Pilar Hellín

Berry Flesh and Skin Ripening Features in Vitis vinifera as Assessed by Transcriptional Profiling

Background Ripening of fleshy fruit is a complex developmental process involving the differentiation of tissues with separate functions. During grapevine berry ripening important processes contributing to table and wine grape quality take place, some of them flesh- or skin-specific. In this study, transcriptional profiles throughout flesh and skin ripening were followed during two different seasons in a table grape cultivar ‘Muscat Hamburg’ to determine tissue-specific as well as common developmental programs. Methodology/Principal Findings Using an updated GrapeGen Affymetrix GeneChip® annotation based on grapevine 12×v1 gene predictions, 2188 differentially accumulated transcripts between flesh and skin and 2839 transcripts differentially accumulated throughout ripening in the same manner in both tissues were identified. Transcriptional profiles were dominated by changes at the beginning of veraison which affect both pericarp tissues, although frequently delayed or with lower intensity in the skin than in the flesh. Functional enrichment analysis identified the decay on biosynthetic processes, photosynthesis and transport as a major part of the program delayed in the skin. In addition, a higher number of functional categories, including several related to macromolecule transport and phenylpropanoid and lipid biosynthesis, were over-represented in transcripts accumulated to higher levels in the skin. Functional enrichment also indicated auxin, gibberellins and bHLH transcription factors to take part in the regulation of pre-veraison processes in the pericarp, whereas WRKY and C2H2 family transcription factors seems to more specifically participate in the regulation of skin and flesh ripening, respectively. Conclusions/Significance A transcriptomic analysis indicates that a large part of the ripening program is shared by both pericarp tissues despite some components are delayed in the skin. In addition, important tissue differences are present from early stages prior to the ripening onset including tissue-specific regulators. Altogether, these findings provide key elements to understand berry ripening and its differential regulation in flesh and skin.

Use of farming and agro-industrial wastes as versatile barriers in reducing pesticide leaching through soil columns.

Increased interest has been recently focused on assessing the influence of the addition of organic wastes related to movement of pesticides in soils of low organic matter (OM) content. This study reports the effect of two different amendments, animal manure (composted sheep manure) and agro-industrial waste (spent coffee grounds) on the mobility of 10 pesticides commonly used for pepper protection on a clay-loam soil (OM = 0.22%). The tested compounds were azoxystrobin, cyprodinil, fludioxonil, hexaconazole, kresoxim-methyl, pyrimethanil, tebuconazole, and triadimenol (fungicides), pirimicarb (insecticide), and propyzamide (herbicide). Breakthrough curves were obtained from disturbed soil columns. Cumulative curves obtained from unamended soil show a leaching of all pesticides although in different proportions (12-65% of the total mass of compound applied), showing triadimenol and pirimicarb the higher leachability. Significant correlation (r = 0.93, p<0.01) was found between the observed and bibliographical values of GUS index. The addition of the amendments used drastically reduced the movement of the studied pesticides. Only two pesticides were found in leachates from amended soils, pyrimethanil (<1%) for both, and pirimicarb (44%) in the soil amended with spent coffee grounds. A decrease in pesticide leaching was observed with the increase in dissolved organic matter (DOM) of leachates. The results obtained point to the interest in the use of organic wastes in reducing the pollution of groundwater by pesticide drainage.

Photocatalytic degradation of five sulfonylurea herbicides in aqueous semiconductor suspensions under natural sunlight

In the present study, the photocatalytic degradation of five sulfonylurea herbicides (chlorsulfuron, flazasulfuron, nicosulfuron, sulfosulfuron and triasulfuron) has been investigated in aqueous suspensions of zinc oxide (ZnO), tungsten (VI) oxide (WO(3)), tin (IV) oxide (SnO(2)) and zinc sulfide (ZnS) at pilot plant scale under natural sunlight. Photocatalytic experiments, especially those involving ZnO photocatalysis, showed that the addition of semiconductors in tandem with the oxidant (Na(2)S(2)O(8)) strongly enhances the degradation rate of the herbicides in comparisons carried out with photolytic tests. The degradation of the herbicides follows a first order kinetics according to the Langmuir-Hinshelwood model. In our conditions, the amount of time required for 50% of the initial pesticide concentration to dissipate (t(½)) ranged from 8 to 27 min (t(30W)=0.3-1.2 min) for sulfosulfuron and chlorsulfuron, respectively in the ZnO/Na(2)S(2)O(8) system. None of the studied herbicides was found after 120 min of illumination (except chlorsulfuron, 0.2 μg L(-1)).

