Phenotypic and genetic diversity of enterococci isolated from Italian cheeses

Veneto Agricoltura-Istituto per la Qualità e le Tecnologie Agroalimentari, Thiene (VI), Italy.
J Dairy Res (Impact Factor: 1.6). 06/2001; 68(2):303-16.
Source: PubMed


In the present study, 124 enterococcal strains, isolated from traditional Italian cow, goat and buffalo cheeses, were characterized using phenotypic features and randomly amplified polymorphic DNA polymerase chain reaction (RAPD-PCR). The RAPD-PCR profiles obtained with four primers and five different amplification conditions were compared by numerical analysis and allowed an inter- and intraspecific differentiation of the isolates. Whole cell protein analysis by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was used as a reference method for species identification. The strains were identified as Enterococcus faecalis (82 strains), E. faecium (27 strains), E. durans (nine strains), E. gallinarum (four strains) and E. hirae (two strains). Species recognition by means of RAPD-PCR was in agreement with the SDS-PAGE results except for eight strains of E. faecium that clustered in separated groups. On the other hand, phenotypic identification based on carbohydrate fermentation profiles, using the rapid ID 32 STREP galleries, gave different results from SDS-PAGE in 12.1% of the cases. The majority of the strains had weak acidifying and proteolytic activities in milk. One E. faecium strain showed vanA (vancomycin resistance) genotype while four strains showed a beta-haemolytic reaction on human blood. Several strains showed antagonistic activity towards indicator strains of Listeria innocua, Clostridium tyrobutyricam and Propionibacterium freudenreichii subsp. shermanii.

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    • "RAPD-PCR was carried out using the primer M13 (5 -GAG GGT GGC GGT TCT-3 ) (Huey and Hall, 1989). PCR reactions and amplification conditions were performed as described by Andrighetto et al. (2001) and Suzzi et al. (2000). Rep-PCR analysis was carried out using the primer GTG 5 (5 -GTG GTG GTG GTG GTG-3 ) and amplification conditions as previously described by Gevers et al. (2001). "
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    • "Thus, a comparison of RAPD-PCR data obtained with a combination of several primers designed for either conserved or variable regions of bacterial genome makes it possible to get an idea on the interspecific diversity of S. thermophilus. This modified PCR program used for RAPD PCR was found to be more efficient than that described by Botina et al. (2007) and Andrighetto et al. (2001, 2002). On the basis of RAPD, no clear correlation can be deduced between RAPD clusters and folate production by isolates. "
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    • "). DNA was amplified using methods and amplification conditions described by Andrighetto et al. (2001) for primer M13 and Ben Omar et al. (2000) for primer Lb2. Rep-PCR was performed using the primer GTG5 (5 0 -GTG GTG GTG GTG GTG-3 0 ) and methods as previously described by Gevers et al. (2001). "
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