Abstract
Mozzarella (M), Grana Padano (GP) and Parmigiano Reggiano (PR) are three of the most important traditional Italian cheeses. In the three cheese manufactures the initial fermentation is carried out by adding natural whey cultures (NWCs) according to a back-slopping procedure. In this study, NWCs and the corresponding curds from M, GP and PR manufactures were analyzed by culture-independent pyrosequencing of the amplified V1–V3 regions of the 16S rRNA gene, in order to provide insights into the microbiota involved in the curd acidification. Moreover, culture-independent high-throughput sequencing of lacS gene amplicons was carried out to evaluate the biodiversity occurring within the S. thermophilus species. Beta diversity analysis showed a species-based differentiation between GP-PR and M manufactures indicating differences between the preparations. Nevertheless, all the samples shared a naturally-selected core microbiome, that is involved in the curd acidification. Type-level variability within S. thermophilus species was also found and twenty-eight lacS gene sequence types were identified. Although lacS gene did not prove variable enough within S. thermophilus species to be used for quantitative biotype monitoring, the possibility of using non rRNA targets for quantitative biotype identification in food was highlighted.
Highlights
Mozzarella (M), Grana Padano (GP) and Parmigiano Reggiano (PR) are three of the most important traditional Italian cheeses
Samples from M manufactures were characterized by abundance of S. thermophilus, while Lb. delbruekii and Lb. helveticus were present at lower extent compared to GP and PR samples (Figure 1)
Methods allowing direct DNA analysis from these environments are valuable in order to avoid biases of the culture-dependent approach. 16S rRNA pyrosequencing has been recently applied to analysis of microbiota in dairy matrices [3,35,36,37] and it is regarded as an extremely sensitive tool, able to reveal subdominant OTUs [38]
Summary
Mozzarella (M), Grana Padano (GP) and Parmigiano Reggiano (PR) are three of the most important traditional Italian cheeses They are all protected designation of origin (PDO) cheeses and the technology of manufacture, as well as the microbiota involved, have been described in previous works [1,2,3,4,5,6]. Streptococcus thermophilus is one of the most important bacteria in the dairy industry and it occurs in NWCs used in the manufacture of several traditional cheeses [3,4,5,8,16,17] Monitoring of this species at biotype level is an important target of the dairy industry and several molecular methods can be used [18,19,20,21,22]. The 250 bp variable region upstream from the lacS gene showed a high level of heterogeneity in a previous study [23], making it a good candidate for sequencing-based biotype monitoring of S. thermophilus
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