Trace analysis of sulfonylurea herbicides in water samples by solid-phase extraction and liquid chromatography-tandem mass spectrometry.

A sensitive method for the simultaneous determination of 30 sulfonylurea herbicides in tap and leaching waters has been developed. Liquid chromatography tandem-mass spectrometry (LC-MS(2)) in electrospray ionization positive mode was used for the separation, identification and quantification of these compounds. The procedure involves a preconcentration step based on solid-phase extraction with a silica-based bonded C(18) cartridge (Sep-Pak Plus) and a N-vinyl-pyrrolidone polymer cartridge (Oasis HLB). The best results were obtained with Oasis HLB using methanol as elution solvent. Average recoveries of 30 analytes from water samples were in the range of 79-115% with a relative standard deviation of <6.1%. The limits of quantification (LOQs) obtained in tap and leaching water samples were in the range of 0.1-5.9 and 0.4-5.8 ng L(-1), respectively. The proposed method was used to determine sulfonylurea herbicide levels in leaching water samples taken from three lysimeters located in an experimental greenhouse.

Reduction of the movement and persistence of pesticides in soil through common agronomic practices.

Laboratory and field studies were conducted in order to determine the leaching potential of eight pesticides commonly used during pepper cultivation by use of disturbed soil columns and field lysimeters, respectively. Two soils with different organic matter content (soils A and B) were used. Additionally, soil B was amended with compost (sheep manure). The tested compounds were cypermethrin, chlorpyrifos-methyl, bifenthrin, chlorpyrifos, cyfluthrin, endosulfan, malathion and tolclofos-methyl. In soil B (lower organic matter content), only endosulfan sulphate, malathion and tolclofos-methyl were found in leachates. For the soil A (higher organic matter content) and amended soil B, pesticide residues were not found in the leachates. In addition, this paper reports on the use of common agronomic practices (solarization and biosolarization) to enhance degradation of these pesticides from polluted soil A. The results showed that both solarization and biosolarization enhanced the degradation rates of endosulfan, bifenthrin and tolclofos-methyl compared with the control. Most of the studied pesticides showed similar behavior under solarization and biosolarization conditions. However, chlorpyrifos was degraded to a greater extent in the solarization than in biosolarization treatment. The results obtained point to the interest in the use of organic amendment in reducing the pollution of groundwater by pesticide drainage and in the use of solarization and biosolarization in reducing the persistence of pesticides in soil.

Prediction of Muscat aroma in table grape by analysis of rose oxide.

Aroma is an important quality characteristic in Muscat grapes and constitutes a major concern for viticulturist and grapevine breeders. For this reason, Muscat aroma variability was characterised in a segregating progeny and in a collection of table grapes, to assess the usefulness of the presence or absence of rose oxide for predicting Muscat genotypes. Simple tasting and an analysis of free and bound aroma compounds, including rose oxide, linalool oxide, linalool, α-terpineol, citronellol, nerol, geraniol, benzyl alcohol and 2-phenylethanol, were carried out. The association between Muscat score and the compounds considered as active odorants according to their odour activity values was also evaluated. The results obtained pointed to a highly significant correlation between the presence/absence of rose oxide in grapes and the presence/absence of Muscat aroma. Thus, this analysis could be a useful tool for identifying Muscat cultivars in a more objective way than sensory analysis.

Leaching potential of several insecticides and fungicides through disturbed clay-loam soil columns

A laboratory study was conducted in order to determine the leaching potential of five insecticides and six fungicides commonly used during pepper cultivation by use of disturbed soil columns. The tested compounds were pyridaben, pyriproxyfen, tebufenpyrad, buprofezin and pirimicarb (insecticides/acaricides) and azoxyxtrobin, kresoxim-methyl, hexaconazole, tebuconazole, triadimenol, and pyrimethanil (fungicides). For this purpose, 100 µg of each pesticide were added to columns (n = 5) filled with 150 g of a clay loam soil and leached with 600 mL of 0.01 M CaCl2 during 10 days. Finally, leachates and soil fractions were analysed for pesticide residues. For insecticides only pirimicarb was found in leachates (48% of the initial amount) while 55% was recovered from the soil layers. For the other studied insecticides, the percentage remaining in the top soil fraction was nearly the total amount added and therefore showing ‘non-leaching’ behaviour. In the case of fungicides 41% of triadimenol and 6% of pyrimethanil were found in leachates. The amount recovered in leachates for the other fungicides was lower than 2%, hexaconazole and tebuconazole exhibiting the higher retention on the upper layer of soil. Therefore, is extremely important to propose methods and conduct to avoid the potential adverse effect of pesticides behaving as ‘leacher’ compounds.

