Yeast Diversity and Persistence in Botrytis-Affected Wine Fermentations

Abstract

Culture-dependent and -independent methods were used to examine the yeast diversity present in botrytisaffected ("botrytized") wine fermentations carried out at high (ϳ30°C) and ambient (ϳ20°C) temperatures. Fermentations at both temperatures possessed similar populations of Saccharomyces, Hanseniaspora, Pichia, Metschnikowia, Kluyveromyces, and Candida species. However, higher populations of non-Saccharomyces yeasts persisted in ambient-temperature fermentations, with Candida and, to a lesser extent, Kluyveromyces species remaining long after the fermentation was dominated by Saccharomyces. In general, denaturing gradient gel electrophoresis profiles of yeast ribosomal DNA or rRNA amplified from the fermentation samples correlated well with the plating data. The direct molecular methods also revealed a Hanseniaspora osmophila population not identified in the plating analysis. rRNA analysis also indicated a large population (>10 6 cells per ml) of a nonculturable Candida strain in the high-temperature fermentation. Monoculture analysis of the Candida isolate indicated an extreme fructophilic phenotype and correlated with an increased glucose/fructose ratio in fermentations containing higher populations of Candida. Analysis of wine fermentation microbial ecology by using both culture-dependent and -independent methods reveals the complexity of yeast interactions enriched during spontaneous fermentations.

Yeast populations associated with Ghanaian cocoa fermentations analysed using denaturing gradient gel electrophoresis (DGGE)

“…A fragment most closely related to the recently described wine-associated yeast Candida zemplinina (Sipiczki, 2003) was only detected using DGGE ( Figure 1, band 7; Table 1). Candida zemplinina has been proposed to form a viable but not culturable (VBNC) population during wine fermentations (Mills et al, 2002). It is possible that C. zemplinina was not detected using culture-based methods because it constitutes a VBNC population during cocoa fermentations.…”

“…The ability to detect VBNC organisms is one of the advantages of the DGGE technique (Muyzer and Smalla, 1998). Cocoa and grape juice both constitute an environment characterized by a relatively low pH and high sugar content, possibly offering an ecological niche for C. zemplinina (Mills et al, 2002;Sipiczki, 2003). Candida stellimalicola was only detected by DGGE after 3 h of fermentation, and by DGGE, as well as by culture-based isolation after 24 h ( Figure 1, band 4; Table 1).…”

“…in the intestinal colon (Tannock, 1999;Nielsen et al, 2003) and soil (Muyzer et al, 1993;Muyzer and Smalla, 1998). During recent years the method has also been applied to investigate yeast populations in milk (Cocolin et al, 2002a), sourdough (Meroth et al, 2003;Gatto and Torriani, 2004) and during coffee (Masoud et al, 2004) and wine fermentations (Cocolin et al, 2000(Cocolin et al, , 2001(Cocolin et al, , 2002bHernán-Gómez et al, 2000;Mills et al, 2002;Cocolin and Mills, 2003;Prakitchaiwattana et al, 2004).…”

Isolation and Characterization of New Metschnikowia pulcherrima Strains as Producers of the Antimicrobial Pigment Pulcherrimin

“…The natural habitats of M. pulcherrima strains are fully matured fruits, especially grapes (Mills et al, 2002). M. pulcherrima produces pulcherrimin, which is a red pigment that accumulates in the growth medium and, clearly visible, around the M. pulcherrima colonies on plates (Kluyver et al, 1953).…”

PCR and DGGE Detection Limits for Wine Spoilage Microbes

“…Genetic fingerprinting of complex microbial populations (Muyzer, 1999) is currently used extensively to study the microbial ecology of wine fer mentations (Cocolin et al, 2000;Mills et al, 2002;Renouf et al, 2007). Polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) analysis is also used, since it allows the detection and identification of individual species, as well as the overall profiling of microbial populations (Stahl & Chapman, 1994).…”