Postharvest Withering Research Articles

From bioprotective effects to diversification of wine aroma: Expanding the knowledge on Metschnikowia pulcherrima oenological potential

Microbial diseases are of major concern in vitiviniculture as they cause grape losses and wine alterations, but the prevention with chemical substances represents a risk to human health and agricultural ecosystem. A promising alternative is the biocontrol and bioprotection activity of non-Saccharomyces yeasts, such as Metschnikowia pulcherrima, which also presents positive oenological traits when used in multistarter fermentations. The aim of this study was to assess the impact of a selected M. pulcherrima strain in the post-harvest withering and vinification of Garganega grapes to produce the sweet ‘passito’ wine Recioto di Gambellara DOCG (Italy). M. pulcherrima was firstly inoculated on grape at the beginning of the withering process, and afterwards in must for multistarter sequential microfermentation trials with Saccharomyces cerevisiae. Microbiological, chemical, and sensory analyses were carried out to monitor the vinification of treated and control grapes. Grape bunches during withering were a suitable environment for the colonization by M. pulcherrima, which effectively prevented growth of molds. Differences in grape must composition were observed, and the diverse inoculation strategies caused noticeable variations of fermentation kinetics, main oenological parameters, wine aroma profile, and sensory perception. M. pulcherrima proved effective to protect grapes against fungal infections during withering and contribute to alcoholic fermentation generating wine with distinguished aromatic characteristics.

Temperature affects organic acid, terpene and stilbene metabolisms in wine grapes during postharvest dehydration.

The partial dehydration of grapes after harvest is a traditional practice in several winegrowing regions that leads to the production of high quality wines. Postharvest dehydration (also known as withering) has a significant impact on the overall metabolism and physiology of the berry, yielding a final product that is richer in sugars, solutes, and aroma compounds. These changes are, at least in part, the result of a stress response, which is controlled at transcriptional level, and are highly dependent on the grape water loss kinetics and the environmental parameters of the facility where grapes are stored to wither. However, it is difficult to separate the effects driven by each single environmental factor from those of the dehydration rate, especially discerning the effect of temperature that greatly affects the water loss kinetics. To define the temperature influence on grape physiology and composition during postharvest dehydration, the withering of the red-skin grape cultivar Corvina (Vitis vinifera) was studied in two conditioned rooms set at distinct temperatures and at varying relative humidity to maintain an equal grape water loss rate. The effect of temperature was also studied by withering the grapes in two unconditioned facilities located in geographic areas with divergent climates. Technological, LC-MS and GC-MS analyses revealed higher levels of organic acids, flavonols, terpenes and cis- and trans-resveratrol in the grapes withered at lower temperature conditions, whereas higher concentrations of oligomeric stilbenes were found in the grapes stored at higher temperatures. Lower expression of the malate dehydrogenase and laccase, while higher expression of the phenylalanine ammonia-lyase, stilbene synthase and terpene synthase genes were detected in the grapes withered at lower temperatures. Our findings provide insights into the importance of the temperature in postharvest withering and its effect on the metabolism of the grapes and on the quality of the derived wines.

The Tea Plant Leaf Cuticle: From Plant Protection to Tea Quality.

Camellia sinensis (tea tree) is a perennial evergreen woody crop that has been planted in more than 50 countries worldwide; its leaves are harvested to make tea, which is one of the most popular nonalcoholic beverages. The cuticle is the major transpiration barrier to restrict nonstomatal water loss and it affects the drought tolerance of tea plants. The cuticle may also provide molecular cues for the interaction with herbivores and pathogens. The tea-making process almost always includes a postharvest withering treatment to reduce leaf water content, and many studies have demonstrated that withering treatment-induced metabolite transformation is essential to shape the quality of the tea made. Tea leaf cuticle is expected to affect its withering properties and the dynamics of postharvest metabolome remodeling. In addition, it has long been speculated that the cuticle may contribute to the aroma quality of tea. However, concrete experimental evidence is lacking to prove or refute this hypothesis. Even though its relevance to the abiotic and biotic stress tolerance and postharvest processing properties of tea tree, tea cuticle has long been neglected. Recently, there are several studies on the tea cuticle regarding its structure, wax composition, transpiration barrier organization, environmental stresses-induced wax modification, and structure–function relations. This review is devoted to tea cuticle, the recent research progresses were summarized and unresolved questions and future research directions were also discussed.

Effects of Traditional and Modern Post-Harvest Withering Processes on the Composition of the Vitis v. Corvina Grape and the Sensory Profile of Amarone Wines.

