Comparison of RAPDs, AFLPs and SSR markers for the genetic analysis of yeast strains of Saccharomyces cerevisiae

We evaluated the usefulness of different molecular techniques for the genetic analysis of Saccharomyces cerevisiae strains. Three commonly used PCR-derived genetic methods, random amplified polymorphic DNA (RAPDs), amplified fragment length polymorphism (AFLPs) and simple sequence repeats (SSRs; microsatellites), were used to characterize 27 wine yeast strains of S. cerevisiae from the “Denominación de Origen Vinos de Madrid” (Spain). Using these methods, we were able to overcome certain limitations associated with classical taxonomic methods. Based on the presence or absence of amplified fragments for each genotype, AFLPs and SSRs showed a similar discriminatory power superior to that of the RAPDs. Genetic relationships between strains were also estimated using the three methods. In general, very poor correlations were found, reflecting the different genomic regions for which the methods are screened. Results are discussed in terms of which molecular technique is most appropriate for use with a particular aspect of genetic evaluation.     

Microbiological evaluation and molecular characterization of bifidobacteria strains in commercial fermented milks

The aim of this study was to evaluate the presence of Bifidobacterium animalis subsp. lactis in commercial dairy products using different molecular techniques. We analyzed the microbiological composition of 13 commercial fermented milks available in the Spanish market. Thirteen strains of genus Bifidobacterium were isolated from these products and were identified by genus-specific PCR, by fluorescence in situ hybridization (FISH), by multiplex PCR and amplified ribosomal DNA restriction analysis (ARDRA). The same sets of strains were typed by randomly amplified polymorphic DNA (RAPD) analysis and by amplified fragment length polymorphism technique (AFLP). All strains were identified as B. animalis subsp. lactis using ARDRA and multiplex PCR techniques. Similarity between strains was evaluated based on RAPD and AFLP profiles. The isolated strains showed similar profiles by using these techniques, revealing the reduced genetic variability existing among commercial strains, and all these profiles were reproducible in repeated analysis. ARDRA and multiplex PCR are techniques that allow differentiation of the bifidobacteria at genus and species level, but do not indicate if they are different strains, for which reason the RAPD technique is very useful. All bifidobacteria isolated from commercial fermented milks in Spain belong to the same species B. animalis subsp. lactis. Our results demonstrate the necessity to control the presence of bifidobacteria in commercial fermented milks, not only at species level but also at strain level. Multiplex PCR and RAPDs are the most suitable, rapid and precise techniques to identify all bifidobacteria contained in fermented milk products at genus-, species-, and strain levels.