On March 5, 2019, Penn State researchers and Extension personnel presented research findings and provided five-minute overviews of upcoming studies at the 2019 Wine Marketing & Research Board Symposium, held in conjunction with the Pennsylvania Winery Association Annual Conference.
In this post, we have included short summaries of what each presenter discussed during their session along with a PDF/access to their presentation.
Under-vine cover crops: Can they mitigate vine vigor and control weeds while maintaining vine productivity?
Presented by Michela Centinari, Assistant Professor of Viticulture, Suzanne Fleishman, Ph.D. Candidate, and Kathy Kelley, Professor of Horticultural Marketing and Business Management
Michela, Suzanne, and Kathy discussed research conducted at Penn State related to the use of under-vine cover crops as a management practice alternative to herbicide or soil cultivation. Michela reviewed potential benefits of under-vine cover crops, such as reduction of excessive vegetative growth, weed suppression, and reduced soil erosion. She showed how the selection of cover crop species depends on the production goals of a vineyard, climate, vine age, and rootstock. Suzanne presented results from her research project. She is investigating above- and belowground effects of competition between a red fescue cover crop and Noiret grapevines, comparing responses between vines grafted to 101-14 Mgt vs Riparia rootstocks. Surveys will be administered to Pennsylvania grape growers and wine consumers in the Mid-Atlantic region. Growers will be asked to respond to questions about interest in using cover crops and benefits that could encourage their use. The consumer survey will focus on learning whether cover crops use would impact their purchasing decision and if they would be willing to pay a price premium for a bottle of wine to offset additional production costs.
Impact of two frost avoidance strategies that delay budburst on grape productivity, chemical and sensory wine quality.
Presented by Michela Centinari, Assistant professor of Viticulture
Crop losses and delays in fruit ripening caused by spring freeze damage represent an enormous challenge for wine grape producers around the world. This multi-year study aims to compare the effectiveness of two frost avoidance strategy (application of a food grade vegetable oil-based adjuvant and delayed winter pruning) on delaying the onset of budburst, thus reducing the risk of spring freeze damage. Our objectives are to: i) evaluate if the delay in budburst impacts grape production and fruit maturity at harvest, as well as chemical and sensory wine properties; ii) elucidate the mechanism of action of the vegetable oil-based adjuvant through an examination of bud respiration and potential phytotoxic effects; and iii) assess the impact of the two frost avoidance strategies on carbohydrate reserve storage and bud freeze tolerance during the dormant season.
Toward the development of a varietal plan for Pennsylvania wine grape growers.
Presented by Claudia Schmidt, Assistant Professor of Agricultural Economics, and Michela Centinari, Assistant Professor of Viticulture
Claudia Schmidt is a new Assistant Professor of Agricultural Economics with an extension appointment at Penn State. Claudia used the opportunity of the symposium to introduce herself to the industry. In her presentation, she first gave an overview on what and where Pennsylvanians buy their wines and spirits. She then talked about the research needed to develop a varietal plan for the Pennsylvania grape and wine industry to match existing and future grape production and variety suitability with anticipated consumer demand. The immediate next steps on her research agenda are to develop a baseline survey of grape production in Pennsylvania and, in collaboration with Michela Centinari, region specific cost of production of grapes.
Survey for grapevine leafroll viruses in Pennsylvania: How common is it, and how is it effecting production and quality?
Presented by Bryan Hed, Research Technologist
This is a continuing project funded by the PA Wine Marketing and Research Board, that has focused on the determination of the incidence of grapevine leafroll associated virus 1 and 3 (the two most economically important and widely distributed of the leafroll viruses) in commercial vineyard blocks of Cabernet franc, Pinot noir, Chardonnay, Riesling, and Chambourcin, across the Commonwealth. Over two years, the survey has shown that grapevine leafroll associated viruses 1 and/or 3, were present in about a third of the vineyard blocks examined. Infection of grapevines by grapevine leafroll-associated viruses can have serious consequences on yield, vigor, cold hardiness, and most notably fruit/wine quality. Bryan also discussed a second phase of the project, anticipated to continue for at least another two years within 6 vineyard blocks of Cabernet franc, identified in the survey. In these vineyards, we plan to plot the spread of these viruses, examine and report their effects on grapevine vegetative growth, yield, and fruit chemistry, and characterize the influence of inter- and intra-seasonal weather conditions on virus-infected grapevine performance.
Integrating the new pest, spotted lanternfly, to your grape pest management program.