In the Valpolicella area (Verona, Italy) Vitis vinifera cv. Corvina is the main grape variety used to produce Amarone wine. Before starting the winemaking process, the Corvina grapes are stored in a withering (i.e., dehydrating) warehouse until about 30% of the berry weight is lost (WL). This practice is performed to concentrate the metabolites in the berry and enrich the Amarone wine in aroma and antioxidant compounds. In compliance with the guidelines and strict Amarone protocol set by the Consorzio of Amarone Valpolicella, withering must be carried out by setting the grapes in a suitable environment, either under controlled relative air humidity (RH) conditions and wind speed (WS)—no temperature modification is to be applied—or, following the traditional methods, in non-controlled environmental conditions. In general, the two processes have different dehydration kinetics due to the different conditions in terms of temperature, RH, and WS, which affect the accumulation of sugars and organic acids and the biosynthesis of secondary metabolites such as stilbenes and glycoside aroma precursors. For this study, the two grape-withering processes were carried out under controlled (C) and non-controlled (NC) conditions, and the final compositions of the Corvina dried grapes were compared also to evaluate the effects on the organoleptic characteristics of Amarone wine. The findings highlighted differences between the two processes mainly in terms of the secondary metabolites of the dried grapes, which affect the organoleptic characteristics of Amarone wine. Indeed, by the sensory evaluation, wines produced by adopting the NC process were found more harmonious, elegant, and balanced. Finally, we can state how using a traditional system, grapes were characterised by higher levels of VOCs (volatile compounds), whilst wines had a higher and appreciable complexity and finesse.

Not just shrivelling: time-series profiling of the biochemical changes in Corvina (<i>Vitis vinifera</i> L.) berries subjected to post-harvest withering

The grape berry withering process is often seen as a means to concentrate the constituents of the berry via water removal, however, recent molecular studies have indicated that the reprogramming of biosynthetic pathways that impart the unique aroma and flavour of the final wine also occur. Metabolic analysis was performed using GC-MS and LC-MS on Corvina berry dehydrated for up to 108 days. The temporal pattern of metabolic changes and time-series relationship between various classes of metabolites were investigated on berries sampled at 23-time points. Principal component analysis of both GC-MS and LC-MS datasets revealed three distinct phases of post-harvest withering; early (day 0–28), mid (day 35–66) and late (day 69–108) stages. Stress-associated amino acids such as proline, serine, ethanolamine, and leucine accumulated significantly during the mid and late stages of the drying process while phosphorylated glucose and fructose, sucrose, kestose and resveratrol increased massively across time. Unlike most of the identified metabolites, low molecular weight flavanols exhibited a consistent pattern of decline during the withering period. Our network analysis revealed that increased metabolic network connectivity occurred during the middle stage of withering, thus reflecting more coordinated metabolite changes while reduced network connectivity at early and late stages indicates minimal metabolite perturbation during these stages. The current prolonged berry withering experiment revealed the timing of metabolite interconversions in the central metabolism and provided critical clues that link the concentration effect, protein degradation, and the onset of stress-like conditions in drying berries.

Changes in water loss and cell wall metabolism during postharvest withering of tobacco (Nicotiana tabacum L.) leaves using tandem mass tag-based quantitative proteomics approach

Withering is an important biological process accompanied by dehydration and cell wall metabolism in postharvest plant organs during curing/processing and storage. However, dynamics involved in cell wall metabolism and resultant water loss during withering in postharvest tobacco leaves is not well-documented. Here, tandem mass tag (TMT)-based quantitative proteomic analysis in postharvest tobacco leaves (cultivar K326) under different withering conditions was performed. In total, 11,556 proteins were detected, among which 496 differentially abundant proteins (DAPs) were identified. To elucidate the withering mechanism of tobacco leaves, 27 DAPs associated with cell wall metabolism were screened. In particular, pectin acetylesterases, glucan endo-1,3-beta-glucosidases, xyloglucan endotransglucosylase/hydrolase, alpha-xylosidase 1-like, probable galactinol-sucrose galactosyltransferases, endochitinase A, chitotriosidase-1-like and expansin were the key proteins responsible for the withering of postharvest tobacco leaves. These DAPs were mainly involved in pectin metabolism, cellulose, hemicellulose and galactose metabolism, amino sugar and nucleotide sugar metabolism as well as cell wall expansion. Furthermore, relative water content and softness values were significantly and positively correlated. Thus, dehydration and cell wall metabolism were crucial for tobacco leaf withering under different conditions. Nine candidate DAPs were confirmed by parallel reaction monitoring (PRM) technique. These results provide new insights into the withering mechanism underlying postharvest physiological regulatory networks in plants/crops.

