OENO IVAS 2019 banner
IVES 9 IVES Conference Series 9 OENO IVAS 9 OENO IVAS 2019 9 Grape and wine microorganisms: diversity and adaptation 9 Application of high-throughput sequencing tools for characterisation of microbial communities during alcoholic fermentation

Application of high-throughput sequencing tools for characterisation of microbial communities during alcoholic fermentation

Abstract

Developments in high-throughput sequencing (HTS) technologies allow us to obtain large amounts of microbial information from wine and must samples. Thus approaches, that are aimed at characterising the microbial diversity during fermentation, can be enhanced, or possibly even replaced, with HTS-based metabarcoding. To reduce experimental biases and increase data reproducibility, we compared 3 DNA extraction methods by evaluating differences in the fungal diversity with Riesling alcoholic fermentation samples at four different vineyards. The fungal diversity profiling was done using the genetic markers ITS2 and D2 using metabarcoding. The extraction methods compared consisted of a commercial kit, a recently published protocol that includes a DNA enhancer, and a protocol based on a buffer containing common inhibitor removal reagents. All methods were able to distinguish vineyard effects on the fungal diversity, but the results differed quantitatively. 

From the results of extraction methods, we applied the chosen methods and further combined the HTS tools of metabarcoding and metagenomics, to characterise how microbial communities of those samples, and their subsequent spontaneously fermented derivatives, vary. We specifically explored microbial community variation related to vineyard level, and during alcoholic fermentation. The vineyard was shown to be strongly influencing the microbial communities. Functional analyses were additionally included to investigate the microbial interactions. An increase in non-Saccharomycetaceae fungal functions and a decrease in bacterial functions were also observed during the early fermentation stage. Overall, our results highlight the importance of standardizing DNA extraction methods when characterising fungal diversity from wine and related samples, and showcase how metagenomic functional analysis offer possibilities to improve our insights into the wine alcoholic fermentation process, including highlighting microbe interactions.

DOI:

Publication date: June 23, 2020

Issue: OENO IVAS 2019

Type: Article

Authors

Sarah Siu Tze Mak, Kimmo Sirén, Christian Carøe, Ulrich Fischer, M. Thomas P. Gilbert 

Section for Evolutionary Genomics, Natural History Museum of Denmark, University of Copenhagen, Copenhagen, Denmark 
Institute for Viticulture and Oenology, Dienstleistungszentrum Ländlicher Raum Rheinpfalz, Neustadt an der Weinstraße, Germany 

Contact the author

Keywords

Riesling, Metabarcoding, Metagenomics, DNA extraction

Tags

IVES Conference Series | OENO IVAS 2019

Citation

Related articles…

Making sense of available information for climate change adaptation and building resilience into wine production systems across the world

Effects of climate change on viticulture systems and winemaking processes are being felt across the world. The IPCC 6thAssessment Report concluded widespread and rapid changes have occurred, the scale of recent changes being unprecedented over many centuries to many thousands of years. These changes will continue under all emission scenarios considered, including increases in frequency and intensity of hot extremes, heatwaves, heavy precipitation and droughts. Wine companies need tools and models allowing to peer into the future and identify the moment for intervention and measures for mitigation and/or avoidance. Previously, we presented conceptual guidelines for a 5-stage framework for defining adaptation strategies for wine businesses. That framework allows for direct comparison of different solutions to mitigate perceived climate change risks. Recent global climatic evolution and multiple reports of severe events since then (smoke taint, heatwave and droughts, frost, hail and floods, rising sea levels) imply urgency in providing effective tools to tackle the multiple perceived risks. A coordinated drive towards a higher level of resilience is therefore required. Recent publications such as the Australian Wine Future Climate Atlas and results from projects such as H2020 MED-GOLD inform on expected climate change impacts to the wine sector, foreseeing the climate to expect at regional and vineyard scale in coming decades. We present examples of practical application of the Climate Change Adaptation Framework (CCAF) to impacts affecting wine production in two wine regions: Barossa (Australia) and Douro (Portugal). We demonstrate feasibility of the framework for climate adaptation from available data and tools to estimate historical climate-induced profitability loss, to project it in the future and to identify critical moments when disruptions may occur if timely measures are not implemented. Finally, we discuss adaptation measures and respective timeframes for successful mitigation of disruptive risk while enhancing resilience of wine systems.

The combined effects of climate, soils, and deficit irrigation on yield and quality of Touriga Nacional under high atmospheric demand in the Douro Region

Global warming is one of the biggest environmental, social and economic threats in several viticultural regions. In the Douro Valley, changes are expected in the coming years, namely an increase in temperature and a decrease in precipitation. These changes are likely to have consequences for the production and quality of wine.
The aim of this study was to explore the effects of different soil characteristics combined with several deficit irrigation strategies, managed throughout ETc references and predawn leaf water potentials thresholds, on physiology, yield, and qualitative attributes on the Touriga Nacional variety under years of mild to severe water and heat stress.
The studies were conducted over seven years (2015 to 2021) in two plots of a commercial vineyard located at Quinta do Ataíde (Symington Family Estates) planted in 2011 and 2014 at 170 meters elevation, growing under three water regimes: non-irrigated (NI) and two deficit irrigation strategies (30% and 60% ETc) assessed weekly by Ψpd. The site has an annual rainfall below 500 mm, with high atmospheric demand. Climate data was collected from a weather station, located on site. Berry ripening was followed weekly for fruit analysis. At harvest, yield, vigour and pruning weight per vine were determined from 90 vines by treatment. Each season at veraison the NDVI Index was accessed by a drone. The soils physic-chemistry in the experimental blocs were analysed and grouped by SWHC. Delta C-13 analyses were also performed per treatment in two years.Irrigation had a positive effect on yield per vine, mostly due to an increase in berry and cluster weight, and fertility index through the years. A significant increase in sugar content, colour and phenols was observed with deficit irrigation in some years, but vine vigour related to soil characteristics had by far the greatest impact on quality.