Presented by Heather Leach, Extension Associate
Spotted lanternfly (SLF) is a new invasive planthopper in the Northeast U.S. that threatens grape production. Heather covered the basic biology, identification, and current distribution of SLF. She also presented on the economic impact of SLF in the grape industry and ways to manage SLF in your vineyard. SLF can feed heavily on vines causing sap depletion in the fall which has resulted in death of vines, or failure of vines to set fruit in the following year. While biological controls such as pathogens and natural enemies along with trapping and behaviorally based methods are being researched, our current management strategy relies on using insecticides sprayed in the vineyard. Heather showed results from the 2018 insecticide trials conducted against SLF, with efficacy from several products including bifenthrin, dinotefuran, thiamethoxam, carbaryl, and zeta-cypermethrin. You can read more about the results from this trial here: https://extension.psu.edu/updated-insecticide-recommendations-for-spotted-lanternfly-on-grape
Five-minute research project overviews
Impact of spotted lanternfly on Pennsylvania wine quality.
Presented by Molly Kelly, Extension Enologist
The Spotted Lanternfly (SLF) presents a severe problem both due to direct damage to grapevines as well as their potential to impact wine quality. Insects are known to produce or sequester toxic alkaloid compounds. The objectives of this study include characterizing the chemical compounds in SLF and production of wines with varying degrees of SLF infestation. We can then provide winegrowers with recommendations for production of wine from infested fruit. Toxicity studies will be conducted to determine the levels of toxic compounds in finished wine, if any, using a mouse bioassay.
Exploring the microbial populations and wild yeast diversity in a Chambourcin wine model system.
Presented by Chun Tang Feng, M.S. Candidate, and Josephine Wee, Assistant Professor of Food Science
In Dr. Josephine Wee’s lab, we are interested in the microbial population and diversity associated with winemaking. When it comes to wine fermentation, not only are commercial yeasts involved in this process, but also many indigenous yeasts. Our research goal is to isolate the wild yeasts and assess their feasibility of wine fermentation. We are expecting to explore the unique yeast strains from local PA which are able to make a positive impact on wine flavor.
Rotundone as a potential impact compound for Pennsylvania wines
Presented by Jessica Gaby, Post-Doctoral Scholar and John Hayes, Associate Professor of Food Science
This study will examine Pennsylvania consumers’ perceptions of rotundone with the goal of determining whether a rotundone-heavy wine would do well on the local market. This will be examined from several different perspectives, including sensory testing of rotundone olfactory thresholds, liking and rejection thresholds for rotundone in red wine, and PA consumer focus groups. The ultimate aim of the study is to determine the ideal concentration of rotundone in a locally-produced wine that would appeal to PA consumers.
Defining regional typicity of Grüner Veltliner wines
Presented by Stephanie Keller, M.S. Candidate, Michela Centinari, Assistant Professor of Viticulture, and Kathy Kelley,
Grüner Veltliner(GV) is a relatively new grape variety to Pennsylvania, and while climatic conditions are favorable to its growth, the Pennsylvania wine industry is still becoming familiar with the varietal characteristics of GV grown and produced throughout the state. This study focuses on defining typicity of Pennsylvania-grown GV wines. Typicity is described as the perceived representativeness of a wine produced from a designated area, and defining typicity can improve wine marketing strategies. This study uses multiple experimental sites across the state to create wines from a standardized vinification method. The wines will be analyzed using both instrumental and human sensory methods.Surveys will be administered to Pennsylvania grape growers and white wine consumers in the Mid-Atlantic region. Growers will be asked their interest in growing GV and what perceived and real barriers may impact their decision to grow the variety. The consumer survey will focus on understating how to introduce them to a wine varietal they may be less aware of and what promotional methods may encourage them to purchase the wine.
Boosting polyfunctional thiols and other aroma compounds in white hybrid wines through foliar nitrogen and sulfur application?
Presented by Ryan Elias, Associate Professor of Food Science, Helene Hopfer, Assistant Professor of Food Science, Molly Kelly, Extension Enologist, and Michela Centinari, Assistant Professor of Viticulture
The quality of aromatic white wines is heavily influenced by the presence of low molecular weight, volatile compounds that often have exceedingly low aroma threshold values. Polyfunctional varietal thiols are an important category of these compounds. This project aims to provide research-based viticultural practices that could lead to increases in beneficial varietal thiols in white hybrid grapes. The expected increase in overall wine quality will be validated both by measuring the concentrations of these desirable compounds (i.e., thiols) in finished wines using instrumental analysis and by human sensory evaluation, thus providing a link between the viticultural practice of foliar spraying and the improvement of overall wine quality.
