Macrowine 2021
IVES 9 IVES Conference Series 9 Ochratoxin a degradation by Botrytis cinerea laccase: effect of oenological factors and redox mediators

Ochratoxin a degradation by Botrytis cinerea laccase: effect of oenological factors and redox mediators

Abstract

AIM: This study evaluates the effect of different oenological factors and natural mediators on the degradation of Ochratoxin A (OTA) using Botrytis cinerea laccase. Because of its risk to human health, different detoxification techniques have been developed in various kinds of foodstuffs. The use of fungal or bacterial laccases is a biological method to decrease the OTA concentration [1, 2]. Laccases can oxidize a wide range of substrates, some of which cannot be oxidized directly by these enzymes and require the use of redox mediators [3]. Due to this, several natural mediators present in wine and different SO2 and ethanol concentrations were tested in the current work.

METHODS: The ability of laccase to degrade OTA was studied by incubation of the enzyme in acetate buffer pH 4.0 and model wine, with OTA and mediators at 28 ºC during 24 h. To determine the impact of SO2 and ethanol on the OTA degradation caused by laccase, different concentrations of SO2 (10, 20 and 30 mg/L) and ethanol (5, 10 and 15% v/v) were used. The quantification of this mycotoxin was carried out in a HPLC-QTOF-MS system.

RESULTS: Under these conditions, OTA cannot be oxidized directly by laccase from Botrytis cinerea and the use of redox mediators is required. Among natural mediators tested, (-)-epicatechin and (+)-catechin were the phenolic compounds with higher impact on the biodegradation of this mycotoxin, achieving a decrease of OTA concentration over 50%. The degradation of OTA was completely inhibed by 30 mg of SO2/L,  while 20 mg of SO2/L reduced lacasse activity by a half and 10 mg of SO2/L hardly caused any effect on the biodegradation of this mycotoxin. A concentration of 15% of ethanol led to a 50% reduction in the activity of laccase over OTA. 

CONCLUSIONS

These preliminary results may be a first step in finding biological alternative strategies to eliminate undesirable substances such as mycotoxins (OTA) present in wine.

DOI:

Publication date: September 14, 2021

Issue: Macrowine 2021

Type: Article

Authors

osé Pérez-Navarro

Regional Institute for Applied Scientific Research (IRICA), University of Castilla-La Mancha, Av. Camilo José Cela 10, 13071 Ciudad Real, Spain. Higher Technical School of Agronomic Engineering, University of Castilla-La Mancha, Ronda de Calatrava 7, 13071 Ciudad Real, Spain.,Tania, PANIAGUA MARTÍNEZ, Faculty of Chemical Sciences and Technologies, University of Castilla-La Mancha, Av. Camilo José Cela, 10, 13071 Ciudad Real, Spain. Pol, GIMÉNEZ, Faculty of Oenology, University of Rovira I Virgili, C/Marcel.li Domingo s/n, 43007 Tarragona, Spain. Joan Miquel, CANALS, Faculty of Oenology, University of Rovira I Virgili, C/Marcel.li Domingo s/n, 43007 Tarragona, Spain. Fernando, ZAMORA, Faculty of Oenology, University of Rovira I Virgili, C/Marcel.li Domingo s/n, 43007 Tarragona, Spain. Sergio, GÓMEZ-ALONSO, Regional Institute for Applied Scientific Research (IRICA), University of Castilla-La Mancha, Av. Camilo José Cela 10, 13071 Ciudad Real, Spain. Faculty of Chemical Sciences and Technologies, University of Castilla-La Mancha, Av. Camilo José Cela, 10, 13071 Ciudad Real, Spain.

