Assessment of the dynamics of the physiological states of Lactococcus lactis ssp. cremoris SK11 during growth by flow cytometry.

AIMS: The aim of this study was to improve knowledge about the dynamics of the physiological states of Lactococcus lactis ssp. cremoris SK11, a chain-forming bacterium, during growth, and to evaluate whether flow cytometry (FCM) combined with fluorescent probes can assess these different physiological states. METHODS AND RESULTS: Cellular viability was assessed using double labelling with carboxyfluorescein diacetate and propidium iodide. FCM makes it possible to discriminate between three cell populations: viable cells, dead cells and cells in an intermediate physiological state. During exponential and stationary phases, the cells in the intermediate physiological state were culturable, whereas this population was no longer culturable at the end of the stationary phase. CONCLUSIONS, AND IMPACT OF THE STUDY: We introduced a new parameter, the ratio of the means of the fluorescence cytometric index to discriminate between viable culturable and viable nonculturable cells. Finally, this work confirms the relevance of FCM combined with two fluorescent stains to evaluate the physiological states of L. lactis SK11 cells during their growth and to distinguish viable cells from viable but not culturable cells.

Analysis of fungal diversity of grapes by application of temporal temperature gradient gel electrophoresis - potentialities and limits of the method.

AIMS: Fungi could be responsible for several problems in wines but the fungal ecosystem of grapes remains little known. The use of traditional methods does not allow to describe quickly this ecosystem. Therefore, we need to improve the knowledge about these fungi to prevent defects in wine. This study aims at evaluating the potentialities of the temporal temperature gradient gel electrophoresis (TTGE) method to describe the fungal ecosystem of grapes. METHODS AND RESULTS: The internal transcribed spacer (ITS) region was amplified and analysed using TTGE. A reference database of 56 fungal species was set up to evaluate the discrimination power of the method. The database was used for the direct identification of the fungal species present in complex samples. The sensitivity of the method is below 10(4) spores per ml. CONCLUSIONS: This method allows to describe the fungal diversity of grapes, but does not always allow to directly identify all fungal species, because of the taxonomic resolution of the ITS sequences. However, this identification strategy is less time consuming than traditional analysis by cloning and sequencing the bands. SIGNIFICANCE AND IMPACT OF THE STUDY: With this method, it will be possible to compare the fungal species present in different vineyards and to connect the presence of some fungi with particular defects in wine.