OWRI Updates

Dr. Laurent Deluc, Associate Professor, Dept. of Horticulture, OSU Oregon Wine Research Institute

As part of the research project studying the role of the regulatory protein Auxin-Response Factor 4, namely ARF4, on the ripening initiation of grape berries, our group has lately invested time and research efforts in promoting the microvine system at OSU, which was developed by our Australian partner Dr. Mark Thomas at the Commonwealth Scientific and Industrial Research Organization (CSIRO) (Chaïb et al. 2010). In early May, we received the first plant materials from Australia (see Figure 1 & 2), which includes in vitro material from white and red-grape varieties. One question that we are often asked about the microvine system is “what is the microvine system and why is it important to research?” If recent advances in sequencing technologies and genomic tools are very helpful to build new hypotheses on complex molecular processes, such hypotheses still need to be validated in planta in order to prove the concept. When one gene from genomic data has been identified as potential link with a trait of interest (fruit quality, disease resistance, stress tolerance), one way to prove the relationship between the genetic marker and the trait of interest is to perform “genetic engineering” or molecular breeding. To do so, this approach requires the use of a reliable model system that must combine several advantages that include small space requirements for growth, short generation time (constant flowering trait), tractable system for genetic engineering (genetic transformation), and small-size genome. The microvine system offers all of these advantages.

Dr. Satyanarayana Gouthu, Research Associate in the Deluc lab, is currently visiting Dr. Mark Thomas’s lab at the CSIRO in Australia to receive the necessary training for the different steps related to microvine propagation and genetic transformation. From my interaction with Dr. Gouthu, it is clear to me that he is learning a lot about microvine, which is essential for him to “master” when he eventually initiates the genetic engineering work at OSU. Meanwhile, another aspect of his research project is also in its final phase. By the end of the summer, we hope to identify a series of potential “interactors” with ARF4. This information is necessary to understand how a protein (ARF4) is regulated and what ARF4 interacts with during the process of berry ripening? We also made significant progress in terms of adapting a new method called Atmospheric Pressure Gas Chromatography Mass Spectrometry (APGC-MS) for metabolite identification in grape berries, in collaboration with the OSU Mass Spectrometry Center. Our colleague from the center has built a database containing around 75 individual analytes belonging to different classes of metabolites (organic acids, amino acids, sugar-related compounds, and pigment- related compounds). Our goal is not only to use this database for routine metabolite analyses in our lab, but also over time to improve the depth of the database by adding new metabolites. We are currently running samples from another experiment with promising results. We are very excited using this new analytical method for our current research project on ARF4 and future research projects as well.

By using the microvine, we expect to connect the function of proteins to important traits for grapevine production. Our goal is to specifically connect the timing of ripening initiation to the protein ARF4. This information could be useful by providing the industry with a potential genetic marker associated with véraison that could serve be used to validate new practices in the field, and to identify new or existing cultivars/clones for advanced or delayed ripening more amenable to local changes in the environment due to climate change. We hope to collaborate with OWRI, OSU and other external partners in pursuit of our research objectives.

Literature cited:

Chaïb J, Torregrosa L, Mackenzie D, Corena P, Bouquet A, Thomas MR. 2010. The grape microvine- a model system for rapid forward and reverse genetics of grapevines. Plant J. 62(6):1083-92. doi: 10.1111/j.1365-313X.2010.04219.x

From OSU to the Cellar Floor- Three OSU Grads Take Charge at Alexana Winery

It’s a sunny Thursday at Alexana winery in Newberg, Oregon. Bryan, Jamie, and Matt gather in the lab to explore new software designed to track blending, lab analysis and winemaking, allowing the team to spend less time on administration and more time in the vineyard and the winery. This is just one challenge this crew of OSU alumni share. They also share a passion for wine, which guided them through Oregon State University and eventually landed them at Alexana.

Job descriptions vary in any profession. In the wine industry, they serve as a loose guide rather than a hard and fast standard, making flexibility a necessary skill. This team of three navigates everything from tasting grapes with their vineyard manager to discussions with the consulting winemaker. So what is a “typical day” for the folks at Alexana Winery?
As the Cellar Master, Matt Stickle’s job follows the seasonality of production. He monitors temperatures and closures; starting and ending each day by checking tank lids, bungs, and temperatures of tanks and the barrel bays, while also cleaning and gathering samples for experimental wines and lab analysis. Matt utilizes  a vine-to-wine philosophy, similar to what researches follow at OSU. He participates in each step of the process; following the grapes from the vineyard to a finished wine being shipped off for storage gives him a sense of satisfaction. “Developing the wine from vine to bottle and sharing the finished product with our consumers gives me a sense of great accomplishment.”

