Sulphur dioxide (SO2) addition is a well-established practice in winemaking for controlling the growth of yeasts and bacterial populations. However, SO2 is a potential allergen and can exacerbate asthmatic symptoms. Ultraviolet radiation (UV)-C (254 nm) may be a potential safe alternative to SO2 for inactivating microorganisms in wine. Indeed, UV-C processing has been reported to reduce microbial counts from 2 to 6 log depending on the UV-C dosage and absorptive properties of the wine. The traditional cultivation-based methods for enumerating wine microorganisms are time-consuming and labour-intensive and are not able to detect cells in VBNC status. Therefore, in this study Propidium MonoAzide (PMA)-qPCR technique was applied to quantify yeasts, lactic acid bacteria and acetic acid bacteria. Amplification was obtained with universal primers and detection of viable cells was possible thanks to the presence of PMA, a dye able to penetrate into dead cells and bind to dsDNA with high affinity, thus blocking subsequent PCR. In this way, only DNA isolated from viable cells can be quantified. UV-C- treated and control white wine samples were analysed by PMA-qPCR and standard plating methods. Results were concordant and, in some cases, the molecular method was able to detect lower quantities of cells. Results confirm previous observations on the efficacy of UV-C treatment but, remarkably, PMA-qPCR give results in 2 days versus the 8 usually necessary for traditional counting.
Effect of UV-C treatment on microbial populations of white wines as revealed by PMA-qPCR
FRACCHETTI, Fabio;DEL CASALE, Antonio;FELIS, Giovanna;TORRIANI, Sandra
2013-01-01
Abstract
Sulphur dioxide (SO2) addition is a well-established practice in winemaking for controlling the growth of yeasts and bacterial populations. However, SO2 is a potential allergen and can exacerbate asthmatic symptoms. Ultraviolet radiation (UV)-C (254 nm) may be a potential safe alternative to SO2 for inactivating microorganisms in wine. Indeed, UV-C processing has been reported to reduce microbial counts from 2 to 6 log depending on the UV-C dosage and absorptive properties of the wine. The traditional cultivation-based methods for enumerating wine microorganisms are time-consuming and labour-intensive and are not able to detect cells in VBNC status. Therefore, in this study Propidium MonoAzide (PMA)-qPCR technique was applied to quantify yeasts, lactic acid bacteria and acetic acid bacteria. Amplification was obtained with universal primers and detection of viable cells was possible thanks to the presence of PMA, a dye able to penetrate into dead cells and bind to dsDNA with high affinity, thus blocking subsequent PCR. In this way, only DNA isolated from viable cells can be quantified. UV-C- treated and control white wine samples were analysed by PMA-qPCR and standard plating methods. Results were concordant and, in some cases, the molecular method was able to detect lower quantities of cells. Results confirm previous observations on the efficacy of UV-C treatment but, remarkably, PMA-qPCR give results in 2 days versus the 8 usually necessary for traditional counting.
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