Resident lactic acid bacteria in raw milk Canestrato Pugliese cheese

Aims: Investigation of the autochthonous lactic acid bacteria (LAB) population of the raw milk Protected Designation of Origin Canestrato Pugliese cheese using phenotypic and genotypic methodologies. Methods and Results: Thirty phenotypic assays and three molecular techniques (restriction fragment length polymorphism, partial sequencing of the 16S rRNA gene and recA multiplex PCR assay) were applied to the identification of 304 isolates from raw milk Canestrato Pugliese cheese. As a result, 168 of 207 isolates identified were ascribed to genus Enterococcus, 25 to Lactobacillus, 13 to Lactococcus and one to Leuconostoc. More in details, among the lactobacilli, the species Lactobacillus brevis and Lactobacillus plantarum were predominant, including 13 and 10 isolates respectively, whereas among the lactococci, Lactococcus cremoris subsp. cremoris was the species more frequently detected (seven isolates). Conclusions: Except for the enterococci, phenotypic tests were not reliable enough for the identification of the isolates, if not combined to the genotype-based molecular techniques. The polyphasic approach utilized allowed 10 different LAB species to be detected; thus suggesting the appreciable LAB diversity of the autochthonous microbial population of the Canestrato Pugliese cheese. Significance and Impact of the Study: A comprehensive study of the resident raw milk Canestrato Pugliese cheese microbial population has been undertaken.