Matt also enjoys the connections he has made with other industry members, and the brainstorming and ideas that are generated by working with them. “There are countless ways to make wine. Discussing different methods with other production people, you learn so much about what has worked, and what we wish had worked.”

As the enologist, Jamie Rauch monitors quality and follows the grapes from vine to finished wines. Her focus changes depending on the time of year and the stage of the wine’s evolution.  Jamie chose to develop expertise working in a lab because she states: “I love the precision, organization, and attention to detail necessary to run a successful lab, and the sensory side of monitoring wines is fascinating. Every decision we make boils down to using our sensory skills to make the final decision on what a wine needs.” Her days are never dull – the flexibility of her position gives her the opportunity to gain experience in every aspect of winemaking. She’s gone from driving a forklift to pouring wines for consumers in the tasting room. Jamie also appreciates having Bryan as a mentor. He facilitates a learning environment where all are gaining experience and trying to make the best possible wines every day.

As the winemaker, Bryan Weil’s day begins with blending, tasting, writing work orders, doing compliance/tracking, observing vineyards, doing lab work, travelling, conducting consumer tastings/wine events, and all of the other day-to-day winemaking jobs. His work also includes lots of cleaning, troubleshooting/fixing equipment, and physical labor. In the tasting room, Bryan enjoys interacting with customers and discussing the wine. He is immensely passionate about the rigors and dedication necessary to produce quality wine. He spends time in the vineyard and consults with the vineyard manager to ensure quality grapes for his wine, and that dedication transcends into the lab and the cellar.

The pressure and quest for perfection challenges Bryan. “We only have one chance to make these wines. We can’t remake the wines again if we make mistakes.  OSU provided me with scientific and practical knowledge to ensure I have the best chance possible to make a quality product year after year,” he says.

Another vine that weaves this team of three together is the connections they have to the Oregon and worldwide wine industry and their fellow OSU grads. Bryan states: “It is amazing to me how many OSU alumni are in the Oregon and Washington wine industries and all over the world. I am fortunate to have a group of OSU alumni that I keep in contact with and conduct tasting groups with. We discuss everything from vineyards, winemaking, and the business of making and selling wine. It’s great to get everyone together because of all of our different experiences we have had in the industry, but at the same time we all came from the same great university in the beginning of our wine careers.” This built-in network has been instrumental in the success of these former students.

Because, after all, networking is everything.

Dr. James Osborne, Enology Extension Specialist and Associate Professor

Harvest is here and in the winery there are many things to prepare for before the fruit starts arriving. One key area to prepare for is yeast and nutrient management. While yeast and nutrient management are always key factors in conducting successful fermentations, extra care is needed in years like this where grape composition may lend itself to more problematic fermentations. Because of the warm and dry growing season fruit may contain high °Brix and low nutrients. This fruit chemistry can cause problems with alcoholic fermentations as yeast need to metabolize a greater amount of sugar with a lower amount of nutrients in a high alcohol environment. The end result is often a very slow/sluggish fermentation or fermentations that do not complete fermentation but rather stall out with a few Brix still remaining. One key factor in preventing stuck/sluggish fermentations is ensuring there is sufficient yeast nutrients present during the fermentation. Yeast assimilable nitrogen (YAN) is one of these key nutrients and insufficient amounts can result in stuck fermentations as well as increased production of hydrogen sulfide.  YAN is composed of nitrogen from ammonia (inorganic nitrogen) and nitrogen from primary amino acids (organic nitrogen). Luckily, we have a number of tools at our disposal to supplement YAN but how and when to perform this supplementation is a little more complicate.

The first step is measuring how much YAN is present in the juice/must. While you want to add enough YAN for a complete and clean fermentation, you do not want to add excessive nutrients as this can also cause problems. Large additions of YAN early in the ferment may lead to over vigorous fermentations and alter the aroma compounds produced by the yeast. In addition, residual nutrients in the wine may contribute to microbial spoilage during aging. So how much YAN do you need? Well, it depends. The general recommendation is between 150-250 mg/L for a 21-23°Brix must. If you have a higher °Brix must or are using a high nutrient demand yeast strain then you may want to consider higher YAN levels. These are not hard and fast rules but are YAN levels that have been reported by researchers and yeast manufacturers to result in fermentations with good kinetics. Aside from nitrogen, the other nutrients that are essential factors for yeast growth are the micronutrients such as the vitamins biotin, pantothenic acid, and thiamin. If you just want to increase YAN then DAP is an efficient way to do this. However, DAP does not contain any micronutrients so in addition to DAP you also should use a complex yeast nutrient that contains a blend of organic nitrogen (amino acids, peptides) and micronutrients. A balanced approach of both DAP and complex nutrients works best if you need to significantly increase your YAN levels. Nutrient additions should be carefully monitored and recorded as there are legal limits to the concentrations that can be added. For example there are limits to the amount of DAP (0.96 g/L), thiamin (0.60 mg/L), and pantothenic acid (0.048 mg/L) that can be added. For complex yeast nutrients carefully read the manufacturer’s instructions carefully to determine the max concentration of the product that can be added.

