[go: up one dir, main page]
More Web Proxy on the site http://driver.im/ Skip to main content

Advertisement

Springer Nature Link
Log in

RFLP analysis of the ribosomal internal transcribed spacers and the 5.8S rRNA gene region of the genus Saccharomyces: a fast method for species identification and the differentiation of flor yeasts

  • Published:
Antonie van Leeuwenhoek Aims and scope Submit manuscript

Abstract

The PCR amplification and subsequent restriction analysis of the region spanning the internal transcribed spacers (ITS1 and ITS2) and the 5.8S rRNA gene was applied to the identification of yeasts belonging to the genus Saccharomyces. This methodology has previously been used for the identification of some species of this genus, but in the present work, this application was extended to the identification of new accepted Saccharomyces species (S. kunashirensis, S. martiniae, S. rosinii, S. spencerorum, and S. transvaalensis), as well as to the differentiation of an interesting group of Saccharomyces cerevisiae strains, known as flor yeasts, which are responsible for ageing sherry wine. Among the species of the Saccharomyces sensu lato complex, the high diversity observed, either in the length of the amplified region (ranged between 700 and 875 bp) or in their restriction patterns allows the unequivocal identification of these species. With respect to the four sibling species of the Saccharomyces sensu stricto complex, only two of them, S. bayanus and S. pastorianus, cannot be differentiated according to their restriction patterns, which is in accordance with the hybrid origin (S. bayanus × S. cerevisiae) of S. pastorianus. The flor S. cerevisiae strains exhibited restriction patterns different from those typical of the species S. cerevisiae. These differences can easily be used to differentiate this interesting group of strains. We demonstrate that the specific patterns exhibited by flor yeasts are due to the presence of a 24-bp deletion located in the ITS1 region and that this could have originated as a consequence of a slipped-strand mispairing during replication or be due to an unequal crossing-over. A subsequent restriction analysis of this region from more than 150 flor strains indicated that this deletion is fixed in flor yeast populations.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
£29.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price includes VAT (United Kingdom)

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Barnett, JA (1992) The taxonomy of the genus Saccharomyces Meyen ex Ress: a short review for non-taxonomists. Yeast 8: 1–23

    Google Scholar 

  • Barnett JA, Payne RW & Yarrow D (1990) Yeast: Characteristic and Identification, 2nd edn. Cambridge University Press, Cambridge, England

    Google Scholar 

  • Cai J, Roberts IN & Collins MD (1996) Phylogenetic relationships among members of the ascomycetous yeast genera Brettanomyces, Debaryomyces, Dekkera and Kluyveromyces deduced by small subunit rRNA gene sequences. Int. J. Syst. Bacteriol. 46: 542–549

    Google Scholar 

  • Clemons KV, McCusker JH, Davis RW & Stevens DA (1994) Comparative pathogenesis of clinical and nonclinical isolates of Saccharomyces cerevisiae. J. Infect. Dis. 169: 859–867

    Google Scholar 

  • de Barros Lopes M, Soden A, Martens AL, Henschke PA & Langridge P (1998) Differentiation and species identification of yeasts using PCR. Int. J. Syst. Bacteriol. 48: 279–286

    Google Scholar 

  • Esteve-Zarzoso B, Belloch C, Uruburu F & Querol A (1999) Identification of yeasts by RFLP analysis of the 5.8S rRNA gene and the two ribosomal internal transcribed spacers. Int. J. Syst. Bacteriol. 49: 329–337

    Google Scholar 

  • Felsenstein J (1993) PHYLIP: phylogeny inference package, v. 3.5c. University of Washington, Seattle, Washington

    Google Scholar 

  • Fitch WM (1971) Toward defining the course of evolution: minimum change for a specified tree topology. Syst. Zool. 20: 406–416

    Google Scholar 

  • Goddard MR & Burt A (1999) Recurrent invasion and extinction of a selfish gene. Proc. Natl. Acad. Sci. USA 96: 13880–13885

    Google Scholar 

  • Guido CG & Carnacini A (1996) Volatile composition of Vernaccia de Oristano sherry-like wine as affected by biological ageing. J. Sci. Food Agric. 70: 44–50