By: Conor McCaney, Graduate Assistant, Department of Food Science & Technology
The winemaking process is a dynamic one: from crush, to fermentation, on to post fermentation cellar procedures, aging, and bottling. Each step along the way allows for the potential ingress of oxygen, whether wanted or not. While oxygen is considered by many to be the enemy of wine, this is not always the case. In fact proper use of enological oxygen at crucial steps in the winemaking process is paramount to wine development. That said, many winemakers dutifully aim to eliminate it from the process altogether particularly in partial tank headspace. Proper gassing regimens and selection of the correct gas for a particular application is something that many do not do well and fail to fully understand the principals at play. Managing proper inert gas procedures is tricky. Most protocols are generally arbitrary ones copied from bad information and the proliferation of poor techniques passed on anecdotally from winemaker to winemaker. In general it is a procedure that is often over looked and never given much thought. This usually means the use of a high pressure cylinder (most often nitrogen), and a ¼” or ½” hose that is allowed to run for an arbitrary amount of time, generally 15 to 20 minutes. The results are the improper use of inert gases from the failure to measure gas volumes delivered (using a flowmeter), monitoring results with the use of a dissolved oxygen meter, using an under or oversized delivery system and unsubstantiated cost analysis pertaining to gas type and volume needed.
Typical gas choices are: carbon dioxide, nitrogen, and argon. Most wineries choose to use carbon dioxide and nitrogen because they believe it provides the best cost-benefit in terms of oxygen displacement per unit cost. This is not the case. To understand this, we must first delve into some fundamental principles of gases. In the wine industry, we typically use gas by volume, either in standard cubic feet or molar volume delivered from a standard steel pressurized cylinder in which the gas is compressed. These gas volumes are usually measured at 25°C and 1 atm. If you happen to purchase gas by the pound it is necessary to divide the gas by its molecular weight before you can compare gases to one another. The approximate molecular weights are: 40 g/mole for argon (Ar), 44 g/mole for carbon dioxide (CO2), 28 g/mole for nitrogen (N2), and 29 g/mole for air. One mole of any of these gases measured at standard pressure (1atm) and temperature (25°C) occupies one molar volume, roughly equivalent to 22.4 liters, 5.92 gallons, or 0.8 standard cubic feet. Using the ideal gas law PV = nRT the behavior of gases can be described in which pressure and volume is a fixed proportion in relation to the number of moles of gas at absolute temperature. This indicates that gas molecules take up the same amount of space regardless of their mass when they are at the same temperature and pressure (Avogadro’s Law). Thus one mole of any gas contains the same number of molecules (i.e., 6.02 x 1023). This also indicates that the head space in a tank, barrel, or other container will fluctuate regularly throughout the day in response to temperature and pressure changes. Tanks that are kept outside experience greater temperature changes throughout the day compared to a tank kept inside at a constant temperature. Changes in barometric pressure and temperature can cause the headspace in a tank to pump 3% to 7% of its volume in and out daily. This ultimately means that the headspace in a tank is not a static system and could be constantly changing.
Air is roughly composed of 78% nitrogen, 21% oxygen, and 1% argon, so in essence nitrogen is air without the oxygen. In any gassing procedure it is ideal to reduce the percentage of oxygen in the headspace to below 1% or even below 0.5% to inhibit the growth of aerobic microbes and prevent wine oxidation. The most commonly used gas in winemaking is nitrogen (N2) with a molecular weight (MW) of 28 g/mole making it moderately lighter (less dense) than air at 29 g/mole MW. Graham’s law of diffusion (also known as Graham’s law of effusion) states that the rate of effusion of a gas is inversely proportional to the square root of its molar mass at constant temperature and pressure. This principle is often used to compare the diffusion rates of two gasses such as nitrogen and air. The diffusion rates of nitrogen and air are almost identical meaning that nitrogen does not provide adequate layering, but rather readily mixes with air and does not remain in contact with the wine surface for an extended period of time. This also means that in order to reduce the O2level from 21% to less than 1%, the headspace needs to be flushed with a volume of nitrogen that is five times the volume of the headspace. So if the tank has a 100 gallons of head space it would take 500 gallons of nitrogen to reduce the O2level from 21% to below 1%. The cost of nitrogen is approximately $0.05 per cubic foot (Praxair, Inc). However, because nitrogen requires five times the volume equivalents to reduce the O2percentage from 21% to less than 1%, the cost to gas a barrel (60 gallons) is $2.00, 100 gallons of headspace is $3.34 and 1,000 gallons of headspace is $33.42. This is significantly higher than the cost of using argon for the same O2reduction in the equivalent headspace volumes. This is why headspace gassing with nitrogen requires a substantial effort and time commitment on the part of the winemaking team to be effective. It takes substantially more nitrogen and a greater application time compared to argon to achieve the same reduction in oxygen percentage with a shorter effective shelf life.
