Did you know that genetically modified (GM) wine has been marketed in the United States for the past three years?
In 2003, the United States Food and Drug Administration (FDA) designated the GM yeast strain Saccharomyces cerevisae ML01 ‘generally recognized as safe’ (GRAS). The strain included a gene for malolactic enzyme from the bacterium Oenococcus oeni and a malate permease gene from the fission yeast Schizosaccharomyces pombe. Wine making involves alcoholic fermentation via the metabolic pathways of yeast and the malolactic pathway to convert malic acid to lactic acid, to reduce the acidity of the wine. Malolactic acid fermentation is usually achieved using lactic acid bacteria, which have a permease for malic acid. Putting the two fermentation pathways in one organism seems a good idea.
The yeast ML01 was modified using a shuttle vector containing a chromosome integration cassette with genes for malolactic enzyme, malate transporter (permease), regulatory genes and a sequence directing homologous recombination at a chromosomal locus (not specified in the FDA report), and the antibiotic phleomycin gene was used as a selectable marker via another plasmid. After culturing the selected antibiotic resistant lactic-acid producing yeast, a phleomycin-sensitive lactic acid producing strain was isolated and found to contain the integrated malolactic-malate transporter genes. The original anti-phleomycin plasmid did not contain a sequence allowing it to be integrated into the yeast chromosome and the plasmid was therefore unstable and frequently lost from the yeast cell . The company distributing the GM yeast, Springer Oenologie, Lesaffre Group of North America, stated that the malate transporter gene and malolactic gene were both controlled by the phosphoglycerate kinase gene (PGK1) promoter and transcription terminator. The recombinant yeast softened the wine’s ‘mouth feel’ by decreasing its acidity, and also reduces buttery flavours (diactyl) due to lactic acid secondary metabolism .
Genetic modification of yeast and bacteria differs fundamentally from modification of plants. The modification of bacteria and yeast is based on homologous recombination while modification of plants is based on illegitimate recombination. In plants the recombinant gene insertions are not precise and disrupt genes that are not specifically targeted, while in yeast, gene insertions disrupt genes that are targeted. For yeast genetic engineering ‘shuttle’ vectors are used, which are propagated in bacteria for insertion in yeast. The shuttle vector may also replicate autonomously in the yeast nucleus and express genes equipped with promoter and terminator genes. The expression vector comes equipped with a sequence homologous with a yeast chromosomal gene. Recombination between vector and chromosomal gene disrupts the target chromosomal gene and inserts the transgene, and frequently also a selectable marker, into the yeast target locus .
The yeast released for commercial wine production, ML01, was found to be only somewhat substantially equivalent to unmodified wine yeast, as a cytochrome p450 enzyme protein appeared to have been altered from the parental strain based on a comprehensive analysis of the yeast cell proteins, and a number of codon changes were observed in the inserted malolactic gene cassette, but those changes were not considered significant. Strain ML01 was claimed to reduce levels of amines such as putrescine and cadaverine, preventing unpleasant side effects of wine drinking .
Wine yeasts genetically unstable and DNA persists in wine
Neither the FDA document  nor the publications on ML01  considered important complications associated with wine yeast. Wine yeasts are unstable and sudden losses in heterozygosity have been observed. Such abrupt changes in the phenotype of wine yeasts are commonplace . Numerous translocations have been observed uniquely in wine yeasts and such chromosome rearrangements involving transgenes can lead to unexpected toxicity in the final product . Yeast cells in wine were found to be hyperactive in mitotic recombination, contributing to the observed instability of wine yeasts .
How much yeast nucleic acid (DNA and RNA) is carried over into wine? Autolysis of wine yeast releases nucleic acids that persist in the wine for at least nine years and contribute to the flavour of wine . The fate of yeast and plant DNA, as monitored by yeast chitinase gene and the plant chlorophyll a/b binding protein gene and micro-satellite markers showed that large DNA markers were present in must (the starting fermentation mixture of mainly yeast and grape juice), while the 250 base pair micro-satellites were present in both must and young wine for up to six months .
Culturable yeast cells were isolated from two out of five bottled wines that had been filtered prior to bottling, and originating from different locations in Greece. Unfiltered wines stored on oak barrels between 1998 and 2002 all contained live yeast . The FDA letter designating wine yeast ML01 to be GRAS indicated that the distributor of GM yeast believed that final wines were free of yeast and yeast DNA, but no data were provided to support that conclusion.
The dissemination and survival of commercial wine yeast in the vineyard was studied over a period of three year, which indicated that the strains were mainly recovered at fairly close proximity to the winery, up to 200 metres. The yeast was largely disseminated by water runoff. The commercial strains tended to appear and disappear from the vicinity of the winery .
The yeasts from a winery abandoned in 1914 differed from those isolated in a modern winery; with the genetic characteristics of the yeast in the abandoned winery persisting for over ninety years . A study of wine jars from the tombs of ancient Egypt showed that S. cerevisiae had been used in winemaking by at least 3150 BC . Regulators should take note of the time that GM yeast may persist.
FDA reviews reads like pr on behalf of GM wine
In the United States, approval of GM plants such as grapes is granted by USDA/APHIS and those reviews provide fairly full information that is made accessible to the public. FDA alone reviews and approves GM microbes such as yeast used in food products. Their full reviews including all required support information does not appear to be readily accessible, and their approval reports, such as the GRAS notice on GM wine yeast reads more like a public relations release on behalf of the promoters of GM wine yeast . The FDA review did not consider the environmental and human health consequences of marketing and consumption of GM wine. The view that the yeast and its autolysis products including DNA, RNA, proteins and carbohydrates are somehow lost from the wine is not supported by scientific evidence. The GM wine yeast does not appear to have been tested for toxicity in animal feeding experiments, nor was the must and finished wine.
A medical journal, The Lancet, pointed out that international faith in the FDA is fast eroding because approvals are frequently influenced by political pressure , and the approval of wine yeast certainly left fundamental questions unanswered. It is surely premature to market GM wine yeast, and as the wines produced using GM yeast are not labelled, it is only prudent for consumers to avoid US wines unless there is information available indicating that GM wine yeasts have not been used.
Van Vuuren, one of the patent holders on GM yeast, was recently quoted as saying that ,“several wineries were using the ML01 yeast as of a few years ago”, but “it is not known if they or others are using it today or exporting wines containing it.” Furthermore, “The company marketing these GE yeast has no legal obligation to identify their customers and has not done so.” Van Vuuren also said “he had heard that some European wineries were buying the yeast in California and shipping it to Europe.”
The Pew Report on GM Wine and Grapes asks : “Are some US and European wineries using this GM yeast for commercial production, or merely for research purposes? If the yeast is being used in producing commercial wines, and these wines are being exported to countries in which it has not yet been approved for use, what can or should the wine industry do about it?”
Currently there are no validated methods to detect GM wine made using GM yeast ML01, and it would be necessary to have develop one.
The Industrial College of the Armed Forces (USA) indicated that the “biotechnology industry is a critical element of national power” . I hope that military force won’t be used to make people drink the GM wine. Institute of Science in Society