Specific molecular interactions between Vitis vinifera and Botrytis cinerea are required for noble rot development in grape berries

Under peculiar climatic conditions, the beneficial form of the necrotrophic fungus Botrytis cinerea can develop on grape berries as a latent infection, known as noble rot, which induces positive biochemical and metabolic changes in the berries, including an increase in the sugar content and the production of aromatic compounds that improve wine quality. The infected berries undergo rapid withering, which is required to produce famous sweet white wines such as Sauternes and Tokaj. To gain insight into the molecular interactions between grapevine berries (Vitis vinifera) and B. cinerea during the establishment of noble rot, we prepared a large-scale transcriptomics dataset representing noble rot development and carried out a comparative meta-analysis with gray mold infection and natural post-harvest withering. In particular, we artificially induced berry botrytization of two grape varieties (Garganega and Möller-Thurgau) and sampled them at different stages of noble rot for comparative whole-transcriptome analysis, highlighting important common transcriptional reprogramming in both varieties reflecting an accelerated withering process. Simultaneously, we analyzed the modulation of B. cinerea genes and compared the expression profile during noble rot development with the previously reported gray mold infection profile, revealing the onset of an infection process by the fungus in its beneficial form associated with reduced virulence. This, together with the restrained plant defense response observed in botrytized berries, may favour the development of noble rot instead of gray mold. Finally, the comprehensive meta-analysis of gene expression during noble rot infection, gray mold and post-harvest withering led to the identification of key genes specifically modulated during noble rot infection.

Expression patterns of alpha-amylase and beta-amylase genes provide insights into the molecular mechanisms underlying the responses of tea plants (Camellia sinensis) to stress and postharvest processing treatments.

The alpha-amylase and beta-amylase genes have been identified from tea plants, and their bioinformatic characteristics and expression patterns provide a foundation for further studies to elucidate their biological functions. Alpha-amylase (AMY)- and beta-amylase (BAM)-mediated starch degradation plays central roles in carbohydrate metabolism and participates extensively in the regulation of a wide range of biological processes, including growth, development and stress response. However, the AMY and BAM genes in tea plants (Camellia sinensis) are poorly understood, and the biological functions of these genes remain to be elucidated. In this study, three CsAMY and nine CsBAM genes from tea plants were identified based on genomic and transcriptomic database analyses, and the genes were subjected to comprehensive bioinformatic characterization. Phylogenetic analysis showed that the CsAMY proteins could be clustered into three different subfamilies, and nine CsBAM proteins could be classified into four groups. Putative catalytically active proteins were identified based on multiple sequence alignments, and the tertiary structures of these proteins were analyzed. Cis-element analysis indicated that CsAMY and CsBAM were extensively involved in tea plant growth, development and stress response. In addition, the CsAMY and CsBAM genes were differentially expressed in various tissues and were regulated by stress treatments (e.g., ABA, cold, drought and salt stress), and the expression patterns of these genes were associated with the postharvest withering and rotation processes. Taken together, our results will enhance the understanding of the roles of the CsAMY and CsBAM gene families in the growth, development and stress response of tea plants and of the potential functions of these genes in determining tea quality during the postharvest processing of tea leaves.

Active rearrangements in the cell wall follow polymer concentration during postharvest withering in the berry skin of Vitis vinifera cv. Corvina

During grape postharvest withering, a worldwide practice used to produce important high-quality wines, the solute concentration increases due to dehydration, and many organoleptic and quality traits, especially related to the berry skin, are affected in a cultivar-specific manner. Nevertheless, a complete comprehension of the underlying processes is still lacking.In this work, we applied ATR-FTIR micro-spectroscopy combined with PCA to monitor cell wall biochemical changes at three stages during postharvest withering on the internal and external sides of the berry skin of the Vitis vinifera cv. Corvina, an important local variety of the Verona province in Italy. The obtained results were integrated by profiling xylogucans and pectins through immunohistochemistry and by genome-wide transcriptomic analysis performed at the same withering stages.Our analysis indicates a gradual passive polymer concentration due to water loss in the first two months of postharvest withering, followed by active structural modifications in the last month of the process. Such rearrangements involve xyloglucans in the internal surface, cuticle components and cellulose in the external surface, and pectins in both surfaces. Moreover, by investigating the expression trend of cell wall metabolism-related genes, we identified several putative molecular markers associated to the polymer dynamics.The present study represents an important step towards an exhaustive comprehension of the postharvest withering process, which is of great interest from both the biological and technological points of view.

The Induction of Noble Rot (Botrytis cinerea) Infection during Postharvest Withering Changes the Metabolome of Grapevine Berries (Vitis vinifera L., cv. Garganega).