Impact of geographical location on the phenolic profile of minority varieties grown in Spain. II: red grapevines

Because terroir and cultivar are drivers of wine quality, is essential to investigate theirs effects on polyphenolic profile before promoting the implantation of a red minority variety in a specific area. This work, included in MINORVIN project, focuses in the polyphenolic profile of 7 red grapevines minority varieties of Vitis vinifera L. (Morate, Sanguina, Santafe, Terriza Tinta Jeromo Tortozona Tinta) and Tempranillo) from six typical viticulture Spanish areas: Aragón (A1), Cataluña (A2), Castilla la Mancha (A3), Castilla –León (A4), Madrid (A5) and Navarra (A6) of 2020 season. Polyphenolic substances were extracted from grapes. 35 compounds were identified and quantified (mg subtance/kg fresh berry) by HPLC and grouped in anthocyanins (ANT) flavanols (FLAVA), flavonols (FLAVO), hydroxycinnamic (AH), benzoic (BA) acids and stilbenes (ST). Antioxidant activity (AA, mmol TE /g fresh berry) was determined by DPPH method. The results were submitted to a two-way ANOVA to investigate the influence of variety, area and their interaction for each polyphenolic family and cluster analysis was used to construct hierarchical dendrograms, searching the natural groupings among the samples. Sanguina (A3) had the most of total polyphenols while Tempranillo (A5) those of ANT. Sanguina (A2) and (A3) reached the highest values of FLAVO, FLAVA and AA. These two last samples had also the maximum of AA. The effect cultivar and area were significant for all polyphenolic families analyzed. A high variability due to variety (>50%) was observed in FLAVA and the maximum value of variability due to growing area was detected in AA (86.41%), ANT and FLAVO (51%); the interaction variety*zone was significant only for ANT, FLAVO, EST and AA. Finally, dendrograms presented five cluster: i) Sanguina (A2); ii) Sanguina (A3); iii) Tempranillo (A5); iv) Tempranillo (A3); Terriza (A3,A5), Morate (A5,A6); v) Santafé (A1,A6); Tortozona tinta (A1,A3,A6); Tinta Jeromo (A3,A4).

Downscaling of remote sensing time series: thermal zone classification approach in Gironde region

In viticulture, the challenges of local climate modelling are multiple: taking into account the local environment, fine temporal and spatial scales, reliable time series of climate data, ease of implementation and reproducibility of the method. At the local scale, recent studies have demonstrated the contribution of spatialization methods for ground-based climate observation data considering topographic factors such as altitude, slope, aspect, and geographic coordinates (Le Roux et al, 2017; De Rességuier et al, 2020). However, these studies have shown questions in terms of the reproducibility and sustainability of this type of climate study. In this context, we evaluated the potential of MODIS thermal satellite images validated with ground-based climate data (Morin et al, 2020). Previous studies have been encouraging, but questions remain to be explored at the regional scale, particularly in the dynamics of the massive use of bioclimatic indices to classify the climate of wine regions. The results at the local scale were encouraging, but this approach was tested in the current study at the regional scale. Several objectives were set: 1) to evaluate the downscaling method for land surface temperature time series, 2) to identify regional thermal structure variations. We used weekly minimum and maximum surface temperature time series acquired by MODIS satellites at a spatial resolution of 1000 m and downscaled at 500 m using topographical variables. Two types of analyses were performed:

Co-design and evaluation of spatially explicit strategies of adaptation to climate change in a Mediterranean watershed

Climate change challenges differently wine growing systems, depending on their biophysical, sociological and economic features. Therefore, there is a need to locally design and evaluate adaptation strategies combining several technical options, and considering the local opportunities and constraints (e.g. water access, wine typicity). The case study took place in a typical and heterogeneous Mediterranean vineyard of 1,500 ha in the South of France. We developed a participatory modeling approach to (1) conceptualize local climate change issues and design spatially explicit adaptation strategies with stakeholders, (2) numerically evaluate their effects on phenology, yield and irrigation needs under the high-emissions climate change scenario RCP 8.5, and (3) collectively discuss simulation results. We organized five sets of workshops, with in-between modeling phases. A process-based model was developed that allowed to evaluate the effects of six technical options (late varieties, irrigation, water saving by reducing canopy size, adjusting cover cropping, reducing density, and shading) with various distributions in the watershed, as well as vineyard relocation. Overall, we co-designed three adaptation strategies. Delay harvest strategy with late varieties showed little effects on decreasing air temperature during ripening. Water constraint limitation strategy would compensate for production losses if disruptive adaptations (e.g. reduced density) were adopted, and more land got access to irrigation. Relocation strategy would foster high premium wine production in the constrained mountainous areas where grapevine is less impacted by climate change. This research shows that a spatial distribution of technical changes gives room for adaptation to climate change, and that the collaboration with local stakeholders is a key to the identification of relevant adaptation. Further research should explore the potential of adaptation strategies based on soil quality improvement and on water stress tolerant varieties.