Population Dynamics and Yeast Diversity in Early Winemaking Stages without Sulfites Revealed by Three Complementary Approaches

Abstract

Nowadays, the use of sulfur dioxide (SO2) during the winemaking process is a controversial societal issue. In order to reduce its use, various alternatives are emerging, in particular bioprotection by adding yeasts, with different impacts on yeast microbiota in early winemaking stages. In this study, quantitative-PCR and metabarcoding high-throughput sequencing (HTS) were combined with MALDI-TOF-MS to monitor yeast population dynamic and diversity in the early stages of red winemaking process without sulfites and with bioprotection by Torulaspora delbrueckii and Metschnikowia pulcherrima addition. By using standard procedures for yeast protein extraction and a laboratory-specific database of wine yeasts, identification at species level of 95% of the isolates was successfully achieved by MALDI-TOF-MS, thus confirming that it is a promising method for wine yeast identification. The different approaches confirmed the implantation and the niche occupation of bioprotection leading to the decrease of fungal communities (HTS) and Hanseniaspora uvarum cultivable population (MALDI-TOF MS). Yeast and fungi diversity was impacted by stage of maceration and, to a lesser extent, by bioprotection and SO2, resulting in a modification of the nature and abundance of the operational taxonomic units (OTUs) diversity.

Yeast and Filamentous Fungi Microbial Communities in Organic Red Grape Juice: Effect of Vintage, Maturity Stage, SO2, and Bioprotection

“…One hypothesis to explain this result is the very high initial population level of indigenous Hanseniaspora spp. (>10 6 cells/ml) compared to our previous study in the 2018 vintage with merlot grapes from the Bordeaux area (10 4 cells/mL) (Windholtz et al, 2021a).…”

“…In a preliminary experiment on merlot, we reported on the occupation of the must by Torulaspora delbrueckii and Metschnikowia pulcherrima used as bioprotection, limiting the relative abundance of filamentous fungi, and in particular of Aureobasidium and Botrytis (Windholtz et al, 2021a). Our present data confirm that the use of these two species led to a decrease in the number of OTUs (Observed and Chao1), and especially in the numbers of OTUs of filamentous fungi that are systematically associated with a decline in grape must quality, such as Aspergillus spp.…”

“…Simonin et al (2018) showed the effectiveness of Torulaspora delbrueckii in colonizing the medium in an aligoté must, a white Burgundy grape variety. In a preliminary study, the use of two species as bioprotection agents was tested in a context of Bordeaux region: Torulaspora delbrueckii and Metschnikowia pulcherrima in merlot must (Windholtz et al, 2021a). Their implantation was confirmed as limiting the abundance of filamentous fungi.…”

Soil composition and rootstock genotype drive the root associated microbial communities in young grapevines

“…Each fresh isolate was smeared on MSP96 target polished steel BC plate and overlaid with 1 μl of 70% formic acid. Once dried at room temperature, samples were overlaid with MALDI matrix (1 μl, 10 mg/ml of α-cyano-4-hydroxycinnamic acid in 50% acetonitrile/2.5% trifluoroacetic acid) for crystallization ( Windholtz et al, 2021 ). Once dried, the target plate was submitted to MALDI-TOF-MS analysis using Microflex MALDI-TOF (Bruker Daltonik GmbH, Leipzig, Germany) bench-top mass spectrometer scanned with laser wavelength of 337 nm and acceleration voltage of 20 kV.…”

Fungal Diversity and Composition of the Continental Solar Saltern in Añana Salt Valley (Spain)

“…In fact, Metschnikowia has been recovered from grapevine phyllospheres, fruit flies, grapes, and wine ferments, as part of the resident microbiota of wineries and winemaking equipment [43], while Hanseniaspora has been isolated in grape must, and in the winery environment [44]. Both Candida and Taphrina genera have been described as being associated with the grape berry microbial community, albeit as a minor percentage [45]. The presence of winemaking-related fungal genera could be the result of contamination via agriculture, possibly due to the proximity of significant cereal-and wine-producing areas of La Rioja, which borders the Añana Salt Valley.…”