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 Use of antisense RNA technology to modulate gene expression in Œnococcus oeni

Use of antisense RNA technology to modulate gene expression in Œnococcus oeni

Abstract

Œnococcus oeni is a wine-associated lactic acid bacterium performs the malolactic fermentation, which improves the taste and aromatic complexity of many wine. Although, wine exhibits harsh and challenging conditions (low pH, low temperature, nutrient-poor and presence of ethanol), O. oeni possesses a remarkable adaptability to those physiochemical conditions.

Mechanisms for responding to environmental changes are universally present in living beings and are essential for coping with the stress and for adapting to the new conditions. O. oeni tolerance to low pH and ethanol make its an interesting bacteria model for investigating stress response mechanism in lactic acid bacteria. However, lack of appropriate techniques to manipulate O. oeni genome has long delay molecular study of this fastidious bacterium. To get around the lack of genetic tool for gene replacement, we focused our work on gene inactivation by using antisense RNA approach to modulate gene expression. With the goal to understanding the function of O. oeni hsp genes in vivo, we produce antisense RNA targeting genes encoding: a small Hsp (hsp18)1, the master regulator of stress response (ctsR)2 and two caseinolytic protease L members of the HSP100 chaperone family (clpL1, clpL2). Thereby, we highlighted that in vivo inhibition of the expression of some of these genes strongly affects the survival of O. oeni in stress conditions.

This study presents an elegant approach providing access to an in vivo study of gene function in O. oeni.

References:

1. Darsonval, M., Msadek, T., Alexandre, H. & Grandvalet, C. The Antisense RNA Approach: a New Application for In Vivo Investigation of the Stress Response of Oenococcus oeni, a Wine-Associated Lactic Acid Bacterium. Appl. Environ. Microbiol. 82, 18–26 (2016).

2. Darsonval, M., Julliat, F., Msadek, T., Alexandre, H. & Grandvalet, C. CtsR, the Master Regulator of Stress-Response in Oenococcus oeni, Is a Heat Sensor Interacting With ClpL1. Front Microbiol 9, (2018).

DOI:

Publication date: June 10, 2020

Issue: OENO IVAS 2019

Type: Article

Authors

Cosette Grandvalet, Frédérique Julliat, Maud Darsonval, Tarek Msadek, Hervé Alexandre

UMR A 02.102 Procédés Alimentaires et Microbiologiques, AgroSup Dijon – Université de Bourgogne Franche-Comté, Dijon, FRANCE.
Unité de Biologie des Bactéries Pathogènes àGram Positif, Institut Pasteur, Paris, FRANCE.
CNRS ERL 6002, Paris, FRANCE.

Contact the author

Keywords

Oenococcus oeni, lactic acid bacteria, antisense RNA , stress response 

Tags

IVES Conference Series | OENO IVAS 2019

Citation

Related articles…

The impact of leaf canopy management on eco-physiology, wood chemical properties and microbial communities in root, trunk and cordon of Riesling grapevines (Vitis vinifera L.)

In the last decades, climate change required already adaptation of vineyard management. Increase in temperature and unexpected weather events cause changes in all phenological stages requiring new management tools. For example, defoliation can be a useful tool to reduce the sugar content in the berries creating differences in the wine profiles. In a ten-year field experiment using Riesling (Vitis vinifera L, planted 1986, Geisenheim, Germany), various mechanical defoliation strategies and different intensities were trialed until 2016 before the vineyard was uprooted. Wood was sampled from the plant compartments root, trunk, cordon and shoot for analyses of physicochemical properties (e.g. lignin and element content, pH, diameter), nonstructural carbohydrates and the microbial communities. The aim of the study was to investigate the influence of reduced canopy leaf area on the sink-source allocation into different compartments and potential changes of the fungal and prokaryotic wood-inhabiting community using a metabarcoding approach. Severe summer pruning (SSP) of the canopy and mechanical defoliation (MDC) above the bunch zone decreased the leaf area by 50% compared to control (C). SSP reduced the photosynthetic capacity, which resulted in an altered source-sink allocation and carbohydrate storage. With lower leaf area, less carbohydrates are allocated. This for example resulted in a decreased trunk diameter. Further, it affected the composition of the grapevine wood microbiota. SSP and MDC management changed significantly the prokaryotic community composition in wood of the root samples, but had no effect in other compartments. In general, this study found strong compartment and less management effects of the microbial community composition and associated physicochemical properties. The highest microbial diversities were identified in the wood of the trunk, and several species were recorded the first time in grapevine.