High performance liquid chromatography–tandem mass spectrometry method for quantifying phenylurea herbicides and their main metabolites in amended and unamended soils

A sensitive multiresidue method for the simultaneous determination of sixteen phenylurea herbicides and their main metabolites in amended soils has been developed. Liquid chromatography tandem-mass spectrometry (LC-MS²) in electrospray ionization positive mode was used for the separation, identification and quantification of these compounds. The procedure involves initial single phase extraction of soil sample with acetonitrile by sonication, followed by liquid-liquid partitioning formed by addition of NaCl. The average recovery by the LC-MS² method obtained for these compounds varied from 76.2 to 107.9% with a relative standard deviation ranging from 2.1 to 5.8%. The method presents good linearity (R²>0.99) over the range assayed 10-1000 μg L⁻¹ (except N-phenylurea 50-1000 μg L⁻¹). The detection limits for the compounds studied varied from 0.1 to 9.0 ng g⁻¹.

Microbial Quality and Bioactive Constituents of Sweet Peppers from Sustainable Production Systems

Integrated, organic, and soil-less production systems are the principal production practices that have emerged to encourage more sustainable agricultural practices and safer edible plants, reducing inputs of plaguicides, pesticides, and fertilizers. Sweet peppers grown commercially under integrated, organic, and soil-less production systems were compared to study the influence of these sustainable production systems on the microbial quality and bioactive constituents (vitamin C, individual and total carotenoids, hydroxycinnamic acids, and flavonoids). The antioxidant composition of peppers was analyzed at green and red maturity stages and at three harvest times (initial, middle, and late season). Irrigation water, manure, and soil were shown to be potential transmission sources of pathogens to the produce. Coliform counts of soil-less peppers were up to 2.9 log units lower than those of organic and integrated peppers. Soil-less green and red peppers showed maximum vitamin C contents of 52 and 80 mg 100 g(-1) fresh weight (fw), respectively, similar to those grown in the organic production system. Moreover, the highest content of total carotenoids was found in the soil-less red peppers, which reached a maximum of 148 mg 100 g(-1) fw, while slightly lower contents were found in integrated and organic red peppers. Hydroxycinnamic acids and flavonoids represented 15 and 85% of the total phenolic content, respectively. Total phenolic content, which ranged from 1.2 to 4.1 mg 100 g(-1) fw, was significantly affected by the harvest time but not by the production system assayed. Soil-less peppers showed similar or even higher concentrations of bioactive compounds (vitamin C, provitamin A, total carotenoid, hydroxycinnamic acids, and flavonoids) than peppers grown under organic and integrated practices. Therefore, in the commercial conditions studied, soil-less culture was a more suitable alternative than organic or integrated practices, because it improved the microbial safety of sweet peppers without detrimental effects on the bioactive compound content.

Heterogeneous photocatalytic oxidation of cyprodinil and fludioxonil in leaching water under solar irradiation.

The efficiency of ZnO and TiO(2) suspensions in the photocatalytic degradation of two fungicides (cyprodinil and fludioxonil) in leaching water was investigated. The experiments were carried out at pilot plant scale using compound parabolic collectors under natural sunlight. The blank experiments for both irradiated compounds solutions showed that both oxides strongly enhanced the removal of the fungicides. The addition of an oxidant (Na(2)S(2)O(8)) to the ZnO or TiO(2) increased the rate of photooxidation. The degradation of cyprodinil and fludioxonil followed first order kinetics according to the Langmuir-Hinshelwood model. Complete degradation of both fungicides was achieved within 4 h (t(30W)=18 min) when treated with illuminated ZnO. The disappearance time (DT(75)), when referred to the normalized illumination time (t(30W)), was lower than 40 and 550 min (t(30W)=2 and 40 min) for both fungicides using ZnO or TiO(2), respectively. ZnO appeared to be more effective in cyprodinil and fludioxonil oxidation than TiO(2) probably due to its nonstoichiometry.