In contrast to nitrogen is carbon dioxide (CO2), which is significantly heavier than air at 44 g/mole compared to 29 g/mole and by Graham’s law has a much slower rate of diffusion compared to air. This allows for a more significant displacement of air compared to nitrogen. However, when CO2is delivered from a compressed tank, it is difficult to achieve the desired laminar flow necessary for successful layering. This results in substantial mixing of CO2and air. A more effective alternative for CO2delivery is dry ice (solid CO2) which leads to more efficient layering of CO2and subsequent displacement of air but does not form a permanent layer. However, it should be noted that CO2cannot be considered inert in the same way as nitrogen and argon. Because of Henry’s Law, which states that the solubility of a gas is directly proportional to the partial pressure of the gas above the solution, CO2readily dissolves into wine under standard conditions and its solubility can be increased or decreased with changes in pressure. This dissolution of CO2into the wine causes the pressure in the tank to fluctuate and results in the intake of air from the outside environment through an airlock to replace the lost volume of gaseous CO2. If there is no vacuum release valve on the tank, this could cause the tank to implode. Carbon dioxide dissolved in the wine will also alter the acid, flavor, and textural profile of the final wine. Carbon dioxide is much more effective when deployed early in the winemaking process at juice stage or when the wine is young as there will be substantial time to allow excess dissolved CO2to come out of solution. The use of dry ice to protect grape must is an effective way to protect wine must from excess oxygen exposure, deter fruit flies, and subsequently cool the must.
This leaves argon with a molecular weight of 40 g/mole, making it substantially heavier than air (29 g/mole) and similar in weight to CO2but more inert. A major opposition to the use of argon regularly in wine production is because it is significantly more expensive compared to the other two gases. It is true that when purchasing gas by volume argon is roughly three times as expensive as nitrogen or carbon dioxide. However it is much more effective at displacing air and creating a more permanent blanket that remains in contact with the wine surface longer while also remaining inert compared to CO2. Less volume is also needed to achieve the same desired results. At approximately $0.11 per cubic foot (Praxair, Inc) not including daily tank rental fee, a barrel (60 gallons) can be completely gassed with argon for $0.88, 100 gallons of head space for $1.47, and 1,000 gallons of headspace for $14.71. This cost is relatively insignificant to a winery’s bottom line in terms of the degree of quality preservation that argon can provide.
When using any of the gases discussed previously, it is important to select the proper pressure gauge, hose diameter, hose length, flowrate, and the use of a t-valve in order to deliver the gas under laminar conditions. The use of a lower velocity, will encourage laminar flow delivery and reduce any chance of turbulence and subsequent mixing with air, thus creating a more layered effect.
It is ideal to keep the flow velocity to 1 meter per sec or less. To determine the velocity divide the volumetric flow rate in cubic meters per second by the cross sectional area in meters of the hose being used. If using cubic feet instead of cubic meters, perform the same calculation but convert the units from cubic meters to cubic feet and meters to feet. Table 1 shows that it is best to use a 1.5” or 2” diameter line with a t-valve to deliver an adequate amount of gas in a reasonable amount of time. This will require the use of an oversized regulator compared to the typical 0.25” regulator used on most compressed gas cylinders.
In essence it is best practice to recommend the use of argon as the headspace gas for the majority of wine production processes. Carbon dioxide and nitrogen have their respective roles but when it comes to headspace gassing argon it the number one choice. In the production of high quality wine, it is imperative to establish proper gassing procedures. This includes the successful training of staff in all aspects of gassing procedures and the selection of the correct gas for the appropriate task. This also requires selecting the correct regulator size, hose diameter and length, the use of T-valves, measuring gas flow using a flowmeter, and finally verifying results with the use of a dissolved oxygen meter to monitor oxygen levels in the tank headspace pre and post gassing. The proper investment of time and resources in this often overlooked area of winemaking can have a profound effect on wine quality and preservation in the long run. It can also reduce long term costs by reducing the amount of gas and time required to achieve the desired reduction in the amount of oxygen present in a tank headspace.
By Dr. Molly Kelly, Enology Extension Educator, Department of Food Science
As harvest comes to a close we have planned which wines will be going through malolactic fermentation (MLF). This article provides some information to assist you in dealing with a potentially difficult MLF.
Malolactic fermentation (MLF) is a process of chemical change in wine in which L-malic acid is converted to L-lactic acid and carbon dioxide. This process is normally conducted by lactic acid bacteria (LAB) including Oenococcus oeni, Lactobacillus spp. and Pediococcus spp. O.oeni is the organism typically used to conduct MLF due to its tolerance to low pH, high ethanol and SO2. Most commercial strains are designed to produce favorable flavor profiles.
Although inoculation with a commercial starter is recommended, MLF may occur spontaneously. The lag phase associated with spontaneous MLF may increase the risk of spoilage organisms as well as the production of volatile acidity. Inoculation with a LAB culture can help avoid these problems by providing the cell population needed to successfully conduct MLF (more than 2×106 cells/mL). The compatibility of yeast and LAB should be taken into account since failed MLF may be due to incompatibility between these two organisms.