    Google Scholar 

  • Guijo S, Mauricio JC, Salmon JM & Ortega JM (1997) Determination of the relative ploidy in different Saccharomyces cerevisiae strains used for fermentation and flor film ageing of dry sherrytype wines. Yeast 13: 101–117

    Google Scholar 

  • Guillamón JM, Sabaté J, Barrio E, Cano J & Querol A (1998) Rapid identification of wine yeast species based on RFLP analysis of the ribosomal internal transcribed spacer (ITS) region. Arch. Microbiol. 169: 387–392

    Google Scholar 

  • Hopfer RL, Walden P, Setterquist S & Highsmith WE (1993) Detection and differentiation of fungi in clinical specimens using polymerase chain reaction (PCR) amplification and restriction enzyme analysis. J. Med. Vet. Mycol. 31: 65–75

    Google Scholar 

  • Huffman JL, Molina FI & Jong S-C (1992) Authentication of ATCC strains in the Saccharomyces cerevisiae complex by PCR fingerprinting. Exp. Mycol. 16: 316–319

    Google Scholar 

  • Ibeas JI, Lozano I, Perdigones F & Jimenez J (1997) Dynamics of flor yeast populations during the biological aging of sherry wines. Am. J. Enol. Vitic. 48: 75–79

    Google Scholar 

  • Ibeas JI & Jimenez J (1997) Mitochondrial DNA loss caused by ethanol in Saccharomyces flor yeasts. Appl. Environ. Microbiol. 63: 7–12

    Google Scholar 

  • James SA, Collins MD & Roberts IN (1996) Use of an rRNA internal transcribed spacer region to distinguish phylogenetically closely related species of the genera Zygosaccharomyces and Torulaspora. Int. J. Syst. Bacteriol. 46: 189–194

    Google Scholar 

  • James SA, Cai J, Roberts IN & Collins MD (1997) A phylogenetic analysis of the genus Saccharomyces based on 18S rRNA gene sequences: description of Saccharomyces kunashirensis sp. nov. and Saccharomyces martiniae sp. nov. Int. J. Syst. Bacteriol. 47: 453–460

    Google Scholar 

  • Kreger-van Rij NJW (1984) The yeasts: A taxonomic Study, 3rd ed. Elsevier Science Publisher B.V., Amsterdam, The Netherlands

    Google Scholar 

  • Kurtzman CP (1992) rRNA sequence comparisons for assessing phylogenetic relationships among yeasts. Int. J. Syst. Bacteriol. 42: 1–6

    Google Scholar 

  • Kurtzman CP (1993) Systematics of the ascomycetous yeasts assessed from ribosomal RNA sequence divergence. Antonie van Leeuwenhoek 63: 165–174

    Google Scholar 

  • Kurtzman CP & Robnett CJ (1998) Identification and phylogeny of ascomycetous yeasts from analysis of nuclear large subunit (26S) ribosomal DNA partial sequences. Antonie van Leeuwenhoek 73: 331–371

    Google Scholar 

  • Li WH (1997) Molecular evolution, Sinauer Associates, Sunderland, MA

    Google Scholar 

  • Martínez P, Codón AC, Pérez L & Benítez T (1995) Physiological and molecular characterization of flor yeasts: polymorphism of flor yeast populations. Yeast 11: 1399–1411

    Google Scholar 

  • Martínez P, Rodríguez P & Benítez T (1997) Velum formation by flor yeasts isolated from sherry wine. Am. J. Enol. Vitic. 48: 55–62

    Google Scholar 

  • McCullough MJ, Clemons KV, McCusker JH & Stevens DA (1998) Intergenic transcribed spacer PCR ribotyping for differentiation of Saccharomyces species and interspecific hybrids. J. Clin. Microbiol. 36: 1035–1038

    Google Scholar 

  • McCusker JH, Clemons KV, Stevens DA & Davis RW (1994) Saccharomyces cerevisiae virulence phenotype as determined with CD-1 mice is associated with the ability to grow at 42 °C and form pseudohyphae. Infect. Immun. 62: 5447–5455