The timing of nutrient additions is important for successful fermentations. Yeast preferentially up-take ammonia (DAP) before amino acids. Therefore, one large addition of DAP at the beginning of fermentation may delay/inhibit uptake of amino acids and cause problems later in fermentation. It is therefore recommended to perform multiple additions of nutrients during the early to mid-fermentation stage. For example, add half the nutrients 12-24 hours after inoculation followed by the remainder of the nutrients around 1/3 sugar depletion. Adding nutrient supplements all at once can lead to a fast fermentation rate, and an imbalance in uptake and usage of nitrogen compounds. Alternatively, supplements added too late in the fermentation (after 2/3 fermentation) may not be utilized by the yeasts. This is because as the fermentation proceeds ethanol concentrations reach a point it impacts the yeast membrane and reduces the ability of the yeast to uptake nutrients.

Aside from YAN, the other yeast nutrient that can play a critical role in conducting successful alcoholic fermentations is oxygen. During the early stages of alcoholic fermentation Saccharomyces can use oxygen for the production of sterols. These sterols are a key component of the yeast cell membrane and will help the yeast resist osmotic stress at the beginning of fermentation and ethanol toxicity near the end of fermentation. There are two times during the fermentation where oxygen addition has been shown to be beneficial. First, once the fermentation has become active and a 1-3 ⁰Brix drop has occurred. The second time is at about 1/3 ⁰Brix drop. Addition of oxygen to the ferment after this time is not recommended. Pumping over or racking and returning can supply some oxygen to the ferment but using a macro-oxygenator or micro-oxygenator at a high rate is a more reliable way to provide air to the ferment.

Aside from adjusting your nutrient regime, you should also consider the yeast strains that you use for your fermentations. In warmer years with high °Brix grapes the choice of yeast strain can make a difference in the prevention of stuck fermentations. Saccharomyces cerevisiae strains vary in their ethanol tolerance as well as their preference for consuming glucose or fructose. In a typical grape juice glucose and fructose is present in an equivalent amount. However, most Saccharomyces cerevisiae yeast strains used in winemaking preferentially consume glucose so that near the end of a fermentation the majority of the remaining sugar will be fructose. This is why it is recommended that a fructophilic (fructose loving) yeast should be used to restart stuck fermentations. These yeast strains can also be used as the primary yeast for fermentation and may be a valuable tool when fermenting lots that have historically been problematic or that are high °Brix ferments. It is recommended that you talk with your yeast supplier about fructophilic yeast strains and their use.

Warmer years also typically present us with fruit containing lower acid. If acid additions are to be performed it is important to measure a few different components of acidity. The typical measurements of acidity are pH and titratable acidity (TA). The pH and TA of your juice/grapes will be impacted primarily by the concentrations of tartaric and malic acid. These acids have different strengths and so a different ratio of these acids will impact both pH and TA independently. For example, you can have two juices with very similar TAs but quite different pH values if their tartaric and malic acid concentrations differ. pH is also impacted by the buffering capacity of the juice/must. The major component of grapes that impacts buffering capacity is potassium. Grapes with high potassium concentrations can be resistant to pH change from acid additions because of this buffering capacity. For example, you may make an acid addition to a set TA level but not see the expected decrease in pH if the juice/must contains high amounts of potassium. Bench-top trials for acid adjustments are recommended so that you can an accurately determine how much acid will be needed to achieve a certain pH and what the resulting TA will be.

One additional note when it comes to calculating pre-fermentation adjustments (YAN and acid). When taking grape and juice samples for analysis, the more closely these samples represent the grape/juice in the tank the better. When assessing red grapes I would recommend taking the grape samples and crushing them by hand in a zip lock bag and letting the juice soak on the skins for a few hours (in the fridge). This will give you a more accurate pH value because the grape skins contain a significant amount of potassium that will soak out during this time. If you analyze the juice immediately after crushing the grapes then you will not account for this potassium. Grape skins also contain some amino acids and so soaking the grapes will give a better estimation of the YAN content of the grapes.

If you have additional questions please contact me at 541-737-6494 or email james.osborne@oregonstate.edu

Dr. Patty Skinkis, Viticulture Extension Specialist & Associate Professor

Excessive plant water stress can cause damage to grapevines, particularly young vines. The recent hot, dry seasons of 2014 and 2015 and the west coast drought drew awareness to water stress and its potential detrimental impacts. However, water stress can occur in any season, and an understanding of how to monitor and manage it is important to the production of healthy vineyards in any region.

