RAPID AND SENSITIVE IN-SITU HYBRIDIZATION METHOD FOR DETECTING AND IDENTIFYING LACTIC-ACID BACTERIA IN WINE

A rapid and sensitive in situ hybridization method, using total genomi c DNA probes, can specifically detect and identify the lactic acid bac teria in wine. The total genomic DNA probes were labelled with digoxyg enin (DIG) by random priming and hybridized with the genomic DNA of th e bacteria to be identified. The hybrids were detected with fluorescen t anti-DIG Fab-fragments or with enzyme-conjugated anti-DIG Fab-fragme nts. The cell wail of the Gram-positive lactic acid bacteria was made permeable with a lysozyme treatment to allow the accessibility of the probe to the target DNA and the penetration of anti-DIG antibodies whi ch specifically bound to their antigens. The total genomic DNA probes exhibited the same specificity with dot-blot and in situ hybridization . The in situ hybridization technique made possible the control of the bacterial population in wine at different stages of vinification of s torage, and the identification of lactic acid bacteria which can cause wine spoilage. (C) 1998 Academic Press.