14A-36 |
Evaluation of methods for genus-level identification of Bifidobacterium species |
E. P. BRICZINSKI and R. F. Roberts. Dept. of Food Science, Pennsylvania State Univ., 121 Borland Lab., University Park, PA 16802-2504 Increasing application of probiotics in food highlights the need for reliable methods for probiotic identification. Common methods used to verify bacterial isolates as members of the genus Bifidobacterium, based on culture methods and cell morphology, are often unreliable. Other techniques to identify Bifidobacterium include assaying for fructose-6-phosphoketolase (F6PPK), PCR analysis of 16S rDNA and determination of organic acids in spent culture media. This study evaluated the presence of F6PPK, PCR amplification of a 16S rDNA sequence, growth on various media, production of organic acids, and cellular morphology as means of identifying Bifidobacterium strains. Seven Bifidobacterium type strains were obtained from ATCC and characterized for growth in media commonly used for bifidobacteria isolation and identification. In addition, the presence of a genus-specific sequence of 16S rDNA was assayed using PCR, organic acids produced were quantified by HPLC, cellular morphology was evaluated and presence of F6PPK was determined. The presence of F6PPK was considered definitive when assigning a strain to the genus Bifidobacterium. A collection of 30 commercial strains, identified as Bifidobacterium species by the supplier, were obtained and assayed as described above. All ATCC bifidobacteria possessed the F6PPK enzyme, the characteristic 16S rDNA sequence, and the ability to produce both lactic and acetic acids. Eleven of the commercial strains were not Bifidobacterium, based on both F6PPK and PCR results. Bifidobacteria could not be distinguished from non-bifid strains by morphology or growth on various media. However, production of acetic acid was detected only with Bifidobacterium-positive strains. The non-bifid strains produced lactic acid without acetic acid. The presence of F6PPK activity and PCR-based methods are reliable for genus-level identification of bifidobacteria. However, these methods are complicated and time-consuming. Production of acetic and lactic acids represents a simple, rapid, qualitative means of identifying Bifidobacterium commercial samples.
Session 14A, Dairy Foods: General developments in dairy technology I
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