This article briefly describes ways to determine whether vines are under water stress. This is the first step used in determining when to begin irrigation in response to stress. For sites that are not irrigated, the methods may be used to determine whether management tactics should be employed to conserve soil moisture or reduce soil water competition. The three most common methods to determine water stress include 1) visual symptoms, 2) leaf or stem water potential and 3) stomatal conductance.

Visual symptoms may be used to determine whether a plant is under stress. It requires training and understanding of the vine’s lifecycle, as symptoms may also be caused by other factors. When visually assessing canopies, it is important to look at shoot tips, tendrils, leaves, clusters, and overall growth. When vines are under water stress, tendrils become limp, shoot tips begin to flop downward, leaves begin to bend (petiole juncture at leaf blade begins to form a shepherds hook) and berries may begin to shrivel. Under prolonged water stress shoot tips stop growing and abscise, tendrils dry up and fall off, and basal leaves may turn yellow and abscise. It is important to consider that tendrils and shoot tips may dry up and fall off as a result of the natural cessation of growth late season, in preparation for dormancy and may not indicate water stress. Visual symptoms are important to document and are strengthened by quantitative measures. Keep in mind that water stress can lead to nutrient deficiencies  which may lead to other visual symptoms.

Leaf water potential is the most common measure conducted by commercial vineyards to determine whether vines are under water stress. A pressure chamber is used to measure the amount of pressure required to push water out of the cut end of the petiole and reflects the amount of water potential (or tension) of the water column in the leaf. It is a measure of negative pressure (- bars), although the gauge on most pressure chambers does not indicate a negative number. Typically, irrigation is initiated when leaf water potential readings reach -12 bars, which is considered moderate stress. Stem water potential, also uses a leaf to measure plant water stress, but it requires additional steps in the process, including covering a leaf with a special reflective bag for at least one hour prior to measurement. Covering the leaf limits transpiration, equalizing the water potential in the leaf close to what is experienced in the shoot (or stem). Stem water potential is usually 1-2 bars less negative as shoots are under less tension than leaves. For example, if leaf water potential readings are at -12 bars, the stem water potential on the same plant may be at -10 bars, and the leaf reading would suggest that you need to start irrigating while the stem reading does not. More information about monitoring plant water stress can be found here.

Above: The pressure chamber. The leaf is placed inside a sealed chamber and pressurized gas is added to the chamber slowly. As the pressure increases water will be forced out of the xylem and will be visible at the cut end of the stem. Note the magnifying glass for easier viewing.

Above: Stem water potential, also uses a leaf to measure plant water stress, requires additional steps in the process, including covering a leaf with a special reflective bag for at least one hour prior to measurement. Covering the leaf limits transpiration, equalizing the water potential in the leaf close to what is experienced in the shoot (or stem).

Above: When measuring water or leaf potential, make sure to cut end of the petiole for an accurate measurement.

Above: A leaf attached to a petiole is placed inside a sealed chamber and pressurised gas is added to the chamber slowly.

Some find the pressure chamber to be cumbersome due to its bulky size and have been interested in using a portable leaf porometer. A leaf porometer measures the rate of water that moves out of the stomata, known as stomatal conductance. When a vine is under water stress, stomata close and stomatal conductance of water is reduced. A vine that is not under any water stress can have stomatal conductance of >250 mmol H₂0/m²/s while a vine under moderate stress has <150 mmol H₂0/m²/s. There is a porometer on the market that is affordable for commercial vineyard use, and the cost is comparable to a pressure chamber. It may be small, but it requires training, attention to detail in collecting the data, and requires a calibration step before using the meter. This calibration needs to be done each day, before sampling begins, and again hen environmental conditions change (approximately every hour).

Both leaf water potential and stomatal conductance measures require the right equipment and training. Both tools should be used on clear, cloudless days within 1 hour before and after solar noon and not under extreme heat events, to give the most accurate readings. We have tested both pressure chambers and porometers in various trials under western Oregon conditions to monitor plant water stress (Skinkis and Schreiner Labs), and both can be good tools to determine when vines are under stress. If you are interested in those findings, contact Patty Skinkis or Paul Schreiner for more information. Whenever using quantitative measure of plant water stress, it is important to make note of visual symptoms, as they may help interpret conditions of the vines.

NOTE:  The reference to porometers and pressure chambers are for local companies that have developed these devices. There may be other companies that provide similar products, and the mention of these two companies does not imply recommendation or endorsement of those products over any other similar product.

Additional Reading

The Pressure Chamber (The Bomb) – UC Davis Fruit & Nut Research Information

Measuring Water Status Using a Pressure Chamber – eXtension.org

Grapevine Management under Drought Conditions

Irrigation Basics for Eastern Washington Vineyards

Dr. Patty Skinkis, Viticulture Extension Specialist & Associate Professor