Microbial composition and viability of natural whey starters used in PDO Comté cheese-making.

Natural whey starters (NWS) are cultures with undefined multiple-strains species commonly used to speed up the fermentation process of cheeses. The aim of this study was to explore the diversity and the viability of Comté cheese NWS microbiota. Culture-dependent methods, i.e. plate counting and genotypic characterization, and culture-independent methods, i.e. qPCR, viability-qPCR, fluorescence microscopy and DNA metabarcoding, were combined to analyze thirty-six NWS collected in six Comté cheese factories at two seasons. Our results highlighted that NWS were dominated by Streptococcus thermophilus (ST) and thermophilic lactobacilli. These species showed a diversity of strains based on Rep-PCR. The dominance of Lactobacillus helveticus (LH) over Lactobacillus delbrueckii (LD) varied depending on the factory and the season. This highlighted two types of NWS: the type-ST/LD (LD > LH) and the type-ST/LH (LD < LH). The microbial composition varied depending on cheese factory. One factory was distinguished by its level of culturable microbial groups (ST, enterococci and yeast) and its fungi diversity. The approaches used to estimate the viability showed that most NWS cells were viable. Further investigations are needed to understand the microbial diversity of these NWS.

Cultivable microbial communities in raw cow milk and potential transfers from stables of sixteen French farms.

The indigenous microflora in raw milk plays an important role in the diversity of cheese flavours and may protect against the growth of pathogens, but the sources of contamination and the factors that might affect the microbial communities in milk are not well known. The objectives of this study were to broaden knowledge of the microbial composition of milk and to assess microbial transfers from the stable to the milk. Air (collected in milking parlour and stable), dust (passively collected using plastic box), cow teat surface, and hay and milk samples were collected in 16 French farms with either stanchion barn or freestall barn configurations and plated on various culture media. Bacterial and fungal colonies were identified using phenotypic and DNA sequencing methods. Results showed that most of the fungal species and environmental bacteria found in the milk were also found in the stable and the milking parlour environments, indicating large microbial transfer from stable to milking parlour then to milk. However, milk from the stanchion barns were more contaminated than milk from freestall barns. Contrasting with other bacterial and fungal species, useful cheese-making bacteria--lactobacilli and PAB--were frequently identified in the milk and on the teat surface but were rarely found in other environments. In conclusion, milk contamination by the stable environment is considerable, even if it is lower in farms with a milking parlour. Besides this environmental contamination, the teat surface remains the main source of useful cheese-making bacteria.

Effect of mesophilic lactobacilli and enterococci adjunct cultures on the final characteristics of a microfiltered milk Swiss-type cheese.

The effect of four associations of adjunct cultures composed of mesophilic lactobacilli and enterococci, either solely or combined, on the microbiological, biochemical and sensory characteristics of Swiss-type cheese made using microfiltered cows' milk and supplemented with propionibacteria was studied. The global pattern of growth was similar to that generally observed in raw milk cheese and interactions between microflora were highlighted during ripening. Enterococci, which negatively affected the survival of streptococci starters, seemed to play a limited role in the formation of volatile compounds, probably due to their low levels throughout ripening. On the contrary, mesophilic lactobacilli, which affected the evolution of propionibacteria, enterococci and L. delbrueckii subsp. lactis starter counts, modified free amino acid content, production of volatile compounds and organoleptic properties of mature cheese. This population appeared to be of major importance in the formation of cheese flavor as it was positively related to numerous potential flavor compounds such as alcohols and their corresponding esters, acetaldehyde and 4-methyl-4-heptanone. The original mesophilic lactobacilli present in milk could play an important role in the sensorial diversity of raw milk Swiss-type cheeses such as Comte.

Use of PCR-based methods and PFGE for typing and monitoring homofermentative lactobacilli during Comté cheese ripening.

This study investigated the genotypic characteristics of selected and wild homofermentative thermophilic lactobacilli strains during ripening of Comté cheeses, made into two cheese plants. Both amplification and restriction analysis of the 16S rRNA gene (PCR-ARDRA) and classical biochemical tests were used to identify isolates. Diversity within homofermentative lactobacilli was not found in their species composition since the same two species Lactobacillus helveticus and L. delbrueckii susbp. lactis were isolated from cheeses. In cheeses made with natural whey starter, it appeared that the most likely sources of L. helveticus and L. delbrueckii susbp. lactis were the starter and raw milk, respectively. The examination of RAPD profiles of lactobacilli strains revealed 19 RAPD groups among 50 isolates, which were different from selected starter strains. Using RAPD, REP-PCR, and PFGE to identify selected starter strains during cheese ripening, we showed that L. helveticus decreased quickly while L. delbrueckii susbp. lactis sustained high viability during ripening. The use of selected L. delbrueckii susbp. lactis strains diminished the genetic diversity among strains isolated from cheese, probably in preventing the raw milk microflora from growing.