Contact the author

Keywords

mycotoxin, enzyme, biodetoxification, fungi, SO2, ethanol

Citation

Related articles…

Projected changes in vine phenology of two varieties with different thermal requirements cultivated in La Mancha DO (Spain) under climate change scenarios

The aim of this work was to analyze the phenology variability of Tempranillo and Chardonnay cultivars, related to the climatic characteristics in La Mancha Designation of Origin, and their potential changes under climate change scenarios. Phenological dates referred to budbreak, flowering, veraison and harvest were analyzed for the period 2000-2019. The weather conditions at daily time scale, recorded during the same period, were also evaluated. The thermal requirements to reach each of these phenological stages were calculated and expressed as the GDD accumulated from DOY=60. Changes in phenology were projected by 2050 and 2070 taking into account those values and the projected temperatures and precipitation, simulated under two Representative Concentration Pathway (RCP) scenarios –RCP4.5 and RCP8.5– using an ensemble of models. The average phenological dates during the period under study were, April 16th ± 6.6 days and April 5th ± 6.0 days for budbreak, May 31st ± 6.0 days and May 27th ± 5.3 days for flowering, July 26th ± 5.6 days and July 25th ± 5.8 days for veraison, and Ago 23rd ± 10.8 days and Ago 17th ± 9.0 days for harvest, respectively, for Tempranillo and Chardonnay. The projected changes in temperature imply an average change in the maximum growing season (April-August) temperatures of 1.2 and 1.9°C by 2050, and 1.6 and 2.6°C by 2070, under the RCP4.5 and RCP8.5 scenarios, respectively. A reduction in precipitation is predicted, which vary between 15% for 2050 under RCP4.5 scenario and up to 30% by 2070 under RCP8.5. The advance of the phenological dates for 2050, could be of 6, 7, 7, and 8 days for Tempranillo and 4, 6, 6 and 9 days for Chardonnay, respectively for budbreak, flowering, veraison and harvest under the RCP4.5 scenario. Under the RCP8.5 emission scenario, the advance could be up to 30% higher.

Investigating the impact of grape exposure and UV radiations on rotundone in Vitis vinifera L. Tardif grapes under field trial conditions

Rotundone is the main aroma compound responsible for peppery notes in wines whose biosynthesis is negatively affected by heat and drought. Through the alteration of precipitation regime and the increase in temperature during maturation, climate change is expected to affect wine peppery typicality. In this context there is a demand for developing sustainable viticultural strategies to enhance rotundone accumulation or limit its degradation. It was recently proposed that ultraviolet (UV) radiations could stimulate rotundone production. The aim of this study was to investigate under field trial conditions the impact of grape exposure and UV treatments on rotundone in Vitis vinifera L. Tardif, an almost extinct grape variety from south-west France that can express particularly high rotundone levels. Four different treatments were compared in 2021 to a control treatment using a randomised complete block design with three replications per treatment. Grape exposure was manipulated through early or late defoliation. Leaf and laterals shoots were removed at Eichorn Lorenz growth stages 32 or 34 on the morning-sun side of the canopy. During grape maturation, UV radiations were either reduced by 99% by installing UV radiation-shielding sheets, or applied four times using the Boxilumix™ non thermal device (Asclepios Tech, Tournefeuille) with the aim of activating plant signalling pathway. Loggers displayed in solar radiation shields were used to assess the effect of such shielding sheets on air temperature within the bunch zone. The composition of grapes subjected to these treatments will be soon analysed for their rotundone content and basic classical laboratory analyses. Grapes will be harvested to elaborate wines under standardized small-scale vinification conditions (60kg) that will be assessed by a trained sensory panel.

Using δ13C and hydroscapes as a tool for discriminating cultivar specific drought response

Measurement of carbon isotope discrimination in berry juice sugars at maturity (δ13C) provides an integrated assessment of water use efficiency (WUE) during the period of berry ripening, and when collected over multiple seasons can be used as an indication of drought stress response. Berry juice δ13C measurements were carried out on 48 different varieties planted in a common garden experiment in Bordeaux, France from 2014 through 2021 and were paired with midday and predawn leaf water potential measurements on the same vines in a subset of six varieties. The aim was to discriminate a large panel of varieties based on their stomatal behaviour and potentially identify hydraulic traits characterizing drought tolerance by comparing δ13C and hydroscapes (the visualisation of plant stomatal behaviour as a response to predawn water potential). Cluster analysis found that δ13C values are likely affected by the differing phenology of each variety, resulting in berry ripening of different varieties taking place under different stress conditions within the same year. We accounted for these phenological differences and found that cluster analysis based on specific δ13C metrics created a classification of varieties that corresponds well to our current empirical understanding of their relative drought tolerances. In addition, we analysed the water potential regulation of the subset of six varieties (using the hydroscape approach) and found that it was well correlated with some δ13C metrics. Surprisingly, a variety’s water potential regulation (specifically its minimum critical leaf water potential under water deficit) was strongly correlated to δ13C values under well-watered conditions, suggesting that base WUE may have a stronger impact on drought tolerance than WUE under water deficit. These results give strong insights on the innate WUE of a very large panel of varieties and suggest that studies of drought tolerance should include traits expressed under non-limiting conditions.