The natural or induced development of noble rot caused by the fungus Botrytis cinerea during the late stages of grapevine (Vitis vinifera L.) berry ripening is used in some traditional viticulture areas to produce high-quality wines such as Sauternes and Tokaji. In this research, we wanted to verify if by changing the environmental conditions during post-harvest withering we could induce the noble rot development on harvested berries in order to positively change the wine produced from withered Garganega berries. Therefore, we exposed the berries to postharvest withering under normal or artificially humid conditions, the latter to induce noble rot. The presence of noble rot symptoms was associated with the development of B. cinerea in the berries maintained under humid conditions. The composition of infected and non-infected berries was investigated by untargeted metabolomics using liquid chromatography/mass spectrometry. We also explored the effects of the two withering methods on the abundance of volatile organic compounds in wine by yeast-inoculated micro-fermentation followed by targeted gas chromatography/mass spectrometry. These experiments revealed significant metabolic differences between berries withered under normal and humid conditions, indicating that noble rot affects berry metabolism and composition. As well as well-known botrytization markers, we detected two novel lipids that have not been observed before in berries infected with noble rot. Unraveling the specific metabolic profile of berries infected with noble rot may help to determine the compounds responsible for the organoleptic quality traits of botrytized Garganega wines.

Application of enzyme preparations for extraction of berry skin phenolics in withered winegrapes

Postharvest withering of grapes strongly affects the content and extractability of phenolic compounds in the production of sfursat, fortified and passito wines. This work evaluated the effectiveness of enzymes applied individually and/or in multi-enzyme blends, on the extraction of anthocyanins, oligomeric flavanols and polymeric flavanols from withered grape skins during simulated maceration. The study was performed on Vitis vinifera L. cv. Nebbiolo and Barbera because of their different skin phenolic profile and cell wall composition. Our findings highlight that the relationship between skin mechanical properties (berry skin break force and energy) and extraction yield of phenolic compounds is variety dependent. Significant correlations were found between the skin softening associated with cell wall degradation and the extraction of anthocyanins and flavanols in Nebbiolo, for which polygalacturonase, individually or in multi-enzyme blends, plays an important role. In Barbera, the extractability of phenolic compounds was not affected by the presence of exogenous enzymes.

Management of postharvest grape withering to optimise the aroma of the final wine: A case study on Amarone

Amarone wine is different from regular dry wine due to the postharvest withering of Corvina, Corvinone and Rondinella grapes. Grapes were withered in a commercial facility with variability in terms of temperature and relative humidity (R.H.). Sugar content reached 230–240gL−1 and 280gL−1 at 20% and 30% mass loss, respectively. Most of VOCs (volatile organic compounds) decreased during withering but few VOCs increased during withering and we considered as markers; in Corvinone they were methylhexanoate, dimethylsuccinate, nerol, nonanoic acid, and benzyl alcohol; in Corvina, benzyl alcohol, isoamyl alcohol, 1-hexanol, p-cymen-8-ol, 2,3 pinanediol, 3-oxo-ionol and 3-methyl-1-pentanol, coumaran and damascenone; in Rondinella, hexanol, nonanoic acid, methyl vanillate, damascenone, 3-oxo-ionol, eugenol, p-cymen-8-ol, 2,3 pinanediol, coumaran and raspberry keton. Olfactive descriptors of the wines and the potential aroma of the combination of Corvina wine with the wines of the other two varieties at different percentages of mass loss are reported.

Assessment of Postharvest Dehydration Kinetics and Skin Mechanical Properties of “Muscat of Alexandria” Grapes by Response Surface Methodology

The dipping of berries in a dilute solution of sodium hydroxide during a short time was evaluated as pretreatment undertaken prior to convective dehydration of wine grapes. The impact of the sodium hydroxide content and dipping time on weight loss (WL) at different dehydration times was thoroughly assessed using central composite design (CCD) and response surface methodology (RSM). Furthermore, the effects of these two variables were also investigated on the skin mechanical properties of dehydrated grapes. The effect of these two pretreatment factors on the dehydration kinetics and skin hardness was satisfactorily fitted to regression models. The berry pretreatment with low sodium hydroxide contents (from 10 to 20 g/L) facilitated the dehydration process during the first 5 days when dipping times longer than 300 s were used. From the seventh day of dehydration, at which time the average WL% was close to 50, the highest values of WL% were obtained using intermediate sodium hydroxide contents and dipping times (around 45 g/L and 185 s, respectively). Because skin hardness affects the dehydration kinetics during postharvest withering, the strongest decrease in skin hardness corresponded to these last berry pretreatment conditions, whereas the greatest increase required the highest sodium hydroxide contents and longest dipping times. The quality of berries dehydrated may be influenced by the pretreatment conditions used, and the present study contributes to increase the knowledge on this effect to a better management of the alkaline pretreatment and dehydration process.