    Google Scholar 

  • Messner R & Prillinger H (1995) Saccharomyces species assignment by long range ribotyping. Antonie van Leeuwenhoek 67: 363–370

    Google Scholar 

  • Molina FI, Inoue T & Jong S-C (1992) Restriction polymorphisms in the internal transcribed spacers and 5.8S rDNA of Saccharomyces. Curr. Microbiol. 25: 251–255

    Google Scholar 

  • Molina FI, Jong S-C & Huffman JL (1993) PCR amplification of the 3' external transcribed and intergenic spacer of the ribosomal DNA repeat unit in three species of Saccharomyces. FEMS Microbiol. Lett. 108: 259–264

    Google Scholar 

  • Molnár O, Messner R, Prillinger H, Stahl U & Slavikova E (1995a) Genotypic identification of Saccharomyces species using random amplified polymorphic DNA analysis. Syst. Appl.Microbiol. 18: 136–145

    Google Scholar 

  • Molnár O, Messner R, Prillinger H, Scheide K, Stahl U, Silberhumer H, Wunderer W (1995b) Genotypische identifizierung von Saccharomyces-arten aus der getränkeindustrie mit hilfe der zufallsprimer-abhängigen polymerase-kettenreaktion (RAPD-PCR). Mitt. Klosterneuburg 45: 113–122

    Google Scholar 

  • Montrocher R, Verner M-C, Briolay J, Gautier C & Marmeisse R (1998) Phylogenetic analysis of the Saccharomyces cerevisiae group based on polymorphisms of rDNA spacer sequences. Int. J. Syst. Bacteriol. 48: 295–303

    Google Scholar 

  • Naumov GI, Naumova ES & Gaillardin C (1993) Genetic and karyotypic identification of wine Saccharomyces bayanus yeasts isolated in France and Italy. Syst. Appl. Microbiol. 16: 274–279

    Google Scholar 

  • Oda Y, Yabuki M, Tonomura K & Fukunaga M (1999) Sequence analysis of 18S-28S rRNA spacer regions from Saccharomyces kunashirensis, S. martiniae, S. rosinii and S. transvaalensis. Curr. Microbiol. 38: 61–63

    Google Scholar 

  • Querol A, Barrio E, Huerta T & Ramón D (1992) Molecular monitoring of wine fermentations conducted by active dry yeast strains. Appl. Environ. Microbiol. 58: 2948–2953

    Google Scholar 

  • Redecker D, Thierfelder H, Walker C & Werner D (1997) Restriction analysis of PCR-amplified internal transcribed spacers of ribosomal DNA as a tool for species identification in different genera of the order Glomales. Appl. Environ. Microbiol. 63: 1756–1761

    Google Scholar 

  • Rosini G, Federici F, Vaughan AE & Martini A. (1982) Systematics of the species of the yeast genus Saccharomyces associated with the fermentation industry. Eur. J. Appl. Microbiol. Biotechnol. 15: 188–193

    Google Scholar 

  • Smole Mozina S, Dlauchy D, Deak T & Raspor P (1997) Identification of Saccharomyces sensu stricto and Torulaspora yeasts by PCR ribotyping. Lett. Appl. Microbiol. 24: 311–315

    Google Scholar 

  • Santa María J (1959) Oxidación de alcohol etílico a ácido acético por levaduras vívas. I. Saccharomyces aceti nov. spec. y Saccharomyces oxidans, nov. spec., nuevas especies aisladas de vino. An. Inst. Nac. Invest. Agron. 8: 713–735

    Google Scholar 

  • Santa María J (1970) Saccharomyces gaditensis y Saccharomyces cordubensis, dos nuevas especies de levaduras de ‘flor’. Bol. Inst. Nac. Invest. Agron. 62: 57–66

    Google Scholar 

  • Tamai, Y, Momma, T, Yoshimoto, H and Kaneko, Y (1998) Coexistence of two types of chromosome in the bottom fermenting yeast, S. pastorianus. Yeast 14: 923–933

    Google Scholar 

  • Thompson JD, Higgins DG & Gibson TJ (1994) Clustal W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, positions-specific gap penalties and weight matrix choice. Nucl. Acid Res. 22: 4673–4680