The key to a successful MLF is to manage the process and to monitor the progress. Although there has been extensive research on the MLF process, it may still be difficult to initiate at times. The possible causes of difficult MLF have been studied less extensively than those of stuck/sluggish alcoholic fermentation. In this article, factors that may influence the start and successful completion of MLF will be discussed.
The main chemical properties that influence MLF are well known: pH, temperature, ethanol and SO2 concentration. A study by Vaillant et al (1995) investigating the effects of 11 physico-chemical parameters, identified ethanol, pH and SO2 as having the greatest inhibitory effect on the growth of LAB in wine.
Generally, LAB prefer increased pH’s and usually, minimal growth occurs at pH 3.0. Under winemaking conditions, pH’s above 3.2 are advised. The pH will determine the dominant species of LAB in the must or wine. At a low pH (3.2 to 3.4) O. oeni is the most abundant LAB species, while at higher pH (3.5 to 4.0), Lactobacillus and Pediococcus will out-number Oenococcus.
MLF is generally inhibited by low temperatures. Research demonstrates that MLF occurs faster at temperatures of 200 C (68˚F) and above versus 150C (59˚F) and below. In the absence of SO2 the optimum temperature range for MLF is 23-250C (73.4˚F-77˚F) with maximum malic acid conversion taking place at 20-250C (68˚F-77˚F). However, with increasing SO2 levels, these temperatures decrease and 200C (68˚F) may be more acceptable.
LAB are ethanol-sensitive with slow or no growth occurring at approximately 13.5%. Commercial O. oeni strains are preferred starter cultures due to tolerance to ethanol. The fatty acid composition of the cell membrane of LAB can be impacted by ethanol content.
LAB may be inhibited by the SO2 produced by yeast during alcoholic fermentation. A total SO2 concentration of more than 50 ppm generally limits LAB growth, especially at lower pH where a larger portion of SO2 is in the antimicrobial form. Generally, it is not recommended to add SO2 after alcoholic fermentation if MLF is desired.
Some of the lesser known factors impacting MLF are discussed below.
MLF can be inhibited by medium chain fatty acids (octanoic and decanoic acids) produced by yeast. It is difficult to finish MLF when octanoic acid content is over 25 mg/L and/or decanoic acid is over 5 mg/L. Bacterial strains that tolerate high concentrations of octanoic and decanoic acids may be important in successful MLF. It is important to check your supplier regarding strain specifications. Yeast hulls may be added before the bacteria are inoculated (0.2g/L) to bind fatty acids. Yeast hulls may also supply unsaturated fatty acids, amino acids and assist with CO2 release.
Some fungicide and pesticide residues may negatively impact malolactic bacteria. Residues of systemic pesticides used in humid years to control botrytis can be most detrimental. Care should be taken in harvest years with high incidence of botrytis. Winegrowers should be familiar with sprays used on incoming fruit and also adhere to pre-harvest intervals.
Lees found at the bottom of a tank can become compacted due to hydrostatic pressure, resulting in yeast, bacteria and nutrients being confined to the point that they cannot function properly. Larger tank sizes may contribute to increased delays in the start of MLF. This inhibition of the start of MLF can be remedied by pumping over either on the day of inoculation or on the second day after inoculation of the bacteria.
Alternatively, contact with yeast lees can have a stimulating effect on MLF. Yeast autolysis releases amino acids and vitamins which may serve as nutrients for LAB. Yeast polysaccharides may also detoxify the medium by adsorbing inhibitory compounds. A general recommendation is to stir lees at least weekly to keep LAB and nutrients in suspension.
Residual levels of lysozyme may impact MLF. Follow the supplier’s recommendations regarding the required time delay between lysozyme additions and the inoculation of the commercial MLF culture. Strains of O. oeni are more sensitive to the effects of lysozyme compared to strains of Lactobacillus or Pediococcus.
Malic acid concentration
Malic acid concentrations vary between grape cultivars and may also differ from year to year in the same grape cultivar. MLF becomes increasingly difficult in wines with levels of malic acid below 0.8g/L. In this case a ML starter culture with high malate permease activity or a short activation protocol is recommended. Check with your supplier to ensure that the chosen strain has these attributes if needed.
Wines with levels above 5 g/L malic acid may start MLF, but may not go to completion. This may be due to inhibition of the bacteria by increasing concentrations of L-lactic acid derived from the MLF itself.
Difficult MLF can result from insufficient nutrients necessary for LAB growth. Since yeast can reduce available nutrients for LAB, time of inoculation is important to avoid competition for nutrients. The addition of nutrients when inoculating for MLF is especially important if the must and wine has low nutrient status or if yeast strains with high nutritional requirements are used. The addition of bacterial nutrients can help ensure a rapid start and successful completion of MLF.