Characterization and pathogenicity of Alternaria spp. strains associated with grape bunch rot during post-harvest withering

Alternaria is a fungal agent of grape bunch rot which occurs during withering, a process which produces passito style wines. Seven isolates of Alternaria spp. were characterized using morphological examination, genotypic analysis and pathogenicity. Six of these isolates produced conidiophores and conidia displaying sporulation patterns typical of the Alternaria alternata species-group. Variability in colony morphology and growth on different media was observed. Phylogenetic analysis of internal transcribed spacer (ITS) sequences clustered all isolates within a monophyletic clade, while intergenic spacer region (IGS)-RFLP profiles were congruent with those of A. alternata and Alternaria arborescens. RAPD-PCR proved helpful in discriminating between strains. To assay strain pathogenicity, grape berries were infected while undergoing withering conditions at different temperatures. Disease capacity was found to be strain dependent and varied consistently between the most and least aggressive strains. This study has provided interesting information on polymorphism within Alternaria spp. populations in withered grapes and on understanding the saprophytic role of this fungus during the post-harvest dehydrating process.

Influence of Different Thermohygrometric Conditions on Changes in Instrumental Texture Properties and Phenolic Composition during Postharvest Withering of ‘Corvina’ Winegrapes (Vitis vinifera L.)

‘Corvina’ winegrapes were postharvest withered under three controlled thermohygrometric conditions. Sampling was done on different days of withering and the mechanical properties, technological ripeness parameters, phenolic composition, and anthocyanin profile were determined. Depending on the condition, skin break force and pedicel detachment force significantly decreased during the dehydration process with a maximum variation of −0.258 and −1.306 N, respectively. Instead, total flavonoids of skin and seeds, and proanthocyanidins and low-weight flavanols of seeds increased, with a maximum variation of +1,483, +733, +1,022, and +469 mg/kg, respectively. The skin hardness was confirmed to be an important parameter for the dehydration kinetics.

Two-Dimensional Differential in Gel Electrophoresis (2D-DIGE) Analysis of Grape Berry Proteome during Postharvest Withering

The practice of postharvest withering is commonly used to correct quality traits and sugar concentration of high quality wines. To date, changes in the metabolome during the berry maturation process have been well documented; however, the biological events which occur at the protein level have yet to be fully investigated. To gain insight into the postharvest withering process, we studied the protein expression profiles of grape (Corvina variety) berry development focusing on withering utilizing a two-dimensional differential in gel electrophoresis (2D-DIGE) proteomics approach. Comparative analysis revealed changes in the abundance of numerous soluble proteins during the maturation and withering processes. On a total of 870 detected spots, 90 proteins were differentially expressed during berry ripening/withering and 72 were identified by MS/MS analysis. The majority of these proteins were related to stress and defense activity (30%), energy and primary metabolism (25%), cytoskeleton remodelling (7%), and secondary metabolism (5%). Moreover, this study demonstrates an active modulation of metabolic pathways throughout the slow dehydration process, including de novo protein synthesis in response to the stress condition and further evolution of physiological processes originated during ripening. These data represent an important insight into the withering process in terms of both Vitis germplasm characterization and knowledge which can assist quality improvement.

Molecular analysis of post-harvest withering in grape by AFLP transcriptional profiling

Post-harvest withering of grape berries is used in the production of dessert and fortified wines to alter must quality characteristics and increase the concentration of simple sugars. The molecular processes that occur during withering are poorly understood, so a detailed transcriptomic analysis of post-harvest grape berries was carried out by AFLP-transcriptional profiling analysis. This will help to elucidate the molecular mechanisms of berry withering and will provide an opportunity to select markers that can be used to follow the drying process and evaluate different drying techniques. AFLP-TP identified 699 withering-specific genes, 167 and 86 of which were unique to off-plant and on-plant withering, respectively. Although similar molecular events were revealed in both withering processes, it was apparent that off-plant withering induced a stronger dehydration stress response resulting in the high level expression of genes involved in stress protection mechanisms, such as dehydrin and osmolite accumulation. Genes involved in hexose metabolism and transport, cell wall composition, and secondary metabolism (particularly the phenolic and terpene compound pathways) were similarly regulated in both processes. This work provides the first comprehensive analysis of the molecular events underpinning post-harvest withering and could help to define markers for different withering processes.