Reverse transcription quantitative PCR revealed persistency of thermophilic lactic acid bacteria metabolic activity until the end of the ripening of Emmental cheese

For Emmental manufacture two kinds of adjunct culture are added: (i) thermophilic lactic acid bacteria (starters) such as Lactobacillus helveticus (LH), and Streptococcus thermophilus (ST) growing the first day of the manufacture and (ii) ripening culture. ST and LH have a key role in curd acidification and proteolysis at the beginning of the manufacture but are considered to be lyzed for a great part of them at the ripening step. The aim of this work was to assess the metabolic activity of these bacteria throughout manufacture and ripening. During Emmental cheesemaking, LH and ST were subjected to i) population quantification by numerations and by quantitative PCR (qPCR) ii) reverse transcription (RT) Temporal Temperature Gel Electrophoresis (TTGE) iii) transcript quantification by RT-qPCR targeting 16S rRNA, tuf and groL mRNAs to evaluate bacterial metabolic activity. During ripening, ST and LH numerations showed a 2.5 log10 loss of culturability whereas qPCR on pelleted cells revealed only one log10 of decrease for both of these species. 109 ST and 108 LH cells/g of cheese still remained. They contained a stable number of 16S transcript and at least 106 copies of mRNAs per 109 cells until the end of ripening. These results prove the unexpected persistency of thermophilic lactic acid bacteria starters (ST and LH) metabolic activity until the end of ripening and open new perspectives in term of their involvement in the quality of cheeses during ripening.

► Emmental cheeses were manufactured and ripened for 2 months.
► Thermophilic starter bacteria (ST, LH) were counted throughout manufacture by Q PCR.
► Lysis during manufacture appeared 1.5 log10 less than suspected with plate count.
► More than 7 log10 thermophilic bacteria/g are peletable throughout the ripening.
► rRNA, mRNA quantifications showed metabolic activity of ST and LH during ripening.