Comparison of imputation methods in long and varied phenological series. Application to the Conegliano dataset, including observations from 1964 over 400 grape varieties

A large varietal collection including over 1700 varieties was maintained in Conegliano, ITA, since the 1950s. Phenological data on a subset of 400 grape varieties including wine grapes, table grapes, and raisins were acquired at bud break, flowering, veraison, and ripening since 1964. Despite the efforts in maintaining and acquiring data over such an extensive collection, the data set has varying degrees of missing cases depending on the variety and the year. This is ubiquitous in phenology datasets with significant size and length. In this work, we evaluated four state-of-the-art methods to estimate missing values in this phenological series: k-Nearest Neighbour (kNN), Multivariate Imputation by Chained Equations (mice), MissForest, and Bidirectional Recurrent Imputation for Time Series (BRITS). For each phenological stage, we evaluated the performance of the methods in two ways. 1) On the full dataset, we randomly hold-out 10% of the true values for use as a test set and repeated the process 1000 times (Monte Carlo cross-validation). 2) On a reduced and almost complete subset of varieties, we varied the percentage of missing values from 10% to 70% by random deletion. In all cases, we evaluated the performance on the original values using normalized root mean squared error. For the full dataset we also obtained performance statistics by variety and by year. MissForest provided average errors of 17% (3 days) at budbreak, 14% (4 days) at flowering, 14.5% (7 days) at veraison, and 17% (3 days) at maturity. We completed the imputations of the Conegliano dataset, one of the world’s most extensive and varied phenological time series and a steppingstone for future climate change studies in grapes. The dataset is now ready for further analysis, and a rigorous evaluation of imputation errors is included.

Effect of multi-level and multi-scale spectral data source on vineyard state assessment

Currently, the main goal of agriculture is to promote the resilience of agricultural systems in a sustainable way through the improvement of use efficiency of farm resources, increasing crop yield and quality under climate change conditions. This last is expected to drastically modify plant growth, with possible negative effects, especially in arid and semi-arid regions of Europe on the viticultural sector. In this context, the monitoring of spatial behavior of grapevine during the growing season represents an opportunity to improve the plant management, winegrowers’ incomes, and to preserve the environmental health, but it has additional costs for the farmer. Nowadays, UAS equipped with a VIS-NIR multispectral camera (blue, green, red, red-edge, and NIR) represents a good and relatively cheap solution to assess plant status spatial information (by means of a limited set of spectral vegetation indices), representing important support in precision agriculture management during the growing season. While differences between UAS-based multispectral imagery and point-based spectroscopy are well discussed in the literature, their impact on plant status estimation by vegetation indices is not completely investigated in depth. The aim of this study was to assess the performance level of UAS-based multispectral (5 bands across 450-800nm spectral region with a spatial resolution of 5cm) imagery, reconstructed high-resolution satellite (Sentinel-2A) multispectral imagery (13 bands across 400-2500 nm with spatial resolution of <2 m) through Convolutional Neural Network (CNN) approach, and point-based field spectroscopy (collecting 600 wavelengths across 400-1000 nm spectral region with a surface footprint of 1-2 cm) in a plant status estimation application, and then, using Bayesian regularization artificial neural network for leaf chlorophyll content (LCC) and plant water status (LWP) prediction. The test site is a Greco vineyard of southern Italy, where detailed and precise records on soil and atmosphere systems, in-vivo plant monitoring of eco-physiological parameters have been conducted.