Research demonstrates that the longer it takes to initiate MLF, there is a greater risk for Brettanomyces growth. Some inoculate during alcoholic fermentation (AF) to avoid this problem. Co-inoculation involves adding malolactic starter 24 hours after AF starts. By controlling microbial populations, the growth of spoilage organisms such as Brettanomyces may be inhibited.
Note that inorganic nitrogen (diammonium phosphate) cannot be used by LAB. Check with your supplier for the optimum nutrient product for your particular MLF needs.
Malolactic bacteria are sensitive to excessive amounts of oxygen. The bacteria should not be exposed to large amounts of oxygen after AF is complete. Micro-oxygenation may have a positive impact on the completion of MLF. This impact may be due to the gentle stirring associated with micro-oxygenation that keeps LAB and nutrients in suspension rather than the exposure to oxygen itself.
Some red grape cultivars may have difficulty completing a successful MLF. Some varieties that may experience increased MLF problems include Merlot, Tannat and Zinfandel. This may be related to certain grape tannins negatively impacting the growth and survival of LAB.
Polyphenols can have either stimulatory or inhibitory effects on the growth of wine LAB. This effect depends on the type and concentration of polyphenols as well as on the LAB strain. The tannin fraction of wine tends to complex with other compounds, minimizing their inhibitory effects on MLF. However, in wines that contain a large amount of condensed tannins only, LAB are increasingly inhibited.
MLF nutrients containing polysaccharides have been shown to minimize this effect. This may be due to interactions between the polysaccharides and tannins.
MLF difficulties are usually due to a combination of factors. A stuck or sluggish MLF is usually not the result of one factor alone. It is important, therefore, to both understand and manage the MLF process at each step of the winemaking process. Proper measurement of the process is also vital to be aware when MLF is not proceeding as desired.
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By Dr. Molly Kelly, Enology Extension Educator, Department of Food Science
In a previous post, Bryan Hed discussed early fruit zone leaf removal and its effects on the development of Botrytis bunch rot and sour rot. This is a good time to review the implications of molds and fruit rots on wine composition and quality. I will also discuss remedial actions in the winery.
Here we will focus on the most common bunch rot pathogen of mature berries, Botrytis cinerea. How severe can Botrytis bunch rot be before wine quality is impacted? This will depend on the type of rot as well as winemaking techniques however, even low levels of infection have been shown to negatively impact wine quality. Red wine quality was shown to be affected by as low as a 5% infection rate of B. cinerea. Extended skin contact in red winemaking can increase the effect of bunch rots on the finished wine. While B. cinerea can be linked with sour rot, it is more commonly associated with other fungi including Aspergillus spp. Sour rot is caused by yeast, acetic acid and other bacterial growth. When acetic acid bacteria, yeast and filamentous fungi are present together, high levels of acetic acid can result. Berries infected with sour rot have a distinct vinegar smell that may be combined with the presence of ethyl acetate. Ethyl acetate is an ester described as smelling like nail polish remover.
Laccases are enzymes produced by fungi. They break down anthocyanins and proanthocyanidins which are important phenolic compounds that contribute to palate structure and wine color. In white wines, some aromatic compounds can be oxidized resulting in the production of earthy aromas.
The largest change in must chemistry as a result of Botrytis growth is seen in amounts of sugars and organic acids. Up to 70 to 90% of tartaric and 50-70% of malic acid can be metabolized by the mold. Resulting changes in the tartaric:malic ratio cause titratable acidity to decrease and pH to increase.
There may also be clarification issues as a result of infection. The fungi produce polysaccharides including β1-3 and β1-6 glucans as well as pectins as a result of the production of enzymes capable of degrading the cell wall. In the presence of alcohol, pectins and glucans aggregate causing filtration difficulties. To mitigate this issue, pectinolytic and glucanase enzymes can be used. When adding enzymes allow at least six hours prior to bentonite additions.
Botrytis cinerea strains differ in the amount of laccase produced. This enzyme can lead to oxidation of aroma/flavor compounds and browning reactions. It can be resistant to sulfur dioxide and not easily removed with fining agents. Bentonite may remove enough laccase to minimize oxidative problems. For varieties where the potential for oxidation is increased, ascorbic acid additions can be added to juice. Since Botrytis uses ammonia nitrogen there is less available for yeast metabolism. Vitamins B1 and B6 are also depleted. Therefore supplementation with nitrogen and a complex nutrient is required. Yeast assimilable nitrogen (YAN) should be measured and adjusted accordingly to avoid stuck fermentations and production of hydrogen sulfide. Also consider inoculating with low nitrogen-dependent yeast and use more than the standard amount of 2 lbs. /1000 gallons.
