Evaluation of microbial diversity during the manufacture of Fior di Latte di Agerola, a traditional raw milk pasta-filata cheese of the Naples area.

Microbial diversity of the raw milk for the production of Fior di Latte di Agerola and its changes during cheesemaking were studied. Viable counts showed that at the end of curd ripening, loads of lactic acid bacteria, both mesophilic and thermophilic rods and cocci, higher than those commonly evidenced in similar cheeses produced by using natural or commercial starters, were detected. Identification of 272 isolates, supported by molecular diagnostic aids, evidenced representative cultures of a high number of bacterial taxa of interest as participating in the process, although most of the isolates belonged to Lactococcus lactis and Lactobacillus helveticus species. RAPD-PCR and REA-PFGE biotyping were performed for the isolates of the above species and it was shown that most of the strains isolated from the raw milk occurred during the whole cheesemaking process, and an active role of these strains in the fermentation was supposed. The results offer further proof of the importance of the raw milk as source of technologically interesting strains of lactic acid bacteria capable of driving the fermentation of traditional cheeses.

16S-23S rDNA intergenic spacer region polymorphism of Lactococcus garvieae, Lactococcus raffinolactis and Lactococcus lactis as revealed by PCR and nucleotide sequence analysis.

The intergenic spacer region (ISR) between the 16S and 23S rRNA genes was tested as a tool for differentiating lactococci commonly isolated in a dairy environment. 17 reference strains, representing 11 different species belonging to the genera Lactococcus, Streptococcus, Lactobacillus, Enterococcus and Leuconostoc, and 127 wild streptococcal strains isolated during the whole fermentation process of "Fior di Latte" cheese were analyzed. After 16S-23S rDNA ISR amplification by PCR, species or genus-specific patterns were obtained for most of the reference strains tested. Moreover, results obtained after nucleotide analysis show that the 16S-23S rDNA ISR sequences vary greatly, in size and sequence, among Lactococcus garvieae, Lactococcus raffinolactis, Lactococcus lactis as well as other streptococci from dairy environments. Because of the high degree of inter-specific polymorphism observed, 16S-23S rDNA ISR can be considered a good potential target for selecting species-specific molecular assays, such as PCR primer or probes, for a rapid and extremely reliable differentiation of dairy lactococcal isolates.