Characterization of non-starter lactic acid bacteria from Italian ewe cheeses based on phenotypic, genotypic, and cell wall protein analyses

Non-starter lactic acid bacteria (NSLAB) were isolated from 12 Italian ewe cheeses representing six different types of cheese, which in several cases were produced by different manufacturers. A total of 400 presumptive Lactob acillus isolates mere obtained, and 123 isolates and 10 type strains were s ubjected to phenotypic, genetic, and cell wall protein characterization ana lyses. Phenotypically, the cheese isolates included 32% Lactobacillus plant arum isolates, 15% L, brevis isolates, 12% L, paracasei subsp, paracasei is olates, 9% L. curvatus isolates, 6% L, fermentum isolates, 6% L. casei subs p, casei isolates, 5% L, pentosus isolates, 3% L. casei subsp, pseudoplanta rum isolates, and 1% L, rhamnosus isolates, Eleven percent of the isolates mere not phenotypically identified, Although a randomly amplified polymorph ic DNA (RAPD) analysis based on three primers and clustering by the unweigh ted pair group method with arithmetic average (UPGMA) was useful for partia lly differentiating the 10 type strains, it did not provide a species-speci fic DNA band or a combination of bands which permitted complete separation of all the species considered. In contrast, sodium dodecyl sulfate-polyacry lamide gel electrophoresis cell wall protein profiles clustered by UPGMA we re species specific and resolved the NSLAB. The only exceptions were isolat es phenotypically identified as L, plantarum and L, pentosus or as L. casei subsp, casei and L, paracasei subsp, paracasei, which were grouped togethe r. Based on protein profiles, Italian ewe cheeses frequently contained four different species and 3 to 16 strains, In general, the cheeses produced fr om raw ewe milk contained a larger number of more diverse strains than the cheeses produced from pasteurized milk, The same cheese produced in differe nt factories contained different species, as well as strains that belonged to the same species but grouped in different RAPD clusters.