Lachancea fermentati Strains Isolated From Kombucha: Fundamental Insights, and Practical Application in Low Alcohol Beer Brewing.

Kristoffer Krogerus,Konstantin Bellut,Elke K Arendt

doi:10.3389/fmicb.2020.00764

Abstract

With a growing interest in non-alcoholic and low alcohol beer (NABLAB), researchers are looking into non-conventional yeasts to harness their special metabolic traits for their production. One of the investigated species is Lachancea fermentati, which possesses the uncommon ability to produce significant amounts of lactic acid during alcoholic fermentation, resulting in the accumulation of lactic acid while exhibiting reduced ethanol production. In this study, four Lachancea fermentati strains isolated from individual kombucha cultures were investigated. Whole genome sequencing was performed, and the strains were characterized for important brewing characteristics (e.g., sugar utilization) and sensitivities toward stress factors. A screening in wort extract was performed to elucidate strain-dependent differences, followed by fermentation optimization to enhance lactic acid production. Finally, a low alcohol beer was produced at 60 L pilot-scale. The genomes of the kombucha isolates were diverse and could be separated into two phylogenetic groups, which were related to their geographical origin. Compared to a Saccharomyces cerevisiae brewers’ yeast, the strains’ sensitivities to alcohol and acidic conditions were low, while their sensitivities toward osmotic stress were higher. In the screening, lactic acid production showed significant, strain-dependent differences. Fermentation optimization by means of response surface methodology (RSM) revealed an increased lactic acid production at a low pitching rate, high fermentation temperature, and high extract content. It was shown that a high initial glucose concentration led to the highest lactic acid production (max. 18.0 mM). The data indicated that simultaneous lactic acid production and ethanol production occurred as long as glucose was present. When glucose was depleted and/or lactic acid concentrations were high, the production shifted toward the ethanol pathway as the sole pathway. A low alcohol beer (<1.3% ABV) was produced at 60 L pilot-scale by means of stopped fermentation. The beer exhibited a balanced ratio of sweetness from residual sugars and acidity from the lactic acid produced (13.6 mM). However, due to the stopped fermentation, high levels of diacetyl were present, which could necessitate further process intervention to reduce concentrations to acceptable levels.

Highlights

Humans have utilized yeasts for the preparation of their foods and beverages long before they even knew of their existence, and beer brewing has been a human activity ever since the Neolithic period (Meussdoerffer, 2009)
There are two fundamentally different approaches when it comes to NABLAB production: physical dealcoholization by means of thermal or membrane methods to remove the ethanol after its formation (Müller et al, 2017), and biological methods like stopped fermentation to limit ethanol production in the first place (Brányik et al, 2012)
The kombucha isolate originating from Australia, KBI 5.3, and the CBS707 type strain had around 20,000 single nucleotide polymorphisms (SNP) compared to the reference genome

Summary

Introduction

Humans have utilized yeasts for the preparation of their foods and beverages long before they even knew of their existence, and beer brewing has been a human activity ever since the Neolithic period (Meussdoerffer, 2009). There are two fundamentally different approaches when it comes to NABLAB production: physical dealcoholization by means of thermal or membrane methods to remove the ethanol after its formation (Müller et al, 2017), and biological methods like stopped fermentation to limit ethanol production in the first place (Brányik et al, 2012) Another old, biological method for NABLAB production has seen a revival in recent years: the application of nonSaccharomyces yeasts ( called non-conventional yeasts) with limited ability to ferment wort sugars, resulting in a low ethanol production. For example, grapes and wine (Saerens and Swiegers, 2014; Estela-Escalante et al, 2016), honey (De Francesco et al, 2015), glaciers in Italy and the Antarctica (Thomas-Hall et al, 2010; De Francesco et al, 2018), Japanese miso (Sohrabvandi et al, 2009; Mohammadi et al, 2011) and, more recently, kombucha (Bellut et al, 2018, 2019a)

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