Wine off-flavors and aromas result from a number of compounds when made from grapes with Botrytis(and other bunch rot organisms). Descriptors include mushroom and earthy odors from compounds such as 1-octen-3-one, 2-heptanol and geosmin. Since fruitiness can be decreased, the use of mutés (unfermented juice) from clean fruit can be added to the base wine to improve aroma. Botrytis also secretes esterases that may hydrolyze fermentation esters. Monoterpenes found in varieties such as Muscat, Riesling and Gewürztraminer can also be diminished.
When Botrytis infection is present, consider the following processing practices in addition to those mentioned above.
- Remove as much rot as possible in the field and sort fruit once it arrives at the winery. Using sorting tables is a great way to improve overall wine quality.
- Whole-cluster press whites, using very light pressure, and discard the initial juice.
- Harvest fruit cool and process quickly. Sulfur dioxide can be added to harvest bins to inhibit acetic acid bacteria.
- Enological tannin additions will bind rot-produced enzymes. They can also bind with protein and decrease the bentonite needed to achieve protein stability. Note: Remember to not add tannins and commercial enzymes at the same time since tannins are known enzyme inhibitors. After an enzyme addition allow six to eight hours before adding tannins.
- Minimize oxygen uptake since laccase activity is inhibited in the absence of oxygen. Inert gas can be used at press, during transfers and to gas headspace.
- Use a commercial yeast strain that will initiate a rapid fermentation. The resulting carbon dioxide will help to protect against oxidation.
- Once fermentation is complete, rack right away. Both Botrytis and laccase settle in the lees.
- Phenolic compounds are the main substrate for fungal enzyme activity. Removal of undesirable phenolic compounds can be achieved using protein fining agents (ex: gelatin, casein, isinglass). The synthetic polymer PVPP can also be used in juice or wine to remove oxidized phenolic compounds.
- Only cold soak clean fruit. Avoid cold soak and extended maceration on Botrytisinfected fruit as this may encourage fungal and bacterial growth.
As always, it is best to avoid rot-compromised fruit, however, using these practical winemaking tips should help to minimize negative impacts on wine production and quality.
DeMarsay, A. Managing Summer Bunch Rots on Wine Grapes, Maryland Cooperative Extension.http://extension.umd.edu/sites/extension.umd.edu/files/_docs/programs/viticulture/ManagingSummerBunchRots.pdf. Accessed 7 May 2018.
Ribereau-Gayon, P. 1988. Botrytis: Advantages and Disadvantages for Producing Quality Wines. Proceedings of the Second International Cool Climate Viticulture and Oenology Symposium. Auckland, New Zealand, pp. 319-323.
Steel, C., J. Blackman, and L. Schmidtke. 2013. Grapevine Bunch Rots: Impacts on Wine Composition, Quality, and Potential Procedures for the Removal of Wine Faults. J. Agric. Food Chem. 61: 5189-5206.
Zoecklein, B. 2014. Fruit Rot in the Mid-Atlantic Region, On-line Winemaking Certificate Program, Wine Enology Grape Chemistry Group, Virginia Tech. http://www.vtwines.info/. Accessed 16 April 2018.
Zoecklein, B. 2014. Grape Maturity, On-line Winemaking Certificate Program, Wine Enology Grape Chemistry Group, Virginia Tech.http://www.vtwines.info/. Accessed 16 April 2018.
By Molly Kelly
There are a number of spoilage microorganisms and yeasts that we are concerned with as winemakers. Two of the most common spoilage yeasts include Kloeckera apiculata and Brettanomyces bruxellensis. The most common form of yeast spoilage is due to Brettanomyces bruxellensis. Although mature grapes may harbor this spoilage yeast, the bigger problem can occur when winery equipment is infected due to poor sanitation practices. This yeast produces volatile phenols and acetic acid. Examples of wine flaws include aromas described as “medicinal” in white wines and “leather” or “horse sweat” in red wines. Other aromas descriptors include barnyard, wet dog, tar, tobacco, creosote, plastic and band aids.
The two major groups of wine spoilage bacteria can be placed in either the acetic acid bacteria (AAB) group or the lactic acid bacteria (LAB) group. The AAB includes the genera Acetobacter and Gluconobacter. Both have aerobic (requiring oxygen) metabolisms and thus their growth generally occurs on wine surfaces as a translucent film that tends to separate into a patchy appearance. In contrast, the LAB require low oxygen conditions for growth (i.e. they are microaerophilic to facultative anaerobic micro-organisms). The LAB includes the genera Lactobacillus, Pediococcus and Oenococcus.
During fermentation the presence of such microbes may be indicated by a spontaneous or sluggish fermentation, or a spontaneous malolactic fermentation (MLF); or the presence of ethyl acetate, volatile acidity (VA) or other off-odors.
Winery Microbiology Laboratory
Because of these possible faults arising due to the presence of spoilage organisms, some wineries have incorporated sanitation monitoring and microbiological techniques into their production practices. Some considerations when planning a winery microbiology laboratory are: space considerations, availability of trained staff to perform testing, willingness to maintain adequate record-keeping, equipment costs as well as the cost of consumables.
