Practical Winery
65 Mitchell Blvd, San Rafael, CA 94903
phone: 415-453-9700 ext 102
1 · 2 · 3 · 4 · Microbes Part 2
September/October 2009
Monitoring microbes during cellaring/bottling
Acetobacter in wine
Non-Saccharomyces yeasts from grapes
There are yeasts on grapes, though very few are Saccharomyces. Kloeckera apiculata (teleomorph Hanseniaspora uvarum) is the major species, and there are quite a few other species also. Some are sensitive to alcohol, or are aerobic so they cannot grow during fermentation, but they may grow before or during the start of fermentation. Other species may survive until middle to late in the fermentation process.
If odors of ethyl acetate (airplane glue/nail polish remover), amyl acetate (banana skin), or other offodors appear in juice or must, examine immediately under a phase-contrast microscope for yeasts that are not the round/ovoid shape of Saccharomyces. Non-Saccharomyces yeasts are sometimes round, but more often have other appearances: apiculate (pointy), elongated, or bizarrely shaped, and they may be unusually small or large.
Monitoring microbes during cellaring/bottling
Brettanomyces in stuck wine.
Kloeckera is especially problematic for several reasons. Their SO2 sensitivity is comparable to that of Saccharomyces, and they are naturally coldtolerant so they grow readily in cool must (10 to 15°C). They can survive much farther into the fermentation than most other non-Saccharomyces species, sometimes all the way to the end.
Sometimes a thick, slimy scum forms, or stringy clumps that fall out later as fluffy lees. Kloeckera can make prodigious amounts of ethyl acetate and amyl acetate, and is quite efficient in quickly depleting nutrients, particularly thiamine and other vitamins. Microscopically, it resembles bowling pins or thin lemons with a knob on each end.
Kloeckera is easy to culture because it is cycloheximide-resistant and grows in one to two days, unlike Brettanomyces (three to seven days), which Kloeckera resembles. There are also PCR primers for Kloeckera, and for Pichia membranifaciens, which could be used at this stage, though not for other non-Saccharomyces vineyard species, so those will not be detected by PCR.
However, the question is notwhether there are Kloeckera and other vineyard yeasts in themust (there are), but howto monitor their population, and how to handle themust so that they do not grow too extensively. For this purpose, following the must microscopically, estimating percentages of non- Saccharomyces species, is more useful. Before fermentation begins, if more than 5 or 10 non-Saccharomyces yeasts are seen in a 40x field, a problem is developing.
Examine some musts microscopically every day, especially any must over 10°C that are soaking on skins or warming up from soaking/settling, or musts that start fermenting spontaneously. Also examine microscopically if acetate or amyl acetate odor is noted.
Microbes to watch out for: Non- Saccharomyces yeasts, lactic acid bacteria
Danger signals: Spontaneous fermentation, ethyl acetate or other off-odor, sluggish fermentation, spontaneous malolactic fermentation (MLF), low viability of yeast inoculum, VAincrease
Monitoring methods: Microscopic exam, sensory cues, chemical tests, PCR-based genetic testing
Monitoring microbes during cellaring/bottling
Yeast and Lactobacilli in stuck wine
(by Patricia Roca)
When preparing dry yeast, check viability with methylene blue stain to make sure that enough survived transport, storage, and rehydration in the winery. Approximately half of the rehydrated yeasts should be viable, as evidenced by lack of blue staining. Counting yeasts with a hemocytometer also helps to monitor the population to be sure that it reaches 108 cells/ml, or a stuck ferment is likely.
Some winemakers encourage the growth of some non-Saccharomyces vineyard species to contribute to aromatics, complexity, andmouthfeel before Saccharomyces, which must become dominant for the fermentation to go to completion, takes over. Potential problems with this approach include depletion of vitamins and nutrients (natural and added) that Saccharomyces will need, production of off-characters such as H2S, ethyl acetate, and acetic acid, and possibly inhibition of Saccharomyces. It is very important for winemakers encouraging participation by vineyard yeast species to guard vigilantly against their extensive growth. Many stuck fermentations have their origins early on, as growth of competing species did not allow Saccharomyces to achieve a large population of healthy yeasts. The winemaker may look for a cause late in fermentation, but what happens at the beginning can set the process in motion.
Spontaneous fermentations, whether intentional or not, should be checked microscopically until Saccharomyces becomes the dominant yeast. If Saccharomyces does not take over within the
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