Exploitation of a polyphasic PCR-DGGE approach forthe investigation of the microbiota of traditional Italian raw milk cheeses.

INTRODUCTION Since the last decade there has been an increasing demand for artisan cheeses manufactured with raw milk and traditional procedures. In raw milk cheeses, fermentation and ripening are carried out by a heterogeneous microbial community, which includes starter lactic acid bacteria (LAB), able to rapidly convert lactose into lactic acid, and non starter LAB (NSLAB) mainly contributing to the development of cheese flavour and aroma. Materials & methods Four traditional cheeses manufactured with local raw milk (Marche region, central Italy) were analysed for physico-chemical (pH, aw), compositional (NaCl, raw protein, fat content) and microbiological traits (occurrence of pathogens and their toxins, and viable counts of lactobacilli, lactococci and thermophilic streptococci). The composition of the LAB population was investigated with a poorly exploited polyphasic PCR-DGGE approach relying on the analysis of the bacterial DNA extracted either from the cheeses or the bulk cells harvested from all the dilution plates used for LAB viable counting. Results & discussion The results of traditional microbiological analyses fully complied with the European legislation [Regulation EC No. 2073/2005]. A high cheese diversity was revealed by physico-chemical, microbiological and PCR-DGGE analyses. As the fingerprints obtained with the two approaches were compared, the analysis of the cultivable communities allowed a significantly higher diversity than the direct approach to be disclosed, although in some cases, species not detected in the bulk cells were identified by analysing the DNA extracted directly from the cheeses. CONCLUSIONS The overall PCR-DGGE results clearly demonstrated that although either approach, taken singly, gives an incomplete picture of the real microbial diversity by introducing bias, their combination allows a greater pool of information to be collected.