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WINEMAKING Leveau tried the technique out on the material sampled from the surface of grapevine leaves and grape berries, and not surpris- ingly he found a huge number of different life forms. What was not so predictable was finding, in addition to standard vineyard microbes, a large amount of novel stuff not found in any current DNA database. The rRNA gave clues about the biological fam- ily and even the genus of these strangers, but then, that only gets us back to the Mona Lisa/baby conundrum. Friendly bugs? Bad bugs? Do we really know what's on our grapevines? Mills put the machinery to work on samples taken at various points during and shortly after a fermentation, and again he found not only standard microbes but a considerable number of things not covered in any enology text and not well described in broader literature. Maybe most intriguing, Mills' readouts showed some interlopers that seemed to grow in numbers over time, running against the trend of most mi- crobes to die off in the presence of rising ethanol. These are simply trial runs on limited samples and are more proof of concept than ecosystem profiles. The news here is not the identifica- tion of hitherto unexpected strains of Polysyllabicus obfuscata, but the demonstration of the power of the techniques. Leveau said that what he would like to do with this technology is track the entire collection of microbiota from leaf and berry into the cellar and in the bottle over time. No one has come close to doing that yet; indeed, one of the few published studies using comprehensive PCR to look at any kind of fermented product focused on pearl millet slurries. Yum! PCR and pyrosequencing in all their forms have limitations, of course. One important gap, according to Mills, is that "none of these methods differentiate between live and dead cells. There is no pub- lished, peer-reviewed evidence that standard quantitative PCR can distinguish live and dead." As you might imagine, people are work- ing on this. Another limitation is that comprehensive PCR only gives the relative numbers of different microbes, not the actual, absolute counts, though that can often be derived from going back and using quantitative PCR to search for a specific critter. Leveau notes that sample preparation is critical to ensure, for example, that the analy- sis is performed on things sitting on leaves and berries, not things inside them. At this point, there are no universal protocols for sample preparation, which gets done case by case. The ability to interpret results from a PCR run depends on the quality of the reference databases used to match the findings. With the explosion of technology and the multiplication of the teams of researchers using it, Leveau says dealing with the huge amount of data is a struggle in itself. And one more problem: While these methods have gotten dra- matically cheaper, they are not free. Putting one of these high- powered machines to work on a set of samples for an eight-hour sequencing search costs several thousand dollars. Systems like the Roche 454 are quite expensive; Davis, for example, doesn't have one, and relies on a centralized analytical services provider to do the actual runs. Tapping the potential of these techniques requires major financial resources, and it would behoove the wine industry to make sure Leveau, Mills and their kin spend their time search- ing for microbes, not for donations. In the world of research, timing isn't everything; funding is. Tim Patterson is the author of "Home Winemaking for Dummies." He writes about wine and makes his own in Berkeley, Calif. Years of ex- perience as a journalist, combined with a contrarian streak, make him interested in getting to the bottom of wine stories, casting a critical eye on conventional wisdom in the process. Skolnik_Dir10_Jan10.qxd 10/19/09 10:04 AM Page 1 STAINLESS STEEL COOPERAGE Providing leading wine makers with stainless steel barrels of high quality, durability, and design. SKOLNIK INDUSTRIES, INC. 4900 SOUTH KILBOURN AVENUE CHICAGO IL 60632-4593 PHONE 773.735.0700 FAX 773.735.7257 TOLL FREE 1.800.441.8780 HTTP://WINEDRUM.SKOLNIK.COM EMAIL: SALES@SKOLNIK.COM Wines & Vines JULY 2011 53