Abstract
The use of selected autochthonous mesophilic lactic acid bacteria as starter cultures was investigated according to the traditional protocol for making Pecorino Crotonose (PC). Leuconostoc mesenteroides subsp. mesenteroides 2A, Lactobacillus casei 23C and Lactobacillus plantarum 18C (Autochthonous Starter, AS1) and Leuc. mesenteroides subsp. mesenteroides 2A, and L. casei 25D and 16A (AS2) were isolated and identified from aged ewes' milk PC cheeses, selected based on several enzymatic activities, and used as starter cultures. As shown by the in vitro kinetic of acidification, selected starter cultures had suitable capabilities to acidify. The manufacture of PC cheeses was carried out at an industrial plant scale. A control cheese (CC) was also made, using commercial starters consisting of mesophilic and thermophilic species. Ripening lasted 105 days at 10 °C. A poly-phasic approach was used to compare cheeses during manufacture and ripening, mainly based on pyrosequencing of the 16S rRNA targeting DNA, proteolysis and volatile component analyses. Compared to CC, both autochthonous starter cultures slightly affected the gross chemical composition of PC cheese. The cell density of thermophilic starters of CC progressively decreased throughout ripening. Plate count and RAPD-PCR showed that the cell number of autochthonous lactobacilli cultures of PC cheeses, made with AS1 and AS2, was almost constant throughout ripening and abundantly higher than that observed in CC. As shown by culture-independent analysis, the OTUs found during ripening varied depending on the manufacture with or without autochthonous starter cultures. The major chemical differences among cheeses were the concentration of free amino acids and the synthesis of some key volatile components (e.g., 2-methyl-1-propanol, 2-methyl-1-butanol, isobutyric, isovaleric, and isocaproic acids). Compared to CC, the use of AS1 positively affected the overall cheese quality.