Investigating the influence of a wine yeast consortium on population dynamics, alcoholic and malolactic fermentation

Janse van Rensburg, Philippe Jacques (2018-12)

Thesis (MSc)--Stellenbosch University, 2018.

Thesis

ENGLISH ABSTRACT: Non-Saccharomyces (NS) starter cultures of species such as Metschnikowia pulcherrima, Torulaspora delbrueckii, Pichia kluyveri and Lachancea thermotolerans, have received attention for their desirable properties and contributions in winemaking. These include the production of volatile compounds or enzymes to modify organoleptic attributes. Most studies have focussed on the use of single NS at a time with Saccharomyces cerevisiae to study the above contributions and properties; including effects on malolactic fermentation (MLF). However, there is growing interest in using complex multi-starter cultures to enhance wine aroma. Indeed, commercial products such as Anchor Alchemy II (Anchor Yeast), which comprises of different strains of S. cerevisiae, and Melody™ (CHR Hansen), comprising of S. cerevisiae, L. thermotolerans and T. delbrueckii in different ratios, are available. To develop such products, the understanding of genetic and phenotypic characteristics of strains and interactions amongst the different organisms is important. Consequently, some studies have investigated yeast-yeast interactions and their mechanisms, such as antagonistic interactions mediated by direct cell contact or through growth inhibitory metabolites. This, results in a decline of certain non-Saccharomyces like Hanseniaspora and therefore effect the final aroma composition. Conversely, synergistic effects can be observed in which species may be promoted to persist longer and therefore they contribute more to aroma. However, knowledge on population dynamics in multi-species starter cultures and their influence on alcoholic fermentation (AF) and MLF remains limited. The current study investigated a multi-species yeast consortium during AF and its effects on Oenococcus oeni viability during MLF. The consortium comprised of M. pulcherrima, L. thermotolerans, T. delbrueckii and S. cerevisiae. Fermentations were conducted in Chenin blanc and Pinotage at 15°C and 25°C, respectively. In all trials M. pulcherrima declined rapidly, while L. thermotolerans persisted until mid-fermentation. The best growth was observed for T. delbrueckii and it was able to persist until late fermentation stages. Fermentations that contained L. thermotolerans produced L-lactic acid in the Pinotage, but not in the Chenin blanc. There were no negative impacts observed on O. oeni populations during MLF for Pinotage and Chenin blanc wines. MLF kinetics were similar in all the Pinotage wines. In the Chenin blanc, the fastest L- malic acid consumption was displayed in wines that were fermented by L. thermotolerans and T. delbrueckii co-inoculations with S. cerevisiae. Different chemical profiles were detected using attenuated total reflection infrared (ATR-IR) spectroscopy. Mostly Chenin blanc wines were found to be significantly different from S. cerevisiae controls. Using gas chromatography, fold changes were observed for many volatile compounds. In conclusion, it is possible to predict a consortium population dynamic based on individual yeast performances in mixed fermentations. The volatile profiles are not additive between treatments and will be unique for each inoculation scheme. MLF seems to not be detrimentally affected by a consortium so long as each strain is regarded as compatible with lactic acid bacteria. Future work should include the evaluation of more yeast species and at alternate inoculation levels. Additionally, the inclusion of Lactobacillus plantarum for MLF needs to be investigated and quantification of detected volatiles should be performed.

AFRIKAANSE OPSOMMING: Nie-Saccharomyces (NS) aanvangskulture van spesies soos Metschnikowia pulcherrima, Torulaspora delbrueckii, Pichia kluyveri en Lachancea thermotolerans, het aansienlike aandag ontvang as gevolg van hulle eienskappe vir wynbereiding. Dit sluit die bydrae tot wynaroma in deur die produksie van vlugtige verbindings soos esters, hoër alkohole en vetsure of die vrystelling van terpene tot hul aktiewe sensoriese status. Verder is die verlaging van alkohol of asynsuur asook die toename in gliserol of melksuur vlakke van belang. Die meeste studies het voorheen gefokus op die gebruik van enkel NS, soos T. delbrueckii of M. pulcherrima, wat agtermekaar of mede ingeënt is met Saccharomyces cerevisiae om bogenoemde bydraes en eienskappe te bestudeer; insluitend die effek op appelmelksuurgisting (AMG). Daar is egter toenemende belangstelling in die gebruik van komplekse multi-spesies aanvangskulture om wynaroma te verbeter. Kommersiële produkte soos Anchor Alchemy II (Anchor Yeast), wat bestaan uit verskillende stamme van S. cerevisiae of Melody™ (CHR Hansen), wat bestaan uit S. cerevisiae, L. thermotolerans en T. delbrueckii in verskillende verhoudings, is beskikbaar. Om sulke produkte te ontwikkel, is die begrip van genetiese en fenotipiese eienskappe van stamme en interaksies tussen die verskillende organismes, belangrik. Gevolglik het sommige studies gis-gis interaksies en hul meganismes ondersoek, soos antagonistiese interaksies bewerkstellig deur direkte sel kontak of deur groei-inhibitoriese metaboliete. Dit lei gevolglik tot 'n afname van sekere nie- Saccharomyces soos Hanseniaspora en beïnvloed dus die finale aroma samestelling. Omgekeerd, kan sinergistiese effekte waargeneem word in spesies wat bevorder kan word om langer te oorleef tydens fermentasie en daarom dra hulle meer by tot aroma. Die kennis oor populasie dinamika in kulture met meer spesies, hul invloed op alkoholiese fermentasie (AF) en AMG, bly egter beperk. Die huidige studie het 'n multi-spesies gis konsortium ondersoek tydens AF en die effek daarvan op Oenococcus oeni se lewensvatbaarheid gedurende AMG. Die konsortium bestaan uit M. pulcherrima, L. thermotolerans, T. delbrueckii en S. cerevisiae. Fermentasies is onderskeidelik in Chenin blanc en Pinotage by 15 ° C en 25 ° C uitgevoer. In alle proewe het M. pulcherrima vinnig gedaal, terwyl L. thermotolerans tot mid-fermentasie oorleef het. Die beste groei is waargeneem vir T. delbrueckii en dit kon oorleef tot laat in die fermentasie. Fermentasies wat L. thermotolerans bevat, het L-melksuur in die Pinotage, maar nie in die Chenin blanc bevat nie. Daar was geen negatiewe effekte op O. oeni populasies tydens AMG vir beide Pinotage en Chenin blanc wyne waargeneem nie. AMG-kinetika was soortgelyk in al die Pinotage-wyne. In die Chenin blanc is die vinnigste L-appelsuur verbruik vertoon in wyne wat gefermenteer is deur L. thermotolerans of T. delbrueckii saam met S. cerevisiae. Verskillende chemiese profiele is waargeneem met behulp van verswakte totale refleksie infrarooi (ATR-IR) spektroskopie. Chenin Blanc-wyne is meestal aansienlik anders as die S. cerevisiae-kontrole in vergelyking met Pinotage. Met behulp van gaschromatografie is veelvoud veranderinge waargeneem vir baie vlugtige verbindings.

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