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…

Impact of climate variability and change on grape yield in Italy

Viticulture is entangled with weather and climate. Therefore, areas currently suitable for grape production can be challenged by climate change. Winegrowers in Italy already experiences the effect of climate change, especially in the form of warmer growing season, more frequent drought periods, and increased frequency of weather extremes. The aim of this study is to investigate the impact of climate variability and change on grape yield in Italy to provide winegrowers the information needed to make their business more sustainable and resilient to climate change. We computed a specific range of bioclimatic indices, selected by the International Organisation of Vine and Wine (OIV), and correlated them to grape yield data. We have worked in collaboration with some wine consortiums in northern and central Italy, which provided grape yield data for our analysis. Using climate variables from the E-OBS dataset we investigate how the bioclimatic indices changed in the past, and the impact of this change on grape productivity in the study areas. The climate impact on productivity is also investigated by using high-resolution convection-permitting models (CPMs – 2.2 horizontal resolution), with the purpose of estimating productivity in future emission scenarios. The CPMs are likely the best available option for this kind of impact studies since they allow a better representation of small-scale processes and features, explicitly resolve deep convection, and show an improved representation of extremes. In our study, we also compare CPMs with regional climate models (RCMs – 12 km horizontal resolution) to assess the added value of high-resolution models for impact studies. Further development of our study will lead to assessing the future suitability for vine cultivation and could lead to the construction of a statistical model for future projection of grape yield.

Mesoclimate impact on Tannat in the Atlantic terroir of Uruguay

The study of climate is relevant as an element conditioning the typicity of a product, its quality and sustainability over the years. The grapevine development and growth and the final grape and wine composition are closely related to temperature, while climate components vary at mesoscale according to topography and/or proximity to large bodies of water. The objective of this work is to assess the mesoclimate of the Atlantic region of Uruguay and to determine the effect of topography and the ocean on temperature and consequently on Tannat grapevine behavior.

Sustaining wine identity through intra-varietal diversification

With contemporary climate change, cultivated Vitis vinifera L. is at risk as climate is a critical component in defining ecologically fitted plant materiel. While winegrowers can draw on the rich diversity among grapevine varieties to limit expected impacts (Morales-Castilla et al., 2020), replacing a signature variety that has created a sense of local distinctiveness may lead to several challenges. In order to sustain wine identity in uncertain climate outcomes, the study of intra-varietal diversity is important to reflect the adaptive and evolutionary potential of current cultivated varieties. The aim of this ongoing study is to understand to what extent can intra-varietal diversity be a climate change adaptation solution. With a focus on early (Sauvignon blanc, Riesling, Grolleau, Pinot noir) to moderate late (Chenin, Petit Verdot, Cabernet franc) ripening varieties, data was collected for flowering and veraison for the various studied accessions (from conservatory plots) and clones. For these phenological growing stages, heat requirements were established using nearby weather stations (adapted from the GFV model, Parker et al., 2013) and model performances were verified. Climate change projections were then integrated to predict the future behaviour of the intra-varietal diversity. Study findings highlight the strong phenotypic diversity of studied varieties and the importance of diversification to enhance climate change resilience. While model performances may require improvements, this study is the first step towards quantifying heat requirements of different clones and how they can provide adaptation solutions for winegrowers to sustain local wine identity in a global changing climate. As genetic diversity is an ongoing process through point mutations and epigenetic adaptations, perspective work is to explore clonal data from a wide variety of geographic locations.

Evolution of the amino acids content through grape ripening: Effect of foliar application of methyl jasmonate with or without urea

The parameters that determine the grape quality, and therefore the optimal harvest time, suffer variations during berry ripening, related to climate change, with the widely known problem of the gap between technological and phenolic maturities. However, there are few studies about its incidence on grape nitrogen composition. For this reason, the use of an elicitor, methyl jasmonate (MeJ), alone or with urea, is proposed as a tool to reduce climatic decoupling, allowing to establish the harvest time in order to achieve the optimum grape quality. The aim was to study the effect of MeJ and MeJ+Urea foliar applications on the evolution of Tempranillo amino acids content throughout the grape maturation. Three treatments were foliarly applied, at veraison and 7 days later: control (water), MeJ (10 mM) and MeJ+Urea (10 mM+6 kg N/ha). Grape samples were taken at five stages of maturation: day before the first and second applications, 15 days after the second application (pre-harvest), harvest day, and 15 days after harvest (post-harvest). The amino acids analysis of the samples was carried out by HPLC. Results showed that the evolution of amino acids was similar regardless of the treatment; however, foliar applications influenced the nitrogen compounds content, i.e., there was no qualitative effect but quantitative one. Most of the amino acids reached their maximum concentration in pre-harvest, being higher in grapes from the treatments than in the control. In general, no differences in grape amino acids content were observed between MeJ and MeJ+Urea treatments. Foliar applications with MeJ and MeJ+Urea enhanced the grape amino acids content, without affecting their profile, helping to optimize their quality and allowing to establish a more complete grape ripening standard. Therefore, MeJ and MeJ+Urea foliar applications can be a simple agronomic practice, which has shown promising results in order to enhance the grape quality.

Copper contamination in vineyard soils of Bordeaux: spatial risk assessment for the replanting of vines and crops

Copper (Cu) is widely and historically used in viticulture as a fungicide against mildew. Cu has a strong affinity for soil organic matter and accumulates in topsoil horizons. Thus, Cu may negatively affect soil organisms and plants, consequently reducing soil fertility and productivity. The Bordeaux vineyards have the largest vineyard surfaces (26%) within French controlled appellation and a great proportion of French wine production (around 5 million hl per year). Considering the local context of vineyard surfaces decreasing (vine uprooting) and possible new crop plantation, the issue of Cu potential toxicity rises. Therefore, the aims of this work are firstly to evaluate the Cu contamination in vineyard soils of Bordeaux, secondly to produce a risk assessment map for new vine or crop plantation. We used soil analyses from several local studies to build a database with 4496 soil horizon samples. The database was enhanced by means of pedotransfer functions in order to estimate the bioaccessible (EDTA-extractable) Cu in soils of samples without measurements. From this database, 1797 georeferenced samples with CuEDTA concentrations in the topsoil (0-50 cm depth) were used for kriging interpolation in order to produce the spatial distribution map of CuEDTA in vineyard soils. Then, the spatial distribution of Cu was crossed with vine uprooting surfaces and municipality boundaries. CuEDTAconcentrations ranged from 0.52 to 459 mg/kg and showed clear anomalies. Our results from spatial analysis showed that almost 50% of vineyard soil surfaces have CuEDTA concentrations higher than 30 mg/kg (moderate risk for new plantation) and 20% with concentrations higher than 50 mg/kg (high risk for new plantation). A decision-support map based on municipalities was realised to provide a simple tool to stakeholders concerned by land use management.