A microscope capable of 1000x magnification is needed to view bacteria and yeast. These can cost anywhere from $1000-$3000 or more but bargains can be found on used microscopes. A phase-contrast microscope requires no staining of slides due to enhanced differences in refractive index between the microorganisms and surrounding medium. This feature also allows for rapid detection and response. The staff in the microbiology lab should have training in the proper use of a microscope as well as identification of microorganisms.
In addition to identifying spoilage organisms, a microscope can be used to monitor yeast populations. By using a simple methylene blue stain, yeast viability can be determined.
Bacterial culture media is available for the growth of spoilage organisms for identification. This requires additional equipment including an incubator. This also requires further training in sterile technique and organism identification techniques. Several types of culture media exist for the detection of the organism of interest. For example, media used to plate for Brettanomyces contains chloramphenicol (200 mg/L) to prevent bacterial growth while others may contain cyclohexamide to prevent Saccharomyces growth. Common media used in culturing juice, wine and environmental samples include WL and WL-differential agar.
Membrane Filter Method
The membrane filter method can be used to isolate small numbers of microbes from a liquid sample. A sterile cellulose nitrate membrane (0.45 microns for bacteria, 0.65-8 microns for yeasts) is placed on a vacuum flask and filtered. Using sterile technique, the membrane is placed on the culture plate and monitored for growth. This method could be used to check bottle sterility.
The swab test method is used for semi-quantitative analysis. Moist sterile cotton swabs are used to monitor dry areas (moistened with sterile saline or water). Dry swabs can be used to test moist areas. The swabs can then be used to inoculate the proper agar medium, depending on the organism of interest. Agar plates can also be used to detect airborne organisms at critical winery locations. Plates are left open for 30 minutes to 2 hours and then incubated. Airborne organisms that settle on the plate will grow and can be further identified.
Monitoring systems exist that utilize bioluminescence technology to measure adenosine triphosphate (ATP). ATP is found in all plant, animal and microbial cells and is the prime energy currency that fuels metabolic processes. It is therefore possible to detect and measure biological matter that should not be present if proper sanitation practices are followed. One system by Hygiena™ uses an enzyme found in fireflies (luciferase). In the presence of ATP, an oxidation reaction occurs that results in light formation that is directly proportional to the amount of ATP present. Results are numeric and expressed as relative light units (RLU).
It should be stressed that cellar hygiene is critical in maintaining wine integrity and quality. Poor wine quality is usually due to poor sanitation practices. Areas of spoilage organism build-up include: the vineyard, second-hand barrels, imported bulk wine and areas of the winery that are difficult to reach.
There are commercial enology laboratories that provide all of the microbiological services discussed here. For more information please contact Molly Kelly at email@example.com.
Crowe, A. August 2007. Avoiding Stuck Ferments. Wine Business Monthly
Just, E. and H. Regnery. 2008. Microbiology and wine preventive care and monitoring in the wine industry. Sartorius Stedim Biotech.
Margalit, Y. 1996. Winery Technology and Operations. The Wine Appreciation Guild, San Francisco.
Ritchie, G. 2006. Stuck Fermentations. Fundamentals of Wine Chemistry and Microbiology. Napa Valley College.
Specht, G. Sept/Oct 2003. Overcoming Stuck and Sluggish Fermentations. Practical Winery and Vineyard Journal.
Van de Water, L. Sept/Oct 2009. Monitoring microbes during fermentation. Practical Winery and Vineyard Journal.
Zoecklein, B., Fugelsang, K.C., Gump, B.H. and Nury, F.S. 1999. Wine Analysis and Production, Kluwer Academic/Plenum Publishers, New York.
Zoecklein, B. 2002. Enology Notes #65, Enology-Grape Chemistry Group, Virginia Tech.
Please join us in welcoming Dr. Molly Kelly as our new Penn State Enology Extension Educator. In this role, Molly will support the technical needs of the Pennsylvania wine industry and lead educational programming focusing on wine quality. She has lead workshops, including winery sanitation, filtration, microscopy, wine analysis and berry sensory analysis. Molly’s research has focused on the effect of nitrogen and sulfur applications on Petit Manseng wine aroma and flavor. Her current research includes a pre-harvest, on-the-vine dehydration study in collaboration with Virginia Tech University.
Prior to this position, Molly was the Enology Extension Specialist at Virginia Polytechnic Institute and State University in Blacksburg, Virginia. She also held the position of enology instructor at Surry Community College in Dobson, N.C., where she developed the enology curriculum and managed all aspects of the college’s 1,000-case bonded winery. Under her direction, Surry produced numerous international award-winning wines. Prior to her position at Surry, she was a biodefense team microbiologist with the New York State Department of Health.