hybrid were selected, at different stages of crop growth. Regression analysis of LA versus measures L, W and LW were performed by linear and quadratic models. The variable that best estimated LA was the relationship between LW with linear model (r2=0.98). Measurements of length and width of leaves presented adjustment to quadratic model r2=0.93 and r2=0.91, respectively. The integrator model for all samples and both hybrids was: LA=0.57×LW. The destructive method of leaf discs showed high correlation (r2=0.99) with the non-destructive method of ImageJ(r) software.

RESUMO: A determinação da área foliar é importante, em estudos agronômicos e fisiológicos, para a obtenção de informações sobre o metabolismo vegetal e as técnicas utilizadas devem ser simples. O objetivo deste estudo foi desenvolver modelos polinomiais para estimar a área foliar (AF). Foram utilizados os métodos de discos foliares e software ImageJ, com as medidas do comprimento (C) e da largura (L), e a relação entre o comprimento vezes a largura (CL) em folhas da cultura do pimentão, em dois ensaios com dois híbridos, Arcade e Impacto, e correlacionar os valores encontrados pelos dois métodos. Através de amostragem aleatória simples, foram selecionadas 560 folhas de cada híbrido, em diferentes estágios de crescimento da cultura. A análise de regressão da área foliar versus as medidas de C, L e CL foi realizada por modelos lineares e quadráticos. A variável que melhor estimou a AF foi a relação entre o CL, pelo modelo linear com r2=0,98. As medidas do comprimento e da largura das folhas apresentou ajustamento do modelo quadrática de r2=0,93 e r2=0,91, respectivamente. O modelo integrador de todas as amostras, para ambos os híbridos, foi: AF=0,57×CL. O método destrutivo por discos mostrou alta correlação (r2=0,99) com o método não destrutivo pelo ImageJ.