Révision d’un stock congelé de souches de bactéries lactiques: Contrôle de la viabilité et de l’identification.

Abstract

The viability and stability of industrial strains during long-term storage is one of the key properties that characterize the quality of these strains, with a view to sustainable exploitation. In this study, a stock of 32 isolates of lactic acid bacteria stored at -20 °C in the presence of 30% glycerol for many years was revised. This revision concerned firstly, direct evaluation of cell viability of isolates, after thawing, by direct microscopic count method, using “Petit Salumbeni” counting cell. Results of this evaluation based on the calculation of the decline rate (in Log CFU / ml) and the survival rate (in %) revealed a change in the number of isolates compared to their initial number before storage. Likewise, great variability in resistance degree to frozen storage was observed among isolates. The decline rate was between 0,2 and 2,5 Log CFU/ml and isolate No. 6 was distinguished by its high stability, showing a decrease of 0,2 Log CFU/ ml against the other isolates and in particular isolate No. 31 which showed out to be the least resistant with a decrease of 2,5 Log CFU/ml. In addition, the calculated survival rate (%) confirmed the results previously observed, revealing a survival of 67% and 0%, respectively for isolates 6 and 31. Secondly, the work focused on reactivation and control of the identification of isolates. Twelve isolates out of the 32 studied were able to grow on MRS broth, after reactivation at 37°C for 24hours or after prolongation up to 72hours. The first steps of identification concerned only, the determination of the macroscopic and microscopic aspects. The reactivated isolates developed on MRS-agar colonies with variable appearances characteristic to the lactic acid bacteria. The wet drop microscopic examination revealed cocoid and bacillary forms, with a dominance of the bacillary form. Finally, the scientific papers of Fonseca et al., (2006), Cabeza et al., (2011) and O'Brien et al., (2016) presented in this work propose different protocols for studying the viability of lactic acid bacteria cultures, subjected to different preservation methods and under the influence of several factors. The study by O’Brien et al., (2016) pointed out the importance of freezing in preserving the probiotic strains contained in traditional kefir. That of Cabeza et al., (2011) had indicated that freeze-drying is the most appropriate method for a more or less efficient preservation and finally Fonseca et al., (2006) had noted that the cooling rate and the freezing temperature have a great influence on the viability and acidifying activity of Lactobacillus delbrueckii subsp. bulgaricus.