Use of directed evolution to generate multiple-stress tolerant Oenococcus oeni for enhanced malolactic fermentation
type: CreativeWork
description: This study aimed to optimise Oenococcus oeni for more efficient malolactic fermentation in wine with multiple stressors. First, a previously evolved ethanol tolerant strain, A90, was characterised for resistance to combined pH and ethanol stress in both MRSAJ and Red Fermented Chemically Defined Grape Juice Medium (RFCDGJM). A90 showed a similar viability in RFCDGJM compared to its parent, SB3, indicating the need for further improvement. With the success of the previous proof-of-concept directed evolution (DE) in O. oeni, a new DE was carried out to determine 1) if DE can be applied to further improve A90 in a wine-like environment using combinations of stressors to generate more superior strains with better general stress resistance; 2) how much further can A90 be developed, and how stable the new phenotype would be; 3) possible new patterns of stress response through study of the genetic basis for the superior phenotype. A continuous culture of A90 was established in a bioreactor and grown in a wine-like environment for approximately 350 generations with increasing ethanol and sulfur dioxide (SO₂), and decreasing pH over time. Samples of the population in the bioreactor were collected at three significant times during the DE to screen for improved isolates based on L-malic acid consumption and growth. Three strains, namely 1-161, 2-49 and 3-83, outperformed from a total of 378 isolates. With a view to applying these strains to the industry, in-depth physiological characterisations were undertaken. Aspects examined included tolerance to various oenologically related stressors such as ethanol, pH, SO₂ and medium chain fatty acids, as well as phenotype stability and fermentation ability under more realistic winemaking conditions, i.e. un-filtered wine and winery scale fermentation. Overall, 2-49 and 3-83 constantly displayed better growth and malolactic activity than the parent strain A90 in either lab-scale or winery-scale trials. Finally, whole genome sequencing o…
abstract: This study aimed to optimise Oenococcus oeni for more efficient malolactic fermentation in wine with multiple stressors. First, a previously evolved ethanol tolerant strain, A90, was characterised for resistance to combined pH and ethanol stress in both MRSAJ and Red Fermented Chemically Defined Grape Juice Medium (RFCDGJM). A90 showed a similar viability in RFCDGJM compared to its parent, SB3, indicating the need for further improvement. With the success of the previous proof-of-concept directed evolution (DE) in O. oeni, a new DE was carried out to determine 1) if DE can be applied to further improve A90 in a wine-like environment using combinations of stressors to generate more superior strains with better general stress resistance; 2) how much further can A90 be developed, and how stable the new phenotype would be; 3) possible new patterns of stress response through study of the genetic basis for the superior phenotype. A continuous culture of A90 was established in a bioreactor and grown in a wine-like environment for approximately 350 generations with increasing ethanol and sulfur dioxide (SO₂), and decreasing pH over time. Samples of the population in the bioreactor were collected at three significant times during the DE to screen for improved isolates based on L-malic acid consumption and growth. Three strains, namely 1-161, 2-49 and 3-83, outperformed from a total of 378 isolates. With a view to applying these strains to the industry, in-depth physiological characterisations were undertaken. Aspects examined included tolerance to various oenologically related stressors such as ethanol, pH, SO₂ and medium chain fatty acids, as well as phenotype stability and fermentation ability under more realistic winemaking conditions, i.e. un-filtered wine and winery scale fermentation. Overall, 2-49 and 3-83 constantly displayed better growth and malolactic activity than the parent strain A90 in either lab-scale or winery-scale trials. Finally, whole genome sequencing o…
author: Jiang, Jiao
keywords: directed evolution, Oenococcus oeni, multiple-stress, malolactic fermentation, whole genome sequencing, Research by Publication
date published: 2017
date modified: 2026-07-24T00:50:53Z
provider: University of Adelaide
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: University of Adelaide

PropertyValue
property ID: Handle
value: http://hdl.handle.net/2440/114505

PropertyValue
property ID: DOI
value: 10.25909/5ba1ef1c145f6