    Google Scholar 

  • Valente P, Gouveia FC, de Lemos GA, Pimentel D, van Elsas JD, Mendoça-Hagler LC & Hagler AN (1996) PCR amplification for differentiation of Saccharomyces cultures. FEMS Microbiol. Lett. 137: 253–256

    Google Scholar 

  • van der Walt JP (1970) Saccharomyces Meyen emend. Reess. In: Lodder E (Ed.) The Yeast: A Taxonomic Study, 2nd edn. (pp 555–718). Elsevier, Amsterdam

    Google Scholar 

  • Vaughan-Martini A (1995) Saccharomyces barnettii and Saccharomyces spencerorum: two new species of Saccharomyces sensu lato (van der Walt). Antonie van Leeuwenhoek 68: 111–118

    Google Scholar 

  • Vaughan-Martini A & Martini A (1987) Three newly delimited species of Saccharomyces sensu stricto. Antonie van Leeuwenhoek 53: 77–84

    Google Scholar 

  • Vaughan-Martini A & Martini A (1989) A proposal for correct nomenclature of the domesticated species of the genus Saccharomyces. In: Cantarelli C & Lanzarini G (Eds) Biotechnology Applications in Beverage Production (pp 1–16). Elsevier Applied Science, Barking

    Google Scholar 

  • Vaughan-Martini A & Martini A (1993) A taxonomic key for the genus Saccharomyces. Syst. Appl. Microbiol. 16: 113–119

    Google Scholar 

  • Vaughan-Martini A & Martini A (1998) Saccharomyces Meyen ex Reess. In: Kurtzman CP & Fell JW (Eds) The Yeasts: A Taxonomic Study, 4th edn. (pp 358–371) Elsevier, Amsterdam

    Google Scholar 

  • Vaughan-Martini A, Martini A & Cardinali G (1993) Electrophoretic karyotyping as a taxonomic tool in the genus Saccharomyces. Antonie van Leeuwenhoek 63: 145–156

    Google Scholar 

  • White TJ, Bruns T, Lee E & Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innis MA, Gelfand DH, Sninsky JJ & White TJ (Eds) PCR Protocols: A Guide to Methods and Applications (pp 315–322). Academic Press, San Diego

    Google Scholar 

  • Wyder MT & Puhan Z (1997) A rapid method for identification of yeasts from Kefyr at species level. Milchwissenschaft. 52: 327–330

    Google Scholar 

  • Yamagishi, H. and Ogata, T. (1999). Chromosomal structures of bottom fermenting yeasts. Syst. Appl. Microbiol. 22: 341–353

    Google Scholar 

  • Yeh L-CC & Lee JC (1991) Higher-order structure of the 5.8S rRNA sequence within the yeasts 355 precursor ribosomal RNA syntetized in vitro. J. Mol. Biol. 217: 649–659

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Departamento de Biotecnología, Instituto de Agroquímica y Tecnología de Alimentos (CSIC), PO Box 73, 46100, Burjassot, València, Spain

    M. Teresa Fernández-Espinar, Braulio Esteve-Zarzoso & Amparo Querol

  2. Colección Española de Cultivos Tipo, Universitat de València, Edificio de Investigación, Dr. Moliner, 50, 46100, Burjassot, València, Spain

    Braulio Esteve-Zarzoso

  3. Unitat de Genètica Evolutiva, Institut "Cavanilles" de Biodiversitat i Biologia Evolutiva, Universitat de València, Edificio de Institutos, Campus de Paterna, PO Box 2085, 46071, València, Spain

    Eladio Barrio

Authors
  1. M. Teresa Fernández-Espinar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Braulio Esteve-Zarzoso
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Amparo Querol
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Eladio Barrio
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Eladio Barrio.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Teresa Fernández-Espinar, M., Esteve-Zarzoso, B., Querol, A. et al. RFLP analysis of the ribosomal internal transcribed spacers and the 5.8S rRNA gene region of the genus Saccharomyces: a fast method for species identification and the differentiation of flor yeasts. Antonie Van Leeuwenhoek 78, 87–97 (2000). https://doi.org/10.1023/A:1002741800609

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1002741800609

Search

Navigation