Understanding and manipulating small signalling molecules to affect the yield/flavour (‘quality’) nexus FINAL REPORT to WINE AUSTRALIA Project Number: CSP 1401 Principal Investigator: Christopher Davies Research Organisation: CSIRO Agriculture & Food Date: December 2017 Project title: Understanding and manipulating small signalling molecules to affect the yield/flavour (‘quality’) nexus Author details: Dr Christopher Davies Team Leader CSIRO Agriculture & Food PO Box 350 Glen Osmond SA 5064 Australia Tel: (08) 8303 8628 Fax: (08) 8303 8601 Email: [email protected] Date report completed: 12/12/2017 Publisher: CSIRO Copyright: © 2017 CSIRO. To the extent permitted by law, all rights are reserved and no part of this publication covered by copyright may be reproduced or copied in any form or by any means without the written permission of CSIRO. Important Disclaimer: CSIRO advises that the information contained in this publication comprises general statements based on scientific research. The reader is advised and needs to be aware that such information may be incomplete or unable to be used in any specific situation. No reliance or actions must therefore be made on that information without seeking prior expert professional, scientific and technical advice. To the extent permitted by law, CSIRO (including its employees and consultants) exclude all liability for any consequences, including, but not limited to all losses, damages, costs, expenses and any other compensation, arising directly or indirectly from using this publication (in part or in whole) and any information or materials contained in it. Table of Contents 1. Abstract ................................................................................................................................. 4 2. Executive summary ............................................................................................................... 5 3. Background ........................................................................................................................... 7 4. Project Outputs and Activities .............................................................................................. 9 5. Method ................................................................................................................................ 14 6. Results ................................................................................................................................. 28 7. Outcomes and conclusions ................................................................................................ 103 8. Recommendations ............................................................................................................. 104 Appendix 1: Communication ................................................................................................. 105 Appendix 2: Intellectual Property .......................................................................................... 108 Appendix 3: References ......................................................................................................... 108 Appendix 4: Staff ................................................................................................................... 119 Appendix 5: Supplemental material ....................................................................................... 119 Appendix 6: Budget reconciliation ........................................................................................ 119 3 1. Abstract The objective was to identify small signaling molecules pivotal to berry development, to determine their mechanisms of action in controlling berry ripening. This knowledge was used to develop methods to mitigate the detrimental effects of climate change on berry composition and the timing of veraison and, consequently, harvest. Extensive vineyard and laboratory studies using techniques including sensory analysis and cutting edge analytics showed that it is possible to alter veraison and harvest timing through in-vineyard treatments without yield loss and with little, or no, impact on wine, the exception being increased pepper notes in some Shiraz plots. 4 2. Executive Summary Grape berry development, and in particular the ripening phase, is a complex process our understanding of which is increasing but there is still much to learn to help sustain grape and wine quality and value in a competitive and ever-changing world. Techniques to better manage current and future challenges need to be derived from an increased knowledge of berry growth and maturation. For example, the environmental conditions under which grapes are grown are continuing to change as a result of global climate change. Greenhouse gas emissions, particularly the increase in carbon dioxide levels, are driving climate change by trapping heat within the atmosphere. The resulting increased temperatures and changing weather patterns have profound effects on plant growth and therefore on viticulture and wine production. The dynamic nature of this problem and its importance make it crucial that we generate methods to mitigate its effects. Grapevines and grapes are very sensitive and responsive to temperature and the observed increase in temperatures is having a profound effect on the rate and timing of berry ripening and on the composition of the fruit used for winemaking, through changes in metabolism. The significant changes to berry ripening that are problematic for the grape and wine industry include: earlier onset of veraison and rapid ripening over a shorter period, rapid sugar accumulation ahead of flavour development, reduced colour/flavour development and compressed harvest seasons. We have targeted the small signaling molecules that control grape berry development as the keys to better understanding berry development (in particular berry ripening) and as tools to manage the ripening process to the benefit of industry. The aim is to better manage the ripening process to improve grape and wine quality and harvest timing without reducing yield. We have demonstrated previously that the small, plant growth regulator (PGR) molecule 1-naphthaleneacetic acid (NAA) has the ability to delay grape berry ripening when applied to pre-veraison berries under ‘cool’ climate conditions. This work was furthered in the current project where we showed that NAA is effective in delaying ripening and therefore harvest under warm climate conditions (McLaren Vale), making possible its use in controlling harvest timing under a broader range of conditions. To further test the potential of NAA under different conditions an experiment was conducted in the Eden Valley where veraison was significantly delayed. Post-veraison treatments with NAA were also trialed to determine whether NAA could delay ripening once it had commenced rather than delaying the onset with pre-veraison treatments. However, the progress of development, as measured by berry weight and Brix, was not altered in fruit treated post-veraison with NAA. NAA is used in other horticultural industries as it is an effective and safe plant growth regulator and is more stable than the auxin, indole-3-acetic acid (IAA), normally found in fruit. IAA is synthesised in grapevine from the amino acid tryptophan and a sustained synthesis of IAA might be initiated if tryptophan levels were increased. The application of tryptophan did not delay ripening and it was also confirmed that the application of IAA itself was ineffective. The lack of effectiveness of IAA is almost certainly due to its rapid metabolism by specialised enzymes within the berry. Field trials demonstrated that another auxin, 4-chloroindole acetic acid, normally found in seeds and for which some evidence suggested that it may be more resistant to rapid degradation and therefore more stable within the berry, was also ineffective in delaying ripening/harvest. NAA is therefore the PGR of choice to delay berry ripening/harvest. Control over the length of the delay in veraison and harvest is also a practical consideration. A comparison of 50 mg/L and 250 mg/L applications 5 showed that higher concentrations increased the delay in ripening and harvest of Shiraz fruit, demonstrating that the length of the delay could be manipulated through dosage levels. In general, the treatments with NAA that delayed ripening had little effect on both the wine volatile metabolite profiles and sensory properties. Longer delays of 2-3 weeks in ripening/harvest tended to increase the levels of some fruity esters with some minor differences in sensory properties and tasters showed no particular preference. In Shiraz, these longer delays increased the peppery character of wines with a coincident increase in the levels of rotundone, the metabolite responsible. The reasons for this specific increase are not yet known. Other PGRs are involved in berry development and might be targets for manipulating berry quality. Cytokinins, for example, have been suggested as potential delayers of ripening. We discovered that the cytokinin isopentenyl adenine (iP), increased from veraison to high levels at harvest, the final level varying considerably between wine grape cultivars. This indicates that iP might be involved in some aspect of berry ripening. Field experiments suggested that it is not involved in the timing of ripening as its application to the low iP cultivar Pinot Noir did not affect the timing or progression of berry development. Although iP probably has some ripening-related role, perhaps related to post-veraison