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Application of temperature gradient gel electrophoresis to the study of yeast diversity in the estuary of the Tagus river,Portugal
Institution:1. Centro de Química Estrutural, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, 1049-001 Lisboa, Portugal;2. IPMA-Instituto Português do Mar e Atmosfera, Rua Alfredo Magalhães Ramalho, 6, 1495-006 Lisboa, Portugal;3. Environment and Climate Change Canada, Aquatic Contaminants Research Division, Water Science and Technology Directorate, Montréal, QC, Canada;4. Acadia University, Department of Earth and Environmental Science, K.C. Irving Environmental Science Center, Wolfville, NS, Canada;1. Instituto do Mar e da Atmosfera (IPMA), Rua Alfredo Magalhães Ramalho, 6, 1495-006 Algés, Lisboa, Portugal;2. MARE — Marine and Environmental Sciences Centre, Faculty of Sciences, University of Lisbon, Campo Grande, 1749-016 Lisbon, Portugal;3. Centro de Química Estrutural, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, Lisboa 1, 1049-001 Lisboa, Portugal;4. Water Quality Centre, Trent University, 1600 West Bank Drive, Peterborough, ON K9J 0G2, Canada;5. Institute of Inorganic and Analytical Chemistry, University of Muenster, Schlossplatz 2, 48149 Munster, Germany
Abstract:Temperature gradient gel electrophoresis (TGGE) was employed for the assessment of yeast diversity in the estuary of the Tagus river (Portugal). The molecular detection of yeasts was carried out directly from water samples and, in parallel, a cultivation approach by means of an enrichment step was employed. A nested PCR was employed to obtain a fungal amplicon containing the D2 domain of the 26S rRNA gene. For identification the TGGE bands were extracted, re-amplified, and sequenced. Fourteen fungal taxa were detected and all except one were yeasts. Most yeast sequences corresponded to members of the Ascomycota and only three belonged to the Basidiomycota. Five yeasts (four ascomycetes and one basidiomycete) could not be identified to the species level due to the uniqueness of their sequences. The number of species detected after enrichment was higher than the number of taxa found using the direct detection method. This suggests that some yeast populations are present in densities that are below the detection threshold of the method. With respect to the analysis of the yeast community structure, our results indicate that the dominant populations belong to Debaryomyces hansenii, Rhodotorula mucilaginosa, Cryptococcus longus, and to an uncultured basidiomycetous yeast phylogenetically close to Cr. longus. The combined analysis of direct detection and cultivation approaches indicates a similar community structure at the two sampled sites since nine species were present at both localities.
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