Influence of agronomic practices in soil water content in mid-mountain vineyards

In the context of LIFE project MIDMACC (LIFE18 CCA/ES/001099), several pilots have been installed in vineyards in mid mountain areas of Catalonia (NE Spain) to test well stablished agronomic practices to increase the adaptation of Mediterranean mid mountain to climate change. Soil water content (SWC) at three different depths (15, 30 and 45cm) was measured in continuum from August 2020. One pilot (WC) included a well-established green cover (GC), a new GC (NC) and a conventional soil management (CM, tilling+herbicides). NC presented an intermediate state between WC and CM, responding similarly to CM in autumn but quickly reaching similar SWC to WC, then following the same evolution till next spring, with CM presenting lower values along autumn and winter. Then vegetation activation decreased SWC in all plots, (much slower in CM, lacking GC). Sensibility to spring rains is again intermediate for NC, which joins SWC evolution of CM by the end of spring till next autumn. It is expected that NC will resemble WC more and more as its GC develops. In the pilot combining vine training (VSP vs Gobelet) and hillside management (slope vs terrace), no clear pattern could be related with these conditions. However, both terraces seem to be more sensitive to spring rains. A third pilot included new vineyards (7 and 1 year old). In the new vineyard (N), higher canopy development, a spontaneous green cover and row straw resulted in a slower SWC dynamic, not so sensitive to rains but conserving more soil water in spring and most of summer, even with presumably a higher water extraction by vines. In the newest vineyard (VN) the deepest sensor is still sensitive to rain events all over the year and SWC is always highest at this depth, revealing small water capture by vines.

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:

Pruned vine biomass exclusion from a clay loam vineyard soil – examining the impact on physical/chemical properties

The wine industry worldwide faces increasing challenges to achieve sustainable levels of carbon emission mitigation. This project seeks to establish the feasibility of harvesting winter pruned vineyard biomass (PVB) for potential use in carbon footprint reduction, through its use as a renewable biofuel for energy production. In order to make this recommendation, technical issues such as the potential environmental impact, chemical composition and fuel suitability, and logistical challenges of harvesting biomass needs to be understood to compare with the results from similar studies. Of particular interest is the role PVB plays as a carbon source in vineyard soils and what effect annual removal might have on soil carbon sequestration. A preliminary trial was established in the Waite Campus vineyard (University of Adelaide) to test current management strategies. Vines are grown in a Eutrophic, Red Dermosol clay loam soil with well managed midrow swards. A comparison was undertaken of mid-row treatments in two 0.25 Ha blocks (Shiraz and Semillon), including annual cultivation for seed bed preparation, the deliberate exclusion of PVB (25 years) and incorporation of PVB (13 years) at an average of 3.4 and 5.5 Mg/Ha-1 for Shiraz and Semillon respectively. In both 0-10cm and 10-30cm soil core sample depths, combined soil carbon % measures in the desired range of 1.80 to 3.50, were not significantly different between treatments or cultivars and yielded an estimated 42 Mg/ha-1 of sequestered soil carbon. Other key physical and chemical measures were likewise not significantly different between treatments. Preliminary results suggest that in a temperate zone vineyard, managed such as the one used in this study, there is no long term negative impact on soil carbon sequestration through removing PVB. This implies that growers could confidently harvest PVB for use in several end fates including as a bio fuel.

Frost risk projections in a changing climate are highly sensitive in time and space to frost modelling approaches

Late spring frost is a major challenge for various winegrowing regions across the world, its occurrence often leading to important yield losses and/or plant failure. Despite a significant increase in minimum temperatures worldwide, the spatial and temporal evolution of spring frost risk under a warmer climate remains largely uncertain. Recent projections of spring frost risk for viticulture in Europe throughout the 21st century show that its evolution strongly depends on the model approach used to simulate budburst. Furthermore, the frost damage modelling methods used in these projections are usually not assessed through comparison to field observations and/or frost damage reports.
The present study aims at comparing frost risk projections simulated using six spring frost models based on two approaches: a) models considering a fixed damage threshold after the predicted budburst date (e.g BRIN, Smoothed-Utah, Growing Degree Days, Fenovitis) and b) models considering a dynamic frost sensitivity threshold based on the predicted grapevine winter/spring dehardening process (e.g. Ferguson model). The capability of each model to simulate an actual frost event for the Vitis vinifera cv. Chadonnay B was previously assessed by comparing simulated cold thermal stress to reports of events with frost damage in Chablis, the northernmost winegrowing region of Burgundy. Models exhibited scores of κ > 0.65 when reproducing the frost/non-frost damage years and an accuracy ranging from 0.82 to 0.90.
Spring frost risk projections throughout the 21st century were performed for all winegrowing subregions of Bourgogne-Franche-Comté under two CMIP5 concentration pathways (4.5 and 8.5) using statistically downscaled 8×8 km daily air temperature and humidity of 13 climate models. Contrasting results with region-specific spring frost risk trends were observed. Three out of five models show a decrease in the frequency of frost years across the whole study area while the other two show an increase that is more or less pronounced depending on winegrowing subregion. Our findings indicate that the lack of accuracy in grapevine budburst and dehardening models makes climate projections of spring frost risk highly uncertain for grapevine cultivation regions.